实时定量PCR测定苏云金芽胞杆菌质粒pBMB67的拷贝数  

Determination of the Copy-number of the Resident Plasmid p BMB67 from Bacillus thuringiensis Strain YBT-1520 by Quantitative Real-time PCR

作　　者：钟春英[1] 孙明[2] 

机构地区：[1]湖北第二师范学院化学与生命科学学院/植物抗癌活性物质提纯与应用湖北省重点实验室,武汉430205 [2]华中农业大学生命科学技术学院/农业微生物国家重点实验室,武汉430070

出　　处：《湖北农业科学》2015年第8期2001-2006,共6页Hubei Agricultural Sciences

基　　金：湖北省自然科学基金项目(2014CFB566)

摘　　要：根据苏云金芽胞杆菌(Bacillus thuringiensis)YBT-1520的全基因组测序结果,在内生质粒p BMB67和染色体上分别选取单拷贝片段,利用实时荧光定量PCR法测得质粒p BMB67的拷贝数为21。此外,在内生质粒p BMB67的3个不同位点设计引物,分别以不同的靶片段作为实时荧光定量PCR的检测对象,测得p BMB67质粒的拷贝数分别为20.1、21.0和21.3,表明单拷贝靶片段的选择对测定质粒的拷贝数没有影响,进一步证明了实时荧光定量PCR测定质粒拷贝数的可行性。According to the whole genome sequence of Bacillus thuringiensis strain YBT-1520,a specific single-copy fragment was selected from the resident plasmid pBMB67 and chromosome separately. And the copy number （PCN） of pBMB67 was estimated as 21 by quantitative real-time PCR. In addition,three pairs of primers were designed at three different fragments of pBMB67 as the targets of quantitative real-time PCR detection,and the result showed that the PCN of pBMB67 was 21.3,20.1, and 21.0,respectively. This finding indicated that the difference of target fragments on plasmid for quantitative real-time PCR detection would not affect the value of PCN,and demonstrated that the quantitative real-time PCR method is reliable to determine the PCN.

关 键 词：苏云金芽胞杆菌(Bacillus thuringiensis) 质粒 质粒拷贝数 实时荧光定量PCR 

分 类 号：Q782[生物学—分子生物学]