H4 Histone modifications in CME W1 cl.8+ cells

H4 Histone modifications in CME W1 cl.8+ cells (Karpen project, Kuroda subgroup)

Nuclei were isolated from the Drosophila CME-W1-Cl.8+ wing imaginal disc cell line that had been crosslinked with formaldehyde. After lysis and sonication the chromatin was immunoprecipitated with an affinity-purified antibody that recognizes histone H4 acetylated on lysine 8. After PCR amplification and labeling the DNA was used to probe Affymetrix microarrays (a sample of the DNA input for the ChIP was processed in parallel and used to probe another copy of the array). The experiment was performed in triplicate, and normalized, averaged data is shown. For tracks displaying binding peaks, normalized signal intensities from three averaged experimental replicates were analyzed using a peak-calling algorithm that calculates genomic regions of significant enrichment (or depletion).

General Description

We aim to determine the locations of the major histone modifications
across the Drosophila melanogaster genome. The modifications under
study are involved in basic chromosomal functions such as DNA
replication, gene expression, gene silencing, and inheritance. We will
perform Chromatin ImmunoPrecipitation (ChIP) using genomic tiling
arrays. We will initially assay localizations using chromatin from
three cell lines and two embryonic stages, and will then extend the
analysis of a subset of proteins to four additional animal tissues/stages.

Protocols

  1. Growth and isolation: Chromatin prep from cultured cells, Crosslinking of cultured cells
  2. Sample preparation: Hybridization to Affy arrays, WGA, ChIP, Array Scanning Protocol
  3. Data Analysis: Smoothed M-value enrichment profiles, M-value normalization, Regions of significant enrichment, Smoothed M-value enrichment profiles
  4. Other Protocols: Regions of significant enrichment

Experimental Reagents

  1. Antibodies: H4K16ac(L)

Sample Details

  1. Animals/Lines: CME W1 Cl.8+

  1. Arrays: Affymetrix Drosophila Tiling Arrays v2.0R


Release Date: 2011-04-26