首页分享转基因花卉的DNA提取与多重PCR分析

转基因花卉的DNA提取与多重PCR分析

来源：花匠小妙招时间：2024-11-11 02:04

（1.福建出入境检验检疫局，福建福州 350001；2.厦门北大之路生物工程有限公司，福建厦门 361009； 3. 厦门大学生物系，福建厦门 361005）

摘要：分别以CTAB法、试剂盒提取菊花、矮牵牛、非洲菊3种花卉的基因组DNA，凝胶电泳结果表明试剂盒提取的效果较好。设计合成了3对引物，分别用于扩增花椰菜花叶病毒（Cauliflower mosaic virus，CaMV）35S启动子、根癌农杆菌胭脂碱合成酶基因（NOS）终止子、大肠杆菌K12菌株新霉素磷酸转移酶II（NPTII）基因。普通PCR检测结果表明，3种花卉6个样品中仅矮牵牛的1个样品含有这3种转基因成分，酶切鉴定结果验证了PCR产物为非假阳性。尝试以多重PCR同时检测转基因矮牵牛与对照阳性质粒中的2个或3个转基因成分，得到预期结果。

关键词：多重PCR；35S启动子；NOS终止子；NPTII基因

中图分类号： S727-31；Q78 文献标识码：A

DNA Extraction and Multiplex Polymerase Chain Reaction Analysis of Transgenic Flowers

SHAO Bi-ying1，CHEN Wen-bing1，LI Shou-song1，LI Shi-cheng2，YE Wen-biao2，CHEN Liang3
（1. Fujian Entry-Exit Inspection and Quarantine Bureau，Fuzhou 350001，China；
2. Xiamen Bioway biology engineering company，Xiamen 361009，China；
3. Dept. of Biol.，Xiamen Univ.，Xiamen 361005，China）

Abstract：The DNAs of chrysanthemum, petunia and Gerbera were extracted by CTAB method and DNA extraction kit, respectively. The gel electrophoresis result showed that the DNA extraction kit is more effective. Three pairs of primes were designed and synthesized to amplify Cauliflower mosaic virus(CaMV) 35S promoter, Agrobacterium tumefaciens nopaline synthase terminator and Escherichia coli strain K12 neomycin phosphotransferase II gene, respectively. The general PCR detection results suggested that only one sample of petunia contained the three genetically modified ingredients among six samples of the three flowers. The restriction enzyme analysis result suggested the PCR products were not false positive. The multiplex polymerase chain reaction（MPCR）was attempted to detect two or three genetically modified ingredients of transgenic petunia and positive control plasmid at the same time, and obtained the same results as expected.

Key words：multiplex polymerase chain reaction；35S promotor；NOS terminator；NPTII gene