ChIP-chip: Histone Modifications in L2

L2 larvae H3K9me3 ChIP-chip (White project)

Details

Nuclei were isolated from staged Drosophila (y; bw cn sp) L2 larvae, and cross-linked with formaldehyde. After lysis and sonication the chromatin was immunoprecipitated with affinity-purified antibodies recognizing histone H3 trimethylated on lysine 9. The DNA was amplified, labeled, and hybridized to Agilent microarrays (mock IP DNA was labelled in parallel and hybridized simultaneously to the same chip). The Joint Binding Deconvolution algorithm was used to predict binding peaks in normalized data from three experimental replicates.

General Description

The White Lab is aiming to map the association of all the Transcription Factors (TF) and DNA associated proteins on the genome of Drosophila melanogaster. The main technique that will be used for this purpose is chromatin immunoprecipitation-on-chip (ChIP-on-chip) utilizing whole-genome tiling arrays. The data generated by ChIP-chip experiments consist basically of a plot of signal intensity across the genome. The highest signals correspond to positions in the genome occupied by the tested TF.
ChIP-seq, meaning direct sequencing of the ChIP sample on the Solexa platform, has also been used to validate some datasets and will become- as the project advances- the main production platform.

To support ChIP-chip and ChIP-seq datasets, and in order to link association of factors to the DNA with regulation of transcription, several RNA-seq (direct sequencing of the transcriptome on the Solexa platform) datasets will be genrated for the time-points under scrutiny.

Protocols

  1. Growth and isolation: L2 collection histone timecourse:KW:1
  2. Sample preparation: White_Lab/IP, Whole Genome Amplification:KW:1, Agilent 244K ChIP Labeling:KW:1, Array Hybridization for ChIP-chip:KW:1, Array Scanning
  3. Data Analysis: Agilent ChIP data processing:KW:1, Agilent ChIP peak finding:KW:1
  4. Other Protocols: Cross-linking

Experimental Reagents

  1. Antibodies: H3K9me3, No Antibody Control
  2. Arrays: Agilent Drosophila tiling array 1 of 3, Agilent Drosophila tiling array 2 of 3, Agilent Drosophila tiling array 3 of 3

Sample Details

  1. Animals/Lines: L2, Drosophila melanogaster, y[1]; Gr22b[1] Gr22d[1] cn[1] CG33964[R4.2] bw[1] sp[1]; LysC[1] MstProx[1] GstD5[1] Rh6[1]
  2. External Links: GSM384760, GSM384761, GSM384762, GSM384763, GSM384764, GSM384765, GSM384766, GSM384767, GSM384768


Release Date: 2009-04-08