ChIP-seq Analysis of Histone Modifications

Snake forebrain, anterior trunk and posterior trunk samples as well as mouse anterior trunk samples were dissected and fixed in 1% formaldehyde for 10 min. For each ChIP-seq experiment approximately 100 ng of tissue were used and processed according to (Lee et al., 2006 (link)) or the ChIP-IT High Sensitivity (Active motif) specifications. H3K27me3 antibody (Millipore, 17–622), H3K27ac antibody (Abcam ab4729) and H3K9me3 (Abcam ab8898) were used. Sequencing was performed using 100 bp single-end reads in the Illumina HiSeq system according to manufacturer’s instructions. The reads obtained from the sequencing were mapped to ENSEMBL Mouse assembly NCBIM37 (mm9) or to the corn snake scaffold using the HTSstation mapping pipeline (http://htsstation.epfl.ch) (David et al., 2014 (link)). All ChIP-seq mappings were normalized to total input chromatin using the bamCompare software from the deepTools Galaxy web server (http://deeptools.ie-freiburg.mpg.de) (Ramirez et al., 2014 (link)). Peak calling was done using MACS (Zhang et al., 2008 (link)).