添加扩增内标的PCR方法快速检测食品中的沙门氏菌  

Establishment of a PCR method with an internal amplication control for rapid detection of Salmonlla in foods

作　　者：张德福[1,2] 付绪磊 汤轶伟[1] 白凤翎[1] 殷喆[2] 杨文慧[2] 赵丽红[1] 张义全[3] 李春[4] 励建荣[1] 

机构地区：[1]渤海大学食品科学与工程学院辽宁省食品安全重点实验室生鲜农产品贮藏加工及安全控制技术国家地方联合工程研究中心,辽宁锦州121013 [2]军事医学科学院微生物流行病研究所病原微生物生物安全国家重点实验室,北京100071 [3]江苏大学医学院,江苏镇江212013 [4]渤海大学数理学院,辽宁锦州121013

出　　处：《食品工业科技》2015年第24期54-57,63,共5页Science and Technology of Food Industry

基　　金："十二五"国家科技支撑计划项目(2012BAD29B06);辽宁省重点实验室开放课题(LNSAKF2013018);辽宁省高等学校创新团队项目(LT2014024);病原微生物生物安全国家重点实验室开放课题(SKLPBS1417;SKLPBS1438);渤海大学博士科研启动项目(BSQD022)

摘　　要：为了建立一种含扩增内标的沙门氏菌PCR检测方法,以细菌16S r RNA为扩增内标对照,以沙门氏菌inv A基因为靶基因设计了一对引物,并优化了PCR反应体系。通过对20种细菌进行PCR检测显示,该方法对沙门氏菌具有良好的特异性。灵敏度实验表明,该检测方法对沙门氏菌纯DNA模板的检测灵敏度为61.1 fg/μL,对沙门氏菌纯培养物的检测灵敏度为2×102 cfu/m L。对人工污染蛋清的检测实验显示,沙门氏菌接种量为2 cfu/25 m L的鸡蛋清样品经过8 h增菌培养后,可被该方法检出。结果表明,该检测方法特异性强、灵敏度高,能排除沙门氏菌PCR检测方法中可能出现的假阴性现象,适用于鸡蛋等食品中沙门氏菌的快速检测。A PCR assay was developed for the rapid detection of Salmonlla using 16S rRNA as internal amplification control. Primers were designed to detect the gene of inv A in Salmonlla and the assay was optimized and assessment. Detection of various bacterias by PCR indicated that this PCR assay was specific for Salmonlla.The sensitivity of detection for Salmonella purified genomic DNA was 61.1 fg/μL and 2 ×102cfu/m L for pure cultures. The detection for artificially contaminated egg white showed that Salmonlla could be detected after eight hours enrichment culture when the Sallmonla inoculation was 2 cfu/25 mL. This detection method had a good specificity and sensitivity,and could eliminate false-negative results in the PCR detection method for Salmonella,and suitable for rapid detection of Salmonella in food.

关 键 词：沙门氏菌 PCR检测 扩增内标对照 假阴性 

分 类 号：TS201.6[轻工技术与工程—食品科学]