Construction And Expression Of Sheep -β-defensin-1 Recombinant Prokaryotic Expression Vector

Beta defense is an important member of the family of antimicrobial peptides, which is widely present in the mucosal epithelial tissues of various organs of birds and mammals. As the body of the first immune defense, the beta defense element biological activity are widely, can directly kill the pathogen with a broad spectrum antibacterial, antiviral, anti cell toxicity and regulate immune function, animal organism to resist the invasion of pathogenic microorganisms of effective natural barrier, at that moment as a substitute for antibiotics, in the prevention and treatment of animal inflammatory hot, is expected to become new anti infective drugs. In this paper, we constructed and prokaryotic expression and protein purification of sheep beta-defense mature peptide msBD-1 by genetic engineering.For the expression of sheep-β-defensin-1 mature peptide (msBD-1) protein, the study according to the article published in GenBank msBD-1 sequence ends with restriction sites BamHI and Xhol, codon optimization, using PCR technology to amplified fragment msBD-1 and cloned into the vector pGEX-4T-1, and successively by bacteria PCR, restriction enzyme digestion and sequencing, fragment confirmed successful cloning. Further, the recombinant plasmid was transformed into transB (DE3) competent cells. After IPTG induction, the recombinant plasmid was highly fused and expressed msBD-1 protein. To increase the amount of protein expression, experiments by varying the concentration of IPTG induced and induction time, to determine the best conditions for inducing protein expression. At the same time, by Western Blot analysis derived recombinant proteins and GST-tag antibody can produce specific reaction. The results show the success of the msBD-1 protein expression, and the msBD-1 protein was purified.