The function of a lysogenic rhizobiophage in the symbiosis between Rhizobium and leguminous plants.

温和噬菌体PHIU通过噬菌体（attP）和细菌（attB）位点之间的位点特异性重组使三叶草根瘤菌4S溶原化。在噬菌体DNA的6.0 kb EcoRI片段上鉴定attP位点，并将其克隆到自杀质粒pSUP 202上的EcoRI位点中。重组质粒pCI 6能以4.40 × 10 ~（-6）的频率整合到4S菌株的染色体上<-3>。pCI 6也能整合到对噬菌体PHIU不敏感的豌豆根瘤菌K5的染色体上。以attP片段为探针的Southern杂交结果表明，pCI 6在染色体上的整合位点在菌株4S和K5之间完全相同。以pSUP 203为载体，构建了含有噬菌体PHIU attP和三叶苜蓿根瘤菌（R.leguminosarum bv.trifolii）基因的融合质粒pCINodl。将携带pCINod 1的大肠杆菌S17-1与菌株4S的共生缺陷突变株H1接合，接种于白色三叶草（Trifolium repens cv.Ladino）的根部形成根瘤。用透射电镜观察结节。根瘤菌通过侵染丝侵入寄主植物细胞，并转化为类杆菌。从根瘤中分离出接合子，其中一株命名为CNH 1。用attP探针进行Southern杂交，结果表明，pCINod 1与pCI 6一样整合到CNH 1菌株的染色体上。这些结果表明，整合到染色体上的反义基因在根瘤菌中有效地发挥功能。

The temperate phage PHIU lysogenizes Rhizobium leguminosarum bv.trifolii 4S by site specific recombination between phage (attP) and bacterial (attB) sites. The attP site was identified on a 6.0 kb EcoRI fragment of the phage DNA and cloned into the EcoRI site on the suicide plasmid pSUP202. Plasmid pCI6, the resulting recombinant plasmid, could integrate into the chromosome of strain 4S at high frequency of 4.40 X 10^<-3>. The pCI6 could also integrate into the chromosome of R.leguminosarum bv.viciae K5 which was not sensitive to phage PHIU infection. Southern hybridization probed with attP fragment revealed that the integration site of pCI6 on the chromosome was completely identical between strains 4S and K5. It is suggested that pCI6 may be a useful chromosome integrative vector for gene transfer on Rhizobium.The fusion plasmid carrying the attP of phage PHIU and the nod-genes derived from R.leguminosarum bv.trifolii was constructed based on pSUP203 and designated as pCINodl. E.coli S17-1 harboring pCINod1 was conjugated with strain H1, a symbiotic defective mutant of strain 4S.The inoculation of the mating mixture resulted in nodule formation on the roots of white clover (Trifolium repens cv.Ladino). The nodules were observed by transmission electron microscopy. Rhizobium invaded into the host plant cells through infection threads and changed their form to bacteroids. Transconjugants were isolated from the nodules and one of them was designated as strain CNH1. Southern hybridization probed with attP revealed that pCINod1 integrated into the chromosome of strain CNH1 as same as pCI6. These results indicate that nod-genes integrated into the chromosome functioned efficiently in Rhizobium.