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DNA polymerase activity of Polα is needed for Rad1 chromatin binding. Xenopus sperm chromatin (10,000 nuclei/μL) was preincubated in cytosol containing the indicated concentrations of aphidicolin for 30 min, then mixed with an equal volume of NPE. The chromatin was isolated 50 min later and bound proteins were analyzed by immunoblotting with the indicated antibodies. Prior to chromatin isolation, samples were taken and immunoblotted with antibodies to phospho-Chk1 (S344) and Chk1.
