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Sandmann, T., Jakobsen, J.S., Furlong, E.E. (2006). ChIP-on-chip protocol for genome-wide analysis of transcription factor binding in Drosophila melanogaster embryos.  Nat. Protoc. 1(6): 2839--2855.
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FBrf0202481
Publication Type
Research paper
Abstract

This protocol describes a method to detect in vivo associations between proteins and DNA in developing Drosophila embryos. It combines formaldehyde crosslinking and immunoprecipitation of protein-bound sequences with genome-wide analysis using microarrays. After crosslinking, nuclei are enriched using differential centrifugation and the chromatin is sheared by sonication. Antibodies specifically recognizing wild-type protein or, alternatively, a genetically encoded epitope tag are used to enrich for specifically bound DNA sequences. After purification and polymerase chain reaction-based amplification, the samples are fluorescently labeled and hybridized to genomic tiling microarrays. This protocol has been successfully used to study different tissue-specific transcription factors, and is generally applicable to in vivo analysis of any DNA-binding proteins in Drosophila embryos. The full protocol, including the collection of embryos and the collection of raw microarray data, can be completed within 10 days.

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    Language of Publication
    English
    Additional Languages of Abstract
    Parent Publication
    Publication Type
    Journal
    Abbreviation
    Nat. Protoc.
    Title
    Nature Protocols
    Publication Year
    2006--
    ISBN/ISSN
    1754-2189 1750-2799
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