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Molecular cloning and expression of two isoforms of Vasa gene in Southern catfish Silurus meridionalis

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Author(s): HU Chong-Jiang | WU Feng-Rui | LIU Zhi-Hao | HUANG Bao-Feng | ZHANG Yao-Guang | WANG De-Shou

Journal: Acta Zoologica Sinica
ISSN 0001-7302

Volume: 54;
Issue: 6;
Start page: 1051;
Date: 2008;
Original page

Keywords: Southern catfish | Vasa | Gene cloning | Alternative splicing | Expression | RT-PCR | in situ hybridization

ABSTRACT
Two isoforms of Vasa cDNA, derived from the 5′alternative splicing of the same gene, were isolated and characterized in Southern catfish, Silurus meridionalis, by reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) methods. Analysis of the nucleotide sequences revealed that the full length cDNA of Southern catfish Vasa (scVasa) comprises 2525 base pairs (bp) with an open reading frame (ORF) of 1989 bp, encoding 662 amino acids, while that of Vasa short form (scVasa-s) comprises 2438 bp with an ORF of 1926 bp, encoding 641 amino acids. Except the difference in 5′-untranslated region, scVasa-s also lacks a part of the 5′-ORF region found in the scVasa. Both of the two deduced amino acid sequences contain all the eight conserved motifs characteristic to the DEAD-box protein family and four arginine-glycine-glycine motifs unique to Vasa proteins. scVasa showed the highest similarity to Vasa homolog of giebel carp (73.3%). Tissue distribution analysis by RT-PCR revealed that these two isoforms were exclusively expressed in the gonads of both sexes. In adult fish, scVasa was found to be mainly expressed in the primary oocytes at phase Ⅰ and Ⅱ in the ovary while in the spermatogonia and primary spermatocytes in the testis by in situ hybridization. Semi-quantitative RT-PCR analysis showed that the expressions of both scVasa isoforms were much higher in ovarian recrudescent stage (mainly with phase Ⅱ oocytes) than in vitellogenic stage (mainly with phase Ⅲ and Ⅳ oocytes) [Acta Zoologica Sinica 54(6):1051–1060, 2008].
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