Construction and initial characterization of yeast strains with Dyskeratosis congenita mutations
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Date
2017
Authors
Ogailan, Abeer Abdullah
University of Lethbridge. Faculty of Arts and Science
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Publisher
Lethbridge, Alta : University of Lethbridge, Dept. of Chemistry and Biochemistry
Abstract
The effect that Dyskeratosis congenita mutations have on ribosome biogenesis has not been fully investigated. Here, I am studying the effect of Dyskeratosis congenita mutations using a yeast model system with the goal of identifying impacts on ribosome biogenesis. I constructed six haploid yeast strains with a single mutation in the CBF5 gene each using site-directed mutagenesis and chromosomal integration. First, I screened for phenotypic effects on growth on solid and in liquid medium at different temperatures; these experiments show normal growth behavior of the mutant strains compared to wild type. Second, Cbf5p expression levels were examined in whole-cell extract via western blotting analysis, which again did not reveal any difference to wild type. Finally, progression through the cell cycle was investigated in synchronized cells. Together, this study indicates that individual Dyskeratosis congenita mutations are well tolerated in the unicellular eukaryote Saccharomyces cerevisiae.
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Keywords
CBF5 gene , Dyskeratosis congenita mutations , H/ACA RNA , H/ACA sRNP , ribosome biogenesis , yeast strains