基于微滴式数字PCR的饮料中嗜酸乳杆菌定量检测  

Quantitative Detection of Lactobacillus acidophilus in Beverage by Droplet Digital Polymerase Chain Reaction

作　　者：刘洋[1] 张娜娜 窦同海 俞漪[1] 徐琼[1] 曲勤凤[1] 赵渝[3] 杨捷琳 朱春梅 LIU Yang;ZHANG Nana;DOU Tonghai;YU Yi;XU Qiong;QU Qinfeng;ZHAO Yu;YANG Jielin;ZHU Chunmei(Shanghai Institute of Quality Inspection and Technical Research/National Center of Supervision and Inspection on Food Products Quality(Shanghai),Shanghai 200033,China;Shanghai Amplicon-gene Bioscience Co.Ltd.,Shanghai 201114,China;College of Life Sciences,Shanghai Normal University,Shanghai 200234,China;Technical center for animal,plant and food inspection and quarantine,Shanghai 200135,China;DuPont Nutrition&Biosciences,Danisco(China)Holding Co.Ltd.,Shanghai 200335,China)

机构地区：[1]上海市质量监督检验技术研究院,国家食品质量监督检验中心(上海),上海200233 [2]上海昂朴生物科技有限公司,上海201114 [3]上海师范大学生命科学学院,上海200234 [4]上海海关动植物与食品检验检疫技术中心,上海200135 [5]杜邦营养与生物科技丹尼斯克(中国)投资有限公司,上海200335

出　　处：《中国乳品工业》2020年第10期47-51,56,共6页China Dairy Industry

基　　金：上海市市场监督管理局科研项目(2018-32);国家市场监督管理总局科研项目(2017QK167,2019YJ018);国家自然科学基金项目(31301486)。

摘　　要：基于微滴式数字PCR技术(droplet digital polymerase chain reaction,ddPCR)建立饮料中嗜酸乳杆菌(Lactobacillus acidophilus strains)的检测方法。以嗜酸乳杆菌NCFM的SPIDR区域的重复序列设计并合成的引物,对其进行条件优化、特异性检测、优化反应体系,并对体系的灵敏度、重复性、抗干扰能力进行验证,最后对市售的实际样品进行检测。结果表明该方法的特异性很好;方法的灵敏度达到了0.25 pg/μL;重复性的标准偏差在2.34%~6.1%之间;在抗干扰检测中,实验组与对照组结果一致性好,提示该方法抗干扰能力较好;对市售的11批次样品进行检测,标记含有嗜酸乳杆菌的样品中均检出嗜酸乳杆菌且含量为1.6×10^3~2.3×10^6copies/g。研究建立的ddPCR方法可用于饮料中的嗜酸乳杆菌的快速和精确定量。Development of the method for detecting Lactobacillus acidophilus in beverage based on droplet digital polymerase chain reaction(ddPCR).Methods:Firstly,the primers were designed and synthesized based on the repeat sequence of the spidr region of Lactobacillus acidophilus NCFM.The conditions,specificity and reaction system were optimized,and the sensitivity,repeatability and anti-interference ability of the system were verified.Finally,the actual samples were tested.The results showed that the specificity of the method was very good;the sensitivity of the method reached 0.25 pg/μL;the standard deviation of repeatability was between 2.34%and 6.1%;in the anti-interference test,the results of the experimental group and the control group were consistent,indicating that the method had good anti-interference ability;11 batches of samples were tested,and Lactobacillus acidophilus was detected in the samples marked with Lactobacillus acidophilus and the content was 1.6×10^3~2.3×10^6 copies/g.The DDPCR method established in this study can be used for rapid and accurate quantification of Lactobacillus acidophilus in beverage.

关 键 词：ddPCR 嗜酸乳杆菌 定量 检测 

分 类 号：TS252.7[轻工技术与工程—农产品加工及贮藏工程]