The Cloning, Expression Analysis And Construction Of A Gene Targeting Vector Of Myostatin (MSTN) From Lateolabrax Japonicus

In this study, MSTN gene was cloned from sea perch by homology cloning and genomic walking. In the 5396 bp genomic sequences, three exons, two introns, 5'and 3'flanking sequences were identified. The sea perch MSTN gene encodes a 374 amino acid protein, including a signal peptide, conserved cysteine residues and a RXXR proteolytic cleavage domain. Expression analysis of MSTN revealed that MSTN was expressed highly in muscle, brain and eyes, intermediately in intestine, and weakly in gill, spleen, liver, and heart. In early embryonic stages, the transcript was undetectable till 15 days larvae after hatching by RT-PCR. It was demonstrated that MSTN mRNA was highly expressed in embryonic stem cell line (LJES1), but it was undetectable in several types of somatic cell lines from sea perch, including fibroblast-like cell, epithelioid cells, lymphocyte-like cells. Furthermore, a pMSTN-GFP plasmid driven by the 1125 bp MSTN 5'-UTR was constructed, which could express green fluorescent protein (GFP) in LJES1 cells. By microinjection, pMSTN-GFP plasmids were transferred into one-cell embryo of zebrafish, and green fluorescent signals were detected in trunk of 15 d fry.For improving the transformation efficiency of exogenous gene into ES cells, a simple and effective method of gene transfer was established with several lipsomes-mediated. Transformation efficiencies of three lipsomes-mediated...