Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of mouse ck-OBF-1 transgene

ABSTRACT: The Lck-OBF-1 construct used to generate transgenic mice contains an N-terminally HA epitope-tagged human OBF-1 cDNA under the control of the murine proximal lck promoter (-3100 to +23 relative to the transcription start site) that was purified by electroelution. Transgenic mouse lines were obtained and bred in B6CF1 x C57BL/6 background. For the analysis, age- and sex-matched littermates were used. The mice were kept in a standard mouse facility. Mice were sacrificed, the thymi were taken, rinsed in PBS and immediately homogenized in TRIZOL reagent (Life Technologies). Total RNA was prepared using TRIZOL reagent and purified on RNeasy Miniprep columns (Qiagen) according to the manufacturers' instructions. Array set (A1): Equal amounts of RNA from four young adult male mice (6.5 weeks old) per genotype were purified as stated above and pooled in equal amounts. Array set (A2): Equal amounts of total RNA from four young adult mice (6.5 weeks old; 2 males and 2 females each time) per genotype were purified as stated above and pooled in equal amounts. Both the WT and the transgenic pool were hybridized to Affymetrix GeneChips in duplicate. Array set (B): RNA was prepared from sorted thymic cell populations (CD4+ CD8+ CD25- and CD4+ CD8+ CD25+) from a pool of 15 young adult (mixed gender, 6 to 9 weeks of age) transgenic mice as stated above. Cell sorting was performed on a MoFlo (DakoCytomation) with a purity of at least 94%.

ORGANISM(S): Mus musculus

SUBMITTER: Edward Oakeley

PROVIDER: E-MEXP-85 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

The Ets factor Spi-B is a direct critical target of the coactivator OBF-1.

Bartholdy Boris B Du Roure Camille C Bordon Alain A Emslie Dianne D Corcoran Lynn M LM Matthias Patrick P

Proceedings of the National Academy of Sciences of the United States of America 20060721 31

OBF-1 (Bob.1, OCA-B) is a lymphoid-specific transcriptional coactivator that associates with the transcription factors Oct-1 or Oct-2 on the conserved octamer element present in the promoters of several ubiquitous and lymphoid-specific genes. OBF-1-deficient mice have B cell-intrinsic defects, lack germinal centers, and have severely impaired immune responses to T cell-dependent antigens. Crucial genes that are regulated by OBF-1 and that might explain the observed phenotype of OBF-1 deficiency ...[more]

PMID: 16861304

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Project description:4C-Seq has proven to be a powerful technique to identify genome-wide interactions with a single locus of interest (or "bait") that can be important for gene regulation. However, analysis of 4C-Seq data is complicated by the many biases inherent to the technique. An important consideration when dealing with 4C-Seq data is the differences in resolution of signal across the genome that result from differences in 3D distance separation from the bait. This leads to the highest signal in the region immediately surrounding the bait and increasingly lower signals in far-cis and trans. Another important aspect of 4C-Seq experiments is the resolution, which is greatly influenced by the choice of restriction enzyme and the frequency at which it can cut the genome. Thus, it is important that a 4C-Seq analysis method is flexible enough to analyze data generated using different enzymes and to identify interactions across the entire genome. Current methods for 4C-Seq analysis only identify interactions in regions near the bait or in regions located in far-cis and trans, but no method comprehensively analyzes 4C signals of different length scales. In addition, some methods also fail in experiments where chromatin fragments are generated using frequent cutter restriction enzymes. Here, we describe 4C-ker, a Hidden-Markov Model based pipeline that identifies regions throughout the genome that interact with the 4C bait locus. In addition, we incorporate methods for the identification of differential interactions in multiple 4C-seq datasets collected from different genotypes or experimental conditions. Adaptive window sizes are used to correct for differences in signal coverage in near-bait regions, far-cis and trans chromosomes. Using several datasets, we demonstrate that 4C-ker outperforms all existing 4C-Seq pipelines in its ability to reproducibly identify interaction domains at all genomic ranges with different resolution enzymes. 4C-Seq experiments from Igh and Cd83 bait in activated B cells and Tcrb (Eb) bait in double negative T cells and immature B cells. RNA-Seq and ATAC-Seq experiments in DN and Immature B cells.

Dataset Information

Transcription profiling of mouse ck-OBF-1 transgene

Publications

The Ets factor Spi-B is a direct critical target of the coactivator OBF-1.

Similar Datasets

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