高山被孢霉ATCC16266Δ~6-脂肪酸脱氢酶基因在酿酒酵母中的表达  

Expression of Δ~6-fatty Acid Desaturase Gene from Mortierella alpina in Saccharomyces cerevisiae

作　　者：刘莉[1] 李明春[1] 胡国武[1] 葛军[1] 张丽[1] 程志晖[1] 邢来君[1] 

机构地区：[1]南开大学微生物系,天津300071

出　　处：《生物工程学报》2001年第2期161-164,共4页Chinese Journal of Biotechnology

基　　金：国家自然科学基金!项目 (39870 0 2 0 );高等学校骨干教师资助计划项目

摘　　要：Δ6 脂肪酸脱氢酶是形成γ 亚麻酸的关键酶。从含有高山被孢霉Δ6 脂肪酸脱氢酶基因的重组质粒pT MACL6中 ,酶切出 1 4kb的目的片段 ,亚克隆到大肠杆菌和酿酒酵母的穿梭表达载体 pYES2 .0 ,在大肠杆菌中筛选到含有目的基因的重组质粒 pYMAD6 ,用醋酸锂方法转化到酿酒酵母的缺陷型菌株INCSc1中 ,在SC Ura合成培养基中 ,选择得到酿酒酵母工程株YMAD6。在合适的培养基及培养条件下 ,加入外源底物亚油酸 ,经半乳糖诱导后 ,收集菌体。通过GC MS对酵母工程株进行脂肪酸色谱分析 ,结果表明 ,产生了 31 6 %的γ 亚麻酸。这是迄今为止 ,国内外Δ6 脂肪酸脱氢酶基因在酿酒酵母中表达量最高的报道。fatty acid desaturase is the rate\|limiting enzyme of the desaturation of linoleic acid in the production of an essential fatty acid,γ\|linolenic acid.The 1.4kb fragment in plasmid pTMACL6 encoding Δ 6 fatty acid desaturase from \%Mortieralla alpina\% ATCC16266 was subcloned into the yeast\|\%E.coli\% shuttle vector pYES2.0,thus an expression recombinant plasmid pYMAD6 contatining target gene was constructed and obtained in the SC\|Ura media.The pYMAD6 was introduced into defective mutant INCSc1 of \%Saccharomyces cerevisiae \%by LiAc method.When linoleic acid was provided as an exogenous substrate to the yeast cultures expressing Δ 6\|fatty acid desaturase activity under appropriate media and temperature condition,the level of γ\|linolenic acid reached 31.6% of the total yeast fatty acids by GC\|MS detecting,which is the highest report of Δ 6 fatty\|acid desaturase gene in \%S.cerevisiae.\%

关 键 词：△^6-脂肪酸脱氢酶基因 Γ-亚麻酸 酿酒酵母 表达 高山被孢霉 

分 类 号：Q786[生物学—分子生物学]