Gene prediction and verification in a compact genome with numerous small introns.

Journal Article (Journal Article)

The genomes of clusters of related eukaryotes are now being sequenced at an increasing rate, creating a need for accurate, low-cost annotation of exon-intron structures. In this paper, we demonstrate that reverse transcription-polymerase chain reaction (RT-PCR) and direct sequencing based on predicted gene structures satisfy this need, at least for single-celled eukaryotes. The TWINSCAN gene prediction algorithm was adapted for the fungal pathogen Cryptococcus neoformans by using a precise model of intron lengths in combination with ungapped alignments between the genome sequences of the two closely related Cryptococcus varieties. This approach resulted in approximately 60% of known genes being predicted exactly right at every coding base and splice site. When previously unannotated TWINSCAN predictions were tested by RT-PCR and direct sequencing, 75% of targets spanning two predicted introns were amplified and produced high-quality sequence. When targets spanning the complete predicted open reading frame were tested, 72% of them amplified and produced high-quality sequence. We conclude that sequencing a small number of expressed sequence tags (ESTs) to provide training data, running TWINSCAN on an entire genome, and then performing RT-PCR and direct sequencing on all of its predictions would be a cost-effective method for obtaining an experimentally verified genome annotation.

Full Text

Duke Authors

Cited Authors

  • Tenney, AE; Brown, RH; Vaske, C; Lodge, JK; Doering, TL; Brent, MR

Published Date

  • November 2004

Published In

Volume / Issue

  • 14 / 11

Start / End Page

  • 2330 - 2335

PubMed ID

  • 15479946

Pubmed Central ID

  • PMC525692

International Standard Serial Number (ISSN)

  • 1088-9051

Digital Object Identifier (DOI)

  • 10.1101/gr.2816704


  • eng

Conference Location

  • United States