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Methods Enzymol. 2018;601:391-418. doi: 10.1016/bs.mie.2017.11.037. Epub 2018 Feb 26.

Analysis of Meiotic Double-Strand Break Initiation in Mammals.

Author information

1
Genetics & Biochemistry Branch, NIDDK, National Institutes of Health, Bethesda, MD, United States.
2
Uniformed Services University of the Health Sciences, Bethesda, MD, United States.
3
Genetics & Biochemistry Branch, NIDDK, National Institutes of Health, Bethesda, MD, United States. Electronic address: rc10d@nih.gov.
4
Uniformed Services University of the Health Sciences, Bethesda, MD, United States. Electronic address: galina.petukhova@usuhs.edu.

Abstract

The repair of programmed DNA double-strand breaks (DSBs) physically tethers homologous chromosomes in meiosis to allow for accurate segregation through meiotic cell divisions. This process, known as recombination, also results in the exchange of alleles between parental chromosomes and contributes to genetic diversity. In mammals, meiotic DSBs occur predominantly in a small fraction of the genome, at sites known as hotspots. Studies of the formation and repair of meiotic DSBs in mammals are challenging, because few cells undergo meiotic DSB formation at a given time. To better understand the initiation and control of meiotic recombination in mammals, we have devised a highly sensitive method to map the sites of meiotic DSBs genome wide. Our method first isolates DNA bound to DSB repair proteins and then specifically sequences the associated single-stranded DNA. This protocol has generated the first meiotic DSB maps in several mammals and the only map of meiotic DSBs in humans.

KEYWORDS:

DSB; Double-strand break; Genomics; Hotspot; Meiosis; Recombination; Single-stranded DNA sequencing

PMID:
29523240
PMCID:
PMC6463303
DOI:
10.1016/bs.mie.2017.11.037
[Indexed for MEDLINE]
Free PMC Article

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