Creating the fbiA-fbiB- Double Null Mutant in Dictyostelium discoideum

Avolio, Theodore

Creating the fbiA-fbiB- Double Null Mutant in Dictyostelium discoideum

Persistent URL

http://hdl.handle.net/10456/42882

Author(s)

Avolio, Theodore

Date Issued

April 3, 2017

Abstract

Dictyostelium discoideum is a soil-dwelling social slime mold that serves as a remarkably useful model organism for studying signal transduction pathways, cell-type proportioning, and the ubiquitin-proteasome system (UPS). In the UPS, proteins are marked for ubiquitin-mediated degradation; therefore, it is a powerful regulatory pathway. FbxA is a protein that has been identified as a part of an SCF ubiquitin-ligase complex. FbxA likely targets another protein known as FbiA for ubiquitin-mediated proteolysis. In fbiA- null mutants, the developing organisms exhibit small prespore cell populations and increased prestalk populations. Thus, FbiA likely promotes the prespore pathway while inhibiting the prestalk pathway. A protein homologous to FbiA has been identified via sequence analysis, and it is suitably called FbiB. Mutants of the fbiB- null variety show a less severe phenotype relative to fbiA- nulls; however, they are comparable. Because of this less severe phenotype, it is suspected that the two proteins “rescue”, consequently demonstrating functional redundancy. Therefore, it is of interest to create an fbiA-fbiB- double null mutant in order to study this relationship. This study took steps toward confirming existing pools of double mutant candidates; however, no conclusive evidence emerged from various PCR analyses to verify genuine double mutants in those pools. As a result, a fresh attempt at creating the fbiA-fbiB- double knockout was commenced, which began by transforming Blasticidin sensitive fbiB- single null mutants. A PCR analysis of newly created double knockout candidates proved to be extraordinarily promising.

Major

Biology

First Reader(s)

Nelson, Margaret K.