深黄被孢霉Δ~6-脂肪酸脱氢酶基因在大豆中的表达

Heteologous Expression of Mortierella isabellina Δ~6-Fatty Acid Desaturase Gene in Soybean

为在传统的油料作物大豆中产生γ 亚麻酸 ,从深黄被孢霉中克隆的Δ6 脂肪酸脱氢酶基因与植物表达载体pBI12 1连接 ,构建了重组质粒pBMICL6 ,采用农杆菌介导的大豆子叶节转化系统成功的将该基因导入到栽培大豆吉林 35、吉林 4 3、吉林 4 7、绥农 10、绥农 14和黑农 37等品种中 ,获得一批转基因植株。经PCR检测和Southern杂交分析 ,证明外源基因已导入并整合到大豆的基因组中。Northern杂交结果表明该基因在转基因大豆的mRNA水平上获得表达。对转基因大豆种子进行脂肪酸成分分析 ,结果表明Δ6 脂肪酸脱氢酶基因获得表达 ,产生了γ 亚麻酸 ,其含量最高可达 2 7 0 6 7% ,这是国内外深黄被孢霉Δ6

linolenic acid (GLA, C18∶3Δ^(6,9,12)) is one of nutritionally important polyunsaturated fatty acids in human and animal diets. Vast majority oilseeds crops do not produce GLA. Δ~6-fatty acid desaturase (D6D) is the rate-limiting enzyme, which catalyzes the conversion of linoleic acid (LA) and α-linolenic acid (ALA) to GLA and octadecatetraenoic acid (OTA). In this study, a Δ~6-fatty acid desaturase gene isolated from Mortierella isabellina was transformed into some soybean cultivars Jilin 35, Jilin 47, Suinong 10, Suinong 14 and Heinong 37 successfully by agrobacterium-mediated cotylendon node transformation system. The results of PCR analysis, Southern blotting and Northern blotting with transgenic plants indicated that target gene was integrated into transgenic soybean genome and expressed at the level of RNA. Meanwhile, total fatty acids were extracted from transgenic seeds and methyl-esterified. Analysis of gas chromatograph (GC) showed that a novel peak corresponding to the standard of GLA methyl ester was detected. the highest percentage of γ-lenolenic acid to total fatty acids in transgenic seeds was 27.067%, which indicated that Mortierella isabellina Δ~6-fatty acid desaturase gene was expressed in transgenic soybean. It is so far the first report on the expression of Δ~6-fatty acid desaturase gene in transgenic soybean in the world.