杨梅SRAP-PCR反应体系的建立与优化

为了建立适宜杨梅基因组DNA的SRAP-PCR扩增体系。以杨梅基因组DNA为模板，通过正交试验设计，从Mg2+、模板DNA、dNTPs、Tap DNA聚合酶和引物5种因素4个水平对杨梅SRAP-PCR反应体系进行优化。各因素对杨梅SRAP-PCR反应的影响程度从大到小依次为：Mg2+，模板DNA，dNTP，引物和Taq DNA聚合酶；建立的杨梅SRAP-PCR最佳反应体系为25μL反应体系中含2.5 mmol/L Mg2+、50 ng DNA模板、0.25 mmol/L dNTPs、0.15 μmol/L引物和1.5 U Taq DNA聚合酶。这一体系的建立为今后利用SRAP-PCR技术开展杨梅分子遗传学研究打下了基础。

Optimization of Sequence-Related Amplified Polymorphism Amplification System for Chinese Bayberry

The aim was to establish SRAP-PCR amplification system which was suitable for Chinese Bayberry genome DNA. The optimized SRAP-PCR amplification system for Chinese bayberry was established by the orthogonal design. The results suggested that the order of factors which affect on the result of SRAP-PCR were Mg^2＋, template DNA, dNTPs, primers and Taq DNA polymerase. A suitable SRAP-PCR system for Chinese bayberry was that total 25 μL reaction system containing 2.5 mmol/L Mg^2＋, 50 ng template DNA, 0.25 mmol/L dNTPs, 0.15 μmol/L primers and 1.5 U Taq DNA polymerase could be able to amplified the most rich polymorphism and clear bands. The optimized SRAP-PCR system was tested on twelve Chinese bayberry germplasms and shown to be steady and reliable for molecular genetics research.