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首页> 外文期刊>Applied and Environmental Microbiology >Detection and Characterization of Plasmid pJP4 Transfer to Indigenous Soil Bacteria
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Detection and Characterization of Plasmid pJP4 Transfer to Indigenous Soil Bacteria

机译：质粒pJP4转移至土生细菌的检测与鉴定

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Prior to gene transfer experiments performed with nonsterile soil, plasmid pJP4 was introduced into a donor microorganism,Escherichia coli ATCC 15224, by plate mating withRalstonia eutropha JMP134. Genes on this plasmid encode mercury resistance and partial 2,4-dichlorophenoxyacetic acid (2,4-D) degradation. The E. coli donor lacks the chromosomal genes necessary for mineralization of 2,4-D, and this fact allows presumptive transconjugants obtained in gene transfer studies to be selected by plating on media containing 2,4-D as the carbon source. Use of this donor counterselection approach enabled detection of plasmid pJP4 transfer to indigenous populations in soils and under conditions where it had previously not been detected. In Madera Canyon soil, the sizes of the populations of presumptive indigenous transconjugants were 107 and 108 transconjugants g of dry soil?1 for samples supplemented with 500 and 1,000 μg of 2,4-D g of dry soil?1, respectively. Enterobacterial repetitive intergenic consensus PCR analysis of transconjugants resulted in diverse molecular fingerprints. Biolog analysis showed that all of the transconjugants were members of the genusBurkholderia or the genus Pseudomonas. No mercury-resistant, 2,4-D-degrading microorganisms containing large plasmids or the tfdB gene were found in 2,4-D-amended uninoculated control microcosms. Thus, all of the 2,4-D-degrading isolates that contained a plasmid whose size was similar to the size of pJP4, contained the tfdB gene, and exhibited mercury resistance were considered transconjugants. In addition, slightly enhanced rates of 2,4-D degradation were observed at distinct times in soil that supported transconjugant populations compared to controls in which no gene transfer was detected.
机译：在用非无菌土壤进行基因转移实验之前，通过与富营养小球藻JMP134的平板交配，将质粒pJP4引入供体微生物大肠杆菌ATCC 15224中。该质粒上的基因编码耐汞性和部分2,4-二氯苯氧基乙酸（2,4-D）降解。大肠杆菌供体缺乏2,4-D矿化所必需的染色体基因，这一事实使得可以通过在含有2,4-D作为碳源的培养基上进行铺板来选择在基因转移研究中获得的推定结合缀合物。这种供体反选择方法的使用使得能够检测质粒pJP4转移到土壤中和在以前未检测到的条件下转移到土著人群中。在马德拉峡谷土壤中，假定补充了500和1,000μg2,4-D g干燥土壤-1的样品，假定的土著超结合物的种群大小分别为107和108 g干燥土壤γ-1。肠缀合物的肠细菌重复基因间共识PCR分析导致了多种分子指纹。 Biolog分析表明，所有转导结合体都是伯克霍尔德氏菌属或假单胞菌属的成员。在经2,4-D修饰的未接种对照微膜中未发现含有大质粒或tfdB基因的抗汞的2,4-D降解微生物。因此，所有包含2,4-D降解分离物的质粒均与pJP4的大小相似，包含tfdB基因，并显示出耐汞性，被认为是转接合剂。此外，与未检测到基因转移的对照相比，在支持转导结合体种群的土壤中，在不同时间观察到2,4-D降解的速率略有提高。

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