大豆微粒体油酸盐脱饱和酶FAD2基因的克隆及其植物表达载体的构建

Cloning of Glycine Max FAD2 Gene and Construction of Plant Expression Vector

从大豆种子中提取RNA,通过逆转录PCR的方法扩增出1307bp的大豆微粒体油酸盐脱饱和酶FAD2基因,并将其克隆到pMD18-T vector,DNA测序分析表明克隆的片段与原序列相似性达到99.9%,蛋白质相似性达到100%,显示其与同科植物花生同源性最高,与其它植物FAD2的氨基酸序列同源性较低。实验室前期从锦橙中克隆了种子球蛋白基因启动子ssp,转化烟草证明具有种子特异性调控表达的特性。本研究利用ssp启动子和大豆的FAD2基因构建了在种子专一性启动子驱动下的植物表达载体p3300-ssp-FAD2-Tons,为进一步进行油料作物的脂肪酸基因工程奠定了基础。

The Glycine max FAD2 Gene was amplified from Glycine max total RNA by reverse transeription PCR, and was cloned into pMD18-T vector. DNA sequence analysis indicated that the cloned 1307bp DNA fragment has the highest homology compared with the published data, showing 99.9% and 100% homology at the nucleotide and at the amino acid sequence respectively. The results of sequence analysis indicate that Glycine max FAD2 gene is was most similar to archis hypogaea and the comparability is lower compared with amino acid of others plants. Our laboratory has cloned promoter of seed globin gene in Citrus sinensis Osbeck cv. It has been proved that the promoter has the characteristic of seed speciality expression in transformed tobacco. FAD2 gene plant expression vector has been constructed. It is the basis of fatty acid gene engineering of oil crop.