采用多重PCR技术检测转基因番木瓜华农一号

Detection of transgenic papaya Huanong No.1 by universal multiple-primer PCR

为了建立针对转基因番木瓜华农一号简便、快速、高灵敏度的多重PCR技术,用DP305植物基因组试剂盒方法提取3种转基因番木瓜(Event 55-1、GM-YK和华农一号)和非转基因番木瓜(台农2号)样品DNA,以木瓜蛋白酶基因(Papain)作为内参基因,进行了单重PCR技术引物特异性验证,多重PCR方法条件摸索及引物的灵敏度确定。结果表明∶5对特异性引物(CP、35s、NPTII、Papain、RP)得到相应的不同大小单一条带,其大小分别103、166、213、281、369 bp,说明引物具有特异性;建立了检测转基因番木瓜华农一号的五重PCR检测技术,检测限为0.2 ng。

In order to establish a simple,rapid,high sensitivity multiplex PCR technique for transgenic papaya Huanong No. 1,the study used the DP305 Plant Genomic DNA Kit to extract DNA from three kinds of transgenic papaya including Event 55-1,GM-YK,Huanong No. 1 and non-GM papaya( Tainong No. 2). The specificity of different primers of single PCR was verified with Papain gene as reference gene. The amplified condition and sensitivity of multiplex PCR technology was determined. In the result,the corresponding single band of different sizes,which was obtained from five pairs of specific primers CP,35 s,NPTII,Papain and RP,were 103 bp,166 bp,213 bp,281 bp and 369 bp,respectively. The five-fold PCR technology was establisehd for transgenic papaya Huanong No. 1. The detection limit was 0. 2 ng.