RNA Expression Test

April 13, 2022

Our RNA was injected into Xenopus laevis oocytes by Ami Patel. Our first experiment with electrophysiology was not successful, as the electrode repeatedly became jammed with debris.

April 15, 2022

Ami Patel injected our KCNJ5 RNA into the oocytes again at a concentration of 450pg/nL and was incubated for 72 hours. This experiment was much more successful, and the data indicated that our RNA expressed successfully.

Frog Ringer's Solution (FR) is an isotonic solution to the frog oocytes at physiological pH. It is also used in many other experiments, which makes it a good point of comparison. The oocyte was bathed in FR for 5 minutes to establish baseline conductance. It was then placed in high K+ (90%) FR to see if the RNA expressed. The increased conductance indicates that our RNA expressed. We kept the oocyte in the 90% K+ FR for 10 minutes to establish the new baseline conductance. The oocyte was then put in a 1uM solution of DAMGO for 1.5 minutes. The large (>20uS) response to DAMGO indicated that the KCNJ5 RNA expressed KCNJ5 channels. It was then placed back in the 90% K+ FR, baseline was reestablished, and then placed back in normal FR. Details are depicted below.