中国玉米新品种DNA指纹库建立系列研究 Ⅲ.多重PCR技术在玉米SSR引物扩增中的应用

Series of Research on Establishing DNA Fingerprinting Database of Chinese New Maize Cultivars Ⅲ.Multiplex PCR Applied in Maize SSR Amplification

根据21对玉米SSR引物扩增片段大小范围组成不同的引物组合进行多重PCR反应的研究。在与单一PCR相同的反应条件下,47个两重PCR组合出现5种不同的扩增情况,其中30个组合扩增正常;10个三重PCR组合中有9个组合扩增正常;两个四重PCR组合中有1个扩增正常,这些筛选出的正常扩增的组合可应用于今后玉米DNA指纹库构建研究中。研究表明,应用与单一PCR完全相同的反应条件,大部分能够获得正常扩增的多重PCR组合,在此基础上提出了简化的多重PCR优化程序。

Twenty-one pairs of maize SSR primers were used for building various multiplex-PCR-sets accord-ing to their amplified bands' size range.Under the same conditions as single PCR reaction,47two-PCR-sets ap-peared five different amplification behaviors,of which30were amplified normally.Nine of10three-PCR-sets and one of2four-PCR-sets were amplified normally.These screened normally-amplified multiplex-PCR-sets could be applied in maize DNA fingerprinting database.The results showed that most multiplex-PCR-sets could be amplified normally under just the same conditions as single PCR.Based on our experience,we proposed a simplified protocol for developing a multiplex PCR assay.