热胁迫下中甸角蒿叶片SSH文库的构建及初步分析

Construction of Leave Library by SSH and Preliminary Analysis of Genes Responsible for Heat Sress in Incarvillea zhongdiannensis

高温可能是限制中甸角蒿(Incarvillea zhongdiannensis)向低海拔地区引种驯化的主要因素之一。为了从基因表达水平研究中甸角蒿高温胁迫响应的分子机制,本研究利用SSH技术构建了中甸角蒿对高温(30℃)处理响应的正反向抑制性差减杂交文库。文库质量检测表明抑制性差减杂交效率较高,质量较好。通过对正反向文库中部分EST进行序列测定,获得了60条高质量的表达序列标签,平均长度为537bp。对序列进行BLAST比对及功能注释,50条EST为功能已知的基因,分别参与信号转导与转录、植物抗逆性反应、光合作用、代谢与能量、蛋白质合成与转运、蛋白质命运、细胞结构和细胞生长等过程。6个EST与功能未知基因的同源性较高。获得的4个未匹配的EST推测为新基因,可能在植物热耐受性方面具有重要的作用。

High temperature might be one of the main limiting factors to be introduced and domesticated into lower altitude for Incarvillea zhongdiannensis. To further reveal the underlying molecular mechanism in I.zhongdiannensis responding to high temperature, the suppression subtractive hybridization approach （SSH） was used in this study. The results showed the quality of forward or reverse subtraction cDNA library is good enough for further SSH experiments. Total sixty expressed tags （ESTs） with high quality were randomly obtained by SSH and sequencing. According to BLAST screening and functional annotation, 50 ESTs with known function were classified into 9 broad categories such as defense/stress, photosynthesis, metabolism, energy, signal transduction and transcription, protein fate, transporter, biosynthesis, cell structure and cell growth. There were six ESTs significantly matched with hypothetical proteins. Four ESTs with no hits possibly encode novel genes and play important role in I.zhongdiannensis responding to high temperature stress.