GSM4905384: H2B8_sperm_input_rep1; Arabidopsis thaliana; ChIP-Seq
Pollen was collected in Galbraith buffer (45 mM MgCl2, 30 mM Sodium Citrate, 20 mM MOPS, 1% Triton-100, pH 7.0) from open flowers of a H2B.8p::H2B.8-eGFP line. Nuclei were released from pollen by vortexing and subsequent debris was removed by filtration. Chromatin was fragmented by micrococcal nuclease (MNase, New England Biolabs) digestion. For IP samples, fragmented chromatin was immunoprecipitated with GFP-Trap beads. For input, no immunoprecipitation was undertaken. Protein and RNA was removed by digestion with Proteinase K and RNase A, respectively. Phenol-chloroform extraction was used to obtain DNA. Libraries were prepared using the Ovation Ultralow System V2 (Nugen, 0344) and sequenced on the NextSeq 500 (Illumina) with 2 × 38 bp paired end reads at the John Innes Centre, Norwich, UK.