ChIP-Seq analysis on FACS-enriched nuclei – an alternative approach for detecting genome-wide meiotic recombination hotspots

Authors

• KG Lam
• F Pratto
• K Brick
• RD Camerini-Otero

Abstract

Meiotic DNA double strand breaks (DSBs) are clustered at hotspots across the genome. These hotspots have been mapped in mouse and human spermatocytes, by an approach of chromatin immunoprecipitation (ChIP) with antibodies to DMC1 protein followed by single-stranded DNA sequencing (SSDS). DMC1 proteins bind specifically to single-stranded DNA (ssDNA) resected from meiotic DSBs. Thus, mapping meiosis-specific ssDNA identifies the locations of DSBs. Since DSB-stage spermatocytes are rare (50-fold to a homogeneous nucleus population, reveals that specific ssDNA signals can be effectively detected without removing non-specific dsDNA. This method should be specially useful for experiments limited by the amount of starting material and for ChIP-Seq analyses on proteins that bind to both ssDNA and dsDNA molecules.

Scientific Focus Area: Genetics and Genomics

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