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Analysis of DNA Double-Strand Break End Resection and Single-Strand Annealing in S. pombe

  • Zhenxin Yan
  • Sandeep Kumar
  • Grzegorz IraEmail author
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Part of the Methods in Molecular Biology book series (MIMB, volume 2153)

Abstract

DNA double-strand break (DSB) end resection is an essential step for homologous recombination. It generates 3′ single-stranded DNA needed for the loading of the strand exchange proteins and DNA damage checkpoint proteins. To study the mechanism of end resection in fission yeast, we apply a robust, quantitative and inducible assay. Resection is followed at a single per genome DSB synchronously generated by the tet-inducible I-PpoI endonuclease. An additional assay to follow resection involves recombination between two direct repeats by single-strand annealing (SSA), since SSA requires extensive resection to expose two single-strand repeats for annealing. The kinetics of resection and SSA repair are then measured using Southern blots.

Key words

DNA double-strand break End resection Single-strand annealing Southern blotting 

Notes

Acknowledgements

This work was supported by NIH grants GM080600 and GM125650.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2021

Authors and Affiliations

  1. 1.Department of Molecular and Human GeneticsBaylor College of MedicineHoustonUSA

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