User:Karmella Haynes/Notebook/Polycomb project/2010/02/02

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02/02/10

 * &#x2713; Minipreps: KAH139-141/V0201
 * &#x2713; Transfection: Gal4-EED expression constructs
 * &#x2713; Thaw cells: HEK293 Luc #4 (to test Gal4-EED function)

Minipreps > Check with XbaI digest

--> Combine duplicate samples, measure conc., and use for Lipo transfection

Lipo transfection > Details

> Add Lipo to 250 μL Opti-MEM --> R.T/ 5 min. > Add DNA to 250 μL Opti-MEM > Add DNA mix to Lipo mix --> R.T./ 20 min. > Starting vol. is 3.5 mL, no need to remove 500 μL; Add 500 μL complexes to each well (4 ml med. each); Grow cells at 37&deg;C > Refresh medium after 5 hours (ab-free) > Add neomycin to samples 2-6

Day 2 > Do protein preps on all samples (for EED Western blot)


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