关键词: 沙门氏菌, 荧光定量PCR, SYBR Green I, DNA提取, 生乳, 乳制品

Abstract: Raw milk was susceptible to Salmonella spp., which was reproduced rapidly. Nevertheless, there was only the potential of very small amount of Salmonella spp. in milk products since sterilization and other treatments though the bacterial population decreased. In this paper, we studied the pretreatment methods of raw milk and milk products according to their contamination levels and optimized the extraction conditions of bacterial total DNA by the boiling lysis procedure. Furthermore, we set up a systemic detection technology for milk-borne Salmonella spp., based on SYBR Green I real\|time PCR targeting the invA gene. The developed methods possessed specificity for Salmonella spp. As far as raw milk was concerned, the limit of detection of the method was 102 cfu·mL-1, the linear range of the standard curve was 102-108 cfu·mL-1, correlation coefficient(R2) was 0.999, and amplification efficiency was 103%. For milk products, 1 cfu of Salmonella spp. present in 25 g (mL) milk samples was shown detectable by this method with an enrichment step of 16 h. This technology could be used to quantitatively detect Salmonella spp. in raw milk only for 3 h, and qualitatively assay milk products only for 1 d. All in all, our work provided a high\|throughout detection technology of Salmonella spp. for dairy enterprises, which was proved rapid, simple, sensitive and accurate with a good reproducibility

Key words: Salmonella spp., real-time PCR, SYBR Green I, DNA extraction, raw milk, milk products