Cited from Cell Type-Specific Gene Expression Profiling Using Fluorescence-Activated Nuclear Sorting
Fluorescence-activated cell sorting (FACS) is a powerful method for the
analysis of cell type-specific transcriptome profiles, DNA or histone
modifications, and chemical compounds.
However, many tissues are recalcitrant to cell separation and are
therefore not readily accessible for FACS analysis. Here, we lay out a
detailed protocol for the generation of transcriptional profiles from
fluorescently labeled nuclei.
Here, the nuclei of cells of interest are labeled by transgenic
expression of nuclear-localized fluorescent protein constructs and are
separated from surrounding unlabeled nuclei from a crude extract by a
cell sorter.
Most techniques of nuclear sorting use some kind of tissue fixation. The
isolated polyadenylated RNA therefore most likely is composed of newly
transcribed mRNA about to be exported from the nucleus as well as
cytoplasmic RNA that was cross-linked to the nuclear envelope.
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