Lissa1: August6-August14
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August 6
August 7
- Pour new SUMO gel -DONE
- Pour new small gels -DONE
- Set up overnights for the following experiments:
- Nocodazole arrest -DONE
- Induction experiment -DONE
- Make more unarrested 403 for phosphatase Western -DONE
- Make media for the nocodazole arrest. -DONE
- Plan EVERYTHING -CHECK
- Cloning
- Finalized experiments
- What to do about luminol/ECF deal? - OOPS, stil don't know
- Integrate new-found numbers/data in to model -DONE
- Maybe update model to include Michaelis-Menton kinetics -NOT YET, talk to Ty
August 8
- Do nocodazole arrest! -CELLS WON'T GROW:(
- Do induction experiment! -DONE
- Set up overnights for the following experiments: -MOVED UNTIL TOMORROW
- Fus3/Active Fus3 gels
- Phosphatase control
- PCR up pGEV out of ACLY700 -DONE
- Run a gel of the PCR -DONE
- If there's time, run a second gel and extract and purify the pGEV (which might not be pGEV, based on the sequencing data) -DONE
August 9
- PCR amplify the pGEV band I purified yesterday -DONE
- PCR out GEV insert from the original pGEV DNA -DONE
- Run gels of these PCRs and do gel extraction -DONE
- Digest pGEV - no time
- Setup overnight of pRS-405 in DH5alpha, in LB + Amp -DONE
- Streak new plate -DONE
- Setup overnight of samples for Fus3, etc -DONE
- Unfortunately, I've got a cold and need to rest:(
August 10
- Prep Fus3 samples (all the way to sample buffer) and store -DOne
- Get concentrations on all DNA - DONE
- Miniprep 405 out of E. coli -DONE
- Streak out new 403 plate from freezer stock -DONE
- Digest pGEV, run on gel to check -DONE
- Design diagnostics for prs405 DNA -DONE
- Digest 405 -DONE
- Do PCR cleanup - DONE
August 11
- Make new SCR-u-h media!
- Do native extraction and phosphatase experiment on Noc. arrested yeast
- Load SUMO gel and start run
- PCR up GEV insert out of miniprepped pGEV DNA
August 12
- Gel-extract and purify all digests/PCRs. -DONE
- Check on SUMO gel l- SOMETHING IS VERY WRONG. CHANGED ALL BUFFERS, CHECKED ALL CONNECTIONS, SET UP AGAIN. DID NOT SEE BUBBLES AT ALL. 1 HR LATER IT HAD MOVED A LITTLE BIT....
August 13
- The plan was to set up the transfer of my gel today, but whatever problem was present yesterday is also present today. I think a wire must have snapped inside the gel box? It's giving 400 V but only 2 mA of current...
August 13, 2006
- Digest my new PCR-ed fragments that hopefully are pGEV with the double digest -DONE
- At the end of the day, do the ligation
- Think about this week's very complicated schedule -DONE
- Set up overnight of my newly-streaked-out 403 cells to do noc. arrest and also to set up new samples for phosphatase control -DONE
- Get opinions on induction experiment -DONE
- Set up new overnight for another induction
- Scrap old SUMO gel and pour a new one
- Prep and run 2 new small Fus3/etc gels -DONE