Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

ModENCODE_Lieb and White Lab: unltra-deep sequencing data of genomic DNA, chromatin input, and ChIP of Su(Hw) and H3K36me3 from S2 cells on Illumina Genome Analyzer.

ABSTRACT: This is a dataset generated by the modENCODE Project. It contains both the pair-end and single-end sequencing data of genomic DNA, chromatin input and the ChIP of the factors Su(Hw) and H3K36me3 from Drosophila S2 cells and it was generated on Illumina Genome Analyzer. The goal was to sequence ~120M tags of each sample and used this ultra-deep dataset to systematically evaluate both experimental and computational factors that influence the results of ChIP-seq experiments. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf S2 cells from the modENCODE batch were amplified before transferring to the plates. 15 plates were put in culture until the cells reached the appropriate concentration. Cells were harvested from 2 plates to extract the genomic DNA. The remaining 13 plates were then treated with formaldehyde at the same time and the chromatin were extracted. 3 plates were used to produce an Input sample corresponding to the chromatin DNA of the treated cells. Each plate corresponding to an eppendorff tube of chromatin, ChIP experiments were performed on 5 tubes for H3K36me3 and on 5 tubes for Su(Hw). ChIP and DNA extraction were performed independently on each tube. After the purification of the DNA, the tubes corresponding to the same sample (Su(Hw), H3K36me3, Input and gDNA) were pooled together. After mixing, each sample was aliquoted again into 5 tubes. 1 aliquot of each was used for ChIP-chip on Affymetrix tiling arrays for quality control, 1 aliquot was used for single-end sequencing at the University of Chicago HGAC, 1 aliquot was used for single-end sequencing at UNC, 1 aliquot was used for paired-end sequencing at UNC and the remaining aliquot was kept as a back-up of the original samples. For each sample, >100M single-end tags were produced (~5 runs), along with the tags from 1 lane of paired-end sequencing experiment. Besides several experimental factors, the performance of multiple peak callers were evaluated at different sequencing depths.

ORGANISM(S): Drosophila melanogaster

SUBMITTER: Yiwen Chen

PROVIDER: E-GEOD-27679 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Systematic evaluation of factors influencing ChIP-seq fidelity.

Chen Yiwen Y Negre Nicolas N Li Qunhua Q Mieczkowska Joanna O JO Slattery Matthew M Liu Tao T Zhang Yong Y Kim Tae-Kyung TK He Housheng Hansen HH Zieba Jennifer J Ruan Yijun Y Bickel Peter J PJ Myers Richard M RM Wold Barbara J BJ White Kevin P KP Lieb Jason D JD Liu X Shirley XS

Nature methods 20120422 6

We evaluated how variations in sequencing depth and other parameters influence interpretation of chromatin immunoprecipitation-sequencing (ChIP-seq) experiments. Using Drosophila melanogaster S2 cells, we generated ChIP-seq data sets for a site-specific transcription factor (Suppressor of Hairy-wing) and a histone modification (H3K36me3). We detected a chromatin-state bias: open chromatin regions yielded higher coverage, which led to false positives if not corrected. This bias had a greater effe ...[more]

PMID: 22522655

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Project description:Insulators delimit independent transcriptional domains within genomes by constraining enhancer and silencer action. These transcriptional effects depend upon DNA recognition by insulator binding proteins that recruit partners that protect against inappropriate long range modulation of non-target promoters. Insulator binding proteins are broadly expressed during development, with largely constitutive binding to thousands of genomic sites. Yet, tissue-specific transcriptional changes result from the loss of individual insulator binding proteins. To understand the molecular basis for such effects, we are studying the classic Drosophila insulator protein Suppressor of Hairy-wing [Su(Hw)]. Genetic studies show that loss of this broadly expressed insulator protein prevents oocyte development. To determine the basis for the block in oogenesis, we coupled transcriptional analyses in su(Hw) mutant ovaries with genome-wide definition of Su(Hw) binding in this tissue. These studies identified 71 direct targets of Su(Hw) regulation, with nearly 70% of these genes showing increased RNA accumulation when Su(Hw) is lost. Surprisingly, derepressed Su(Hw) target genes correspond to genes normally highly expressed in neural tissues, suggesting that Su(Hw) has a critical role in silencing neural genes in the ovary. Support for this postulate was obtained by genetic studies. We found that oocyte production was restored in su(Hw) mutant females that carry a deletion of one allele of the elav family RNA binding protein 9 (Rbp9) gene. These su(Hw) null oocytes can be fertilized, with evidence that embryos lacking Su(Hw) show compromised development. Our studies extend the known transcriptional activities of Su(Hw), indicating that Su(Hw) can function as an insulator, activator and repressor, the latter function being essential for oogenesis. These findings highlight that insulator proteins are versatile transcriptional regulatory proteins, suggesting that tissue specific contributions to transcription result from direct regulation of individual genes.

Dataset Information

ModENCODE_Lieb and White Lab: unltra-deep sequencing data of genomic DNA, chromatin input, and ChIP of Su(Hw) and H3K36me3 from S2 cells on Illumina Genome Analyzer.

Publications

Systematic evaluation of factors influencing ChIP-seq fidelity.

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