Chen Kang, Jiang Chao, Yuan Yuan, Huang Lu-Qi, Li Man
Zhongguo Zhong Yao Za Zhi. 2014 Oct;39(19):3673-7.
To obtained an accurate, rapid and efficient method for authenticate medicinal snakes listed in Chinese Pharmacopoeia (Zaocysd humnades, Bungarus multicinctus, Agkistrodon acutus), a rapid PCR method for authenticate snakes and its adulterants was established based on the classic molecular authentication methods. DNA was extracted by alkaline lysis and the specific primers were amplified by two-steps PCR amplification method. The denatured and annealing temperature and cycle numbers were optimized. When 100 x SYBR Green I was added in the PCR product, strong green fluorescence was visualized under 365 nm UV whereas adulterants without. The whole process can complete in 30-45 minutes. The established method provides the technical support for authentication of the snakes on field.
为获得一种准确、快速且高效的方法来鉴别《中国药典》中收录的药用蛇类（乌梢蛇、金钱白花蛇、蕲蛇），基于经典分子鉴别方法建立了一种快速PCR鉴别蛇类及其混伪品的方法。采用碱裂解法提取DNA，通过两步PCR扩增法扩增特异性引物。对变性温度、退火温度和循环次数进行了优化。当在PCR产物中加入100倍的SYBR Green I时，在365nm紫外光下可观察到强烈的绿色荧光，而混伪品则无此现象。整个过程可在30 - 45分钟内完成。所建立的方法为野外蛇类鉴别提供了技术支持。
