TaqMan荧光定量PCR在饲料沙门氏菌检测中的应用评估  

Assessments of a TaqMan based real-time PCR assay for the detection of Salmonella in feeds

作　　者：佘容 罗丹[1] 刘耀敏 范秀丽 颜其贵 

机构地区：[1]通威股份有限公司检测中心,四川成都610041 [2]四川农业大学动物医学院,四川温江611130 [3]四川农业大学动物疫病与人类健康四川省重点实验室,四川温江611130

出　　处：《中国预防兽医学报》2015年第12期947-951,共5页Chinese Journal of Preventive Veterinary Medicine

基　　金：教育部长江学者和创新团队发展计划项目(IRTO848);四川省科技支撑计划项目(2014NZ0003)

摘　　要：为评估Taq Man荧光定量PCR应用于饲料沙门氏菌检测的可行性,本研究以沙门氏菌JEO402-1基因为靶基因,并采用已建立的沙门氏菌Taq Man荧光定量PCR检测方法,进行了特异性、灵敏度、检测限等相关方法学验证。结果显示:Taq Man荧光定量PCR法特异性良好,在饲料加标样本检测中的最低检测限为2×10^(-1)cfu/m L。重复性试验表明,在不同浓度样本中的变异系数均低于1.0%,具有较高的重复性和稳定性。利用该方法和传统培养法对72份饲料样本进行检测,结果显示两种方法检测结果符合率为65.3%,Taq Man荧光定量PCR法阳性检出率比传统培养法高34.7%;传统培养法检测阳性样本,PCR法检测结果均为阳性。以上结果表明,本研究建立的Taq Man荧光定量PCR法可以应用于饲料样本的沙门氏菌检测。To evaluate the specificity and sensitivity of TaqMan real-time PCR for the detection of Salmonella in feed, the JEO402-1 gene of Salmonella was chosen as the target gene, and the optimized method was applied for the detection of Salmonella from Salmonella CMCC10115 strain artificially Salmonella-contaminated and real feed samples. The results showed that TaqMan real-time PCR showed high specificity and sensitivity, and the detection limit was as low as 2 xl0-1 cfu/mL in artificially Salmonella-contaminated feed samples. Meanwhile, the coefficient of variation was less than 1.0% in the replicate test, indicating the assay was stable, reliability and reproducibility. With 72 real feed samples, TaqMan real-time PCR exhibited as 65.3% consistence as traditional culturing method. Moreover, TaqMan real-time PCR showed higher sensitivity, which was supported by the higher detectable rate than that of traditional method. These results suggest that the TaqMan real-time PCR with JEO402-1 as the target gene could be applied for the detection of Salmonella in feed samples.

关 键 词：沙门氏菌 TaqMan荧光定量PCR法 检测限 

分 类 号：S852.61[农业科学—基础兽医学]