【摘要】：本研究基于實驗室前期野生大豆堿脅迫轉(zhuǎn)錄組數(shù)據(jù),篩選出一個堿脅迫下上調(diào)表達(dá)的假定蛋白基因,暫命名為GsARHP(alkali stress related hypothetical protein gene)。首先利用Real-time PCR方法驗證了GsARHP基因受堿脅迫誘導(dǎo)表達(dá)。生物信息學(xué)分析表明,該基因編碼一個含有130個氨基酸的親水蛋白,含有信號肽但無跨膜結(jié)構(gòu)域;構(gòu)建了GsARHP植物超量表達(dá)載體,利用農(nóng)桿菌介導(dǎo)的子葉節(jié)侵染法轉(zhuǎn)化肇東紫花苜蓿,通過PCR,Southern Blot和RT-PCR方法檢測獲得了3個超量表達(dá)GsARHP基因的轉(zhuǎn)基因株系,并對其耐堿性進行了分析。結(jié)果表明,在0,100和150mmol/L NaHCO3處理14d后,非轉(zhuǎn)基因株系明顯萎蔫、黃化甚至死亡,而轉(zhuǎn)基因株系則長勢良好;進一步分析其生理指標(biāo)顯示,相對質(zhì)膜透性與丙二醛含量均顯著低于非轉(zhuǎn)基因株系(P0.01),而葉綠素含量與CAT活性顯著高于非轉(zhuǎn)基因株系(P0.01),說明GsARHP基因的超量表達(dá)可以增強紫花苜蓿的耐堿能力。
[Abstract]:In this study, based on the data of alkaline stress transcriptional group in early laboratory of wild soybean, a hypothesis protein gene up regulation under alkali stress was screened and named GsARHP (alkali stress related hypothetical protein gene). First, Real-time PCR method was used to verify the expression of GsARHP based on alkali stress. Bioinformatics analysis table This gene encodes a hydrophilic protein containing 130 amino acids, containing a signal peptide but without a transmembrane domain, and constructs a GsARHP plant overexpression vector, which uses Agrobacterium mediated cotyledon node infection to convert alfalfa in Zhaodong, and 3 overexpressed GsARHP genes are detected by PCR, Southern Blot and RT-PCR method. The results showed that after 0100 and 150mmol/L NaHCO3 treatment of 14d, the non transgenic lines were obviously wilted, yellow and even died, while the transgenic lines had good growth. Further analysis of the physiological indexes showed that the relative plasma membrane permeability and the content of prop two aldehyde were significantly lower than those of the non transgenic lines (P0.01). The content of green pigment and CAT activity were significantly higher than those of non-transgenic lines (P 0.01), indicating that overexpression of GsARHP gene could enhance alkali tolerance 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
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本文編號：2172277