转基因小麦B73-6-1外源基因拷贝数的确定  

Determination of Exogenous Gene Copy Number for Transgenic Wheat B73-6-1

作　　者：甄贞[1] 段俊枝[2] 于艳波[1] 曲波[1] 袁肖寒[1] 高学军[1] 栾凤侠 张明辉[1] 

机构地区：[1]东北农业大学生命科学学院/东北农业大学农业生物功能基因重点实验室/农业部转基因生物产品成分监督检验测试中心,黑龙江哈尔滨150030 [2]河南省农业科学院农业经济与信息研究所,河南郑州450002 [3]黑龙江出入境检验检疫局,黑龙江哈尔滨150030

出　　处：《河南农业科学》2016年第4期19-22,30,共5页Journal of Henan Agricultural Sciences

基　　金：黑龙江省教育厅科学技术研究(面上)项目(12521018)

摘　　要：为了确定转基因小麦B73-6-1中外源基因HMW-GS的拷贝数,以期为HMW-GS基因的整合位点和遗传表达研究奠定基础,构建了含有外源基因HMW-GS的标准品,并以wx012基因作为内源参照,通过实时荧光定量PCR方法,进行了3次重复试验。结果表明,在3组试验中得到的HMW-GS基因分子数分别为8.62×1018、4.34×1018、1.10×1018个,wx012基因分子数分别为6.27×1017、3.14×1017、7.84×1017个,经计算HMW-GS基因拷贝数分别为13.75、13.84、14.01个,由此确定转基因小麦B73-6-1中HMW-GS外源基因的拷贝数为14个。To reveal the copy number of exogenous gene HMW-GS in wheat B73-6-1,a real-time PCR technique for estimating transgene copy number in transgenic wheat B73-6-1 was built,so as to provide the basis for the analysis of integration sites and genetic expression of HMW-GS. In this study,the standard plasmid containing exogenous gene HMW-GS was constructed to detect copy number of HMW-GS in transgenic wheat B73-6-1. The SYBR Green I real-time PCR results showed that the molecule number of the exogenous gene HMW-GS were 8. 62 × 10^18,4. 34 × 10^18,1. 10 × 10^18,and the molecule number of the endogenous reference gene wx012 were 6. 27 × 10^17,3. 14 × 10^17,7. 84 × 10^17,respectively in three repeat experiments. The copy number of HMW-GS in transgenic wheat B73-6-1 were 13. 75,13. 84,14. 01 by caculated. So this assay revealed that transgenic wheat B73-6-1 carried 14 copies of the exogenous gene HMW-GS.

关 键 词：转基因小麦 B73-6-1 拷贝数 标准品 外源基因 

分 类 号：S512.1[农业科学—作物学]