荧光定量PCR检测原料乳中粘质沙雷氏菌

Fluorescence Quantitative PCR Detection of Serratia marcescens in Raw Milk

为建立检测原料乳中粘质沙雷氏菌(S.marcescens)定性定量的检测方法,本研究利用S. marcescens S-核糖基高半胱氨酸酶(luxS)基因片段设计1对特异性引物,设计并建立了S. marcescens luxS基因的SYBR GreenⅠ实时荧光定量PCR的检测方法。结果显示,该方法可特异性检测粘质沙雷氏菌的存在,检测的最低限为6.9×10~2cfu/mL。与其他原料乳中常见的微生物均无交叉反应,比常规PCR的检测限(6.9×10~3cfu/mL)要广。对2015年11月~2016年1月长江下游地区8个牧场的原料乳进行检验,结果表明本检测方法具有较好的敏感性、特异性和实用性。

For creating a qualitative and quantitative detection method to test Serratia marcescens in raw milk,this study created a real time fluorescence quantitative PCR detection method based on a pair of specific primers of S.marcescens luxS gene.The results showed that,this method could specifically detect the presence of S.marcescens without cross reaction to other common microorganisms in raw milk and the measurement sensitivity was 6.9×10^2 cfu/mL.The detection limit of S.marcescens in artificial positive milk samples was 6.9 ×10^3 cfu/mL.Finally,the raw milk of 8 farms in the lower reaches of the Yangtze River from November 2015 to January 2016 was monitored by this method.The results showed good sensitivity,specificity and practicality of this detection method.This study can rapidly detect S.marcescens in raw milk,which effectively improving the quality and safety of raw milk.It can also promote healthy and sustainable development of dairy industry.