应用水稻叶片直接PCR扩增的方法

The Method of Direct-PCR Amplification in Rice Leaves(Oryza Sativa L)

本研究以新鲜和冷冻的水稻幼苗叶片为材料,探索水稻叶片直接作为模板进行PCR扩增的方法。结果表明,与对照SDS法提取的DNA相比,应用新鲜或冷冻的1~2 mm2水稻叶片直接作为模板在缓冲液为10 mmol/L KCl,2.5 mmol/L Mg Cl2,25 mmol/L(NH4)2SO4,40 mmol/L Tris-HCl(p H 9.0),0.8%PCR增强剂的反应体系中扩增,PCR扩增效果无明显差异;研究结果表明,该方法省去DNA体外提取过程,节约成本,操作简便,可以直接利用水稻叶片作为DNA模板进行PCR扩增。

In this research fresh and frozen rice seedling leaves were used as experimental materials, the method of direct-PCR amplification in rice leaves was developed and analyzed. The results showed that compared to PCR amplification with template DNA extracted by SDS method, there was no significant difference in the amplification effect by directly using fresh or frozen 1~2 mm2 rice leaf as template in mixed DNA extracting and PCR amplifying buffer of 10 mmol/L KCl, 2.5 mmol/L Mg Cl2, 25mmol/L(NH4)2SO4, 40 mmol/L Tris-HCl(p H 9.0)and 0.8% PCR enhancer. The results indicated that the rice leaf could be directly used as template for PCR-amplification, which eliminates the process of DNA extraction, saves the cost, and operates easy.