乌梢蛇DNA检测试剂盒的研制与应用
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摘要
目的研制乌梢蛇DNA提取和检测试剂,对试剂盒的特异性、稳定性和重复性进行评价,并对市售乌梢蛇药材进行质量检测,最终开发乌梢蛇DNA检测试剂盒。方法应用现代的DNA指纹技术,对2015年版《中国药典》乌梢蛇DNA检测方法进行了优化和改良,研制了乌梢蛇DNA提取和检测试剂,开发了乌梢蛇PCR检测试剂盒,并对试剂盒检测参数进行评价。应用该试剂盒对随机抽取北京、天津、长春和吉林4个城市的18个乌梢蛇样品,进行检测,同时与药典DNA方法进行比对试验。结果乌梢蛇DNA检测试剂盒经反复冻融5、10和20次后依然有效。18个市售乌梢蛇样品中14个样品为正品,4个样品为伪品。多次试验特异性、稳定性和重复性结果完全相同。试剂盒法与药典法检测结果一致。且试剂盒法可在普通实验室内完成DNA检测任务。结论乌梢蛇DNA检测试剂盒对乌梢蛇药材鉴别简单方便、准确有效。适用于乌梢蛇DNA的快速检测。
OBJECTIVE To develop Zaocys dhumnades DNA extraction and detection kit,evaluate the specificity,stability and repeatability of the kit,and investigate the quality of commercial black tip snake products using the kit. METHODS Using the modern DNA fingerprint technology,the pharmacopoeia Zaocys dhumnades DNA detection method was optimized and improved,and Zaocys dhumnades DNA extraction and detection reagent was developed to obtain the tip snake PCR detection kit. The performance parameters of the kit were evaluated. The kit was used on 18 commercial black tip snake products randomly sampled from Beijing,Tianjin,Changchun and Jilin City. The results were compared with those got by using pharmacopoeia method of DNA test. RESULTS Zaocys dhumnades DNA testing kit was still valid after repeated freezing and thawing for 5,10 and 20 times. Among the 18 commercial black tip snake samples 14 were quality goods,and four were adulterants. Repeated experiments of specificity,stability and repeatability displayed exactly the same results. The test result of the kit method was consistent with the pharmacopoeia method. And the kit method could fulfil the DNA testing task in ordinary laboratories. CONCLUSION Zaocys dhumnades DNA testing kit can identify tip snake products conveniently,accurately and effectively,which can be applied in the rapid detection of Zaocys dhumnades DNA.
OBJECTIVE To develop Zaocys dhumnades DNA extraction and detection kit,evaluate the specificity,stability and repeatability of the kit,and investigate the quality of commercial black tip snake products using the kit. METHODS Using the modern DNA fingerprint technology,the pharmacopoeia Zaocys dhumnades DNA detection method was optimized and improved,and Zaocys dhumnades DNA extraction and detection reagent was developed to obtain the tip snake PCR detection kit. The performance parameters of the kit were evaluated. The kit was used on 18 commercial black tip snake products randomly sampled from Beijing,Tianjin,Changchun and Jilin City. The results were compared with those got by using pharmacopoeia method of DNA test. RESULTS Zaocys dhumnades DNA testing kit was still valid after repeated freezing and thawing for 5,10 and 20 times. Among the 18 commercial black tip snake samples 14 were quality goods,and four were adulterants. Repeated experiments of specificity,stability and repeatability displayed exactly the same results. The test result of the kit method was consistent with the pharmacopoeia method. And the kit method could fulfil the DNA testing task in ordinary laboratories. CONCLUSION Zaocys dhumnades DNA testing kit can identify tip snake products conveniently,accurately and effectively,which can be applied in the rapid detection of Zaocys dhumnades DNA.
引文
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[5]LIU Y,YANG M Y,ZHANG L H.Identification and characterization of DNA finger print between penis ettestis of cervusnippon temm ink and bullwhip[J].Lishizhen Med Mater Med Res(时珍国医国药),2010,21(4):993-995.
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