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. 2017 Feb 8;56(13):1824–1835. doi: 10.1021/acs.biochem.6b01213

Figure 1.

Figure 1

DNA substrates utilized in this study. The sequence of the primer (29 nucleotides) and template (62 nucleotides) strands comprising each DNA substrate are identical. When annealed, each DNA duplex mimics a blocked P/T junction.42 The size of the double-stranded P/T region (29 bp) is in agreement with the requirements for assembly of a PCNA ring onto DNA by RFC.41,45 The single-stranded DNA (ssDNA) region (33 nucleotides) adjacent to the 3′-end of the P/T junction is consistent with the footprint of RPA (22–30 nucleotides).39 When prebound to Neutravidin, the biotin attached to the 3′-end of a template strand prevents loaded PCNA from sliding off the dsDNA, i.e., duplex, end of the substrate. A Cy3 dye attached to the 5′-end of a primer strand serves as a FRET donor.