快速PCR技术鉴别中药材蛤蚧的方法研究

摘要

目的:建立简单快速鉴别蛤蚧的特异性PCR方法.方法:对比蛤蚧及其伪品细胞色素C氧化酶Ⅰ亚基基因(CO Ⅰ)序列,设计蛤蚧特异性PCR鉴别引物,优化特异性PCR条件,对蛤蚧及其7种常见混淆品进行扩增及荧光检测.结果:扩增产物经琼脂糖凝胶电泳和荧光检测,所有蛤蚧药材均能扩增出约400 bp的特异性条带,加入SYBR Green Ⅰ染料后,在365 nm下出现强烈绿色荧光,混伪品不具特异条带和绿色荧光,鉴别操作可在30 min内完成.结论:快速PCR结合荧光染料检测可快速鉴别蛤蚧及其常见伪品,为蛤蚧的快速鉴定提供了新的思路和方法.%Objective:To establish a rapid specific PCR method to authenticate gecko and its adulterants.Methods:Specific PCR primers were designed based on CO Ⅰ sequences and the annealing temperature of PCR reaction conditions was optimized,which was performed to authenticate gecko and its 7 adulterants combined with fluorescence analysis.Results:Gecko could obtain a 400 bp specific band in electrophoresis and a great green fluorescence was also observed under 365 nm UV lamp,while the adulterants did not.Conclusion:The results indicated that rapid specific PCR combined with SYBR green Ⅰ fluorescence detection could authenticate gecko and its adulterants rapidly and simply,which provided a new idea and way for gecko rapid identification.