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. 2003 Nov 1;31(21):6117–6126. doi: 10.1093/nar/gkg814

Figure 4.

Figure 4

DNA polymerase activity of POLQ. (A) Purified recombinant POLQ. Silver-stained gel showing molecular weight markers (lane 1) and 30 ng of purified POLQ (lane 2). Proteins were separated by electrophoresis on an SDS 4–15% polyacrylamide gradient gel. Two separate samples of 5 ng of POLQ were loaded onto similar gels, immunoblotted with a 1:1000 dilution of POLQ antibody 1 (lane 3) or antibody 2 (lane 4), and detected with secondary reagents as described in Materials and Methods. (B) The hairpin primer–template for the primer extension assay. The self- complementary oligonucleotide was labeled with 32P at the 5′ end. (C) The primer–template (16 fmol) was incubated with 1.25 ng of POLQ in the presence of a single dNTP (A, T, G or C, lanes 3–6), two dNTPs (lane 7), three dNTPs (lane 8) or all four dNTPs (lane 9). EDTA was added before incubation for the reaction in lane 1, and dNTPs were omitted in lane 2. The reaction mixture in lane 10 included all four dNTPs and 6 × 10 –4 U of Taq DNA polymerase (Promega), incubated for 30 min at 74°C.