RAPD-PCR快速鉴定啤酒污染菌的研究

本研究主要评价RAPD-PCR技术在快速鉴定啤酒污染菌中的应用。首先评价了CTAB法和试剂盒提取DNA模板对RAPD指纹稳定性的影响以及乳酸菌专一性BP引物扩增的16SrDNA的5’末端740bp序列用于鉴定菌种的可行性,采用PCR产物直接测序鉴定分离的污染菌,构建标准污染菌库。对分离菌M-13的RAPD指纹聚类分析表明,相同来源的同一种菌能很好地归类在一起。根据建立的快速鉴定流程和初步确定的菌种相似性阈值(SCC),对8株分离菌的鉴定表明,RAPD-PCR技术是一项简单、快捷、可靠性强的鉴定技术,为研究啤酒酿造过程的污染菌提供了一个非常有用的快速鉴定工具。

Study on Application of RAPD Fingerprint on Rapid Identification for Beer Spoilage

The application of RAPD (random amplified polymorphic DNA) as a method for rapid identification for beer spoilage was evaluated in the paper. The effects of CTAB (cetyltrimethylammonium bromide) DNA extraction method and commercial kit on the fingerprint reproducibility were compared and it showned that the variability of RAPD fingerprint was small. The possibility to identify isolates with the 740bp sequence from 5' end of 16SrDNA was evaluated, which could be amplified with primer BP. The isolates were identified by PCR (polymerase chain reaction) product direct sequencing method. A small library was implemented by cluster analysis of RAPD fingerprint of those strains with primer M-13, which was proved to be useful for the isolates classification. Rapid identification of eight isolates was carried out according to the identification procedure and cut- off species similarity coefficient. It suggested that the RAPD-PCR technique is simple, fast and reliable for identification, proved to be an useful tool for studying the beer spoilage in the process of brewing.