豌豆根瘤菌共生基因pJB5JI与华癸中生根瘤菌7653R共生质粒的相互关系

The Interaction Between the Symbiotic Genes pJB5JI of Rhizobium Leguminosarum bv. Viciae and the Symbiotic Plasmid of Mesorhizobium huakuii 7653R

摘要:

华癸中生根瘤菌(Mesorhizobium huakuii) 7653R是分离自我国南方水稻田的一株根瘤菌, 含有2个内源质粒: p7653Ra和p7653Rb, 其中7653Rb是共生质粒。通过Tn5-sacB的插入方法来消除质粒, 获得7653Rb消除的突变株7653RD。将豌豆根瘤菌T83K3的共生质粒pJB5JI导入7653R和7653RD中, 盆栽结果表明含有pJB5JI的转移接合子7653R-197的竞争结瘤能力和共生固氮能力均高于7653R。pJB5JI不能恢复7653RD在紫云英上的结瘤能力。含有pJB5JI的7653RD可以在豌豆上结无效瘤, 表明pJB5JI可以在7653R的染色体背景下表达其功能。对转移接合子中的质粒稳定性进行检测, 结果表明pJB5JI在人工传代的情况下可以稳定存在, 但经过共生之后发生了遗传分离, 对转移接合子和出发菌株及分离菌株进行kan基因的PCR扩增, 除了受体菌外其他菌株都可得到PCR产物, 由此推测, pJB5JI可能部分或全部整合到了受体菌的染色体基因组中。

Abstract:

Mesorhizobium huakuii strain 7653R, isolated from nodules of A. sinicus L, contains two indigenous plasmids, p7653Ra and p7653Rb, the latter being the symbiotic plasmid. We eliminated the plasmids via Tn5-sacB insertion and obtained its symbiotic plasmid-cured derivative 7653RD. Then, we transferred the symbiotic plasmid pJB5JI of Rhizobium leguminosarum bv. Viciae T83K3 into 7653R and 7653RD. The pot plant test showed an increase in competitive ability and symbiotic nitrogen fixation of transconjugant 7653R-197 (pJB5JI) compared to 7653R. pJB5JI could not restore the ability of 7653RD to nodulate Astragalus sinicus. 7653RD (pJB5JI) could form ineffective nodules on peas, implying that the symbiotic plasmid pJB5JI could express its function at the chromosomal background of Mesorhizobium huakuii 7653R. We checked the stability of plasmid in transconjugants under free-living and during symbiosis. The results indicated pJB5JI could not be detected in some nodule isolates. We amplified kan resistance gene from all transconjugants and nodule isolates which suggested that pJB5JI might fully or partially integrated into the chromosome of recipients.