Genes Essential for Nod Factor Production and Nodulation Are Located on a Symbiotic Amplicon (AMPRtrCFN299pc60) in Rhizobium tropici
Genes Essential for Nod Factor Production and Nodulation Are Located on a Symbiotic Amplicon (AMPRtrCFN299pc60) in Rhizobium tropici
复制标题
结瘤因子产生和结瘤所必需的基因位于热带根瘤菌的共生扩增子 (AMPRtrCFN299pc60) 上
作者：
P. Mavingui;T. Laeremans;M. Flores;D. Romero;E. Martínez;R. Palacios
摘要在etli根瘤菌基因组中发现了可扩增的DNA区域（扩增子）。本文报道了热带根瘤菌共生扩增子的分离和分子特征。为了寻找共生扩增子，将含有卡那霉素抗性标记的盒插入R.热带CFN 299。通过增加细胞培养物中卡那霉素的浓度来选择具有扩增的衍生物。将扩增的DNA区域移动到大肠杆菌中，然后移动到根癌农杆菌中。60-kb的共生扩增子，我们称之为AMPRtrCFN 299 pc60，包含几个固氮和固氮基因，两侧是一个新的插入序列ISRtr 1。通过ISRtr 1重复序列之间的同源重组扩增AMPRtrCFN 299 pc60增加了结瘤因子的量。引人注目的是，扩增子接合转移到无质粒A中。tumefaciens菌株赋予接合子产生R. tropici Nod因子和菜豆（Phaseolusvulgaris）结瘤的能力，表明R.对转化过程必需的营养基因被限制在pSym的可重复DNA区域。
ABSTRACT Amplifiable DNA regions (amplicons) have been identified in the genome of Rhizobium etli. Here we report the isolation and molecular characterization of a symbiotic amplicon of Rhizobium tropici. To search for symbiotic amplicons, a cartridge containing a kanamycin resistance marker that responds to gene dosage and conditional origins of replication and transfer was inserted in the nodulation region of the symbiotic plasmid (pSym) of R. tropici CFN299. Derivatives harboring amplifications were selected by increasing the concentration of kanamycin in the cell culture. The amplified DNA region was mobilized into Escherichia coli and then into Agrobacterium tumefaciens. The 60-kb symbiotic amplicon, which we termed AMPRtrCFN299pc60, contains several nodulation and nitrogen fixation genes and is flanked by a novel insertion sequence ISRtr1. Amplification of AMPRtrCFN299pc60 through homologous recombination between ISRtr1 repeats increased the amount of Nod factors. Strikingly, the conjugal transfer of the amplicon into a plasmidlessA. tumefaciens strain confers on the transconjugant the ability to produce R. tropici Nod factors and to nodulatePhaseolus vulgaris, indicating that R. tropicigenes essential for the nodulation process are confined to an ampliable DNA region of the pSym.