中药材蛤蚧的特异性PCR鉴定

Authentication of Gecko by Allele-specific PCR

以线粒体Cytb、COI、12SrRNA、16SrRNA基因序列为基础,探讨特异性PCR技术鉴定蛤蚧及伪品的可行性。本研究以所扩增的16条COI序列为引物设计依据序列设计了1对位点特异性引物COISF,COISR,同时从Gen-Bank上下载30条序列,设计了CytbSF,CytbSR;16S rRNASF,16S rRNASR;12S rRNASF,12S rRNASR另外3对位点特异性引物,因此共用4对位点特异性引物分别扩增蛤蚧及伪品实验样本。结果表明在复性温度为65℃时,4对引物都出现了理想的结果,即蛤蚧正品出现了扩增条带,而伪品没有扩增条带。同时对市售的蛤蚧商品进行了检测,在所供的7号标本中,有4号为蛤蚧正品,其余3号为蛤蚧伪品。本研究所设计的位点特异性引物可快捷、准确的鉴定蛤蚧及伪品,且在药检工作中具有极大的应用前景。

To explore the practicability of allele-specific PCR in identifying gecko,we designed a pair of primer COISF,COISR based on 16 sequences.Meanwhile,we downloaded another 30 sequences from the GenBank to design other three pairs of primers CytbSF,CytbSR;16S rRNASF,16S rRNASR;12S rRNASF,12S rRNASR,respectively.At the annealing temperature 65℃,all the genuine gecko present electrophoretic bands,while the adulterants show no electrophoretic bands.Therefore,the four pairs of primers are effective in identifying gecko.Moreover,we test the primers using seven samples from market,the results showed that four of them is genuine,the rest are adulterants.In conclusion,the DNA barcode-specific PCR is effective for identifying gecko and its adulterant.