Single stranded dna

3′-Radiolabelled substrates were made by labelling 10 pmol of single-stranded DNA (Eurofins MWG Operon, Germany) with 3.3 pmol of α-³²P-dATP (PerkinElmer) by incubation with terminal deoxynucleotidyl transferase (TdT, 20 U; ThermoFisher Scientific), at 37 °C for 1 hour, before being passed through a P6 Micro Bio-Spin chromatography column (BioRad), as reported. See ESI Table 1† for oligonucleotide sequences used in this work.

10 pmol of single-stranded DNA (Eurofins MWG Operon, Germany) were labelled with 3.3 pmol of α-³²P-dATP (Perkin Elmer) by terminal deoxynucleotidyl transferase (TdT, 20 U; ThermoFisher Scientific), incubated together at 37°C for 1 h. This solution was passed through a P6 Micro Bio-Spin chromatography column (BioRad), and the radiolabelled DNA was annealed with the appropriate unlabelled oligonucleotides (1:1.5 molar ratio of labelled to unlabelled oligonucleotide) (Supplementary Table S1 for sequences) by heating to 95°C for 5 min, and cooling to below 30°C in annealing buffer (10 mM Tris–HCl; pH 7.5, 100 mM NaCl, 0.1 mM EDTA). Annealed substrates were validated by non-denaturing PAGE (Supplementary Figure S3).

10 pmol of single-stranded DNA (Eurofins MWG Operon, Germany) was labelled with 3.3 pmol of α-³²P-dATP (Perkin Elmer) by incubation with terminal deoxynucleotidyl transferase (TdT, 20 U; ThermoFisher Scientific), at 37°C for 1 h. This solution was then passed through a P6 Micro Bio-Spin chromatography column (BioRad), and the radiolabeled DNA was annealed with the appropriate unlabeled oligonucleotides (1:1.5 molar ratio of labelled to unlabeled oligonucleotide) (Supplementary Table S1 for sequences) by heating to 95°C for 5 min and gradual cooling to below 30°C in annealing buffer (10 mM Tris–HCl; pH 7.5, 100 mM NaCl, 0.1 mM EDTA).