By Vageesha Herath, Biochemistry
Advisor: Annette Rowe
Abstract: The genetic manipulation of organism such as E. coli has led to our advanced understanding of this organism's physiology. E. coli is capable of a fairly restricted range of metabolisms compared with other microbes, however genetic approaches in the majority of microbes remain unstudied and unknown. One such microbe, Thioclava electrotropha ElOx9, is an electro-trophic bacterium. The mechanism or genes involved in this process remain unknown. While the overall objective of studying this organism is to understand its capacity for electron uptake, this current work investigates the feasibility of genetically engineering T. electrotropha. Previous work in T. electrotropha has demonstrated that this organism is capable of acquiring a plasmid (small circular DNA found in bacteria that is separate from the central chromosomal DNA) via conjugation. This work investigated the potential for T. electrotropha using mating independent approaches-specifically transformation and electroporation. The plasmid used for this work, pMiniHimar, contains an antibiotic resistance (Kanamycin) that allowed us to screen for acquisition of the plasmid by various methods. Under specific conditions, Thioclava electrotropha bacterial cells can be induced to up-take pMinihimar plasmids, which contain the gene for Kanamycin resistance. This supports the genetic tractability of this organism through conjugation independent methods.