GSM1843695: EO027_rep1_IgG; Caenorhabditis elegans; ChIP-Seq
Flash frozen larvae were physically homogenized in a mixer mill, fixed in 1 % formaldehyde for 10 minutes, quenched with 50 mM glycin for 5 min, sheared by sonication in a Bioruptor, and the resulting chromatin was extracted. Immunoprecipitation was performed using either 5 mg a-ELT-2 (455-2A4) antibody, 5 mg a-H3K4me3 antibody (WAKO 305-34819), or mock conditions (IgG only). The anti-ELT-2 monoclonal antibodies was produce by the Southern Alberta Cancer Research Institute Antibody Services by using a purified poly-histidine tagged full-length ELT-2 protein produced in E. coli as an antigen to produce anti-ELT-2 monoclonal antibody 455-2A4 (isotype IgG1). Antibodies concentrated from hybridoma supernatants concentrated ~10-fold by pressure filtration, dialyzed extensively against PBS and then concentrated a further 2-3 fold by dialysis against PBS-50 % glycerol. ChIP-recovered DNA and input DNA were ligated to sequencing adapters using containing an eight-base multiplexing barcode sequence. DNA libraries were sequenced on either the Hi-Seq2000 or Hi-Seq2500 Illumina sequencers with 50 rounds of single-end sequencing, 8 cycles of which were used for multiplex barcode indices.