【摘要】 选用酵母偏好密码子,设计合成一种酸性β-半乳糖苷酶基因,并研究其在毕赤酵母中的高效表达。优化后,密码子适应指数(CAI)由原来的0.61提升到0.85。在基因的5’端融合编码α信号肽序列,在3’端融合编码6个组氨酸标签序列,并在N端编码α信号肽和编码成熟酸性β半乳糖苷酶基因间分别引入编码kex2和Ste13裂解信号序列。基因克隆到pPICZα-A表达载体,构建成分泌型重组酵母表达载体pPICZα-β-Gal。在醇氧化酶(AOX1)启动子调控下,酸性β-半乳糖苷酶得到高效分泌表达,达到508 mg/L,比优化前提高3倍。重组酸性β-半乳糖苷酶具有天然的酸性β半乳糖苷酶相同的酶活性。更多还原

【Abstract】 An acid Lac Z was designed by using yeast preference codon to study its expression in Pichia Pastoris Vector.After optimizing,the codon adaptation index(CAI) rose from 0.61 to 0.85.In this experiment,we linked fusion coding α signal peptide sequence to 5’-end of the gene,and linked six His tag sequences to 3’-end of the gene.In addition,at N-end,kex2 and Ste13 cracking signal sequences were inserted between α signal peptide sequence and acid Lac Z gene.This gene was cloned to pPICZα-A expression vector to be a secretory restructuring yeast expression vector.Under regulate and control of alcohol oxidase(AOX1) promoter,acid β-galactosidase is obtained high expression to 508 mg/L which is three times than before.This recombination acid β-galactosidase owns the same enzymatic activity as the nature one.更多还原