Endosperm-specific chromatin profiling by fluorescence-activated nuclei sorting and ChIP-on-chip.
Cell-type-specific analysis of gene expression and chromatin profiling requires the isolation of discrete cell populations from complex pools. However, until now this most critical step has been labor intensive and technical challenging. Here, we describe a rapid protocol based on fluorescence-activated cell sorting (FACS) for cell-type-specific RNA and chromatin profiling. We detail how to isolate nuclei from Arabidopsis inflorescence and silique homogenates and how to purify endosperm nuclei labeled by nuclear-targeted green fluorescent protein using FACS. The purified fluorescent endosperm nuclei can be further used for chromatin immunoprecipitation (ChIP) followed by hybridization to high-resolution whole-genome tiling microarrays (ChIP-on-chip) or transcriptional profiling.
Department of Biology and Zurich-Basel Plant Science Center, Swiss Federal Institute of Technology, ETH Centre, Zurich, Switzerland.
Isabelle
I
Weinhofer
Claudia
C
Köhler
D002467:Cell Nucleus / Q000235:genetics*; D002843:Chromatin / Q000235:genetics*; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D056625:Endosperm / Q000166:cytology*; D005434:Flow Cytometry / Q000379:methods*; D020869:Gene Expression Profiling; D016133:Polymerase Chain Reaction; D046888:Tissue Array Analysis / Q000379:methods*
D002843:Chromatin