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作物学报 ›› 2015, Vol. 41 ›› Issue (11): 1663-1670.doi: 10.3724/SP.J.1006.2015.01663

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

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

刘芳**,刘睿洋**,彭烨,官春云*   

  1. 湖南农业大学农学院 / 国家油料改良中心湖南分中心, 湖南长沙 410128
  • 收稿日期:2015-04-04 修回日期:2015-07-20 出版日期:2015-11-12 网络出版日期:2015-08-11
  • 基金资助:

    本研究由湖南省科技创新基金项目(CX2013A012)和湖南省科技重大专项(2014FJ1006)资助。

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

LIU Fang**,LIU Rui-Yang**,PENG Ye,GUAN Chun-Yun*   

  1. College of Agronomy, Hunan Agricultural University / National Oilseed Crops Improvement Center in Hunan, Changsha 410128, China
  • Received:2015-04-04 Revised:2015-07-20 Published:2015-11-12 Published online:2015-08-11

摘要:

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。参照前人研究方法,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BnFAD2-C1,其开放阅读框为1155 bp。采用RACE (rapid-amplification of cDNA 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,BnFAD2-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BnFAD2-C1基因的启动子(promoter)和内含子(intron)序列,并通过PLACE和PlantCARE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,BnFAD2-C1基因表达量发生变化,推断茉莉酸在BnFAD2-C1基因的表达过程中可能发挥一定的调控作用。

关键词: 甘蓝型油菜, BnFAD2-C1基因, RACE, 生物信息学分析, 茉莉酸

Abstract:

Fatty acid desaturase gene (FAD2) is a key factor in regulating oleic acid content. There are four copies located on chromosomes A1, C1, A5, and C5 in Brassica napus. One copy containing 1155 bp open reading frame was cloned with previous research method and named as BnFAD2-C1 which was location on chromosome C1 based on the genome database information of oleracea and oilseed. Then 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 of BnFAD2-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 regulating BnFAD2-C1 gene transcription. At the same time, jasmonic acid (JA) was inferred to make certain contributions to regulate BnFAD2-C1 gene expression showing a changeable expression quantity when treated with Jasmonic acid.

Key words: Brassica napus, BnFAD2-C1, RACE, Bioinformatics analysis, Jasmonic acid

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