| 副溶血弧菌EMA-PCR检测技术的建立 |
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| 引用本文： | 伦镜盛,夏常艳,罗鹏,梁嘉明,黄通旺,胡忠.副溶血弧菌EMA-PCR检测技术的建立[J].微生物学通报,2011,38(6):952-956. |
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| 作者姓名： | 伦镜盛 夏常艳 罗鹏 梁嘉明 黄通旺 胡忠 |
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| 作者单位： | 1. 汕头大学生物系,广东汕头,515063
2. 中国科学院南海海洋研究所,广东广州,510301 |
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| 基金项目： | 广东省科技计划项目(No. 2006B20501008, 2009B020309007); 广东省海洋渔业科技推广项目(No. A200901H06) |
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| 摘 要： | PCR技术被广泛应用于副溶血弧菌的检测中, 但传统的PCR技术无法区分样品中的死细菌与活细菌, 往往使检测结果出现较高的假阳性。因此, 将叠氮溴乙锭(Ethidium monoazide bromide, EMA)与PCR技术结合, 建立一种快速、准确的副溶血弧菌检测方法。以dnaJ基因为检测副溶血弧菌的靶基因, 分别用副溶血弧菌的纯培养细胞及其基因组DNA作模板进行PCR检测, 灵敏度分别为2.5×104 CFU/mL和6×102 fg/μL。在检测样品前处理过程中加入EMA, 当EMA的浓度小于5 mg/L时, EMA对活菌靶基因的扩增没有明显的抑制; 而终浓度为2 mg/L的EMA, 能有效抑制1×108 CFU/mL副溶血弧菌死菌的扩增。活菌和死菌混合体系的PCR结果表明, EMA-PCR能有效降低副溶血弧菌检测过程中的假阳性。
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| 关 键 词： | 副溶血弧菌 EMA-PCR 死菌与活菌 |
Establishment of EMA-PCR detection method of Vibrio parahaemolyticus |
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| LUN Jing-Sheng,XIA Chang-Yan,LUO Peng,LIANG Jia-Ming,HUANG Tong-Wang and HU Zhong.Establishment of EMA-PCR detection method of Vibrio parahaemolyticus[J].Microbiology,2011,38(6):952-956. |
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| Authors: | LUN Jing-Sheng XIA Chang-Yan LUO Peng LIANG Jia-Ming HUANG Tong-Wang and HU Zhong |
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| Institution: | 1. Department of Biology, Shantou University, Shantou, Guangdong 515063, China;1. Department of Biology, Shantou University, Shantou, Guangdong 515063, China;2. South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou, Guangdong 510301, China;1. Department of Biology, Shantou University, Shantou, Guangdong 515063, China;1. Department of Biology, Shantou University, Shantou, Guangdong 515063, China;1. Department of Biology, Shantou University, Shantou, Guangdong 515063, China |
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| Abstract: | PCR technology has been widely used in the detection of Vibrio parahaemolyticus.However,traditional PCR appeared higher false-positive result because of the lack of differentiation between DNA from viable and dead microorganisms.Therefore,Ethidium Monoazide Bromide(EMA) was added in the process of PCR to establish a rapid and accurate detection method of V.parahaemolyticus.The dnaJ gene was used as the target gene for PCR detection of V.parahaemolyticus by utilizing its pure isolates and genomic DNA as the ... |
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| Keywords: | V parahaemolyticus EMA-PCR Viable and dead bacteria |
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