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2020是一株分离自中国南方水稻田里的华癸中生根瘤菌(Mesorhizobium huakuii),有3个内源质粒,分别命名为p2020a,p2020b和p2020c.用Tn5-sacB插入突变的方法对2020进行质粒消除,得到了两株质粒缺失突变株2020D29和2020D8.缺失了第一大质粒p2020c的突变株2020D29的结瘤固氮能力有显著的提高;而缺失了第二大质粒p2020b的突变株2020D8失去了在紫云英(Astragalus sinicus)上结瘤的能力;第三大质粒很难被消除,原因可能是该质粒上含有菌株生长所必需的基因.然后将豌豆根瘤菌(Rhizobium leguminosarum)的共生质粒pJB5JI转入2020及其质粒缺失突变株中,盆栽结果显示,2020-137(pJB5JI)的竞争结瘤能力和固氮能力显著高于2020.但是pJB5JI不能恢复2020D8在紫云英上的结瘤能力.2020D8-8(pJB5JI)可以在豌豆(Pisum sativum Linn)上形成无效瘤,这说明pJB5JI的功能可以在2020的遗传背景下进行表达.对pJB5JI在受体菌中的稳定性进行检测,结果发现在人工传代的情况下pJB5JI可以稳定的存在,但经过与植物共生之后只能在部分根瘤分离物中检测到pJB5JI,对这些转移接合子和出发菌株及分离菌株进行Km基因的PCR扩增,除了出发的受体菌外其余的菌株都可以得到PCR产物.由此推断,在没有检测到pJB5JI的分离株中,pJB5JI可能部分或全部整合到了受体根瘤菌的染色体DNA中.
2020 is a Mesorhizobium huakuii isolated from rice fields in southern China and has three endogenous plasmids named p2020a, p2020b and p2020c, respectively. 2020 was carried out by inserting a mutation into Tn5-sacB The plasmids were removed and two plasmid deletion mutants, 2020D29 and 2020D8, were obtained.The nodulation and nitrogen fixation ability of the mutant strain 2020D29, which lacked the first large plasmid p2020c, was significantly increased, while the mutant strain 2020D8, which lacked the second largest plasmid p2020b, lost The ability to nodulate on Astragalus sinicus; the third largest plasmid is difficult to eliminate due to the fact that the plasmid contains the genes necessary for the growth of the strain, and the symbiotic plasmid pJB5JI from Rhizobium leguminosarum The results of pot culture showed that the competitive nodulation ability and nitrogen fixation ability of 2020-137 (pJB5JI) were significantly higher than those of 2020. However, pJB5JI could not restore the nodulation ability of 2020D8 on aspartame.2020D8 -8 (pJB5JI) can form noxious tumors on pea (Pisum sativum Linn), indicating that the function of pJB5JI can be expressed in the genetic background of 2020. The stability of pJB5JI in recipient bacteria The results showed that pJB5JI could be stably existed in the case of artificial passage, but pJB5JI could only be detected in some nodule isolates after being symbiotic with the plant. PCR was performed on these metastatic adapters and the starting strains and isolates for Km gene PCR The PCR product was obtained by amplification of the rest of the strains except for the starting recipient strain, suggesting that pJB5JI may be partially or fully incorporated into the chromosome DNA of the recipient rhizobium in isolates that did not detect pJB5JI.