- Wash and image Western on Saturday morning
July 3, 2006
- Start SUMO gel with ATP analog/arrested yeast from Friday. - DONE
- SO FEW YEAST - is something wrong? What protocol to use? Same one?
- Who is "the boy who lived" on my pGEV transformation?
- Work on biobricking PPL parts (now that I have Endy Lab membership)
- What if the pGEV we have is on a CEN/ARS plasmid with a Trp marker, like the paper says? Is this even a remote possibility?
- Come up with schedule for rest of week... July 4 vacation day? - DONE
- Colony PCR of "the boy who lived" - DONE
- Transform new pGEV in to E. coli - DONE
July 4, 2006
- Day off? Institute holiday...
July 5, 2006
- Native extraction! using unarrested 403 - DONE
- Pour new sumo gel for native extraction samples - DONE
- Transfer and overnight incubation of Western from July 3 - DONE
- Run a gel of the colony PCR from Monday - DONE
- If it works, and pGEV is there, grow up overnight culture from new plate
- If it works, and pGEV isn't there...
- If it doesn't work, try again (see July 6).
- Conclusion: something is happening, but it isn't either bad nor good. Mixed bag. Try again.
- Order anti-Fus3 antibody from Santa Cruz, order Rainbow Marker from GE Healthcare. - DONE
- Set up overnight culture of pGEV E. coli in LB+Amp. - Had to plate anew (1:200) instead, but that is DONE
- Set up overnight of 403-1030-1018 in SCD -u -h. -DONE
July 6, 2006
- Wash and image Western - DONE
- Try colony PCR again (AM) and run gel - DONE
- Grow up overnight culture of pGEV404 in DH5alpha in LB+Amp. -DONE
- Grow up overnight culture of 403-1030-1018 in SCR -u -h. - DONE
- Set up freezer stock (the kind in glycerol) of 403-1030-1018 from yesterday's overnight. -DONE
- Debug peristaltics - DONE
July 7, 2006
- Miniprep pGEV out of E. coli, do characteristic digest (run gel), linearize pGEV with SnaBI (run gel). -DONE
- Transfer and overnight incubation of Western - set up transfer
- If sumo gel with noc-arrested samples didn't work, re-arreast cells. - DONE, even though it worked
- If there's time, do a PCR of the yeast genome with Trp1_A and Trp1_D.- MOVED TO MONDAY
- Transform pGEV in to 1030-1018 using green star EZ method - DONE
July 8, 2006
- Set up overnight incubation of Western - DONE
July 9, 2006
- Imaging of Western -DONE
- Pour new 12% SUMO gel - DONE
- Prep arrested cells from Friday and load on gel - DONE