Rep-PCR应用于快速鉴定啤酒污染菌的研究

为评价rep-PCR在快速鉴定啤酒污染菌中的应用,首先比较了DNA提取方法,确定CTAB法作为制备rep-PCR的DNA模板的方法。并通过PCR产物直接测序的方法,从分离菌中鉴定得到11种常见的啤酒污染菌。用BOXA1R和(GTG)_5引物扩增分离菌,采用Gel ComparⅡ软件处理电泳图,构建污染菌的标准指纹图库。经过聚类分析表明,BOXA1R和(GTG)5对Lactobacillusbrevis、L.buchneri、L.casei/paracasei、L.plantarum和L.fermentum的聚类效果具有互补性,并首次提出指纹比对快速鉴定的相似系数阈值的概念。对来自三个不同来源的9株乳酸菌的快速鉴定结果表明,rep-PCR鉴定技术简单、快速、可靠,在快速鉴定方面将具有重要的应用价值。

Study on Application of rep-PCR Fingerprint in Rapid Identification of Beer-spoilager

The application potential of rep-PCR in typing beer-spoilage isolates was studied. The effects of different factors, including DNA templates and primers, on the quality and reproducibility of fingerprints were investigated. The CTAB protocol was shown to be the feasible method for DNA extraction. Primers BOXA1R and (GTG)5 were used in rep-PCR, and the PCR products were sequenced to identify strains isolated from two breweries. Rep-PCR fingerprint profiles were obtained by using GelCompar II software. Cluster analysis showed that the isolates belonging to Lactobacillus brevis, L. buchneri, L. casei/paracasei, L. plantarum are divided into 2 or 3 subgroups. In addition, the two rep-PCR fingerprint profiles complemented with each other in typing these isolates. Combining the similarity coefficient cut-off (SCC) of species, 9 unknown isolates were identified rapidly by using both fingerprint databases. The results indicate that rep-PCR is a simple, reliable and promising method for rapid identification of beer-spoilager.