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Briefly, 2 g of purified GST or GST-tagged protein kinases were incubated with 2 g His-tagged ILK5 protein as a substrate in a 25 L reaction buffer containing 20 mM Tris-HCl (pH 7.4), 5 mM MgCl 2 or MnCl 2 , 100 mM NaCl, 2 mM ATP, with or without 0.2 L radioactive [- 32 P] ATP for 3 h at 30C