9:00 A – 6:50P, 8:20P -11:15P
- discuss CMAKE minGW sys with JM
- 9:30 A flip collection plate
- Working on BioPhys J review.
- 11:30 A – check samples from oven
- substantial issues with resin quality: 1 sample R solid at 70C. Other three samples (2 S 1R) very soft, easily squeezable. This is probably a substantial source of etching issues. Should do more pure resin changes.
- previously have observed soft resin at top and hard resin at bottom. could be that EtOH resin is harder and that fresh resin added at the top polymerizes more slowly, or that resin separates over time. Can test this with some straight resin samples. (try tonight)
- 2 samples pure resin, 2 samples resin + 15uL EtOH, 2 samples resin + 50 uL EtOH, baking O/N at 70C, 9:00PM
- 1:15 PM remove samples from oven, let cool.
- 4:30 P, samples much stiffer when cool, gel-like samples now flaky if scratched.
- 12:00P microscope seminar BL1080
- Working on En locus data analysis from 09-12-12. (manual inspection of different nuclei using conventional overlay + switching cycles to ID labeled loci).
- sectioned hard sample R (rabbit-H3 direct label). 3 coverslips. Some notable number of burs on shared knife. Sections cutting excellent ribbons.
- Cut sections of DNA FISH+protein: lamin still present in Cy3B, no 488 nuclear stain detectable, no 405 stain detectable from 750-H3. Try stronger/longer postfix?
Thoughts on Insulator experiments
Why is en not derepressed in Esc mutants, but AbdB is?
- bistable Trx/Pc system tipped far enough at AbdB locus but not at Pc
- different components initiate silencing
- different components maintain silent state (PRC1 vs PRC2 thing as alternatives rather than cooperatives?)
- differences in timing to establish silent state.
- Pc/Scm double mutants de-repress both.