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United States Department of Agriculture

Agricultural Research Service

Research Project: IMPROVING GREAT LAKES AQUACULTURE PRODUCTION Title: Analysis of Expressed Sequence Tags from the Liver and Brain of Estrogen Treated Yellow Perch (Perca flavescens)

Authors
item Goetz, F - GREAT LAKES WATER INST
item Binkowski, F - GREAT LAKES WATER INST
item Shepherd, Brian
item Goetz, G - GREAT LAKES WATER INST
item Rise, M - GREAT LAKES WATER INST

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: August 31, 2004
Publication Date: N/A

Technical Abstract: Over the last decade, there has been strong interest in the aquaculture of yellow perch (Perca flavescens) in North America. We have been developing genetically defined yellow perch broodstocks for aquaculture using genetic analyses and selection methods. However, aside from microsatellites, there are very few genomic resources for P. flavescens. To address this, we have initiated an expressed sequence tag (EST) analysis of genes derived from the liver and brain of control and estradiol 17 treated yellow perch. Estrogens have been previously reported to stimulate growth in yellow perch and, therefore, should also impact gene expression in relevant tissues. Juvenile yellow perch (10 gm) were treated in the diet with estradiol 17 (15 g/gm) or ethanol vehicle (control) for 9 weeks. At this time, livers, brain and white muscle were removed, flash frozen in liquid nitrogen and stored at -80oC. RNA was extracted from equal numbers of male and female perch livers and brain, and used to construct separate estrogen and control cDNA libraries in Zap Express (liver; pBK-CMV) and Zap (brain; pBluescript SK). Following in vivo excision, libraries were randomly plated and colonies were picked, grown and plasmid cDNA prepared for automated sequencing. We have sequenced 4,000 ESTs per library (estrogen-treated liver and brain and control liver and brain) resulting in a total of 16,000 ESTs. Sequencing reactions were run on an ABI 3730, and sequence chromatogram files were trimmed for quality using phred, vector screened using cross match, and analyzed locally using blastx against the NCBI nonredundant (nr) protein database and blastn against the NCBI nucleotide (nt) and EST (dbEST) databases. All sequences were also analyzed for redundancy using CAP3, and both contigs and singletons were annotated by blastx against the Gene Ontology (GO) database. Dietary estrogen treatment significantly enhanced growth of perch above that observed in controls and the enhancement was greater for females than males. When the ESTs were analyzed globally across estrogen-treated and control tissues, there were obvious differences in the appearance of certain genes and in the number of genes categorized under specific GO terms. For example, in the livers of estrogen-treated perch there was a large increase in lipid transporter and sugar binding genes, and a reduction in serine protease, oxidoreductase, and calcium binding genes when compared to control liver ESTs. Besides changes in GO term gene numbers, there were also very specific differences in genes observed between estrogen-treated and untreated fish. For example, from contig analysis, specific genes were observed in livers from estrogen-treated fish and include vitellogenin, chitinase, desaturase and serum lectin. The ESTs from the liver and brain, when combined with an additional 8,000 ESTs that we have sequenced from yellow perch ovaries, will begin to populate the gene database for yellow perch and facilitate efforts to genetically select for desirable traits in this species.

Last Modified: 4/23/2014
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