2013 URM Students

Jorge Felipe Ortiz-Carpena  
Mentor: Dr. Joshua J. Rosenthal

Deciphering the Functional Effects of RNA-editing in Squid Kv1.3 Channels

RNA editing describes a set of post-transcriptional processes that modify the nucleotide sequence of RNA’s from that encoded by their genes. In a sense, this molecular process provides eukaryotes a selectivity filter that affects the flow of genetic information, enabling an organism to remove deleterious mutations from the genome or create novel transcripts. Several model organisms have been used to study RNA editing, however squids use this process extensively to modify their protein function in order to better adapt to their dynamic environments. By studying the squid model, we can potentially gain insights into the specific mechanisms underlying editing. Recent work from our laboratory has identified a novel squid potassium channel (SqKv1.3) from a newly generated transcriptome from the squid giant axon system. Although this channel is clearly a member of the Kv1 subfamily, its sequence reveals a number of unusual features. For example, its voltage sensor lacks one of the conserved positive charges used to gate. Jorge’s studies have focused on cloning SqKv1.3, identifying the RNA-editing sites in SqKv1.3 and characterizing the effects of these molecular changes on channel function. So far, Jorge has cloned SqKv1.3 and identified 12 novel RNA-editing sites within its messages. Recently he has begun preliminary expression studies of SqKv1.3 in Xenopus oocytes. By studying its electrophysiological properties using a two-electrode voltage clamp, he hopes to better understand how this channel’s novel structural properties affect function.

Francisco A. Montalvo Soto  
Mentor: Dr. Joshua J. Rosenthal

Design and Construction of a Sensitive Low Cost Fluorometer

Fluorometers have become indispensable tools for FRET based experiments in cellular and molecular biology. The cost of such an instrument is usually very high. However the basic functionality of a flourometer makes it easy to replicate at a lower cost utilizing basic electronics. A flourometer works by transferring light energy to a sample containing a chromophore. Chromophores are part of a molecule that absorb certain wavelengths of visible light, emit others and reflect others. The excited sample’s chromophore will emit the energy in the form of a light wavelength. Finally this emission is picked up by a sensor that interprets it in the form of voltage changes. The voltage signaled is then amplified and noise filtered before being picked up by an oscilloscope. The voltage is directly proportional to the amount of fluorescence emitted by the sample. By adding fluorescent sequences in genes that add chromophoric motifs we can determine if a target protein is being produced or edited. Currently, iour lab works on RNA repair. By using this device we can determine if the RNA is being repaired in the desired cell. For this purpose the prototype design will be focused on the detection of MCHERRY and eGFP excitation emission frequencies to verify if the target protein is being produced.