6/13
Today I began to make my own growth media for my Clone A cells (a type of colon cancer). I retrieved a bottle of media from the cold room and added in 50 mL of FBS (Fetal Bovine Serum) and 50 mL of Pen/strep (Penicillin/Streptomycin). After I formed this media, I applied it to Clone A cells in a 10cm dish and also to cells in a 6-well plate, which will be used for a death curve. After this, I went to the computer and changed the sequence of some RNA to form a hairpin loop and ordered these new sequences off of the internet.
6/14
I began today by helping Brian (my mentor) run a Western blot to test for the expression of Fascin in certain lysates. I loaded the lysates into a 12% gel and ran electricity through it. After, we took the gel into a dark room and developed the image using special chemicals (called PICO) and put the image onto a film. Next, I added .5µg/mL Puromycin (a poison) to the Clone A cells in the 6-well plate. Leaving the first well empty as a control, I added Puromycin to the remaining 5 wells, gradually increasing the amount. This test would determine the minimal amount of Puromycin necessary to kill Clone A cells.
6/15
I came in today and observed my death curve. My results concluded that 5µg of Puromycin would efficiently kill Clone A cells. I replaced the media and puromycin in the 6-well plate, then split the cells in the 10cm dish by centrifuging them into a pellet, re-suspending them in media and then only adding half of that media to the plate.
