Cell synchronization

500,000 cells were plated in 6-well plates and synchronized at various stages of the cell cycle: For G1 arrest, cells were treated with mimosine (http://www.sigmaaldrich.com/catalog/product/sigma/m0253), 400 uM, 24 hours, or placed in serum free F-media with no insulin, adenine or EGF for 48 hours. For S phase synchronization cells were place in thymidine (http://www.sigmaaldrich.com/catalog/product/sigma/t1895), 2 ug/ml, 12 hours followed by release for 10 hours followed by either another round of thymidine or aphidicolin (http://www.pfstar.com/Fisher-Scientific-BP615–1-APHIDICOLIN-1MG-EA_p_274776.html), 500 uM for 12 hours. Aphidicolin was also used alone. For mitotic arrest, cells were treated with nocadazole (http://www.sigmaaldrich.com/catalog/product/sigma/m1404), 2 ug/ml, 24 hours or colcemid (https://www.lifetechnologies.com/order/catalog/product/15212012), 20 nM, 16 hours, or subjected to a mitotic shake off.