Effects of stress and other environmental factors on horizontal plasmid transfer assessed by direct quantification of discrete transfer events
Effects of stress and other environmental factors on horizontal plasmid transfer assessed by direct quantification of discrete transfer events
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DOI：
10.1111/j.1574-6941.2006.00230.x
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发表时间：
2007-03-01
影响因子：
4.2
通讯作者：
Kroer, Niels
中科院分区：
文献类型：
--
作者：
Johnsen, Anders R.;Kroer, Niels
选择压力可能影响质粒的水平转移。无法区分基因转移和接合子的生长使测试复杂化。我们已经开发出一种方法，使离散的传输事件的量化。它使用微量滴定威尔斯孔中的大量重复配对（192或384）以及转移阳性和转移阴性威尔斯孔的计数。我们应用该方法研究了IncP 1质粒pRO 103在大肠杆菌和恶臭假单胞菌菌株之间的转移。pRO 103编码对汞和四环素的抗性以及对2，4-二氯苯氧乙酸（2，4-D）的部分降解。结果表明，转移与供体代谢活性呈正相关，转移的最佳温度为29 ℃。在刺激供体活性时，最适温度降至24.5 ℃。HgCl 2以上1.0 μ g L-1的负面影响转移，而2，4-D高达0.3 mM没有影响。汞的负面影响被证明是一个结果，强调的收件人。通过传统的过滤器配对，无法检测到汞对转移的影响。因此，该方法上级于过滤器交配，并且由于实验设计允许操作个体参数，因此其对于评估和比较环境因素对质粒转移的影响是理想的。
Selection pressure may affect the horizontal transfer of plasmids. The inability to distinguish between gene transfer and the growth of transconjugants complicates testing. We have developed a method that enables the quantification of discrete transfer events. It uses large numbers of replicate matings (192 or 384) in microtiter wells and the counting of transfer-positive and transfer-negative wells. We applied the method to study the transfer of the IncP1 plasmid pRO103 between Escherichia coli and Pseudomonas putida strains. pRO103 encodes resistance to mercury and tetracycline and partial degradation of 2,4-dichlorophenoxyacetic acid (2,4-D). The results showed positive correlation between transfer and donor metabolic activity, and an optimal temperature for transfer of 29 degrees C. On stimulation of donor activity, the optimal temperature was decreased to 24.5 degrees C. HgCl2 above 1.0 mu g L-1 negatively affected transfer, whereas 2,4-D up to 0.3 mM had no effect. The negative effect of mercury was shown to be a result of stressing of the recipient. No effects of mercury on transfer could be detected by traditional filter mating. Thus, the method is superior to filter mating and, as the experimental design allows the manipulation of individual parameters, it is ideal for the assessment and comparison of effects of environmental factors on plasmid transfer.