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地鳖的环介导等温扩增快速鉴别

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  • 万方数据
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目的:建立基于环介导等温扩增技术(LAMP)的地鳖分子鉴定方法.方法:针对地鳖CO Ⅰ序列设计特异性引物2对(外引物DB-F3、DB-B3和内引物DB-FIP、DB-BIP),在含有Bst DNA聚合酶和甲基红-苯酚红指示剂的反应体系下于63℃下恒温扩增90 min,通过目测检视判断反应结果,与DNA条形码和聚合酶链式反应(PCR)鉴定方法进行对比,考察方法专属性和灵敏度.结果:11批地鳖样品均由紫红色变为橙黄色,1批冀地鳖、9批金边土鳖和1批伪品龙虱均为紫红色,鉴别结果与DNA条形码,PCR结果一致,LAMP检出限为0.984 ng·mL-1.结论:本项目建立的LAMP检测方法准确,灵敏度高,操作简便快捷,对设备要求低,可应用于地鳖鉴别的快速筛查.
Rapid authentication of Eupolyphaga sinensis by loop-mediated isothermal amplification
Objective:To establish an authentication method for Eupolyphaga sinensis based on loop-mediated i-sothermal amplification(LAMP).Methods:Two pairs of primers(outer primer DB-F3,DB-B3 and inner primer DB-FIP,DB-BIP)were designed according to Eupolyphaga sinensis CO Ⅰ specific loci.The reaction system containing template,primers,Bst DNA polymerase and methyl red-phenol red indicator amplified at 63℃ for 90 min.The LAMP results were determined by visual observation and compared with DNA barcoding and pol-ymerase chain reaction(PCR)methods to investigate the specificity and sensitivity.Results:All 11 batches of the authentic Eupolyphaga sinensis sample tubes colour changed from purple red to orange yellow,while sample tubes of 1 batch of Steleophaga plancyi,9 batches Opisthoplatia orientalis and 1 batch of the adulterant,Cybister tripunctatus orientalis,remained purple red after the reaction.The LAMP results were consistent with those of DNA barcoding and PCR.The detection limit of LAMP was 0.984 ng·mL-1.Conclusion:The established LAMP method is ac-curate,sensitive and easy to operate,low equipment required,and can be applied to the rapid screening of the authentication of Eupolyphaga sinensis.

Eupolyphaga sinensisloop-mediated isothermal amplificationDNA barcodingpolymerase chain reactionauthentication of Chinese medicinal materialsvisual checkrapid identification

陈伟韬、李养学、江洁怡、谢灿辉、贾德政、刘晓彤、廖惠娜、李素梅

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广东省第二中医院(广东省中医药工程技术研究院)广东省中医药研究开发重点实验室,广州 510095

广州中医药大学第五临床医学院,广州 510405

广州新华学院药学院,广州 510520

地鳖 环介导等温扩增技术 DNA条形码 聚合酶链式反应 中药材鉴别 目视检查 快速鉴别

广东省中医药局科研项目2021年省属科研机构稳定性支持项目

20244001粤财科教[2021]113号

2024

10.16155/j.0254-1793.2024-0035

药物分析杂志
中国药学会

药物分析杂志

CSTPCD北大核心
影响因子：1.039
ISSN：0254-1793
年,卷(期)：2024.(9)
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