Monday 06/02/14

9:45am – 4:45pm,


  • Set up RT reactions Lib3 samples B10-B12, C09-C12, D01-D08
  • Wash newly stained samples (B08, B09, E11-F01, + PH immuno-FISH)
  • set up STORM imaging of PH immuno-FISH

Lab Meeting


  • letterhead and signature for letter for NF

PH Project

  • test GA-postfixed cells. no staining of chromatin loci. higher 561/488 background (probably from GA).

Chromatin project

Probe making

setup RT reactions

  • per reaction
    • 4 uL of P1 A405.
    • 5 uL of RT buffer
    • 2 uL dNTP
    • 14 uL RNA
    • 1 uL RNase inhibitor RNAsin
    • .25 uL Maxima
  • master mix 16x
    • 64 uL P1 A405
    • 80 uL RT buffer
    • 32 uL dNTP
    • 16 uL RNase inhbitor
    • 4 uL Maxima


  • ran RT reactions (30 uL scale, 400 uL primer, 15 uL RNA). (B10-B12, C09-C12, D01-D08).
  • poor incorporation and some smearing I think I’m getting substantial RNase degradation. Maybe should run the T7 reactions shorter and let it sit for less long before running RT.
  • probably still decent enough for a staining, we’ll check the concentrations and probably dilute a bit less onto cells.

Left B10-B12, C09-C12, Right D01-D08.


  • STORM imaging of Y B08 region.
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