药材与资源石豆兰属植物rDNA ITS序列的克隆与分析

Cloning and analysis of rDNA ITS sequences from plants in Bulbophyllum Thouars

目的通过测定11种石豆兰属植物的ITS序列,为石豆兰属植物分子鉴定和遗传多样性研究提供依据。方法利用PCR法从叶片基因组DNA中克隆ITS序列,并借助生物信息学软件对其进行分析。结果 11种石豆兰属植物的ITS全长为633～645 bp,5.8 S序列长度162 bp,ITS1和ITS2序列变异位点丰富,共168个,其中信息位点95个,序列存在大量的转换、颠换和缺失;5.8 S序列较为保守,仅含6个变异位点和2个信息位点;11种石豆兰属植物的遗传距离为0.002 2～0.212 0,其中齿瓣石豆兰与斑唇石豆兰之间的遗传距离最小,亲缘关系最为接近。序列已上传至NCBI,登录号为JN619409～JN619419。结论获得了11种石豆兰属植物的rDNA ITS序列,为石豆兰属植物分子鉴定和遗传多样性研究奠定了基础。

Objective To provide the evidence for molecular identification and genetic diversity studies, sequencing and comparison of ITS sequences from 11 plants in Bulbophyllum Thouars were performed. Methods ITS sequences were cloned from leaf genomic DNA by PCR and were analyzed using bioinformatics softwares. Results The full ITS and 5.8 S sequences of 11 plants in Bulbophyllum Thouars were 633-645 and 162 bp in length, respectively. Variable sites were rich in ITS1 and ITS2 sequences with 168 variable sites among which 95 were parsimony information sites. Many transitions, transversions, and deletions were observed. 5.8 S sequences were conserved and there were only six variable sites and two parsimony information ones. The genetic distance of 11 plants in Bulbophyllum Thouars ranged from 0.002 2 to 0.212 0. The smallest genetic distance was observed between B. levinei and B. pectenveneris revealing their closest relationship. Sequences were submitted to the NCBI database under the accession numbers of JN619409-JN619419. Conclusion rDNA ITS sequences of 11 plants in Bulbophyllum Thouars are obtained which could provide a foundation for molecular identification and genetic diversity studies.