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目的研究温郁金Curcuma wenyujin的SRAP-PCR扩增体系最适宜的条件。方法以提取纯化温郁金基因组DNA为模板,利用PCR扩增技术,对SRAP反应中主要影响因素Mg2+浓度、d NTP浓度、模板DNA质量浓度、引物浓度、Taq聚合酶浓度进行优化。结果根据实验确定的最佳体系为25μL SRAP反应体系,其中Mg2+2.0 mmol/L,Taq聚合酶2.0 U,引物浓度1.6μmol/L,模板DNA 0.4 ng/μL,d NTP 2.0 mmol/L,10×PCR缓冲液2.5μL。结论通过扩增条件优化实验,扩增的产物条带清晰明亮、重复性好,确定的反应体系及反应程序适用于温郁金的SRAP分子标记,为今后温郁金的遗传多样性研究奠定了基础。
Objective To study the optimal conditions of SRAP-PCR amplification of Curcuma wenyujin. Methods The purified genomic DNA from P. tepidum was used as a template to optimize the factors affecting the SRAP reaction such as Mg2 + concentration, dNTP concentration, DNA template concentration, primer concentration and Taq polymerase concentration by PCR amplification. Results The optimum system was 25μL SRAP reaction system, including 2.0 mmol / L Mg2 +, 2.0 U / L Taq polymerase, 1.6 μmol / L primer, 0.4 ng / μL template DNA and 2.0 mmol / L dNTP × PCR Buffer 2.5 μL. CONCLUSION: The amplified product bands are clear and bright with good reproducibility through the optimization of amplification conditions. The determined reaction system and reaction procedure are suitable for the SRAP molecular marker of warm turmeric, which lays the foundation for the future research on the genetic diversity of turmeric.