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油酸去饱和酶FAD2(fatty acid desaturase2)在油酸中引第二个双键生成亚油酸,是负责植物体内产生多不饱和脂肪酸的第一步关键酶。本研究采用RT-PCR和RACE(rapid amplification of cDNA ends)技术,以中间锦鸡儿未成熟种子为材料,克隆了三个fad2基因(分别命名为fad2-2A、fad2-1A和fad2-1B)。将这三个基因与汪阳东从中间锦鸡儿枝叶中克隆的fad2-2B(CaFAD2基因,GenBank登录号AY957394)一起进行了序列比对和分析。序列同源性比较结果表明,fad2-1A与fad2-1B同源性很高,fad2-1A编码的前283个氨基酸与fad2-1B的同源性高达98.9%,fad2-2B与fad2-1A的氨基酸序列同源性最低,只有64.7%。这四个基因的成功克隆,为进一步研究基因分工、调控方式打下了基础,为实现对锦鸡儿属植物脂肪酸成分直接、精确调控提供了保证,也是从一个物种中克隆四个fad2基因的第一次报道。
Oleic acid desaturase2 (FAD2) leads the second double bond in oleic acid to generate linoleic acid, which is the first key enzyme responsible for the production of polyunsaturated fatty acids in plants. In this study, we cloned three fad2 genes (named as fad2-2A, fad2-1A and fad2-1B respectively) by using RT-PCR and RACE (rapid amplification of cDNA ends). The three genes were sequenced and analyzed with fad2-2B (CaFAD2 gene, GenBank accession number AY957394) cloned by Wang Yangdong from Caragana intermedia leaves. Comparison of sequence homology showed that fad2-1A had high homology with fad2-1B, the first 283 amino acids encoded by fad2-1A shared 98.9% homology with fad2-1B, the highest homology between fad2-2B and fad2-1A The amino acid sequence homology lowest, only 64.7%. The successful cloning of these four genes laid the foundation for further research on gene division and regulation and provided the guarantee for the direct and precise regulation of fatty acid composition of Caragana plants. It was also the first to clone four fad2 genes from one species Times reported.