基于生药学和DNA条形码技术的丁药鉴定研究

Identification of Ding Medicine based on Pharmacognosy and DNA Barcoding

目的:基于生药学鉴别和psbA-trnH序列,采用DNA条形码技术对云贵地区民间丁药药材样品进行基原鉴定研究。方法:通过采集丁药药材样品,运用植物学性状分析、显微镜观察和薄层色谱技术,筛选出关键鉴别特征。同时提取样本基因组总DNA,基于psbA-trnH序列设计特异性引物扩增丁药基因序列,经测序、拼接和校对后,通过美国国立生物技术信息中心数据库的局部序列对比工具进行比对,鉴定其物种来源。结果:丁药原植物形态,药材粉末与解离组织的显微镜特征明显,其薄层色谱呈现出专属的斑点,具备有效鉴别价值。进一步的psbA-trnH序列分析显示,丁药与Orixa japonicac Thunb.(臭常山)的psbA-trnH序列相似度极高,达到99.80%。结论:根据生药学特点和psbA-trnH序列作为DNA条形码能够对丁药的基原植物进行鉴别,鉴定其为臭常山,这为保障云贵地区丁药药材来源的科学鉴定提供参考。

Objective:To conduct a basic identification study of folk Ding medicine herb samples in Yunnan and Guizhou regions with DNA barcoding technology,based on the pharmacognosy identification and psbAtrnH sequences.Methods:By collecting the samples of Ding medicine herb,the key identification features were screened out by using botanical trait analysis,microscopic observation,and thin-layer chromatography(TLC)technique.Meanwhile,the total genomic DNA of the samples was extracted,and specific primers were designed to amplify the gene sequences of Ding medicine based on psbA-trnH sequences,after sequencing,splicing and proofreading,the species origin was identified by BLAST comparison in the NCBI database.Results:The original botanical form of Ding medicine,the microscopic features of powdered and dissociated tissues of the herb were obvious,and its thin-layer chromatography showed exclusive spots that possessed effective identification value.Further psbA-trnH sequence analysis showed that the psbA-trnH sequence similarity between Ding medicine and Orixa japonica Thunb.was extremely high,reaching 99.80%.Conclusion:Based on the pharmacognosy characteristics and psbA-trnH sequences as DNA barcodes,it is possible to identify the basal plant of Ding medicine as Orixa japonica Thunb.which provides a reference to ensure the scientific identification of the source of Ding medicine in Yunnan and Guizhou regions.