Theses and Dissertations


Title: Development of expressed sequence tags (ESTs) from eastern oyster (Crassostrea virginica): Lessons learned from previous efforts

Name: Peatman, Eric J.

Degree: MS

Chair: Zhanjiang (John) Liu

Resides:

University: Auburn University

Location: Auburn, Alabama

Date: 2004

Pages: 69

Keywords: EST, oyster, crassostrea virginica

Abstract:

Marine organisms respond to the environment by regulating their gene expression. Pollutants may induce changes in a characteristic set of genes resulting in unique genomic expression signatures (GES). To identify marine biomarkers, a project was initiated to develop ESTs from the eastern oyster (Crassostrea virginica). A total of 4,572 cDNA clones were sequenced generating 4,348 ESTs (95.1% success rate). All these ESTs have been submitted to dbEST. The ESTs had an average length of 641 bp. Combined with the existing oyster ESTs, a total of 5,206 ESTs are now available in the dbEST database from C. virginica. This work accounted for over 83% of ESTs available from the eastern oyster. Cluster analysis of the 4,348 ESTs indicated the presence of 243 contigs and 1,673 singletons together representing 1,916 unique sequences. Low sequencing redundancy was achieved by overgo hybridization subtraction of mitochondrial genes that are expressed abundantly in the gill and gonad tissues from which the ESTs were sequenced. Of the 1,916 unique sequences, 875 were identified as known genes (46%) by BLASTX similarity searches and 1,041 were unknown gene sequences. This rate of gene identification was low in comparison to vertebrates, but was reasonably high for oyster ESTs as relatively low numbers of known genes are available in the GenBank from invertebrates, especially the molluscs. The largest number of significant hits came from comparisons with Drosophila genes. A search of the ESTs using 1,395 gene ontology terms indicated the presence of a number of genes involved in stress biology including several heat shock proteins, stress-induced proteins, and an antimicrobial peptide, defensin B. These ESTs should be useful for studies of environmental pollution and stress biology. cDNA microarrays are being developed for the assessment of differential gene expression under various environmental conditions.

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