Degree Type

Dissertation

Date of Award

1993

Degree Name

Doctor of Philosophy

Department

Veterinary Microbiology and Preventive Medicine

First Advisor

Peter A. Pattee

Abstract

Tandem chromosomal duplications offer a means of genetic and physical mapping in microorganisms which lack defined conjugal systems. To determine if tandem chromosomal duplications occur in S. aureus, two libraries of transposon-induced auxotrophic mutations with different resistance phenotypes were needed;To complement the existing library of Tn551-induced auxotrophs, two Tn917lac derivatives were constructed: Tn917G, which contains the gentamicin-resistance gene of Tn4001, and Tn917T, which contains the tetracycline-resistance gene of pT181 and sequences of the Gram-negative cloning vector p34H. Transposon mutagenesis performed with Tn917G and Tn917T demonstrated that these two constructs are useful tools in the genetic analysis of the S. aureus chromosome;Tandem duplications of the S. aureus chromosome were detected in two different regions by transduction of Tn551-induced auxotrophic mutations. The frequency of putative tandem duplications detected in this manner was 0.98% for the tyrA region and 12.54% for the ilv region. The ends of the tandem duplications were estimated by transducing Tn917T-induced auxotrophic mutations into strains containing the chromosomal tandem duplication and by pulsed-field gel electrophoresis (PFGE). The tyrA duplication was only about 25 kb, and only included trp among those markers examined. The larger of the two ilv duplications included lys and thus included about 27% of the 2800-kb S. aureus chromosome. The unique junction fragments and the ends of the tandem duplications were defined by PFGE. All of the tandem duplications were transducible, indicating that the marker used for selection of transductants is close to the unique junction fragment. Segregation to the wild-type parental phenotype occurred most frequently with the ilv tandem duplications, which is attributed to their greater size.

DOI

https://doi.org/10.31274/rtd-180813-9604

Publisher

Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/

Copyright Owner

Laura Lea Hruska

Language

en

Proquest ID

AAI9334990

File Format

application/pdf

File Size

168 pages

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