益生菌制品中植物乳杆菌ST-III快速定性、定量检测方法的建立

【目的】建立快速定性、定量检测益生菌产品中植物乳杆菌菌株ST-III的方法，为其实际应用提供参考。【方法】根据植物乳杆菌菌株ST-III与NCBI库中3株已知序列的植物乳杆菌全基因组序列差异，设计菌株特异性引物，进行菌株特异性PCR验证和定性检测；然后建立荧光定量PCR，并检测益生菌制品中菌株ST-III含量。【结果】设计的引物具有良好的菌株特异性；经PCR扩增，含有ST-III的检验样本出现预期特征条带，与预期目的片段（242 bp）相近。建立的荧光定量PCR标准曲线方程为y=-3.369lgx+39.84，R2为0.9990，符合Real-time PCR定量的要求，其检测限为2.85×103 CFU/mL。经检测，样品中植物乳杆菌ST-III的数量分别为3.25×105、1.28×106、4.55×106 CFU/mL。【结论】建立的植物乳杆菌菌株ST-III定性、定量检测方法快速灵敏、测定时间短，可以在实际生产中应用。

Development on rapid qualitative and quantitative detection methods for Lactobacillus plantarum ST-Ⅲ in probiotics products

【Objective】In order to provide references for the reasonable use of L. plantarum ST-III， rapid qualitative and quantitative methods for ST-III detection in probiotics products were established through this research.【Method】The differences in the whole genome sequences between ST-III and the other three Lactobacillus plantarum strains accessed from the NCBI database were compared and the strain specificity primers of ST-III was designed accordingly in order to carry out strain-specific PCR analysis and fluorescent quantitative real-time PCR detection. The established real-time PCR method was used to detect the quantity of ST-III in probiotics products. 【Result】The designed primers had good specificity for ST-III strain. The size of the expected specific band presented in the ST-III samples using PCR amplification was similar to that of the aiming band. The curve of established fluorescent quantitative real-time PCR was y=
-3.369lgx+39.84（R2=0.9990）， which could demand the usage of real-time PCR， and its detection limit was 2.85×103 CFU/mL. By this detection method， the quantity of ST-III in probiotics products was 3.25×105， 1.28×106， and 4.55×106 CFU/mL， respectively. 【Conclusion】The established methods for ST-III detection were rapid， accurate， and time-saving， therefore it could be used in relevant production processes.