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The reaction was then quenched by the addition of 5x reducing buffer (50% glycerol, 250 mM Tris-Cl [pH 6.9], 10% SDS, 0.05% bromophenol blue, 6.25% beta-mercaptoethanol), boiled and separated by sodium dodecyl sulfate polyacrylamide (SDS-PAGE) on 15% polyacrylamide gels