Amplicon SMF for Pol II occupancy project
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Illumina MiSeq
ORGANISM(S): Mus musculus
SUBMITTER:
PROVIDER: E-MTAB-14461 | biostudies-arrayexpress |
SECONDARY ACCESSION(S): ERP164179
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:To quantify the contribution of specific transcription factor binding versus chromatin context in chromatin opening, we inserted libraries containing hundreds of CREs into a landing pad within a neutral chromatin environment, devoid of activating or repressive chromatin modifications, in mESCs using Recombination-Mediated Cassette Exchange (RMCE). Chromatin accessibility of the inserted fragments was then profiled by targeted SMF using PCR primers that anneal to a synthetic flanking sequence, allowing unambiguous differentiation of the ectopic site from its endogenous counterpart. The dataset generated includes amplicon-based SMF data from the ectopic site under steady-state conditions, in mouse cells (i.e., TC-1 knock-out of the three DNA methyl transferases (DNMT TKO) mESCs). Two biological replicates were generated. In summary, cells were collected for SMF, which marks accessible cytosines via recombinant methyltransferases, followed by bisulfite sequencing to infer protein-DNA interactions and chromatin accessibility at single-molecule resolution. The sequencing library was prepared using the NEBNext DNA Ultra II Library Prep Kit and sequenced on an Illumina platform, using either a MiSeq 250 bp paired-end run, a MiSeq i100 250 bp paired-end run, or a NextSeq 2000 P1 300 bp paired-end run. Reads were pre-processed with TrimGalore and a custom R script was used to trim the plasmid backbone from the reads. After, pre-processed reads were aligned using QuasR. Further analyses were conducted using custom scripts available at https://github.com/Krebslabrep/TF-chromatin.git.
2026-05-21 | E-MTAB-16658 | biostudies-arrayexpress
Project description:To directly test how individual TFs combine their functions to open chromatin at specific loci, we used synthetic DNA sequences. We generated a library containing all combinations of TF-motif deletions for three CREs: a CTCF-bound region as a positive control (found in \"MutLib1\"), an enhancer with five KLF4 motifs (found in \"MutLib2\"), and a promoter with YY1, NRF1, and MYC motifs (found in \"MutLib1\"). We then inserted the library into a landing pad within a neutral chromatin environment, devoid of activating or repressive chromatin modifications, in mESCs using Recombination-Mediated Cassette Exchange (RMCE). Chromatin accessibility of the inserted fragments was then profiled by targeted SMF using PCR primers that anneal to a synthetic flanking sequence, allowing unambiguous differentiation of the ectopic site from its endogenous counterpart. The dataset generated includes amplicon-based SMF data from the ectopic site under steady-state conditions, in mouse cells (i.e., TC-1 knock-out of the three DNA methyl transferases (DNMT TKO) mESCs). Two biological replicates were generated. In summary, cells were collected for SMF, which marks accessible cytosines via recombinant methyltransferases, followed by bisulfite sequencing to infer protein-DNA interactions and chromatin accessibility at single-molecule resolution. The sequencing library was prepared using the NEBNext DNA Ultra II Library Prep Kit and sequenced on an Illumina platform, using either a MiSeq 250 bp paired-end run, a MiSeq i100 250 bp paired-end run, or a NextSeq 2000 P1 300 bp paired-end run. Reads were pre-processed with TrimGalore and a custom R script was used to trim the plasmid backbone from the reads. After, pre-processed reads were aligned using QuasR. Further analyses were conducted using custom scripts available at https://github.com/Krebslabrep/TF-chromatin.git.
2026-05-21 | E-MTAB-16657 | biostudies-arrayexpress
Project description:To quantify the contribution of specific transcription factor binding versus chromatin context in chromatin opening, we inserted libraries containing hundreds of CREs into a landing pad within a neutral chromatin environment, devoid of activating or repressive chromatin modifications, in mESCs using Recombination-Mediated Cassette Exchange (RMCE). Chromatin accessibility of the inserted fragments was then profiled by targeted SMF using PCR primers that anneal to a synthetic flanking sequence, allowing unambiguous differentiation of the ectopic site from its endogenous counterpart. To investigate whether H3K27Ac contributes to chromatin accessibility at enhancers, we globally reduced H3K27Ac levels by chemically inhibiting the histone acetyltransferase p300 with the small molecule A-485 (24 hours treatment, final concentration: 3 μM). This was performed in the same mESCs carrying the CREs at the landing pad. The dataset generated includes amplicon-based SMF data from the ectopic site in DMSO- and A-485-treated conditions in mouse cells (i.e., TC-1 knock-out of the three DNA methyl transferases (DNMT TKO) mESCs). Two biological replicates were generated for each condition. In summary, cells were collected for SMF, which marks accessible cytosines via recombinant methyltransferases, followed by bisulfite sequencing to infer protein-DNA interactions and chromatin accessibility at single-molecule resolution. The sequencing library was prepared using the NEBNext DNA Ultra II Library Prep Kit and sequenced on an Illumina platform, using either a MiSeq 250 bp paired-end run, a MiSeq i100 250 bp paired-end run, or a NextSeq 2000 P1 300 bp paired-end run. Reads were pre-processed with TrimGalore and a custom R script was used to trim the plasmid backbone from the reads. After, pre-processed reads were aligned using QuasR. Further analyses were conducted using custom scripts available at https://github.com/Krebslabrep/TF-chromatin.git.
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Project description:Bait-capture based Single Molecule Footprinting (SMF) data from Kreibich et al., 2022. SMF data is obtained by treating extracted nuclei with a GpC methyltransferase, where binding of proteins on DNA, e.g. nucleosomes and transcription factors (TFs), leave behind unmethylated GpCs as footprints. Data in this experiment comprises SMF data obtained from WT embryonic stem cells (ES), DNMT TKO ES, TET TKO ES, F1 hybrid ES (129/CAST), neural progenitor (NP),�myoblast (C2C12) and�murine erythroleukemia (MEL)�cells. These data were generated by employing Agilent Sure-Select Mouse Methyl-Seq kit, enriching the sample for cis-regulatory regions of the mouse genome prior to library preparation. Thus, these data contain high coverage accessibility information at regulatory loci in different cell types. The SMF procedure maintains the endogenous DNA methtylation in CpG context, allowing the simultaneous detection of chromatin accessibility, TF binding and endogenous DNA methylation.
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Project description:Bait-capture based Single Molecule Footprinting (SMF) data. SMF data is obtained by treating extracted nuclei with a GpC and a CpG methyltransferase, where binding of proteins on DNA, e.g. nucleosomes and transcription factors (TFs), leave behind unmethylated cytosines as footprints. Data in this experiment comprises SMF data obtained from a previously reported Sox2 degron mESC line (see PMID: 33318687). Sequencing libraries were prepared using Agilent Sure-Select Mouse Methyl-Seq kit, enriching the sample for cis-regulatory regions of the mouse genome prior to library preparation. Thus, these data contain high coverage accessibility information at regulatory loci in different cell types.
2026-01-26 | E-MTAB-15014 | biostudies-arrayexpress
Project description:Single Molecule Footprinting (SMF) data from Sonmezer et al., 2020. SMF data is obtained by treating isolated nuclei with methyltransferases, where binding of proteins on DNA, e.g. nucleosomes and TFs, leave behind unmethylated cytosines as footprints. Data in this experiment comprises SMF data obtained from ES cells and various derivatives of ES cells, such DNMT-null, REST-knockout ES cells, neural progenitors and ES cells treated with NRF1 sirNA.
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Project description:Amplicon SMF for Pol II occupancy project
| PRJEB80138 | ENA
Project description:Bait-capture based Single Molecule Footprinting (SMF) data. SMF data is obtained by treating extracted nuclei with a GpC and a CpG methyltransferase, where binding of proteins on DNA, e.g. nucleosomes and transcription factors (TFs), leave behind unmethylated cytosines as footprints. Data in this experiment comprises SMF data obtained from two F1 hybrid embryonic stem cells (ES) obtained by crossing C57BL/6NJ with CAST/EiJ and SPRET/EiJ, respectively. The resulting cell line were underwent triple genetic knockout for the endogenous methyltransferase enzymes (DNMT TKO). Sequencing libraries were prepared using Agilent Sure-Select Mouse Methyl-Seq kit, enriching the sample for cis-regulatory regions of the mouse genome prior to library preparation. Thus, these data contain high coverage accessibility information at regulatory loci in different cell types.
2026-01-26 | E-MTAB-14460 | biostudies-arrayexpress
Project description:Bait-capture based Single Molecule Footprinting (SMF) data from Sonmezer et al., 2020. SMF data is obtained by treating isolated nuclei with methyltransferases, where binding of proteins on DNA, e.g. nucleosomes and TFs, leave behind unmethylated cytosines as footprints. Data in this experiment comprises SMF data obtained from ES, DNMT-TKO, and neural progenitor (NP) cells. These data were generated by employing Agilent Sure-Select Mouse Methyl-Seq kit, enriching the sample for regulatory regions of mouse genome prior to library preparation. Thus, these data contain high coverage accessibility information at regulatory loci in different cell types.
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