E-MTAB-4585 - Time course Pho/dSfmbt Chromatin Immunoprecipitation on Drosophila melanogaster embryos during embryogenesis
Last updated on 11 April 2017, released on 11 April 2017
Here we improved BiTS-ChIP (Bonn et al, Nature Protocols 7, 978-994 (2012)) to identify active enhancer and promoter elements genome wide in the 104 cardiomyocytes that constitute the Drosophila heart tube and represents only ~0.5% of the total cell content of the embryo. A transgenic Drosophila strain expressing nuclear GFP under the control of a cardiac specific enhancer (TinC*>GFP) was used for staged embryo collections at stages 13-14 (10-13h of development). After embryo fixation and dissociation, intact fixed nuclei were fluorescent labelling. Purification of this rare nuclear population was achieved by a two-step sorting procedure, yielding ~98% purity. Chromatin was extracted and used for immunoprecipitation and sequencing (ChIP-seq) to analyze chromatin modifications at promoters (H3K4me3 and H3K27ac) and enhancers (H3K27ac). Two independent biological replicates (from FACS sorting, chromatin preparations and ChIP-Seq) were performed for each mark and sequenced using Illumina HiSeq.
ChIP-seq, binding site identification design, development or differentiation design, reference design, replicate design