生物素标记庆大霉素耐药基因探针

采用低熔点琼脂糖挖块法回收源自澳大利亚的pDG0103的2．0kb的BamHI—HindIII片段(携带2”-0-腺苷转移酶ANT(2”)]基因)和自建的pBY102的4．9kb的Pst1-EcoRI片段。以缺口平移法,用生物素-7-dATP进行标记,制备成探针。通过southern印迹杂交和菌落原位杂交,证明澳源的Gm—DNA探针与美国的探针同源,而与作者构建的Gm-DNA探针不同源。再以菌落原位杂交法,用生物素标记的上述两种探针分别检测1 06株庆大霉素(Gm)耐药的细菌,结果表明这些菌株携带的Gm钝化酶基因的类型不止一种。

Two biotin-labeled probes of gentamicin resistance genes

A 2.0 kb BamHI-HindIII fragment of pDG0103 from Australia containing gentamicin 2"-o-adenylytransferase ANT(2")] gene and a 4.9 kb PstI-EcoRI fragment of pBY102 were recovered from low-temperature-melting agarose by the slot method. Both fragments were labeled with biotin-7-dATP by nick translation with a commercial kit. The result of colony and Southern hybridization was that: the 2.0 kb probe from Australia hybridized with that containing ANT(2") from America, while no hybridization occurred between the 2.0 kb probe and the 4.9 kb probe constructed in our lab. Furthermore, the above two fragments were used as probes for detection of 106 strains of gentamicin resistant Enterobacteriaceae. It revealed that there were more than one gentamicin resistance gene in the tested strains.