Figure 1.

TLS by yPol η, hPol η, yPol ζ, hPol ι, and hPol κ on UV-irradiated template. (A) Illustration of experimental design. See main text for explanation. (B) Primer extension by yPol δ alone. A ssDNA template (Figure 2A) was irradiated with UVB (302 nm; 0, 0.5, 1, 2.5, 5 and 10 kJ/m² in lanes 2–7) or UVC (254 nm; 0, 0.1, 0.2, 0.5, 1, and 2 kJ/m² in lanes 9–14), and used in primer extension by yPol δ. Products were analysed by electrophoresis through a polyacrylamide gel containing 7M urea (sequencing gel). Potential sites of pyrimidine dimers (indicated by ‘]’) and major sites that blocked the extension (dots) were indicated on the template sequence. (C) Quantification of relative amount of full-length DNA products from panel B. (D) Primer extension experiments were carried out using yPol δ and the indicated second DNA polymerases (second Pol; ‘yη’ = yPol η, ‘ζ’ = yPol ζ, ‘hη’ = hPol η, ‘ι’ = hPol ι, ‘κ’ = hPol κ). Samples were withdrawn from the reaction at indicated times and analysed by the gel electrophoresis. (E) Similar experiments to D with indicated second DNA polymerases were repeated and relative formation of full-length product after 30 min of reaction were calculated (mean + SEM, n = 3, ***P < 0.001, ****P < 0.0001, ^nsp ≥ 0.05; One-way ANOVA with Bonferroni's multiple comparison test to the ‘δ’ data that is shown as a white bar).