摘要
目的建立多重实时聚合酶链式反应(polymerase chain reaction,PCR)法快速检测食品中的产志贺毒素大肠埃希氏菌(Shiga toxin-producing Escherichiacoli,STEC)的方法。方法以产志贺毒素大肠埃希氏菌携带的毒力基因stx1、stx2和黏附基因eae靶基因引物探针建立多重实时PCR体系,并采用原位冻干技术将PCR反应体系和阳性质控品进行了预先分装冻干,制成稳定、便捷的即用型反应体系,随后对方法的灵敏性、特异性和稳定性进行评价。结果所建立的多重实时PCR法检测eae、stx1、stx2基因的灵敏性为102 CFU/mL。与32种非STEC菌均无交叉反应。整体检测时长可控制在1 h以内,冻干体系可在常温条件下保存1年以上。结论本方法快速、简便,适用于测定食品中的STEC。
Objective To establish a multiplex real-time polymerase chain reaction(PCR)method for rapid detection of Shiga toxin-producing Escherichia coli(STEC).Methods A multiplex PCR system was established using target gene primers of virulence genes stx1,stx2 and adhesion gene eae carried by STEC.The PCR reaction system and positive control samples were pre-packed and lyophilized using in situ lyophilization technology to create a stable,convenient,ready-to-use reaction system.Subsequently,the sensitivity,specificity and stability of the method were evaluated.Results The sensitivity of the multiplex fluorescent PCR method established for detecting eae,stx1,and stx2 genes was 102 CFU/mL.There was no cross reaction with 32 non-STEC bacteria.The overall detection time can be controlled within 1 h,and the lyophilized system can be stored at room temperature for more than 1 year.Conclusions The method is rapid,simple and suitable for the determination of STEC in food.
作者
赵芳
牛娜
刘莹
涂晓波
刘慧玲
金晓蕾
吕敬章
ZHAO Fang;NIU Na;LIU Ying;TU Xiao-Bo;LIU Hui-Ling;JIN Xiao-Lei;LV Jing-Zhang(Shenzhen Customs,People’s Republic of China,Shenzhen 518045,China;Shenzhen Academy of Inspection and Quarantine,Shenzhen 518045,China)
出处
《食品安全质量检测学报》
CAS
2024年第21期294-300,共7页
Journal of Food Safety and Quality
基金
海关总署科研项目(2022HK026、2024HK010)。
关键词
快速检测
多重实时聚合酶链反应技术
产志贺毒素大肠埃希氏菌
冻干
rapid detection
multiplex real-time polymerase chain reaction
Shiga toxin-producing Escherichia coli
freeze drying
