叠氮溴化丙锭-qPCR定量检测发酵乳中动物双歧杆菌乳亚种BB-12和植物乳植杆菌ST-Ⅲ  

Quantitative determination of Bifidobacterium animalis subsp.lactis BB-12 and Lactobacillus plantarum ST-Ⅲin fermented milk by propidium monoazide-qPCR

作　　者：张玉律 包静 王琳玲 ZHANG Yu-Lv;BAO Jing;WANG Lin-Ling(State Key Laboratory of Dairy Biotechnology,Shanghai Engineering Research Center of Dairy Biotechnology,Dairy Research Institute of Bright Dairy Co.,Ltd.,Shanghai 200436,China;Centre Testing International Group Co.,Ltd.,Shenzhen 518101,China)

机构地区：[1]乳业生物技术国家重点实验室,上海乳业生物工程技术研究中心,光明乳业股份有限公司乳业研究院,上海200436 [2]华测检测认证集团股份有限公司,深圳518101

出　　处：《食品安全质量检测学报》2025年第7期255-261,共7页Journal of Food Safety and Quality

基　　金：国家重点研发计划项目(2022YFD2100705);市国资委能级提升项目(2022013)。

摘　　要：目的将叠氮溴化丙锭(propidium monoazide,PMA)与实时荧光定量聚合酶链式反应(real-time fluorescence quantitative polymerase chain reaction,qPCR)联合,建立一种发酵乳体系中动物双歧杆菌乳亚种BB-12和植物乳植杆菌ST-Ⅲ的活菌定量检测方法。方法首先设计动物双歧杆菌乳亚种BB-12和植物乳植杆菌ST-Ⅲ的特异性基因序列的引物探针,验证引物探针特异性,优化PMA反应条件,建立标准曲线,验证灵敏度,最后定量检测发酵乳样品中动物双歧杆菌乳亚种BB-12和植物乳植杆菌ST-Ⅲ菌株的数量。结果动物双歧杆菌乳亚种BB-12与植物乳植杆菌ST-Ⅲ经PMA处理受损菌的最佳质量浓度为10μg/mL,处理菌液样品的最佳曝光时间为10 min,PMA-qPCR方法能够准确检测发酵乳体系中的动物双歧杆菌乳亚种BB-12与植物乳植杆菌ST-Ⅲ。结论本研究建立了一种快速、准确PMA-qPCR检测方法,用于检测发酵乳体系中的动物双歧杆菌乳亚种BB-12与植物乳植杆菌ST-Ⅲ,为发酵乳产品中益生菌的定量检测提供一定的借鉴意义。Objective To establish a quantitative detection method for the quantitative determination of the viable bacteria number of Bifidobacterium animalis subsp.lactis BB-12 and Lactiplantibacillus plantarum ST-Ⅲ in fermented milk,by combining propidium monoazide(PMA)with real-time fluorescence quantitative polymerase chain reaction(qPCR).Methods Firstly,the specific primer probe of Bifidobacterium animalis subsp.lactis BB-12 and Lactiplantibacillus plantarum ST-Ⅲ was designed and verified,the standard curve was established using standard strains,and the reaction conditions of the PMA were optimized.Secondly,the quantities of Bifidobacterium animalis subsp.lactis BB-12 and Lactiplantibacillus plantarum ST-Ⅲ in the fermented milk samples were quantitatively detected.Results The optimal mass concentration of damaged bacteria of Bifidobacterium animalis subsp.lactis BB-12 and Lactiplantibacillus plantarum ST-Ⅲ was 10μg/mL,and the optimal exposure time was 10 min.The PMA-qPCR detection method could accurately detect Bifidobacterium animalis subsp.lactis BB-12 and Lactiplantibacillus plantarum ST-Ⅲ in fermented milk.Conclusion In this study,a rapid and accurate PMA-qPCR detection method is established for the detection of Bifidobacterium animalis subsp.lactis BB-12 and Lactiplantibacillus plantarum ST-Ⅲ in fermented milk system,which provides certain reference significance for the quantitative detection of probiotics in fermented milk products.

关 键 词：叠氮溴化丙锭 实时荧光定量聚合酶链式反应 动物双歧杆菌乳亚种BB-12 植物乳植杆菌ST-Ⅲ 

分 类 号：TS2[轻工技术与工程—食品科学与工程]