Figure 7.

Analysis of APE activity in S.pombe extract. Wild-type and mutant cells were broken by French press and centrifuged. Crude cell extract (1 µg) was mixed with 50 fmol of the oligonucleotide containing the AP site in reaction mixture (20 µl) containing 20 mM Tris pH 8.0, 100 mM NaCl, 1 mM MgCl₂, 1 mM DTT and 100 µg/ml BSA. After incubation at 37°C, the 5′-labeled 30mer product was identified with PhosphorImager (Molecular Dynamics) after electrophoretic separation on 20% polyacrylamide gels containing 7 M urea (26,45).