用Digoxigenin标记重组质粒DNA探针特异性的改良

IMPROVING THE SPECIFICITY OF THE RECOMBINANT PLASMlD DNA PROBE LABELED WITH DIGOXIGENIN

鸡马立克病病毒基因组DNA的BanHI-K_3片断克隆的重组质粒DNA及从质粒上切下的经电泳和电洗脱提纯的BamHI-K_2DNA片断,在用非放射性Digoxigenin([dig])标记后,分别用作检测马立克病病毒DNA的探针。对两种探针特异性的比较表明,BamHI-K_3克隆的全质粒DNA探针对其它质粒来源的NDA仍表现出很强的交叉反应性,而用经电泳电洗脱提纯的BamHI-K_3DNA片断制备的探针,则仅对含同源DNA片断的质粒提取物或菌苔裂解物产生明显呈色反应,但对不含同源DNA的其它质粒DNA提取物几乎完全不呈现交叉反应。

Marck＇s disease virus gcnomic DNA clone BamHI-K_3 samples, which in the forms of the total recombinant plasmid DNA or only the BamHI-K_3 fragment purified by digestion, clectrophoresis and elcctroelution from the total recombinant plasmid, were labeled with nonradioactive digoxigenin respectively and used as probes for dctcction of Marek＇s disease virus DNA. The comparison of two probes for their specificities has indicated that the probe of the total recombinant plasmid DNA gave a strong cross-reaction to other heterogeneous DNA in the another kind of plasmid. When the elcctroclution-purificd BamHI-K_3 was used as probe, it gave specific reactions only to plasmid preparalion of homologous DNA clones or E. coli colonies with homologous DNA clones, it did not show any cross-reaction to other plasmid preparation without homologous DNA fragment.