Mentor: Carole Dabney-Smith, Ph.D.

Cyanobacteria are model organisms to study protein sorting and transport because of the presence of both the thylakoid and plasma membranes. This experiment investigates if the tatC gene is vital to the functioning of the TAT pathway as a whole in Cyanobacteria by using knockout mutations. The tatC gene knockout mutants are made using CRISPR-mediated plasmids ligated via a Gibson Assembly, cloned in E. coli, and transformed into Synechocystis PCC 6803. The cyanobacteria cells are grown in BG-11 media with chloramphenicol over multiple generations. The genomic DNA is extracted and used in conformation PCRs to ensure the correct transformation of the mutant cyanobacteria. The mutant cells with the knockout present as albino, whereas the native cells are the typical green. The PCR results and phenotypic changes in the mutant cells indicate that the TAT pathway is not functional without the TatC gene. Repression studies should be conducted as a next step to identify what levels of TatC gene expression are required for the pathway to function. Additionally, similar methodology should be used to investigate the other TAT components present in Synechocystis PCC 6803.

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