仿刺参(Apostichopus japonicus) mtDNA三个基因片段的序列分析

采用PCR技术对中国烟台、威海和莱州三个地点仿刺参的16S rRNA、COI、IrRNA-COI三个mtDNA基因片段进行了扩增和测序,分别得到了长度约为570bp、640bp和900bp的三段序列,分析了三个采样点样品的遗传多样性。结果表明,每段基因序列扩增均获得两种单倍型,已在Genbank中注册(注册号码:AY852278—AY852283)。通过比较,三个地点样品的序列差异不显著,无缺失、插入、颠换的现象,只有个别位点出现转换,其中16S rRNA序列最为保守,16S rRNA、COI、IrRNA-COI三段序列A T的平均含量为56·2%、59·2%、61·8%,均大于G C含量。三个采样点样品同源性为99·84%—99·96%。将获得的仿刺参DNA序列和Genbank中的来自东太平洋8个外群进行基因序列比较,结果发现,不同目、科的海参的序列差异显著。采用DNAsp统计了各类位点数;MEGA2·1分析了碱基组成和遗传距离,构建了系统树。系统树体现的分类关系与这些物种的形态学分类关系完全一致。

SEQUENCE ANALYSIS OF THREE MITOCHONDRIAL GENES OF SEA CUCUMBER (APOSTICHOPUS JAPONICUS)

Fragments from three mitochondrial genes of Apostichopus japonicus sampled from three sites (Yantai, Weihai, Laizhou, all in Shandong, China) were amplified and sequenced. 570bp 16S rRNA, 640bp COI and 900bp IrRNA-COI fragments were obtained. Analysis indicated that: 1) two haplotypes were identified from every mtDNA fragment, and the number of accession was from AY852278 to AY852283 in Genbank; 2) there was no prominent difference among the three groups and no insertion, deletion or transversions among them except for a few transitions; 3) the mean A T contents of 16S rRNA, COI and IrRNA-COI gene fragments were 56.2%, 59.2% and 61.8%, respectively, which exceeded the contents of G C; 4) the homology among specimens was from 99.84% to 99.96%. The sequences of A. japonicus in this study were compared with those of 8 related outgroups coming from Genbank. Prominent difference existed among different orders or families. The sequences in the samples have higher homology with Parastichopus californicus and P. parvimensis, and significant difference with that of five Dendrochirotida groups. Our comparative study showed 324 sites of insertion or deletion, 388 monomorphic sites, 291 polymorphic sites and 243 parsimony informative sites in IrRNA-COI sequence; 399 monomorphic sites, 241 polymorphic sites and 186 parsimony informative sites in total 641 sites of COI sequence. In the sequences of 16S rRNA, 363 sites of insertion or deletion, 154 monomorphic sites; 125 polymorphic sites; 110 parsimony informative sites were observed. MEGA 2.1 was used to analyze the base composition and genetic distance, and the NJ and MP phylogenetic trees were constructed. The mean A T contents exceeded the contents of G C, and the contents of G were lower than other bases. The evolutionary relationship of 11 groups in phylogenetic NJ and MP tree supports the existing morphology taxon, so did the genetic distance. The specimens in this study belonged to the Aspidochirotida Stichopodidae, so they clustered with P. californicus and P. parvimensis at first.