UNC-55 Combined (GFP ChIP)

Identification of Transcription Factor RPC-1::RPC-1 Binding Regions in Young Adults (Snyder project, Snyder subgroup)

Synchronized L2 larvae from C. elegans strain NC1369 (a transgenic strain engineered to express a gene fusion between unc-55 and GFP) were treated with the cross-linking reagent formaldehyde. After lysis and sonication, the chromatin was immunoprecipitated with an affinity-purified antibody that recognizes GFP. The bound DNA was purified and sequenced in an Illumina GA-2. A sample of the input DNA was sequenced in parallel. The ChIP-seq data generated by this experiment was analyzed using the PeakSeq peak-calling algorithm to predict protein binding sites in the C. elegans genome.

Details

General Description

We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals. Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology.

Protocols

  1. Growth and isolation: Plated Worm Growth and Harvest
  2. Sample preparation: ChIP, Illumina Deep Sequencing
  3. Data Analysis: Illumina Data Merging
  4. Other Protocols: Illumina Data Analysis, ChIP-seq replicate verification, Peak Calling

Experimental Reagents

  1. Antibodies: eGFP

Sample Details

  1. Animals/Lines: Caenorhabditis elegans


Release Date: 2013-04-24