Cloning of genes involved in myo-inositol transport in a Pseudomonas sp.
A soil isolate of a Pseudomonas sp. can utilize myo-inositol (MI) as the sole carbon source. In this strain, MI is transported through the membrane by a high-affinity transport system in which a periplasmic binding protein is involved. Mutants impaired in the transport system were obtained by mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine and subsequently identified by their slow growth rate at low MI concentrations. Strains with a low linear initial rate of MI uptake were analyzed. Using a broad-host-range cosmid cloning system, we have constructed a gene bank of the wild-type Pseudomonas sp. in an Escherichia coli recA-host. A rapid mating technique enabled us to screen the gene library for clones which are able to restore the active transport of MI in the mutant. An 11.5-kilobase segment containing genes involved in the MI transport has been isolated, and its restriction enzyme cleavage map has been determined.
D001692:Biological Transport; D003001:Cloning, Molecular; D005798:Genes, Bacterial; D007294:Inositol / Q000378:metabolism*; D009154:Mutation; D011549:Pseudomonas / Q000235:genetics* / Q000378:metabolism
D007294:Inositol