Abstract
The publication of a draft sequence of a third mammalian genome--that of the rat--suggests a need to rethink genome annotation. New mammalian sequences will not receive the kind of labor-intensive annotation efforts that are currently being devoted to human. In this paper, we demonstrate an alternative approach: reverse transcription-polymerase chain reaction (RT-PCR) and direct sequencing based on dual-genome de novo predictions from TWINSCAN. We tested 444 TWINSCAN-predicted rat genes that showed significant homology to known human genes implicated in disease but that were partially or completely missed by methods based on protein-to-genome mapping. Using primers in exons flanking a single predicted intron, we were able to verify the existence of 59% of these predicted genes. We then attempted to amplify the complete predicted open reading frames of 136 genes that were verified in the single-intron experiment. Spliced sequences were amplified in 46 cases (34%). We conclude that this procedure for elucidating gene structures with native cDNA sequences is cost-effective and will become even more so as it is further optimized.
| Originalsprog | Engelsk |
|---|---|
| Tidsskrift | Genome Research |
| Vol/bind | 14 |
| Udgave nummer | 4 |
| Sider (fra-til) | 665-71 |
| Antal sider | 7 |
| ISSN | 1088-9051 |
| DOI | |
| Status | Udgivet - 2004 |
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