植物乳杆菌的DPO引物实时荧光PCR检测方法  

Real-time PCR for the detection of Lactobacillus plantarum based on dual priming oligonucleotide system

作　　者：孙凯[1] 魏霜[2] 刘玉莉[2] 许如苏[2] 刘中勇[2] 鄞杰平[2] 袁俊杰 吴希阳[1] 

机构地区：[1]暨南大学理工学院食品科学与工程系 [2]汕头出入境检验检疫局,广东汕头515041 [3]湛江出入境检验检疫局,广东湛江524000

出　　处：《食品与发酵工业》2016年第9期178-183,共6页Food and Fermentation Industries

基　　金：国家质量监督检验检疫总局科技计划项目(2015IK078)

摘　　要：根据已报道的植物乳杆菌（Lactobacillus plantarum）的23S rRNA基因保守序列,设计用于检测植物乳杆菌的特异性DPO（dual priming oligonucleotide）引物,结合SYBR GreenⅠ实时荧光PCR技术,建立植物乳杆菌的DPO引物实时荧光PCR检测方法,对其特异性和灵敏度进行了评价,并将建立的检测方法用于实际样品的检测中。结果显示,该方法对2株植物乳杆菌能得到阳性扩增,其余11株乳酸菌及阴性对照没有扩增曲线,灵敏度高达0.001 ng/μL,而且在退火温度为50~60℃范围内不影响其特异性,表明该方法对退火温度不敏感;用微生物菌制剂和酸奶共13种益生菌产品进行了验证,检出情况与产品标识一致。因此,该研究建立的SYBR GreenⅠ实时荧光PCR能够快速、准确、高效地检测微生物菌制剂及其他益生菌产品中的植物乳杆菌。The species-specific DPO（ dual priming oligonucleotide） primers were designed basing on the 23 S rRNA gene sequences of Lactobacillus plantarum. A SYBR GreenⅠreal-time PCR assay based on DPO primers was developed for the detection of L. plantarum. The specificity and sensitivity of the assay have been estimated. The results showed that this method was of high specificity at annealing temperature range from 50 to 60 ℃,and only two L.plantarum strains could be amplified. The other 10 Lactobacillus. spp. strains gave negative results,and the sensitivity of this method reached 0. 001 ng / μL. The specificity and sensitivity of the assay was evaluated using 13 probiotics products including microbiological preparation and yogurt,and the results were in consistent with the product identification. Therefore,L. plantarum can be detected rapidly,accurately and efficiently by this method.

关 键 词：植物乳杆菌 实时荧光PCR DPO引物 检测 

分 类 号：TS201.3[轻工技术与工程—食品科学]