羊源杀鲑气单胞菌16S rRNA基因的克隆测序及分析

Cloning and Sequence Analysis of the 16S rRNA Gene of Aeromonas salmonicida in Goat

从重庆市丰都县无菌采集山羊血液,提取全血基因组,用16SrRNA基因的引物进行PCR扩增,得到2条长约1 500bp的扩增片段,将其克隆到pMD19-T载体后进行测序,并与3条豚鼠气单胞菌,3条嗜水气单胞菌,3条中间气单胞菌,4条杀鲑气单胞菌,2条温和气单胞菌,3条维氏气单胞菌和5条舒氏气单胞菌16SrRNA基因序列进行系统发育分析.结果表明:所克隆的基因片段长度均为1 507bp,GenBank登录号为JF823571和JF823572.系统发育分析发现2条序列均被聚类到杀鲑气单胞菌群,并与温和气单胞菌聚类到一个大的分支.同源性比对结果显示所获得的序列与杀鲑气单胞菌法国株(ATCC33658T)X74681和阿根廷株AF134065的同源性最高,均为99.6%,表明重庆地区存在受杀鲑气单胞菌感染的山羊.

Blood samples were collected from goats in Fengdu,Chongqing and the genome DNA was extracted from the whole blood.The 16S rRNA gene of Aeromonas salmonicida was PCR-amplified and two fragments about 1 500 bp in length were obtained,which were then cloned to pMD19-T vector and sequenced.The sequences were submitted to GenBank,the accession number being JF823571 and JF823572,respectively.Phylogenetic analysis of 16S rRNA sequences with 3 homologous sequences from A.caviae,3 from A.hydrophila,3 from A.media,4 from A.salmonicida,2 from A.sobria,3 from A.veroni and 2 from A.schubertil showed that 16S rRNA gene of A.salmonicida spp.had highest homology（99.6%） with that of France（ATCC33658T）X74681 and Argentina AF134065,and those species were more closely related to A.salmonicida than to any other Aeromonas species,indicating that there exists A.salmonicida infection in goats in Chongqing.