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. 2005 Mar;25(6):2158–2168. doi: 10.1128/MCB.25.6.2158-2168.2005

FIG. 2.

FIG. 2.

Validation of the overhang protection assay. (A) The overhang protection assay was performed on single-stranded oligonucleotides containing (TTAGGG)n repeats (n = 6, 9, 16, 32, 48, and 64). Oligonucleotides were either annealed directly to the C-rich probe (annealed) or underwent the overhang protection assay (protected). Weighted mean sizes were calculated after quantitating the signals in each lane with ImageQuant software between molecular size (M) marker positions of 36 to 384 bp. (B) Results of the assay in panel A are plotted. (C) The overhang protection assay was performed on the model template pBSK-Rep4, which contained an ∼450-nt 3′ (TTAGGG)n overhang after 3 kb of double-stranded pBSK vector sequences. Prior to the assay, a different amount of pBSK-Rep4 was mixed with or without HeLa genomic DNA. (D) Whole genomic DNA from HeLa cells was digested with T7 gene 6 exonuclease (a 5′-to-3′ exonuclease) for the indicated times and then treated with or without Exo I before being analyzed in the overhang protection assay. Black lines indicate positions of the bulk of the overhang DNA.