利用TAIL-PCR克隆耐盐基因及其分析  

Screening of Salt-tolerance rhizobium Strains and Cloning and Analysis of Salt-tolerance Gene

作　　者：张伟涛[1,2] 程国军[2] 周俊初[2] 

机构地区：[1]中南林业科技大学生物环境科学与技术研究所,长沙410004 [2]华中农业大学农业微生物国家重点实验室,武汉430070

出　　处：《生物技术通报》2010年第11期166-170,176,共6页Biotechnology Bulletin

基　　金：国家"973"项目(001CB1089);国家微生物资源平台建设项目(2005DKA21208-6)

摘　　要：通过大量筛选得到一株高耐盐的豇豆根瘤菌WME7,最高可耐15g/L NaCl,利用Tn5-sacB转座子对该菌株进行随机插入突变,从突变子中筛选获得30个共生缺陷型突变株。利用TAIL-PCR(thermal asymmetric interlaced PCR)方法克隆了突变株Tn5-sacB侧翼序列,通过BLAST发现有1个突变株的插入失活基因与鼠伤寒沙门氏菌抗银的结合蛋白SilE有94%同源性,表明有关Na+离子的抗性基因可能与Ag+离子的抗性基因有某种关系。该基因在其它菌中也能抗其它金属离子(铜、锌、钴、铬)。A high salt-tolerance rhizobium strain was screened,which can grown on 15 g/L NaCl YMA medium at most.Using Tn5-sacB insertion mutagenesis,30 salt-tolerance-defective mutants were screened by salt-tolerance experiments.The DNA sequences of the contiguous region from the Tn5 insertion site were determined by thermal asymmetric interlaced PCR（TAIL-PCR）.A new gene was cloned and BLAST analysis of the gene showed a sequence similarity（94%）to Salmonella typhimurium silver binding protein SilE gene,indicating there are certain relationship between gene of resistance to Ag＋ and Na＋.BLAST indicating this gene can resist copper,cadmium,zinc and cobalt et al.

关 键 词：筛选 耐盐根瘤菌 TAIL-PCR 耐盐基因 

分 类 号：Q78[生物学—分子生物学]