期刊论文

中文

华北农学报

1000-7091

2017

32

6

121-125

10.7668/hbnxb.2017.06.018

湖南农业大学生物科学技术学院大学生创新项目

本校为第一机构

为了以酿酒酵母S78为宿主菌异源高效表达糖化酶基因,进一步扩大糖化酶基因在工业生产上的应用。运用RT-PCR法从黑曲霉中克隆得到糖化酶基因(glaA) cDNA,去除其信号肽编码区后的序列重组到酵母表达载体 pVT102U/α ADH1强启动子下游,并与α因子分泌肽信号序列融合。用PEG/LiAc法将构建的重组表达载体转入酿酒酵母S78菌株,筛选出的转化菌点种到可溶性淀粉平板上培养,用碘染法鉴定重组基因的表达情况。鉴定出了典型水解圈的酵母转化子,转化子接种到YPD培养基中摇瓶培养后,取发酵上清液经SDS-PAGE检测到分子量大小约为80 kDa的目标蛋白带,上清液用DNS法检测其淀粉酶最高酶活为28. 86 IU/mL。表明糖化...

In order to express the glucoamylase gene efficiently with Saccharomyces cerevisiae S78 as a host strain and to further expand the application of glaA gene in industrial production. The glaA gene was cloned from Aspergillus niger by RT-PCR,and the sequence removed the natural signal peptide coding region was recombined in-to the downstream of the yeast expression vector pVT102 U/αADH1 strong promoter and fused with the sequence ofα factor secretion peptide signal. The recombinant expression vector was transformed into Saccharomyces cerevisiae...