title: Cloning and partial characterization of two chromosomal loci from Bacteroides ovatus that contain genes essential for growth on guar gum.
abstract: Previously, we isolated three transposon insertion mutants of Bacteroides ovatus (M-4, M-5, and M-7) that were unable to grow on the branched polysaccharide guar gum. In this study, we used a tetracycline resistance gene on the transposon to clone chromosomal DNA adjacent to the transposon insertions in each of the three mutants. Restriction analysis of the flanking chromosomal DNA in M-4 and M-7 revealed that the insertions in these two mutants were in the same location. The cloned DNA adjacent to the insertions in M-5 and M-7 was used as a hybridization probe to clone the wild-type loci. Two clones of about 10 kbp in size were obtained. Restriction analysis showed that these two clones did not overlap. The clone of the M-5 locus appeared to contain all of the genes affected by the M-5 insertion, but we were unable to demonstrate complementation of the M-5 mutation because of the instability of the clone in this background. Analysis of the clone of the M-7 locus showed that it contained a guar gum-regulated promoter, but the transcript originating from this promoter was not affected by the transposon insertion. Thus, the M-7 locus apparently contains at least two separate transcriptional units, the one defined by this promoter and the one interrupted by the transposon insertion. Insertion mutations downstream of the guar gum-regulated promoter demonstrated that there were essential guar gum utilization genes in this region. The M-7 mutant was eliminated by the wild type in the intestinal tracts of germfree mice.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Microbiology, University of Illinois, Urbana 61801.
authors.forename: P J
authors.initials: PJ
authors.lastname: Valentine
authors.forename: P
authors.initials: P
authors.lastname: Arnold
authors.forename: A A
authors.initials: AA
authors.lastname: Salyers
mesh_terms: D000818:Animals; D001439:Bacteroides / Q000235:genetics* / Q000254:growth & development; D002876:Chromosomes, Bacterial / Q000737:chemistry*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000302:isolation & purification; D015180:Electrophoresis, Gel, Two-Dimensional; D005685:Galactans / Q000378:metabolism*; D005798:Genes, Bacterial / Q000502:physiology*; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D008351:Mannans / Q000378:metabolism*; D051379:Mice; D016254:Mutagenesis, Insertional; D053149:Plant Gums; D014158:Transcription, Genetic
chemical_list: D004269:DNA, Bacterial; D005685:Galactans; D008351:Mannans; D053149:Plant Gums; C007894:guar gum