基于线粒体16S rDNA序列的深圳水库区双脐螺分子鉴定

摘要

Objective To identify the species of Biomphalaria snails collected in Shenzhen reservoir,based on the mitochondrial 16S rDNA sequences.Methods The 16S rDNA fragments were amplified by PCR from the genome DNA of Biomphalaria snails,and inserted in plasmid pMD-18T for sequencing.The sequence of 16S rDNA fragment and its phylogenetic relationships with those of other species of Biomphalaria snails were analyzed with BLAST and MEGA4 software.Results The amplified 16S rDNA fragment of the Biomphalaria snails was about 466 bp in length.As aligned with the corresponding sequences of the related Biomphalaria species,the identity of nucleotides was 99％ with 1 isolate of Biomphaltria straminea (B.straminea),98％ with 3 isolates of B.kuhniana,95％ with 1 isolate of B.intermedia,and 94％ with 1 isolate of B.edisoni.Based on the 16S rDNA sequence,the results of phylogenetic analysis with neighbor-joining (NJ) and unweighted pair-group method with arithmetic means (UPGMA) indicated that the snails had close genetic relationships with the B.straminea isolate (Genbank accession NO.AY030213.1) Conclusion The Biomphalaria snails collected in Shenzhen reservoir could be classified as B.straminea based on the characteristics of 16S rDNA sequence.%目的 基于线粒体16S rDNA序列对深圳水库区双脐螺进行分子鉴定.方法 以双脐螺基因组DNA为模板,聚合酶链反应扩增16S rDNA目的基因片段,并以pMD18-T载体克隆后进行序列测定,采用BLAST和MEGA4软件分析序列特征.结果 双脐螺线粒体16S rDNA扩增片段大小约为466 bp;测定序列与GenBank中1株巴西藁杆双脐螺(Biomphalaria straminea,B.straminea)同源性为99％;与3株库恩双脐螺(B.kuhniana)同源性均为98％;与l株中介双脐螺(B.intermedia)和l株双脐螺(B.edisoni)的同源性分别为95％和94％.深圳水库区2株双脐螺与巴西藁杆双脐螺(序列号AY030213.1)的遗传距离为0;基于16S rDNA序列,采用MEGA4软件作系统发生分析,以邻位连接法(NJ)和非加权组平均法(UPGMA)构建的系统发生树,水库区双脐螺与该藁杆双脐螺属于同一个分支.结论 根据16S rDNA序列特征,深圳水库区双脐螺可归类为藁杆双脐螺.