本文選題：大豆 + MsDREB　； 參考：《植物生理學(xué)報》2017年08期

【摘要】：本研究目的是利用轉(zhuǎn)基因技術(shù)改良大豆(Glycine max)的耐旱性,并研究rd29A和CaMV-35S兩類啟動子的驅(qū)動效果。利用構(gòu)建的轉(zhuǎn)基因載體p CAMBIA-rd29A-MsDREB1和p CAMBIA-35S-MsDREB1,通過農(nóng)桿菌(Agrobacterium tumefaciens)介導(dǎo)法,將苜蓿(Medicago sativa)基因DREB1導(dǎo)入大豆品種‘中黃13號’,獲得rd29A和CaMV-35S兩類啟動子驅(qū)動的MsDREB1轉(zhuǎn)基因大豆。對T_1至T_2代植株進行PCR、Southern blot分析,分別篩選到9和12個轉(zhuǎn)基因大豆株系,各隨機選擇兩個轉(zhuǎn)基因株系作為研究對象。正常水分狀態(tài)下初花期統(tǒng)計大豆株高及葉面積。苗齡30 d的植株在不同干旱脅迫條件下,用逆轉(zhuǎn)錄定量PCR(RT-q PCR)分析基因表達差異,測定葉綠素含量、丙二醛含量、相對含水量及植株干重,并分析各株系干旱后復(fù)水的成活率。結(jié)果表明,兩種啟動子對MsDREB1表達的調(diào)控存在明顯差異,在非脅迫下35S啟動子調(diào)控的MsDREB1為超量表達,而rd29A啟動子調(diào)控的MsDREB1表達量較低;在嚴(yán)重干旱脅迫下,rd29A:MsDREB1表達量高于35S:MsDREB1表達量;MsDREB1超量表達抑制植株正常生長。兩種啟動子各轉(zhuǎn)基因株系均有一定耐旱能力,但存在差異。MsDREB1誘導(dǎo)表達耐旱性效果更明顯,在中度干旱脅迫下,其植株相對含水量、葉綠素含量、單株干重均顯著高于MsDREB1超量表達,而丙二醛含量顯著低于MsDREB1超量表達。結(jié)果說明MsDREB1作為轉(zhuǎn)錄調(diào)節(jié)因子參與了植物的干旱調(diào)節(jié)。該研究為MsDREB1基因在大豆耐旱基因工程中的應(yīng)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.
【作者單位】： 黃淮學(xué)院生物與食品工程學(xué)院;
【基金】：河南省科技發(fā)展計劃項目(112300410042)~~
【分類號】：S565.1

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