一个植物乳杆菌新质粒的序列分析与归类

Sequence analysis and classification of a new plasmid isolated from Lactobacillus plantarum

[目的]分离鉴定植物乳杆菌PC518的质粒并分析滚环复制p C194家族复制起点特征。[方法]从植物乳杆菌PC518中提取质粒,HindⅢ单酶切后克隆测序,然后用反向PCR方法验证质粒序列的完整性。使用DNAMAN V6. 0软件和MEGA X软件对43个p C194家族质粒的复制起点序列和复制蛋白进行比对分析。[结果]分离得到一个3 325 bp的新质粒p LP325。43个p C194家族质粒复制起点中:24个在nick上、下游均有反向重复序列,12个只在nick上游有反向重复序列,4个只在下游有反向重复序列。复制蛋白的聚类与复制起点中反向重复序列的位置是对应的。[结论]p LP325的复制方式推定为滚环复制,属于p C194家族。p C194家族复制起点的bind以反向重复序列为特征,位于nick上游或下游。

[Objective] To isolate and identify plasmids from Lactobacillus plantarum PC518. The origin of replication characterized of rolling-circle replication p C194 family were analyzed.[Method]Plasmids were extracted from L. plantarum PC518 and digested by HindⅢ. The digested fragment was cloned and sequenced. After sequencing,the integrity of the plasmid sequence was verified by reverse PCR. DNA MAN V6. 0 software and MEGAX software were used to compare and analyze the origin of replication sequence and replication protein of 43 p C194 family plasmids.[Result]A new 3 325 bp plasmid p LP325 was isolated and purified. Among the 43 plasmid origin of replication of p C194 family,24 plasmid had reverse repeat sequences in both upstream and downstream of nick,12 had reverse repeat sequences only in upstream of nick and 4 had reverse repeat sequences only in downstream of nick. Clustering of replicating proteins corresponded to the location of reverse repeat sequences in the origin of replication.[Conclusion] Plasmid p LP325 should belong to p C194 rolling-circle family. The bind of origin of replication belonged to p C194 family replication was characterized by reverse repeat sequence,which could be located upstream or downstream of nick.