紫苏FAD3基因的克隆与表达分析

Cloning and Expression Analysis of FAD3 from Perilla frutescens

ω-3脂肪酸脱氢酶(ω-3 FAD)是植物脂肪酸合成途径的关键限速酶,通过调节该酶的过量表达,可以使植物中的α-亚麻酸(ALA)含量增加。本研究采用RT-PCR的方法,以紫苏L1品系为材料,克隆得到一个ω-3脂肪酸脱氢酶基因PfFAD3。生物信息学分析结果显示PfFAD3开放阅读框为1 176 bp,编码319个氨基酸,推测其蛋白分子量为44.7 k D,等电点为9.19;PfFAD3较为保守,具有四个富含组氨酸的保守区域。q RT-PCR结果表明,PfFAD3基因具有组织表达特异性,在种子中的表达量远高于其它器官。同时,研究了外源Me JA和温度对紫苏茎和叶中PfFAD3基因表达的影响,结果显示外源Me JA能够快速诱导叶片中PfFAD3基因表达量的上调,而抑制PfFAD3基因在茎中的表达;低温能够诱导叶片中PfFAD3基因表达量上调,而高温则抑制叶与茎中PfFAD3基因的表达,表明PfFAD3基因可能参与紫苏的防御反应。

ω-3 fatty acid desaturase(ω-3 FAD) is a key rate-limiting enzyme of plant fatty acid synthesis pathway,by adjusting the overexpression of the enzyme, can make plants α- linolenic acid(ALA) content increasement. A c DNA sequence of ω-3 fatty acid desaturase from Perilla frutescens L1 was cloned by RT-PCR, named as PfFAD3.Bioinformatics analysis showed that PfFAD3 contains a 1 176 bp open reading frame(ORF) encoding a deduced protein of 391 amino acids with the estimated molecular weight and isoelectric point(PI) of the putative protein being 44.7 k D and 9.19, respectively; and it was quite conservative that contained four histidine-rich conserved regions. q RT-PCR analysis proved that PfFAD3 expressed in all tested organs, but the expression in seeds was much higher than other organs.Under exogenous Me JA and temperature treatment, we analyzed the expression of PfFAD3 in stems and leaves of Perilla frutescens, results indicated that exogenous Me JA could induce rapidly increase PfFAD3 expression in leaf, but inhibite PfFAD3 expression in stem.Low temperature can up-regulate PfFAD3 expression in leaf, but high temperature may down-regulate PfFAD3 expression in leaf and stem,suggesting PfFAD3 genes may involved in defense responses of Perilla frutescen.