TaqMan探针实时荧光定量聚合酶链式反应检测低温酸乳中常见的霉菌和酵母

doi:10.7506/rykxyjs1671-5187-20240703-049

乳业科学与技术 ›› 2024, Vol. 47 ›› Issue (5): 20-24.DOI: 10.7506/rykxyjs1671-5187-20240703-049

TaqMan探针实时荧光定量聚合酶链式反应检测低温酸乳中常见的霉菌和酵母

张捷，陈勃旭，杜海宽，张子仑，严陶陶，陈佳，周巍

（1.河北省食品检验研究院，河北省食品安全重点实验室，国家市场监督管理总局重点实验室（特殊食品监管技术），特殊食品安全与健康河北省工程研究中心，河北?石家庄 050227；2.石家庄学院化工学院，河北?石家庄 050035）

出版日期:2024-09-01 发布日期:2024-11-06
基金资助:
河北省市场监督管理局科技计划项目（2023ZD20）；河北省高等学校科学技术研究项目（ZC2023112）；国家市场监督管理总局科技创新人才计划项目（QNBJ202323）

Detection of Common Molds and Yeasts in Low-Temperature Yogurt by TaqMan probe Real-Time Fluorescence Quantitative Polymerase Chain Reaction

ZHANG Jie, CHEN Boxu, DU Haikuan, ZHANG Zilun, YAN Taotao, CHEN Jia, ZHOU Wei

(1. Hebei Food Safety Key Laboratory, Key Laboratory of Special Food Supervision Technology, State Administration for Market Regulation, Hebei Engineering Research Center for Special Food Safety and Health, Hebei Food Inspection and Research Institute, Shijiazhuang 050227, China; 2. College of Chemical Engineering, Shijiazhuang University, Shijiazhuang 050035, China)

Published:2024-09-01 Online:2024-11-06

摘要/Abstract

摘要： 为提高低温酸乳中霉菌和酵母菌检测效率，根据保守基因延伸因子1α（elongation factor 1α，EF-1α）序列分别针对霉菌与酵母菌设计引物与探针，利用实时荧光定量聚合酶链式反应，基于TaqMan探针分别建立霉菌和酵母菌检测方法。以低温酸乳中常见真菌与乳酸菌基因组DNA为扩增模板进行特异性检验，并对方法灵敏度与重复性进行检验，同时构建标准曲线。结果表明：根据EF-1α基因设计的引物与探针特异性良好；所构建的标准曲线相关系数均大于0.99；该方法检测酵母菌的灵敏度为101?CFU/mL，检测霉菌的灵敏度为102?CFU/mL；重复性检验中组内与组间的变异系数均小于2%，表明方法具有良好的可靠性与稳定性，适用于低温酸乳中霉菌与酵母菌的快速检测。

关键词: TaqMan探针；霉菌；酵母；实时荧光定量聚合酶链式反应；低温酸乳

Abstract: In order to improve the detection efficiency of molds and yeasts in low-temperature yogurt, primers and probes were designed according to the sequences of the conserved gene elongation factor-1α (EF-1α) for molds and yeasts, respectively. A real-time fluorescence quantitative polymerase chain reaction (real-time PCR) method with TaqMan probe was established for the detection of molds and yeasts, respectively. The genomic DNA of fungi and lactic acid bacteria commonly found in low-temperature yogurt, were used as amplification templates for specificity testing. The sensitivity and repeatability were also tested, and the standard curves were constructed. The results showed that the designed primers and probes had good specificity; the sensitivity of the method was 101 and 102 CFU/mL for yeasts and molds, respectively; the coefficients of variation within and between groups in the repeatability test were less than 2%, indicating good reliability and stability; the correlation coefficients of the standard curves were higher than 0.99. Therefore, the real-time PCR method enables the rapid detection of molds and yeasts in low-temperature yogurt.

Key words: TaqMan probe; mold; yeast; real-time fluorescence quantitative polymerase chain reaction; low-temperature yogurt

中图分类号:

TS252.54

张捷，陈勃旭，杜海宽，张子仑，严陶陶，陈佳，周巍. TaqMan探针实时荧光定量聚合酶链式反应检测低温酸乳中常见的霉菌和酵母[J]. 乳业科学与技术, 2024, 47(5): 20-24.

ZHANG Jie, CHEN Boxu, DU Haikuan, ZHANG Zilun, YAN Taotao, CHEN Jia, ZHOU Wei. Detection of Common Molds and Yeasts in Low-Temperature Yogurt by TaqMan probe Real-Time Fluorescence Quantitative Polymerase Chain Reaction[J]. Journal of Dairy Science and Technology, 2024, 47(5): 20-24.

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TaqMan探针实时荧光定量聚合酶链式反应检测低温酸乳中常见的霉菌和酵母

Detection of Common Molds and Yeasts in Low-Temperature Yogurt by TaqMan probe Real-Time Fluorescence Quantitative Polymerase Chain Reaction

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