Dammann-Kalinowski T, Niemann S, Keller M, Selbitschka W, Tebbe C C, Pühler A
Lehrstuhl für Genetik, Fakultät für Biologie, Universität Bielefeld, Germany.
Appl Microbiol Biotechnol. 1996 May;45(4):509-12. doi: 10.1007/BF00578463.
The deliberate release of genetically engineered microorganisms requires a thorough characterization of the microbes in question. For the two bioluminescent Rhizobium meliloti strains, L1 and L33 [Selbitschka et al. (1992) Mol Ecol 1:9-19; Selbitschka et al. (1995) FEMS Microbiol Ecol 16:223-232], designated for field release, the sites of genetic modifications in the chromosomes were sequenced from amplified genomic DNA. This indicated no unexpected alterations in the nucleotide sequence. The bioluminescent phenotype was stably inherited over more than 100 generations in liquid cultures. The presence of the luciferase gene in both strains did not have secondary effects on a variety of metabolic pathways as assessed by the Biolog GN system. A specific polymerase chain reaction amplification, based on the chromosomal insertion site of the luc cassette, allowed the discrimination between the two strains and thus simplifies monitoring. The RecA-deficient strain L1 showed a strongly (more than 90%) reduced ability to nodulate alfalfa in competition with its parent strain R. meliloti 2011 and its RecA+ counterpart L33.
故意释放基因工程微生物需要对相关微生物进行全面的特性描述。对于指定用于田间释放的两种发光苜蓿根瘤菌菌株L1和L33[Selbitschka等人（1992年）《分子生态学》1:9 - 19；Selbitschka等人（1995年）《FEMS微生物生态学》16:223 - 232]，从扩增的基因组DNA对染色体上的基因修饰位点进行了测序。这表明核苷酸序列没有意外改变。在液体培养中，发光表型在100多代中稳定遗传。通过Biolog GN系统评估，两种菌株中荧光素酶基因的存在对多种代谢途径没有次生影响。基于luc盒染色体插入位点的特异性聚合酶链反应扩增能够区分这两种菌株，从而简化了监测。与亲本菌株苜蓿根瘤菌2011及其RecA +对应菌株L33竞争时，RecA缺陷菌株L1结瘤苜蓿的能力大幅降低（超过90%）。
