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Denaturing PAGE (8%) showing the impact of absence/presence of PCNA on processive leading strand synthesis by wild type Pol ε and its variants. Extension of a 5′-TET-labeled 35–mer oligo annealed to M13mp18ssDNA (NEB). Reactions were performed at 40-fold molar excess of DNA substrate over polymerase to meet single-hit criteria. Individual reactions were stopped at 0, 2, 5, and 15 min, respectively.

Journal: Nucleic Acids Research

Article Title: A thumb-domain insertion balances processivity and fidelity in DNA polymerase ε

doi: 10.1093/nar/gkag282

Figure Lengend Snippet: Denaturing PAGE (8%) showing the impact of absence/presence of PCNA on processive leading strand synthesis by wild type Pol ε and its variants. Extension of a 5′-TET-labeled 35–mer oligo annealed to M13mp18ssDNA (NEB). Reactions were performed at 40-fold molar excess of DNA substrate over polymerase to meet single-hit criteria. Individual reactions were stopped at 0, 2, 5, and 15 min, respectively.

Article Snippet: The substrate for processivity assays was prepared by annealing M13mp18 single-stranded DNA (NEB) with a 5′-TET–labeled 35-mer oligonucleotide (TET-CCCAGTCACGACGTTGTAAAACGACGGCCAGTGCC) in equimolar amounts in 125 mM NaAc (pH 7.8).

Techniques: Labeling