Thursday, March 4, 2010

ChIP-Seq used to examine diverse roles of transcription factors

Zhong et al. (Yale University)* developed an experimental pipeline in C. elegans  to identify transcription factor binding sites, using chromatin-immunoprecipitation and deep sequencing (ChIP-Seq). They studied the tissue-specific transcription factor PHA-4 and found distinct sets of PHA-4 targets under conditions of embryonic development and environmental stress (starvation).

The researchers used a GFP-tagged fosmid clone, constructed using the pCC1FOS Vector, to create transgenic worms expressing PHA-4. For starvation assays using RNAi, transcripts were prepared using the AmpliScribe™ T7 High Yield Transcription Kit. The DNA isolated after ChIP was treated with the End-It™ DNA End-Repair Kit before A-tailing, adaptor ligation, and sequencing.
*Zhong M, et al. (2010). Genome-wide identification of binding sites defines distinct functions for Caenorhabditis elegans PHA-4/FOXA in development and environmental response. PLoS genetics, 6 (2) PMID: 20174564

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