本文關(guān)鍵詞： 野生大豆 GsWRKY 肇東苜蓿 農(nóng)桿菌 耐堿性　出處：《作物學(xué)報》2017年09期 　論文類型：期刊論文

【摘要】：WRKY蛋白屬于鋅指型轉(zhuǎn)錄調(diào)控因子,能夠參與植物多種逆境響應(yīng)。本研究利用前期野生大豆鹽堿脅迫RNA-seq測序數(shù)據(jù),從構(gòu)建的堿脅迫基因調(diào)控網(wǎng)絡(luò)中篩選并克隆到GsWRKY15基因。分析GsWRKY15在堿脅迫下野生大豆根中的表達模式,發(fā)現(xiàn)該基因受堿脅迫誘導(dǎo)顯著上調(diào)表達,且在脅迫后1 h表達量最高。分析GsWRKY15基因在野生大豆各組織中的表達特異性,發(fā)現(xiàn)該基因在各組織中均有表達,花中表達量最高。采用根癌農(nóng)桿菌侵染苜蓿子葉節(jié)方法,將GsWRKY15轉(zhuǎn)化肇東苜蓿,獲得39株抗性植株。通過PCR、Southern blot和RT-PCR方法分析抗性植株,獲得了超量表達GsWRKY15基因的轉(zhuǎn)基因株系并對其進行了耐堿性分析。在150 mmol L 1 Na HCO3處理2周后轉(zhuǎn)基因苜蓿生長狀態(tài)良好,而非轉(zhuǎn)基因苜蓿出現(xiàn)萎蔫、變黃,甚至死亡;非轉(zhuǎn)基因苜蓿的相對質(zhì)膜透性和丙二醛含量顯著高于轉(zhuǎn)基因苜蓿,而葉綠素含量顯著低于轉(zhuǎn)基因苜蓿;同時分析堿脅迫下轉(zhuǎn)基因植株中脅迫相關(guān)基因的表達模式,發(fā)現(xiàn)H+-Ppase、NADP-ME、KIN1、RD29A基因的表達量高于非轉(zhuǎn)基因苜蓿。結(jié)果表明GsWRKY15基因的超量表達能夠顯著增強苜蓿的耐堿能力。
[Abstract]:WRKY protein is a zinc-finger transcription regulator and can participate in many stress responses of plants. In this study, RNA-seq sequencing data of wild soybean saline-alkali stress were used. The GsWRKY15 gene was screened and cloned from the constructed alkali stress gene regulatory network. The expression pattern of GsWRKY15 in the root of wild soybean under alkali stress was analyzed. It was found that the gene was significantly up-regulated by alkali stress. After 1 hour of stress, the expression of GsWRKY15 gene was the highest in all tissues of wild soybean. The results showed that the GsWRKY15 gene was expressed in all tissues and the highest in flower. Agrobacterium tumefaciens was used to infect cotyledons of alfalfa by Agrobacterium tumefaciens (Agrobacterium tumefaciens). 39 resistant plants were obtained by transforming GsWRKY15 into alfalfa, and the resistant plants were analyzed by Southern blot and RT-PCR. Transgenic lines with overexpression of GsWRKY15 gene were obtained and their alkaline tolerance was analyzed. After treatment with 150 mmol L ~ (-1) Na HCO3 for 2 weeks, transgenic alfalfa grew well, while non-transgenic alfalfa showed wilting, yellowing and even death. The relative plasma membrane permeability and malondialdehyde content of non-transgenic alfalfa were significantly higher than that of transgenic alfalfa, while chlorophyll content was significantly lower than that of transgenic alfalfa. It was found that the expression of RD29A gene of NADP-MEKIN1 was higher than that of non-transgenic alfalfa. The results showed that the overexpression of GsWRKY15 gene could significantly enhance the alkali-tolerant ability of alfalfa.
【作者單位】： 東北農(nóng)業(yè)大學(xué)農(nóng)業(yè)生物功能基因重點實驗室;
【基金】：國家自然科學(xué)基金項目(31171578) 黑龍江省高�？萍紕�(chuàng)新團隊建設(shè)計劃項目(2011TD005) 東北農(nóng)業(yè)大學(xué)學(xué)科團隊建設(shè)項目(團隊1)資助~~
【分類號】：Q943.2;S541.9
，

本文編號：1497743