基于16SrRNA和gyrB基因串联DNA特征序列的副溶血性弧菌鉴定

Identification of Vibrio parahaemolyticus based on the concatenated DNA sequencing of 16S rRNA and gyrB gene

【目的】建立基于16S核糖体RNA(16S rRNA)和DNA促旋酶亚基B(gyrB)基因串联DNA特征序列的副溶血性弧菌鉴定方法,为菌株鉴定和分类提供研究思路和方法依据。【方法】选择副溶血性弧菌、溶藻弧菌以及弧菌属的其他代表性菌株,分别以16S rRNA和gyrB基因为研究靶点,设计引物进行特异性核酸序列扩增和核酸测序,将串联后的核酸序列进行聚类比对分析,判断鉴定结果。【结果】副溶血性弧菌鉴定的16S rRNA+gyrB基因串联DNA特征序列单独聚类为1个分支,在种的鉴定水平具有较好的分辨力。【结论】基于16S rRNA+gyrB串联DNA特征序列,可将副溶血性弧菌鉴定到种的水平,是该菌株污染物的系统发育分析及在食品药品安全监管中的风险溯源的方法与依据。

[Objective]To establish the concatenated DNA sequencing of 16S ribosomal RNA(16S rRNA)and DNA gyrase subunit B(gyrB)gene,and provide evidence for the identification and classification of Vibrio parahaemolyticus(V.parahaemolyticus).[Methods]Typical strains in the genus of Vibrio spp.was selected,such as V.parahaemolyticus,V.alginolyticus and other species for examination of 16S rRNA and gyrB gene as target.Primers were separately designed to amplify these two nucleotide fragments.Phylogenetic analysis was performed using the concatenated sequences.[Results]The concatenated 16S rRNA+gyrB nucleotide sequence of V.parahaemolyticus formed a single cluster in the phylogenetic analysis,which identified the typical strains of Vibro spp.at the species level.[Conclusion]In our study,an identification method of V.parahaemolyticus is established based on concatenated 16S rRNA+gyrB nucleotide sequencing.It can identify the strains of V.parahaemolyticus at the species level,which may be applied in phylogenetic analysis and contamination tracing of V.parahaemolyticus in food and drug control.