细菌来源甲基对硫磷水解酶基因在酵母中的优化与表达
首发时间:2016-05-17
摘要:对Plesiomonas sp.M6来源的甲基对硫磷水解酶基因进行去掉N端信号肽序列和密码子优化设计,人工合成该基因,通过生物砖技术分别构建该基因的1-4拷贝毕赤酵母表达载体,获得的载体通过电转化方式导入毕赤酵母细胞。通过对表达产物的鉴定和酶学性质研究结果表明:经优化改造后的甲基对硫磷水解酶基因在毕赤酵母细胞成功表达,摇瓶发酵结果显示,该酶表达量达到1.9U/ml,是已报导水平的10倍。酶活性达到15.83U/mg。携带不同拷贝甲基对硫磷水解酶基因的酵母表达菌株在该酶的表达水平上无显著差异。
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Optimization and expression of mph from Plesiomonas sp.M6 in Pichia pastoris
Abstract:In this paper, the modified mph of Plesiomonas sp.M6 was synthesized which codon was optimized and the N-signal peptide sequence was removed. The expressing vector of P.pastoris carrying 1-4 mph expressing cassettes seprately,have been constructed by biological brick technology, and the recombinant P.pastoris have been obtained by electroporation. Results showed that the modified mph had been expressed in P.pastoris by identifying the expression product.The expression level reached 1.9U/ml by shaking culture and the level of enzyme activity reached 15.83U/mg, which is 10 times comparing to the public reported . The expressing levels of mph showed no obvious difference among the recombinant strains carrying different copies of mph expression cassettes .
Keywords: methyl parathion hydrolase Pichia pastoris optimization and expression
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No.4688834987145146****
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