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re-examination of the structures of the lipopolysaccharide core oligosaccharides from rhizobium leguminosarum biovar phaseoli. 19911811857
the region for exopolysaccharide synthesis in rhizobium leguminosarum biovar trifolii is located on the non-symbiotic plasmid.an exo- mutant of rhizobium leguminosarum biovar trifolii was isolated which did not produce acidic exopolysaccharide and induced defective, non-fixing nodules on clover plants. the nodules were defective at a late stage of development, they contained infection threads and bacteria were released into the host cells. cosmid parf136 capable of complementing the exo- mutation was isolated from a cosmid bank made from total r. trifolii dna. hybridization between dna of parf136 and plasmids of r. tri ...19911814135
sequence and structural organization of a nif a-like gene and part of a nifb-like gene of herbaspirillum seropedicae strain z78.the deduced amino acid sequence derived from the sequence of a fragment of dna from the free-living diazotroph herbaspirillum seropedicae was aligned to the homologous protein sequences encoded by the nifa genes from azorhizobium caulinodans, rhizobium leguminosarum, rhizobium meliloti and klebsiella pneumoniae. high similarity was found in the central domain and in the c-terminal region. the h. seropedicae putative nifa sequence was also found to contain an interdomain linker similar to that co ...19911840608
identification of nolr, a negative transacting factor controlling the nod regulon in rhizobium meliloti.in rhizobium meliloti, expression of the nodulation genes (nod and nol genes) is under both positive and negative controls. these genes are activated by the products of the three related nodd genes, in conjunction with signal molecules from the host plants. we showed that negative regulation is mediated by a repressor protein, binding to the overlapping nodd1 and noda as well as to the nodd2 promoters. the encoding gene, termed nolr, was identified and cloned from strain 41. by subcloning, delet ...19911840615
comparative response of pisum sativum nodulated with indigenous soil rhizobium populations and/or co-inoculated with a rhizobium leguminosarum strain. i. acetylene-reducing, dihydrogen- and carbon dioxide-evolving activities.no significant differences in the acetylene-reducing activity and evolution of h2 and co2 from nodulated roots of pisum sativum inoculated with soil rhizobium populations from two soils with different acidities (ruzynĕ soil 7.6; lukavec soil 4.9) were observed. rhizobium population from lukavec soil formed nodules, exhibiting a higher h2 evolution. co-inoculation with the hup+ strain 128c30 (7 x 10(7) cells per seedling) eliminated, to some extent, the effect of soil populations on physiological ...19911841862
inoculation of vicia sativa subsp. nigra roots with rhizobium leguminosarum biovar viciae results in release of nod gene activating flavanones and chalcones.flavonoids released by roots of vicia sativa subsp. nigra (v. sativa) activate nodulation genes of the homologous bacterium rhizobium leguminosarum biovar viciae (r. l. viciae). inoculation of v. sativa roots with infective r. l. viciae bacteria largely increases the nod gene-inducing ability of v. sativa root exudate (a.a.n. van brussel et al., j bact 172: 5394-5401). the present study showed that, in contrast to sterile roots and roots inoculated with r. l. viciae cured of its sym plasmid, roo ...19911859867
nodt, a positively-acting cultivar specificity determinant controlling nodulation of trifolium subterraneum by rhizobium leguminosarum biovar trifolii.rhizobium leguminosarum biovar trifolii strain ta1 nodulates a range of trifolium plants including red, white and subterranean clovers. nitrogen-fixing nodules are promptly initiated on the tap roots of these plants at the site of inoculation. in contrast to these associations, strain ta1 has a 'nod-' phenotype on a particular cultivar of subterranean clover called woogenellup (a.h. gibson, aust j agric sci 19: (1968) 907-918) where it induces rare, poorly developed, slow-to-appear and ineffecti ...19911868196
rhizobium lipopolysaccharide modulates infection thread development in white clover root hairs.the interaction between rhizobium lipopolysaccharide (lps) and white clover roots was examined. the limulus lysate assay indicated that rhizobium leguminosarum bv. trifolii (hereafter called r. trifolii) released lps into the external root environment of slide cultures. immunofluorescence and immunoelectron microscopy showed that purified lps from r. trifolii 0403 bound rapidly to root hair tips and infiltrated across the root hair wall. infection thread formation in root hairs was promoted by p ...19911885517
regulation of phenolic catabolism in rhizobium leguminosarum biovar trifolii.in members of the family rhizobiaceae, many phenolic compounds are degraded by the protocatechuate branch of the beta-ketoadipate pathway. in this paper we describe a novel pattern of induction of protocatechuate (pca) genes in rhizobium leguminosarum biovar trifolii. isolation of pca mutant strains revealed that 4-hydroxybenzoate, quinate, and 4-coumarate are degraded via the protocatechuate pathway. at least three inducers govern catabolism of 4-hydroxybenzoate to succinyl coenzyme a and acety ...19911885531
characterization of reca genes and reca mutants of rhizobium meliloti and rhizobium leguminosarum biovar viciae.dna fragments carrying the reca genes of rhizobium meliloti and rhizobium leguminosarum biovar viciae were isolated by complementing a uv-sensitive reca- escherichia coli strain. sequence analysis revealed that the coding region of the r. meliloti reca gene consists of 1044 bp coding for 348 amino acids whereas the coding region of the r. leguminosarum bv. viciae reca gene has 1053 bp specifying 351 amino acids. the r. meliloti and r. leguminosarum bv. viciae reca genes show 84.8% homology at th ...19911896024
six nodulation genes of nod box locus 4 in rhizobium meliloti are involved in nodulation signal production: nodm codes for d-glucosamine synthetase.the nucleotide sequence of the nod box locus n4 in rhizobium meliloti was determined and revealed six genes organized in a single transcriptional unit, which are induced in response to a plant signal such as luteolin. mutations in these genes influence the early steps of nodule development on medicago, but have no detectable effect on melilotus, another host for r. meliloti. based on sequence homology, the first open reading frame (orf) corresponds to the nodm gene and the last to the nodn gene ...19911909418
a novel highly unsaturated fatty acid moiety of lipo-oligosaccharide signals determines host specificity of rhizobium.in rhizobium leguminosarum biovar viciae, the nodabc and nodfel operons are involved in the production of lipo-oligosaccharide signals which mediate host specificity. the structure of these metabolites and those produced in nod mutants links the node and nodl genes to specific chemical features of the signal molecules. a node-determined, highly unsaturated fatty acid and a nodl-determined o-acetyl substituent are essential for the ability of the signals to induce nodule meristems on the host pla ...19911944592
polypeptide composition of bacterial cyclic diguanylic acid-dependent cellulose synthase and the occurrence of immunologically crossreacting proteins in higher plants.to comprehend the catalytic and regulatory mechanism of the cyclic diguanylic acid (c-di-gmp)-dependent cellulose synthase of acetobacter xylinum and its relatedness to similar enzymes in other organisms, the structure of this enzyme was analyzed at the polypeptide level. the enzyme, purified 350-fold by enzyme-product entrapment, contains three major peptides (90, 67, and 54 kda), which, based on direct photoaffinity and immunochemical labeling and amino acid sequence analysis, are constituents ...19911647035
polygalacturonase is a virulence factor in agrobacterium tumefaciens biovar 3.agrobacterium tumefaciens biovar 3 causes both crown gall and root decay of grapes. all biovar 3 strains, regardless of their tumorigenicity, produce in culture a single polygalacturonase with a pi around 4.5. a. tumefaciens biovar 3 strain cg49 was mutagenized with tn5 by using psup2021 as a suicide vector. a mutant strain, cg50, lacking polygalacturonase activity was isolated. the mutation was due to a single tn5 insertion in an 8.5-kb ecori fragment that also contained the polygalacturonase s ...19911655716
a cultivar-specific interaction between rhizobium leguminosarum bv. trifolii and subterranean clover is controlled by nodm, other bacterial cultivar specificity genes, and a single recessive host gene.insertion mutagenesis identified two negatively acting gene loci which restrict the ability of rhizobium leguminosarum bv. trifolii ta1 to infect the homologous host trifolium subterraneum cv. woogenellup. one locus was confirmed by dna sequence analysis as the nodm gene, while the other locus, designated csn-1 (cultivar-specific nodulation), is not located on the symbiosis plasmid. the presence of these cultivar specificity loci could be suppressed by the introduction of the nodt gene from anu8 ...19911673458
novel organization of the common nodulation genes in rhizobium leguminosarum bv. phaseoli strains.nodulation by rhizobium, bradyrhizobium, and azorhizobium species in the roots of legumes and nonlegumes requires the proper expression of plant genes and of both common and specific bacterial nodulation genes. the common nodabc genes form an operon or are physically mapped together in all species studied thus far. rhizobium leguminosarum bv. phaseoli strains are classified in two groups. the type i group has reiterated nifhdk genes and a narrow host range of nodulation. the type ii group has a ...19911991718
high-frequency rearrangements in rhizobium leguminosarum bv. phaseoli plasmids.high-frequency genomic rearrangements affecting the plasmids of rhizobium leguminosarum bv. phaseoli cfn42 were analyzed. this strain contains six large plasmids ranging in size from 200 to 600 kb. in the absence of any selective pressure, we found 11 strains from 320 analyzed colonies that presented different kinds of plasmid-borne rearrangements, including sequence amplification, deletion, cointegration, and loss of plasmids. these data support the concept that the r. leguminosarum bv. phaseol ...19911991727
occurrence of lipid a variants with 27-hydroxyoctacosanoic acid in lipopolysaccharides from members of the family rhizobiaceae.lipopolysaccharides (lpss) isolated from several strains of rhizobium, bradyrhizobium, agrobacterium, and azorhizobium were screened for the presence of 27-hydroxyoctacosanoic acid. the lpss from all strains, with the exception of azorhizobium caulinodans, contained various amounts of this long-chain hydroxy fatty acid in the lipid a fractions. analysis of the lipid a sugars revealed three types of backbones: those containing glucosamine (as found in rhizobium meliloti and rhizobium fredii), tho ...19912007543
construction of a mobilizable cloning vector for site-directed mutagenesis of gram-negative bacteria: application to rhizobium leguminosarum.a mobilizable cloning vector was constructed from defined fragments to serve as a suicide plasmid for site-directed mutagenesis. the new vector, pkok4, closely resembles plasmid pbr325. however, the inverted duplication existing in the latter was not introduced. the useful cloning sites of pbr325 (ecori, hindiii, ecorv, bamhi, sali, psti and pvui) were retained and are located in one of the three resistance markers, apr, cmr or tcr, respectively. also, in pkok4 the cmr gene retains its own promo ...19912013412
structural complexity of the symbiotic plasmid of rhizobium leguminosarum bv. phaseoli.the complete physical map of the symbiotic plasmid of rhizobium leguminosarum bv. phaseoli strain cfn42 was established. the data support the concept that rhizobium symbiotic genes are part of a complex genomic structure which contains a large amount of reiterated dna sequences. this plasmid is a circular structure of 390 kb with approximately 10 families of internally reiterated dna sequences of two to three elements each. one family includes two directly oriented nitrogenase operons situated 1 ...19912013564
partial deletion of the rhizobium phaseoli cfn23 symbiotic plasmid implies a concomitant amplification of plasmid dna sequences.rhizobium leguminosarum biovar phaseoli cfn23 loses its ability to nodulate beans at a high frequency because of a deletion of part of its symbiotic (psym) plasmid (soberón-chávez et al., 1986). we report here that at least 80 kb of psym are deleted upon loss of the symbiotic phenotype; the deletion removes the nitrogenase structural nifhdk and the common nodabc genes. the size of the deleted psym is not reduced, since it is accompanied by an amplification of other psym plasmid sequences. this g ...19912014006
genetic and physical analysis of the nodd3 region of rhizobium meliloti.the nodulation (nod) genes of the symbiont rhizobium meliloti are transcriptionally controlled by protein activators in the nodd gene family. while nodd1 and nodd2 act in concert with small molecular weight inducers provided by the host legume plant, nodd3 is an inducer-independent activator of the nod promoters. we determined the sequence of the nodd3 gene, confirmed the expression of a 35 kda protein in vitro, and determined the insertion points of five tn5 insertions in the region of the nodd ...19912017373
isolation of the rhizobium leguminosarum nodf nodulation protein: nodf carries a 4'-phosphopantetheine prosthetic group.rhizobium species produce a protein product of the nodf gene that has a limited but recognizable homology to the well-characterized acyl carrier protein (acp) of escherichia coli. nodf functions together with node in generating a host-specific response to the plant host in the interchange of signals leading to the effective nodulation of roots (h.p. spaink, j. weinman, m.a. djordjevic, c.a. wijffelman, r.j.h. okker, and b. j.j. lugtenberg, embo j. 8:2811-2818, 1989; b. scheres, c. van de wiel, a ...19912019559
the degree of esterification and points of substitution by o-acetyl and o-(3-hydroxybutanoyl) groups in the acidic extracellular polysaccharides secreted by rhizobium leguminosarum biovars viciae, trifolii, and phaseoli are not related to host range.rhizobium leguminosarum biovars viciae, trifolii, and phaseoli have been grown in the presence and absence of 4',5,7-trihydroxyflavonone (naringenin) or 4',5,7-trihydroxyflavone (apigenin), which induce the expression of nodulation genes of the bacteria. the acidic polysaccharides secreted by the rhizobium were isolated from the culture media and purified. the polysaccharides were cleaved with a bacteriophage enzyme and the octasaccharide repeating units formed were isolated. the glycosyl sequen ...19912033051
distribution of o-acetyl groups in the exopolysaccharide synthesized by rhizobium leguminosarum strains is not determined by the sym plasmid.the patterns of o-acetylation of the exopolysaccharide (eps) from the sym plasmid-cured derivatives of rhizobium leguminosarum bv. trifolii strain lpr5, r. leguminosarum bv. trifolii strain anu843 and r. leguminosarum bv. viciae strain 248 were determined by 1h and 13c nmr spectroscopy. beside a site indicative of the chromosomal background, these strains have one site of o-acetylation in common, namely residue b of the repeating unit. the o-acetyl esterification pattern of eps of the sym plasmi ...19912033052
genetic structure of a soil population of nonsymbiotic rhizobium leguminosarum.the genetic structure of a population of nonsymbiotic rhizobium leguminosarum strains was determined by the electrophoretic mobilities of eight metabolic enzymes. nonsymbiotic strains were isolated from the rhizosphere of bean plants and characterized by growth on differential media and at different temperatures, intrinsic antibiotic resistance, the lack of homology to a nifh probe, and their inability to form nodules on bean roots. all the isolates clustered with r. leguminosarum bv. phaseoli r ...19911707606
antigenic changes in lipopolysaccharide i of rhizobium leguminosarum bv. viciae in root nodules of vicia sativa subsp. nigra occur during release from infection threads.three different monoclonal antibodies raised against the o antigen-containing lipopolysaccharide (lps i) of free-living cells were used in an immunocytochemical study to follow the fate of lps i on the outer membrane of rhizobium leguminosarum bv. viciae 248 during the nodulation of vicia sativa subsp. nigra. after immunogold labeling, the lps i epitopes were detected on the outer membrane of bacteria present in infection threads throughout the nodule. epitopes were not detectable on bacteria re ...19911708764
suppression of nodulation gene expression in bacteroids of rhizobium leguminosarum biovar viciae.the expression of nod genes of rhizobium leguminosarum bv. viciae in nodules of pisum sativum was investigated at both the translational and transcriptional levels. by using immunoblots, it was found that the levels of noda, nodi, node, and nodo proteins were reduced at least 14-fold in bacteriods compared with cultured cells, whereas nodd protein was reduced only 3-fold. northern (rna) blot hybridization, rnase protection assays, and in situ rna hybridization together showed that, except for th ...19911712355
rhizobium tropici, a novel species nodulating phaseolus vulgaris l. beans and leucaena sp. trees.a new rhizobium species that nodulates phaseolus vulgaris l. and leucaena spp. is proposed on the basis of the results of multilocus enzyme electrophoresis, dna-dna hybridization, an analysis of ribosomal dna organization, a sequence analysis of 16s rdna, and an analysis of phenotypic characteristics. this taxon, rhizobium tropici sp. nov., was previously named rhizobium leguminosarum biovar phaseoli (type ii strains) and was recognized by its host range (which includes leucaena spp.) and nif ge ...19911715738
the oac gene encoding a lipopolysaccharide o-antigen acetylase maps adjacent to the integrase-encoding gene on the genome of shigella flexneri bacteriophage sf6.lysogens of shigella flexneri harbouring the temperate bacteriophage, sf6, have been previously shown to undergo a serotype conversion due to o-acetylation of the o-antigen of the lipopolysaccharide. a partial physical map of the phage genome has been constructed. analysis of the phage dna suggests that the phage packages by a headful mechanism and that the mature dna molecules are terminally redundant. cloning of the psti fragments of sf6 enabled the region encoding the serotype conversion to b ...19911720755
biological activity of rhizobial siderophore.non-nodulating mutant of rhizobium leguminosarum biovar trifolli produces the phenolate type of siderophore consisting of 2,3-dihydroxybenzoic acid and threonine. the activity of this compound against the various bacteria was tested. only, the growth of r. leguminosarum strains was stimulated by siderophore. the other species of rhizobium, especially r. meliloti, were sensitive to this agent. the growth of r. meliloti was also inhibited by agrobactin and pseudobactin. this effect was reversed by ...19911726625
transposon tn5 as an identifiable marker in rhizobia: survival and genetic stability of tn5 mutant bean rhizobia under temperature stressed conditions in desert soils.five transposon tn5 insertion mutants of a beanrhizobium strain (rhizobium leguminosarum b. v.phaseoli) were used in an ecological study to evaluate the extent to which transposon tn5 was stable to serve as an identifiable marker in rhizobia under a high temperature stress condition in two sonoran desert soils. all the mutants possessed single chromosomal insertions of the transposon. in both soils, under the temperature stress conditions that were employed (40°c), both wild type and mutant popu ...199124194199
differential accumulation of hydroxyproline-rich glycoproteins in bean root nodule cells infected with a wild-type strain or a c4-dicarboxylic acid mutant of rhizobium leguminosarum bv. phaseoli.an antiserum raised against deglycosylated hydroxyproline-rich glycoproteins (hpgps) from melon (cucumis melo l.) was used to study the relationship between rhizobium infection and induction of hrgps in bean (phaseolus vulgaris l.) root nodule cells infected with either the wild-type or a c4-dicarboxylic acid mutant strain of rhizobium leguminosarum bv. phaseoli. in effective nodules, where fixation of atmospheric dinitrogen is taking place, hrgps were found to accumulate mainly in the walls of ...199124194237
assessment of faba bean (vicia faba) response to inoculation with rhizobium leguminosarum in clay loam nile delta soil.a field experiment was conducted to assess the response to inoculation with rhizobia in a clay loam soil of the nile delta using faba bean (vicia faba) for two successive winter seasons (1985/6 and 1986/7). three selected strains of rhizobium leguminosarum, tal 634, nrc 65 and tal 1400, were used singly or in combination as peat-based inocula in 1985/6 winter season. strain tal 1400 was replaced by strain f9 in the 1986/7 winter season. a significant seed yield response was obtained only with st ...199124424931
distribution of glucose/mannose-specific isolectins in pea (pisum sativum l.) seedlings.we report on the distribution and initial characterization of glucose/mannose-specific isolectins of 4- and 7-d-old pea (pisum sativum l.) seedlings grown with or without nitrate supply. particular attention was payed to root lectin, which probably functions as a determinant of host-plant specificity during the infection of pea roots by rhizobium leguminosarum bv. viciae. a pair of seedling cotyledons yielded 545±49 μg of affinity-purified lectin, approx. 25% more lectin than did dry seeds. shoo ...199024196924
the occurrence of unusual laminated structures rich in β-1,4-glucans in plastids of phaseolus vulgaris root-nodule cells infected by an ineffective c4-dicarboxylic-acid mutant of rhizobium leguminosarum bv. phaseoli.root nodules induced in phaseolus vulgaris l. by the wild-type (wt) and a c4-dicarboxylic-acid mutant strain of rhizobium leguminosarum biovar. phaseoli were compared on the basis of ultrastructure and cytochemistry of cellulose subunits. the mutant bacteroids failed to colonize infected host cells in a normal manner, and presented a premature degenerative appearance. starch granules, rough endoplasmic reticulum and mitochondria were found to accumulate in the ineffective nodules. the most strik ...199024202009
transcriptional analysis of the glnb-glna region of rhizobium leguminosarum biovar viciae.we report that the glnb and glna genes of rhizobium leguminosarum biovar viciae are preceded by promoters located upstream of each gene. we find the presence of a glnb-glna and a glna mrna whose intracellular concentration changes two- to three-fold when r. leguminosarum is grown on different nitrogen sources. primer extension analysis shows unique transcriptional initiation sites upstream of glnb and glna. the glnb promoter is rpon(ntra)-dependent, while the glna promoter does not contain a typ ...19902077357
identification and cloning of nodulation genes and host specificity determinants of the broad host-range rhizobium leguminosarum biovar phaseoli strain ciat899.rhizobium leguminosarum biovar phaseoli type ii strain ciat899 nodulates a wide range of hosts: phaseolus vulgaris (beans), leucaena esculenta (leucaena) and macroptilium atropurpureum (siratro). a nodulation region from the symbiotic plasmid has been isolated and characterized. this region, which is contained in the overlapping cosmid clones pcv38 and pcv117, is able to induce nodules in beans, leucaena and siratro roots when introduced in strains cured for the symbiotic plasmid, psym. in addit ...19902082147
nucleotide sequence of the hydrogenase structural genes from rhizobium leguminosarum. 19902103457
a new family of rsf1010-derived expression and lac-fusion broad-host-range vectors for gram-negative bacteria.a series of broad-host-range expression and lac fusion vectors, based on rsf1010 derivatives, was constructed. the expression vectors contain various promoters (pnm, plac, ptac and ps1) for expression of foreign genes. the efficiency of the promoters was determined in escherichia coli, rhizobium meliloti, rhizobium leguminosarum and pseudomonas putida by beta-galactosidase activity measurements. of the promoters assayed in e. coli, the most effective is the tac promoter, whereas in soil bacteria ...19902115488
excessive excretion of cyclic beta-(1,2)-glucan by rhizobium trifolii ta-1.at 25 degrees c, the optimal temperature for growth of rhizobium trifolii ta-1, extracellular and capsular polysaccharide (eps and cps) were the main carbohydrate products synthesized in mannitol-rich medium (10 g of mannitol and 1 g of glutamic acid per liter). in the same medium at 33 degrees c, eps and cps production was inhibited, and up to 3.9 g of cyclic beta-(1,2)-glucan was produced during an incubation period of 20 days with a total biomass of 0.55 g of protein. in a medium containing 5 ...19902117876
regulatory functions of the three nodd genes of rhizobium leguminosarum biovar phaseoli.the three nodd genes of a strain of rhizobium leguminosarum biovar phaseoli were cloned to study their effects on transcription of themselves and of the nodc genes of biovars phaseoli and viciae. efficient transcription of nodd1 required nodd1 and was enhanced by exposure of the cells to bean exudate consistent with the presence of a nod-box preceding the noie-nodd1 operon. transcription of nodd2 and nodd3 was constitutive. nodc of r. leguminosarum biovar phaseoli was activated by each of the no ...19902120543
identification of nods and nodu, two inducible genes inserted between the bradyrhizobium japonicum nodyabc and nodij genes.the so-called common nodulation (nod) gene cluster of bradyrhizobium japonicum is characterized by a unique composition of genes that are arranged in the following order: nody, noda, nodb, nodc, nods, nodu, nodi, nodj. as reported here, the identification of the two new genes nods and nodu resulted from the dna sequencing of a 4.5-kilobase nodc-downstream region covering nods, nodu, nodi, and nodj. the predicted nods, nodu, nodi, and nodj proteins had the following respective amino acid (aa) len ...19902134855
two genes that regulate exopolysaccharide production in rhizobium sp. strain ngr234: dna sequences and resultant phenotypes.two closely linked genes involved in the regulation of exopolysaccharide (eps) production in rhizobium sp. strain ngr234, exox and exoy, were sequenced, and their corresponding phenotypes were investigated. inhibition of eps synthesis occurred in wild-type strains when extra copies of exox were introduced, but only when exoy had been deleted or mutated or was present at a lower copy number. normal eps synthesis occurred in rhizobium sp. when both exox and exoy were introduced on the same replico ...19902152899
two plasmids other than the nodulation plasmid are necessary for formation of nitrogen-fixing nodules by rhizobium leguminosarum.a system which allows direct selection for curing of plasmids in gram-negative bacteria was used to generate derivatives of rhizobium leguminosarum vf39 cured of each of six plasmids present in this strain. phenotypes could be correlated with the absence of five of the six plasmids. the smallest plasmid, prlevf39a, carries genes for the production of a melanin-like pigment as has been previously reported. plasmid prlevf39d carries nodulation and nitrogen fixation genes. curing of the plasmids pr ...19902161988
differential expression of hydrogen uptake (hup) genes in vegetative and symbiotic cells of rhizobium leguminosarum.the genetic determinants responsible for h2-uptake (hup genes) in rhizobium leguminosarum are organized in six transcriptional units, designated regions hupi to hupvi, with region hupi coding for the hydrogenase structural genes (leyva et al. 1990). regulation of the expression of hup genes from r. leguminosarum was examined by using hup-lacz fusions and mrna dot-blot analysis. none of the six hup regions is transcribed in vegetative cells grown under normal aerobic conditions, whereas all six r ...19902166228
an fnr-like protein encoded in rhizobium leguminosarum biovar viciae shows structural and functional homology to rhizobium meliloti fixk.a 1.9 kb dna region of rhizobium leguminosarum biovar viciae strain vf39 capable of promoting microaerobic and symbiotic induction of the rhizobium meliloti fixn gene was identified by heterologous complementation. sequence analysis of this dna region revealed the presence of two complete open reading frames, orf240 and orf114. the deduced amino acid sequence of orf240 showed significant homology to escherichia coli fnr and r. meliloti fixk. the major difference between orf240 and fixk is the pr ...19902175385
the nifh promoter region of rhizobium leguminosarum: nucleotide sequence and promoter elements controlling activation by nifa protein.the nucleotide (nt) sequence of the rhizobium leguminosarum nifh promoter region contains a consensus promoter, a consensus upstream activator sequence (uas), a pseudo (psi) promoter and a psi uas. we mapped the transcription start point for the consensus promoter sequence by primer extension. this promoter differs from the consensus in one of the four supposedly invariant nt and can be activated by the klebsiella pneumoniae nifa product in escherichia coli. under these conditions the psi promot ...19902185138
rhizobium meliloti and rhizobium leguminosarum dctd gene products bind to tandem sites in an activation sequence located upstream of sigma 54-dependent dcta promoters.free-living rhizobia transport external c4-dicarboxylates to use as sole carbon sources, and uptake of these compounds is essential for nitrogen fixation by rhizobial bacteroids. in both rhizobium leguminosarum and rhizobium meliloti, the genes dctb and dctd are believed to form an ntrb/ntrc-like two-component system which regulates the synthesis of a c4-dicarboxylate transport protein encoded by dcta. here we confirm the identity of sigma 54-dependent promoters previously hypothesized for the r ...19902193923
subcellular localization of the rhizobium leguminosarum nodi gene product.by the use of antibodies raised against a fusion protein of lacz'-nodi (produced in escherichia coli) which specifically react with nodi protein, it was shown that in wild-type rhizobium leguminosarum biovar viciae nodi protein (i) is recovered with the cytoplasmic membrane fraction and (ii) is translated as part of the nodabcij operon. in addition, it was found that the bacterial chromosomal background strongly influences the expression of several nod genes.19902203755
analysis of three nodd genes in rhizobium leguminosarum biovar phaseoli; nodd1 is preceded by noie, a gene whose product is secreted from the cytoplasm.in a strain of rhizobium leguminosarum biovar phaseoli, three copies of the regulatory nodulation gene nodd were identified on the sym plasmid and sequenced. two were closely linked to each other and the third was near, but not adjacent, to the nodabc genes. each of these nodd genes could correct the nod- defect of a nodd mutant strain of r. leguminosarum biovar viciae on peas. a truncated form of nodd2 could also correct this mutant, indicating that the c-terminus of nodd2 is not needed for ind ...19902215216
either of two nod gene loci can complement the nodulation defect of a nod deletion mutant of rhizobium leguminosarum bv viciae.a deletion mutant of rhizobium leguminosarum biovar viciae lacking the host-specific nodulation (nod) gene region (nodfel nodmnt and nodo) but retaining the other nod genes (nodd nodabcij) was unable to nodulate peas or vicia hirsuta, although it did induce root hair deformation. the mutant appeared to be blocked in its ability to induce infection threads and could be rescued for nodulation of v. hirsuta in mixed inoculation experiments with an exopolysaccharide deficient mutant (which is also n ...19902233683
rhizobium leguminosarum exob mutants are deficient in the synthesis of udp-glucose 4'-epimerase.rhizobium leguminosarum bv. viciae exo- mutant strains rbl5523,exo7::tn5,rbl5523,exo8::tn5 and rbl5523,exo52::tn5 are affected in nodulation and in the syntheses of lipopolysaccharide, capsular polysaccharide, and exocellular polysaccharide. these mutants were complemented for nodulation and for the syntheses of these polysaccharides by plasmid pmp2603. the gene in which these mutants are defective is functionally homologous to the exob gene of rhizobium meliloti. the repeating unit of the resid ...19902250016
rleai: a novel class-iis restriction endonuclease from rhizobium leguminosarum recognizing 5'-cccaca(n)12-3' 3'-gggtgt(n)9-5'. 19902253885
[construction of frankia genomic libraries and isolation of clones homologous to nodulation genes from rhizobium leguminosarum].high molecular genomic dnas were isolated by using the lysozyme plus achromopeptidase system from frankia strains at4, ccol and hr16, the root nodule endophytes of alnus, casuarina and hippophae respectively, and used to construct genomic libraries in plafr1, a broad host range cosmid vector within many gram-negative hosts. the genomic libraries were screened by in situ colony hybridization to identify clones homologous to common nodulation genes of rhizobium leguminosarum, based on the sequence ...19902268450
structure of the extracellular polysaccharide secreted by rhizobium leguminosarum var. phaseoli ciat 899.the structure of the extracellular polysaccharide secreted by rhizobium leguminosarum var. phaseoli ciat 899 has been studied by methylation analysis. 1h-n.m.r. spectroscopy, and partial acid hydrolysis. the repeating unit is an octasaccharide made up of d-glucose, d-galactose, pyruvic acid, and acetic acid in the molar ratios 6:2:1.5:1.5. half of the terminal gal groups are 4,6-substituted by pyruvic acid acetal groups and the other half by o-acetyl groups at position 3. also, one of the 3-link ...19902279241
the enod12 gene product is involved in the infection process during the pea-rhizobium interaction.the pea cdna clone ppsenod12 represents a gene involved in the infection process during pisum sativum l.-rhizobium leguminosarum bv. viciae symbiosis. the enod12 protein is composed of pentapeptides containing two hydroxyprolines. the expression of the enod12 gene is induced in cells through which the infection thread is migrating, but also in cells that do not yet contain an infection thread. soluble compounds from rhizobium are involved in eliciting enod12 gene expression. rhizobium common and ...19902297789
expression of rhizobium leguminosarum cfn42 genes for lipopolysaccharide in strains derived from different r. leguminosarum soil isolates.two mutant derivatives of rhizobium leguminosarum anu843 defective in lipopolysaccharide (lps) were isolated. the lps of both mutants lacked o antigen and some sugar residues of the lps core oligosaccharides. genetic regions previously cloned from another rhizobium leguminosarum wild-type isolate, strain cfn42, were used to complement these mutants. one mutant was complemented to give lps that was apparently identical to the lps of strain anu843 in antigenicity, electrophoretic mobility, and sug ...19902298697
the rhizobium nodulation gene nodo encodes a ca2(+)-binding protein that is exported without n-terminal cleavage and is homologous to haemolysin and related proteins.nodulation and host-specific recognition of legumes such as peas and vicia spp. are encoded by the nodulation (nod) genes of rhizobium leguminosarum biovar viciae. one of these genes, nodo, has been shown to encode an exported protein that contains a multiple tandem repeat of a nine amino acid domain. this domain was found to be homologous to repeated sequences in a group of bacterial exported proteins that includes haemolysin, cyclolysin, leukotoxin and two proteases. these proteins are secrete ...19902303029
immunochemical analysis of lipopolysaccharides from free-living and endosymbiotic forms of rhizobium leguminosarum.rhizobium leguminosarum b556 and 8002 differ only with respect to carrying symbiotic plasmids with specificity for pisum or phaseolus hosts, respectively. protease-treated samples derived from free-living cultures of both strains revealed a ladder of lipopolysaccharide (lps-1) bands after periodate-silver staining of sodium dodecyl sulfate-polyacrylamide gels. these bands were arranged as doublets. after western (immuno-) blotting, all lps-1 bands reacted with monoclonal antibody jim 21, whereas ...19902318803
molecular characterization of the nodulation gene, nodt, from two biovars of rhizobium leguminosarum.dna sequencing of the nodij region from rhizobium leguminosarum biovar trifolii revealed the nodt gene immediately downstream of nodj. dna hybridizations using a nodt-specific probe showed that nodt is present in several r. leguminosarum strains. interestingly, a flavonoid-inducible nodt gene homologue in r. leguminosarum bv. viciae is not in the nodabcij operon but is located downstream of nodmn. the sequence of the nodt gene from bv. viciae was determined and a comparison of the predicted amin ...19902338917
a biovar-specific signal of rhizobium leguminosarum bv. viciae induces increased nodulation gene-inducing activity in root exudate of vicia sativa subsp. nigra.flavonoids in root exudate of leguminous plants activate the transcription of rhizobium genes involved in the formation of root nodules (nod genes). we report that inoculation with the homologous symbiont r. leguminosarum bv. viciae results in an increased nod gene-inducing activity (ini) in root exudate of v. sativa subsp. nigra, whereas inoculation with heterologous rhizobium strains results in exudates with nod gene-inducing activity comparable to that of uninfected plants. ini can be demonst ...19902394688
dna sequence encoding the two structural genes for the uptake hydrogenase of rhizobium leguminosarum bv. viciae b10. 19902402452
a rhizobium leguminosarum mutant defective in symbiotic iron acquisition.iron acquisition by symbiotic rhizobium spp. is essential for nitrogen fixation in the legume root nodule symbiosis. rhizobium leguminosarum 116, an ineffective mutant strain with a defect in iron acquisition, was isolated after nitrosoguanidine mutagenesis of the effective strain 1062. the pop-1 mutation in strain 116 imparted to it a complex phenotype, characteristic of iron deficiency: the accumulation of porphyrins (precursors of hemes) so that colonies emitted a characteristic pinkish-red f ...19902404949
determination of viability within serotypes of a soil population of rhizobium leguminosarum bv. trifolii.concern has been raised about the percentage of viable cells within soil rhizobia populations measured by the immunofluorescence direct count method. the purpose of this study was to evaluate a direct viable count technique which is based on the fact that viable bacteria in natural populations undergo cell elongation when they are exposed to a combination of substrate and the inhibitor of dna gyrase, nalidixic acid. a soil extraction procedure was developed to recover a high proportion of soil b ...19902407187
genetic organization of the hydrogen uptake (hup) cluster from rhizobium leguminosarum.in symbiosis with peas, rhizobium leguminosarum upm791 induces the synthesis of a hydrogen uptake (hup) system that recycles hydrogen generated in nodules by nitrogenase. a cosmid (pal618) containing hup genes from this strain on a 20-kilobase-pair (kb) dna insert has previously been isolated in our laboratory (a. leyva, j. m. palacios, t. mozo, and t.ruiz-argüeso, j. bacteriol. 169:4929-4934, 1987). here we show that cosmid pal618 contains all of the genetic information required to confer high ...19902407728
production and epitope analysis of monoclonal antibodies against a rhizobium leguminosarum biovar trifolii strain.heat-treated cells of rhizobium leguminosarum biovar trifolii strain 162x95 were used to produce monoclonal antibodies (mabs). the fusion produced three cross-reactive mabs and eight mabs specific for the immunizing strain and a group of five other r. trifolii strains from the same geographic region where 162x95 was isolated (california). seven mabs were analyzed by competitive enzyme-linked immunosorbent assay to determine the number of different epitopes detectable on strain 162x95. the result ...19901697150
a previously unrecognized glutamine synthetase expressed in klebsiella pneumoniae from the glnt locus of rhizobium leguminosarum.using glnt dna of rhizobium meliloti as a hybridization probe we identified a r. leguminosarum biovar phaseoli (r. l. phaseoli) locus (glnt) expressing a glutamine synthetase activity in klebsiella pneumoniae. a 2.2 kb dna fragment from r. l. phaseoli was cloned to give plasmid pmw5a, which shows interspecific complementation of a k. pneumoniae glna mutant. the cloned sequence did not show cross-hybridization to glna or glnii, the genes coding for two glutamine synthetase isozymes of rhizobium s ...19901980142
construction of a symbiotically effective strain of rhizobium leguminosarum bv. trifolii with increased nodulation competitiveness.genes involved in nodulation competitiveness (tfx) were inserted by marker exchange into the genome of the effective strain rhizobium leguminosarum bv. trifolii ta1. isogenic strains of ta1 were constructed which differed only in their ability to produce trifolitoxin, an antirhizobial peptide. trifolitoxin production by the ineffective strain r. leguminosarum bv. trifolii t24 limited nodulation of clover roots by trifolitoxin-sensitive strains of r. leguminosarum bv. trifolii. the trifolitoxin-p ...199016348109
role of microniches in protecting introduced rhizobium leguminosarum biovar trifolii against competition and predation in soil.the importance of microniches for the survival of introduced rhizobium leguminosarum biovar trifolii cells was studied in sterilized and recolonized sterilized loamy sand and silt loam. the recolonized soils contained several species of soil microorganisms but were free of protozoa. part of these soil samples was inoculated with the flagellate bodo saltans, precultured on rhizobial cells. the introduced organisms were enumerated in different soil fractions by washing the soil, using a standardiz ...199016348125
isolation of rhizobium leguminosarum (biovar trifolii) strains from ethiopian soils and symbiotic effectiveness on african annual clover species.strains of rhizobium leguminosarum (biovar trifolii) isolated from two ethiopian soils or obtained from a commercial source were evaluated for symbiotic effectiveness on five african annual clover species. numerous rhizobium trifolii strains that exhibited varying levels of symbiotic effectiveness were isolated from both soils (a nitosol and a vertisol), and it was possible to identify strains that were highly effective for each clover species. the soil isolates were, as a group, superior to the ...199016348157
purification and properties of catechol 1,2-dioxygenase from rhizobium leguminosarum biovar viceae usda 2370.the catechol 1,2-dioxygenase of rhizobium leguminosarum biovar viceae usda 2370 was purified 296-fold, yielding a homogeneous preparation with a specific activity of 51.1 u mg of protein. the molecular weight of the native protein was 70,000, with two identical subunits of 34,500 and 1 g-atom of iron per mol. the optimum ph for catalytic activity was 9.0 to 9.5.199016348234
new field isolates of rhizobium leguminosarum biovar viciae that nodulate the primitive pea cultivar afghanistan in addition to modern cultivars.a collection of 13 field isolates of rhizobium leguminosarum bv. viciae that have the ability to nodulate the roots of current north american cultivars of peas as well as a "primitive" cultivar, afghanistan, was examined. these isolates originated in diverse geographical regions of the world, which indicates that this phenotype is not restricted to isolates from any one region. when subclones of the nodulation region from one plasmid were used to examine ecori-fragment-length polymorphisms in th ...199016348240
increased bean (phaseolus vulgaris l.) nodulation competitiveness of genetically modified rhizobium strains.rhizobium leguminosarum bv. phaseoli strain collections harbor heterogeneous groups of bacteria in which two main types of strains may be distinguished, differing both in the symbiotic plasmid and in the chromosome. we have analyzed under laboratory conditions the competitive abilities of the different types of rhizobium strains capable of nodulating phaseolus vulgaris l. bean. r. leguminosarum bv. phaseoli type i strains (characterized by nif gene reiterations and a narrow host range) are more ...199016348252
development and partial characterization of nearly isogenic pea lines (pisum sativum l.) that alter uptake hydrogenase activity in symbiotic rhizobium.some rhizobium bacteria have h(2)-uptake (hup) systems that oxidize h(2) evolved from nitrogenase in leguminous root nodules. pea (pisum sativum l.) cultivars ;ji1205' and ;alaska' produce high hup (hup(++)) and moderate hup (hup(+)) phenotypes, respectively, in rhizobium leguminosarum 128c53. the physiological significance and biochemical basis of this host plant genetic effect are unknown. the purpose of this investigation was to advance basic hup studies by developing nearly isogenic lines of ...199016667415
products of dark co(2) fixation in pea root nodules support bacteroid metabolism.products of the nodule cytosol in vivo dark [(14)c]co(2) fixation were detected in the plant cytosol as well as in the bacteroids of pea (pisum sativum l. cv "bodil") nodules. the distribution of the metabolites of the dark co(2) fixation products was compared in effective (fix(+)) nodules infected by a wild-type rhizobium leguminosarum (mnf 300), and ineffective (fix(-)) nodules of the r. leguminosarum mutant mnf 3080. the latter has a defect in the dicarboxylic acid transport system of the bac ...199016667422
sym 13-a gene conditioning ineffective nodulation in pisum sativum.treatment of pisum sativum (l.) cv. ;sparkle' with ethyl methanesulfonic acid (ems) produced a stable mutant, e135f, which forms small, white, ineffective nodules. these nodules exhibit histological zonation typical of an indeterminant nodule, e.g. meristematic, early symbiotic, late symbiotic, and senescent zones. compared with the nitrogen fixing nodules of the parent, the zones are smaller and the nodules senesce prematurely. bacteroids in e135f are less elongated and less differentiated than ...199016667870
diversity within serogroups of rhizobium leguminosarum biovar viceae in the palouse region of eastern washington as indicated by plasmid profiles, intrinsic antibiotic resistance, and topography.serology, plasmid profiles, and intrinsic antibiotic resistance (iar) were determined for 192 isolates of rhizobium leguminosarum biovar viceae from nodules of peas (pisum sativum l.) grown on the south slope and bottomland topographic positions in eastern washington state. a total of 3 serogroups and 18 plasmid profile groups were identified. nearly all isolates within each plasmid profile group were specific for one of the three serogroups. cluster analysis of iar data showed that individual c ...198916347814
use of the chrome azurol s agar plate technique to differentiate strains and field isolates of rhizobium leguminosarum biovar trifolii.identification of rhizobium and bradyrhizobium strains and especially of indigenous isolates continues to be one of the major difficulties associated with competition studies. because there is no universally accepted method, the method of choice depends on preference, experience, and equipment. here, an agar plate technique was used to distinguish strains and field isolates of rhizobium leguminosarum biovar trifolii to provide a basis for identifying nodule occupants in further competition studi ...198916347877
population size and distribution of rhizobium leguminosarum bv. trifolii in relation to total soil bacteria and soil depth.bacterial cells small enough to pass through 0.4-mum-pore-size filters made up 5 to 9% of the indigenous bacterial population in 0- to 20-cm-depth samples of abiqua silty clay loam. within the same soil samples, cells of a similar dimension were stained with fluorescent antibodies specific to each of four antigenically distinct indigenous serogroups of rhizobium leguminosarum bv. trifolii and made up 22 to 34% of the soil population of the four serogroups. despite the extensive contribution of s ...198916347896
expression by soil bacteria of nodulation genes from rhizobium leguminosarum biovar trifolii.gram-negative, rod-shaped bacteria from the soil of white clover-ryegrass pastures were screened for their ability to nodulate white clover (trifolium repens) cultivar grasslands huia and for dna homology with genomic dna from rhizobium leguminosarum biovar trifolii icmp2668 (nzp582). of these strains, 3.2% were able to hybridize with strain icmp2668 and nodulate white clover and approximately 19% hybridized but were unable to nodulate. strains which nodulated but did not hybridize with strain i ...198916347936
competition among strains of rhizobium leguminosarum biovar trifolii and use of a diallel analysis in assessing competition.competition between indigenous rhizobium leguminosarum biovar trifolii strains and inoculant strains or between mixtures of inoculant strains was assessed in field and growth-room studies. strain effectiveness under competition was compared with strain performance in the absence of competition. field inoculation trials were conducted at elora, ontario, canada, with soil containing indigenous r. leguminosarum biovar trifolii. the indirect fluorescent-antibody technique was used for the identifica ...198916347952
localization of bacteria and hemoglobin in root nodules of parasponia andersonii containing both bradyrhizobium strains and rhizobium leguminosarum biovar trifolii.dual occupancy of parasponia andersonii nodules with different bradyrhizobium strains and rhizobium leguminosarum biovar trifolii was frequently obtained when two strains were inoculated into plants grown aseptically in tubes. since reisolates of bradyrhizobium strains from dually occupied nodules acquired the ability to nodulate trifolium repens, the spatial relationship of the two species of bacteria during nodule initiation and development was investigated and their proximity was demonstrated ...198916347995
developmentally regulated expression of the gene family for cytosolic glutamine synthetase in pisum sativum.in pisum sativum, two classes of genes encode distinct isoforms of cytosolic glutamine synthetase (gs). the first class comprises two nearly identical or "twin" gs genes (gs341 and gs132), while the second comprises a single gs gene (gs299) distinct in both coding and noncoding regions from the "twin" gs genes. gene-specific analyses were used to monitor the individual contribution of each gene for cytosolic gs during root nodule development and in cotyledons during germination, two contexts whe ...198916667090
evidence for divalent cation (ca2+)-stabilized oligomeric proteins and covalently bound protein-peptidoglycan complexes in the outer membrane of rhizobium leguminosarum.two unusual characteristics of some outer membrane proteins of rhizobium leguminosarum are described. first, most of the major outer membrane proteins could only be visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis after lysozyme treatment of the isolated cell envelopes, suggesting a very strong, possibly covalent, interaction of these proteins with the peptidoglycan. these peptidoglycan-associated outer membrane proteins belonged to two distinct groups of immunologically r ...19892500420
construction of a lacz-kanamycin-resistance cassette, useful for site-directed mutagenesis and as a promoter probe.a lacz gene without a promoter, but containing its ribosome-binding site, was cloned next to the kanamycin-resistance (kmr) gene of plasmid puc4k, yielding a lacz-kmr cassette. from the resulting plasmid, pkok5, the lacz-kmr cassette was recloned by means of bamhi into plasmid pkok4, a mobilizable derivative of pbr322 which mediates ampicillin, chloramphenicol and tetracycline resistance. the lacz-kmr cassette can be excised from pkok5 or pkok6 by digestion with bamhi, sali or psti. it can be us ...19892515118
analysis of the major inducers of the rhizobium noda promoter from vicia sativa root exudate and their activity with different nodd genes.root exudate of vicia sativa contains 7 inducers for the noda promoter of rhizobium leguminosarum biovar viciae. six of these inducers are flavanones. one inducer was identified as 3,5,7,3'-tetrahydroxy-4'-methoxyflavanone, and a second inducer most likely is 7,3'-dihydroxy-4'-methoxyflavanone. the inducing activity of these compounds and the other inducers depends on the nodd gene present in the test strains, which originated either from r. leguminosarum biovars viciae or trifolii, or from r. m ...19892519112
extension of host range of rhizobium leguminosarum bv. trifolii caused by point mutations in nodd that result in alterations in regulatory function and recognition of inducer molecules.the positive activation of several nodulation genes in strain anu843 of rhizobium leguminosarum biovar trifolii is mediated by the product of the nodd gene and by the interaction of nodd with plant-secreted inducer and anti-inducer compounds. we have mutagenized the nodd gene of strain anu843 with nitrosoguanidine and have found that the ability of the mutated nodd products to interact with inducer and anti-inducer compounds is affected by the amino acid sequence in at least two key regions, inc ...19892520822
isolation and characterization of mutants of rhizobium leguminosarum bv. viciae 248 with altered lipopolysaccharides: possible role of surface charge or hydrophobicity in bacterial release from the infection thread.effects of alterations in lipopolysaccharide (lps) structure of rhizobium leguminosarum bv. viciae on effective symbiosis and on a number of cell surface characteristics were studied. tn5 mutants with altered lpss were screened for their inability to bind monoclonal antibody 3, one of three monoclonal antibodies to the tentative o-antigenic part of the wild-type lps of strain 248. ten class i lps mutants completely lacked the o-antigen-containing lps species. the class ii lps mutant had a severe ...19892536673
symbiotic properties of rhizobia containing a flavonoid-independent hybrid nodd product.a hybrid nodd gene consisting of 75% of the nodd1 gene of rhizobium meliloti at the 5' end and 27% of the nodd gene of rhizobium trifolii at the 3' end activates the six tested inducible nod promoters of rhizobium leguminosarum, r. trifolii, or r. meliloti to maximal levels, even in the absence of flavonoids. in strains containing such a constitutive activating nodd gene, transcription of nod genes started at the same site as in flavonoid-induced strains containing a wild-type nodd gene. in cont ...19892544568
the rhizobium meliloti host range nodq gene encodes a protein which shares homology with translation elongation and initiation factors.the rhizobium meliloti nod region iib is involved in host-range determination: (i) the presence of region iib is necessary for transfer of alfalfa root hair curling ability to rhizobium leguminosarum biovar trifolii; (ii) a mutation in region iib extends the r. meliloti infection host range to vicia sativa nigra; (iii) dominance of r. meliloti nod genes over r. leguminosarum biovar viciae nod genes is abolished by mutations in region iib. the nucleotide sequence of this region has been determine ...19892546009
omegon-km: a transposable element designed for in vivo insertional mutagenesis and cloning of genes in gram-negative bacteria.to combine the features of the omega interposons with the advantages of in vivo transposition mutagenesis, we have constructed an artificial transposon, called omegon-km. the omegon-km transposon is carried on the plasmid pjff350 which can be conjugally mobilized into a broad range of gram-negative bacteria. omegon-km is flanked, in inverted orientation, by synthetic 28-bp repeats derived from the ends of is1. in addition, each end of omegon-km has the very efficient transcription and translatio ...19892546859
direct selection for curing and deletion of rhizobium plasmids using transposons carrying the bacillus subtilis sacb gene.we have constructed derivatives of the transposon tn5 carrying the mob site (orit) of plasmid rp4, and an npti-sacb-sacr cassette [ried and collmer, gene 57 (1987) 239-246]. the mob site, in conjunction with the antibiotic-resistance markers carried on the transposons, allows identification of transposon inserts in cryptic plasmids by mobilisation to other strains. the sacb-sacr genes allow direct selection for the loss or curing of plasmids, because only strains which no longer contain an activ ...19892548927
rhizobium leguminosarum exopolysaccharide mutants: biochemical and genetic analyses and symbiotic behavior on three hosts.ten independently generated mutants of rhizobium leguminosarum biovar phaseoli cfn42 isolated after tn5 mutagenesis formed nonmucoid colonies on all agar media tested and lacked detectable production of the normal acidic exopolysaccharide in liquid culture. the mutants were classified into three groups. three mutants harbored tn5 insertions on a 3.6-kilobase-pair ecori fragment and were complemented to have normal exopolysaccharide production by cosmids that shared an ecori fragment of this size ...19892549002
comparative study of the symbiotic plasmid dna in free living bacteria and bacteroids of rhizobium leguminosarum.the symbiotic plasmid (psym) dna present in bacteroids of strain rcr1001 of rhizobium leguminosarum biovar viceae has been compared qualitatively and quantitatively with that present in free living bacteria by hybridization experiments with appropriate probes. a decrease in the relative amount of psym dna was observed in bacteroids as compared to bacteria. no rearrangements of the symbiotically expressed psym borne genes were detected in bacteroids.19892551770
the nifa gene product from rhizobium leguminosarum biovar trifolii lacks the n-terminal domain found in other nifa proteins.the nifa gene has been identified between the fixx and nifb genes in the clover microsymbiont rhizobium leguminosarum biovar trifolii (r.i. bv. trifolii) strain anu843. expression of the nifa gene is induced in the symbiotic state and site-directed mutagenesis experiments indicate that nifa expression is essential for symbiotic nitrogen fixation. interestingly, the predicted r.i. bv. trifolii nifa protein lacks an n-terminal domain that is present in the homologous proteins from r.i. bv. viciae, ...19892552256
genes involved in lipopolysaccharide production and symbiosis are clustered on the chromosome of rhizobium leguminosarum biovar viciae vf39.four mutants of rhizobium leguminosarum biovar viciae vf39 altered in lipopolysaccharide (lps) synthesis were isolated upon random tn5 mutagenesis. these mutants produced matt colonies on ty medium and showed autoagglutination and loss of motility. on sodium dodecyl sulfate-polyacrylamide gels, they lacked a slow-migrating carbohydrate band, corresponding to the complete lps (lpsi). all four mutants formed small white nodules on vicia hirsuta. these nodules were infected but showed no nitrogen-f ...19892553672
characterization and nucleotide sequence of a novel gene fixw upstream of the fixabc operon in rhizobium leguminosarum.on the rhizobium leguminosarum pre sym plasmid, fixabc and a novel gene fixw were identified upstream of the regulatory gene nifa. the molecular masses of fixabc, 29, 44 and 50 kda respectively, were estimated by polyacrylamide gel electrophoresis (page) and of fixw, 25 kda, by page and nucleotide sequencing. hybridization studies using bacteroid mrna as a probe showed that fixabc is one operon which can be transcribed independently of fixw. nucleotide sequencing revealed that both fixw and fixa ...19892555670
transposable elements for efficient manipulation of a wide range of gram-negative bacteria: promoter probes and vectors for foreign genes.we describe here the construction and use of a series of modified transposons based on the insertion sequence is1. like their parent, omegon-km [fellay et al., gene 76 (1989) 215-226], these elements permit efficient insertional mutagenesis of a variety of gram-negative bacteria. the presence of a functional pbr322 origin of replication within the transposable element facilitates subsequent cloning of the mutated gene. the omegon-km system was previously shown to function in pseudomonas putida, ...19892559879
transposon mutagenesis and complementation of the fructokinase gene in rhizobium leguminosarum biovar trifolii.transposon tn5 was used to generate a fructokinase mutation in rhizobium leguminosarum biovar trifolii bal. the section of the genome containing tn5 was cloned into the ecori site of the vector phc79 and isolated by direct selection on medium containing kanamycin and tetracycline. total ecori digestion was used to obtain a single fragment containing tn5 and flanking dna sequences. the flanking dna was used as a probe to isolate an intact fructokinase gene from a plafr1 cosmid clone bank of the p ...19892561289
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