US20080097089A1 - siRNA targeting deubiqutination enzymes - Google Patents

siRNA targeting deubiqutination enzymes Download PDF

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US20080097089A1
US20080097089A1 US11/977,558 US97755807A US2008097089A1 US 20080097089 A1 US20080097089 A1 US 20080097089A1 US 97755807 A US97755807 A US 97755807A US 2008097089 A1 US2008097089 A1 US 2008097089A1
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sirna
seq
sense strand
base
sirnas
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Anastasia Khvorova
Angela Reynolds
Devin Leake
William Marshall
Steven Read
Stephen Scaringe
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Dharmacon Inc
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Assigned to DHARMACON, INC. reassignment DHARMACON, INC. ASSIGNMENT OF ASSIGNORS INTEREST (SEE DOCUMENT FOR DETAILS). Assignors: MARSHALL, WILLIAM, READ, STEVEN, SCARINGE, STEPHEN, KHVOROVA, ANASTASIA, LEAKE, DEVIN, REYNOLD, ANGELA
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    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
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    • C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
    • C12N15/1135—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing against oncogenes or tumor suppressor genes
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    • C12N2320/10—Applications; Uses in screening processes
    • C12N2320/11—Applications; Uses in screening processes for the determination of target sites, i.e. of active nucleic acids

Definitions

  • the present invention relates to RNA interference (“RNAi”).
  • RNAi RNA interference
  • dsRNA double stranded RNA
  • Double stranded RNA induced gene silencing can occur on at least three different levels: (i) transcription inactivation, which refers to RNA guided DNA or histone methylation; (ii) siRNA induced mRNA degradation; and (iii) mRNA induced transcriptional attenuation.
  • RNAi RNA induced silencing
  • siRNAs small inhibitory RNAs
  • RNA binding Protein RDE-4 Interacts with RDE-1, DCR-1 and a DexH-box Helicase to Direct RNAi in C. elegans, Cell 109(7):861-71; Ketting et al. (2002) Dicer Functions in RNA Interference and in Synthesis of Small RNA Involved in Developmental Timing in C. elegans ; Martinez et al., Single-Stranded Antisense siRNAs Guide Target RNA Cleavage in RNAi, Cell 110(5):563; Hutvagner & Zamore (2002) A microRNA in a multiple-turnover RNAi enzyme complex, Science 297:2056.
  • Dicer Type III endonuclease known as Dicer.
  • Dicer a Type III endonuclease known as Dicer.
  • Dicer a ribonuclease-III-like enzyme, processes the dsRNA into 19-23 base pair short interfering RNAs with characteristic two base 3′ overhangs. Bernstein, Caudy, Hammond, & Hannon (2001) Role for a bidentate ribonuclease in the initiation step of RNA interference, Nature 409:363.
  • RNA-induced silencing complex RISC
  • one or more helicases unwind the siRNA duplex, enabling the complementary antisense strand to guide target recognition.
  • Nykanen, Haley, & Zamore ATP requirements and small interfering RNA structure in the RNA interference pathway, Cell 107:309.
  • one or more endonucleases within the RISC cleaves the target to induce silencing.
  • Elbashir, Lendeckel, & Tuschl (2001) RNA interference is mediated by 21- and 22-nucleotide RNAs, Genes Dev. 15:188, FIG. 1 .
  • RNAi exhibits sequence specificity. Kisielow, M. et al. (2002) Isoform-specific knockdown and expression of adaptor protein ShcA using small interfering RNA, J. Biochem. 363:1-5. Thus, the RNAi machinery can specifically knock down one type of transcript, while not affecting closely related mRNA. These properties make siRNA a potentially valuable tool for inhibiting gene expression and studying gene function and drug target validation. Moreover, siRNAs are potentially useful as therapeutic agents against: (1) diseases that are caused by over-expression or misexpression of genes; and (2) diseases brought about by expression of genes that contain mutations.
  • RNAi RNA-dependent gene silencing depends on a number of factors.
  • One of the most contentious issues in RNAi is the question of the necessity of siRNA design, i.e., considering the sequence of the siRNA used.
  • long dsRNA molecules are cleaved into siRNA by Dicer, thus generating a diverse population of duplexes that can potentially cover the entire transcript.
  • siRNA design is not a crucial element of RNAi.
  • others in the field have begun to explore the possibility that RNAi can be made more efficient by paying attention to the design of the siRNA.
  • none of the reported methods have provided a satisfactory scheme for reliably selecting siRNA with acceptable levels of functionality. Accordingly, there is a need to develop rational criteria by which to select siRNA with an acceptable level of functionality, and to identify siRNA that have this improved level of functionality, as well as to identify siRNAs that are hyperfunctional.
  • the present invention is directed to increasing the efficiency of RNAi, particularly in mammalian systems. Accordingly, the present invention provides kits, siRNAs and methods for increasing siRNA efficacy.
  • the present invention provides a kit for gene silencing, wherein said kit is comprised of a pool of at least two siRNA duplexes, each of which is comprised of a sequence that is complementary to a portion of the sequence of one or more target messenger RNA, and each of which is selected using non-target specific criteria.
  • the present invention provides a method for selecting an siRNA, said method comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; and determining the relative functionality of the at least two siRNAs.
  • the present invention also provides a method for selecting an siRNA wherein said selection criteria are embodied in a formula comprising: ( ⁇ 14)*G 13 ⁇ 13*A 1 ⁇ 12*U 7 ⁇ 11*U 2 ⁇ 10*A 11 ⁇ 10*U 4 ⁇ 10*C 3 ⁇ 10*C 5 ⁇ 10*C 6 ⁇ 9*A 10 ⁇ 9*U 9 ⁇ 9*C 18 ⁇ 8*G 10 ⁇ 7*U 1 ⁇ 7*U 16 ⁇ 7*C 17 ⁇ 7*C 19 +7*U 17 +8*A 2 +8*A 4 +8*A 5 +8*C 4 +9*G 8 +10*A 7 +10*U 18 +11*A 19 +11*C 9 +15*G 1 +18*A 3 +19*U 10 ⁇ Tm ⁇ 3*(GC total ) ⁇ 6*(GC 15-19 ) ⁇ 30*X; or Formula VIII ( ⁇ 8)*A1+( ⁇ 1)*A2+(12)*A3+(7)
  • the invention provides a method for developing an algorithm for selecting siRNA, said method comprising: (a) selecting a set of siRNA; (b) measuring gene silencing ability of each siRNA from said set; (c) determining relative functionality of each siRNA; (d) determining improved functionality by the presence or absence of at least one variable selected from the group consisting of the presence or absence of a particular nucleotide at a particular position, the total number of As and Us in positions 15-19, the number of times that the same nucleotide repeats within a given sequence, and the total number of Gs and Cs; and (e) developing an algorithm using the information of step (d).
  • the present invention provides a kit, wherein said kit is comprised of at least two siRNAs, wherein said at least two siRNAs comprise a first optimized siRNA and a second optimized siRNA, wherein said first optimized siRNA and said second optimized siRNA are optimized according a formula comprising Formula X.
  • the present invention also provides a method for identifying a hyperfunctional siRNA, comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; determining the relative functionality of the at least two siRNAs and assigning each of the at least two siRNAs a functionality score; and selecting siRNAs from the at least two siRNAs that have a functionality score that reflects greater than 80 percent silencing at a concentration in the picomolar range, wherein said greater than 80 percent silencing endures for greater than 120 hours.
  • the present invention provides a hyperfunctional siRNA that is capable of silencing Bcl2.
  • the present invention provides a method for developing an siRNA algorithm for selecting functional and hyperfunctional siRNAs for a given sequence.
  • the method comprises:
  • step (e) developing an algorithm using the information of step (d).
  • the set of siRNAs comprises at least 90 siRNAs from at least one gene, more preferably at least 180 siRNAs from at least two different genes, and most preferably at least 270 and 360 siRNAs from at least three and four different genes, respectively. Additionally, in step (d) the determination is made with preferably at least two, more preferably at least three, even more preferably at least four, and most preferably all of the variables. The resulting algorithm is not target sequence specific.
  • the present invention provides rationally designed siRNAs identified using the formulas above.
  • the present invention is directed to hyperfunctional siRNA.
  • siRNAs that target nucleotide sequences for deubiquitination enzymes are provided.
  • the siRNAs are rationally designed.
  • the siRNAs are functional or hyperfunctional.
  • an siRNA that targets nucleotide sequence for a deubiquitination enzyme is provided, wherein the siRNA is selected from the group consisting of various siRNA sequences targeting nucleotide sequences for deubiquitination enzymes that are disclosed herein.
  • the siRNA sequence is selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • siRNA comprising a sense region and an antisense region are provided, said sense region and said antisense region together form a duplex region comprising 18-30 base pairs, and said sense region comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of siRNA sequences targeting nucleotide sequences for deubiquitination enzymes that are disclosed herein.
  • the siRNA sequence is selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • an siRNA comprising a sense region and an antisense region
  • said sense region and said antisense region together form a duplex region comprising 18-30 base pairs
  • said sense region comprises a sequence that is identical to a contiguous stretch of at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • the duplex region is 19-30 base pairs
  • the sense region comprises a sequence that is identical to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • a pool of at least two siRNAs comprises a first siRNA and a second siRNA, said first siRNA comprising a duplex region of length 18-30 base pairs that has a first sense region that is at least 90% similar to 18 bases of a first sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, and said second siRNA comprises a duplex region of length 18-30 base pairs that has a second sense region that is at least 90% similar to 18 bases of a second sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, wherein said first sense region and said second sense region are not identical.
  • the first sense region comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940
  • said second sense region comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940
  • the duplex of said first siRNA is 19-30 base pairs
  • said first sense region comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940
  • said duplex of said second siRNA is 19-30 base pairs and comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • the duplex of said first siRNA is 19-30 base pairs and said first sense region comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, and said duplex of said second siRNA is 19-30 base pairs and said second region comprises a sequence that is identical to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • FIG. 1 shows a model for siRNA-RISC interactions.
  • RISC has the ability to interact with either end of the siRNA or miRNA molecule. Following binding, the duplex is unwound, and the relevant target is identified, cleaved, and released.
  • FIG. 2 is a representation of the functionality of two hundred and seventy siRNA duplexes that were generated to target human cyclophilin, human diazepam-binding inhibitor (DB), and firefly luciferase.
  • FIG. 3 a is a representation of the silencing effect of 30 siRNAs in three different cells lines, HEK293, DU145, and Hela.
  • FIG. 3 b shows the frequency of different functional groups (>95% silencing (black), >80% silencing (gray), >50% silencing (dark gray), and ⁇ 50% silencing (white)) based on GC content. In cases where a given bar is absent from a particular GC percentage, no siRNA were identified for that particular group.
  • FIG. 3 c shows the frequency of different functional groups based on melting temperature (Tm).
  • FIG. 4 is a representation of a statistical analysis that revealed correlations between silencing and five sequence-related properties of siRNA: (A) an A at position 19 of the sense strand, (B) an A at position 3 of the sense strand, (C) a U at position 10 of the sense strand, (D) a base other than G at position 13 of the sense strand, and (E) a base other than C at position 19 of the sense strand. All variables were correlated with siRNA silencing of firefly luciferase and human cyclophilin. siRNAs satisfying the criterion are grouped on the left (Selected) while those that do not, are grouped on the right (Eliminated). Y-axis is “% Silencing of Control.” Each position on the X-axis represents a unique siRNA.
  • FIGS. 5A and 5B are representations of firefly luciferase and cyclophilin siRNA panels sorted according to functionality and predicted values using Formula VIII.
  • the siRNA found within the circle represent those that have Formula VIII values (SMARTSCORESTM, or siRNA rank) above zero. siRNA outside the indicated area have calculated Formula VIII values that are below zero.
  • Y-axis is “Expression (% Control).” Each position on the X-axis represents a unique siRNA.
  • FIG. 6A is a representation of the average internal stability profile (AISP) derived from 270 siRNAs taken from three separate genes (cyclophilin B, DBI and firefly luciferase). Graphs represent AISP values of highly functional, functional, and non-functional siRNA.
  • FIG. 6B is a comparison between the AISP of naturally derived GFP siRNA (filled squares) and the AISP of siRNA from cyclophilin B, DBI, and luciferase having >90% silencing properties (no fill) for the antisense strand. “DG” is the symbol for ⁇ G, free energy.
  • FIG. 7 is a histogram showing the differences in duplex functionality upon introduction of base pair mismatches.
  • the X-axis shows the mismatch introduced in the siRNA and the position it is introduced (e.g., 8C>A reveals that position 8 (which normally has a C) has been changed to an A).
  • the Y-axis is “% Silencing (Normalized to Control).”
  • the samples on the X-axis represent siRNAs at 100 nM and are, reading from left to right: 1A to C, 1A to G, 1A to U; 2A to C, 2A to G, 2A to U; 3A to C, 3A to G, 3A to U; 4G to A, 4G to C; 4G to U; 5U to A, 5U to C, 5U to G; 6U to A, 6U to C, 6U to G; 7G to A, 7G to C, 7G to U; 8C to A, 8C to G, 8C to U; 9G to A, 9G to C, 9G to U; 10C to A, 10C to G, 10C to U; 11G to A, 11G to C, 11G to U; 12G to A, 12G to C, 12G to U; 13A to C, 13A to G, 13A to U; 14G to A, 14G to C,
  • FIG. 8 is histogram that shows the effects of 5′sense and antisense strand modification with 2′-O-methylation on functionality.
  • FIG. 9 shows a graph of SMARTSCORESTM, or siRNA rank, versus RNAi silencing values for more than 360 siRNA directed against 30 different genes.
  • SiRNA to the right of the vertical bar represent those siRNA that have desirable SMARTSCORESTM, or siRNA rank.
  • FIGS. 10 A-E compare the RNAi of five different genes (SEAP, DBI, PLK, Firefly Luciferase, and Renilla Luciferase) by varying numbers of randomly selected siRNA and four rationally designed (SMART-selected) siRNA chosen using the algorithm described in Formula VIII.
  • RNAi induced by a pool of the four SMART-selected siRNA is reported at two different concentrations (100 and 400 nM).
  • 10 F is a comparison between a pool of randomly selected EGFR siRNA (Pool 1) and a pool of SMART-selected EGFR siRNA (Pool 2). Pool 1, S1-S4 and Pool 2 S1-S4 represent the individual members that made up each respective pool. Note that numbers for random siRNAs represent the position of the 5′ end of the sense strand of the duplex.
  • the Y-axis represents the % expression of the control(s).
  • the X-axis is the percent expression of the control.
  • FIG. 11 shows the Western blot results from cells treated with siRNA directed against twelve different genes involved in the clathrin-dependent endocytosis pathway (CHC, DynII, CALM, CLCa, CLCb, Eps15, Eps15R, Rab5a, Rab5b, Rab5c, P2 subunit of AP-2 and EEA.1).
  • siRNA were selected using Formula VIII.
  • FIG. 12 is a representation of the gene silencing capabilities of rationally-selected siRNA directed against ten different genes (human and mouse cyclophilin, C-myc, human lamin A/C, QB (ubiquinol-cytochrome c reductase core protein I), MEK1 and MEK2, ATE1 (arginyl-tRNA protein transferase), GAPDH, and Eg5).
  • the Y-axis is the percent expression of the control. Numbers 1, 2, 3 and 4 represent individual rationally selected siRNA. “Pool” represents a mixture of the four individual siRNA.
  • FIG. 13 is the sequence of the top ten Bcl2 siRNAs as determined by Formula VIII. Sequences are listed 5′ to 3′.
  • FIG. 14 is the knockdown by the top ten Bcl2 siRNAs at 100 nM concentrations.
  • the Y-axis represents the amount of expression relative to the non-specific (ns) and transfection mixture control.
  • FIG. 15 represents a functional walk where siRNA beginning on every other base pair of a region of the luciferase gene are tested for the ability to silence the luciferase gene.
  • the Y-axis represents the percent expression relative to a control.
  • the X-axis represents the position of each individual siRNA. Reading from left to right across the X-axis, the position designations are 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIGS. 16A and 16B are histograms demonstrating the inhibition of target gene expression by pools of 2 ( 16 A) and 3 ( 16 B) siRNA duplexes taken from the walk described in FIG. 15 .
  • the Y-axis in each represents the percent expression relative to control.
  • the X-axis in each represents the position of the first siRNA in paired pools, or trios of siRNAs. For instance, the first paired pool contains siRNAs 1 and 3.
  • the second paired pool contains siRNAs 3 and 5.
  • Pool 3 (of paired pools) contains siRNAs 5 and 7, and so on.
  • the X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIGS. 17A and 17B are histograms demonstrating the inhibition of target gene expression by pools of 4 ( 17 A) and 5 ( 17 B) siRNA duplexes.
  • the Y-axis in each represents the percent expression relative to control.
  • the X-axis in each represents the position of the first siRNA in each pool.
  • the X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIGS. 18A and 18B are histograms demonstrating the inhibition of target gene expression by siRNAs that are ten ( 18 A) and twenty ( 18 B) base pairs base pairs apart.
  • the Y-axis represents the percent expression relative to a control.
  • the X-axis represents the position of the first siRNA in each pool.
  • the X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIG. 19 shows that pools of siRNAs (dark gray bar) work as well (or better) than the best siRNA in the pool (light gray bar).
  • the Y-axis represents the percent expression relative to a control.
  • the X-axis represents the position of the first siRNA in each pool.
  • the X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIG. 20 shows that the combination of several semifunctional siRNAs (dark gray) result in a significant improvement of gene expression inhibition over individual (semi-functional; light gray) siRNA.
  • the Y-axis represents the percent expression relative to a control.
  • FIGS. 21A, 21B and 21 C show both pools (Library, Lib) and individual siRNAs in inhibition of gene expression of Beta-Galactosidase, Renilla Luciferase and SEAP (alkaline phosphatase).
  • Numbers on the X-axis indicate the position of the 5′-most nucleotide of the sense strand of the duplex.
  • the Y-axis represents the percent expression of each gene relative to a control.
  • Libraries contain 19 nucleotide long siRNAs (not including overhangs) that begin at the following nucleotides: SEAP: Lib 1: 206, 766, 812, 923, Lib 2: 1117, 1280, 1300, 1487, Lib 3: 206, 766, 812, 923, 1117, 1280, 1300, 1487, Lib 4: 206, 812, 1117, 1300, Lib 5: 766, 923, 1280, 1487, Lib 6: 206, 1487; Bgal: Lib 1: 979, 1339, 2029, 2590, Lib 2: 1087, 1783, 2399, 3257, Lib 3: 979, 1783, 2590, 3257, Lib 4: 979, 1087, 1339, 1783, 2029, 2399, 2590, 3257, Lib 5: 979, 1087, 1339, 1783, Lib 6: 2029, 2399, 2590, 3257; Renilla : Lib 1
  • FIG. 22 shows the results of an EGFR and TfnR internalization assay when single gene knockdowns are performed.
  • the Y-axis represents percent internalization relative to control.
  • FIG. 23 shows the results of an EGFR and TfnR internalization assay when multiple genes are knocked down (e.g., Rab5a, b, c).
  • the Y-axis represents the percent internalization relative to control.
  • FIG. 24 shows the simultaneous knockdown of four different genes.
  • siRNAs directed against G6PD, GAPDH, PLK, and UQC were simultaneously introduced into cells. Twenty-four hours later, cultures were harvested and assayed for mRNA target levels for each of the four genes. A comparison is made between cells transfected with individual siRNAs vs. a pool of siRNAs directed against all four genes.
  • FIG. 25 shows the functionality of ten siRNAs at 0.3 nM concentrations.
  • Complementary refers to the ability of polynucleotides to form base pairs with one another. Base pairs are typically formed by hydrogen bonds between nucleotide units in antiparallel polynucleotide strands. Complementary polynucleotide strands can base pair in the Watson-Crick manner (e.g., A to T, A to U, C to G), or in any other manner that allows for the formation of duplexes.
  • Watson-Crick manner e.g., A to T, A to U, C to G
  • uracil rather than thymine is the base that is considered to be complementary to adenosine.
  • a U is denoted in the context of the present invention, the ability to substitute a T is implied, unless otherwise stated.
  • Perfect complementarity or 100% complementarity refers to the situation in which each nucleotide unit of one polynucleotide strand can hydrogen bond with a nucleotide unit of a second polynucleotide strand.
  • Less than perfect complementarity refers to the situation in which some, but not all, nucleotide units of two strands can hydrogen bond with each other. For example, for two 20-mers, if only two base pairs on each strand can hydrogen bond with each other, the polynucleotide strands exhibit 10% complementarity. In the same example, if 18 base pairs on each strand can hydrogen bond with each other, the polynucleotide strands exhibit 90% complementarity.
  • deoxynucleotide refers to a nucleotide or polynucleotide lacking a hydroxyl group (OH group) at the 2′ and/or 3′ position of a sugar moiety. Instead, it has a hydrogen bonded to the 2′ and/or 3′ carbon.
  • deoxynucleotide refers to the lack of an OH group at the 2′ position of the sugar moiety, having instead a hydrogen bonded directly to the 2′ carbon.
  • deoxyribonucleotide and “DNA” refer to a nucleotide or polynucleotide comprising at least one sugar moiety that has an H, rather than an OH, at its 2′ and/or 3′position.
  • duplex region refers to the region in two complementary or substantially complementary polynucleotides that form base pairs with one another, either by Watson-Crick base pairing or any other manner that allows for a stabilized duplex between polynucleotide strands that are complementary or substantially complementary.
  • a polynucleotide strand having 21 nucleotide units can base pair with another polynucleotide of 21 nucleotide units, yet only 19 bases on each strand are complementary or substantially complementary, such that the “duplex region” has 19 base pairs.
  • the remaining bases may, for example, exist as 5′ and 3′ overhangs.
  • 100% complementarity is not required; substantial complementarity is allowable within a duplex region.
  • Substantial complementarity refers to 79% or greater complementarity. For example, a mismatch in a duplex region consisting of 19 base pairs results in 94.7% complementarity, rendering the duplex region substantially complementary.
  • filter refers to one or more procedures that are performed on sequences that are identified by the algorithm.
  • filtering includes in silico procedures where sequences identified by the algorithm can be screened to identify duplexes carrying desirable or undesirable motifs. Sequences carrying such motifs can be selected for, or selected against, to obtain a final set with the preferred properties.
  • filtering includes wet lab experiments. For instance, sequences identified by one or more versions of the algorithm can be screened using any one of a number of procedures to identify duplexes that have hyperfunctional traits (e.g., they exhibit a high degree of silencing at subnanomolar concentrations and/or exhibit high degrees of silencing longevity).
  • gene silencing refers to a process by which the expression of a specific gene product is lessened or attenuated. Gene silencing can take place by a variety of pathways. Unless specified otherwise, as used herein, gene silencing refers to decreases in gene product expression that results from RNA interference (RNAi), a defined, though partially characterized pathway whereby small inhibitory RNA (siRNA) act in concert with host proteins (e.g., the RNA induced silencing complex, RISC) to degrade messenger RNA (mRNA) in a sequence-dependent fashion.
  • RNAi RNA interference
  • host proteins e.g., the RNA induced silencing complex, RISC
  • the level of gene silencing can be measured by a variety of means, including, but not limited to, measurement of transcript levels by Northern Blot Analysis, B-DNA techniques, transcription-sensitive reporter constructs, expression profiling (e.g., DNA chips), and related technologies.
  • the level of silencing can be measured by assessing the level of the protein encoded by a specific gene. This can be accomplished by performing a number of studies including Western Analysis, measuring the levels of expression of a reporter protein that has e.g., fluorescent properties (e.g., GFP) or enzymatic activity (e.g., alkaline phosphatases), or several other procedures.
  • fluorescent properties e.g., GFP
  • enzymatic activity e.g., alkaline phosphatases
  • microRNA refers to microRNA.
  • nucleotide refers to a ribonucleotide or a deoxyribonucleotide or modified form thereof, as well as an analog thereof.
  • Nucleotides include species that comprise purines, e.g., adenine, hypoxanthine, guanine, and their derivatives and analogs, as well as pyrimidines, e.g., cytosine, uracil, thymine, and their derivatives and analogs.
  • Nucleotide analogs include nucleotides having modifications in the chemical structure of the base, sugar and/or phosphate, including, but not limited to, 5-position pyrimidine modifications, 8-position purine modifications, modifications at cytosine exocyclic amines, and substitution of 5-bromo-uracil; and 2′-position sugar modifications, including but not limited to, sugar-modified ribonucleotides in which the 2′-OH is replaced by a group such as an H, OR, R, halo, SH, SR, NH 2 , NHR, NR 2 , or CN, wherein R is an alkyl moiety.
  • Nucleotide analogs are also meant to include nucleotides with bases such as inosine, queuosine, xanthine, sugars such as 2′-methyl ribose, non-natural phosphodiester linkages such as methylphosphonates, phosphorothioates and peptides.
  • Modified bases refer to nucleotide bases such as, for example, adenine, guanine, cytosine, thymine, uracil, xanthine, inosine, and queuosine that have been modified by the replacement or addition of one or more atoms or groups.
  • nucleotide bases such as, for example, adenine, guanine, cytosine, thymine, uracil, xanthine, inosine, and queuosine that have been modified by the replacement or addition of one or more atoms or groups.
  • Some examples of types of modifications that can comprise nucleotides that are modified with respect to the base moieties include but are not limited to, alkylated, halogenated, thiolated, aminated, amidated, or acetylated bases, individually or in combination.
  • More specific examples include, for example, 5-propynyluridine, 5-propynylcytidine, 6-methyladenine, 6-methylguanine, N,N,-dimethyladenine, 2-propyladenine, 2-propylguanine, 2-aminoadenine, 1-methylinosine, 3-methyluridine, 5-methylcytidine, 5-methyluridine and other nucleotides having a modification at the 5 position, 5-(2-amino)propyl uridine, 5-halocytidine, 5-halouridine, 4-acetylcytidine, 1-methyladenosine, 2-methyladenosine, 3-methylcytidine, 6-methyluridine, 2-methylguanosine, 7-methylguanosine, 2,2-dimethylguanosine, 5-methylaminoethyluridine, 5-methyloxyuridine, deazanucleotides such as 7-deaza-adenosine, 6-azouridine, 6-azocytidine, 6-azo
  • Modified nucleotides also include those nucleotides that are modified with respect to the sugar moiety, as well as nucleotides having sugars or analogs thereof that are not ribosyl.
  • the sugar moieties may be, or be based on, mannoses, arabinoses, glucopyranoses, galactopyranoses, 4′-thioribose, and other sugars, heterocycles, or carbocycles.
  • nucleotide is also meant to include what are known in the art as universal bases.
  • universal bases include but are not limited to 3-nitropyrrole, 5-nitroindole, or nebularine.
  • nucleotide is also meant to include the N3′ to P5′ phosphoramidate, resulting from the substitution of a ribosyl 3′ oxygen with an amine group.
  • nucleotide also includes those species that have a detectable label, such as for example a radioactive or fluorescent moiety, or mass label attached to the nucleotide.
  • off-target silencing and “off-target interference” are defined as degradation of mRNA other than the intended target mRNA due to overlapping and/or partial homology with secondary mRNA messages.
  • polynucleotide refers to polymers of nucleotides, and includes but is not limited to DNA, RNA, DNA/RNA hybrids including polynucleotide chains of regularly and/or irregularly alternating deoxyribosyl moieties and ribosyl moieties (i.e., wherein alternate nucleotide units have an —OH, then and —H, then an —OH, then an —H, and so on at the 2′ position of a sugar moiety), and modifications of these kinds of polynucleotides, wherein the attachment of various entities or moieties to the nucleotide units at any position are included.
  • polyribonucleotide refers to a polynucleotide comprising two or more modified or unmodified ribonucleotides and/or their analogs.
  • polyribonucleotide is used interchangeably with the term “oligoribonucleotide.”
  • ribonucleotide and the phrase “ribonucleic acid” (RNA), refer to a modified or unmodified nucleotide or polynucleotide comprising at least one ribonucleotide unit.
  • a ribonucleotide unit comprises an hydroxyl group attached to the 2′ position of a ribosyl moiety that has a nitrogenous base attached in N-glycosidic linkage at the 1′ position of a ribosyl moiety, and a moiety that either allows for linkage to another nucleotide or precludes linkage.
  • siRNA refers to small inhibitory RNA duplexes that induce the RNA interference (RNAi) pathway. These molecules can vary in length (generally 18-30 base pairs) and contain varying degrees of complementarity to their target mRNA in the antisense strand. Some, but not all, siRNA have unpaired overhanging bases on the 5′ or 3′ end of the sense strand and/or the antisense strand.
  • siRNA includes duplexes of two separate strands, as well as single strands that can form hairpin structures comprising a duplex region.
  • siRNA may be divided into five (5) groups (non-functional, semi-functional, functional, highly functional, and hyper-functional) based on the level or degree of silencing that they induce in cultured cell lines. As used herein, these definitions are based on a set of conditions where the siRNA is transfected into said cell line at a concentration of 100 nM and the level of silencing is tested at a time of roughly 24 hours after transfection, and not exceeding 72 hours after transfection. In this context, “non-functional siRNA” are defined as those siRNA that induce less than 50% ( ⁇ 50%) target silencing. “Semi-functional siRNA” induce 50-79% target silencing. “Functional siRNA” are molecules that induce 80-95% gene silencing.
  • “Highly-functional siRNA” are molecules that induce greater than 95% gene silencing. “Hyperfunctional siRNA” are a special class of molecules. For purposes of this document, hyperfunctional siRNA are defined as those molecules that: (1) induce greater than 95% silencing of a specific target when they are transfected at subnanomolar concentrations (i.e., less than one nanomolar); and/or (2) induce functional (or better) levels of silencing for greater than 96 hours. These relative functionalities (though not intended to be absolutes) may be used to compare siRNAs to a particular target for applications such as functional genomics, target identification and therapeutics.
  • SMARTSCORETM or “siRNA rank” refers to a number determined by applying any of the formulas to a given siRNA sequence.
  • SMART-selected or “rationally selected” or “rational selection” refers to siRNA that have been selected on the basis of their SMARTSCORESTM, or siRNA ranking.
  • substantially similar refers to a similarity of at least 90% with respect to the identity of the bases of the sequence.
  • target is used in a variety of different forms throughout this document and is defined by the context in which it is used.
  • Target mRNA refers to a messenger RNA to which a given siRNA can be directed against.
  • Target sequence and “target site” refer to a sequence within the mRNA to which the sense strand of an siRNA shows varying degrees of homology and the antisense strand exhibits varying degrees of complementarity.
  • siRNA target can refer to the gene, mRNA, or protein against which an siRNA is directed.
  • target silencing can refer to the state of a gene, or the corresponding mRNA or protein.
  • transfection refers to a process by which agents are introduced into a cell.
  • the list of agents that can be transfected is large and includes, but is not limited to, siRNA, sense and/or anti-sense sequences, DNA encoding one or more genes and organized into an expression plasmid, proteins, protein fragments, and more.
  • methods for transfecting agents into a cell including, but not limited to, electroporation, calcium phosphate-based transfections, DEAE-dextran-based transfections, lipid-based transfections, molecular conjugate-based transfections (e.g., polylysine-DNA conjugates), microinjection and others.
  • the present invention is directed to improving the efficiency of gene silencing by siRNA. Through the inclusion of multiple siRNA sequences that are targeted to a particular gene and/or selecting an siRNA sequence based on certain defined criteria, improved efficiency may be achieved.
  • the present invention is directed to increasing the efficiency of RNAi, particularly in mammalian systems. Accordingly, the present invention provides kits, siRNAs and methods for increasing siRNA efficacy.
  • the present invention provides a kit for gene silencing, wherein said kit is comprised of a pool of at least two siRNA duplexes, each of which is comprised of a sequence that is complementary to a portion of the sequence of one or more target messenger RNA, and each of which is selected using non-target specific criteria.
  • Each of the at least two siRNA duplexes of the kit complementary to a portion of the sequence of one or more target mRNAs is preferably selected using Formula X.
  • the present invention provides a method for selecting an siRNA, said method comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; and determining the relative functionality of the at least two siRNAs.
  • the present invention also provides a method wherein said selection criteria are embodied in a formula comprising: ( ⁇ 14)*G 13 ⁇ 13*A 1 ⁇ 12*U 7 ⁇ 11*U 2 ⁇ 10*A 11 ⁇ 10*U 4 ⁇ 10*C 3 ⁇ 10*C 5 ⁇ 1*C 6 ⁇ 9*A 10 ⁇ 9*U 9 ⁇ 9*C 18 ⁇ 8*G 10 ⁇ 7*U 1 ⁇ 7*U 16 ⁇ 7*C 17 ⁇ 7*C 19 +7*U 17 +8*A 2 +8*A 4 +8*A 5 +8*C 4 +9*G 8 +10*A 7 +10*U 18 +11*A 19 +11*C 9 +15*G 1 +18*A 3 +19*U 10 ⁇ Tm ⁇ 3*(GC total ) ⁇ 6*(GC 15-19 ) ⁇ 30*X; or Formula VIII ( ⁇ 8)*A1+( ⁇ 1)*A2+(12)*A3+(7)*A4+(18)*
  • position numbering begins at the 5′-most position of a sense strand
  • a 1 1 if A is the base at position 1 of the sense strand, otherwise its value is 0;
  • a 2 1 if A is the base at position 2 of the sense strand, otherwise its value is 0;
  • a 3 1 if A is the base at position 3 of the sense strand, otherwise its value is 0;
  • a 4 1 if A is the base at position 4 of the sense strand, otherwise its value is 0;
  • a 5 1 if A is the base at position 5 of the sense strand, otherwise its value is 0;
  • a 6 1 if A is the base at position 6 of the sense strand, otherwise its value is 0;
  • a 7 1 if A is the base at position 7 of the sense strand, otherwise its value is 0;
  • a 10 1 if A is the base at position 10 of the sense strand, otherwise its value is 0;
  • a 11 1 if A is the base at position 11 of the sense strand, otherwise its value is 0;
  • a 13 1 if A is the base at position 13 of the sense strand, otherwise its value is 0;
  • a 19 1 if A is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • C 19 1 if C is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • G 1 1 if G is the base at position 1 on the sense strand, otherwise its value is 0;
  • G 2 1 if G is the base at position 2 of the sense strand, otherwise its value is 0;
  • G 8 1 if G is the base at position 8 on the sense strand, otherwise its value is 0;
  • G 10 1 if G is the base at position 10 on the sense strand, otherwise its value is 0;
  • GC 15-19 the number of G and C bases within positions 15-19 of the sense strand, or within positions 15-18 if the sense strand is only 18 base pairs in length;
  • GC total the number of G and C bases in the sense strand
  • Tm 100 if the siRNA oligo has the internal repeat longer then 4 base pairs, otherwise its value is 0;
  • X the number of times that the same nucleotide repeats four or more times in a row.
  • any of the methods of selecting siRNA in accordance with the invention can further comprise comparing the internal stability profiles of the siRNAs to be selected, and selecting those siRNAs with the most favorable internal stability profiles. Any of the methods of selecting siRNA can further comprise selecting either for or against sequences that contain motifs that induce cellular stress. Such motifs include, for example, toxicity motifs. Any of the methods of selecting siRNA can further comprise either selecting for or selecting against sequences that comprise stability motifs.
  • the present invention provides a method of gene silencing, comprising introducing into a cell at least one siRNA selected according to any of the methods of the present invention.
  • the siRNA can be introduced by allowing passive uptake of siRNA, or through the use of a vector.
  • the invention provides a method for developing an algorithm for selecting siRNA, said method comprising: (a) selecting a set of siRNA; (b) measuring gene silencing ability of each siRNA from said set; (c) determining relative functionality of each siRNA; (d) determining improved functionality by the presence or absence of at least one variable selected from the group consisting of the presence or absence of a particular nucleotide at a particular position, the total number of As and Us in positions 15-19, the number of times that the same nucleotide repeats within a given sequence, and the total number of Gs and Cs; and (e) developing an algorithm using the information of step (d).
  • the invention provides a method for selecting an siRNA with improved functionality, comprising using the above-mentioned algorithm to identify an siRNA of improved functionality.
  • the present invention provides a kit, wherein said kit is comprised of at least two siRNAs, wherein said at least two siRNAs comprise a first optimized siRNA and a second optimized siRNA, wherein said first optimized siRNA and said second optimized siRNA are optimized according a formula comprising Formula X.
  • the present invention provides a method for identifying a hyperfunctional siRNA, comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; determining the relative functionality of the at least two siRNAs and assigning each of the at least two siRNAs a functionality score; and selecting siRNAs from the at least two siRNAs that have a functionality score that reflects greater than 80 percent silencing at a concentration in the picomolar range, wherein said greater than 80 percent silencing endures for greater than 120 hours.
  • the invention provides kits and/or methods wherein the siRNA are comprised of two separate polynucleotide strands; wherein the siRNA are comprised of a single contiguous molecule such as, for example, a unimolecular siRNA (comprising, for example, either a nucleotide or non-nucleotide loop); wherein the siRNA are expressed from one or more vectors; and wherein two or more genes are silenced by a single administration of siRNA.
  • a unimolecular siRNA comprising, for example, either a nucleotide or non-nucleotide loop
  • the present invention provides a hyperfunctional siRNA that is capable of silencing Bcl2.
  • the present invention provides a method for developing an siRNA algorithm for selecting functional and hyperfunctional siRNAs for a given sequence.
  • the method comprises:
  • step (e) developing an algorithm using the information of step (d).
  • the set of siRNAs comprises at least 90 siRNAs from at least one gene, more preferably at least 180 siRNAs from at least two different genes, and most preferably at least 270 and 360 siRNAs from at least three and four different genes, respectively. Additionally, in step (d) the determination is made with preferably at least two, more preferably at least three, even more preferably at least four, and most preferably all of the variables. The resulting algorithm is not target sequence specific.
  • the present invention provides rationally designed siRNAs identified using the formulas above.
  • the present invention is directed to hyperfunctional siRNA.
  • the methods disclosed herein can be used in conjunction with comparing internal stability profiles of selected siRNAs, and designing an siRNA with a desirable internal stability profile; and/or in conjunction with a selection either for or against sequences that contain motifs that induce cellular stress, for example, cellular toxicity.
  • siRNA(s) can be introduced into the cell by any method known in the art, including passive uptake or through the use of one or more vectors.
  • any of the methods and kits disclosed herein can employ either unimolecular siRNAs, siRNAs comprised of two separate polynucleotide strands, or combinations thereof. Any of the methods disclosed herein can be used in gene silencing, where two or more genes are silenced by a single administration of siRNA(s).
  • the siRNA(s) can be directed against two or more target genes, and administered in a single dose or single transfection, as the case may be.
  • the present invention provides a method for improving the effectiveness of gene silencing for use to silence a particular gene through the selection of an optimal siRNA.
  • An siRNA selected according to this method may be used individually, or in conjunction with the first embodiment, i.e., with one or more other siRNAs, each of which may or may not be selected by this criteria in order to maximize their efficiency.
  • an siRNA is selected for a given gene by using a rational design. That said, rational design can be described in a variety of ways. Rational design is, in simplest terms, the application of a proven set of criteria that enhance the probability of identifying a functional or hyperfunctional siRNA. In one method, rationally designed siRNA can be identified by maximizing one or more of the following criteria:
  • a low GC content preferably between about 30-52%.
  • a Tm which refers to the character of the internal repeat that results in inter- or intramolecular structures for one strand of the duplex, that is preferably not stable at greater than 50° C., more preferably not stable at greater than 37° C., even more preferably not stable at greater than 30° C. and most preferably not stable at greater than 20° C.
  • a C base at position 10 of the sense strand makes a minor contribution to duplex functionality.
  • the absence of a C at position 3 of the sense strand is very important. Accordingly, preferably an siRNA will satisfy as many of the aforementioned criteria as possible.
  • GC content as well as a high number of AU in positions 15-19 of the sense strand, may be important for easement of the unwinding of double stranded siRNA duplex.
  • Duplex unwinding has been shown to be crucial for siRNA functionality in vivo.
  • the internal structure is measured in terms of the melting temperature of the single strand of siRNA, which is the temperature at which 50% of the molecules will become denatured.
  • the positions refer to sequence positions on the sense strand, which is the strand that is identical to the mRNA.
  • At least criteria 1 and 8 are satisfied. In another preferred embodiment, at least criteria 7 and 8 are satisfied. In still another preferred embodiment, at least criteria 1, 8 and 9 are satisfied.
  • the base pair that is not present is the base pair that is located at the 3′ of the sense strand.
  • additional bases are added at the 5′ end of the sense chain and occupy positions ⁇ 1 to ⁇ 11.
  • SEQ. ID NO. 0001 NNANANNNNUCNAANNNNA and SEQ. ID NO. 0028 GUCNNANANNNNUCNAANNNNA both would have A at position 3, A at position 5, U at position 10, C at position 11, A and position 13, A and position 14 and A at position 19.
  • SEQ. ID NO. 0028 would also have C at position ⁇ 1, U at position ⁇ 2 and G at position ⁇ 3.
  • N is any base, A, C, G, or U: SEQ. ID NO. 0001.
  • NNANANNNNUCNAANNNNA SEQ. ID NO. 0002.
  • NNANANNNNUGNAANNNNA SEQ. ID NO. 0003.
  • NNANANNNNUUNAANNNNA SEQ. ID NO. 0004.
  • NNANANNNNUCNCANNNNA SEQ. ID NO. 0005.
  • NNANANNNNUGNCANNNNA SEQ. ID NO. 0006.
  • NNANANNNNUCNUANNNNA SEQ. ID NO. 0008.
  • NNANANNNNUGNUANNNNA SEQ.
  • NNANANNNNUUNUANNNNA SEQ. ID NO. 0010.
  • NNANCNNNNUCNAANNNNA SEQ. ID NO. 0011.
  • NNANCNNNNUGNAANNNNA SEQ. ID NO. 0012.
  • NNANCNNNNUUNAANNNNA SEQ. ID NO. 0013.
  • NNANCNNNNUCNCANNNNA SEQ. ID NO. 0014.
  • NNANCNNNNUGNCANNNNA SEQ. ID NO. 0015.
  • NNANCNNNNUUNCANNNNA SEQ. ID NO. 0016.
  • NNANCNNNNUCNUANNNNA SEQ. ID NO. 0017.
  • NNANCNNNNNNUGNUANNNNA SEQ. ID NO. 0018.
  • NNANCNNNNUUNUANNNNA SEQ. ID NO. 0019.
  • NNANGNNNNUCNAANNNNA SEQ. ID NO. 0020.
  • NNANGNNNNUGNAANNNNA SEQ. ID NO. 0021.
  • NNANGNNNNUUNAANNNNA SEQ. ID NO. 0022.
  • NNANGNNNNUCNCANNNNA SEQ. ID NO. 0023.
  • NNANGNNNNUGNCANNNNA SEQ. ID NO. 0024.
  • NNANGNNNNUUNCANNNNA SEQ. ID NO. 0025.
  • NNANGNNNNUCNUANNNNA SEQ. ID NO. 0026.
  • NNANGNNNNUGNUANNNNA SEQ. ID NO. 0027.
  • NNANGNNNNNUNUANNNNA SEQ. ID NO. 0019.
  • NNANGNNNNUCNAANNNNA SEQ. ID NO. 0020.
  • NNANGNNNNUGNAANNNNA
  • a 19 1 if A is the base at position 19 on the sense strand, otherwise its value is 0,
  • AU 15-19 0-5 depending on the number of A or U bases on the sense strand at positions 15-19;
  • G 13 1 if G is the base at position 13 on the sense strand, otherwise its value is 0;
  • GC the number of G and C bases in the entire sense strand
  • Tm 20° C. 1 if the Tm is greater than 20° C.
  • a 3 1 if A is the base at position 3 on the sense strand, otherwise its value is 0;
  • U 10 1 if U is the base at position 10 on the sense strand, otherwise its value is 0;
  • a 14 1 if A is the base at position 14 on the sense strand, otherwise its value is 0;
  • a 11 1 if A is the base at position 11 of the sense strand, otherwise its value is 0.
  • Formulas I-VII provide relative information regarding functionality. When the values for two sequences are compared for a given formula, the relative functionality is ascertained; a higher positive number indicates a greater functionality. For example, in many applications a value of 5 or greater is beneficial.
  • formulas V and VI might provide greater insights into duplex functionality.
  • formula II might by used in situations where hairpin structures are not observed in duplexes, and formula IV might be applicable for sequences that have higher AU content.
  • application of a given algorithm may identify an unusually large number of potential siRNA sequences, and in those cases, it may be appropriate to re-analyze that sequence with a second algorithm that is, for instance, more stringent.
  • analysis of a sequence with a given formula yields no acceptable siRNA sequences (i.e. low SMARTSCORESTM, or siRNA ranking).
  • analysis of a single sequence with two separate formulas may give rise to conflicting results (i.e.
  • one formula generates a set of siRNA with high SMARTSCORESTM, or siRNA ranking, while the other formula identifies a set of siRNA with low SMARTSCORESTM, or siRNA ranking).
  • weighted factor(s) e.g. GC content
  • the sequence could be analyzed by a third, fourth, or fifth algorithm to identify a set of rationally designed siRNA.
  • GC refers to criteria that select siRNA solely on the basis of GC content.
  • siRNAs that produce ⁇ 70% silencing drops from 23% to 8% and the number of siRNA duplexes that produce >80% silencing rises from 50% to 88.5%.
  • siRNA duplexes with >80% silencing a larger portion of these siRNAs actually silence >95% of the target expression (the new criteria increases the portion from 33% to 50%).
  • the new criteria increases the portion from 33% to 50%.
  • Table II similarly shows the particularly beneficial results of pooling in combination with the aforementioned criteria. However, Table II, which takes into account each of the aforementioned variables, demonstrates even a greater degree of improvement in functionality.
  • the above-described algorithms may be used with or without a computer program that allows for the inputting of the sequence of the mRNA and automatically outputs the optimal siRNA.
  • the computer program may, for example, be accessible from a local terminal or personal computer, over an internal network or over the Internet.
  • RNA duplex of 18-30 base pairs is selected such that it is optimized according a formula selected from: ( ⁇ 14)*G 13 ⁇ 13*A 1 ⁇ 12*U 7 ⁇ 11*U 2 ⁇ 10*A 11 ⁇ 10*U 4 ⁇ 10*C 3 ⁇ 10*C 5 ⁇ 10*C 6 ⁇ 9*A 10 ⁇ 9*U 9 ⁇ 9*C 18 ⁇ 8*G 10 ⁇ 7*U 1 ⁇ 7*U 16 ⁇ 7*C 17 ⁇ 7*C 19 +7*U 17 +8*A 2 +8*A 4 +8*A 5 +8*C 4 +9*G 8 +10*A 7 +10*U 18 +11*A 19 +11*C 9 +15*G 1 +18*A 3 +19*U 10 ⁇ Tm ⁇ 3*(GC total) ⁇ 6*(GC 15-19 ) ⁇ 30*X; and Formula VIII (14.1)*A 3 +(14.
  • a 1 1 if A is the base at position 1 of the sense strand, otherwise its value is 0;
  • a 2 1 if A is the base at position 2 of the sense strand, otherwise its value is 0;
  • a 3 1 if A is the base at position 3 of the sense strand, otherwise its value is 0;
  • a 4 1 if A is the base at position 4 of the sense strand, otherwise its value is 0;
  • a 5 1 if A is the base at position 5 of the sense strand, otherwise its value is 0;
  • a 6 1 if A is the base at position 6 of the sense strand, otherwise its value is 0;
  • a 7 1 if A is the base at position 7 of the sense strand, otherwise its value is 0;
  • a 10 1 if A is the base at position 10 of the sense strand, otherwise its value is 0;
  • a 11 1 if A is the base at position 11 of the sense strand, otherwise its value is 0;
  • a 13 1 if A is the base at position 13 of the sense strand, otherwise its value is 0;
  • a 19 1 if A is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • C 19 1 if C is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • G 1 1 if G is the base at position 1 on the sense strand, otherwise its value is 0;
  • G 2 1 if G is the base at position 2 of the sense strand, otherwise its value is 0;
  • G 8 1 if G is the base at position 8 on the sense strand, otherwise its value is 0;
  • G 10 1 if G is the base at position 10 on the sense strand, otherwise its value is 0;
  • G 13 1 if G is the base at position 13 on the sense strand, otherwise its value is 0;
  • G 19 1 if G is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • U 1 1 if U is the base at position 1 on the sense strand, otherwise its value is 0;
  • U 2 1 if U is the base at position 2 on the sense strand, otherwise its value is 0;
  • U 3 1 if U is the base at position 3 on the sense strand, otherwise its value is 0;
  • U 4 1 if U is the base at position 4 on the sense strand, otherwise its value is 0;
  • U 7 1 if U is the base at position 7 on the sense strand, otherwise its value is 0;
  • U 9 1 if U is the base at position 9 on the sense strand, otherwise its value is 0;
  • U 10 1 if U is the base at position 10 on the sense strand, otherwise its value is 0;
  • U 15 1 if U is the base at position 15 on the sense strand, otherwise its value is 0;
  • U 16 1 if U is the base at position 16 on the sense strand, otherwise its value is 0;
  • U 17 1 if U is the base at position 17 on the sense strand, otherwise its value is 0;
  • U 18 1 if U is the base at position 18 on the sense strand, otherwise its value is 0;
  • GC 15-19 the number of G and C bases within positions 15-19 of the sense strand, or within positions 15-18 if the sense strand is only 18 base pairs in length;
  • GC total the number of G and C bases in the sense strand
  • Tm 100 if the siRNA oligo has the internal repeat longer then 4 base pairs, otherwise its value is 0;
  • X the number of times that the same nucleotide repeats four or more times in a row.
  • formulas VIII, IX, and X provide methods for selecting siRNA in order to increase the efficiency of gene silencing.
  • a subset of variables of any of the formulas may be used, though when fewer variables are used, the optimization hierarchy becomes less reliable.
  • a single letter of A or C or G or U followed by a subscript refers to a binary condition.
  • the binary condition is that either the particular base is present at that particular position (wherein the value is “1”) or the base is not present (wherein the value is “0”). Because position 19 is optional, i.e., there might be only 18 base pairs, when there are only 18 base pairs, any base with a subscript of 19 in the formulas above would have a zero value for that parameter.
  • each variable is a number followed by *, which indicates that the value of the variable is to be multiplied or weighed by that number.
  • the numbers preceding the variables A, or G, or C, or U in Formulas VIII, IX, and X were determined by comparing the difference in the frequency of individual bases at different positions in functional siRNA and total siRNA. Specifically, the frequency in which a given base was observed at a particular position in functional groups was compared with the frequency that that same base was observed in the total, randomly selected siRNA set. If the absolute value of the difference between the functional and total values was found to be greater than 6%, that parameter was included in the equation.
  • the inventors When developing a means to optimize siRNAs, the inventors observed that a bias toward low internal thermodynamic stability of the duplex at the 5′-antisense (AS) end is characteristic of naturally occurring miRNA precursors. The inventors extended this observation to siRNAs for which functionality had been assessed in tissue culture.
  • AS 5′-antisense
  • a value of 0-5 will be ascribed depending on the number of G or C bases at positions 15 to 19. If there are only 18 base pairs, the value is between 0 and 4.
  • GC total content a number from 0-30 will be ascribed, which correlates to the total number of G and C nucleotides on the sense strand, excluding overhangs.
  • significance of the GC content (as well as AU content at positions 15-19, which is a parameter for formulas III-VII) relates to the easement of the unwinding of a double-stranded siRNA duplex.
  • Duplex unwinding is believed to be crucial for siRNA functionality in vivo and overall low internal stability, especially low internal stability of the first unwound base pair is believed to be important to maintain sufficient processivity of RISC complex-induced duplex unwinding.
  • RISC is a complex of approximately twelve proteins; Dicer is one, but not the only, helicase within this complex. Accordingly, although the GC parameters are believed to relate to activity with Dicer, they are also important for activity with other RISC proteins.
  • the value of the parameter Tm is 0 when there are no internal repeats longer than (or equal to) four base pairs present in the siRNA duplex; otherwise the value is 1.
  • the value will be one (1).
  • the value will be zero (0).
  • RNA the “target RNA” or “target molecule”
  • a computer program to evaluate the criteria for every sequence of 18-30 base pairs or only sequences of a fixed length, e.g., 19 base pairs.
  • the computer program is designed such that it provides a report ranking of all of the potential siRNAs 18-30 base pairs, ranked according to which sequences generate the highest value. A higher value refers to a more efficient siRNA for a particular target gene.
  • the computer program that may be used may be developed in any computer language that is known to be useful for scoring nucleotide sequences, or it may be developed with the assistance of commercially available product such as Microsoft's PRODUCT.NET.
  • BLAST Basic Local Alignment Search Tool
  • Formulas I-VII either Formula VIII, Formula IX, or Formula X may be used for a given mRNA target sequence. However, it is possible that according to one or the other formula more than one siRNA will have the same value. Accordingly, it is beneficial to have a second formula by which to differentiate sequences.
  • Formulas IX and X were derived in a similar fashion as Formula VIII, yet used a larger data set and thus yields sequences with higher statistical correlations to highly functional duplexes.
  • the sequence that has the highest value ascribed to it may be referred to as a “first optimized duplex.”
  • the sequence that has the second highest value ascribed to it may be referred to as a “second optimized duplex.”
  • the sequences that have the third and fourth highest values ascribed to them may be referred to as a third optimized duplex and a fourth optimized duplex, respectively.
  • each of them may, for example, be referred to as first optimized duplex sequences or co-first optimized duplexes.
  • Formula X is similar to Formula IX, yet uses a greater numbers of variables and for that reason, identifies sequences on the basis of slightly different criteria.
  • siRNA sequences identified using Formula VIII and Formula X are contained within the sequence listing.
  • the data included in the sequence listing is described more fully below.
  • the sequences identified by Formula VIII and Formula X that are disclosed in the sequence listing may be used in gene silencing applications.
  • Formulas I-X may be used to select or to evaluate one, or more than one, siRNA in order to optimize silencing.
  • at least two optimized siRNAs that have been selected according to at least one of these formulas are used to silence a gene, more preferably at least three and most preferably at least four.
  • the siRNAs may be used individually or together in a pool or kit. Further, they may be applied to a cell simultaneously or separately. Preferably, the at least two siRNAs are applied simultaneously. Pools are particularly beneficial for many research applications. However, for therapeutics, it may be more desirable to employ a single hyperfunctional siRNA as described elsewhere in this application.
  • siRNAs When planning to conduct gene silencing, and it is necessary to choose between two or more siRNAs, one should do so by comparing the relative values when the siRNA are subjected to one of the formulas above. In general a higher scored siRNA should be used.
  • Useful applications include, but are not limited to, target validation, gene functional analysis, research and drug discovery, gene therapy and therapeutics. Methods for using siRNA in these applications are well known to persons of skill in the art.
  • siRNA Because the ability of siRNA to function is dependent on the sequence of the RNA and not the species into which it is introduced, the present invention is applicable across a broad range of species, including but not limited to all mammalian species, such as humans, dogs, horses, cats, cows, mice, hamsters, chimpanzees and gorillas, as well as other species and organisms such as bacteria, viruses, insects, plants and C. elegans.
  • the present invention is also applicable for use for silencing a broad range of genes, including but not limited to the roughly 45,000 genes of a human genome, and has particular relevance in cases where those genes are associated with diseases such as diabetes, Alzheimer's, cancer, as well as all genes in the genomes of the aforementioned organisms.
  • siRNA selected according to the aforementioned criteria or one of the aforementioned algorithms are also, for example, useful in the simultaneous screening and functional analysis of multiple genes and gene families using high throughput strategies, as well as in direct gene suppression or silencing.
  • siRNA panel consisting of 270 siRNAs targeting three genes, Human Cyclophilin, Firefly Luciferase, and Human DBI. In all three cases, siRNAs were directed against specific regions of each gene. For Human Cyclophilin and Firefly Luciferase, ninety siRNAs were directed against a 199 bp segment of each respective mRNA. For DBI, 90 siRNAs were directed against a smaller, 109 base pair region of the mRNA. The sequences to which the siRNAs were directed are provided below.
  • t is present. This is because many databases contain information in this manner. However, the t denotes a uracil residue in mRNA and siRNA. Any algorithm will, unless otherwise specified, process at in a sequence as a u.
  • the set of duplexes was analyzed to identify correlations between siRNA functionality and other biophysical or thermodynamic properties.
  • siRNA panel was analyzed in functional and non-functional subgroups, certain nucleotides were much more abundant at certain positions in functional or non-functional groups. More specifically, the frequency of each nucleotide at each position in highly functional siRNA duplexes was compared with that of nonfunctional duplexes in order to assess the preference for or against any given nucleotide at every position.
  • the data set was also analyzed for distinguishing biophysical properties of siRNAs in the functional group, such as optimal percent of GC content, propensity for internal structures and regional thermodynamic stability. Of the presented criteria, several are involved in duplex recognition, RISC activation/duplex unwinding, and target cleavage catalysis.
  • FIG. 2 The original data set that was the source of the statistically derived criteria is shown in FIG. 2 . Additionally, this figure shows that random selection yields siRNA duplexes with unpredictable and widely varying silencing potencies as measured in tissue culture using HEK293 cells.
  • duplexes are plotted such that each x-axis tick-mark represents an individual siRNA, with each subsequent siRNA differing in target position by two nucleotides for Human Cyclophilin B and Firefly Luciferase, and by one nucleotide for Human DBI.
  • the y-axis denotes the level of target expression remaining after transfection of the duplex into cells and subsequent silencing of the target.
  • FIG. 3 a shows the evaluation of thirty siRNAs targeting the DBI gene in three cell lines derived from different tissues.
  • Each DBI siRNA displays very similar functionality in HEK293 (ATCC, CRL-1573, human embryonic kidney), HeLa (ATCC, CCL-2, cervical epithelial adenocarcinoma) and DU145 (HTB-81, prostate) cells as determined by the B-DNA assay.
  • HEK293 ATCC, CRL-1573, human embryonic kidney
  • HeLa ATCC, CCL-2, cervical epithelial adenocarcinoma
  • DU145 HTB-81, prostate
  • the complementary sequence of the silencing siRNA may be present in more than one gene. Accordingly, in these circumstances, it may be desirable not to use the siRNA with highest SMARTSCORETM, or siRNA ranking. In such circumstances, it may be desirable to use the siRNA with the next highest SMARTSCORETM, or siRNA ranking.
  • the G/C content of each duplex in the panel was calculated and the functional classes of siRNAs ( ⁇ F50, ⁇ F50, ⁇ F80, ⁇ F95 where F refers to the percent gene silencing) were sorted accordingly.
  • the group with extremely low GC content (26% or less) contained a higher proportion of non-functional siRNAs and no highly-functional siRNAs.
  • the G/C content range of 30%-52% was therefore selected as Criterion I for siRNA functionality, consistent with the observation that a G/C range 30%-70% promotes efficient RNAi targeting.
  • the siRNA panel presented here permitted a more systematic analysis and quantification of the importance of this criterion than that used previously.
  • a relative measure of local internal stability is the A/U base pair (bp) content; therefore, the frequency of A/U bp was determined for each of the five terminal positions of the duplex (5′ sense (S)/5′ antisense (AS)) of all siRNAs in the panel. Duplexes were then categorized by the number of A/U bp in positions 1-5 and 15-19 of the sense strand. The thermodynamic flexibility of the duplex 5′-end (positions 1-5; S) did not appear to correlate appreciably with silencing potency, while that of the 3′-end (positions 15-19; S) correlated with efficient silencing. No duplexes lacking A/U bp in positions 15-19 were functional.
  • the complementary strands of siRNAs that contain internal repeats or palindromes may form internal fold-back structures. These hairpin-like structures exist in equilibrium with the duplexed form effectively reducing the concentration of functional duplexes.
  • the propensity to form internal hairpins and their relative stability can be estimated by predicted melting temperatures. High Tm reflects a tendency to form hairpin structures. Lower Tm values indicate a lesser tendency to form hairpins.
  • FIG. 4 shows the results of these queries and the subsequent resorting of the data set (from FIG. 2 ).
  • the data is separated into two sets: those duplexes that meet the criteria, a specific nucleotide in a certain position—grouped on the left (Selected) and those that do not—grouped on the right (Eliminated).
  • the duplexes are further sorted from most functional to least functional with the y-axis of FIG.
  • FIG. 4 and Table IV show quantitative analysis for the following five sequence-related properties of siRNA: (A) an A at position 19 of the sense strand; (B) an A at position 3 of the sense strand; (C) a U at position 10 of the sense strand; (D) a base other than G at position 13 of the sense strand; and (E) a base other than C at position 19 of the sense strand.
  • Another sequence-related property correlated with silencing was the presence of an A in position 3 of the sense strand ( FIG. 4 b ). Of the siRNAs with A3, 34.4% were F95, compared with 21.7% randomly selected siRNAs. The presence of a U base in position 10 of the sense strand exhibited an even greater impact ( FIG. 4 c ). Of the duplexes in this group, 41.7% were F95. These properties became criteria V and VI, respectively.
  • FIG. 4 Two negative sequence-related criteria that were identified also appear on FIG. 4 .
  • lack of a C at position 19 of the sense strand also correlated with functionality ( FIG. 4 e ).
  • position 19 was most likely occupied by A, and rarely occupied by C.
  • siRNA ranking a score (referred to as a SMARTSCORETM, or siRNA ranking) according to the values derived from the formulas.
  • all duplexes scoring lower than 0 and ⁇ 20 (minus 20) according to formulas VIII and IX, respectively contained some functional siRNAs but included all non-functional siRNAs.
  • the difference in the frequency of a given attribute e.g., GC content, base preference
  • individual functional groups e.g., ⁇ F50
  • the total siRNA population studied e.g., 270 siRNA molecules selected randomly.
  • Criterion I (30%-52% GC content) members of the ⁇ F50 group were observed to have GC contents between 30-52% in 16.4% of the cases.
  • the total group of 270 siRNAs had GC contents in this range, 20% of the time.
  • the >F95 group contained a “U” at this position 41.7% of the time.
  • the total group of 270 siRNAs had a “U” at this position 21.7% of the time, thus the improvement over random is calculated to be 20% (or 41.7%-21.7%).
  • siRNAs derived from the cyclophilin B, the diazepam binding inhibitor (DBI), and the luciferase gene were individually transfected into HEK293 cells and tested for their ability to induce RNAi of the respective gene. Based on their performance in the in vivo assay, the sequences were then subdivided into three groups, (i) >95% silencing; (ii) 80-95% silencing; and (iii) less than 50% silencing. Sequences exhibiting 51-84% silencing were eliminated from further consideration to reduce the difficulties in identifying relevant thermodynamic patterns.
  • siRNA molecules that were critical for successful gene silencing.
  • highly functional siRNA >95% gene silencing, see FIG. 6 a , >F95
  • SAP internal stability
  • low-efficiency siRNA i.e., those exhibiting less than 50% silencing, ⁇ F50
  • siRNAs with poor silencing capabilities show a distinctly different profile. While the AISP value at position 12 is nearly identical with that of strong siRNAs, the values at positions 7 and 8 rise considerably, peaking at a high of ⁇ 9.0 kcal/mol. In addition, at the 5′ end of the molecule the AISP profile of strong and weak siRNA differ dramatically. Unlike the relatively strong values exhibited by siRNA in the >95% silencing group, siRNAs that exhibit poor silencing activity have weak AISP values ( ⁇ 7.6, ⁇ 7.5, and ⁇ 7.5 kcal/mol for positions 1, 2 and 3 respectively).
  • siRNA that have strong or even stronger gene-specific silencing effects might have exaggerated ⁇ G values (either higher or lower) at key positions.
  • ⁇ G values either higher or lower
  • the 5′-most position of the sense strand position 19
  • position 12 and position 7 could have values above 8.3 kcal/mol and below 7.7 kcal/mole, respectively, without abating the silencing effectiveness of the molecule.
  • a stabilizing chemical modification e.g., a chemical modification of the 2′ position of the sugar backbone
  • a stabilizing chemical modification e.g., a chemical modification of the 2′ position of the sugar backbone
  • mismatches similar to those described previously could be introduced that would lower the AG values at that position.
  • non-functional siRNA are defined as those siRNA that induce less than 50% ( ⁇ 50%) target silencing
  • siRNA induce 50-79% target silencing
  • functional siRNA are molecules that induce 80-95% gene silencing
  • highly-functional siRNA are molecules that induce great than 95% gene silencing.
  • siRNA that reduces gene activity by only 30%. While this level of gene silencing may be “non-functional” for, e.g., therapeutic needs, it is sufficient for gene mapping purposes and is, under these uses and conditions, “functional.” For these reasons, functional siRNA can be defined as those molecules having greater than 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% silencing capabilities at 100 nM transfection conditions. Similarly, depending upon the needs of the study and/or application, non-functional and semi-functional siRNA can be defined as having different parameters.
  • semi-functional siRNA can be defined as being those molecules that induce 20%, 30%, 40%, 50%, 60%, or 70% silencing at 100 nM transfection conditions.
  • non-functional siRNA can be defined as being those molecules that silence gene expression by less than 70%, 60%, 50%, 40%, 30%, or less. Nonetheless, unless otherwise stated, the descriptions stated in the “Definitions” section of this text should be applied.
  • Functional attributes can be assigned to each of the key positions in the AISP of strong siRNA.
  • the low 5′ (sense strand) AISP values of strong siRNAs may be necessary for determining which end of the molecule enters the RISC complex.
  • the high and low AISP values observed in the central regions of the molecule may be critical for siRNA-target mRNA interactions and product release, respectively.
  • siRNA functionality is likely influenced by specific biophysical and molecular properties that promote efficient interactions within the context of the multi-component complexes. Indeed, the systematic analysis of the siRNA test set identified multiple factors that correlate well with functionality. When combined into a single algorithm, they proved to be very effective in selecting active siRNAs.
  • RNAi may also be predictive of key functional associations important for each step in RNAi.
  • the potential formation of internal hairpin structures correlated negatively with siRNA functionality.
  • Complementary strands with stable internal repeats are more likely to exist as stable hairpins thus decreasing the effective concentration of the functional duplex form.
  • the duplex is the preferred conformation for initial pre-RISC association.
  • the effective concentration required is at least two orders of magnitude higher than that of the duplex form.
  • siRNA-pre-RISC complex formation is followed by an ATP-dependent duplex unwinding step and “activation” of the RISC.
  • the siRNA functionality was shown to correlate with overall low internal stability of the duplex and low internal stability of the 3′ sense end (or differential internal stability of the 3′ sense compare to the 5′ sense strand), which may reflect strand selection and entry into the RISC.
  • Overall duplex stability and low internal stability at the 3′ end of the sense strand were also correlated with siRNA functionality.
  • siRNAs with very high and very low overall stability profiles correlate strongly with non-functional duplexes.
  • One interpretation is that high internal stability prevents efficient unwinding while very low stability reduces siRNA target affinity and subsequent mRNA cleavage by the RISC.
  • Base preferences for A at position 19 of the sense strand but not C are particularly interesting because they reflect the same base preferences observed for naturally occurring miRNA precursors. That is, among the reported miRNA precursor sequences 75% contain a U at position 1 which corresponds to an A in position 19 of the sense strand of siRNAs, while G was under-represented in this same position for miRNA precursors.
  • the functional interpretation of the predominance of a U/A base pair is that it promotes flexibility at the 5′antisense ends of both siRNA duplexes and miRNA precursors and facilitates efficient unwinding and selective strand entrance into an activated RISC.
  • RISC preferentially cleaves target mRNA between nucleotides 10 and 11 relative to the 5′ end of the complementary targeting strand. Therefore, it may be that U, the preferred base for most endoribonucleases, at this position supports more efficient cleavage.
  • a U/A bp between the targeting siRNA strand and its cognate target mRNA may create an optimal conformation for the RISC-associated “slicing” activity.
  • the output of any one of the formulas previously listed can be filtered to remove or select for siRNAs containing undesirable or desirable motifs or properties, respectively.
  • sequences identified by any of the formulas can be filtered to remove any and all sequences that induce toxicity or cellular stress.
  • Introduction of an siRNA containing a toxic motif into a cell can induce cellular stress and/or cell death (apoptosis) which in turn can mislead researchers into associating a particular (e.g., nonessential) gene with, e.g., an essential function.
  • sequences generated by any of the before mentioned formulas can be filtered to identify and retain duplexes that contain toxic motifs.
  • duplexes may be valuable from a variety of perspectives including, for instance, uses as therapeutic molecules.
  • a variety of toxic motifs exist and can exert their influence on the cell through RNAi and non-RNAi pathways. Examples of toxic motifs are explained more fully in commonly assigned U.S. Provisional Patent Application Ser. No. 60/538,874, entitled “Identification of Toxic Sequences,” filed Jan. 23, 2004. Briefly, toxic motifs include A/G UUU A/G/U, G/C AAA G/C, and GCCA, or a complement of any of the foregoing.
  • sequences identified by any of the before mentioned formulas can be filtered to identify duplexes that contain motifs (or general properties) that provide serum stability or induce serum instability.
  • duplexes targeting disease-associated genes will be introduced into patients intravenously.
  • post-algorithm filters designed to select molecules that contain motifs that enhance duplex stability in the presence of serum and/or (conversely) eliminate duplexes that contain motifs that destabilize siRNA in the presence of serum, would be beneficial.
  • sequences identified by any of the before mentioned formulas can be filtered to identify duplexes that are hyperfunctional.
  • Hyperfunctional sequences are defined as those sequences that (1) induce greater than 95% silencing of a specific target when they are transfected at subnanomolar concentrations (i.e., less than one nanomolar); and/or (2) induce functional (or better) levels of silencing for greater than 96 hours.
  • Filters that identify hyperfunctional molecules can vary widely. In one example, the top ten, twenty, thirty, or forty siRNA can be assessed for the ability to silence a given target at, e.g., concentrations of 1 nM and 0.5 nM to identify hyperfunctional molecules.
  • the present invention provides a pool of at least two siRNAs, preferably in the form of a kit or therapeutic reagent, wherein one strand of each of the siRNAs, the sense strand comprises a sequence that is substantially similar to a sequence within a target mRNA.
  • the opposite strand, the antisense strand will preferably comprise a sequence that is substantially complementary to that of the target mRNA.
  • one strand of each siRNA will comprise a sequence that is identical to a sequence that is contained in the target mRNA.
  • each siRNA will be 19 base pairs in length, and one strand of each of the siRNAs will be 100% complementary to a portion of the target mRNA.
  • siRNAs directed to a particular target By increasing the number of siRNAs directed to a particular target using a pool or kit, one is able both to increase the likelihood that at least one siRNA with satisfactory functionality will be included, as well as to benefit from additive or synergistic effects. Further, when two or more siRNAs directed against a single gene do not have satisfactory levels of functionality alone, if combined, they may satisfactorily promote degradation of the target messenger RNA and successfully inhibit translation. By including multiple siRNAs in the system, not only is the probability of silencing increased, but the economics of operation are also improved when compared to adding different siRNAs sequentially. This effect is contrary to the conventional wisdom that the concurrent use of multiple siRNA will negatively impact gene silencing (e.g., Holen, T. et al. (2003) Similar behavior of single strand and double strand siRNAs suggests they act through a common RNAi pathway. NAR 31: 2401-21407).
  • the kit is comprised of at least three siRNAs, wherein one strand of each siRNA comprises a sequence that is substantially similar to a sequence of the target mRNA and the other strand comprises a sequence that is substantially complementary to the region of the target mRNA.
  • one strand will comprise a sequence that is identical to a sequence that is contained in the mRNA and another strand that is 100% complementary to a sequence that is contained in the mRNA.
  • the kit is comprised of at least four siRNAs, wherein one strand of each siRNA comprises a sequence that is substantially similar to a region of the sequence of the target mRNA, and the other strand comprises a sequence that is substantially complementary to the region of the target mRNA.
  • one strand of each of the siRNA duplexes will comprise a sequence that is identical to a sequence that is contained in the mRNA, and another strand that is 100% complementary to a sequence that is contained in the mRNA.
  • kits and pools with at least five, at least six, and at least seven siRNAs may also be useful with the present invention.
  • pools of five siRNA induced 95% gene silencing with 77% probability and 80% silencing with 98.8% probability.
  • pooling of siRNAs together can result in the creation of a target-specific silencing reagent with almost a 99% probability of being functional.
  • the fact that such high levels of success are achievable using such pools of siRNA enables one to dispense with costly and time-consuming target-specific validation procedures.
  • each of the siRNAs within a pool will preferably comprise 18-30 base pairs, more preferably 18-25 base pairs, and most preferably 19 base pairs.
  • at least 18 contiguous bases of the antisense strand will be 100% complementary to the target mRNA. More preferably, at least 19 contiguous bases of the antisense strand will be 100% complementary to the target mRNA.
  • there may be overhangs on either the sense strand or the antisense strand and these overhangs may be at either the 5′ end or the 3′ end of either of the strands, for example there may be one or more overhangs of 1-6 bases.
  • overhangs When overhangs are present, they are not included in the calculation of the number of base pairs.
  • the two nucleotide 3′ overhangs mimic natural siRNAs and are commonly used but are not essential.
  • the overhangs should consist of two nucleotides, most often dTdT or UU at the 3′ end of the sense and antisense strand that are not complementary to the target sequence.
  • the siRNAs may be produced by any method that is now known or that comes to be known for synthesizing double stranded RNA that one skilled in the art would appreciate would be useful in the present invention.
  • the siRNAs will be produced by Dharmacon's proprietary ACE® technology.
  • siRNAs are well known to persons skilled in the art and include, but are not limited to, any chemical synthesis of RNA oligonucleotides, ligation of shorter oligonucleotides, in vitro transcription of RNA oligonucleotides, the use of vectors for expression within cells, recombinant Dicer products and PCR products.
  • siRNA duplexes within the aforementioned pools of siRNAs may correspond to overlapping sequences within a particular mRNA, or non-overlapping sequences of the mRNA. However, preferably they correspond to non-overlapping sequences. Further, each siRNA may be selected randomly, or one or more of the siRNA may be selected according to the criteria discussed above for maximizing the effectiveness of siRNA.
  • siRNAs that contain substituted and/or labeled nucleotides that may, for example, be labeled by radioactivity, fluorescence or mass.
  • the most common substitutions are at the 2′ position of the ribose sugar, where moieties such as H (hydrogen) F, NH 3 , OCH 3 and other O— alkyl, alkenyl, alkynyl, and orthoesters, may be substituted, or in the phosphorous backbone, where sulfur, amines or hydrocarbons may be substituted for the bridging of non-bridging atoms in the phosphodiester bond.
  • H (hydrogen) F NH 3 , OCH 3 and other O— alkyl, alkenyl, alkynyl, and orthoesters
  • sulfur, amines or hydrocarbons may be substituted for the bridging of non-bridging atoms in the phosphodiester bond.
  • the cell type into which the siRNA is introduced may affect the ability of the siRNA to enter the cell; however, it does not appear to affect the ability of the siRNA to function once it enters the cell.
  • Methods for introducing double-stranded RNA into various cell types are well known to persons skilled in the art.
  • the presence of proteins such as RdRP, the RNA-dependent RNA polymerase may catalytically enhance the activity of the siRNA.
  • RdRP propagates the RNAi effect in C. elegans and other non-mammalian organisms.
  • the siRNA may be inherited.
  • Two other proteins that are well studied and known to be a part of the machinery are members of the Argonaute family and Dicer, as well as their homologues.
  • the RISC complex might be associated with the ribosome so the more efficiently translated mRNAs will be more susceptible to silencing than others.
  • siRNA localization Another very important factor in the efficacy of siRNA is mRNA localization. In general, only cytoplasmic mRNAs are considered to be accessible to RNAi to any appreciable degree. However, appropriately designed siRNAs, for example, siRNAs modified with internucleotide linkages or 2′-O-methyl groups, may be able to cause silencing by acting in the nucleus. Examples of these types of modifications are described in commonly assigned U.S. patent application Ser. Nos. 10/431,027 and 10/613,077.
  • the effectiveness of the two may be greater than one would predict based on the effectiveness of two individual siRNAs.
  • This additive or synergistic effect is particularly noticeable as one increases to at least three siRNAs, and even more noticeable as one moves to at least four siRNAs.
  • the pooling of the non-functional and semi-functional siRNAs, particularly more than five siRNAs can lead to a silencing mixture that is as effective if not more effective than any one particular functional siRNA.
  • each siRNA will be present in a concentration of between 0.001 and 200 ⁇ M, more preferably between 0.01 and 200 nM, and most preferably between 0.1 and 10 nM.
  • kits of the present invention will also preferably comprise a buffer to keep the siRNA duplex stable.
  • the buffer may be comprised of 100 mM KCl, 30 mM HEPES-pH 7.5, and 1 mM MgCl 2 .
  • kits might contain complementary strands that contain any one of a number of chemical modifications (e.g., a 2′-O-ACE) that protect the agents from degradation by nucleases. In this instance, the user may (or may not) remove the modifying protective group (e.g., deprotect) before annealing the two complementary strands together.
  • kits may be organized such that pools of siRNA duplexes are provided on an array or microarray of wells or drops for a particular gene set or for unrelated genes.
  • the array may, for example, be in 96 wells, 384 wells or 1284 wells arrayed in a plastic plate or on a glass slide using techniques now known or that come to be known to persons skilled in the art.
  • controls such as functional anti-lamin A/C, cyclophilin and two siRNA duplexes that are not specific to the gene of interest.
  • siRNA pools may be retained in lyophilized form at minus twenty degrees ( ⁇ 20° C.) until they are ready for use. Prior to usage, they should be resuspended; however, even once resuspended, for example, in the aforementioned buffer, they should be kept at minus twenty degrees, ( ⁇ 20° C.) until used.
  • the aforementioned buffer, prior to use, may be stored at approximately 4° C. or room temperature. Effective temperatures at which to conduct transfections are well known to persons skilled in the art and include for example, room temperature.
  • kits may be applied either in vivo or in vitro.
  • the siRNA of the pools or kits is applied to a cell through transfection, employing standard transfection protocols. These methods are well known to persons skilled in the art and include the use of lipid-based carriers, electroporation, cationic carriers, and microinjection. Further, one could apply the present invention by synthesizing equivalent DNA sequences (either as two separate, complementary strands, or as hairpin molecules) instead of siRNA sequences and introducing them into cells through vectors. Once in the cells, the cloned DNA could be transcribed, thereby forcing the cells to generate the siRNA.
  • vectors suitable for use with the present application include but are not limited to the standard transient expression vectors, adenoviruses, retroviruses, lentivirus-based vectors, as well as other traditional expression vectors. Any vector that has an adequate siRNA expression and procession module may be used. Furthermore, certain chemical modifications to siRNAs, including but not limited to conjugations to other molecules, may be used to facilitate delivery. For certain applications it may be preferable to deliver molecules without transfection by simply formulating in a physiological acceptable solution.
  • another embodiment includes the use of multiple siRNA targeting multiple genes. Multiple genes may be targeted through the use of high- or hyper-functional siRNA. High- or hyper-functional siRNA that exhibit increased potency, require lower concentrations to induce desired phenotypic (and thus therapeutic) effects. This circumvents RISC saturation. It therefore reasons that if lower concentrations of a single siRNA are needed for knockout or knockdown expression of one gene, then the remaining (uncomplexed) RISC will be free and available to interact with siRNA directed against two, three, four, or more, genes. Thus in this embodiment, the authors describe the use of highly functional or hyper-functional siRNA to knock out three separate genes.
  • siRNA of this type could be used to knockout or knockdown the expression of six or more genes.
  • hyperfunctional siRNA describes a subset of the siRNA population that induces RNAi in cells at low- or sub-nanomolar concentrations for extended periods of time. These traits, heightened potency and extended longevity of the RNAi phenotype, are highly attractive from a therapeutic standpoint. Agents having higher potency require lesser amounts of the molecule to achieve the desired physiological response, thus reducing the probability of side effects due to “off-target” interference. In addition to the potential therapeutic benefits associated with hyperfunctional siRNA, hf-siRNA are also desirable from an economic perspective. Hyperfunctional siRNA may cost less on a per-treatment basis, thus reducing overall expenditures to both the manufacturer and the consumer.
  • Identification of hyperfunctional siRNA involves multiple steps that are designed to examine an individual siRNA agent's concentration- and/or longevity-profiles.
  • a population of siRNA directed against a single gene are first analyzed using the previously described algorithm (Formula VIII). Individual siRNA are then introduced into a test cell line and assessed for the ability to degrade the target mRNA. It is important to note that when performing this step it is not necessary to test all of the siRNA. Instead, it is sufficient to test only those siRNA having the highest SMARTSCORESTM, or siRNA ranking (i.e., SMARTSCORESTM, or siRNA ranking > ⁇ 10). Subsequently, the gene silencing data is plotted against the SMARTSCORESTM, or siRNA rankings (see FIG.
  • siRNAs that (1) induce a high degree of gene silencing (i.e., they induce greater than 80% gene knockdown) and (2) have superior SMARTSCORESTM (i.e., a SMARTSCORETM, or siRNA ranking, of > ⁇ 10, suggesting a desirable average internal stability profile) are selected for further investigations designed to better understand the molecule's potency and longevity.
  • an siRNA is introduced into one (or more) cell types in increasingly diminishing concentrations (e.g., 3.0 ⁇ 0.3 nM).
  • siRNA that exhibit hyperfunctional potency i.e., those that induce 80% silencing or greater at, e.g., picomolar concentrations
  • siRNA having high (> ⁇ 10) SMARTSCORESTM, or siRNA rankings and greater than 80% silencing are examined.
  • siRNA are introduced into a test cell line and the levels of RNAi are measured over an extended period of time (e.g., 24-168 hrs).
  • siRNAs that exhibit strong RNA interference patterns i.e., >80% interference
  • periods of time greater than, e.g., 120 hours are thus identified.
  • siRNAs While the example(s) given above describe one means by which hyperfunctional siRNA can be isolated, neither the assays themselves nor the selection parameters used are rigid and can vary with each family of siRNA. Families of siRNA include siRNAs directed against a single gene, or directed against a related family of genes.
  • siRNA The highest quality siRNA achievable for any given gene may vary considerably.
  • rigorous studies such as those described above may enable the identification of an siRNA that, at picomolar concentrations, induces 99 + % silencing for a period of 10 days.
  • Yet identical studies of a second gene may yield an siRNA that at high nanomolar concentrations (e.g., 100 nM) induces only 75% silencing for a period of 2 days.
  • Both molecules represent the very optimum siRNA for their respective gene targets and therefore are designated “hyperfunctional.” Yet due to a variety of factors including but not limited to target concentration, siRNA stability, cell type, off-target interference, and others, equivalent levels of potency and longevity are not achievable.
  • the parameters described in the before mentioned assays can vary. While the initial screen selected siRNA that had SMARTSCORESTM above ⁇ 10 and a gene silencing capability of greater than 80%, selections that have stronger (or weaker) parameters can be implemented. Similarly, in the subsequent studies designed to identify molecules with high potency and longevity, the desired cutoff criteria (i.e., the lowest concentration that induces a desirable level of interference, or the longest period of time that interference can be observed) can vary. The experimentation subsequent to application of the rational criteria of this application is significantly reduced where one is trying to obtain a suitable hyperfunctional siRNA for, for example, therapeutic use. When, for example, the additional experimentation of the type described herein is applied by one skilled in the art with this disclosure in hand, a hyperfunctional siRNA is readily identified.
  • the siRNA may be introduced into a cell by any method that is now known or that comes to be known and that from reading this disclosure, persons skilled in the art would determine would be useful in connection with the present invention in enabling siRNA to cross the cellular membrane.
  • These methods include, but are not limited to, any manner of transfection, such as, for example, transfection employing DEAE-Dextran, calcium phosphate, cationic lipids/liposomes, micelles, manipulation of pressure, microinjection, electroporation, immunoporation, use of vectors such as viruses, plasmids, cosmids, bacteriophages, cell fusions, and coupling of the polynucleotides to specific conjugates or ligands such as antibodies, antigens, or receptors, passive introduction, adding moieties to the siRNA that facilitate its uptake, and the like.
  • siRNA nomenclature All siRNA duplexes are referred to by sense strand.
  • the first nucleotide of the 5′-end of the sense strand is position 1, which corresponds to position 19 of the antisense strand for a 19-mer.
  • silencing was determined by measuring specific transcript mRNA levels or enzymatic activity associated with specific transcript levels, 24 hours post-transfection, with siRNA concentrations held constant at 100 nM. For all experiments, unless otherwise specified, transfection efficiency was ensured to be over 95%, and no detectable cellular toxicity was observed.
  • the following system of nomenclature was used to compare and report siRNA-silencing functionality: “F” followed by the degree of minimal knockdown. For example, F50 signifies at least 50% knockdown, F80 means at least 80%, and so forth. For this study, all sub-F50 siRNAs were considered non-functional.
  • HEK293 cells or HEK293Lucs or any other cell type of interest are released from their solid support by trypsinization, diluted to 3.5 ⁇ 10 5 cells/ml, followed by the addition of 100 ⁇ L of cells/well. Plates are then incubated overnight at 37° C., 5% CO 2 . Transfection procedures can vary widely depending on the cell type and transfection reagents.
  • a transfection mixture consisting of 2 mL Opti-MEM I (Gibco-BRL), 80 ⁇ l Lipofectamine 2000 (Invitrogen), 15 ⁇ L SUPERNasin at 20 U/R1 (Ambion), and 1.5 ⁇ l of reporter gene plasmid at 1 ⁇ g/ ⁇ l is prepared in 5-ml polystyrene round bottom tubes.
  • One hundred ⁇ l of transfection reagent is then combined with 100 ⁇ l of siRNAs in polystyrene deep-well titer plates (Beckman) and incubated for 20 to 30 min at room temperature.
  • Opti-MEM Five hundred and fifty microliters of Opti-MEM is then added to each well to bring the final siRNA concentration to 100 nM. Plates are then sealed with parafilm and mixed. Media is removed from HEK293 cells and replaced with 95 ⁇ l of transfection mixture. Cells are incubated overnight at 37° C., 5% CO 2 .
  • Quantification of gene knockdown A variety of quantification procedures can be used to measure the level of silencing induced by siRNA or siRNA pools.
  • QuantiGene branched-DNA (bDNA) kits (Bayer) (Wang, et al, Regulation of insulin preRNA splicing by glucose . Proc. Natl. Acad. Sci. USA 1997, 94:4360.) are used according to manufacturer instructions.
  • bDNA QuantiGene branched-DNA kits
  • media is removed from HEK293 cells 24 hrs post-transfection, and 50 ⁇ l of Steady-GLO reagent (Promega) is added. After 5 minutes, plates are analyzed on a plate reader.
  • Anti-Firefly and anti-Cyclophilin siRNAs panels ( FIG. 5 a, b ) sorted according to using Formula VIII predicted values. All siRNAs scoring more than 0 (formula VIII) and more then 20 (formula IX) are fully functional. All ninety sequences for each gene (and DBI) appear below in Table III. TABLE III Cyclo 1 SEQ. ID 0032 GUUCCAAAAACAGUGGAUA Cyclo 2 SEQ. ID 0033 UCCAAAAACAGUGGAUAAU Cyclo 3 SEQ. ID 0034 CAAAAACAGUGGAUAAUUU Cyclo 4 SEQ. ID 0035 AAAACAGUGGAUAAUUUUG Cyclo 5 SEQ.
  • siRNAs for five genes, human DBI, firefly luciferase (fLuc), renilla luciferase (rLuc), human PLK, and human secreted alkaline phosphatase (SEAP).
  • fLuc firefly luciferase
  • rLuc renilla luciferase
  • SEAP human secreted alkaline phosphatase
  • clathrin-mediated endocytosis pathway Components of clathrin-mediated endocytosis pathway are key to modulating intracellular signaling and play important roles in disease. Chromosomal rearrangements that result in fusion transcripts between the Mixed-Lineage Leukemia gene (MLL) and CALM (clathrin assembly lymphoid myeloid leukemia gene) are believed to play a role in leukemogenesis. Similarly, disruptions in Rab7 and Rab9, as well as HIP1 (Huntingtin-interacting protein), genes that are believed to be involved in endocytosis, are potentially responsible for ailments resulting in lipid storage, and neuronal diseases, respectively. For these reasons, siRNA directed against clathrin and other genes involved in the clathrin-mediated endocytotic pathway are potentially important research and therapeutic tools.
  • siRNAs directed against genes involved in the clathrin-mediated endocytosis pathways were selected using Formula VIII.
  • the targeted genes were clathrin heavy chain (CHC, accession # NM 004859), clathrin light chain A (CLCa, NM — 001833), clathrin light chain B (CLCb, NM — 001834), CALM (U45976), ⁇ 2 subunit of AP-2 ( ⁇ 2, NM — 001282), Eps15 (NM — 001981), Eps15R(NM — 021235), dynamin II (DYNII, NM — 004945), Rab5a (BC001267), Rab5b (NM-002868), Rab5c (AF141304), and EEA.1 (XM — 018197).
  • siRNAs duplexes with the highest scores were selected and a BLAST search was conducted for each of them using the Human EST database. In order to minimize the potential for off-target silencing effects, only those sequences with more than three mismatches against un-related sequences were used. All duplexes were synthesized at Dharmacon, Inc. as 21-mers with 3′-UU overhangs using a modified method of 2′-ACE chemistry, Scaringe (2000) Advanced 5′-silyl-2′-orthoester approach to RNA oligonucleotide synthesis, Methods Enzymol. 317:3, and the antisense strand was chemically phosphorylated to insure maximized activity.
  • HeLa cells were grown in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum, antibiotics and glutamine.
  • siRNA duplexes were resuspended in 1 ⁇ siRNA Universal buffer (Dharmacon, Inc.) to 20CM prior to transfection.
  • HeLa cells in 12-well plates were transfected twice with 4 ⁇ l of 20 ⁇ M siRNA duplex in 3 ⁇ l Lipofectamine 2000 reagent (Invitrogen, Carlsbad, Calif., USA) at 24-hour intervals. For the transfections in which 2 or 3 siRNA duplexes were included, the amount of each duplex was decreased, so that the total amount was the same as in transfections with single siRNAs.
  • Cells were plated into normal culture medium 12 hours prior to experiments, and protein levels were measured 2 or 4 days after the first transfection.
  • Equal amounts of lysates were resolved by electrophoresis, blotted, and stained with the antibody specific to targeted protein, as well as antibodies specific to unrelated proteins, PP1 phosphatase and Tsg101 (not shown).
  • the cells were lysed in Triton X-100/glycerol solubilization buffer as described previously. Tebar, Bohlander, & Sorkin (1999) Clathrin Assembly Lymphoid Myeloid Leukemia (CALM) Protein: Localization in Endocytic-coated Pits, Interactions with Clathrin, and the Impact of Overexpression on Clathrin-mediated Traffic, Mol. Biol. Cell, 10:2687.
  • CALM Clathrin Assembly Lymphoid Myeloid Leukemia
  • the antibodies to assess the levels of each protein by Western blot were obtained from the following sources: monoclonal antibody to clathrin heavy chain (TD. 1) was obtained from American Type Culture Collection (Rockville, Md., USA); polyclonal antibody to dynamin II was obtained from Affinity Bioreagents, Inc. (Golden, Colo., USA); monoclonal antibodies to EEA. 1 and Rab5a were purchased from BD Transduction Laboratories (Los Angeles, Calif., USA); the monoclonal antibody to Tsg101 was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, Calif., USA); the monoclonal antibody to GFP was from ZYMED Laboratories Inc.
  • FIG. 11 demonstrates the in vivo functionality of 48 individual siRNAs, selected using Formula VIII (most of them will meet the criteria incorporated by Formula IX as well) targeting 12 genes.
  • Various cell lines were transfected with siRNA duplexes (Dup1-4) or pools of siRNA duplexes (Pool), and the cells were lysed 3 days after transfection with the exception of CALM (2 days) and ⁇ 2 (4 days).
  • CALM has two splice variants, 66 and 72 kD.
  • Eps15R a doublet of ⁇ 130 kD
  • Eps15R immunoprecipitates shown by arrows. The cells were lysed 3 days after transfection.
  • Equal amounts of lysates were resolved by electrophoresis and blotted with the antibody specific to a targeted protein (GFP antibody for YFP fusion proteins) and the antibody specific to unrelated proteins PP1 phosphatase or ⁇ -actinin, and TSG1101.
  • the amount of protein in each specific band was normalized to the amount of non-specific proteins in each lane of the gel. Nearly all of them appear to be functional, which establishes that Formula VIII and IX can be used to predict siRNAs' functionality in general in a genome wide manner.
  • YFP-Rab5b or YFP-Rab5c a DNA fragment encoding the full-length human Rab5b or Rab5c was obtained by PCR using Pfu polymerase (Stratagene) with a SacI restriction site introduced into the 5′ end and a KpnI site into the 3′ end and cloned into pEYFP-C1 vector (CLONTECH, Palo Alto, Calif., USA).
  • GFP-CALM and YFP-Rab5a were described previously (Tebar, Bohlander, & Sorkin (1999) Clathrin Assembly Lymphoid Myeloid Leukemia (CALM) Protein: Localization in Endocytic-coated Pits, Interactions with Clathrin, and the Impact of Overexpression on Clathrin-mediated Traffic, Mol. Biol. Cell 10:2687).
  • FIG. 12 illustrates four siRNAs targeting 10 different genes (Table V for sequence and accession number information) that were selected according to the Formula VIII and assayed as individuals and pools in HEK293 cells.
  • the level of siRNA induced silencing was measured using the B-DNA assay.
  • Bcl-2 is a ⁇ 25 kD, 205-239 amino acid, anti-apoptotic protein that contains considerable homology with other members of the BCL family including BCLX, MCL1, BAX, BAD, and BIK.
  • the protein exists in at least two forms (Bcl2a, which has a hydrophobic tail for membrane anchorage, and Bcl2b, which lacks the hydrophobic tail) and is predominantly localized to the mitochondrial membrane. While Bcl2 expression is widely distributed, particular interest has focused on the expression of this molecule in B and T cells.
  • Bcl2 expression is down-regulated in normal germinal center B cells yet in a high percentage of follicular lymphomas, Bcl2 expression has been observed to be elevated.
  • Bcl-2 translocation makes this gene an attractive target for RNAi.
  • Identification of siRNA directed against the bcl2 transcript (or Bcl2-IgH fusions) would further our understanding Bcl2 gene function and possibly provide a future therapeutic agent to battle diseases that result from altered expression or function of this gene.
  • Bcl-2 siRNAs having the top ten SMARTSCORESTM, or siRNA rankings were selected and tested in a functional assay to determine silencing efficiency.
  • each of the ten duplexes were synthesized using 2′-O-ACE chemistry and transfected at 100 nM concentrations into cells. Twenty-four hours later assays were performed on cell extracts to assess the degree of target silencing. Controls used in these experiments included mock transfected cells, and cells that were transfected with a non-specific siRNA duplex.
  • siRNA 1 GGGAGAUAGUGAUGAAGUA SEQ. ID NO. 302
  • siRNA 2 GAAGUACAUCCAUUAUAAG SEQ. ID NO. 303
  • siRNA 3 GUACGACAACCGGGAGAUA SEQ. ID NO. 304
  • siRNA 4 AGAUAGUGAUGAAGUACAU SEQ. ID NO. 305
  • siRNA 5 UGAAGACUCUGCUCAGUUU SEQ. ID NO.
  • siRNA 6 GCAUGCGGCCUCUGUUUGA SEQ. ID NO. 307 siRNA 7 UGCGGCCUCUGUUUGAUUU SEQ. ID NO. 308 siRNA 8 GAGAUAGUGAUGAAGUACA SEQ. ID NO. 309 siRNA 9 GGAGAUAGUGAUGAAGUAC SEQ. ID NO. 310 siRNA 10 GAAGACUCUGCUCAGUUUG SEQ. ID NO. 311
  • Bcl2 siRNA Sense Strand, 5′ ⁇ 3′
  • siRNAs listed in the sequence listing may potentially act as therapeutic agents.
  • a number of prophetic examples follow and should be understood in view of the siRNA that are identified in the sequence listing.
  • the appropriate message sequence for each gene is analyzed using one of the before mentioned formulas (preferably formula VIII) to identify potential siRNA targets. Subsequently these targets are BLAST'ed to eliminate homology with potential off-targets.
  • siRNA duplexes were synthesized using Dharmacon proprietary ACE® chemistry against one of the standard reporter genes: firefly luciferase. The duplexes were designed to start two base pairs apart and to cover approximately 180 base pairs of the luciferase gene (see sequences in Table III). Subsequently, the siRNA duplexes were co-transfected with a luciferase expression reporter plasmid into HEK293 cells using standard transfection protocols and luciferase activity was assayed at 24 and 48 hours.
  • FIG. 15 represents a typical screen of ninety siRNA duplexes (SEQ. ID NO. 0032-0120) positioned two base pairs apart.
  • SEQ. ID NO. 0032-0120 ninety siRNA duplexes positioned two base pairs apart.
  • the functionality of the siRNA duplex is determined more by a particular sequence of the oligonucleotide than by the relative oligonucleotide position within a gene or excessively sensitive part of the mRNA, which is important for traditional anti-sense technology.
  • FIGS. 16A and B When two continuous oligonucleotides were pooled together, a significant increase in gene silencing activity was observed (see FIGS. 16A and B). A gradual increase in efficacy and the frequency of pools functionality was observed when the number of siRNAs increased to 3 and 4 ( FIGS. 16A, 16B , 17 A, and 17 B). Further, the relative positioning of the oligonucleotides within a pool did not determine whether a particular pool was functional (see FIGS. 18A and 18B , in which 100% of pools of oligonucleotides distanced by 2, 10 and 20 base pairs were functional).
  • siRNA are positioned continuously head to toe (5′ end of one directly adjacent to the 3′ end of the others).
  • siRNA pools that were tested performed at least as well as the best oligonucleotide in the pool, under the experimental conditions whose results are depicted in FIG. 19 .
  • siRNA duplexes were pooled together in groups of five at a time, a significant functional cooperative action was observed (see FIG. 20 ).
  • pools of semi-active oligonucleotides were 5 to 25 times more functional than the most potent oligonucleotide in the pool. Therefore, pooling several siRNA duplexes together does not interfere with the functionality of the most potent siRNAs within a pool, and pooling provides an unexpected significant increase in overall functionality
  • siRNA sequences for the human cyclophilin B protein listed in Table III above lists the siRNA sequences for the human cyclophilin B protein. A particularly functional siRNA may be selected by applying these sequences to any of Formula I to VII above.
  • kits for silencing a gene Preferably, within the kit there would be at least one sequence that has a relatively high predicted functionality when any of Formulas I-VII is applied.
  • siRNA may be used as both research or diagnostic tools and therapeutic agents, either individually or in pools. Genes involved in signal transduction, the immune response, apoptosis, DNA repair, cell cycle control, and a variety of other physiological functions have clinical relevance and therapeutic agents that can modulate expression of these genes may alleviate some or all of the associated symptoms. In some instances, these genes can be described as a member of a family or class of genes and siRNA (randomly, conventionally, or rationally designed) can be directed against one or multiple members of the family to induce a desired result.
  • siRNA having heightened levels of potency can be identified by testing each of the before mentioned duplexes at increasingly limiting concentrations.
  • siRNA having increased levels of longevity can be identified by introducing each duplex into cells and testing functionality at 24, 48, 72, 96, 120, 144, 168, and 192 hours after transfection.
  • Agents that induce >95% silencing at sub-nanomolar concentrations and/or induce functional levels of silencing for >96 hours are considered hyperfunctional.
  • Two or more genes having similar, overlapping functions often leads to genetic redundancy. Mutations that knockout only one of, e.g., a pair of such genes (also referred to as homologs) results in little or no phenotype due to the fact that the remaining intact gene is capable of fulfilling the role of the disrupted counterpart. To fully understand the function of such genes in cellular physiology, it is often necessary to knockout or knockdown both homologs simultaneously. Unfortunately, concomitant knockdown of two or more genes is frequently difficult to achieve in higher organisms (e.g., mice) thus it is necessary to introduce new technologies dissect gene function. One such approach to knocking down multiple genes simultaneously is by using siRNA. For example, FIG.
  • siRNA directed against a number of genes involved in the clathrin-mediated endocytosis pathway resulted in significant levels of protein reduction (e.g., >80%).
  • internalization assays were performed using epidermal growth factor and transferrin. Specifically, mouse receptor-grade EGF (Collaborative Research Inc.) and iron-saturated human transferrin (Sigma) were iodinated as described previously (Jiang, X., Huang, F., Marusyk, A. & Sorkin, A. (2003) Mol Biol Cell 14, 858-70).
  • HeLa cells grown in 12-well dishes were incubated with 125 I-EGF (1 ng/ml) or 125 I-transferrin (1 ⁇ g/ml) in binding medium (DMEM, 0.1% bovine serum albumin) at 37° C., and the ratio of internalized and surface radioactivity was determined during 5-min time course to calculate specific internalization rate constant k c as described previously (Jiang, X et al.).
  • the measurements of the uptakes of radiolabeled transferrin and EGF were performed using short time-course assays to avoid influence of the recycling on the uptake kinetics, and using low ligand concentration to avoid saturation of the clathrin-dependent pathway (for EGF Lund, K. A., Opresko, L. K., Strarbuck, C., Walsh, B. J. & Wiley, H. S. (1990) J. Biol. Chem. 265, 15713-13723).
  • siRNA or siRNA pools directed against a collection of genes are simultaneously transfected into cells and cultured for twenty-four hours.
  • mRNA is harvested from treated (and untreated) cells and labeled with one of two fluorescent probes dyes (e.g., a red fluorescent probe for the treated cells, a green fluorescent probe for the control cells.).
  • Equivalent amounts of labeled RNA from each sample is then mixed together and hybridized to sequences that have been linked to a solid support (e.g., a slide, “DNA CHIP”). Following hybridization, the slides are washed and analyzed to assess changes in the levels of target genes induced by siRNA.
  • the ten rationally designed Bcl2 siRNA (identified in FIGS. 13, 14 ) were tested to identify hyperpotent reagents. To accomplish this, each of the ten Bcl-2 siRNA were individually transfected into cells at a 300 pM (0.3 nM) concentrations. Twenty-four hours later, transcript levels were assessed by B-DNA assays and compared with relevant controls. As shown in FIG. 25 , while the majority of Bcl-2 siRNA failed to induce functional levels of silencing at this concentration, siRNA 1 and 8 induced >80% silencing, and siRNA 6 exhibited greater than 90% silencing at this subnanomolar concentration.
  • RNAi RNAi reverse transcriptase
  • the selection of a cell line is usually determined by the desired application. The most important feature to RNAi is the level of expression of the gene of interest. It is highly recommended to use cell lines for which siRNA transfection conditions have been specified and validated.
  • siRNA re-suspension Add 20 ⁇ l siRNA universal buffer to each siRNA to generate a final concentration of 50 ⁇ M.
  • Transfection Create a Mixture 1 by combining the specified amounts of OPTI-MEM serum free media and transfection reagent in a sterile polystyrene tube.
  • Create a Mixture 2 by combining specified amounts of each siRNA with OPTI-MEM media in sterile 1 ml tubes.
  • Create a Mixture 3 by combining specified amounts of Mixture 1 and Mixture 2. Mix gently (do not vortex) and incubate at room temperature for 20 minutes.
  • Create a Mixture 4 by combining specified amounts of Mixture 3 to complete media. Add appropriate volume to each cell culture well. Incubate cells with transfection reagent mixture for 24-72 hours at 37° C. This incubation time is flexible. The ratio of silencing will remain consistent at any point in the time period.
  • Assay for gene silencing using an appropriate detection method such as RT-PCR, Western blot analysis, immunohistochemistry, phenotypic analysis, mass spectrometry, fluorescence, radioactive decay, or any other method that is now known or that comes to be known to persons skilled in the art and that from reading this disclosure would useful with the present invention.
  • the optimal window for observing a knockdown phenotype is related to the mRNA turnover of the gene of interest, although 24-72 hours is standard.
  • Final Volume reflects amount needed in each well for the desired cell culture format. When adjusting volumes for a Stock Mix, an additional 10% should be used to accommodate variability in pipetting, etc. Duplicate or triplicate assays should be carried out when possible.
  • siRNAs that target nucleotide sequences for deubiquitination enzymes with the NCBI accession numbers denoted below and having sequences generated in silico by the algorithms herein, are provided.
  • the siRNAs are rationally designed.
  • the siRNAs are functional or hyperfunctional.
  • the present invention provides an siRNA that targets a nucleotide sequence for a deubiquitination enzymes, wherein the siRNA is selected from the group consisting of SEQ. ID NOs. 438-3940.
  • an siRNA comprising a sense region and an antisense region, wherein said sense region and said antisense region are at least 90% complementary, said sense region and said antisense region together form a duplex region comprising 18-30 base pairs, and said sense region comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • an siRNA comprising a sense region and an antisense region, wherein said sense region and said antisense region are at least 90% complementary, said sense region and said antisense region together form a duplex region comprising 18-30 base pairs, and said sense region comprises a sequence that is identical to a contiguous stretch of at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • an siRNA comprising a sense region and an antisense region, wherein said sense region and said antisense region are at least 90% complementary, said sense region and said antisense region together form a duplex region comprising 19-30 base pairs, and said sense region comprises a sequence that is identical to a contiguous stretch of at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • a pool of at least two siRNAs comprising a first siRNA and a second siRNA
  • said first siRNA comprises a duplex region of length 18-30 base pairs that has a first sense region that is at least 90% similar to 18 bases of a first sequence selected from the group consisting of: SEQ. ID NOs 438-3940
  • said second siRNA comprises a duplex region of length 18-30 base pairs that has a second sense region that is at least 90% similar to 18 bases of a second sequence selected from the group consisting of: SEQ. ID NOs 438-3940 and wherein said first sense region and said second sense region are not identical.
  • a pool of at least two siRNAs comprising a first siRNA and a second siRNA
  • said first siRNA comprises a duplex region of length 18-30 base pairs that has a first sense region that is identical to at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940
  • the second siRNA comprises a second sense region that comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • a pool of at least two siRNAs comprising a first siRNA and a second siRNA
  • said first siRNA comprises a duplex region of length 19-30 base pairs and has a first sense region comprising a sequence that is at least 90% similar to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940
  • said duplex of said second siRNA is 19-30 base pairs and comprises a second sense region that comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • a pool of at least two siRNAs comprising a first siRNA and a second siRNA
  • said first siRNA comprises a duplex region of length 19-30 base pairs and has a first sense region comprising a sequence that is identical to at least 18 bases of a sequence selected the group consisting of: SEQ. ID NOs 438-3940
  • said duplex of said second siRNA is 19-30 base pairs and comprises a second sense region comprising a sequence that is identical to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • the antisense region is at least 90% complementary to a contiguous stretch of bases of one of the NCBI sequences identified in Example XVII; each of the recited NCBI sequences is incorporated by reference as if set forth fully herein. In some embodiments, the antisense region is 100% complementary to a contiguous stretch of bases of one of the NCBI sequences identified in Example XVII.
  • the antisense region is 20-30 bases in length

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Abstract

Efficient sequence specific gene silencing is possible through the use of siRNA technology. By selecting particular siRNAs by rational design, one can maximize the generation of an effective gene silencing reagent, as well as methods for silencing genes. Methods, compositions, and kits generated through rational design of siRNAs are disclosed including those directed to nucleotide sequences for deubiquitination enzymes.

Description

    CROSS-REFERENCE TO RELATED APPLICATIONS
  • This application is a continuation-in-part of U.S. Ser. No. 10/714,333, filed Nov. 14, 2003, which claims the benefit of U.S. Provisional Application No. 60/426,137, filed Nov. 14, 2002, and also claims the benefit of U.S. Provisional Application No. 60/502,050, filed Sep. 10, 2003; this application is also a continuation-in-part of U.S. Ser. No. 10/940,892, filed Sep. 14, 2004, which is a continuation of PCT Application No. PCT/US04/14885, international filing date May 12, 2004. The disclosures of the priority applications, including the sequence listings and tables submitted in electronic form in lieu of paper, are incorporated by reference into the instant specification.
    • SEQUENCE LISTING
  • The sequence listing for this application has been submitted in accordance with 37 CFR § 1.52(e) and 37 CFR § 1.821 on CD-ROM in lieu of paper on a disk containing the sequence listing file entitled “DHARMA—2100-US79_CRF.txt” created Sep. 28, 2007, 668 kb. Applicants hereby incorporate by reference the sequence listing provided on CD-ROM in lieu of paper into the instant specification.
    • FIELD OF INVENTION
  • The present invention relates to RNA interference (“RNAi”).
    • BACKGROUND OF THE INVENTION
  • Relatively recently, researchers observed that double stranded RNA (“dsRNA”) could be used to inhibit protein expression. This ability to silence a gene has broad potential for treating human diseases, and many researchers and commercial entities are currently investing considerable resources in developing therapies based on this technology.
  • Double stranded RNA induced gene silencing can occur on at least three different levels: (i) transcription inactivation, which refers to RNA guided DNA or histone methylation; (ii) siRNA induced mRNA degradation; and (iii) mRNA induced transcriptional attenuation.
  • It is generally considered that the major mechanism of RNA induced silencing (RNA interference, or RNAi) in mammalian cells is mRNA degradation. Initial attempts to use RNAi in mammalian cells focused on the use of long strands of dsRNA. However, these attempts to induce RNAi met with limited success, due in part to the induction of the interferon response, which results in a general, as opposed to a target-specific, inhibition of protein synthesis. Thus, long dsRNA is not a viable option for RNAi in mammalian systems.
  • More recently it has been shown that when short (18-30 bp) RNA duplexes are introduced into mammalian cells in culture, sequence-specific inhibition of target mRNA can be realized without inducing an interferon response. Certain of these short dsRNAs, referred to as small inhibitory RNAs (“siRNAs”), can act catalytically at sub-molar concentrations to cleave greater than 95% of the target mRNA in the cell. A description of the mechanisms for siRNA activity, as well as some of its applications are described in Provost et al (2002) Ribonuclease Activity and RNA Binding of Recombinant Human Dicer, EMBO J. 21(21): 5864-5874; Tabara et al. (2002) The dsRNA Binding Protein RDE-4 Interacts with RDE-1, DCR-1 and a DexH-box Helicase to Direct RNAi in C. elegans, Cell 109(7):861-71; Ketting et al. (2002) Dicer Functions in RNA Interference and in Synthesis of Small RNA Involved in Developmental Timing in C. elegans; Martinez et al., Single-Stranded Antisense siRNAs Guide Target RNA Cleavage in RNAi, Cell 110(5):563; Hutvagner & Zamore (2002) A microRNA in a multiple-turnover RNAi enzyme complex, Science 297:2056.
  • From a mechanistic perspective, introduction of long double stranded RNA into plants and invertebrate cells is broken down into siRNA by a Type III endonuclease known as Dicer. Sharp, RNA interference—2001, Genes Dev. 2001, 15:485. Dicer, a ribonuclease-III-like enzyme, processes the dsRNA into 19-23 base pair short interfering RNAs with characteristic two base 3′ overhangs. Bernstein, Caudy, Hammond, & Hannon (2001) Role for a bidentate ribonuclease in the initiation step of RNA interference, Nature 409:363. The siRNAs are then incorporated into an RNA-induced silencing complex (RISC) where one or more helicases unwind the siRNA duplex, enabling the complementary antisense strand to guide target recognition. Nykanen, Haley, & Zamore (2001) ATP requirements and small interfering RNA structure in the RNA interference pathway, Cell 107:309. Upon binding to the appropriate target mRNA, one or more endonucleases within the RISC cleaves the target to induce silencing. Elbashir, Lendeckel, & Tuschl (2001) RNA interference is mediated by 21- and 22-nucleotide RNAs, Genes Dev. 15:188, FIG. 1.
  • The interference effect can be long lasting and may be detectable after many cell divisions. Moreover, RNAi exhibits sequence specificity. Kisielow, M. et al. (2002) Isoform-specific knockdown and expression of adaptor protein ShcA using small interfering RNA, J. Biochem. 363:1-5. Thus, the RNAi machinery can specifically knock down one type of transcript, while not affecting closely related mRNA. These properties make siRNA a potentially valuable tool for inhibiting gene expression and studying gene function and drug target validation. Moreover, siRNAs are potentially useful as therapeutic agents against: (1) diseases that are caused by over-expression or misexpression of genes; and (2) diseases brought about by expression of genes that contain mutations.
  • Successful siRNA-dependent gene silencing depends on a number of factors. One of the most contentious issues in RNAi is the question of the necessity of siRNA design, i.e., considering the sequence of the siRNA used. Early work in C. elegans and plants circumvented the issue of design by introducing long dsRNA (see, for instance, Fire, A. et al. (1998) Nature 391:806-811). In this primitive organism, long dsRNA molecules are cleaved into siRNA by Dicer, thus generating a diverse population of duplexes that can potentially cover the entire transcript. While some fraction of these molecules are non-functional (i.e., induce little or no silencing) one or more have the potential to be highly functional, thereby silencing the gene of interest and alleviating the need for siRNA design. Unfortunately, due to the interferon response, this same approach is unavailable for mammalian systems. While this effect can be circumvented by bypassing the Dicer cleavage step and directly introducing siRNA, this tactic carries with it the risk that the chosen siRNA sequence may be non-functional or semi-functional.
  • A number of researches have expressed the view that siRNA design is not a crucial element of RNAi. On the other hand, others in the field have begun to explore the possibility that RNAi can be made more efficient by paying attention to the design of the siRNA. Unfortunately, none of the reported methods have provided a satisfactory scheme for reliably selecting siRNA with acceptable levels of functionality. Accordingly, there is a need to develop rational criteria by which to select siRNA with an acceptable level of functionality, and to identify siRNA that have this improved level of functionality, as well as to identify siRNAs that are hyperfunctional.
    • SUMMARY OF THE INVENTION
  • The present invention is directed to increasing the efficiency of RNAi, particularly in mammalian systems. Accordingly, the present invention provides kits, siRNAs and methods for increasing siRNA efficacy.
  • According to a first embodiment, the present invention provides a kit for gene silencing, wherein said kit is comprised of a pool of at least two siRNA duplexes, each of which is comprised of a sequence that is complementary to a portion of the sequence of one or more target messenger RNA, and each of which is selected using non-target specific criteria.
  • According to a second embodiment, the present invention provides a method for selecting an siRNA, said method comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; and determining the relative functionality of the at least two siRNAs.
  • According to a third embodiment, the present invention also provides a method for selecting an siRNA wherein said selection criteria are embodied in a formula comprising:
    (−14)*G13−13*A1−12*U7−11*U2−10*A11−10*U4−10*C3−10*C5−10*C6−9*A10−9*U9−9*C18−8*G10−7*U1−7*U16−7*C17−7*C19+7*U17+8*A2+8*A4+8*A5+8*C4+9*G8+10*A7+10*U18+11*A19+11*C9+15*G1+18*A3+19*U10−Tm−3*(GCtotal)−6*(GC15-19)−30*X; or  Formula VIII
    (−8)*A1+(−1)*A2+(12)*A3+(7)*A4+(18)*A5+(12)*A6+(19)*A7+(6)*A8+(−4)*A9+(−5)*A10+(−2)*A11+(−5)*A12+(17)*A13+(−3)*A14+(4)*A15+(2)*A16+(8)*A17+(11)*A18+(30)*A19+(−13)*U1+(−10)*U2+(2)*U3+(−2)*U4+(−5)*U5+(5)*U6+(−2)*U7+(−10)*U8+(−5)*U9+(15)*U10+(−1)*U11+(0)*U12+(10)*U13+(−9)*U14+(−13)*U15+(−10)*U16+(3)*U17+(9)*U18+(9)*U19+(7)*C1+(3)*C2+(−21)*C3+(5)*C4+(−9)*C5+(−20)*C6+(−18)*C7+(−5)*C8+(5)*C9+(1)*C10+(2)*C11+(−5)*C12+(−3)*C13+(−6)*C14+(−2)*C15+(−5)*C16+(−3)*C17+(−12)*C18+(−18)*C19+(14)*G1+(8)*G2+(7)*G3+(−10)*G4+(−4)*G5+(2)*G6+(1)*G7+(9)*G8+(5)*G9+(−11)*G10+(1)*G11+(9)*G12+(−24)*G13+(18)*G14+(11)*G15+(13)*G16+(−7)*G17+(−9)*G18+(−22)*G19+6*(number of A+U in position 15-19)-3*(number of G+C in whole siRNA),  Formula X
    wherein position numbering begins at the 5′-most position of a sense strand, and
    A1=1 if A is the base at position 1 of the sense strand, otherwise its value is 0;
    A2=1 if A is the base at position 2 of the sense strand, otherwise its value is 0;
    A3=1 if A is the base at position 3 of the sense strand, otherwise its value is 0;
    A4=1 if A is the base at position 4 of the sense strand, otherwise its value is 0;
    A5=1 if A is the base at position 5 of the sense strand, otherwise its value is 0;
    A6=1 if A is the base at position 6 of the sense strand, otherwise its value is 0;
    A7=1 if A is the base at position 7 of the sense strand, otherwise its value is 0;
    A10=1 if A is the base at position 10 of the sense strand, otherwise its value is 0;
    A11=1 if A is the base at position 11 of the sense strand, otherwise its value is 0;
    A13=1 if A is the base at position 13 of the sense strand, otherwise its value is 0;
    A19=1 if A is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
    C3=1 if C is the base at position 3 of the sense strand, otherwise its value is 0;
    C4=1 if C is the base at position 4 of the sense strand, otherwise its value is 0;
    C5=1 if C is the base at position 5 of the sense strand, otherwise its value is 0;
    C6=1 if C is the base at position 6 of the sense strand, otherwise its value is 0;
    C7=1 if C is the base at position 7 of the sense strand, otherwise its value is 0;
    C9=1 if C is the base at position 9 of the sense strand, otherwise its value is 0;
    C17=1 if C is the base at position 17 of the sense strand, otherwise its value is 0;
    C18=1 if C is the base at position 18 of the sense strand, otherwise its value is 0;
    C19=1 if C is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
    G1=1 if G is the base at position 1 on the sense strand, otherwise its value is 0;
    G2=1 if G is the base at position 2 of the sense strand, otherwise its value is 0;
    G8=1 if G is the base at position 8 on the sense strand, otherwise its value is 0;
    G10=1 if G is the base at position 10 on the sense strand, otherwise its value is 0;
    G13=1 if G is the base at position 13 on the sense strand, otherwise its value is 0;
    G19=1 if G is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
    U1=1 if U is the base at position 1 on the sense strand, otherwise its value is 0;
    U2=1 if U is the base at position 2 on the sense strand, otherwise its value is 0;
    U3=1 if U is the base at position 3 on the sense strand, otherwise its value is 0;
    U4=1 if U is the base at position 4 on the sense strand, otherwise its value is 0;
    U7=1 if U is the base at position 7 on the sense strand, otherwise its value is 0;
    U9=1 if U is the base at position 9 on the sense strand, otherwise its value is 0;
    U10=1 if U is the base at position 10 on the sense strand, otherwise its value is 0;
    U15=1 if U is the base at position 15 on the sense strand, otherwise its value is 0;
    U16=1 if U is the base at position 16 on the sense strand, otherwise its value is 0;
    U17=1 if U is the base at position 17 on the sense strand, otherwise its value is 0;
    U18=1 if U is the base at position 18 on the sense strand, otherwise its value is 0.
    GC5-19=the number of G and C bases within positions 15-19 of the sense strand, or within positions 15-18 if the sense strand is only 18 base pairs in length;
    GCtotal=the number of G and C bases in the sense strand;
    Tm=100 if the siRNA oligo has the internal repeat longer then 4 base pairs, otherwise its value is 0; and
    X=the number of times that the same nucleotide repeats four or more times in a row.
  • According to a fourth embodiment, the invention provides a method for developing an algorithm for selecting siRNA, said method comprising: (a) selecting a set of siRNA; (b) measuring gene silencing ability of each siRNA from said set; (c) determining relative functionality of each siRNA; (d) determining improved functionality by the presence or absence of at least one variable selected from the group consisting of the presence or absence of a particular nucleotide at a particular position, the total number of As and Us in positions 15-19, the number of times that the same nucleotide repeats within a given sequence, and the total number of Gs and Cs; and (e) developing an algorithm using the information of step (d).
  • According to a fifth embodiment, the present invention provides a kit, wherein said kit is comprised of at least two siRNAs, wherein said at least two siRNAs comprise a first optimized siRNA and a second optimized siRNA, wherein said first optimized siRNA and said second optimized siRNA are optimized according a formula comprising Formula X.
  • The present invention also provides a method for identifying a hyperfunctional siRNA, comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; determining the relative functionality of the at least two siRNAs and assigning each of the at least two siRNAs a functionality score; and selecting siRNAs from the at least two siRNAs that have a functionality score that reflects greater than 80 percent silencing at a concentration in the picomolar range, wherein said greater than 80 percent silencing endures for greater than 120 hours.
  • According to a sixth embodiment, the present invention provides a hyperfunctional siRNA that is capable of silencing Bcl2.
  • According to a seventh embodiment, the present invention provides a method for developing an siRNA algorithm for selecting functional and hyperfunctional siRNAs for a given sequence. The method comprises:
  • (a) selecting a set of siRNAs;
  • (b) measuring the gene silencing ability of each siRNA from said set;
  • (c) determining the relative functionality of each siRNA;
  • (d) determining the amount of improved functionality by the presence or absence of at least one variable selected from the group consisting of the total GC content, melting temperature of the siRNA, GC content at positions 15-19, the presence or absence of a particular nucleotide at a particular position, relative thermodynamic stability at particular positions in a duplex, and the number of times that the same nucleotide repeats within a given sequence; and
  • (e) developing an algorithm using the information of step (d).
  • According to this embodiment, preferably the set of siRNAs comprises at least 90 siRNAs from at least one gene, more preferably at least 180 siRNAs from at least two different genes, and most preferably at least 270 and 360 siRNAs from at least three and four different genes, respectively. Additionally, in step (d) the determination is made with preferably at least two, more preferably at least three, even more preferably at least four, and most preferably all of the variables. The resulting algorithm is not target sequence specific.
  • In another embodiment, the present invention provides rationally designed siRNAs identified using the formulas above.
  • In yet another embodiment, the present invention is directed to hyperfunctional siRNA.
  • The ability to use the above algorithms, which are not sequence or species specific, allows for the cost-effective selection of optimized siRNAs for specific target sequences. Accordingly, there will be both greater efficiency and reliability in the use of siRNA technologies.
  • In various embodiments, siRNAs that target nucleotide sequences for deubiquitination enzymes are provided. In various embodiments, the siRNAs are rationally designed. In various embodiments, the siRNAs are functional or hyperfunctional.
  • In various embodiments, an siRNA that targets nucleotide sequence for a deubiquitination enzyme is provided, wherein the siRNA is selected from the group consisting of various siRNA sequences targeting nucleotide sequences for deubiquitination enzymes that are disclosed herein. In various embodiments, the siRNA sequence is selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • In various embodiments, siRNA comprising a sense region and an antisense region are provided, said sense region and said antisense region together form a duplex region comprising 18-30 base pairs, and said sense region comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of siRNA sequences targeting nucleotide sequences for deubiquitination enzymes that are disclosed herein. In various embodiments, the siRNA sequence is selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • In various embodiments, an siRNA comprising a sense region and an antisense region is provided, said sense region and said antisense region together form a duplex region comprising 18-30 base pairs, and said sense region comprises a sequence that is identical to a contiguous stretch of at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940. In various embodiments, the duplex region is 19-30 base pairs, and the sense region comprises a sequence that is identical to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • In various embodiments, a pool of at least two siRNAs is provided, wherein said pool comprises a first siRNA and a second siRNA, said first siRNA comprising a duplex region of length 18-30 base pairs that has a first sense region that is at least 90% similar to 18 bases of a first sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, and said second siRNA comprises a duplex region of length 18-30 base pairs that has a second sense region that is at least 90% similar to 18 bases of a second sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, wherein said first sense region and said second sense region are not identical.
  • In various embodiments, the first sense region comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, and said second sense region comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940. In various embodiments, the duplex of said first siRNA is 19-30 base pairs, and said first sense region comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, and said duplex of said second siRNA is 19-30 base pairs and comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • In various embodiments, the duplex of said first siRNA is 19-30 base pairs and said first sense region comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940, and said duplex of said second siRNA is 19-30 base pairs and said second region comprises a sequence that is identical to a sequence selected from the group consisting of SEQ ID NO. 438 to SEQ ID NO. 3940.
  • For a better understanding of the present invention together with other and further advantages and embodiments, reference is made to the following description taken in conjunction with the examples, the scope of which is set forth in the appended claims.
    • BRIEF DESCRIPTION OF THE FIGURES
  • FIG. 1 shows a model for siRNA-RISC interactions. RISC has the ability to interact with either end of the siRNA or miRNA molecule. Following binding, the duplex is unwound, and the relevant target is identified, cleaved, and released.
  • FIG. 2 is a representation of the functionality of two hundred and seventy siRNA duplexes that were generated to target human cyclophilin, human diazepam-binding inhibitor (DB), and firefly luciferase.
  • FIG. 3 a is a representation of the silencing effect of 30 siRNAs in three different cells lines, HEK293, DU145, and Hela. FIG. 3 b shows the frequency of different functional groups (>95% silencing (black), >80% silencing (gray), >50% silencing (dark gray), and <50% silencing (white)) based on GC content. In cases where a given bar is absent from a particular GC percentage, no siRNA were identified for that particular group. FIG. 3 c shows the frequency of different functional groups based on melting temperature (Tm).
  • FIG. 4 is a representation of a statistical analysis that revealed correlations between silencing and five sequence-related properties of siRNA: (A) an A at position 19 of the sense strand, (B) an A at position 3 of the sense strand, (C) a U at position 10 of the sense strand, (D) a base other than G at position 13 of the sense strand, and (E) a base other than C at position 19 of the sense strand. All variables were correlated with siRNA silencing of firefly luciferase and human cyclophilin. siRNAs satisfying the criterion are grouped on the left (Selected) while those that do not, are grouped on the right (Eliminated). Y-axis is “% Silencing of Control.” Each position on the X-axis represents a unique siRNA.
  • FIGS. 5A and 5B are representations of firefly luciferase and cyclophilin siRNA panels sorted according to functionality and predicted values using Formula VIII. The siRNA found within the circle represent those that have Formula VIII values (SMARTSCORES™, or siRNA rank) above zero. siRNA outside the indicated area have calculated Formula VIII values that are below zero. Y-axis is “Expression (% Control).” Each position on the X-axis represents a unique siRNA.
  • FIG. 6A is a representation of the average internal stability profile (AISP) derived from 270 siRNAs taken from three separate genes (cyclophilin B, DBI and firefly luciferase). Graphs represent AISP values of highly functional, functional, and non-functional siRNA. FIG. 6B is a comparison between the AISP of naturally derived GFP siRNA (filled squares) and the AISP of siRNA from cyclophilin B, DBI, and luciferase having >90% silencing properties (no fill) for the antisense strand. “DG” is the symbol for ΔG, free energy.
  • FIG. 7 is a histogram showing the differences in duplex functionality upon introduction of base pair mismatches. The X-axis shows the mismatch introduced in the siRNA and the position it is introduced (e.g., 8C>A reveals that position 8 (which normally has a C) has been changed to an A). The Y-axis is “% Silencing (Normalized to Control).” The samples on the X-axis represent siRNAs at 100 nM and are, reading from left to right: 1A to C, 1A to G, 1A to U; 2A to C, 2A to G, 2A to U; 3A to C, 3A to G, 3A to U; 4G to A, 4G to C; 4G to U; 5U to A, 5U to C, 5U to G; 6U to A, 6U to C, 6U to G; 7G to A, 7G to C, 7G to U; 8C to A, 8C to G, 8C to U; 9G to A, 9G to C, 9G to U; 10C to A, 10C to G, 10C to U; 11G to A, 11G to C, 11G to U; 12G to A, 12G to C, 12G to U; 13A to C, 13A to G, 13A to U; 14G to A, 14G to C, 14G to U; 15G to A, 15G to C, 15G to U; 16A to C, 16A to G, 16A to U; 17G to A, 17G to C, 17G to U; 18U to A, 18U to C, 18U to G; 19U to A, 19U to C, 19U to G; 20 wt; Control.
  • FIG. 8 is histogram that shows the effects of 5′sense and antisense strand modification with 2′-O-methylation on functionality.
  • FIG. 9 shows a graph of SMARTSCORES™, or siRNA rank, versus RNAi silencing values for more than 360 siRNA directed against 30 different genes. SiRNA to the right of the vertical bar represent those siRNA that have desirable SMARTSCORES™, or siRNA rank.
  • FIGS. 10A-E compare the RNAi of five different genes (SEAP, DBI, PLK, Firefly Luciferase, and Renilla Luciferase) by varying numbers of randomly selected siRNA and four rationally designed (SMART-selected) siRNA chosen using the algorithm described in Formula VIII. In addition, RNAi induced by a pool of the four SMART-selected siRNA is reported at two different concentrations (100 and 400 nM). 10F is a comparison between a pool of randomly selected EGFR siRNA (Pool 1) and a pool of SMART-selected EGFR siRNA (Pool 2). Pool 1, S1-S4 and Pool 2 S1-S4 represent the individual members that made up each respective pool. Note that numbers for random siRNAs represent the position of the 5′ end of the sense strand of the duplex. The Y-axis represents the % expression of the control(s). The X-axis is the percent expression of the control.
  • FIG. 11 shows the Western blot results from cells treated with siRNA directed against twelve different genes involved in the clathrin-dependent endocytosis pathway (CHC, DynII, CALM, CLCa, CLCb, Eps15, Eps15R, Rab5a, Rab5b, Rab5c, P2 subunit of AP-2 and EEA.1). siRNA were selected using Formula VIII. “Pool” represents a mixture of duplexes 1-4. Total concentration of each siRNA in the pool is 25 nM. Total concentration=4×25=100 nM.
  • FIG. 12 is a representation of the gene silencing capabilities of rationally-selected siRNA directed against ten different genes (human and mouse cyclophilin, C-myc, human lamin A/C, QB (ubiquinol-cytochrome c reductase core protein I), MEK1 and MEK2, ATE1 (arginyl-tRNA protein transferase), GAPDH, and Eg5). The Y-axis is the percent expression of the control. Numbers 1, 2, 3 and 4 represent individual rationally selected siRNA. “Pool” represents a mixture of the four individual siRNA.
  • FIG. 13 is the sequence of the top ten Bcl2 siRNAs as determined by Formula VIII. Sequences are listed 5′ to 3′.
  • FIG. 14 is the knockdown by the top ten Bcl2 siRNAs at 100 nM concentrations. The Y-axis represents the amount of expression relative to the non-specific (ns) and transfection mixture control.
  • FIG. 15 represents a functional walk where siRNA beginning on every other base pair of a region of the luciferase gene are tested for the ability to silence the luciferase gene. The Y-axis represents the percent expression relative to a control. The X-axis represents the position of each individual siRNA. Reading from left to right across the X-axis, the position designations are 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIGS. 16A and 16B are histograms demonstrating the inhibition of target gene expression by pools of 2 (16A) and 3 (16B) siRNA duplexes taken from the walk described in FIG. 15. The Y-axis in each represents the percent expression relative to control. The X-axis in each represents the position of the first siRNA in paired pools, or trios of siRNAs. For instance, the first paired pool contains siRNAs 1 and 3. The second paired pool contains siRNAs 3 and 5. Pool 3 (of paired pools) contains siRNAs 5 and 7, and so on. For each of 16A and 16B, the X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIGS. 17A and 17B are histograms demonstrating the inhibition of target gene expression by pools of 4 (17A) and 5 (17B) siRNA duplexes. The Y-axis in each represents the percent expression relative to control. The X-axis in each represents the position of the first siRNA in each pool. For each of 17A and 17B, the X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIGS. 18A and 18B are histograms demonstrating the inhibition of target gene expression by siRNAs that are ten (18A) and twenty (18B) base pairs base pairs apart. The Y-axis represents the percent expression relative to a control. The X-axis represents the position of the first siRNA in each pool. For each of 18A and 18B, the X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIG. 19 shows that pools of siRNAs (dark gray bar) work as well (or better) than the best siRNA in the pool (light gray bar). The Y-axis represents the percent expression relative to a control. The X-axis represents the position of the first siRNA in each pool. The X-axis from left to right reads 1, 3, 5, 7, 9, 11, 13, 15, 17, 19, 21, 23, 25, 27, 29, 31, 33, 35, 37, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 73, 75, 77, 79, 81, 83, 85, 87, 89, and Plasmid.
  • FIG. 20 shows that the combination of several semifunctional siRNAs (dark gray) result in a significant improvement of gene expression inhibition over individual (semi-functional; light gray) siRNA. The Y-axis represents the percent expression relative to a control.
  • FIGS. 21A, 21B and 21C show both pools (Library, Lib) and individual siRNAs in inhibition of gene expression of Beta-Galactosidase, Renilla Luciferase and SEAP (alkaline phosphatase). Numbers on the X-axis indicate the position of the 5′-most nucleotide of the sense strand of the duplex. The Y-axis represents the percent expression of each gene relative to a control. Libraries contain 19 nucleotide long siRNAs (not including overhangs) that begin at the following nucleotides: SEAP: Lib 1: 206, 766, 812, 923, Lib 2: 1117, 1280, 1300, 1487, Lib 3: 206, 766, 812, 923, 1117, 1280, 1300, 1487, Lib 4: 206, 812, 1117, 1300, Lib 5: 766, 923, 1280, 1487, Lib 6: 206, 1487; Bgal: Lib 1: 979, 1339, 2029, 2590, Lib 2: 1087, 1783, 2399, 3257, Lib 3: 979, 1783, 2590, 3257, Lib 4: 979, 1087, 1339, 1783, 2029, 2399, 2590, 3257, Lib 5: 979, 1087, 1339, 1783, Lib 6: 2029, 2399, 2590, 3257; Renilla: Lib 1: 174, 300, 432, 568, Lib 2: 592, 633, 729, 867, Lib 3: 174, 300, 432, 568, 592, 633, 729, 867, Lib 4: 174, 432, 592, 729, Lib 5: 300, 568, 633, 867, Lib 6: 592, 568.
  • FIG. 22 shows the results of an EGFR and TfnR internalization assay when single gene knockdowns are performed. The Y-axis represents percent internalization relative to control.
  • FIG. 23 shows the results of an EGFR and TfnR internalization assay when multiple genes are knocked down (e.g., Rab5a, b, c). The Y-axis represents the percent internalization relative to control.
  • FIG. 24 shows the simultaneous knockdown of four different genes. siRNAs directed against G6PD, GAPDH, PLK, and UQC were simultaneously introduced into cells. Twenty-four hours later, cultures were harvested and assayed for mRNA target levels for each of the four genes. A comparison is made between cells transfected with individual siRNAs vs. a pool of siRNAs directed against all four genes.
  • FIG. 25 shows the functionality of ten siRNAs at 0.3 nM concentrations.
    • DETAILED DESCRIPTION Definitions
  • Unless stated otherwise, the following terms and phrases have the meanings provided below:
  • Complementary
  • The term “complementary” refers to the ability of polynucleotides to form base pairs with one another. Base pairs are typically formed by hydrogen bonds between nucleotide units in antiparallel polynucleotide strands. Complementary polynucleotide strands can base pair in the Watson-Crick manner (e.g., A to T, A to U, C to G), or in any other manner that allows for the formation of duplexes. As persons skilled in the art are aware, when using RNA as opposed to DNA, uracil rather than thymine is the base that is considered to be complementary to adenosine. However, when a U is denoted in the context of the present invention, the ability to substitute a T is implied, unless otherwise stated.
  • Perfect complementarity or 100% complementarity refers to the situation in which each nucleotide unit of one polynucleotide strand can hydrogen bond with a nucleotide unit of a second polynucleotide strand. Less than perfect complementarity refers to the situation in which some, but not all, nucleotide units of two strands can hydrogen bond with each other. For example, for two 20-mers, if only two base pairs on each strand can hydrogen bond with each other, the polynucleotide strands exhibit 10% complementarity. In the same example, if 18 base pairs on each strand can hydrogen bond with each other, the polynucleotide strands exhibit 90% complementarity.
  • Deoxynucleotide
  • The term “deoxynucleotide” refers to a nucleotide or polynucleotide lacking a hydroxyl group (OH group) at the 2′ and/or 3′ position of a sugar moiety. Instead, it has a hydrogen bonded to the 2′ and/or 3′ carbon. Within an RNA molecule that comprises one or more deoxynucleotides, “deoxynucleotide” refers to the lack of an OH group at the 2′ position of the sugar moiety, having instead a hydrogen bonded directly to the 2′ carbon.
  • Deoxyribonucleotide
  • The terms “deoxyribonucleotide” and “DNA” refer to a nucleotide or polynucleotide comprising at least one sugar moiety that has an H, rather than an OH, at its 2′ and/or 3′position.
  • Duplex Region
  • The phrase “duplex region” refers to the region in two complementary or substantially complementary polynucleotides that form base pairs with one another, either by Watson-Crick base pairing or any other manner that allows for a stabilized duplex between polynucleotide strands that are complementary or substantially complementary. For example, a polynucleotide strand having 21 nucleotide units can base pair with another polynucleotide of 21 nucleotide units, yet only 19 bases on each strand are complementary or substantially complementary, such that the “duplex region” has 19 base pairs. The remaining bases may, for example, exist as 5′ and 3′ overhangs. Further, within the duplex region, 100% complementarity is not required; substantial complementarity is allowable within a duplex region. Substantial complementarity refers to 79% or greater complementarity. For example, a mismatch in a duplex region consisting of 19 base pairs results in 94.7% complementarity, rendering the duplex region substantially complementary.
  • Filters
  • The term “filter” refers to one or more procedures that are performed on sequences that are identified by the algorithm. In some instances, filtering includes in silico procedures where sequences identified by the algorithm can be screened to identify duplexes carrying desirable or undesirable motifs. Sequences carrying such motifs can be selected for, or selected against, to obtain a final set with the preferred properties. In other instances, filtering includes wet lab experiments. For instance, sequences identified by one or more versions of the algorithm can be screened using any one of a number of procedures to identify duplexes that have hyperfunctional traits (e.g., they exhibit a high degree of silencing at subnanomolar concentrations and/or exhibit high degrees of silencing longevity).
  • Gene Silencing
  • The phrase “gene silencing” refers to a process by which the expression of a specific gene product is lessened or attenuated. Gene silencing can take place by a variety of pathways. Unless specified otherwise, as used herein, gene silencing refers to decreases in gene product expression that results from RNA interference (RNAi), a defined, though partially characterized pathway whereby small inhibitory RNA (siRNA) act in concert with host proteins (e.g., the RNA induced silencing complex, RISC) to degrade messenger RNA (mRNA) in a sequence-dependent fashion. The level of gene silencing can be measured by a variety of means, including, but not limited to, measurement of transcript levels by Northern Blot Analysis, B-DNA techniques, transcription-sensitive reporter constructs, expression profiling (e.g., DNA chips), and related technologies. Alternatively, the level of silencing can be measured by assessing the level of the protein encoded by a specific gene. This can be accomplished by performing a number of studies including Western Analysis, measuring the levels of expression of a reporter protein that has e.g., fluorescent properties (e.g., GFP) or enzymatic activity (e.g., alkaline phosphatases), or several other procedures.
  • miRNA
  • The term “miRNA” refers to microRNA.
  • Nucleotide
  • The term “nucleotide” refers to a ribonucleotide or a deoxyribonucleotide or modified form thereof, as well as an analog thereof. Nucleotides include species that comprise purines, e.g., adenine, hypoxanthine, guanine, and their derivatives and analogs, as well as pyrimidines, e.g., cytosine, uracil, thymine, and their derivatives and analogs.
  • Nucleotide analogs include nucleotides having modifications in the chemical structure of the base, sugar and/or phosphate, including, but not limited to, 5-position pyrimidine modifications, 8-position purine modifications, modifications at cytosine exocyclic amines, and substitution of 5-bromo-uracil; and 2′-position sugar modifications, including but not limited to, sugar-modified ribonucleotides in which the 2′-OH is replaced by a group such as an H, OR, R, halo, SH, SR, NH2, NHR, NR2, or CN, wherein R is an alkyl moiety. Nucleotide analogs are also meant to include nucleotides with bases such as inosine, queuosine, xanthine, sugars such as 2′-methyl ribose, non-natural phosphodiester linkages such as methylphosphonates, phosphorothioates and peptides.
  • Modified bases refer to nucleotide bases such as, for example, adenine, guanine, cytosine, thymine, uracil, xanthine, inosine, and queuosine that have been modified by the replacement or addition of one or more atoms or groups. Some examples of types of modifications that can comprise nucleotides that are modified with respect to the base moieties include but are not limited to, alkylated, halogenated, thiolated, aminated, amidated, or acetylated bases, individually or in combination. More specific examples include, for example, 5-propynyluridine, 5-propynylcytidine, 6-methyladenine, 6-methylguanine, N,N,-dimethyladenine, 2-propyladenine, 2-propylguanine, 2-aminoadenine, 1-methylinosine, 3-methyluridine, 5-methylcytidine, 5-methyluridine and other nucleotides having a modification at the 5 position, 5-(2-amino)propyl uridine, 5-halocytidine, 5-halouridine, 4-acetylcytidine, 1-methyladenosine, 2-methyladenosine, 3-methylcytidine, 6-methyluridine, 2-methylguanosine, 7-methylguanosine, 2,2-dimethylguanosine, 5-methylaminoethyluridine, 5-methyloxyuridine, deazanucleotides such as 7-deaza-adenosine, 6-azouridine, 6-azocytidine, 6-azothymidine, 5-methyl-2-thiouridine, other thio bases such as 2-thiouridine and 4-thiouridine and 2-thiocytidine, dihydrouridine, pseudouridine, queuosine, archaeosine, naphthyl and substituted naphthyl groups, any O- and N-alkylated purines and pyrimidines such as N6-methyladenosine, 5-methylcarbonylmethyluridine, uridine 5-oxyacetic acid, pyridine-4-one, pyridine-2-one, phenyl and modified phenyl groups such as aminophenol or 2,4,6-trimethoxy benzene, modified cytosines that act as G-clamp nucleotides, 8-substituted adenines and guanines, 5-substituted uracils and thymines, azapyrimidines, carboxyhydroxyalkyl nucleotides, carboxyalkylaminoalkyl nucleotides, and alkylcarbonylalkylated nucleotides. Modified nucleotides also include those nucleotides that are modified with respect to the sugar moiety, as well as nucleotides having sugars or analogs thereof that are not ribosyl. For example, the sugar moieties may be, or be based on, mannoses, arabinoses, glucopyranoses, galactopyranoses, 4′-thioribose, and other sugars, heterocycles, or carbocycles.
  • The term nucleotide is also meant to include what are known in the art as universal bases. By way of example, universal bases include but are not limited to 3-nitropyrrole, 5-nitroindole, or nebularine. The term “nucleotide” is also meant to include the N3′ to P5′ phosphoramidate, resulting from the substitution of a ribosyl 3′ oxygen with an amine group.
  • Further, the term nucleotide also includes those species that have a detectable label, such as for example a radioactive or fluorescent moiety, or mass label attached to the nucleotide.
  • Off-Target Silencing and Off-Target Interference
  • The phrases “off-target silencing” and “off-target interference” are defined as degradation of mRNA other than the intended target mRNA due to overlapping and/or partial homology with secondary mRNA messages.
  • Polynucleotide
  • The term “polynucleotide” refers to polymers of nucleotides, and includes but is not limited to DNA, RNA, DNA/RNA hybrids including polynucleotide chains of regularly and/or irregularly alternating deoxyribosyl moieties and ribosyl moieties (i.e., wherein alternate nucleotide units have an —OH, then and —H, then an —OH, then an —H, and so on at the 2′ position of a sugar moiety), and modifications of these kinds of polynucleotides, wherein the attachment of various entities or moieties to the nucleotide units at any position are included.
  • Polyribonucleotide
  • The term “polyribonucleotide” refers to a polynucleotide comprising two or more modified or unmodified ribonucleotides and/or their analogs. The term “polyribonucleotide” is used interchangeably with the term “oligoribonucleotide.”
  • Ribonucleotide and Ribonucleic Acid
  • The term “ribonucleotide” and the phrase “ribonucleic acid” (RNA), refer to a modified or unmodified nucleotide or polynucleotide comprising at least one ribonucleotide unit. A ribonucleotide unit comprises an hydroxyl group attached to the 2′ position of a ribosyl moiety that has a nitrogenous base attached in N-glycosidic linkage at the 1′ position of a ribosyl moiety, and a moiety that either allows for linkage to another nucleotide or precludes linkage.
  • siRNA
  • The term “siRNA” refers to small inhibitory RNA duplexes that induce the RNA interference (RNAi) pathway. These molecules can vary in length (generally 18-30 base pairs) and contain varying degrees of complementarity to their target mRNA in the antisense strand. Some, but not all, siRNA have unpaired overhanging bases on the 5′ or 3′ end of the sense strand and/or the antisense strand. The term “siRNA” includes duplexes of two separate strands, as well as single strands that can form hairpin structures comprising a duplex region.
  • siRNA may be divided into five (5) groups (non-functional, semi-functional, functional, highly functional, and hyper-functional) based on the level or degree of silencing that they induce in cultured cell lines. As used herein, these definitions are based on a set of conditions where the siRNA is transfected into said cell line at a concentration of 100 nM and the level of silencing is tested at a time of roughly 24 hours after transfection, and not exceeding 72 hours after transfection. In this context, “non-functional siRNA” are defined as those siRNA that induce less than 50% (<50%) target silencing. “Semi-functional siRNA” induce 50-79% target silencing. “Functional siRNA” are molecules that induce 80-95% gene silencing. “Highly-functional siRNA” are molecules that induce greater than 95% gene silencing. “Hyperfunctional siRNA” are a special class of molecules. For purposes of this document, hyperfunctional siRNA are defined as those molecules that: (1) induce greater than 95% silencing of a specific target when they are transfected at subnanomolar concentrations (i.e., less than one nanomolar); and/or (2) induce functional (or better) levels of silencing for greater than 96 hours. These relative functionalities (though not intended to be absolutes) may be used to compare siRNAs to a particular target for applications such as functional genomics, target identification and therapeutics.
  • SMARTSCORE™, or siRNA rank
  • The term “SMARTSCORE™”, or “siRNA rank” refers to a number determined by applying any of the formulas to a given siRNA sequence. The term “SMART-selected” or “rationally selected” or “rational selection” refers to siRNA that have been selected on the basis of their SMARTSCORES™, or siRNA ranking.
  • Substantially Similar
  • The phrase “substantially similar” refers to a similarity of at least 90% with respect to the identity of the bases of the sequence.
  • Target
  • The term “target” is used in a variety of different forms throughout this document and is defined by the context in which it is used. “Target mRNA” refers to a messenger RNA to which a given siRNA can be directed against. “Target sequence” and “target site” refer to a sequence within the mRNA to which the sense strand of an siRNA shows varying degrees of homology and the antisense strand exhibits varying degrees of complementarity. The phrase “siRNA target” can refer to the gene, mRNA, or protein against which an siRNA is directed. Similarly, “target silencing” can refer to the state of a gene, or the corresponding mRNA or protein.
  • Transfection
  • The term “transfection” refers to a process by which agents are introduced into a cell. The list of agents that can be transfected is large and includes, but is not limited to, siRNA, sense and/or anti-sense sequences, DNA encoding one or more genes and organized into an expression plasmid, proteins, protein fragments, and more. There are multiple methods for transfecting agents into a cell including, but not limited to, electroporation, calcium phosphate-based transfections, DEAE-dextran-based transfections, lipid-based transfections, molecular conjugate-based transfections (e.g., polylysine-DNA conjugates), microinjection and others.
  • The present invention is directed to improving the efficiency of gene silencing by siRNA. Through the inclusion of multiple siRNA sequences that are targeted to a particular gene and/or selecting an siRNA sequence based on certain defined criteria, improved efficiency may be achieved.
  • The present invention will now be described in connection with preferred embodiments. These embodiments are presented in order to aid in an understanding of the present invention and are not intended, and should not be construed, to limit the invention in any way. All alternatives, modifications and equivalents that may become apparent to those of ordinary skill upon reading this disclosure are included within the spirit and scope of the present invention.
  • Furthermore, this disclosure is not a primer on RNA interference. Basic concepts known to persons skilled in the art have not been set forth in detail.
  • The present invention is directed to increasing the efficiency of RNAi, particularly in mammalian systems. Accordingly, the present invention provides kits, siRNAs and methods for increasing siRNA efficacy.
  • According to a first embodiment, the present invention provides a kit for gene silencing, wherein said kit is comprised of a pool of at least two siRNA duplexes, each of which is comprised of a sequence that is complementary to a portion of the sequence of one or more target messenger RNA, and each of which is selected using non-target specific criteria. Each of the at least two siRNA duplexes of the kit complementary to a portion of the sequence of one or more target mRNAs is preferably selected using Formula X.
  • According to a second embodiment, the present invention provides a method for selecting an siRNA, said method comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; and determining the relative functionality of the at least two siRNAs.
  • In one embodiment, the present invention also provides a method wherein said selection criteria are embodied in a formula comprising:
    (−14)*G13−13*A1−12*U7−11*U2−10*A11−10*U4−10*C3−10*C5−1*C6−9*A10−9*U9−9*C18−8*G10−7*U1−7*U16−7*C17−7*C19+7*U17+8*A2+8*A4+8*A5+8*C4+9*G8+10*A7+10*U18+11*A19+11*C9+15*G1+18*A3+19*U10−Tm−3*(GCtotal)−6*(GC15-19)−30*X; or  Formula VIII
    (−8)*A1+(−1)*A2+(12)*A3+(7)*A4+(18)*A5+(12)*A6+(19)*A7+(6)*A8+(−4)*A9+(−5)*A10+(−2)*A11+(−5)*A12+(17)*A13+(−3)*A14+(4)*A15+(2)*A16+(8)*A17+(11)*A18+(30)*A19+(−13)*U1+(−10)*U2+(2)*U3+(−2)*U4+(−5)*U5+(5)*U6+(−2)*U7+(−10)*U8+(−5)*U9+(15)*U10+(−1)*U11+(0)*U12+(10)*U13+(−9)*U14+(−13)*U15+(−10)*U16+(3)*U17+(9)*U18+(9)*U19+(7)*C1+(3)*C2+(−21)*C3+(5)*C4+(−9)*C5+(−20)*C6+(−18)*C7+(−5)*C8+(5)*C9+(1)*C10+(2)*C11+(−5)*C12+(−3)*C13+(−6)*C14+(−2)*C15+(−5)*C16+(−3)*C17+(−12)*C18+(−18)*C19+(14)*G1+(8)*G2+(7)*G3+(−10)*G4+(−4)*G5+(2)*G6+(1)*G7+(9)*G8+(5)*G9+(−11)*G10+(1)*G11+(9)*G12+(−24)*G13+(18)*G14+(11)*G15+(13)*G16+(−7)*G17+(−9)*G18+(−22)*G19+6*(number of A+U in position 15-19)−3*(number of G+C in whole siRNA),  Formula X
  • wherein position numbering begins at the 5′-most position of a sense strand, and
  • A1=1 if A is the base at position 1 of the sense strand, otherwise its value is 0;
  • A2=1 if A is the base at position 2 of the sense strand, otherwise its value is 0;
  • A3=1 if A is the base at position 3 of the sense strand, otherwise its value is 0;
  • A4=1 if A is the base at position 4 of the sense strand, otherwise its value is 0;
  • A5=1 if A is the base at position 5 of the sense strand, otherwise its value is 0;
  • A6=1 if A is the base at position 6 of the sense strand, otherwise its value is 0;
  • A7=1 if A is the base at position 7 of the sense strand, otherwise its value is 0;
  • A10=1 if A is the base at position 10 of the sense strand, otherwise its value is 0;
  • A11=1 if A is the base at position 11 of the sense strand, otherwise its value is 0;
  • A13=1 if A is the base at position 13 of the sense strand, otherwise its value is 0;
  • A19=1 if A is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • C3=1 if C is the base at position 3 of the sense strand, otherwise its value is 0;
  • C4=1 if C is the base at position 4 of the sense strand, otherwise its value is 0;
  • C5=1 if C is the base at position 5 of the sense strand, otherwise its value is 0;
  • C6=1 if C is the base at position 6 of the sense strand, otherwise its value is 0;
  • C7=1 if C is the base at position 7 of the sense strand, otherwise its value is 0;
  • C9=1 if C is the base at position 9 of the sense strand, otherwise its value is 0;
  • C17=1 if C is the base at position 17 of the sense strand, otherwise its value is 0;
  • C18=1 if C is the base at position 18 of the sense strand, otherwise its value is 0;
  • C19=1 if C is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • G1=1 if G is the base at position 1 on the sense strand, otherwise its value is 0;
  • G2=1 if G is the base at position 2 of the sense strand, otherwise its value is 0;
  • G8=1 if G is the base at position 8 on the sense strand, otherwise its value is 0;
  • G10=1 if G is the base at position 10 on the sense strand, otherwise its value is 0;
      • G13=1 if G is the base at position 13 on the sense strand, otherwise its value is 0;
        G19=1 if G is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
        U1=1 if U is the base at position 1 on the sense strand, otherwise its value is 0;
        U2=1 if U is the base at position 2 on the sense strand, otherwise its value is 0;
        U3=1 if U is the base at position 3 on the sense strand, otherwise its value is 0;
        U4=1 if U is the base at position 4 on the sense strand, otherwise its value is 0;
        U7=1 if U is the base at position 7 on the sense strand, otherwise its value is 0;
        U9=1 if U is the base at position 9 on the sense strand, otherwise its value is 0;
        U10=1 if U is the base at position 10 on the sense strand, otherwise its value is 0;
        U15=1 if U is the base at position 15 on the sense strand, otherwise its value is 0;
        U16=1 if U is the base at position 16 on the sense strand, otherwise its value is 0;
        U17=1 if U is the base at position 17 on the sense strand, otherwise its value is 0;
        U18=1 if U is the base at position 18 on the sense strand, otherwise its value is 0.
  • GC15-19=the number of G and C bases within positions 15-19 of the sense strand, or within positions 15-18 if the sense strand is only 18 base pairs in length;
  • GCtotal=the number of G and C bases in the sense strand;
  • Tm=100 if the siRNA oligo has the internal repeat longer then 4 base pairs, otherwise its value is 0; and
  • X=the number of times that the same nucleotide repeats four or more times in a row.
  • Any of the methods of selecting siRNA in accordance with the invention can further comprise comparing the internal stability profiles of the siRNAs to be selected, and selecting those siRNAs with the most favorable internal stability profiles. Any of the methods of selecting siRNA can further comprise selecting either for or against sequences that contain motifs that induce cellular stress. Such motifs include, for example, toxicity motifs. Any of the methods of selecting siRNA can further comprise either selecting for or selecting against sequences that comprise stability motifs.
  • In another embodiment, the present invention provides a method of gene silencing, comprising introducing into a cell at least one siRNA selected according to any of the methods of the present invention. The siRNA can be introduced by allowing passive uptake of siRNA, or through the use of a vector.
  • According to a third embodiment, the invention provides a method for developing an algorithm for selecting siRNA, said method comprising: (a) selecting a set of siRNA; (b) measuring gene silencing ability of each siRNA from said set; (c) determining relative functionality of each siRNA; (d) determining improved functionality by the presence or absence of at least one variable selected from the group consisting of the presence or absence of a particular nucleotide at a particular position, the total number of As and Us in positions 15-19, the number of times that the same nucleotide repeats within a given sequence, and the total number of Gs and Cs; and (e) developing an algorithm using the information of step (d).
  • In another embodiment, the invention provides a method for selecting an siRNA with improved functionality, comprising using the above-mentioned algorithm to identify an siRNA of improved functionality.
  • According to a fourth embodiment, the present invention provides a kit, wherein said kit is comprised of at least two siRNAs, wherein said at least two siRNAs comprise a first optimized siRNA and a second optimized siRNA, wherein said first optimized siRNA and said second optimized siRNA are optimized according a formula comprising Formula X.
  • According to a fifth embodiment, the present invention provides a method for identifying a hyperfunctional siRNA, comprising applying selection criteria to a set of potential siRNA that comprise 18-30 base pairs, wherein said selection criteria are non-target specific criteria, and said set comprises at least two siRNAs and each of said at least two siRNAs contains a sequence that is at least substantially complementary to a target gene; determining the relative functionality of the at least two siRNAs and assigning each of the at least two siRNAs a functionality score; and selecting siRNAs from the at least two siRNAs that have a functionality score that reflects greater than 80 percent silencing at a concentration in the picomolar range, wherein said greater than 80 percent silencing endures for greater than 120 hours.
  • In other embodiments, the invention provides kits and/or methods wherein the siRNA are comprised of two separate polynucleotide strands; wherein the siRNA are comprised of a single contiguous molecule such as, for example, a unimolecular siRNA (comprising, for example, either a nucleotide or non-nucleotide loop); wherein the siRNA are expressed from one or more vectors; and wherein two or more genes are silenced by a single administration of siRNA.
  • According to a sixth embodiment, the present invention provides a hyperfunctional siRNA that is capable of silencing Bcl2.
  • According to a seventh embodiment, the present invention provides a method for developing an siRNA algorithm for selecting functional and hyperfunctional siRNAs for a given sequence. The method comprises:
  • (a) selecting a set of siRNAs;
  • (b) measuring the gene silencing ability of each siRNA from said set;
  • (c) determining the relative functionality of each siRNA;
  • (d) determining the amount of improved functionality by the presence or absence of at least one variable selected from the group consisting of the total GC content, melting temperature of the siRNA, GC content at positions 15-19, the presence or absence of a particular nucleotide at a particular position, relative thermodynamic stability at particular positions in a duplex, and the number of times that the same nucleotide repeats within a given sequence; and
  • (e) developing an algorithm using the information of step (d).
  • According to this embodiment, preferably the set of siRNAs comprises at least 90 siRNAs from at least one gene, more preferably at least 180 siRNAs from at least two different genes, and most preferably at least 270 and 360 siRNAs from at least three and four different genes, respectively. Additionally, in step (d) the determination is made with preferably at least two, more preferably at least three, even more preferably at least four, and most preferably all of the variables. The resulting algorithm is not target sequence specific.
  • In another embodiment, the present invention provides rationally designed siRNAs identified using the formulas above.
  • In yet another embodiment, the present invention is directed to hyperfunctional siRNA.
  • The ability to use the above algorithms, which are not sequence or species specific, allows for the cost-effective selection of optimized siRNAs for specific target sequences. Accordingly, there will be both greater efficiency and reliability in the use of siRNA technologies.
  • The methods disclosed herein can be used in conjunction with comparing internal stability profiles of selected siRNAs, and designing an siRNA with a desirable internal stability profile; and/or in conjunction with a selection either for or against sequences that contain motifs that induce cellular stress, for example, cellular toxicity.
  • Any of the methods disclosed herein can be used to silence one or more genes by introducing an siRNA selected, or designed, in accordance with any of the methods disclosed herein. The siRNA(s) can be introduced into the cell by any method known in the art, including passive uptake or through the use of one or more vectors.
  • Any of the methods and kits disclosed herein can employ either unimolecular siRNAs, siRNAs comprised of two separate polynucleotide strands, or combinations thereof. Any of the methods disclosed herein can be used in gene silencing, where two or more genes are silenced by a single administration of siRNA(s). The siRNA(s) can be directed against two or more target genes, and administered in a single dose or single transfection, as the case may be.
  • Optimizing siRNA
  • According to one embodiment, the present invention provides a method for improving the effectiveness of gene silencing for use to silence a particular gene through the selection of an optimal siRNA. An siRNA selected according to this method may be used individually, or in conjunction with the first embodiment, i.e., with one or more other siRNAs, each of which may or may not be selected by this criteria in order to maximize their efficiency.
  • The degree to which it is possible to select an siRNA for a given mRNA that maximizes these criteria will depend on the sequence of the mRNA itself. However, the selection criteria will be independent of the target sequence. According to this method, an siRNA is selected for a given gene by using a rational design. That said, rational design can be described in a variety of ways. Rational design is, in simplest terms, the application of a proven set of criteria that enhance the probability of identifying a functional or hyperfunctional siRNA. In one method, rationally designed siRNA can be identified by maximizing one or more of the following criteria:
  • (1) A low GC content, preferably between about 30-52%.
  • (2) At least 2, preferably at least 3 A or U bases at positions 15-19 of the siRNA on the sense strand.
  • (3) An A base at position 19 of the sense strand.
  • (4) An A base at position 3 of the sense strand.
  • (5) A U base at position 10 of the sense strand.
  • (6) An A base at position 14 of the sense strand.
  • (7) A base other than C at position 19 of the sense strand.
  • (8) A base other than G at position 13 of the sense strand.
  • (9) A Tm, which refers to the character of the internal repeat that results in inter- or intramolecular structures for one strand of the duplex, that is preferably not stable at greater than 50° C., more preferably not stable at greater than 37° C., even more preferably not stable at greater than 30° C. and most preferably not stable at greater than 20° C.
  • (10) A base other than U at position 5 of the sense strand.
  • (11) A base other than A at position 11 of the sense strand.
  • (12) A base other than an A at position 1 of the sense strand.
  • (13) A base other than an A at position 2 of the sense strand.
  • (14) An A base at position 4 of the sense strand.
  • (15) An A base at position 5 of the sense strand.
  • (16) An A base at position 6 of the sense strand.
  • (17) An A base at position 7 of the sense strand.
  • (18) An A base at position 8 of the sense strand.
  • (19) A base other than an A at position 9 of the sense strand.
  • (20) A base other than an A at position 10 of the sense strand.
  • (21) A base other than an A at position 11 of the sense strand.
  • (22) A base other than an A at position 12 of the sense strand.
  • (23) An A base at position 13 of the sense strand.
  • (24) A base other than an A at position 14 of the sense strand.
  • (25) An A base at position 15 of the sense strand
  • (26) An A base at position 16 of the sense strand.
  • (27) An A base at position 17 of the sense strand.
  • (28) An A base at position 18 of the sense strand.
  • (29) A base other than a U at position 1 of the sense strand.
  • (30) A base other than a U at position 2 of the sense strand.
  • (31) A U base at position 3 of the sense strand.
  • (32) A base other than a U at position 4 of the sense strand.
  • (33) A base other than a U at position 5 of the sense strand.
  • (34) A U base at position 6 of the sense strand.
  • (35) A base other than a U at position 7 of the sense strand.
  • (36) A base other than a U at position 8 of the sense strand.
  • (37) A base other than a U at position 9 of the sense strand.
  • (38) A base other than a U at position 11 of the sense strand.
  • (39) A U base at position 13 of the sense strand.
  • (40) A base other than a U at position 14 of the sense strand.
  • (41) A base other than a U at position 15 of the sense strand.
  • (42) A base other than a U at position 16 of the sense strand.
  • (43) A U base at position 17 of the sense strand.
  • (44) A U base at position 18 of the sense strand.
  • (45) A U base at position 19 of the sense strand.
  • (46) A C base at position 1 of the sense strand.
  • (47) A C base at position 2 of the sense strand.
  • (48) A base other than a C at position 3 of the sense strand.
  • (49) A C base at position 4 of the sense strand.
  • (50) A base other than a C at position 5 of the sense strand.
  • (51) A base other than a C at position 6 of the sense strand.
  • (52) A base other than a C at position 7 of the sense strand.
  • (53) A base other than a C at position 8 of the sense strand.
  • (54) A C base at position 9 of the sense strand.
  • (55) A C base at position 10 of the sense strand.
  • (56) A C base at position 11 of the sense strand.
  • (57) A base other than a C at position 12 of the sense strand.
  • (58) A base other than a C at position 13 of the sense strand.
  • (59) A base other than a C at position 14 of the sense strand.
  • (60) A base other than a C at position 15 of the sense strand.
  • (61) A base other than a C at position 16 of the sense strand.
  • (62) A base other than a C at position 17 of the sense strand.
  • (63) A base other than a C at position 18 of the sense strand.
  • (64) A G base at position 1 of the sense strand.
  • (65) A G base at position 2 of the sense strand.
  • (66) A G base at position 3 of the sense strand.
  • (67) A base other than a G at position 4 of the sense strand.
  • (68) A base other than a G at position 5 of the sense strand.
  • (69) A G base at position 6 of the sense strand.
  • (70) A G base at position 7 of the sense strand.
  • (71) A G base at position 8 of the sense strand.
  • (72) A G base at position 9 of the sense strand.
  • (73) A base other than a G at position 10 of the sense strand.
  • (74) A G base at position 11 of the sense strand.
  • (75) A G base at position 12 of the sense strand.
  • (76) A G base at position 14 of the sense strand.
  • (77) A G base at position 15 of the sense strand.
  • (78) A G base at position 16 of the sense strand.
  • (79) A base other than a G at position 17 of the sense strand.
  • (80) A base other than a G at position 18 of the sense strand.
  • (81) A base other than a G at position 19 of the sense strand.
  • The importance of various criteria can vary greatly. For instance, a C base at position 10 of the sense strand makes a minor contribution to duplex functionality. In contrast, the absence of a C at position 3 of the sense strand is very important. Accordingly, preferably an siRNA will satisfy as many of the aforementioned criteria as possible.
  • With respect to the criteria, GC content, as well as a high number of AU in positions 15-19 of the sense strand, may be important for easement of the unwinding of double stranded siRNA duplex. Duplex unwinding has been shown to be crucial for siRNA functionality in vivo.
  • With respect to criterion 9, the internal structure is measured in terms of the melting temperature of the single strand of siRNA, which is the temperature at which 50% of the molecules will become denatured. With respect to criteria 2-8 and 10-11, the positions refer to sequence positions on the sense strand, which is the strand that is identical to the mRNA.
  • In one preferred embodiment, at least criteria 1 and 8 are satisfied. In another preferred embodiment, at least criteria 7 and 8 are satisfied. In still another preferred embodiment, at least criteria 1, 8 and 9 are satisfied.
  • It should be noted that all of the aforementioned criteria regarding sequence position specifics are with respect to the 5′ end of the sense strand. Reference is made to the sense strand, because most databases contain information that describes the information of the mRNA. Because according to the present invention a chain can be from 18 to 30 bases in length, and the aforementioned criteria assumes a chain 19 base pairs in length, it is important to keep the aforementioned criteria applicable to the correct bases.
  • When there are only 18 bases, the base pair that is not present is the base pair that is located at the 3′ of the sense strand. When there are twenty to thirty bases present, then additional bases are added at the 5′ end of the sense chain and occupy positions −1 to −11. Accordingly, with respect to SEQ. ID NO. 0001 NNANANNNNUCNAANNNNA and SEQ. ID NO. 0028 GUCNNANANNNNUCNAANNNNA, both would have A at position 3, A at position 5, U at position 10, C at position 11, A and position 13, A and position 14 and A at position 19. However, SEQ. ID NO. 0028 would also have C at position −1, U at position −2 and G at position −3.
  • For a 19 base pair siRNA, an optimal sequence of one of the strands may be represented below, where N is any base, A, C, G, or U:
    SEQ. ID NO. 0001. NNANANNNNUCNAANNNNA
    SEQ. ID NO. 0002. NNANANNNNUGNAANNNNA
    SEQ. ID NO. 0003. NNANANNNNUUNAANNNNA
    SEQ. ID NO. 0004. NNANANNNNUCNCANNNNA
    SEQ. ID NO. 0005. NNANANNNNUGNCANNNNA
    SEQ. ID NO. 0006. NNANANNNNUUNCANNNNA
    SEQ. ID NO. 0007. NNANANNNNUCNUANNNNA
    SEQ. ID NO. 0008. NNANANNNNUGNUANNNNA
    SEQ. ID NO. 0009. NNANANNNNUUNUANNNNA
    SEQ. ID NO. 0010. NNANCNNNNUCNAANNNNA
    SEQ. ID NO. 0011. NNANCNNNNUGNAANNNNA
    SEQ. ID NO. 0012. NNANCNNNNUUNAANNNNA
    SEQ. ID NO. 0013. NNANCNNNNUCNCANNNNA
    SEQ. ID NO. 0014. NNANCNNNNUGNCANNNNA
    SEQ. ID NO. 0015. NNANCNNNNUUNCANNNNA
    SEQ. ID NO. 0016. NNANCNNNNUCNUANNNNA
    SEQ. ID NO. 0017. NNANCNNNNUGNUANNNNA
    SEQ. ID NO. 0018. NNANCNNNNUUNUANNNNA
    SEQ. ID NO. 0019. NNANGNNNNUCNAANNNNA
    SEQ. ID NO. 0020. NNANGNNNNUGNAANNNNA
    SEQ. ID NO. 0021. NNANGNNNNUUNAANNNNA
    SEQ. ID NO. 0022. NNANGNNNNUCNCANNNNA
    SEQ. ID NO. 0023. NNANGNNNNUGNCANNNNA
    SEQ. ID NO. 0024. NNANGNNNNUUNCANNNNA
    SEQ. ID NO. 0025. NNANGNNNNUCNUANNNNA
    SEQ. ID NO. 0026. NNANGNNNNUGNUANNNNA
    SEQ. ID NO. 0027. NNANGNNNNNUNUANNNNA
  • In one embodiment, the sequence used as an siRNA is selected by choosing the siRNA that score highest according to one of the following seven algorithms that are represented by Formulas I-VII:
    Relative functionality of siRNA=−(GC/3)+(AU 15-19)−(Tm 20° C.)*3−(G 13)*3−(C 19)+(A 19)*2+(A 3)+(U 10)+(A 14)−(U 5)−(A 11)  Formula I
    Relative functionality of siRNA=−(GC/3)−(AU 15-19)*3−(G 13)*3−(C 19)+(A 19)*2+(A 3)  Formula II
    Relative functionality of siRNA=−(GC/3)+(AU 15-19)−(Tm 20° C.)*3  Formula III
    Relative functionality of siRNA=−GC/2+(AU 15-19)/2−( Tm 20° C.)*2−(G 13)*3−(C 19)+(A 19)*2+(A 3)+(U 10)+(A 14)−(U 5)−(A 11)  Formula IV
    Relative functionality of siRNA=−(G 13)*3−(C 19)+(A 19)*2+(A 3)+(U 10)+(A 14)−(U 5)−(A 11)  Formula V
    Relative functionality of siRNA=−(G 13)*3−(C 19)+(A 19)*2+(A 3)  Formula VI
    Relative functionality of siRNA=−(GC/2)+(AU 15-19)/2−(Tm 20° C.)*1−(G 13)*3−(C 19)+(A 19)*3+(A 3)*3+(U 10)/2+(A 14)/2−(U 5)/2−(A 11)/2  Formula VII
  • In Formulas I-VII:
  • wherein A19=1 if A is the base at position 19 on the sense strand, otherwise its value is 0,
  • AU15-19=0-5 depending on the number of A or U bases on the sense strand at positions 15-19;
  • G13=1 if G is the base at position 13 on the sense strand, otherwise its value is 0;
  • C19=1 if C is the base at position 19 of the sense strand, otherwise its value is 0;
  • GC=the number of G and C bases in the entire sense strand;
  • Tm20° C.=1 if the Tm is greater than 20° C.;
  • A3=1 if A is the base at position 3 on the sense strand, otherwise its value is 0;
  • U10=1 if U is the base at position 10 on the sense strand, otherwise its value is 0;
  • A14=1 if A is the base at position 14 on the sense strand, otherwise its value is 0;
  • U5=1 if U is the base at position 5 on the sense strand, otherwise its value is 0; and
  • A11=1 if A is the base at position 11 of the sense strand, otherwise its value is 0.
  • Formulas I-VII provide relative information regarding functionality. When the values for two sequences are compared for a given formula, the relative functionality is ascertained; a higher positive number indicates a greater functionality. For example, in many applications a value of 5 or greater is beneficial.
  • Additionally, in many applications, more than one of these formulas would provide useful information as to the relative functionality of potential siRNA sequences. However, it is beneficial to have more than one type of formula, because not every formula will be able to help to differentiate among potential siRNA sequences. For example, in particularly high GC mRNAs, formulas that take that parameter into account would not be useful and application of formulas that lack GC elements (e.g., formulas V and VI) might provide greater insights into duplex functionality. Similarly, formula II might by used in situations where hairpin structures are not observed in duplexes, and formula IV might be applicable for sequences that have higher AU content. Thus, one may consider a particular sequence in light of more than one or even all of these algorithms to obtain the best differentiation among sequences. In some instances, application of a given algorithm may identify an unusually large number of potential siRNA sequences, and in those cases, it may be appropriate to re-analyze that sequence with a second algorithm that is, for instance, more stringent. Alternatively, it is conceivable that analysis of a sequence with a given formula yields no acceptable siRNA sequences (i.e. low SMARTSCORES™, or siRNA ranking). In this instance, it may be appropriate to re-analyze that sequences with a second algorithm that is, for instance, less stringent. In still other instances, analysis of a single sequence with two separate formulas may give rise to conflicting results (i.e. one formula generates a set of siRNA with high SMARTSCORES™, or siRNA ranking, while the other formula identifies a set of siRNA with low SMARTSCORES™, or siRNA ranking). In these instances, it may be necessary to determine which weighted factor(s) (e.g. GC content) are contributing to the discrepancy and assessing the sequence to decide whether these factors should or should not be included. Alternatively, the sequence could be analyzed by a third, fourth, or fifth algorithm to identify a set of rationally designed siRNA.
  • The above-referenced criteria are particularly advantageous when used in combination with pooling techniques as depicted in Table I:
    TABLE I
    FUNCTIONAL PROBABILITY
    OLIGOS POOLS
    CRITERIA >95% >80% <70% >95% >80% <70%
    CURRENT 33.0 50.0 23.0 79.5 97.3 0.3
    NEW 50.0 88.5 8.0 93.8 99.98 0.005
    (GC) 28.0 58.9 36.0 72.8 97.1 1.6
  • The term “current” used in Table I refers to Tuschl's conventional siRNA parameters (Elbashir, S. M. et al. (2002) “Analysis of gene function in somatic mammalian cells using small interfering RNAs” Methods 26: 199-213). “New” refers to the design parameters described in Formulas I-VII. “GC” refers to criteria that select siRNA solely on the basis of GC content.
  • As Table I indicates, when more functional siRNA duplexes are chosen, siRNAs that produce <70% silencing drops from 23% to 8% and the number of siRNA duplexes that produce >80% silencing rises from 50% to 88.5%. Further, of the siRNA duplexes with >80% silencing, a larger portion of these siRNAs actually silence >95% of the target expression (the new criteria increases the portion from 33% to 50%). Using this new criteria in pooled siRNAs, shows that, with pooling, the amount of silencing >95% increases from 79.5% to 93.8% and essentially eliminates any siRNA pool from silencing less than 70%.
  • Table II similarly shows the particularly beneficial results of pooling in combination with the aforementioned criteria. However, Table II, which takes into account each of the aforementioned variables, demonstrates even a greater degree of improvement in functionality.
    TABLE II
    FUNCTIONAL PROBABILITY
    OLIGOS POOLS
    NON- NON-
    FUNCTIONAL AVERAGE FUNCTIONAL FUNCTIONAL AVERAGE FUNCTIONAL
    RANDOM 20 40 50 67 97 3
    CRITERIA 1 52 99 0.1 97 93 0.0040
    CRITERIA 4 89 99 0.1 99 99 0.0000
  • The terms “functional,” “Average,” and “Non-functional” used in Table II, refer to siRNA that exhibit >80%, >50%, and <50% functionality, respectively. Criteria 1 and 4 refer to specific criteria described above.
  • The above-described algorithms may be used with or without a computer program that allows for the inputting of the sequence of the mRNA and automatically outputs the optimal siRNA. The computer program may, for example, be accessible from a local terminal or personal computer, over an internal network or over the Internet.
  • In addition to the formulas above, more detailed algorithms may be used for selecting siRNA. Preferably, at least one RNA duplex of 18-30 base pairs is selected such that it is optimized according a formula selected from:
    (−14)*G13−13*A1−12*U7−11*U2−10*A11−10*U4−10*C3−10*C5−10*C6−9*A10−9*U9−9*C18−8*G10−7*U1−7*U16−7*C17−7*C19+7*U17+8*A2+8*A4+8*A5+8*C4+9*G8+10*A7+10*U18+11*A19+11*C9+15*G1+18*A3+19*U10−Tm−3*(GC total)−6*(GC15-19)−30*X; and  Formula VIII
    (14.1)*A3+(14.9)*A6+(17.6)*A13+(24.7)*A19+(14.2)*U10+(10.5)*C9+(23.9)*G1+(16.3)*G2+(−12.3)*A11+(−19.3)*U1+(−12.1)*U2+(−11)*U3+(−15.2)*U15+(−11.3)*U16+(−11.8)*C3+(−17.4)*C6+(−10.5)*C7+(−13.7)*G13+(−25.9)*G19−Tm−3*(GCtotal)−6*(GC15-19)−30*X; and  Formula IX
    (−8)*A1+(−1)*A2+(12)*A3+(7)*A4+(18)*A5+(12)*A6+(19)*A7+(6)*A8+(−4)*A9+(−5)*A10+(−2)*A11+(−5)*A12+(17)*A13+(−3)*A14+(4)*A15+(2)*A16+(8)*A17+(11)*A18+(30)*A19+(−13)*U1+(−10)*U2+(2)*U3+(−2)*U4+(−5)*U5+(5)*U6+(−2)*U7+(−10)*U8+(−5)*U9+(15)*U10+(−1)*U11+(0)*U12+(10)*U13+(−9)*U14+(−13)*U15+(−10)*U16+(3)*U17+(9)*U18+(9)*U19+(7)*C1+(3)*C2+(−21)*C3+(5)*C4+(−9)*C5+(−20)*C6+(−18)*C7+(−5)*C8+(5)*C9+(1)*C10+(2)*C11+(−5)*C12+(−3)*C13+(−6)*C14+(−2)*C15+(−5)*C16+(−3)*C17+(−12)*C18+(−18)*C19+(14)*G1+(8)*G2+(7)*G3+(−10)*G4+(−4)*G5+(2)*G6+(1)*G7+(9)*G8+(5)*G9+(−11)*G10+(1)*G11+(9)*G12+(−24)*G13+(18)*G14+(11)*G15+(13)*G16+(−7)*G17+(−9)*G18+(−22)*G19+6*(number of A+U in position 15-19)−3*(number of G+C in whole siRNA).  Formula X
  • wherein
  • A1=1 if A is the base at position 1 of the sense strand, otherwise its value is 0;
  • A2=1 if A is the base at position 2 of the sense strand, otherwise its value is 0;
  • A3=1 if A is the base at position 3 of the sense strand, otherwise its value is 0;
  • A4=1 if A is the base at position 4 of the sense strand, otherwise its value is 0;
  • A5=1 if A is the base at position 5 of the sense strand, otherwise its value is 0;
  • A6=1 if A is the base at position 6 of the sense strand, otherwise its value is 0;
  • A7=1 if A is the base at position 7 of the sense strand, otherwise its value is 0;
  • A10=1 if A is the base at position 10 of the sense strand, otherwise its value is 0;
  • A11=1 if A is the base at position 11 of the sense strand, otherwise its value is 0;
  • A13=1 if A is the base at position 13 of the sense strand, otherwise its value is 0;
  • A19=1 if A is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • C3=1 if C is the base at position 3 of the sense strand, otherwise its value is 0;
  • C4=1 if C is the base at position 4 of the sense strand, otherwise its value is 0;
  • C5=1 if C is the base at position 5 of the sense strand, otherwise its value is 0;
  • C6=1 if C is the base at position 6 of the sense strand, otherwise its value is 0;
  • C7=1 if C is the base at position 7 of the sense strand, otherwise its value is 0;
  • C9=1 if C is the base at position 9 of the sense strand, otherwise its value is 0;
  • C17=1 if C is the base at position 17 of the sense strand, otherwise its value is 0;
  • C18=1 if C is the base at position 18 of the sense strand, otherwise its value is 0;
  • C19=1 if C is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • G1=1 if G is the base at position 1 on the sense strand, otherwise its value is 0;
  • G2=1 if G is the base at position 2 of the sense strand, otherwise its value is 0;
  • G8=1 if G is the base at position 8 on the sense strand, otherwise its value is 0;
  • G10=1 if G is the base at position 10 on the sense strand, otherwise its value is 0;
  • G13=1 if G is the base at position 13 on the sense strand, otherwise its value is 0;
  • G19=1 if G is the base at position 19 of the sense strand, otherwise if another base is present or the sense strand is only 18 base pairs in length, its value is 0;
  • U1=1 if U is the base at position 1 on the sense strand, otherwise its value is 0;
  • U2=1 if U is the base at position 2 on the sense strand, otherwise its value is 0;
  • U3=1 if U is the base at position 3 on the sense strand, otherwise its value is 0;
  • U4=1 if U is the base at position 4 on the sense strand, otherwise its value is 0;
  • U7=1 if U is the base at position 7 on the sense strand, otherwise its value is 0;
  • U9=1 if U is the base at position 9 on the sense strand, otherwise its value is 0;
  • U10=1 if U is the base at position 10 on the sense strand, otherwise its value is 0;
  • U15=1 if U is the base at position 15 on the sense strand, otherwise its value is 0;
  • U16=1 if U is the base at position 16 on the sense strand, otherwise its value is 0;
  • U17=1 if U is the base at position 17 on the sense strand, otherwise its value is 0;
  • U18=1 if U is the base at position 18 on the sense strand, otherwise its value is 0;
  • GC15-19=the number of G and C bases within positions 15-19 of the sense strand, or within positions 15-18 if the sense strand is only 18 base pairs in length;
  • GCtotal=the number of G and C bases in the sense strand;
  • Tm=100 if the siRNA oligo has the internal repeat longer then 4 base pairs, otherwise its value is 0; and
  • X=the number of times that the same nucleotide repeats four or more times in a row.
  • The above formulas VIII, IX, and X, as well as formulas I-VII, provide methods for selecting siRNA in order to increase the efficiency of gene silencing. A subset of variables of any of the formulas may be used, though when fewer variables are used, the optimization hierarchy becomes less reliable.
  • With respect to the variables of the above-referenced formulas, a single letter of A or C or G or U followed by a subscript refers to a binary condition. The binary condition is that either the particular base is present at that particular position (wherein the value is “1”) or the base is not present (wherein the value is “0”). Because position 19 is optional, i.e., there might be only 18 base pairs, when there are only 18 base pairs, any base with a subscript of 19 in the formulas above would have a zero value for that parameter. Before or after each variable is a number followed by *, which indicates that the value of the variable is to be multiplied or weighed by that number.
  • The numbers preceding the variables A, or G, or C, or U in Formulas VIII, IX, and X (or after the variables in Formula I-VII) were determined by comparing the difference in the frequency of individual bases at different positions in functional siRNA and total siRNA. Specifically, the frequency in which a given base was observed at a particular position in functional groups was compared with the frequency that that same base was observed in the total, randomly selected siRNA set. If the absolute value of the difference between the functional and total values was found to be greater than 6%, that parameter was included in the equation. Thus, for instance, if the frequency of finding a “G” at position 13 (G13) is found to be 6% in a given functional group, and the frequency of G13 in the total population of siRNAs is 20%, the difference between the two values is 6%-20%=−14%. As the absolute value is greater than six (6), this factor (−14) is included in the equation. Thus, in Formula VIII, in cases where the siRNA under study has a G in position 13, the accrued value is (−14)*(1)=−14. In contrast, when a base other than G is found at position 13, the accrued value is (−14)*(0)=0.
  • When developing a means to optimize siRNAs, the inventors observed that a bias toward low internal thermodynamic stability of the duplex at the 5′-antisense (AS) end is characteristic of naturally occurring miRNA precursors. The inventors extended this observation to siRNAs for which functionality had been assessed in tissue culture.
  • With respect to the parameter GC15-19, a value of 0-5 will be ascribed depending on the number of G or C bases at positions 15 to 19. If there are only 18 base pairs, the value is between 0 and 4.
  • With respect to the criterion GCtotal content, a number from 0-30 will be ascribed, which correlates to the total number of G and C nucleotides on the sense strand, excluding overhangs. Without wishing to be bound by any one theory, it is postulated that the significance of the GC content (as well as AU content at positions 15-19, which is a parameter for formulas III-VII) relates to the easement of the unwinding of a double-stranded siRNA duplex. Duplex unwinding is believed to be crucial for siRNA functionality in vivo and overall low internal stability, especially low internal stability of the first unwound base pair is believed to be important to maintain sufficient processivity of RISC complex-induced duplex unwinding. If the duplex has 19 base pairs, those at positions 15-19 on the sense strand will unwind first if the molecule exhibits a sufficiently low internal stability at that position. As persons skilled in the art are aware, RISC is a complex of approximately twelve proteins; Dicer is one, but not the only, helicase within this complex. Accordingly, although the GC parameters are believed to relate to activity with Dicer, they are also important for activity with other RISC proteins.
  • The value of the parameter Tm is 0 when there are no internal repeats longer than (or equal to) four base pairs present in the siRNA duplex; otherwise the value is 1. Thus for example, if the sequence ACGUACGU, or any other four nucleotide (or more) palindrome exists within the structure, the value will be one (1). Alternatively if the structure ACGGACG, or any other 3 nucleotide (or less) palindrome exists, the value will be zero (0).
  • The variable “X” refers to the number of times that the same nucleotide occurs contiguously in a stretch of four or more units. If there are, for example, four contiguous As in one part of the sequence and elsewhere in the sequence four contiguous Cs, X=2. Further, if there are two separate contiguous stretches of four of the same nucleotides or eight or more of the same nucleotides in a row, then X=2. However, X does not increase for five, six or seven contiguous nucleotides.
  • Again, when applying Formula VIII, Formula IX, or Formula X, to a given mRNA, (the “target RNA” or “target molecule”), one may use a computer program to evaluate the criteria for every sequence of 18-30 base pairs or only sequences of a fixed length, e.g., 19 base pairs. Preferably the computer program is designed such that it provides a report ranking of all of the potential siRNAs 18-30 base pairs, ranked according to which sequences generate the highest value. A higher value refers to a more efficient siRNA for a particular target gene. The computer program that may be used may be developed in any computer language that is known to be useful for scoring nucleotide sequences, or it may be developed with the assistance of commercially available product such as Microsoft's PRODUCT.NET. Additionally, rather than run every sequence through one and/or another formula, one may compare a subset of the sequences, which may be desirable if for example only a subset are available. For instance, it may be desirable to first perform a BLAST (Basic Local Alignment Search Tool) search and to identify sequences that have no homology to other targets. Alternatively, it may be desirable to scan the sequence and to identify regions of moderate GC context, then perform relevant calculations using one of the above-described formulas on these regions. These calculations can be done manually or with the aid of a computer.
  • As with Formulas I-VII, either Formula VIII, Formula IX, or Formula X may be used for a given mRNA target sequence. However, it is possible that according to one or the other formula more than one siRNA will have the same value. Accordingly, it is beneficial to have a second formula by which to differentiate sequences. Formulas IX and X were derived in a similar fashion as Formula VIII, yet used a larger data set and thus yields sequences with higher statistical correlations to highly functional duplexes. The sequence that has the highest value ascribed to it may be referred to as a “first optimized duplex.” The sequence that has the second highest value ascribed to it may be referred to as a “second optimized duplex.” Similarly, the sequences that have the third and fourth highest values ascribed to them may be referred to as a third optimized duplex and a fourth optimized duplex, respectively. When more than one sequence has the same value, each of them may, for example, be referred to as first optimized duplex sequences or co-first optimized duplexes. Formula X is similar to Formula IX, yet uses a greater numbers of variables and for that reason, identifies sequences on the basis of slightly different criteria.
  • It should also be noted that the output of a particular algorithm will depend on several of variables including: (1) the size of the data base(s) being analyzed by the algorithm, and (2) the number and stringency of the parameters being applied to screen each sequence. Thus, for example, in U.S. patent application Ser. No. 10/714,333, entitled “Functional and Hyperfunctional siRNA,” filed Nov. 14, 2003, Formula VIII was applied to the known human genome (NCBI REFSEQ database) through ENTREZ (EFETCH). As a result of these procedures, roughly 1.6 million siRNA sequences were identified. Application of Formula VIII to the same database in March of 2004 yielded roughly 2.2 million sequences, a difference of approximately 600,000 sequences resulting from the growth of the database over the course of the months that span this period of time. Application of other formulas (e.g., Formula X) that change the emphasis of, include, or eliminate different variables can yield unequal numbers of siRNAs. Alternatively, in cases where application of one formula to one or more genes fails to yield sufficient numbers of siRNAs with scores that would be indicative of strong silencing, said genes can be reassessed with a second algorithm that is, for instance, less stringent.
  • siRNA sequences identified using Formula VIII and Formula X (minus sequences generated by Formula VIII) are contained within the sequence listing. The data included in the sequence listing is described more fully below. The sequences identified by Formula VIII and Formula X that are disclosed in the sequence listing may be used in gene silencing applications.
  • It should be noted that for Formulas VIII, IX, and X all of the aforementioned criteria are identified as positions on the sense strand when oriented in the 5′ to 3′ direction as they are identified in connection with Formulas I-VII unless otherwise specified.
  • Formulas I-X, may be used to select or to evaluate one, or more than one, siRNA in order to optimize silencing. Preferably, at least two optimized siRNAs that have been selected according to at least one of these formulas are used to silence a gene, more preferably at least three and most preferably at least four. The siRNAs may be used individually or together in a pool or kit. Further, they may be applied to a cell simultaneously or separately. Preferably, the at least two siRNAs are applied simultaneously. Pools are particularly beneficial for many research applications. However, for therapeutics, it may be more desirable to employ a single hyperfunctional siRNA as described elsewhere in this application.
  • When planning to conduct gene silencing, and it is necessary to choose between two or more siRNAs, one should do so by comparing the relative values when the siRNA are subjected to one of the formulas above. In general a higher scored siRNA should be used.
  • Useful applications include, but are not limited to, target validation, gene functional analysis, research and drug discovery, gene therapy and therapeutics. Methods for using siRNA in these applications are well known to persons of skill in the art.
  • Because the ability of siRNA to function is dependent on the sequence of the RNA and not the species into which it is introduced, the present invention is applicable across a broad range of species, including but not limited to all mammalian species, such as humans, dogs, horses, cats, cows, mice, hamsters, chimpanzees and gorillas, as well as other species and organisms such as bacteria, viruses, insects, plants and C. elegans.
  • The present invention is also applicable for use for silencing a broad range of genes, including but not limited to the roughly 45,000 genes of a human genome, and has particular relevance in cases where those genes are associated with diseases such as diabetes, Alzheimer's, cancer, as well as all genes in the genomes of the aforementioned organisms.
  • The siRNA selected according to the aforementioned criteria or one of the aforementioned algorithms are also, for example, useful in the simultaneous screening and functional analysis of multiple genes and gene families using high throughput strategies, as well as in direct gene suppression or silencing.
  • Development of the Algorithms
  • To identify siRNA sequence features that promote functionality and to quantify the importance of certain currently accepted conventional factors—such as G/C content and target site accessibility—the inventors synthesized an siRNA panel consisting of 270 siRNAs targeting three genes, Human Cyclophilin, Firefly Luciferase, and Human DBI. In all three cases, siRNAs were directed against specific regions of each gene. For Human Cyclophilin and Firefly Luciferase, ninety siRNAs were directed against a 199 bp segment of each respective mRNA. For DBI, 90 siRNAs were directed against a smaller, 109 base pair region of the mRNA. The sequences to which the siRNAs were directed are provided below.
  • It should be noted that in certain sequences, “t” is present. This is because many databases contain information in this manner. However, the t denotes a uracil residue in mRNA and siRNA. Any algorithm will, unless otherwise specified, process at in a sequence as a u.
  • Human Cyclophilin: 193-390, M60857
    SEQ. ID NO. 29:
    gttccaaaaa cagtggataa ttttgtggcc ttagctacag
    gagagaaagg atttggctac aaaaacagca aattccatcg
    tgtaatcaag gacttcatga tccagggcgg agacttcacc
    aggggagatg gcacaggagg aaagagcatc tacggtgagc
    gcttccccga tgagaacttc aaactgaagc actacgggcc
    tggctggg
  • Firefly Luciferase: 1434-1631, U47298 (pGL3, Promega)
    SEQ. ID NO. 30:
    tgaacttccc gccgccgttq ttgttttgga gcacggaaag
    acgatgacgg aaaaagagat cgtggattac gtcgccagtc
    aagtaacaac cgcgaaaaag ttgcgcggag gagttgtgtt
    tgtggacgaa gtaccgaaag gtcttaccgg aaaactcgac
    gcaagaaaaa toagagagat cctcataaag gccaagaagg
  • DBI, NM—020548 (202-310) (Every Position)
    SEQ. ID NO. 0031:
    acgggcaagg ccaagtggga tgcctggaat gagctgaaag
    ggacttccaa ggaagatgcc atgaaagctt acatcaacaa
    agtagaagag ctaaagaaaa aatacggg
  • A list of the siRNAs appears in Table III (see Examples Section, Example II)
  • The set of duplexes was analyzed to identify correlations between siRNA functionality and other biophysical or thermodynamic properties. When the siRNA panel was analyzed in functional and non-functional subgroups, certain nucleotides were much more abundant at certain positions in functional or non-functional groups. More specifically, the frequency of each nucleotide at each position in highly functional siRNA duplexes was compared with that of nonfunctional duplexes in order to assess the preference for or against any given nucleotide at every position. These analyses were used to determine important criteria to be included in the siRNA algorithms (Formulas VIII, IX, and X).
  • The data set was also analyzed for distinguishing biophysical properties of siRNAs in the functional group, such as optimal percent of GC content, propensity for internal structures and regional thermodynamic stability. Of the presented criteria, several are involved in duplex recognition, RISC activation/duplex unwinding, and target cleavage catalysis.
  • The original data set that was the source of the statistically derived criteria is shown in FIG. 2. Additionally, this figure shows that random selection yields siRNA duplexes with unpredictable and widely varying silencing potencies as measured in tissue culture using HEK293 cells. In the figure, duplexes are plotted such that each x-axis tick-mark represents an individual siRNA, with each subsequent siRNA differing in target position by two nucleotides for Human Cyclophilin B and Firefly Luciferase, and by one nucleotide for Human DBI. Furthermore, the y-axis denotes the level of target expression remaining after transfection of the duplex into cells and subsequent silencing of the target.
  • siRNA identified and optimized in this document work equally well in a wide range of cell types. FIG. 3 a shows the evaluation of thirty siRNAs targeting the DBI gene in three cell lines derived from different tissues. Each DBI siRNA displays very similar functionality in HEK293 (ATCC, CRL-1573, human embryonic kidney), HeLa (ATCC, CCL-2, cervical epithelial adenocarcinoma) and DU145 (HTB-81, prostate) cells as determined by the B-DNA assay. Thus, siRNA functionality is determined by the primary sequence of the siRNA and not by the intracellular environment. Additionally, it should be noted that although the present invention provides for a determination of the functionality of siRNA for a given target, the same siRNA may silence more than one gene. For example, the complementary sequence of the silencing siRNA may be present in more than one gene. Accordingly, in these circumstances, it may be desirable not to use the siRNA with highest SMARTSCORE™, or siRNA ranking. In such circumstances, it may be desirable to use the siRNA with the next highest SMARTSCORE™, or siRNA ranking.
  • To determine the relevance of G/C content in siRNA function, the G/C content of each duplex in the panel was calculated and the functional classes of siRNAs (<F50, ≧F50, ≧F80, ≧F95 where F refers to the percent gene silencing) were sorted accordingly. The majority of the highly-functional siRNAs (≧F95) fell within the G/C content range of 36%-52% (FIG. 3B). Twice as many non-functional (<F50) duplexes fell within the high G/C content groups (>57% GC content) compared to the 36%-52% group. The group with extremely low GC content (26% or less) contained a higher proportion of non-functional siRNAs and no highly-functional siRNAs. The G/C content range of 30%-52% was therefore selected as Criterion I for siRNA functionality, consistent with the observation that a G/C range 30%-70% promotes efficient RNAi targeting. Application of this criterion alone provided only a marginal increase in the probability of selecting functional siRNAs from the panel: selection of F50 and F95 siRNAs was improved by 3.6% and 2.2%, respectively. The siRNA panel presented here permitted a more systematic analysis and quantification of the importance of this criterion than that used previously.
  • A relative measure of local internal stability is the A/U base pair (bp) content; therefore, the frequency of A/U bp was determined for each of the five terminal positions of the duplex (5′ sense (S)/5′ antisense (AS)) of all siRNAs in the panel. Duplexes were then categorized by the number of A/U bp in positions 1-5 and 15-19 of the sense strand. The thermodynamic flexibility of the duplex 5′-end (positions 1-5; S) did not appear to correlate appreciably with silencing potency, while that of the 3′-end (positions 15-19; S) correlated with efficient silencing. No duplexes lacking A/U bp in positions 15-19 were functional. The presence of one A/U bp in this region conferred some degree of functionality, but the presence of three or more A/Us was preferable and therefore defined as Criterion II. When applied to the test panel, only a marginal increase in the probability of functional siRNA selection was achieved: a 1.8% and 2.3% increase for F50 and F95 duplexes, respectively (Table IV).
  • The complementary strands of siRNAs that contain internal repeats or palindromes may form internal fold-back structures. These hairpin-like structures exist in equilibrium with the duplexed form effectively reducing the concentration of functional duplexes. The propensity to form internal hairpins and their relative stability can be estimated by predicted melting temperatures. High Tm reflects a tendency to form hairpin structures. Lower Tm values indicate a lesser tendency to form hairpins. When the functional classes of siRNAs were sorted by Tm (FIG. 3 c), the following trends were identified: duplexes lacking stable internal repeats were the most potent silencers (no F95 duplex with predicted hairpin structure Tm>60° C.). In contrast, about 60% of the duplexes in the groups having internal hairpins with calculated Tm values less than 20° C. were F80. Thus, the stability of internal repeats is inversely proportional to the silencing effect and defines Criterion III (predicted hairpin structure Tm≦20° C.).
  • Sequence-Based Determinants of siRNA Functionality
  • When the siRNA panel was sorted into functional and non-functional groups, the frequency of a specific nucleotide at each position in a functional siRNA duplex was compared with that of a nonfunctional duplex in order to assess the preference for or against a certain nucleotide. FIG. 4 shows the results of these queries and the subsequent resorting of the data set (from FIG. 2). The data is separated into two sets: those duplexes that meet the criteria, a specific nucleotide in a certain position—grouped on the left (Selected) and those that do not—grouped on the right (Eliminated). The duplexes are further sorted from most functional to least functional with the y-axis of FIG. 4 a-e representing the % expression i.e., the amount of silencing that is elicited by the duplex (Note: each position on the X-axis represents a different duplex). Statistical analysis revealed correlations between silencing and several sequence-related properties of siRNAs. FIG. 4 and Table IV show quantitative analysis for the following five sequence-related properties of siRNA: (A) an A at position 19 of the sense strand; (B) an A at position 3 of the sense strand; (C) a U at position 10 of the sense strand; (D) a base other than G at position 13 of the sense strand; and (E) a base other than C at position 19 of the sense strand.
  • When the siRNAs in the panel were evaluated for the presence of an A at position 19 of the sense strand, the percentage of non-functional duplexes decreased from 20% to 11.8%, and the percentage of F95 duplexes increased from 21.7% to 29.4% (Table IV). Thus, the presence of an A in this position defined Criterion IV.
  • Another sequence-related property correlated with silencing was the presence of an A in position 3 of the sense strand (FIG. 4 b). Of the siRNAs with A3, 34.4% were F95, compared with 21.7% randomly selected siRNAs. The presence of a U base in position 10 of the sense strand exhibited an even greater impact (FIG. 4 c). Of the duplexes in this group, 41.7% were F95. These properties became criteria V and VI, respectively.
  • Two negative sequence-related criteria that were identified also appear on FIG. 4. The absence of a G at position 13 of the sense strand, conferred a marginal increase in selecting functional duplexes (FIG. 4 d). Similarly, lack of a C at position 19 of the sense strand also correlated with functionality (FIG. 4 e). Thus, among functional duplexes, position 19 was most likely occupied by A, and rarely occupied by C. These rules were defined as criteria VII and VIII, respectively.
  • Application of each criterion individually provided marginal but statistically significant increases in the probability of selecting a potent siRNA. Although the results were informative, the inventors sought to maximize potency and therefore consider multiple criteria or parameters. Optimization is particularly important when developing therapeutics. Interestingly, the probability of selecting a functional siRNA based on each thermodynamic criteria was 2%-4% higher than random, but 4%-8% higher for the sequence-related determinates. Presumably, these sequence-related increases reflect the complexity of the RNAi mechanism and the multitude of protein-RNA interactions that are involved in RNAi-mediated silencing.
    TABLE IV
    PERCENT IMPROVEMENT
    CRITERION FUNCTIONAL OVER RANDOM (%)
    I. 30%-52% G/C Content <F50 16.4 −3.6
    ≧F50 83.6 3.6
    ≧F80 60.4 4.3
    ≧F95 23.9 2.2
    II. At least 3 A/U bases at <F50 18.2 −1.8
    positions 15-19 of the sense ≧F50 81.8 1.8
    strand ≧F80 59.7 3.6
    ≧F95 24.0 2.3
    III. Absence of internal <F50 16.7 −3.3
    repeats, as measured by Tm of ≧F50 83.3 3.3
    secondary structure ≦20° C. ≧F80 61.1 5.0
    ≧F95 24.6 2.9
    IV. An A base at position 19 <F50 11.8 −8.2
    of the sense strand ≧F50 88.2 8.2
    ≧F80 75.0 18.9
    ≧F95 29.4 7.7
    V. An A base at position 3 of <F50 17.2 −2.8
    the sense strand ≧F50 82.8 2.8
    ≧F80 62.5 6.4
    ≧F95 34.4 12.7
    VI. A U base at position 10 <F50 13.9 −6.1
    of the sense strand ≧F50 86.1 6.1
    ≧F80 69.4 13.3
    ≧F95 41.7 20
    VII. A base other than C at <F50 18.8 −1.2
    position 19 of the sense strand ≧F50 81.2 1.2
    ≧F80 59.7 3.6
    ≧F95 24.2 2.5
    VIII. A base other than G at <F50 15.2 −4.8
    position 13 of the sense strand ≧F50 84.8 4.8
    ≧F80 61.4 5.3
    ≧F95 26.5 4.8

    The siRNA Selection Algorithm
  • In an effort to improve selection further, all identified criteria, including but not limited to those listed in Table IV were combined into the algorithms embodied in Formula VIII, Formula IX, and Formula X. Each siRNA was then assigned a score (referred to as a SMARTSCORE™, or siRNA ranking) according to the values derived from the formulas. Duplexes that scored higher than 0 or −20 (unadjusted), for Formulas VIII and IX, respectively, effectively selected a set of functional siRNAs and excluded all non-functional siRNAs. Conversely, all duplexes scoring lower than 0 and −20 (minus 20) according to formulas VIII and IX, respectively, contained some functional siRNAs but included all non-functional siRNAs. A graphical representation of this selection is shown in FIG. 5. It should be noted that the scores derived from the algorithm can also be provided as “adjusted” scores. To convert Formula VIII unadjusted scores into adjusted scores it is necessary to use the following equation:
    (160+unadjusted score)/2.25
  • When this takes place, an unadjusted score of “0” (zero) is converted to 75. Similarly, unadjusted scores for Formula X can be converted to adjusted scores. In this instance, the following equation is applied:
    (228+unadjusted score)/3.56
  • When these manipulations take place, an unadjusted score of 38 is converted to an adjusted score of 75.
  • The methods for obtaining the seven criteria embodied in Table IV are illustrative of the results of the process used to develop the information for Formulas VIII, IX, and X. Thus similar techniques were used to establish the other variables and their multipliers. As described above, basic statistical methods were use to determine the relative values for these multipliers.
  • To determine the value for “Improvement over Random” the difference in the frequency of a given attribute (e.g., GC content, base preference) at a particular position is determined between individual functional groups (e.g., <F50) and the total siRNA population studied (e.g., 270 siRNA molecules selected randomly). Thus, for instance, in Criterion I (30%-52% GC content) members of the <F50 group were observed to have GC contents between 30-52% in 16.4% of the cases. In contrast, the total group of 270 siRNAs had GC contents in this range, 20% of the time. Thus for this particular attribute, there is a small negative correlation between 30%-52% GC content and this functional group (i.e., 16.4%-20%=−3.6%). Similarly, for Criterion VI, (a “U” at position 10 of the sense strand), the >F95 group contained a “U” at this position 41.7% of the time. In contrast, the total group of 270 siRNAs had a “U” at this position 21.7% of the time, thus the improvement over random is calculated to be 20% (or 41.7%-21.7%).
  • Identifying the Average Internal Stability Profile of Strong siRNA
  • In order to identify an internal stability profile that is characteristic of strong siRNA, 270 different siRNAs derived from the cyclophilin B, the diazepam binding inhibitor (DBI), and the luciferase gene were individually transfected into HEK293 cells and tested for their ability to induce RNAi of the respective gene. Based on their performance in the in vivo assay, the sequences were then subdivided into three groups, (i) >95% silencing; (ii) 80-95% silencing; and (iii) less than 50% silencing. Sequences exhibiting 51-84% silencing were eliminated from further consideration to reduce the difficulties in identifying relevant thermodynamic patterns.
  • Following the division of siRNA into three groups, a statistical analysis was performed on each member of each group to determine the average internal stability profile (AISP) of the siRNA. To accomplish this the Oligo 5.0 Primer Analysis Software and other related statistical packages (e.g., Excel) were exploited to determine the internal stability of pentamers using the nearest neighbor method described by Freier et al., (1986) Improved free-energy parameters for predictions of RNA duplex stability, Proc Natl. Acad. Sci. USA 83(24): 9373-7. Values for each group at each position were then averaged, and the resulting data were graphed on a linear coordinate system with the Y-axis expressing the AG (free energy) values in kcal/mole and the X-axis identifying the position of the base relative to the 5′ end.
  • The results of the analysis identified multiple key regions in siRNA molecules that were critical for successful gene silencing. At the 3′-most end of the sense strand (5′antisense), highly functional siRNA (>95% gene silencing, see FIG. 6 a, >F95) have a low internal stability (AISP of position 19=˜−7.6 kcal/mol). In contrast low-efficiency siRNA (i.e., those exhibiting less than 50% silencing, <F50) display a distinctly different profile, having high ΔG values (˜−8.4 kcal/mol) for the same position. Moving in a 5′ (sense strand) direction, the internal stability of highly efficient siRNA rises (position 12=˜−8.3 kcal/mole) and then drops again (position 7=˜−7.7 kcal/mol) before leveling off at a value of approximately −8.1 kcal/mol for the 5′ terminus. siRNA with poor silencing capabilities show a distinctly different profile. While the AISP value at position 12 is nearly identical with that of strong siRNAs, the values at positions 7 and 8 rise considerably, peaking at a high of ˜−9.0 kcal/mol. In addition, at the 5′ end of the molecule the AISP profile of strong and weak siRNA differ dramatically. Unlike the relatively strong values exhibited by siRNA in the >95% silencing group, siRNAs that exhibit poor silencing activity have weak AISP values (−7.6, −7.5, and −7.5 kcal/mol for positions 1, 2 and 3 respectively).
  • Overall the profiles of both strong and weak siRNAs form distinct sinusoidal shapes that are roughly 180° out-of-phase with each other. While these thermodynamic descriptions define the archetypal profile of a strong siRNA, it will likely be the case that neither the ΔG values given for key positions in the profile or the absolute position of the profile along the Y-axis (i.e., the ΔG-axis) are absolutes. Profiles that are shifted upward or downward (i.e., having on an average, higher or lower values at every position) but retain the relative shape and position of the profile along the X-axis can be foreseen as being equally effective as the model profile described here. Moreover, it is likely that siRNA that have strong or even stronger gene-specific silencing effects might have exaggerated ΔG values (either higher or lower) at key positions. Thus, for instance, it is possible that the 5′-most position of the sense strand (position 19) could have ΔG values of 7.4 kcal/mol or lower and still be a strong siRNA if, for instance, a G-C→G-T/U mismatch were substituted at position 19 and altered duplex stability. Similarly, position 12 and position 7 could have values above 8.3 kcal/mol and below 7.7 kcal/mole, respectively, without abating the silencing effectiveness of the molecule. Thus, for instance, at position 12, a stabilizing chemical modification (e.g., a chemical modification of the 2′ position of the sugar backbone) could be added that increases the average internal stability at that position. Similarly, at position 7, mismatches similar to those described previously could be introduced that would lower the AG values at that position.
  • Lastly, it is important to note that while functional and non-functional siRNA were originally defined as those molecules having specific silencing properties, both broader or more limiting parameters can be used to define these molecules. As used herein, unless otherwise specified, “non-functional siRNA” are defined as those siRNA that induce less than 50% (<50%) target silencing, “semi-functional siRNA” induce 50-79% target silencing, “functional siRNA” are molecules that induce 80-95% gene silencing, and “highly-functional siRNA” are molecules that induce great than 95% gene silencing. These definitions are not intended to be rigid and can vary depending upon the design and needs of the application. For instance, it is possible that a researcher attempting to map a gene to a chromosome using a functional assay, may identify an siRNA that reduces gene activity by only 30%. While this level of gene silencing may be “non-functional” for, e.g., therapeutic needs, it is sufficient for gene mapping purposes and is, under these uses and conditions, “functional.” For these reasons, functional siRNA can be defined as those molecules having greater than 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% silencing capabilities at 100 nM transfection conditions. Similarly, depending upon the needs of the study and/or application, non-functional and semi-functional siRNA can be defined as having different parameters. For instance, semi-functional siRNA can be defined as being those molecules that induce 20%, 30%, 40%, 50%, 60%, or 70% silencing at 100 nM transfection conditions. Similarly, non-functional siRNA can be defined as being those molecules that silence gene expression by less than 70%, 60%, 50%, 40%, 30%, or less. Nonetheless, unless otherwise stated, the descriptions stated in the “Definitions” section of this text should be applied.
  • Functional attributes can be assigned to each of the key positions in the AISP of strong siRNA. The low 5′ (sense strand) AISP values of strong siRNAs may be necessary for determining which end of the molecule enters the RISC complex. In contrast, the high and low AISP values observed in the central regions of the molecule may be critical for siRNA-target mRNA interactions and product release, respectively.
  • If the AISP values described above accurately define the thermodynamic parameters of strong siRNA, it would be expected that similar patterns would be observed in strong siRNA isolated from nature. Natural siRNAs exist in a harsh, RNase-rich environment and it can be hypothesized that only those siRNA that exhibit heightened affinity for RISC (i.e., siRNA that exhibit an average internal stability profile similar to those observed in strong siRNA) would survive in an intracellular environment. This hypothesis was tested using GFP-specific siRNA isolated from N. benthamiana. Llave et al. (2002) Endogenous and Silencing-Associated Small RNAs in Plants, The Plant Cell 14, 1605-1619, introduced long double-stranded GFP-encoding RNA into plants and subsequently re-isolated GFP-specific siRNA from the tissues. The AISP of fifty-nine of these GFP-siRNA were determined, averaged, and subsequently plotted alongside the AISP profile obtained from the cyclophilin B/DBI/luciferase siRNA having >90% silencing properties (FIG. 6 b). Comparison of the two groups show that profiles are nearly identical. This finding validates the information provided by the internal stability profiles and demonstrates that: (1) the profile identified by analysis of the cyclophilin B/DBI/luciferase siRNAs are not gene specific; and (2) AISP values can be used to search for strong siRNAs in a variety of species.
  • Both chemical modifications and base-pair mismatches can be incorporated into siRNA to alter the duplex's AISP and functionality. For instance, introduction of mismatches at positions 1 or 2 of the sense strand destabilized the 5′ end of the sense strand and increases the functionality of the molecule (see Luc, FIG. 7). Similarly, addition of 2′-O-methyl groups to positions 1 and 2 of the sense strand can also alter the AISP and (as a result) increase both the functionality of the molecule and eliminate off-target effects that results from sense strand homology with the unrelated targets (FIG. 8).
  • Rationale for Criteria in a Biological Context
  • The fate of siRNA in the RNAi pathway may be described in 5 major steps: (1) duplex recognition and pre-RISC complex formation; (2) ATP-dependent duplex unwinding/strand selection and RISC activation; (3) mRNA target identification; (4) mRNA cleavage, and (5) product release (FIG. 1). Given the level of nucleic acid-protein interactions at each step, siRNA functionality is likely influenced by specific biophysical and molecular properties that promote efficient interactions within the context of the multi-component complexes. Indeed, the systematic analysis of the siRNA test set identified multiple factors that correlate well with functionality. When combined into a single algorithm, they proved to be very effective in selecting active siRNAs.
  • The factors described here may also be predictive of key functional associations important for each step in RNAi. For example, the potential formation of internal hairpin structures correlated negatively with siRNA functionality. Complementary strands with stable internal repeats are more likely to exist as stable hairpins thus decreasing the effective concentration of the functional duplex form. This suggests that the duplex is the preferred conformation for initial pre-RISC association. Indeed, although single complementary strands can induce gene silencing, the effective concentration required is at least two orders of magnitude higher than that of the duplex form.
  • siRNA-pre-RISC complex formation is followed by an ATP-dependent duplex unwinding step and “activation” of the RISC. The siRNA functionality was shown to correlate with overall low internal stability of the duplex and low internal stability of the 3′ sense end (or differential internal stability of the 3′ sense compare to the 5′ sense strand), which may reflect strand selection and entry into the RISC. Overall duplex stability and low internal stability at the 3′ end of the sense strand were also correlated with siRNA functionality. Interestingly, siRNAs with very high and very low overall stability profiles correlate strongly with non-functional duplexes. One interpretation is that high internal stability prevents efficient unwinding while very low stability reduces siRNA target affinity and subsequent mRNA cleavage by the RISC.
  • Several criteria describe base preferences at specific positions of the sense strand and are even more intriguing when considering their potential mechanistic roles in target recognition and mRNA cleavage. Base preferences for A at position 19 of the sense strand but not C, are particularly interesting because they reflect the same base preferences observed for naturally occurring miRNA precursors. That is, among the reported miRNA precursor sequences 75% contain a U at position 1 which corresponds to an A in position 19 of the sense strand of siRNAs, while G was under-represented in this same position for miRNA precursors. These observations support the hypothesis that both miRNA precursors and siRNA duplexes are processed by very similar if not identical protein machinery. The functional interpretation of the predominance of a U/A base pair is that it promotes flexibility at the 5′antisense ends of both siRNA duplexes and miRNA precursors and facilitates efficient unwinding and selective strand entrance into an activated RISC.
  • Among the criteria associated with base preferences that are likely to influence mRNA cleavage or possibly product release, the preference for U at position 10 of the sense strand exhibited the greatest impact, enhancing the probability of selecting an F80 sequence by 13.3%. Activated RISC preferentially cleaves target mRNA between nucleotides 10 and 11 relative to the 5′ end of the complementary targeting strand. Therefore, it may be that U, the preferred base for most endoribonucleases, at this position supports more efficient cleavage. Alternatively, a U/A bp between the targeting siRNA strand and its cognate target mRNA may create an optimal conformation for the RISC-associated “slicing” activity.
  • Post Algorithm Filters
  • According to another embodiment, the output of any one of the formulas previously listed can be filtered to remove or select for siRNAs containing undesirable or desirable motifs or properties, respectively. In one example, sequences identified by any of the formulas can be filtered to remove any and all sequences that induce toxicity or cellular stress. Introduction of an siRNA containing a toxic motif into a cell can induce cellular stress and/or cell death (apoptosis) which in turn can mislead researchers into associating a particular (e.g., nonessential) gene with, e.g., an essential function. Alternatively, sequences generated by any of the before mentioned formulas can be filtered to identify and retain duplexes that contain toxic motifs. Such duplexes may be valuable from a variety of perspectives including, for instance, uses as therapeutic molecules. A variety of toxic motifs exist and can exert their influence on the cell through RNAi and non-RNAi pathways. Examples of toxic motifs are explained more fully in commonly assigned U.S. Provisional Patent Application Ser. No. 60/538,874, entitled “Identification of Toxic Sequences,” filed Jan. 23, 2004. Briefly, toxic motifs include A/G UUU A/G/U, G/C AAA G/C, and GCCA, or a complement of any of the foregoing.
  • In another instance, sequences identified by any of the before mentioned formulas can be filtered to identify duplexes that contain motifs (or general properties) that provide serum stability or induce serum instability. In one envisioned application of siRNA as therapeutic molecules, duplexes targeting disease-associated genes will be introduced into patients intravenously. As the half-life of single and double stranded RNA in serum is short, post-algorithm filters designed to select molecules that contain motifs that enhance duplex stability in the presence of serum and/or (conversely) eliminate duplexes that contain motifs that destabilize siRNA in the presence of serum, would be beneficial.
  • In another instance, sequences identified by any of the before mentioned formulas can be filtered to identify duplexes that are hyperfunctional. Hyperfunctional sequences are defined as those sequences that (1) induce greater than 95% silencing of a specific target when they are transfected at subnanomolar concentrations (i.e., less than one nanomolar); and/or (2) induce functional (or better) levels of silencing for greater than 96 hours. Filters that identify hyperfunctional molecules can vary widely. In one example, the top ten, twenty, thirty, or forty siRNA can be assessed for the ability to silence a given target at, e.g., concentrations of 1 nM and 0.5 nM to identify hyperfunctional molecules.
  • Pooling
  • According to another embodiment, the present invention provides a pool of at least two siRNAs, preferably in the form of a kit or therapeutic reagent, wherein one strand of each of the siRNAs, the sense strand comprises a sequence that is substantially similar to a sequence within a target mRNA. The opposite strand, the antisense strand, will preferably comprise a sequence that is substantially complementary to that of the target mRNA. More preferably, one strand of each siRNA will comprise a sequence that is identical to a sequence that is contained in the target mRNA. Most preferably, each siRNA will be 19 base pairs in length, and one strand of each of the siRNAs will be 100% complementary to a portion of the target mRNA.
  • By increasing the number of siRNAs directed to a particular target using a pool or kit, one is able both to increase the likelihood that at least one siRNA with satisfactory functionality will be included, as well as to benefit from additive or synergistic effects. Further, when two or more siRNAs directed against a single gene do not have satisfactory levels of functionality alone, if combined, they may satisfactorily promote degradation of the target messenger RNA and successfully inhibit translation. By including multiple siRNAs in the system, not only is the probability of silencing increased, but the economics of operation are also improved when compared to adding different siRNAs sequentially. This effect is contrary to the conventional wisdom that the concurrent use of multiple siRNA will negatively impact gene silencing (e.g., Holen, T. et al. (2003) Similar behavior of single strand and double strand siRNAs suggests they act through a common RNAi pathway. NAR 31: 2401-21407).
  • In fact, when two siRNAs were pooled together, 54% of the pools of two siRNAs induced more than 95% gene silencing. Thus, a 2.5-fold increase in the percentage of functionality was achieved by randomly combining two siRNAs. Further, over 84% of pools containing two siRNAs induced more than 80% gene silencing.
  • More preferably, the kit is comprised of at least three siRNAs, wherein one strand of each siRNA comprises a sequence that is substantially similar to a sequence of the target mRNA and the other strand comprises a sequence that is substantially complementary to the region of the target mRNA. As with the kit that comprises at least two siRNAs, more preferably one strand will comprise a sequence that is identical to a sequence that is contained in the mRNA and another strand that is 100% complementary to a sequence that is contained in the mRNA. During experiments, when three siRNAs were combined together, 60% of the pools induced more than 95% gene silencing and 92% of the pools induced more than 80% gene silencing.
  • Further, even more preferably, the kit is comprised of at least four siRNAs, wherein one strand of each siRNA comprises a sequence that is substantially similar to a region of the sequence of the target mRNA, and the other strand comprises a sequence that is substantially complementary to the region of the target mRNA. As with the kit or pool that comprises at least two siRNAs, more preferably one strand of each of the siRNA duplexes will comprise a sequence that is identical to a sequence that is contained in the mRNA, and another strand that is 100% complementary to a sequence that is contained in the mRNA.
  • Additionally, kits and pools with at least five, at least six, and at least seven siRNAs may also be useful with the present invention. For example, pools of five siRNA induced 95% gene silencing with 77% probability and 80% silencing with 98.8% probability. Thus, pooling of siRNAs together can result in the creation of a target-specific silencing reagent with almost a 99% probability of being functional. The fact that such high levels of success are achievable using such pools of siRNA, enables one to dispense with costly and time-consuming target-specific validation procedures.
  • For this embodiment, as well as the other aforementioned embodiments, each of the siRNAs within a pool will preferably comprise 18-30 base pairs, more preferably 18-25 base pairs, and most preferably 19 base pairs. Within each siRNA, preferably at least 18 contiguous bases of the antisense strand will be 100% complementary to the target mRNA. More preferably, at least 19 contiguous bases of the antisense strand will be 100% complementary to the target mRNA. Additionally, there may be overhangs on either the sense strand or the antisense strand, and these overhangs may be at either the 5′ end or the 3′ end of either of the strands, for example there may be one or more overhangs of 1-6 bases. When overhangs are present, they are not included in the calculation of the number of base pairs. The two nucleotide 3′ overhangs mimic natural siRNAs and are commonly used but are not essential. Preferably, the overhangs should consist of two nucleotides, most often dTdT or UU at the 3′ end of the sense and antisense strand that are not complementary to the target sequence. The siRNAs may be produced by any method that is now known or that comes to be known for synthesizing double stranded RNA that one skilled in the art would appreciate would be useful in the present invention. Preferably, the siRNAs will be produced by Dharmacon's proprietary ACE® technology. However, other methods for synthesizing siRNAs are well known to persons skilled in the art and include, but are not limited to, any chemical synthesis of RNA oligonucleotides, ligation of shorter oligonucleotides, in vitro transcription of RNA oligonucleotides, the use of vectors for expression within cells, recombinant Dicer products and PCR products.
  • The siRNA duplexes within the aforementioned pools of siRNAs may correspond to overlapping sequences within a particular mRNA, or non-overlapping sequences of the mRNA. However, preferably they correspond to non-overlapping sequences. Further, each siRNA may be selected randomly, or one or more of the siRNA may be selected according to the criteria discussed above for maximizing the effectiveness of siRNA.
  • Included in the definition of siRNAs are siRNAs that contain substituted and/or labeled nucleotides that may, for example, be labeled by radioactivity, fluorescence or mass. The most common substitutions are at the 2′ position of the ribose sugar, where moieties such as H (hydrogen) F, NH3, OCH3 and other O— alkyl, alkenyl, alkynyl, and orthoesters, may be substituted, or in the phosphorous backbone, where sulfur, amines or hydrocarbons may be substituted for the bridging of non-bridging atoms in the phosphodiester bond. Examples of modified siRNAs are explained more fully in commonly assigned U.S. patent application Ser. No. 10/613,077, filed Jul. 1, 2003.
  • Additionally, as noted above, the cell type into which the siRNA is introduced may affect the ability of the siRNA to enter the cell; however, it does not appear to affect the ability of the siRNA to function once it enters the cell. Methods for introducing double-stranded RNA into various cell types are well known to persons skilled in the art.
  • As persons skilled in the art are aware, in certain species, the presence of proteins such as RdRP, the RNA-dependent RNA polymerase, may catalytically enhance the activity of the siRNA. For example, RdRP propagates the RNAi effect in C. elegans and other non-mammalian organisms. In fact, in organisms that contain these proteins, the siRNA may be inherited. Two other proteins that are well studied and known to be a part of the machinery are members of the Argonaute family and Dicer, as well as their homologues. There is also initial evidence that the RISC complex might be associated with the ribosome so the more efficiently translated mRNAs will be more susceptible to silencing than others.
  • Another very important factor in the efficacy of siRNA is mRNA localization. In general, only cytoplasmic mRNAs are considered to be accessible to RNAi to any appreciable degree. However, appropriately designed siRNAs, for example, siRNAs modified with internucleotide linkages or 2′-O-methyl groups, may be able to cause silencing by acting in the nucleus. Examples of these types of modifications are described in commonly assigned U.S. patent application Ser. Nos. 10/431,027 and 10/613,077.
  • As described above, even when one selects at least two siRNAs at random, the effectiveness of the two may be greater than one would predict based on the effectiveness of two individual siRNAs. This additive or synergistic effect is particularly noticeable as one increases to at least three siRNAs, and even more noticeable as one moves to at least four siRNAs. Surprisingly, the pooling of the non-functional and semi-functional siRNAs, particularly more than five siRNAs, can lead to a silencing mixture that is as effective if not more effective than any one particular functional siRNA.
  • Within the kits of the present invention, preferably each siRNA will be present in a concentration of between 0.001 and 200 μM, more preferably between 0.01 and 200 nM, and most preferably between 0.1 and 10 nM.
  • In addition to preferably comprising at least four or five siRNAs, the kits of the present invention will also preferably comprise a buffer to keep the siRNA duplex stable. Persons skilled in the art are aware of buffers suitable for keeping siRNA stable. For example, the buffer may be comprised of 100 mM KCl, 30 mM HEPES-pH 7.5, and 1 mM MgCl2. Alternatively, kits might contain complementary strands that contain any one of a number of chemical modifications (e.g., a 2′-O-ACE) that protect the agents from degradation by nucleases. In this instance, the user may (or may not) remove the modifying protective group (e.g., deprotect) before annealing the two complementary strands together.
  • By way of example, the kits may be organized such that pools of siRNA duplexes are provided on an array or microarray of wells or drops for a particular gene set or for unrelated genes. The array may, for example, be in 96 wells, 384 wells or 1284 wells arrayed in a plastic plate or on a glass slide using techniques now known or that come to be known to persons skilled in the art. Within an array, preferably there will be controls such as functional anti-lamin A/C, cyclophilin and two siRNA duplexes that are not specific to the gene of interest.
  • In order to ensure stability of the siRNA pools prior to usage, they may be retained in lyophilized form at minus twenty degrees (−20° C.) until they are ready for use. Prior to usage, they should be resuspended; however, even once resuspended, for example, in the aforementioned buffer, they should be kept at minus twenty degrees, (−20° C.) until used. The aforementioned buffer, prior to use, may be stored at approximately 4° C. or room temperature. Effective temperatures at which to conduct transfections are well known to persons skilled in the art and include for example, room temperature.
  • The kits may be applied either in vivo or in vitro. Preferably, the siRNA of the pools or kits is applied to a cell through transfection, employing standard transfection protocols. These methods are well known to persons skilled in the art and include the use of lipid-based carriers, electroporation, cationic carriers, and microinjection. Further, one could apply the present invention by synthesizing equivalent DNA sequences (either as two separate, complementary strands, or as hairpin molecules) instead of siRNA sequences and introducing them into cells through vectors. Once in the cells, the cloned DNA could be transcribed, thereby forcing the cells to generate the siRNA. Examples of vectors suitable for use with the present application include but are not limited to the standard transient expression vectors, adenoviruses, retroviruses, lentivirus-based vectors, as well as other traditional expression vectors. Any vector that has an adequate siRNA expression and procession module may be used. Furthermore, certain chemical modifications to siRNAs, including but not limited to conjugations to other molecules, may be used to facilitate delivery. For certain applications it may be preferable to deliver molecules without transfection by simply formulating in a physiological acceptable solution.
  • This embodiment may be used in connection with any of the aforementioned embodiments. Accordingly, the sequences within any pool may be selected by rational design.
  • Multigene Silencing
  • In addition to developing kits that contain multiple siRNA directed against a single gene, another embodiment includes the use of multiple siRNA targeting multiple genes. Multiple genes may be targeted through the use of high- or hyper-functional siRNA. High- or hyper-functional siRNA that exhibit increased potency, require lower concentrations to induce desired phenotypic (and thus therapeutic) effects. This circumvents RISC saturation. It therefore reasons that if lower concentrations of a single siRNA are needed for knockout or knockdown expression of one gene, then the remaining (uncomplexed) RISC will be free and available to interact with siRNA directed against two, three, four, or more, genes. Thus in this embodiment, the authors describe the use of highly functional or hyper-functional siRNA to knock out three separate genes. More preferably, such reagents could be combined to knockout four distinct genes. Even more preferably, highly functional or hyperfunctional siRNA could be used to knock out five distinct genes. Most preferably, siRNA of this type could be used to knockout or knockdown the expression of six or more genes.
  • Hyperfunctional siRNA
  • The term hyperfunctional siRNA (hf-siRNA) describes a subset of the siRNA population that induces RNAi in cells at low- or sub-nanomolar concentrations for extended periods of time. These traits, heightened potency and extended longevity of the RNAi phenotype, are highly attractive from a therapeutic standpoint. Agents having higher potency require lesser amounts of the molecule to achieve the desired physiological response, thus reducing the probability of side effects due to “off-target” interference. In addition to the potential therapeutic benefits associated with hyperfunctional siRNA, hf-siRNA are also desirable from an economic perspective. Hyperfunctional siRNA may cost less on a per-treatment basis, thus reducing overall expenditures to both the manufacturer and the consumer.
  • Identification of hyperfunctional siRNA involves multiple steps that are designed to examine an individual siRNA agent's concentration- and/or longevity-profiles. In one non-limiting example, a population of siRNA directed against a single gene are first analyzed using the previously described algorithm (Formula VIII). Individual siRNA are then introduced into a test cell line and assessed for the ability to degrade the target mRNA. It is important to note that when performing this step it is not necessary to test all of the siRNA. Instead, it is sufficient to test only those siRNA having the highest SMARTSCORES™, or siRNA ranking (i.e., SMARTSCORES™, or siRNA ranking >−10). Subsequently, the gene silencing data is plotted against the SMARTSCORES™, or siRNA rankings (see FIG. 9). siRNA that (1) induce a high degree of gene silencing (i.e., they induce greater than 80% gene knockdown) and (2) have superior SMARTSCORES™ (i.e., a SMARTSCORE™, or siRNA ranking, of >−10, suggesting a desirable average internal stability profile) are selected for further investigations designed to better understand the molecule's potency and longevity. In one, non-limiting study dedicated to understanding a molecule's potency, an siRNA is introduced into one (or more) cell types in increasingly diminishing concentrations (e.g., 3.0→0.3 nM). Subsequently, the level of gene silencing induced by each concentration is examined and siRNA that exhibit hyperfunctional potency (i.e., those that induce 80% silencing or greater at, e.g., picomolar concentrations) are identified. In a second study, the longevity profiles of siRNA having high (>−10) SMARTSCORES™, or siRNA rankings and greater than 80% silencing are examined. In one non-limiting example of how this is achieved, siRNA are introduced into a test cell line and the levels of RNAi are measured over an extended period of time (e.g., 24-168 hrs). siRNAs that exhibit strong RNA interference patterns (i.e., >80% interference) for periods of time greater than, e.g., 120 hours, are thus identified. Studies similar to those described above can be performed on any and all of the >106 siRNA included in this document to further define the most functional molecule for any given gene. Molecules possessing one or both properties (extended longevity and heightened potency) are labeled “hyperfunctional siRNA,” and earmarked as candidates for future therapeutic studies.
  • While the example(s) given above describe one means by which hyperfunctional siRNA can be isolated, neither the assays themselves nor the selection parameters used are rigid and can vary with each family of siRNA. Families of siRNA include siRNAs directed against a single gene, or directed against a related family of genes.
  • The highest quality siRNA achievable for any given gene may vary considerably. Thus, for example, in the case of one gene (gene X), rigorous studies such as those described above may enable the identification of an siRNA that, at picomolar concentrations, induces 99+% silencing for a period of 10 days. Yet identical studies of a second gene (gene Y) may yield an siRNA that at high nanomolar concentrations (e.g., 100 nM) induces only 75% silencing for a period of 2 days. Both molecules represent the very optimum siRNA for their respective gene targets and therefore are designated “hyperfunctional.” Yet due to a variety of factors including but not limited to target concentration, siRNA stability, cell type, off-target interference, and others, equivalent levels of potency and longevity are not achievable. Thus, for these reasons, the parameters described in the before mentioned assays can vary. While the initial screen selected siRNA that had SMARTSCORES™ above −10 and a gene silencing capability of greater than 80%, selections that have stronger (or weaker) parameters can be implemented. Similarly, in the subsequent studies designed to identify molecules with high potency and longevity, the desired cutoff criteria (i.e., the lowest concentration that induces a desirable level of interference, or the longest period of time that interference can be observed) can vary. The experimentation subsequent to application of the rational criteria of this application is significantly reduced where one is trying to obtain a suitable hyperfunctional siRNA for, for example, therapeutic use. When, for example, the additional experimentation of the type described herein is applied by one skilled in the art with this disclosure in hand, a hyperfunctional siRNA is readily identified.
  • The siRNA may be introduced into a cell by any method that is now known or that comes to be known and that from reading this disclosure, persons skilled in the art would determine would be useful in connection with the present invention in enabling siRNA to cross the cellular membrane. These methods include, but are not limited to, any manner of transfection, such as, for example, transfection employing DEAE-Dextran, calcium phosphate, cationic lipids/liposomes, micelles, manipulation of pressure, microinjection, electroporation, immunoporation, use of vectors such as viruses, plasmids, cosmids, bacteriophages, cell fusions, and coupling of the polynucleotides to specific conjugates or ligands such as antibodies, antigens, or receptors, passive introduction, adding moieties to the siRNA that facilitate its uptake, and the like.
  • Having described the invention with a degree of particularity, examples will now be provided. These examples are not intended to and should not be construed to limit the scope of the claims in any way.
    • EXAMPLES General Techniques and Nomenclatures
  • siRNA nomenclature. All siRNA duplexes are referred to by sense strand. The first nucleotide of the 5′-end of the sense strand is position 1, which corresponds to position 19 of the antisense strand for a 19-mer. In most cases, to compare results from different experiments, silencing was determined by measuring specific transcript mRNA levels or enzymatic activity associated with specific transcript levels, 24 hours post-transfection, with siRNA concentrations held constant at 100 nM. For all experiments, unless otherwise specified, transfection efficiency was ensured to be over 95%, and no detectable cellular toxicity was observed. The following system of nomenclature was used to compare and report siRNA-silencing functionality: “F” followed by the degree of minimal knockdown. For example, F50 signifies at least 50% knockdown, F80 means at least 80%, and so forth. For this study, all sub-F50 siRNAs were considered non-functional.
  • Cell culture and transfection. 96-well plates are coated with 50 μl of 50 mg/ml poly-L-lysine (Sigma) for 1 hr, and then washed 3× with distilled water before being dried for 20 min. HEK293 cells or HEK293Lucs or any other cell type of interest are released from their solid support by trypsinization, diluted to 3.5×105 cells/ml, followed by the addition of 100 μL of cells/well. Plates are then incubated overnight at 37° C., 5% CO2. Transfection procedures can vary widely depending on the cell type and transfection reagents. In one non-limiting example, a transfection mixture consisting of 2 mL Opti-MEM I (Gibco-BRL), 80 μl Lipofectamine 2000 (Invitrogen), 15 μL SUPERNasin at 20 U/R1 (Ambion), and 1.5 μl of reporter gene plasmid at 1 μg/μl is prepared in 5-ml polystyrene round bottom tubes. One hundred μl of transfection reagent is then combined with 100 μl of siRNAs in polystyrene deep-well titer plates (Beckman) and incubated for 20 to 30 min at room temperature. Five hundred and fifty microliters of Opti-MEM is then added to each well to bring the final siRNA concentration to 100 nM. Plates are then sealed with parafilm and mixed. Media is removed from HEK293 cells and replaced with 95 μl of transfection mixture. Cells are incubated overnight at 37° C., 5% CO2.
  • Quantification of gene knockdown. A variety of quantification procedures can be used to measure the level of silencing induced by siRNA or siRNA pools. In one non-limiting example: to measure mRNA levels 24 hrs post-transfection, QuantiGene branched-DNA (bDNA) kits (Bayer) (Wang, et al, Regulation of insulin preRNA splicing by glucose. Proc. Natl. Acad. Sci. USA 1997, 94:4360.) are used according to manufacturer instructions. To measure luciferase activity, media is removed from HEK293 cells 24 hrs post-transfection, and 50 μl of Steady-GLO reagent (Promega) is added. After 5 minutes, plates are analyzed on a plate reader.
    • Example I Sequences Used to Develop the Algorithm
  • Anti-Firefly and anti-Cyclophilin siRNAs panels (FIG. 5 a, b) sorted according to using Formula VIII predicted values. All siRNAs scoring more than 0 (formula VIII) and more then 20 (formula IX) are fully functional. All ninety sequences for each gene (and DBI) appear below in Table III.
    TABLE III
    Cyclo
    1 SEQ. ID 0032 GUUCCAAAAACAGUGGAUA
    Cyclo
    2 SEQ. ID 0033 UCCAAAAACAGUGGAUAAU
    Cyclo
    3 SEQ. ID 0034 CAAAAACAGUGGAUAAUUU
    Cyclo
    4 SEQ. ID 0035 AAAACAGUGGAUAAUUUUG
    Cyclo
    5 SEQ. ID 0036 AACAGUGGAUAAUUUUGUG
    Cyclo
    6 SEQ. ID 0037 CAGUGGAUAAUUUUGUGGC
    Cyclo
    7 SEQ. ID 0038 GUGGAUAAUUUUGUGGCCU
    Cyclo
    8 SEQ. ID 0039 GGAUAAUUUUGUGGCCUUA
    Cyclo
    9 SEQ. ID 0040 AUAAUUUUGUGGCCUUAGC
    Cyclo
    10 SEQ. ID 0041 AAUUUUGUGGCCUUAGCUA
    Cyclo
    11 SEQ. ID 0042 UUUUGUGGCCUUAGCUACA
    Cyclo
    12 SEQ. ID 0043 UUGUGGCCUUAGCUACAGG
    Cyclo
    13 SEQ. ID 0044 GUGGCCUUAGCUACAGGAG
    Cyclo
    14 SEQ. ID 0045 GGCCUUAGCUACAGGAGAG
    Cyclo
    15 SEQ. ID 0046 CCUUAGCUACAGGAGAGAA
    Cyclo
    16 SEQ. ID 0047 UUAGCUACAGGAGAGAAAG
    Cyclo
    17 SEQ. ID 0048 AGCUACAGGAGAGAAAGGA
    Cyclo
    18 SEQ. ID 0049 CUACAGGAGAGAAAGGAUU
    Cyclo
    19 SEQ. ID 0050 ACAGGAGAGAAAGGAUUUG
    Cyclo
    20 SEQ. ID 0051 AGGAGAGAAAGGAUUUGGC
    Cyclo
    21 SEQ. ID 0052 GAGAGAAAGGAUUUGGCUA
    Cyclo
    22 SEQ. ID 0053 GAGAAAGGAUUUGGCUACA
    Cyclo
    23 SEQ. ID 0054 GAAAGGAUUUGGCUACAAA
    Cyclo 24 SEQ. ID 0055 AAGGAUUUGGCUACAAAAA
    Cyclo
    25 SEQ. ID 0056 GGAUUUGGCUACAAAAACA
    Cyclo
    26 SEQ. ID 0057 AUUUGGCUACAAAAACAGC
    Cyclo 27 SEQ. ID 0058 UUGGCUACAAAAACAGCAA
    Cyclo
    28 SEQ. ID 0059 GGCUACAAAAACAGCAAAU
    Cyclo 29 SEQ. ID 0060 CUACAAAAACAGCAAAUUC
    Cyclo
    30 SEQ. ID 0061 ACAAAAACAGCAAAUUCCA
    Cyclo
    31 SEQ. ID 0062 AAAAACAGCAAAUUCCAUC
    Cyclo
    32 SEQ. ID 0063 AAACAGCAAAUUCCAUCGU
    Cyclo 33 SEQ. ID 0064 ACAGCAAAUUCCAUCGUGU
    Cyclo 34 SEQ. ID 0065 AGCAAAUUCCAUCGUGUAA
    Cyclo 35 SEQ. ID 0066 CAAAUUCCAUCGUGUAAUC
    Cyclo
    36 SEQ. ID 0067 AAUUCCAUCGUGUAAUCAA
    Cyclo 37 SEQ. ID 0068 UUCCAUCGUGUAAUCAAGG
    Cyclo 38 SEQ. ID 0069 CCAUCGUGUAAUCAAGGAC
    Cyclo 39 SEQ. ID 0070 AUCGUGUAAUCAAGGACUU
    Cyclo
    40 SEQ. ID 0071 CGUGUAAUCAAGGACUUCA
    Cyclo 41 SEQ. ID 0072 UGUAAUCAAGGACUUCAUG
    Cyclo
    42 SEQ. ID 0073 UAAUCAAGGACUUCAUGAU
    Cyclo 43 SEQ. ID 0074 AUCAAGGACUUCAUGAUCC
    Cyclo 44 SEQ. ID 0075 CAAGGACUUCAUGAUCCAG
    Cyclo 45 SEQ. ID 0076 AGGACUUCAUGAUCCAGGG
    Cyclo 46 SEQ. ID 0077 GACUUCAUGAUCCAGGGCG
    Cyclo
    47 SEQ. ID 0078 CUUCAUGAUCCAGGGCGGA
    Cyclo 48 SEQ. ID 0079 UCAUGAUCCAGGGCGGAGA
    Cyclo 49 SEQ. ID 0080 AUGAUCCAGGGCGGAGACU
    Cyclo
    50 SEQ. ID 0081 GAUCCAGGGCGGAGACUUC
    Cyclo 51 SEQ. ID 0082 UCCAGGGCGGAGACUUCAC
    Cyclo
    52 SEQ. ID 0083 CAGGGCGGAGACUUCACCA
    Cyclo 53 SEQ. ID 0084 GGGCGGAGACUUCACCAGG
    Cyclo 54 SEQ. ID 0085 GCGGAGACUUCACCAGGGG
    Cyclo 55 SEQ. ID 0086 GGAGACUUCACCAGGGGAG
    Cyclo
    56 SEQ. ID 0087 AGACUUCACCAGGGGAGAU
    Cyclo
    57 SEQ. ID 0088 ACUUCACCAGGGGAGAUGG
    Cyclo
    58 SEQ. ID 0089 UUCACCAGGGGAGAUGGCA
    Cyclo 59 SEQ. ID 0090 CACCAGGGGAGAUGGCACA
    Cyclo
    60 SEQ. ID 0091 CCAGGGGAGAUGGCACAGG
    Cyclo 61 SEQ. ID 0092 AGGGGAGAUGGCACAGGAG
    Cyclo 62 SEQ. ID 0093 GGGAGAUGGCACAGGAGGA
    Cyclo
    63 SEQ. ID 0094 GAGAUGGCACAGGAGGAAA
    Cyclo 64 SEQ. ID 0095 GAUGGCACAGGAGGAAAGA
    Cyclo 65 SEQ. ID 0096 UGGCACAGGAGGAAAGAGC
    Cyclo
    66 SEQ. ID 0097 GCACAGGAGGAAAGAGCAU
    Cyclo 67 SEQ. ID 0098 ACAGGAGGAAAGAGCAUCU
    Cyclo
    68 SEQ. ID 0099 AGGAGGAAAGAGCAUCUAC
    Cyclo 69 SEQ. ID 0100 GAGGAAAGAGCAUCUACGG
    Cyclo
    70 SEQ. ID 0101 GGAAAGAGCAUCUACGGUG
    Cyclo 71 SEQ. ID 0102 AAAGAGCAUCUACGGUGAG
    Cyclo
    72 SEQ. ID 0103 AGAGCAUCUACGGUGAGCG
    Cyclo 73 SEQ. ID 0104 AGCAUCUACGGUGAGCGCU
    Cyclo 74 SEQ. ID 0105 CAUCUACGGUGAGCGCUUC
    Cyclo 75 SEQ. ID 0106 UCUACGGUGAGCGCUUCCC
    Cyclo
    76 SEQ. ID 0107 UACGGUGAGCGCUUCCCCG
    Cyclo 77 SEQ. ID 0108 CGGUGAGCGCUUCCCCGAU
    Cyclo
    78 SEQ. ID 0109 GUGAGCGCUUCCCCGAUGA
    Cyclo
    79 SEQ. ID 0110 GAGCGCUUCCCCGAUGAGA
    Cyclo
    80 SEQ. ID 0111 GCGCUUCCCCGAUGAGAAC
    Cyclo
    81 SEQ. ID 0112 GCUUCCCCGAUGAGAACUU
    Cyclo
    82 SEQ. ID 0113 UUCCCCGAUGAGAACUUCA
    Cyclo 83 SEQ. ID 0114 CCCCGAUGAGAACUUCAAA
    Cyclo 84 SEQ. ID 0115 CCGAUGAGAACUUCAAACU
    Cyclo
    85 SEQ. ID 0116 GAUGAGAACUUCAAACUGA
    Cyclo 86 SEQ. ID 0117 UGAGAACUUCAAACUGAAG
    Cyclo
    87 SEQ. ID 0118 AGAACUUCAAACUGAAGCA
    Cyclo
    88 SEQ. ID 0119 AACUUCAAACUGAAGCACU
    Cyclo
    89 SEQ. ID 0120 CUUCAAACUGAAGCACUAC
    Cyclo
    90 SEQ. ID 0121 UCAAACUGAAGCACUACGG
    DB
    1 SEQ. ID 0122 ACGGGCAAGGCCAAGUGGG
    DB
    2 SEQ. ID 0123 CGGGCAAGGCCAAGUGGGA
    DB
    3 SEQ. ID 0124 GGGCAAGGCCAAGUGGGAU
    DB
    4 SEQ. ID 0125 GGCAAGGCCAAGUGGGAUG
    DB
    5 SEQ. ID 0126 GCAAGGCCAAGUGGGAUGC
    DB
    6 SEQ. ID 0127 CAAGGCCAAGUGGGAUGCC
    DB
    7 SEQ. ID 0128 AAGGCCAAGUGGGAUGCCU
    DB
    8 SEQ. ID 0129 AGGCCAAGUGGGAUGCCUG
    DB
    9 SEQ. ID 0130 GGCCAAGUGGGAUGCCUGG
    DB
    10 SEQ. ID 0131 GCCAAGUGGGAUGCCUGGA
    DB
    11 SEQ. ID 0132 CCAAGUGGGAUGCCUGGAA
    DB
    12 SEQ. ID 0133 CAAGUGGGAUGCCUGGAAU
    DB
    13 SEQ. ID 0134 AAGUGGGAUGCCUGGAAUG
    DB
    14 SEQ. ID 0135 AGUGGGAUGCCUGGAAUGA
    DB
    15 SEQ. ID 0136 GUGGGAUGCCUGGAAUGAG
    DB
    16 SEQ. ID 0137 UGGGAUGCCUGGAAUGAGC
    DB
    17 SEQ. ID 0138 GGGAUGCCUGGAAUGAGCU
    DB
    18 SEQ. ID 0139 GGAUGCCUGGAAUGAGCUG
    DB
    19 SEQ. ID 0140 GAUGCCUGGAAUGAGCUGA
    DB
    20 SEQ. ID 0141 AUGCCUGGAAUGAGCUGAA
    DB
    21 SEQ. ID 0142 UGCCUGGAAUGAGCUGAAA
    DB
    22 SEQ. ID 0143 GCCUGGAAUGAGCUGAAAG
    DB
    23 SEQ. ID 0144 CCUGGAAUGAGCUGAAAGG
    DB 24 SEQ. ID 0145 CUGGAAUGAGCUGAAAGGG
    DB
    25 SEQ. ID 0146 UGGAAUGAGCUGAAAGGGA
    DB
    26 SEQ. ID 0147 GGAAUGAGCUGAAAGGGAC
    DB 27 SEQ. ID 0148 GAAUGAGCUGAAAGGGACU
    DB
    28 SEQ. ID 0149 AAUGAGCUGAAAGGGACUU
    DB 29 SEQ. ID 0150 AUGAGCUGAAAGGGACUUC
    DB
    30 SEQ. ID 0151 UGAGCUGAAAGGGACUUCC
    DB
    31 SEQ. ID 0152 GAGCUGAAAGGGACUUCCA
    DB
    32 SEQ. ID 0153 AGCUGAAAGGGACUUCCAA
    DB 33 SEQ. ID 0154 GCUGAAAGGGACUUCCAAG
    DB 34 SEQ. ID 0155 CUGAAAGGGACUUCCAAGG
    DB 35 SEQ. ID 0156 UGAAAGGGACUUCCAAGGA
    DB
    36 SEQ. ID 0157 GAAAGGGACUUCCAAGGAA
    DB 37 SEQ. ID 0158 AAAGGGACUUCCAAGGAAG
    DB 38 SEQ. ID 0159 AAGGGACUUCCAAGGAAGA
    DB 39 SEQ. ID 0160 AGGGACUUCCAAGGAAGAU
    DB
    40 SEQ. ID 0161 GGGACUUCCAAGGAAGAUG
    DB 41 SEQ. ID 0162 GGACUUCCAAGGAAGAUGC
    DB
    42 SEQ. ID 0163 GACUUCCAAGGAAGAUGCC
    DB 43 SEQ. ID 0164 ACUUCCAAGGAAGAUGCCA
    DB 44 SEQ. ID 0165 CUUCCAAGGAAGAUGCCAU
    DB 45 SEQ. ID 0166 UUCCAAGGAAGAUGCCAUG
    DB 46 SEQ. ID 0167 UCCAAGGAAGAUGCCAUGA
    DB
    47 SEQ. ID 0168 CCAAGGAAGAUGCCAUGAA
    DB 48 SEQ. ID 0169 CAAGGAAGAUGCCAUGAAA
    DB 49 SEQ. ID 0170 AAGGAAGAUGCCAUGAAAG
    DB
    50 SEQ. ID 0171 AGGAAGAUGCCAUGAAAGC
    DB 51 SEQ. ID 0172 CGAAGAUGCCAUGAAAGCU
    DB
    52 SEQ. ID 0173 GAAGAUGCCAUGAAAGCUU
    DB 53 SEQ. ID 0174 AAGAUGCCAUGAAAGCUUA
    DB 54 SEQ. ID 0175 AGAUGCCAUGAAAGCUUAC
    DB 55 SEQ. ID 0176 GAUGCCAUGAAAGCUUACA
    DB
    56 SEQ. ID 0177 AUGCCAUGAAAGCUUACAU
    DB
    57 SEQ. ID 0178 UGCCAUGAAAGCUUACAUC
    DB
    58 SEQ. ID 0179 GCCAUGAAAGCUUACAUCA
    DB 59 SEQ. ID 0180 CCAUGAAAGCUUACAUCAA
    DB
    60 SEQ. ID 0181 CAUGAAAGCUUACAUCAAC
    DB 61 SEQ. ID 0182 AUGAAAGCUUACAUCAACA
    DB 62 SEQ. ID 0183 UGAAAGCUUACAUCAACAA
    DB
    63 SEQ. ID 0184 GAAAGCUUACAUCAACAAA
    DB 64 SEQ. ID 0185 AAAGCUUACAUCAACAAAG
    DB 65 SEQ. ID 0186 AAGCUUACAUCAACAAAGU
    DB
    66 SEQ. ID 0187 AGCUUACAUCAACAAAGUA
    DB 67 SEQ. ID 0188 GCUUACAUCAACAAAGUAG
    DB
    68 SEQ. ID 0189 CUUACAUCAACAAAGUAGA
    DB 69 SEQ. ID 0190 UUACAUCAACAAAGUAGAA
    DB
    70 SEQ. ID 0191 UACAUCAACAAAGUAGAAG
    DB 71 SEQ. ID 0192 ACAUCAACAAAGUAGAAGA
    DB
    72 SEQ. ID 0193 CAUCAACAAAGUAGAAGAG
    DB 73 SEQ. ID 0194 AUCAACAAAGUAGAAGAGC
    DB 74 SEQ. ID 0195 UCAACAAAGUAGAAGAGCU
    DB 75 SEQ. ID 0196 CAACAAAGUAGAAGAGCUA
    DB
    76 SEQ. ID 0197 AACAAAGUAGAAGAGCUAA
    DB 77 SEQ. ID 0198 ACAAAGUAGAAGAGCUAAA
    DB
    78 SEQ. ID 0199 CAAAGUAGAAGAGCUAAAG
    DB
    79 SEQ. ID 0200 AAAGUAGAAGAGCUAAAGA
    DB
    80 SEQ. ID 0201 AAGUAGAAGAGCUAAAGAA
    DB
    81 SEQ. ID 0202 AGUAGAAGAGCUAAAGAAA
    DB
    82 SEQ. ID 0203 GUAGAAGAGCUAAAGAAAA
    DB 83 SEQ. ID 0204 UAGAAGAGCUAAAGAAAAA
    DB 84 SEQ. ID 0205 AGAAGAGCUAAAGAAAAAA
    DB
    85 SEQ. ID 0206 GAAGAGCUAAAGAAAAAAU
    DB 86 SEQ. ID 0207 AAGAGCUAAAGAAAAAAUA
    DB
    87 SEQ. ID 0208 AGAGCUAAAGAAAAAAUAC
    DB
    88 SEQ. ID 0209 GAGCUAAAGAAAAAAUACG
    DB
    89 SEQ. ID 0210 AGCUAAAGAAAAAAUACGG
    DB
    90 SEQ. ID 0211 GCUAAAGAAAAAAUACGGG
    Luc
    1 SEQ. ID 0212 AUCCUCAUAAAGGCCAAGA
    Luc
    2 SEQ. ID 0213 AGAUCCUCAUAAAGGCCAA
    Luc
    3 SEQ. ID 0214 AGAGAUCCUCAUAAAGGCC
    Luc
    4 SEQ. ID 0215 AGAGAGAUCCUCAUAAAGG
    Luc
    5 SEQ. ID 0216 UCAGAGAGAUCCUCAUAAA
    Luc
    6 SEQ. ID 0217 AAUCAGAGAGAUCCUCAUA
    Luc
    7 SEQ. ID 0218 AAAAUCAGAGAGAUCCUCA
    Luc
    8 SEQ. ID 0219 GAAAAAUCAGAGAGAUCCU
    Luc
    9 SEQ. ID 0220 AAGAAAAAUCAGAGAGAUC
    Luc
    10 SEQ. ID 0221 GCAAGAAAAAUCAGAGAGA
    Luc
    11 SEQ. ID 0222 ACGCAAGAAAAAUCAGAGA
    Luc
    12 SEQ. ID 0223 CGACGCAAGAAAAAUCAGA
    Luc
    13 SEQ. ID 0224 CUCGACGCAAGAAAAAUCA
    Luc
    14 SEQ. ID 0225 AACUCGACGCAAGAAAAAU
    Luc
    15 SEQ. ID 0226 AAAACUCGACGCAAGAAAA
    Luc
    16 SEQ. ID 0227 GGAAAACUCGACGCAAGAA
    Luc
    17 SEQ. ID 0228 CCGGAAAACUCGACGCAAG
    Luc
    18 SEQ. ID 0229 UACCGGAAAACUCGACGCA
    Luc
    19 SEQ. ID 0230 CUUACCGGAAAACUCGACG
    Luc
    20 SEQ. ID 0231 GUCUUACCGGAAAACUCGA
    Luc
    21 SEQ. ID 0232 AGGUCUUACCGGAAAACUC
    Luc
    22 SEQ. ID 0233 AAAGGUCUUACCGGAAAAC
    Luc
    23 SEQ. ID 0234 CGAAAGGUCUUACCGGAAA
    Luc 24 SEQ. ID 0235 ACCGAAAGGUCUUACCGGA
    Luc
    25 SEQ. ID 0236 GUACCGAAAGGUCUUACCG
    Luc
    26 SEQ. ID 0237 AAGUACCGAAAGGUCUUAC
    Luc 27 SEQ. ID 0238 CGAAGUACCGAAAGGUCUU
    Luc
    28 SEQ. ID 0239 GACGAAGUACCGAAAGGUC
    Luc 29 SEQ. ID 0240 UGGACGAAGUACCGAAAGG
    Luc
    30 SEQ. ID 0241 UGUGGACGAAGUACCGAAA
    Luc
    31 SEQ. ID 0242 UUUGUGGACGAAGUACCGA
    Luc
    32 SEQ. ID 0243 UGUUUGUGGACGAAGUACC
    Luc 33 SEQ. ID 0244 UGUGUUUGUGGACGAAGUA
    Luc 34 SEQ. ID 0245 GUUGUGUUUGUGGACGAAG
    Luc 35 SEQ. ID 0246 GAGUUGUGUUUGUGGACGA
    Luc
    36 SEQ. ID 0247 AGGAGUUGUGUUUGUGGAC
    Luc 37 SEQ. ID 0248 GGAGGAGUUGUGUUUGUGG
    Luc 38 SEQ. ID 0249 GCGGAGGAGUUGUGUUUGU
    Luc 39 SEQ. ID 0250 GCGCGGAGGAGUUGUGUUU
    Luc
    40 SEQ. ID 0251 UUGCGCGGAGGAGUUGUGU
    Luc 41 SEQ. ID 0252 AGUUGCGCGGAGGAGUUGU
    Luc
    42 SEQ. ID 0253 AAAGUUGCGCGGAGGAGUU
    Luc 43 SEQ. ID 0254 AAAAAGUUGCGCGGAGGAG
    Luc 44 SEQ. ID 0255 CGAAAAAGUUGCGCGGAGG
    Luc 45 SEQ. ID 0256 CGCGAAAAAGUUGCGCGGA
    Luc 46 SEQ. ID 0257 ACCGCGAAAAAGUUGCGCG
    Luc
    47 SEQ. ID 0258 CAACCGCGAAAAAGUUGCG
    Luc 48 SEQ. ID 0259 AACAACCGCGAAAAAGUUG
    Luc 49 SEQ. ID 0260 GUAACAACCGCGAAAAAGU
    Luc
    50 SEQ. ID 0261 AAGUAACAACCGCGAAAAA
    Luc 51 SEQ. ID 0262 UCAAGUAACAACCGCGAAA
    Luc
    52 SEQ. ID 0263 AGUCAAGUAACAACCGCGA
    Luc 53 SEQ. ID 0264 CCAGUCAAGUAACAACCGC
    Luc 54 SEQ. ID 0265 CGCCAGUCAAGUAACAACC
    Luc 55 SEQ. ID 0266 GUCGCCAGUCAAGUAACAA
    Luc
    56 SEQ. ID 0267 ACGUCGCCAGUCAAGUAAC
    Luc
    57 SEQ. ID 0268 UUACGUCGCCAGUCAAGUA
    Luc
    58 SEQ. ID 0269 GAUUACGUCGCCAGUCAAG
    Luc 59 SEQ. ID 0270 UGGAUUACGUCGCCAGUCA
    Luc
    60 SEQ. ID 0271 CGUGGAUUACGUCGCCAGU
    Luc 61 SEQ. ID 0272 AUCGUGGAUUACGUCGCCA
    Luc 62 SEQ. ID 0273 AGAUCGUGGAUUACGUCGC
    Luc
    63 SEQ. ID 0274 AGAGAUCGUGGAUUACGUC
    Luc 64 SEQ. ID 0275 AAAGAGAUCGUGGAUUACG
    Luc 65 SEQ. ID 0276 AAAAAGAGAUCGUGGAUUA
    Luc
    66 SEQ. ID 0277 GGAAAAAGAGAUCGUGGAU
    Luc 67 SEQ. ID 0278 ACGGAAAAAGAGAUCGUGG
    Luc
    68 SEQ. ID 0279 UGACGGAAAAAGAGAUCGU
    Luc 69 SEQ. ID 0280 GAUGACGGAAAAAGAGAUC
    Luc
    70 SEQ. ID 0281 ACGAUGACGGAAAAAGAGA
    Luc 71 SEQ. ID 0282 AGACGAUGACGGAAAAAGA
    Luc
    72 SEQ. ID 0283 AAAGACGAUGACGGAAAAA
    Luc 73 SEQ. ID 0284 GGAAAGACGAUGACGGAAA
    Luc 74 SEQ. ID 0285 ACGGAAAGACGAUGACGGA
    Luc 75 SEQ. ID 0286 GCACGGAAAGACGAUGACG
    Luc
    76 SEQ. ID 0287 GAGCACGGAAAGACGAUGA
    Luc 77 SEQ. ID 0288 UGGAGCACGGAAAGACGAU
    Luc
    78 SEQ. ID 0289 UUUGGAGCACGGAAAGACG
    Luc
    79 SEQ. ID 0290 GUUUUGGAGCACGGAAAGA
    Luc
    80 SEQ. ID 0291 UUGUUUUGGAGCACGGAAA
    Luc
    81 SEQ. ID 0292 UGUUGUUUUGGAGCACGGA
    Luc
    82 SEQ. ID 0293 GUUGUUGUUUUGGAGCACG
    Luc 83 SEQ. ID 0294 CCGUUGUUGUUUUGGAGCA
    Luc 84 SEQ. ID 0295 CGCCGUUGUUGUUUUGGAG
    Luc
    85 SEQ. ID 0296 GCCGCCGUUGUUGUUUUGG
    Luc 86 SEQ. ID 0297 CCGCCGCCGUUGUUGUUUU
    Luc
    87 SEQ. ID 0298 UCCCGCCGCCGUUGUUGUU
    Luc
    88 SEQ. ID 0299 CUUCCCGCCGCCGUUGUUG
    Luc
    89 SEQ. ID 0300 AACUUCCCGCCGCCGUUGU
    Luc
    90 SEQ. ID 0301 UGAACUUCCCGCCGCCGUU
    • Example II Validation of the Algorithm Using DBI, Luciferase, PLK, EGFR, and SEAP
  • The algorithm (Formula VIII) identified siRNAs for five genes, human DBI, firefly luciferase (fLuc), renilla luciferase (rLuc), human PLK, and human secreted alkaline phosphatase (SEAP). Four individual siRNAs were selected on the basis of their SMARTSCORES™ derived by analysis of their sequence using Formula VIII (all of the siRNAs would be selected with Formula IX as well) and analyzed for their ability to silence their targets' expression. In addition to the scoring, a BLAST search was conducted for each siRNA. To minimize the potential for off-target silencing effects, only those target sequences with more than three mismatches against un-related sequences were selected. Semizarov, et al. (2003)
  • Specificity of short interfering RNA determined through gene expression signatures, Proc. Natl. Acad. Sci. USA, 100:6347. These duplexes were analyzed individually and in pools of 4 and compared with several siRNAs that were randomly selected. The functionality was measured as a percentage of targeted gene knockdown as compared to controls. All siRNAs were transfected as described by the methods above at 100 nM concentration into HEK293 using Lipofectamine 2000. The level of the targeted gene expression was evaluated by B-DNA as described above and normalized to the non-specific control. FIG. 10 shows that the siRNAs selected by the algorithm disclosed herein were significantly more potent than randomly selected siRNAs. The algorithm increased the chances of identifying an F50 siRNA from 48% to 91%, and an F80 siRNA from 13% to 57%. In addition, pools of SMART siRNA silence the selected target better than randomly selected pools (see FIG. 10F).
    • Example III Validation of the Algorithm Using Genes Involved in Clathrin-Dependent Endocytosis
  • Components of clathrin-mediated endocytosis pathway are key to modulating intracellular signaling and play important roles in disease. Chromosomal rearrangements that result in fusion transcripts between the Mixed-Lineage Leukemia gene (MLL) and CALM (clathrin assembly lymphoid myeloid leukemia gene) are believed to play a role in leukemogenesis. Similarly, disruptions in Rab7 and Rab9, as well as HIP1 (Huntingtin-interacting protein), genes that are believed to be involved in endocytosis, are potentially responsible for ailments resulting in lipid storage, and neuronal diseases, respectively. For these reasons, siRNA directed against clathrin and other genes involved in the clathrin-mediated endocytotic pathway are potentially important research and therapeutic tools.
  • siRNAs directed against genes involved in the clathrin-mediated endocytosis pathways were selected using Formula VIII. The targeted genes were clathrin heavy chain (CHC, accession # NM 004859), clathrin light chain A (CLCa, NM—001833), clathrin light chain B (CLCb, NM—001834), CALM (U45976), β2 subunit of AP-2 (β2, NM—001282), Eps15 (NM—001981), Eps15R(NM—021235), dynamin II (DYNII, NM—004945), Rab5a (BC001267), Rab5b (NM-002868), Rab5c (AF141304), and EEA.1 (XM—018197).
  • For each gene, four siRNAs duplexes with the highest scores were selected and a BLAST search was conducted for each of them using the Human EST database. In order to minimize the potential for off-target silencing effects, only those sequences with more than three mismatches against un-related sequences were used. All duplexes were synthesized at Dharmacon, Inc. as 21-mers with 3′-UU overhangs using a modified method of 2′-ACE chemistry, Scaringe (2000) Advanced 5′-silyl-2′-orthoester approach to RNA oligonucleotide synthesis, Methods Enzymol. 317:3, and the antisense strand was chemically phosphorylated to insure maximized activity.
  • HeLa cells were grown in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum, antibiotics and glutamine. siRNA duplexes were resuspended in 1× siRNA Universal buffer (Dharmacon, Inc.) to 20CM prior to transfection. HeLa cells in 12-well plates were transfected twice with 4 μl of 20 μM siRNA duplex in 3 μl Lipofectamine 2000 reagent (Invitrogen, Carlsbad, Calif., USA) at 24-hour intervals. For the transfections in which 2 or 3 siRNA duplexes were included, the amount of each duplex was decreased, so that the total amount was the same as in transfections with single siRNAs. Cells were plated into normal culture medium 12 hours prior to experiments, and protein levels were measured 2 or 4 days after the first transfection.
  • Equal amounts of lysates were resolved by electrophoresis, blotted, and stained with the antibody specific to targeted protein, as well as antibodies specific to unrelated proteins, PP1 phosphatase and Tsg101 (not shown). The cells were lysed in Triton X-100/glycerol solubilization buffer as described previously. Tebar, Bohlander, & Sorkin (1999) Clathrin Assembly Lymphoid Myeloid Leukemia (CALM) Protein: Localization in Endocytic-coated Pits, Interactions with Clathrin, and the Impact of Overexpression on Clathrin-mediated Traffic, Mol. Biol. Cell, 10:2687. Cell lysates were electrophoresed, transferred to nitrocellulose membranes, and Western blotting was performed with several antibodies followed by detection using enhanced chemiluminescence system (Pierce, Inc). Several x-ray films were analyzed to determine the linear range of the chemiluminescence signals, and the quantifications were performed using densitometry and AlphaImager v5.5 software (Alpha Innotech Corporation). In experiments with Eps15R-targeted siRNAs, cell lysates were subjected to immunoprecipitation with Ab860, and Eps15R was detected in immunoprecipitates by Western blotting as described above.
  • The antibodies to assess the levels of each protein by Western blot were obtained from the following sources: monoclonal antibody to clathrin heavy chain (TD. 1) was obtained from American Type Culture Collection (Rockville, Md., USA); polyclonal antibody to dynamin II was obtained from Affinity Bioreagents, Inc. (Golden, Colo., USA); monoclonal antibodies to EEA. 1 and Rab5a were purchased from BD Transduction Laboratories (Los Angeles, Calif., USA); the monoclonal antibody to Tsg101 was purchased from Santa Cruz Biotechnology, Inc. (Santa Cruz, Calif., USA); the monoclonal antibody to GFP was from ZYMED Laboratories Inc. (South San Francisco, Calif., USA); the rabbit polyclonal antibodies Ab32 specific to α-adaptins and Ab20 to CALM were described previously (Sorkin et al. (1995) Stoichiometric Interaction of the Epidermal Growth Factor Receptor with the Clathrin-associated Protein Complex AP-2, J. Biol. Chem., 270:619), the polyclonal antibodies to clathrin light chains A and B were kindly provided by Dr. F. Brodsky (UCSF); monoclonal antibodies to PP1 (BD Transduction Laboratories) and α-Actinin (Chemicon) were kindly provided by Dr. M. Dell'Acqua (University of Colorado); Eps15 Ab577 and Eps15R Ab860 were kindly provided by Dr. P. P. Di Fiore (European Cancer Institute).
  • FIG. 11 demonstrates the in vivo functionality of 48 individual siRNAs, selected using Formula VIII (most of them will meet the criteria incorporated by Formula IX as well) targeting 12 genes. Various cell lines were transfected with siRNA duplexes (Dup1-4) or pools of siRNA duplexes (Pool), and the cells were lysed 3 days after transfection with the exception of CALM (2 days) and β2 (4 days).
  • Note a β1-adaptin band (part of AP-1 Golgi adaptor complex) that runs slightly slower than β2 adaptin. CALM has two splice variants, 66 and 72 kD. The full-length Eps15R (a doublet of ˜130 kD) and several truncated spliced forms of ˜100 kD and ˜70 kD were detected in Eps15R immunoprecipitates (shown by arrows). The cells were lysed 3 days after transfection. Equal amounts of lysates were resolved by electrophoresis and blotted with the antibody specific to a targeted protein (GFP antibody for YFP fusion proteins) and the antibody specific to unrelated proteins PP1 phosphatase or α-actinin, and TSG1101. The amount of protein in each specific band was normalized to the amount of non-specific proteins in each lane of the gel. Nearly all of them appear to be functional, which establishes that Formula VIII and IX can be used to predict siRNAs' functionality in general in a genome wide manner.
  • To generate the fusion of yellow fluorescent protein (YFP) with Rab5b or Rab5c (YFP-Rab5b or YFP-Rab5c), a DNA fragment encoding the full-length human Rab5b or Rab5c was obtained by PCR using Pfu polymerase (Stratagene) with a SacI restriction site introduced into the 5′ end and a KpnI site into the 3′ end and cloned into pEYFP-C1 vector (CLONTECH, Palo Alto, Calif., USA). GFP-CALM and YFP-Rab5a were described previously (Tebar, Bohlander, & Sorkin (1999) Clathrin Assembly Lymphoid Myeloid Leukemia (CALM) Protein: Localization in Endocytic-coated Pits, Interactions with Clathrin, and the Impact of Overexpression on Clathrin-mediated Traffic, Mol. Biol. Cell 10:2687).
    • Example IV Validation of the Algorithm Using Eg5, GADPH, ATE1, MEK2, MEK1, QB, LaminA/C, C-myc, Human Cyclophilin, and Mouse Cyclophilin
  • A number of genes have been identified as playing potentially important roles in disease etiology. Expression profiles of normal and diseased kidneys has implicated Edg5 in immunoglobulin A neuropathy, a common renal glomerular disease. Myc1, MEK1/2 and other related kinases have been associated with one or more cancers, while lamins have been implicated in muscular dystrophy and other diseases. For these reasons, siRNA directed against the genes encoding these classes of molecules would be important research and therapeutic tools.
  • FIG. 12 illustrates four siRNAs targeting 10 different genes (Table V for sequence and accession number information) that were selected according to the Formula VIII and assayed as individuals and pools in HEK293 cells. The level of siRNA induced silencing was measured using the B-DNA assay. These studies demonstrated that thirty-six out of the forty individual SMART-selected siRNA tested are functional (90%) and all 10 pools are fully functional.
    • Example V Validation of the Algorithm Using Bcl2
  • Bcl-2 is a ˜25 kD, 205-239 amino acid, anti-apoptotic protein that contains considerable homology with other members of the BCL family including BCLX, MCL1, BAX, BAD, and BIK. The protein exists in at least two forms (Bcl2a, which has a hydrophobic tail for membrane anchorage, and Bcl2b, which lacks the hydrophobic tail) and is predominantly localized to the mitochondrial membrane. While Bcl2 expression is widely distributed, particular interest has focused on the expression of this molecule in B and T cells. Bcl2 expression is down-regulated in normal germinal center B cells yet in a high percentage of follicular lymphomas, Bcl2 expression has been observed to be elevated. Cytological studies have identified a common translocation ((14; 18)(q32; q32)) amongst a high percentage (>70%) of these lymphomas. This genetic lesion places the Bcl2 gene in juxtaposition to immunoglobulin heavy chain gene (IgH) encoding sequences and is believed to enforce inappropriate levels of gene expression, and resistance to programmed cell death in the follicle center B cells. In other cases, hypomethylation of the Bcl2 promoter leads to enhanced expression and again, inhibition of apoptosis. In addition to cancer, dysregulated expression of Bcl-2 has been correlated with multiple sclerosis and various neurological diseases.
  • The correlation between Bcl-2 translocation and cancer makes this gene an attractive target for RNAi. Identification of siRNA directed against the bcl2 transcript (or Bcl2-IgH fusions) would further our understanding Bcl2 gene function and possibly provide a future therapeutic agent to battle diseases that result from altered expression or function of this gene.
  • In Silico Identification of Functional siRNA
  • To identify functional and hyperfunctional siRNA against the Bcl2 gene, the sequence for Bcl-2 was downloaded from the NCBI Unigene database and analyzed using the Formula VIII algorithm. As a result of these procedures, both the sequence and SMARTSCORES™, or siRNA rankings of the Bcl2 siRNA were obtained and ranked according to their functionality. Subsequently, these sequences were BLAST'ed (database) to insure that the selected sequences were specific and contained minimal overlap with unrealated genes. The SMARTSCORES™, or siRNA rankings for the top 10 Bcl-2 siRNA are identified in FIG. 13.
  • In Vivo Testing of Bcl-2 SiRNA
  • Bcl-2 siRNAs having the top ten SMARTSCORES™, or siRNA rankings were selected and tested in a functional assay to determine silencing efficiency. To accomplish this, each of the ten duplexes were synthesized using 2′-O-ACE chemistry and transfected at 100 nM concentrations into cells. Twenty-four hours later assays were performed on cell extracts to assess the degree of target silencing. Controls used in these experiments included mock transfected cells, and cells that were transfected with a non-specific siRNA duplex.
  • The results of these experiments are presented below (and in FIG. 14) and show that all ten of the selected siRNA induce 80% or better silencing of the Bcl2 message at 100 nM concentrations. These data verify that the algorithm successfully identified functional Bcl2 siRNA and provide a set of functional agents that can be used in experimental and therapeutic environments.
    siRNA 1 GGGAGAUAGUGAUGAAGUA SEQ. ID NO. 302
    siRNA 2 GAAGUACAUCCAUUAUAAG SEQ. ID NO. 303
    siRNA 3 GUACGACAACCGGGAGAUA SEQ. ID NO. 304
    siRNA 4 AGAUAGUGAUGAAGUACAU SEQ. ID NO. 305
    siRNA 5 UGAAGACUCUGCUCAGUUU SEQ. ID NO. 306
    sIRNA 6 GCAUGCGGCCUCUGUUUGA SEQ. ID NO. 307
    siRNA 7 UGCGGCCUCUGUUUGAUUU SEQ. ID NO. 308
    siRNA 8 GAGAUAGUGAUGAAGUACA SEQ. ID NO. 309
    siRNA 9 GGAGAUAGUGAUGAAGUAC SEQ. ID NO. 310
    siRNA 10 GAAGACUCUGCUCAGUUUG SEQ. ID NO. 311
  • Bcl2 siRNA: Sense Strand, 5′→3′
    • Example VI Sequences Selected by the Algorithm
  • Sequences of the siRNAs selected using Formulas (Algorithms) VIII and IX with their corresponding ranking, which have been evaluated for the silencing activity in vivo in the present study (Formula VIII and IX, respectively) are shown in Table V. It should be noted that the “t” residues in Table V, and elsewhere, when referring to siRNA, should be replaced by “u” residues.
    TABLE V
    SEQ.
    ID FORMULA FORMULA
    GENE Name NO. FTLLSEQTENCE VIII IX
    CLTC NM_004859 0312 GAAAGAATCTGTAGAGAAA 76 94.2
    CLTC NM_004859 0313 GCAATGAGCTGTTTGAAGA 65 39.9
    CLTC NM_004859 0314 TGACAAAGGTGGATAAATT 57 38.2
    CLTC NM_004859 0315 GGAAATGGATCTCTTTGAA 54 49.4
    CLTA NM_001833 0316 GGAAAGTAATGGTCCAACA 22 55.5
    CLTA NM_001833 0317 AGACAGTTATGCAGCTATT 4 22.9
    CLTA NM_001833 0318 CCAATTCTCGGAAGCAAGA 1 17
    CLTA NM_001833 0319 GAAAGTAATGGTCCAACAG −1 −13
    CLTB NM_001834 0320 GCGCCAGAGTGAACAAGTA 17 57.5
    CLTB NM_001834 0321 GAAGGTGGCCCAGCTATGT 15 −8.6
    CLTB NM_001834 0322 GGAACCAGCGCCAGAGTGA 13 40.5
    CLTB NM_001834 0323 GAGCGAGATTGCAGGCATA 20 61.7
    CALM U45976 0324 GTTAGTATCTGATGACTTG 36 −34.6
    CALM U45976 0325 GAAATGGAACCACTAAGAA 33 46.1
    CALM U45976 0326 GGAAATGGAACCACTAAGA 30 61.2
    CALM U45976 0327 CAACTACACTTTCCAATGC 28 6.8
    EPS15 NM_001981 0328 CCACCAAGATTTCATGATA 48 25.2
    EPS15 NM_001981 0329 GATCGGAACTCCAACAAGA 43 49.3
    EPS15 NM_001981 0330 AAACGGAGCTACAGATTAT 39 11.5
    EPS15 NM_001981 0331 CCACACAGCATTCTTGTAA 33 −23.6
    EPS15R NM_021235 0332 GAAGTTACCTTGAGCAATC 48 33
    EPS15R NM_021235 0333 GGACTTGGCCGATCCAGAA 27 33
    EPS15R NM_021235 0334 GCACTTGGATCGAGATGAG 20 1.3
    EPS15R NM_021235 0335 CAAAGACCAATTCGCGTTA 17 27.7
    DNM2 NM_004945 0336 CCGAATCAATCGCATCTTC 6 −29.6
    DNM2 NM_004945 0337 GACATGATCCTGCAGTTCA 5 −14
    DNM2 NM_004945 0338 GAGCGAATCGTCACCACTT 5 24
    DNM2 NM_004945 0339 CCTCCGAGCTGGCGTCTAC −4 −63.6
    ARF6 AF93885 0340 TCACATGGTTAACCTCTAA 27 −21.1
    ARF6 AF93885 0341 GATGAGGGACGCCATAATC 7 −38.4
    ARF6 AF93885 0342 CCTCTAACTACAAATCTTA 4 16.9
    ARF6 AF93885 0343 GGAAGGTGCTATCCAAAAT 4 11.5
    RAB5A BC001267 0344 GCAAGCAAGTCCTAACATT 40 25.1
    RAB5A BC001267 0345 GGAAGAGGAGTAGACCTTA 17 50.1
    RAB5A BC001267 0346 AGGAATCAGTGTTGTAGTA 16 11.5
    RAB5A BC001267 0347 GAAGAGGAGTAGACCTTAC 12 7
    RAB5B NM_002868 0348 GAAAGTCAAGCCTGGTATT 14 18.1
    RAB5B NM_002868 0349 AAAGTCAAGCCTGGTATTA 6 −17.8
    RAB5B NM_002868 0350 GCTATGAACGTGAATGATC 3 −21.1
    RAB5B NM_002868 0351 CAAGCCTGGTATTACGTTT −7 −37.5
    RAB5C AF141304 0352 GGAACAAGATCTGTCAATT 38 51.9
    RAB5C AF141304 0353 GCAATGAACGTGAACGAAA 29 43.7
    RAB5C AF141304 0354 CAATGAACGTGAACGAAAT 18 43.3
    RAB5C AF141304 0355 GGACAGGAGCGGTATCACA 6 18.2
    EEA1 XM_018197 0356 AGACAGAGCTTGAGAATAA 67 64.1
    EEA1 XM_018197 0357 GAGAAGATCTTTATGCAAA 60 48.7
    EEA1 XM_018197 0358 GAAGAGAAATCAGCAGATA 58 45.7
    EEA1 XM_018197 0359 GCAAGTAACTCAACTAACA 56 72.3
    AP2B1 NM_001282 0360 GAGCTAATCTGCCACATTG 49 −12.4
    AP2B1 NM_001282 0361 GCAGATGAGTTACTAGAAA 44 48.9
    AP2B1 NM_001282 0362 CAACTTAATTGTCCAGAAA 41 28.2
    AP2B1 NM_001282 0363 CAACACAGGATTCTGATAA 33 −5.8
    PLK NM_005030 0364 AGATTGTGCCTAAGTCTCT −35 −3.4
    PLK NM_005030 0365 ATGAAGATCTGGAGGTGAA 0 −4.3
    PLK NM_005030 0366 TTTGAGACTTCTTGCCTAA −5 −27.7
    PLK NM_005030 0367 AGATCACCCTCCTTAAATA 15 72.3
    GAPDH NM_002046 0368 CAACGGATTTGGTCGTATT 27 −2.8
    GAPDH NM_002046 0369 GAAATCCCATCACCATCTT 24 3.9
    GAPDH NM_002046 0370 GACCTCAACTACATGGTTT 22 −22.9
    GAPDH NM_002046 0371 TGGTTTACATGTTCCAATA 9 9.8
    c-Myc 0372 GAAGAAATCGATGTTGTTT 31 −11.7
    c-Myc 0373 ACACAAACTTGAACAGCTA 22 51.3
    c-Myc 0374 GGAAGAAATCGATGTTGTT 18 26
    c-Myc 0375 GAAACGACGAGAACAGTTG 18 −8.9
    MAP2K1 NM_002755 0376 GCACATGGATGGAGGTTCT 26 16
    MAP2K1 NM_002755 0377 GCAGAGAGAGCAGATTTGA 16 0.4
    MAP2K1 NM_002755 0378 GAGGTTCTCTGGATCAAGT 14 15.5
    MAP2K1 NM_002755 0379 GAGCAGATTTGAAGCAACT 14 18.5
    MAP2K2 NM_030662 0380 CAAAGACGATGACTTCGAA 37 26.4
    MAP2K2 NM_030662 0381 GATCAGCATTTGCATGGAA 24 −0.7
    MAP2K2 NM_030662 0382 TCCAGGAGTTTGTCAATAA 17 −4.5
    MAP2K2 NM_030662 0383 GGAAGCTGATCCACCTTGA 16 59.2
    KNSL1(EG5) NM_004523 0384 GCAGAAATCTAAGGATATA 53 35.8
    KNSL1(EG5) NM_004523 0385 CAACAAGGATGAAGTCTAT 50 18.3
    KNSL1(EG5) NM_004523 0386 CAGCAGAAATCTAAGGATA 41 32.7
    KNSL1(EG5) NM_004523 0387 CTAGATGGCTTTCTCAGTA 39 3.9
    CyclophilinA NM_021130 0388 AGACAAGGTCCCAAAGACA −16 58.1
    CyclophilinA NM_021130 0389 GGAATGGCAAGACCAGCAA −6 36
    CyclophilinA NM_021130 0390 AGAATTATTCCAGGGTTTA −3 16.1
    CyclophilinA NM_021130 0391 GCAGACAAGGTCCCAAAGA 8 8.9
    LAMIN A/C NM_170707 0392 AGAAGCAGCTTCAGGATGA 31 38.8
    LAMIN A/C NM_170707 0393 GAGCTTGACTTCCAGAAGA 33 22.4
    LAMIN A/C NM_170707 0394 CCACCGAAGTTCACCCTAA 21 27.5
    LAMIN A/C NM_170707 0395 GAGAAGAGCTCCTCCATCA 55 30.1
    CyclophilinB M60857 0396 GAAAGAGCATCTACGGTGA 41 83.9
    CyclophilinB M60857 0397 GAAAGGATTTGGCTACAAA 53 59.1
    CyclophilinB M60857 0398 ACAGCAAATTCCATCGTGT −20 28.8
    CyclophilinB M60857 0399 GGAAAGACTGTTCCAAAAA 2 27
    DBI1 NM_020548 0400 CAACACGCCTCATCCTCTA 27 −7.6
    DBI2 NM_020548 0401 CATGAAAGCTTACATCAAC 25 −30.8
    DBI3 NM_020548 0402 AAGATGCCATGAAAGCTTA 17 22
    DBI4 NM_020548 0403 GCACATACCGCCTGAGTCT 15 3.9
    rLUC1 0404 GATCAAATCTGAAGAAGGA 57 49.2
    rLUC2 0405 GCCAAGAAGTTTCCTAATA 50 13.7
    rLUC3 0406 CAGCATATCTTGAACCATT 41 −2.2
    rLUC4 0407 GAACAAAGGAAACGGATGA 39 29.2
    SeAP1 NM_031313 0408 CGGAAACGGTCCAGGCTAT 6 26.9
    SeAP2 NM_031313 0409 GCTTCGAGCAGACATGATA 4 −11.2
    SeAP3 NM_031313 0410 CCTACACGGTCCTCCTATA 4 4.9
    SeAP4 NM_031313 0411 GCCAAGAACCTCATCATCT 1 −9.9
    fLUC1 0412 GATATGGGCTGAATACAAA 54 40.4
    fLUC2 0413 GCACTCTGATTGACAAATA 47 54.7
    fLUC3 0414 TGAAGTCTCTGATTAAGTA 46 34.5
    fLUC4 0415 TCAGAGAGATCCTCATAAA 40 11.4
    mCyclo_1 NM_008907 0416 GCAAGAAGATCACCATTTC 52 46.4
    mCyclo_2 NM_008907 0417 GAGAGAAATTTGAGGATGA 36 70.7
    mCyclo_3 NM_008907 0418 GAAAGGATTTGGCTATAAG 35 −1.5
    mCyclo_4 NM_008907 0419 GAAAGAAGGCATGAACATT 27 10.3
    BCL2_1 NM_000633 0420 GGGAGATAGTGATGAAGTA 21 72
    BCL2_2 NM_000633 0421 GAAGTACATCCATTATAAG 1 3.3
    BCL2_3 NM_000633 0422 GTACGACAACCGGGAGATA 1 35.9
    BCL2_4 NM_000633 0423 AGATAGTGATGAAGTACAT −12 22.1
    BCL2_5 NM_000633 0424 TGAAGACTCTGCTCAGTTT 36 19.1
    BCL2_6 NM_000633 0425 GCATGCGGCCTCTGTTTGA 5 −9.7
    QB1 NM_003365.1 0426 GCACACAGCUUACUACAUC 52 −4.8
    QB2 NM_003365.1 0427 GAAAUGCCCUGGUAUCUCA 49 22.1
    QB3 NM_003365.1 0428 GAAGGAACGUGAUGUGAUC 34 22.9
    QB4 NM_003365.1 0429 GCACUACUCCUGUGUGUGA 28 20.4
    ATE1-1 NM_007041 0430 GAACCCAGCUGGAGAACUU 45 15.5
    ATE1-2 NM_007041 0431 GAUAUACAGUGUGAUCUUA 40 12.2
    ATE1-3 NM_007041 0432 GUACUACGAUCCUGAUUAU 37 32.9
    ATE1-4 NM_007041 0433 GUGCCGACCUUUACAAUUU 35 18.2
    EGFR-1 NM_005228 0434 GAAGGAAACTGAATTCAAA 68 79.4
    EGFR-1 NM_005228 0435 GGAAATATGTACTACGAAA 49 49.5
    EGFR-1 NM_005228 0436 CCACAAAGCAGTGAATTTA 41 7.6
    EGFR-1 NM_005228 0437 GTAACAAGCTCACGCAGTT 40 25.9
  • Many of the genes to which the described siRNA are directed play critical roles in disease etiology. For this reason, the siRNAs listed in the sequence listing may potentially act as therapeutic agents. A number of prophetic examples follow and should be understood in view of the siRNA that are identified in the sequence listing. To isolate these siRNAs, the appropriate message sequence for each gene is analyzed using one of the before mentioned formulas (preferably formula VIII) to identify potential siRNA targets. Subsequently these targets are BLAST'ed to eliminate homology with potential off-targets.
    • Example VII Evidence for the Benefits of Pooling
  • Evidence for the benefits of pooling have been demonstrated using the reporter gene, luciferase. Ninety siRNA duplexes were synthesized using Dharmacon proprietary ACE® chemistry against one of the standard reporter genes: firefly luciferase. The duplexes were designed to start two base pairs apart and to cover approximately 180 base pairs of the luciferase gene (see sequences in Table III). Subsequently, the siRNA duplexes were co-transfected with a luciferase expression reporter plasmid into HEK293 cells using standard transfection protocols and luciferase activity was assayed at 24 and 48 hours.
  • Transfection of individual siRNAs showed standard distribution of inhibitory effect. Some duplexes were active, while others were not. FIG. 15 represents a typical screen of ninety siRNA duplexes (SEQ. ID NO. 0032-0120) positioned two base pairs apart. As the figure suggests, the functionality of the siRNA duplex is determined more by a particular sequence of the oligonucleotide than by the relative oligonucleotide position within a gene or excessively sensitive part of the mRNA, which is important for traditional anti-sense technology.
  • When two continuous oligonucleotides were pooled together, a significant increase in gene silencing activity was observed (see FIGS. 16A and B). A gradual increase in efficacy and the frequency of pools functionality was observed when the number of siRNAs increased to 3 and 4 (FIGS. 16A, 16B, 17A, and 17B). Further, the relative positioning of the oligonucleotides within a pool did not determine whether a particular pool was functional (see FIGS. 18A and 18B, in which 100% of pools of oligonucleotides distanced by 2, 10 and 20 base pairs were functional).
  • However, relative positioning may nonetheless have an impact. An increased functionality may exist when the siRNA are positioned continuously head to toe (5′ end of one directly adjacent to the 3′ end of the others).
  • Additionally, siRNA pools that were tested performed at least as well as the best oligonucleotide in the pool, under the experimental conditions whose results are depicted in FIG. 19. Moreover, when previously identified non-functional and marginally (semi) functional siRNA duplexes were pooled together in groups of five at a time, a significant functional cooperative action was observed (see FIG. 20). In fact, pools of semi-active oligonucleotides were 5 to 25 times more functional than the most potent oligonucleotide in the pool. Therefore, pooling several siRNA duplexes together does not interfere with the functionality of the most potent siRNAs within a pool, and pooling provides an unexpected significant increase in overall functionality
    • Example VIII Additional Evidence of the Benefits of Pooling
  • Experiments were performed on the following genes: P-galactosidase, Renilla luciferase, and Secreted alkaline phosphatase, which demonstrates the benefits of pooling. (see FIGS. 21A, 21B and 21C). Individual and pools of siRNA (described in Figure legends 21A-C) were transfected into cells and tested for silencing efficiency. Approximately 50% of individual siRNAs designed to silence the above-specified genes were functional, while 100% of the pools that contain the same siRNA duplexes were functional.
    • Example IX Highly Functional siRNA
  • Pools of five siRNAs in which each two siRNAs overlap to 10-90% resulted in 98% functional entities (>80% silencing). Pools of siRNAs distributed throughout the mRNA that were evenly spaced, covering an approximate 20-2000 base pair range, were also functional. When the pools of siRNA were positioned continuously head to tail relative to mRNA sequences and mimicked the natural products of Dicer cleaved long double stranded RNA, 98% of the pools evidenced highly functional activity (>95% silencing).
    • Example X Human Cyclophilin B
  • Table III above lists the siRNA sequences for the human cyclophilin B protein. A particularly functional siRNA may be selected by applying these sequences to any of Formula I to VII above.
  • Alternatively, one could pool 2, 3, 4, 5 or more of these sequences to create a kit for silencing a gene. Preferably, within the kit there would be at least one sequence that has a relatively high predicted functionality when any of Formulas I-VII is applied.
    • Example XI Sample Pools of siRNAs and Their Application to Human Disease
  • The genetic basis behind human disease is well documented and siRNA may be used as both research or diagnostic tools and therapeutic agents, either individually or in pools. Genes involved in signal transduction, the immune response, apoptosis, DNA repair, cell cycle control, and a variety of other physiological functions have clinical relevance and therapeutic agents that can modulate expression of these genes may alleviate some or all of the associated symptoms. In some instances, these genes can be described as a member of a family or class of genes and siRNA (randomly, conventionally, or rationally designed) can be directed against one or multiple members of the family to induce a desired result.
  • To identify rationally designed siRNA to each gene, the sequence was analyzed using Formula VIII or Formula X to identify rationally designed siRNA. To confirm the activity of these sequences, the siRNA are introduced into a cell type of choice (e.g., HeLa cells, HEK293 cells) and the levels of the appropriate message are analyzed using one of several art proven techniques. siRNA having heightened levels of potency can be identified by testing each of the before mentioned duplexes at increasingly limiting concentrations. Similarly, siRNA having increased levels of longevity can be identified by introducing each duplex into cells and testing functionality at 24, 48, 72, 96, 120, 144, 168, and 192 hours after transfection. Agents that induce >95% silencing at sub-nanomolar concentrations and/or induce functional levels of silencing for >96 hours are considered hyperfunctional.
    • Example XII Validation of Multigene Knockout Using Rab5 and Eps
  • Two or more genes having similar, overlapping functions often leads to genetic redundancy. Mutations that knockout only one of, e.g., a pair of such genes (also referred to as homologs) results in little or no phenotype due to the fact that the remaining intact gene is capable of fulfilling the role of the disrupted counterpart. To fully understand the function of such genes in cellular physiology, it is often necessary to knockout or knockdown both homologs simultaneously. Unfortunately, concomitant knockdown of two or more genes is frequently difficult to achieve in higher organisms (e.g., mice) thus it is necessary to introduce new technologies dissect gene function. One such approach to knocking down multiple genes simultaneously is by using siRNA. For example, FIG. 11 showed that rationally designed siRNA directed against a number of genes involved in the clathrin-mediated endocytosis pathway resulted in significant levels of protein reduction (e.g., >80%). To determine the effects of gene knockdown on clathrin-related endocytosis, internalization assays were performed using epidermal growth factor and transferrin. Specifically, mouse receptor-grade EGF (Collaborative Research Inc.) and iron-saturated human transferrin (Sigma) were iodinated as described previously (Jiang, X., Huang, F., Marusyk, A. & Sorkin, A. (2003) Mol Biol Cell 14, 858-70). HeLa cells grown in 12-well dishes were incubated with 125I-EGF (1 ng/ml) or 125I-transferrin (1 μg/ml) in binding medium (DMEM, 0.1% bovine serum albumin) at 37° C., and the ratio of internalized and surface radioactivity was determined during 5-min time course to calculate specific internalization rate constant kc as described previously (Jiang, X et al.). The measurements of the uptakes of radiolabeled transferrin and EGF were performed using short time-course assays to avoid influence of the recycling on the uptake kinetics, and using low ligand concentration to avoid saturation of the clathrin-dependent pathway (for EGF Lund, K. A., Opresko, L. K., Strarbuck, C., Walsh, B. J. & Wiley, H. S. (1990) J. Biol. Chem. 265, 15713-13723).
  • The effects of knocking down Rab5a, 5b, 5c, Eps, or Eps 15R (individually) are shown in FIG. 22 and demonstrate that disruption of single genes has little or no effect on EGF or Tfn internalization. In contrast, simultaneous knock down of Rab5a, 5b, and 5c, or Eps and Eps 15R, leads to a distinct phenotype (note: total concentration of siRNA in these experiments remained constant with that in experiments in which a single siRNA was introduced, see FIG. 23). These experiments demonstrate the effectiveness of using rationally designed siRNA to knockdown multiple genes and validates the utility of these reagents to override genetic redundancy.
    • Example XIII Validation of Multigene Targeting Using G6PD, GAPDH, PLK, and UQC
  • Further demonstration of the ability to knock down expression of multiple genes using rationally designed siRNA was performed using pools of siRNA directed against four separate genes. To achieve this, siRNA were transfected into cells (total siRNA concentration of 100 nM) and assayed twenty-four hours later by B-DNA. Results shown in FIG. 24 show that pools of rationally designed molecules are capable of simultaneously silencing four different genes.
    • Example XIV Validation of Multigene Knockouts as Demonstrated by Gene Expression Profiling, a Prophetic Example
  • To further demonstrate the ability to concomitantly knockdown the expression of multiple gene targets, single siRNA or siRNA pools directed against a collection of genes (e.g., 4, 8, 16, or 23 different targets) are simultaneously transfected into cells and cultured for twenty-four hours. Subsequently, mRNA is harvested from treated (and untreated) cells and labeled with one of two fluorescent probes dyes (e.g., a red fluorescent probe for the treated cells, a green fluorescent probe for the control cells.). Equivalent amounts of labeled RNA from each sample is then mixed together and hybridized to sequences that have been linked to a solid support (e.g., a slide, “DNA CHIP”). Following hybridization, the slides are washed and analyzed to assess changes in the levels of target genes induced by siRNA.
    • Example XV Identifying Hyperfunctional siRNA
  • Identification of Hyperfunctional Bcl-2 siRNA
  • The ten rationally designed Bcl2 siRNA (identified in FIGS. 13, 14) were tested to identify hyperpotent reagents. To accomplish this, each of the ten Bcl-2 siRNA were individually transfected into cells at a 300 pM (0.3 nM) concentrations. Twenty-four hours later, transcript levels were assessed by B-DNA assays and compared with relevant controls. As shown in FIG. 25, while the majority of Bcl-2 siRNA failed to induce functional levels of silencing at this concentration, siRNA 1 and 8 induced >80% silencing, and siRNA 6 exhibited greater than 90% silencing at this subnanomolar concentration.
  • By way of prophetic examples, similar assays could be performed with any of the groups of rationally designed genes described in the Examples. Thus for instance, rationally designed siRNA sequences directed against a gene of interest could be introduced into cells at increasingly limiting concentrations to determine whether any of the duplexes are hyperfunctional.
    • Example XVI Gene Silencing: Prophetic Example
  • Below is an example of how one might transfect a cell.
  • Select a cell line. The selection of a cell line is usually determined by the desired application. The most important feature to RNAi is the level of expression of the gene of interest. It is highly recommended to use cell lines for which siRNA transfection conditions have been specified and validated.
  • Plate the cells. Approximately 24 hours prior to transfection, plate the cells at the appropriate density so that they will be approximately 70-90% confluent, or approximately 1×105 cells/ml at the time of transfection. Cell densities that are too low may lead to toxicity due to excess exposure and uptake of transfection reagent-siRNA complexes. Cell densities that are too high may lead to low transfection efficiencies and little or no silencing. Incubate the cells overnight. Standard incubation conditions for mammalian cells are 37° C. in 5% CO2. Other cell types, such as insect cells, require different temperatures and CO2 concentrations that are readily ascertainable by persons skilled in the art. Use conditions appropriate for the cell type of interest.
  • siRNA re-suspension. Add 20 μl siRNA universal buffer to each siRNA to generate a final concentration of 50 μM.
  • siRNA-lipid complex formation. Use RNase-free solutions and tubes. Using the following table, Table XI:
    TABLE XI
    96-WELL 24-WELL
    MIXTURE 1 (TRANSIT-TKO-PLASMID DILUTION MIXTURE)
    Opti-MEM 9.3 μl 46.5 μl
    TransIT-TKO (1 μg/μl) 0.5 μl 2.5 μl
    MIXTURE
    1 FINAL VOLUME 10.0 μl 50.0 μl
    MIXTURE 2 (SIRNA DILUTION MIXTURE)
    Opti-MEM 9.0 μl 45.0 μl
    siRNA (1 μM) 1.0 μl 5.0 μl
    MIXTURE
    2 FINAL VOLUME 10.0 μl 50.0 μl
    MIXTURE 3 (SIRNA-TRANSFECTION REAGENT MIXTURE)
    Mixture 1 10 μl 50 μl
    Mixture
    2 10 μl 50 μl
    MIXTURE
    3 FINAL VOLUME 20 μl 100 μl
    Incubate
    20 minutes at room temperature
    MIXTURE 4 (MEDIA-SIRNA/TRANSFECTION REAGENT
    MIXTURE)
    Mixture 3 20 μl 100 μl
    Complete media 80 μl 400 μl
    MIXTURE
    4 FINAL VOLUME 100 μl 500 μl
    Incubate 48 hours at 37° C.
  • Transfection. Create a Mixture 1 by combining the specified amounts of OPTI-MEM serum free media and transfection reagent in a sterile polystyrene tube. Create a Mixture 2 by combining specified amounts of each siRNA with OPTI-MEM media in sterile 1 ml tubes. Create a Mixture 3 by combining specified amounts of Mixture 1 and Mixture 2. Mix gently (do not vortex) and incubate at room temperature for 20 minutes. Create a Mixture 4 by combining specified amounts of Mixture 3 to complete media. Add appropriate volume to each cell culture well. Incubate cells with transfection reagent mixture for 24-72 hours at 37° C. This incubation time is flexible. The ratio of silencing will remain consistent at any point in the time period. Assay for gene silencing using an appropriate detection method such as RT-PCR, Western blot analysis, immunohistochemistry, phenotypic analysis, mass spectrometry, fluorescence, radioactive decay, or any other method that is now known or that comes to be known to persons skilled in the art and that from reading this disclosure would useful with the present invention. The optimal window for observing a knockdown phenotype is related to the mRNA turnover of the gene of interest, although 24-72 hours is standard. Final Volume reflects amount needed in each well for the desired cell culture format. When adjusting volumes for a Stock Mix, an additional 10% should be used to accommodate variability in pipetting, etc. Duplicate or triplicate assays should be carried out when possible.
    • Example XVII siRNAs That Target Deubiquitination Enzymes
  • siRNAs that target nucleotide sequences for deubiquitination enzymes with the NCBI accession numbers denoted below and having sequences generated in silico by the algorithms herein, are provided. In various embodiments, the siRNAs are rationally designed. In various embodiments, the siRNAs are functional or hyperfunctional. These siRNA that have been generated by the algorithms of the present invention include:
    SEQ +HL,26
    ID
    NO Name siRNASense Accession
     438 AMSH AGAAGGAAGCAGAGGAAUU NM_006463
     439 AMSH CGGUAGAGGUGAAUGAAGA NM_006463
     440 AMSH GGGCAUCACCUGAGAAAGA NM_006463
     441 AMSH GAAGAAGGAAGCAGAGGAA NM_006463
     442 AMSH CAAUAUGAAUGGAGCUUAU NM_006463
     443 AMSH GCUCUGGAGUUGAGAUUAU NM_006463
     444 AMSH GUGGAAAACUGAUGAGGAA NM_006463
     445 AMSH GAAAGACACAGUAAAGAAA NM_006463
     446 AMSH ACACAGAGAACGAAGAAGA NM_006463
     447 AMSH AGAAAGACACAGUAAAGAA NM_006463
     448 AMSH GAAAAGAAAGACACAGUAA NM_006463
     449 AMSH GAAAUUAAGUAGCUCAGAA NM_006463
     450 AMSH CAACAGAAGCAGCAGCAAU NM_006463
     451 AMSH GAACAUGGCCAUCCAGCAA NM_006463
     452 AMSH GCUAACACAUCCCGAAGAA NM_006463
     453 AMSH GGAGAUUGCAUUUCCCAAA NM_006463
     454 AMSH CAACUGUAACUCAGAAAUU NM_006463
     455 AMSH UUACAAAUCUGCUGUCAUU NM_006463
     456 AMSH CUAGAAAGCUUUGGAAGUU NM_006463
     457 AMSH GAGUUGAGAUUAUCCGAAU NM_006463
     458 AMSH CAACACAGAGAACGAAGAA NM_006463
     459 AMSH-LP GAGUAGAGAUGGAGAGGAU NM_020799
     460 AMSH-LP AAUAGAAACCUGUGGAAUA NM_020799
     461 AMSH-LP UGGAGAAUGUAGAGGAAUU NM_020799
     462 AMSH-LP UGAUAGAGGCAGAAAGGAA NM_020799
     463 AMSH-LP UGAAGAAACUGAAGGAGAU NM_020799
     464 AMSH-LP GAAGAAGGAAAUUUGGAAA NM_020799
     465 AMSH-LP UAAUACAGUGAGAGGAAUA NM_020799
     466 AMSH-LP GAAUGUAUUUGCAGAUCAA NM_020799
     467 AMSH-LP UGGAAUACUCUGUGGAAAA NM_020799
     468 AMSH-LP GGACCAGACUAUUGUGACA NM_020799
     469 AMSH-LP UCUAAUACAGUGAGAGGAA NM_020799
     470 AMSH-LP ACGUAGAAUACCAAGAAUA NM_020799
     471 AMSH-LP CGUAGAAUACCAAGAAUAU NM_020799
     472 AMSH-LP AGAGUUAGCCCGAGGUCAA NM_020799
     473 AMSH-LP AGAGAUUGAUAGAGGCAGA NM_020799
     474 AMSH-LP UGUAUUUGGAAGAAGGAAA NM_020799
     475 AMSH-LP CAUGGAGAAUGUAGAGGAA NM_020799
     476 AMSH-LP AUGAAGAAACUGAAGGAGA NM_020799
     477 AMSH-LP GGAAGAAGGAAAUUUGGAA NM_020799
     478 AMSH-LP GGAAUAGAAACCUGUGGAA NM_020799
     479 AMSH-LP UCUAAGUGCUGUUCAGAAU NM_020799
     480 AMSH-LP GGAGAAUGUAGAGGAAUUA NM_020799
     481 AMSH-LP UCACCAAAGCAUAAAGACA NM_020799
     482 AMSH-LP GCAUAAAGACACUGGCAUC NM_020799
     483 AMSH-LP GUGGAAUACUCUGUGGAAA NM_020799
     484 AMSH-LP AAUUGGAGCAUCAGAGAUU NM_020799
     485 AMSH-LP GCAGAAAGGAAGCGGAUUG NM_020799
     486 AMSH-LP GGUCUGGAGUAGAGAUGGA NM_020799
     487 AMSH-LP CUGAAAAGCAGGAUAUUAU NM_020799
     488 AMSH-LP CUUCCUAACCAUCGAGAUU NM_020799
     489 BAP1 GAGCAAAGGAUAUGCGAUU NM_004656
     490 BAP1 CCACAAGUCUCAAGAGUCA NM_004656
     491 BAP1 GAGAAGAGGAAGAAGUUCA NM_004656
     492 BAP1 GGGUGCAAGUGGAGGAGAU NM_004656
     493 BAP1 CGUGAUUGAUGAUGAUAUU NM_004656
     494 BAP1 UCUACGACCUUCAGAGCAA NM_004656
     495 BAP1 GGGUCAUCAUGGAGCGUAU NM_004656
     496 BAP1 GAGGUAGAGAAGAGGAAGA NM_004656
     497 BAP1 UCAAAGAGUCCCAGAAGGA NM_004656
     498 BAP1 CCAUCAACGUCUUGGCUGA NM_004656
     499 BAP1 AGGAUGACGAGGAGGAUGA NM_004656
     500 BAP1 AGGAGGUAGAGAAGAGGAA NM_004656
     501 BAP1 AGGCUGAGAUUGCAAACUA NM_004656
     502 BAP1 UCAAGGAGGAGGUAGAGAA NM_004656
     503 BAP1 UGGAUGGGCUGAAGGUCUA NM_004656
     504 BAP1 UGUCAGUGCUGCAGCCCAA NM_004656
     505 BAP1 CCAACCUAGUGGAGCAGAA NM_004656
     506 BAP1 GUAGAGAAGAGGAAGAAGU NM_004656
     507 BAP1 AGGAUGACUAUGAGGAUGA NM_004656
     508 BAP1 CAGAUGAUGAGGAUGACUA NM_004656
     509 BAP1 AGAGAAGGACCCACAACUA NM_004656
     510 BAP1 CGGUUCUGCUGAUGGGCAA NM_004656
     511 BAP1 AGGAAGAAGUUCAAGAUUG NM_004656
     512 BAP1 AGGAAGGCAUGCUGGCCAA NM_004656
     513 BAP1 CUGGAUCAAUGAAUGAAUA NM_004656
     514 BAP1 AACCCAAGCUAGUGGUGAA NM_004656
     515 BAP1 CCAAGGAGCUGCUGGCACU NM_004656
     516 BAP1 AGGUAUAAGGGGAAGGGAA NM_004656
     517 BAP1 GCAGAGACAGGGUUGCUUA NM_004656
     518 BAP1 CCUACAAGGCCAAGCGCCA NM_004656
     519 C10ORF29 GAUGAUGACUGGAGGCAAA NM_152586
     520 C10ORF29 UAACCUAUCCAGAGAGAAA NM_152586
     521 C10ORF29 GAUAGAAGUUUGUCAGGUA NM_152586
     522 C10ORF29 GUGAGGAAGCCUUUGGAAA NM_152586
     523 C10ORF29 AGUACAGGUUUCAAGGAUA NM_152586
     524 C10ORF29 CAGGAAGGACUUUGAACUA NM_152586
     525 C10ORF29 GCAGUACAGUGCAGAGAAU NM_152586
     526 C10ORF29 GCAGCACAGUCAAUGGUAA NM_152586
     527 C10ORF29 CCUCAGAAUUGGAGUCUCU NM_152586
     528 C10ORF29 GAGCAGGAGACCUCAGAAU NM_152586
     529 C10ORF29 AGACAUAUACCAAGAGAAG NM_152586
     530 C10ORF29 CAGGGUGGACUGAGAAGAA NM_152586
     531 C10ORF29 CCAAACAGGCUCUGCAGAU NM_152586
     532 C10ORF29 UCAGGUAGUCUAAGGAAGA NM_152586
     533 C12ORF6 UCACAAAGCAGAAGAAUUA NM_020367
     534 C12orf6 CAAGAUAGACUUUGCAGAA NM_020367
     535 C12ORF6 AGAUAGACUUUGCAGAAAU NM_020367
     536 C12orf6 CAAACAAUGAAGUGGAUGA NM_020367
     537 C12orf6 GUGAAUUUGUGGAAGCAAU NM_020367
     538 C12ORF6 ACAAUGAAGUGGAUGACAU NM_020367
     539 C12ORF6 GAGAUUACAUAAACGGAGA NM_020367
     540 C12ORF6 GUUUCUUGCUCGAGUGCUA NM_020367
     541 C12ORF6 GAUGAUACCUGGAACCCAA NM_020367
     542 C12orf6 CAGAAAUGAAGCAAAUGAA NM_020367
     543 C12ORF6 GCAGAAAUGAAGCAAAUGA NM_020367
     544 C12ORF6 GGAGCUAUGUGAAUUUAUA NM_020367
     545 C12orf6 CAGCAGUGAAUUUGUGGAA NM_020367
     546 C12orf6 GGGAGAAUGUGAAUACUCA NM_020367
     547 C12ORF6 CAACAAACAAUGAAGUGGA NM_020367
     548 C12orf6 GAAGCAAUCUGCAUUCAUA NM_020367
     549 C12ORF6 GCACAAUCAAACACAUGAA NM_020367
     550 C12ORF6 UCUGCAAAGAUGACAUAAA NM_020367
     551 C12ORF6 UGACAUAAAGCAUGGGAAC NM_020367
     552 C12ORF6 CAAAGAUGACAUAAAGCAU NM_020367
     553 C12orf6 GCAAAGAUGACAUAAAGCA NM_020367
     554 C12ORF6 GAACCUAUUUUGCUAGAGA NM_020367
     555 C12ORF6 GCCUCAGAUUAAUGAACAA NM_020367
     556 C12orf6 AAGAUAGACUUUGCAGAAA NM_020367
     557 C12orf6 CAAGGAAGCUUUAUUCUUU NM_020367
     558 C12ORF6 GACAUGGACACGUCAGAUA NM_020367
     559 C12ORF6 CCUGAGUACUUGAUAGACU NM_020367
     560 C12ORF6 CUUCCAAAGACGGGAGCUA NM_020367
     561 C12ORF6 GGAGUUCUUUUGCAGGAAA NM_020367
     562 C12ORF6 UUGCUAAUCUCUUUGGGAA NM_020367
     563 C14orf137 CAGACUGGCUGGAGAAAUU NM_023112
     564 C14orf137 GGGAAAACAUAGUGGAUGA NM_023112
     565 C14orf137 UGAGAUUCCUUGUGUGAAA NM_023112
     566 C14orf137 GAACAAUUAGCAACAGAAA NM_023112
     567 C14orf137 ACAAUUAGCAACAGAAAGA NM_023112
     568 C14orf137 GAGGAAAAGCUCAAGGUUA NM_023112
     569 C14orf137 GCCGAUAAACAUUGAUUAA NM_023112
     570 C14orf137 CCACCAUGGUCUAUGAGAA NM_023112
     571 C14orf137 AAACAUUCUCCUAGCAUUA NM_023112
     572 C14orf137 CAACAAUACCUAUGUGUCA NM_023112
     573 C14orf137 CCACCUGGACAGAGGCUUA NM_023112
     574 C140RF137 AGGAUUUACCGGAGGAAAA NM_023112
     575 C140RF137 UGGUGGAACUGGUAGAGAA NM_023112
     576 C14orf137 UCACUGGAUUAUUGGUUAA NM_023112
     577 C140RF137 GGGAGAUCUUCAAGUUCAA NM_023112
     578 C140RF137 GGAGGAAAAUCGAGGAACU NM_023112
     579 C14orf137 GGGAACAAUUAGCAACAGA NM_023112
     580 C14orf137 CCAAUAGCACUCUCAGAUA NM_023112
     581 C14orf137 UCUGGAUACUUCAAGGAUA NM_023112
     582 C14or1137 CCGUUUACCUGCUCUAUAA NM_023112
     583 C140RF137 UCUACAGGGCCUUGGGCUA NM_023112
     584 C140RF137 UGGAACUGGUAGAGAAGGA NM_023112
     585 C14orf137 GGACAAAUGCUUUCUAACU NM_023112
     586 C14orf137 GGACAGAGGCUUACAAGAC NM_023112
     587 C14orf137 GGCCAUACCCCUUAAAUAA NM_023112
     588 C14orf137 CCUCAGGACCUUCAAGAUU NM_023112
     589 C14orf137 CUGCAGGGAUUUACUGGAA NM_023112
     590 C14orf137 GGAACAAUUAGCAACAGAA NM_023112
     591 C14orf137 GGUUCUAACUUCAGCAUUC NM_023112
     592 C14orf137 CAAUAGCACUCUCAGAUAU NM_023112
     593 C15ORF16 GGAAAGACGACAACGAUAA NM_130901
     594 C15ORF16 GGGAAAGACGACAACGAUA NM_130901
     595 C15ORF16 AGGACAAGGAGAAGGAGAA NM_130901
     596 C15ORF16 GCAAGGAGAAGAAGGCCAA NM_130901
     597 C15ORF16 GGACCUGGUGUUACGGAAA NM_130901
     598 C15ORF16 AGGCAACAAUGGUGGCUUU NM_130901
     599 C15ORF16 GCGUGUACAGCGAGGAUUU NM_130901
     600 C15ORF16 AGAAGGAGAAGCAGCGCAA NM_130901
     601 C15ORF16 CCACGCAGCUGGUGCUCAA NM_130901
     602 C15ORF16 CCAAGCAGCCAGAGCGAGA NM_130901
     603 C15ORF16 GAACAGAGAGACCAGCAAA NM_130901
     604 C15ORF16 AGACCAGCAAAGAGAACAA NM_130901
     605 C15ORF16 CCACGUACCCGCAGCAGAA NM_130901
     606 C15ORF16 ACGGCAAGGACAAGGAGAA NM_130901
     607 C15ORF16 GAGACCUGCUGGAAGGCAA NM_130901
     608 C15ORF16 GUUAAGAGACUCAGGUGGA NM_130901
     609 C15ORF16 ACGAGGAGAUGAUCGGCUA NM_130901
     610 C15ORF16 GGCAAGAACGGCAAGGACA NM_130901
     611 C15ORF16 UGACGGAUUCUGAGCACAA NM_130901
     612 C15ORF16 CGCUAGAAGCCAAGCUGAA NM_130901
     613 C15ORF16 GGUUCUGGCCUAUGAUCAA NM_130901
     614 C15ORF16 GAGAAGCAGCGCAAGGAGA NM_130901
     615 C6.1A GAAUAAAUCAGGAGACAAA NM_024332
     616 C6.1A CCACAAAACUCAACAAUAA NM_024332
     617 C6.1A GGGAAAGAGUUUAUGAAUU NM_024332
     618 C6.1A AGGAAAGGUAUGAGAGAAU NM_024332
     619 C6.1A CAACAGAGGCAGAGAGGUU NM_024332
     620 C6.1A GGAACUAACUGUUGAUACA NM_024332
     621 C6.1A GAGAGAACCCAGAAAUAAA NM_024332
     622 C6.1A GAAUAAAGUCUGUAGUCUA NM_024332
     623 C6.1A AGGAAGAGCUUAUGCAAGA NM_024332
     624 C6.1A GCACAGAGAAGGAGGAAGU NM_024332
     625 C6.1A GCAAUUCAAUGGAGGAAGA NM_024332
     626 C6.1A GAGGAAGGACCGAGUAGAA NM_024332
     627 C6.1A GGACCUACAUACACACAAA NM_024332
     628 C6.1A CGUCAGAAUUGUUCACAUU NM_024332
     629 C6.1A AAGAUAAGCUACAGAUUGA NM_024332
     630 C6.1A AGAAGGAAGAGGAAAGGUA NM_024332
     631 C6.1A GAAGAUAAGAACACAAAGA NM_024332
     632 C6.1A GCAUAUACUGGAACUGAAA NM_024332
     633 C6.1A CAGAGAAGGAGGAAGUAAU NM_024332
     634 C6.1A AGGACAGACUGGAGCAAAA NM_024332
     635 C6.1A GCAGAGGAACAGACACAUA NM_024332
     636 C6.1A GAAUAGCCUUUCUGACAAA NM_024332
     637 C6.1A GAACAGACACAUACGUCAA NM_024332
     638 C6.1A GAGGAAAGGUAUGAGAGAA NM_024332
     639 C6.1A AGGUAUGAGAGAAUCGAAA NM_024332
     640 C6.1A GACAUUCAUUCCAGAGAAA NM_024332
     641 C6.1A GGAAGGACCGAGUAGAAAU NM_024332
     642 C6.1A GUACAGCCACUCUGGGAAA NM_024332
     643 C6.1A GGGCCAGAAGGAAGAGGAA NM_024332
     644 C6.1A UAGAAUAAAUCAGGAGACA NM_024332
     645 CEZANNE CAAAGAUGAUAGUGACAAU NM_020205
     646 CEZANNE GCACUGAGACACUGGAGAA NM_020205
     647 CEZANNE UGGCAGAACAGAAGCAGAA NM_020205
     648 CEZANNE CAGCUGAGUCUGUUGGUAA NM_020205
     649 CEZANNE CUGAGACACUGGAGAAGAA NM_020205
     650 CEZANNE GGGCAAGGAGGCUAAACAA NM_020205
     651 CEZANNE GCAGAGAGGAAGAUCAUGA NM_020205
     652 CEZANNE GGAAAGAAUUGGGAUGUGA NM_020205
     653 CEZANNE AGAAGGGAGUUGAGAAGGA NM_020205
     654 CEZANNE CAAAGGAGGUUCUCAGUCA NM_020205
     655 CEZANNE CCGAUUGGCCAGUGUAAUU NM_020205
     656 CGI-77 CAUUAUAGUUGGUGAAGAA NM_016023
     657 CGI-77 CAGAAGAAUUUCAGAAGUA NM_016023
     658 CGI-77 ACAGAAACAUAGAGAGGAA NM_016023
     659 CGI-77 GAGAGGAACUGGAGCAAUU NM_016023
     660 CGI-77 ACAUAUGGAAAGUGAGAAA NM_016023
     661 CGI-77 GCACAAAAGAGACGGGAAA NM_016023
     662 CGI-77 ACAAACACCAAUAGAGAUA NM_016023
     663 CGI-77 GUAUAAAGCCAUUGAAGAU NM_016023
     664 CGI-77 GCAAAGAGAAGAAGGAGUU NM_016023
     665 CGI-77 GGAAAAGGAGCGAGAAGAA NM_016023
     666 CGI-77 CGAAGAGCUUGAUGAGGAA NM_016023
     667 CGI-77 GCAGCUAGACAGUUAGAAA NM_016023
     668 CGI-77 GAAGAUCAACUGAAAGAAA NM_016023
     669 CG1-77 CCGAAGAGCUUGAUGAGGA NM_016023
     670 CGI-77 AAGAAAUGGAACAGAAACA NM_016023
     671 CGI-77 AAAGAAAGCUGCAUUGGAA NM_016023
     672 CGI-77 CAGAAGUACUGUGAAGAUA NM_016023
     673 CGI-77 GAAAUGGAACAGAAACAUA NM_016023
     674 CGI-77 GGAGCGAGAAGAACGGAUA NM_016023
     675 CGI-77 GAGACAUGCAUAUGGCUUA NM_016023
     676 CGI-77 AGACAUAUGGAAAGUGAGA NM_016023
     677 CGI-77 GCUCAAAUAUUGGCAGCUA NM_016023
     678 CGI-77 GGUAUUGACCGAAGAGCUU NM_016023
     679 CGI-77 AGAAGGAGUUGCAAGCCAA NM_016023
     680 CGI-77 ACUUGGUGCUUGAGAAUCA NM_016023
     681 CGI-77 CAAUUGAAGCUGACUACUA NM_016023
     682 CGI-77 CCAGACAUAUGGAAAGUGA NM_016023
     683 CGI-77 GCUGAGAAGGCAUCGCAAA NM_016023
     684 CGI-77 UGACUACUAAGGAGAAUAA NM_016023
     685 CGI-77 CUAAGGAGAAUAAGAUAGA NM_016023
     686 COPS5 GGACAUACCCAAAGGGCUA NM_006837
     687 COPS5 CUACAAACCUCCUGAUGAA NM_006837
     688 COPS5 UGGAAUAAAUACUGGGUGA NM_006837
     689 COPS5 UGAAAGACAGUAUGAGAAA NM_006837
     690 COPS5 GGACUAAGGAUCACCAUUA NM_006837
     691 COPS5 ACAAAUACGACAAGAAACA NM_006837
     692 COPS5 GGUGAAACCAUGAUCAUUA NM_006837
     693 COPS5 CCGAAAAUCAGAAGACAAA NM_006837
     694 COPS5 GGAUUGAUGUCUCAGGUUA NM_006837
     695 COPS5 AACAAUAUCCGCAGGGAAA NM_006837
     696 COPS5 CUACAAAUACGACAAGAAA NM_006837
     697 COPS5 GUGAAUACGUUGAGUUCUU NM_006837
     698 COPS5 UAGGAAAGGUGGAUGGUGA NM_006837
     699 COPS5 GAACAAUAUCCGCAGGGAA NM_006837
     700 COPS5 AUACAUGGCUGCAUACAUA NM_006837
     701 COPS5 ACAAGAAACAGCAGCAAGA NM_006837
     702 COPS5 CUGAAAGACAGUAUGAGAA NM_006837
     703 COPS5 GAGAAGUACUUUACCUGAA NM_006837
     704 COPS5 AAGAAACAGCAGCAAGAAA NM_006837
     705 COPS5 AGAAACAGCAGCAAGAAAU NM_006837
     706 COPS5 CUUGAGCUGUUGUGGAAUA NM_006837
     707 COPS5 GGAGUUUCAUGUUGGGUUU NM_006837
     708 COPS5 CGGCGAAGCCCUGGACUAA NM_006837
     709 COPS5 CCUUAGAAGUCUCAUAUUU NM_006837
     710 COPS5 UAGAAACGCAUGACCGAAA NM_006837
     711 COPS5 CCUGAAAGACAGUAUGAGA NM_006837
     712 COPS5 GAAUAAAUACUGGGUGAAU NM_006837
     713 COPS5 AUGAAUACAUGGCUGCAUA NM_006837
     714 COPS5 GGAAUAAAUACUGGGUGAA NM_006837
     715 COPS5 GGUGAUUGAUCCAACAAGA NM_006837
     716 CYLD UGAAAGAAUUGGAGUGAUA NM_015247
     717 CYLD GGAAGAAGGUCGUGGUCAA NM_015247
     718 CYLD GGGUAGAACCUUUGCUAAA NM_015247
     719 CYLD CUUCAAGAAUGCAGCGUUA NM_015247
     720 CYLD CCGAAGAGGCUGAAUCAUA NM_015247
     721 CYLD CAUAAUAAACCAAAGGCUA NM_015247
     722 CYLD GCUCAUUGGCUGAAGUUAA NM_015247
     723 CYLD AAAGGAUGUUGUAGAGAUA NM_015247
     724 CYLD AUGUAUGAGUGUAGAGAAU NM_015247
     725 CYLD CCGGAUAUGUGGAGGGCUU NM_015247
     726 CYLD CUGUAAUGGAAGAGCUAAA NM_015247
     727 CYLD GUGAAGUAGUAGAAGAAAA NM_015247
     728 CYLD CGAAGAGGCUGAAUCAUAA NM_015247
     729 CYLD UGGCUGAAGUUAAGGAGAA NM_015247
     730 CYLD AAGUAAAGGCCUCCAAAUA NM_015247
     731 CYLD AGGAUGUUGUAGAGAUAAA NM_015247
     732 CYLD UGGGAGAGAUGGAUAUUUA NM_015247
     733 CYLD CAGAAACAGACUAAGUAAA NM_015247
     734 CYLD GAAAAGAAAGCUUAGGAUA NM_015247
     735 CYLD GAGUUGAAUUGCUGGAAGA NM_015247
     736 CYLD GGAUUUACCUCUGAAGAAA NM_015247
     737 CYLD GGGAAGUAUAGGACAGUAU NM_015247
     738 CYLD CACAAUGAGUUCAGGCUUA NM_015247
     739 CYLD AGAGAUAUCUACAGACUUU NM_015247
     740 CYLD UAGUGAAACCCAAGAGCUA NM_015247
     741 CYLD AAGAAGGCUUGGAGAUAAU NM_015247
     742 CYLD CAGAGUUACUUUUGGCAAU NM_015247
     743 CYLD AAGUGAAGUAGUAGAAGAA NM_015247
     744 CYLD AGAAAAUACUCCACCAAAA NM_015247
     745 CYLD AGAUAAUGAUUGGGAAGAA NM_015247
     746 DKFZP434D0127 GAAGGAUACUAAUGGGUAA NM_032147
     747 DKFZP434D0127 CCAACGAGUUACUGAGAAU NM_032147
     748 DKFZP434D0127 GGGACUGACCUCUGAGUUA NM_032147
     749 DKFZP434D0127 UCACAGAAGCCCAGAAACA NM_032147
     750 DKFZP434D0127 AAACGAAGGCCAAUAGUAA NM_032147
     751 DKFZP434D0127 GUAGAAAGAUGGAACUUAU NM_032147
     752 DKFZP434D0127 UGAAAGAGGUUGAUUUGUA NM_032147
     753 DKFZP434D0127 GAAAGAGGUUGAUUUGUAA NM_032147
     754 DKFZP434D0127 GGAAUUUCUUUGUGAACUU NM_032147
     755 DKFZP434D0127 CCAGAAACAACUUAUGAUA NM_032147
     756 DKFZP434D0127 GCAUUUGCCCUGAUGGAAA NM_032147
     757 DKFZP434D0127 AAUAGUAACUCCUGGUGUA NM_032147
     758 DKFZP434D0127 GUAAUAACCGAGAGAAGAU NM_032147
     759 DKFZP761A052 CAACACAUUCCAUGGGAUA NM_017602
     760 DKFZP761A052 UGUUUUAGCUCUUGCCAAA NM_017602
     761 DKFZP761A052 UGCAUGAGGUUGUGCGAAA NM_017602
     762 DKFZP761A052 UGGACUAUCUGAUGAAGAA NM_017602
     763 DKFZP761A052 CCACCUACAUUAACAGGAA NM_017602
     764 DKFZP761A052 CGGAAUAUCCACUAUAAUU NM_017602
     765 DKFZP761A052 UGAUGAAGAAUGCCAUAAA NM_017602
     766 DKFZP761A052 CAACAGUGAGGACGAGUAU NM_017602
     767 DKFZP761A052 CAGCACGCAUCGAGGCUAU NM_017602
     768 DKFZP761A052 GAGCAGUCUCUGAUGAAGA NM_017602
     769 DKFZP761A052 GCUACAACAGUGAGGACGA NM_017602
     770 DKFZP761A052 ACAGUGAGGACGAGUAUGA NM_017602
     771 DKFZP761A052 CAACAGGAAUACCUAGACA NM_017602
     772 DKFZP761A052 CAGCAAGCGGCGACGUCAA NM_017602
     773 DKFZP761A052 CAGGAGCAUUGGUUUGAAA NM_017602
     774 DKFZP761A052 AAAACAAAGUGCACAGAGA NM_017602
     775 DKFZP761A052 CUGUGGAGGUGUACCAGUA NM_017602
     776 DKFZP761A052 GCAUGGACUAUCUGAUGAA NM_017602
     777 DKFZP761A052 CUGCAGUGGUUGCGGGAUC NM_017602
     778 DKFZP761A052 GAUGCUAGAAGACAAGAAA NM_017602
     779 DKFZP761A052 ACAUUAACAGGAAGCGGAA NM_017602
     780 DKFZP761A052 CUAGACAGUAUGAAGAAAA NM_017602
     781 DKFZP761A052 CCGACUACUUCUCCAACUA NM_017602
     782 DKFZP761A052 CCUAGACAGUAUGAAGAAA NM_017602
     783 DKFZP761A052 AUUAACAGGAAGCGGAAAA NM_017602
     784 DKFZP761A052 GUUAGCUACCAUCGGAAUA NM_017602
     785 DKFZP761A052 CUGACCAGGUGUAUGGAGA NM_017602
     786 DKFZP761A052 AGUCAUGGAUUGAACAGCA NM_017602
     787 DKFZP761A052 UGAUGAAGAAUGCCGACUA NM_017602
     788 DKFZP761A052 GCAACCACAUUGAGAUGCA NM_017602
     789 E2-EPF CCUCCAACUCUGUCUCUAA NM_014501
     790 E2-EPF GGCACUGGGACCUGGAUUU NM_014501
     791 E2-EPF ACAAGGAGGUGACGACACU NM_014501
     792 E2-EPF GGAGAACUACGAGGAGUAU NM_014501
     793 E2-EPF UCAAGUGCCUGCUGAUCCA NM_014501
     794 E2-EPF GACACGUACUGCUGACCAU NM_014501
     795 E2-EPF CCGCAGCCAUGAACUCCAA NM_014501
     796 E2-EPF GCUACUUCCUGACCAAGAU NM_014501
     797 E2-EPF GGAGGUCUGUUCCGCAUGA NM_014501
     798 E2-EPF ACUGGGACCUGGAUUUGUU NM_014501
     799 E2-EPF UCAAGGUCUUUCCCAACGA NM_014501
     800 E2-EPF GCACUGGGACCUGGAUUUG NM_014501
     801 E2-EPF GUACUGCUGACCAUCAAGU NM_014501
     802 FBXO7 GGAAAGAACUGUACAGGAA NM_012179
     803 FBXO7 GUGCUGAUCUCGAGUGUUA NM_012179
     804 FBXO7 CCAGAAACCUUUUAAGAGA NM_012179
     805 FBXO7 GAAUAUGACCAAAGACCAA NM_012179
     806 FBXO7 AGGAAGAGGCACAUACAAA NM_012179
     807 FBXO7 UGAGAUUAGAAGUGUGAAA NM_012179
     808 FBXO7 ACAAAGAUCUUCAGAAACU NM_012179
     809 FBXO7 CCGGAGAAGUGGAAGUUGA NM_012179
     810 FBXO7 GCGUGAUUUUCGAGACAAU NM_012179
     811 FBXO7 CCUGUGUGCCUUUGGGAAA NM_012179
     812 FBXO7 ACUCCAGAAUAAUGAGCAA NM_012179
     813 FBXO7 GCACAUACAAAGAAAAGAA NM_012179
     814 FBXO7 CCUUGAUAGUGUUGAUACA NM_012179
     815 FBXO7 CAUUAGAGACCUUGUAUCA NM_012179
     816 FBXO7 CCGAAAGGGCGGUUUGUGA NM_012179
     817 FBXO7 AGACUAGCAUACAGGAUGA NM_012179
     818 FBXO7 UAGCAUACAGGAUGAACAA NM_012179
     819 FBXO7 GAAACCUGAUUGUUGUAAA NM_012179
     820 FBXO7 CAAAGAGAAACUAGGGGAA NM_012179
     821 FBXO7 GAACCUACCAGAUGUAUUU NM_012179
     822 FBXO7 AAAGAACUGUACAGGAAGA NM_012179
     823 FBXO7 CUGAGUCAAUUCAAGAUAA NM_012179
     824 FBXO7 CUGUACAGGAAGAGGCACA NM_012179
     825 FBXO7 GUGAAUCGGUGGAAGGGCA NM_012179
     826 FBXO7 CCACAGAUUCAGAGCAUUC NM_012179
     827 FBXO7 GAAUGACGACAGUAUGUUA NM_012179
     828 FBXO7 GAUGAACAACCAAGUGAUU NM_012179
     829 FBXO7 AGACACAGAUUGGAAAGAA NM_012179
     830 FBXO7 AGAGGCACAUACAAAGAAA NM_012179
     831 FBXO7 GGGUGUAUAAGCUGCAGUA NM_012179
     832 FBXO8 CGAAAGAACAGGAAGGAUU NM_012180
     833 FBXO8 CCAAAUGCACUGAGAGAAU NM_012180
     834 FBXO8 UGGCAAGGGUUGUGCAAAU NM_012180
     835 FBXO8 GGAGAAUGGCUGCGAGCAA NM_012180
     836 FBXO8 GGAGAGAUGUCUUGGAUGA NM_012180
     837 FBXO8 CCAAGGAGGCAUUGACAUA NM_012180
     838 FBXO8 UGAAGAGCGUGGAGAGUAU NM_012180
     839 FBXO8 GAAAUCAGUUCUUGCCAAA NM_012180
     840 FBXO8 CAGAUGAGGGAGUGAACUA NM_012180
     841 FBXO8 GCAAGGAAAUCGAAAGAAC NM_012180
     842 FBXO8 GAACCCACCUUUAGGAUUU NM_012180
     843 FBXO8 CGCCAAAGGAAAUAGCAAA NM_012180
     844 FBXO8 GAUGAUUCGCCAAAGGAAA NM_012180
     845 FBXO8 GCUGUUAACGAAUGAUAAA NM_012180
     846 FBXO8 AAGCAGAGCCACUGUCUAA NM_012180
     847 FBXO8 GCAUUAGAUUUGCCUGAAA NM_012180
     848 FBXO8 GAAGAGUGUUGAUUUGUUA NM_012180
     849 FBXO8 GAGAAUCUAUCUUGAUGAA NM_012180
     850 FBXO8 UGUGGAGAGUGGUCAGAAA NM_012180
     851 FBXO8 GGUCAAGGGUUGUGGAGAG NM_012180
     852 FBXO8 GGACAAGUAAAGUUUGAAU NM_012180
     853 FBXO8 AGGCAAGGAAAUCGAAAGA NM_012180
     854 FBXO8 CACCAAUCAUCGUAAACAA NM_012180
     855 FBXO8 CAAUAAGAACCCACCUUUA NM_012180
     856 FBXO8 GCCCUCAUGUGAAGAAUAA NM_012180
     857 FBXO8 GGUAAGUCCUUUAAACCAU NM_012180
     858 FBXO8 UGAUUUAAUGCGAGAACUU NM_012180
     859 FBXO8 CCAAGGACUUAGCAGAUAU NM_012180
     860 FBXO8 CAGGAAGGAUUCAUUAAUU NM_012180
     861 FBXO8 GAACAGGAAGGAUUCAUUA NM_012180
     862 FBXW3 GGGAAGAGUACAAGGACAA NM_012165
     863 FBXW3 GGGCAAGGCGGAAGAGGAA NM_012165
     864 FBXW3 GGAUGCAGCUAGAGGAUGA NM_012165
     865 FBXW3 GAAAGAGGCACAAGGAAGA NM_012165
     866 FBXW3 CAUCCAGACUGAAGACCAA NM_012165
     867 FBXW3 GGCACAAGGAAGAAAGUAU NM_012165
     868 FBXW3 GGGAAAGAGGCACAAGGAA NM_012165
     869 FBXW3 UGACACACUUGGACAGAGA NM_012165
     870 FBXW3 GGACAAGGAGGUCAGGUUU NM_012165
     871 FBXW3 GGAAGAUUGGCCUUGGUAA NM_012165
     872 FBXW3 GGAUGAUGCUUUGUACAUA NM_012165
     873 FBXW3 GAAGGAAGGGGAAGAGGAA NM_012165
     874 FBXW3 ACACUUGGACAGAGACUUU NM_012165
     875 FBXW3 CAGUGAAGGUGUCUCAGAA NM_012165
     876 FBXW3 GAUGAUGCUUUGUACAUAU NM_012165
     877 FBXW3 GAAGAAAGUAUGGGAAGGA NM_012165
     878 FBXW3 CCGCACCAGUGUCCGGAAA NM_012165
     879 FBXW3 AGGAGGAGAUGGGAAGAUU NM_012165
     880 FBXW3 UGAACUGUGUGGAUUGCAA NM_012165
     881 FBXW3 GAAGAUUGGCCUUGGUAAG NM_012165
     882 FBXW3 AAGUGGAGAUGCAGUCAGA NM_012165
     883 FBXW3 GAACUGUGUGGAUUGCAAA NM_012165
     884 FBXW3 GCACAAGGAAGAAAGUAUG NM_012165
     885 FBXW3 GGAGGUGAACUGUGUGGAU NM_012165
     886 FBXW3 GUACAUAUCCCAGGCUAAU NM_012165
     887 FBXW3 CGGGAAAGAGGCACAAGGA NM_012165
     888 FBXW3 GAGGUGAACUGUGUGGAUU NM_012165
     889 FBXW3 GGCGAGAGGCAUCAUCAAA NM_012165
     890 FBXW3 CCAGAUGGUGCCAGCUUGA NM_012165
     891 FBXW3 GAUUGGCCUUGGUAAGAUU NM_012165
     892 FLJ11807 AGACCAAGAUCCAGAAAGA NM_024954
     893 FLJ11807 CGGGAUGAGUUCUGGGACA NM_024954
     894 FLJ11807 CUGAAGAAAGAGCGGCUUA NM_024954
     895 FLJ11807 GCAAUGAGCCCCUGAAGAA NM_024954
     896 FLJ11807 GCUACGAUGAGCUGGGCAA NM_024954
     897 FLJ11807 GAGCAAACGGGAUGAGUUC NM_024954
     898 FLJ11807 CCAGAAAGAUUUUGUCAUC NM_024954
     899 FLJ11807 ACCAAGAUCCAGAAAGAUU NM_024954
     900 FLJ11807 CCAAGAUCCAGAAAGAUUU NM_024954
     901 FLJ11807 CUUGCAACCUUGUCAGAGA NM_024954
     902 FLJ11807 CUUAAGUGGAAGAGCGACU NM_024954
     903 FLJ11807 GGAGCAAACGGGAUGAGUU NM_024954
     904 FLJ11807 CAGCACAAUUGGCAGAGAU NM_024954
     905 FLJ11807 CCGGGAAGCUGCUCACAGA NM_024954
     906 FLJ11807 CAAGCGAGCAGGACGCAAU NM_024954
     907 FLJ11807 UGAAGAAAGAGCGGCUUAA NM_024954
     908 FLJ11807 ACACGGAGGAGGAGAGCCU NM_024954
     909 FLJ11807 AAGAGCGGCUUAAGUGGAA NM_024954
     910 FLJ11807 GCUACCAGCUGCCCAUCUA NM_024954
     911 FLJ11807 UCAUCCAGGUCAUCAUCAA NM_024954
     912 FLJ11807 GGAGACCAAGAUCCAGAAA NM_024954
     913 FLJ11807 AAGAAAGAGCGGCUUAAGU NM_024954
     914 FLJ11807 GCACAAUUGGCAGAGAUGA NM_024954
     915 FLJ11807 UUGCAACCUUGUCAGAGAA NM_024954
     916 FLJ11807 GCCGUGAGUUCCCGCUGAA NM_024954
     917 FLJ11807 CAUGGCACCCUCUGUGAAU NM_024954
     918 FLJ11807 GCAAACGGGAUGAGUUCUG NM_024954
     919 FLJ11807 CCACACACGGGCCUUGCAA NM_024954
     920 FLJ11807 AGAAAGAGCGGCUUAAGUG NM_024954
     921 FLJ11807 ACACGGGCCUUGCAACCUU NM_024954
     922 FLJ12552 GGUGAUUGAUGUAGGGAAA NM_022832
     923 FLJ12552 GGGAGAAUGUGUUGGCAUA NM_022832
     924 FLJ12552 AGUCAUAAGUUCAGGAUAU NM_022832
     925 FLJ12552 GGGAAAUGUUUGUACUAUA NM_022832
     926 FLJ12552 GGAACUGAUUCAUAAGAAA NM_022832
     927 FLJ12552 CAGAAGAAUUGGGUAGAUA NM_022832
     928 FLJ12552 GCAGGAUGCUCAUGAAUUU NM_022832
     929 FLJ12552 GGGUGCAAGUAUAGCUUUA NM_022832
     930 FLJ12552 CCAUGAAACUUACGCAGUA NM_022832
     931 FLJ12552 UGAGUAACCUCGAUAUUAA NM_022832
     932 FLJ12552 UUGGAAGGCUGUAGAAGUA NM_022832
     933 FLJ12552 CAGCAUUCAUGGAUAGAUU NM_022832
     934 FLJ12552 GCCCAAAGGCUCUGCAGUA NM_022832
     935 FLJ12552 CAAGAGAGUAACUGAAAGA NM_022832
     936 FLJ12552 GAGCAGAAUACAUCCAUUA NM_022832
     937 FLJ12552 GAAACAGGGUGAAGAGAAG NM_022832
     938 FLJ12552 CAAACAAGAAGCCCAGAAA NM_022832
     939 FLJ12552 GUAGCAAAGAUGAAGAUUU NM_022832
     940 FLJ12552 UGGAGCAGCUGCACAGAUA NM_022832
     941 FLJ14981 GGAGUGAGGUCGUGGGUUA NM_032868
     942 FLJ14981 CCAUCGAAGAGGAGAUCUA NM_032868
     943 FLJ14981 UGGAAGAGGAGGAGGAGGA NM_032868
     944 FLJ14981 GGAAGCGGGUGGACAGCAA NM_032868
     945 FLJ14981 GGGCUCAGGUAAUAAAGAA NM_032868
     946 FLJ14981 AGGAGUUGCUGAUGGAAGA NM_032868
     947 FLJ14981 GGGUCUCAGCAGAGGACAA NM_032868
     948 FLJ14981 GGACAAGAGUCGGAGACCA NM_032868
     949 FLJ14981 AGAAACUGGACAAGUACAA NM_032868
     950 FLJ14981 GCAAGAAGCUGGUGAACCC NM_032868
     951 FLJ14981 UGAUGGAAGAGGAGGAGGA NM_032868
     952 FLJ14981 CGGAUGGGCUCAGGUAAUA NM_032868
     953 FLJ14981 GGGCCUCUGUCAAGUACAA NM_032868
     954 FLJ14981 AGUACAAAGGCCAGAAACU NM_032868
     955 FLJ14981 CUUAAGAUCUCCUUGGCCA NM_032868
     956 FLJ14981 AGUCCAAGAUCUCGCCUUU NM_032868
     957 FLJ14981 UCAACAAGUUCCAGCCGUU NM_032868
     958 FLJ14981 CAGCCAGACGGAAGGAUCA NM_032868
     959 FLJ14981 CCGAGAUGCUGCUGGUGGA NM_032868
     960 FLJ14981 AGGAGGAGUUGCUGAUGGA NM_032868
     961 FLJ14981 CUGCAGGGCUCCAGCAAUG NM_032868
     962 FLJ14981 AGAUGCUGCUGGUGGAGUU NM_032868
     963 FLJ14981 CAAGUACAAAGGCCAGAAA NM_032868
     964 FLJ14981 ACGCACAGAUGGACUACCA NM_032868
     965 FLJ14981 UCGACAAGCUUAAGAUCUC NM_032868
     966 FLJ14981 ACACCUACCUCGACAAGCU NM_032868
     967 FLJ14981 GGUGGAAGUAACAUCCUUU NM_032868
     968 FLJ14981 GGAGCCAGGAGCACACCUA NM_032868
     969 FLJ14981 GGAUGGGCUCAGGUAAUAA NM_032868
     970 FLJ14981 GCUCAGGUAAUAAAGAAAC NM_032868
     971 FLJ20113 ACGAUAUCCUCUACAAAUA NM_017670
     972 FLJ20113 GAUCAAGGACCUCCACAAA NM_017670
     973 FLJ20113 GGUUGUAAAUGGUCCUAUU NM_017670
     974 FLJ20113 AGGAGUAUGCUGAAGAUGA NM_017670
     975 FLJ20113 CAAUUGAGGAUUUCCACAA NM_017670
     976 FLJ20113 GCAAGGAGAGCGACCACAU NM_017670
     977 FLJ20113 CCUAUCUACUCCUGAGCUU NM_017670
     978 FLJ20113 UGGAGGCACUGCUGGAUGA NM_017670
     979 FLJ20I13 UCUAUGGGAUCCUGGAAGU NM_017670
     980 FLJ20113 ACUACGAUAUCCUCUACAA NM_017670
     981 FLJ20113 GACCGAAUUCAGCAAGAGA NM_017670
     982 FLJ20113 CAACAUCUAUCAACAGAAG NM_017670
     983 FLJ20113 GCCAGGCGCUAGACAUGUA NM_017670
     984 FLJ20727 AGAUAAAGAUGGUGAACAA NM_017944
     985 FLJ20727 CGAGAGAAGCUUAGUGAAA NM_017944
     986 FLJ20727 GGAUAAGACAUUAAAGGAA NM_017944
     987 FLJ20727 CAACAUGUCAGCAGGAUAA NM_017944
     988 FLJ20727 GGAUGGAGCACCAAAUAAA NM_017944
     989 FLJ20727 GAGAAGUGAUGGUGAAAGU NM_017944
     990 FLJ20727 GGAAUCAUGUUGUGCACUA NM_017944
     991 FLJ20727 CCAUUUACCUGCUGAAACA NM_017944
     992 FLJ20727 CAGUAGAAAUGGCUUAUAA NM_017944
     993 FLJ23251 GGGAGAAGGACAAGGCAUA NM_024818
     994 FLJ23251 GAACAUACUCUGAGGAACA NM_024818
     995 FLJ23251 GCAUACAGCUUAUUGAUUA NM_024818
     996 FLJ23251 CAAAGGAUAUUGUAGGAUA NM_024818
     997 FLJ23251 AGACUUACCUGAAGGAAUU NM_024818
     998 FLJ23251 GCUCAGAGGUGGUGGAUUC NM_024818
     999 FLJ23251 AAGAAAACCUCAAAGGAUA NM_024818
    1000 FLJ23251 GAUUAGAGCUGGCAAGCAU NM_024818
    1001 FLJ23251 GGACAAAUCUGAUCCUGUA NM_024818
    1002 FLJ23251 GGACAUGGACAAUAAAGUA NM_024818
    1003 FLJ23251 AGUAUAUGAUCCUCAGAUA NM_024818
    1004 FLJ23251 UGUAAGCGACUAUGAGAAA NM_024818
    1005 FLJ23251 GGACAAGGCAUACAGCUUA NM_024818
    1006 FLJ23251 GGAACUAGCCAAUAUGAAU NM_024818
    1007 FLJ23251 GAAAUAACAUCCUCUCAAA NM_024818
    1008 FLJ23251 AUGAGGACAUGGACAAUAA NM_024818
    1009 FLJ23277 CAGAAAGGAUGGAGAACAU NM_032236
    1010 FLJ23277 CAAUAAAGCAUGAAAGUGA NM_032236
    1011 FLJ23277 GGUGAAUGGUAUAAGUUUA NM_032236
    1012 FLJ23277 CAUCAGACUUGAAGGAAAU NM_032236
    1013 FLJ23277 AGACGAAACUGCAAAGGAA NM_032236
    1014 FLJ23277 GUAGAAGAGUGGCGGAAAU NM_032236
    1015 FLJ23277 CCAAAUGCCUUAUGUCAAA NM_032236
    1016 FLJ23277 GAAAUUACAACUAGGGAUU NM_032236
    1017 FLJ23277 GGAAGAAAAUGAAGCCUUA NM_032236
    1018 FLJ23277 GCAAGAUGGUGAUGCAGAA NM_032236
    1019 FLJ23277 GAUGUAAACCCUUCAGAAA NM_032236
    1020 FLJ23277 GAGGAGUGGUGUAUUGAAA NM_032236
    1021 FLJ23277 GAACAAAGCAACGGAAAGA NM_032236
    1022 FLJ23277 GCAUAUUGCGUCUGAAGAA NM_032236
    1023 FLJ25157 GCAAGAACCAGAAAGAGAA NM_152653
    1024 FLJ25157 AGAAGGAACUUGCAGAAAU NM_152653
    1025 FLJ25157 CCAAAGGAGACAACAUUUA NM_152653
    1026 FLJ25157 CUGUAGUGCUGGACCCAAA NM_152653
    1027 FLJ25157 ACAAAGAGUUGAUGACAGU NM_152653
    1028 FLJ25157 CCAACAGAGCAGAGCAUGA NM_152653
    1029 FLJ25157 GAAUGGAGGUCAACUAUAU NM_152653
    1030 FLJ25157 CGAUGGAGAUCAACGUGAA NM_152653
    1031 FLJ25157 CCACACAGUACAUGACCAA NM_152653
    1032 FLJ25157 GCUGCUAAAUUGUCAACUA NM_152653
    1033 FLJ25157 GAUGACAGUCCAAGCACUA NM_152653
    1034 FLJ25157 GGAGACAACAUUUAUGAAU NM_152653
    1035 FLJ25157 CUAAAAGAAUUCAGAAGGA NM_152653
    1036 FLJ25157 GAAGGAACUUGCAGAAAUC NM_152653
    1037 FLJ25157 CAGAAGGAACUUGCAGAAA NM_152653
    1038 FLJ25157 UCAGCAAGAACCAGAAAGA NM_152653
    1039 FLJ25157 CCACUGAGGCACAAAGAGU NM_152653
    1040 FLJ25157 UCAGAAGGAACUUGCAGAA NM_152653
    1041 FLJ25157 CAAGAAUCUAUCACUGUAA NM_152653
    1042 FLJ25157 GAUGGAGAUCAACGUGAAA NM_152653
    1043 FLJ25157 UAUGAAUGGAGGUCAACUA NM_152653
    1044 FLJ25157 GCACAAAGAGUUGAUGACA NM_152653
    1045 FLJ25157 UCACCAGACUAUCCGUUUA NM_152653
    1046 FLJ25157 AGAUCAACGUGAAAGUGUU NM_152653
    1047 FLJ25157 ACAAGAAUCUAUCACUGUA NM_152653
    1048 FLJ25157 CAACUGGAGUCCGGCUUUA NM_152653
    1049 FLJ25157 GAGAUCAACGUGAAAGUGU NM_152653
    1050 FLJ25157 UGAAUGGAGGUCAACUAUA NM_152653
    1051 FLJ25157 GCAAAACCGCUGCUAAAUU NM_152653
    1052 FLJ25157 CAACUAGUGCUAAAAGAAU NM_152653
    1053 FLJ30626 GAAGAUGGUUGCAGAGGAA NM_153210
    1054 FLJ30626 GCAAGGAAACUCAAGGAAA NM_153210
    1055 FLJ30626 CAAGGAAACUCAAGGAAAA NM_153210
    1056 FLJ30626 GUGAAAGGCAGAAGCAUUA NM_153210
    1057 FLJ30626 CGUGAAAGGCAGAAGCAUU NM_153210
    1058 FLJ30626 ACAUCAAGCUUCCCAGAAA NM_153210
    1059 FLJ30626 CAAUAUAGAUCUUCCUUGA NM_153210
    1060 FLJ30626 GAGAACAGGAGGAAUGAGA NM_153210
    1061 FLJ30626 GCACUGGGCAUGUCACAAA NM_153210
    1062 FLJ30626 GGCAGAAGCAUUAGCAUGA NM_153210
    1063 FLJ30626 GGGCUUAUAUCCUGUUCUA NM_153210
    1064 FLJ30626 AGAGGGAGAUAAUGUGUAU NM_153210
    1065 FLJ30626 GAAGGCUGCUCGAGGAACA NM_153210
    1066 FLJ30626 GGAUGGUGAAGCUGAGUUU NM_153210
    1067 FLJ30626 GCAGCAGGCUAGCAGGUUU NM_153210
    1068 FLJ30626 GCACUGCGGGUGAGGAUGA NM_153210
    1069 FLJ37318 GCUCAGUCGUCAAUGUAAA NM_152586
    1070 FLJ37318 GGGAAGAGUCCACUGAACA NM_152586
    1071 FLJ37318 GGAAGAGUCCACUGAACAU NM_152586
    1072 FLJ37318 AAAGAUGAAUCCAUGGUAU NM_152586
    1073 HP43.8KD GGUAAUAGAUUUUCCAGAA NM_032557
    1074 HP43.8KD GGAAGUAAUUUUCGUGCUA NM_032557
    1075 HP43.8KD CAAAAUGCAUCAUGGAAAA NM_032557
    1076 HP43.8KD CCAGUGAAGAGAAGAUUAA NM_032557
    1077 HP43.8KD CCAAUGGAUUUGAUGACAA NM_032557
    1078 HP43.8KD UGUUUGAACUGCAGGAGUA NM_032557
    1079 HP43.8KD AAAGAGAGCUGCGGGAAUA NM_032557
    1080 HP43.8KD UAUACAAGAUGGUGGUCUA NM_032557
    1081 HP43.8KD UUAGAAGAAUUCAGAGGAA NM_032557
    1082 HP43.8KD GAGUAUAUCUUCUGUGGUU NM_032557
    1083 HP43.8KD GAUCAGAAGUAUCAUGUGA NM_032557
    1084 HP43.8KD AGACCCACCUCUACAGAAA NM_032557
    1085 HP43.8KD GAUGAAGCUUCCUGCACAA NM_032557
    1086 HP43.8KD UCACACAAGCCUUCUGAAA NM_032557
    1087 HP43.8KD AGAAGAACCAGUAGUUUAU NM_032557
    1088 HSA243666 GGACAUAAAGAAAGGGAAU NM_017582
    1089 HSA243666 AUGCUGAGGUUGAGAUAAA NM_017582
    1090 HSA243666 AGAGAAACUUGGAGCUGAA NM_017582
    1091 HSA243666 CAGUCGAACUCGUGAAUGA NM_017582
    1092 HSA243666 GAGAUAAAGACGGGACAUA NM_017582
    1093 HSA243666 GGAUGGUGAUCAUUGGUAA NM_017582
    1094 HSA243666 CCAAAGGAUGAAACAAAUA NM_017582
    1095 HSA243666 CUAGAAAGGGUUAGCAUUU NM_017582
    1096 HSA243666 UACCAAAGGAUGAAACAAA NM_017582
    1097 HSA243666 GCAACAUCACGGAGUCAUA NM_017582
    1098 HSA243666 ACACAGAAGACUUAGAUCA NM_017582
    1099 HSA243666 GCAUGGAACUUCUCACCAA NM_017582
    1100 HSA243666 AGAUAAAGACGGGACAUAA NM_017582
    1101 HSA243666 CUGAAACGAUGGUGUUAAU NM_017582
    1102 HSA243666 CAACAAAUCUCAAUACAGU NM_017582
    1103 HSA243666 AGAAAGGGUUAGCAUUUUA NM_017582
    1104 HSA243666 GGGAAAUUAUGGAUUCUUU NM_017582
    1105 HSA243666 CCAAAGAUGGCCAAGUGAA NM_017582
    1106 HSHIN1 GGAAGUAGCUGAUGAAGAU NM_017493
    1107 HSHIN1 GGGUAGGACAAGUGGAAAU NM_199324
    1108 HSHIN1 UCGAGAGAACAGAGAGAAA NM_017493
    1109 HSHIN1 GAGAGAACAGAGAGAAAUU NM_017493
    1110 HSHIN1 GAUGAAGAUAACAGUGAAA NM_017493
    1111 HSHIN1 AGAGAAAUUUGAAGCGUUU NM_017493
    1112 HSHIN1 GGAUGACAGUUGCAAGUAA NM_017493
    1113 HSHIN1 AUGAAUUGCUGUAUGAGAA NM_017493
    1114 HSHIN1 GAGAGAAAUUUGAAGCGUU NM_199324
    1115 HSHIN1 AGAAGGAUCAUUUGAAGAA NM_017493
    1116 HSHIN1 UAUCAGAUUCAGAGGAUGA NM_017493
    1117 HSHIN1 UCACAUACCCAGUAAUGAA NM_017493
    1118 HSHIN1 UGCCAUACAAGGAAAGUUA NM_017493
    1119 HSHIN1 UGGAAGUAGCUGAUGAAGA NM_017493
    1120 HSHIN1 ACAGGAAUGGGUAGGACAA NM_017493
    1121 HSHIN1 GGUAGGACAAGUGGAAAUA NM_017493
    1122 HSHIN1 CUUCGAGAGAACAGAGAGA NM_017493
    1123 HSHIN1 GAACAGAGAGAAAUUUGAA NM_017493
    1124 HSHIN1 CGUUGGAAGUAGCUGAUGA NM_017493
    1125 HSHIN1 GGGACUGUUUGCUUUGAAA NM_017493
    1126 HSHIN1 CAUACCCAGUAAUGAAAUA NM_017493
    1127 HSHIN1 CUUAUGUACAGGAAAGAUU NM_199324
    1128 HSHIN1 CUGAUGAAGAUAACAGUGA NM_017493
    1129 HSHIN1 UCACUAUCUUCGAGAGAAC NM_017493
    1130 HSHIN1 AAUAUCAGAUUCAGAGGAU NM_017493
    1131 HSHIN1 GUGUAUCCCAUAAAGUAUA NM_017493
    1132 HSHIN1 UGAUGAAGAUAACAGUGAA NM_017493
    1133 HSHIN1 CGAGAGAACAGAGAGAAAU NM_017493
    1134 HSHIN1 AAGUAGCUGAUGAAGAUAA NM_199324
    1135 HSHIN1 CUUCACAAGUAACAGAAAA NM_199324
    1136 KIAA0063 CAGAAGAGGUAGAGGCUCA NM_014876
    1137 KIAA0063 GCAAAUGGCACUUGAAAGA NM_014876
    1138 KIAA0063 GGACAAAAGAGAAGCCAAA NM_014876
    1139 KIAA0063 GGAAAGGAGACAAGGCCAA NM_014876
    1140 KIAA0063 CGGCAGAGUUUUAUAAUGA NM_014876
    1141 KIAA0063 AGAAGAGCAUGCUGGGAAA NM_014876
    1142 KIAA0063 GCAUGUUGAUCCUGAGCAA NM_014876
    1143 KIAA0063 UGAAGGACCCAGAGCAUAA NM_014876
    1144 KIAA0063 CGUUAUAUCUUCUCAGCAA NM_014876
    1145 KIAA0063 GCAAAGACGCUUAAACAUU NM_014876
    1146 KIAA0063 AGACAGGCUUCUUGGAGUA NM_014876
    1147 KIAA0063 ACAAAUCUACCAUGAGAAA NM_014876
    1148 KIAA0063 AAACAUCAUUUGCGAGGAA NM_014876
    1149 KIAA0063 GGAUAUGCAAAGACGCUUA NM_014876
    1150 KIAA0063 CCACAAAUCUACCAUGAGA NM_014876
    1151 KIAA0063 AUAAAGGUCUCUCAGGAAA NM_014876
    1152 KIAA0063 CAUGGACACCAGAGAAUAA NM_014876
    1153 KIAA0063 UCACUAACGUCAUGGGCUU NM_014876
    1154 KIAA0063 CAUAAAGGUCUCUCAGGAA NM_014876
    1155 KIAA0063 CCAAGCGGCUACUGCGUUA NM_014876
    1156 KIAA0063 AAGAAGAGCAUGCUGGGAA NM_014876
    1157 KIAA0063 GGAAACUCCUCUCCACAAA NM_014876
    1158 KIAA0063 UGGCAGCACUUCAGACCAA NM_014876
    1159 KIAA0063 AAAUCUACCAUGAGAAACA NM_014876
    1160 KIAA0063 UGACAGUGGUAGCAAAAUA NM_014876
    1161 KIAA0063 GCAACUACGAUGUGAAUGU NM_014876
    1162 KIAA0063 GGAAAGUAUUGUGAAGACA NM_014876
    1163 KIAA0063 GGGAGGGAAUCCUAGCUUA NM_014876
    1164 KIAA0063 CGGGAUACGCUGCAAGAGA NM_014876
    1165 KIAA0063 CCACUGGCACUGUCAGAUA NM_014876
    1166 KIAA0710 CAAGAUGGCAGUAAAGAAA NM_014871
    1167 KIAA0710 AGGAGCAGGUGGUGGAUUA NM_014871
    1168 KIAA0710 GAAUUGACCCAGAUGGAAA NM_014871
    1169 KIAA0710 GGGUAUACAUUGUGCCUUU NM_014871
    1170 KIAA0710 GGUCACAGAUGGUGCUAUU NM_014871
    1171 KIAA0710 CCAAACAAGUCCCAAGAAU NM_014871
    1172 KIAA0710 UGACCCAGAUGGAAAGUAA NM_014871
    1173 KIAA0710 CACAGUAGUUCAAGACUUA NM_014871
    1174 KIAA0710 CAUCAAAUAUUCCAAGCUA NM_014871
    1175 KIAA0710 CCACAGUAGUUCAAGACUU NM_014871
    1176 KIAA0710 GAUACAACCUGAACAUCAA NM_014871
    1177 KIAA0710 GCUAUUAAUUGAACUGGAA NM_014871
    1178 KIAA0710 CCUGAUGGCUACUGUGGUA NM_014871
    1179 KIAA0710 AAGAACAACCUCAAGUAUA NM_014871
    1180 KIAA0710 GAUGGAAAGUAAUUGGUAU NM_014871
    1181 KIAA1203 GGUCAGUGUUGUUGGAAUA NM_020718
    1182 KIAA1203 AGAAGGAAAUCUUGGAGAA NM_020718
    1183 KIAA1203 UGAAAUUUGGCUUGGAUUA NM_020718
    1184 KIAA1203 AGAAAGGAGUGAAGAUGAU NM_020718
    1185 KIAA1203 CAGAUUGUGUUAACAGAAA NM_020718
    1186 KIAA1203 UGGAGAAGAUGAAGUAUUU NM_020718
    1187 KIAA1203 GAACCAAGCGACAGUCAUA NM_020718
    1188 KIAA1203 CAUCGUUCCUUUUGUGAUA NM_020718
    1189 KIAA1203 AGAAAGUGUUCGUCUGCAA NM_020718
    1190 KIAA1203 GAAGCAGUGUCUAUGGAAA NM_020718
    1191 KIAA1203 GUAUAUUCCUGAUGCAGAA NM_020718
    1192 KIAA1203 UGGCACUGCUCAUGUGAAA NM_020718
    1193 KIAA1203 CUUUGAGACUCCCGAAAUA NM_020718
    1194 KIAA1203 GAGAAAGGAGUGAAGAUGA NM_020718
    1195 KIAA1203 UGAAGAUGAUGGAGGCUUU NM_020718
    1196 KIAA1203 GACAAGGAGACAAGAGAUU NM_020718
    1197 KIAA1203 GCAGGGAAGCAUUACGUUA NM_020718
    1198 KIAA1203 CCUCAAACCUGCACUUUAU NM_020718
    1199 KIAA1203 CAGGGAAGCAUUACGUUAA NM_020718
    1200 KIAA1203 UGUAGUGUAUCAAGGCAAA NM_020718
    1201 KIAA1203 CAUGAAAGCGACUGCAUUU NM_020718
    1202 KIAA1203 CGUCAGAGUUUGUCAUCCA NM_020718
    1203 KIAA1453 CCGUAUAUGUCCCAGAAUA NM_025090
    1204 KIAA1453 GCACACAGCCACAGGUGAA NM_025090
    1205 KIAA1453 GAACAUCGGCAAUGGGAUU NM_025090
    1206 KIAA1915 GGACUGAGAAACAGAGCAA XM_055481
    1207 KIAA1915 GGUUAUAAGUGAGGAAAUU XM_055481
    1208 KIAA1915 AAGCUAAAUACCAGAGUUA XM_055481
    1209 KIAA191S GUUAGAGGCUUCAGUGUUA XM_055481
    1210 KIAA1915 AAAUGAAGAUAAAGGGACA XM_055481
    1211 KIAA1915 GGAUAAAGAAACACCAAAU XM_055481
    1212 KIAA1915 GGUUAGAAUUCAAGUAGUU XM_055481
    1213 KIAA1915 GAAAAGACAAGAUGGAUAA XM_055481
    1214 KIAA1915 AGGAAAGCCAUGAGGAAGA XM_055481
    1215 KIAA1915 CAGAAAAGAUGCAGUAGAA XM_055481
    1216 KIAA1915 GUACAGGACUACAGUGUGA XM_055481
    1217 KIAA1915 AAAGAAGAGAAGAGGAAAA XM_055481
    1218 KIAA1915 UGAAAAGACAAGAUGGAUA XM_055481
    1219 KIAA1915 AGAGGAAAGCCAUGAGGAA XM_055481
    1220 KIAA1915 GUGCAAAGUUCAUUGGGAU XM_055481
    1221 KIAA1915 GCCACAAACAGUUGACAAA XM_055481
    1222 KIAA1915 GCCAAAUGGUAGAGGAAAG XM_055481
    1223 KIAA1915 ACUCAGAAGUUGAUAAAGU XM_055481
    1224 KIAA1915 CAAAGGACUUAGAAGGACA XM_055481
    1225 KIAA1915 GAAAUAAUCCCUUACCAUA XM_055481
    1226 KIAA1915 GGCCAUAAUCUUCAAGUUA XM_055481
    1227 KIAA1915 CUGCUGAGGAGUUGGCAAA XM_055481
    1228 KIAA1915 ACAGAGCAAUGGUGACAAA XM_055481
    1229 KIAA1915 GCUUUAUGGCUGAAGAAUU XM_055481
    1230 KIAA1915 UCAAAUGCGGUCUGGAUAA XM_055481
    1231 KIAA1915 CUACAAAACCAGCCAGUUA XM_055481
    1232 KIAA1915 GAUAAAGGGACAAAGGCAU XM_055481
    1233 KIAA1915 UGAGGAAGACUCUGAGCAA XM_055481
    1234 KIAA1915 CAACCAAGAGAAUGGAGUA XM_055481
    1235 KIAA1915 AGUGAAGAGUUAUGCAAGA XM_055481
    1236 LOC161725 GCAAGAACGGCAAGGACAA NM_130901
    1237 LOC161725 CAGCAGACGCAGCAGAAUA NM_130901
    1238 LOC161725 GAUACAAUGUUAAGAGACU NM_130901
    1239 LOC161725 GCAAGGACAAGGAGAAGGA NM_130901
    1240 LOC161725 GUUCUACCUUCGAGGAUUU NM_130901
    1241 LOC161725 ACAAGGAGAAGGAGAAGCA NM_130901
    1242 LOC161725 AGAAGCAGCGCAAGGAGAA NM_130901
    1243 LOC161725 AGAGAGACCAGCAAAGAGA NM_130901
    1244 LOC220213 CAAACAAACUCAAGUGCAA XM_166659
    1245 LOC220213 CGACGAAGAACUUGCCAAA XM_166659
    1246 LOC220213 GAGUAAUGGCCACAGGAAA XM_166659
    1247 LOC220213 AGACAGAACACCAGCUAAU XM_166659
    1248 LOC220213 UGGAGGAGCACUUGACAAA XM_166659
    1249 LOC220213 CCAGCUAAUGAAAGAGUUA XM_166659
    1250 LOC220213 AUGAAAGAGUUAAGCGUGA XM_166659
    1251 LOC220213 GGGCAGAUGCUGAAUGUGA XM_166659
    1252 LOC220213 CCUAAGUGAUACUGCAUUU XM_166659
    1253 LOC220213 UGGAGAAGCAGGACAAGUA XM_166659
    1254 LOC220213 GCAAAGGAAACGCGACGAA XM_166659
    1255 LOC220213 GGAUGGGCCCUAAGUGAUA XM_166659
    1256 LOC220213 GGUCAGAGGUGCAAAGUUG XM_166659
    1257 LOC220213 GAGGAUUCCCUGAGGCCUA XM_166659
    1258 LOC220213 GCGAAGAGCACUUGGCGGA XM_166659
    1259 LOC220213 CAACAGAUGCUCAACAAAU XM_166659
    1260 LOC220213 ACGAGAAGCUGGCCCUAUA XM_166659
    1261 LOC220213 GGGAGCAGACGGUGCACUA XM_166659
    1262 LOC220213 GAGGAGCACUUGACAAAGA XM_166659
    1263 LOC220213 AGGUGGAGAAGCAGGACAA XM_166659
    1264 LOC220213 CAGUUAGGCUGGAGAAAUG XM_166659
    1265 LOC220213 UGGCCGAGGUGGAGAAGCA XM_166659
    1266 LOC220213 AAACUCAAGUGCAAAGGAA XM_166659
    1267 LOC220213 AAGGAAACGCGACGAAGAA XM_166659
    1268 LOC220213 GGAGAAAUGAGACAGAACA XM_166659
    1269 LOC220213 GACAACUGGUGCAAACAAA XM_166659
    1270 LOC220213 GCUCAGUAACGGACACUAU XM_166659
    1271 LOC220213 ACACUAUGAUGCUGUAUUU XM_166659
    1272 LOC220213 CCACCUACCUUGAGUAAUG XM_166659
    1273 LOC220213 GAAAUGAGACAGAACACCA XM_166659
    1274 MGC10702 AGAAAUAACUCCCAAACAA NM_032663
    1275 MGC10702 AGGCAACAAUUUAGAAGAA NM_032663
    1276 MGC10702 GUGCAAGUCUGAAGAAUGA NM_032663
    1277 MGC10702 GAAAGAAAGAAGCGUAGAA NM_032663
    1278 MGC10702 CCAAGAAGUUACUGAUGAU NM_032663
    1279 MGC10702 GGAAGACUCACUAGUAAUA NM_032663
    1280 MGC10702 ACAGGAUGCUCACGAAUUA NM_032663
    1281 MGC10702 CAAAUUACCUGCCGCACAA NM_032663
    1282 MGC10702 GGAGCAGCAGUCAGAAAUA NM_032663
    1283 MGC10702 GGGAGGUACCUUUGUCUAA NM_032663
    1284 MGC10702 CAGCAGGAAUAUAUGUUAU NM_032663
    1285 MGC10702 UGACAACUGUACAAAGAUU NM_032663
    1286 MGC10702 CAACACAACCCUAAACUGA NM_032663
    1287 MGC10702 AGGUGGUUCUGUUGUGUUA NM_032663
    1288 MGC10702 AUGGAAGACUCACUAGUAA NM_032663
    1289 MGC10702 CCACACCAGUUCUGAAUCA NM_032663
    1290 MGC14793 UGAUAUUGCUGUAGGUUUA NM_032929
    1291 MGC14793 CAUGAAACUUUGUGAAGAA NM_032929
    1292 MGC14793 GGUGAAAGAUCCAACUAAA NM_032929
    1293 MGC14793 GGAAAUGGAGACACAGAAA NM_032929
    1294 MGC14793 AGAAAGUACCGUAAGAUAA NM_032929
    1295 MGC14793 GCAAAUAGAGUUUAAGUGA NM_032929
    1296 MGC14793 GAGCAAUAGCUGAGAAUCU NM_032929
    1297 MGC14793 GCACAGACUUAUACUCUUA NM_032929
    1298 MGC14793 GUGAAUUUCUCAAGGUAUU NM_032929
    1299 MGC14793 GAUCAAAGAAAGUAGUACA NM_032929
    1300 MGC14793 GGAGGAAAAUGCAGAAAUU NM_032929
    1301 MGC14793 UAGAAAGUACCGUAAGAUA NM_032929
    1302 MGC14793 GCUCUAUGCUUUAUGAAAA NM_032929
    1303 MGC14793 AUAAGGAAAUGGAGACACA NM_032929
    1304 MGC14793 CUCAGAAUGUUUAGAAGAA NM_032929
    1305 MGC14793 AAAUGGAGACACAGAAAUU NM_032929
    1306 MGC14793 CGUGAAUCAUGUAAAGAGA NM_032929
    1307 MGC14793 GCUCAGAAUGUUUAGAAGA NM_032929
    1308 MGC14793 GAGAAUCUGUGGUCAGUUU NM_032929
    1309 MGC14793 GGGUGAAAGAUCCAACUAA NM_032929
    1310 MGC14793 GAGAAAGCCAAAAGAAGUA NM_032929
    1311 MGC14793 CAGCUUAUCCUCAUAUAUA NM_032929
    1312 MGC14793 UGAGGCAAAUAGAGUUUAA NM_032929
    1313 MGC14793 UCUAGAAAGUACCGUAAGA NM_032929
    1314 MGC14793 GCCUGGAAAUAAACACAUA NM_032929
    1315 MGC14793 GAACAGAGCCCCAUUGUAU NM_032929
    1316 MGC14793 UUACUGAUCUGAUGAAUGA NM_032929
    1317 MGC14793 UGUUACAGCUCUAUGCUUU NM_032929
    1318 MGC14793 GUAAUACCAUAAAGUGAGA NM_032929
    1319 MGC14793 CAGAAUGUUUAGAAGAAAG NM_032929
    1320 MJD CCAAAGAGGCAUUCAGCAA NM_004993
    1321 MJD CAAACAAAAUGAUGGGAAA NM_030660
    1322 MJD CUUUAGAAACUGUCAGAAA NM_004993
    1323 MJD GUUCAACAGUCCAGAGUAU NM_030660
    1324 MJD UCAAAGAGAUGAGGAAAUA NM_030660
    1325 MJD GCACUAAGUCGCCAAGAAA NM_004993
    1326 MJD ACGAAGAUGAGGAGGAUUU NM_030660
    1327 MJD GAAACAGCCUUCUGGAAAU NM_030660
    1328 MJD CCGCAGGGCUAUUCAGCUA NM_004993
    1329 MJD CAAGGUAGUUCCAGAAACA NM_004993
    1330 MJD CAAAUUAACCUUUCAGGAA NM_030660
    1331 MJD CAGGAAUGUUAGACGAAGA NM_004993
    1332 MJD AGUAAUGGUUCUAGAAGGA NM_004993
    1333 MJD AAUUACAACAGGAAGGUUA NM_030660
    1334 MJD CGAGAAGCCUACUUUGAAA NM_030660
    1335 MJD UGACAUGGAAGAUGAGGAA NM_030660
    1336 MJD UGGAGAAGAAUUAGCACAA NM_030660
    1337 MJD CUUGACGGGUCCAGAAUUA NM_030660
    1338 MJD CAACAGAUGCAUCGACCAA NM_004993
    1339 MJD ACUAAGUCGCCAAGAAAUU NM_004993
    1340 MJD GAUCACAACUUUUCUGCUA NM_004993
    1341 MJD GCUCAACAUUGCCUGAAUA NM_004993
    1342 MJD AGACCUGGAACGAGUGUUA NM_030660
    1343 MJD GGUAAUGUGUCAAAGAGAU NM_030660
    1344 MJD GGAAGAGACGAGAAGCCUA NM_030660
    1345 MJD CCAAGAAAUUGACAUGGAA NM_030660
    1346 MJD UGAAAUCAGCCUUGCACAA NM_030660
    1347 MJD ACAGGAAGGUUAUUCUAUA NM_004993
    1348 MJD AAGAAAUUGACAUGGAAGA NM_030660
    1349 MJD AGGUAGUUCCAGAAACAUA NM_030660
    1350 NY-REN-60 GGGAAGAAAUGGAAAGAAU NM_032582
    1351 NY-REN-60 GCUAAGAUCUCAAGUAAAA NM_032582
    1352 NY-REN-60 UCUCAAAGGCUGCGCAUUA NM_032582
    1353 NY-REN-60 AGGAAAGGGUUGUAGAUGA NM_032582
    1354 OTUB1 CCGACUACCUUGUGGUCUA NM_017670
    1355 OTUB1 CCGAAUUCAGCAAGAGAUU NM_017670
    1356 OTUB1 AGACCAGGCCUGACGGCAA NM_017670
    1357 OTUB1 CUGCCAAGAGCAAGGAAGA NM_017670
    1358 OTUB1 AGGUGGAGCCCAUGUGCAA NM_017670
    1359 OTUB1 GAGCAGGUGGAGAGGCAGA NM_017670
    1360 OTUB1 AGGAAGACCUGGUGUCCCA NM_017670
    1361 OTUB1 CGGCCUGGACACUACGAUA NM_017670
    1362 OTUB1 AAGAGAUUGCUGUGCAGAA NM_017670
    1363 OTUB1 AGAUCAAGGACCUCCACAA NM_017670
    1364 OTUB1 UCUAUCAACAGAAGAUCAA NM_017670
    1365 OTUBI ACAAGGAGUAUGCUGAAGA NM_017670
    1366 OTUB1 CAAGGAGUAUGCUGAAGAU NM_017670
    1367 OTUB1 GGAGUAUGCUGAAGAUGAC NM_017670
    1368 OTUB1 CCACCAAUCCGCACAUCUU NM_017670
    1369 OTUB1 GGCCUGGACACUACGAUAU NM_017670
    1370 OTUB1 CCGAAGGUGUUAACUGUCU NM_017670
    1371 POH1 CAAUAAGGCUGUAGAAGAA NM_005805
    1372 POH1 GGCAUUAAUUCAUGGACUA NM_005805
    1373 POH1 CUGAACAGCUGGCAAUAAA NM_005805
    1374 POH1 CAGAAGAUGUUGCUAAAUU NM_005805
    1375 POH1 ACAAUAAGGCUGUAGAAGA NM_005805
    1376 POH1 AAGGAAAGGUUGUUAUUGA NM_005805
    1377 POH1 CCAGAAAUAUGGACAGACU NM_005805
    1378 POH1 GUUUGACACUUCAGGACUA NM_005805
    1379 POH1 CUUAAGAGUUGUAGUUACU NM_005805
    1380 PRPF8 AGGAGAAGCUGCAGGAGAA NM_006445
    1381 PRPF8 GGGCCAAGUUCCUGGACUA NM_006445
    1382 PRPF8 GGAUAUGGCCGGAGUGUUU NM_006445
    1383 PRPF8 UGGCAAAGACAGUAACAAA NM_006445
    1384 PRPF8 GCUCAAAAGAAGAGGUAUU NM_006445
    1385 PRPF8 UGACAGACUUGGUGGAUGA NM_006445
    1386 PRPF8 CCAAGAUCAUGAAGGCAAA NM_006445
    1387 PRPF8 CAACAUGAAAUAUGAGCUA NM_006445
    1388 PRPF8 GGAUCAAGGUCGAGGUGCA NM_006445
    1389 PRPF8 CGGAUGAUGAUGAGGAAUU NM_006445
    1390 PRPF8 GAUGAAGACUGGAAUGAAU NM_006445
    1391 PRPF8 GGAAAUUGAGACAGUACAA NM_006445
    1392 PRPF8 CAGAAAUACUGGAUUGACA NM_006445
    1393 PRPF8 AGGAAUGAGCCAUGAAGAA NM_006445
    1394 PRPF8 GCAGAUGGAUUGCAGUAUA NM_006445
    1395 PRPF8 AACCAAGGAAAGAAAGAAA NM_006445
    1396 PRPF8 ACACAUCACUGGAGCCAUU NM_006445
    1397 PRPF8 ACAAAGAGUUCUACCACGA NM_006445
    1398 PRPF8 GAUUAAGCCUGCAGACACA NM_006445
    1399 PRPF8 GGACAUGAACCAUACGAAU NM_006445
    1400 PRPF8 CCAAAUUGCAGGAUACCUA NM_006445
    1401 PRPF8 CCAAGAAUGUGCUUAAGAA NM_006445
    1402 PRPF8 GAUAAGGGCUGGCGUGUCA NM_006445
    1403 PRPF8 CAGACUUGGUGGAUGACAA NM_006445
    1404 PRPF8 AGAACAACGUCGUCAUCAA NM_006445
    1405 PRPF8 CCUAUAAGCAUGACACCAA NM_006445
    1406 PRPF8 CAAUGUAUAUGUAGGCUUU NM_006445
    1407 PRPF8 AAGACUGAGUGGAGGGUCA NM_006445
    1408 PRPF8 GAAUCUAUGAAGUGGAAGA NM_006445
    1409 PRPF8 CGAGUGAAGGCGAGUGCAA NM_006445
    1410 PSMD14 GGACAUGAACCAAGACAAA NM_005805
    1411 PSMD14 CAAUAAAGAAUGUUGGCAA NM_005805
    1412 PSMD14 CUGUAGAAGAAGAAGAUAA NM_005805
    1413 PSMD14 CAAUGGAAGUUAUGGGUUU NM_005805
    1414 PSMD14 ACAAUGAAUCAGUGGUAAA NM_005805
    1415 PSMD14 GGACAGACUUCUUAGACUU NM_005805
    1416 PSMD14 UAAAGGAGAUGUUGGAAUU NM_005805
    1417 PSMD14 GAUUAUACCGUCAGAGUGA NM_005805
    1418 PSMD14 GUAAAGGAGAUGUUGGAAU NM_005805
    1419 PSMD14 ACACAAUGAAUCAGUGGUA NM_005805
    1420 PSMD14 CAAAUAUUGUCCAGUGUUU NM_005805
    1421 PSMD14 GUACUUAUGACCUCAAAUA NM_005805
    1422 PSMD14 AAUCAGUGGUAAAGGAGAU NM_005805
    1423 PSMD14 CAGCAGAACAAGUCUAUAU NM_005805
    1424 PSMD14 GGUCUUAGGACAUGAACCA NM_005805
    1425 PSMD14 AAGAAGAGUUGGAUGGAAG NM_005805
    1426 PSMD14 GAAGAAGAAGAUAAGAUGA NM_005805
    1427 PSMD14 AGAGUUGGAUGGAAGGUUU NM_005805
    1428 PSMD14 GAACCAAGACAAACAACUU NM_005805
    1429 PSMD14 GGGUUUGAUGCUUGGAGAA NM_005805
    1430 PSMD14 CAGAGUGAUUGAUGUGUUU NM_005805
    1431 AD1 UGUUAGAGGUGGAGGAUUU NM_006590
    1432 AD1 GGACUUUGACUUUGAGAAA NM_006590
    1433 AD1 ACAUAAAGGCCAAUGAUUA NM_006590
    1434 AD1 UUGGAGAGCUGAUGAGAAA NM_006590
    1435 AD1 AGGCAAAUGGUAUGAAUUA NM_006590
    1436 AD1 CUGAAGAAGUACACAAGAA NM_006590
    1437 AD1 GGAACUACUUUCUGGAAGA NM_006590
    1438 AD1 GAAGAAGUACACAAGAAUA NM_006590
    1439 AD1 GAGAAGGAAUAUAAGACUU NM_006590
    1440 AD1 GGCUGAUGAUGGUAAAUAA NM_006590
    1441 AD1 GAGAUAAUGAUGAAACCAA NM_006590
    1442 AD1 ACAGAUAAUUCCUUCCAAA NM_006590
    1443 AD1 GGAAGAGGCGAGAUAAUGA NM_006590
    1444 AD1 GGGAGUAGUUGAAGAACAG NM_006590
    1445 AD1 GAGGCGAGAUAAUGAUGAA NM_006590
    1446 AD1 GCAAAUGGUAUGAAUUACA NM_006590
    1447 AD1 GUAAAUAAGAACACAGAAG NM_006590
    1448 AD1 GCAAGCAGAAAGCCAGUAA NM_006590
    1449 SBBI54 CCAAGGAGGUGGAGGAGAA NM_138334
    1450 SBBI54 UGACCAAGGAGGUGGAGGA NM_138334
    1451 SBBI54 GGGAAAGGCCAGCACUUCA NM_138334
    1452 SBBI54 CGGCAACUAUGAUGUCAAU NM_138334
    1453 SBBI54 GCGAGGUGCUGCUGGUAGU NM_138334
    1454 SBBI54 GCAACUAUGAUGUCAAUGU NM_138334
    1455 SBBI54 UGGACGGUGUCUACUACAA NM_138334
    1456 SBBI54 GGAAAGGCCAGCACUUCAU NM_138334
    1457 SBBI54 ACUAUGAUGUCAAUGUGAU NM_138334
    1458 SBBI54 CCGCUGCUGCCUCAAUAAA NM_138334
    1459 SBBI54 GGAGGCUGCCGAUGAGAUC NM_138334
    1460 SBBI54 GCACCGGCAACUAUGAUGU NM_138334
    1461 SBBI54 GCUGCUGCCUCAAUAAAUC NM_138334
    1462 SBBI54 CAACUAUGAUGUCAAUGUG NM_138334
    1463 SBBI54 CCGAUGAGAUCUGCAAGAG NM_138334
    1464 SBBI54 GGACGGUGUCUACUACAAC NM_138334
    1465 SBBI54 UGCCGCUGCUGCCUCAAUA NM_138334
    1466 SBBI54 GGUGGACGGUGUCUACUAC NM_138334
    1467 SBBI54 GAGGUGGUGGUGGUAGUGA NM_138334
    1468 SBBI54 GAAAGGCCAGCACUUCAUG NM_138334
    1469 SENP2 GCGAAUUACUCGAGGAGAU NM_021627
    1470 SENP2 CCUCAUGCAUUGUGGGUUA NM_021627
    1471 SENP2 GAUUAUAUUUCUAGGGACA NM_021627
    1472 SENP2 GGAGUGGACUGGAGCGUAA NM_021627
    1473 SENP2 GAACAAACGCUAACUAAUA NM_021627
    1474 SENP2 AAGAAGAUGGUGUGGGAAA NM_021627
    1475 SENP2 GGUAAAUCUCUUUGAACAA NM_021627
    1476 SENP2 GGGUAAAUCUCUUUGAACA NM_021627
    1477 SENP2 CAGAGAAGAUGGUCGGAAU NM_021627
    1478 SENP2 GGAAGAAGAUGGUGUGGGA NM_021627
    1479 SENP2 GGAGAUAUUCAGACAUUAA NM_021627
    1480 SENP2 GGACAAACCUAUCACAUUU NM_021627
    1481 SENP2 CAAAGAAGUCAGAUGGACA NM_021627
    1482 SENP2 GGAAAUCAGUAAUGCCCUA NM_021627
    1483 SENP2 AGAGAAGUACCGAAAGUUA NM_021627
    1484 SENP2 GAGGAGAUAUUCAGACAUU NM_021627
    1485 SENP2 CCACAAAGCCCAUGGUAAC NM_021627
    1486 SENP2 GCUGAAACUGGGUAAUAAA NM_021627
    1487 SENP2 GGAACAACAUGCUGAAACU NM_021627
    1488 SENP2 AAAGAGAGGGACAGAAGAA NM_021627
    1489 SENP2 GGAGGAAAGUGUCAAUAAU NM_021627
    1490 SENP2 AAAGAGAGAAGUACCGAAA NM_021627
    1491 SENP2 GCAAAAUCACGGAGUCAAA NM_021627
    1492 SENP2 UUACAGAGGACAUGGAAAA NM_021627
    1493 SENP2 UGGAAAAGGAAAUCAGUAA NM_021627
    1494 SENP2 GAUGAAAGUAAGACCAAAA NM_021627
    1495 SENP2 CAGUAAUGCCCUAGGCCAU NM_021627
    1496 SENP2 CAUGAAACCACACGAGAUU NM_021627
    1497 SENP2 AAGAGGAAAGAGAGAAGUA NM_021627
    1498 SENP2 UGAUGAAAUACCAGCCAAA NM_021627
    1499 TAMBP CAGAAGAGCUGAAGGCAGA NM_201647
    1500 TAMBP CCAGCUGGGUAGUGCGGUA NM_006463
    1501 TAMBP UGGAAUUCUCUGUGGAAAA NM_006463
    1502 TAMBP UGUCAUUCCUGAAAAGAAA NM_006463
    1503 TAMBP CCAAAGCAGAAGAGCUGAA NM_201647
    1504 TAMBP GAAGGUAGACCCUGGCCUA NM_006463
    1505 TAMBP AGUGGAGACAUGUGGAAUU NM_006463
    1506 TAMBP CCUUCAUCCUCUAUAACAA NM_201647
    1507 TAMBP UCACACAACUGUAAGGCCA NM_201647
    1508 TNFAIP3 GGGAAGAUUUGAAGACUUA NM_006290
    1509 TNFAIP3 GCACCAUGUUUGAAGGAUA NM_006290
    1510 TNFAIP3 GAGCAGGAGAGGAAAGAUA NM_006290
    1511 TNFAIP3 CAUAUUUGCUCUAGAAGAA NM_006290
    1512 TNFAIP3 GCGGAAAGCUGUGAAGAUA NM_006290
    1513 TNFAIP3 UCACAAGAGUCAACAUUAA NM_006290
    1514 TNFAIP3 CAGCAUGAGUACAAGAAAU NM_006290
    1515 TNFAIP3 GCUAAGAAGUUUGGAAUCA NM_006290
    1516 TNFAIP3 GGGACGAGCAAGUGCAGAA NM_006290
    1517 TNFAIP3 CAGACUUGGUACUGAGGAA NM_006290
    1518 TNFAIP3 CAAAGUUGGAUGAAGCUAA NM_006290
    1519 TNFAIP3 GGAAUUGCAUCCAAGGUAU NM_006290
    1520 TNFAIP3 UACAAGAAAUGGCAGGAAA NM_006290
    1521 TNFAIP3 CAAAAGGACAGAAGAGCAA NM_006290
    1522 TNFAIP3 ACAAGAAAUGGCAGGAAAA NM_006290
    1523 TNFAIP3 GAAAAUGAGAUGAAGGAGA NM_006290
    1524 TNFAIP3 CAGAAGAGCAACUGAGAUC NM_006290
    1525 TNFAIP3 GCACAAUGGCUGAACAAGU NM_006290
    1526 TNFAIP3 GCUCAAGGAAACAGACACA NM_006290
    1527 TNFAIP3 AAAAUGAGAUGAAGGAGAA NM_006290
    1528 TNFAIP3 CAAUAGGAAGGCUAAAUAA NM_006290
    1529 TNFAIP3 CAACUCACUGGAAGAAAUA NM_006290
    1530 TNFAIP3 CCAGUAACCAUGAGUAUGA NM_006290
    1531 TNFAIP3 UUUGAAAGUGGGUGGAAUU NM_006290
    1532 TNFAIP3 GAAUUGCAUCCAAGGUAUA NM_006290
    1533 TNFAIP3 AGUACUUAAUGGUGAUAGA NM_006290
    1534 TNFAIP3 AGGCUUUGUAUUUGAGCAA NM_006290
    1535 TNFAIP3 AAACGAACGGUGACGGCAA NM_006290
    1536 TNFAIP3 AGAGAUUUCAUGAGGCCAA NM_006290
    1537 TNFAIP3 UCAUUGAAGCUCAGAAUCA NM_006290
    1538 TRABID GGGAGAAACUUUAGGAUAU NM_017580
    1539 TRABID GGUAAUAGCCAAAGGAGAU NM_017580
    1540 TRABID GGAGCUAGGUAAUGAGGAA NM_017580
    1541 TRABID GCAUGCAUCUUUUGAGAAA NM_017580
    1542 TRABID GGUUGUAGAAGGUGAUUUA NM_017580
    1543 TRABID UGUAAUGACCCUAAAGUUA NM_017580
    1544 TRABID GGAGAAACUUUAGGAUAUA NM_017580
    1545 TRABID CAGCAGAUAUUGAAGAUUU NM_017580
    1546 TRABID GAAGAAGAAUCUCCAAUUA NM_017580
    1547 TRABID GAAUUUAUCUCCAGUGUUU NM_017580
    1548 TRABID GGAAGUAGUCCUUUGAUAU NM_017580
    1549 TRABID ACAGAGAACUUUUGUUGAA NM_017580
    1550 TRABID CGUUAUAUCUCCUGCCUAU NM_017580
    1551 TRABID UCAAACAGCAGAAGACCAA NM_017580
    1552 TRABID GGAAGAUGAGGAUGAUGAA NM_017580
    1553 TRFP CAGAGAUGGUCUUGGAGAA NM_004275
    1554 TRFP GAAGGCAUCUAAAGAGAAA NM_004275
    1555 TRFP AGGCACAACUGGUUUGAUA NM_004275
    1556 TRFP CCUCAAGAAUGGUAAUUAU NM_004275
    1557 TRFP ACUACAAGAUGAAAGACAA NM_004275
    1558 TRFP GCAGUCAUGUGCUGAGUUA NM_004275
    1559 TRFP UAUGAAGAGUGGAGAAGUU NM_004275
    1560 TRFP ACAUGGAACUCUUCAACAA NM_004275
    1561 TRFP UCUACAAGGUUAUCAGGAA NM_004275
    1562 TRFP UCCUAGAGAUGGAGUAGAA NM_004275
    1563 TRFP GUUAAUGGCUUCUGUUACA NM_004275
    1564 TRFP GGAUUGUGCUGUAAUAAGA NM_004275
    1565 TRFP CCUGUUUGCUGAAGUGAUU NM_004275
    1566 TRFP GCUCAGUUCAGUACCAUAA NM_004275
    1567 TRFP GUGAAUGUAUGUACUGUAU NM_004275
    1568 TRFP GCUAUAAGAUCGAAGUUUG NM_004275
    1569 TRFP GAGGUAAUCAGAUGGUCUA NM_004275
    1570 TRFP GAAAUAUUCAUCCAGGUUA NM_004275
    1571 TRFP GAUCUUAGGGUUAGAAUAC NM_004275
    1572 TRFP AGGCCAAACUGCUAUAAGA NM_004275
    1573 TRFP AAAGAGAUGCUGGGACAUA NM_004275
    1574 TRFP CAAGUCAGGUGGUGGUCAU NM_004275
    1575 TRFP GAUCAAAGCAGACUCAUCA NM_004275
    1576 TRFP GCAAGAGUGUUCAGCAAAC NM_004275
    1577 UBE1 GCUCAGACCUGCAAGAGAA NM_003334
    1578 UBE1 GGGAGGAGGACAUCGGUAA NM_003334
    1579 UBE1 CGGCAGUGAAGCAGACAUA NM_153280
    1580 UBE1 GGUCAAGGCUGUUACCCUA NM_003334
    1581 UBE1 CAGCAGAACUGGUAGCCUU NM_153280
    1582 UBE1 UUUCAGAAGUACAGGGCAU NM_153280
    1583 UBE1 CCUUCUACCUUGUUUGAAA NM_153280
    1584 UBE1 CCUUAUACCUUUAGCAUCU NM_003334
    1585 UBE1 GGAGGAGGACAUCGGUAAA NM_003334
    1586 UBE1 GGAAAUCAGCCCAUGGAGA NM_003334
    1587 UBE1 CCAUUGACUUUGAGAAGGA NM_153280
    1588 UBE1 GAGAAAUCAUCGUUACAGA NM_153280
    1589 UBE1 GCUAUGGUUUCUAUGGUUA NM_153280
    1590 UBE1 AGUCAAAUCUGAAUCGACA NM_153280
    1591 UBE1 UCAAAGUACCUAAGAAGAU NM_003334
    1592 UBE1 AGAAGGAUGAUGACAGCAA NM_153280
    1593 UBE1 CAGACAAGCUCCCUGGAUU NM_153280
    1594 UBE1 GAAAUGAUCCUCACAGAUU NM_153280
    1595 UBE1 GCGUGGAGAUCGCUAAGAA NM_003334
    1596 UBE1 UCAAACAGUUCCUCGACUA NM_003334
    1597 UBE1 CCUGAAUCCUAAUAAAGAA NM_153280
    1598 UBE1 GGUCAAAGUACCUAAGAAG NM_153280
    1599 UBE1 UGCAAGAGAAGCUGGGCAA NM_003334
    1600 UBE1 UGGCCAAUGCCCUGGACAA NM_003334
    1601 UBE1 CAGAAAAUGUCAACCAGUA NM_003334
    1602 UBE1 GGGAUGAGUUUGAAGGCCU NM_153280
    1603 UBE1 GGGAUGUCACGAAGUUAAA NM_153280
    1604 UBE1 CGGCGAGGAUGUCGAGGUU NM_003334
    1605 UBE1 GAGUGGACAUUGUGGGAUC NM_003334
    1606 UBE1 GAAAUCAGCCCAUGGAGAU NM_153280
    1607 UBE1C AGAGAGAGAUUAUGAGCAA NM_003968
    1608 UBE1C GCGAGGAGCCGGAGAAGAA NM_003968
    1609 UBE1C GCCUAAAGAUAUUGGAAGA NM_003968
    1610 UBE1C AAUGAUACCUGGAGAGAGA NM_003968
    1611 UBE1C CAUUUGAAGCAGAAAGAAA NM_198197
    1612 UBE1C CAGAAGGUUUUAAAGGAAA NM_198197
    1613 UBE1C ACAGAAGGUUUUAAAGGAA NM_003968
    1614 UBE1C CAUUGGAGCUGGCGGCUUA NM_003968
    1615 UBE1C UGAAGUUGCUGCAGAAUUU NM_003968
    1616 UBE1C UACAGGAGGUUUUGGAUUA NM_003968
    1617 UBE1C GCUAAGAAGUUGAAUCGAU NM_003968
    1618 UBE1C GGAGCAGCCUUUUGGAGAA NM_003968
    1619 UBE1C GCUGAAUACCUGUGCAUGA NM_003968
    1620 UBE1C AAGCAGAAAGAAAGGAAAA NM_003968
    1621 UBE1C GAAGCAGAAAGAAAGGAAA NM_003968
    1622 UBE1C GCUGAUAUCUCUUCUAAAU NM_003968
    1623 UBE1C CAAUAGUGCUUCUCUGCAA NM_003968
    1624 UBE1C AGAGAGAGCAUCACAAUAU NM_003968
    1625 UBE1C GGUGUUACGUAUAGGCUCA NM_198195
    1626 UBE1C AGAUGAUCCUGAACAUAUA NM_003968
    1627 UBE1C CAGCUAAACUACAGGAGGU NM_003968
    1628 UBE1C GAAGAUGGAUAAAUGGCAU NM_003968
    1629 UBE1C GGCUGAAGUUGCUGCAGAA NM_003968
    1630 UBE1C GGUAACCUCUAUUGAAGAA NM_003968
    1631 UBE1C GGAACCAUGUAAAGAAGUU NM_003968
    1632 UBE1C AGAACACUGUAUUGAGUAU NM_003968
    1633 UBE1C GCUGGAACCAUGUAAAGAA NM_003968
    1634 UBE1C GAGAUGAUCCUGAACAUAU NM_003968
    1635 UBE1C GAAGAAAAGAAGGAGAAUA NM_003968
    1636 UBE1C GAAGAACGAACAAGGCCAA NM_198195
    1637 UBE1DC1 GGACAAACAUGGAUGGAAU NM_024818
    1638 UBE1DC1 GGAAGCAGCAGGAGGAAUA NM_024818
    1639 UBE1DC1 GCUUAUAAUUCCUGGAGAA NM_198329
    1640 UBE1DC1 AAUAAGUAAUGGUGGGUUA NM_198329
    1641 UBE1DC1 ACAAGAAGAGGAAGAGAUA NM_024818
    1642 UBE1DC1 GGUUAUACAAGAAGAGGAA NM_198329
    1643 UBE1DC1 AAGAAGAGGAAGAGAUAAU NM_198329
    1644 UBE1DC1 UGGAAGACCUCAUGGCCAA NM_198329
    1645 UBE1DC1 AGGAAAACCUGUUGAUCUA NM_024818
    1646 UBE1DC1 GCUCGAAUGACAAUAAAUA NM_198329
    1647 UBE1DC1 GAACUAGCCAAUAUGAAUA NM_024818
    1648 UBE1DC1 GGGAAUUGUAAGCGACUAU NM_024818
    1649 UBE1DC1 AAGUAAAGUUCAAGCAGCA NM_024818
    1650 UBE1DC1 ACAAGAGGUUAUACAAGAA NM_198329
    1651 UBE1L CUGCAAAGCUGGAGGAGCA NM_003335
    1652 UBE1L AGGAAGAGCCACUGGAAGA NM_003335
    1653 UBE1L AGGAAGUGCUGAAGGCAAU NM_003335
    1654 UBE1L CAGCACGGGUUGAGGGUGA NM_003335
    1655 UBE1L CUUCGAAGCUACUGGAUGA NM_003335
    1656 UBE1L UGGAAGAGCCACUGGAUGA NM_003335
    1657 UBE1L GGACAGAGGAAGAGCCACU NM_003335
    1658 UBE1L CCACAGAACUGGCAAGACU NM_003335
    1659 UBE1L CCCAAGACUGUGAGACAUA NM_003335
    1660 UBE1L CAGGAAGUGCUGAAGGCAA NM_003335
    1661 UBE1L GGGCCUGAACCCAGACUUA NM_003335
    1662 UBE1L UAAUAAAGUGCUUGAGGAU NM_003335
    1663 UBE1L UCUCGGGAAUUGAGGGAAU NM_003335
    1664 UBE1L GACCCAAGACUGUGAGACA NM_003335
    1665 UBE1L CAUCUUUGCUAGUAAUCUA NM_003335
    1666 UBE1L UGUCAUAAGCAUGGAGUUU NM_003335
    1667 UBE1L GGGCAGUGCUACAGUAUUC NM_003335
    1668 UBE1L AGCAGAAGGAACUGAACAA NM_003335
    1669 UBE1L GGGCUAUCACUGAAGUCAA NM_003335
    1670 UBE1L UGAACAAAGCCCUGGAAGU NM_003335
    1671 UBE1L CUAAUAAAGUGCUUGAGGA NM_003335
    1672 UBE1L GCAGGUGUCUUGAGCCCUA NM_003335
    1673 UBE1L GAAUUGAGGGAAUGGUUGA NM_003335
    1674 UBE1L CCUGGAGAUUGGAGACACA NM_003335
    1675 UBE1L GCGAAUUGUGGGCCAGAUU NM_003335
    1676 UBE1L CUACAGUAUUCAUGCCACA NM_003335
    1677 UBE1L AUGGAGACUUGGUGACUUU NM_003335
    1678 UBE1L CCAUGUGGACUUUGUGGUA NM_003335
    1679 UBE1L CACAGUAGGCACUCAAUAA NM_003335
    1680 UBE2A AUGCCAAGUUUCAGAAUUA NM_003336
    1681 UBE2A CUAAAGGAGUACAGCAAUU NM_003336
    1682 UBE2A GGAAUAUGGCCUACAGAGA NM_003336
    1683 UBE2A GCAGAGGAAUGGAAACAAU NM_003336
    1684 UBE2A GAACAAACGGGAAUAUGAA NM_003336
    1685 UBE2A GGAAGGGAGCAUAGCAUAU NM_003336
    1686 UBE2A GCAAGAAGGAGAAAGUUGA NM_003336
    1687 UBE2A GUUGAAGGACUCAGCUAAA NM_003336
    1688 UBE2A CAGGAGAACAAACGGGAAU NM_003336
    1689 UBE2A GAGCAGAGGAAUGGAAACA NM_003336
    1690 UBE2A CUAUGCAGAUGGUAGUAUA NM_003336
    1691 UBE2A GGGAGCAGAGGAAUGGAAA NM_003336
    1692 UBE2A AAUAUGGCCUACAGAGAAU NM_003336
    1693 UBE2A AAUGAUUGCUGAAGUGUUU NM_003336
    1694 UBE2A ACAAGUAACCCAUGUAAAA NM_003336
    1695 UBE2A GAAUAUGGCCUACAGAGAA NM_003336
    1696 UBE2A CAGCAAUUGUAGUAACUGA NM_003336
    1697 UBE2A GUGAAUGUGUUUGGAAUAU NM_003336
    1698 UBE2A GCAUAUCUGUGGCAAACUA NM_003336
    1699 UBE2A GGAUGGAACAUUUAAACUU NM_003336
    1700 UBE2A GAGAAUAGAAACAAAUCCA NM_003336
    1701 UBE2A AAUAAACCACCUACAGUUA NM_003336
    1702 UBE2A AGAACAAACGGGAAUAUGA NM_003336
    1703 UBE2A AGCUGUACCAGGAGAACAA NM_003336
    1704 UBE2A ACUCAAUUGUCCAUCUUUA NM_003336
    1705 UBE2A AGUUAUUGCUGCAUGCUUU NM_003336
    1706 UBE2A GCACAAGUAACCCAUGUAA NM_003336
    1707 UBE2A CAAGAAGGAGAAAGUUGAA NM_003336
    1708 UBE2A UCACUGAAGAAUAUCCAAA NM_003336
    1709 UBE2A GAUACUAAGAUCUCAGUCA NM_003336
    1710 UBE2B GAACAAAGCUGGAAUGAUU NM_003337
    1711 UBE2B AUACAUAACUUCAGUGCAA NM_003337
    1712 UBE2B CAAACGAGAAUAUGAGAAA NM_003337
    1713 UBE2B AGGCUGAGGUGGCAGAAUA NM_003337
    1714 UBE2B GGAGGGAAAUCUUGGUGUA NM_003337
    1715 UBE2B CAAGAGACUUUGUCACUUA NM_003337
    1716 UBE2B GGAAAACAAACGAGAAUAU NM_003337
    1717 UBE2B CAUCUUAGUUUACUGGAUA NM_003337
    1718 UBE2B GGCAAAGGCGGGAGGAUCA NM_003337
    1719 UBE2B GCUUGUGUAUCUUGAUUAA NM_003337
    1720 UBE2B AGACAUAACUGGUUUGACU NM_003337
    1721 UBE2B GGUAGCAUAUGUUUAGAUA NM_003337
    1722 UBE2B AGUUAUAUUUGGACCAGAA NM_003337
    1723 UBE2B GCACAGCUUUAUCAGGAAA NM_003337
    1724 UBE2B GCAGUGGAAUGCAGUUAUA NM_003337
    1725 UBE2B GUAUUUAGGCCAUUUGUUA NM_003337
    1726 UBE2B CAGGAAAACAAACGAGAAU NM_003337
    1727 UBE2B ACACAGAGAACACAAAUUU NM_003337
    1728 UBE2B ACAACAUCAUGCAGUGGAA NM_003337
    1729 UBE2B GCACAUAUUGGAGGGAAAU NM_003337
    1730 UBE2B GUUUAGAUAUCCUUCAGAA NM_003337
    1731 UBE2B UAUCCUAUGCCUUCAAAUA NM_003337
    1732 UBE2B GAGGGAAAUCUUGGUGUAA NM_003337
    1733 UBE2B GAGUAAUUCUAGACAUAAC NM_003337
    1734 UBE2B GAGCUGUGAUUAUGCCAUU NM_003337
    1735 UBE2B GCACUUGCCUGUAGUCUCA NM_003337
    1736 UBE2B GGGAGACGGGAUAGUGUUU NM_003337
    1737 UBE2B ACUAAGUUAUUGCUGCAUA NM_003337
    1738 UBE2B ACAAACGAGAAUAUGAGAA NM_003337
    1739 UBE2B AGCAAUACCCUGUCUUUAA NM_003337
    1740 UBE2C GGUAUAAGCUCUCGCUAGA NM_007019
    1741 UBE2C AAGAAGUACCUGCAAGAAA NM_007019
    1742 UBE2C CGGUUGAGCCCUUGUAUAU NM_007019
    1743 UBE2C CCACAGCUUUUAAGAAGUA NM_007019
    1744 UBE2C CCAUGGAGCAGCUGGAACA NM_007019
    1745 UBE2C GGAGAACCCAACAUUGAUA NM_007019
    1746 UBE2C GUAUAGGACUCUUUAUCUU NM_007019
    1747 UBE2C CUGGAACAGUAUAUGAAGA NM_007019
    1748 UBE2C UGGAGCAGCUGGAACAGUA NM_007019
    1749 UBE2C UAAAUUAAGCCUCGGUUGA NM_007019
    1750 UBE2C AGAAGUACCUGCAAGAAAC NM_007019
    1751 UBE2C UUAAGAAGUACCUGCAAGA NM_007019
    1752 UBE2C UAUCUUGAGCUGUGGUAUU NM_007019
    1753 UBE2C GUUGAGCCCUUGUAUAUUA NM_007019
    1754 UBE2C UGCAAGAAACCUACUCAAA NM_007019
    1755 UBE2C CAGACAACCUUUUCAAAUG NM_007019
    1756 UBE2C UAGGAGAACCCAACAUUGA NM_007019
    1757 UBE2C CAAGAAACCUACUCAAAGC NM_181799
    1758 UBE2C CAGCUGGAACAGUAUAUGA NM_007019
    1759 UBE2C GCAAGAAACCUACUCAAAG NM_007019
    1760 UBE2C GAGCAGCUGGAACAGUAUA NM_007019
    1761 UBE2C GAGCCCUUGUAUAUUAAAU NM_007019
    1762 UBE2C CGAGCGAGUUCCUGUCUCU NM_007019
    1763 UBE2C GGACCAUUCUGCUCUCCAU NM_007019
    1764 UBE2C GGUCUGCCCUGUAUGAUGU NM_007019
    1765 UBE2C UACAAUGCGCCCACAGUGA NM_007019
    1766 UBE2C UCUAGGAGAACCCAACAUU NM_007019
    1767 UBE2C UAACAUAUGCCUGGACAUC NM_007019
    1768 UBE2C ACAAUGCGCCCACAGUGAA NM_007019
    1769 UBE2C GGUUGAGCCCUUGUAUAUU NM_007019
    1770 UBE2D1 GAGAAUGGACUCAGAAAUA NM_003338
    1771 UBE2D1 CAACAGACAUGCAAGAGAA NM_003338
    1772 UBE2D1 GCAUUGAGAAAGACAUUUA NM_003338
    1773 UBE2D1 CAGAAAGAAUUGAGUGAUC NM_003338
    1774 UBE2D1 GGAAGUAUUUGUCUCGAUA NM_003338
    1775 UBE2D1 GAAAGAAUUGAGUGAUCUA NM_003338
    1776 UBE2D1 UUGUAUGCAUUGAGAAAGA NM_003338
    1777 UBE2D1 CCAUGAAACCAUUUGAUUU NM_003338
    1778 UBE2D1 CAGCUGGACCUGUGGGAGA NM_003338
    1779 UBE2D1 CAUAAACAGUAAUGGAAGU NM_003338
    1780 UBE2D1 AGGAAGAUGUGUAACUUUU NM_003338
    1781 UBE2D1 CUAAGUAGGAAGAUGUGUA NM_003338
    1782 UBE2D1 UGAAUUAAUUGCACUGCUA NM_003338
    1783 UBE2D1 AGGAUAUUCUGUAGAUUGA NM_003338
    1784 UBE2D1 GUAGAUUGAUUGCAGAUUU NM_003338
    1785 UBE2D1 CUGAAAAGCAACCAAAUUA NM_003338
    1786 UBE2D1 AAGAGAAUGGACUCAGAAA NM_003338
    1787 UBE2D1 GCAAGAGAAUGGACUCAGA NM_003338
    1788 UBE2D1 GCACAAAUCUAUAAAUCAG NM_003338
    1789 UBE2D1 CAUCCAAACAUAAACAGUA NM_003338
    1790 UBE2D1 AGAGAAUGGACUCAGAAAU NM_003338
    1791 UBE2D1 AGUACCAGAUAUUGCACAA NM_003338
    1792 UBE2D1 CCAUGGCGCUGAAGAGGAU NM_003338
    1793 UBE2D1 GGAAGAUGUGUAACUUUUA NM_003338
    1794 UBE2D1 CAACAUUAGCAGUAAAUUG NM_003338
    1795 UBE2D1 UGUCUAAGAUGUCAGUUUU NM_003338
    1796 UBE2D1 CCCAACGGCUGAUAAUUAA NM_003338
    1797 UBE2D1 GAUCUACAAUGCAGCUGAA NM_003338
    1798 UBE2D1 GAAAUGAUCUUUACACUGU NM_003338
    1799 UBE2D1 CAAUGCAGCUGAAAAGCAA NM_003338
    1800 UBE2D2 UGGAUAACCUCUACAAAUA NM_003339
    1801 UBE2D2 GGGAAUGGACUCAGAAGUA NM_003339
    1802 UBE2D2 CAGCAUUUGUCUUGAUAUU NM_003339
    1803 UBE2D2 GAGAAAAGUACAACAGAAU NM_003339
    1804 UBE2D2 CUAUCAGGGUGGAGUAUUU NM_003339
    1805 UBE2D2 CAACAGAAUAGCUCGGGAA NM_003339
    1806 UBE2D2 AGAGAAUCCACAAGGAAUU NM_003339
    1807 UBE2D2 CUACAAAACAGAUAGAGAA NM_003339
    1808 UBE2D2 GAAGAGAAUCCACAAGGAA NM_003339
    1809 UBE2D2 AGAGAAAAGUACAACAGAA NM_003339
    1810 UBE2D2 AGAAGUAUGCGAUGUAAUU NM_003339
    1811 UBE2D2 GGAUAACCUCUACAAAUAA NM_003339
    1812 UBE2D2 AAGAGAAUCCACAAGGAAU NM_003339
    1813 UBE2D2 CAAAACAGAUAGAGAAAAG NM_003339
    1814 UBE2D2 GGAAUGGACUCAGAAGUAU NM_003339
    1815 UBE2D2 CUACAAUAAUGGGGCCAAA NM_003339
    1816 UBE2D2 GGAGAUGAUAUGUUCCAUU NM_003339
    1817 UBE2D2 CAAAAGUACUCUUGUCCAU NM_003339
    1818 UBE2D2 ACAAGGAAUUGAAUGAUCU NM_003339
    1819 UBE2D2 UGAAGAGAAUCCACAAGGA NM_003339
    1820 UBE2D2 UUGGAUAACCUCUACAAAU NM_003339
    1821 UBE2D2 CAGAAUAGCUCGGGAAUGG NM_181838
    1822 UBE2D2 GAAAAGUACAACAGAAUAG NM_003339
    1823 UBE2D2 GCUCGGGAAUGGACUCAGA NM_003339
    1824 UBE2D2 CAAUCCAGAUGAUCCUUUA NM_003339
    1825 UBE2D2 GCAUUUGUCUUGAUAUUCU NM_003339
    1826 UBE2D2 GAAGUAUGCGAUGUAAUUA NM_003339
    1827 UBE2D2 CACAAGGAAUUGAAUGAUC NM_003339
    1828 UBE2D2 CAGCACUAACUAUUUCAAA NM_003339
    1829 UBE2D2 GGCAGCAUUUGUCUUGAUA NM_003339
    1830 UBE2D3 UGGCAGAGCUGGUGUGAGA NM_003340
    1831 UBE2D3 GCUCUAACAUGCUGAAGAA NM_003340
    1832 UBE2D3 GAGCAUACACCGAGAGAGU NM_003340
    1833 UBE2D3 GCUUGGAGCUAUUAGUUAA NM_003340
    1834 UBE2D3 GGAAAUUGGAAGUCAAACA NM_003340
    1835 UBE2D3 AGAGAUAAGUACAACAGAA NM_003340
    1836 UBE2D3 GAGAUAAGUACAACAGAAU NM_003340
    1837 UBE2D3 GCGCUGAAACGGAUUAAUA NM_003340
    1838 UBE2D3 GAACAGAAAAUGUGAUGUA NM_003340
    1839 UBE2D3 GCAUACACCGAGAGAGUGA NM_003340
    1840 UBE2D3 GUGAGGAGCCAGACGACAA NM_003340
    1841 UBE2D3 AAAUGGAGCAUGUGUAUUA NM_003340
    1842 UBE2D3 CCGGGAUAAUCAAGAGUUU NM_003340
    1843 UBE2D3 CUGAAUAAAUGAUGCAAGU NM_003340
    1844 UBE2D3 CAACAGAAUAUCUCGGGAA NM_181887
    1845 UBE2D3 GAUGAGUGAUCAACUAAUA NM_003340
    1846 UBE2D3 GAUUUGAGGUUACAUGAUA NM_003340
    1847 UBE2D3 CAGCAUUUGUCUCGAUAUU NM_003340
    1848 UBE2D3 GAGAAUGGAAAUUGGAAGU NM_003340
    1849 UBE2D3 AUCAAAGGAUACAGCAUUA NM_003340
    1850 UBE2D3 GAAACGGAUUAAUAAGGAA NM_003340
    1851 UBE2D3 GCAAGUUGUCAAUGGAUGA NM_003340
    1852 UBE2D3 UGUUAGAGAUUUGAGGUUA NM_003340
    1853 UBE2D3 CAGACAGAGAUAAGUACAA NM_181886
    1854 UBE2D3 AAACAGACAGAGAUAAGUA NM_003340
    1855 UBE2D3 GGUUACAUGAUAUGCUUUA NM_003340
    1856 UBE2D3 CAUCAAAGGAUACAGCAUU NM_003340
    1857 UBE2D3 CUAACAUGCUGAAGAAAUC NM_003340
    1858 UBE2D3 UGAUAUGUUUCAUUGGCAA NM_181888
    1859 UBE2E1 CCAAGAAGAAGGAGAGUAA NM_003341
    1860 UBE2E1 GAGCAAACCGAGAAAGAAA NM_182666
    1861 UBE2E1 GUGAAAACCUGUAGUGAAA NM_003341
    1862 UBE2E1 GAGCAGAACAUGACAGAAU NM_182666
    1863 UBE2E1 GGUUCUAUGUUGUGGACUA NM_003341
    1864 UBE2E1 UGUCAUUAGUUCUGCAAUA NM_003341
    1865 UBE2E1 CGGAGGAGCCAGACACAAA NM_003341
    1866 UBE2E1 AGAGAUGAGUAGUGCGUUU NM_003341
    1867 UBE2E1 GGUAAAGAGUAGGGUAUUU NM_003341
    1868 UBE2E1 GCUUGGACAUAUUGAAAGA NM_003341
    1869 UBE2E1 AGAAGGAGCUGGCGGACAU NM_003341
    1870 UBE2E1 GACAGUGGACCAAGAGAUA NM_003341
    1871 UBE2E1 GGAAGGGAACAUUGAUAUU NM_003341
    1872 UBE2E1 CCAUAUAAGAGAUGAGUAG NM_003341
    1873 UBE2E1 GCUUGUAGUCUGUAAAUUU NM_003341
    1874 UBE2E1 UGAAAUACCUUAAGCUGUU NM_003341
    1875 UBE2E1 UGAAUCAGGACUUGUGAAA NM_003341
    1876 UBE2E1 CCAAGAGAAUUCAGAAGGA NM_003341
    1877 UBE2E1 GAGGGUGGGAGUUGGUAAA NM_003341
    1878 UBE2E1 AAACCGAGAAAGAAACAAA NM_003341
    1879 UBE2E1 GUAAAGUCAGCAUGAGCAA NM_003341
    1880 UBE2E1 GAAAACCUGUAGUGAAAUA NM_003341
    1881 UBE2E1 GAGAUACGCUACAUAAAUU NM_003341
    1882 UBE2E1 GGGAGUUGGUAAAGAGUAG NM_003341
    1883 UBE2E1 GAAGAGAGCUGCUUAUGAU NM_003341
    1884 UBE2E1 GAGUAGGGUAUUUCUAUAA NM_003341
    1885 UBE2E1 CAACCAGCAAACCGAGAAA NM_003341
    1886 UBE2E1 GAGAGUAAAGUCAGCAUGA NM_003341
    1887 UBE2E1 AAGAAGGAGAGUAAAGUCA NM_182666
    1888 UBE2E1 CAAGAGAUACGCUACAUAA NM_182666
    1889 UBE2E3 AAGAACAAGAGGAAAGAAA NM_006357
    1890 UBE2E3 AUAUGAAGGUGGUGUGUUU NM_006357
    1891 UBE2E3 GAGCAGAACACGACAGGAU NM_006357
    1892 UBE2E3 ACCAAGAUGUCCAGUGAUA NM_006357
    1893 UBE2E3 AGAAGGAGCUAGCUGAAAU NM_006357
    1894 UBE2E3 GGUAAAUGCUAUCAAGAGU NM_006357
    1895 UBE2E3 AGAGAUAACUUCACCAAGA NM_006357
    1896 UBE2E3 CUGAAGAACAAGAGGAAAG NM_182678
    1897 UBE2E3 AAGCAUAGCCACUCAGUAU NM_006357
    1898 UBE2E3 ACACCAAACUCUCUAGCAA NM_006357
    1899 UBE2E3 UGUAUUAAACCCAGAUCUA NM_006357
    1900 UBE2E3 GCCUGAAGAACAAGAGGAA NM_182678
    1901 UBE2E3 CAAGAGAUACGCAACAUAA NM_006357
    1902 UBE2E3 CAAUAAACAUGCUCCUGAA NM_006357
    1903 UBE2E3 GAAGGAGCUAGCUGAAAUA NM_006357
    1904 UBE2E3 GCUAUCAAGAGUAGAACUU NM_006357
    1905 UBE2E3 GGCAAAGGUCCGAUGAUGA NM_006357
    1906 UBE2E3 GCAAAUCUUUAUAGCCUUU NM_006357
    1907 UBE2E3 CCUAAAGGAGAUAACAUUU NM_006357
    1908 UBE2E3 CUCCAGAGCCUGAAGAACA NM_006357
    1909 UBE2E3 CCAAGAGAUACGCAACAUA NM_006357
    1910 UBE2E3 GCCUAAAGGAGAUAACAUU NM_006357
    1911 UBE2E3 UGAAAUAACCCUUGAUCCU NM_006357
    1912 UBE2E3 GUAAAUGCUAUCAAGAGUA NM_006357
    1913 UBE2E3 AGAACAAGAGGAAAGAAAA NM_182678
    1914 UBE2E3 CACCAAACUCUCUAGCAAA NM_006357
    1915 UBE2E3 GCAGUGUGAAGGAGCAGAA NM_006357
    1916 UBE2E3 ACUCAGUAUUUGACCAACA NM_006357
    1917 UBE2G1 GAGAAGUGGUUUAGGAAAA NM_003342
    1918 UBE2G1 GGAUAGAACUUGAGACAGU NM_003342
    1919 UBE2G1 GGAAGUAGUCUUUGGCUUA NM_003342
    1920 UBE2G1 GGGAAGAUAAGUAUGGUUA NM_003342
    1921 UBE2G1 ACAAAGAAAUUGCGAUGUA NM_003342
    1922 UBE2G1 CUACUUAACUUCUGGGUUU NM_003342
    1923 UBE2G1 GCACCCAAAUGUUGAUAAA NM_003342
    1924 UBE2G1 GGAAGAUAGAAAUGGAGAA NM_003342
    1925 UBE2G1 GGGUAAAUGCUUUGCUAUU NM_003342
    1926 UBE2G1 GGUGUUUGAUUGUGAGAAU NM_003342
    1927 UBE2G1 GAUUAAUGUUUGAGCUUCA NM_003342
    1928 UBE2G1 GUAUAGAUCCCGUCACUAA NM_003342
    1929 UBE2G1 CAAUAAAACAUGCCAGUUA NM_003342
    1930 UBE2G1 GAGAAAUGCUUUGGCAGAA NM_003342
    1931 UBE2G1 GUGGAAACCAUCAUGAUUA NM_003342
    1932 UBE2G1 GAAGAUAGAAAUGGAGAAU NM_003342
    1933 UBE2G1 GGAACUGGGCUGCAAUAAA NM_003342
    1934 UBE2G1 GCAGAAGAGUGAAAGAACU NM_003342
    1935 UBE2G1 UAAUGUUGAUGCUGCGAAA NM_003342
    1936 UBE2G1 UCAUGUACAUCCACAAAUA NM_003342
    1937 UBE2G1 GGGAAGAUAGAAAUGGAGA NM_003342
    1938 UBE2G1 GUACAUAGCACAACAUGAU NM_003342
    1939 UBE2G1 AGAGACUGCUUUUGAGUGA NM_003342
    1940 UBE2G1 GCUAGUAACUUCACUUAUU NM_003342
    1941 UBE2G1 GCACAACAUGAUCCGGAUA NM_003342
    1942 UBE2G1 AUGCACAGCUACAGGCUUU NM_003342
    1943 UBE2G1 GUUGAUUCCUUAUGCAAAU NM_003342
    1944 UBE2G1 GCUACAGGCUUUCUACUUA NM_003342
    1945 UBE2G1 AAUGGAGGGAAGAUAGAAA NM_003342
    1946 UBE2G1 CCAGAGAAGUGGUUUAGGA NM_003342
    1947 UBE2G2 ACAAAGGGCUUCAGGUAGA NM_003343
    1948 UBE2G2 GAUCACAAGGUCAGGAAAU NM_003343
    1949 UBE2G2 CCAUGAAUGAAGAGAACUU NM_003343
    1950 UBE2G2 UCACAAAGGGCUUCAGGUA NM_003343
    1951 UBE2G2 GAAAUGGGGUAAAUAGAAA NM_003343
    1952 UBE2G2 GCUCUGUGCUGGUACCAAA NM_003343
    1953 UBE2G2 UGACGAAAGUGGAGCUAAC NM_003343
    1954 UBE2G2 UUACAAACCAUCACACUUA NM_003343
    1955 UBE2G2 GGCCAAGGCAGGUAGAUCA NM_003343
    1956 UBE2G2 UUACAGUAUUCCUCACAAA NM_003343
    1957 UBE2G2 CCAAGCAGAUCGUCCAGAA NM_003343
    1958 UBE2G2 GAGAUUUACCUGUGAGAUG NM_003343
    1959 UBE2G2 GGCAGAUGCUUUUCUUAUA NM_003343
    1960 UBE2G2 GGGAGCAGUUCUAUAAGAU NM_003343
    1961 UBE2G2 GCAGAUGCUUUUCUUAUAA NM_003343
    1962 UBE2G2 GCGAUGACCCCAUGGGCUA NM_182688
    1963 UBE2G2 ACAGCUUACUGCAGUUUUA NM_003343
    1964 UBE2G2 GUCAGGAAAUUGAGACCAU NM_003343
    1965 UBE2G2 CAAGGCAGGUAGAUCACAA NM_003343
    1966 UBE2G2 UGGCAGAGCCCAAUGACGA NM_182688
    1967 UBE2G2 GCGAUGACCGGGAGCAGUU NM_003343
    1968 UBE2G2 CCACUUGAUUACCCGUUAA NM_182688
    1969 UBE2G2 GGCAACAGAGCGAGACUCA NM_003343
    1970 UBE2G2 GAUCAUGGGCCCAGAAGAC NM_182688
    1971 UBE2G2 GAGCAGUUCUAUAAGAUUG NM_003343
    1972 UBE2G2 ACCAUCACACUUAGAAAUA NM_003343
    1973 UBE2G2 CAACAGAGCGAGACUCAGU NM_003343
    1974 UBE2G2 CAUCACACUUAGAAAUACU NM_003343
    1975 UBE2G2 GGUUGUACCUGCCAGACUU NM_003343
    1976 UBE2G2 GGAGCAGUUCUAUAAGAUU NM_182688
    1977 UBE2H GGAAAUAGCCGCUCUGAUA NM_003344
    1978 UBE2H AAGAGUACAUCCAGAAAUA NM_003344
    1979 UBE2H GGACAGCACAGGAGGAGAA NM_003344
    1980 UBE2H GAUAGGUAGAUGUGAGUAA NM_003344
    1981 UBE2H CAGAAGAAUACAAGCAGAA NM_003344
    1982 UBE2H GGGAGGACUUAAUGAAUUU NM_003344
    1983 UBE2H CGGAGGAGGCGCUGAAAGA NM_003344
    1984 UBE2H CAUGAGAAGCAGACUAUAA NM_003344
    1985 UBE2H GGCAAGAGGCGGAUGGACA NM_003344
    1986 UBE2H AAGAUGAGGCCCAGGAUAU NM_003344
    1987 UBE2H ACUGAACUGUCGAAGGAAA NM_003344
    1988 UBE2H UUAGAAAGGUUGCAGAUUU NM_003344
    1989 UBE2H GGGAGCUAAUUAAGAGUAU NM_003344
    1990 UBE2H UCAAGCUCAUCGAGAGUAA NM_003344
    1991 UBE2H GUAUAAAGCAGGGAGCUAA NM_003344
    1992 UBE2H AUAUGGAGUUGUAGUAGAA NM_003344
    1993 UBE2H GCAGAAAAUUAAAGAGUAC NM_003344
    1994 UBE2H CCAGAAGAAUACAAGCAGA NM_003344
    1995 UBE2H GAAGAUGAGGCCCAGGAUA NM_003344
    1996 UBE2H CCAAUAUAUUUGAGUCCUU NM_182697
    1997 UBE2H CGCUGAAAGAACAGGAAGA NM_003344
    1998 UBE2H GAUAUGGAGUUGUAGUAGA NM_003344
    1999 UBE2H GAGAAGCAGACUAUAAUAU NM_003344
    2000 UBE2H GAAGAAUACAAGCAGAAAA NM_182697
    2001 UBE2H GAGAGUAAACAUGAGGUUA NM_003344
    2002 UBE2H CUACUGAACUGUCGAAGGA NM_003344
    2003 UBE2H GAGUGGACCUACCUGAUAA NM_182697
    2004 UBE2H UGGGAGGACUUAAUGAAUU NM_182697
    2005 UBE2H AGAAAUACGCCACGGAGGA NM_182697
    2006 UBE2H AGUUAUUGGCCUAUCCUAA NM_003344
    2007 UBE2I GAGGAAAGCAUGGAGGAAA NM_003345
    2008 UBE21 GGGAAGGAGGCUUGUUUAA NM_003345
    2009 UBE21 CCAUCUUAGAGGAGGACAA NM_194260
    2010 UBE2I GAAAAGGGUCCGAGCACAA NM_003345
    2011 UBE2I GAGCACAAGCCAAGAAGUU NM_003345
    2012 UBE2I AGGAAAGCAUGGAGGAAAG NM_003345
    2013 UBE2I GGAGGAACGUGUGGAGUUU NM_003345
    2014 UBE2I AAUCUAAAGUUGCUCCAUA NM_003345
    2015 UBE2I CAGCUCAAGCAGAGGCCUA NM_003345
    2016 UBE2I GAAUACAGGAACUUCUAAA NM_003345
    2017 UBE2I CAGAGUGGAGUACGAGAAA NM_003345
    2018 UBE2I GCAGAGGCCUACACGAUUU NM_003345
    2019 UBE2I GGGCCUGUGUCUUGGACUU NM_003345
    2020 UBE2I ACAGGAACUUCUAAAUGAA NM_003345
    2021 UBE2I GGAUCUGUGUUUGGUGAAA NM_003345
    2022 UBE2I AGAGGAAAGCAUGGAGGAA NM_003345
    2023 UBE2I CUGUGUUGCUGUUUAGGUA NM_003345
    2024 UBE2I GAGGAAAGACCACCCAUUU NM_003345
    2025 UBE2I GGAAGGAGGCUUGUUUAAA NM_003345
    2026 UBE2I UAAAGUUGCUCCAUACAAU NM_003345
    2027 UBE2I AGAAGGAAGGGAUUGGUUU NM_003345
    2028 UBE2I GGGAUUGGUUUGGCAAGAA NM_003345
    2029 UBE2I UGGAGGAACGUGUGGAGUU NM_003345
    2030 UBE2I ACAGAGUGGAGUACGAGAA NM_194259
    2031 UBE2I GCCAAAACAGAGUGGAGUA NM_194259
    2032 UBE2I GCCCAGGAGAGGAAAGCAU NM_194259
    2033 UBE2I GGAGGAAAGACCACCCAUU NM_194260
    2034 UBE2I AAGCAGAGGCCUACACGAU NM_003345
    2035 UBE2I GAGAGGAAAGCAUGGAGGA NM_194259
    2036 UBE2I UAUCCAAGACCCAGCUCAA NM_194259
    2037 UBE2J1 UGUAGAAUGUAUAGGGAUA NM_016021
    2038 UBE2J1 GAGUAUAAGGACAGCAUUA NM_016336
    2039 UBE2J1 AAGAAGAGCACUUGCCAAA NM_016336
    2040 UBE2J1 GCAAAUAAGCUUUAAGGCA NM_016021
    2041 UBE2J1 GCUUUAAGGCAGAAGUCAA NM_016021
    2042 UBE2J1 CAGCCUUCGUGGAGUAUAA NM_016021
    2043 UBE2J1 UGAAAGAAGCGGCAGAAUU NM_016021
    2044 UBE2J1 CCAAAGAACUGGCUAGGCA NM_016021
    2045 UBE2J1 GAAUAUAUCUGGCAAACGA NM_016021
    2046 UBE2J1 GAGAAUUACUAGUUACUCA NM_016336
    2047 UBE2J1 GGGUAUAAUCUAAGAAUUG NM_016336
    2048 UBE2J1 GUAUAGGGAUAGAAGAGUU NM_016021
    2049 UBE2J1 CGACGAAUAUAUCUGGCAA NM_016021
    2050 UBE2J1 AAAUAAGCUUUAAGGCAGA NM_016021
    2051 UBE2J1 UAGGAGAUGUACUGGAAUU NM_016021
    2052 UBE2J1 CUAUAUUGCCCUUUGAAAU NM_016336
    2053 UBE2J1 CAGAGAAGGUUGUCUACUU NM_016021
    2054 UBE2J1 CAGCAGAGUCAGAGAAGGU NM_016021
    2055 UBE2J1 CAUAGGAGAUGUACUGGAA NM_016021
    2056 UBE2J1 CCUAAACAUUCUUCAGUGA NM_016021
    2057 UBE2J1 AAGCAAAACUCCUCAAGUA NM_016021
    2058 UBE2J1 GUCAAACAUUUAUGAGGAA NM_016021
    2059 UBE2J1 GAAUGGCACUUCACGGUUA NM_016336
    2060 UBE2J1 AGUCAGAGAAGGUUGUCUA NM_016021
    2061 UBE2J1 CCAUAGGUUCUCUAGAUUA NM_016021
    2062 UBE2J1 GAAAGAAGCGGCAGAAUUG NM_016021
    2063 UBE2J1 CAAAUAAGCUUUAAGGCAG NM_016021
    2064 UBE2J1 GGCUAAUGGUCGAUUUGAA NM_016021
    2065 UBE2J1 UGGCAGCCUUCGUGGAGUA NM_016021
    2066 UBE2J1 GCUAAGAAUACCUCCAUGA NM_016021
    2067 UBE2J2 ACAGAAAGCACAAGACGAA NM_058167
    2068 UBE2J2 GCUCCUAGGUUUAGCUUUU NM_058167
    2069 UBE2J2 UCAGUGAGUUGAAAGGAAA NM_058167
    2070 UBE2J2 CUGAAGUCGUGGAGGAGAU NM_194457
    2071 UBE2J2 GUUGAAAGGAAAUGUGUUU NM_058167
    2072 UBE2J2 UAUUAGAUGUGGUCACUUA NM_058167
    2073 UBE2J2 ACAAAUGUGUUUACAGUCA NM_058167
    2074 UBE2J2 GUAUAGAGACGUCGGACUU NM_058167
    2075 UBE2J2 CCACCAGGCUCCUAGGUUU NM_058167
    2076 UBE2J2 GUGGAGGAGAUUAAACAAA NM_194458
    2077 UBE2J2 GUGCUGGAUUUGUAGCUUA NM_058167
    2078 UBE2J2 CGAACUUGUUUGUGAUAGU NM_194458
    2079 UBE2J2 AAGUCGUGGAGGAGAUUAA NM_058167
    2080 UBE2J2 CAGAGAAUUUCCUUUCAAA NM_194315
    2081 UBE2J2 GUGCAGAGUUUAGCAUUUA NM_194457
    2082 UBE2J2 AGUGCAGAGUUUAGCAUUU NM_058167
    2083 UBE2J2 GGGUUUGGUCUCAGCAUUU NM_058167
    2084 UBE2J2 ACGGGAGGUUUAAGUGCAA NM_194457
    2085 UBE2J2 UGGAGGAGAUUAAACAAAA NM_194458
    2086 UBE2J2 UGGCUAUUAUCAUGGAAAA NM_058167
    2087 UBE2J2 CACCAGGCUCCUAGGUUUA NM_058167
    2088 UBE2J2 UGAGCUUCAUGGUGGAGAA NM_058167
    2089 UBE2J2 GCACAAGACGAACUCAGUA NM_058167
    2090 UBE2J2 GGCCUUGUGUGCUGGAUUU NM_058167
    2091 UBE2J2 GUGUUUACAGUCAGAAUGA NM_058167
    2092 UBE2J2 UUAUGAAGGUGGCUAUUAU NM_058167
    2093 UBE2J2 AGACGAACUCAGUAGCAGA NM_194315
    2094 UBE2J2 GUGGCUAUUAUCAUGGAAA NM_194316
    2095 UBE2J2 CCAGAGAAUUUCCUUUCAA NM_194315
    2096 UBE2J2 CCCAGUAUCUAUAUGAUCA NM_194458
    2097 UBE2L3 CCUCAGUUCUUGUGUGAAA NM_003347
    2098 UBE2L3 UGAAGGAGCUUGAAGAAAU NM_003347
    2099 UBE2L3 UAAGAAUGCUGAAGAGUUU NM_003347
    2100 UBE2L3 GCUGAAGAGUUUACAAAGA NM_003347
    2101 UBE2L3 CCGCAAAUGUGGGAUGAAA NM_003347
    2102 UBE2L3 UGAUGAAUCUCGCCAGAAA NM_003347
    2103 UBE2L3 ACAAAGAAAUAUGGGGAAA NM_003347
    2104 UBE2L3 CUGAAGAGUUUACAAAGAA NM_003347
    2105 UBE2L3 GAAAAGUGAUGAUGGAUUU NM_003347
    2106 UBE2L3 CCAGAAUGUCCGUUUGAAA NM_003347
    2107 UBE2L3 GUGGAGAAGAUGACUUAAA NM_003347
    2108 UBE2L3 GAUGAAGGAGCUUGAAGAA NM_198157
    2109 UBE2L3 GACUAGAUCCUGUGGAGAA NM_003347
    2110 UBE2L3 CAGAGAAGCCCUAUAAUCA NM_003347
    2111 UBE2L3 GCACAGAGAAGCCCUAUAA NM_003347
    2112 UBE2L3 CCACCGAAGAUCACAUUUA NM_003347
    2113 UBE2L3 GCAGAAAAGUGAUGAUGGA NM_003347
    2114 UBE2L3 CCUCAUAGCACUGGUGAAU NM_003347
    2115 UBE2L3 GUAAGAAUGCUGAAGAGUU NM_003347
    2116 UBE2L3 UGACCUAGCUGAAGAAUAC NM_003347
    2117 UBE2L3 CCUUCAGAAUCGAAAUCAA NM_003347
    2118 UBE2L3 GAGAAGAUGACUUAAACUU NM_003347
    2119 UBE2L3 GCAGGAGGCUGAUGAAGGA NM_003347
    2120 UBE2L3 GCCAGUAAUUAGUGCCGAA NM_003347
    2121 UBE2L3 GGGCUGACCUAGCUGAAGA NM_003347
    2122 UBE2L3 ACCCAAACAUCGACGAAAA NM_003347
    2123 UBE2L3 AAGCAGGACUCUGUGGAAA NM_003347
    2124 UBE2L3 GCUUGAAGAAAUCCGCAAA NM_003347
    2125 UBE2L3 CCAGGUUGAUGAAGCUAAU NM_003347
    2126 UBE2L3 GGUUGAUGAAGCUAAUUUA NM_003347
    2127 UBE2L6 GGACGAGAACGGACAGAUU NM_004223
    2128 UBE2L6 GUGAAUAGACCGAAUAUCA NM_004223
    2129 UBE2L6 UGAAGGAGCUGGAGGAUCU NM_004223
    2130 UBE2L6 CCUCAGACUGUGAAGUAUA NM_004223
    2131 UBE2L6 GAAAGAAUGCCGAAGAGUU NM_004223
    2132 UBE2L6 AAACCAUGUUCUUGCUUAA NM_004223
    2133 UBE2L6 GAGUUUAAGUUUGCAGUUA NM_004223
    2134 UBE2L6 CUUCUUAGGUUGUUAGUCA NM_004223
    2135 UBE2L6 GGUUGUUAGUCAUUAGUUU NM_004223
    2136 UBE2L6 GCACUGGAUCCUCGGCAUA NM_004223
    2137 UBE2L6 AGUGAGAACUGGAAGCCUU NM_004223
    2138 UBE2L6 GAGUGUGUGUGUUGUGUAU NM_004223
    2139 UBE2L6 GUCACAAUCUGAAGAAUCA NM_004223
    2140 UBE2L6 AUCCGGAGCUGUUCAGAAA NM_004223
    2141 UBE2L6 UGAUGGGAAUAUACAGAUU NM_004223
    2142 UBE2L6 CCGGAGCUGUUCAGAAAGA NM_004223
    2143 UBE2L6 GCUGGUGAAUAGACCGAAU NM_004223
    2144 UBE2L6 GUUCAGAAAGAAUGCCGAA NM_004223
    2145 UBE2L6 GUUUAAGUUUGCAGUUACA NM_004223
    2146 UBE2L6 UCAGAAAGAAUGCCGAAGA NM_004223
    2147 UBE2L6 CCAAGCCACUGAUGGGAAU NM_004223
    2148 UBE2L6 AAAGAAUGCCGAAGAGUUC NM_004223
    2149 UBE2L6 GGCCUCAGACUGUGAAGUA NM_004223
    2150 UBE2L6 UGAUCAAAUUCACAACCAA NM_004223
    2151 UBE2L6 GGACCAGGCCUCAGACUGU NM_004223
    2152 UBE2L6 GAAACCAUGUUCUUGCUUA NM_004223
    2153 UBE2L6 CAUCAUCAGCAGUGAGAAC NM_004223
    2154 UBE2L6 GCAUGCGAGUGGUGAAGGA NM_004223
    2155 UBE2L6 GGAGCUGUUCAGAAAGAAU NM_004223
    2156 UBE2L6 GUAUAUAUCCUCCAGCAUU NM_004223
    2157 UBE2M ACAUUGACCUCGAGGGCAA NM_003969
    2158 UBE2M CAGAGGUCCUGCAGAACAA NM_003969
    2159 UBE2M CCGAGGACCCACUGAACAA NM_003969
    2160 UBE2M GGAAGUUUGUGUUCAGUUU NM_003969
    2161 UBE2M AGGUGAAGUGUGAGACAAU NM_003969
    2162 UBE2M CUACAAGAGUGGGAAGUUU NM_003969
    2163 UBE2M GGAUCCAGAAGGACAUAAA NM_003969
    2164 UBE2M GAAAUAGGGUUGGCGCAUA NM_003969
    2165 UBE2M CUGCCCAAGACGUGUGAUA NM_003969
    2166 UBE2M UUAAGGUGGGCCAGGGUUA NM_003969
    2167 UBE2M AGAAGAAGGAGGAGGAGUC NM_003969
    2168 UBE2M AGCCAGUCCUUACGAUAAA NM_003969
    2169 UBE2M GAUGAGGGCUUCUACAAGA NM_003969
    2170 UBE2M AGGACAUAAACGAGCUGAA NM_003969
    2171 UBE2M CAAGGUGAAGUGUGAGACA NM_003969
    2172 UBE2M CUGCGGAUCCAGAAGGACA NM_003969
    2173 UBE2M AAGCCAGUCCUUACGAUAA NM_003969
    2174 UBE2M GAAGCCAGUCCUUACGAUA NM_003969
    2175 UBE2M GAGCUGAACCUGCCCAAGA NM_003969
    2176 UBE2M CAGACGACCUCCUCAACUU NM_003969
    2177 UBE2M GCACCAAGGGCAGCAGCAA NM_003969
    2178 UBE2M GCCUCAACAUCCUCAGAGA NM_003969
    2179 UBE2M UGAAGCAGCAGAAGAAGGA NM_003969
    2180 UBE2M AAAUAGGGUUGGCGCAUAC NM_003969
    2181 UBE2M GAUAUCAGCUUCUCAGAUC NM_003969
    2182 UBE2M GCCCAAGACGUGUGAUAUC NM_003969
    2183 UBE2M UAAUUUAUGGCCUGCAGUA NM_003969
    2184 UBE2M GGACAUAAACGAGCUGAAC NM_003969
    2185 UBE2M CCACGUGCCCCUAGUUAUU NM_003969
    2186 UBE2M UUUAACACCAGGCUAACUA NM_003969
    2187 UBE2N CAACGAAGCCCAAGCCAUA NM_003348
    2188 UBE2N GGACACAGUCUUAGAAACA NM_003348
    2189 UBE2N GGCUAUAUGCCAUGAAUAA NM_003348
    2190 UBE2N AGACAAGUUGGGAAGAAUA NM_003348
    2191 UBE2N GAAGGUAGUUGUCAGGUUA NM_003348
    2192 UBE2N CAUCUGGAUUGUUGUGAAA NM_003348
    2193 UBE2N CAUCCUGGUUUAAGUAUAA NM_003348
    2194 UBE2N GGGAGGUAGUUUAAUUUUA NM_003348
    2195 UBE2N CCGAACCAGAUGAGAGCAA NM_003348
    2196 UBE2N CAGAUGAUCCAUUAGCAAA NM_003348
    2197 UBE2N GGACUAGGCUAUAUGCCAU NM_003348
    2198 UBE2N GUAGACAAGUUGGGAAGAA NM_003348
    2199 UBE2N UAUAAAGCACUGUGAAUGA NM_003348
    2200 UBE2N GCAUCAAAGCCGAACCAGA NM_003348
    2201 UBE2N CAUCAAAGCCGAACCAGAU NM_003348
    2202 UBE2N GGGAGGGACUUUUAAACUU NM_003348
    2203 UBE2N GGGAAGAAUAUGUUUAGAU NM_003348
    2204 UBE2N ACACAGUCUUAGAAACAUU NM_003348
    2205 UBE2N UCUGGAAACUUCUGUAAAU NM_003348
    2206 UBE2N CCAGAUGAUCCAUUAGCAA NM_003348
    2207 UBE2N CUAAUAACAUUGCUGUCAA NM_003348
    2208 UBE2N AGCCCAAGCCAUAGAAACA NM_003348
    2209 UBE2N CAAAUGAUGUAGCGGAGCA NM_003348
    2210 UBE2N CCAUCCUGGUUUAAGUAUA NM_003348
    2211 UBE2N GAUCUAAUUGCAUUGGUUA NM_003348
    2212 UBE2N GAGCAGAGGCUAGAAGUAU NM_003348
    2213 UBE2N UAACCAGGUCUUUAGAAUA NM_003348
    2214 UBE2N AUAGAAACAGCUAGAGCAU NM_003348
    2215 UBE2N AGAUGAUCCAUUAGCAAAU NM_003348
    2216 UBE2N AACCAGGUCUUUAGAAUAU NM_003348
    2217 UBE2Q AUGAAGGAGCUCAGGGAUA NM_017582
    2218 UBE2Q CAGAAGACUUAGAUCACUA NM_017582
    2219 UBE2Q CCAUAGAGUCAGUGAUCAU NM_017582
    2220 UBE2Q AUGAAAUGAAAGAGGAAGA NM_017582
    2221 UBE2Q CUAUGAAAUGAAAGAGGAA NM_017582
    2222 UBE2Q CCAGAUCCUCAAAGAGAAA NM_017582
    2223 UBE2Q UGGAGAGGCUGGUGGACAU NM_017582
    2224 UBE2Q CCUCAAAGAGAAAGAAGGA NM_017582
    2225 UBE2Q AAGAUGAAGAUGAGGAGAU NM_017582
    2226 UBE2Q GCUGGUGGACAUAAAGAAA NM_017582
    2227 UBE2Q AGAUGAUGGCAUUGGAAAA NM_017582
    2228 UBE2Q CUUAAAUGGUGCAGUGUCU NM_017582
    2229 UBE2R2 GGAAAUGGAGAGACAGUAA NM_017811
    2230 UBE2R2 CCACAACCCUGGCGGAAUA NM_017811
    2231 UBE2R2 CAGUAAAGGAAAAGACAAA NM_017811
    2232 UBE2R2 CGACAUUGAUGAUGAAGAU NM_017811
    2233 UBE2R2 GGAGAGACAGUAAAGGAAA NM_017811
    2234 UBE2R2 GGAUCGAAAGCCUGAAAUA NM_017811
    2235 UBE2R2 GCAUAUACUGGGUAGCAAA NM_017811
    2236 UBE2R2 GUGCUAAGCCUGAUGAAAU NM_017811
    2237 UBE2R2 AGACAAAGAAUAUGCUGAA NM_017811
    2238 UBE2R2 AGGAAGAUGCCGACUGUUA NM_017811
    2239 UBE2R2 AAGAUGAGGAGGAGGAAGA NM_017811
    2240 UBE2R2 CCACUAAGGCCGAAGCAGA NM_017811
    2241 UBE2R2 GAGAGACAGUAAAGGAAAA NM_017811
    2242 UBE2R2 GUUCACAAGUGCUGCAUAU NM_017811
    2243 UBE2R2 UCAGUUAUGUUCAGGAAAU NM_017811
    2244 UBE2R2 GACCUUUAAUGGAGAGAGA NM_017811
    2245 UBE2R2 UCUGAAAGGUGGAAUCCUA NM_017811
    2246 UBE2R2 AGGUUUAGCUGCUCAUUUA NM_017811
    2247 UBE2R2 GAUCGAAAGCCUGAAAUAA NM_017811
    2248 UBE2R2 CAUCAGGGACUUUGUGCUA NM_017811
    2249 UBE2R2 GAGUAACCCUCCACAGAAU NM_017811
    2250 UBE2R2 GCGGAAUACUGCAUCAAAA NM_017811
    2251 UBE2R2 CCUUCAGAUUCUUGACCAA NM_017811
    2252 UBE2R2 UUAUGAGAAUGGAGAUGUA NM_017811
    2253 UBE2R2 AUCGAAAGCCUGAAAUAAA NM_017811
    2254 UBE2R2 UCGAAAGCCUGAAAUAAAU NM_017811
    2255 UBE2R2 ACAGAAUGUUCACAGCAAA NM_017811
    2256 UBE2R2 GUGGAAUCCUACUCAGAAU NM_017811
    2257 UBE2R2 CCAUGAAACCAUCGUAACA NM_017811
    2258 UBE2R2 GCUGAAAUUAUUAGGAAAC NM_017811
    2259 UBE2S UGGAGAACUACGAGGAGUA NM_014501
    2260 UBE2S GCAUCAAGGUCUUUCCCAA NM_014501
    2261 UBE2S CCAAGAAGCAUGCUGGCGA NM_014501
    2262 UBE2S UCAACGUGCUCAAGAGGGA NM_014501
    2263 UBE2V2 GGAAAUAGGUGUAUGGAUA NM_003350
    2264 UBE2V2 GGACAAACAUACAACAAUU NM_003350
    2265 UBE2V2 GAUGAAAGCGUGUGGAGAA NM_003350
    2266 UBE2V2 CUAAUGAUGUCCAAAGAAA NM_003350
    2267 UBE2V2 GGAAGAACUUGAAGAAGGA NM_003350
    2268 UBE2V2 CAGAAUAUAUAGCCUGAAA NM_003350
    2269 UBE2V2 CGAAAUAGAAUUCAGGUUU NM_003350
    2270 UBE2V2 GUGUACUUCUUGUGAAUUA NM_003350
    2271 UBE2V2 AGUAGAAUGUGGACCUAAA NM_003350
    2272 UBE2V2 AGUUGUACUUCAAGAGCUA NM_003350
    2273 UBE2V2 CCUCAGAGACUGUGCCAUU NM_003350
    2274 UBE2V2 GAGUUAAAGUUCCUCGUAA NM_003350
    2275 UBE2V2 AAAGAGAGCUGCAGUUGAA NM_003350
    2276 UBE2V2 AGUUAAAGUUCCUCGUAAU NM_003350
    2277 UBE2V2 GUUCAGGAUUGUUUUGAUA NM_003350
    2278 UBE2V2 GCACUGUCAUUUAAACAUA NM_003350
    2279 UBE2V2 AGGCAUGAUUAUUGGGCCA NM_003350
    2280 UBE2V2 GAAAUAGGUGUAUGGAUAU NM_003350
    2281 UBE2V2 GUUGGAAGAACUUGAAGAA NM_003350
    2282 UBE2V2 GAUGAAGAUAUGACACUUA NM_003350
    2283 UBE2V2 AUAAAUAAUUCCAGUGGGA NM_003350
    2284 UBE2V2 CCAAGGACAAAUUAUGAAA NM_003350
    2285 UBE2V2 GCAUACCAGUGUUAGCAAA NM_003350
    2286 UBE2V2 AAGCUGUUCUUGUGUGUUA NM_003350
    2287 UBE2V2 CUGGACAUUUGUAAGAAUA NM_003350
    2288 UBE2V2 AGAACUUGAAGAAGGACAA NM_003350
    2289 UBE2V2 ACAGAAAUGGCAUGCUUUA NM_003350
    2290 UBE2V2 CUUACAAGGUGGACAGGCA NM_003350
    2291 UBE2V2 AAGCAUGUGUGUUUCUAAA NM_003350
    2292 UBE2V2 AGUGGAAGCAUGUGUGUUU NM_003350
    2293 UBE3A AGACAAAGAUGAAGAUGAA NM_000462
    2294 UBE3A GUAGAGAAAGAGAGGAUUA NM_130838
    2295 UBE3A CCUCAGAACUUUAGUAACA NM_130838
    2296 UBE3A UAACAGAAGAGAAGGUAUA NM_130839
    2297 UBE3A AAGUAGAAACAGAGAACAA NM_000462
    2298 UBE3A GGAAUUUGAAGGAGAACAA NM_130839
    2299 UBE3A GGGAGAAAUUGGAAUGUUU NM_130839
    2300 UBE3A GUAAUUAAAGUGAGGAGAA NM_000462
    2301 UBE3A GGAGAAGAAAGAAGAAACA NM_000462
    2302 UBE3A CAACAAAGUUAGGAAGUUU NM_000462
    2303 UBE3A CAAAUGAAAUGGUAGUCAA NM_000462
    2304 UBE3A CAAGAAAGGCGCUAGAAUU NM_130839
    2305 UBE3A AGCAAAAGAUGAAGACAAA NM_000462
    2306 UBE3A AGGCAUUGGUACAGAGCUU NM_000462
    2307 UBE3A GCACCAGUGUUAUUGGAAA NM_000462
    2308 UBE3A GGAAGAAGACUCAGAAGCA NM_130838
    2309 UBE3A GAAGAGAAGGUAUAUGAAA NM_130839
    2310 UBE3A GUUCAAGGCUUUUCGGAGA NM_130839
    2311 UBE3A UCAGAAGUUUGGCGAAAUA NM_130839
    2312 UBE3A CAGUCGAAAUCUAGUGAAU NM_000462
    2313 UBE3A GCGUGAAAGUGUUACAUAU NM_000462
    2314 UBE3A UGAAUGAGGUUCUAGAAAU NM_000462
    2315 UBE3A GAGAUGAUCGCUAUGGAAA NM_000462
    2316 UBE3A AGAUGAAGACAAAGAUGAA NM_000462
    2317 UBE3A GAAGAAAGAAGAAACAAGA NM_130838
    2318 UBE3A GGUGAUUGAUUGAUUGAUU NM_130838
    2319 UBE3A AUUAGGGAGUUCUGGGAAA NM_130839
    2320 UBE3A CAAUGAAGAAGAUGAUGAA NM_000462
    2321 UBE3A AAGCAAAAGAUGAAGACAA NM_000462
    2322 UBE3A GCUCACAGGGAGACAACAA NM_130839
    2323 UBE3B AAGAAAGGCUUGUGCAGAA NM_130466
    2324 UBE3B GUGCAAAUAUAAUGGGACA NM_183415
    2325 UBE3B GGAGAGAGAUUGAUGACUU NM_130466
    2326 UBE3B CCACAGUCCUUCAGAGUUU NM_130466
    2327 UBE3B CAAAGAGGAUAAUGAGAGA NM_130466
    2328 UBE3B AGAGAUAUCAGGAGAGAGA NM_130466
    2329 UBE38 GCACAGGGCUGCAGAAAAU NM_130466
    2330 UBE3B GGAACAAACACUAACCUAA NM_130466
    2331 UBE3B CCUACAUCCAUGAGAAUUA NM_130466
    2332 UBE3B CAGAUGGGUUCGUGAGUUU NM_130466
    2333 UBE3B GGAAACAUGGCAAGGAAUU NM_130466
    2334 UBE3B UCAGAAUCAAAGAGGAUAA NM_130466
    2335 UBE3B UGGAAGAGCUGGUCACUAU NM_130466
    2336 UBE3B GGGAGAGAGUGAUUUAUAA NM_130466
    2337 UBE3B GGAUGGAAUUGUAGAGAAC NM_183414
    2338 UBE3B GGGAUGGAAUUGUAGAGAA NM_130466
    2339 UBE3B GCAGAGAGAUAUCAGGAGA NM_183414
    2340 UBE38 UGUUAGAGGAGGAGACAGA NM_183415
    2341 UBE3B GCAGAAGUCCAGAAGGUUU NM_130466
    2342 UBE3B AGAUAUCAGGAGAGAGAUU NM_183414
    2343 UBE3B CCGUCAGGCACGAGAAGAA NM_183414
    2344 UBE3B GGGAAAAGGUGAAAGUCUU NM_183414
    2345 UBE3B GAAUCAAAGAGGAUAAUGA NM_183414
    2346 UBE3B CAACAGAUCAAGAACAUUU NM_130466
    2347 UBE3B CCAGAGUGUUAGAGGAGGA NM_130466
    2348 UBE3B GUGUGAAGUUUGUCAAUGA NM_130466
    2349 UBE3B CAACAUGCCAGGUGACAUU NM_130466
    2350 UBE3B UCAAGAACAUUUUGUGGUA NM_183414
    2351 UBE3B CAGCAUGGGUUGAGUGUAC NM_130466
    2352 UBE3B CAGAAGAAGUCCAACCUGA NM_183414
    2353 UBE4A GGAGAAAACUGGAGGAAAA NM_004788
    2354 UBE4A UGACAGACCAGGAGAAUAA NM_004788
    2355 UBE4A GGAAAUGAACCUAGAAACA NM_004788
    2356 UBE4A AGAAAUGGCAGUAGAGCUA NM_004788
    2357 UBE4A GAAUAGAUACUGUGAACUA NM_004788
    2358 UBE4A CAGUAGAGCUAGAAGAUCA NM_004788
    2359 UBE4A CAUUAAUGCUGCUGAAUAU NM_004788
    2360 UBE4A GCAACUUGGCAGAGAGAAU NM_004788
    2361 UBE4A CAGAGUAACUUCACAUAAA NM_004788
    2362 UBE4A GCAGGAAAUAUGUGAGCAA NM_004788
    2363 UBE4A GCAUAGAAAGAAUGAAGAA NM_004788
    2364 UBE4A GGAGCAACUUGAAUAGAUA NM_004788
    2365 UBE4A GAGCAGACUUUCUAACAUA NM_004788
    2366 UBE4A GAUAAUAGCGUGUCAGAGA NM_004788
    2367 UBE4A UGAGAAGGAUCGAGGUGAA NM_004788
    2368 UBE4A GGAAUAUGAUUAUGGCUUU NM_004788
    2369 UBE4A GGGAGAGCAUUAAGGAUUU NM_004788
    2370 UBE4A CGGGAGAGCAUUAAGGAUU NM_004788
    2371 UBE4A AUAAAUAAGCCUGGGAAUA NM_004788
    2372 UBE4A GGGCAAAUGUACCAGAAGA NM_004788
    2373 UBE4A CCAAGAGGCUAAUACUAAA NM_004788
    2374 UBE4A GAGGAUACCUUAAAGUGAA NM_004788
    2375 UBE4A GCAUUAAGGAUUUGGCUGA NM_004788
    2376 UBE4A GGAGUUGAAUGAUGAAGAA NM_004788
    2377 UBE4A UCAAAGUACAGGAGGCCAA NM_004788
    2378 UBE4A ACAACAACAUCUCAAGUAA NM_004788
    2379 UBE4A AGUCAUGAUUCCAGUGUUU NM_004788
    2380 UBE4A GGCCAAACACAGAACUAAA NM_004788
    2381 UBE4A UAACUUACCAAGAGGCUAA NM_004788
    2382 UBE4A UGUGUAGACUUCUGAAUAA NM_004788
    2383 UBE4B AGGAAGAGAUGAAGAACAA NM_006048
    2384 UBE4B CAAUAGAACUGUAGAAGAU NM_006048
    2385 UBE4B CCGAGAAAGUGGAGGAGAU NM_006048
    2386 UBE4B ACAGAAAUCUCUUGCUAAA NM_006048
    2387 UBE4B GAAAAUGAUCGAAGAGAAA NM_006048
    2388 UBE4B AGACAGAUAUGCUGAACUA NM_006048
    2389 UBE4B CAAAGAAGCUGUUGGACCA NM_006048
    2390 UBE4B UUGAAAACCCUGAGAAAUA NM_006048
    2391 UBE4B AGAAGAUGUUGCAGAAUUU NM_006048
    2392 UBE4B CGAUGGAAGAUGUGAAUGA NM_006048
    2393 UBE4B CGGCAGACGCUGACAGAGA NM_006048
    2394 UBE4B GAAAGGACAUGGAUGAGAA NM_006048
    2395 UBE4B GGAGCAAGUAAUUGGGAUU NM_006048
    2396 UBE4B GAGUUGGAAUAGAGGAAAA NM_006048
    2397 UBE4B GCAACUAGACACCGCGAAA NM_006048
    2398 UBE4B GGAGAUGAGGGUUGGGUUU NM_006048
    2399 UBE4B UGUAGAAGAUGUUGCAGAA NM_006048
    2400 UBE4B CGAGAAAGUGGAGGAGAUA NM_006048
    2401 UBE4B UAAUGAAAGCCAAUGGAAA NM_006048
    2402 UBE4B UGACACACGUUCAGAAGAA NM_006048
    2403 UBE4B GGACAGACCUCUCAGCCAA NM_006048
    2404 UBE4B CCGAGUUGGAAUAGAGGAA NM_006048
    2405 UBE4B UGGAGGAGAUAGUGGCCAA NM_006048
    2406 UBE4B AAGCGGAGCCUCAGUGAUA NM_006048
    2407 UBE4B CUGCAAUGCUGAACUUUAA NM_006048
    2408 UBE4B GAACUUUAAUCUUCAGCAA NM_006048
    2409 UBE4B UGAAAAUGAUCGAAGAGAA NM_006048
    2410 UBE4B UUGAAAUGAUUGAGAACCA NM_006048
    2411 UBE4B GCUCAAUAGAACUGUAGAA NM_006048
    2412 UBE4B ACAUGGAGGUUGAUGAAAA NM_006048
    2413 UBL3 GAGAGUAAUUGUUGUGUAA NM_007106
    2414 UBL3 AGACAUUACCAGAGCCAAA NM_007106
    2415 UBL3 GAAAUAAACUGGUUCACUA NM_007106
    2416 UBL3 UGAGAAGACUGGAGAGAGU NM_007106
    2417 UBL3 CCAAAUAUUCUACGACUUA NM_007106
    2418 UBL3 GUACAUAAAGCCUGAAUGA NM_007106
    2419 UBL3 GGUCAGAGGAAUCGUGAGA NM_007106
    2420 UBL3 GCACAGAAUACUAAACUGA NM_007106
    2421 UBL3 GCACAGCACUAAAGCAUGA NM_007106
    2422 UBL3 UCACAACAAUGGAUAGAAU NM_007106
    2423 UBL3 GCAUAGAUCCAGUAAUGUA NM_007106
    2424 UBL3 GUAUAUGACAAUUGGCCAA NM_007106
    2425 UBL3 GUAAAUAGCACUAAACGUU NM_007106
    2426 UBL3 GUUAAUUUGUGGACAGUCA NM_007106
    2427 UBL3 GAGCAGAGUUUUAAAAUGA NM_007106
    2428 UBL3 CAUAAAGCCUGAAUGAUGA NM_007106
    2429 UBL3 GAUAUUAGUGCUUGCAAUU NM_007106
    2430 UBL3 AAACAACAGUGAUGCAUUU NM_007106
    2431 UBL3 GAAAGAAUUACGUCGCUUA NM_007106
    2432 UBL3 GUAACUGAGUGUUUGUUUA NM_007106
    2433 UBL3 AGAAGACUGGAGAGAGUAA NM_007106
    2434 UBL3 GGUCAGCAGUCCAAAUAUU NM_007106
    2435 UBL3 CAGCGUUGUUUAAAUGUAA NM_007106
    2436 UBL3 GUAAACACAUGGAACUGAA NM_007106
    2437 UBL3 UGUCACAUUAGGAGCAUUA NM_007106
    2438 UBL3 GGUGAGAGCUGCAUAGAUC NM_007106
    2439 UBL3 CAAAGCAUGUAUAUGACAA NM_007106
    2440 UBL3 CGACAUUGCUUCAGAAACC NM_007106
    2441 UBL3 GAGUAAUUGUUGUGUAAUC NM_007106
    2442 UBL3 UGUGUGGUAUCUAGAAAUA NM_007106
    2443 UBL4 GGAAACGACUCUCGGAUUA NM_014235
    2444 UBL4 AGACAAUGGAGAAGGGCUU NM_014235
    2445 UBL4 CUGAAGUGACUGAGACAAU NM_014235
    2446 UBL4 GAACAGCUACAGAGGGAUU NM_014235
    2447 UBL4 UGGAACAGCUACAGAGGGA NM_014235
    2448 UBL4 UGACUGAGACAAUGGAGAA NM_014235
    2449 UBL4 GGCCCUGGCAGAUGGGAAA NM_014235
    2450 UBL4 GGGAAACGACUCUCGGAUU NM_014235
    2451 UBL4 CUACAGAGGGAUUACGAGA NM_014235
    2452 UBL4 GAAGGUGCUACUAGAAGAA NM_014235
    2453 UBL4 AGAAGGUGCUACUAGAAGA NM_014235
    2454 UBL4 AGCUCAACCUAGUGGUCAA NM_014235
    2455 UBL4 GGAGAAGGUGCUACUAGAA NM_014235
    2456 UBL4 UAGAAGAAGGCGAGGCCCA NM_014235
    2457 UBL4 UGGAGAAGGUGCUACUAGA NM_014235
    2458 UBL4 CAGCUGAUCUCCAAAGUCU NM_014235
    2459 UBL4 ACUCCAAGCUCAACCUAGU NM_014235
    2460 UBL4 GAAACGACUCUCGGAUUAU NM_014235
    2461 UBL4 GCAGAUGGGAAACGACUCU NM_014235
    2462 UBL4 GAGAAGGUGCUACUAGAAG NM_014235
    2463 UBL4 AAACGACUCUCGGAUUAUA NM_014235
    2464 UBL4 UCUGGCAGCUGAUCUCCAA NM_014235
    2465 UBL4 UGAAGUGACUGAGACAAUG NM_014235
    2466 UBL4 GCUGAUCUCCAAAGUCUUG NM_014235
    2467 UBL4 GGCAGCUGAUCUCCAAAGU NM_014235
    2468 UBL4 GAAGUGACUGAGACAAUGG NM_014235
    2469 UBL4 CAACUCCAAGCUCAACCUA NM_014235
    2470 UBL5 GGAUGAACCUGGAGCUUUA NM_024292
    2471 UBL5 GGGAAGAAGGUCCGCGUUA NM_024292
    2472 UBL5 GAUAGAUGCUUGUUUGUAA NM_024292
    2473 UBL5 GAAGAAGGUCCGCGUUAAA NM_024292
    2474 UBL5 GGAAGAAGGUCCGCGUUAA NM_024292
    2475 UBL5 CUGAAGAAGUGGUACACGA NM_024292
    2476 UBL5 AAGAAGUGGUACACGAUUU NM_024292
    2477 UBL5 UGAUCGAGGUUGUUUGCAA NM_024292
    2478 UBL5 CUAGGAUGAUCGAGGUUGU NM_024292
    2479 UBL5 GGGAUGAACCUGGAGCUUU NM_024292
    2480 UBL5 GCAACACGGAUGAUACCAU NM_024292
    2481 UBL5 UGAAGAAGUGGUACACGAU NM_024292
    2482 UBL5 GAACCUGGAGCUUUAUUAU NM_024292
    2483 UBL5 UUAAAUGCAACACGGAUGA NM_024292
    2484 UBL5 CCUGGAGCUUUAUUAUCAA NM_024292
    2485 UBL5 GAAGAAGUGGUACACGAUU NM_024292
    2486 UBL5 GGUACCCGUUGGAACAAGA NM_024292
    2487 UBL5 GGAUAGAUGCUUGUUUGUA NM_024292
    2488 UBL5 AAGAUUGUCCUGAAGAAGU NM_024292
    2489 UBL5 ACCUGGAGCUUUAUUAUCA NM_024292
    2490 UBL5 AAAUGCAACACGGAUGAUA NM_024292
    2491 UBL5 UGGGAUAGAUGCUUGUUUG NM_024292
    2492 UBL5 AGAUUGUCCUGAAGAAGUG NM_024292
    2493 UBL5 UGAACCUGGAGCUUUAUUA NM_024292
    2494 UBL5 GACCUUAAGAAGCUGAUUG NM_024292
    2495 UBL5 AGAAGGUCCGCGUUAAAUG NM_024292
    2496 UBL5 AAGAAGGUCCGCGUUAAAU NM_024292
    2497 UBL5 GAAGGUCCGCGUUAAAUGC NM_024292
    2498 UBL5 GAUGAUCGAGGUUGUUUGC NM_024292
    2499 UBR1 GGAUGAAUAUGGAGAAACA NM_174916
    2500 UBR1 UCAAAUAGCAUCAAGGAAA NM_174916
    2501 UBR1 GAUUAUGGCUCAUCAGAAA NM_174916
    2502 UBR1 GUGAAGCGAUUAAGAGAAA NM_174916
    2503 UBR1 CUGCAGAUGAUGAGCGAAA NM_174916
    2504 UBR1 GGACAAAUUGGGAAGAGUA NM_174916
    2505 UBR1 GGACAGAAAGAUUAAGAAU NM_174916
    2506 UBR1 AGAAAUGACUAUAUGGGAA NM_174916
    2507 UBR1 CUUGGAAAGUGGAGAAUAU NM_174916
    2508 UBR1 GAACAUAUGCAGAAGAAAA NM_174916
    2509 UBR1 AAGAAGAACAGGAGGUGAA NM_174916
    2510 UBR1 CCAGAAAUCUACUGCCUUA NM_174916
    2511 UBR1 GCUUAGAGAAUGUCAUAAA NM_174916
    2512 UBR1 GGGAAGAGUAUAUGCAGUA NM_174916
    2513 UBR1 GGUCCUGGUUGAAGGUAAA NM_174916
    2514 UBR1 CAAAUAGCAUCAAGGAAAU NM_174916
    2515 UBR1 CUAUGGAAUUUGUGAAGUA NM_174916
    2516 UBR1 GGAGAAAACAAGAAAACAA NM_174916
    2517 UBR1 UAGAGGUACUAGUGGAAUA NM_174916
    2518 UBR1 ACAAAGUCCUACAGAGUAU NM_174916
    2519 UBR1 AGGAAGAUAUAAAGAGUCA NM_174916
    2520 UBR1 AUAAACAACUGCAGAAAGA NM_174916
    2521 UBR1 GCUGAGAUGUGGCGAAGAA NM_174916
    2522 UBR1 CCAAAGAAGAGGUCACAAU NM_174916
    2523 UBR1 CCAAAUGGCUUUUCAUAUU NM_174916
    2524 UBR1 CUAUAUGGGAAGAGGAAAA NM_174916
    2525 UBR1 UUUCAAAAGUGGAGAGACA NM_174916
    2526 UBR1 CAAUAUGGACAGAAAGAUU NM_174916
    2527 UBR1 GGUACUGAGAGGAUGGAAA NM_174916
    2528 UBR1 CCGAAGACAGGUUGGGCAA NM_174916
    2529 UCHL1 GGCCAAUAAUCAAGACAAA NM_004181
    2530 UCHL1 CAUGAGAACUUCAGGAAAA NM_004181
    2531 UCHL1 UUGAAGAGCUGAAGGGACA NM_004181
    2532 UCHL1 GGGUAGAUGACAAGGUGAA NM_004181
    2533 UCHL1 CCGCGAAGAUGCAGCUCAA NM_004181
    2534 UCHL1 GCCAAGGUCUGCAGAGAAU NM_004181
    2535 UCHL1 GAAGCAGACCAUUGGGAAU NM_004181
    2536 UCHL1 GCAUGAGAACUUCAGGAAA NM_004181
    2537 UCHL1 AGACCUUGGAUGUGGUUUA NM_004181
    2538 UCHL1 GCUGAAGGGACAAGAAGUU NM_004181
    2539 UCHL1 CUAUGAACUUGAUGGACGA NM_004181
    2540 UCHL1 CUAAAAUGCUUCAGUACUU NM_004181
    2541 UCHL1 GGAUGUGGUUUAAUUGUUU NM_004181
    2542 UCHL1 CCAAUAAUCAAGACAAACU NM_004181
    2543 UCHL1 CGGCCCAGCAUGAGAACUU NM_004181
    2544 UCHL1 UAGAUGACAAGGUGAAUUU NM_004181
    2545 UCHL1 CCCUGAAGACAGAGCAAAA NM_004181
    2546 UCHL1 CAGCUCAAGCCGAUGGAGA NM_004181
    2547 UCHL1 CCGAGAUGCUGAACAAAGU NM_004181
    2548 UCHL1 GGGAUUUGAGGAUGGAUCA NM_004181
    2549 UCHL1 GGUUUCUGUCUGUAAGUUA NM_004181
    2550 UCHL1 GUUUCUGUCUGUAAGUUAA NM_004181
    2551 UCHL1 CCUUGGAUGUGGUUUAAUU NM_004181
    2552 UCHL1 GGAGGGACUUUGCUGAUUU NM_004181
    2553 UCHL1 AAGCAGACCAUUGGGAAUU NM_004181
    2554 UCHL1 ACGCAGUGGCCAAUAAUCA NM_004181
    2555 UCHL1 UUAACAACGUGGAUGGCCA NM_004181
    2556 UCHL1 CAGAGGACACCCUGCUGAA NM_004181
    2557 UCHL1 CUGAAGGGACAAGAAGUUA NM_004181
    2558 UCHL1 GGUGUGAGCUUCAGAUGGU NM_004181
    2559 UCHL3 CAGCAUAGCUUGUCAAUAA NM_006002
    2560 UCHL3 GAACAGAAGAGGAAGAAAA NM_006002
    2561 UCHL3 CAGCAAUGCCUGUGGAACA NM_006002
    2562 UCHL3 UGGUGAAACUAGUGAUGAA NM_006002
    2563 UCHL3 GGCAAUUCGUUGAUGUAUA NM_006002
    2564 UCHL3 AUUCAGAACAGAAGAGGAA NM_006002
    2565 UCHL3 CUGAAGAACGAGCCAGAUA NM_006002
    2566 UCHL3 CCUGAUGAACUAAGAUUUA NM_006002
    2567 UCHL3 CCAAUUAACCAUGGUGAAA NM_006002
    2568 UCHL3 GAACUAAGAUUUAAUGCGA NM_006002
    2569 UCHL3 CCAUAGAAGUUUGCAAGAA NM_006002
    2570 UCHL3 CAGUAUAUUUCAUGAAGCA NM_006002
    2571 UCHL3 AAAUAAAAUCUCAGGGACA NM_006002
    2572 UCHL3 GAAGUUUGCAAGAAGUUUA NM_006002
    2573 UCHL3 ACCAAGUAUAGAUGAGAAA NM_006002
    2574 UCHL3 GCGCGACCCUGAUGAACUA NM_006002
    2575 UCHL3 UGGAACAAUUGGACUGAUU NM_006002
    2576 UCHL3 UCAGAACAGAAGAGGAAGA NM_006002
    2577 UCHL3 GUCAAUGAGCCCUGAAGAA NM_006002
    2578 UCHL3 CAAGUAUAGAUGAGAAAGU NM_006002
    2579 UCHL3 CAAUAAUGGAAACACCAAA NM_006002
    2580 UCHL3 GCACUUUGAAUCUGGAUCA NM_006002
    2581 UCHL3 ACAAUAAAGACAAGAUGCA NM_006002
    2582 UCHL3 GCAAUUCGUUGAUGUAUAU NM_006002
    2583 UCHL3 GUAUAGAUGAGAAAGUAGA NM_006002
    2584 UCHL3 CACCAAGUAUAGAUGAGAA NM_006002
    2585 UCHL3 GAACAAUUGGACUGAUUCA NM_006002
    2586 UCHL3 UCGUUGAUGUAUAUGGAAU NM_006002
    2587 UCHL3 UGAAACUAGUGAUGAAACU NM_006002
    2588 UCHL3 AGACUGAGGCACCAAGUAU NM_006002
    2589 UCHL5 GUUUAGAGCCUGAGAAUUU NM_015984
    2590 UCHL5 GCAGAAGAUAGCAGAGUUA NM_015984
    2591 UCHL5 GGAUACAGAUCAAGGUAAU NM_015984
    2592 UCHL5 GCAAAGAAAGCUCAGGAAA NM_015984
    2593 UCHL5 UGAAGAAGAAGUACAGAAA NM_015984
    2594 UCHL5 GUACAGUGAAGGUGAAAUU NM_015984
    2595 UCHL5 GUUAAUACCACUAGUAGAA NM_015984
    2596 UCHL5 GCUUAUUGAAGAAGAAGUA NM_015984
    2597 UCHL5 GUAGAAAAGGCAAAAGAAA NM_015984
    2598 UCHL5 GAGCCCAAGUAGAAGAAAU NM_015984
    2599 UCHL5 GCACAAACGAUAUUCCUUA NM_015984
    2600 UCHL5 UAGCAGAGUUACAAAGACA NM_015984
    2601 UCHL5 UAAAGAACUUAGAGCAACA NM_015984
    2602 UCHL5 CAAAGAAAGCUCAGGAAAC NM_015984
    2603 UCHL5 GAUUAAACUGGUUGUCUUA NM_015984
    2604 UCHL5 CCGAGGAGCCCAAGUAGAA NM_015984
    2605 UCHL5 GAGUUUAGAGCCUGAGAAU NM_015984
    2606 UCHL5 AGCCAUAGUGAGUGUGUUA NM_015984
    2607 UCHL5 GAGGAGGAACCCAUGGAUA NM_015984
    2608 UCHL5 AGAAGGACCGAUUGAUUUA NM_015984
    2609 UCHL5 GAGCAAUUCAGAUGUGAUU NM_015984
    2610 UCHL5 GUUAAGUGCUAUUCAGUCA NM_015984
    2611 UCHL5 AGCCCAAGUAGAAGAAAUA NM_015984
    2612 UCHL5 UCAGAUGCUUAUUGAAGAA NM_015984
    2613 UCHL5 UGAUAUAUGAGCAGAAGAU NM_015984
    2614 UCHL5 GAGAAGGACCGAUUGAUUU NM_015984
    2615 UCHL5 AGAUGUGAUUCGACAAGUA NM_015984
    2616 UCHL5 CAGUAAGGCCUGUCAUAGA NM_015984
    2617 UCHL5 GAUGCUUAUUGAAGAAGAA NM_015984
    2618 UCHL5 GAUACGAAGACAUCAGCAA NM_015984
    2619 UEV3 GGGCAAAUCAUGAGAAUAA NM_018314
    2620 UEV3 AGAAAGACCUGCUGAAUUU NM_018314
    2621 UEV3 CAACAGGGAUAUUAUGAUA NM_018314
    2622 UEV3 CGAGCAAGGAGAAGACAAA NM_018314
    2623 UEV3 GCAAAGAAGUAUGGGUUAU NM_018314
    2624 UEV3 AAGAAGAAGUAGUGAGUCU NM_018314
    2625 UEV3 GGCACAGACUUCAGGCAAA NM_018314
    2626 UEV3 GGACCUAACUGUGGAAGAA NM_018314
    2627 UEV3 GGCAAAGAAGUAUGGGUUA NM_018314
    2628 UEV3 UCACAGAGAUUACAGUAUA NM_018314
    2629 UEV3 GAUCGGAAUUGGAUGUAAU NM_018314
    2630 UEV3 UGGGAAUCUUAGUCGGAAA NM_018314
    2631 UEV3 UUACAAACUGCUUGGUUAA NM_018314
    2632 UEV3 CAGUGGAAAUCAUGACCUA NM_018314
    2633 UEV3 GUACAGAGCAAUGUGGAUA NM_018314
    2634 UEV3 GUAGAGGGUUUCUUGAUUA NM_018314
    2635 UEV3 CAGAAAGACCUGCUGAAUU NM_018314
    2636 UEV3 CGGUAUUGCCUGCACAUUA NM_018314
    2637 UEV3 GCUCAAGGCAGAAUAUAUU NM_018314
    2638 UEV3 UUUACAAACUGCUUGGUUA NM_018314
    2639 UEV3 AGAAAUGAUUGCCAAGUUU NM_018314
    2640 UEV3 CAUCUCAACCAGUGGAAAU NM_018314
    2641 UEV3 CCAAGAAGAAGUAGUGAGU NM_018314
    2642 UEV3 AAUUCAAAGAGCUGGGCAA NM_018314
    2643 UEV3 AGGCAAAGAAGUAUGGGUU NM_018314
    2644 UEV3 GAAAGAAGAUACAGUUACU NM_018314
    2645 UEV3 GGUUGGAGGUGGAGAACUC NM_018314
    2646 UEV3 UUGAUGUGGUACAGAGCAA NM_018314
    2647 UEV3 CCAAUUCGUUUCUGGAUUU NM_018314
    2648 UEV3 GCAGAAUAUAUUUGCCCUA NM_018314
    2649 UFD1L GCAAUAGACUGGAUGGAAA NM_005659
    2650 UFD1L GGAUGAAGCUGGAGGCAGA NM_005659
    2651 UFD1L GAGAAAGGAGGGAAGAUAA NM_005659
    2652 UFD1L ACAAAGAACCCGAAAGACA NM_005659
    2653 UFD1L GGAGAAAGGAGGGAAGAUA NM_005659
    2654 UFD1L AAUCAAGCCUGGAGAUAUU NM_005659
    2655 UFD1L GGAAGAAAGCCCUAAGUGA NM_005659
    2656 UFD1L CUACAAAGAACCCGAAAGA NM_005659
    2657 UFD1L CAAAGAACCCGAAAGACAA NM_005659
    2658 UFD1L GGACAGUCAUUGCGUAAAA NM_005659
    2659 UFD1L AGAAAGGAGGGAAGAUAAU NM_005659
    2660 UFD1L GGUCAGAUGUGGAGAAAGG NM_005659
    2661 UFD1L GGAGAUAUUAAAAGAGGAA NM_005659
    2662 UFD1L CCAAUCAAGCCUGGAGAUA NM_005659
    2663 UFD1L CGACAGAAGGUGAAGCCGA NM_005659
    2664 UFD1L GCCAUCAACUAUAAUGAAA NM_005659
    2665 UFD1L UGUUCAAACUGACCAAUAA NM_005659
    2666 UFD1L AAAGACAAGUCCAGCAUGA NM_005659
    2667 UFD1L AAAGGAGGGAAGAUAAUUA NM_005659
    2668 UFD1L CCAAAGCCGUAUUAGAAAA NM_005659
    2669 UFD1L AAGGACAGUCAUUGCGUAA NM_005659
    2670 UFD1L CAAACUGACCAAUAAGAAU NM_005659
    2671 UFD1L ACUGUUGGCUGAUUGGAAA NM_005659
    2672 UFD1L CAGGUCAGAUGUGGAGAAA NM_005659
    2673 UFD1L GGUAAGAUAACUUUCAUCA NM_005659
    2674 UFD1L CCCAAAGCCGUAUUAGAAA NM_005659
    2675 UFD1L CGAAAGACAAGUCCAGCAU NM_005659
    2676 UFD1L GUGACAUGAACGUGGACUU NM_005659
    2677 UFD1L UGGAAAGAAGAAAGGGGUA NM_005659
    2678 UFD1L CAUGAGGAGUCGACAGAAG NM_005659
    2679 USP1 AAACAAAGGUGAAGAACAA NM_003368
    2680 USP1 GAAGAGAACCAGAGACAAA NM_003368
    2681 USP1 GCAUAGAGAUGGACAGUAU NM_003368
    2682 USP1 AGAAAGGAUUGUAGGAGAA NM_003368
    2683 USP1 AAUUAGAGUUGGUGGAAAU NM_003368
    2684 USP1 AGAAAUACCUCAUCCGAAA NM_003368
    2685 USP1 GAAAGAAGCUCUAAAGGAU NM_003368
    2686 USP1 GAUUGUAGGAGAAGAUAAA NM_003368
    2687 USP1 GGGAAACAUUCAAGAAACA NM_003368
    2688 USP1 GUUUUGAGCUAGUGGAGAA NM_003368
    2689 USP1 UGACCAAAUGUGUGAAAUA NM_003368
    2690 USP1 GGUUAAAGUCUGCAACUAA NM_003368
    2691 USP1 CCAAGGAGUCAAAGGACAA NM_003368
    2692 USP1 GGGAAGUGUGAAAGUGAUA NM_003368
    2693 USP1 GGUUUUAAAUCUGGAGUAA NM_003368
    2694 USP1 GUGCAAAGCUUAAAGGAGU NM_003368
    2695 USP1 CUAGAAGAAUGGAGCACAA NM_003368
    2696 USP1 GAGACAAACUAGAUCAAAA NM_003368
    2697 USP1 GAAUAUAGAGCAUCUGAAA NM_003368
    2698 USP1 UGGAAGAGAACCAGAGACA NM_003368
    2699 USP1 UGUAGAAGCUAUUGGACUU NM_003368
    2700 USP1 CAAAGCAGAUUAUGAGCUA NM_003368
    2701 USP1 CUGAAGACUUUAAAGAGAA NM_003368
    2702 USP1 GGGAAAUUGCAAAGAAGAU NM_003368
    2703 USP1 AGUCAAAGGACAAUCUAAA NM_003368
    2704 USP1 GGAAAUACACAGCCAAGUA NM_003368
    2705 USP1 UGUCAUACCUAGUGAAAGU NM_003368
    2706 USP1 UAGAAAGGAUUGUAGGAGA NM_003368
    2707 USP1 GGAAGAAAGAAGCUCUAAA NM_003368
    2708 USP1 GGGGUGUGGUUGAGAAUUA NM_003368
    2709 USP10 AAGAAGAUGCUGAGGAAUA NM_005153
    2710 USP10 CAAACAAGAGGUUGAGAUA NM_005153
    2711 USP10 GAUAAAAUCGUGAGGGAUA NM_005153
    2712 USP10 ACAAGAAGAUGCUGAGGAA NM_005153
    2713 USP10 GAGAUAAGAAAGUGGAUUU NM_005153
    2714 USP10 CCACAGAAGCCCUGGUCAA NM_005153
    2715 USP10 GAGGAAAUGUUGAACCUAA NM_005153
    2716 USP10 GAGAUAAGUCGAAGAGUGA NM_005153
    2717 USP10 GGACAAAGGGAGCGUAAAA NM_005153
    2718 USP10 GGACAAGAAUAUCAGAGAA NM_005153
    2719 USP10 GACAAGAAUAUCAGAGAAU NM_005153
    2720 USP10 AAGAAGAGCAGGAAGAACA NM_005153
    2721 USP10 AGGAAAUGUUGAACCUAAA NM_005153
    2722 USP10 CCAUAAAGAUUGCAGAGUU NM_005153
    2723 USP10 GAGAAUCUGUCCAAGGUUA NM_005153
    2724 USP10 AUGCAGAAUUUAUGGGUGA NM_005153
    2725 USP10 CAGCCUACCUCCUGUAUUA NM_005153
    2726 USP10 UUGCAGAGUUGCUGGAGAA NM_005153
    2727 USP10 UAAACUACCUGAUGGACAA NM_005153
    2728 USP10 AUGAAGAAGAGCAGGAAGA NM_005153
    2729 USP10 GCAGGAAGAACAAGGUGAA NM_005153
    2730 USP10 GGAAGAACAAGGUGAAGGA NM_005153
    2731 USP10 GGGAACUGGUGCUACAUUA NM_005153
    2732 USP10 UGGAGUUGCUAAUGGACAA NM_005153
    2733 USP10 CAGCAGAGUUCAAAAGAAU NM_005153
    2734 USP10 GGAAAUUAGUAAAGAACUG NM_005153
    2735 USP10 UGAAAAGGGUCGACAAGAA NM_005153
    2736 USP10 CCACAUAUAUUUACAGACU NM_005153
    2737 USP10 GGACUUGGAAAUUAGUAAA NM_005153
    2738 USP10 GUGAAGGAAGCGAGGAUGA NM_005153
    2739 USP11 AGGUAGAGGCUGAGGGCUA NM_004651
    2740 USP11 CGGAAGAGGAUGAGGACUU NM_004651
    2741 USP11 GGGUGAAGAAGAAGGAGUA NM_004651
    2742 USP11 AAGAUGACGAGGAGGAUAA NM_004651
    2743 USP11 GUGGAGAAGCACUGGUAUA NM_004651
    2744 USP11 UCAGAAGGCUCUUUGGAUA NM_004651
    2745 USP11 GCACAGGAGGCAUGGCAAA NM_004651
    2746 USP11 UGGUGGAAGGCGAGGAUUA NM_004651
    2747 USP11 CAGAGAUGAAGAAGCGUUA NM_004651
    2748 USP11 GGGAUGAGAAAGAAGAUGA NM_004651
    2749 USP11 GAACAAGGUUGGCCAUUUU NM_004651
    2750 USP11 GGACGAUGGGGAUGAGAAA NM_004651
    2751 USP11 GAGAUAAACUGGCGCCUCA NM_004651
    2752 USP11 UGUAUAACGUCCUGAUGUA NM_004651
    2753 USP11 AGAAAGAAGAUGACGAGGA NM_004651
    2754 USP11 ACAACAACAUGUCGGAAGA NM_004651
    2755 USP11 AGAAUCAGAUCGAGUCCAA NM_004651
    2756 USP11 GCUGGUUCCUUGUGGAGAA NM_004651
    2757 USP11 GGCACAAUGAUUUGGGCAA NM_004651
    2758 USP11 AUGACGAGGUAGAGGCUGA NM_004651
    2759 USP11 CCGAGUACUUCCUCAACAA NM_004651
    2760 USP11 GCUCUGAGUUCAUGGAUGU NM_004651
    2761 USP11 AUGCAGACCUGGUGAAGCA NM_004651
    2762 USP11 GGCACUACUUUGAUGACAA NM_004651
    2763 USP11 CUGGUAUGGUCUAGAGCAU NM_004651
    2764 USP11 AAGAAGAUGACGAGGAGGA NM_004651
    2765 USP11 GGGCAUGAAGGGUGAGAUC NM_004651
    2766 USP11 GUUACUAUGACGAGGUAGA NM_004651
    2767 USP11 GUUCAUGGAUGUUAAUUGA NM_004651
    2768 USP11 GUGAGAUCGCAGAGGCCUA NM_004651
    2769 USP12 CAGCAAAGAUGAAGAUUUU NM_182488
    2770 USP12 CCUAAGAAGUUCAUCACAA NM_182488
    2771 USP12 GAAGAGAGAAAGCAGGAAA NM_182488
    2772 USP12 CAUCACAAGAUUACGGAAA NM_182488
    2773 USP12 UUGCAAUAGUUAAGAGUCA NM_182488
    2774 USP12 CCACCAAUCCAGACAGAAU NM_182488
    2775 USP12 GCAGCAAAGAUGAAGAUUU NM_182488
    2776 USP12 ACAAGAUUACGGAAAGAAA NM_182488
    2777 USP12 UGCACAAGCUAUUGAAGAA NM_182488
    2778 USP12 GGAUCAACUUCAUCGAUAU NM_182488
    2779 USP12 CACAAACGGAUGAAAGUUA NM_182488
    2780 USP12 GAAAGAGAUUGGUCCAGAA NM_182488
    2781 USP12 ACAAACGGAUGAAAGUUAA NM_182488
    2782 USP12 GCGUAUAAGAGUCAACCUA NM_182488
    2783 USP12 CAGAACAGUUUCCGGUCAA NM_182488
    2784 USP12 CAUCAGAUAUCUCAAAGAA NM_182488
    2785 USP12 GGCAUUAGAGAAAGAGAUU NM_182488
    2786 USP12 GCCCAUGAUUCUAGCUCUA NM_182488
    2787 USP12 GCAAACAGGAAGCACACAA NM_182488
    2788 USP12 GCAACAAGAUGCCCAUGAA NM_182488
    2789 USP12 UGGAUCAACUUCAUCGAUA NM_182488
    2790 USP12 GCAUAGCCACUCAGAAGAA NM_182488
    2791 USP12 GUUUAACACUUCAGGUGAU NM_182488
    2792 USP12 CAAACAGGAAGCACACAAA NM_182488
    2793 USP12 GAUCAACUUCAUCGAUAUA NM_182488
    2794 USP12 GCAGCAAACAGGAAGCACA NM_182488
    2795 USP12 UGAAGAGUGUCGCAGCAAA NM_182488
    2796 USP12 UUGAAGAAUUCUACGGGUU NM_182488
    2797 USP12 CGGCAUUAGAGAAAGAGAU NM_182488
    2798 USP12 UCAAUUACUCACUGCUUAA NM_182488
    2799 USP13 UGAUUGAGAUGGAGAAUAA NM_003940
    2800 USP13 GCACGAAACUGAAGCCAAU NM_003940
    2801 USP13 GGGAACAUGUUGAAAGACA NM_003940
    2802 USP13 GGUCAUUACAUUUGCCAUA NM_003940
    2803 USP13 AGAAUGGGCUCCAGGACAA NM_003940
    2804 USP13 AGACAGUGAUUUUGUGAUU NM_003940
    2805 USP13 UGAUGAAGGAGGAGCACAA NM_003940
    2806 USP13 AUAUGAAGAUGAAGCCAAA NM_003940
    2807 USP13 GAAGGGAAGCAGAAGCAAA NM_003940
    2808 USP13 GCGACAGGGUCUACAAGAA NM_003940
    2809 USP13 UGAAAUAGCACUACCAAAU NM_003940
    2810 USP13 GGACAAUGGAGUCAGGAUU NM_003940
    2811 USP13 ACACGGAGAGGGUGGAUUA NM_003940
    2812 USP13 UGAAGAAGACAGUGAUUUU NM_003940
    2813 USP13 GAGAAUAAUGCCAAUGCAA NM_003940
    2814 USP13 GGAUGGAUCUGGAACAUAU NM_003940
    2815 USP13 ACGAAUAAUAACCUGGAAA NM_003940
    2816 USP13 AGAAUAAUGCCAAUGCAAA NM_003940
    2817 USP13 ACUUGGUAGUGCAGAUAAA NM_003940
    2818 USP13 AGGAUGAACUGAUCGCUUA NM_003940
    2819 USP13 GGGCAGAUGUUUAUUCUUU NM_003940
    2820 USP13 GUUACAGCCAGGAGAGGAA NM_003940
    2821 USP13 GCACUGGAUUGGAUCUUUA NM_003940
    2822 USP13 GGAAGAUGGCUGCAGGAGA NM_003940
    2823 USP13 UGAACUAACGAGAAGGGAA NM_003940
    2824 USP13 GAAGAUGGGUGAUUUACAA NM_003940
    2825 USP13 AGGAGGAAUUCCAAGAUUU NM_003940
    2826 USP13 CAGAGGAAAUCGUAGCUAU NM_003940
    2827 USP13 GCUUAUGAACUAACGAGAA NM_003940
    2828 USP13 GGAAGGAAGAUGGGUGAUU NM_003940
    2829 USP14 GGAGAAAUUUGAAGGUGUA NM_005151
    2830 USP14 GGAUUAAGUUUGAUGAUGA NM_005151
    2831 USP14 UGAAAUAAUGGAAGAGGAA NM_005151
    2832 USP14 GAAACAAGAUGAAUGGAUU NM_005151
    2833 USP14 AGAGGAAAGUGAACAGUAA NM_005151
    2834 USP14 GGUGAAAGGAGGAACGCUA NM_005151
    2835 USP14 GCCAAAUACAAGUGACAAA NM_005151
    2836 USP14 GAGUAUUGCAACAGAAAUU NM_005151
    2837 USP14 ACCAGAACUUCAAGAGAAA NM_005151
    2838 USP14 CCAGAAAGAAGUUAAGUAU NM_005151
    2839 USP14 GAGUUGAAAUAAUGGAAGA NM_005151
    2840 USP14 ACAGAAAGUUAUGGUGAAA NM_005151
    2841 USP14 CAACAGAAAUUGGAAGCAA NM_005151
    2842 USP14 CGAGAAAGGUGAACAAGGA NM_005151
    2843 USP14 AUCCAGAGCUUUAGAGGAA NM_005151
    2844 USP14 AGAAGAACCCUCAGCCAAA NM_005151
    2845 USP14 GAGGAACGCUAAAGGAUGA NM_005151
    2846 USP14 CACCAGAACUUCAAGAGAA NM_005151
    2847 USP14 GACAGAAAGUUAUGGUGAA NM_005151
    2848 USP14 GCUGAAGACAUGUUUAAUA NM_005151
    2849 USP14 GCUAAUGAAUGUUGGAUAC NM_005151
    2850 USP14 GCAAAGAAAUGCCUUGUAU NM_005151
    2851 USP14 CAUGAAAUGUACAGAAUCU NM_005151
    2852 USP14 GAAAUUUGAAGGUGUAGAA NM_005151
    2853 USP14 GCAGGUGCCUUGAGAGCUU NM_005151
    2854 USP14 CCAAAGUUCUUAAGGAUGU NM_005151
    2855 USP14 UCUAUAAUCCAGAGCUUUA NM_005151
    2856 USP14 GCGAGUAUUGCAACAGAAA NM_005151
    2857 USP14 CUUGAGAGCUUCAGGGGAA NM_005151
    2858 USP14 GGAUACAAAUGAUGCGAGU NM_005151
    2859 USP15 GGAACAAAUACAUGAGUAA NM_006313
    2860 USP15 GCAGAUAAGAUGAUAGUUA NM_006313
    2861 USP15 AGGAAUGAGAGGUGAAAUA NM_006313
    2862 USP15 GCAUAAACACCUUGAAUUU NM_006313
    2863 USP15 GAAUUGAAGCUAUGUGAAA NM_006313
    2864 USP15 AUGAAAGUGUGGAGUAUAA NM_006313
    2865 USP15 GAUUUUAACUUGUGCAGUA NM_006313
    2866 USP15 GGAAACAGGUUCUAGAUUU NM_006313
    2867 USP15 ACAAAUACCAGAUGGGAGA NM_006313
    2868 USP15 CAUUUGAACCACUGAAUAA NM_006313
    2869 USP15 GAAACAAUACUGAAGACAA NM_006313
    2870 USP15 UAGCAAAGCUGACACAAUA NM_006313
    2871 USP15 GAUGUGAGUUGAUGACAAA NM_006313
    2872 USP15 CUGCAAAGUAGAAGUAUAU NM_006313
    2873 USP15 GGAGAUAAUGAUUCUGAAA NM_006313
    2874 USP15 AUACAGAGCACGUGAUUAU NM_006313
    2875 USP15 ACACAUUGAUGGAAGGUCA NM_006313
    2876 USP15 ACUGCAAAGUAGAAGUAUA NM_006313
    2877 USP15 CCUGUUUGCCUAAGAGAAA NM_006313
    2878 USP15 GAUGGAAGGUCAAGAGCCA NM_006313
    2879 USP15 GAGACCAGAUUGUGGAACA NM_006313
    2880 USP15 UGUUGUAACUCGAAGAUUU NM_006313
    2881 USP15 AAGCAAAUGUGGUCUGGAA NM_006313
    2882 USP15 ACAAUUGGCUAUAAAGGUA NM_006313
    2883 USP15 UCAGAUAACCGAAUGUAAA NM_006313
    2884 USP15 GAAUAUUCGCUAUGGAUGA NM_006313
    2885 USP15 GCAGAUGGAAGGCCAGAUA NM_006313
    2886 USP15 AGGACAGGUAUUAGUGAUA NM_006313
    2887 USP15 UCAGCAAGCCACAAAGAAA NM_006313
    2888 USP15 CCGAACUGAUCAAGCAAAU NM_006313
    2889 USP16 AGAUAAAGCUGAAGAAGAA NM_006447
    2890 USP16 GGAACAAGGUAAUUUGAAA NM_006447
    2891 USP16 AAACAUGGCUAAAGAGAAU NM_006447
    2892 USP16 AGAUCAAGAUAGUGAGGAA NM_006447
    2893 USP16 CAGAGAAAGAUAAUGGAAA NM_006447
    2894 USP16 AAGAGUGAGUAAAGGAAUA NM_006447
    2895 USP16 CCACAGAGGAAGUAGAUAU NM_006447
    2896 USP16 AGAAUGAACAAGAGAGAGA NM_006447
    2897 USP16 UGAUCAGAGUGGUAAGAAA NM_006447
    2898 USP16 CAUCUUUGGUGGUGAACUA NM_006447
    2899 USP16 AGAAACGGACAAAGGGAAA NM_006447
    2900 USP16 GUUCAAACCAGUUGGGUCA NM_006447
    2901 USP16 GACCAAAGGCAAAUAUAAA NM_006447
    2902 USP16 GGACCAAAGGCAAAUAUAA NM_006447
    2903 USP16 GGAUGAAGAUCAAGAUAGU NM_006447
    2904 USP16 UGAAGAAGAAACAGAAGAA NM_006447
    2905 USP16 GUAAUGGACCAAAGGCAAA NM_006447
    2906 USP16 GUGAAUGUGGAAUGGAAUA NM_006447
    2907 USP16 UAAGAAUGUUGCAGAAGAA NM_006447
    2908 USP16 AGUGAAAGAUAAAGCUGAA NM_006447
    2909 USP16 GAACAAAGGUGUAUGAGGU NM_006447
    2910 USP16 GAACCAACGAAGACAACAA NM_006447
    2911 USP16 GUGAAAGGACUCAGUAAUU NM_006447
    2912 USP16 CAGAGGAAGUAGAUAUGAA NM_006447
    2913 USP16 GGAAGGGAGCAAUGGAGAA NM_006447
    2914 USP16 AAGAUAAAGCUGAAGAAGA NM_006447
    2915 USP16 AGAGAGAAAAGAAGGAAAA NM_006447
    2916 USP16 GAAAGAUAAAGCUGAAGAA NM_006447
    2917 USP16 CGAAUAAACUGCUUUGUGA NM_006447
    2918 USP16 CUGUAAGACUGACAAUAAA NM_006447
    2919 USP18 GAAGAAGACCCGUGGGAAA NM_017414
    2920 USP18 GGACAGACCUGCUGCCUUA NM_017414
    2921 USP18 CUGCAUAUCUUCUGGUUUA NM_017414
    2922 USP18 ACAUGAAGAUGGAGUGCUA NM_017414
    2923 USP18 CCAUAUGAAUCAAGUGUUU NM_017414
    2924 USP18 GCAUGGCGCUUGAGAGAUU NM_017414
    2925 USP18 GGAAUGCUGUGGAUGGAAA NM_017414
    2926 USP18 UCCAUAAGAUAGUGUGAUA NM_017414
    2927 USP18 AGAAGGAAGAAGACAGCAA NM_017414
    2928 USP18 GGAAACAGGUCUUGAAGCU NM_017414
    2929 USP18 GGAAUUCACAGACGAGAAA NM_017414
    2930 USP18 UAAACACGGUCAUGAAUAA NM_017414
    2931 USP18 CCCAAAACCUUCAGAGAUU NM_017414
    2932 USP18 GGAAAAUGGUCUUGCUUCA NM_017414
    2933 USP18 ACAGCAACAUGAAGAGAGA NM_017414
    2934 USP18 GGGAAGACAUCCAGUGUAC NM_017414
    2935 USP18 GACAUUAGCCCUUUAGUUA NM_017414
    2936 USP18 GGAAAAUGGUUCUGCUUCA NM_017414
    2937 USP18 GCUGACGAGCAGAGGAGAA NM_017414
    2938 USP18 AAGAAAAGAAGGAAGAAGA NM_017414
    2939 USP18 CCAGGAUAUUGAAGAGGAU NM_017414
    2940 USP18 UGAUAAACACGGUCAUGAA NM_017414
    2941 USP18 CCAACAUGAUGCUGCCCAA NM_017414
    2942 USP18 GGAGAAGCAUUGUUUUCAA NM_017414
    2943 USP18 CCAGGGAGUUAUCAAGCAA NM_017414
    2944 USP18 AGGAAUUCACAGACGAGAA NM_017414
    2945 USP18 GGGAAGAAGACCCGUGGGA NM_017414
    2946 USP18 GAGAAGCAUUGUUUUCAAA NM_017414
    2947 USP18 UCGUAAUGAAUGUGGACUU NM_017414
    2948 USP18 AGGAUAUUGAAGAGGAUCA NM_017414
    2949 USP2 CGACAGAUGUGGAGAAAAU NM_004205
    2950 USP2 CUGAAUACCUGGUCGACUA NM_004205
    2951 USP2 AGACCCAGAUCCAGAGAUA NM_004205
    2952 USP2 UCGACUACCUGGAGAACUA NM_004205
    2953 USP2 CCAAAGAGGAUGUGCUUGA NM_004205
    2954 USP2 GAAGGGACAUCUUUGGAAA NM_004205
    2955 USP2 GGGAAGAGACGGCAUGAAU NM_004205
    2956 USP2 ACACAAACCUGACAAGAGA NM_004205
    2957 USP2 CGAGAAAGGCCGACAGAUG NM_004205
    2958 USP2 CUGCAGUGCCUGAGCAACA NM_004205
    2959 USP2 GCAUGAGGCUCUUCACCAA NM_171997
    2960 USP2 ACACAGCCCUCGUGGAAGA NM_004205
    2961 USP2 GACCUGGACUUAAGAGAAU NM_004205
    2962 USP2 GGAAGAGACGGCAUGAAUU NM_004205
    2963 USP2 AGGAUGUGCUUGAUGGAGA NM_171997
    2964 USP2 CGGCAUGAAUUCUAAGAGU NM_004205
    2965 USP2 CUGAAGCGCUACACAGAAU NM_004205
    2966 USP2 GAAAAUAUCUAGAACGGGA NM_004205
    2967 USP2 UCGCUGACGUGUACAGAUU NM_004205
    2968 USP2 UGACAUUAAUGGACUGCAU NM_004205
    2969 USP2 ACAAACCUGACAAGAGAAA NM_004205
    2970 USP2 GGCAGAAAACGGUGUAUAA NM_004205
    2971 USP2 UAACUAAAGUGUUCAGACU NM_004205
    2972 USP2 CAUCUGAGUUCAAGACCCA NM_004205
    2973 USP2 GCCGACAGAUGUGGAGAAA NM_004205
    2974 USP2 GGAUGUGCUUGAUGGAGAU NM_004205
    2975 USP2 ACACACAGCCCUCGUGGAA NM_171997
    2976 USP2 AGAAAACGGUGUAUAAAGA NM_171997
    2977 USP2 CGAGGUGAACCGAGUGACA NM_171997
    2978 USP2 UCAACAAAGCCAAGAAUUC NM_171997
    2979 USP20 GGACAAUGAUGCUCACCUA NM_006676
    2980 USP20 CUGAUGAGUUAAAGGGUGA NM_006676
    2981 USP20 GGUCAAAGGAAGCGGCCAU NM_006676
    2982 USP20 CCAUAGGAGAGGUGACCAA NM_006676
    2983 USP20 UCAAGAAAGCCCAGGUAUU NM_006676
    2984 USP20 GGGAGUAGCUGCUGCUUCA NM_006676
    2985 USP20 GUACAAACUCGGAGCAAGU NM_006676
    2986 USP20 GAAAGGAGGACCUGGCCAA NM_006676
    2987 USP20 AGUUAAAGGGUGACAACAU NM_006676
    2988 USP20 AUGAGAAACGGGAGGGUGA NM_006676
    2989 USP20 GAGUGACACGGAUGAGAAA NM_006676
    2990 USP20 GCAUGAAGAACCUCGGGAA NM_006676
    2991 USP20 CGCCAGAGCCGGACAAUGA NM_006676
    2992 USP20 CGUCGUACGUGCUCAAGAA NM_006676
    2993 USP20 GGUGUAAGAAGCUGCGGAA NM_006676
    2994 USP20 GGUACGAGUUUGAUGACCA NM_006676
    2995 USP20 UGCAGAACGCCGAGGGCUA NM_006676
    2996 USP20 GGGCAGAUUUCGGAGGAGA NM_006676
    2997 USP20 GUAAAGAGGCAGAAAAGUU NM_006676
    2998 USP20 CCUGUGAGAAGGAGGUAUU NM_006676
    2999 USP20 GGAACCGGAUGAUGAAACA NM_006676
    3000 USP20 GGACCAAACCUAUGGGCCU NM_006676
    3001 USP20 CCAUUCAUGCACAGGCAAA NM_006676
    3002 USP20 UCUGAAAGCUGUUCCUAUU NM_006676
    3003 USP20 AAGCACAACUUGACCGUGA NM_006676
    3004 USP20 ACGAGACGGUGGUGCAGAA NM_006676
    3005 USP20 CUGCUCAAAUCUAAGGGAA NM_006676
    3006 USP20 GAGCUGAUGCUAAAGAAGA NM_006676
    3007 USP20 CGAGUGACACGGAUGAGAA NM_006676
    3008 USP20 UCUCUGAGGUCUGGCAUAA NM_006676
    3009 USP21 GCUAGAAGAACCUGAGUUA NM_012475
    3010 USP21 GAGCUGUCUUCCAGAAAUA NM_012475
    3011 USP21 GAGCCAACCUAAUGUGGAA NM_016572
    3012 USP21 GUGCCAGGCCUGUGGGUAU NM_012475
    3013 USP21 AAGAAGAGCUAGAGUCGGA NM_016572
    3014 USP21 GUUUCAACCUUUUCACUAA NM_012475
    3015 USP21 GGUUGUAGCUCCAUUAUUU NM_016572
    3016 USP21 CACUAAGGAAGAAGAGCUA NM_012475
    3017 USP21 CUAAGGAAGAAGAGCUAGA NM_016572
    3018 USP21 CGGCAGAAAACUCGAAGUA NM_012475
    3019 USP21 GUACAAAGAUUCCCUCGAA NM_012475
    3020 USP21 UGAUGAACGGCUCAAGAAA NM_012475
    3021 USP21 CCACCCACUUUGAGACGUA NM_012475
    3022 USP21 GCGAGAGGACAGCAAGAUU NM_016572
    3023 USP21 CUGUGAAGCCCUUUAAACA NM_012475
    3024 USP21 GUUAAGUGAUGAUGACCGA NM_016572
    3025 USP21 GAGCGAGAGGACAGCAAGA NM_012475
    3026 USP21 CCUCAGAGACGUCUAUUUU NM_012475
    3027 USP21 GCAGCAAGGAGCGCAGAAA NM_012475
    3028 USP21 GCGCAGAAACCCAGCCUCU NM_012475
    3029 USP21 UAGAAGAACCUGAGUUAAG NM_012475
    3030 USP21 CUUCGAAACCUGGGAAACA NM_012475
    3031 USP21 GAACCUGAGUUAAGUGAUG NM_012475
    3032 USP21 UGGCAUCCAGCGAGGGCUA NM_012475
    3033 USP21 AGUUGACAGUACAAAGAUU NM_016572
    3034 USP21 UUCAGUAGGUGUAGACUUU NM_016572
    3035 USP21 GCAGGAUGCCCAAGAGUUC NM_012475
    3036 USP21 CGGGAUUGUUUCAACCUUU NM_016572
    3037 USP21 CUGAUGAACGGCUCAAGAA NM_016572
    3038 USP21 AUGAACGGCUCAAGAAACU NM_016572
    3039 USP22 CUGCAAAGGUGAUGACAAU XM_042698
    3040 USP22 CCAAGGAGGAGCAGCGAAA XM_042698
    3041 USP22 UGACAAAGACAUGGAAAUA XM_042698
    3042 USP22 GGACUACAUCUAUGACAAA XM_042698
    3043 USP22 CCACAAAGCAGCUCACUAU XM_042698
    3044 USP22 CAGCGAAAAGCUUGGAAAA XM_042698
    3045 USP22 GCAAACCGGUUCAGUGCUA XM_042698
    3046 USP22 AUGACAAAGACAUGGAAAU XM_042698
    3047 USP22 GCGAGGGCAACGUGGUAAA XM_042698
    3048 USP22 AGGACUACAUCUAUGACAA XM_042698
    3049 USP22 CAACAAUGACAACAAGUAU XM_042698
    3050 USP22 ACUGAGAGCCUAUGACAAU XM_042698
    3051 USP22 CAACCAAACGGGAGCUUGA XM_042698
    3052 USP22 CCGAAAAGGAGAAAGAUCA XM_042698
    3053 USP22 GGAGAAAGAUCACCUCGAA XM_042698
    3054 USP22 GUGGACAACUGGAAGCAGA XM_042698
    3055 USP22 AGCAAAGAGAGCAGGAUGA XM_042698
    3056 USP22 GCAAAGAGAGCAGGAUGAA XM_042698
    3057 USP22 GCAGCAGCGCCAAGAUCAA XM_042698
    3058 USP22 GACAAAGACAUGGAAAUAA XM_042698
    3059 USP22 UCACAAAGAAGCAUAUUCA XM_042698
    3060 USP22 CGGACAGUCUCAACAAUGA XM_042698
    3061 USP22 GCAAGGCGUUGGAGAGAAG XM_042698
    3062 USP22 GCAGCUUCAAGGUGGACAA XM_042698
    3063 USP22 CCUCACUGUUUCAGGAGUU XM_042698
    3064 USP22 AAAUAAUCGCCAAGGAGGA XM_042698
    3065 USP22 GCAUCAUAGACCAGAUCUU XM_042698
    3066 USP22 UCAACAAUGACAACAAGUA XM_042698
    3067 USP22 GAAGCAUAUUCACGAGCAU XM_042698
    3068 USP22 GAGCGAGGGCAACGUGGUA XM_042698
    3069 USP24 CCAACUACUUCAUGAAAUA XM_165973
    3070 USP24 CCUCAUGAGUUAAAGAAUA XM_165973
    3071 USP24 CUAUAGAGGCUCACAGUAA XM_165973
    3072 USP24 UGACACAGUUAUAGAAGAA XM_165973
    3073 USP24 GAACAAAUCCUUACAGUGA XM_165973
    3074 USP24 GAAUGUUGCUCCUGGCAUA XM_165973
    3075 USP24 GGACGAGAAUUGAUAAAGA XM_165973
    3076 USP24 GAGAUUACUUGGCUGGAUA XM_165973
    3077 USP24 GAGAACAGCAGGAUGCAUA XM_165973
    3078 USP24 GCCAAAUCCUUCUGUGAAA XM_165973
    3079 USP24 GGGAGUAGCCAGCCAAUUA XM_165973
    3080 USP24 GGAUGAAUACCUCAAGAAA XM_165973
    3081 USP24 AAAGCAAGCUGCAGUACUA XM_165973
    3082 USP24 GAAAAUGCCUGCUCGAAUA XM_165973
    3083 USP24 CUACGAUGCUUGAAGAUGA XM_165973
    3084 USP24 GAAUAGAGAUGUAUACAGU XM_165973
    3085 USP24 AGAAGAAGAUGUCAGAACA XM_165973
    3086 USP24 GGAAAUGCUUGGUUCAUCA XM_165973
    3087 USP24 GGGAAUCUGUGUUCGACAA XM_165973
    3088 USP24 CAACUUGAUUCUUUAGGAA XM_165973
    3089 USP24 UUGAUCAGAUGGAUGAAUA XM_165973
    3090 USP24 UGACAGUAAUGAAACCAUA XM_165973
    3091 USP24 GUUCUAAGCUCCAAACUCA XM_165973
    3092 USP24 CGAAACAGACUCAGCAGUU XM_165973
    3093 USP24 GAAAAGGACCUGUAUUAAA XM_165973
    3094 USP24 AAACAAACCCAUACACUGA XM_165973
    3095 USP24 GAAAAGAGAAUAACAGUGA XM_165973
    3096 USP24 AGGGAAACCUUACCUGUUA XM_165973
    3097 USP24 CCACAGCUUUGUUGAAUGA XM_165973
    3098 USP24 UUACCAAGGAGUUUGAUUA XM_165973
    3099 USP25 GCACAGAAACAGAGAAAUA NM_013396
    3100 USP25 UGGAAUAACUGAUGAGGAA NM_013396
    3101 USP25 GGGAAGAGCUAGUGAGGGA NM_013396
    3102 USP25 GGGAGUACUUGAAGGUAAA NM_013396
    3103 USP25 CUGUAGAAGAUAUGAGAAA NM_013396
    3104 USP25 GGGAAGAGAUCAAGAGACU NM_013396
    3105 USP25 GCUAAAGAAUGUUGGCAAU NM_013396
    3106 USP25 GAAAGUUGCUCAAGCCAAA NM_013396
    3107 USP25 GAUAAAACCUGAAGAAGUA NM_013396
    3108 USP25 AACAAAGGCUAGAAAGAUA NM_013396
    3109 USP25 GGCAAUACUUGUUGGUUUA NM_013396
    3110 USP25 UAUAGGAAAUUCAGGGAAA NM_013396
    3111 USP2S CAUCAGGAUUAUAGGAAAU NM_013396
    3112 USP25 CAUUAUUACUGGUGGAUGA NM_013396
    3113 USP25 CAGAAAGCUUUGCAGGAAA NM_013396
    3114 USP25 ACAGAUACAUGCACAGAAA NM_013396
    3115 USP25 GCAGAUGGAUGAAGUACAA NM_013396
    3116 USP25 GCAUCAGGAUUAUAGGAAA NM_013396
    3117 USP25 GGACAGAAAUAGAAAAUGA NM_013396
    3118 USP25 CCAGCAUAGCAGAGAAUAA NM_013396
    3119 USP25 GGUAAAUCCAGGAGUGUAU NM_013396
    3120 USP25 UGGAGAGGAUCGAGAAGUA NM_013396
    3121 USP25 UGGGAGUACUUGAAGGUAA NM_013396
    3122 USP25 CCACCGGAUUUGAGAGAUU NM_013396
    3123 USP25 CGUGAAAGCAGAUGGAUGA NM_013396
    3124 USP25 GAGCAUUGGUUUACUGAAU NM_013396
    3125 USP25 GGAAUUUGCCUCAAGUAAA NM_013396
    3126 USP25 AGGCAAUUAAGUUGGAAUA NM_013396
    3127 USP25 GAACCAGGCACCAAAGAAA NM_013396
    3128 USP25 GAGCAAUUGCCUUGAGUUU NM_013396
    3129 USP26 CCACAAAGCUGGAGGUAAA NM_031907
    3130 USP26 CCACAAAGUUGAUGAGAAA NM_031907
    3131 USP26 CAGAAGAGCUUGAGUAUAA NM_031907
    3132 USP26 AAGCUAUAAUCGAGAGAAA NM_031907
    3133 USP26 ACAAAGGAAUCCAAAGAUA NM_031907
    3134 USP26 GAACGAUGCUCAUGAGUUU NM_031907
    3135 USP26 GAGUGAGGAUGGAGAAAUU NM_031907
    3136 USP26 GGUUACACAAAGUGGGAUA NM_031907
    3137 USP26 UAUCAGAAUUCCAGAAAGA NM_031907
    3138 USP26 GCACAACACAGAAGGAAAU NM_031907
    3139 USP26 CCUAUGACUUUGAGAAACA NM_031907
    3140 USP26 GCAAAAGUACCUUGGAAAA NM_031907
    3141 USP26 ACACAAAGUGGGAUAAAUU NM_031907
    3142 USP26 CAGCACAAGAAGAGGAAAA NM_031907
    3143 USP26 CCAAUAAGAAUCCAAGAAA NM_031907
    3144 USP26 CCACAGAUGCUGAACAAUU NM_031907
    3145 USP26 GCUAUAAUCGAGAGAAACA NM_031907
    3146 USP26 UUGAAGCAGUGGAAAGAAA NM_031907
    3147 USP26 CCUAAGACCUACAAAAUUA NM_031907
    3148 USP26 CUGAAAGAUAACAUGGAAA NM_031907
    3149 USP26 CAGAGAUGAAUGAGGAAUU NM_031907
    3150 USP26 GAGAAAUCAAGUAGCAAAU NM_031907
    3151 USP26 GAAAUGAUGAUAAGGGAGA NM_031907
    3152 USP26 GAGAGAAACAAUUGAAGUU NM_031907
    3153 USP26 UGGAAAGAAAGAAGAAAGA NM_031907
    3154 USP26 CAACAAAGGAAUCCAAAGA NM_031907
    3155 USP26 CUGUUUAGAUCAACUGAAA NM_031907
    3156 USP26 GAGCAGAAGAGCUUGAGUA NM_031907
    3157 USP26 CCGAAAAUCCAAAACGAAA NM_031907
    3158 USP26 GCAGAAGACAGCAGCAAAA NM_031907
    3159 USP28 GGGAAGAAGUUGAAAGAGA NM_020886
    3160 USP28 CCAUUGAGGUCAUGAGAAA NM_020886
    3161 USP28 GGACAAGAGCGUUGGUUUA NM_020886
    3162 USP28 UGGAAAGGUAUGUGAAAUA NM_020886
    3163 USP28 UCACAGAUGAGGAGAUAAA NM_020886
    3164 USP28 AAGCGAAACUGAAGGAAAU NM_020886
    3165 USP28 GCAAGGAGCUUAUUCGAAA NM_020886
    3166 USP28 GGCAAUACAUGUUGGUUUA NM_020886
    3167 USP28 GGAACAGUUUGCAGAUAAA NM_020886
    3168 USP28 GAAGGUGGCUCAAGCGAAA NM_020886
    3169 USP28 GAAAUAAGAGAGAGUGUAU NM_020886
    3170 USP28 CUGUGGAACUCAAGCAUUA NM_020886
    3171 USP28 GAAAGUUGAAGGAGGAAAU NM_020886
    3172 USP28 GGAAGAAGUUGAAAGAGAU NM_020886
    3173 USP28 GGCAGAAAGCAGUGUAUUA NM_020886
    3174 USP28 AGUUGAAGGAGGAAAUAAA NM_020886
    3175 USP28 GACCAGACAUCCAAGGAAA NM_020886
    3176 USP28 ACAAGGAAGUAUUAGCAAA NM_020886
    3177 USP28 AGAAGUGGCAUGAAGAUUA NM_020886
    3178 USP28 GAGAGAAAUCACAGGCAUU NM_020886
    3179 USP28 UGUCGAAGUCAUACAGAAA NM_020886
    3180 USP28 CAAGCGAAACUGAAGGAAA NM_020886
    3181 USP28 CAGCAAGAUGUGAGUGAAU NM_020886
    3182 USP28 UGGAACAGUUUGCAGAUAA NM_020886
    3183 USP28 GAUUAUAGUUUGUUCCGAA NM_020886
    3184 USP28 UGAAUAUGGAAGAGUACAA NM_020886
    3185 USP28 CUAAACGCUCAAAGAGAAA NM_020886
    3186 USP28 GGAGUGAGAUUGAACAAGA NM_020886
    3187 USP28 AAUUCAAGCUGAUGGAAGA NM_020886
    3188 USP28 GCAACUUAGACGAGUGUUU NM_020886
    3189 USP29 GUACAAAGAUCAAGAGAGA NM_020903
    3190 USP29 GGAAUAUGCUGAAGGAAAU NM_020903
    3191 USP29 AGGAAUAUGCUGAAGGAAA NM_020903
    3192 USP29 AGACAGAUUCCUUGAAAUA NM_020903
    3193 USP29 ACAAAGAUCAAGAGAGAAU NM_020903
    3194 USP29 AUACAAAGCAAUAGGAAGA NM_020903
    3195 USP29 CCAAGGAACUAACAAGAAA NM_020903
    3196 USP29 CAUCAAGUUUAGAGGAUUU NM_020903
    3197 USP29 CAUGAAAGUUUGAAGAACA NM_020903
    3198 USP29 GGAUGAAAUCUCAAUGAAA NM_020903
    3199 USP29 UGAAAGAAGACAUGGAAAA NM_020903
    3200 USP29 GAUGAUAUCUCUAAAGGUA NM_020903
    3201 USP29 CAAUCAGUCUACAGAAUUA NM_020903
    3202 USP29 GGUGAAGAAUAACGAGCAA NM_020903
    3203 USP29 GUGCAAAGACAAAAGGAAA NM_020903
    3204 USP29 UUGAGGAGCUGUUAAGAAA NM_020903
    3205 USP29 GCUAGGGACUGAUUAGAAA NM_020903
    3206 USP29 GCUGAAGGAAAUUGACAAA NM_020903
    3207 USP29 GAAACAGUGCAAAGACAAA NM_020903
    3208 USP29 GGUUGCUGGUGAAGAAUAA NM_020903
    3209 USP29 AAGAAGAAGAGCUUGAAUA NM_020903
    3210 USP29 UUGUAAAGCUUGUGGUCAU NM_020903
    3211 USP29 AGAAGAAGAGCUUGAAUAU NM_020903
    3212 USP29 GAGAGAAUUACUUGGGAAU NM_020903
    3213 USP29 GUAUGAAGAUGGAGGGAAG NM_020903
    3214 USP29 CUUUAAAGAAGAAGAGCUU NM_020903
    3215 USP29 CCAGAGAGACUGUGGAGAU NM_020903
    3216 USP29 CAGAAGAUGCCUUUGUUUA NM_020903
    3217 USP29 CUGAAAGAAGACAUGGAAA NM_020903
    3218 USP29 GGAAAAUGGCUCUGCACUA NM_020903
    3219 USP3 GUAGAAGAGUUUAGAAAGA NM_006537
    3220 USP3 GGAGUUAAGGAAUGGGAAA NM_006537
    3221 USP3 GUGAUGAUUUUGUGGUUAA NM_006537
    3222 USP3 UGGUAGAAGAGUUUAGAAA NM_006537
    3223 USP3 GGACAGCAUAUUUAAGAAA NM_006537
    3224 USP3 AGAAGAACUUGAUGAGACA NM_006537
    3225 USP3 GGGACAGAAUCUAGAAAGU NM_006537
    3226 USP3 CACAAUGACUGCUGAGGAA NM_006537
    3227 USP3 UAACAAAGGGCAUCAUUAA NM_006537
    3228 USP3 CCAAGUCAGUUCAGAAGUA NM_006537
    3229 USP3 CCACAAAAUUUCACAAUGA NM_006537
    3230 USP3 GGCCAAAGCUGGAUCGGAU NM_006537
    3231 USP3 CUAGAAAGUUUGAUCCAUU NM_006537
    3232 USP3 GACAGAAUCUAGAAAGUUU NM_006537
    3233 USP3 GAGAACACUUACAGAACUU NM_006537
    3234 USP3 GCUCUAAGAAUCAAGAAAA NM_006537
    3235 USP3 ACUCAACACUAAACAGCAA NM_006537
    3236 USP3 GGCUAUAUUCGGAGGCAUU NM_006537
    3237 USP3 AGACUGUGGUGAAGGCGAA NM_006537
    3238 USP3 CCAUGAAUUCAAUGCGCUA NM_006537
    3239 USP3 GGGAUAACAAUGUGUCUUU NM_006537
    3240 USP3 AGAAGAGUUUAGAAAGACA NM_006537
    3241 USP3 UUGCAUAAAUGGAGCAUCU NM_006537
    3242 USP3 ACGAGGAGACUGUGGUGAA NM_006537
    3243 USP3 GUACAGAAAGUCAGAGAAC NM_006537
    3244 USP3 UUGAUGAGACAGAGUUAUA NM_006537
    3245 USP3 GCUGAAAGCAAGAUGUGUU NM_006537
    3246 USP3 ACACUAAACAGCAAGUUAU NM_006537
    3247 USP3 CCAAAGCUGGAUCGGAUAA NM_006537
    3248 USP3 CGUAGAAUUUCCACUGAGA NM_006537
    3249 USP30 GUACAAAGAUUGAAGCCAA NM_032663
    3250 USP30 GAAAUAACUCCCAAACAAA NM_032663
    3251 USP30 GGGAGUUAUAGGUGGAAUU NM_032663
    3252 USP30 CAUUGGAAGAUGAGCGAGA NM_032663
    3253 USP30 GGACAUAAACCUAGUCAAC NM_032663
    3254 USP30 ACAGAAAGAAAGAAGCGUA NM_032663
    3255 USP30 CAUCAGAAUCAGUGCGGGA NM_032663
    3256 USP30 GCGCCUACCUGCUGUUCUA NM_032663
    3257 USP30 GAACAAGAACCCAGGGCCU NM_032663
    3258 USP30 UGAUGGACAUUUACAAGUA NM_032663
    3259 USP30 UGUUGGAUGUCUUAAGAAU NM_032663
    3260 USP30 AAGAAAGAAGCGUAGAAAA NM_032663
    3261 USP30 AGAAAGAAGCGUAGAAAAG NM_032663
    3262 USP30 GCCAAGAAGUUACUGAUGA NM_032663
    3263 USP31 GGAUAAAGGAAAAGCAAAA NM_032236
    3264 USP31 GUGCAAGAUUUUAGAAAGA NM_032236
    3265 USP31 CAAAUUUGUUCCAGGAUAA NM_032236
    3266 USP31 GGACAAGGAAGAAGCUAAA NM_032236
    3267 USP31 GAAGAAGGGUUUAAAGGUA NM_032236
    3268 USP31 GAAAAGAAGAGGAGGAAUU NM_032236
    3269 USP31 CAGUAAAGGGCGAUGGAUU NM_032236
    3270 USP31 GCAGGAUGCUCAAGAAUUU NM_032236
    3271 USP31 CCAAAUUUGUUCCAGGAUA NM_032236
    3272 USP31 CCAUAAAGUUGUGGAUAAU NM_032236
    3273 USP31 GCAAACUGGACAUAAGAAA NM_032236
    3274 USP31 GUUUAAAGGUACUGGUCUU NM_032236
    3275 USP31 AAGAAGAGGAGGAAUUAAA NM_032236
    3276 USP31 CUGAAACAGAGGAGGACAA NM_032236
    3277 USP31 GGCGGAAAUUUGUUAGAAA NM_032236
    3278 USP31 AGUAGAACGUUGUCGCAUA NM_032236
    3279 USP31 ACAGCAAGUCUGAGGACAA NM_020718
    3280 USP31 AUGCAAAUACAACGGGAAA NM_020718
    3281 USP31 CUUCAAGACUAUUGUGUCA NM_020718
    3282 USP31 UGUCAGAAACAGAGCAACA NM_020718
    3283 USP31 UCGUAGAUACACAGAGCAA NM_020718
    3284 USP31 CAAAGAAACCAGAGAGCAC NM_020718
    3285 USP31 UUGUAAAUACUGAGGAUGA NM_020718
    3286 USP31 AUUUAGGCCUGAAGGAAUU NM_020718
    3287 USP32 AGGCAAAGAUGAAGAGAAA NM_032582
    3288 USP32 GGAGAUAUCCUGUGGGUUA NM_032582
    3289 USP32 GUAAAUGGUCGGUGGAUAA NM_032582
    3290 USP32 GAUCAAUUGUUGUGGAUUU NM_032582
    3291 USP32 GGACAAAUUAUUAGAGGAU NM_032582
    3292 USP32 GGAAGAAGAAGGACAAAUU NM_032582
    3293 USP32 GAAUACUACAGAAGAGAAA NM_032582
    3294 USP32 GCAUUAAAGAGGAAGAUAU NM_032582
    3295 USP32 CAGCUAAGAUCUCAAGUAA NM_032582
    3296 USP32 GCAUAUAAGUGUCCGAUUU NM_032582
    3297 USP32 GGACAAAUCCCAUUGGUAU NM_032582
    3298 USP32 GUACAUGGUUCCAACAUAA NM_032582
    3299 USP32 GAGAAACGCUAUUGGUUAU NM_032582
    3300 USP32 AGUAAAGGCUACAUCAUUA NM_032582
    3301 USP32 UGGCAAGGGAGGUGAAGAA NM_032582
    3302 USP32 UGGCAACAGUGGAAAGAAU NM_032582
    3303 USP32 ACUAGAAGGAGGACGAUUA NM_032582
    3304 USP32 GAACUAUGUUAUACGGGAA NM_032582
    3305 USP32 UCACAAAUGGAAUGGUUAA NM_032582
    3306 USP32 UAUCAAACAUCCCAGGAAA NM_032582
    3307 USP32 CAGUAUGGAUGAAGACUUU NM_032582
    3308 USP32 GGACAACCAUCUAAGAAGA NM_032582
    3309 USP32 GUGGAAAGAAUAUGUCAAA NM_032582
    3310 USP32 UGAUAUUGUAGAAGGCAUA NM_032582
    3311 USP32 AUGCAGACCAUCAAGGAAA NM_032582
    3312 USP32 ACGAUUUGAUUUAGAGACA NM_032582
    3313 USP33 CAGCAGAGCUUCAGAAUAU NM_015017
    3314 USP33 UUUUGAAAGUCUUGGGAAA NM_015017
    3315 USP33 CAACAGUGAUAGAGCAGAA NM_015017
    3316 USP33 AAAGAGAAGAUGUGCAAUA NM_015017
    3317 USP33 AGACAAUGGAAGAAGACAA NM_015017
    3318 USP33 AGACAUAUUUGAUGGAACA NM_015017
    3319 USP33 GGAAAUACUUGUUACAUGA NM_015017
    3320 USP33 CAGCUCAAAUUGUGACAUA NM_015017
    3321 USP33 UGGAAUAUGUGAAGAGGUU NM_015017
    3322 USP33 AGUCAUGGAAGUAGAAGAA NM_015017
    3323 USP33 AAAGAGAGGAGAAGGAUAU NM_015017
    3324 USP33 UCAGAAAUGUCAAAGGAUU NM_015017
    3325 USP33 AGUAAAUUCUGAAGGCGAA NM_015017
    3326 USP33 CGACAGUGGCUUAAUAAAU NM_015017
    3327 USP33 CUGAAACAGUCGACUUAAA NM_015017
    3328 USP33 GGACUAGCUCGAACAGAUA NM_015017
    3329 USP33 GGGCAUGUCUGGAGAAUAG NM_015017
    3330 USP33 UCAUGGAAGUAGAAGAAGA NM_015017
    3331 USP33 GGAGAAGGAUAUCAAAUUU NM_015017
    3332 USP33 UCAUGUUGAUCCAGAUAUA NM_015017
    3333 USP33 UGGAAGAAGACAAGAGCCA NM_201624
    3334 USP33 AUAUAGAAGCGGAUGAAGA NM_015017
    3335 USP33 GGAAACUGUCAAAGUGCAA NM_201624
    3336 USP33 GAUCAUGUGGCGAAGCAUA NM_015017
    3337 USP33 CAAUGUUAAUUCAGGAUGA NM_015017
    3338 USP34 GGAUAUAGUCCAAGAUGAA NM_014709
    3339 USP34 GCGGAAAGCCAUAGGAAAA NM_014709
    3340 USP34 CUACAAAGCUAGUGCCCUA NM_014709
    3341 USP34 CAGGUUAAGUUUAGAGCAA NM_014709
    3342 USP34 AUAGAAAGUUAGAGAGUCA NM_014709
    3343 USP34 GGGAAGAAGAAGAGGAGGA NM_014709
    3344 USP34 GGGCAUGUUUUAAGAAAUU NM_014709
    3345 USP34 CAAGCUAAUAGGAGGGAAA NM_014709
    3346 USP34 GCAUGGAACCAGAGGAAGA NM_014709
    3347 USP34 GGUUCAAAGUCAAGUGUUU NM_014709
    3348 USP34 GGGAAAGAGUGAGAGGAAA NM_014709
    3349 USP34 GAAGAAAGUACGAGCUGAA NM_014709
    3350 USP34 CGGAAUAGAAAGUUAGAGA NM_014709
    3351 USP34 GAGCAAGAAGCCAAAGAAA NM_014709
    3352 USP34 CCUAAUAUGUUGAUGGCAU NM_014709
    3353 USP34 GCAUAUAAUCCUAGACCUU NM_014709
    3354 USP34 GGAUAUGGGAGGUGAUUGA NM_014709
    3355 USP34 UGGGAAAGAGUGAGAGGAA NM_014709
    3356 USP34 GAGAAGAAGAAUUAGAAGA NM_014709
    3357 USP34 AUUAAAGCACUGUGGAAUA NM_014709
    3358 USP34 CAUAUUUAAUGGAGAGUGA NM_014709
    3359 USP34 GAACCAAACAUGUGCAACA NM_014709
    3360 USP34 AGAUAUGAGAGAAGAAGAA NM_014709
    3361 USP34 CAAAAGACUCAGAGAGCUA NM_014709
    3362 USP34 CAGAAAGGCAUUUGACAUU NM_014709
    3363 USP34 GAGCUUACGAUGUGAAGAA NM_014709
    3364 USP34 CAUCUGAGACAGUGGCAAA NM_014709
    3365 USP34 CCAAGUAUUCAGAGGAUAU NM_014709
    3366 USP34 GGAAAGAGUGAGAGGAAAG NM_014709
    3367 USP34 UCUCAUGGUUAUAGAAAGA NM_014709
    3368 USP35 GGAUAGAGAGGGAGGAAGA XM_290527
    3369 USP35 AAGAAGAGAAGGUGGAGAA XM_290527
    3370 USP35 AGGAAAGGAUAGAGAGGGA XM_290527
    3371 USP35 GUGGAGAAGGAGACAGAAA XM_290527
    3372 USP35 UGGAAGAGGAAGAAGAGAA XM_290527
    3373 USP35 AAGAGAAGGUGGAGAAGGA XM_290527
    3374 USP35 AAGACAAGGAUGAGGAUGA XM_290527
    3375 USP35 GGAAGGAGAGAGGGAGAAA XM_290527
    3376 USP35 AGAAAGAGGAGGAGGUGGA XM_290527
    3377 USP35 GGAAAGCACCAGAGGGGAA XM_290527
    3378 USP35 GUUAAGAAGUUCAGCAUCU XM_290527
    3379 USP35 GGGAGAAAGAGGAGGAGGU XM_290527
    3380 USP35 AAGGAGAGAGGGAGAAAGA XM_290527
    3381 USP35 CCGAGAAGGUGGUGGAGCU XM_290527
    3382 USP35 AAGAGGAGGAGGUGGAAGA XM_290527
    3383 USP35 GCUCGGAGUAUCUGAAGUA XM_290527
    3384 USP35 AGAGCGAGCUGGCGGGUUU XM_290527
    3385 USP35 AGGAAGAAGGGAAGGAGGA XM_290527
    3386 USP35 GGAGGAGGUGGAAGAGGAA XM_290527
    3387 USP35 AGGAGGAGGUGGAAGAGGA XM_290527
    3388 USP35 AGAGGGAGGAAGAAGGGAA XM_290527
    3389 USP35 AGGAAGAAGAGAAGGUGGA XM_290527
    3390 USP35 AGGAGGUGGAAGAGGAAGA XM_290527
    3391 USP35 AGACAAGGAUGAGGAUGAA XM_290527
    3392 USP35 GGAAGAGGAAGAAGAGAAG XM_290527
    3393 USP35 AGAGAACGGAGAAGGAAGA XM_290527
    3394 USP35 GAGAAGGUGGAGAAGGAGA XM_290527
    3395 USP35 AGACAGAAAAGGAGGCUGA XM_290527
    3396 USP35 AGGUGGAGAAGGAGACAGA XM_290527
    3397 USP35 AGGUGGAAGAGGAAGAAGA XM_290527
    3398 USP36 GGGAAGAGGAAGAGGAAGA NM_025090
    3399 USP36 GGAAGAGUCUCCAAGGAAA NM_025090
    3400 USP36 UGAUAAAGCUUACGGGAGA NM_025090
    3401 USP36 CGUAUAUGUCCCAGAAUAA NM_025090
    3402 USP36 GGAAGAGGAAGAAGAAGAA NM_025090
    3403 USP36 CUGGAAAGAAGGUGAAGAA NM_025090
    3404 USP36 AGGAAGAGGAAGAAGAAGA NM_025090
    3405 USP36 AAAGAAAGCAGGAGACACA NM_025090
    3406 USP36 GGACUGAGACCGUGGUUGA NM_025090
    3407 USP36 GCACACCACUGAAGAGAUU NM_025090
    3408 USP36 UGUCCUGAGUGGAGAGAAU NM_025090
    3409 USP36 GUGCUAAAUGCAAGAAGAA NM_025090
    3410 USP36 AAGAGAGAGAAGAGGAGAA NM_025090
    3411 USP36 GGGAAGGAAAAGAAAAUUA NM_025090
    3412 USP36 AGAGAGAAGAGGAGAAACU NM_025090
    3413 USP36 CCAAGAAGAACAUCGGCAA NM_025090
    3414 USP36 GGAAAGGAGCAGAAGGUCU NM_025090
    3415 USP36 GGACAGUGGUACCAGAUGA NM_025090
    3416 USP36 CACCAAGGAUGUAGGCUAU NM_025090
    3417 USP36 GAAAGGAGCAGAAGGUCUU NM_025090
    3418 USP36 AGCAGAAGGUCUUGGUGAA NM_025090
    3419 USP36 UCACCAAGGAUGUAGGCUA NM_025090
    3420 USP36 AGAGAGAGAAGAGGAGAAA NM_025090
    3421 USP36 AGAGGAAGAGGAAGAAGAA NM_025090
    3422 USP36 GAGGAGGAAAGGAGCAGAA NM_025090
    3423 USP36 CGACAAGACUCUGGGACGA NM_025090
    3424 USP36 CUCAAAUACUCAUCUGAUA NM_025090
    3425 USP37 CUACAAUACUGGAGGAAUU NM_020935
    3426 USP37 GAAGAUUACCCUAAGGAAA NM_020935
    3427 USP37 CAAAAGAGCUACCGAGUUA NM_020935
    3428 USP37 GAAAAGAAAUGCUGAGACA NM_020935
    3429 USP37 GCUCAGAAUUGAAUGAAGA NM_020935
    3430 USP37 AAGUAAGGAUGCAGAGGAA NM_020935
    3431 USP37 GCGUGAAAGGGAAGAGCAA NM_020935
    3432 USP37 GUGAAAGGGAAGAGCAAGA NM_020935
    3433 USP37 GUAAGGAUGCAGAGGAAAU NM_020935
    3434 USP37 GGAAAUACCUGCUAUAUGA NM_020935
    3435 USP37 CCGAAGAACUGGAGUAUUC NM_020935
    3436 USP37 GAAAAGAGGAAAAGAAUGA NM_020935
    3437 USP37 GAGAAAAGCAGCUGAGUUU NM_020935
    3438 USP37 CUGAAAGAAGAUAUGGAAA NM_020935
    3439 USP37 GAUUAAGACUGUAGCAGGA NM_020935
    3440 USP37 UCGAAAAGUUCUUGGUAAU NM_020935
    3441 USP37 CAGCUAAGUCAUAACAUUA NM_020935
    3442 USP37 GAAGAAAAUUCACCAGAUA NM_020935
    3443 USP37 ACUCAGGAUUUGACAGAAU NM_020935
    3444 USP37 AGAUCAGGGUUGCUAGAAA NM_020935
    3445 USP37 GAAACAGUCAGAAGAGAAU NM_020935
    3446 USP37 GGGCCGAAGAACUGGAGUA NM_020935
    3447 USP37 UGACAGAAUGAGCGAAGAA NM_020935
    3448 USP37 AGCCAGAGACUUUGUGAAA NM_020935
    3449 USP37 GAGAUAAGUAAGAGAGAUG NM_020935
    3450 USP37 GGGAACAGAAAGAAGAUGA NM_020935
    3451 USP37 CUGAGGAACUGAAAAGAAA NM_020935
    3452 USP37 GGAGGAAUUCCAAGGAUAU NM_020935
    3453 USP37 UGUAGCAGGAAGUGGAAUA NM_020935
    3454 USP37 GAAUAGGACAUCAGGGCUU NM_020935
    3455 USP38 GCAUAGUACUAAUGGUUUA NM_032557
    3456 USP38 CAGAAGAACCAGUAGUUUA NM_032557
    3457 USP38 ACACAAGCCUUCUGAAAUU NM_032557
    3458 USP38 GGAAGUAGCUAGUAAAGCA NM_032557
    3459 USP38 GAAAGAGAGCUGCGGGAAU NM_032557
    3460 USP38 CAGCAAGACUGUUCUGAAU NM_032557
    3461 USP38 ACUAAUGGUUUAAGUGGUA NM_032557
    3462 USP38 UCAGAAACCAGGAGGUGAA NM_032557
    3463 USP38 GGUAAUUGCACUCCUGAAA NM_032557
    3464 USP38 GUAACUUGCUGCAGAACAU NM_032557
    3465 USP38 GAGUUUUACUGUUCUGAAA NM_032557
    3466 USP38 CUGGAUAAAUGGAGACCCA NM_032557
    3467 USP38 GUAUCAUGUGAGAAGGAAA NM_032557
    3468 USP38 UUUAAUGACAGUAGAGUGA NM_032557
    3469 USP38 AGGUAGAGGUGUUACGGAU NM_032557
    3470 USP39 UGAAUAACAUAAAGGCCAA NM_006590
    3471 USP39 GAGAAUACUUGUCUGAAGA NM_006590
    3472 USP39 CCAUGAGGAUCUUCACUAA NM_006590
    3473 USP39 GCAUCACUGAGAAGGAAUA NM_006590
    3474 USP39 ACGAGUACCAGGAGACAAU NM_006590
    3475 USP39 CAAAUGUGGAUCUGAGAGA NM_006590
    3476 USP39 UCACUGAGAAGGAAUAUAA NM_006590
    3477 USP39 AGACUUACAAGGAGAACUU NM_006590
    3478 USP39 CUGAAUAACAUAAAGGCCA NM_006590
    3479 USP39 GGGUAUUGUGGGACUGAAU NM_006590
    3480 USP39 UGGCUAAGUUCAAUGGCAU NM_006590
    3481 USP39 UCUUCAACAUCCUGGCUAA NM_006590
    3482 USP4 CCAAAUGGAUGAAGGUUUA NM_003363
    3483 USP4 AGAACAAACUGAAUGGUAA NM_003363
    3484 USP4 GAACAAACUGAAUGGUAAA NM_003363
    3485 USP4 GGAACAAAUACAUGAGCAA NM_199443
    3486 USP4 AGGAAGAAAUGGAGCAUCA NM_003363
    3487 USP4 GGAAGAAGUAUGUGGGCUU NM_003363
    3488 USP4 GAGAAGAUGAGGAAGAAAU NM_003363
    3489 USP4 CUGCAUAUGCGAAGAACAA NM_003363
    3490 USP4 CCUCAGAAGAAGAAGAAGA NM_003363
    3491 USP4 AACAGAUACUGGAGGGAUA NM_003363
    3492 USP4 GCAAAUGGUGAUAGCACUA NM_003363
    3493 USP4 GGGAAGAUGAGCCAGGAAA NM_003363
    3494 USP4 CCUACGAGCAGUUGAGCAA NM_003363
    3495 USP4 GAACAGCUGUGAAGGAGAA NM_003363
    3496 USP4 GGAAAUUGCAGAAGCCUAU NM_003363
    3497 USP4 GGAAGAUGAGCCAGGAAAU NM_003363
    3498 USP4 AAGGAGAAGAUGAGGAAGA NM_199443
    3499 USP4 GGGAUAAGCUCGACACAGU NM_003363
    3500 USP4 GCAUCAGGAAGAAGGCAAA NM_003363
    3501 USP4 GGGAAAUUGCAGAAGCCUA NM_199443
    3502 USP4 ACGAGAAGCAUGUGAGCAU NM_003363
    3503 USP4 CAGAUGCGGUGGUGGCAAA NM_003363
    3504 USP4 GAGAGGAAGUCCAGGCCAU NM_003363
    3505 USP4 GCAAAGUCGAGGUGUAUUU NM_003363
    3506 USP4 GAAGAAAGAUCGAGUUAUG NM_003363
    3507 USP4 ACUGCAAAGUCGAGGUGUA NM_003363
    3508 USP4 GCAAAGGAAGCCUGGGAGA NM_003363
    3509 USP4 GAACUGAAGCUCUGUGAGA NM_003363
    3510 USP4 UGAAGGAGAAGAUGAGGAA NM_003363
    3511 USP4 CAGAAGAAGAAGAAGACCA NM_199443
    3512 USP40 GGGAAGAAAUUAAAGACUA NM_018218
    3513 USP40 GCUUUUAAUUGAAGGACAA NM_018218
    3514 USP40 CAAAUAACCAAGAGGAAAA NM_018218
    3515 USP40 GGAAAAGGGAAGAAAUUAA NM_018218
    3516 USP40 AAACAAUAUCUGUGAGAGA NM_018218
    3517 USP40 AGGAUAAACCCGAUGCAAA NM_018218
    3518 USP40 CCAAGGAAGACAUGAGGAA NM_018218
    3519 USP40 CAAUCUUAUUAGAGGAGAA NM_018218
    3520 USP40 GCUCAGAAAUGGAAGCUCA NM_018218
    3521 USP40 AGUUAGAAUCUGAAGAGAA NM_018218
    3522 USP40 ACAAAUAACCAAGAGGAAA NM_018218
    3523 USP40 GGACCAGUAAUGAGGAAAU NM_018218
    3524 USP40 CUGAAGAGAAGCAAGUUAA NM_018218
    3525 USP40 CCAGAGUGAAGAAGAGAUU NM_018218
    3526 USP40 AAGGAAGACAUGAGGAAGA NM_018218
    3527 USP40 UGAAAUUGCUGAUGGGGAA NM_018218
    3528 USP40 UGUAAGAACGUUAGCGAGA NM_018218
    3529 USP40 UGGAAAUCGUAGUAGAAGA NM_018218
    3530 USP40 CUAUGAAGCUGGAGAGCCU NM_018218
    3531 USP40 GGGACUGUGUCUUGGAAAA NM_018218
    3532 USP40 AAGUAAACCAGAUGUGAAU NM_018218
    3533 USP40 CCUGGACGGUGGAGAGGAA NM_018218
    3534 USP40 GGAUUAAUCUCAAGCCCUU NM_018218
    3535 USP40 CUGAUAUGUUCUGGAGAUA NM_018218
    3536 USP40 GCAGAAUUGAAGAUGGGAA NM_018218
    3537 USP40 AGUUAAUGCUGAAGAAAUC NM_018218
    3538 USP40 AAUCAUCCCUUUACAGUUA NM_018218
    3539 USP40 GGGAAAAGGAUAUUGAACA NM_018218
    3540 USP40 AUGUUGAUCAUUUGGGAAA NM_018218
    3541 USP40 CGGAGAUACUAUUGGUGUU NM_018218
    3542 USP42 CGGAACAGCUUGAUGGAGA XM_374396
    3543 USP42 GGGCAAGGAGAAUGGGAUU XM_374396
    3544 USP42 CUAGAAGAGCCUAAAGCAA XM_374396
    3545 USP42 UCUGAAACGUUUUGCAAAU XM_376571
    3546 USP42 GGAGCAAGACUGAGGGCCA XM_374396
    3547 USP42 CAGCAAUAAAUUAGACAGA XM_376571
    3548 USP42 CAUAGUAAUUCUUUGGAGA XM_376571
    3549 USP42 CAAGAAAAUCAGAGGACUU XM_376571
    3550 USP42 AGUCAAAGGGGCUGGGCAA XM_374396
    3551 USP42 CUGAAAGGCUCGACGGAUG XM_374396
    3552 USP42 GCUUCAAAGAGGUUCACUA XM_374396
    3553 USP42 CAGUCAUGUUGAAAAGAAA XM_374396
    3554 USP42 CUUGAAUGGCAGCAAUAAA XM_374396
    3555 USP42 GCUCCCAGCCCGUGAUGAA XM_374396
    3556 USP42 CAGUGAUAUUAGAUCGGUA XM_374396
    3557 USP42 CCAAAGACAAACACCGAGA XM_374396
    3558 USP42 GUGCAGACAGCGACAGUGA XM_374396
    3559 USP42 ACAAAAGGAUCAAGCCCUA XM_374396
    3560 USP42 CUCCAGAAUUUGGGCAAUA XM_376571
    3561 USP42 GCAACAAACUGAAAGGCUC XM_376571
    3562 USP42 CCUGGAACGUACAGCUCUA XM_376571
    3563 USP42 GCUUAGCAACAAACUGAAA XM_374396
    3564 USP42 UCAACAAGGCAUUGGAGCA XM_374396
    3565 USP42 CUUCAGGCUUAGCAACAAA XM_374396
    3566 USP42 GGCCAUUACUUCUGCUACA XM_374396
    3567 USP42 AAACACUUACGGAUGGAAA XM_376571
    3568 USP42 CUAAAGCAAAGAAGCACAA XM_376571
    3569 USP42 GGAAAGUCCCGGAAACGGA XM_374396
    3570 USP42 CACUCUUGUUUGUCAGAUA XM_374396
    3571 USP42 CAGUCUACCUCGAACGCAU XM_374396
    3572 USP43 GCCAUGAACUGGAAGGAGA XM_371015
    3573 USP43 GCUUGAAGAACCACGGCAA XM_371015
    3574 USP43 GCUCUCAGUUCCAAGGCAA XM_371015
    3575 USP43 AGACGAGGUUCUUGAGUGU XM_371015
    3576 USP43 GCGCUCAGGGCUUGAAGAA XM_371015
    3577 USP43 GGAAGAUGGUUGCAGAGGA XM_371015
    3578 USP43 CGGAAGAAGGAGAACAGGA XM_371015
    3579 USP43 GAGAUAAUGUGUAUGCCUU XM_371015
    3580 USP43 GUGCAGUGUCUCAGCAACA XM_371015
    3581 USP43 CAGCAAAGACAGUCGCCGA XM_371015
    3582 USP43 AUACCAUCGCAGAGGGAGA XM_371015
    3583 USP43 UGAACUGGAAGGAGAGCUU XM_371015
    3584 USP43 GAGGAUGAGAAGUCAGCAU XM_371015
    3585 USP43 CAGGUGGGCGAGAGAAGAA XM_371015
    3586 USP44 GUAACAGGAUUGAGAAAUU NM_032147
    3587 USP44 AAGCAGAAUUGGAAAGUAU NM_032147
    3588 USP44 GAUUGAGAAAUUUGGGAAA NM_032147
    3589 USP44 GUAUCAAGUUAAAGCAGAA NM_032147
    3590 USP44 GUGUAACAGGAUUGAGAAA NM_032147
    3591 USP44 GGAAAUACUUGCUAUAUGA NM_032147
    3592 USP44 AAACUAAGCAUGUGCACUA NM_032147
    3593 USP44 ACUAAGUGGUGGAGCAUCA NM_032147
    3594 USP44 UCUUAAACAUGGAGCCCUA NM_032147
    3595 USP44 AUGAAUGAAUGUCAGGAAA NM_032147
    3596 USP44 CUGAAGCUUUAGAAGGAAA NM_032147
    3597 USP44 GGUAAAAUCUUUCGAACAU NM_032147
    3598 USP44 GAAUUGGAGUAUCAAGUUA NM_032147
    3599 USP44 CUGAAAUGUUGGCCAAAUU NM_032147
    3600 USP44 GCUCUUUGGCACAGGAGAA NM_032147
    3601 USP44 CCAUGUUGCCUGUGGAAGA NM_032147
    3602 USP44 GGACGUAAUAACCGAGAGA NM_032147
    3603 USP46 CAGAAAAGGAUGAGGGUAA NM_022832
    3604 USP46 CAGCAAAAGAAGAAGGAAA NM_022832
    3605 USP46 UGAUGACAUUGUAGAGAAA NM_022832
    3606 USP46 CCAAAGAAGUUCAUUUCAA NM_022832
    3607 USP46 CUAUCAGUCAAGAGAGUAA NM_022832
    3608 USP46 GAGGAGAAGAAACAGGAAA NM_022832
    3609 USP46 AGCAAAAGAAGAAGGAAAA NM_022832
    3610 USP46 GGUCAAUUUUGGAAACACA NM_022832
    3611 USP46 AGGAGAAGAAACAGGAAAA NM_022832
    3612 USP46 CUAAGAGACUUCAGCAACA NM_022832
    3613 USP46 CAUACAAGGCCCAGCAAAA NM_022832
    3614 USP47 GAACAGAAAUGGAAGCAAA NM_017944
    3615 USP47 GAAAGCAAAUGAAGGGAAA NM_017944
    3616 USP47 CCUGAAAGCUGAAGGAUUU NM_017944
    3617 USP47 GCGCAAUACAUGCAAGAUA NM_017944
    3618 USP47 GCUUAUAAGAUGAUGGAUU NM_017944
    3619 USP47 AGGAAUGACUGUACGGCAA NM_017944
    3620 USP47 CAGCAAAAGUACUGAGACA NM_017944
    3621 USP47 GAAAAGAGACAACGAGAAA NM_017944
    3622 USP47 GGAUAUUAUUCUAGUGCUU NM_017944
    3623 USP47 GCUCCGAGACUUUGGAUUA NM_017944
    3624 USP47 CGCAAUACAUGCAAGAUAA NM_017944
    3625 USP47 UGACAGAUGAGCAAAGAAA NM_017944
    3626 USP47 GUGAAUAAUGACAGGAGUA NM_017944
    3627 USP47 GGAUAACACAAGAGGACAU NM_017944
    3628 USP47 AGAGAGAGUUGGAAGAACA NM_017944
    3629 USP47 GCAUUAUAUAAGUGGGAAU NM_017944
    3630 USP47 GGAUGAUGACUGUGAAAGA NM_017944
    3631 USP47 AAGGCAAGCUGAAGGACUA NM_017944
    3632 USP47 AAAGGAGAAUACAGAGUUA NM_017944
    3633 USP47 AGUAGAAGAACGAAAGCAA NM_017944
    3634 USP47 GUAUAAAGUCAUUCAGUGA NM_017944
    3635 USP49 GGGCAAGAGUGUUCAGCAA NM_004275
    3636 USP49 GAGAAUGGCCCUUGCCUUA NM_004275
    3637 USP49 GGGCAGAGAAGCAAGGAAC NM_004275
    3638 USP49 GUGUGGACUGUGAGACUUA NM_004275
    3639 USP49 UCGAGUUCCUACAGAGUUU NM_004275
    3640 USP49 GGCAAGAGUGUUCAGCAAA NM_004275
    3641 USP5 CCACAGAGAAGGUGAAGUA NM_003481
    3642 USP5 UGGAUAAGCUGGAGAAGAU NM_003481
    3643 USP5 UCAACAUGGUGGAGAGGAA NM_003481
    3644 USP5 AGAGGAAGUAUGUGGAUAA NM_003481
    3645 USP5 UGACUGAGUUGGAGAUAGA NM_003481
    3646 USP5 AAGAGGAGCUUCUGGAGUA NM_003481
    3647 USP5 GAGCAGAGGGGCAGCGAUA NM_003481
    3648 USP5 GGAUGGUCCUGGAAAGUAU NM_003481
    3649 USP5 UGGAGUACGAGGAGAAGAA NM_003481
    3650 USP5 CGAGGAGAAGUUUGAAUUA NM_003481
    3651 USP5 GGAUGCAGCCCUUAACAAA NM_003481
    3652 USP5 AGAGAGAACCUGUGGCUCA NM_003481
    3653 USP5 AGGAGAAGUUUGAAUUAGA NM_003481
    3654 USP5 GAGCUGACGUGUACUCAUA NM_003481
    3655 USP5 CAGAGGAAGUAUGUGGAUA NM_003481
    3656 USP5 GCCUCAAGCAGUUGGACAA NM_003481
    3657 USP5 GUGGAGAGACAUUUCAAUA NM_003481
    3658 USP5 CAUCAAGAAAGAAGGCAGA NM_003481
    3659 USP5 AAAGAAGGCAGAUGGGUGA NM_003481
    3660 USP5 GCGAGGAGAAGUUUGAAUU NM_003481
    3661 USP5 UGGAGAGACAUUUCAAUAA NM_003481
    3662 USP5 AACAGUAUGUGGAGAGACA NM_003481
    3663 USP5 CAAAAUACACGAUGUGAAU NM_003481
    3664 USP5 CAGACAAGACGAUGACUGA NM_003481
    3665 USP5 AAGUGUGACAUGAGAGAGA NM_003481
    3666 USP5 AGUUGGAGAUAGACAUGAA NM_003481
    3667 USP5 AAGCCGAAGAGGAGAAGAU NM_003481
    3668 USP5 GAUCUACAAUGACCAGAAA NM_003481
    3669 USP5 CCAAGUGUGACAUGAGAGA NM_003481
    3670 USP5 GCUUCUUGGUGGAGGAAAG NM_003481
    3671 USP52 GCAGAAAGAUGGACUGGAA NM_014871
    3672 USP52 GGGAAAUCUCCAAGAACAA NM_014871
    3673 USP52 GAGUCAAGUUUGUGGGUCA NM_014871
    3674 USP52 UGGAAAUACCACAGGCUUA NM_014871
    3675 USP52 CCAUUGAGGAGUUGAAGAA NM_014871
    3676 USP52 AGAACAACCUCAAGUAUAU NM_014871
    3677 USP52 UGGAUGAGAAUGAGGAUAU NM_014871
    3678 USP52 GCACGGAAGCAGCGGAAAA NM_014871
    3679 USP52 CCAUGAAGAAGGUGGGCUU NM_014871
    3680 USP52 CCUUCAAGAUGGCAGUAAA NM_014871
    3681 USP52 GCACUGAGCCUGAGUCUUU NM_014871
    3682 USP52 CAUCAAAGUUGGAGAGACC NM_014871
    3683 USP52 GUGUAUGACCUGAUGGCUA NM_014871
    3684 USP52 GGGUCUGGAUGCUGAGUUU NM_014871
    3685 USP52 GGGAAACCCAUGACAGUAU NM_014871
    3686 USP53 CCUAAGAACUGUUGGGUUA XM_052597
    3687 USP53 GCAAUGAGGUUGAAAGAAU XM_052597
    3688 USP53 GGAAGAGAGUGAACAGUAA XM_052597
    3689 USP53 CAGCAUAGUCCAAGACAUA XM_052597
    3690 USP53 CAGGCAAAGCAGAGAGAAA XM_052597
    3691 USP53 GAUUGGAACUAGAUGGAAA XM_052597
    3692 USP53 CGACGAAGCUUGCGGGUUU XM_052597
    3693 USP53 GGAUAUCAGUGGUGUUAAA XM_052597
    3694 USP53 UGGAAAAGGAGCAGAGAAA XM_052597
    3695 USP53 AGCCAAGAUUCUAGGGAUA XM_052597
    3696 USP53 GAUAAUUGGCAGAUGCAAA XM_052597
    3697 USP53 ACAGAUGACUAUAGGAAAU XM_052597
    3698 USP53 CCAUAAUGCAAGAGAACAU XM_052597
    3699 USP53 GUUGAAAGAAUGUUGGAAA XM_052597
    3700 USP53 GUUAAAUGAACCAGGACAA XM_052597
    3701 USP53 UGGAAACCUAUGAGAGAAA XM_052597
    3702 USP53 UUACAGAAUUUGUGCGGUA XM_052597
    3703 USP53 GGAAAGAUGUUGUCUCCAA XM_052597
    3704 USP53 GGGAAGAGAGUGAACAGUA XM_052597
    3705 USP53 ACGGAAACCUGGUGGCAAU XM_052597
    3706 USP53 GAUGAAAUGAAGCAGGAAA XM_052597
    3707 USP53 CGACAAGCAACCUAAAUAA XM_052597
    3708 USP53 GGGACAAAGAAAAGAUUUA XM_052597
    3709 USP53 UGAAGACAAUGGAAAGUUA XM_052597
    3710 USP53 GAUCAAAGGGAAAAGAUAA XM_052597
    3711 USP53 GCAUAGUCCAAGACAUAAA XM_052597
    3712 USP53 UGACAAUGGCACUGGAUAU XM_052597
    3713 USP53 CAACAGAUGACUAUAGGAA XM_052597
    3714 USP53 GGGAAAAGAUAAAAGACAU XM_052597
    3715 USP53 UCACAUUGAUCAAAGGGAA XM_052597
    3716 USP54 ACAAGAAGGAAGAGGCAUU NM_152586
    3717 USP54 AGGAAGAGGCAUUGCUCAA NM_152586
    3718 USP54 CUCACAGGGUCAAGAGAAA NM_152586
    3719 USP54 GGACAGAGGCAGUGAGGAG NM_152586
    3720 USP54 CCACAAGAAGGAAGAGGCA NM_152586
    3721 USP54 AGUACAGUGCAGAGAAUUU NM_152586
    3722 USP54 CAAUAGACUCCCAGGAACU NM_152586
    3723 USP54 GGCCAUGGGCAAAGCAACA NM_152586
    3724 USP54 GGUGUCUCCAUGAGGGAUA NM_152586
    3725 USP54 GUGAGAGAUGUUAGGUCUA NM_152586
    3726 USP54 AUUCAUCAAACGUGAGGAA NM_152586
    3727 USP54 AGAGAGCAGAUCAGGGCUA NM_152586
    3728 USP6 AGGAAAGGGUUGUAGAUAA NM_004505
    3729 USP6 GUAAAUGAUCAGUGGAUAA NM_004505
    3730 USP6 GCUAAGAUCUCAAGUCAAA NM_004505
    3731 USP6 GCGGAGAGGUUCACAACAA NM_004505
    3732 USP6 GGAUGGAAAUGCUGGGAGA NM_004505
    3733 USP6 CGAAGAAACUAACAAGGAA NM_004505
    3734 USP6 GAACAAAUAUGUAGUGAGU NM_004505
    3735 USP6 GUACAUGAUUCCAACAUAA NM_004505
    3736 USP6 CAAGAGAGGUGAAGAAAGU NM_004505
    3737 USP6 GGAGAAUGGGAGACAUAUA NM_004505
    3738 USP6 GCACAGGAGCGGAAGGACA NM_004505
    3739 USP6 GGAGAAAGCAAGAUCAUGA NM_004505
    3740 USP6 GGGCAGUUGUGGAAAGUGA NM_004505
    3741 USP6 GGAUGGACAUGGUAGAGAA NM_004505
    3742 USP6 ACGAGCAAGUGGAUGGAAA NM_004505
    3743 USP6 UGGGAGAAUGGGAGACAUA NM_004505
    3744 USP6 CCAUUAGCCUGUAAACAAA NM_004505
    3745 USP6 GGAAAGGGUUGUAGAUAAG NM_004505
    3746 USP6 GCUAAAUGCUAUGGUGAUU NM_004505
    3747 USP6 GGACCUACCCAAACCAAUA NM_004505
    3748 USP6 GCUCUAAGGGCUAUAAAUU NM_004505
    3749 USP6 CAACAAAGAAGCUGGAUCU NM_004505
    3750 USP6 GUCCAGAUAUGAACAAAUA NM_004505
    3751 USP6 GCACAGUAGCAAACUCAUA NM_004505
    3752 USP6 AGACAGCACUGAUGACCAA NM_004505
    3753 USP6 CCAUGUGGCAUCAGGACAA NM_004505
    3754 USP7 GGAGAAAGCAUCAGGGAAA NM_003470
    3755 USP7 GGACAUAGACAAAGAGAAU NM_003470
    3756 USP7 UGAUAAACCUGUAGGAACA NM_003470
    3757 USP7 GCGAUUACAAGAAGAGAAA NM_003470
    3758 USP7 GAGAAGUGAUGAAGCGAAU NM_003470
    3759 USP7 UGAUAAAGCCCUUGAUGAA NM_003470
    3760 USP7 CGAAUUUAACAGAGAGAAU NM_003470
    3761 USP7 GGUUCAUAGUGGAGAUAAU NM_003470
    3762 USP7 CAGAGAAAGGUGUGAAAUU NM_003470
    3763 USP7 GUGUAAAGAAGUAGACUAU NM_003470
    3764 USP7 CAGAGAGAAUUCAGGACUA NM_003470
    3765 USP7 GGGCAUAUCUACACACCAA NM_003470
    3766 USP7 AGAAGGAGUUUGAGAAGUU NM_003470
    3767 USP7 GAGAACAGGCGAAGUUUUA NM_003470
    3768 USP7 CAGCAAUGUUAGAUAAUGA NM_003470
    3769 USP7 GAAUUACAGCAUAGUGAUA NM_003470
    3770 USP7 GGUAAUCCUCUUAGACAUA NM_003470
    3771 USP7 AAACUUAGGCUGCUAGAAA NM_003470
    3772 USP7 ACAUAAAUGAAGACGAGUA NM_003470
    3773 USP7 ACAUAGACAAAGAGAAUGA NM_003470
    3774 USP7 AGAUAAUCAUGGUGGACAU NM_003470
    3775 USP7 CUACAACUGAUGAGAUUUA NM_003470
    3776 USP7 CUUACAGGAAGCAGAGAAA NM_003470
    3777 USP7 GGAAAUAACACUAUAUCCA NM_003470
    3778 USP7 AGGAGGACAUGGAGGAUGA NM_003470
    3779 USP7 GGGAGAAAGCAUCAGGGAA NM_003470
    3780 USP7 GGAACAUGGCUUACAGGAA NM_003470
    3781 USP7 GUGCAUCUGUUAAGGCAAA NM_003470
    3782 USP7 CCAAUUUAGGGAAGAGGAA NM_003470
    3783 USP7 UGACGUGUCUCUUGAUAAA NM_003470
    3784 USP8 AAGAAGAAAUGGAGAAGAA NM_005154
    3785 USP8 CGAAAGAAAUAAAGCUCAA NM_005154
    3786 USP8 GGACAGGACAGUAUAGAUA NM_005154
    3787 USP8 CCUGAAGAGCAUAGAAUAA NM_005154
    3788 USP8 CUUUAAAGCUGCAGAACAU NM_005154
    3789 USP8 AAAUAAAGCUCAACGAGAA NM_005154
    3790 USP8 ACACAAUGAUGACGGAUAA NM_005154
    3791 USP8 GAAAUGGAGAAGAAAGAAA NM_005154
    3792 USP8 GAUAAUCGGAAGAGAUAUA NM_005154
    3793 USP8 GUACAAACCAUGAGCAACA NM_005154
    3794 USP8 GGAAACAGGAAGAGAGGAU NM_005154
    3795 USP8 AGAAAGAGAAACUGAGGAA NM_005154
    3796 USP8 GAAACAAGAAGCUGAAGAA NM_005154
    3797 USP8 GAAGGAAGAACAAGAACAA NM_005154
    3798 USP8 GCAAAGAGGGGCAAAGAAA NM_005154
    3799 USP8 GGAAAGGGCCUAUGUACUA NM_005154
    3800 USP8 CAAAGAAAUAACAGGAGUA NM_005154
    3801 USP8 CCAAGAAAGAAGAUAAAGA NM_005154
    3802 USP8 CUGAUAAUCGGAAGAGAUA NM_005154
    3803 USP8 AGAAGUUAAACCAGAGAAA NM_005154
    3804 USP8 ACAAGAAGCUGAAGAAAAU NM_005154
    3805 USP8 GCAAUGAGCCUUUGGUUUU NM_005154
    3806 USP8 GUGAACAGGCCAAGAAAGA NM_005154
    3807 USP8 GGCCAAGAAAGAAGAUAAA NM_005154
    3808 USP8 AGGCCAAGAAAGAAGAUAA NM_005154
    3809 USP8 GAGCAAUGGUGAAAAGAAU NM_005154
    3810 USP8 GAAGAGAAGAGGAAGCCAA NM_005154
    3811 USP8 GAUUAAAGGACAACCAGAA NM_005154
    3812 USP8 CAGACGAUACCGAAAGAAA NM_005154
    3813 USP8 GGAAAGGCAGCAAGAGGAA NM_005154
    3814 USP9X UGAGAGAAGUGUACGGAAA NM_021906
    3815 USP9X CCUUAGAGAUGGAGCAAGA NM_004652
    3816 USP9X ACAAAUGACAAAUGGGUUA NM_004652
    3817 USP9X GGGAUGAUGUAUUUGGAUA NM_004652
    3818 USP9X GUGGAGAUGGUGAGAGAAA NM_004652
    3819 USP9X GAGAGAAAUCGCUGGUAUA NM_021906
    3820 USP9X GAGCACAGCAAGAGAGAGA NM_004652
    3821 USP9X GGGUCGUGAUUCAGAGUAA NM_004652
    3822 USP9X AGACACAGCUUCUGAAAUU NM_021906
    3823 USP9X CCAAGAUGCUCCAGAUGAA NM_004652
    3824 USP9X GAACAAGUUAUGUGAAGAA NM_004652
    3825 USP9X GGGAUGAGAAGCAGGACAA NM_004652
    3826 USP9X CCAAAGGAAUGGUGGAGAU NM_004652
    3827 USP9X CAGAAUGGAUACAGCAGAA NM_004652
    3828 USP9X GCAAAGGUGUAUAUAGUAU NM_004652
    3829 USP9X CCUCAAUGCUCUUAAAAUA NM_004652
    3830 USP9X UGACAAAGACAGUGUUAAU NM_004652
    3831 USP9X CUGCAGUAAUUCAGAGGAA NM_004652
    3832 USP9X ACACGAUGCUUUAGAAUUU NM_021906
    3833 USP9X AGGCACAGGUAGUGAUGUA NM_004652
    3834 USP9X CAGAAAACCUUGUAACUUA NM_004652
    3835 USP9X GGAAUGGCUUGGAGAUGAA NM_004652
    3836 USP9X CUAUACAACUAAAGCGAUU NM_021906
    3837 USP9X ACGAAUGGCAGAAUGGAUA NM_004652
    3838 USP9X AGACUUAGAUCCUGAUAUU NM_004652
    3839 USP9X GGACAAGAAGAAACUGUUA NM_004652
    3840 USP9X UAUUAAACCAUUUGGGCAA NM_004652
    3841 USP9X AGAAAUGGUUCCACAGUUU NM_004652
    3842 USP9X GGUGGAUAGUUUAGAUGAA NM_004652
    3843 USP9X GAUGAGGCUUCAAGAUAUA NM_021906
    3844 USP9Y GGUGGUAACUUUUGAAUUA NM_004654
    3845 USP9Y GGAAAGAGAAUGUGCAAUU NM_004654
    3846 USP9Y UUUAAGAAGUGGAGAACUA NM_004654
    3847 USP9Y ACAAAUGACAAGUGGGUAA NM_004654
    3848 USP9Y CAGCAAGAGAGAAGGGUAA NM_004654
    3849 USP9Y GAAAUAACUUCUUGCCAAA NM_004654
    3850 USP9Y AGGCACAGGUAGUGAUUUA NM_004654
    3851 USP9Y UCGAAUGGUUAGAGUAUUA NM_004654
    3852 USP9Y GCAAUAAGCUGGAGGUGGA NM_004654
    3853 USP9Y GAACUUAGCUUCAAGAAUU NM_004654
    3854 USP9Y CCAAAUACAGAUAUGGAAA NM_004654
    3855 USP9Y GCGAAUGGCAGAAUGGAUA NM_004654
    3856 USP9Y GAGAGAGUGUAGUGAUUAA NM_004654
    3857 USP9Y GGAAUGAAAUGCUUUGAAA NM_004654
    3858 USP9Y GGUUAUAUCUAGUGUAUCA NM_004654
    3859 USP9Y GAACAGUAUUCUUGCAAUU NM_004654
    3860 USP9Y GCAGAGAGCUUGGAGAUAA NM_004654
    3861 USP9Y CAGAAGAGGUGGUGGAAUG NM_004654
    3862 USP9Y GUAUUUAAGAAGUGGAGAA NM_004654
    3863 USP9Y GAGAUGAUGUAUUUGGAUA NM_004654
    3864 USP9Y GGCAAAGAAUGAAGCCAAA NM_004654
    3865 USP9Y AAUUAGGGCUAUACAGAAA NM_004654
    3866 USP9Y UAACAGAGCUAUAGAUCUU NM_004654
    3867 USP9Y AAACACAGCUUCUGAAAUU NM_004654
    3868 USP9Y CAGCAUUAAUUGUGCAAGA NM_004654
    3869 USP9Y GGACAAGAUGAGACUAUAA NM_004654
    3870 USP9Y CCAAGUUACUCAUGAUCAA NM_004654
    3871 USP9Y AGGCAAAGAAUGAAGCCAA NM_004654
    3872 VCIP135 CAGAAGGACUGGAGUGAUA NM_025054
    3873 VCIP135 GGGACAGACUUUAGUAAUA NM_025054
    3874 VCIP135 GAUCCAAGAGCUAGGGAAA NM_025054
    3875 VCIP135 GGAAGAGUGGUCAGAGAAA NM_025054
    3876 VCIP135 CAGCAUGGCGACAGAAUUA NM_025054
    3877 VCIP135 CGACAGAAUUACAAUAGAA NM_025054
    3878 VCIP135 GCGAAAGGUCAGAGGAGAU NM_025054
    3879 VCIP135 AGUAAUAGUUCCACCUAAA NM_025054
    3880 VCIP135 AAACAGAAGUUGUGAGUUC NM_025054
    3881 VCIP135 CAAUGAAACUUGUUACCAA NM_025054
    3882 VCIP135 GGAUAAUCGCCUUCACAAA NM_025054
    3883 VCIP135 GGUCAGAGAAACAGUAUAU NM_025054
    3884 VCIP135 AGAGAAGUGCACUGGGAAA NM_025054
    3885 VCIP135 CCUCAUAGAACCAGAGCAU NM_025054
    3886 VCIP135 GACAGAAGUUUGCAAGAUA NM_025054
    3887 VCIP135 GGUGAAUUUGGGAGUGAAA NM_025054
    3888 VCIP135 UGGAGAUGUUCAAGGACAA NM_025054
    3889 VCIP135 CAACAUUCCUCCAUAUUUA NM_025054
    3890 VCIP135 UGGCAUGCCUUAAGAGAGA NM_025054
    3891 VCIP135 GAAAGUAUAGCCAGAGAAU NM_025054
    3892 VCIP135 GAGAAGAAGAUCCGAAUCA NM_025054
    3893 VCIP135 UGGUUGAGGCCCAGCGAAA NM_025054
    3894 VCIP135 CCGACAACUUCUAAGGAGA NM_025054
    3895 VCIP135 GGUCAGAGGAGAUGGGUCU NM_025054
    3896 VCIP135 CAGACUAUGGAAUGAGUAA NM_025054
    3897 VCIP135 UUGAAGAGAUGGAUAGUCA NM_025054
    3898 VCIP135 GAGUAGUAACAAUGAGAGA NM_025054
    3899 VCIP135 GUGGAAGAGUGGUCAGAGA NM_025054
    3900 VCIP135 GGAUGGUGGUUGUGUUAUU NM_025054
    3901 VCIP135 AGGAGUUAAACAUGAGUAA NM_025054
    3902 VDU1 CCUCAGAACAUUUGGGAUA NM_015017
    3903 VDU1 UGAAAGUAGUAGUCAGAAA NM_015017
    3904 VDU1 GAGAAGAUGUGCAAUAAGA NM_015017
    3905 VDU1 GGAUGAAGAAGAUGAACUU NM_015017
    3906 VDU1 GCUAAAGCAAUGUUGUUAA NM_015017
    3907 ZA20D1 GAUCAUGAAUGGAGGAAUA NM_020205
    3908 ZA20D1 GCAGCAAGCUCAAGAAGAA NM_020205
    3909 ZA20D1 UGAAAGUACUUGAGGAUCA NM_020205
    3910 ZA20D1 AGAAGGAGGCAGAGAGGAA NM_020205
    3911 ZA20D1 AGGAAAUGAUCCAGCGCUA NM_020205
    3912 ZA20D1 CGUUUGAACUGGUGGGUGA NM_020205
    3913 ZA20D1 CCAGCAGAGUCGAGGGCAA NM_020205
    3914 ZA20D1 CCAUGGAGCAGAAGGAGAA NM_020205
    3915 ZA20D1 GCUGAAAGUACUUGAGGAU NM_020205
    3916 ZA20D1 GGACAAGAAGAGAGCAGAU NM_020205
    3917 ZA20D1 CAGCAGACACAGCAGAAUA NM_020205
    3918 ZA20D1 ACUUACAGAUUCAGAGUAU NM_020205
    3919 ZA20D1 GGGACAGGGUUGGGAGGAA NM_020205
    3920 ZA20D1 CCAUAGUCGUCGUGGCAGA NM_020205
    3921 ZA20D1 AGGAGAAGUCAAAGCGAGA NM_020205
    3922 ZA20D1 GGGACUUGAUGCUGCGGAA NM_020205
    3923 ZA20D1 GCAGCUUCAUAGAGAGAGA NM_020205
    3924 ZA20D1 GGAGAAUACCAAGGAACAA NM_020205
    3925 ZA20D1 AGGAGAAUACCAAGGAACA NM_020205
    3926 RANGB1 GAACAAAUCCGGAGAGAGA NM_017580
    3927 RANGB1 CCAAAGACCUAGUGGAACA NM_017580
    3928 RANGB1 GAGAGGAAGAUGAGGAUGA NM_017580
    3929 RANGB1 GCCCAAAGACCUAGUGGAA NM_017580
    3930 RANGB1 ACAAAUCCGGAGAGAGAUA NM_017580
    3931 RANGB1 GGAAGUUGCAGUAGUGGUA NM_017580
    3932 RANGB1 GAGAGAAGAACAAUGGCAA NM_017580
    3933 RANGB1 GGGGAGAAACUUUAGGAUA NM_017580
    3934 RANGB1 GUAAUGAGGAACAGCAAGA NM_017580
    3935 RANGB1 GCAGGAUAUGCUAGCAAUA NM_017580
    3936 RANGB1 UGUCCAGACUCUAGUGCAA NM_017580
    3937 RANGB1 CCUAAUAACAUUGAAGCAA NM_017580
    3938 RANGB1 GGACUCAGUGCUUCGGAAA NM_017580
    3939 RANGB1 GGAUGAUGAAGAUGAAUGA NM_017580
    3940 RANGB1 CCGAGGUGCUGGUGCUAAU NM_017580
  • Thus, consistent with Example XVII, the present invention provides an siRNA that targets a nucleotide sequence for a deubiquitination enzymes, wherein the siRNA is selected from the group consisting of SEQ. ID NOs. 438-3940.
  • In another embodiment, an siRNA is provided, said siRNA comprising a sense region and an antisense region, wherein said sense region and said antisense region are at least 90% complementary, said sense region and said antisense region together form a duplex region comprising 18-30 base pairs, and said sense region comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • In another embodiment, an siRNA is provided wherein the siRNA comprises a sense region and an antisense region, wherein said sense region and said antisense region are at least 90% complementary, said sense region and said antisense region together form a duplex region comprising 18-30 base pairs, and said sense region comprises a sequence that is identical to a contiguous stretch of at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • In another embodiment, an siRNA is provided wherein the siRNA comprises a sense region and an antisense region, wherein said sense region and said antisense region are at least 90% complementary, said sense region and said antisense region together form a duplex region comprising 19-30 base pairs, and said sense region comprises a sequence that is identical to a contiguous stretch of at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • In another embodiment, a pool of at least two siRNAs is provided, wherein said pool comprises a first siRNA and a second siRNA, said first siRNA comprises a duplex region of length 18-30 base pairs that has a first sense region that is at least 90% similar to 18 bases of a first sequence selected from the group consisting of: SEQ. ID NOs 438-3940 and said second siRNA comprises a duplex region of length 18-30 base pairs that has a second sense region that is at least 90% similar to 18 bases of a second sequence selected from the group consisting of: SEQ. ID NOs 438-3940 and wherein said first sense region and said second sense region are not identical.
  • In another embodiment, a pool of at least two siRNAs is provided, wherein said pool comprises a first siRNA and a second siRNA, said first siRNA comprises a duplex region of length 18-30 base pairs that has a first sense region that is identical to at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940 and wherein the second siRNA comprises a second sense region that comprises a sequence that is identical to at least 18 bases of a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • In another embodiment, a pool of at least two siRNAs is provided, wherein said pool comprises a first siRNA and a second siRNA, said first siRNA comprises a duplex region of length 19-30 base pairs and has a first sense region comprising a sequence that is at least 90% similar to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940, and said duplex of said second siRNA is 19-30 base pairs and comprises a second sense region that comprises a sequence that is at least 90% similar to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • In another embodiment, a pool of at least two siRNAs is provided, wherein said pool comprises a first siRNA and a second siRNA, said first siRNA comprises a duplex region of length 19-30 base pairs and has a first sense region comprising a sequence that is identical to at least 18 bases of a sequence selected the group consisting of: SEQ. ID NOs 438-3940 and said duplex of said second siRNA is 19-30 base pairs and comprises a second sense region comprising a sequence that is identical to a sequence selected from the group consisting of: SEQ. ID NOs 438-3940.
  • In each of the aforementioned embodiments, preferably the antisense region is at least 90% complementary to a contiguous stretch of bases of one of the NCBI sequences identified in Example XVII; each of the recited NCBI sequences is incorporated by reference as if set forth fully herein. In some embodiments, the antisense region is 100% complementary to a contiguous stretch of bases of one of the NCBI sequences identified in Example XVII.
  • Further, in some embodiments that are directed to siRNA duplexes in which the antisense region is 20-30 bases in length, preferably there is a stretch of 19 bases that is at least 90%, more preferably 100% complementary to the recited sequence id number and the entire antisense region is at least 90% and more preferably 100% complementary to a contiguous stretch of bases of one of the NCBI sequences identified in Example XVII.
  • While the invention has been described in connection with specific embodiments thereof, it will be understood that it is capable of further modifications and this application is intended to cover any variations, uses, or adaptations of the invention following, in general, the principles of the invention and including such departure from the present disclosure as come within known or customary practice within the art to which the invention pertains and as may be applied to the essential features hereinbefore set forth and as follows in the scope of the appended claims.

Claims (9)

1. An siRNA comprising a sense region and an antisense region, wherein said sense region and said antisense region together form a duplex region, said antisense region and said sense region are each 18-30 nucleotides in length and said antisense region comprises a sequence that is at least 90% complementary to a sequence selected from the group consisting of SEQ. ID NOs. 438-3940.
2. An siRNA comprising a sense region and an antisense region, wherein said sense region and said antisense region together form a duplex region and said sense region and said antisense region are each 18-30 nucleotides in length, and said antisense region comprises a sequence that is 100% complementary to a contiguous stretch of at least 18 bases of a sequence selected from the group consisting of SEQ. ID NOs. 438-3940.
3. The siRNA of claim 2, wherein each of said antisense region and said sense region are 19-30 nucleotides in length, and said antisense region comprises a sequence that is 100% complementary to said sequence selected from the group consisting of: SEQ. ID NOs. 438-3940.
4. A pool of at least two siRNAs, wherein said pool comprises a first siRNA and a second siRNA, said first siRNA comprises a first antisense region and a first sense region that together form a first duplex region and each of said first antisense region and said first sense region are 18-30 nucleotides in length and said first antisense region is at least 90% complementary to 18 bases of a first sequence selected from the group consisting of: SEQ. ID NOs. 438-3940 and said second siRNA comprises a second antisense region and a second sense region that together form a second duplex region and each of said second antisense region and said second sense region are 18-30 nucleotides in length and said second antisense region is at least 90% complementary to 18 bases of a second sequence selected from the group consisting of: SEQ. ID NOs. 438-3940, wherein said first antisense region and said second antisense region are not identical.
5. The pool of claim 4, wherein said first antisense region comprises a sequence that is 100% complementary to at least 18 bases of said first sequence, and said second antisense region comprises a sequence that is 100% complementary to at least 18 bases of said second sequence.
6. The pool of claim 4, wherein said first siRNA is 19-30 nucleotides in length and said first antisense region comprises a sequence that is at least 90% complementary to said first sequence, and second siRNA is 19-30 nucleotides in length and said second antisense region comprises a sequence that is at least 90% complementary to said second sequence.
7. The pool of claim 4, wherein said first antisense region is 19-30 nucleotides in length and said first antisense region comprises a sequence that is 100% complementary to at least 18 bases of said first sequence, and said second antisense region is 19-30 nucleotides in length and said second antisense region comprises a sequence that is 100% complementary to said second sequence.
8. The siRNA of claim 1, wherein said antisense region and said sense region are each 19-25 nucleotides in length.
9. The siRNA of claim 4, wherein said first antisense region, said first sense region, said second sense region and said second antisense region are each 19-25 nucleotides in length.
US11/977,558 2002-11-14 2007-10-25 siRNA targeting deubiqutination enzymes Abandoned US20080097089A1 (en)

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US42613702P 2002-11-14 2002-11-14
US50205003P 2003-09-10 2003-09-10
US10/714,333 US8090542B2 (en) 2002-11-14 2003-11-14 Functional and hyperfunctional siRNA
PCT/US2004/014885 WO2006006948A2 (en) 2002-11-14 2004-05-12 METHODS AND COMPOSITIONS FOR SELECTING siRNA OF IMPROVED FUNCTIONALITY
US10/940,892 US20120052487A9 (en) 2002-11-14 2004-09-14 Methods and compositions for selecting sirna of improved functionality
US11/977,558 US20080097089A1 (en) 2002-11-14 2007-10-25 siRNA targeting deubiqutination enzymes

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US10/940,892 Continuation-In-Part US20120052487A9 (en) 2002-11-14 2004-09-14 Methods and compositions for selecting sirna of improved functionality

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Family Applications (118)

Application Number Title Priority Date Filing Date
US11/593,100 Expired - Fee Related US7615541B2 (en) 2002-11-14 2006-11-03 siRNA targeting TIE-2
US11/594,530 Expired - Fee Related US7608706B2 (en) 2002-11-14 2006-11-08 siRNA targeting ras-related nuclear protein
US11/594,666 Abandoned US20070128641A1 (en) 2002-11-14 2006-11-08 siRNA targeting hypoxia-inducible factor 1
US11/598,179 Expired - Fee Related US7541453B2 (en) 2002-11-14 2006-11-09 siRNA targeting aquaporin 4
US11/595,698 Expired - Fee Related US7598369B2 (en) 2002-11-14 2006-11-09 siRNA targeting histamine receptor H1
US11/729,388 Abandoned US20070185317A1 (en) 2002-11-14 2007-03-28 siRNA targeting HtrA serine peptidase 1
US11/729,924 Abandoned US20080015114A1 (en) 2002-11-14 2007-03-29 siRNA targeting connective tissue growth factor (CTGF)
US11/731,894 Expired - Fee Related US7521191B2 (en) 2002-11-14 2007-03-30 siRNA targeting connexin 43
US11/731,875 Abandoned US20070299253A1 (en) 2002-11-14 2007-03-30 siRNA targeting vacuolar ATPase
US11/731,843 Expired - Fee Related US7569684B2 (en) 2002-11-14 2007-03-30 siRNA targeting gremlin
US11/731,890 Abandoned US20080045703A1 (en) 2002-11-14 2007-03-30 siRNA targeting platelet-derived growth factor receptor beta polypeptide (PDGFR)
US11/732,457 Expired - Fee Related US7638621B2 (en) 2002-11-14 2007-04-03 siRNA targeting insulin-like growth factor 1 receptor (IGF-1R)
US11/732,413 Abandoned US20070238868A1 (en) 2002-11-14 2007-04-03 siRNA targeting chemokine (C-X-C motif) receptor 4 (CXCR4)
US11/732,809 Abandoned US20070255046A1 (en) 2002-11-14 2007-04-04 siRNA targeting spectrin SH3 domain binding protein 1 (SSH3BP1)
US11/732,810 Abandoned US20070219362A1 (en) 2002-11-14 2007-04-04 siRNA targeting azurocidin 1 (Cartionic Antimicrobial protein 37)
US11/784,536 Abandoned US20070179286A1 (en) 2002-11-14 2007-04-06 siRNA targeting testis-specific serine kinase 4
US11/784,559 Abandoned US20070213520A1 (en) 2002-11-14 2007-04-06 siRNA targeting calcium/calmodulin dependent protein kinase IV (CAMK4)
US11/784,752 Abandoned US20070213521A1 (en) 2002-11-14 2007-04-09 siRNA targeting cell division cycle 25 (CDC25C)
US11/784,755 Expired - Fee Related US7550572B2 (en) 2002-11-14 2007-04-09 siRNA targeting cell division cycle 20 homolog (CDC20)
US11/784,756 Abandoned US20070232797A1 (en) 2002-11-14 2007-04-09 siRNA targeting cell division cycle 2-like 5(CDC2L5)
US11/807,486 Active 2024-07-25 US7745610B2 (en) 2002-11-14 2007-05-29 siRNA targeting cyclin dependent kinase 11 (CDK11)
US11/807,530 Abandoned US20070255047A1 (en) 2002-11-14 2007-05-29 siRNA targeting cell division cycle 6 homolog (CDC6)
US11/807,577 Abandoned US20070260048A1 (en) 2002-11-14 2007-05-29 siRNA targeting centromere protein E, 312kDa (CENPE)
US11/809,909 Abandoned US20070255048A1 (en) 2002-11-14 2007-06-01 siRNA targeting discoidin domain receptor family, member 1 (DDR1)
US11/810,074 Abandoned US20070276135A1 (en) 2002-11-14 2007-06-04 siRNA targeting dual specificity phosphate 5 (DUSP5)
US11/810,383 Expired - Fee Related US7595388B2 (en) 2002-11-14 2007-06-05 siRNA targeting EPH receptor A3 (EPHA3)
US11/810,384 Abandoned US20070260049A1 (en) 2002-11-14 2007-06-05 siRNA targeting MAD2 mitotic arrest deficient-like (MAD2L2)
US11/810,382 Abandoned US20070260047A1 (en) 2002-11-14 2007-06-05 siRNA targeting EPH receptor A4 (EPHA4)
US11/810,673 Abandoned US20070293664A1 (en) 2002-11-14 2007-06-06 siRNA targeting minichromosome maintenance deficient 5 (MCM5)
US11/810,672 Abandoned US20070255050A1 (en) 2002-11-14 2007-06-06 siRNA targeting minichromosome maintenance deficient 2, mitotin (MCM2)
US11/811,003 Abandoned US20070287833A1 (en) 2002-11-14 2007-06-07 siRNA targeting minichromosome maintenance deficient 6 (MCM6)
US11/811,005 Abandoned US20070265437A1 (en) 2002-11-14 2007-06-07 siRNA targeting testes development-related NYD-SP21 (NYD-SP21)
US11/811,012 Abandoned US20070260050A1 (en) 2002-11-14 2007-06-07 siRNA targeting minichromosome maintenance deficient 7 (MCM7)
US11/811,209 Abandoned US20080085997A1 (en) 2002-11-14 2007-06-08 siRNA targeting phosphoinositide-3-kinase, class 2, beta polypeptide (PIK3C2B)
US11/811,423 Expired - Lifetime US7645870B2 (en) 2002-11-14 2007-06-08 siRNA targeting proteasome 26S subunit, non-ATPase, 10 (Gankyrin or PSMD10)
US11/811,424 Abandoned US20070244312A1 (en) 2002-11-14 2007-06-08 siRNA targeting phosphoinositide-3-kinase, class 2, alpha polypeptide (PIK3C2A)
US11/811,954 Abandoned US20070249819A1 (en) 2002-11-14 2007-06-12 siRNA targeting WEE1 homolog (WEE1)
US11/811,929 Abandoned US20070255051A1 (en) 2002-11-14 2007-06-12 siRNA targeting serine/threonine kinase 22B (STK22B)
US11/811,950 Abandoned US20070260052A1 (en) 2002-11-14 2007-06-12 siRNA targeting RAD1 homolog (RAD1)
US11/811,925 Abandoned US20070260051A1 (en) 2002-11-14 2007-06-12 siRNA targeting pituitary tumor-transforming 1 (PTTG1)
US11/818,547 Abandoned US20070255052A1 (en) 2002-11-14 2007-06-14 siRNA targeting v-myc myelocytomatosis viral oncogene homolog (MYC)
US11/818,938 Expired - Lifetime US7678896B2 (en) 2002-11-14 2007-06-15 siRNA targeting serine/threonine kinase 12 (STK12 or aurora B kinase)
US11/818,936 Expired - Lifetime US7598370B2 (en) 2002-11-14 2007-06-15 siRNA targeting polo-like kinase-1 (PLK-1)
US11/880,624 Abandoned US20080027215A1 (en) 2002-11-14 2007-07-23 siRNA targeting vascular endothelial growth factor (VEGF)
US11/880,965 Expired - Fee Related US7579458B2 (en) 2002-11-14 2007-07-25 siRNA targeting synuclein, alpha (SNCA-1)
US11/881,772 Abandoned US20080027216A1 (en) 2002-11-14 2007-07-27 siRNA targeting sodium channel, voltage-gated, type X, alpha (SCN10A)
US11/881,767 Abandoned US20080039617A1 (en) 2002-11-14 2007-07-27 siRNA targeting neuropeptide Y (NPY)
US11/975,902 Abandoned US20080097091A1 (en) 2002-11-14 2007-10-22 siRNA targeting TNFalpha
US11/977,128 Abandoned US20080097092A1 (en) 2002-11-14 2007-10-23 siRNA targeting kinases
US11/977,347 Abandoned US20080076908A1 (en) 2002-11-14 2007-10-24 siRNA targeting nuclear receptors
US11/977,558 Abandoned US20080097089A1 (en) 2002-11-14 2007-10-25 siRNA targeting deubiqutination enzymes
US11/977,675 Abandoned US20080071073A1 (en) 2002-11-14 2007-10-25 siRNA targeting ubiquitin ligases
US11/978,107 Expired - Fee Related US7605252B2 (en) 2002-11-14 2007-10-26 siRNA targeting kinase insert domain receptor (KDR)
US11/978,106 Expired - Lifetime US7655789B2 (en) 2002-11-14 2007-10-26 siRNA targeting transient receptor potential cation channel, subfamily V, member 1 (TRPV1)
US11/978,097 Expired - Fee Related US7638622B2 (en) 2002-11-14 2007-10-26 SiRNA targeting intercellular adhesion molecule 1 (ICAM1)
US11/978,070 Expired - Fee Related US7582746B2 (en) 2002-11-14 2007-10-26 siRNA targeting complement component 3 (C3)
US11/978,120 Abandoned US20080081904A1 (en) 2002-11-14 2007-10-26 siRNA targeting carbonic anhydrase 4(CA4)
US11/978,125 Abandoned US20080086002A1 (en) 2002-11-14 2007-10-26 siRNA targeting secreted frizzled-related protein 1 (sFRP1)
US11/978,475 Abandoned US20080113372A1 (en) 2002-11-14 2007-10-29 siRNA targeting glucagon receptor (GCGR)
US11/978,398 Expired - Lifetime US7709629B2 (en) 2002-11-14 2007-10-29 siRNA targeting diacylglycerol O-acyltransferase homolog 2 (DGAT2)
US11/978,457 Abandoned US20080113371A1 (en) 2002-11-14 2007-10-29 siRNA targeting beta secretase (BACE)
US11/978,476 Expired - Fee Related US7635771B2 (en) 2002-11-14 2007-10-29 siRNA targeting amyloid beta (A4) precursor protein (APP)
US11/978,487 Abandoned US20080113374A1 (en) 2002-11-14 2007-10-29 siRNA targeting fructose-1,6-bisphosphatase 1 (FBP1)
US11/978,455 Expired - Fee Related US7795421B2 (en) 2002-11-14 2007-10-29 siRNA targeting apolipoprotein B (APOB)
US11/978,518 Expired - Fee Related US7632938B2 (en) 2002-11-14 2007-10-29 siRNA targeting superoxide dismutase 1 (SOD1)
US11/980,263 Expired - Fee Related US7632939B2 (en) 2002-11-14 2007-10-30 siRNA targeting proto-oncogene MET
US11/980,102 Expired - Fee Related US7662950B2 (en) 2002-11-14 2007-10-30 siRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US11/980,300 Expired - Fee Related US7592443B2 (en) 2002-11-14 2007-10-30 siRNA targeting interleukin-1 receptor-associated kinase 4 (IRAK4)
US12/321,749 Expired - Fee Related US7666853B2 (en) 2002-11-14 2009-01-23 siRNA targeting connective tissue growth factor (CTGF)
US12/322,387 Expired - Fee Related US7589191B2 (en) 2002-11-14 2009-02-02 siRNA targeting hypoxia-inducible factor 1
US12/455,098 Expired - Fee Related US7741470B2 (en) 2002-11-14 2009-05-28 siRNA targeting gremlin
US12/460,876 Abandoned US20100004141A1 (en) 2002-11-14 2009-07-24 siRNA targeting polo-like Kinase-1 (PLK-1)
US12/462,029 Expired - Fee Related US7745612B2 (en) 2002-11-14 2009-07-28 siRNA targeting interleukin-1 receptor-associated kinase 4 (IRAK4)
US12/462,420 Expired - Fee Related US7737267B2 (en) 2002-11-14 2009-08-04 siRNA targeting hypoxia-inducible factor 1
US12/462,820 Expired - Fee Related US8022198B2 (en) 2002-11-14 2009-08-10 siRNA targeting histamine receptor H1
US12/584,352 Expired - Fee Related US8222395B2 (en) 2002-11-14 2009-09-03 siRNA targeting kinase insert domain receptor (KDR)
US12/584,850 Expired - Fee Related US7897754B2 (en) 2002-11-14 2009-09-11 SiRNA targeting ras-related nuclear protein RAN
US12/586,167 Expired - Fee Related US7855186B2 (en) 2002-11-14 2009-09-17 siRNA targeting TIE-2
US12/589,879 Expired - Fee Related US8039610B2 (en) 2002-11-14 2009-10-29 siRNA targeting superoxide dismutase 1 (SOD1)
US12/590,097 Expired - Fee Related US7816512B2 (en) 2002-11-14 2009-11-02 siRNA targeting proto-oncogene MET
US12/590,252 Expired - Fee Related US7829696B2 (en) 2002-11-14 2009-11-04 siRNA targeting amyloid beta (A4) precursor protein (APP)
US12/592,872 Active 2024-09-09 US8304528B2 (en) 2002-11-14 2009-12-03 SiRNA targeting fructose-1, 6-bisphosphatase 1 (FBP1)
US12/653,120 Expired - Fee Related US8022199B2 (en) 2002-11-14 2009-12-08 SiRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US12/653,402 Expired - Fee Related US7807820B2 (en) 2002-11-14 2009-12-11 siRNA targeting beta secretase (BACE)
US12/655,107 Expired - Fee Related US7833989B2 (en) 2002-11-14 2009-12-23 siRNA targeting connective tissue growth factor (CTGF)
US12/657,448 Expired - Lifetime US8067576B2 (en) 2002-11-14 2010-01-21 siRNA targeting serine/threonine kinase 12 (STK12 or aurora B kinase)
US12/660,582 Expired - Lifetime US8247169B2 (en) 2002-11-14 2010-03-01 SiRNA targeting diacylglycerol O-acyltransferase homolog 2 (DGAT2)
US12/798,603 Expired - Fee Related US8030476B2 (en) 2002-11-14 2010-04-07 siRNA targeting gremlin
US12/798,802 Expired - Fee Related US7935813B2 (en) 2002-11-14 2010-04-12 siRNA target hypoxia-inducible factor 1
US12/798,906 Active 2024-06-21 US8236942B2 (en) 2002-11-14 2010-04-13 SiRNA targeting glucagon receptor (GCGR)
US12/799,758 Expired - Lifetime US8217162B2 (en) 2002-11-14 2010-04-30 siRNA targeting interleukin-1 receptor-associated kinase 4(IRAK4)
US12/799,975 Abandoned US20100267587A1 (en) 2002-11-14 2010-05-05 siRNA targeting cyclin dependent kinase 11 (CDK11)
US12/804,014 Expired - Fee Related US8071754B2 (en) 2002-11-14 2010-07-12 siRNA targeting apolipoprotein B (APOB)
US12/806,570 Expired - Fee Related US7999097B2 (en) 2002-11-14 2010-08-17 siRNA targeting beta secretase (BACE)
US12/807,526 Expired - Fee Related US8222396B2 (en) 2002-11-14 2010-09-08 SiRNA targeting proto-oncogene MET
US12/924,078 Expired - Lifetime US8268985B2 (en) 2002-11-14 2010-09-20 siRNA targeting amyloid beta (A4) precursor protein (APP)
US12/924,653 Expired - Fee Related US8138329B2 (en) 2002-11-14 2010-10-01 siRNA targeting connective tissue growth factor (CTGF)
US12/927,144 Expired - Fee Related US8314229B2 (en) 2002-11-14 2010-11-08 siRNA targeting tie-2
US13/135,336 Expired - Lifetime US8293887B2 (en) 2002-11-14 2011-07-01 SiRNA targeting beta secretase (BACE)
US13/136,780 Expired - Fee Related US8633306B2 (en) 2002-11-14 2011-08-10 SiRNA targeting histamine receptor H1
US13/136,812 Expired - Fee Related US8426579B2 (en) 2002-11-14 2011-08-11 SiRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US13/199,240 Abandoned US20110319297A1 (en) 2002-11-14 2011-08-23 siRNA targeting gremlin
US13/199,946 Abandoned US20120015850A1 (en) 2002-11-14 2011-09-13 siRNA targeting Superoxide
US13/317,752 Expired - Lifetime US8232386B2 (en) 2002-11-14 2011-10-27 SiRNA targeting apolipoprotein B (APOB)
US13/385,320 Expired - Fee Related US8461326B2 (en) 2002-11-14 2012-02-14 SiRNA targeting connective tissue growth factor (CTGF)
US13/489,725 Abandoned US20120252873A1 (en) 2002-11-14 2012-06-06 siRNA Targeting Interleukin-1 Receptor-associated Kinase 4 (IRAK4)
US13/494,360 Abandoned US20120258888A1 (en) 2002-11-14 2012-06-12 siRNA Targeting Proto-oncogene MET
US13/524,015 Expired - Fee Related US8575329B2 (en) 2002-11-14 2012-06-15 siRNA targeting kinase insert domain receptor (KDR)
US13/536,005 Expired - Fee Related US8445668B2 (en) 2002-11-14 2012-06-28 SiRNA targeting apolipoprotein (APOB)
US13/539,630 Abandoned US20120270751A1 (en) 2002-11-14 2012-07-02 siRNA Targeting Diacylglycerol O-Acyltransferase Homolog 2 (DGAT2)
US13/542,332 Abandoned US20120283311A1 (en) 2002-11-14 2012-07-05 siRNA Targeting Glucagon Receptor (GCCR)
US13/551,794 Expired - Fee Related US8658784B2 (en) 2002-11-14 2012-07-18 siRNA targeting amyloid beta (A4) precursor protein (APP)
US13/613,910 Abandoned US20130023446A1 (en) 2002-11-14 2012-09-13 siRNA Targeting Beta Secretase (BACE)
US13/632,519 Abandoned US20130059760A1 (en) 2002-11-14 2012-10-01 siRNA Targeting Fructose-1, 6-bisphosphatase 1 (FBP1)
US13/647,869 Expired - Fee Related US8658785B1 (en) 2002-11-14 2012-10-09 siRNA targeting tie-2
US13/847,544 Expired - Fee Related US8883998B2 (en) 2002-11-14 2013-03-20 siRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US13/867,175 Abandoned US20130225447A1 (en) 2002-11-14 2013-04-22 siRNA Targeting Apolipoprotein B (APOB)
US14/099,339 Expired - Fee Related US8907077B2 (en) 2002-11-14 2013-12-06 siRNA targeting TIE-2

Family Applications Before (50)

Application Number Title Priority Date Filing Date
US11/593,100 Expired - Fee Related US7615541B2 (en) 2002-11-14 2006-11-03 siRNA targeting TIE-2
US11/594,530 Expired - Fee Related US7608706B2 (en) 2002-11-14 2006-11-08 siRNA targeting ras-related nuclear protein
US11/594,666 Abandoned US20070128641A1 (en) 2002-11-14 2006-11-08 siRNA targeting hypoxia-inducible factor 1
US11/598,179 Expired - Fee Related US7541453B2 (en) 2002-11-14 2006-11-09 siRNA targeting aquaporin 4
US11/595,698 Expired - Fee Related US7598369B2 (en) 2002-11-14 2006-11-09 siRNA targeting histamine receptor H1
US11/729,388 Abandoned US20070185317A1 (en) 2002-11-14 2007-03-28 siRNA targeting HtrA serine peptidase 1
US11/729,924 Abandoned US20080015114A1 (en) 2002-11-14 2007-03-29 siRNA targeting connective tissue growth factor (CTGF)
US11/731,894 Expired - Fee Related US7521191B2 (en) 2002-11-14 2007-03-30 siRNA targeting connexin 43
US11/731,875 Abandoned US20070299253A1 (en) 2002-11-14 2007-03-30 siRNA targeting vacuolar ATPase
US11/731,843 Expired - Fee Related US7569684B2 (en) 2002-11-14 2007-03-30 siRNA targeting gremlin
US11/731,890 Abandoned US20080045703A1 (en) 2002-11-14 2007-03-30 siRNA targeting platelet-derived growth factor receptor beta polypeptide (PDGFR)
US11/732,457 Expired - Fee Related US7638621B2 (en) 2002-11-14 2007-04-03 siRNA targeting insulin-like growth factor 1 receptor (IGF-1R)
US11/732,413 Abandoned US20070238868A1 (en) 2002-11-14 2007-04-03 siRNA targeting chemokine (C-X-C motif) receptor 4 (CXCR4)
US11/732,809 Abandoned US20070255046A1 (en) 2002-11-14 2007-04-04 siRNA targeting spectrin SH3 domain binding protein 1 (SSH3BP1)
US11/732,810 Abandoned US20070219362A1 (en) 2002-11-14 2007-04-04 siRNA targeting azurocidin 1 (Cartionic Antimicrobial protein 37)
US11/784,536 Abandoned US20070179286A1 (en) 2002-11-14 2007-04-06 siRNA targeting testis-specific serine kinase 4
US11/784,559 Abandoned US20070213520A1 (en) 2002-11-14 2007-04-06 siRNA targeting calcium/calmodulin dependent protein kinase IV (CAMK4)
US11/784,752 Abandoned US20070213521A1 (en) 2002-11-14 2007-04-09 siRNA targeting cell division cycle 25 (CDC25C)
US11/784,755 Expired - Fee Related US7550572B2 (en) 2002-11-14 2007-04-09 siRNA targeting cell division cycle 20 homolog (CDC20)
US11/784,756 Abandoned US20070232797A1 (en) 2002-11-14 2007-04-09 siRNA targeting cell division cycle 2-like 5(CDC2L5)
US11/807,486 Active 2024-07-25 US7745610B2 (en) 2002-11-14 2007-05-29 siRNA targeting cyclin dependent kinase 11 (CDK11)
US11/807,530 Abandoned US20070255047A1 (en) 2002-11-14 2007-05-29 siRNA targeting cell division cycle 6 homolog (CDC6)
US11/807,577 Abandoned US20070260048A1 (en) 2002-11-14 2007-05-29 siRNA targeting centromere protein E, 312kDa (CENPE)
US11/809,909 Abandoned US20070255048A1 (en) 2002-11-14 2007-06-01 siRNA targeting discoidin domain receptor family, member 1 (DDR1)
US11/810,074 Abandoned US20070276135A1 (en) 2002-11-14 2007-06-04 siRNA targeting dual specificity phosphate 5 (DUSP5)
US11/810,383 Expired - Fee Related US7595388B2 (en) 2002-11-14 2007-06-05 siRNA targeting EPH receptor A3 (EPHA3)
US11/810,384 Abandoned US20070260049A1 (en) 2002-11-14 2007-06-05 siRNA targeting MAD2 mitotic arrest deficient-like (MAD2L2)
US11/810,382 Abandoned US20070260047A1 (en) 2002-11-14 2007-06-05 siRNA targeting EPH receptor A4 (EPHA4)
US11/810,673 Abandoned US20070293664A1 (en) 2002-11-14 2007-06-06 siRNA targeting minichromosome maintenance deficient 5 (MCM5)
US11/810,672 Abandoned US20070255050A1 (en) 2002-11-14 2007-06-06 siRNA targeting minichromosome maintenance deficient 2, mitotin (MCM2)
US11/811,003 Abandoned US20070287833A1 (en) 2002-11-14 2007-06-07 siRNA targeting minichromosome maintenance deficient 6 (MCM6)
US11/811,005 Abandoned US20070265437A1 (en) 2002-11-14 2007-06-07 siRNA targeting testes development-related NYD-SP21 (NYD-SP21)
US11/811,012 Abandoned US20070260050A1 (en) 2002-11-14 2007-06-07 siRNA targeting minichromosome maintenance deficient 7 (MCM7)
US11/811,209 Abandoned US20080085997A1 (en) 2002-11-14 2007-06-08 siRNA targeting phosphoinositide-3-kinase, class 2, beta polypeptide (PIK3C2B)
US11/811,423 Expired - Lifetime US7645870B2 (en) 2002-11-14 2007-06-08 siRNA targeting proteasome 26S subunit, non-ATPase, 10 (Gankyrin or PSMD10)
US11/811,424 Abandoned US20070244312A1 (en) 2002-11-14 2007-06-08 siRNA targeting phosphoinositide-3-kinase, class 2, alpha polypeptide (PIK3C2A)
US11/811,954 Abandoned US20070249819A1 (en) 2002-11-14 2007-06-12 siRNA targeting WEE1 homolog (WEE1)
US11/811,929 Abandoned US20070255051A1 (en) 2002-11-14 2007-06-12 siRNA targeting serine/threonine kinase 22B (STK22B)
US11/811,950 Abandoned US20070260052A1 (en) 2002-11-14 2007-06-12 siRNA targeting RAD1 homolog (RAD1)
US11/811,925 Abandoned US20070260051A1 (en) 2002-11-14 2007-06-12 siRNA targeting pituitary tumor-transforming 1 (PTTG1)
US11/818,547 Abandoned US20070255052A1 (en) 2002-11-14 2007-06-14 siRNA targeting v-myc myelocytomatosis viral oncogene homolog (MYC)
US11/818,938 Expired - Lifetime US7678896B2 (en) 2002-11-14 2007-06-15 siRNA targeting serine/threonine kinase 12 (STK12 or aurora B kinase)
US11/818,936 Expired - Lifetime US7598370B2 (en) 2002-11-14 2007-06-15 siRNA targeting polo-like kinase-1 (PLK-1)
US11/880,624 Abandoned US20080027215A1 (en) 2002-11-14 2007-07-23 siRNA targeting vascular endothelial growth factor (VEGF)
US11/880,965 Expired - Fee Related US7579458B2 (en) 2002-11-14 2007-07-25 siRNA targeting synuclein, alpha (SNCA-1)
US11/881,772 Abandoned US20080027216A1 (en) 2002-11-14 2007-07-27 siRNA targeting sodium channel, voltage-gated, type X, alpha (SCN10A)
US11/881,767 Abandoned US20080039617A1 (en) 2002-11-14 2007-07-27 siRNA targeting neuropeptide Y (NPY)
US11/975,902 Abandoned US20080097091A1 (en) 2002-11-14 2007-10-22 siRNA targeting TNFalpha
US11/977,128 Abandoned US20080097092A1 (en) 2002-11-14 2007-10-23 siRNA targeting kinases
US11/977,347 Abandoned US20080076908A1 (en) 2002-11-14 2007-10-24 siRNA targeting nuclear receptors

Family Applications After (67)

Application Number Title Priority Date Filing Date
US11/977,675 Abandoned US20080071073A1 (en) 2002-11-14 2007-10-25 siRNA targeting ubiquitin ligases
US11/978,107 Expired - Fee Related US7605252B2 (en) 2002-11-14 2007-10-26 siRNA targeting kinase insert domain receptor (KDR)
US11/978,106 Expired - Lifetime US7655789B2 (en) 2002-11-14 2007-10-26 siRNA targeting transient receptor potential cation channel, subfamily V, member 1 (TRPV1)
US11/978,097 Expired - Fee Related US7638622B2 (en) 2002-11-14 2007-10-26 SiRNA targeting intercellular adhesion molecule 1 (ICAM1)
US11/978,070 Expired - Fee Related US7582746B2 (en) 2002-11-14 2007-10-26 siRNA targeting complement component 3 (C3)
US11/978,120 Abandoned US20080081904A1 (en) 2002-11-14 2007-10-26 siRNA targeting carbonic anhydrase 4(CA4)
US11/978,125 Abandoned US20080086002A1 (en) 2002-11-14 2007-10-26 siRNA targeting secreted frizzled-related protein 1 (sFRP1)
US11/978,475 Abandoned US20080113372A1 (en) 2002-11-14 2007-10-29 siRNA targeting glucagon receptor (GCGR)
US11/978,398 Expired - Lifetime US7709629B2 (en) 2002-11-14 2007-10-29 siRNA targeting diacylglycerol O-acyltransferase homolog 2 (DGAT2)
US11/978,457 Abandoned US20080113371A1 (en) 2002-11-14 2007-10-29 siRNA targeting beta secretase (BACE)
US11/978,476 Expired - Fee Related US7635771B2 (en) 2002-11-14 2007-10-29 siRNA targeting amyloid beta (A4) precursor protein (APP)
US11/978,487 Abandoned US20080113374A1 (en) 2002-11-14 2007-10-29 siRNA targeting fructose-1,6-bisphosphatase 1 (FBP1)
US11/978,455 Expired - Fee Related US7795421B2 (en) 2002-11-14 2007-10-29 siRNA targeting apolipoprotein B (APOB)
US11/978,518 Expired - Fee Related US7632938B2 (en) 2002-11-14 2007-10-29 siRNA targeting superoxide dismutase 1 (SOD1)
US11/980,263 Expired - Fee Related US7632939B2 (en) 2002-11-14 2007-10-30 siRNA targeting proto-oncogene MET
US11/980,102 Expired - Fee Related US7662950B2 (en) 2002-11-14 2007-10-30 siRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US11/980,300 Expired - Fee Related US7592443B2 (en) 2002-11-14 2007-10-30 siRNA targeting interleukin-1 receptor-associated kinase 4 (IRAK4)
US12/321,749 Expired - Fee Related US7666853B2 (en) 2002-11-14 2009-01-23 siRNA targeting connective tissue growth factor (CTGF)
US12/322,387 Expired - Fee Related US7589191B2 (en) 2002-11-14 2009-02-02 siRNA targeting hypoxia-inducible factor 1
US12/455,098 Expired - Fee Related US7741470B2 (en) 2002-11-14 2009-05-28 siRNA targeting gremlin
US12/460,876 Abandoned US20100004141A1 (en) 2002-11-14 2009-07-24 siRNA targeting polo-like Kinase-1 (PLK-1)
US12/462,029 Expired - Fee Related US7745612B2 (en) 2002-11-14 2009-07-28 siRNA targeting interleukin-1 receptor-associated kinase 4 (IRAK4)
US12/462,420 Expired - Fee Related US7737267B2 (en) 2002-11-14 2009-08-04 siRNA targeting hypoxia-inducible factor 1
US12/462,820 Expired - Fee Related US8022198B2 (en) 2002-11-14 2009-08-10 siRNA targeting histamine receptor H1
US12/584,352 Expired - Fee Related US8222395B2 (en) 2002-11-14 2009-09-03 siRNA targeting kinase insert domain receptor (KDR)
US12/584,850 Expired - Fee Related US7897754B2 (en) 2002-11-14 2009-09-11 SiRNA targeting ras-related nuclear protein RAN
US12/586,167 Expired - Fee Related US7855186B2 (en) 2002-11-14 2009-09-17 siRNA targeting TIE-2
US12/589,879 Expired - Fee Related US8039610B2 (en) 2002-11-14 2009-10-29 siRNA targeting superoxide dismutase 1 (SOD1)
US12/590,097 Expired - Fee Related US7816512B2 (en) 2002-11-14 2009-11-02 siRNA targeting proto-oncogene MET
US12/590,252 Expired - Fee Related US7829696B2 (en) 2002-11-14 2009-11-04 siRNA targeting amyloid beta (A4) precursor protein (APP)
US12/592,872 Active 2024-09-09 US8304528B2 (en) 2002-11-14 2009-12-03 SiRNA targeting fructose-1, 6-bisphosphatase 1 (FBP1)
US12/653,120 Expired - Fee Related US8022199B2 (en) 2002-11-14 2009-12-08 SiRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US12/653,402 Expired - Fee Related US7807820B2 (en) 2002-11-14 2009-12-11 siRNA targeting beta secretase (BACE)
US12/655,107 Expired - Fee Related US7833989B2 (en) 2002-11-14 2009-12-23 siRNA targeting connective tissue growth factor (CTGF)
US12/657,448 Expired - Lifetime US8067576B2 (en) 2002-11-14 2010-01-21 siRNA targeting serine/threonine kinase 12 (STK12 or aurora B kinase)
US12/660,582 Expired - Lifetime US8247169B2 (en) 2002-11-14 2010-03-01 SiRNA targeting diacylglycerol O-acyltransferase homolog 2 (DGAT2)
US12/798,603 Expired - Fee Related US8030476B2 (en) 2002-11-14 2010-04-07 siRNA targeting gremlin
US12/798,802 Expired - Fee Related US7935813B2 (en) 2002-11-14 2010-04-12 siRNA target hypoxia-inducible factor 1
US12/798,906 Active 2024-06-21 US8236942B2 (en) 2002-11-14 2010-04-13 SiRNA targeting glucagon receptor (GCGR)
US12/799,758 Expired - Lifetime US8217162B2 (en) 2002-11-14 2010-04-30 siRNA targeting interleukin-1 receptor-associated kinase 4(IRAK4)
US12/799,975 Abandoned US20100267587A1 (en) 2002-11-14 2010-05-05 siRNA targeting cyclin dependent kinase 11 (CDK11)
US12/804,014 Expired - Fee Related US8071754B2 (en) 2002-11-14 2010-07-12 siRNA targeting apolipoprotein B (APOB)
US12/806,570 Expired - Fee Related US7999097B2 (en) 2002-11-14 2010-08-17 siRNA targeting beta secretase (BACE)
US12/807,526 Expired - Fee Related US8222396B2 (en) 2002-11-14 2010-09-08 SiRNA targeting proto-oncogene MET
US12/924,078 Expired - Lifetime US8268985B2 (en) 2002-11-14 2010-09-20 siRNA targeting amyloid beta (A4) precursor protein (APP)
US12/924,653 Expired - Fee Related US8138329B2 (en) 2002-11-14 2010-10-01 siRNA targeting connective tissue growth factor (CTGF)
US12/927,144 Expired - Fee Related US8314229B2 (en) 2002-11-14 2010-11-08 siRNA targeting tie-2
US13/135,336 Expired - Lifetime US8293887B2 (en) 2002-11-14 2011-07-01 SiRNA targeting beta secretase (BACE)
US13/136,780 Expired - Fee Related US8633306B2 (en) 2002-11-14 2011-08-10 SiRNA targeting histamine receptor H1
US13/136,812 Expired - Fee Related US8426579B2 (en) 2002-11-14 2011-08-11 SiRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US13/199,240 Abandoned US20110319297A1 (en) 2002-11-14 2011-08-23 siRNA targeting gremlin
US13/199,946 Abandoned US20120015850A1 (en) 2002-11-14 2011-09-13 siRNA targeting Superoxide
US13/317,752 Expired - Lifetime US8232386B2 (en) 2002-11-14 2011-10-27 SiRNA targeting apolipoprotein B (APOB)
US13/385,320 Expired - Fee Related US8461326B2 (en) 2002-11-14 2012-02-14 SiRNA targeting connective tissue growth factor (CTGF)
US13/489,725 Abandoned US20120252873A1 (en) 2002-11-14 2012-06-06 siRNA Targeting Interleukin-1 Receptor-associated Kinase 4 (IRAK4)
US13/494,360 Abandoned US20120258888A1 (en) 2002-11-14 2012-06-12 siRNA Targeting Proto-oncogene MET
US13/524,015 Expired - Fee Related US8575329B2 (en) 2002-11-14 2012-06-15 siRNA targeting kinase insert domain receptor (KDR)
US13/536,005 Expired - Fee Related US8445668B2 (en) 2002-11-14 2012-06-28 SiRNA targeting apolipoprotein (APOB)
US13/539,630 Abandoned US20120270751A1 (en) 2002-11-14 2012-07-02 siRNA Targeting Diacylglycerol O-Acyltransferase Homolog 2 (DGAT2)
US13/542,332 Abandoned US20120283311A1 (en) 2002-11-14 2012-07-05 siRNA Targeting Glucagon Receptor (GCCR)
US13/551,794 Expired - Fee Related US8658784B2 (en) 2002-11-14 2012-07-18 siRNA targeting amyloid beta (A4) precursor protein (APP)
US13/613,910 Abandoned US20130023446A1 (en) 2002-11-14 2012-09-13 siRNA Targeting Beta Secretase (BACE)
US13/632,519 Abandoned US20130059760A1 (en) 2002-11-14 2012-10-01 siRNA Targeting Fructose-1, 6-bisphosphatase 1 (FBP1)
US13/647,869 Expired - Fee Related US8658785B1 (en) 2002-11-14 2012-10-09 siRNA targeting tie-2
US13/847,544 Expired - Fee Related US8883998B2 (en) 2002-11-14 2013-03-20 siRNA targeting myeloid differentiation primary response gene (88) (MYD88)
US13/867,175 Abandoned US20130225447A1 (en) 2002-11-14 2013-04-22 siRNA Targeting Apolipoprotein B (APOB)
US14/099,339 Expired - Fee Related US8907077B2 (en) 2002-11-14 2013-12-06 siRNA targeting TIE-2

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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2013184524A2 (en) * 2012-06-04 2013-12-12 Academia Sinica Usp37 inactivation as a treatment for plzf/rara-associated acute promyelocytic leukemia
US20150344887A1 (en) * 2014-05-29 2015-12-03 Korea Institute Of Science And Technology siRNA FOR INHIBITION OF USP15 EXPRESSION AND PHARMACEUTICAL COMPOSITION CONTAINING THE SAME
US10125369B2 (en) 2012-12-05 2018-11-13 Alnylam Pharmaceuticals, Inc. PCSK9 iRNA compositions and methods of use thereof
US10851377B2 (en) 2015-08-25 2020-12-01 Alnylam Pharmaceuticals, Inc. Methods and compositions for treating a proprotein convertase subtilisin kexin (PCSK9) gene-associated disorder

Families Citing this family (242)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US2950073A (en) * 1957-02-18 1960-08-23 Lockheed Aircraft Corp Hoist mechanism
ATE508188T1 (en) 2002-02-01 2011-05-15 Life Technologies Corp OLIGONUCLEOTIDE COMPOSITIONS WITH IMPROVED EFFECTIVENESS
US20060009409A1 (en) 2002-02-01 2006-01-12 Woolf Tod M Double-stranded oligonucleotides
WO2003064621A2 (en) 2002-02-01 2003-08-07 Ambion, Inc. HIGH POTENCY siRNAS FOR REDUCING THE EXPRESSION OF TARGET GENES
AU2003276666A1 (en) * 2002-06-12 2003-12-31 Ambion, Inc. Methods and compositions relating to polypeptides with rnase iii domains that mediate rna interference
US20040248094A1 (en) * 2002-06-12 2004-12-09 Ford Lance P. Methods and compositions relating to labeled RNA molecules that reduce gene expression
US20100075423A1 (en) * 2002-06-12 2010-03-25 Life Technologies Corporation Methods and compositions relating to polypeptides with rnase iii domains that mediate rna interference
US9719092B2 (en) 2002-11-14 2017-08-01 Thermo Fisher Scientific Inc. RNAi targeting CNTD2
US8198427B1 (en) 2002-11-14 2012-06-12 Dharmacon, Inc. SiRNA targeting catenin, beta-1 (CTNNB1)
US9879266B2 (en) 2002-11-14 2018-01-30 Thermo Fisher Scientific Inc. Methods and compositions for selecting siRNA of improved functionality
US9839649B2 (en) 2002-11-14 2017-12-12 Thermo Fisher Scientific Inc. Methods and compositions for selecting siRNA of improved functionality
US20100113307A1 (en) * 2002-11-14 2010-05-06 Dharmacon, Inc. siRNA targeting vascular endothelial growth factor (VEGF)
US7691998B2 (en) * 2002-11-14 2010-04-06 Dharmacon, Inc. siRNA targeting nucleoporin 62kDa (Nup62)
WO2006006948A2 (en) * 2002-11-14 2006-01-19 Dharmacon, Inc. METHODS AND COMPOSITIONS FOR SELECTING siRNA OF IMPROVED FUNCTIONALITY
US7781575B2 (en) 2002-11-14 2010-08-24 Dharmacon, Inc. siRNA targeting tumor protein 53 (p53)
US9228186B2 (en) 2002-11-14 2016-01-05 Thermo Fisher Scientific Inc. Methods and compositions for selecting siRNA of improved functionality
US10011836B2 (en) 2002-11-14 2018-07-03 Thermo Fisher Scientific Inc. Methods and compositions for selecting siRNA of improved functionality
US20090227780A1 (en) * 2002-11-14 2009-09-10 Dharmacon, Inc. siRNA targeting connexin 43
US9771586B2 (en) 2002-11-14 2017-09-26 Thermo Fisher Scientific Inc. RNAi targeting ZNF205
PT2284266E (en) * 2002-11-14 2013-12-17 Thermo Fisher Scient Biosciences Inc Sirna targeting tp53
US7592442B2 (en) * 2002-11-14 2009-09-22 Dharmacon, Inc. siRNA targeting ribonucleotide reductase M2 polypeptide (RRM2 or RNR-R2)
US9719094B2 (en) 2002-11-14 2017-08-01 Thermo Fisher Scientific Inc. RNAi targeting SEC61G
US7612196B2 (en) 2002-11-14 2009-11-03 Dharmacon, Inc. siRNA targeting cyclin-dependent kinase inhibitor 1B (p27, Kip1) (CDKN1B)
US7951935B2 (en) 2002-11-14 2011-05-31 Dharmacon, Inc. siRNA targeting v-myc myelocytomatosis viral oncogene homolog (MYC)
US7829694B2 (en) * 2002-11-26 2010-11-09 Medtronic, Inc. Treatment of neurodegenerative disease through intracranial delivery of siRNA
US7618948B2 (en) * 2002-11-26 2009-11-17 Medtronic, Inc. Devices, systems and methods for improving and/or cognitive function through brain delivery of siRNA
US7595306B2 (en) * 2003-06-09 2009-09-29 Alnylam Pharmaceuticals Inc Method of treating neurodegenerative disease
WO2005035561A1 (en) * 2003-10-14 2005-04-21 University College Cork - National University Of Ireland, Cork Igf-i responsive gene and use thereof
US8088902B2 (en) * 2004-04-05 2012-01-03 The Rockefeller University DNA virus microRNA and methods for inhibiting same
AU2005276245C1 (en) 2004-08-23 2015-02-26 Sylentis S.A.U. Treatment of eye disorders characterized by an elevated introacular pressure by siRNAs
MX2007003795A (en) * 2004-09-28 2007-07-11 Quark Biotech Inc Oligoribonucleotides and methods of use thereof for treatment of alopecia, acute renal failure and other diseases.
US20060142228A1 (en) 2004-12-23 2006-06-29 Ambion, Inc. Methods and compositions concerning siRNA's as mediators of RNA interference
DE102005003788A1 (en) * 2005-01-19 2006-07-20 Eberhard-Karls-Universität Tübingen Universitätsklinikum siRNA molecules for the treatment of blood vessels
JP5964538B2 (en) * 2005-02-03 2016-08-03 コーダ セラピューティクス， インコーポレイテッド Anti-connexin compounds and their use
JP4131271B2 (en) * 2005-03-30 2008-08-13 ソニー株式会社 Information processing apparatus and method, and program
JP5368097B2 (en) * 2005-09-20 2013-12-18 ジェイ・アール・シンプロット・カンパニー Low acrylamide food
US8258109B2 (en) * 2005-10-20 2012-09-04 Isis Pharmaceuticals, Inc. Compositions and methods for modulation of LMNA expression
GB0521351D0 (en) * 2005-10-20 2005-11-30 Genomica Sau Modulation of TRPV expression levels
US20070135518A1 (en) * 2005-12-09 2007-06-14 Marta Weinstock-Rosin Use of low-dose ladostigil for neuroprotection
AU2006337093B2 (en) * 2005-12-22 2013-03-14 Opko Pharmaceuticals, Llc. Compositions and methods for regulating complement system
EP2674493B1 (en) 2006-04-13 2017-12-27 Arrowhead Research Corporation RNAi-mediated inhibition of histamine receptor H1-related conditions
TW201336514A (en) 2006-04-13 2013-09-16 Alcon Res Ltd RNAi-mediated inhibition of spleen tyrosine kinase-related inflammatory conditions
WO2007147067A2 (en) 2006-06-14 2007-12-21 Rosetta Inpharmatics Llc Methods and compositions for regulating cell cycle progression
MX2009001896A (en) * 2006-08-24 2009-04-17 Alcon Res Ltd Rnai-mediated inhibition of gremlin for treatment of iop-related conditions.
WO2008067382A2 (en) * 2006-11-28 2008-06-05 Alcon Research, Ltd. Rnai-mediated inhibition of aquaporin 4 for treatment of iop-related conditions
KR100981289B1 (en) * 2006-12-18 2010-09-10 주식회사 파나진 Peptide nucleic acid oligomers comprising universal bases, preparation methods thereof, and kits, devices and methods for the analysis, detection or modulation of nucleic acids using the same
EP2126081A2 (en) * 2007-03-02 2009-12-02 MDRNA, Inc. Nucleic acid compounds for inhibiting hif1a gene expression and uses thereof
RS53165B (en) 2007-03-14 2014-06-30 Bionsil S.R.L. Btk inhibitors for use in treating chemotherapeutic drug-resistant epithelial tumours
US8843675B2 (en) * 2007-03-15 2014-09-23 Broadcom Corporation Pipelined buffer interconnect
US8906632B2 (en) * 2007-03-23 2014-12-09 Korea University Research & Business Foundation Use of inhibitors of leukotriene B4 receptor BLT2 for treating asthma
US8785408B2 (en) 2007-06-27 2014-07-22 Quark Pharmaceuticals, Inc. Compositions and methods for reducing or protecting against delayed graft function (DGF)
TW200911290A (en) * 2007-07-02 2009-03-16 Alcon Res Ltd RNAI-mediated inhibition of HTRA1 for treatment of macular degeneration
EA201000085A1 (en) 2007-07-05 2010-10-29 Новартис Аг dsPHK INTRODUCED FOR THE TREATMENT OF VIRAL INFECTION
US7910722B2 (en) 2007-07-05 2011-03-22 Florida State University Research Foundation RNAi therapeutic for treatment of hepatitis C infection
WO2009008990A2 (en) * 2007-07-06 2009-01-15 Intradigm Corporation Methods and compositions for treatment of cancer and other angiogenesis - related diseases
US20110160279A1 (en) * 2007-08-13 2011-06-30 Board Of Trustees Of Southern Illinois University METHODS FOR TREATMENT AND PREVENTION OF OTOTOXICITY BY siRNA
WO2009079399A2 (en) * 2007-12-14 2009-06-25 Alnylam Pharmaceuticals, Inc. Method of treating neurodegenerative disease
EP2237786A2 (en) 2007-12-21 2010-10-13 Coda Therapeutics, Inc. Use of inhibitors of connexin43 for treatment of fibrotic conditions
EP2237795A2 (en) * 2007-12-21 2010-10-13 Coda Therapeutics, Inc. Use of anti-connexin polypeptide agent in combination with anti-connexin polynucleotide agent for the treatment of fibrotic conditions
JP2011507859A (en) * 2007-12-21 2011-03-10 コーダ セラピューティクス， インコーポレイテッド Use of anti-connexin polynucleotides for the treatment of surgical adhesions
AU2008343839A1 (en) * 2007-12-21 2009-07-09 Coda Therapeutics, Inc. Use of anti-connexin peptides, alone or in combination with anti-connexin polynucleotides, for the treatment of surgical adhesions
EP2077335A1 (en) * 2007-12-22 2009-07-08 Universitätsklinikum Schleswig-Holstein EXO1 promoter polymorphism associated with exceptional life expectancy in humans
ES2535419T3 (en) * 2007-12-27 2015-05-11 Protiva Biotherapeutics Inc. Polo kinase expression silencing using interfering RNA
EP2075333A1 (en) * 2007-12-28 2009-07-01 Qiagen GmbH Positive controls for expression modulating experiments
CA2711635A1 (en) * 2008-01-07 2009-08-06 Coda Therapeutics, Inc. Wound healing compositions and treatments
WO2009102931A1 (en) * 2008-02-15 2009-08-20 Alcon Research, Ltd. Rnai-mediated inhibition of connexin 43 for treatment of iop-related conditions
WO2009113579A1 (en) * 2008-03-11 2009-09-17 学校法人埼玉医科大学 Double-stranded nucleic acid molecule suitable for prevention or treatment of cancer, cancer cell proliferation inhibitor, and pharmaceutical preparation
DK2279254T3 (en) 2008-04-15 2017-09-18 Protiva Biotherapeutics Inc PRESENT UNKNOWN LIPID FORMS FOR NUCLEIC ACID ADMINISTRATION
US8962614B2 (en) * 2008-04-17 2015-02-24 The Johns Hopkins University ON01910.Na enhances chemotherapeutic agent activity in drug-resistant tumors
US8217161B2 (en) * 2008-04-22 2012-07-10 Clemson University Research Foundation Methods of inhibiting multiple cytochrome P450 genes with siRNA
DE102008029669A1 (en) * 2008-05-16 2009-11-19 Schlaak, Jörg Friedrich, Prof. Dr. med. New therapeutics for hepatitis therapy
WO2009158432A2 (en) 2008-06-27 2009-12-30 Amgen Inc. Ang-2 inhibition to treat multiple sclerosis
US8946172B2 (en) * 2008-08-25 2015-02-03 Excaliard Pharmaceuticals, Inc. Method for reducing scarring during wound healing using antisense compounds directed to CTGF
MX2011002143A (en) 2008-08-25 2011-07-20 Excaliard Pharmaceuticals Inc Antisense oligonucleotides directed against connective tissue growth factor and uses thereof.
WO2010027279A2 (en) * 2008-09-04 2010-03-11 Genesis Research And Development Corporation Limited Compositions and methods for the treatment and prevention of neoplastic disorders
EP3336188B1 (en) 2008-09-22 2020-05-06 Phio Pharmaceuticals Corp. Reduced size self-delivering rnai compounds
EP3025727A1 (en) 2008-10-02 2016-06-01 The J. David Gladstone Institutes Methods of treating liver disease
US9095592B2 (en) * 2008-11-07 2015-08-04 The Research Foundation For The State University Of New York Bruton's tyrosine kinase as anti-cancer drug target
US20100233270A1 (en) 2009-01-08 2010-09-16 Northwestern University Delivery of Oligonucleotide-Functionalized Nanoparticles
US20120016011A1 (en) * 2009-03-19 2012-01-19 Merck Sharp & Dohme Corp. RNA Interference Mediated Inhibition of Connective Tissue Growth Factor (CTGF) Gene Expression Using Short Interfering Nucleic Acid (siNA)
US20110045080A1 (en) * 2009-03-24 2011-02-24 William Marsh Rice University Single-Walled Carbon Nanotube/Bioactive Substance Complexes and Methods Related Thereto
WO2010111497A2 (en) * 2009-03-27 2010-09-30 Merck Sharp & Dohme Corp. RNA INTERFERENCE MEDIATED INHIBITION OF THE INTERCELLULAR ADHESION MOLECULE 1 (ICAM-1)GENE EXPRESSION USING SHORT INTERFERING NUCLEIC ACID (siNA)
EP2414524B1 (en) * 2009-04-03 2017-08-23 Centre National De La Recherche Scientifique Gene transfer vectors comprising genetic insulator elements and methods to identify genetic insulator elements
US20110038852A1 (en) * 2009-06-10 2011-02-17 3-V Biosciences, Inc. Antivirals that target transporters, carriers, and ion channels
CN101851625B (en) * 2009-06-25 2012-07-18 上海交通大学 RNA interference sequences of glucagon receptor gene
CN101586103B (en) * 2009-06-25 2010-12-08 上海交通大学 RNA interference sequence of glucagon receptor gene
CN101851624B (en) * 2009-06-25 2012-04-18 上海交通大学 RNA interference sequences of glucagon receptor gene
EP2449114B9 (en) 2009-07-01 2017-04-19 Protiva Biotherapeutics Inc. Novel lipid formulations for delivery of therapeutic agents to solid tumors
US10165286B2 (en) 2009-07-08 2018-12-25 Dejero Labs Inc. System and method for automatic encoder adjustment based on transport data
EP2480668A2 (en) * 2009-09-23 2012-08-01 Protiva Biotherapeutics Inc. Compositions and methods for silencing genes expressed in cancer
US9799416B2 (en) * 2009-11-06 2017-10-24 Terrapower, Llc Methods and systems for migrating fuel assemblies in a nuclear fission reactor
US20110152343A1 (en) * 2009-12-22 2011-06-23 Functional Genetics, Inc. Protease inhibitors and broad-spectrum antiviral
US20130023578A1 (en) * 2009-12-31 2013-01-24 Samyang Biopharmaceuticals Corporation siRNA for inhibition of c-Met expression and anticancer composition containing the same
CN111700901A (en) 2010-01-08 2020-09-25 Ionis制药公司 Modulation of angiopoietin-like 3 expression
CN102102101A (en) * 2010-03-05 2011-06-22 重庆医科大学 SiRNA for inhibiting expression of Plk1 and use thereof
FR2957426B1 (en) * 2010-03-12 2012-03-02 Thales Sa AUTONOMOUS PSEUDOLITE POSITIONING SYSTEM IN THE CONSTRAINED ZONE AND METHOD OF IMPLEMENTING THE SAME
EP2550001B1 (en) 2010-03-24 2019-05-22 Phio Pharmaceuticals Corp. Rna interference in ocular indications
KR20180044433A (en) 2010-03-24 2018-05-02 알엑스아이 파마슈티칼스 코포레이션 Rna interference in dermal and fibrotic indications
EP2380595A1 (en) 2010-04-19 2011-10-26 Nlife Therapeutics S.L. Compositions and methods for selective delivery of oligonucleotide molecules to specific neuron types
WO2011133658A1 (en) 2010-04-22 2011-10-27 Boston Medical Center Corporation Compositions and methods for targeting and delivering therapeutics into cells
EP2575799A4 (en) * 2010-05-19 2013-11-13 Beth Israel Hospital Methods for treating inflammatory autoimmune disorders
EP2390327A1 (en) * 2010-05-27 2011-11-30 Sylentis S.A. siRNA and their use in methods and compositions for the treatment and/or prevention of eye conditions
US9074721B2 (en) 2010-06-09 2015-07-07 Alex Lau Support system
US9316346B2 (en) 2010-06-09 2016-04-19 Colebrook Bosson Saunders (Products) Limited Support system
PT2585596T (en) * 2010-06-23 2021-03-23 Curna Inc Treatment of sodium channel, voltage-gated, alpha subunit (scna) related diseases by inhibition of natural antisense transcript to scna
EP2598639A4 (en) 2010-07-28 2014-08-13 Alcon Res Ltd Sirna targeting vegfa and methods for treatment in vivo
USD684982S1 (en) 2010-08-11 2013-06-25 Colebrook Bosson Saunders (Products) Limited Display support with indicator window
WO2012027713A2 (en) * 2010-08-26 2012-03-01 Alnylam Pharmaceuticals, Inc. Compositions and methods for inhibition of snca
WO2012034942A1 (en) 2010-09-13 2012-03-22 Santaris Pharma A/S Compounds for the modulation of aurora kinase b expression
ES2732929T3 (en) 2010-10-22 2019-11-26 Olix Pharmaceuticals Inc Nucleic acid molecules that induce RNA interference and uses thereof
US9587033B2 (en) 2010-11-15 2017-03-07 University Of Florida Research Foundation, Inc. Therapeutic and diagnostic applications targeting TNK-1
AU2011329777B2 (en) * 2010-11-17 2016-06-09 Ionis Pharmaceuticals, Inc. Modulation of alpha synuclein expression
US20150247148A1 (en) * 2011-01-31 2015-09-03 Kyowa Hakko Kirin Co., Ltd. Composition for suppressing expression of target gene
KR101697396B1 (en) 2011-02-02 2017-01-17 엑스칼리아드 파마슈티컬즈, 인코포레이티드 Method of treating keloids or hypertrophic scars using antisense compounds targeting connective tissue growth factor (ctgf)
KR101839177B1 (en) 2011-04-13 2018-03-15 아이오니스 파마수티컬즈, 인코포레이티드 Antisense modulation of ptpib expression
WO2012151199A1 (en) 2011-05-02 2012-11-08 Immunomedics, Inc. Ultrafiltration concentration of allotype selected antibodies for small-volume administration
US8802839B2 (en) 2011-07-15 2014-08-12 Fibrogen, Inc. Connective tissue growth factor antisense oligonucleotides
CA2842034C (en) 2011-07-18 2023-09-05 University Of Kentucky Research Foundation Protection of cells from alu-rna-induced degeneration and inhibitors for protecting cells
CN104024413B (en) * 2011-10-19 2016-08-17 苏州瑞博生物技术有限公司 SiRNA and application thereof and the method for suppression plk1 gene expression
EP2776130A1 (en) 2011-11-07 2014-09-17 Institut National de la Sante et de la Recherche Medicale (INSERM) A ddr1 antagonist or an inhibitor of ddr1 gene expression for use in the prevention or treatment of crescentic glomerulonephritis
US9006199B2 (en) * 2011-11-14 2015-04-14 Silenseed Ltd. Methods and compositions for treating prostate cancer
EP2782930B1 (en) 2011-11-27 2018-07-11 Yeda Research and Development Co. Ltd. Methods of regulating angiogenesis and compositions capable of same
US9707235B1 (en) 2012-01-13 2017-07-18 University Of Kentucky Research Foundation Protection of cells from degeneration and treatment of geographic atrophy
KR20220045089A (en) 2012-02-24 2022-04-12 아뷰터스 바이오파마 코포레이션 Trialkyl cationic lipids and methods of use thereof
WO2013123996A1 (en) * 2012-02-24 2013-08-29 Astrazeneca Uk Limited Novel sirna inhibitors of human icam-1
AU2013225619A1 (en) 2012-02-29 2014-10-16 Benitec Biopharma Limited Pain treatment
TR201905395T4 (en) * 2012-03-15 2019-05-21 Hyun Kee Kim Anti-gremlin-1 antibody.
DE202012003753U1 (en) * 2012-04-13 2013-07-17 Joseph Vögele AG Cross distribution arrangement for a paver
CN102690826B (en) * 2012-04-19 2014-03-05 山西医科大学 shRNA for specifically reducing human Aurora-A gene expression and application thereof
ES2716818T3 (en) * 2012-05-22 2019-06-17 Olix Pharmaceuticals Inc Nuclear interference-inducing nucleic acid molecule capable of penetrating cells and using it
CN109513003A (en) 2012-08-14 2019-03-26 Ibc药品公司 T- cell for treating disease redirects bispecific antibody
JP2015533792A (en) 2012-09-05 2015-11-26 シレンティス・エセ・ア・ウ siRNA and its use in methods and compositions for the treatment and / or prevention of ocular conditions
GB201215857D0 (en) 2012-09-05 2012-10-24 Sylentis Sau siRNA and their use in methods and compositions for the treatment and/or prevention of eye conditions
JP6386461B2 (en) * 2012-10-26 2018-09-05 エヌライフ、セラピューティックス、ソシエダッド、リミターダＮｌｉｆｅ Ｔｈｅｒａｐｅｕｔｉｃｓ， Ｓ．Ｌ． Compositions and methods for the treatment of Parkinson's disease by selective delivery of oligonucleotide molecules to specific neuronal species
US20150258127A1 (en) 2012-10-31 2015-09-17 INSERM (Institut National de la Santé et de la Recherche Médicale) Methods for preventing antiphospholipid syndrome (aps)
WO2014110006A1 (en) 2013-01-10 2014-07-17 Ge Healthcare Dharmacon, Inc. Templates, libraries, kits and methods for generating molecules
US10227589B2 (en) * 2013-01-28 2019-03-12 Council Of Scientific And Industrial Research Method for inhibiting tumor growth through RNA-interference using liposomally associated CDC20 siRNA
US9777274B2 (en) 2013-03-15 2017-10-03 Techulon Inc. Antisense molecules for treatment of staphylococcus aureus infection
EP2970976A4 (en) 2013-03-15 2016-11-02 Techulon Inc Antisense molecules for treatment of staphylococcus aureus infection
LT2999785T (en) 2013-05-22 2018-07-25 Alnylam Pharmaceuticals, Inc. SERPINA1 iRNA COMPOSITIONS AND METHODS OF USE THEREOF
EP3017047A4 (en) 2013-07-03 2017-06-14 Dicerna Pharmaceuticals Inc. Methods and compositions for the specific inhibition of alpha-1 antitrypsin by double-stranded rna
RU2656154C2 (en) * 2013-07-05 2018-05-31 Байонир Корпорейшн Gen-specific sirna related to respiratory disease, two-spiral construct of oligo-rna comprising sirna, and containing it composition for prevention or treatment of respiratory disease
KR20150006742A (en) * 2013-07-09 2015-01-19 (주)바이오니아 Liver cancer related genes-specific siRNA, double-stranded oligo RNA molecules comprising the siRNA, and composition for the prevention or treatment of cancer comprising the same
TW201536329A (en) 2013-08-09 2015-10-01 Isis Pharmaceuticals Inc Compounds and methods for modulation of dystrophia myotonica-protein kinase (DMPK) expression
CA2921839A1 (en) 2013-08-28 2015-03-05 Ionis Pharmaceuticals, Inc. Modulation of prekallikrein (pkk) expression
US10150965B2 (en) 2013-12-06 2018-12-11 Dicerna Pharmaceuticals, Inc. Methods and compositions for the specific inhibition of transthyretin (TTR) by double-stranded RNA
IL294470A (en) * 2013-12-12 2022-09-01 Alnylam Pharmaceuticals Inc Complement component irna compositions and methods of use thereof
PT3087183T (en) 2013-12-24 2020-10-08 Ionis Pharmaceuticals Inc Modulation of angiopoietin-like 3 expression
EP3581654B1 (en) 2013-12-27 2021-03-31 Dicerna Pharmaceuticals, Inc. Methods and compositions for the specific inhibition of glycolate oxidase (hao1) by double-stranded rna
WO2015116696A1 (en) 2014-01-28 2015-08-06 Massachusetts Institute Of Technology Combination therapies and methods of use thereof for treating cancer
MX2016010683A (en) 2014-02-21 2017-05-11 Ibc Pharmaceuticals Inc Disease therapy by inducing immune response to trop-2 expressing cells.
WO2015130416A1 (en) 2014-02-25 2015-09-03 Immunomedics, Inc. Humanized rfb4 anti-cd22 antibody
WO2015132303A1 (en) 2014-03-04 2015-09-11 Sylentis Sau Sirnas and their use in methods and compositions for the treatment and/or prevention of eye conditions
AU2015240761B2 (en) 2014-04-01 2019-09-12 Biogen Ma Inc. Compositions for modulating SOD-1 expression
WO2016005514A1 (en) * 2014-07-10 2016-01-14 Stichting Katholieke Universiteit Antisense oligonucleotides for the treatment of usher syndrome type 2
EP3176269B1 (en) * 2014-07-29 2020-12-02 Wellmarker Bio Co., Ltd. Inhibitors of met and igsf1 for treating cancer
JP6683686B2 (en) 2014-08-22 2020-04-22 オークランド ユニサービシーズ リミティド Channel regulator
CN104258377B (en) * 2014-09-10 2017-06-30 中南大学湘雅医院 Application of the PIK3C2A albumen in liver-cancer medicine is treated
EP3666896A1 (en) 2014-10-10 2020-06-17 Dicerna Pharmaceuticals, Inc. Therapeutic inhibition of lactate dehydrogenase and agents therefor
EP3212794B1 (en) 2014-10-30 2021-04-07 Genzyme Corporation Polynucleotide agents targeting serpinc1 (at3) and methods of use thereof
SG11201703419UA (en) 2014-11-14 2017-05-30 Voyager Therapeutics Inc Modulatory polynucleotides
CN107109407A (en) 2014-11-14 2017-08-29 沃雅戈治疗公司 Treat the composition and method of amyotrophic lateral sclerosis (ALS)
CA2968531A1 (en) 2014-11-21 2016-05-26 Northwestern University The sequence-specific cellular uptake of spherical nucleic acid nanoparticle conjugates
CN104450710B (en) * 2014-11-28 2018-06-05 广州市锐博生物科技有限公司 Inhibit nucleic acid oligomer and its application of MYD88 genes
US10036017B2 (en) 2015-02-17 2018-07-31 Dicerna Pharmaceuticals, Inc. Methods and compositions for the specific inhibition of complement component 5(C5) by double-stranded RNA
US11129844B2 (en) 2015-03-03 2021-09-28 Ionis Pharmaceuticals, Inc. Compositions and methods for modulating MECP2 expression
CN104673799B (en) * 2015-03-09 2019-02-05 浙江大学 It targets the siRNA of people RAN and its is preparing the purposes in anti-viral hepatitis type C drug
US10689647B2 (en) 2015-05-08 2020-06-23 Dicerna Pharmaceuticals, Inc. Methods and compositions for the specific inhibition of antithrombin 3 (AT3) by double-stranded RNA
EP4285912A2 (en) 2015-09-25 2023-12-06 Ionis Pharmaceuticals, Inc. Compositions and methods for modulating ataxin 3 expression
KR20180071362A (en) 2015-11-16 2018-06-27 올릭스 주식회사 Treatment of age-related macular degeneration with RNA complex targeting MyD88 or TLR3
SG11201804729RA (en) 2015-12-07 2018-07-30 Genzyme Corp Methods and compositions for treating a serpinc1-associated disorder
WO2017100193A1 (en) * 2015-12-10 2017-06-15 Fibrogen, Inc. Methods for treatment of motor neuron diseases
CN105462978B (en) * 2015-12-18 2018-05-15 中国科学院北京基因组研究所 One species specificity suppresses siRNA and its application of MAGEA1 gene expressions
CN105462977B (en) * 2015-12-18 2018-05-11 中国科学院北京基因组研究所 One species specificity suppresses siRNA and its application of MAGEA1 gene expressions
US10478503B2 (en) 2016-01-31 2019-11-19 University Of Massachusetts Branched oligonucleotides
KR20180104075A (en) 2016-02-02 2018-09-19 올릭스 주식회사 Treatment of atopic dermatitis and asthma using RNA complexes targeting IL4Ra, TRPA1, or F2RL1
KR20180104692A (en) 2016-02-02 2018-09-21 올릭스 주식회사 Treatment of angiogenesis related diseases using RNA complexes targeting ANGPT2 and PDGFB
US10301628B2 (en) 2016-04-11 2019-05-28 Olix Pharmaceuticals, Inc. Treatment of idiopathic pulmonary fibrosis using RNA complexes that target connective tissue growth factor
CN110214187B (en) 2016-05-18 2024-01-30 沃雅戈治疗公司 Regulatory polynucleotides
CN109312403B (en) * 2016-06-17 2023-06-27 豪夫迈·罗氏有限公司 In vitro nephrotoxicity screening assay
KR101916652B1 (en) 2016-06-29 2018-11-08 올릭스 주식회사 Compounds improving RNA interference of small interfering RNA and use thereof
WO2018039629A2 (en) 2016-08-25 2018-03-01 Northwestern University Micellar spherical nucleic acids from thermoresponsive, traceless templates
JOP20190104A1 (en) 2016-11-10 2019-05-07 Ionis Pharmaceuticals Inc Compounds and methods for reducing atxn3 expression
TWI788312B (en) 2016-11-23 2023-01-01 美商阿尼拉製藥公司 SERPINA1 iRNA COMPOSITIONS AND METHODS OF USE THEREOF
WO2018106959A1 (en) 2016-12-07 2018-06-14 Progenity Inc. Gastrointestinal tract detection methods, devices and systems
AU2017378393B2 (en) 2016-12-14 2023-03-09 Biora Therapeutics, Inc. Treatment of a disease of the gastrointestinal tract with a TNF inhibitor
CN110770343A (en) 2017-02-10 2020-02-07 成均馆大学校产学协力团 Long double-stranded RNA for RNA interference
DE102017206836A1 (en) * 2017-04-24 2018-10-25 Zf Friedrichshafen Ag Transmission for a motor vehicle
JP2020517714A (en) 2017-04-28 2020-06-18 オークランド ユニサービシズ リミテッド Treatment method and new construct
EP3618839A4 (en) 2017-05-05 2021-06-09 Voyager Therapeutics, Inc. Compositions and methods of treating amyotrophic lateral sclerosis (als)
WO2018209270A1 (en) 2017-05-11 2018-11-15 Northwestern University Adoptive cell therapy using spherical nucleic acids (snas)
BR112020003126A2 (en) 2017-09-14 2020-10-13 Arrowhead Pharmaceuticals, Inc. rnai agents and compositions for inhibiting the expression of angiopoietin type 3 (angptl3), and methods of use
CN111182913A (en) * 2017-10-02 2020-05-19 西奈医疗中心 Methods and compositions for effective delivery through multiple biological barriers
KR102609396B1 (en) 2017-10-13 2023-12-01 노보 노르디스크 헬스 케어 악티엔게젤샤프트 Methods and compositions for inhibiting expression of LDHA
AU2018352236A1 (en) 2017-10-16 2020-04-23 The Curators Of The University Of Missouri Treatment of amyotrophic lateral sclerosis (ALS)
WO2019079242A1 (en) 2017-10-16 2019-04-25 Voyager Therapeutics, Inc. Treatment of amyotrophic lateral sclerosis (als)
TWI809004B (en) 2017-11-09 2023-07-21 美商Ionis製藥公司 Compounds and methods for reducing snca expression
JP2021504415A (en) 2017-12-01 2021-02-15 スーチョウ リボ ライフ サイエンス カンパニー、リミテッドＳｕｚｈｏｕ Ｒｉｂｏ Ｌｉｆｅ Ｓｃｉｅｎｃｅ Ｃｏ．， Ｌｔｄ． Compositions and complexes containing double-stranded oligonucleotides, double-stranded oligonucleotides, and preparation methods and uses.
WO2019105419A1 (en) 2017-12-01 2019-06-06 苏州瑞博生物技术有限公司 Nucleic acid, composition and conjugate containing same, preparation method, and use
MX2020005860A (en) 2017-12-06 2020-09-09 Avidity Biosciences Inc Compositions and methods of treating muscle atrophy and myotonic dystrophy.
BR112020012523A2 (en) 2017-12-21 2020-11-24 F. Hoffmann-La Roche Ag complementary diagnosis for htra1 rna antagonists
CN116375774A (en) 2017-12-29 2023-07-04 苏州瑞博生物技术股份有限公司 Conjugate, preparation method and application thereof
BR112020013848A2 (en) 2018-01-08 2020-12-01 Iovance Biotherapeutics, Inc. methods for expanding tumor-infiltrating lymphocytes and for treating an individual with cancer, tumor-infiltrating lymphocyte population, and, method for evaluating transcription factors
WO2019136459A1 (en) 2018-01-08 2019-07-11 Iovance Biotherapeutics, Inc. Processes for generating til products enriched for tumor antigen-specific t-cells
US11713446B2 (en) 2018-01-08 2023-08-01 Iovance Biotherapeutics, Inc. Processes for generating TIL products enriched for tumor antigen-specific T-cells
CR20200301A (en) 2018-01-12 2020-10-26 Roche Innovation Ct Copenhagen As Alpha-synuclein antisense oligonucleotides and uses thereof
WO2019169243A1 (en) 2018-03-02 2019-09-06 Ionis Pharmaceuticals, Inc. Compounds and methods for the modulation of amyloid-beta precursor protein
JP2021522800A (en) 2018-05-09 2021-09-02 アイオーニス ファーマシューティカルズ， インコーポレーテッドＩｏｎｉｓ Ｐｈａｒｍａｃｅｕｔｉｃａｌｓ，Ｉｎｃ． Compounds and methods for reducing ATXN3 expression
US11162083B2 (en) 2018-06-14 2021-11-02 University Of South Carolina Peptide based inhibitors of Raf kinase protein dimerization and kinase activity
WO2019246313A1 (en) 2018-06-20 2019-12-26 Progenity, Inc. Treatment of a disease of the gastrointestinal tract with a tnf inhibitor
MX2020013653A (en) 2018-06-27 2021-03-02 Ionis Pharmaceuticals Inc Compounds and methods for reducing lrrk2 expression.
WO2020010042A1 (en) 2018-07-02 2020-01-09 Voyager Therapeutics, Inc. Treatment of amyotrophic lateral sclerosis and disorders associated with the spinal cord
US11918600B2 (en) 2018-08-21 2024-03-05 Suzhou Ribo Life Science Co., Ltd. Nucleic acid, pharmaceutical composition and conjugate containing nucleic acid, and use thereof
EP3862024A4 (en) 2018-09-30 2022-08-17 Suzhou Ribo Life Science Co., Ltd. Sirna conjugate, preparation method therefor and use thereof
KR20210095165A (en) 2018-11-19 2021-07-30 프로제너티, 인크. Methods and devices for treating diseases with biopharmaceuticals
WO2020104669A1 (en) * 2018-11-23 2020-05-28 Silence Therapeutics Gmbh Nucleic acids for inhibiting expression of c3 in a cell
EP3956450A4 (en) 2019-07-26 2022-11-16 Ionis Pharmaceuticals, Inc. Compounds and methods for modulating gfap
EP4022060A1 (en) 2019-08-27 2022-07-06 Silence Therapeutics GmbH Nucleic acids for inhibiting expression of c3 in a cell
CN112442500A (en) * 2019-08-30 2021-03-05 恩智（广州）医药科技有限公司 siRNA for inhibiting MCM7, composition and application thereof
US20220290155A1 (en) * 2019-08-30 2022-09-15 Agency For Science, Technology And Research A Method Of Promoting Survival And/Or Function Of A Motor Neuron And Related Agents, Uses And Methods
CN110938691B (en) * 2019-12-03 2023-07-07 兰州大学 Application of human DUS4L gene and related products
EP4309722A2 (en) 2019-12-13 2024-01-24 Biora Therapeutics, Inc. Ingestible device for delivery of therapeutic agent to the gastrointestinal tract
AU2021237465A1 (en) 2020-03-19 2022-10-13 Avidity Biosciences, Inc. Compositions and methods of treating Facioscapulohumeral muscular dystrophy
CN113444723A (en) * 2020-03-27 2021-09-28 北京键凯科技股份有限公司 Interfering RNA for inhibiting vascular endothelial growth factor receptor 2 gene expression and application thereof
US11446387B2 (en) 2020-03-27 2022-09-20 Avidity Biosciences, Inc. Compositions and methods of treating muscle dystrophy
BR112022020020A2 (en) 2020-04-04 2022-11-22 Pfizer METHODS OF TREATMENT OF CORONAVIRUS DISEASE 2019
EP4159858A1 (en) * 2020-05-26 2023-04-05 Olix Pharmaceuticals, Inc. Rnai agent targeting myd88 and use thereof
US11459567B2 (en) 2020-06-24 2022-10-04 Patricia Virginia Elizalde Specific siRNA molecules, composition and use thereof for the treatment of triple negative breast cancer
WO2022006134A2 (en) 2020-06-29 2022-01-06 Ionis Pharmaceuticals, Inc. Compounds and methods for modulating plp1
CN111849992A (en) * 2020-08-17 2020-10-30 南通大学 siRNA molecule of targeted c-Met gene and application thereof
TW202227627A (en) 2020-09-11 2022-07-16 美商愛羅海德製藥公司 Rnai agents for inhibiting expression of dux4, compositions thereof, and methods of use
JP2023550485A (en) * 2020-11-23 2023-12-01 ユニバーシティー オブ マサチューセッツ Oligonucleotides for DGAT2 regulation
WO2022136466A1 (en) * 2020-12-23 2022-06-30 Argonaute RNA Limited Treatment of cardiovascular disease
EP4291654A2 (en) 2021-02-12 2023-12-20 Alnylam Pharmaceuticals, Inc. Superoxide dismutase 1 (sod1) irna compositions and methods of use thereof for treating or preventing superoxide dismutase 1- (sod1-) associated neurodegenerative diseases
KR102644654B1 (en) 2021-04-19 2024-03-07 노보 노르디스크 에이/에스 Compositions and methods for inhibiting nuclear receptor subfamily 1 group H member 3 (NR1H3) expression
JP7463621B2 (en) 2021-05-28 2024-04-08 ノヴォ ノルディスク アー／エス Compositions and methods for inhibiting mitochondrial amidoxime reducing component 1 (MARC1) expression
US11833221B2 (en) 2021-09-01 2023-12-05 Ionis Pharmaceuticals, Inc. Oligomeric compounds for reducing DMPK expression
WO2023122805A1 (en) 2021-12-20 2023-06-29 Vestaron Corporation Sorbitol driven selection pressure method
WO2023133639A1 (en) * 2022-01-13 2023-07-20 University Health Network Targeting the ythdf1 - arhgef2 axis for cancer treatment
WO2023144798A1 (en) 2022-01-31 2023-08-03 Genevant Sciences Gmbh Ionizable cationic lipids for lipid nanoparticles
WO2023158514A1 (en) 2022-02-18 2023-08-24 Massachusetts Institute Of Technology Cancer treatment by combined inhibition of polo-like kinase and microtubule polymerization
WO2023245060A2 (en) 2022-06-15 2023-12-21 Arrowhead Pharmaceuticals, Inc. Rnai agents for inhibiting expression of superoxide dismutase 1 (sod1), compositions thereof, and methods of use

Family Cites Families (175)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US702831A (en) * 1901-08-20 1902-06-17 Samuel Thompson Combined double escutcheon and paint protector.
US760807A (en) * 1903-11-11 1904-05-24 Lee Roberts Cutting-nippers.
US5831066A (en) 1988-12-22 1998-11-03 The Trustees Of The University Of Pennsylvania Regulation of bcl-2 gene expression
US5741671A (en) * 1991-12-12 1998-04-21 The Johns Hopkins University Isolation cloning and expression of transmembrane water channel aquaporin 1(AQP1)
US20040204380A1 (en) 1999-01-07 2004-10-14 Ackermann Elizabeth J Antisense modulation of novel anti-apoptotic bcl-2-related proteins
US6001992A (en) 1999-01-07 1999-12-14 Isis Pharmaceuticals Inc. Antisense modulation of novel anti-apoptotic bcl-2-related proteins
US5981175A (en) * 1993-01-07 1999-11-09 Genpharm Internation, Inc. Methods for producing recombinant mammalian cells harboring a yeast artificial chromosome
US20020086321A1 (en) 1993-02-02 2002-07-04 Craig Ruth W. Myeloid cell leukemia associated gene MCL-1
US5641756A (en) 1993-07-27 1997-06-24 Hybridon, Inc. Modified VEGF oligonucleotides
US6410322B1 (en) 1993-07-27 2002-06-25 Hybridon Inc Antisense oligonucleotide inhibition of vascular endothelial growth factor expression
US5731294A (en) 1993-07-27 1998-03-24 Hybridon, Inc. Inhibition of neovasularization using VEGF-specific oligonucleotides
ATE167518T1 (en) 1994-02-14 1998-07-15 Amgen Inc CELL CYCLE PROTEIN IN MAMMALS
US6150092A (en) * 1994-06-27 2000-11-21 Taogosei Company, Ltd. Antisense nucleic acid compound targeted to VEGF
US5830879A (en) 1995-10-02 1998-11-03 St. Elizabeth's Medical Center Of Boston, Inc. Treatment of vascular injury using vascular endothelial growth factor
US6346398B1 (en) 1995-10-26 2002-02-12 Ribozyme Pharmaceuticals, Inc. Method and reagent for the treatment of diseases or conditions related to levels of vascular endothelial growth factor receptor
WO1997020925A1 (en) 1995-12-08 1997-06-12 Hybridon, Inc. Modified vegf antisense oligonucleotides for treatment of skin disorders
JP2000501941A (en) 1995-12-08 2000-02-22 ハイブリドン・インコーポレイテッド Modified VEGF antisense oligonucleotide
US6716575B2 (en) 1995-12-18 2004-04-06 Sugen, Inc. Diagnosis and treatment of AUR1 and/or AUR2 related disorders
EP0910634A2 (en) 1996-04-17 1999-04-28 Hoechst Marion Roussel Deutschland GmbH ANTISENSE INHIBITORS OF VASCULAR ENDOTHELIAL GROWTH FACTOR (VEgF/VPF) EXPRESSION
US20050261485A1 (en) * 1996-05-23 2005-11-24 Toagosei Co., Ltd., A Japan Corporation Method of producing antisense oligonucleotide
CA2259123C (en) 1996-07-01 2003-10-21 Jim A. Wright Oligonucleotides from the untranslated regions of ribonucleotide reductase and their use to modulate cell growth
AU6237198A (en) 1996-12-19 1998-07-15 Isis Pharmaceuticals, Inc. Large-scale purification of full length oligonucleotides by solid-liquid affinity extraction
US6800744B1 (en) * 1997-07-02 2004-10-05 Genome Therapeutics Corporation Nucleic acid and amino acid sequences relating to Streptococcus pneumoniae for diagnostics and therapeutics
JPH1142091A (en) 1997-07-25 1999-02-16 Toagosei Co Ltd Anti-sense nucleic acid compound
CA2248762A1 (en) * 1997-10-22 1999-04-22 University Technologies International, Inc. Antisense oligodeoxynucleotides regulating expression of tnf-.alpha.
US6506559B1 (en) 1997-12-23 2003-01-14 Carnegie Institute Of Washington Genetic inhibition by double-stranded RNA
WO1999038965A1 (en) 1998-01-29 1999-08-05 The Trustees Of Columbia University In The City Of New York Human hairless gene, protein and uses thereof
US6111086A (en) 1998-02-27 2000-08-29 Scaringe; Stephen A. Orthoester protecting groups
US20030228597A1 (en) 1998-04-13 2003-12-11 Cowsert Lex M. Identification of genetic targets for modulation by oligonucleotides and generation of oligonucleotides for gene modulation
CA2326823A1 (en) 1998-04-20 1999-10-28 Ribozyme Pharmaceuticals, Inc. Nucleic acid molecules with novel chemical compositions capable of modulating gene expression
WO1999055910A1 (en) 1998-04-24 1999-11-04 Arizona Board Of Regents Method of inducing apoptosis in a target cell
DE69919869T2 (en) 1998-06-10 2005-09-29 Biognostik Gesellschaft für Biomolekulare Diagnostik mbH STIMULATION OF THE IMMUNE SYSTEM
EP1093373A4 (en) 1998-07-02 2005-04-13 Univ Columbia OLIGONUCLEOTIDE INHIBITORS OF bcl-xL
US6228642B1 (en) 1998-10-05 2001-05-08 Isis Pharmaceuticals, Inc. Antisense oligonucleotide modulation of tumor necrosis factor-(α) (TNF-α) expression
US6172216B1 (en) 1998-10-07 2001-01-09 Isis Pharmaceuticals Inc. Antisense modulation of BCL-X expression
WO2000021559A2 (en) 1998-10-09 2000-04-20 Musc Foundation For Research Development Blocking factor b to treat complement-mediated immune disease
PL348163A1 (en) 1998-11-06 2002-05-06 Basf Ag Inhibition of the formation of vascular hyperpermeability
US20040235041A1 (en) * 1998-11-17 2004-11-25 Shimkets Richard A. Nucleic acids containing single nucleotide polymorphisms and methods of use thereof
EP1147189A2 (en) 1998-12-04 2001-10-24 Immusol, Inc. Ribozyme therapy for the treatment and/or prevention of restenosis
US5958773A (en) 1998-12-17 1999-09-28 Isis Pharmaceuticals Inc. Antisense modulation of AKT-1 expression
CN1166249C (en) 1999-01-19 2004-09-08 西门子公司 Method for the time synchronisation of a computer network and computer network with time synchronisation
DE19956568A1 (en) 1999-01-30 2000-08-17 Roland Kreutzer Method and medicament for inhibiting the expression of a given gene
ES2265948T3 (en) 1999-06-14 2007-03-01 Cancer Research Technology Limited THERAPY FOR CANCER.
DE19928367A1 (en) * 1999-06-21 2000-12-28 Will E C H Gmbh & Co Handling of stacked sheet materials has a facility for stacking and separating into selected stack size
US6770633B1 (en) 1999-10-26 2004-08-03 Immusol, Inc. Ribozyme therapy for the treatment of proliferative skin and eye diseases
GB9927444D0 (en) 1999-11-19 2000-01-19 Cancer Res Campaign Tech Inhibiting gene expression
DE10160151A1 (en) 2001-01-09 2003-06-26 Ribopharma Ag Inhibiting expression of target gene, useful e.g. for inhibiting oncogenes, by administering double-stranded RNA complementary to the target and having an overhang
US7179796B2 (en) 2000-01-18 2007-02-20 Isis Pharmaceuticals, Inc. Antisense modulation of PTP1B expression
US20070026394A1 (en) 2000-02-11 2007-02-01 Lawrence Blatt Modulation of gene expression associated with inflammation proliferation and neurite outgrowth using nucleic acid based technologies
US8202979B2 (en) * 2002-02-20 2012-06-19 Sirna Therapeutics, Inc. RNA interference mediated inhibition of gene expression using chemically modified short interfering nucleic acid
WO2003070918A2 (en) 2002-02-20 2003-08-28 Ribozyme Pharmaceuticals, Incorporated Rna interference by modified short interfering nucleic acid
US20030084471A1 (en) 2000-03-16 2003-05-01 David Beach Methods and compositions for RNA interference
EP1272630A2 (en) 2000-03-16 2003-01-08 Genetica, Inc. Methods and compositions for rna interference
ES2336887T5 (en) 2000-03-30 2019-03-06 Whitehead Inst Biomedical Res Mediators of RNA interference specific to RNA sequences
US6509559B1 (en) * 2000-06-20 2003-01-21 Ppt Vision, Inc. Binary optical grating and method for generating a moire pattern for 3D imaging
US6521258B1 (en) 2000-09-08 2003-02-18 Ferro Corporation Polymer matrices prepared by supercritical fluid processing techniques
US20030186920A1 (en) 2000-10-13 2003-10-02 Sirois Martin G. Antisense oligonucleotide directed toward mammalian vegf receptor genes and uses thereof
AU2002220070A1 (en) 2000-11-01 2002-05-15 Bki Holding Corporation Cellulose ethers and method of preparing the same
EP1873259B1 (en) 2000-12-01 2012-01-25 Max-Planck-Gesellschaft zur Förderung der Wissenschaften e.V. RNA interference mediated by 21 and 22nt RNAs
US20020150945A1 (en) 2000-12-28 2002-10-17 Cell Therapeutics, Inc. Methods for making polynucleotide libraries, polynucleotide arrays, and cell libraries for high-throughput genomics analysis
JP2004532616A (en) 2000-12-28 2004-10-28 ジョンソン・アンド・ジョンソン・リサーチ・ピー・ティー・ワイ・リミテッド Double-stranded RNA-mediated gene suppression
US20030143597A1 (en) 2000-12-28 2003-07-31 Finney Robert E. Methods for making polynucleotide libraries, polynucleotide arrays, and cell libraries for high-throughput genomics analysis
EP1353676A4 (en) 2000-12-29 2006-05-31 Alteon Inc Method for treating fibrotic diseases or other indications
WO2003035869A1 (en) 2001-10-26 2003-05-01 Ribopharma Ag Use of a double-stranded ribonucleic acid for specifically inhibiting the expression of a given target gene
KR100489509B1 (en) * 2001-03-22 2005-05-16 마쯔시다덴기산교 가부시키가이샤 Positive-electrode active material and nonaqueous-electrolyte secondary battery containing the same
WO2002085285A2 (en) 2001-04-18 2002-10-31 Wyeth Methods and reagents for regulating bone and cartilage formation
WO2002085308A2 (en) 2001-04-24 2002-10-31 Epigenesis Pharmaceuticals, Inc. Antisense and anti-inflammatory based compositions to treat respiratory disorders
WO2002085309A2 (en) 2001-04-24 2002-10-31 Epigenesis Pharmaceuticals, Inc. Composition, formulations & kits for treatment of respiratory & lung disease with anti-sense oligonucleotides & a bronchodilating agent
US20050239731A1 (en) 2001-05-18 2005-10-27 Sirna Therapeutics, Inc. RNA interference mediated inhibition of MAP kinase gene expression using short interfering nucleic acid (siNA)
US20030124513A1 (en) 2001-05-29 2003-07-03 Mcswiggen James Enzymatic nucleic acid treatment of diseases or conditions related to levels of HIV
US20050176025A1 (en) 2001-05-18 2005-08-11 Sirna Therapeutics, Inc. RNA interference mediated inhibition of B-cell CLL/Lymphoma-2 (BCL-2) gene expression using short interfering nucleic acid (siNA)
US20050148530A1 (en) 2002-02-20 2005-07-07 Sirna Therapeutics, Inc. RNA interference mediated inhibition of vascular endothelial growth factor and vascular endothelial growth factor receptor gene expression using short interfering nucleic acid (siNA)
WO2003070897A2 (en) 2002-02-20 2003-08-28 Ribozyme Pharmaceuticals, Incorporated RNA INTERFERENCE MEDIATED INHIBITION OF TNF AND TNF RECEPTOR SUPERFAMILY GENE EXPRESSION USING SHORT INTERFERING NUCLEIC ACID (siNA)
US20050130181A1 (en) 2001-05-18 2005-06-16 Sirna Therapeutics, Inc. RNA interference mediated inhibition of wingless gene expression using short interfering nucleic acid (siNA)
US20050227935A1 (en) 2001-05-18 2005-10-13 Sirna Therapeutics, Inc. RNA interference mediated inhibition of TNF and TNF receptor gene expression using short interfering nucleic acid (siNA)
WO2003070910A2 (en) 2002-02-20 2003-08-28 Ribozyme Pharmaceuticals, Incorporated INHIBITION OF VASCULAR ENDOTHELIAL GROWTH FACTOR (VEGF) AND VEGF RECEPTOR GENE EXPRESSION USING SHORT INTERFERING NUCLEIC ACID (siNA)
US20050048529A1 (en) 2002-02-20 2005-03-03 Sirna Therapeutics, Inc. RNA interference mediated inhibition of intercellular adhesion molecule (ICAM) gene expression using short interfering nucleic acid (siNA)
US20040219671A1 (en) 2002-02-20 2004-11-04 Sirna Therapeutics, Inc. RNA interference mediated treatment of parkinson disease using short interfering nucleic acid (siNA)
AU2002344237B8 (en) 2001-05-29 2008-11-06 Sirna Therapeutics, Inc. Nucleic Acid Based Modulation of Female Reproductive Diseases and Conditions
ATE479748T1 (en) * 2001-05-29 2010-09-15 Human Cell Systems Inc TESTICULAR CARNITINE TRANSPORTER AND ITS GENE
WO2003006424A1 (en) 2001-07-10 2003-01-23 4Sc Ag Novel compounds as anti-inflammatory, immunomodulatory and anti-proliferatory agents
US6734017B2 (en) * 2001-09-28 2004-05-11 Isis Pharmaceuticals, Inc. Antisense modulation of vascular endothelial growth factor receptor-2 expression
WO2003035870A1 (en) 2001-10-26 2003-05-01 Ribopharma Ag Drug for treating a carcinoma of the pancreas
RU2297451C2 (en) * 2001-10-30 2007-04-20 Дайити Фармасьютикал Ко., Лтд. Method for propagation of hematopoietic stem cells
EP1575976A4 (en) 2001-11-02 2006-08-23 Insert Therapeutics Inc Methods and compositions for therapeutic use of rna interference
US20040063654A1 (en) 2001-11-02 2004-04-01 Davis Mark E. Methods and compositions for therapeutic use of RNA interference
FR2832154B1 (en) 2001-11-09 2007-03-16 Centre Nat Rech Scient OLIGONUCLEOTIDES INHIBITORS AND THEIR USE FOR SPECIFICALLY REPRESSING A GENE
US20030186903A1 (en) 2001-11-23 2003-10-02 Isis Pharmaceuticals Inc. Antisense modulation of MyD88 expression
EP2014674B1 (en) 2001-11-26 2014-02-12 Laboratoire Biodim Protein-protein interactions in human immunodeficiency virus
US6965025B2 (en) 2001-12-10 2005-11-15 Isis Pharmaceuticals, Inc. Antisense modulation of connective tissue growth factor expression
KR100441894B1 (en) * 2002-01-26 2004-07-27 한국전자통신연구원 Micro-integrated near-field optical recording head and optical recording system using the same
ATE508188T1 (en) * 2002-02-01 2011-05-15 Life Technologies Corp OLIGONUCLEOTIDE COMPOSITIONS WITH IMPROVED EFFECTIVENESS
EP1442143A4 (en) 2002-02-20 2005-02-16 Sirna Therapeutics Inc RNA INTERFERENCE MEDIATED INHIBITION OF BCL2 GENE EXPRESSION USING SHORT INTERFERING NUCLEIC ACID (siNA)
AU2003207708A1 (en) 2002-02-20 2003-09-09 Sirna Therapeutics, Inc. Rna interference mediated inhibition of map kinase genes
JP2005521393A (en) 2002-03-20 2005-07-21 マサチューセッツ インスティテュート オブ テクノロジー HIV treatment
KR20050002917A (en) 2002-03-29 2005-01-10 윤 엔 Human ribonucleotide reductase m2 subunit
US20030224512A1 (en) 2002-05-31 2003-12-04 Isis Pharmaceuticals Inc. Antisense modulation of beta-site APP-cleaving enzyme expression
WO2003094789A1 (en) 2002-05-13 2003-11-20 Salviac Limited Retrieval catheter for an embolic filter
US20040101857A1 (en) 2002-11-23 2004-05-27 Isis Pharmaceuticals Inc. Modulation of cytokine-inducible kinase expression
AU2003239897A1 (en) 2002-05-23 2003-12-12 Ceptyr, Inc. Modulation of ptp1b signal transduction by rna interference
US20040102391A1 (en) 2002-11-21 2004-05-27 Isis Pharmaceuticals Inc. Modulation of Gankyrin expression
JP2005529616A (en) 2002-06-18 2005-10-06 アイアールエム エルエルシー Diagnosis and treatment of chemotherapy-resistant tumors
US7148342B2 (en) 2002-07-24 2006-12-12 The Trustees Of The University Of Pennyslvania Compositions and methods for sirna inhibition of angiogenesis
US6906186B1 (en) 2002-07-30 2005-06-14 Isis Pharmaceuticals, Inc. Antisense modulation of polo-like kinase expression
GB0218010D0 (en) * 2002-08-05 2002-09-11 Ciba Spec Chem Water Treat Ltd Production of a fermentation product
US20040029275A1 (en) 2002-08-10 2004-02-12 David Brown Methods and compositions for reducing target gene expression using cocktails of siRNAs or constructs expressing siRNAs
WO2004027030A2 (en) 2002-09-18 2004-04-01 Isis Pharmaceuticals, Inc. Efficient reduction of target rna’s by single- and double-stranded oligomeric compounds
WO2004031237A1 (en) 2002-09-30 2004-04-15 Oncotherapy Science, Inc. Genes and polypeptides relating to human myeloid leukemia
US20070254850A1 (en) 2002-10-30 2007-11-01 Judy Lieberman Methods for Treating and Preventing Apoptosis-Related Diseases Using Rna Interfering Agents
NZ540779A (en) 2002-11-01 2008-05-30 Univ Pennsylvania Compositions and methods for siRNA inhibition of HIF-1 alpha
ES2334125T3 (en) 2002-11-04 2010-03-05 University Of Massachusetts SPECIFIC ARN INTERFERENCE OF ALELOS.
US7619081B2 (en) 2002-11-14 2009-11-17 Dharmacon, Inc. siRNA targeting coatomer protein complex, subunit beta 2 (COPB2)
US7977471B2 (en) 2002-11-14 2011-07-12 Dharmacon, Inc. siRNA targeting TNFα
US7612196B2 (en) 2002-11-14 2009-11-03 Dharmacon, Inc. siRNA targeting cyclin-dependent kinase inhibitor 1B (p27, Kip1) (CDKN1B)
US7250496B2 (en) 2002-11-14 2007-07-31 Rosetta Genomics Ltd. Bioinformatically detectable group of novel regulatory genes and uses thereof
PT2284266E (en) 2002-11-14 2013-12-17 Thermo Fisher Scient Biosciences Inc Sirna targeting tp53
US7635770B2 (en) 2002-11-14 2009-12-22 Dharmacon, Inc. siRNA targeting protein kinase N-3 (PKN-3)
US7691998B2 (en) 2002-11-14 2010-04-06 Dharmacon, Inc. siRNA targeting nucleoporin 62kDa (Nup62)
US7951935B2 (en) 2002-11-14 2011-05-31 Dharmacon, Inc. siRNA targeting v-myc myelocytomatosis viral oncogene homolog (MYC)
US7582747B2 (en) 2002-11-14 2009-09-01 Dharmacon, Inc. siRNA targeting inner centromere protein antigens (INCENP)
US7781575B2 (en) 2002-11-14 2010-08-24 Dharmacon, Inc. siRNA targeting tumor protein 53 (p53)
WO2006006948A2 (en) 2002-11-14 2006-01-19 Dharmacon, Inc. METHODS AND COMPOSITIONS FOR SELECTING siRNA OF IMPROVED FUNCTIONALITY
US7592442B2 (en) 2002-11-14 2009-09-22 Dharmacon, Inc. siRNA targeting ribonucleotide reductase M2 polypeptide (RRM2 or RNR-R2)
AU2003295539A1 (en) 2002-11-15 2004-06-15 University Of Massachusetts Allele-targeted rna interference
US7217807B2 (en) 2002-11-26 2007-05-15 Rosetta Genomics Ltd Bioinformatically detectable group of novel HIV regulatory genes and uses thereof
WO2004052920A2 (en) 2002-12-12 2004-06-24 The Burnham Institute Conversion of apoptotic proteins
US20040248299A1 (en) 2002-12-27 2004-12-09 Sumedha Jayasena RNA interference
DE602004015064D1 (en) * 2003-01-06 2008-08-28 Wyeth Corp COMPOSITIONS AND METHODS FOR DIAGNOSIS AND TREATMENT OF COLONY CANCER
FR2835837B1 (en) 2003-02-06 2007-03-16 Centre Nat Rech Scient INHIBITORY OLIGONUCLEOTIDES AND THEIR USE FOR SPECIFICALLY REPRESSING A GENE ENCODING A GROWTH FACTOR
FR2835838B1 (en) 2003-02-06 2007-11-16 Centre Nat Rech Scient OLIGONUCLEOTIDES INHIBITORS AND THEIR USE FOR SPECIFICALLY REPRESSING A GENE ENCODING A TRANSCRIPTION FACTOR
AU2004210986A1 (en) * 2003-02-11 2004-08-26 Wyeth Methods for monitoring drug activities in vivo
WO2004076639A2 (en) 2003-02-26 2004-09-10 Wyeth Use of gene expression profiling in the diagnosis and treatment of lupus nephritis and systemic lupus erythematosus
EP3450559A1 (en) * 2003-03-07 2019-03-06 Alnylam Pharmaceuticals, Inc. Therapeutic compositions
JP4605799B2 (en) 2003-04-02 2011-01-05 ダーマコン， インコーポレイテッド Modified polynucleotides for use in RNA interference
AU2003224132A1 (en) 2003-04-24 2004-11-19 Galapagos Genomics N.V. Effective sirna knock-down constructs
US7399853B2 (en) 2003-04-28 2008-07-15 Isis Pharmaceuticals Modulation of glucagon receptor expression
CA2527301A1 (en) 2003-05-30 2004-12-09 Nippon Shinyaku Co., Ltd. Oligonucleic acid-bearing composite and pharmaceutical composition containing the composite
DK1633767T3 (en) 2003-06-02 2019-03-25 Univ Massachusetts METHODS AND COMPOSITIONS FOR MANAGING THE EFFECT OF RNA SILENCING
US7750144B2 (en) 2003-06-02 2010-07-06 University Of Massachusetts Methods and compositions for enhancing the efficacy and specificity of RNA silencing
ES2357116T5 (en) 2003-06-02 2021-09-03 Univ Massachusetts Methods and compositions to improve the efficacy and specificity of FNAi
EP3604537B1 (en) 2003-06-13 2021-12-08 Alnylam Europe AG Double-stranded ribonucleic acid with increased effectiveness in an organism
US7825235B2 (en) 2003-08-18 2010-11-02 Isis Pharmaceuticals, Inc. Modulation of diacylglycerol acyltransferase 2 expression
DE10341333B4 (en) * 2003-09-08 2006-06-08 Siemens Ag Piezoelectric actuator and method for producing a piezoelectric actuator
CN1886793A (en) 2003-11-27 2006-12-27 皇家飞利浦电子股份有限公司 Method and system for chapter marker and title boundary insertion in dv video
KR100586654B1 (en) * 2003-12-19 2006-06-07 이처닷컴 주식회사 Wireless banking system and wireless banking method using mobile phone
WO2005062937A2 (en) 2003-12-22 2005-07-14 University Of Massachusetts Methods and compositions for enhancing the efficacy and specificity of single and double blunt-ended sirna
JP2005233933A (en) * 2004-01-19 2005-09-02 Nec Electronics Corp Combination test method and testing device
KR101054402B1 (en) * 2004-02-06 2011-08-04 엘지전자 주식회사 Pulley assembly of washing machine
US20050176045A1 (en) 2004-02-06 2005-08-11 Dharmacon, Inc. SNP discriminatory siRNA
AU2005222902B2 (en) 2004-03-12 2010-06-10 Alnylam Pharmaceuticals, Inc. iRNA agents targeting VEGF
KR101147147B1 (en) 2004-04-01 2012-05-25 머크 샤프 앤드 돔 코포레이션 Modified polynucleotides for reducing off-target effects in rna interference
CA2557532A1 (en) 2004-04-23 2005-11-10 Angela M. Christiano Inhibition of hairless protein mrna
WO2005117991A2 (en) 2004-05-04 2005-12-15 Nastech Pharmaceutical Company Inc. Compositions and methods for enhancing delivery of nucleic acids into cells and for modifying expression of target genes in cells
AU2005248147A1 (en) 2004-05-11 2005-12-08 Alphagen Co., Ltd. Polynucleotides for causing RNA interference and method for inhibiting gene expression using the same
US7605250B2 (en) 2004-05-12 2009-10-20 Dharmacon, Inc. siRNA targeting cAMP-specific phosphodiesterase 4D
MX2007003795A (en) * 2004-09-28 2007-07-11 Quark Biotech Inc Oligoribonucleotides and methods of use thereof for treatment of alopecia, acute renal failure and other diseases.
AU2005306533B2 (en) * 2004-11-17 2012-05-31 Arbutus Biopharma Corporation siRNA silencing of apolipoprotein B
US7879992B2 (en) * 2005-01-31 2011-02-01 Isis Pharmaceuticals, Inc. Modification of MyD88 splicing using modified oligonucleotides
US7150451B2 (en) * 2005-02-18 2006-12-19 Gm Global Technology Operations, Inc. Air spring and jounce shock assembly
CN101277704A (en) 2005-04-12 2008-10-01 因特拉迪格姆公司 Composition and methods of rnai therapeutics for treatment of cancer and other neovascularization diseases
US20060286575A1 (en) 2005-04-16 2006-12-21 Cylene Pharmaceuticals, Inc. MCL-1 quadruplex nucleic acids and uses thereof
US8268629B2 (en) * 2005-06-21 2012-09-18 dTEC Systems L.L.C. Method for the measurement of water and water-soluble components in non-aqueous liquids
EP1772274B2 (en) 2005-09-29 2011-05-25 Brother Kogyo Kabushiki Kaisha Ink cartridge, set of ink cartridges and ink jet recording system
US7825099B2 (en) 2006-01-20 2010-11-02 Quark Pharmaceuticals, Inc. Treatment or prevention of oto-pathologies by inhibition of pro-apoptotic genes
US20070259827A1 (en) 2006-01-25 2007-11-08 University Of Massachusetts Compositions and methods for enhancing discriminatory RNA interference
US7910566B2 (en) * 2006-03-09 2011-03-22 Quark Pharmaceuticals Inc. Prevention and treatment of acute renal failure and other kidney diseases by inhibition of p53 by siRNA
AU2007253776B2 (en) 2006-05-19 2012-04-26 Arrowhead Research Corporation RNAi-mediated inhibition of tumor necrosis factor alpha-related conditions
US8523707B2 (en) * 2006-05-31 2013-09-03 Bridgestone Sports Co., Ltd. Multi-piece solid golf ball
US8138160B2 (en) 2006-08-03 2012-03-20 Warsaw Orthopedic, Inc. Reagents, methods and systems to suppress pro-inflammatory cytokines
MX2009001896A (en) * 2006-08-24 2009-04-17 Alcon Res Ltd Rnai-mediated inhibition of gremlin for treatment of iop-related conditions.
JP2010507387A (en) * 2006-10-25 2010-03-11 クアーク・ファーマスーティカルス、インコーポレイテッド Novel siRNA and method of using the same
US8785408B2 (en) 2007-06-27 2014-07-22 Quark Pharmaceuticals, Inc. Compositions and methods for reducing or protecting against delayed graft function (DGF)
US7572747B2 (en) * 2007-09-27 2009-08-11 Alfred University Optical glass
BRPI0817605A2 (en) 2007-10-03 2017-05-09 Quark Pharmaceuticals Inc new siren structures
CN101640467B (en) 2008-07-28 2012-05-30 鸿富锦精密工业（深圳）有限公司 Shrapnel and voice coil motor

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2013184524A2 (en) * 2012-06-04 2013-12-12 Academia Sinica Usp37 inactivation as a treatment for plzf/rara-associated acute promyelocytic leukemia
WO2013184524A3 (en) * 2012-06-04 2014-03-20 Academia Sinica Usp37 inactivation as a treatment for plzf/rara-associated acute promyelocytic leukemia
US10125369B2 (en) 2012-12-05 2018-11-13 Alnylam Pharmaceuticals, Inc. PCSK9 iRNA compositions and methods of use thereof
US20150344887A1 (en) * 2014-05-29 2015-12-03 Korea Institute Of Science And Technology siRNA FOR INHIBITION OF USP15 EXPRESSION AND PHARMACEUTICAL COMPOSITION CONTAINING THE SAME
US10851377B2 (en) 2015-08-25 2020-12-01 Alnylam Pharmaceuticals, Inc. Methods and compositions for treating a proprotein convertase subtilisin kexin (PCSK9) gene-associated disorder

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