User:Robert J. Schiemann/Notebook/Lipsick Lab/2010/02/02

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  • Dissect gfp-PCNA x tdt-Myb Flies
    • Chose orange eyed males
    • Dissect testes in 1x PBS Solution. Store in eppendorf with 1x PBS until dissections are finished.
    • Fix with 4% Formaldehyde PBS Solution
      • 30 minutes
    • Wash 3 x 5 minutes in 1x PBS-T
    • Incubate 30 minutes with PBS 0.6% Triton.
    • Wash once with 1x PBS-T
    • Stain with ToPro3 1:1000 in 2% NGS
      • 30 mins at RT
    • Wash 4 x 5 minutes in PBST
    • Mount in Vectashield
  • Collect Flp-out virgins in case PCR fails again
  • Re-run PCR with new reagents