Role of Hopx in myogenesis and commitment [ChIP-Seq]

Hopx is expressed in cardiomyoblasts, progenitors commited to the myocyte fate. ChIP-seq experiements were performed to understand Hopx occupancy in E9.5 hearts. An epitope tagged allele, in which a 3X-Flag sequence is appeneded to the endogenous Hopx transcript. Our results suggest that Hopx occupancy is enriched in regions close to Wnt family members, confirmed by gene ontology analysis and indepedent expression analysis. E9.5 hearts were microdissected and collected from Hopx3XFlag/+ embryos. Approximately 30-40 (5-6 litters) crosslinked hearts were pooled and sheered. ~10 ug of sheered chromatin was used in ChIP experiments. Each experiment contains a set of ChIP DNA product: input (background control) and IP as the Hopx binding product. Background noise was subtracted and signal was used in analyses. Since Hopx is a cofactor, and does not bind DNA directly, one sample included crosslinking in the presence of EGS (Ethylene glycol bis[succinimidylsuccinate]) in addition to formaldehdye, while the other replicate only included crosslinking in the presence of formaldehyde only.