甘蓝型油菜BnFAD2-C1基因全长序列的克隆、表达及转录调控元件分析

Cloning and Expression of BnFAD2-C1 Gene Involved in Brassica napus and Analysis of Transcription Regulation Elements

脂肪酸去饱和酶基因(FAD2)是控制植物体中油酸含量的关键基因,在甘蓝型油菜中有4个FAD2基因的拷贝,分别定位在A1、C1、A5、C5染色体上。本文克隆了1个FAD2拷贝基因,依据油菜基因组数据库信息,将其定位到C1染色体上,命名为Bn FAD2-C1,其开放阅读框为1155 bp。采用RACE(rapid-amplification of c DNA ends)技术获得了175 bp的5?UTR序列和212 bp的3?UTR序列。采用荧光定量PCR技术研究发现,Bn FAD2-C1在根、花和角果皮中仅保持本底水平的表达,在种子发育中期呈现高效表达,具有种子特异性诱导表达的特征。根据甘蓝和油菜基因组数据库信息,同源克隆到Bn FAD2-C1基因的启动子(promoter)和内含子(intron)序列,并通过PLACE和Plant CARE网站分析,初步预测到调控该基因转录的潜在顺式作用元件。通过茉莉酸诱导处理,Bn FAD2-C1基因表达量发生变化,推断茉莉酸在Bn FAD2-C1基因的表达过程中可能发挥一定的调控作用。

Fatty acid desaturase gene （FAD2） is a key factor in regulating oleic acid content. There are four copies ofFAD2 lo-cated on chromosomes A1, C1, A5, and C5 inBrassica napus. One copy containing 1155 bp openreading frame was cloned with previous research method and named asBnFAD2-C1, which was location on chromosome C1 based on the genome database in-formation of oleracea and oilseed. The untranslated regions （UTR） of 5′ and 3′ end with 175 bp and 212 bp length respectively were cloned by RACE （rapid-amplification of cDNA ends） technique. The expression pattern ofBnFAD2-C1 gene was identified using quantity PCR technique, showing a seed-specific inducible expression in mid developmental seeds and a background-level expression in root, flower and siliqua wall. The promoter and intron region were also cloned and analyzed using PLACE and PlantCARE websites to predict some potential cis-elements in regulatingBnFAD2-C1gene transcription. At the same time, jas-monic acid （JA） was inferred to make certain contributions to regulateBnFAD2-C1gene expression showing a changeable ex-pression quantity when treated with jasmonic acid.