H3 Histone Modifications in BG3 cells

H3K36me2_W.BG3.Solexa (Karpen project, Kuroda subgroup)

Nuclei were isolated from the Drosophila BG3 neuronal cell line that had been crosslinked with formaldehyde. After lysis and sonication the chromatin was immunoprecipitated with an affinity-purified antibody that recognizes histone H3 dimethylated on lysine 36. The resulting fragments were used to create libraries and sequenced on the Illumina platform. Short reads were aligned to the reference genome using Bowtie, and peaks for each replicate were called using SPP (using sequence from the Input material as a comparison control). Statistically significant regions of enrichment from the two biological replicates were then combined and used to call the cumulative peaks.

Details

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 the Illumina
NGS sequencing platform. 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: Crosslinking of cultured cells, Chromatin prep from cultured cells
  2. Sample preparation: ChIP, Illumina sequencing
  3. Data Analysis: Bowtie alignment
  4. Other Protocols: SPP Peak calling

Experimental Reagents

  1. Antibodies: H3K36me2

Sample Details

  1. Animals/Lines: Drosophila melanogaster


Release Date: 2013-03-19