ChIP-Seq: Histone Modifications in L3

L3 larvae H3K4Me3 ChIP-seq (White project)

Details

Staged Drosophila (y; bw cn sp) L3 larvae were crushed and cross-linked with formaldehyde. After lysis and sonication, the chromatin was immunoprecipitated with antibodies recognizing histone H3 trimethylated on lysine 4. Input and ChIP'd DNA was single-end sequenced in two lanes each on the Illumina Genome Analyzer, aligned with Eland. Density ratios were calculated with the R SPP package. MACS software was used to call peaks.

Series 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: L3 collection histone timecourse:KW:1
  2. Sample preparation: White_Lab/IP, Solexa ChIP-seq:KW:1
  3. Data Analysis: Solexa ChIP-seq Analysis:KW:1, ChIP-Seq Peak-calling:KW:1, ChIP-Seq Peak-calling HMMSeg:KW:1
  4. Other Protocols: Formaldehyde crosslinking of chromatin

Experimental Reagents

  1. Antibodies: H3K9Ac, H3K4me1, H3K27Ac, H3K27me3, H3K4me3

Sample Details

  1. Animals/Lines: L3, 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: GSM400661, GSM401418, GSM401417, GSM400661, GSM400660, GSM401416, GSM439440


Release Date: 2010-01-12 Track 797