细辛属8种药用植物rDNA ITS的克隆与序列分析

Cloning and sequence analysis of rDNA ITS from plants in genus Asarum L.

目的克隆和分析8种细辛属药用植物的ITS序列,为开展该属植物分子鉴定和遗传多样性研究奠定基础。方法以基因组DNA为模板,利用PCR法克隆ITS全长序列,借助生物信息学软件对序列进行比对分析,并构建系统发育树。结果测序结果表明,8种细辛属植物的ITS全长为637~646 bp,其中的5.8 S序列最为保守,长度和碱基组成完全一致;ITS1与ITS2均存在一定的变异,它们的长度分别为255~257 bp和226~232 bp,序列中出现大量插入/缺失和转换/颠换现象,ITS1、ITS2的可变位点和简约信息位点数分别为46/30和14/9。系统发育分析结果表明,8种细辛属植物在进化树上可分为4组,与传统分类结果完全一致,I^IV分别对应细辛组、华细辛组、长花组和杜衡组。结论 8种细辛属植物ITS序列具有丰富的信息位点,可用于这些植物的分子鉴定。

Objective To clone and analyze ITS sequences of eight AsarumL. medical plants, and to provide evidence for both molecular identification and genetic diversity studies. Methods PCR amplifications were performed with genomicDNA as templates to isolate ITS sequences, and sequence comparisons as well as phylogenetic tree construction were conducted by bioinformatics software. Results Sequencing results indicated that the full ITS sequences of eight Asarum plants ranged from 637 to 646 bp in length, and 5.8 S sequence was highly conserved with identical sequence length and nucleotide composition. However, sequence variations were observed in ITS1 and ITS2, their sequences were 255—257 bp and 226—232 bp in length, respectively. Insertion/deletion and conversion/transition events were occurred frequently in ITS1 and ITS2 sequences, with variable sites of 46 and 30 and parsimony informative sites of 14 and 9, respectively. Results of the phylogenetic analysis indicated that the eight plants grouped to four different clades, which were completely coincide with traditional plant taxonomy. Groups of I—IV corresponded to section AsarumL., AsiasarumL., Longiflora L., and Heterotropa（Nakai） Hara, respectively. Conclusion ITS sequences of eight Asarum plants were rich in informative sites, which could be applied as molecular marker to identify these species.