基因测序技术在中药质量研究中的应用(Ⅱ)——山药基原的DNA测序鉴别

Application of gene technology in quality control of Chinese materia medica Ⅱ. Identification of Chinese Rhizoma Dioscoreae by DNA sequencing

目的 分析正品山药 Dioscorea polystachya Turcz.与地方习用品广山药 D.persimilis Prain et Burkill、土山药 D. japaonica Thunb.和方山药 D.alata L .的核基因组 18S r RNA基因序列 ,为山药基原鉴别及品质评价提供分子依据。方法 采用 PCR直接测序技术测定山药及其地方习用品的 18S r RNA基因核苷酸序列并作序列同源性分析。结果 山药、广山药和土山药的 18S r RNA序列长度均为 1810 bp,而方山药为 180 7bp。根据排序比较 ,广山药与正品山药的 18S r RNA序列完全相同 ,而与土山药和方山药的序列同源性分别为 99.89%和 97.5 1%。结论

Object To distinguish the genuine Dioscorea polystachya Turcz. from other species of Dioscorea L. such as D. persimilis Prain et Burkill, D. japonica Thunb., and D. alata L., conventionally used in some localities by sequencing their nuclear ribosomal RNA subunit (18S rRNA), to provide mole cular evidences for the quality control of drugs of Dioscorea L. Methods By direct PCR sequencing to detect the homology of 18S rRNA sequence of different species of Dioscorea L.. Results The 18S rRNA gene sequence of genuine D. polystachya, D. persimilis and D. japonica showed identical length of 1810 bp, whereas that of D. alata was 1807 bp. Multiple sequence alignment of 18S rRNA gene showed that the homology was identical between D.persimilis and the genuine D. polystachya; but a difference of 99.89% with D. japonica and 97.51% with D. alata. Conclusion DNA sequencing can provide an accurate and reliable mean for the identification of the origin and genuineness of Dioscorea L..