Figure 4.
Higher percentages of broken forks are detected by DNA combing compared to spreading, and this difference is reduced by addition of proteinase K to the spreading and hybrid spreading protocols. (A) Representative images of DNA fibers obtained with the combing approach. Scale bar is 10 µm. (B) Blue–green–blue versus blue–green forks scored after combing and presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. Statistics: mean ± SEM. Number of biological repeats N = 3. (C) Representative images of DNA fibers obtained with the spreading approach. Scale bar is 10 µm. (D and E) Blue–green–blue versus blue–green forks scored after spreading and presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. D and E indicate results from the Vindigni and Chaudhuri laboratories, respectively. N indicates the number of biological repeats (N = 3 in D and N = 3 in E). Statistics: mean ± SEM. (F) Representative images of DNA fibers obtained with the hybrid spreading approach. Scale bar is 10 µm. (G) Blue–green–blue versus blue–green forks scored after using a hybrid spreading protocol, where the DNA is extracted following the spreading approach and combed on coverslips. Data are presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. Data are from N = 3 independent experiments. Statistics: mean ± SEM. (H) Blue–green–blue versus blue–green forks scored after addition of proteinase K to hybrid spreading protocol. Data are presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. Statistics: mean ± SEM. Number of biological repeats N = 3.

Higher percentages of broken forks are detected by DNA combing compared to spreading, and this difference is reduced by addition of proteinase K to the spreading and hybrid spreading protocols. (A) Representative images of DNA fibers obtained with the combing approach. Scale bar is 10 µm. (B) Blue–green–blue versus blue–green forks scored after combing and presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. Statistics: mean ± SEM. Number of biological repeats N = 3. (C) Representative images of DNA fibers obtained with the spreading approach. Scale bar is 10 µm. (D and E) Blue–green–blue versus blue–green forks scored after spreading and presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. D and E indicate results from the Vindigni and Chaudhuri laboratories, respectively. N indicates the number of biological repeats (N = 3 in D and N = 3 in E). Statistics: mean ± SEM. (F) Representative images of DNA fibers obtained with the hybrid spreading approach. Scale bar is 10 µm. (G) Blue–green–blue versus blue–green forks scored after using a hybrid spreading protocol, where the DNA is extracted following the spreading approach and combed on coverslips. Data are presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. Data are from N = 3 independent experiments. Statistics: mean ± SEM. (H) Blue–green–blue versus blue–green forks scored after addition of proteinase K to hybrid spreading protocol. Data are presented as a percentage of the total scored pulse-labeled tracts. At least 100 tracts were scored for each sample. Numbers indicate the mean tracts %. Statistics: mean ± SEM. Number of biological repeats N = 3.

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