Project description:Estrogen Receptor alpha (ERα) is a key driver of most breast cancers, and it is the target of endocrine therapies used in the clinic to treat women with ERα positive (ER+) breast cancer. The two methods ChIP-seq (chromatin immunoprecipitation coupled with deep sequencing) and RIME (Rapid Immunoprecipitation of Endogenous Proteins) have greatly improved our understanding of ERα function during breast cancer progression and in response to anti-estrogens. A critical component of both ChIP-seq and RIME protocols is the antibody that is used to pull down the bait protein. To date, most of the ChIP-seq and RIME experiments for the study of ERα have been performed using the sc-543 antibody from Santa Cruz Biotechnology. However, this antibody has been discontinued, thereby severely impacting the study of ERα in normal physiology as well as diseases such as breast cancer and ovarian cancer. Here, we compare the sc-543 antibody with other commercially available antibodies, and we show that 06-935 (EMD Millipore) and ab3575 (Abcam) antibodies can successfully replace the sc-543 antibody for ChIP-seq and RIME experiments.
Project description:We performed chromatin immunoprecipitation (ChIP) with two polyclonal anti-ERalpha antibodies directed against the N- and C-terminus of the protein, with or without estrogen treatment for 45 minutes of MCF-7 cells, and the immunoprecipitated DNA was sequenced with the Illumina/Solexa Genome Analyzer.
Project description:This study contains a series of ChIP-seq experiments performed on nuclear material similar to what was used in Tissue- and Stage-specific Capture-C for selected TSS and CRM (BioStudies/ArrayExpress collection E-MTAB-9310). ChIP-seq with chromatin isolated from nuclei isolated from 2-3h whole embryos samples or Mef2-/Elav-sorted nuclei from 6-8h and 10-12h embryo samples was performed using anti-H3K27ac (Anti-Histone H3 (acetyl K27) antibody, Abcam #ab4729, purified polyclonal), anti-CTCF (rabbit anti Drosophila CTCF antibody, Reinkawitz lab, purified polyclonal), anti-Beaf (mouse anti Drosophila BEAF antibody, DSHB #1553420, monoclonal) and anti-Su(Hw) (goat anti Drosophila Su(Hw) antibody, Geyer lab, purified polyclonal) antibodies.
Project description:ChIP on chip assay of AbrC3 to determine its binding sites in S. coelicolor M145 genome using purified anti-AbrC3 polyclonal antibodies. Samples from nutrient broth (NB) cultures were collected at 36, 48 and 60 hours, cross-linked, and finally hybridized to DNA 104K microarrays designed by University of Surrey.
Project description:The WhiG sigma factor gene is required for spore formation is Streptomyces venezuelae. It is similar to the FliA sigma factor of E. coli. WhiG deletion strains are able to make aerial hyphae but are defective in the spore maturation. This ChIP-Seq experiment was carried out to determine all the binding sites WhiG binds to in the genome of Streptomyces venezuelae. Anti-WhiG polyclonal antibodies were used for ChIP-Seq of the wild type (WT) strain after 34 hours of growth in shaken cultures. A WhiG deletion strain was made and anti-WhiG antibodies were used for ChIP-Seq in the deletion strain after 34 hours of growth in shaken cultures. This was used as the negative control and ChIP-Seq peak positions in this were disregarded in the WT.
Project description:ChIP-on-Chip experiment using chromatin from the human adult ovarian granulosa cell tumor derived cell line KGN (Nishi et al, 2001), or from a KGN subclone overexpressing WT FOXL2 (KF3 subclone), and an isovolumic blend of our custom anti-FOXL2 polyclonal antibodies (Cocquet J et al, 2002). Non precipitated sheared matched deproteinized chromatin (Input) was used a control to estimate enrichment peaks (and thus FOXL2 binding sites) from IPed DNA. DNA from three independent ChIP assays (Input extractions) was pooled, and 100ng of DNA was linearly amplified using the Whole Genome Amplification kit (Sigma). The two ChIP-chip experiments serve as biological replicates. Moreover, 42 out of 48 targets promoters chosen from the âenrichment peakâ list were subsequently confirmed as enrichment sites in anti-FOXL2 ChIPed DNA for subsequent experiments.
Project description:ChIP-on-Chip experiment using chromatin from wild-type 5 weeks old swiss mice ovaries (Nishi et al, 2001) and an isovolumic blend of our custom anti-Foxl2 polyclonal antibodies (Cocquet J et al, 2002). Non precipitated sheared matched deproteinized chromatin (Input) was used a control to estimate genomic enrichment peaks (and thus Foxl2 binding sites) from IPed DNA. DNA from three independent ChIP assays (Input extractions) was pooled, and 100ng of DNA was linearly amplified using the Whole Genome Amplification kit (Sigma).
Project description:To determine the influence of BldM and WhiI on the binding of each other to their respective binding sites in the genome of Streptomyces venezuelae, ChIP-Seq experiments were carried out using polyclonal antibodies against the wild type proteins as well as anti-FLAG antibodies against FLAG-tagged proteins. Null mutants of Streptomyces venezuelae lacking WhiI or BldM were used as controls. The wild tpe strain was used as control in experiments using the anti-FLAG antibodies.
Project description:We performed ChIP-seq ananlysis on HDAC1 in MiaPaCa2 using two different antibodies (Santa Cruz Biotechnologies sc-81598 HDAC1(10E2) and Invitrogen HDAC1 Polyclonal Antibody PA1-860), two non-targeting scrableled sgRNA controls and two replicates for each.
