食品中霍乱弧菌的新型检测方法研究  

Research on a new method for the detection of Vibrio cholerae in high-risk sea foods

作　　者：黎明[1] 周琛[2] 阮佳[2] 迪力努尔.多里坤 李永新[2] 

机构地区：[1]成都市疾病预防控制中心,四川成都610041 [2]四川大学,四川成都610041

出　　处：《中国卫生检验杂志》2016年第5期686-688,共3页Chinese Journal of Health Laboratory Technology

摘　　要：目的建立高危海产品中霍乱弧菌的PCR-激光诱导荧光-毛细管电泳检测方法。方法本研究以高危海产品为研究对象,针对霍乱弧菌肠毒素ctx A基因选择引物,利用PCR反应扩增霍乱弧菌的特征基因片段,用高效毛细管电泳-激光诱导荧光快速检测PCR产物。本研究对PCR条件及毛细管电泳分离条件进行优化,从而实现对高危海产品中霍乱弧菌的快速准确检测。结果在优化的PCR条件下,霍乱弧菌的ctx A基因可得到有效扩增,并且无非特异性条带;PCR扩增产物测序后,与基因库中的序列进行同源性比对,匹配度为99%。所建立的方法用于60份高危海产品中霍乱弧菌的检测,结果与普通凝胶电泳法所得结果一致。结论本方法所需样品量少、快速、灵敏、准确,可以成功用于霍乱弧菌的检测。Objective To establish a new method of PCR-capillary electrophoresis with laser-induced fluorescence detector to detect pathogenic Vibrio cholerae（ V. cholerae） in high-risk sea foods. Methods In this study,high risk seafood was selected as the research object. The primers were synthesized according to ctx A gene,and PCR reaction was used for the amplification of gene fragment of Vibrio cholerae. Subsequently,the PCR products were detected by capillary electrophoresis-laser induced fluorescence with fast speed. The parameters of PCR reaction and the separation conditions of CE were optimized,and thus a rapid and accurate detection of V. cholerae was achieved. Results Under the optimized conditions,ctx A gene of Vibrio cholerae can be efficiently amplified and no unspecific products were oberserved. The alignment analysis showed that the PCR products had a good agreement of 99% with the published sequences from Gen Bank. This proposed method was applied for the detection of a total of 60 high-risk sea food samples. The results were consistent with those by agarose gel electrophoresis.Conclusion This method was rapid,sensitive,accurate,and required less amount of sample,and expected to be successfully used for the detection of Vibrio cholera.

关 键 词：霍乱弧菌 聚合酶链式反应 CTX A基因 琼脂糖凝胶电泳 毛细管电泳 

分 类 号：R155.5[医药卫生—营养与食品卫生学]