嗜甲基细菌Serratia Marcescens NH8的筛选及其pqq基因的克隆

以甲醇为唯一碳源，从土壤中筛选获得一株长势优良的嗜甲基营养菌NH8，对其16S rRNA基因序列分析，初步鉴定为粘质沙雷氏菌（Serratia marcescens），命名为Serratia marcescens NH8.通过下载数据库中pqq基因簇序列进行多重比对，设计PCR扩增引物，从Serratia marcescens NH8中成功克隆出吡咯喹啉醌（Pyrroloquinoline Quinone,PQQ）合成相关的pqq基因簇片段.该基因簇序列全长5 535 bp，由6个开放阅读框pqqA，pqqB，pqqC，pqqD，pqqE和pqqF组成，并且与Serratia marcescens WW4的pqq基因簇序列同源性为99%.同时，对pqq基因簇中各基因的结构与功能进行了分析，核酸序列分析结果表明pqqB，pqqC，pqqD，pqqE和pqqF基因构成细菌操纵子发挥作用.

Screening of a Methylotrophic Bacteria Serratia Marcescens NH8 and Isolation of Its Pyrroloquinoline Quinone (pqq) Genes Cluster

In this work,screening by using methanol as the sole carbon source,a methylotrophic bacteria,namely NH8,was isolated from soil samples,and was identified preliminarily as Serratia marcescens based on phylogenetic analysis of 16S rRNA gene sequence.With multiple alignments of nucleotide sequences against pqq genes cluster,some primers amplifying the pqq genes were designed by Primer 5 software.Therefore,the pqq genes cluster,which was related to PQQ biosynthesis,had been isolated from Serratia marcescens NH8 by PCR amplification.The pqq genes fragment of 5 535 bp had six open reading frames including pqqA,pqqB,pqqC,pqqD,pqqE,pqqF genes and showed 99% homology to pqq genes of Serratia marcescens WW4.Meantime,the structures and functions of each gene in pqq genes cluster were analyzed.The result show that 5 genes of pqqB，pqqC，pqqD，pqqE，pqqF construct one operon for regulating PQQ biosynthesis in Serratia marcescens NH8.