荧光定量聚合酶链式反应法检测三七粉掺伪情况的研究

Study on detection of adulteration of Panax notoginseng powder by fluorescence quantitative polymerase chain reaction

目的建立特异性检测三七粉掺伪比例的荧光定量聚合酶链式反应(quantitative polymerase chain reaction,qPCR)方法。方法三七粉经适宜的方法进行核酸提取后,以经过鉴定的正品三七药材粉末核酸提取物作为定量基准,采用特异性引物运用qPCR技术SYBR Green染料法进行扩增,以ΔRn为纵坐标,以百分含量的对数值作为横坐标,绘制标准曲线,建立三七粉掺伪比例相对定量方法。结果在相对含量1%~20%范围内相关系数(r)为0.998,线性关系良好,用人工掺伪25%、50%、75%的样本验证定量方法准确性,准确度在103.2%~122.6%之间,相对标准偏差(relative standard deviation,RSD)均小于15%。结论该方法的线性范围、准确度、精密度均良好,可以用于三七粉掺伪比例的定量测定;另由于伪品没有特异性扩展曲线,该方法还可用于三七粉的真伪鉴别。

Objective To establish a method for the specific detection of adulteration of Panax notoginseng powder by fluorescence quantitative PCR.Methods After nucleic acid extraction of Panax notoginseng powder by an appropriate method,nucleic acid extract of authenticated authentic Panax notoginseng powder was used as the quantitative basis,and the specific primer was used for amplification by fluorescent quantitative PCR technique SYBR Green dye method.The standard curve was drawn withΔRn as the ordinate and the pair value of percentage of content as the horizontal coordinate.A relative quantitative method of adulteration ratio of Panax notoginseng powder was established.Results In the range of relative content of 1%~20%,the correlation coefficient(r)was 0.998,and the linear relationship was good.The accuracy of the quantitative method was verified by artificially adulterated samples of 25%,50%and 75%,and the accuracy was between 103.2%and 122.6%,and the RSD was all less than 15%.Conclusion The linear range,accuracy and precision of the method were good,and it can be used for the quantitative determination of adulteration ratio of Panax notoginseng powder.In addition,the method can also be used to identify the authenticity of Panax notoginseng powder because there was no specific expansion curve.