Date Written

4-28-2019

Document Type

Distinction Paper

Degree Name

Biology-BS

Department

Biology & Earth Science

First Committee Member

Jennifer Bennett, Ph.D.

Second Committee Member

David C. Sheridan, Ph.D.

Third Committee Member

Meredith Meyer, Ph.D.

Keywords

Streptomyces, Bacterial Development, Microbiology

Subject Categories

Biology | Genetics and Genomics

Abstract

Characterization of two developmental genes in Streptomyces was carried out by different genetic manipulation techniques in two different species. The construction of an overexpression mutant for a gene encoding a putative c-di-GMP binding protein in Streptomyces scabies was started. This gene was found to be a homologue of a protein that was found to bind to c-di-GMP in a protein binding assay that consisted of streptavidin coated beads and biotinylated c-di-GMP. Attempts at cloning the gene encoding this protein into an overexpression construct were made. Another developmental gene was also investigated. Gene disruption by PCR targeting was used to isolate a deletion-insertion mutation of ssdA in Streptomyces coelicolor. which is a spore shape determination gene. This gene was previously identified through random transposon insertion mutagenesis and the mutant was observed to be delayed for sporulation as well as having heterogenous shaped and sized spores. The transposon insertion truncated the gene, but left a large portion intact. A complete deletion mutation was isolated by PCR- directed mutagenesis of the wild-type gene on a cosmid to determine if the complete deletion resulted in a similar phenotype compared to the insertion mutation. The gene was replaced with an antibiotic resistance marker and the mutant S. coelicolor strain was attained with the ssdA deletion. The complete deletion was obtained in the wild type strain and preliminary results reveal that the phenotype is similar to the insertion truncation mutant. In the future, the genotype will be verified by PCR.

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