关键词: 烟草, DNA提取, 碱裂解, PCR, 转基因检测

Abstract: To expeditiously extract DNA templates from a large number of transgenic tobacco samples,the alkaline lysis method of extracting DNA from tobacco was improved in this study. Solution of 0.01 mol/L NaOH was used as the extraction buffer and the supernatant could be directly used as a template for PCR reaction,after crushing transgenic tobacco leaf tissues without heating/boiling and neutralizing procedures. The results showed that DNA template produced by NaOH solution of 0.01-0.1 mol/L was amplified successfully. The DNA template produced by the improved method did not rely on particular reaction condition,a variety of types of PCR Mix buffer all amplified successfully. Compared with the traditional SDS extraction method or commercial kits method,PCR results showed no significant differences. In addition,the improved method presented promising results while it was applied to other crops such as wheat,sorghum,rice and maize. The PCR products amplified from the DNA template by the improved method was able to be directly used for sequencing analysis. In conclusion,the improved method is in need of fewer instruments and less sample consumption,rapid,simple,low cost,no cross-contamination,and very efficient to handle a large number of samples,thus,it may rapidly extract and identify the DNA from a large number of transgenic tobacco samples.

Key words: tobacco, DNA extraction, alkaline lysis, PCR, transgenic detection