六株成团肠杆菌 (Enterobacteragglomerans)接合子的分子生物学分析

对 6株成团肠杆菌 (Enterobacteragglomerans)接合子的分子生物学进行了分析 .6株菌与nifHDK基因有杂交 .菌株总DNA经BamHⅠ酶切后与pEA9 DNA进行Southern杂交 ,只有 2株菌具有完整的质粒DNA ,其余菌株质粒DNA发生了 15 3～ 137 7kb不同程度的缺失 .用切割位点较少的限制性内切酶XbaⅠ酶切 6株菌的总DNA ,经脉冲场凝胶电泳 (PFGE)后用pEA9 DNA为探针进行Southern杂交 ,每株菌的pEA9 DNA明显大于用BamHⅠ酶切后的杂交结果 ,表明质粒与染色体发生了整合 .转座子Tn5或插入序列IS 12 2 2和IS 12 71可能参与质粒与染色体的整合过程 .

Molecular Biological Analyses of 6 Enterobacter agglomerans Transconjugants

Molecular biological analyses of 6 integration hybrids of Enterobacter agglomerans were carried out.All of them could hybridize with the nif HDK probe. Hybridization of Bam HⅠ digested genomic DNA of integration hybrids with pEA9 DNA showed that only two of them contained the intact pEA9 DNA. In the remaining integration hybrids fragments of pEA9::Tn5 Mob DNA with a size between 15.3 and 137.7 kb were lost during integration. Hybridization with Xba Ⅰ (rare cutter) digested genomic DNA of integration hybrids showed that all integration hybrids contained at least one overlap fragment (an overlap fragment is defined as a fragment with a size that can only be explained by linking pEA9 DNA to chromosomal DNA). These overlap fragments are strong indicators for the integration status of pEA9 DNA. They contain at least one copy of IS 1222, IS 1271 or Tn5, which might eventually be responsible for the integration of pEA9 DNA in chromosomal DNA of Entherobacter agglomerans .