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通过PCR扩增法制备转基因油菜外源筛选基因FMV35S启动子的DIG-11-dUTP标记DNA探针,与该基因PCR扩增产物进行Southern杂交,通过显色反应对商品化转基因油菜进行检测。本研究对反应基片带正电荷的尼龙膜和硝酸纤维素膜进行了比较,并比较了不同固定条件和杂交条件下的杂交结果。尼龙膜较之硝酸纤维素膜可更好的结合DNA片段,经紫外线照射交联5 分钟,与含甲酰胺的杂交液在45℃杂交30 min结果较为理想。
The DIG-11-dUTP labeled DNA probe of transgenic FMV35S promoter of transgenic rapeseed was prepared by PCR amplification. The amplified product was subjected to Southern hybridization with PCR amplification products, and the commercial transgenic rapeseed was detected by colorimetric reaction. In this study, the positively charged nylon membrane and nitrocellulose membrane of the reaction substrate were compared and the hybridization results under different fixing conditions and hybridization conditions were compared. Compared with nitrocellulose membrane, nylon membrane could bind DNA segment better, cross-linking by ultraviolet radiation for 5 minutes, and hybridization with formamide-containing hybridization solution at 45 ℃ for 30 min.