Tuesday 12/04/12

9:40 A – 7:45P, 9:00P – 1:00A

Dfd/tub analysis

  • Break down O/N confocal run
  • 3 channels, speed 6, 22 positions, 17.5 finished in 11 hour run.  contact zeiss rep Casey about changing channels between z-stacks rather than every section, should be much faster.
  • annotating ages from overview images of last nights’ Dfd tub data
  • Analyzing data (see post).

Probe making,

  • spin down dissolving DNA from pulverized gels, remove supernatant for precipitation, return remaining gel to 55C shaker with fresh ddH2O to elute remaining probe.
  • precipitate eluted probe

In situs Day 2

  •  hot washes
  • block
  • primary antibodies O/N

Ucryo sections on coverglass

  • dk a-rb-488 doesn’t label either, so hopefully not a problem with the abcam dk a-rab 750 I made earlier.
  • reblocked and staining with H3K9me2 again (5:300 of the rediluted in glycerol stock).  It would be really nice if this would work.
Cell culture test 50kb probe
  •  Plate new cells from overgrown 11/20 culture flask
  • Odd colored precpitate observed in mixing flask.  Cells tossed.
  • Contact Nicole about starting a new culture going from frozen stock.  Current line has been in continuous culture since Aug.
  • Fix new plated cells.  (Attachment was poor / or even new small squeeze bottle has too much force, most cells detached).
  • Prehybe 60C, denature probe, denature cells + probe at 92, incubate 37C O/N
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