Djebali, S., Kapranov, P., Foissac, S., Lagarde, J., Reymond, A., Ucla, C., Wyss, C., Drenkow, J., Dumais, E., Murray, R. R., Lin, C., Szeto, D., Denoeud, F., Calvo, M., Frankish, A., Harrow, J., Makrythanasis, P., Vidal, M., Salehi-Ashtiani, K., Antonarakis, S. E., Gingeras, T. R., Guigo, R. (2008) Efficient targeted transcript discovery via array-based normalization of RACE libraries. Nature Methods, 5 (7). pp. 629-35. ISSN 15487091 (ISSN) (Public Dataset)
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Abstract
Rapid amplification of cDNA ends (RACE) is a widely used approach for transcript identification. Random clone selection from the RACE mixture, however, is an ineffective sampling strategy if the dynamic range of transcript abundances is large. To improve sampling efficiency of human transcripts, we hybridized the products of the RACE reaction onto tiling arrays and used the detected exons to delineate a series of reverse-transcriptase (RT)-PCRs, through which the original RACE transcript population was segregated into simpler transcript populations. We independently cloned the products and sequenced randomly selected clones. This approach, RACEarray, is superior to direct cloning and sequencing of RACE products because it specifically targets new transcripts and often results in overall normalization of transcript abundance. We show theoretically and experimentally that this strategy leads indeed to efficient sampling of new transcripts, and we investigated multiplexing the strategy by pooling RACE reactions from multiple interrogated loci before hybridization.
| Item Type: | Paper |
|---|---|
| Additional Information: | 1548-7105 (Electronic) Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't PMC2713501 |
| Uncontrolled Keywords: | Alternative Splicing Chromosomes, Human, Pair 21/genetics Chromosomes, Human, Pair 22/genetics Cloning, Molecular DNA, Complementary/*genetics Exons Gene Expression Profiling/*methods *Gene Library Genome, Human Humans Molecular Sequence Data Nucleic Acid Amplification Techniques/*methods Oligonucleotide Array Sequence Analysis/methods Protein Isoforms/genetics RNA/*genetics Reverse Transcriptase Polymerase Chain Reaction Transcription, Genetic |
| Subjects: | bioinformatics > genomics and proteomics > analysis and processing > DNA RNA Processing bioinformatics > genomics and proteomics bioinformatics > genomics and proteomics > analysis and processing > microarray gene expression processing ?? Q1 ?? bioinformatics > genomics and proteomics > alignment > sequence alignment |
| CSHL Authors: | |
| Communities: | CSHL labs > Gingeras lab |
| Depositing User: | Tom Adams |
| Date: | 2008 |
| Date Deposited: | 13 Jul 2011 17:55 |
| Last Modified: | 12 Jul 2013 18:28 |
| PMCID: | PMC2713501 |
| Related URLs: | |
| Dataset ID: |
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| URI: | https://repository.cshl.edu/id/eprint/7718 |
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