正交优化芒果SRAP扩增体系及引物筛选

Optimized Conditions of Sequence-related Amplified Polymorphism(SRAP) System and Primer Screening for Mango

利用正交实验设计,对芒果SRAP-PCR反应的5因素(Mg2+、dNTPs、模板DNA、引物和Taq DNA聚合酶)4水平的扩增效果进行研究,确立适合芒果SRAP-PCR反应的最佳体系。总体积25μL的SRAP-PCR优化的最佳反应体系中含有:2.5μL 10×buffer、20 ng模板DNA、2.5 mmol/L Mg2+、0.2 mmol/L dNTPs、0.4μmo1/L Primer、Taq DNA聚合酶1.5 U。并运用该体系从153对引物组合中初步筛选出50对SRAP引物组合。建立的SRAP标记可为芒果遗传多样性、分子标记及辅助育种等提供研究基础。

An orthogonal design was used to optimize the SRAP amplification system including five factors （the concentration of Mg2＋, dNTPs, template DNA, primer and Taq DNA polymerase） at four levels for mango. The optimum SRAP-PCR system was established, namely 25 μL reaction system containing 2.5 μL 10×buffer, 20 ng template DNA, 2.5 mmol/L Mg2＋, 0.2 mmol/L dNTPs, 0.4 μmol/L Primer, 1.5 U Tot/DNA polymerase. Under the optimum reaction conditions, 50 out of 153 SRAP primer combinations were selected. The results also indicated that the optimized conditions of SRAP could provide an effective means for the research of genetic diversity, molecular marker and auxiliary breeding in mango.