Use of electroporation to generate a Thiobacillus neapolitanus carboxysome mutant

Abstract

Two cloning vectors designed for use in Escherichia coli and the thiobacilli were constructed by combining a Thiobacillus intermedius plasmid replicon with a multicloning site, lacZ', and either a kanamycin or a streptomycin resistance gene. Conditions necessary for the introduction of DNA into T. intermedius and T. neapolitanus via electroporation were examined and optimized. By using optimal electroporation conditions, the gene encoding a carboxysome shell protein, csoS1A, was insertionally inactivated in T. neapolitanus. The mutant showed a reduced number of carboxysomes and an increased level of CO2 necessary for growth.