TY  - JOUR
A1  - Wu, Jia Qian
A1  - Shteynberg, David
A1  - Arumugam, Manimozhiyan
A1  - Gibbs, Richard A.
A1  - Brent, Michael R.
T1  - Identification of Rat Genes by TWINSCAN Gene Prediction, RT–PCR, and Direct Sequencing
Y1  - 2004/04/01 
JF  - Genome Research 
JO  - Genome Research 
SP  - 665 
EP  - 671 
DO  - 10.1101/gr.1959604 
VL  - 14 
IS  - 4 
UR  - http://genome.cshlp.org/content/14/4/665.abstract 
N2  - 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. 
ER  -