过表达苜蓿MsDREB1基因大豆耐旱性分析

发布时间：2018-06-09 00:01

  本文选题：大豆 + MsDREB　； 参考：《植物生理学报》2017年08期

【摘要】：本研究目的是利用转基因技术改良大豆(Glycine max)的耐旱性,并研究rd29A和CaMV-35S两类启动子的驱动效果。利用构建的转基因载体p CAMBIA-rd29A-MsDREB1和p CAMBIA-35S-MsDREB1,通过农杆菌(Agrobacterium tumefaciens)介导法,将苜蓿(Medicago sativa)基因DREB1导入大豆品种‘中黄13号’,获得rd29A和CaMV-35S两类启动子驱动的MsDREB1转基因大豆。对T_1至T_2代植株进行PCR、Southern blot分析,分别筛选到9和12个转基因大豆株系,各随机选择两个转基因株系作为研究对象。正常水分状态下初花期统计大豆株高及叶面积。苗龄30 d的植株在不同干旱胁迫条件下,用逆转录定量PCR(RT-q PCR)分析基因表达差异,测定叶绿素含量、丙二醛含量、相对含水量及植株干重,并分析各株系干旱后复水的成活率。结果表明,两种启动子对MsDREB1表达的调控存在明显差异,在非胁迫下35S启动子调控的MsDREB1为超量表达,而rd29A启动子调控的MsDREB1表达量较低;在严重干旱胁迫下,rd29A:MsDREB1表达量高于35S:MsDREB1表达量;MsDREB1超量表达抑制植株正常生长。两种启动子各转基因株系均有一定耐旱能力,但存在差异。MsDREB1诱导表达耐旱性效果更明显,在中度干旱胁迫下,其植株相对含水量、叶绿素含量、单株干重均显著高于MsDREB1超量表达,而丙二醛含量显著低于MsDREB1超量表达。结果说明MsDREB1作为转录调节因子参与了植物的干旱调节。该研究为MsDREB1基因在大豆耐旱基因工程中的应用提供方法。
[Abstract]:The purpose of this study was to improve the drought tolerance of Glycine max by transgenic technique and to study the driving effect of rd29A and CaMV-35S promoters. Using the constructed transgenic vectors pCAMBIA-rd29A-MsDREB1 and p CAMBIA-35S-MsDREB1, the gene DREB1 of Medicago sativa was introduced into soybean variety 'Zhonghuang 13' by Agrobacterium tumefaciens (Agrobacterium tumefaciens). Two kinds of promoter driven MsDREB1 soybean, rd29A and CaMV-35S, were obtained. Southern blot analysis was carried out on the plants of T _ 1 to T _ 2, and 9 and 12 transgenic soybean lines were screened, and two transgenic lines were randomly selected as the research objects. The plant height and leaf area of soybean at early flowering stage were calculated under normal water condition. Under different drought stress conditions, the difference of gene expression, chlorophyll content, malondialdehyde content, relative water content and plant dry weight were analyzed by reverse transcription quantitative PCRRT-q PCR, and the survival rate of rehydration after drought was analyzed. The results showed that there was significant difference between the two promoters in regulating the expression of MsDREB1. The expression of MsDREB1 regulated by 35s promoter was overexpressed under non-stress, but the expression of MsDREB1 regulated by rd29A promoter was lower. Under severe drought stress, the expression of rd29A: MsDREB1 was higher than that of 35s: MsDREB1. The overexpression of MsDREB1 inhibited the normal growth of plants. The transgenic lines of the two promoters have some drought tolerance, but there is a difference. MsDREB1 induces the drought tolerance more obviously. Under moderate drought stress, the relative water content and chlorophyll content of the transgenic lines are higher than that of the control. The dry weight per plant was significantly higher than that of MsDREB1, while the malondialdehyde content was significantly lower than that of MsDREB1. The results showed that MsDREB1 was involved in drought regulation as a transcription regulator. This study provides a method for the application of MsDREB1 gene in soybean drought tolerance gene engineering.
【作者单位】： 黄淮学院生物与食品工程学院;
【基金】：河南省科技发展计划项目(112300410042)~~
【分类号】：S565.1

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