干旱胁迫下狗牙根Tifway和C299抑制性消减文库的构建抗旱基因分离
首发时间:2014-08-07
摘要:采用抑制差减杂交技术,构建了草坪草狗牙根Tifway 和C299干旱5d 和10d应答SSH (Suppression Subtractive Hybridization) 文库四个,文库共包含1272个阳性克隆。对质粒的巢式PCR 结果表明,插入片段大小为88--823 bp 。所有阳性克隆进行测序, 利用BLASTn 和BLASTx 进行序列相似性分析,并进行DNAMAN软件比对分析,确定共有757个EST,其中,277个在Genbank中可以查找到相似性较高的含有明确功能的同源蛋白。这些基因中包括了与叶表面蜡质合成基因(CER1 和固醇脱氢酶基因)、 与耐旱相关的基因(脱水素和HVA-22-like蛋白基因)、与抗氧化相关基因(超氧化物歧化酶、脱氢抗坏血酸还原酶和2-半胱氨酸过氧化物酶)、和转录调控因子(EREBP-4 like 蛋白和WRKY)。它们的表达体现出植物在干旱胁迫条件下特异基因表达水平上的反应。
关键词: 狗牙根 干旱胁迫 抑制性消减cDNA文库
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Construction and Characterization of Bermudagrass C299 and Tifway Suppression Subtractive Hybridization (SSH) Library under Drought Stress
Abstract:To gain insights into molecular mechanisms of grass responding to drought stress, we constructed a suppression subtractive cDNA library to identify drought responsive genes for C4 grass species bermudagrass(Cynodon dactylon(L.)Pers.) C299 and bermudagrass(Cynodon dactylon (L.) Pers. × C. Transvaalensis Burtt Davy) Tifway. 1272 clones were confirmed to have inserted cDNA fragment in this library. Nested PCR confirmed inserted cDNA fragment with the length from 100 to 500 bp.The SSH analysis identified 72 non-redundant putative drought-responsive cDNAs, in which 23 genes with homology to known proteins were found in Genbank. This genes are including alcohol dehydrogenase 1,carbonic anhydrase,DNA repair ATPase-related and so on. Their expression in plant shows plants responding to drought at molecular level.
Keywords: bermudagrass drought stress suppression subtractive hybridization library
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干旱胁迫下狗牙根Tifway和C299抑制性消减文库的构建抗旱基因分离
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