Studies on the Vibrio alginolyticus sucrose utilization system cloned into Escherichia coli

Scholle, Renate Regina

Studies on the Vibrio alginolyticus sucrose utilization system cloned into Escherichia coli

Doctoral Thesis

1989

Publisher

University of Cape Town

Department

Department of Molecular and Cell Biology

Faculty

Faculty of Science

Abstract

This dissertation forms part of the study on the molecular biology of the aerobic, collagenolytic, halotolerant, gram-negative organism Vibrio alginolyticus. The cloning, expression and regulation of the v. alginolyticus sucrose utilization system in Escherichia coli is discussed and the results of a molecular analysis of the sucrase gene (scrA) are presented. The clone pVSlOO, containing a sucrose utilization system, was isolated from a genomic library of v. alginolyticus. Plasmid pVSlOO was mapped and the origin of its insert determined by Southern blotting and DNA hybridization. The number and sizes of the polypeptide products encoded by plasmid pVSlOO were determined by DNAdirected cell-free protein synthesis. The capsule, produced by transformed and untransformed E. coli JA221 cells, was shown to be independent of the presence of plasmid pVSlOO. The sucrase activity assay was optimized with respect to time, pH, temperature and salt requirements.