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United States Department of Agriculture

Agricultural Research Service

2009 Annual Report

1a.Objectives (from AD-416)
A major constraint to increasing the production efficiency of the Nation's cool and cold water aquaculture industry is the lack of genetically improved strains of fish for aquaculture. There is only limited genetic information on traits that will enhance production efficiency and yield a better quality fish. Identification and characterization of genes affecting aquaculture production traits will facilitate the development of genetically improved strains to increase aquaculture production efficiency. The objectives of the research outlined in this agreement focus on the identification of genes affecting oocyte maturation and embryogenesis in rainbow trout. Characterization of these genes will result in more in-depth understanding of the biology of reproduction and embryonic development and will impact strategies for improving developmental characteristics in broodstock.

1b.Approach (from AD-416)
Collaborators at the USDA/ARS/NCCCWA and West Virginia University have worked to develop resources for conducting functional genomic research in rainbow trout. Resources which specifically target studies on embryogenesis include large volumes of expressed sequence tag data from oocyte and embryonic developmental stages and a cDNA panel representing oocytes thru day 50 post fertilization for gene expression analyses. Strategies will include analyses of gene expression and proteomic data in NCCCWA Broodstock to identify and characterize novel-oocyte specific genes, similar strategies will be employed to characterize gene expression throughout embryonic development.

3.Progress Report

In an effort to identify genes associated with muscle growth and flesh quality, we characterized the protein expression profile of degenerating muscle in rainbow trout during sexual maturation, the levels of 146 proteins changed significantly.

Micro RNAs (miRNAs) are short RNA sequences which control the expression of genes. We have identified 210 unique sequences representing 54 distinct micro RNAs from rainbow trout somatic tissues, and measured miRNA expression patterns in adult somatic tissues and unfertilized eggs. Single nucleotide polymorphisms (SNPs) were identified in the miRNAs and their target sites in rainbow trout genes. The miRNAs identified and characterized in this study provide a new tool for functional genome research in rainbow trout and other salmonids. To identify oocyte-expressed miRNAs in rainbow trout, we constructed a miRNA library from mature unfertilized oocytes. Expression analysis identified 2 miRNAs (miRNA-451 and a novel miRNA) that are expressed predominantly in mature oocytes and 3 novel miRNAs that were also expressed in somatic tissues. These 5 miRNAs were further analyzed for their expression during ovarian development. The high abundance of miRNA-451 in late vitellogenic oocytes suggests a role for this miRNA in early embryonic development.

Through analysis of ESTs (Expressed Sequence Tags)from an oocyte cDNA library, we identified a novel oocyte-specific transcript. The cDNA is 1966 bp in length containing an open reading frame which encodes a predicted protein of 514 amino acids. RNA expression analysis demonstrated that this novel gene is highly expressed during early previtellogensis. The expression of this gene in oocytes and its developmental pattern of expression suggest a role for this novel gene in the regulation of early oocyte and follicle development. We have also identified a new isoform of the rainbow trout 14-3-3E1 gene, which is lacking 17 amino acid residues at the C terminus. It shows tissue-specific as well as stage-specific RNA expression during ovarian development and early embryogenesis. Analysis by yeast two-hybrid system demonstrated that the new isoform interacts with a number of proteins, indicating that the new isoform is a functional protein. The ADODR is in frequent contact with the cooperator through phone calls, email, and annual site visits in addition to receipt of written reports.

Last Modified: 5/26/2015
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