Click-chemistry, a new tool for microbiology? (2014)

Undergraduate: Mauricio Barreto

Faculty Advisor: Barbara (Dr.) MacGregor
Department: Biology

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We are currently working on a new method that use the affinity of biotin/streptavidin in order to more effectively understand and study the microorganisms that reside in marine sediments. This method could be used to identify genes expressed in response to particular conditions, or ¿ in low-energy environments such as the deep subsurface ¿ to investigate whether RNA is turned over at all. This method uses the ¿click¿ chemistry reaction in order to identify the RNA transcripts being produced at a particular time in marine sediments. This process involves the uptake of the RNA analog 5-ethynyl uridine (EU) into incubated cells. After incubation of samples with EU label, total RNA is isolated, tagged with desthiobiotin-azide, and tagged RNA captured with T1 streptavidin beads for further analysis. Our results have shown that this method has been successful in labeling and capturing RNA in various forms of cultured bacteria including E. Coli and B. Subtilis. While yielded results were not as strong for B. Subtilis, the positive results from E.Coli suggest that this method can indeed be used to label and capture RNA from prokaryotic cells. We are currently applying this method now to real samples to determine the plausibility of it¿s use and are currently sequencing captured RNA to more comprehensively determine the effectiveness of this method and the types of RNA being captured.