Protocol for Chromatin Immunoprecipitation of Meiotic-Stage-Specific Tomato Anthers.

Abstract

Interactions occurring between DNA and proteins across the nuclear genome regulate numerous processes, including meiosis. Meiosis ensures genetic variation and balanced segregation of homologous chromosomes. It involves complex DNA-protein interactions across the entire genome to regulate a broad range of processes, including formation and repair of double-strand DNA breaks (DSBs), chromosome compaction, homolog pairing, synapsis, and homologous recombination. The latter meiotic event, meiotic recombination, often occurs at discrete locations in a genome, within a tight time window. The identification of genomic binding sites of meiotic proteins is a major step toward understanding the molecular mechanisms underlying meiotic recombination and provides important information for plant breeding. Collecting meiotic cells from plants is challenging, tedious, and time consuming, since the meiocyte-producing organs, the anthers, are generally small and limited to certain developmental stages of plants. Here we provide a protocol to isolate meiotic-stage-specific anthers and perform ChIP on this material. We have developed a ChIP protocol specifically suited to (1) small amounts of input material and (2) proteins that bind transiently to chromatin and at very low frequency. © 2018 by John Wiley & Sons, Inc.