WO2011084485A1 - Isolement d'arnm activement traduits d'un petit nombre de cellules - Google Patents

Isolement d'arnm activement traduits d'un petit nombre de cellules Download PDF

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WO2011084485A1
WO2011084485A1 PCT/US2010/060512 US2010060512W WO2011084485A1 WO 2011084485 A1 WO2011084485 A1 WO 2011084485A1 US 2010060512 W US2010060512 W US 2010060512W WO 2011084485 A1 WO2011084485 A1 WO 2011084485A1
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mrna
homo sapiens
chaperone
protein
cells
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Jingfang Ju
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The Research Foundation Of State University Of New York
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/1003—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6806—Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay

Definitions

  • the invention relates to methods for the isolation of actively translated mRNAs from a small number of cells and kits for performing the methods.
  • isolation of polysomes remains a major bottleneck for the investigation of post transcriptional regulation particularly where large numbers of cells are not available (e.g., cell cultures, serum, plasma, lavage, saliva, urine, amniotic fluid, microaspirates, microbiopsies, laser-captured cells, formalin fixed paraffin embedded tissue, CSF, lymphatic fluid, rodent tissues, primate tissues, cultured stem cells, oocytes, sperm, etc.).
  • cells e.g., cell cultures, serum, plasma, lavage, saliva, urine, amniotic fluid, microaspirates, microbiopsies, laser-captured cells, formalin fixed paraffin embedded tissue, CSF, lymphatic fluid, rodent tissues, primate tissues, cultured stem cells, oocytes, sperm, etc.
  • RNAs are associated with multi-subunit ribosomes (polyribosomes or polysomes).
  • MicroRNAs and other non-coding RNAs have also been isolated from polysomes (Brandhorst, B., Isolating DNA, RNA, polysomes and protein. Methods Cell Biol. 2004;74:579-99; Arava, Y., Isolation of polysomal RNA for microarray analysis. Methods Mol Biol. 2003;224:79-87).
  • Chaperones are proteins that assist in the folding and assembly of
  • Chaperones are an essential component of and always found in polysomes (Chen, M.S., Characterization of an hsp70 related clone encoding a 33kDa protein with homology to a protein which associates with polysomes. Biochim. Biophys. Acta. 1996 Oct 17; 1297: 124-6; Ohashi, S. et al., HSP60 interacts with YB-1 and affects its polysome association and subcellular localization. Biochem. Biophys. Res. Commun. 2009 Aug 7;385:545-50).
  • HSPs heat shock proteins
  • HSP60, HSP70 and HSP90 are about 60, 70, and 90 kiloDaltons, respectively.
  • these designations generally represent a family of HSPs. For example, the majority of genomes code for multiple members of the Hsp70 molecular chaperone family, with higher organisms possessing more Hsp70s. In humans, there are 13 known HSP70 proteins, plus four related HSP110 proteins.
  • the present invention provides the capability to isolate mRNAs (and other RNAs), that are associated specifically with polysomes, from limited amounts of biological tissues and fluids.
  • the present method allows the specific isolation from limited source material of polysomal complexes containing actively translated RNA.
  • the invention features a method of isolating an actively translated RNA from a biological sample having the step of contacting a biological sample with a chaperone binding molecule.
  • Actively translated RNAs include, but are not limited to, mRNA, microRNAs, siRNAs.
  • the chaperone binding molecule is an antibody.
  • the chaperone-binding molecule selectively binds a chaperone protein selected from the group consisting of a HSP40 family chaperone protein, a HSP70 family chaperone protein, a HSP90 family chaperone protein, and combinations thereof.
  • the chaperone-binding molecule selectively binds a HSP70 family chaperone protein. In another embodiment the chaperone- binding molecule selectively binds HSC70 or HSP73.
  • the invention features a method of isolating actively translated R A from a biological sample comprising contacting the biological sample with a chaperone binding molecule to form a chaperone binding molecule - chaperone complex, separating the chaperone binding molecule - chaperone complex from the sample, and isolating the actively translated m NA from the chaperone binding molecule - chaperone complex.
  • the chaperone binding molecule - chaperone complex is separated from the sample by binding chaperone-binding molecule to a solid support.
  • the invention includes methods for isolating microRNA and/or other non-coding RNAs associated with actively-transcribed mRNAs in a biological sample.
  • a chaperone binding molecule is mixed with a biological sample, a chaperone binding molecule - chaperone complex is formed, the chaperone binding molecule - chaperone complex is separated from the sample, then microRNA and other non-coding RNAs are purified from the complexes.
  • the antibody selectively binds a chaperone protein selected from the group consisting of a HSP40 family chaperone protein, a HSP70 family chaperone protein, a HSP90 family chaperone protein, a HSP1 10 family chaperone protein, and combinations thereof.
  • a chaperone protein selected from the group consisting of a HSP40 family chaperone protein, a HSP70 family chaperone protein, a HSP90 family chaperone protein, a HSP1 10 family chaperone protein, and combinations thereof.
  • the chapreone-binding molecule selectively binds a HSP70 family chaperone protein.
  • the chapreone-binding molecule selectively binds HSC70/HSP73.
  • the method is used to isolate actively translated RNA from samples containing any number of cells, and is particularly advantageous where the number of cells is small (e.g., 5000 cells or fewer.
  • the biological sample contains less than about 5000 cells.
  • the biological sample contains less that 500 cells.
  • the biological sample contains less than 50, or less than 20, or less than 5 cells.
  • the sample contains one cell.
  • the biological sample contains from 500 cells to 5000 cells. In another embodiment, the biological sample contains from 50 to 500 cells. In another embodiment, the biological sample contains from 20 cells to 50 cells. In yet another embodiment, the biological sample contains from 5 cells to 20 cells. In still another embodiment, the biological sample contains from 1 cell to 5 cells.
  • the invention features a method for analysis of actively translated mRNA from a biological sample comprising contacting the biological sample with an chaperone binding molecule that binds to a chaperone protein to form a chaperone binding molecule - chaperone complex, isolating the antibody - chaperone complex, and analyzing the actively translated mR A from the antibody - chaperone complex.
  • analyzing the actively translated mRNA includes amplification of the isolated actively translated mRNA.
  • analyzing the actively translated mRNA includes microarray analysis of the isolated actively translated mRNA.
  • analyzing the actively translated mRNA includes quantitative RT-PCR of the isolated actively translated mRNA.
  • analyzing the actively translated mRNA includes sequencing the RNA. In one such embodiment, deep sequencing is employed to identify somatic mutations in a population of cells or tissue.
  • the antibody selectively binds a chaperone protein selected from the group consisting of a HSP40 family chaperone protein, a HSP70 family chaperone protein, a HSP90 family chaperone protein, a HSP110 family chaperone protein, and combinations thereof. In another embodiment the antibody selectively binds a HSP70 family chaperone protein. In another embodiment the antibody selectively binds a chaperone protein selected from the group consisting of a HSP40 family chaperone protein, a HSP70 family chaperone protein, a HSP90 family chaperone protein, a HSP110 family chaperone protein, and combinations thereof. In another embodiment the antibody selectively binds a HSP70 family chaperone protein. In another embodiment the antibody selectively binds
  • the biological sample contains less than about 5000 cells. In certain embodiments, the biological sample contains less that 500 cells, or less than 50, or less than 20, or less than 5 cells. In one embodiment, the sample contains one cell. In other embodiments, the biological sample contains from 500 cells to 5000 cells, or from 50 to 500 cells, or from 20 cells to 50 cells, or from 5 cells to 20 cells, or from 1 cell to 5 cells.
  • the invention features a kit for isolating actively translated mRNA from a biological sample comprising: (a) one or more chaperone-binding molecules, and (b) instructions for use.
  • the kit further contains a solid support for immobilizing the one or more chaperone-binding molecules.
  • the one or more chaperone-binding molecules are one or more antibodies.
  • the kit comprises one or more reagents suitable for separating a complex comprising an actively translated mRNA and a chaperone protein from a biological sample.
  • suitable lysis solutions and buffers are those that minimize disruption of protein:protein interactions, particularly disruption of cellular complexes comprising RNA and one or more chaperone proteins.
  • the kit includes reagents for working the invention, such as, e.g., PBS and wash buffer.
  • the one or more antibodies selectively bind a chaperone protein selected from the group consisting of a HSP40 family chaperone protein, a HSP70 family chaperone protein, a HSP90 family chaperone protein, a HSP110 family chaperone protein, and combinations thereof.
  • the antibody selectively binds to a HSP70 family chaperone protein.
  • the antibody selectively binds to HSC70/HSP73.
  • FIG. 1 Schematic diagram of TrIP-Chip approach. Affinity beads with covalently attached anti-Hsp70 antibodies are used to immunoprecipitate a cross-linked complex composed of Hsp70, nascent peptides, polysomes and an actively translating mRNA. The mRNA is used to conduct qPCR or converted to a labeled cRNA in a two step reaction, and hybridized to whole genome microarrays. If translation is not taking place, there are no nascent peptides with which Hsp70 can be associated (A); Schematic diagram of antibody affinity bead preparation and polysome isolation procedure (B).
  • FIG. 1 Western immunoblot analysis of in vitro translated TS protein expression isolated using hsp70/hsc70 antibody affinity capture beads (lane 2); unrelated ⁇ - tubulin antibody beads were used as negative control (lane 1) (A).
  • FIG. 3 HCT-116 colon cancer cells were treated with 400 ⁇ puromycin for 1 hour, and total RNA was isolated with hsp70 antibody affinity capture beads isolated RNP complex. The amount of total RNA was quantified by O.D. measurement at 260 nm using Nanodrop Spectrophotometer. Concentration of control RNA sample was set at 100% compared to puromycin treated sample (A). Real time qRT-PCR analysis of translational initiation factor eIF-4E expression level in both control cells and cells treated with 400 ⁇ puromycin (B).
  • B Western immunoblot analysis of TS and p53 protein expression isolated using hsp70/hsc70s antibody affinity capture beads (C).
  • FIG. 5 Gene Ontology (GO) analysis was performed using the DAVID bioinformatics suite.
  • DAVID Gene Functional Classification algorithm allows a gene list to be condensed into organized classes based on cellular functions. The genes are grouped and clustered based on their cellular and molecular functions which helps to identify functional related genes presented as a fuzzy heat map graphic view. The annotation terms are ordered based on their enrichment scores associated with the group. A bright areas represents a positive association between the gene-term: conversely, black represents an unknown relationship. The scattered pattern shows the functional difference. More than 20 clusters were represented with unique functions such as protein synthesis, cell cycle control, and RNA binding (A). A zoomed in view of cluster one was represented as genes involved in translational initiation (B). The distribution of each set of classified genes is presented in Table 3.
  • FIG. Western immunoblot analysis of PP2A protein expression isolated using hsp70/hsc70 antibody affinity capture beads from control and 5-FU treated HCT116 colon cancer cells (lane 1, control; lane 2, plus 5-FU) (A).
  • Chaperones are proteins that assist in the folding and assembly of
  • chaperone proteins One of the major functions of chaperone proteins is to prevent newly synthesized polypeptide chains from misfolding or mis-associating with other cellular molecules. Proteins may not be able to fold fully until their polypeptide chain and/or other subunits are fully synthesized.
  • the Hsp70 family of heat shock proteins contains multiple homologs ranging in size from 66-78 kDa, and are the eukaryotic equivalents of the bacterial DnaK.
  • the most studied Hsp70 members include the cytosolic stress-induced Hsp70 (Hsp72), the constitutive cytosolic Hsc70 (Hsp73), and the ER-localized BiP (Grp78).
  • Hsp70 family members contain highly conserved N-terminal ATP-ase and C-terminal protein binding domains.
  • the chaperone protein is Hsc70 (Hsp73).
  • Chaperone proteins of the present invention include any chaperone that associates directly or indirectly with a newly translated polypeptide chain including, but not limited to, members of the HSP70, HSP40, HSP90, and HSPl 10 families.
  • the Hsp40 family of heat-shock proteins contain a domain by which they interact with Hsp70s.
  • FIG. 1 A One embodiment of the actively translated R A isolation process is illustrated in Fig. 1 A.
  • Translationally active mRNAs are associated with multiple units of ribosomes and the newly synthesized polypeptides are closely associated with molecular chaperones, such as hsp70s.
  • molecular chaperones which assist in the proper folding of nascent polypeptides into higher ordered structures, provide an anchor for separating translationally active mRNAs associated with polysomes from free mRNAs.
  • Affinity antibody capture beads will capture hsp70s chaperones associated with the polysome complexes so that polysomes can be separated from monosomes and free mRNAs.
  • the isolated translationally active mRNAs can then be used for array based or real time qRT-PCR gene expression analysis.
  • Biological samples may be from any source of cells including samples from a patient and cultured cells, such as fibroblasts, white blood cells, sperm, oocytes, and stem cells.
  • sources of cells include, but are not limited to an aspirate, a microaspirate, a biopsy including an aspiration biopsy or a microbiopsy, a frozen biopsy sample, a lavage, and circulating cancer cells in body fluid or bone marrow.
  • Other sources of cells include a body tissue such as skin, bone marrow, liver, lung, brain, or other body tissue, and body fluids including plasma, serum, blood, amniotic fluid, lymphatic fluid, cerebrospinal fluid, saliva, or urine.
  • the cells can be from any organism, such as a mammal, including but not limited to a human, a primate, a rat, a mouse, or other rodent.
  • the cells are dissected from tissue, such as by microdissection, including laser- capture microdissection.
  • the cells can be fixed cells, including, but not limited to formalin fixed paraffin embedded tissue.
  • the method of the invention may include contacting the biological sample with a crosslinking agent.
  • Crosslinking includes covalently linking macromolecules in a complex and may be accomplished by exposure to agents such as UV, heat, formaldehyde, glutaraldehyde, paraformaldehyde, bissulfosuccinimidyl suberate, disuccinimidyl suberate, and dithiobissuccinimidyl propionate (DSP).
  • agents such as UV, heat, formaldehyde, glutaraldehyde, paraformaldehyde, bissulfosuccinimidyl suberate, disuccinimidyl suberate, and dithiobissuccinimidyl propionate (DSP).
  • DSP dithiobissuccinimidyl propionate
  • the cells may be permeablized and/or lysed by methods known in the art.
  • the cells may be contacted with one or more detergents suitable for immunoprecipitation such as, but not limited to, Triton X-100, Triton X-102, and/or Tween- 20.
  • Preferred buffers are those that minimize disruption of protein:protein interactions.
  • Chaperone-binding molecules include any molecule that can selectively bind chaperone proteins.
  • the chaperone binding molecule is an antibody that selectively binds one or more chaperone proteins.
  • the chaperone binding molecule is an aptamer.
  • Chaperone binding molecules used in the present invention may be linked directly or indirectly to a solid support.
  • the solid support may be linked to a secondary antibody, which binds to the chaperone binding molecule.
  • secondary antibodies are known in the art.
  • Solid supports useful for isolation of the chaperone binding molecule - chaperone protein complex include, but are not limited to, multi-well plates, membranes, affinity columns, agarose beads, sepharose beads, and magnetic beads.
  • a chaperone protein (and any associated molecules) may be immobilized by being bound to a chaperone binding molecule, which is directly or indirectly linked to a solid support.
  • a chaperone binding molecule - chaperone complex to a solid support does not require permanent attachment, but means that the binding is sufficient to allow separation and/or detection of the bound complex.
  • the present invention provides antibodies, fragments, or variants of antibodies that recognize an epitope in a chaperone protein.
  • epitope refers to an antigenic determinant on an antigen to which an antibody binds.
  • Epitopes usually consist of chemically active surface groupings of molecules such as amino acids or sugar side chains, and typically have specific three- dimensional structural characteristics, as well as specific charge characteristics. Epitopes generally have at least five contiguous amino acids.
  • antibody and “antibodies” include polyclonal antibodies, monoclonal antibodies, humanized or chimeric antibodies, and antigen binding fragments of antibodies, including, but not limited to, single chain Fv antibody fragments, Fab fragments, and F(ab') 2 fragments.
  • Polyclonal antibodies are heterogeneous populations of antibody molecules that are specific for a particular antigen, while monoclonal antibodies are homogeneous populations of antibodies to a particular epitope contained within an antigen.
  • Chaperone-binding molecules include any synthetic and naturally occurring molecules that bind specifically to a chaperone protein. Examples include proteins, fragments thereof, polypeptides, polynucleotides, and synthetic analogs. Because the binding substances are simply a source of "binding sites," there is no requirement that the sequences be derived from a natural source, or that the structures resemble any naturally occurring protein. Synthetic analogs include, for instance, polypeptide- and polynucleotide-like polymers having modified backbones, side chains, or bases.
  • Antibodies can be obtained by any technique, for example, from naturally occurring antibodies, or Fab or scFv phage display libraries. Antibodies of the invention can be obtained by standard hybridoma technology (Harlow & Lane, ed., Antibodies: A
  • Antibody fragments that have specific binding affinity for the polypeptide of interest can be generated by known techniques. Such antibody fragments include, but are not limited to, F(ab') 2 fragments that can be produced by pepsin digestion of an antibody molecule, and Fab fragments that can be generated by reducing the disulfide bridges of F(ab') 2 fragments. Alternatively, Fab expression libraries can be constructed. See, for example, Huse et al. (1989) Science 246: 1275-1281. Single chain Fv antibody fragments are formed by linking the heavy and light chain fragments of the Fv region via an amino acid bridge (e.g., 15 to 18 amino acids), resulting in a single chain polypeptide.
  • an amino acid bridge e.g. 15 to 18 amino acids
  • Single chain Fv antibody fragments can be produced through standard techniques, such as those disclosed in U.S. Pat. No. 4,946,778.
  • Antibody fragments further include single domain antibodies, also known as nanobodies. These fragments were developed when it was discovered that camels and llamas possess fully functional antibodies that lack light chains, and thus comprise only one binding domain. It is understood that, to make a single domain antibody from an antibody comprising a VH and a VL domain, certain amino acid substitutions outside the CDRs can be desired to enhance binding, expression or solubility. For example, it can be desirable to modify amino acid residues that would otherwise be buried in the VH-VL interface.
  • antibodies or fragments thereof can be tested for recognition of the target polypeptide by standard immunoassay methods including, for example, enzyme- linked immunosorbent assay (ELISA) or radioimmunoassay assay (RIA). See, Short
  • Useful aptamers include nucleic acid aptamers that have been engineered through repeated rounds of selection in vitro to bind to a molecular target. Aptamers provide binding characteistics similar to antibodies, can be engineered completely in a test tube, and are readily produced by chemical synthesis. Peptide aptamers are also useful as chaperone- binding molecules. Peptide aptamers typically comprise a variable peptide loop of about 10 to 20 amino acids attached at both ends to a protein scaffold. A commonly used scaffold is the bacterial protein Thioredoxin-A.
  • peptides can be generated in numbers several orders of magnitude greater than by conventional one-at-a-time methods by synthesis on polyethylene rods or pins, arranged, for example, in a microtiter plate format.
  • the pin technology is representative of techniques that generate libraries of single compounds in a spatially-differentiated manner.
  • An alternative approach, to rapidly prepare large mixtures of compounds, is the split-pool approach (e.g., Houghten, R.A., 1985, Proc. Natl. Acad. Sci. U.S.A. 82:5131-5135) where a solid support material (e.g., beads) is physically segregated into equal portions for coupling to each of the individual initial reactants.
  • a solid support material e.g., beads
  • peptide libraries are a collection of N-substituted glycines as peptide monomers which are assembled in a modular fashion. (Zuckermann, R.N. et al, 1994, J. Med. Chem. 37:2678-2685.) The structures of the resulting compounds are unique, display unique binding properties, and incorporate the important functionalities of peptides in a novel backbone.
  • Specificity of the chaperone-binding molecules can be determined based on affinity and/or avidity.
  • Affinity represented by the equilibrium constant for the dissociation of an antigen with an antibody (Kd) measures the binding strength between an antigenic determinant and an antibody-binding site.
  • Avidity is a measure of the strength of binding between an antibody with its antigen, which is related to both the affinity between an epitope with its antigen-binding site on the antibody and the valence of the antibody. Valence refers to the number of antigen binding sites of a particular epitope.
  • Antibodies typically bind with a dissociation constant (Kd) of 10 ⁇ 5 to 10 "11 liters/mol. Any Kd less than 10 ⁇ 4 liters/mol is generally considered to indicate nonspecific binding. The lesser the value of the Kd, the stronger the binding strength between an antigenic determinant and the antibody-binding site.
  • the Kd of the chaperone-binding molecule is less than about 10 "8 . In another embodiment, the Kd is less that about 10 "9 . In yet another embodiment, the Kd is less that about 10 "10 . In still another embodiment, Kd is less that about 10 "11 .
  • antibody and antibody fragments includes modifications that retain specificity for a specific antigen.
  • Proteins and non-protein agents may be conjugated to the antibodies by methods that are known in the art. Conjugation methods include direct linkage, linkage via covalently attached linkers, and specific binding pair members (e.g., avidin-biotin). Such methods include, for example, that described by
  • Antibodies and antibody fragments of the present invention further include those for which binding characteristics have been improved by direct mutation, methods of affinity maturation, phage display, or chain shuffling. Affinity and specificity can be modified or improved by mutating CDRs and screening for antigen binding sites having the desired characteristics (see, e.g., Yang et al., J. Mol. Biol, 254: 392-403 (1995)). CDRs are mutated in a variety of ways. One way is to randomize individual residues or combinations of residues so that in a population of otherwise identical antigen binding sites, all twenty amino acids are found at particular positions.
  • mutations are induced over a range of CDR residues by error prone PCR methods (see, e.g., Hawkins et al., J. Mol. Biol, 226: 889-896 (1992)).
  • phage display vectors containing heavy and light chain variable region genes can be propagated in mutator strains of E. coli (see, e.g., Low et al., J. Mol. Biol., 250: 359-368 (1996)).
  • Isolated m NA may be used as a template for amplification using methods known in the art. For example, methods for linear or exponential amplification of the isolated mRNA sequences. Some suitable techniques are described in: Van Gelder et al, (1990) Proc. Natl. Acad. Sci. USA 87, 1663-67; Belyavsky et al, (1989) Nucleic Acids Res. 17, 2919-2932; Tarn et al, (1989) Nucleic Acids Res. 17, 1269; Brady et al, (1989) Meth. Mol. Cell. Biol. 21, 17-2; U.S. Patent Application 6,706,476; U.S.
  • Isolated mRNA may be analyzed by a variety of methods including, but not limited to, quantitative real-time PCR, microarray analysis, and deep sequencing. Suitable techniques are described in: Lipshutz, R.J., Fodor, S.P., Gingeras, T.R.
  • the invention features a kit for isolating actively translated mRNA from a biological sample.
  • the kit is used to perform isolation and purification of actively translated mRNA according to the invention.
  • the kit contains a chaperone -binding molecule, which can be supplied conjugated to a solid support such as a bead or microtiter plate.
  • the chaperone-binding molecule is conjugated to a secondary reagent that can be immobilized and/or detected.
  • the kit also includes instructions for use.
  • the kit may contain one or more additional reagents, including a cross-linking agent, lysis buffer, R ase inhibitor, wash buffer, and a solid support.
  • the methods and kits of the present invention are useful in the identification of genes that are differentially translated in diseased cells or cells of different developmental stages.
  • the methods and kits are useful in detection or identification of genes that are differentially translated in response to external factors such as infectious agents and chemical agents.
  • the methods and kits are useful for prediction, detection and diagnosis of disease.
  • Example 1 Preparation of antibody affinity capture beads.
  • affinity capture beads (chaperone binding molecule linked to a solid support)
  • hsp70/hsc70 antibodies were evaluated to optimize the conditions for binding to the actively translated mR A.
  • HSP70 mouse monoclonal antibody HSC70 (HSP73) rabbit polyclonal antibody (Millipore Inc.); and HSC70 (HSP73) rat monoclonal antibody (Bio Vision Inc.). Beads with antibody were incubated with slow tilt rotation for 24 hours at 2-8°C. The tube was then placed on a magnet for 2 minutes and supernatant was removed with a pipette. Antibody-coated beads were washed three times with washing buffer and used for immunoprecipitation. [0053] Example 2. Translationally regulated genes can be captured by affinity capture beads.
  • TS mRNA expression was determined by real time qRT-PCR analysis and the TS protein was analyzed using Western immunoblot analysis.
  • qRT-PCR Real-time quantitative reverse transcription-PCR analysis was performed on the control mRNAs.
  • PCR primers and probes were purchased from Applied Biosystems (Foster City, CA).
  • qRT-PCR was performed on an ABI 7500HT instrument under the following conditions: 25°C, 10 min and 37°C, 30 min of reverse transcription; up to 45 cycles of PCR at 95°C, 10 min; 95°C, 15 s; 60°C, 1 min per cycle.
  • Example 3 Translationally regulated genes can be captured by affinity capture beads from HCT-116 colon cancer cells.
  • HCT-116 human colon cancer cells were incubated with 400 ⁇ puromycin, a drug that triggers premature polypeptide termination and polysome disassembly.
  • the HCT-116 cell line was obtained from ATCC and maintained at 37°C and 5% CO2 in 75-cm 2 plastic tissue culture flasks with McCoy's 5 A medium containing 10% fetal bovine serum.
  • cytoplasmic extracts To prepare cytoplasmic extracts, cells from 6-well tissue culture plates were harvested with 0.05%> Trypsin-EDTA (Invitrogen, CA) and washed with ice-cold PBS containing 100 ⁇ g/ml cycloheximide (Sigma, MO). Cells were counted and 10,000 cells were incubated with 800 ⁇ 1 of McCoy's 5 A medium containing 10% fetal bovine serum and 100 ⁇ cycloheximide (Sigma, MO) for 5 min at 37°C. After incubation, 200 ⁇ of DSP (PIERCE, IL) was introduced as a cross-linking reagent and incubation was carried out for 5 min at 37°C. The cells were washed twice by centrifugation at 1,000 rpm for 3 min, discarding the supernatant, and rinsing the cell pellets with ice-cold PBS containing
  • cycloheximide 100 ⁇ g/ml cycloheximide (Sigma, MO). The final pellets were swollen for 20 min in 500 ⁇ of low salt buffer (LSB) (20 mM HEPES, pH 7.4, lOOmM KC1 , 2 mM MgCl 2 ) containing 1 mM dithiothreitol and lysed by the addition of 200 ⁇ lysis buffer (1 x LSB containing 1.2% Triton X- 100) (Sigma) followed by brief vortexing. One -tenth (70 ⁇ ) of the above lysate was transferred to the Ig-coated beads, and incubation was carried out for 1-2 hours at 2-8°C.
  • LSB low salt buffer
  • 70 ⁇ 70 ⁇
  • the chaperone complexes containing polysomes and translationally active mRNA transcripts were isolated, and the mRNAs eluted from beads conjugated with HSP70/HSP73 using the Array Pure Nanoscale RNA Purification Kit (EPICENTRE, WI).
  • Example 4 Isolation of translationally active mRNA from cells treated with 5-fluorouracil
  • Translational control plays an important role in chemoresistance, such as acute resistance to 5-fluorouracil (5-FU) treatment.
  • 5-FU 5-fluorouracil
  • Previous studies have identified that the target enzyme of 5-FU, thymidylate synthase (TS), is regulated at the translational level.
  • TS thymidylate synthase
  • TS not only negatively regulates its own synthesis at the translational level, but also interacts directly with p53 mRNA to down-regulate p53 protein expression.
  • HCT- 116 cells at 2x 10 5 density were plated in 6-well tissue culture plates for 24 hrs. Cells were then treated with 5 ⁇ 5-FU for 24 hours; untreated cells were used as controls. Actively translating mRNA was isolated as described in Example 3.
  • Anti-p53 mouse monoclonal antibody was used at a 1 : 1000 dilution and were obtained from Santa Cruz Biotech Inc., CA. [0068] Example 5. Gene expression analysis of 5-FU responsive translationally active genes isolated from polysome complexes in HCT-116 cells.
  • RNA amplification was employed using the Agilent Low RNA Input Linear Amplification Kit (Agilent, CA), which was also used to amplify mRNAs isolated from polysomes.
  • the T7 promoter primer (1.2 ⁇ ) was added and the total reaction volume was adjusted to 11.5 ⁇ with nuclease free water. The primer and template were incubated at 65°C for 10 min and then placed on ice for 5 min.
  • cDNA Master Mix (8.5 ⁇ ) was added to each sample and incubation carried out at 40°C for 2 hours, 65°C for 15 min, and on ice for 5 min. Then 60 ⁇ of amplification reaction buffer was added to 20 ⁇ of cDNA product and incubation proceeded at 40°C for 4 hours. After purification, cRNA products were dried in a Speed Vacuum apparatus and the total volume adjusted to 10.5 ⁇ . Random Hexamers (1.0 ⁇ ) were added to the cRNA products, incubation performed at 65°C for 10 min and the tube chilled to 4°C for 5min. 59 ⁇ of amplification reaction buffer was added to 21 ⁇ of cDNA product and incubation proceeded at 40°C for 4 hours.
  • Hybridization was carried out at 10 rpm at 65°C for 17 hours in a hybridization oven. After hybridization, the arrays were washed with Gene Expression Wash Buffer 1 (Agilent, CA). The slides were then dried and kept in the dark until scanning. Images were captured on an Axon GenePix 4200A scanner.
  • NM_004037 AMPD2 4.639 Homo sapiens adenosine monophosphate deaminase 2
  • transcript variant 1 (isoform L) (AMPD2), transcript variant 1, mRNA
  • NM_001002244 ANAPCl 1 25.86 Homo sapiens APC 11 anaphase promoting complex subunit 11 homolog (yeast) (ANAPCl 1), transcript variant 1, mRNA [NM_001002244]
  • NM_138797 ANKRD54 5.442 Homo sapiens ankyrin repeat domain 54 (ANKRD54), mRNA [NM_138797]
  • NM_001284 AP3S1 32.24 Homo sapiens adaptor-related protein complex 3, sigma 1 subunit (AP3S1), mRNA [NM_001284]
  • NM_001657 AREG 14.87 Homo sapiens amphiregulin (schwannoma-derived growth factor) (AREG), mRNA [NM_001657]
  • NM_001658 ARF1 14.83 Homo sapiens ADP-ribosylation factor 1 (ARF1),
  • transcript variant 4 mRNA [NM_001658]
  • NM_018120 ARMC1 0.17 Homo sapiens armadillo repeat containing 1 (ARMC1), mRNA [NM_018120]
  • NM_004849 ATG5 0.01 Homo sapiens ATG5 autophagy related 5 homolog (S.
  • NM_000701 ATP1A1 34.38 Homo sapiens ATPase, Na+/K+ transporting, alpha 1 polypeptide (ATP1A1), transcript variant 1, mRNA
  • NM_001677 ATP1B1 8.034 Homo sapiens ATPase, Na+/K+ transporting, beta 1 polypeptide (ATP1B1), transcript variant 1, mRNA
  • NM_006886 ATP5E 6.422 Homo sapiens ATP synthase, H+ transporting
  • mitochondrial Fl complex epsilon subunit (ATP5E), nuclear gene encoding mitochondrial protein, mRNA [NM_006886]
  • NM_001002031 ATP5G2 6.354 Homo sapiens ATP synthase, H+ transporting
  • mitochondrial F0 complex subunit C2 (subunit 9) (ATP5G2), nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA [NM_001002031]
  • NM_001002258 ATP5G3 12.48 Homo sapiens ATP synthase, H+ transporting,
  • mitochondrial F0 complex subunit C3 (subunit 9) (ATP5G3), nuclear gene encoding mitochondrial protein, transcript variant 3, mRNA [NM_001002258]
  • NM_006356 ATP5H 31.53 Homo sapiens ATP synthase, H+ transporting,
  • mitochondrial F0 complex subunit d (ATP5H)
  • mitochondrial F0 complex subunit d (ATP5H)
  • nuclear gene encoding mitochondrial protein transcript variant 1
  • transcript variant 1 mRNA [NM_006356]
  • NM_006476 ATP5L 8.456 Homo sapiens ATP synthase, H+ transporting,
  • mitochondrial F0 complex mitochondrial F0 complex
  • subunit G nuclear gene encoding mitochondrial protein
  • NM_001697 ATP50 5.68 Homo sapiens ATP synthase, H+ transporting,
  • mitochondrial F 1 complex O subunit (oligomycin sensitivity conferring protein) (ATP50), nuclear gene encoding mitochondrial protein, mRNA [NM 001697]
  • NM_001694 ATP6V0C 18.3 Homo sapiens ATPase, H+ transporting, lysosomal 16kDa,
  • V0 subunit c (ATP6V0C), mRNA [NM_001694]
  • NM_004231 ATP6V1F 5.796 Homo sapiens ATPase, H+ transporting, lysosomal 14kDa,
  • VI subunit F (ATP6V1F), mRNA [NM_004231]
  • NM_178191 ATPIF1 7.619 Homo sapiens ATPase inhibitory factor 1 (ATPIF1), nuclear gene encoding mitochondrial protein, transcript variant 3, mRNA [NM_178191]
  • ATPIF1 Homo sapiens ATPase inhibitory factor 1
  • NM_003779 B4GALT3 6.752 Homo sapiens UDP-Gal:betaGlcNAc beta 1,4- galactosyltransferase, polypeptide 3 (B4GALT3), mRNA [NM_003779]
  • NM_014417 BBC3 16.84 Homo sapiens BCL2 binding component 3 (BBC3),
  • transcript variant 2 mRNA [NM_001207]
  • NM_006806 BTG3 5.67 Homo sapiens BTG family, member 3 (BTG3), mRNA
  • NM_017589 BTG4 7.105 Homo sapiens B-cell translocation gene 4 (BTG4), mRNA
  • NM_144600 C16orf63 6.026 Homo sapiens chromosome 16 open reading frame 63
  • NM_152350 C17orf45 6.981 Homo sapiens chromosome 17 open reading frame 45
  • NM_012264 C22orf5 17.11 Homo sapiens chromosome 22 open reading frame 5
  • NM_199417 C3orf60 5.886 Homo sapiens chromosome 3 open reading frame 60
  • NM_031897 CACNG6 5.65 Homo sapiens calcium channel, voltage-dependent, gamma subunit 6 (CACNG6), transcript variant 3, mRNA [NM_031897]
  • NM_005831 CALCOC02 7.976 Homo sapiens calcium binding and coiled-coil domain 2
  • calmodulin 2 (phosphorylase kinase, delta)
  • CAMK2N2 10.08 Homo sapiens calcium/calmodulin-dependent protein kinase II inhibitor 2 (CAMK2N2), mRNA [NM_033259]
  • CAMKK2 0.206 Homo sapiens calcium/calmodulin-dependent protein kinase kinase 2, beta (CAMKK2), transcript variant 1, mRNA [NM_006549]
  • NM_001748 CAPN2 6.305 Homo sapiens calpain 2, (m/II) large subunit (CAPN2), mRNA [NM_001748]
  • NM_006135 CAPZA1 12.54 Homo sapiens capping protein (actin filament) muscle Z- line, alpha 1 (CAPZA1), mRNA [NM_006135]
  • NM_024537 CARS2 7.534 Homo sapiens cysteinyl-tRNA synthetase 2, mitochondrial
  • CARS2 (putative) (CARS2), mRNA [NM_024537] NM_016587 CBX3 13.39 Homo sapiens chromobox homolog 3 (HP1 gamma
  • CBX3 Drosophila
  • transcript variant 2 mRNA [NM_016587]
  • NM_012073 CCT5 9.103 Homo sapiens chaperonin containing TCPl, subunit 5
  • CCT7 transcript variant 1, mRNA [NM_006429]
  • CD63 28.53 Homo sapiens CD63 molecule (CD63), transcript variant
  • CD81 7.793 Homo sapiens CD81 molecule (CD81), mRNA
  • CDK2 Homo sapiens cyclin-dependent kinase 2
  • transcript variant 1 mRNA [NM_001798]
  • NM_001800 CDKN2D 4.826 Homo sapiens cyclin-dependent kinase inhibitor 2D (pi, 9 inhibits CDK4) (CDKN2D), transcript variant 1, mRNA [NM_001800]
  • NM_001806 CEBPG 6.633 Homo sapiens CCAAT/enhancer binding protein (C/EBP), gamma (CEBPG), mRNA [NM_001806]
  • NM_014246 0.207 Homo sapiens cadherin, EGF LAG seven-pass G-type receptor 1 (flamingo homolog, Drosophila) (CELSR1), mRNA [NM_014246]
  • NM_016139 CHCHD2 55.82 Homo sapiens coiled-coil-helix-coiled-coil-helix domain containing 2 (CHCHD2), mRNA [NM_016139]
  • NM_016565 CHCHD8 5.027 Homo sapiens coiled-coil-helix-coiled-coil-helix domain containing 8 (CHCHD8), mRNA [NM_016565]
  • NM_007236 CHP 7.844 Homo sapiens calcium binding protein P22 (CHP), mRNA
  • CKAP4 5.756 Homo sapiens cytoskeleton-associated protein 4 (CKAP4), mRNA [NM_006825]
  • NM_001305 CLDN4 5.861 Homo sapiens claudin 4 (CLDN4), mRNA [NM_001305]
  • NM_004859 CLTC 11.95 Homo sapiens clathrin, heavy chain (He) (CLTC), mRNA
  • NM_016057 COPZ1 5.801 Homo sapiens coatomer protein complex, subunit zeta 1
  • NM_006067 COX4NB 17.11 Homo sapiens COX4 neighbor (COX4NB), mRNA
  • NM_004373 COX6A1 20.31 Homo sapiens cytochrome c oxidase subunit Via
  • COX6A1 polypeptide 1
  • NM_004373 nuclear gene encoding mitochondrial protein
  • polypeptide 2 liver
  • COX7A2 mRNA [NM_001865]
  • NM_016190 CR N 0.14 Homo sapiens cornulin (CRNN), mRNA [NM_016190]
  • NM_003798 CTNNAL1 5.217 Homo sapiens catenin (cadherin-associated protein), alphalike 1 (CTNNAL1), mRNA [NM_003798]
  • CYB5R3 15.35 Homo sapiens cytochrome b5 reductase 3 (CYB5R3), transcript variant M, mRNA [NM_000398]
  • CYB5R3 11.23 Homo sapiens cytochrome b5 reductase 3 (CYB5R3), transcript variant M, mRNA [NM_000398]
  • DAP Homo sapiens death-associated protein
  • DAZAP1 Homo sapiens DAZ associated protein 1 (DAZAP1), transcript variant 1, mRNA [NM_170711]
  • DAZAP2 9.077 Homo sapiens DAZ associated protein 2 (DAZAP2), mRNA [NM_014764]
  • DCI (3,2 trans-enoyl-Coenzyme A isomerase) (DCI), nuclear gene encoding mitochondrial protein, mRNA
  • DCP1A cerevisiae
  • NM_018403 mRNA [NM_018403]
  • DCUN1D5 8.175 Homo sapiens DCN1, defective in cullin neddylation 1, domain containing 5 (S. cerevisiae) (DCUN1D5), mRNA [NM_032299]
  • NM_005147 DNAJA3 14.55 Homo sapiens DnaJ (Hsp40) homolog, subfamily A, member 3 (DNAJA3), mRNA [NM_005147]
  • NM_015190 DNAJC9 5.269 Homo sapiens DnaJ (Hsp40) homolog, subfamily C, member 9 (DNAJC9), mRNA [NM_015190]
  • NM_001379 DNMT1 4.616 Homo sapiens DNA (cytosine-5-)-methyltransferase 1
  • NM_012145 DTYMK 6.216 Homo sapiens deoxythymidylate kinase (thymidylate kinase) (DTYMK), mRNA [NM_012145]
  • NM_004417 DUSP1 0.119 Homo sapiens dual specificity phosphatase 1 (DUSP1), mRNA [NM_004417]
  • DUSP5 9.712 Homo sapiens dual specificity phosphatase 5 (DUSP5), mRNA [NM_004419]
  • EBNA1BP2 31.19 Homo sapiens EBNA1 binding protein 2 (EBNA1BP2), mRNA [NM_006824]
  • NM_014329 EDC4 9.509 Homo sapiens enhancer of mRNA decapping 4 (EDC4), mRNA [NM_014329]
  • NM_152991 EED 4.466 Homo sapiens embryonic ectoderm development (EED), transcript variant 2, mRNA [NM_152991]
  • EEF1B2 NM_001959 EEF1B2 117.4 Homo sapiens eukaryotic translation elongation factor 1 beta 2 (EEF1B2), transcript variant 1, mRNA
  • NM_032378 EEF1D 5.24 Homo sapiens eukaryotic translation elongation factor 1 delta (guanine nucleotide exchange protein) (EEF1D), transcript variant 1, mRNA [NM_032378]
  • EIF2S2 5.995 Homo sapiens eukaryotic translation initiation factor 2, subunit 2 beta, 38kDa (EIF2S2), mRNA [NM_003908]
  • NM_003758 EIF3S1 4.192 Homo sapiens eukaryotic translation initiation factor 3, subunit 1 alpha, 35kDa (EIF3S1), mRNA [NM_003758]
  • NM_013234 EIF3S12 7.172 Homo sapiens eukaryotic translation initiation factor 3, subunit 12 (EIF3S12), mRNA [NM_013234]
  • NM_003756 EIF3S3 61.64 Homo sapiens eukaryotic translation initiation factor 3, subunit 3 gamma, 40kDa (EIF3S3), mRNA [NM_003756]
  • EIF3S9 24.08 Homo sapiens eukaryotic translation initiation factor 3, subunit 9 eta, 116kDa (EIF3S9), transcript variant 1, mRNA [NM_003751]
  • EIF4A1 4.515 Homo sapiens eukaryotic translation initiation factor 4A, isoform 1 (EIF4A1), mRNA [NM_001416]
  • NM_006331 EMG1 14.34 Homo sapiens EMG1 nucleolar protein homolog (S.
  • EMG1 cerevisiae
  • NM_001424 EMP2 6.226 Homo sapiens epithelial membrane protein 2 (EMP2), mRNA [NM_001424]
  • NM_001427 EN2 9.828 Homo sapiens engrailed homeobox 2 (EN2), mRNA
  • EP300 8.615 Homo sapiens EIA binding protein p300 (EP300), mRNA
  • EPS8L2 0.01 Homo sapiens EPS8-like 2 (EPS8L2), mRNA
  • ERLIN1 7.225 Homo sapiens ER lipid raft associated 1 (ERLINl), mRNA [NM_006459]
  • NM_004730 ETF1 6.176 Homo sapiens eukaryotic translation termination factor 1
  • NM_000126 ETFA 11.25 Homo sapiens electron-transfer-flavoprotein, alpha
  • NM_152424 FAM123B 19.34 Homo sapiens family with sequence similarity 123B
  • BC064950 FAM134A 5.442 Homo sapiens chromosome 2 open reading frame 17, mRNA (cDNA clone IMAGE:6048607), partial cds. [BC064950]
  • NM_005687 FARSB 5.791 Homo sapiens phenylalanyl-tRNA synthetase, beta subunit
  • FBR-MuSV ubiquitously expressed (fox derived); ribosomal protein S30 (FAU), mRNA [NM_001997]
  • NM_201555 FHL2 50.35 Homo sapiens four and a half LIM domains 2 (FHL2), transcript variant 2, mRNA [NM_201555]
  • NM_032822 FLJ14668 5.612 Homo sapiens hypothetical protein FLJ14668 (FLJ14668), mRNA [NM_032822]
  • NM_024829 FLJ22662 8.429 Homo sapiens hypothetical protein FLJ22662 (FLJ22662), mRNA [NM_024829]
  • NM_173627 FLJ35220 0.108 Homo sapiens hypothetical protein FLJ35220 (FLJ35220), mRNA [NM_173627]
  • NM_001462 FPRL1 0.222 Homo sapiens formyl peptide receptor-like 1 (FPRL1), transcript variant 1, mRNA [NM_001462]
  • FRG1 Homo sapiens FSHD region gene 1 (FRG1), mRNA
  • GABARAPL2 4.457 Homo sapiens GABA(A) receptor-associated protein-like
  • NM_152237 GAS2L1 5.204 Homo sapiens growth arrest-specific 2 like 1 (GAS2L1), transcript variant 3, mRNA [NM_152237]
  • NM_198460 GBP6 16.33 Homo sapiens guanylate binding protein family, member 6
  • NM_006302 GCS1 6.643 Homo sapiens glucosidase I (GCS1), mRNA
  • GCSH aminomethyl carrier
  • NM_001001560 GGA1 5.958 Homo sapiens golgi associated, gamma adaptin ear
  • GGA1 ARF binding protein 1
  • transcript variant 2 mRNA [NM_001001560]
  • NM_014394 GHITM 5.222 Homo sapiens growth hormone inducible transmembrane protein (GHITM), mRNA [NM_014394]
  • NM_016080 GLOD4 8.961 Homo sapiens glyoxalase domain containing 4 (GLOD4), mRNA [NM_016080]
  • GLTSCR2 10.87 Homo sapiens glioma tumor suppressor candidate region gene 2 (GLTSCR2), mRNA [NM_015710]
  • GOLPH2 8.253 Homo sapiens golgi phosphoprotein 2 (GOLPH2)
  • transcript variant 1 mRNA [NM_016548]
  • NM_019858 GPR162 0.131 Homo sapiens G protein-coupled receptor 162 (GPR162), transcript variant A-2, mRNA [NM_019858]
  • nucleotide-gated potassium channel 2 (HCN2)
  • NM_001194 mRNA
  • HEBP1 Homo sapiens heme binding protein 1 (HEBP1), mRNA
  • NM_015382 HECTD1 7.417 Homo sapiens HECT domain containing 1 (HECTD1), mRNA [NM_015382]
  • HERPUD1 5.11 Homo sapiens homocysteine-inducible, endoplasmic reticulum stress-inducible, ubiquitin-like domain member 1 (HERPUD1), transcript variant 1, mRNA [NM_014685]
  • HEBCH hydrolase hydrolase
  • NM_005340 HINT1 14.93 Homo sapiens histidine triad nucleotide binding protein 1
  • NM_033500 HKl 8.496 Homo sapiens hexokinase 1 (HKl), nuclear gene encoding mitochondrial protein, transcript variant 5, mRNA
  • NM_178580 HM13 5.386 Homo sapiens histocompatibility (minor) 13 (HM13), transcript variant 2, mRNA [NM_178580]
  • HMGA2 5.302 Homo sapiens high mobility group AT-hook 2 (HMGA2), transcript variant 1, mRNA [NM_003483]
  • NM_002129 HMGB2 4.671 Homo sapiens high-mobility group box 2 (HMGB2), mRNA [NM_002129]
  • HMHA1 mRNA [NM_012292] ⁇ _001002033 HN1 61.87 Homo sapiens hematological and neurological expressed 1
  • NM_194247 HNRPA3 10.75 Homo sapiens heterogeneous nuclear ribonucleoprotein A3
  • HOXA3 Homo sapiens homeobox A3 (HOXA3), transcript variant
  • HOXB6 Homo sapiens homeobox B6 (HOXB6), mRNA
  • NM_003299 HSP90B1 17.54 Homo sapiens heat shock protein 90kDa beta (Grp94), member 1 (HSP90B1), mRNA [NM_003299]
  • HSPA5 7.297 Homo sapiens heat shock 70kDa protein 5 (glucose- regulated protein, 78kDa) (HSPA5), mRNA [NM_005347]
  • NM_006597 HSPA8 11.35 Homo sapiens heat shock 70kDa protein 8 (HSPA8), transcript variant 1, mRNA [NM_006597]
  • HSPC111 6.091 Homo sapiens hypothetical protein HSPC111 (HSPC111), mRNA [NM_016391]
  • HSPD1 nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA [NM_002156]
  • NM_000871 HTR6 10.07 Homo sapiens 5-hydroxytryptamine (serotonin) receptor 6
  • NM_016400 HYPK 11.26 Homo sapiens Huntingtin interacting protein K (HYPK), mRNA [NM_016400]
  • NM_002165 ID1 18.43 Homo sapiens inhibitor of DNA binding 1 , dominant negative helix-loop-helix protein (ID1), transcript variant 1, mRNA [NM_002165]
  • IER2 Homo sapiens immediate early response 2 (IER2), mRNA
  • NM_016545 IER5 9.981 Homo sapiens immediate early response 5 (IER5), mRNA
  • NM_000597 IGFBP2 4.951 Homo sapiens insulin-like growth factor binding protein 2
  • ribonucleoprotein ribonucleoprotein, homolog (yeast) (IMP3), mRNA
  • J04208 IMPDH2 24.33 Human inosine-5'-monophosphate dehydrogenase (IMP) mRNA, complete cds. [J04208]
  • NM_003870 IQGAP1 5.497 Homo sapiens IQ motif containing GTPase activating protein 1 (IQGAP1), mRNA [NM_003870]
  • NM_194279 ISCA2 10.76 Homo sapiens iron-sulfur cluster assembly 2 homolog (S.
  • NM_014736 KIAAOlOl 9.537 Homo sapiens KIAAOlOl (KIAAOlOl), transcript variant l, mRNA [NM_014736]
  • NM_017641 KIF21A 0.144 Homo sapiens kinesin family member 21 A (KIF21A), mRNA [NM_017641]
  • NM_000421 KRT10 7.118 Homo sapiens keratin 10 (epidermolytic hyperkeratosis;
  • NM_000224 KRT18 8.532 Homo sapiens keratin 18 (KRT18), transcript variant 1, mRNA [NM_000224]
  • NM_005561 LAMP1 9.2 Homo sapiens lysosomal-associated membrane protein 1
  • NM_002296 LBR 22.12 Homo sapiens lamin B receptor (LBR), transcript variant l, mRNA [NM_002296]
  • LMNB2 8.076 Homo sapiens lamin B2 (LMNB2), mRNA [NM_032737]
  • BC062368 LOCI 52217 7.949 Homo sapiens hypothetical protein BC007882, mRNA
  • LSM7 Homo sapiens LSM7 homolog, U6 small nuclear RNA associated (S. cerevisiae) (LSM7), mRNA [NM_016199]
  • NM_001003690 MAD2L1BP 6.979 Homo sapiens MAD2L1 binding protein (MAD2L1BP), transcript variant 1, mRNA [NM_001003690]
  • NM_001017405 MAEA 19.06 Homo sapiens macrophage erythroblast attacher (MAEA), transcript variant 1, mRNA [NM_001017405]
  • NM_002372 MAN2A1 0.212 Homo sapiens mannosidase, alpha, class 2A, member 1
  • NM_152496 MANEAL 5.623 Homo sapiens mannosidase, endo-alpha-like (MANEAL), transcript variant 2, mRNA [NM_152496]
  • NM_002446 MAP3K10 7.627 Homo sapiens mitogen-activated protein kinase kinase kinase 10 (MAP3K10), mRNA [NM_002446]
  • NM_004526 MCM2 4.565 Homo sapiens MCM2 minichromosome maintenance deficient 2, mitotin (S. cerevisiae) (MCM2), mRNA
  • NM_002388 MCM3 4.871 Homo sapiens MCM3 minichromosome maintenance deficient 3 (S. cerevisiae) (MCM3), mRNA [NM_002388]
  • NM_005917 MDH1 15.06 Homo sapiens malate dehydrogenase 1, NAD (soluble)
  • NM_014166 MED4 22.22 Homo sapiens mediator of RNA polymerase II
  • NM_006838 METAP2 20.24 Homo sapiens methionyl aminopeptidase 2 (METAP2), mRNA [NM_006838]
  • transcript variant 1 mRNA
  • NM_203477 MGC70863 5.188 Homo sapiens similar to RPL23AP7 protein (MGC70863), transcript variant 1, mRNA [NM_203477]
  • MIF glycosylation-inhibiting factor
  • NM_014048 MKL2 5.137 Homo sapiens MKL/myocardin-like 2 (MKL2), mRNA
  • leukemia trithorax homolog, Drosophila
  • MLLT11 mRNA [NM_006818]
  • NM_019556 MOSPD1 7.908 Homo sapiens motile sperm domain containing 1
  • MRCL3 4.765 Homo sapiens myosin regulatory light chain MRCL3
  • NM_017840 MRPL16 6.905 Homo sapiens mitochondrial ribosomal protein LI 6
  • MRPL16 nuclear gene encoding mitochondrial protein, mRNA [NM_017840]
  • NM_032112 MRPL43 6.91 Homo sapiens mitochondrial ribosomal protein L43
  • MRPL43 nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA [NM_032112]
  • MRPL46 nuclear gene encoding mitochondrial protein, mRNA [NM_022163]
  • NM_021107 MRPS12 6.026 Homo sapiens mitochondrial ribosomal protein S12
  • MRPS12 nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA [NM_021107]
  • NM_031280 MRPS15 7.642 Homo sapiens mitochondrial ribosomal protein S 15
  • MRPS15 nuclear gene encoding mitochondrial protein, mRNA [NM_031280]
  • MRPS16 nuclear gene encoding mitochondrial protein, mRNA [NM_016065]
  • MRPS18B nuclear gene encoding mitochondrial protein, mRNA [NM_014046]
  • NM_016067 MRPS18C 5.037 Homo sapiens mitochondrial ribosomal protein S 18C
  • MRPS18C nuclear gene encoding mitochondrial protein, mRNA [NM_016067]
  • NM_032014 MRPS24 10.9 Homo sapiens mitochondrial ribosomal protein S24
  • MRPS24 nuclear gene encoding mitochondrial protein, mRNA [NM_032014]
  • NM_021821 MRPS35 8.853 Homo sapiens mitochondrial ribosomal protein S35
  • MRPS35 nuclear gene encoding mitochondrial protein, mRNA [NM_021821]
  • NM_033281 MRPS36 5.685 Homo sapiens mitochondrial ribosomal protein S36
  • MRPS36 nuclear gene encoding mitochondrial protein, mRNA [NM_033281]
  • NM_031902 MRPS5 4.92 Homo sapiens mitochondrial ribosomal protein S5
  • MRPS5 nuclear gene encoding mitochondrial protein, mRNA [NM_031902]
  • NM_182640 MRPS9 4.873 Homo sapiens mitochondrial ribosomal protein S9
  • MRPS9 nuclear gene encoding mitochondrial protein, mRNA [NM_182640]
  • NM_005946 MT1A 4.519 Homo sapiens metallothionein 1A (MT1A), mRNA
  • NM_005947 MT1B 4.879 Homo sapiens metallothionein IB (MT1B), mRNA
  • NM_005947 NM_175617 MT1E 4.98 Homo sapiens metallothionein IE (MT1E), mRNA
  • NM_176870 MT1M 8.142 Homo sapiens metallothionein 1M (MT1M), mRNA
  • NM_005953 MT2A 26.93 Homo sapiens metallothionein 2A (MT2A), mRNA
  • NADP+ dependent 2, methenyltetrahydrofolate cyclohydrolase (MTHFD2), nuclear gene encoding mitochondrial protein, transcript variant 1 , mRNA
  • NM_002466 MYBL2 4.967 Homo sapiens v-myb myeloblastosis viral oncogene homolog (avian)-like 2 (MYBL2), mRNA [NM_002466]
  • NM_002467 MYC 8.723 Homo sapiens v-myc myelocytomatosis viral oncogene homolog (avian) (MYC), mRNA [NM_002467]
  • NM_016132 MYEF2 0.191 Homo sapiens myelin expression factor 2 (MYEF2), mRNA [NM_016132]
  • NM_139207 NAP1L1 6.297 Homo sapiens nucleosome assembly protein 1 -like 1
  • transcript variant 1 mRNA [NM_172164]
  • NM_005381 NCL 19.24 Homo sapiens nucleolin (NCL), mRNA [NM_005381]
  • NCSTN Homo sapiens nicastrin (NCSTN), mRNA [NM_015331]
  • NM_002489 NDUFA4 4.877 Homo sapiens NADH dehydrogenase (ubiquinone) 1 alpha subcomplex, 4, 9kDa (NDUFA4), nuclear gene encoding mitochondrial protein, mRNA [NM_002489]
  • NM_002490 NDUFA6 6.95 Homo sapiens NADH dehydrogenase (ubiquinone) 1 alpha subcomplex, 6, 14kDa (NDUFA6), nuclear gene encoding mitochondrial protein, mRNA [NM_002490]
  • NM_004548 NDUFB10 16.77 Homo sapiens NADH dehydrogenase (ubiquinone) 1 beta subcomplex, 10, 22kDa (NDUFB10), mRNA
  • NM_004547 NDUFB4 6.467 Homo sapiens NADH dehydrogenase (ubiquinone) 1 beta subcomplex, 4, 15kDa (NDUFB4), nuclear gene encoding mitochondrial protein, mRNA [NM_004547]
  • NM_005004 NDUFB8 23.13 Homo sapiens NADH dehydrogenase (ubiquinone) 1 beta subcomplex, 8, 19kDa (NDUFB8), mRNA [NM_005004]
  • NEDD8 developmentally down-regulated 8
  • NM_006156 mRNA [NM_006156]
  • NF2 neuroma
  • transcript variant 8 mRNA
  • NM_181832 NF2 5.332 Homo sapiens neurofibromin 2 (bilateral acoustic
  • NF2 neuroma
  • transcript variant 8 mRNA
  • NM_020529 NFKBIA 5.211 Homo sapiens nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha (NFKBIA), mRNA [NM_020529]
  • NM_016645 NGRN 6.242 Homo sapiens neugrin, neurite outgrowth associated
  • NIPA2 Homo sapiens non imprinted in Prader- Willi/ Angelman syndrome 2 (NIPA2), transcript variant 1, mRNA
  • NM_002512 NME2 7.492 Homo sapiens non-metastatic cells 2, protein (NM23B) expressed in (NME2), transcript variant 1, mRNA
  • COUP transcription factor 1 COUP-TF1
  • COUP-TF I COUP transcription factor 1
  • V-ERBA-related protein EAR-3 V-ERBA-related protein EAR-3
  • NM_021005 NR2F2 13.16 Homo sapiens nuclear receptor subfamily 2, group F, member 2 (NR2F2), mRNA [NM_021005]
  • NTF2-like export factor 1 (NXTl), mRNA
  • NM_001014446 OCIAD2 9.146 Homo sapiens OCIA domain containing 2 (OCIAD2), transcript variant 1, mRNA [NM_001014446]
  • NM_001005203 OR8S1 0.193 Homo sapiens olfactory receptor, family 8, subfamily S, member 1 (OR8S1), mRNA [NM_001005203]
  • succinocarboxamide synthetase PAICS
  • transcript variant 2 mRNA [NM_006452]
  • NM_007262 PARK7 18.16 Homo sapiens Parkinson disease (autosomal recessive, early onset) 7 (PARK7), mRNA [NM_007262]
  • PCBP1 Homo sapiens poly(rC) binding protein 1 (PCBP1)
  • NM_005020 PDE1C 0.196 Homo sapiens phosphodiesterase 1C, calmodulin- dependent 70kDa (PDE1C), mRNA [NM_005020]
  • NM_004911 PDIA4 4.625 Homo sapiens protein disulfide isomerase family A, member 4 (PDIA4), mRNA [NM_004911]
  • NM_014887 PFAAP5 4.26 Homo sapiens phosphonoformate immuno-associated protein 5 (PFAAP5), mRNA [NM_014887]
  • NM_007350 PHLDA1 6.001 Homo sapiens pleckstrin homology-like domain, family A, member 1 (PHLDA1), mRNA [NM_007350]
  • NM_012398 PIP5K1C 4.585 Homo sapiens phosphatidylinositol-4-phosphate 5-kinase, type I, gamma (PIP5K1C), mRNA [NM_012398]
  • NM_015549 PLEKHG3 4.693 Homo sapiens pleckstrin homology domain containing, family G (with RhoGef domain) member 3 (PLEKHG3), mRNA [NM_015549]
  • NM_018049 PLEKHJ1 280.2 Homo sapiens pleckstrin homology domain containing, family J member 1 (PLEKHJ1), mRNA [NM_018049]
  • NM_002668 PLP2 5.112 Homo sapiens proteolipid protein 2 (colonic epithelium- enriched) (PLP2), mRNA [NM_002668]
  • NM_002670 PLS1 4.873 Homo sapiens plastin 1 (I isoform) (PLS1), mRNA
  • NM_020360 PLSCR3 5.118 Homo sapiens phospholipid scramblase 3 (PLSCR3), mRNA [NM_020360]
  • NM_017443 POLE3 6.232 Homo sapiens polymerase (DNA directed), epsilon 3 (pi 7 subunit) (POLE3), mRNA [NM_017443]
  • RNA II DNA directed polypeptide A, 220kDa (POLR2A), mRNA [NM_000937]
  • POLR2J2 polypeptide J-related gene
  • NM_032959 mRNA [NM_032959]
  • NM_005837 POP7 6.522 Homo sapiens processing of precursor 7, ribonuclease P subunit (S. cerevisiae) (POP7), mRNA [NM_005837] NM_021129 PPA1 18.13 Homo sapiens pyrophosphatase (inorganic) 1 (PPA1), mRNA [NM_021129]
  • NM_000943 PPIC 0.0278 Homo sapiens peptidylprolyl isomerase C (cyclophilin C)
  • NM_177983 PPM1G 4.641 Homo sapiens protein phosphatase 1G (formerly 2C), magnesium-dependent, gamma isoform (PPM1G), transcript variant 1, mRNA [NM_177983]
  • NM_138689 PPP1R14B 7.384 Homo sapiens protein phosphatase 1 , regulatory (inhibitor) subunit 14B (PPP1R14B), mRNA [NM_138689]
  • NM_024607 PPP1R3B 11.47 Homo sapiens protein phosphatase 1 , regulatory (inhibitor) subunit 3B (PPP1R3B), mRNA [NM_024607]
  • NM_138558 PPP1R8 8.723 Homo sapiens protein phosphatase 1 , regulatory (inhibitor) subunit 8 (PPP1R8), transcript variant 2, mRNA
  • NM_002715 PPP2CA 5.813 Homo sapiens protein phosphatase 2 (formerly 2A), catalytic subunit, alpha isoform (PPP2CA), mRNA
  • NM_001009552 PPP2CB 10.09 Homo sapiens protein phosphatase 2 (formerly 2A), catalytic subunit, beta isoform (PPP2CB), transcript variant 2, mRNA [NM_001009552]
  • PRDX4 8.136 Homo sapiens peroxiredoxin 4 (PRDX4), mRNA
  • NM_004905 PRDX6 5.192 Homo sapiens peroxiredoxin 6 (PRDX6), mRNA
  • NM_002773 PRSS8 5.035 Homo sapiens protease, serine, 8 (PRSS8), mRNA
  • NM_020200 PRTFDC1 0.16 Homo sapiens phosphoribosyl transferase domain
  • NM_002789 PSMA4 18.14 Homo sapiens proteasome (prosome, macropain) subunit, alpha type, 4 (PSMA4), mRNA [NM_002789]
  • NM_002791 PSMA6 9.663 Homo sapiens proteasome (prosome, macropain) subunit, alpha type, 6 (PSMA6), mRNA [NM_002791]
  • NM_002793 PSMB1 36.45 Homo sapiens proteasome (prosome, macropain) subunit, beta type, 1 (PSMB1), mRNA [NM_002793]
  • NM_002802 PSMC1 5.809 Homo sapiens proteasome (prosome, macropain) 26S subunit, ATPase, 1 (PSMC1), mRNA [NM_002802]
  • NM_005805 PSMD14 4.907 Homo sapiens proteasome (prosome, macropain) 26S subunit, non- ATPase, 14 (PSMD14), mRNA
  • NM_001015509 PTRH2 Homo sapiens peptidyl-tRNA hydrolase 2 (PTRH2), nuclear gene encoding mitochondrial protein, transcript variant 1, mRNA [NM_001015509]
  • NM_004219 PTTG1 8.83 Homo sapiens pituitary tumor-transforming 1 (PTTG1), mRNA [NM_004219]
  • NM_004219 PTTG1 4.869 Homo sapiens pituitary tumor-transforming 1 (PTTG1), mRNA [NM_004219] NM_015317 PUM2 7.61 Homo sapiens pumilio homolog 2 (Drosophila) (PUM2), mRNA [NM_015317]
  • NM_007062 PWP1 9.827 Homo sapiens PWP1 homolog (S. cerevisiae) (PWP1), mRNA [NM_007062]
  • NM_024638 QTRTD1 67.01 Homo sapiens queuine tRNA-ribosyltransferase domain containing 1 (QTRTD1), mRNA [NM_024638]
  • NM_004578 RAB4A 5.418 Homo sapiens RAB4A, member RAS oncogene family
  • NM_004582 RABGGTB 5.997 Homo sapiens Rab geranylgeranyltransferase, beta subunit
  • RANBPl Homo sapiens RAN binding protein 1 (RANBPl), mRNA
  • NM_002887 RARS 7.849 Homo sapiens arginyl-tRNA synthetase (RARS), mRNA
  • NM_005105 RBM8A 15.63 Homo sapiens RNA binding motif protein 8A (RBM8A), mRNA [NM_005105]
  • NM_005669 REEP5 4.716 Homo sapiens receptor accessory protein 5 (REEP5), mRNA [NM_005669]
  • NM_015523 REX02 10.14 Homo sapiens REX2, RNA exonuclease 2 homolog (S.
  • NM_017819 RG9MTD1 6.782 Homo sapiens RNA (guanine-9-) methyltransferase
  • NM_005614 RHEB 4.841 Homo sapiens Ras homolog enriched in brain (RHEB), mRNA [NM_005614]
  • NM_002939 RNH1 6.957 Homo sapiens ribonuclease/angiogenin inhibitor 1
  • NM_000975 RPL11 5.544 Homo sapiens ribosomal protein LI 1 (RPL11), mRNA
  • NM_000976 RPL12 10.66 Homo sapiens ribosomal protein L12 (RPL12), mRNA
  • NM_003973 RPL14 9.297 Homo sapiens ribosomal protein L14 (RPL14), transcript variant 2, mRNA [NM_003973]
  • NM_002948 RPL15 14.38 Homo sapiens ribosomal protein LI 5 (RPL15), mRNA
  • NM_002948 RPL15 6.062 Homo sapiens ribosomal protein LI 5 (RPL15), mRNA
  • NM_000985 RPL17 4.824 Homo sapiens ribosomal protein L17 (RPL17), transcript variant 1, mRNA [NM_000985]
  • NM_000979 RPL18 6.586 Homo sapiens ribosomal protein LI 8 (RPL18), mRNA
  • NM_000981 RPL19 147.7 Homo sapiens ribosomal protein L19 (RPL19), mRNA
  • NM_000983 RPL22 4.747 Homo sapiens ribosomal protein L22 (RPL22), mRNA
  • NM_000984 RPL23A 11.89 Homo sapiens ribosomal protein L23a (RPL23A), mRNA
  • NM_000986 RPL24 19.98 Homo sapiens ribosomal protein L24 (RPL24), mRNA
  • NM_016093 RPL26L1 7.474 Homo sapiens ribosomal protein L26-like 1 (RPL26L1), mRNA [NM_016093]
  • NM_000993 RPL31 17.74 Homo sapiens ribosomal protein L31 (RPL31), mRNA
  • NM_033625 RPL34 59.35 Homo sapiens ribosomal protein L34 (RPL34), transcript variant 2, mRNA [NM_033625]
  • NM_000996 RPL35A 21.23 Homo sapiens ribosomal protein L35a (RPL35A), mRNA
  • NM_015414 RPL36 5.287 Homo sapiens ribosomal protein L36 (RPL36), transcript variant 2, mRNA [NM_015414]
  • NM_021029 RPL36A 17.53 Homo sapiens ribosomal protein L36a (RPL36A), mRNA
  • NM_000999 RPL38 100.7 Homo sapiens ribosomal protein L38 (RPL38), transcript variant 1, mRNA [NM_000999]
  • NM_001000 RPL39 7.078 Homo sapiens ribosomal protein L39 (RPL39), mRNA
  • NM_053275 RPLPO 7.848 Homo sapiens ribosomal protein, large, P0 (RPLPO), transcript variant 2, mRNA [NM_053275]
  • NM_001003 RPLP1 105 Homo sapiens ribosomal protein, large, PI (RPLP1), transcript variant 1, mRNA [NM_001003]
  • NM_001015 RPS11 6.163 Homo sapiens ribosomal protein S 11 (RPS 11), mRNA
  • NM_001016 NM_001017 RPS13 22.56 Homo sapiens ribosomal protein S13 (RPS 13), mRNA
  • NM_005617 RPS14 5.668 Homo sapiens ribosomal protein S14 (RPS 14), transcript variant 3, mRNA [NM_005617]
  • NM_001019 RPS15A 17.61 Homo sapiens ribosomal protein SI 5a (RPS15A), transcript variant 2, mRNA [NM_001019]
  • NM_001020 RPS16 21.56 Homo sapiens ribosomal protein S16 (RPS 16), mRNA
  • NM_002952 RPS2 10.65 Homo sapiens ribosomal protein S2 (RPS2), mRNA
  • NM_001024 RPS21 19.89 Homo sapiens ribosomal protein S21 (RPS21), mRNA
  • NM_001025 RPS23 4.25 Homo sapiens ribosomal protein S23 (RPS23), mRNA
  • NM_001028 RPS25 35.14 Homo sapiens ribosomal protein S25 (RPS25), mRNA
  • NM_001030 RPS27 9.202 Homo sapiens ribosomal protein S27 (metallopanstimulin
  • NM_001032 RPS29 22.91 Homo sapiens ribosomal protein S29 (RPS29), transcript variant 1, mRNA [NM_001032]
  • NM_001005 RPS3 5.221 Homo sapiens ribosomal protein S3 (RPS3), mRNA
  • NM_001011 RPS7 21.93 Homo sapiens ribosomal protein S7 (RPS7), mRNA
  • NM_001033 RRM1 4.609 Homo sapiens ribonucleotide reductase Ml polypeptide
  • NM_001034 RRM2 30.43 Homo sapiens ribonucleotide reductase M2 polypeptide
  • NM_005500 SAE1 15.61 Homo sapiens SUMOl activating enzyme subunit 1
  • NM_005499 SAE2 11.05 Homo sapiens SUMOl activating enzyme subunit 2
  • NM_005505 SCARB1 5.28 Homo sapiens scavenger receptor class B, member 1
  • SCOTIN 9.123 Homo sapiens scotin (SCOTIN), mRNA [NM_016479]
  • NM_182706 SCRIB 5104 Homo sapiens scribbled homolog (Drosophila) (SCRIB), transcript variant 1, mRNA [NM_182706]
  • SELK Homo sapiens selenoprotein K
  • SEPW1 Homo sapiens selenoprotein W, 1 (SEPW1), mRNA
  • NM_004630 SF1 8.327 Homo sapiens splicing factor 1 (SF1), transcript variant 1, mRNA [NM_004630]
  • NM_003017 SFRS3 5.625 Homo sapiens splicing factor, arginine/serine-rich 3
  • NM_003769 SFRS9 18.09 Homo sapiens splicing factor, arginine/serine-rich 9
  • NM_152621 SGMS2 5.131 Homo sapiens sphingomyelin synthase 2 (SGMS2), mRNA [NM_152621]
  • NM_006427 SIVA1 17.05 Homo sapiens SIVA1, apoptosis-inducing factor (SIVA1), transcript variant 1, mRNA [NM_006427]
  • NM_006930 SKP1A 8.761 Homo sapiens S-phase kinase-associated protein 1A
  • NM_006598 SLC12A7 6.025 Homo sapiens solute carrier family 12 (potassium/chloride transporters), member 7 (SLC12A7), mRNA
  • NM_021734 SLC25A19 8.7 Homo sapiens solute carrier family 25 (mitochondrial thiamine pyrophosphate carrier), member 19 (SLC25A19), mRNA [NM_021734]
  • NM_213611 SLC25A3 5.807 Homo sapiens solute carrier family 25 (mitochondrial carrier; phosphate carrier), member 3 (SLC25A3), nuclear gene encoding mitochondrial protein, transcript variant 3, mRNA [NM_213611]
  • NM_001152 SLC25A5 7.17 Homo sapiens solute carrier family 25 (mitochondrial carrier; adenine nucleotide translocator), member 5 (SLC25A5), mRNA [NM_001152]
  • NM_198580 SLC27A1 4.457 Homo sapiens solute carrier family 27 (fatty acid
  • NMJ98580 NM_003041 SLC5A2 0.0193 Homo sapiens solute carrier family 5 (sodium/glucose cotransporter), member 2 (SLC5A2), mRNA
  • NM_003096 SNRPG 44.9 Homo sapiens small nuclear ribonucleoprotein polypeptide
  • NM_000454 SOD1 12.57 Homo sapiens superoxide dismutase 1, soluble
  • NM_014978 SORCS3 0.164 Homo sapiens sortilin-related VPS 10 domain containing receptor 3 (SORCS3), mRNA [NM_014978]
  • NM_003113 SP100 9.236 Homo sapiens SP100 nuclear antigen (SP100), transcript variant 2, mRNA [NM_003113]
  • NM_021199 SQRDL 8.771 Homo sapiens sulfide quinone reductase-like (yeast)
  • SREBF1 transcription factor 1
  • transcript variant 1 mRNA [NM_001005291]
  • NM_003143 SSBP1 4.96 Homo sapiens single-stranded DNA binding protein 1
  • NM_003144 SSR1 7.851 Homo sapiens signal sequence receptor, alpha (translocon-associated protein alpha) (SSR1), mRNA [NM_003144]
  • NM_003145 SSR2 11.51 Homo sapiens signal sequence receptor, beta (translocon-associated protein beta) (SSR2), mRNA [NM_003145]
  • NM_203401 STMN1 24.76 Homo sapiens stathmin 1/oncoprotein 18 (STMN1), transcript variant 1, mRNA [NM_203401]
  • NM_015894 STMN3 6.697 Homo sapiens stathmin-like 3 (STMN3), mRNA
  • NM_007178 STRAP 19.44 Homo sapiens serine/threonine kinase receptor associated protein (STRAP), mRNA [NM_007178]
  • TCEB1 7.365 Homo sapiens transcription elongation factor B (SIII), polypeptide 1 (15kDa, elongin C) (TCEB1), mRNA
  • NM_003200 TCF3 7.147 Homo sapiens transcription factor 3 (E2A immunoglobulin enhancer binding factors E12/E47) (TCF3), mRNA
  • NM_030752 TCP1 13.42 Homo sapiens t-complex 1 (TCP1), transcript variant 1, mRNA [NM_030752]
  • NM_001586 TEX28 319.6 Homo sapiens testis expressed 28 (TEX28), mRNA
  • NM_003236 TGFA 7.588 Homo sapiens transforming growth factor, alpha (TGFA), mRNA [NM_003236]
  • NM_005782 THOC4 14.47 Homo sapiens THO complex 4 (THOC4), mRNA
  • AK025280 THRAP2 5.301 Homo sapiens cDNA: FLJ21627 fis, clone COL08058.
  • NM_003254 TIMP1 4.622 Homo sapiens TIMP metallopeptidase inhibitor 1
  • TRIP1 mRNA [NM_003254]
  • TK1 48.01 Homo sapiens thymidine kinase 1, soluble (TK1), mRNA
  • TMEM4 13.7 Homo sapiens transmembrane protein 4 (TMEM4), mRNA
  • TMEM9B 11.92 Homo sapiens TMEM9 domain family, member B
  • NM_001065 TNFRSF1A 27.93 Homo sapiens tumor necrosis factor receptor superfamily, member 1A (TNFRSF1A), mRNA [NM_001065]
  • NM_003807 TNFSF14 24.95 Homo sapiens tumor necrosis factor (ligand) superfamily, member 14 (TNFSF14), transcript variant 1, mRNA
  • NM_005079 TPD52 5.983 Homo sapiens tumor protein D52 (TPD52), transcript variant 3, mRNA [NM_005079] NM_153649 TPM3 5.735 Homo sapiens tropomyosin 3 (TPM3), transcript variant 2, mRNA [NM_153649]
  • NM_016292 TRAP1 16.4 Homo sapiens TNF receptor-associated protein 1
  • NM_001011658 TRAPPC2 0.176 Homo sapiens trafficking protein particle complex 2
  • NM_016399 TRIAP1 16.27 Homo sapiens TP53 regulated inhibitor of apoptosis 1
  • NM_021158 TRIB3 5.91 Homo sapiens tribbles homolog 3 (Drosophila) (TRJB3), mRNA [NM_021158]
  • NM_003302 TRIP6 7.623 Homo sapiens thyroid hormone receptor interactor 6
  • NM_017646 TRIT1 0.0796 Homo sapiens tRNA isopentenyltransferase 1 (TRIT1), mRNA [NM_017646]
  • NM_006000 TUBA4A 5.129 Homo sapiens tubulin, alpha 4a (TUBA4A), mRNA
  • TP53 Homo sapiens tumor protein p53
  • TUBB6 7.935 Homo sapiens tubulin, beta 6 (TUBB6), mRNA
  • TXNL5 5.082 Homo sapiens thioredoxin-like 5 (TXNL5), mRNA
  • NM_001071 TYMS 4.999 Homo sapiens thymidylate synthetase (TYMS), mRNA
  • NM_003333 UBA52 15.83 Homo sapiens ubiquitin A-52 residue ribosomal protein fusion product 1 (UBA52), transcript variant 2, mRNA [NM_003333]
  • NM_021009 UBC 67.28 Homo sapiens ubiquitin C (UBC), mRNA [NM_021009]
  • transcript variant 2 (7.2 kD) (UCRC), transcript variant 2, mRNA
  • NM_013282 UHRF1 5.665 Homo sapiens ubiquitin-like, containing PHD and RING finger domains, 1 (UHRF1), transcript variant 2, mRNA [NM_013282]
  • NM_003362 UNG 5.482 Homo sapiens uracil-DNA glycosylase (UNG), transcript variant 1, mRNA [NM_003362] NM_006830 UQCR 10.48 Homo sapiens ubiquinol-cytochrome c reductase, 6.4kDa subunit (UQCR), mRNA [NM_006830]
  • NM_006004 UQCRH 3 -Feb Homo sapiens ubiquinol-cytochrome c reductase hinge protein (UQCRH), mRNA [NM_006004]
  • NM_014402 UQCRQ 7.412 Homo sapiens ubiquinol-cytochrome c reductase, complex
  • NM_032747 USMG5 9.398 Homo sapiens upregulated during skeletal muscle growth 5 homolog (mouse) (USMG5), mRNA [NM_032747]
  • ribonucleoprotein ribonucleoprotein, homolog C (yeast) (UTP14C), mRNA [NM_021645]
  • NM_003378 VGF 5.302 Homo sapiens VGF nerve growth factor inducible (VGF), mRNA [NM_003378]
  • VPS24 transcript variant 1, mRNA
  • NM_016303 WBP5 11.53 Homo sapiens WW domain binding protein 5 (WBP5), transcript variant 1, mRNA [NM_016303]
  • NM_030798 WBSCR16 4.879 Homo sapiens Williams-Beuren syndrome chromosome region 16 (WBSCR16), mRNA [NM_030798]
  • NM_052950 WDFY2 0.192 Homo sapiens WD repeat and FYVE domain containing 2
  • NM_145294 WDR90 0.0386 Homo sapiens WD repeat domain 90 (WDR90), mRNA
  • NM_003400 XPOl 5.32 Homo sapiens exportin 1 (CRM1 homolog, yeast) (XPOl), mRNA [NM_003400]
  • NM_004559 YBX1 4.318 Homo sapiens Y box binding protein 1 (YBX1), mRNA
  • NM_017798 YTHDFl Homo sapiens YTH domain family, member 1 (YTHDFl), mRNA [NM_017798]
  • NM_139118 YY1AP1 5.12 Homo sapiens YY1 associated protein 1 (YY1AP1), transcript variant 2, mRNA [NM_139118]
  • NM_004926 ZFP36L1 12.96 Homo sapiens zinc finger protein 36, C3H type-like 1
  • NM_153813 ZFPM1 10.25 Homo sapiens zinc finger protein, multitype 1 (ZFPM1), mRNA [NM_153813]
  • NM_007149 ZNF184 5.475 Homo sapiens zinc finger protein 184 (ZNF184), mRNA
  • NM_152600 ZNF579 11.09 Homo sapiens zinc finger protein 579 (ZNF579), mRNA

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Abstract

La présente invention concerne un procédé de séparation d'un ARN activement traduit d'un échantillon biologique comprenant l'étape consistant à mettre en contact un échantillon biologique avec une molécule de liaison chaperonne. La présente invention permet l'isolement, la détection, la détermination de l'expression, et le profilage de l'ARN activement traduit. L'invention propose également un kit pour mettre l'invention en application.
PCT/US2010/060512 2009-12-15 2010-12-15 Isolement d'arnm activement traduits d'un petit nombre de cellules WO2011084485A1 (fr)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2013170000A1 (fr) * 2012-05-09 2013-11-14 The Rockfeller University Procédés et compositions pour le profilage de transcriptome dépendant de l'activité

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20030031661A1 (en) * 2001-05-01 2003-02-13 Michael Graner Methods of recovering chaperone proteins and complexes thereof
US20040096878A1 (en) * 1999-12-28 2004-05-20 Ribonomics, Inc. Methods for isolating and characterizing endogenous mRNA-protein (mRNP) complexes
US20050009028A1 (en) * 2001-10-29 2005-01-13 Nathaniel Heintz Method for isolating cell-type specific mrnas
US20050100947A1 (en) * 1998-07-14 2005-05-12 Zyomyx, Inc. Array devices and methods of use thereof

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20050100947A1 (en) * 1998-07-14 2005-05-12 Zyomyx, Inc. Array devices and methods of use thereof
US20040096878A1 (en) * 1999-12-28 2004-05-20 Ribonomics, Inc. Methods for isolating and characterizing endogenous mRNA-protein (mRNP) complexes
US20030031661A1 (en) * 2001-05-01 2003-02-13 Michael Graner Methods of recovering chaperone proteins and complexes thereof
US20050009028A1 (en) * 2001-10-29 2005-01-13 Nathaniel Heintz Method for isolating cell-type specific mrnas

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2013170000A1 (fr) * 2012-05-09 2013-11-14 The Rockfeller University Procédés et compositions pour le profilage de transcriptome dépendant de l'activité

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