黄曲霉素产毒真菌ver-1、verB及ITS基因序列多重PCR体系的优化  

Optimization of Multiplex PCR System for Aflatoxin Producing Fungi Based on ver-1,verB and ITS Genes

作　　者：潘锋君[1] 叶垚敏 张晓芹[2] PAN Fengjun;YE Yaomin;ZHANG Xiaoqin(Lishui Municipal Central Hospital,Lishui 323000,China;Lishui Hospital of Traditional Chinese Medicine,Lishui 323000,China)

机构地区：[1]丽水市中心医院,浙江丽水323000 [2]丽水市中医院,浙江丽水323000

出　　处：《中国现代应用药学》2022年第7期912-917,共6页Chinese Journal of Modern Applied Pharmacy

基　　金：浙江省基础公益研究计划项目(LGF19H280002)。

摘　　要：目的对黄曲霉素产毒真菌的多重PCR体系进行优化,确定最佳PCR体系。方法以黄曲霉素产生过程中的主要调控基因ver-1、verB及通用基因片段ITS序列为目的片段,设计多重PCR引物,采用单因素和正交优化试验,对DNA模板、Mg^(2+)浓度、引物用量、dNTPs用量、退火温度等因素进行考察。结果最佳多重PCR体系为:50μL体系中含有引物(5μmol·L^(−1))3μL,dNTPs(2.5 mmol·L^(−1))5μL,Mg^(2+)(2.5 mmol·L^(−1))4μL,DNA浓度10 ng·μL^(−1),退火温度56℃。结论建立的体系可用于黄曲霉素产毒真菌的多重PCR鉴定,对黄曲霉产毒菌的源头鉴定具有一定的意义。OBJECTIVE To optimize the multiplex PCR system of aflatoxin producing fungi and to determine the optimal PCR system.METHODS Multiple PCR primers were designed based on the sequences of ver-1,verB and ITS,which were the main regulatory genes in aflatoxin production,DNA template,Mg^(2+)concentration,primer dosage,dNTPs dosage,annealing temperature and other factors were investigated by single factor and orthogonal test.RESULTS The optimal multiplex PCR system was as follows:50μL system contained primer(5μmol·L^(−1))3μL,dNTPs(2.5 mmol·L^(−1))5μL,Mg^(2+)(2.5 mmol·L^(−1))4μL,DNA concentration 10 ng·μL^(−1),annealing temperature 56℃.CONCLUSION The established system can be used for the identification of aflatoxin producing fungi by multiplex PCR and has certain significance for the source identification of aflatoxin producing fungi.

关 键 词：黄曲霉素 产毒真菌 多重PCR 优化体系 

分 类 号：R927[医药卫生—药学]