以地高辛标记的Southern杂交技术已广泛应用于多个物种中。选择橡胶树为材料，就基因组DNA的提取、探针标记方法的选择以及具体的杂交操作过程等方面对该体系进行了优化。结果表明，至少40μg高质量的DNA在300μL的大酶切体系中，酶切12 h可获得良好效果；PCR法标记的探针杂交条带清晰、背景浅，其效率明显强于随机引物法标记的探针。本研究优化的体系信号强、背景浅和灵敏度高，为橡胶树Southern杂交鉴定分析提供参考。%Southern blot technology based on the DIG-chemiluminescent detection has been widely used in numerous species. Using Hevea brasiliensis as materials, the DIG-labeled Southern blot analysis was improved through optimizing several key steps, including extraction of genome DNA, usage of DNA samples amount, digestion system, the comparison between PCR and random-primed labeled method and a serial of procedure in the process of the hybrid. The results showed that desirable digestion result could be achieved when at least 40μg DNA samples with high quality were enzymed in 300μL reaction system for 12 hours. The efficiency of probe labeled with the method of PCR was higher than that of probe labeled with random primer obviously, while the Southern blot result of PCR labeled was clear, and the background was lightly, which was conducive to read the copy number accurately. Finally, Southern blot with DIG-labeled was optimized, and good result with high sensitivity and low background was achieved.
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