Höfte M, Mergeay M, Verstraete W
Laboratory of Microbial Ecology, Faculty of Agricultural Sciences, State University of Ghent, Belgium.
Appl Environ Microbiol. 1990 Apr;56(4):1046-52. doi: 10.1128/aem.56.4.1046-1052.1990.
The mini Mu element Mu dII1681, which contains the lac operon genes and a kanamycin resistance gene, was inserted in the chromosome of plant growth-beneficial Pseudomonas aeruginosa 7NSK2 to construct a marked strain (MPB1). In MPB1, beta-galactosidase is permanently expressed under the culture conditions used. The MPB1 strain could be recovered with an efficiency of about 100% from a sandy loam soil on 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside medium containing sebacic acid and kanamycin. The limit of detection is about 10 CFU/g of soil. A detailed comparison was made between the wild-type strain 7NSK2 and the Mu dII1681-containing MPB1 strain. The results showed that no genes essential for growth, siderophore production, survival in sterile and nonsterile conditions, plant growth stimulation, or root colonization had been damaged in the MPB1 strain, which means that MPB1 can reliably be used for ecological studies in soil. MPB1 survived well at 4 or 28 degrees C but died off relatively rapidly in air-dried soil or at subzero temperatures. In these conditions, however, the MPB1 strain did not completely disappear from the soil but survived at a very low level of about 100 CFU/g of soil for more than 3 months. This observation stresses the need for very sensitive counting methods for ecological studies and for the evaluation of released microorganisms. Maize was inoculated with MPB1 via seed inoculation or soil inoculation. Upon seed inoculation, only the upper root parts were effectively colonized, while soil inoculation resulted in a complete colonization of the root system.
含有乳糖操纵子基因和卡那霉素抗性基因的微小Mu元件Mu dII1681被插入到对植物生长有益的铜绿假单胞菌7NSK2的染色体中，构建了一个标记菌株（MPB1）。在MPB1中，β-半乳糖苷酶在所用培养条件下持续表达。MPB1菌株在含有癸二酸和卡那霉素的5-溴-4-氯-3-吲哚基-β-D-吡喃半乳糖苷培养基上，能以约100%的效率从砂壤土中回收。检测限约为10 CFU/g土壤。对野生型菌株7NSK2和含有Mu dII1681的MPB1菌株进行了详细比较。结果表明，MPB1菌株中对生长、铁载体产生、在无菌和非无菌条件下存活、植物生长刺激或根部定殖至关重要的基因均未受损，这意味着MPB1可可靠地用于土壤生态学研究。MPB1在4℃或28℃下存活良好，但在风干土壤或零下温度下相对迅速死亡。然而，在这些条件下，MPB1菌株并未从土壤中完全消失，而是在约100 CFU/g土壤的极低水平下存活了3个月以上。这一观察结果强调了在生态学研究和评估释放的微生物时，需要非常灵敏的计数方法。通过种子接种或土壤接种将MPB1接种到玉米上。种子接种时，只有上部根部有效地定殖，而土壤接种则导致根系完全定殖。
