Lissa1: July1-July8

Weekend

1. Wash and image Western on Saturday morning

July 3, 2006

1. Start SUMO gel with ATP analog/arrested yeast from Friday. - DONE
   1. SO FEW YEAST - is something wrong? What protocol to use? Same one?
2. Who is "the boy who lived" on my pGEV transformation?
3. Work on biobricking PPL parts (now that I have Endy Lab membership)
4. 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?
5. Come up with schedule for rest of week... July 4 vacation day? - DONE
6. Colony PCR of "the boy who lived" - DONE
7. Transform new pGEV in to E. coli - DONE

July 4, 2006

1. Day off? Institute holiday...

July 5, 2006

1. Native extraction! using unarrested 403 - DONE
2. Pour new sumo gel for native extraction samples - DONE
3. Transfer and overnight incubation of Western from July 3 - DONE
4. Run a gel of the colony PCR from Monday - DONE
   1. If it works, and pGEV is there, grow up overnight culture from new plate
   2. If it works, and pGEV isn't there...
   3. If it doesn't work, try again (see July 6).
   4. Conclusion: something is happening, but it isn't either bad nor good. Mixed bag. Try again.
5. Order anti-Fus3 antibody from Santa Cruz, order Rainbow Marker from GE Healthcare. - DONE
6. Set up overnight culture of pGEV E. coli in LB+Amp. - Had to plate anew (1:200) instead, but that is DONE
7. Set up overnight of 403-1030-1018 in SCD -u -h. -DONE

July 6, 2006

1. Wash and image Western - DONE
2. Try colony PCR again (AM) and run gel - DONE
3. Grow up overnight culture of pGEV404 in DH5alpha in LB+Amp. -DONE
4. Grow up overnight culture of 403-1030-1018 in SCR -u -h. - DONE
5. Set up freezer stock (the kind in glycerol) of 403-1030-1018 from yesterday's overnight. -DONE
6. Debug peristaltics - DONE

July 7, 2006

1. Miniprep pGEV out of E. coli, do characteristic digest (run gel), linearize pGEV with SnaBI (run gel). -DONE
2. Transfer and overnight incubation of Western - set up transfer
3. If sumo gel with noc-arrested samples didn't work, re-arreast cells. - DONE, even though it worked
4. If there's time, do a PCR of the yeast genome with Trp1_A and Trp1_D.- MOVED TO MONDAY
5. Transform pGEV in to 1030-1018 using green star EZ method - DONE

July 8, 2006

1. Set up overnight incubation of Western - DONE

July 9, 2006

1. Imaging of Western -DONE
2. Pour new 12% SUMO gel - DONE
3. Prep arrested cells from Friday and load on gel - DONE