摘要
为研究油葵FAD2-1基因在脂肪酸合成中的催化功能,利用RT-PCR技术,从油葵品种新葵杂4号未成熟种子中克隆HaFAD2-1基因的全长cDNA序列。将该基因序列构建穿梭表达载体pYES2-HaFAD2-1,并转化到营养缺陷型酿酒酵母菌株INVSc1中。将重组酵母诱导培养后对其总脂肪酸进行GC-MS分析。结果显示,HaFAD2-1基因编码一个长378个氨基酸的脂肪酸去饱和酶蛋白,分子质量43 719,等电点(pI)为8.38,具有3个高度保守的组氨酸簇。脂肪酸甲酯气相色谱分析结果表明HaFAD2-1基因在酿酒酵母中获得表达,能将油酸转化为亚油酸,证明克隆得到的HaFAD2-1具有完整的催化功能。
To study the catalytic function of oil sunflower FAD2-1 gene in fatty acid synthesis, a full length cDNA of FAD2 named HaFAD2-1 was isolated from developing seeds of Helianthus annuus L.Xinza 4 by RT-PCR.The HaFAD2-1 gene encodes a-378aa oleic acid desaturase with predicted molecular weight of 43 719, isoelectric point (pI) value of 8.38 and three his-boxes.HaFAD2-1 gene was sub-cloned into the shuttle vector pYES2 to generate the recombinant vector pYES2-HaFAD2-1 which was transformed into a defective mutant INVSc1 strain of Saccharomyces cerevisiae for expression.The total fatty acids of recombinant yeast were analyzed by GC-MS after induced culture.The result showed that transgenic yeast with the vector pYES2-HaFAD2-1 could catalyze oleic acid into linoleic acid, indicating that HaFAD2-1 is a functional enzyme of FAD2.
出处
《江苏农业学报》
CSCD
北大核心
2017年第2期-,共7页
Jiangsu Journal of Agricultural Sciences
基金
国家自然科学基金项目(31360052)
兵团博士基金项目(2012BB005)
关键词
油葵
脂肪酸去饱和酶
基因克隆
酿酒酵母转化
功能分析
oil sunflower
fatty acid desaturase
gene clone
transgenic Saccharomyces cerevisiae
function analysis
