利用表达载体pYES2表达乙醇脱氢酶2(Alcohol dehydrogenase 2,ADH2)和乙醛脱氢酶6(Acetaldehyde dehydrogenase 6,ALDH6),从而实现将乙醇直接转化为乙酸.以啤酒酵母总DNA为模板,通过PCR获得adh2基因和aldh6基因.分别构建表达载体pYES2-adh2、pYES2-aldh6和pY-ES2-aldh6-adh2,并电击导入宿主菌INVScl中.PCR测序结果上传GenBank获得登录号为JX901290、JX901291,与已公布序列相似性分别为96％和99％.重组菌扩大培养后移入到诱导培养基中,进行诱导表达.在含2％半乳糖诱导培养基诱导下4h酶活达到最高,ADH酶活为0.49U/mg,是原始菌株的2.7倍,ALDH酶活为0.11 U/mg,为原始菌株的2倍.%In order to realize ethanol directly transform to acetic acid,the alcohol dehydrogenase 2 (adh2) and acetaldehyde dehydrogenase 6 (aldh6) genes were cloned and expressed.The genomic DNA of Saccharomyces cerevisiae as template was extracted and adh2 and aldh6 genes were obtained by PCR.Two genes were inserted into expression vector pYES2.Then the expression vectors pYES2-adh2,pYES2-aldh6 and pYES2-aldh6-adh2 were constructed.They were transformed to INVScl.Sequence tests showed that the acquired fragments exhibited 96％ and 99％ homology to adh2 and aldh6 gene sequences from GenBank report.Then these recombinants were grown on glucose and shift to inductive medium.The activities reached to the highest when the recombinants were induced for 4 hours.The activities of adh2 and aldh6 were 2.7-fold and 2-fold higher than those of the wild types,respectively.
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