The ssDNA [30 nucleotides (nt), *5′-ACGCTGCCGAATTCTACCAGTGCCTTGCTA] was labeled at the 5′ end with biotin. HeLa cells transiently transfected with plasmids encoding FLAG-tagged WT RPA1 or RPA1 mutants were lysed in binding buffer [10 mM tris-HCl (pH 7.5), 100 mM NaCl, BSA (10 μg/ml), 10% glycerol, 0.5% NP-40]. Biotinylated ssDNA was incubated with the lysates for 30 min at room temperature. Streptavidin beads were then added followed by further incubation with the lysates for 1 hour at room temperature. The samples were subjected to SDS-PAGE and analyzed by Western blotting.

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