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| 叠氮溴化丙锭结合实时定量聚合酶链式反应快速检测玫瑰酱中戊糖片球菌活菌 |
| Rapid detection of viable Pediococcus pentosaceus in rose jam using propidium monoazide- quantitative real-time polymerase chain reaction |
| 投稿时间：2025-01-10 修订日期：2025-06-10 |
| DOI： |
| 中文关键词: 玫瑰酱 戊糖片球菌 propidium monoazide quantitative real-time polymerase chain reaction 活菌检测 |
| 英文关键词:rose jam Pediococcus pentosaceus propidium monoazide quantitative real-time polymerase chain reaction viable cell detection |
| 基金项目:中央引导地方科技发展资金项目(YDZX2023081) |
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| 中文摘要: |
| 目的 探究基于叠氮溴化丙锭（propidium monoazide，PMA）与实时定量聚合酶链式反应（quantitative real-time polymerase chain reaction，qPCR）技术联用，建立一种快速检测玫瑰酱中戊糖片球菌活菌的方法。方法 根据特异性基因murI设计戊糖片球菌特异性引物，通过优化PMA浓度、暗孵育时间和曝光时间，确定最佳PMA处理条件，并通过PMA-qPCR方法验证不同比例的戊糖片球菌死菌/活菌混合菌液，评估该方法的灵敏度和检出限。结果 设计的引物具有良好的特异性，PMA的最佳处理条件为浓度80 μg/mL，暗孵育时间为15 min，曝光时间为25 min，此条件下，构建的标准曲线相关系数r2为0.9983，最低检出限为3.98×103 CFU/g。结论 本研究建立的PMA-qPCR方法可有效实现玫瑰酱中戊糖片球菌活菌数的快速检测，灵敏度高，为复杂食品基质中微生物的定量检测提供了技术支持。 |
| 英文摘要: |
| Objective To establish a rapid detection method of viable Pediococcus pentosaceus in rose jam based on the combination of propidium monoazide (PMA) and quantitative real-time polymerase chain reaction (qPCR). Methods Primers specific for P. pentosaceus were designed according to the species-specific murI gene. The optimal PMA treatment conditions were determined by optimizing the PMA concentration, dark incubation time, and exposure time. Different ratios of P. pentosaceus dead/live bacterial mixtures were validated by the PMA-qPCR method to assess the sensitivity and detection limit of the technique. Results The designed primers exhibited high specificity, and the optimal PMA treatment conditions were determined as 80 μg/mL PMA, 15 min of dark incubation, and 25 min of exposure. Under this condition, a standard curve with an r2 value of 0.9983 was constructed, and the lowest detection limit was determined to be 3.98×103 CFU/g. Conclusion The PMA-qPCR method established in this study enables the rapid and accurate quantification of viable P.pentosaceus in rose jam with high sensitivity, providing technical support for the quantitative detection of microorganisms in complex food matrices |
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