Graduate Thesis Or Dissertation

 

Analysis of segregants of F'gal heterogenotes in Escherichia coli Public Deposited

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  • The purpose of this study was to isolate and characterize any segregants of the heterogenotes formed from sexduction by an F' donor. Following the mating of F' W4520, carrying an F' gal₁⁺ gal₂⁺ factor, with W3350, a gal₁⁻ gal₂⁻ recipient, a number of recombinant clones were picked and examined for segregants. From 5 such clones, 50 segregants of the gal⁺ phenotype and 50 of the gal⁻ phenotype were isolated. The genotypes of these segregants were determined as follows. The gal⁻ segregants were examined to determine the genotype of any F factors carried, by crosses against three gal⁻ recipient types, which were streptomycin resistant mutants isolated early in the study. The failure of these segregants to produce recombinants in these crosses was taken as evidence that no F factors of the gal⁺ type were carried by these segregants. The gal⁻ segregants were also analyzed for their chromosomal genotype by crosses against two gal⁻ F' homogenotes formed earlier in the study. Again, failure of these crosses to produce gal⁺ recombinants was noted and taken as evidence that these gal⁻ segregants carried chromosomal genotypes of the gal₁⁻ gal₂⁻ type. By a similar procedure, the 50 gal⁺ segregants were examined to determine the genotype of any F factors carried, by crosses against the same 3 gal⁻ recipient types, which in this case, were azide resistant mutants isolated early in the study. The formation of gal⁺ recombinants from these crosses was taken as evidence that these 50 gal⁺ segregants carried F' factors of the type F gal₁⁺ gal₂⁺. The segregants were cured of these F factors with acridine orange and their chromosomal genotypes determined by crosses against the same two gal⁻ F¹ homogenotes used earlier. The absence of gal⁺ recombinant formation from these crosses confirmed that these segregants carried chromosomes of the gal₁⁻ gal₂⁻ type. It was found that no variation of genotype existed among the 50 segregants of each type. The 50 gal⁻ strains were concluded to be either haploid parental types, with the genotype gal₁⁻ gal₂⁻ , resulting from loss of the F factor in the sexductional heterogenotes, or possibly homogenotes of the type gal₁⁻ gal₂⁻/ F gal₁⁻ gal₂⁻ , which were undetectable in this system. The 50 gal⁺ segregants, carrying the genotype gal₁⁻ gal₂⁻ / F gal₁⁺ gal₂⁺ , were of the same type as the heterogenotes from which they were segregated. Since no recombinant types were found, it was concluded that the F' factors, in this system, undergo recombination with their host chromosome, during sexduction, at lower frequencies than expected from analogy with transduction by bacteriophage.
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