WO2006084201A3 - Method of isolating, labeling and profiling small rna and whole-genome transcripts - Google Patents

Method of isolating, labeling and profiling small rna and whole-genome transcripts Download PDF

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Publication number
WO2006084201A3
WO2006084201A3 PCT/US2006/003977 US2006003977W WO2006084201A3 WO 2006084201 A3 WO2006084201 A3 WO 2006084201A3 US 2006003977 W US2006003977 W US 2006003977W WO 2006084201 A3 WO2006084201 A3 WO 2006084201A3
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WO
WIPO (PCT)
Prior art keywords
rna
labeled
microarray
small
labeling
Prior art date
Application number
PCT/US2006/003977
Other languages
French (fr)
Other versions
WO2006084201A2 (en
Inventor
X Xia
Original Assignee
Genosensor Corp
X Xia
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US11/116,915 external-priority patent/US20060248197A1/en
Application filed by Genosensor Corp, X Xia filed Critical Genosensor Corp
Publication of WO2006084201A2 publication Critical patent/WO2006084201A2/en
Publication of WO2006084201A3 publication Critical patent/WO2006084201A3/en

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    • 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
    • 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
    • C12N15/1096—Processes for the isolation, preparation or purification of DNA or RNA cDNA Synthesis; Subtracted cDNA library construction, e.g. RT, RT-PCR
    • 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
    • C12Q1/6813—Hybridisation assays
    • C12Q1/6834—Enzymatic or biochemical coupling of nucleic acids to a solid phase
    • C12Q1/6837—Enzymatic or biochemical coupling of nucleic acids to a solid phase using probe arrays or probe chips

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  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Engineering & Computer Science (AREA)
  • Genetics & Genomics (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biotechnology (AREA)
  • General Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • General Health & Medical Sciences (AREA)
  • Biophysics (AREA)
  • Microbiology (AREA)
  • Analytical Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • Biomedical Technology (AREA)
  • Immunology (AREA)
  • Crystallography & Structural Chemistry (AREA)
  • Plant Pathology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Bioinformatics & Computational Biology (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

Disclosed herein is a method of selectively labeling non-messenger RNA molecules by isolating total RNA from a tissue or cell, dissolving the isolated RNA, blocking 3' ends of the RNA and adding T4 RNA ligase and a labeled nucleic acid adaptor, with the result that the T4 RNA ligase ligates the adaptor only to RNA having a 5' phosphate group such as small RNAs. A method of labeling the 5' end of mRNA isolates total RNA from a tissue or cell, dissolves RNA in RNase-free water, removes a 5' cap structure from the mRNA using tobacco acid pyrophosphatase (TAP), removes the TAP, blocks the 3' end of the RNA molecules; and ligates an adaptor to the RNA by adding T4 RNA ligase and a labeled DNA or RNA adaptor. A method of amplifying and labeling non-capped RNAs and/or capped RNAs is useful in expression analysis of the whole-genome transcripts from cells and tissues. Sense and antisense transcripts are labeled from different experimental approaches. In another embodiment, there is disclosed a method of sequence selection and probe design. Probes are designed complementary to the labeled target RNA. For small RNAs, sequences are selected for detections of mature, counterparts (ig miR*), and precursors. In another embodiment, there is disclosed a method of expression profiling small RNA by separating labeled RNA from capped RNA, providing a microarray comprising a plurality of probes hybridizable to small RNA, incubating the labeled small RNA with the microarray, washing unhybridized RNA from the microarray and drying the microarray, staining hybridized RNA on the microarray; and scanning the labeled microarray to determine the identity and quantity of labeling to the various nu'RNA probe sites and thus providing an expression profile of small RNA.
PCT/US2006/003977 2005-02-04 2006-02-06 Method of isolating, labeling and profiling small rna and whole-genome transcripts WO2006084201A2 (en)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US65003405P 2005-02-04 2005-02-04
US60/650,034 2005-02-04
US11/116,915 US20060248197A1 (en) 2005-04-27 2005-04-27 Adaptive connectionless scheduling protocol
US11/116,915 2005-04-28

Publications (2)

Publication Number Publication Date
WO2006084201A2 WO2006084201A2 (en) 2006-08-10
WO2006084201A3 true WO2006084201A3 (en) 2009-04-16

Family

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Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/US2006/003977 WO2006084201A2 (en) 2005-02-04 2006-02-06 Method of isolating, labeling and profiling small rna and whole-genome transcripts

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107429296A (en) * 2015-03-13 2017-12-01 生命技术公司 Method, composition and the kit of capture, detection and quantitative tiny RNA

Families Citing this family (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2008040355A2 (en) * 2006-10-06 2008-04-10 Exiqon A/S Novel methods for quantification of micrornas and small interfering rnas
WO2009135212A2 (en) * 2008-05-02 2009-11-05 Epicentre Technologies Corporation Selective 5' ligation tagging of rna
EP3260558B1 (en) 2012-11-02 2020-05-13 Life Technologies Corporation Novel compositions and methods for enhancing pcr specificity

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
IGLOI, G.L. ET AL.: "Nonradioactive Labeling of RNA", ANALAYTICAL BIOCHEM., vol. 233, no. 1, 1 January 1996 (1996-01-01), pages 124 - 129 *
KINOSHITA, Y. ET AL.: "Fluorescence-, isotope- or biotin-labeling of the 5'-end of single stranded DNA/RNA using T4 RNa ligase", NUCL. ACIDS RES., vol. 25, no. 18, 15 September 1997 (1997-09-15), pages 3747 - 3748, XP002907774, DOI: doi:10.1093/nar/25.18.3747 *
LIU X. ET AL.: "MApping the 5' and 3' ends of Tetrahymena thermophila mRNAs using RNA ligase mediated amplification of cDNA ends (RLM-RACE)", NUCL. ACIDS RES., vol. 21, no. 21, 25 October 1993 (1993-10-25), pages 4954 - 4960 *
MARUYAMA K. ET AL.: "Oligo-capping: a simple method to replace the cap structure of eukaryotic mRNAs with oligoribonucleotides", GENE, vol. 138, no. 1-2, 28 January 1994 (1994-01-28), pages 171 - 174, XP002915960, DOI: doi:10.1016/0378-1119(94)90802-8 *
VOLLOCH V. ET AL.: "Ligation-mediated amplification of RNA from murine erythroid cells reveals a novel class of beta globin mRNA with an extended 5'-untranslated region.", NUCL. ACIDS RES., vol. 22, no. 13, 11 July 1994 (1994-07-11), pages 2507 - 2511 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107429296A (en) * 2015-03-13 2017-12-01 生命技术公司 Method, composition and the kit of capture, detection and quantitative tiny RNA
CN107429296B (en) * 2015-03-13 2022-01-28 生命技术公司 Methods, compositions and kits for capturing, detecting and quantifying small RNAs

Also Published As

Publication number Publication date
WO2006084201A2 (en) 2006-08-10

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