Chromatin Proteins in Oregon strainChromatin-binding Proteins in Oregon strain (Karpen project, Elgin subgroup)Nuclei were isolated from staged Drosophila Oregon R embryos, 14-16 hr after egg laying that had been crosslinked with formaldehyde. After lysis and sonication the chromatin was immunoprecipitated with an affinity-purified antibody that recognizes chromosomal protein HP1. 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 DescriptionWe aim to determine the locations of 125 chromosomal proteins acrossthe Drosophila melanogaster genome. The proteins 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
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Release Date: 2011-05-12 |