Identification of rat genes by TWINSCAN gene prediction, RT-PCR, and direct sequencing

Jia Qian Wu, David Shteynberg, Manimozhiyan Arumugam, Richard A. Gibbs, Michael R. Bren

Research output: Contribution to journalArticlepeer-review

30 Scopus citations


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.

Original languageEnglish
Pages (from-to)665-671
Number of pages7
JournalGenome research
Issue number4
StatePublished - Apr 2004


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