小拟南芥高盐胁迫诱导cDNA文库的构建与测序分析  

Construction and Sequencing Analysis of cDNA Library Induced by High Salt Stress in Olimarabidopsis pumila

作　　者：院海英[1] 徐芳[1] 吕新华[1] 崔百明[1] 黄先忠[1] 

机构地区：[1]石河子大学生命科学学院/石河子大学农业生物技术重点实验室,石河子832003

出　　处：《石河子大学学报（自然科学版）》2012年第1期1-4,共4页Journal of Shihezi University(Natural Science)

基　　金：国家自然科学基金项目(31060149);石河子大学高层次人才启动项目(RCZX200902)

摘　　要：以新疆小拟南芥为材料,分离了小拟南芥经500mmol/L NaCl胁迫处理的幼苗总RNA,采用改良的SMARTcDNA合成方法合成第一链cDNA,采用LD PCR的方法合成第二链cDNA,经BP反应重组到入门载体pDONR207上,构建了小拟南芥高盐胁迫的入门cDNA文库。文库插入片段平均大小为1.1kb,阳性克隆率为94%。文库经随机挑选克隆测序,生物信息学分析初步获得了10个有功能的基因,其中包含各种酶如水解酶、氧化还原酶等;功能蛋白如离子运输通道蛋白、胁迫诱导蛋白,细胞色素蛋白及激素受体蛋白基因等。可以利用入门文库构建各种目的cDNA文库,本研究为将来从小拟南芥中筛选耐盐基因奠定了基础。Total RNA was isolated from shoot of Olimarabidopsis pumila stressed with 500 mmol/L NaCl.The first-strand cDNA was synthesized with superscript III RTase through an improved SMART cDNA synthesis method,the second-strand was synthesized via LD PCR amiplification,and the double-strand cDNA was recombined into entry vector pDONR207 via Gateway BP Clonase.The entry cDNA library of Olimarabidopsis pumila induced by high salt stress was thus constructed,with an average inserts size of 1.1 kb,and the positive clone rate of 94%.Randomly-selected clones were sequenced,and through bioinformatics analvsis,10 functional genes were obtained,including different enzymes such as oxidoreductase,hydroylase,etc,and function proteins,such as ion transporter proteins,stress induced protein,cytochrome b5 protein,and phytohormone receptor protein,etc.Various expression libraries can be constructed based on the entry library,and this study laid a foundation of mining salt-tolerance genes from Olimarabidopsis pumila.

关 键 词：新疆小拟南芥 耐盐 入门文库 表达文库 BP重组 

分 类 号：Q943.2[生物学—植物学]