Submitted to: Animal Genetics International Conference Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: 7/21/2004
Publication Date: 9/11/2004
Citation: Harhay, G.P., Smith, T.P., Sonstegard, T.S., Clawson, M.L. 2004. Database directed full-length cDNA generation using EST data from normalized bovine cDNA libraries. Proc., 29th International Conference on Animal Genetics (ISAG), Tokyo, Japan. September 11-16, 2004. Abstract A022, p. 37.
Technical Abstract: Full-length cDNA including the 5' untranslated region (UTR), entire coding sequence, and 3' UTR are vitally important in producing accurate gene models and resolving ambiguities in gene models built solely upon computational prediction, genomic and expressed sequence tag (EST) data. Annotation of the impending bovine genome sequence assembly will benefit greatly from full-length cDNA sequence data. Therefore, we developed a database for directed full-length bovine cDNA prediction, sequencing, analysis, and annotation. To facilitate the economical production of bovine full-length cDNA, EST data produced from normalized cDNA libraries were computationally screened for potential full-length clone candidates via similarity to transcripts in the NCBI RefSeq database. Using primer walking, the entire length of the candidate clone inserts were sequenced and assembled in the database pipeline. We routinely sequenced and contiged 384 different candidate clones per plate. The pipeline is tolerant to sequencing failures so that sequencing of different clones with different walk steps may occur simultaneously on the same plate in any given sequencing run. Over 1,000 full-length cDNA have been processed through this analysis and annotation pipeline prior to submission to the NCBI FLIC (Full-Length Insert Clone) database.