Wednesday 11/27/12

10:15 A — 11:15 PM

Polytene cell staining

  •  hot washes in 2x SSCT (45 C in coplin jar in dry oven)
  • check staining — cent probe looks good (rather small foci though)
  • Antibody staining H3K9 did not work.  Should have included Dm01-Cy3B to check antibody penetration for this tissue prep.
  • attempt to permiabilize with MeOH, 10 min at RT.  Relabel with DM01-Cy3B (O/N at 4C).  (Probably just succeeded in removing cent-probe + Hoechst without sufficiently permiabilizing.  Abcam recommends doing this step at -20C.  Could try acetone at -20C for 10 min if this doesn’t work.

Cell culture, 50kb probe test

  • hot washes
  • check staining –> no dots 🙁
  • not enough probe?  Try again, higher concentration of probe
  • restaining at 1:10 probe
Embryo staining 
  • cent probe
  • failed 50kbp from cell culture.

Fly work:

  • missed virgin collection last night
  • collect males and females for later sorting from 2x HbZ crosses
Group meeting to prep: 12/17

Revisions for snail paper.

  • calculate probe concentrations (100-250 ng/uL stocks)
  • Add cell boundaries to Figure 1
  • Compute total mRNA counts for competitively labeled probes.
  •  Send to ML

PCR repeated

  • primer order:
  • SimD-dna: AbdB, F4R1, F2R2, F1R3, F3R3, F2R1, yw-DNA: AbdB, F2R1


This entry was posted in Summaries and tagged , , , . Bookmark the permalink.