Proteomic profiling of cardiac tissue by isolation of nuclei tagged in specific cell types (INTACT)

摘要: The proper dissection of the molecular mechanisms governing specification and differentiation specific cell types requires isolation pure populations from heterogeneous tissues whole organisms. Here, we describe a method for purification nuclei defined or tissue in vertebrate embryos using INTACT (isolation tagged types). This method, previously developed plants, flies worms, utilizes vivo tagging nuclear envelope with biotin subsequent affinity labeled nuclei. In this study successfully purified cardiac skeletal muscle Xenopus strategy. We went on to demonstrate utility approach by coupling liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomic methodologies profile proteins expressed developing hearts. From these studies have identified orthologs 12 human encoded genes, which when mutated lead congenital heart disease. Thus, combining technologies are able identify tissue-specific that required normal organ development.