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. 2016 May 24;44(12):5861–5871. doi: 10.1093/nar/gkw468

Figure 2.

Figure 2.

MGME1 cleaves imprecisely around the flap base. (A) Mapping of MGME1 final cleavage sites shows a pool of products with short flaps, nicks and 1 nt gaps. Nuclease assays were performed on 15 nt flap substrates that had been radioactively labeled at the 3′-terminus on the downstream oligonucleotide. The numbered values in the rightmost lane indicate how close to the base MGME1 can cleave (0 = the whole flap is cleaved off and a ligatable nick is created). (B) Substrates (10 fmol) containing no flaps (lanes 1–6), 1 nt flaps (lanes 7–12) and 30 nt flaps (lanes 13–18) were incubated with increasing amounts of MGME1 (0, 12.5, 50, 250, 1250 fmol) in the presence of T4 DNA ligase. Ligation of the upstream and downstream primers results in an 80 nt product (arrowhead). MM, molecular marker. (C) As in (B), except repeated in the presence of POLγ. Note that in both (A) and (B), reaction buffer included dNTPs and ATP.