Cloning, Expression Of Myostatin And Preparation, Application Of Its Genetically Engineering Vaccine In Swine

As a negative regulator of skeletal muscle, Myostatin could influence the development and growth of muscle. The negative regulation of skeletal muscle should be eliminated for the rapid growth of muscle in the later period of growth. The genetically engineering vaccine of myostatin was prepared by using myostatin as an immunogen from thigh muscle of Shitou goose by engineered means and different immunological adjuvants. The swine was actively immunized by using the vaccine mentioned above to produce anti-myostatin antibody so as to rivalry myostatin in vivo and promote the muscle growth. Immune effects of vaccines with both oil suspension and liposome as adjuvants were compared, respectively in this experiment.1 Molecular Cloning of The Mature Peptide of Shitou Goose Myostatin and Its Expression in Escherichia coliBased on the cDNA nucleotide sequence of Shitou goose myostatin gene and the nucleotide sequence of mature peptide, a pair of primers were designed and synthesized to sub-clone the gene coding Shitou goose myostatin mature protein. The restriction enzyme digestion, BamH I and Sac I were added to the two primers repectively. Then the myostatin mature peptide DNA fragment was cloned into pMD18-T vector. Restriction enzyme digestion and PCR amplification confirmed the inserted fragment was myostatin mature peptide nucleotide sequence. The sub-cloned myostatin mature peptide gene has been successfully inserted to expression vector pRSETA. The result of DNA sequencing confirmed the structrue of recombinant expression plasmid was correct, which had the identities of 92%, 88%, 99% with the myostatin gene mature peptide nucleotide sequence of chicken (AF019621) , swine (AY208121) and domestic goose (AF440862) published in the Genebank respectively. But the amino acid sequence of mature protein translated had the identities of 100%, 100%, 99.1% with that of chicken, swine and domestic goose. The recombinant expression plasmid expressed in E.coli through the inducement of IPTG. The SDS-PAGE electrophoretic analysis showed the prokaryotic expression product was the cloned recombinant protein with molecular weight of 18.5KD.