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The beads were washed 4 times with 1 mL cold wash buffer (100 mM Tris-HCl pH 8, 150 mM NaCl, 10% glycerol, 2 mM DTT, 10 mM NaF, 10 mM Na 2 MoO 4 , 2 mM NaVO 3 , 0.5%(v/v) NP-40, 1%(v/v) plant protease inhibitor cocktail)