食源有害微生物DNA快速提取与多重PCR检测初探  

Preliminary Study on Rapid Extraction of DNA and Multiplex PCR Detection of Harmful Food-Borne Microorganisms

作　　者：张静[1] 黄钢 孙婧 张英洁 付宇浩 李朝阳 菅一鸣 李振庭 ZHANG Jing;HUANG Gang;SUN Jing;ZHANG Yingjie;FU Yuhao;LI Chaoyang;JIAN Yiming;LI Zhenting(School of Life Sciences,Jianghan University,Wuhan 430056,Hubei,China)

机构地区：[1]江汉大学生命科学学院,湖北武汉430056

出　　处：《江汉大学学报（自然科学版）》2020年第4期46-53,共8页Journal of Jianghan University：Natural Science Edition

摘　　要：目的建立一种食源有害微生物快速检测体系。方法以致泻大肠埃希氏菌、金黄色葡萄球菌、志贺氏菌、沙门氏菌、单核细胞增生李斯特氏菌5种有害菌为检测对象,以30 s快速提取基因组DNA及多重PCR的试验技术进行了研究。结果筛选出特异性引物8对;发现致泻大肠埃希氏菌与单核细胞增生李斯特氏菌能够进行DNA快速提取PCR检测;构建了致泻大肠埃希氏菌与单核细胞增生李斯特氏菌DNA快速提取多重PCR检测体系,筛选出多重PCR成功的引物组合4对:rfbE/Hly、fbE/HlyA1、rfbE/HlyA2和eaeA/HlyA2,并对试验体系进行了优化。结论利用DNA快速提取多重PCR检测某些食源有害微生物是可行的。但是,针对具体情况采取措施优化反应十分必要,针对多重PCR设计相应引物,可能是优化关键。Objective To establish a rapid detection system for harmful food-borne microorganisms. Methods In this study,5 kinds of harmful bacteria such as Escherichia coli,Staphylococcus aureus,Shigella,Salmonella,and Listeria monocytogenes were detected.The rapid extraction of genomic DNA in 30 seconds and multiplex PCR were used as experimental techniques. Results 8 pairs of specific primers of the 5 bacteria were screened out,and it was found that Escherichia coli and Listeria monocytogenes could be detected by DNA rapid extraction and PCR. A multiplex PCR system based on DNA rapid extraction of Escherichia coli and Listeria monocytogenes was established. Four pairs of primer combinations:rfbE/Hly,fbE/HlyA1,rfbE/HlyA2,and eaeA/HlyA2,were screened out successfully in multiplex PCR experiments,and the experiments were optimized. Conclusion It is feasible to detect harmful food-borne microorganisms by DNA rapid extraction and multiplex PCR.However,it is necessary to take measures to optimize the experiments. The design of primers for multiplex PCR may be the key to optimize the experiments.

关 键 词：食源有害微生物 DNA快速提取 多重PCR 检测 

分 类 号：TS201.3[轻工技术与工程—食品科学]