畜牧兽医学报 ›› 2009, Vol. 40 ›› Issue (2): 185-190.doi:

• 遗传繁育 • 上一篇    下一篇

转基因小鼠的多重PCR快速检测技术

王趁芳,李新云,王志伟,敖红,崔文涛,李奎*   

  1. 中国农业科学院北京畜牧兽医研究所 中国农业科学院家养动物遗传资源与种质创新重点开放实验室,农业部转基因动物与饲料安全鉴定检验测试中心,北京 100193
  • 收稿日期:1900-01-01 修回日期:1900-01-01 出版日期:2009-02-24 发布日期:2009-02-24
  • 通讯作者: 李奎

Multiplex PCR Detection of Exogenous Gene in Transgenic Mice

WANG Chen-fang,LI Xin-yun,WANG Zhi-wei,AO Hong,CUI Wen-tao,LI Kui*   

  1. The Key Laboratory of Domestic Animal Genetic Resources and Germplasm Innovation of CAAS, Institute of Animal Science, Chinese Academy of Agricultural Sciences, Center for Identification and Detection of Transgenic Animal and Feed Safety of Department of Agriculture, Beijing 100193,China
  • Received:1900-01-01 Revised:1900-01-01 Online:2009-02-24 Published:2009-02-24

摘要: 转基因检测技术的标准化可以为转基因法规的制定和转基因产品的标识提供技术支持。采用碱裂解法快速提取鼠尾基因组DNA,用PCR方法检测其中的外源基因结构如cytomegalovirus (CMV)启动子、hGH polyA终止子及目的基因跨膜蛋白66 (Transmembrane protein 66, Tmem66),筛选到阳性样品。优化了多重 PCR 引物退火温度及缓冲液浓度,并探讨了扩增速率对多重PCR的影响。结果表明,此多重PCR方法可得到很好的扩增效果,可用于转基因小鼠外源基因的检测,同时为哺乳动物检测标准的建立提供参考。

Abstract: Focused on detection method for transgenic animals and its standardizations abroad in this article.It is essential to administrative regulations of transgenic animal and label of transgenic animal food.Extracting genomic DNA from mice tails by alkaline lysis method.The positive mice were screened by detecting of several transgenic elements such as cytomegalovirus (CMV) promoter, hGH polyA terminator, transmembrane protein 66 (Tmem66)target gene with multiplex PCR method, as detecting mice endogenous GAPDH gene. The process of the multiplex PCR reaction was optimized,such as the optimal annealing temperature and the concentration of the PCR buffer, simultaneously studying the effect of reaction rate on multiplex PCR.The results showed that the multiplex PCR method was simple and accurate to detect transgenic elements of transgenic mice, which offered us useful experience for developing the typical testing standard of transgenic mammals and its drived food.