 | Eileen Oni Rowan University BiologyMentor(s)Ron Hart, Ph.D. Jonathan Davila Chris Ricupero Cindy Camarillo, Ph.D. Department of Cell Biology and Neuroscience Mavis Swerdel W.M Keck Center for Collaborative Neuroscience Rutgers University |
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| Identifying microRNA precursors capable of cleavage by Microprocessor in embryonic stem cells | ||
| MicroRNAs, cleaved from hairpin-shaped precursor RNAs by a cellular complex known as the Microprocessor, play an important role in preserving the pluripotency of embryonic stem cells. The interaction between mature microRNAs and the mRNAs responsible for differentiation stimulates translational repression and/or degradation of the target mRNA, reducing protein synthesis. Through previous work, unique microRNA sequences have been identified through deep sequencing techniques. Since it is not fully understood which of these predicted precursor microRNAs are capable of hairpin cleavage, we aim to determine whether they are capable of such processing. Precursor microRNA sequences inserted downstream of a green fluorescent protein (GFP) plasmid by recombination were transfected into HEK-293 cells. For a visual assessment of hairpin cleavage, intensity of GFP fluorescence was noted through photomicroscopy and fluorescence activated sorting (FACS). From the inhibition of GFP fluorescence expression caused by cleavage of hairpin precursors in GFP mRNA, we conclude here that the predicted microRNA precursors are capable of cleavage. The results of these preliminary studies will help to determine, on a larger scale, which predicted microRNAs are capable of hairpin cleavage while understanding a significant participant of pluripotency regulation by producing a gain-of-function assay for newly-discovered microRNAs. |