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. 2009 Nov 11;38(2):467–476. doi: 10.1093/nar/gkp929

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© The Author(s) 2009. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 1. — Generation and characterization of telomeric DNA substrates. (A) pRST5 contains an insert of 98 telomeric repeats (bold), which is excised by BsmBI/HindIII to generate 0.6 kb linear substrates containing exclusively telomeric DNA. Overhangs are filled by T4 DNA polymerase, dA/G/TTP and biotinylated dCTP. After incubation of immobilized substrates for 20 min in cytosolic extracts, proteins in the soluble fraction or bound to DNA are monitored by western blotting. Non-biotinylated DNA is used for replication and NHEJ assays. A NT control plasmid pRST5_NT harbors a random sequence of similar length. (B) Bead-bound NT or telomeric (T) DNA was incubated with cytosolic extracts at 2 × 10⁹ ends/µl. Soluble fractions were separated (lanes 1–3), beads washed and bead-bound fractions (lanes 4–6) analyzed by western blotting. Lanes 3 and 6, beads only.