Journal: Nucleic Acids Research
Article Title: Excision of uracil from DNA by hSMUG1 includes strand incision and processing
doi: 10.1093/nar/gky1184
Figure Lengend Snippet: Trapping experiments for Schiff base intermediate. Left panel, EcFpg (17 pmol) alone as a negative control, and together with EcUng (3 pmol) as a positive control, EcUng as well as hUNG (5 pmol) alone as negative controls, and hSMUG1 (0.3 pmol) alone, were incubated with substrate 2 (1 pmol) and 50 mM NaBH 4 in reaction buffer at 37°C for 1 h (final volume, 10 μl). Right panel, EcFpg (10 pmol) alone as a negative control, and together with EcUng (10 pmol) as a positive control, EcUng as well as hUNG (10 pmol) alone as negative controls, and hSMUG1 (10 pmol) alone, were incubated with substrate 2 (1 pmol) and 50 mM NaBH 4 in reaction buffer at 37°C for 1 h (final volume, 10 μl). In each case (A and B), trapped was separated from un-trapped substrate by denaturing PAGE [10% (w/v)] at 200 V for 1 h. The experiments were performed in triplicate showing the same result.
Article Snippet: Enzymes hSMUG1 (full length) was obtained from NEB (New England BioLabs) and investigated for contaminants by MS analysis (see ) as well as purified by us [see , Production of purified hSMUG1(25–270) and ].
Techniques: Negative Control, Positive Control, Incubation, Polyacrylamide Gel Electrophoresis