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. 2017 Aug 18;7:8729. doi: 10.1038/s41598-017-08663-x

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Rap1 protects the 5′ end when bound to dsDNA 2 nt inwards of the ds-ss junction. (a,d) The substrates used for DEPA experiments. The black bar indicates the guide sequence, and “*” the radioactive label (as described in Fig. 2 legend). (a) The telomere substrate D17S9 contains 17 bp dsDNA and a 9 nt 3′ overhang. The Rap1 MBS (bold text) is located 2 nt inwards of the ds-ss junction. (d) Substrate D17S9m contains 17 bp telomere dsDNA with four mutations in the Rap1 MBS (bold letters, mutations in red) and a 9 nt ssDNA 3′ overhang. (b,e) DEPA sequencing gels of D17S9 (b) and D17S9m (e), showing the loading control (LC) and uncleaved substrate (S) from the Rap1 containing and BSA control reactions at 0; 20; 40; 60; 120 s. Full-size gel showing all the reaction products can be found in Supplementary Fig. S4. (c,f) Quantification of the gels shown in (b) and (e) respectively.