User:Karmella Haynes/Notebook/Polycomb project/2010/08/10

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08/10/10

  • ✓ Transfection: H3me Reporter lines (screen for YFP induction)
  • ✓ Microscopy: Fix cells (KAH130, 131 clones)
  • ✓ Western: protein preps for KAH130, 131 clones



Fugene transfection
> 12-well plate
Plate 1. KAH96 clones (6 total)
Plate 2. KAH146 clones (6 total)
Plate 3. KAH147 clones (6 total)

Wells Plasmid DNA Volume Fugene Opti-MEM
20 total no plasmid --- 0 μL 1.8 μL 18.2 μL
20 total KAH59/pcVN 200 ng 0.6 μL " "
20 total KAH60/pcVN 200 ng 0.4 μL " "

> Master mixes (20x mix, 3 mixes): 36.0 μL Fugene + 364.0 μL Opti-MEM --> R.T/ 5 min.
> Add 20x vol DNA --> R.T./ 20 min.
> Add 20 μL complexes to each well (1 ml med. each); Grow cells at 37°C overnight



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