砂仁及其混伪品益智仁的ISSR分子鉴别

为创建砂仁及其主要混伪品益智仁的ISSR分子鉴别方法,以UBC818为引物,对影响ISSR-PCR反应体系的引物、dNTPs、DNA模板、Taq DNA聚合酶和Mg2+浓度进行5因素4水平正交优化试验,并在此基础上筛选阳春砂ISSR引物及砂仁正伪品的鉴别引物。结果表明,20μL阳春砂ISSR-PCR最佳反应体系包括引物0.5μmol/L、dNTPs 0.25 mmol/L、DNA模板40 ng、Taq DNA聚合酶1.2 U、Mg^2+2.0 mmol/L;从52条ISSR引物中筛选出10条阳春砂ISSR引物,从6条阳春砂和益智的共同引物中,筛选出扩增条带清晰、多态性强的砂仁正伪品的鉴别引物UBC808,利用UBC808对15份样品进行验证试验,结果表明鉴别体系稳定性好,可用于砂仁与益智仁的快速、准确鉴别。

ISSR Molecular Identification of Amomum villosum and Its Adulterant Alpinia oxyphylllae Fructus

To establish a new method for ISSR molecular identification of Amomum villosum and its adulterants Alpinia oxyphylllae fructus,the ISSR-PCR reaction system for Amomum villosum Lour.was studied by orthogonal design with four levels of primers,dNTPs,DNA template,Taq DNA polymerase and Mg2+concentration by using UBC818 as primer.And the Amomum villosum’s ISSR primer and its primer for identification were screened.The optional parameters for 20μL PCR reaction system contained primer 0.5μmol/L,dNTPs 0.25 mmol/L,DNA template 40 ng,Taq DNA polymerase 1.2 U and Mg^2+2.0 mmol/L.10 ISSR primers were screened out from 52 ISSR primers.From the 6 common primers of Amomum villosum Lour.and Alpinia oxyphylllae fructus,UBC808 with clear amplification bands and strong polymorphism was screened out for identification,and 15 samples were validated which showed that the identification system was stable and it could be used for the rapid and accurate identification of Amomum villosum and Alpinia oxyphylllae fructus.