User:Karmella Haynes/Notebook/Polycomb project/2010/08/03

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08/03/10

  • ✓ Transfection: Test H3me Reporter clones
  • ✓ KAH126-3: expand thawed cells
  • ✓ Senescence assay: 6-well plates, -/+ dox, for KAH126-1, 126-3, 132-8, 154-2, FTRx (rotenone, dox, DMSO, u.t.)
  • ✓ Cultures: split confluent plates



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
18 total no plasmid --- 0 μL 1.8 μL 18.2 μL
18 total KAH59/pcVN 200 ng 0.4 μL " "

> Master mixes (20x mix, 2 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


08/04/10
Results: No YFP

08/05/10
Results: Screened for YFP again today and saw lots of YFP+ cells in KAH146-1; re-plate KAH96 & 147 for another transfection


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