摘要
【目的】建立一种快速经济的提取畲药十二时辰基原植物基因组DNA的方法,实现通过分子生药学方法准确鉴别该中草药的真伪,为进一步开发应用该福建地区特色中草药奠定基础。【方法】采用CTAB法、改良CTAB法、试剂盒法、SDS法、高盐低pH法提取十二时辰基原植物叶片基因组DNA,采用琼脂糖凝胶电泳检测DNA的质量和完整性,用优选后的方法提取其基因组DNA,与其混淆品DNA分别进行双向测序,评价该分子生药学方法的鉴别效果。【结果】改良CTAB法提取得到的DNA纯度较高,SSR引物扩增与ITS2序列扩增均可得到清晰明亮的条带,是提取畲药十二时辰基原植物基因组DNA的最佳方法。用该方法提取得到的基因组DNA经PCR扩增后的产物可用于双向测序,进而鉴别它的基源,实现与常见混淆品的准确区分。【结论】本研究建立的改良CTAB法能有效地提取畲药十二时辰的基因组DNA,方法操作简单、提取得到的DNA质量较好,可用于畲药十二时辰与其混淆品的分子生药学鉴别。
【Objective】A rapid and economical method to extract the genomic DNA from Clematis florida Thunb.var.plena D.Don for commercial authentication of the valuable Chinese herbal medicine was established.【Method】CTAB,Testing Kit,SDS,and the high salt/low pH methods,along with a modified CTAB method,were applied to extract genomic DNA from leaves of the herbal plant.The quality and integrity of the extracted DNA samples were compared using agarose gel electrophoresis.Then,by bidirectional sequencing,the selected molecular pharmacognosy method was used to differentiate between the authentic material and a counterfeit for methodology verification.【Result】The modified CTAB method yielded high purity DNA with clear and bright bands on SSR primer and ITS2 sequence amplifications.The PCR amplified products of the extracted DNA were used in the bidirectional sequencing to correctly identify and accurately differentiate the true and faux products.【Conclusion】The improved CTAB method established in this study could effectively extract the genomic DNA of the target plant material.It was simple,rapid and reliable in extracting high quality DNA for molecular pharmacognosy identification on Clematis florida Thunb.var.plena D.Don.
作者
何舒澜
李泳宁
朱扶蓉
HE Shulan;Li Yongning;ZHU Furong(Fujian Health College,Fuzhou,Fujian350101,China)
出处
《福建农业学报》
CAS
CSCD
北大核心
2021年第3期264-270,共7页
Fujian Journal of Agricultural Sciences
基金
福建省卫生计生中青年骨干人才项目(2016-ZQN-75)
福建省中青年教师教育科研项目职业院校专项(JZ180548)
福建卫生职业技术学院科技创新团队项目(2018-1-3)。
关键词
十二时辰
基因组
DNA提取
鉴别
Clematis florida Thunb.var.plena D.Don
genome
DNA extraction
identification
