(A) Representative gel of five independent biological experiments showing that EXO1 detects the 5′ DNA 20-mer flap by tracking and threading. Lanes 1–2 contain substrate GK249, lanes 3–4 substrate GK250, which contain a hairpin loop in the ssDNA (starting at nucleotide position 6 from the 5′ end), and lanes 5–6 substrate GK254 which contains a TEG-biotin, at position 6 nt from 5′ end on the ssDNA and lanes 7–8 substrate GK254 contains in presents of streptavidin. Lanes 2, 4, 6 and 8 are substrates in the presence of EXO1. (B) Statistical analysis of degradation of the different DNA conversions. One-way analysis of variance with Tukey's multiple-comparison test. GK250 shows a trend in versus GK249. GK254 and GK249 show a similar degradation. (C) A schematic model of tracking, where EXO1 detect the junction of the 5′ flap and cleave directly. (D) A schematic model of threading, where EXO1 detects the ssDNA end at the 5′ flap and slide of the ssDNA to the junction of the 5′ flap and cleave.