猪PR-39抗菌肽基因在毕赤酵母中的表达

Expression of pig PR-39 antibacterial peptide gene in Pichia pastoris

[目的]在毕赤酵母中表达抗菌肽PR-39基因,获得有抗菌活性的PR-39。[方法]根据酵母和猪密码子偏好性,对其密码子进行优化改造。将经SOE-PCR获得的PR-39基因与毕赤酵母表达载体pPIC9K连接,构建重组载体pPIC9K-PR-39。经SacⅠ线性化电击转化毕赤酵母GS115,取阳性克隆进行髙拷贝转化子筛选和诱导表达。[结果]pPIC9K-PR-39重组质粒构建成功,pPIC9K-PR-39菌株发酵产物检测结果对DH5α大肠杆菌和金黄色葡萄球菌都有抑菌效果。[结论]获得了PR-39基因的重组酵母,并用毕赤酵母系统成功地分泌表达了具有明显抗菌活性的抗菌肽PR-39。

[ Objective] Expression of pig antimicrobial peptide PR -39 gene in Pichia Pastoris, and obtaining PR -39 with antimicrobial activity. [ Methods ] According to the partiality codon of yeast and pig, the codons were optimized. The PR - 39 gene was amplified by SOE - PCR and ligated to pPIC9K to construct the recombinant expression vector pPIC9K - PR - 39. The recombinant vector was linearized by Sac I ,and transformed into GSll5 by electroporation. The positive clones were screened for high copy transformants and induced by methanol. [ Results] The recombinant carrier pPIC9K -PR -39 has been constructed, and the detection of pPICgK - PR - 39 strain fermentation products shows that the PR - 39 peptide has the antibacterial activity against E. coli and S. aureus. [ Conclusion ] The PR - 39 recombinant yeast has been obtained and the antibacterial peptide PR - 39 with obvious activity has been successfully expressed in Pichia pastoris system.