NIH Research Festival
Post-transcriptional modifications in RNA play numerous roles in enhancing and extending the functionality of the RNA species. In particular, transfer RNA (tRNA) contains a high density and diversity of modifications that are crucial for its structure, function, and stability. Recent evidence suggests that the modification status of tRNA changes in response to fluctuating environmental conditions. To decode these modifications, a number of next-generation sequencing technologies have been developed. We adapted an MTSEA affinity-based enrichment strategy to characterize the presence of the ribonucleotide modification 4-thiouridine (4SU) at the transcriptome-wide level in Escherichia coli. Thiol-modified tRNAs are enriched via MTSEA capture and cDNA libraries are generated for next-generation sequencing. Here, we investigate various metabolic and environmental stress conditions under which 4SU may be added to Escherichia coli BW25113 tRNAs. Future work will expand upon the link between thiol modification status and stress response in other species, and may contribute to our understanding of thiolated RNAs in human disease.
Scientific Focus Area: Molecular Biology and Biochemistry
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