Lissa1: June24-June30

Weekend June 24-25, 2006

 * 1) Maybe pour SUMO gel on Sunday night

Monday, June 26, 2006

 * 1) Brainstorm on why transformations are failing
 * 2) Make new freezer stock of analog-inhibited Cdc28
 * 3) DIAGNOSTICS:
 * 4) Pour more SCD -U-T plates
 * 5) POur more SCD -U-H plates
 * 6) Use yeast freezer stocks to plate test transformations.
 * 7) Re-digest 404 with SnaBI and run an agarose gel to see if it worked.
 * 8) Grow up an overnight of pGEV e. coli so I can miniprep more tomorrow
 * 9) Pour a sumo gel with 12% acrylamide
 * 10) Order restriction enzymes from NEB, and run diagnostic digest on pGEV
 * 11) Grow s288c (non-knockout yeast) on my old plates and see if they can survive at all - if they can, then the plates are ok - if they can't, then the plates are bad.

Tuesday, June 27, 2006

 * 1) Run sumo gel without 379 alpha - and use peristaltic pump (make more buffer)
 * 2) Make new freezer stock of yeast from yesterday's overnight culture
 * 3) Transform SSY403
 * 4) Miniprep pGEV, pRS404 and pRS414 out of E. coli
 * 5) Do characteristic digests of pGEV and run on gel

Wednesday, June 28, 2006

 * 1) TALK AT GROUP MEETING TODAY
 * 2) Check on SUMO GEL
 * 3) FIX PERISTALTIC PUMP
 * 4) Make more anode buffer
 * 5) Learn to use pH meter
 * 6) Check on plates
 * 7) Order antibodies -on hold until I hear from Drew
 * 8) Anti-Fus3
 * 9) Anti-active MAPK
 * 10) What to do about anti-Cdc28 antibodies?
 * 11) When to do beta-estradiol stocks? Probably not until transformation is working.
 * 12) Think about pGEV issue/digest
 * 13) Do another pGEV digest, same as yesterday, just to be sure I've done it right.
 * 14) Run out old gel to get 'exact' length of EcoRV-cut fragment
 * 15) Grow new overnight of yeast?
 * 16) If cells in -80 freezer are not competent, or if we decide the EZ transformation really doesn't work, we'll need new cells.

Thursday, June 29, 2006

 * 1) Do transfer and set up overnight incubation of Western
 * 2) Transform pGEV in to yeast, with 404, 414, and 416.
 * 3) Grow up overnight of 403 that are currently on a plate in the 30 degree room.

Friday, June 30, 2006

 * 1) Make new freezer stock of 403 chemically inhibited yeast.
 * 2) Start Western on overnight incubation
 * 3) Pour new SUMO gel
 * 4) Native extraction!