盐胁迫下北美海蓬子全长cDNA文库的构建及耐盐相关基因的表达分析  

cDNA Library Construction of Salicornia bigelovii Torr.Under Salt Stresses and Analysis of Tolerance Related Gene

作　　者：王忠[1] 汪仁[1] 彭峰[1] 何树兰[1] 郑玉红[1] 

机构地区：[1]江苏省中国科学院植物研究所,南京210014

出　　处：《农业生物技术学报》2013年第5期505-510,共6页Journal of Agricultural Biotechnology

基　　金：江苏省中国科学院植物研究所所青年基金(所201002)

摘　　要：盐生植物北美海蓬子(Salicornia bigelovii Torr.)是世界上最具耐盐性的植物之一,为了研究其耐盐机理和获得新的耐盐基因,本研究利用SMARTTM(switching mechanism at 5'end of RNA transcript)技术构建了北美海蓬子在盐胁迫下的全长cDNA文库。原始文库滴度为5.46×105pfu/mL,重组率为96.5%,插入片段长度在0.5～2kb,1kb以上的占60%。从原始文库中随机挑取100个阳性克隆进行序列分析,获得65条高质量的基因序列。通过检索后发现其中有些是逆境应答相关基因和新基因。此外,以400mmol/LNaCl处理的北美海蓬子为模板,对文库中与耐盐相关的SbX和SbSAMS进行RT-PCR分析,发现随着胁迫时间的延长,海蓬子体内的SbX表达量上调,而SbSAMS的表达量则逐渐减弱。研究结果为植物耐盐性机理、农作物遗传性状改良和种质创新提供基础资料。Salicornia bigekn,ii Torr. is one of the most salt-tolerant plant species in the world. In order to study the mechanism of salt tolerance and isolate new genes related to salt tolerance in S. bigelovii Torr., a normalized full-length cDNA library in S. bigelovii Tort. was constructed by SMART ^（switching mechanism at 5＇ end of RNA transcript） technique. The primary library had a titer of5.46×10^5 pfu/mL, and it＇s recombination rate reached 96.7%. The average size range ofcDNA inserts was 0.5-2 kb, 60% of them more than 1.0 kb. 100 clones were randomly selected and sequenced, and then preliminary analysis was performed using bioinformatics software, 65 gene sequences were generated from the cDNA library. By Blast searches, we found that 35 genes showed significant similarity to the annotated genes, and the remaining 30 genes represented novel genes without any annotation. In addition, we analyzed SbX and SbSAMS mRNA level by semi-quantity RT-PCR in S. bigekn：ii Torr. under 400 mmoFL NaCI. We found that SbX was up-regulated by salt slress, and the expression of SbSAMS was opposite. The results may provide basic information about the mechanism of salt tolerance in plants, improvement and germplasm enhancement of crop genetic traits.

关 键 词：北美海蓬子 盐胁迫 CDNA文库 

分 类 号：S435.621.2[农业科学—农业昆虫与害虫防治]