棉花花药中高温胁迫响应基因的克隆与分析  

Cloning and Analysis of High-Temperature-Stress-Responsive Genes in Cotton Anther

作　　者：田东洋[1] 陈劲[1] 袁小玲[1] 刘志[1] 

机构地区：[1]湖南农业大学生物科学技术学院,长沙410128

出　　处：《分子植物育种》2012年第5期551-558,共8页Molecular Plant Breeding

基　　金：国家自然科学基金项目(30900909);"十二五"863计划(2012AA101108)共同资助

摘　　要：以耐高温和敏感型陆地棉品系HLY11与TS18为材料,采用抑制差减杂交技术(SSH)构建高温胁迫下花药的正向和反向差减文库。共获得了2880个阳性克隆,随机挑取400个克隆进行PCR验证,经测序和序列拼接分析,共获得342条有效EST序列,聚类分析得到278条非冗余的EST(unigenes)。BlastN分析发现,有211条unigenes能在GenBank数据库中找到同源序列,其中164条与已知功能蛋白有较高同源性,47条与未知功能或假定蛋白有较高相似性,其余67条没有同源匹配。KEGG分析将152条unigenes定位到46条代谢途径中,而P<0.05的代谢途径有14条,包含了31个unigenes。初步分析发现,棉花花药中响应高温逆境胁迫的基因涉及热激蛋白、抗氧化酶类、碳水化合物代谢、转录因子和跨膜转运蛋白等过程,其中淀粉和糖代谢、谷胱甘肽代谢、植物病原生物相互作用和氧化磷酸化作用等可能与棉花花药高温逆境耐性的相关性较大。A forward and a reverse eDNA libraries were constructed by suppression subtractive hybridization （SSH） method using the anthers of a tolerant upland cotton （Gossypium hirsutum L.） line HLY11 and a sensitive line TS18 under high temperature stress. A total of 2 880 positive clones were obtained from the eDNA libraries, among which 400 clones were selected randomly and confirmed by PCR, and then sequenced. 342 valid EST sequences were acquired by sequencing and analysis of sequence assembly and 278 non-redundant unigenes were acquired by splitting and clustering analyses of the eDNA sequences. The BlastN analysis showed that 211 unigenes had homologous sequences in GenBank database, among which 164 unigenes had high homology with known functional protein and 47 unigenes have higher similarity with unknown function or putative proteins, while the rest of 67 unigenes had no protein homologous. 152 unigenes were mapped to 46 metabolic pathways by KEGG analyses, in which there were 14 pathways containing 31 unigenes with P-value less than 0.05. Preliminary analysis found that high-temperature-stress-responsive genes in cotton anther involved in the process of heat shock protein, antioxidant enzymes, carbohydrate metabolism, transcriptional factor, trans-membrane protein, of which the starch and sucrose metabolism, glutathione metabolism, plant-pathogen interaction and oxidative phosphorylation may closely correlate with tolerance to high temperature stress in cotton anther.

关 键 词：棉花 花药 抑制消减杂交 高温胁迫响应基因 EST 

分 类 号：S562[农业科学—作物学]