User:Karmella Haynes/Notebook/Polycomb project/2010/07/01

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07/01/10

 * &#x2713; Transfection: FTRx H3me reporter clones

Transfections > U2OS cells + (H3me reporter + Gal-VP64) constructs > Use Fugene, 12-well format > Samples (plated in duplicate, transfect just one): --> KAH96-2, 3, 5, 6 --> KAH142-1, 2, 3, 4, 6, 7, 8

> Master mix (12x): 18 μL Fugene + 582 μL Opti-MEM --> R.T./ 5 min. > Add 14.4 μL DNA --> R.T./ 20 min. > Add 51.2 μL DNA/Fugene mix to each well (2 ml medium/ well); Grow cells at 37&deg;C > Check RFP/ YFP after 18-24 hours

7/02/10 > Good RFP (Gal4-VP64 activator) signal in all Fugene transfected samples, but no YFP induction. The co-transfection experiment showed that Gal4-VP64 does activate H3me reporter, so my conclusion is that these cells are not stably transfected w/ the H3me sensor genes. Repeat transfection of H3me sensors.


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