30,000 CATFISH ESTs: A CASE STUDY FOR A SET OF QUALITY STANDARDS ESTABLISHED FOR THE ANALYSIS OF ESTs

¹ The Fish Molecular Genetics and Biotechnology Laboratory, Department of Fisheries and Allied Aquacultures and Program of Cell and Molecular Biosciences, Aquatic Genomics Unit, Auburn University, Auburn, AL 36849, USA

² The W. M. Keck Center for Comparative and Functional Genomics, Department of Animal Sciences, University of Illinois at Urbana-Champaign, 330 ERML, 1201 W. Gregory Dr., Urbana, IL 61801

³ Department of Animal Sciences, Purdue University, West Lafayette IN 47907

Expressed sequence tags (ESTs) are valuable genome resources. We have generated 31,451 catfish ESTs including 21,147 from channel catfish and 10,304 from blue catfish. With the existing catfish ESTs, this project brings the total number of ESTs of catfish to over 43,000. For the purpose of cDNA microarrays, the channel and blue catfish ESTs were combined because they are extremely similar in sequence identities. Over 20,000 unique sequences have been identified from this project. Catfish ESTs are rich in microsatellites and single nucleotide polymorphisms (SNPs). While SNP analysis is still under way, searches of microsatellites resulted in identification of 4,855 ESTs containing microsatellites, which included 569 contigs and 3,534 singletons and together representing 4,103 unique sequences. These microsatellites should be useful for linkage mapping and for comparative mapping because they are associated with genes. We are currently conducting bioinformatic analysis for annotation, gene ontology, SNPs, and the unigene list. To facilitate downstream applications including cDNA microarrays, full-length cDNA sequencing, and identification of gene-associated markers, we are establishing a set of quality standards for the analysis of ESTs. These standards include: 1. Low redundancy rate/high gene discovery rate; 2. Gene identification rate; 3. average EST length; 4. Rate of complete ORFs among known genes; 5. Ratio of contigs formed within known genes and the number of known genes; 6. Percentage of clean sequences with high quality scores; 7. Genome resource information contents including microsatellites, insertion/deletions, alternative splicing, alternative adenylation, and SNPs.