卷枝毛霉Δ12-脱饱和酶基因的克隆及其在酿酒酵母中的高效表达  

Cloning and high expression of Δ12- desaturase genes from Mucor circinelloides EIM-10

作　　者：刘小琳[1,2] 江贤章[1,2] 张明亮[1,2] 黄建忠[1,2] 

机构地区：[1]福建师范大学闽南科技学院生命科学与化学系,福建泉州362332 [2]福建师范大学生命科学学院福建省现代发酵技术工程研究中心工业微生物教育部工程研究中心,福建福州350108

出　　处：《食品工业科技》2015年第21期196-199,213,共5页Science and Technology of Food Industry

基　　金：福建省发改委产业化项目(闽教发[2010]77号)

摘　　要：Δ12-脱饱和酶是亚油酸合成的关键酶,为提高Δ12-脱饱和酶的活性,本研究利用RT-PCR对Mucor sp.EIM-10的Δ12-脱饱和酶c DNA进行克隆,并导入p YES2.0质粒中,构建重组表达载体。运用醋酸锂转化法将重组质粒导入酿酒酵母,成功构建p YMD12酿酒酵母表达系统。为了进一步提高Δ12-脱饱和酶的表达水平,用GAP启动子替代p YES2.0质粒自带的启动子,成功构建p YGAPMD12重组菌。最终产物经GC-MS检测显示,p YGAPMD12重组菌转化C18∶1的转化率为69.172%,比p YMD12重组菌提高32.771%。本文为进一步提高Δ12-脱饱和酶的表达水平提供参考依据。△12-desaturase plays an important role in the biosynthesis of linoleic acid in Mucor sp ElM-10, To improve enzymatic on the △12-desaturase activity, a recombinant expression vector was constructed.The △12- desaturase gene was cloned by RT-PCR technology.The PCR products were ligated into pYES2.0 plasmid.And then the recombinant plasmid （pYMD12）was transformed into Saccharomyces cerevisiae strain INVSc1 by lithium acetate method.To improve enzymatic activity, built-in promoter of pYES2.0 was replaced by GAP promoter, another new recombinant expression（pYGAPMD12） were constructed in Saccharomyces cerevisiae strain INVScl. Gas chromatography analysis showed that conversion efficiency of pYGAPMD12 was 69.]72% and the conversion efficiency of pYMD12 was 36.4-01%. This paper was provided important reference for further stay of △12- desaturase.

关 键 词：Δ12-脱饱和酶 GAP启动子 酿酒酵母 卷枝毛霉 

分 类 号：TS201.3[轻工技术与工程—食品科学]