MilliporeSigma
  • Home
  • Search Results
  • FACS-based isolation of fixed mouse neuronal nuclei for ATAC-seq and Hi-C.

FACS-based isolation of fixed mouse neuronal nuclei for ATAC-seq and Hi-C.

STAR protocols (2021-07-27)
Ekaterina Eremenko, Anastasia Golova, Daniel Stein, Monica Einav, Ekaterina Khrameeva, Debra Toiber
ABSTRACT

The organization of chromatin structure plays a crucial role in gene expression, DNA replication, and repair. Chromatin alterations influence gene expression, and modifications could be associated with genomic instability in the cells during aging or diseases. Here, we provide a modified protocol to isolate fixed neuronal nuclei from a single mouse cortex to investigate the spatial organization of chromatin structure on a genome-wide scale by ATAC-seq (the assay for transposase-accessible chromatin with high-throughput sequencing) and chromatin conformation by Hi-C (high-throughput chromosome conformation capture).

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Milli-Mark® Anti-NeuN-PE Antibody, clone A60, clone A60, Milli-Mark®, from mouse
Sigma-Aldrich
Magnesium chloride, anhydrous, ≥98%
Sigma-Aldrich
Sucrose, for molecular biology, ≥99.5% (GC)
Sigma-Aldrich
Triton X-100, for molecular biology
Sigma-Aldrich
Goat serum
Sigma-Aldrich
TWEEN® 20, viscous liquid
Sigma-Aldrich
OptiPrep Density Gradient Medium, used for cell and subcellular organelle isolation
Sigma-Aldrich
Potassium chloride, for molecular biology, ≥99.0%
Sigma-Aldrich
Calcium chloride dihydrate, BioXtra, ≥99.0%
Sigma-Aldrich
Phosphatase Inhibitor Cocktail 2, aqueous solution (dark coloration may develop upon storage, which does not affect the activity)
Sigma-Aldrich
DL-Dithiothreitol, ≥98% (HPLC), ≥99.0% (titration)
Sigma-Aldrich
Sodium acetate, anhydrous, for molecular biology, ≥99%