Tandem RNA affinity purifications to study an essential snoRNA in ribosome assembly

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Rocca, Daniel G.
University of Lethbridge. Faculty of Arts and Science
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Lethbridge, Alta. : University of Lethbridge, Dept. of Chemistry and Biochemistry
Eukaryotic ribosome assembly is an essential process involving many protein and RNA assembly factors. Mutations of genes encoding ribosomal proteins or assembly factors can result in diseases in humans. The upregulation of ribosome assembly is also a hallmark of many cancers. snR30 is an interesting assembly factor due to its role as an unusual and essential H/ACA small nucleolar RNA and as it is one of only three essential RNA assembly factors. Here, I have developed a system for the co-expression of aptamer-tagged variants of snR30 and truncations of ribosomal RNA in Saccharomyces cerevisiae. Next, I optimized the tandem purifications of the snR30-bound pre-ribosomes utilizing these aptamers. In conclusion, my system will allow for in-depth characterization of the composition and structure of snR30-bound ribosome precursors. This characterization will improve the understanding of the essential activity of snR30 for ribosome assembly in yeast and inform future studies on the function of the homologous U17 RNA in humans.
snR30 , Ribosome assembly