Analysis of gene transfer and expression mechanisms in aquatic ecosystems using the Aquatron

澄清了可复制的水处理系统的操作条件。在人工培养箱中人工诱导铜绿微囊藻水华。在这些水槽中，通过靶向pT 7 GFP的FISH与荧光抗体偶联的方法，在3至5天内检测到pT 7 GFP从大肠杆菌（pT 7 GFP）向0.1至0.8%的土著细菌的基因转移。结果表明，胞外代谢产物能促进pBHR 1基因从大肠杆菌S17-1（pBHR 1）向施氏假单胞菌的接合转移，并能使含GFP基因质粒的大肠杆菌自溶、噬菌体感染和原虫捕食。对处理后的样品中溶解的DNA进行定量分析，结果表明，来自噬菌体自溶和感染的溶解DNA可能更有助于细菌间的水平基因转移，从世界17个不同地点分离的芽孢杆菌菌株具有相同的mer操纵子和相似的转座子。这一发现表明转座子可能有助于在自然环境中芽孢杆菌菌株中mer操纵子的全球分布和水平传播。将从自然环境中分离的转座子整合到大肠杆菌DH 5 α的不可转化质粒pGEM和可转化质粒pR 388上。将大肠杆菌DH 5 α中的转座子通过接合转移到大肠杆菌S17-1中。这些实验表明，在细胞中存在转座子从pGEM到pR 388的转座。

Operational conditions for replicable aquatrons were clarified. Blooms of cyanobacterium Microcystis aeruginosa was induced artificially in the aquatrons. In these aquatrons, gene transfer of pT7GFP from E.coli (pT7GFP) to 0.1 to 0.8% of indigenous bacteria was detected within 3 to 5 days, by means of FISH targeted to pT7GFP couple with fluorescent antibody method. It was also found that extracellular metabolic products enhanced conjugal gene transfer of pBHR1 from E.coli S17-1(pBHR1) to Pseudomonas stutzeri.E.coli with a plasmid coding GFP gene was exposed to autolysis, infection of phage and predation by protozoa. Quantification of dissolved DNA, as gene source in a natural aquatic environments, in the treated sample, revealed that dissolved DNA from autolysis and infection of phage are likely to more contribute than the other to horizontal gene transfer among bacteria.Bacillus strains isolated from 17 various sites in the world possessed identical mer operon on similar transposons. This finding suggested that transposons may contribute the worldwide distribution and horizontal dissemination of the mer operons among Bacillus strains in natural environments. E.coli DH5α was incorporated with both a transposon, which was isolated from natural environment, on untransformable pGEM and transformable plasmid pR388. The transposon in the E.coli DH5α was transferred to E.coli S17-1 by conjugation. These experiments indicated that there existed transposition of the transposon from pGEM to pR388 in a cell.