摘要
以Bio-ll-dUTP替代dTTP经二步法缺口平移标记DNA片段制得生物素DX_(13)探针。其碱性磷酸酶ABC体系显色灵敏度为100fg(约5×10^(-18)mol);与5μg染色体DNA进行Southern印迹杂交,可得清晰的单拷贝杂交区带。以此探针对一血友病家系做RFLP分析,确定两成员为甲型血友病基因携带者。与32P探针对比,两种探针的杂交和分析结果完全一致。表明高标记的生物素探针可应用于单拷贝基因分析和基因诊断。
Biotin-labeled DX13 probe was prepared by the improved nick translation in the presence of Bio-11-dUTP. A very sensitive probe was obtained, which could be visualized as little as 100fg immobilized on nitrocellulose filter in conjunction with streptav idin-enzyme conjugates. The probe was sensitive enough for detection of single copy gene in 5μg of chromosomal DNA. By the use of this probe a hemophilia A family was analysed and 2 carriers in the family were picked out.
