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目的:以高粱丝黑穗病2381(抗病)、矮四(感病)为材料,以优化SSR反应体系为目的。方法:采用CTAB法提取高粱基因组总DNA,用SSR分子标记技术对其进行多态性扩增,通过对体系中不同的Taq酶浓度、模板浓度、dNTP浓度和引物浓度的梯度分析。结果:建立并优化了的SSR-PCR反应体系为20μl反应体系:dNTP浓度为200μmol/L、Taq酶浓度为1.5U、DNA浓度为100ng和引物浓度为0.4μmol/L。达到了较理想的扩增效果。用该体系对94对SSR引物进行了筛选,其中47对引物扩增出了多态性谱带,其中Xtxp3和Xtxp13在抗感的品种间扩增出了差异谱带。结论:应用优化的SSR反应体系可以对高粱丝黑穗病基因进行分析。
OBJECTIVE: To optimize the SSR reaction system using sorghum smut 2381 (resistant to disease) and dwarf four (susceptible) as materials. Methods: Total DNA of sorghum genomic DNA was extracted by CTAB method and amplified by SSR molecular marker. The concentration of Taq enzyme, template, dNTP and primer were analyzed by gradient analysis. Results: The established SSR-PCR reaction system was 20μl reaction system: dNTP concentration of 200μmol / L, Taq enzyme concentration of 1.5U, DNA concentration of 100ng and primer concentration of 0.4μmol / L. Reached a more ideal amplification effect. Ninety-four pairs of SSR primers were screened by this system. Among them, 47 pairs of primers amplified polymorphic bands, of which Xtxp3 and Xtxp13 amplified different bands among resistant varieties. Conclusion: The sorghum head smut gene can be analyzed by using optimized SSR reaction system.