Zeph L R, Stotzky G
Department of Biology, New York University, New York 10003.
Appl Environ Microbiol. 1989 Mar;55(3):661-5. doi: 10.1128/aem.55.3.661-665.1989.
Presumptive bacteriophage P1 transductants of Escherichia coli, isolated from soil inoculated with lysates of transducing phage P1 and E. coli, were confirmed to be lysogenic for phage P1 by hybridization with a biotinylated DNA probe prepared from the 1.2-kilobase-pair HindIII 3 fragment of bacteriophage P1. No P1 lysogens of indigenous soil bacteria were detected with the DNA probe. The sensitivity and specificity of the DNA probe were assessed with purified and dot blot DNA, respectively. In addition, two techniques for the lysis and deproteinization of bacteria and bacteriophages on nitrocellulose filters were compared. These studies indicated that biotinylated DNA probes may be an effective alternative to conventional radiolabeled DNA probes for detecting specific gene sequences in bacteria indigenous to or introduced into soil.
从接种了转导噬菌体P1和大肠杆菌裂解物的土壤中分离出的推定大肠杆菌噬菌体P1转导子,通过与由噬菌体P1的1.2千碱基对HindIII 3片段制备的生物素化DNA探针杂交,被确认为对噬菌体P1呈溶原性。用该DNA探针未检测到本土土壤细菌的P1溶原菌。分别用纯化的和斑点印迹DNA评估了DNA探针的敏感性和特异性。此外,还比较了两种在硝酸纤维素滤膜上裂解细菌和噬菌体并使其脱蛋白的技术。这些研究表明,生物素化DNA探针可能是传统放射性标记DNA探针的有效替代品,用于检测土壤中本土或引入的细菌中的特定基因序列。