Date of Award

5-2016

Degree Type

Honors College Thesis

Department

Chemistry and Biochemistry

First Advisor

Sabine Heinhorst, Ph.D.

Advisor Department

Chemistry and Biochemistry

Abstract

The purpose of this research was to develop a deletion construct for the chemoautotrophic bacterium Halothiobacillus neapolitanus, which will be used to generate a mutant lacking a carboxysome shell protein gene. The carboxysome is the location of carbon dioxide fixation. The operon that encodes the carboxysome contains three genes for CsoS1 proteins, the major components of the carboxysome shell. The small CsoS1 proteins self-assemble into hexamers with small central pores. The hexamers arrange into the facets of the icosahedral carboxysome shell. The pores are believed to be involved in selective diffusion of materials necessary for carbon dioxide fixation across the shell.

A deletion construct to replace the csoS1C gene with a kanamycin resistance cassette was designed that will allow gene replacement by homologous recombination to determine if the csoS1C paralog is necessary to form functional carboxysomes. This deletion construct will allow the function of this paralog to be studied in the resulting mutant. To develop the construct, primers were designed to amplify the kanamycin resistance gene with short ends that are homologous to regions flanking the csoS1C gene in the H. neapolitanus genome. E. coli DY330 was transformed with the amplified resistance cassette and a plasmid containing the csoS1C region of genomic DNA for homologous recombination that will yield the deletion construct.

Included in

Biochemistry Commons

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