title: liver, Control, long-term,4

value: Control, long-term


title: liver, 0.2CBD, long-term,1

status: Public on Mar 16 2024

type: SRA

city: Kraków

first: Artur

last: Gurgul

address: Rędzina 1C

department: Center for Experimental and Innovative Medicine

country: Poland

institute: University of Agriculture in Krakow

library_source: transcriptomic

data_processing:
Quality control, FastQC, v0.11.9
Filtering, Flexbar, 3.5.0
Alignment, STAR, 2.7.5c
Reads counting, Htseq-count, 1.99.2
Reads normalization, DESeq2 3.16
Assembly: GRCm39
Supplementary files format and content: Processed data files include raw read counts for all samples in tabular format, text files (.txt)

molecule: polyA RNA

organism: Mus musculus

source_name: liver

extract_protocol:
Tissues were collected and immediately submerged in stayRNA™ (A&A Biotechnology) reagent. After 24 of cooling (4C), tissues were frozen at -80C until the time of RNA isolation. For RNA isolation, tissue scrapes (15-30 mg) were homogenized and used for RNA isolation with AllPrep DNA/RNA (Qiagen) kit. Following isolation, RNA was controlled for quality with the TapeStation4150 system and quantified.
Libraries were prepared using CORALL mRNA-Seq V2 (Lexogen) following standard protocol.
RNA-Seq, stranded

treatment_protocol: The experiment was conducted on C57BL/6J laboratory mice strain (n=48), divided into subgroups receiving CBD at doses of 0.2, 10, or 20 mg/kg of body weight, and a control group receiving placebo, administered for 2 (short-term) or 28 (long-term) days. CBD was administered once a day via intraperitoneal injection. RNA-Seq analysis of the liver was conducted to identify alternations in mouse transcriptomes induced by CBD treatment in those organs.

tag: tissue

value: liver

tag: treatment

value: 0.2CBD, long-term


title: liver, Control, long-term,6


title: liver, Control, long-term,3


title: liver, Control, long-term,5


title: liver, 20CBD, short-term,4

value: 20CBD, short-term


title: liver, 20CBD, short-term,5


title: liver, 20CBD, short-term,2


title: liver, 20CBD, short-term,1


title: liver, Control, long-term,2


title: liver, 20CBD, short-term,3


title: liver, Control, long-term,1


title: liver, 20CBD, short-term,6


title: liver, 10CBD, short-term,5

value: 10CBD, short-term


title: liver, 10CBD, short-term,6


title: liver, 10CBD, short-term,4


title: liver, 10CBD, long-term,4

value: 10CBD, long-term


title: liver, 10CBD, short-term,2


title: liver, 10CBD, long-term,3


title: liver, 10CBD, short-term,3


title: liver, 0.2CBD, long-term,5


title: liver, 10CBD, short-term,1


title: liver, 0.2CBD, short-term,5

value: 0.2CBD, short-term


title: liver, 0.2CBD, short-term,6


title: liver, 0.2CBD, short-term,4


title: liver, 0.2CBD, short-term,2


title: liver, 0.2CBD, short-term,1


title: liver, Control, short-term,5

value: Control, short-term


title: liver, Control, short-term,4


title: liver, 0.2CBD, short-term,3


title: liver, Control, short-term,2


title: liver, Control, short-term,3


title: HI5

status: Public on Feb 25 2024

city: Cambridge

first: Vincent

last: Butty

state: MA

address: 77 Massachusetts Avenue Bldg 68-317A

department: Biology / Koch Institute / Bioengineering / CEHS

country: USA

institute: Massachusetts Institute of Technology

description: instrument model: Singular Genomics G4

data_processing:
Fastq files were generated with the Illumina off-line basecaller (olb) v. 1.9.4.
Reads were aligned against mm10 (Feb., 2009) using bwa mem v. 0.7.12-r1039 with flags –t 16 –f, and mapping rates, fraction of multiply-mapping reads, number of unique 20-mers at the 5´ end of the reads, insert size distributions and fraction of ribosomal RNAs were calculated using bedtools v. 2.25.0. In addition, each resulting bam file was randomly down-sampled to a million reads, which were aligned against hg19, and read density across genomic features were estimated for RNA-Seq-specific quality control metrics
Fastq files were processed using the nf-core/rnaseq v 3.11.1 pipeline [https://zenodo.org/records/10171269] in the Nextflow 22.10.4 environment, using the GRCm39 reference genome and ENSEMBL GRCm39.110 murine annotation.
Gene-level expression tables of counts and transcript-per-million (TPM) generated by STAR/Salmon processing were retrieved [Dobin2013, Soneson2015, Patro2017] .
Assembly: GRCm39 / ENSEMBL 110 annotation
Supplementary files format and content: Gene-level raw counts and transcript per million (TPM) data generated by Salmon, tab-separated format

molecule: total RNA

source_name: Nervous

growth_protocol: HBG3 mESC were expanded on a feeder layer in mESC proliferation medium consisting of EmbryoMax ES DMEM with FBS, pen-strep, L-glut, EmbryoMax nucleosides, MEM non-essential amino acids, b-mercaptoethanol and mLIF. Then differentiated using a base of Advanced DMEM F-12 and Neurobasal with L-glutamine, pen-strep, KOSR, and b-mercaptoethanol. After moving into a low adhesion dish, puromorphamine and retinoic acid is added. After 5 days since initiating differentiation, we add ciliary neurotrophic factor and glial derived neurotrophic factor. Around day 8 of differentiation, the cells have aggregated into spheroids within the petri dish and have begun expressing HB9 signaling mature neuronal differentiation.

extract_protocol:
RNeasy Micro Kit Protocol
Sequencing libraries were generated using Takara's SMARTR ZapR v2-based protocol; briefly, 4 ng of each RNA sample were used (corresponding to half of the reaction volume). Full volume was used for the 8 samples with the lowest concentration. RNAs were fragmented for 4 minutes, followed by first-strand cDNA synthesis using the SMARTR Pico v2 protocol. TruSeq-Singular unique dual index (UDI) sequencing adapters were added by PCR (5 cycles). Library fragments corresponding to rRNA molecules were depleted by cleaving with the ZapR v2 and mammalian-specific R-probe v2 kit at full volume, and the second round of PCR was performed using the Singular qPCR primers for 15 cycles. Library sizes were quantified and verified by QPCR and on a Fragment Analyzer before loading for sequencing on a Singular G4 instrument in a 50-base paired-end configuration.

treatment_protocol: Once seeded onto our fibrin platform, myokine-infused medium was added to the spheroids or magnetic matrix actuation was applied for 30 mins at 4 Hz everyday throughout the length of the experiment.

value: Nervous

tag: cell line

value: HBG3 mESC HB9::GFP

tag: cell type

value: Nerve cells

tag: genotype

value: heterozygous blastocysts

value: High intensity biochem


title: C6

description: instrument model: Singular Genomics G4

value: Biochemical control


title: HI3

description: instrument model: Singular Genomics G4


title: HI2

description: instrument model: Singular Genomics G4


title: C5

description: instrument model: Singular Genomics G4


title: HI1

description: instrument model: Singular Genomics G4


title: C3

description: instrument model: Singular Genomics G4


title: C2

description: instrument model: Singular Genomics G4


title: C1

description: instrument model: Singular Genomics G4


title: LI6

description: instrument model: Singular Genomics G4

value: Low intensity biochem


title: LI4

description: instrument model: Singular Genomics G4


title: LI3

description: instrument model: Singular Genomics G4


title: LI5

description: instrument model: Singular Genomics G4


title: LI2

description: instrument model: Singular Genomics G4


title: MC4

description: instrument model: Singular Genomics G4

value: Mechanical control


title: LI1

description: instrument model: Singular Genomics G4


title: liver, Control, short-term,6


title: MC2

description: instrument model: Singular Genomics G4


title: MC1

description: instrument model: Singular Genomics G4


title: liver, Control, short-term,1


title: HI6

description: instrument model: Singular Genomics G4


title: M3

description: instrument model: Singular Genomics G4

value: Mechanical stimulation


title: HI4

description: instrument model: Singular Genomics G4


title: M4

description: instrument model: Singular Genomics G4


title: M1

description: instrument model: Singular Genomics G4


title: C4

description: instrument model: Singular Genomics G4


title: Atrazine_A24_4

status: Public on Feb 01 2024

city: RALEIGH

first: Dereje

last: Jima

state: NC

address: 850 Main Campus Drive

department: CHHE

institute: NCSU

description: Atrazine

data_processing:
Data analysis was performed in consultation with Bioinformatics Core at NCSU Center for Human Health and the Environment. An average of ~62.5 million paired-end raw RNAseq data were generated for each replicate. The quality of sequenced data was assessed using the fastqc application, and 10 poor-quality bases were trimmed from the 5’-end. The remaining good-quality reads were aligned to the X. Laevis reference genome (ZENLA 9.2) database using the STAR (Dobin et al., 2013) aligner. Per-gene counts of uniquely mapped reads for each replicate were calculated using the htseq-count script from the HTSeq (Anders et al., 2015) python package. The count matrix was imported to R statistical computing environment for further analysis. Initially, genes that had no count in most replicate samples were discarded. The remaining count data were normalized for sequencing depth and distortion, and dispersion was estimated using DESeq2 (Love et al., 2014) Bioconductor package in the R statistical computing environment. We fitted a leaner model using the treatment levels, and differentially expressed genes were identified after applying multiple testing corrections using the Benjamini-Hochberg procedure (Benjamini & Hochberg, 1995). The final significant genes were generated using padj<0.05
Assembly: https://download.xenbase.org/pub/Genomics/JGI/Xenla9.2/XENLA_9.2_Xenbase.gtf

organism: Xenopus laevis

source_name: Whole intestine

extract_protocol:
After 24 hours of exposure, intestines from embryos of each treatment group were dissected from anesthetized embryos with sharpened forceps. The anterior portion of the embryo (including the head, heart, and foregut) were removed and discarded, leaving only the mid-and hindgut portion of the intestine. Intestines from multiple embryos were removed, combined within a microcentrifuge tube containing TRIzol™ Reagent (Invitrogen) such that each replicate contained 10 intestines (n = 4). RNA extraction was performed using the TRIzol™ Reagent (Invitrogen) according to the manufacturer's instructions, followed by lithium chloride precipitation. Quality and quantity of RNA yield was confirmed on a Thermo Fisher Scientific Nanodrop 1000 and an Agilent 2100 Bioanalyzer.
Illumina poly-A enriched directional RNA library

treatment_protocol: NF 39 embryos were exposed to 35 mg/L ATR or DMSO control for 24 hours.

tag: developmental stage

value: Nieuwkoop and Faber 40

value: Homogenized whole intestines (N=10)

tag: time

value: 24 hours post-exposure

value: 35 mg/L atrazine


title: M2

description: instrument model: Singular Genomics G4


title: Atrazine_A24_3

description: Atrazine


title: Atrazine_A24_1

description: Atrazine


title: DMSO_D24_4

description: DMSO

value: 0 mg/L atrazine


title: DMSO_D24_2

description: DMSO


title: Skmel doxo 4

city: Barcelona

first: Camille

last: Stephan Otto Attolini

state: Select a State or Province

address: Baldiri Reixac 10

department: Biostatistics and bioinformatics Unit

country: Spain

institute: IRB Barcelona

data_processing:
TruSeq and Nextera adapters were removed using cuadapt (v4.1) with parameters -m 30 -e 0.2 -q 20 and -O 1
Reads were aligned to the mouse reference genome GRCh38 using STAR (v2.7.10a) with default parameters
SAM files were converted to BAM and sorted using sambamba (v2.098.1)
The count matrix was generated with the function featureCounts of the R package Rsubread (v2.8.2) and annotation from ENSEMBL GRCh38.p13
Assembly: GRCh38
Supplementary files format and content: Excel file with raw counts

organism: Homo sapiens

growth_protocol: Cells were grown in DMEM supplemented with 10% FBS and antibiotics.

extract_protocol:
Total RNA was extracted through a Trizol-Chloroform separation step. The acqueous phase was then loaded onto the Qiagen Rneasy Mini Kit (Cat #74106) columns and  the manufacturer's instructions were followed (bypassing the step of cell lysis) in order to finish the RNA extraction.
RNA libraries were prepared for sequencing using the kits Illumina® Stranded mRNA Prep, Ligation (20040534) and IDT® for Illumina® RNA UD Indexes Set A, Ligation (20040553), following manufacturer´s instructions and 10 cycles of amplification.

treatment_protocol: Senescent was induced with doxorubicin (100nM) and palbociclib (1uM SK-MEL-147, 5uM in A549). After 72hrs medium free of drug was used with every change. Diapause-like state was induced with INK128 (100nM in SK-MEL-147 and 200nM in A549). INK128 was replenished every media change. Media was changed every 3days.

tag: replicate

value: doxo


title: Atrazine_A24_2

description: Atrazine


title: Skmel pb 4

value: pb


title: DMSO_D24_1

description: DMSO


title: Skmel ink 4

value: ink


title: Skmel ink 3


title: Skmel ctr 4

value: ctr


title: Skmel pb 3


title: MC3

description: instrument model: Singular Genomics G4

value: Nervous

value: HBG3 mESC HB9::GFP


title: Skmel ctr 3


title: Skmel doxo 3


title: Skmel ink 2


title: Skmel ctr 2


title: Skmel pb 1


title: Skmel doxo 1


title: Skmel pb 2


title: Skmel doxo 2


title: A549 ink 3


title: Skmel ink 1


title: A549 ctr 3


title: A549 doxo 3


title: A549 ink 2


title: A549 pb 2-2


title: A549 pb 3


title: DMSO_D24_3

description: DMSO


title: A549 ctr 2-2


title: A549 ctr 2


title: A549 pb 2


title: A549 ink 2-2


title: A549 doxo 1


title: A549 ink 1


title: A549 ctr 1


title: SK-Mel-147_ink_rep3_H4K20me3

city: Great Abington

first: Laia

last: Richart Gines

state: Cambridgeshire

address: Granta Park

country: United Kingdom

institute: Altos Labs

library_source: genomic

data_processing:
*library strategy: CUT&Tag-Seq
Adapters and low‐quality bases were removed from reads with Cutadapt and TrimGalore.
Trimmed reads were mapped to the human reference genome hg38 with Bowtie2 using default parameters. PCR duplicates were removed with Picard Tools MarkDuplicates.
Deduplicated bam files of biological replicates were merged using SAMtools merge.
BigWig files were created using DeepTools bamCoverage.
Assembly: hg38
Supplementary files format and content: BigWig files of genome-wide enrichment of the probed histone modification.

molecule: genomic DNA

extract_protocol:
After antibody binding and tagmentation, PCI was added and the solution was transferred to a phase-lock tube and centrifuged for 5 min at 16,000xg. Chloroform was added and the acqueous solution was transferred to a new tube containing ethanol. Samples were centrifuged for 15min at 16,000xg. Two additional ethanol washes were performed before drying the pellet and dissolving it in Tris-HCl pH 8 0.1mM EDTA.
Libraries were prepared for sequencing using barcoded primers from Buenrostro et al. 2015. The NEBNext HiFi 2x PCR Master mix was used to perform the reaction.

treatment_protocol: Senescence was induced with doxorubicin (100nM) for 72h. Diapause-like state was induced with INK128 (100nM in SK-Mel-147 and 200nM in A549). INK128 was replenished at every media change every 3 days.

value: Melanoma

tag: antibody


title: SK-Mel-147_ink_rep2_H4K20me3


title: SK-Mel-147_ink_rep1_H4K20me3


title: SK-Mel-147_doxo_rep3_H4K20me3

value: Doxorubicin


title: SK-Mel-147_doxo_rep2_H4K20me3


title: SK-Mel-147_doxo_rep1_H4K20me3


title: Skmel ctr 1


title: SK-Mel-147_ctrl_rep3_H4K20me3


title: SK-Mel-147_ctrl_rep1_H4K20me3


title: A549_ink_rep3_H4K20me3

value: Lung carcinoma


title: A549_ink_rep2_H4K20me3


title: A549 doxo 2-2


title: A549_doxo_rep2_H4K20me3


title: A549 doxo 2


title: A549_ctrl_rep3_H4K20me3


title: A549_doxo_rep1_H4K20me3


title: A549_doxo_rep3_H4K20me3


title: A549_ctrl_rep2_H4K20me3


title: A549_ctrl_rep1_H4K20me3


title: SK-Mel-147_ink_rep3_H3K9me3


title: SK-Mel-147_ink_rep1_H3K9me3


title: A549 pb 1


title: SK-Mel-147_doxo_rep2_H3K9me3


title: SK-Mel-147_ink_rep2_H3K9me3


title: SK-Mel-147_ctrl_rep3_H3K9me3


title: SK-Mel-147_ctrl_rep2_H3K9me3


title: SK-Mel-147_doxo_rep1_H3K9me3


title: SK-Mel-147_ctrl_rep1_H3K9me3


title: A549_ink_rep3_H3K9me3


title: A549_ink_rep2_H3K9me3


title: A549_doxo_rep1_H3K9me3


title: A549_ink_rep1_H3K9me3


title: A549_doxo_rep3_H3K9me3


title: A549_ctrl_rep3_H3K9me3


title: A549_ink_rep1_H4K20me3


title: A549_ctrl_rep1_H3K9me3


title: A549_doxo_rep2_H3K9me3


title: U937

status: Public on Jan 14 2024

city: Charlottesville

first: Kevin

middle: A.

last: Janes

state: VA

address: Box 800759 Health System

department: Department of Biomedical Engineering

institute: University of Virginia

data_processing:
For sequencing data with fastq files, read quality was assessed with FastQC (version 0.11.5). Illumina adapter sequences from raw fastq files were trimmed using Trimmomatic (version 0.39). The trimmed fastq files were aligned to the GRCh38 human reference genome (hg38, release 86) using HISAT2 (version 2.1.0) and gene counts were obtained using featureCounts, downloaded from Subread package. All downstream analyses were performed in R Studio (version, 1.4.1106) using the Bioconductor framework.
Assembly: GrCh38.86
Supplementary files format and content: .csv file contains raw counts for each gene per sample

source_name: Acute myeloid leukemia cell line

growth_protocol: Human AML cell lines were cultured as previously described (Pearson et al. Mol. Cancer Research, 2020).

extract_protocol:
Total RNA was isolated from AML patient PBMCs and AML cell lines by resuspending pellets in Trizol and purifying RNA with Direct-zol RNA Miniprep (Zymo Research # R2052). DNase treatment was done using RNase-Free DNase Set (Qiagen # 79254). RNA integrity was analyzed using the Agilent TapeStation system according to the manufacturer’s protocol. RNA samples with RIN scores higher than 5.3 were sent to Novogene for mRNA library prep and sequencing. Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers followed by the second strand cDNA synthesis. The library was ready after end repair, A-tailing, adapter ligation, size selection, amplification, and purification.
100 ng total RNA isolated using trizol standard protocol (according to the manufacturer’s instructions) was used for sequencing using TruSeq Stranded Total RNA Ribo Zero kit (Human/Mouse/Rat) on NovaSeq6000 S4, 150 bp paired-end.

value: Acute myeloid leukemia cell line


title: Urea TKS400G P13 Thymus KO CTCF

status: Public on Dec 21 2023

city: San Francisco

first: Terumi

last: Kohwi-Shigematsu

state: CA

address: 513 Parnassus Ave (HSW810)

department: Department of Orofacial Sciences

institute: UCSF

description: Urea TKS400G  P13 Thymus KO CTCF

data_processing:
Sequenced reads were quality-checked using FastQC prior to downstream analysis. Unaligned reads were mapped to the mouse reference genome (NCBI37/mm9) using Bowtie2 (version 2.3.1).  Candidate binding sites were called using MACS2 (v2.2.7) using a q-value threshold of 0.01. Coverage tracks are presented in RPM (read pileups per million reads) and utilities from UCSC were used for creating browser tracks.
assembly: mmp9

source_name: Thymus

extract_protocol:
Thymocytes isolated from thymus of Satb1-/- mice (P13)  were cross-linked with 1% formaldehyde for 10 min at room temperature, followed by addition of glycine at 0.125 M for 5 min to quench cross-linking reaction. These cells were washed with 1X PBS, suspended in 1% BSA-PBS and cells were precipitated in a tube.  These cells were re-suspended in lysis buffer (4% SDS, 50 mM Tris-HCl [pH 8.0], 10 mM EDTA, 100 mM NaCl) with Protease inhibitor (Roche) and RNase inhibitor.  The resulting cell lysate was incubated for 30 min at room temperature and loaded on top of 8 M urea followed by ultracentrifugation in a Beckman TL-100 Ultracentrifuge for 4-5 hrs at 45K rpm (181,000g, g=relative centrifugation force or RCF; TLS55 rotor).  Recovered chromatin pellet was sonicated to generate fragment size ranging 300–1,000bp using Branson sonifier (20% power, 10 sec on and 30 sec off for 4-6 cycles) in 300ml of sonication buffer [0.5% Na-laurylsarcosine, 0.1% deoxycholate (DOC), 1mM EDTA, 0.5mM EGTA, 10mM TrisHCl, 100mM NaCl] with Protease inhibitor and RNase inhibitor. Note: urea-purified crosslinked chromatin is highly vulnerable to sonication, therefore, it needs to adjust the sonication condition depending on cell type and/or the size of the precipitate DNA.  The sonicated chromatin with 1% Triton X-100 were dialyzed against dialysis buffer (10 mM Tris-HCl [pH 8.0], 100 mM NaCl, 1 mM EDTA, 5% glycerol), and dialyzed for 16 hrs at 4°C (this dialysis is optional).  The chromatin sample was centrifuged at 15,000g for 5 min at 4°C to remove a trace amount of residual insoluble components.   The test-reverse cross-linking of chromatin aliquot (5-10ml of the final volume) was done at 65°C for 2-3 hrs in buffer containing 1% SDS, 100 mM NaCl, 1 mM EDTA, 10mMTris.  Samples were further treated by 100 µg/ml RNase A at 37°C for 1 hr, 200 µg/ml proteinase K at 60°C for more than 3 hrs, followed by DNA purification using QIAquick PCR purification kit (Qiagen), shearing efficiency was examined by Bioanalyzer 2100 (Agilent Technologies Inc.). For chromatin-immunoprecipitation, 1 µg of an antibody of choice were added to 10 ~ 20 µg of sonicated solubilized chromatin in 350 ml in dilution buffer (1% Triton X-100, 0.01% SDS, 1 mM EDTA, 15 mM Tris-HCl, 100 mM NaCl) and incubated overnight with rotation at 4°C.  At least three chromatin samples were prepared for ChIP-seq from independent experiments to confirm the results. The chromatin fragments bound to antibodies (after incubation overnight) were captured using 20 µl of protein-A, protein-G or protein AG Dynabeads (Invitrogen; pre-washed with 0.1% BSA [Fraction V, Sigma), by 4 hrs incubation time at 4°C.  Dynabeads bound to chromatin fragments were washed twice with Binding buffer (10 mM Tris, pH8.0, 1 mM EDTA, 1 mM EGTA, 0.5% Triton X-100, 0.1% DOC, 100 mM NaCl), twice with LiCl buffer (100 mM Tris [pH 8.0], 1 mM EDTA, 250 mM LiCl, 0.5% NP-40, and 0.5% DOC).  The tubes were changed and washed twice with 0.1M NaCl-TE buffer. The immunoprecipitated samples were eluted from Dynabeads by elution buffer (10 mM Tris-HCl [pH 8.0], 1 mM EDTA, 100 mM NaCl and 1% SDS) at 65oC for 20 min.  After elution, ChIP DNAs was reverse-crosslinked at 65oC for 6 hrs, followed by RNase-A treatment at 37°C for 1 hr and proteinase K treatments for 3 hrs at 60°C. The recovered DNA was further purified with a QIAquick PCR purification kit (Qiagen), with elution in 15µl of Qiagen Elution Buffer (alternatively, Phenol-chloroform extraction and ethanol precipitation).
Briefly for library construction, ChIP DNA fragments were end-repaired and ligated to indexed sequencing adapters (NEB or Rubicon library kit) and amplified with 8-11 rounds of PCR, depending on the DNA amount.  Using SPRI beads (Beckmancoulter), small fragments (<200bp) were removed from the amplified library. The size-selected library was sequenced to 50bp single-end read lengths on an Illumina HiSeq4000 (Vincent J. Coates Genomic Sequencing Laboratory, UC Berkeley).

value: Thymus

value: SATB1 KO

tag: chip antibody

value: CTCF


title: Hindlimb, E11.5, H11^ZRS-mCherry/ZRS404G>A-eGFP (dual-enSERT-1), scRNA-seq

status: Public on Jan 18 2024

city: Irvine

first: Evgeny

last: Kvon

address: 4310 McGaugh Hall

department: Developmental and Cell Biology

institute: University of California, Irvine

library_source: transcriptomic single cell

data_processing:
Fastq files were aligned to the mm39 genome assembly (Ensembl, GCA_000001635.9) and barcodes counted to generate a count matrix using CellRanger (10x Genomics, Cell Ranger 3.1.0). Count matrix data were analyzed using the Seurat R package, version 4.
Assembly: mm39 with custom build for mCherry and GFP

source_name: FVB

extract_protocol:
Hindlimb cells were dissociated with collagenase II into single cells and loaded at a concentration of 10,000 cells.
Recovery of 10,000 cells by the Chromium Next GEM Chip G Single Cell Kit 3’ Gene Expression Kit v3.1 (10X Genomics, cat no. 1000127)
Captured cells were pair-end sequenced on an Illumina NovaSeq 6000 for approximately 50,000 reads per cell.

tag: strain

value: FVB

value: Hindlimb

tag: age


title: Late, Ctrl, replicate2, scATACseq

status: Public on Dec 20 2023

city: Solna

first: Bastien

last: Hervé

address: Solnavägen 9

department: MBB

country: Sweden

institute: Karolinska Institutet

description: 10X Genomics

library_source: genomic single cell

data_processing:
A total of 6 +1 batches were collected from the sequencing facility. Fastq files from 19 +9 samples were processed throughout the 10X genomics standard pipeline. Gene expression and chromatin accessibility libraries were input into cellranger-arc ‘count’ v2.0.2 with default settings to align the biological readouts on the associated mm10 reference genome v2020-A-2.0.0. Sample aggregation of both transcriptomic and genomics metrics was done using the ‘aggr’ of the same cellranger executable file, without normalization. The aggregated count matrix and fragments file were loaded into R v4.3.2. The former was lodged into a Seurat v5.0.3 assay while the latter accommodate into a Signac v1.13.0 chromatin assay associated to an Ensembl base annotation v79 for mouse where UCSC nomenclature was applied to provide gene names readability. If not specifically mentioned, the following depictions of the analysis pipeline belong to either Seurat or Signac packages. Cell free mRNA contamination was removed from each droplet using Cellbender v0.3.0. Genes called in all sample by Cellbender were included into the feature list. Potential cells called in both Cellranger and Cellbender were included into the cell list. A total of 181,674 cells were retrieved. Most of the genes affected by the ambient RNA removal were canonical genes and ribosomal genes. After assessing that each cell could be uniquely identified, quality metrics for both gene expression and peak accessibility were calculated. Among others, the mitochondrial ratio, the cell cycle score, the nucleosome signal, and the Transcription Start Site (TSS) enrichment were generated to support the following quality control cutoffs. Depending of the quality of each sample, for each cell, a maximum of 30,000/150,000 and a minimum of 1,000 ATAC counts, a maximum of 10,000/50,000 and a minimum of 600 RNA counts, a minimum of 600 detected genes, a maximum of 0.8/1.5 nucleosomal signal, a TSS minimum enrichment of 2 and a maximum percentage of mitochondrial information of 15/50 were prerequisite to consider a given cell for the downstream analysis. These thresholds shrinked by around 12.6% the overall number of cells, from 179,078 down to 156,205. While these thresholds are on purpose not too strained, they did not lead to any clustering perturbation downstream.
Assembly: mm10
Supplementary files format and content: EAE_multiomics.rds : Aggregation of cells from all samples in a Seurat object with associated metadata
Supplementary files format and content: EAE_multiomics_annotation.csv : Metadata annotation of the aggregation of cells from all samples in a Seurat object
Supplementary files format and content: EAE_multiomics_fragments.tsv.gz : Aggregation of fragments from all samples
Supplementary files format and content: EAE_multiomics_fragments.tsv.gz.tbi : Index of aggregation of fragments from all samples
Supplementary files format and content: EAE_multiomics_peak_annotation.tsv : Annotation of peaks called by Macs2, following 10Xgenomics cellranger-arc peaks annotation guidelines
Supplementary files format and content: EAE_multiomics.bedpe : Association of peaks to genes (LinkPeaks), peaks to genes (Cicero) and peaks annotation to generate a bedpe file similar to cellranger-arc output
Supplementary files format and content: EAE_multiomics_peaks2genes.rds : Peaks to genes links using LinkPeaks function from Signac package with 500,000bp maximum distance from TSS and minimum 10 cells to validate the link
Supplementary files format and content: EAE_multiomics_peaks2peaks.rds : Cicero connections calculation on all celltypes downsampled to 1000 cells per timepoint, and generation of connection graphs via generate_ccans and ConnectionsToLinks functions
Supplementary files format and content: EAE_multiomics_OLG_peaks2genes : Peaks to genes links on OLGs populations, using LinkPeaks function from Signac package with 50,000bp maximum distance from TSS and minimum 10 cells to validate the link
Supplementary files format and content: EAE_multiomics_OLG_DORCs_peaks2genes : Peaks to genes links on OLGs populations for potential DORC genes, using LinkPeaks function from Signac package with 500,000bp maximum distance from TSS and minimum 10 cells to validate the link
Supplementary files format and content: Astro_Ctrl-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Astrocytes population at Ctrl time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: Astro_Early-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Astrocytes population at Early time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: Astro_Peak-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Astrocytes population at Peak time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: Astro_Late-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Astrocytes population at Late time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: OPC_Ctrl-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of OPC population at Ctrl time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: OPC_Early-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of OPC population at Early time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: OPC_Peak-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of OPC population at Peak time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: OPC_Late-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of OPC population at Late time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: COP_Ctrl-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of COP population at Ctrl time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: COP_Early-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of COP population at Early time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: COP_Peak-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of COP population at Peak time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: COP_Late-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of COP population at Late time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: Micro_Ctrl-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Microglia population at Ctrl time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: Micro_Early-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Microglia population at Early time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: Micro_Peak-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Microglia population at Peak time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: Micro_Late-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of Microglia population at Late time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL1_Ctrl-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL1 population at Ctrl time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL1_Early-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL1 population at Early time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL1_Peak-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL1 population at Peak time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL1_Late-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL1 population at Late time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL2_Ctrl-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL2 population at Ctrl time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL2_Early-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL2 population at Early time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL2_Peak-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL2 population at Peak time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL2_Late-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL2 population at Late time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL56_Ctrl-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL56 population at Ctrl time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL56_Early-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL56 population at Early time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL56_Peak-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL56 population at Peak time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file
Supplementary files format and content: MOL56_Late-TileSize-100-normMethod-rc.bw : Bigwig file of fragments coverage in 100bp window size of MOL56 population at Late time point normalized by relative count (rc) as a scaling factor of 10,000 divided by the total number of fragments involved in this file

source_name: Spinal Cord

extract_protocol:
At early, peak, and late stage, mice were perfused with PBS, and spinal cords were collected. Spinal cord tissues were then dissociated into a single cell suspension according to the manufacturer’s protocol of Adult Brain Dissociation Kit, mouse and rat (Miltenyi Biotec, 130-107-677, we did not perform the red blood cells removal step since majority of the red blood cells had been removed with PBS perfusion). For the debris removal step, majority of the samples were processed with 38% percoll except 1 Naïve sample, 3 CFA-Ctrl samples, 3 EAE early stage samples, 2 + 1 EAE peak stage samples, and 2 + 1 EAE late stage samples were processed debris removal solution (Miltenyi Biotec, 130-107-677) according to the manufacturer’s protocol to enrich more OPCs. Spinal cord single EGFP+ cells were enriched with the BD FACSAria III Cell Sorter (BD Biosciences). The cells were then lysed and washed according to the demonstrated protocol, Nuclei Isolation for Single Cell Multiome ATAC + Gene Expression Sequencing (10x Genomics, CG000365), with modifications: the cells were centrifuged for 10 min at 300 g and 4°C, resuspended in lysis buffer (containing 10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.01% Tween-20, 0.01% IGEPAL (CA-630), 0.001% Digitonin, 1% BSA, 1 mM DTT, 1 U/ul RNase inhibitor) and incubated on ice for 3 min. After the incubation, wash buffer (containing 10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20, 1% BSA, 1 mM DTT, 1 U/ul RNase inhibitor) was added on top without mixing. The nuclei were centrifuged for 5 min at 500 g and 4°C. Nuclei were washed once in wash buffer followed by another wash with diluted 1x Nuclei buffer (10x Genomics, PN-1000283) containing 1% BSA, 1 mM DTT and 1 U/ul RNase inhibitor.
Libraries were sequenced on an Illumina Novaseq 6000 with a 50-8-24-49 read setup for ATAC (minimum 25,000 read pairs per cell) and a 28-10-10-90 read setup for RNA (minimum 20,000 read pairs per cell).
The Chromium Next GEM Single Cell Multiome ATAC + Gene Expression chemistry (10x Genomics, PN-1000283) was applied to create single cells ATAC and RNA libraries. 26 +10 EAE mice (8 +6 mice from early stage, 8 +2 mice from peak stage and 10 +2 mice from late stage), 12 +6 CFA-Ctrl mice (4 +6 mice from early stage, 4 mice from peak stage and 4 mice from late stage) were used for independent replicates.

treatment_protocol: For the EAE induction, animals were injected subcutaneously with an emulsion of MOG35–55 in complete Freund's adjuvant (CFA) (Hooke Laboratories, EK-2110, containing 1 mg MOG35-55/mL emulsion and 2-5 mg killed mycobacterium tuberculosis H37Ra/mL emulsion, Hooke Laboratories adjusted concentrations by lot) followed by the intraperitoneal injection of pertussis toxin in PBS on day 0 and day 1 (200-225 ng per animal, adjusted by lot according to Hooke manufacturer’s instructions). Scores of EAE were graded according to the following criteria: 0, asymptomatic; 1, limp tail or titubation; 2, limp tail and weakness of hindlimbs; 3, limp tail and complete paralysis of hindlimbs; 4, limp tail, complete paralysis of two hindlimbs with forelimb involvement; 5, moribund or dead; 0.5 for intermediate symptoms. CFA-control mice were injected subcutaneously with the control emulsion containing CFA but without MOG35–55 (Hooke Laboratories, CK-2110), and followed by the administration of pertussis toxin in PBS on day 0 and day 1 (200-225 ng per animal, adjusted by lot according to Hooke manufacturer’s instructions). Spinal cords from EAE and CFA-Ctrl were collected at early (day 8-9), peak (day 14-15), and late stage (day 38-40)

tag: timepoint

value: Late

tag: sequencing id

value: Spinal Cord

value: Sox10:Cre-RCE:LoxP (EGFP)

tag: debris removal

value: percoll

tag: batch

tag: Sex

value: pooled male and female

tag: data tissue_collection

tag: number of_mice

value: CFA-Ctrl

tag: eae scores


title: Late, Ctrl, replicate1, scRNAseq

description: 10X Genomics

tag: tissue

tag: cell line

tag: treatment


title: Late, Ctrl, replicate1, scATACseq

description: 10X Genomics

tag: genotype


title: Late, Ctrl, replicate2, scRNAseq

description: 10X Genomics

tag: replicate


title: SK-Mel-147_doxo_rep3_H3K9me3

tag: cell type

tag: antibody


title: Early, Ctrl, replicate2, scATACseq

description: 10X Genomics

value: Early


title: Urea TKS355E P13 Thymus WT CTCF (replica 2)

description: Urea TKS355E P13 Thymus WT CTCF

extract_protocol:
Thymocytes isolated from thymus of Satb1+/+ mice (P13)  were cross-linked with 1% formaldehyde for 10 min at room temperature, followed by addition of glycine at 0.125 M for 5 min to quench cross-linking reaction. These cells were washed with 1X PBS, suspended in 1% BSA-PBS and cells were precipitated in a tube.  These cells were re-suspended in lysis buffer (4% SDS, 50 mM Tris-HCl [pH 8.0], 10 mM EDTA, 100 mM NaCl) with Protease inhibitor (Roche) and RNase inhibitor.  The resulting cell lysate was incubated for 30 min at room temperature and loaded on top of 8 M urea followed by ultracentrifugation in a Beckman TL-100 Ultracentrifuge for 4-5 hrs at 45K rpm (181,000g, g=relative centrifugation force or RCF; TLS55 rotor).  Recovered chromatin pellet was sonicated to generate fragment size ranging 300–1,000bp using Branson sonifier (20% power, 10 sec on and 30 sec off for 4-6 cycles) in 300ml of sonication buffer [0.5% Na-laurylsarcosine, 0.1% deoxycholate (DOC), 1mM EDTA, 0.5mM EGTA, 10mM TrisHCl, 100mM NaCl] with Protease inhibitor and RNase inhibitor. Note: urea-purified crosslinked chromatin is highly vulnerable to sonication, therefore, it needs to adjust the sonication condition depending on cell type and/or the size of the precipitate DNA.  The sonicated chromatin with 1% Triton X-100 were dialyzed against dialysis buffer (10 mM Tris-HCl [pH 8.0], 100 mM NaCl, 1 mM EDTA, 5% glycerol), and dialyzed for 16 hrs at 4°C (this dialysis is optional).  The chromatin sample was centrifuged at 15,000g for 5 min at 4°C to remove a trace amount of residual insoluble components.   The test-reverse cross-linking of chromatin aliquot (5-10ml of the final volume) was done at 65°C for 2-3 hrs in buffer containing 1% SDS, 100 mM NaCl, 1 mM EDTA, 10mMTris.  Samples were further treated by 100 µg/ml RNase A at 37°C for 1 hr, 200 µg/ml proteinase K at 60°C for more than 3 hrs, followed by DNA purification using QIAquick PCR purification kit (Qiagen), shearing efficiency was examined by Bioanalyzer 2100 (Agilent Technologies Inc.). For chromatin-immunoprecipitation, 1 µg of an antibody of choice were added to 10 ~ 20 µg of sonicated solubilized chromatin in 350 ml in dilution buffer (1% Triton X-100, 0.01% SDS, 1 mM EDTA, 15 mM Tris-HCl, 100 mM NaCl) and incubated overnight with rotation at 4°C.  At least three chromatin samples were prepared for ChIP-seq from independent experiments to confirm the results. The chromatin fragments bound to antibodies (after incubation overnight) were captured using 20 µl of protein-A, protein-G or protein AG Dynabeads (Invitrogen; pre-washed with 0.1% BSA [Fraction V, Sigma), by 4 hrs incubation time at 4°C.  Dynabeads bound to chromatin fragments were washed twice with Binding buffer (10 mM Tris, pH8.0, 1 mM EDTA, 1 mM EGTA, 0.5% Triton X-100, 0.1% DOC, 100 mM NaCl), twice with LiCl buffer (100 mM Tris [pH 8.0], 1 mM EDTA, 250 mM LiCl, 0.5% NP-40, and 0.5% DOC).  The tubes were changed and washed twice with 0.1M NaCl-TE buffer. The immunoprecipitated samples were eluted from Dynabeads by elution buffer (10 mM Tris-HCl [pH 8.0], 1 mM EDTA, 100 mM NaCl and 1% SDS) at 65oC for 20 min.  After elution, ChIP DNAs was reverse-crosslinked at 65oC for 6 hrs, followed by RNase-A treatment at 37°C for 1 hr and proteinase K treatments for 3 hrs at 60°C. The recovered DNA was further purified with a QIAquick PCR purification kit (Qiagen), with elution in 15µl of Qiagen Elution Buffer (alternatively, Phenol-chloroform extraction and ethanol precipitation).
Briefly for library construction, ChIP DNA fragments were end-repaired and ligated to indexed sequencing adapters (NEB or Rubicon library kit) and amplified with 8-11 rounds of PCR, depending on the DNA amount.  Using SPRI beads (Beckmancoulter), small fragments (<200bp) were removed from the amplified library. The size-selected library was sequenced to 50bp single-end read lengths on an Illumina HiSeq4000 (Vincent J. Coates Genomic Sequencing Laboratory, UC Berkeley).

value: SATB1 Wild-type

tag: chip antibody


title: Early, Ctrl, replicate2, scRNAseq

description: 10X Genomics


title: Peak, Ctrl, replicate2, scATACseq

description: 10X Genomics

value: Peak


title: Peak, Ctrl, replicate1, scATACseq

description: 10X Genomics

tag: associated geo

tag: associate geo_sample


title: Early, Ctrl, replicate1, scRNAseq

description: 10X Genomics

tag: timepoint

tag: sequencing id


title: Early, Ctrl, replicate1, scATACseq

description: 10X Genomics

tag: debris removal

tag: batch


title: Peak, Ctrl, replicate1, scRNAseq

description: 10X Genomics

tag: Sex

tag: data tissue_collection


title: A549_ctrl_rep2_H3K9me3


title: Late, EAE, replicate3, scATACseq

description: 10X Genomics

tag: number of_mice

value: EAE

tag: eae scores


title: Late, EAE, replicate4, scRNAseq

description: 10X Genomics

tag: tissue

tag: cell line

value: debris

value: male


title: Late, EAE, replicate3, scRNAseq

description: 10X Genomics

tag: genotype

tag: treatment


title: Late, EAE, replicate4, scATACseq

description: 10X Genomics

tag: replicate


title: Late, EAE, replicate2, scATACseq

description: 10X Genomics


title: Late, EAE, replicate5, scATACseq

description: 10X Genomics

value: female


title: Late, EAE, replicate2, scRNAseq

description: 10X Genomics


title: Peak, EAE, replicate4, scATACseq

description: 10X Genomics


title: Peak, EAE, replicate4, scRNAseq

description: 10X Genomics


title: Late, EAE, replicate1, scRNAseq

description: 10X Genomics


title: Late, EAE, replicate1, scATACseq

description: 10X Genomics


title: Peak, EAE, replicate3, scATACseq

description: 10X Genomics


title: Peak, EAE, replicate3, scRNAseq

description: 10X Genomics


title: Peak, EAE, replicate2, scATACseq

description: 10X Genomics


title: Early, EAE, replicate3, scRNAseq

description: 10X Genomics


title: Peak, EAE, replicate2, scRNAseq

description: 10X Genomics


title: Peak, EAE, replicate1, scATACseq

description: 10X Genomics


title: Early, EAE, replicate3, scATACseq

description: 10X Genomics


title: Peak, EAE, replicate1, scRNAseq

description: 10X Genomics


title: Early, EAE, replicate4, scATACseq

description: 10X Genomics


title: Early, EAE, replicate4, scRNAseq

description: 10X Genomics


title: Early, EAE, replicate2, scRNAseq

description: 10X Genomics


title: Peak, Ctrl, replicate2, scRNAseq

description: 10X Genomics


title: OPFR_M_S42-03_S24

status: Public on Dec 13 2023

description: Gene Expression in hippocampus

data_processing:
Illumina Casava1.7 software used for basecalling.
Data analysis was performed in consultation with Bioinformatics Core at NCSU Center for Human Health and the Environment. An average of ~44.7 million paired-end raw RNAseq data were generated for each replicates. The quality of sequenced data was assessed using the fastqc application, and 12 poor-quality bases were trimmed from the 5’-end. The remaining good-quality reads were aligned to the Rat reference genome (mRatBN 7.2 version 106) downloaded from the Ensembl database using STAR (Dobin et al., 2013) aligner. Per-gene counts of uniquely mapped reads for each replicate were calculated using the htseq-count script from the HTSeq (Anders et al., 2015) python package. The count matrix was imported to R statistical computing environment for further analysis. Initially, genes that had no count in most replicate samples were discarded. The remaining count data were normalized for sequencing depth and distortion, and dispersion was estimated using DESeq2 (Love et al., 2014)Bioconductor package in the R statistical computing environment (R Core Team (2022). We fitted a leaner model using the treatment levels, and differentially expressed genes were identified after applying multiple testing corrections using the Benjamini-Hochberg procedure (Benjamini & Hochberg, 1995). The final significant genes were generated using padj<0.05
Assembly: mRatBN 7.2 version 106
Supplementary files format and content: Matrix table with raw gene counts for every gene and every sample

organism: Rattus norvegicus

source_name: Hippocampus Tissue Punches (Brain)

extract_protocol:
6, 1 mm punches were taken from dorsal hippocampus from adult offspring. Punches were kept at -80C until RNA was harvested using Qiagen AllPrep DNA/RNA/miRNA kit.
Purification of messenger RNA (mRNA) was performed using oligo-dT beads provided in the NEBNext Poly(A) mRNA Magnetic Isolation Module (New England Biolabs [NEB]). Complementary DNA (cDNA) libraries for Illumina sequencing were constructed using the NEBNext Ultra Directional RNA Library Prep Kit (NEB) and NEBNext Multiplex Oligos for Illumina (NEB) using the manufacturer-specified protocol. Following adapter ligation, the samples were selected for a final library size (adapters included) of 400–550 bp using sequential AMPure XP bead isolation (Beckman Coulter). Library enrichment was performed and indexes for each sample were added during the protocol-specified PCR amplification. Amplified library fragments were purified and checked for quality and final concentration using an Agilent 2200 Tapestation (D1000 chip, Agilent Technologies) combined with a Qubit fluorometer (ThermoFisher). Libraries were pooled in equimolar amounts for clustering and sequencing
RNA-Seq, Illumina HiSeq 2500 DNA sequencer, utilizing a 125 bp single-end cycle sequencing kit

treatment_protocol: rats exposured  to vehicle or OPFR mixture (3.3 mg/kg bw) daily from conception, through gestation to P10, sacraficed and the brains removed.

tag: strain

value: Wistar Rat

value: 6, 1 mm hippocampal punches

tag: age

value: postnatal day 10

value: 1000 ug OPFR mixture in 20 ul  in ethanol on treat


title: Early, EAE, replicate2, scATACseq

description: 10X Genomics


title: Late, EAE, replicate5, scRNAseq

description: 10X Genomics

value: Late


title: Early, EAE, replicate1, scATACseq

description: 10X Genomics

value: Spinal Cord


title: Early, EAE, replicate1, scRNAseq

description: 10X Genomics

value: Sox10:Cre-RCE:LoxP (EGFP)


title: OPFR_M_S41-01_S22

description: Gene Expression in hippocampus


title: OPFR_F_S41-03_S23

description: Gene Expression in hippocampus


title: OPFR_F_S38-06_S21

description: Gene Expression in hippocampus


title: OPFR_M_S37-01_S19

description: Gene Expression in hippocampus


title: OPFR_M_S34-06_1_S17

description: Gene Expression in hippocampus


title: OPFR_F_S30-06_S15

description: Gene Expression in hippocampus


title: OPFR_F_S29-05_S14

description: Gene Expression in hippocampus


title: OPFR_F_S37-07_S20

description: Gene Expression in hippocampus


title: CNT_M_S14-05_S11

description: Gene Expression in hippocampus

value: 20 ul ethanol on treat administered to mother


title: CNT_F_S12_05_S9

description: Gene Expression in hippocampus


title: OPFR_F_S34-06_2_S18

description: Gene Expression in hippocampus


title: CNT_F_S11_08_S8

description: Gene Expression in hippocampus


title: CNT_F_S14-07_S12

description: Gene Expression in hippocampus


title: OPFR_M_S29-03_S13

description: Gene Expression in hippocampus


title: CNT_M_S13-05_S10

description: Gene Expression in hippocampus


title: CNT_M_S11_06_S7

description: Gene Expression in hippocampus


title: CNT_M_S10_04_S5

description: Gene Expression in hippocampus


title: CNT_F_S09_07_S4

description: Gene Expression in hippocampus


title: Vglut, Sublibrary 3, Replicate 6, DNA, CreON

status: Public on Nov 08 2023

city: St. Louis

first: Joseph

middle: D

last: Dougherty

state: MO

address: 4370 Duncan Ave

department: Genetics

institute: Washington University in St. Louis

description:
Prepared from DNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, amplified with CreON primer.
vglut_combined_umi_counts.csv

data_processing:
UMIs were extracted from the start of Read2 using umi_tools (v1.0.0): umi_tools extract -I <Read2 fastq> --bc-pattern=NNNNNNNNNNNN --read2-in=<Read1 fastq> --stdout=<processed Read2 fastq> --read2-out=<processed Read1 fastq>
Barcode sequences were extracted from reads using cutadapt (v2.1); For libraries primed with the CreOFF RT primer, the following command was run: cutadapt --cores=0 -a "^TAGTAACACGACGCCGACTGTCGAC...GGTACCCGCATCGTTAATCGGCATAACTTCGTATAATGTATGCTATACGAAGTTATGGGTCGATGGTGAGATCTGGACTAGAGGGTCGANNNNNNNNNNNNAGATCGGAAGAGCAC$" -A  "^TCGACCCTCTAGTCCAGATCTCACCATCGACCCATAACTTCGTATAGCATACATTATACGAAGTTATGCCGATTAACGATGCGGGTACC...GTCGACAGTCGGCGTCGTGTTACTAAGATCGGAAGAGCGT$" -o <trimmed Read1 fastq> -p <trimmed Read2 fastq> --pair-filter=any --discard-untrimmed <processed Read1 fastq> <processed Read2 fastq>; For libraries primed with the CreON RT primer, the following command was run: cutadapt --cores=0 -a "^TAGTAACACGACGCCGACTGTCGAC...GGTACCCGCATCGTTAATCGGCATAACTTCGTATAATGTATGCTATACGAAGTTATCATTGGTCACGCACGATTTCNNNNNNNNNNNNAGATCGGAAGAGCACACGTCTGAACTCC$" -A  "^GAAATCGTGCGTGACCAATGATAACTTCGTATAGCATACATTATACGAAGTTATGCCGATTAACGATGCGGGTACC...GTCGACAGTCGGCGTCGTGTTACTAAGATCGGAAGAGCGTCGTGTAGGGAAAG$" -o <trimmed Read1 fastq> -p <trimmed Read2 fastq> --pair-filter=any --discard-untrimmed <processed Read1 fastq> <processed Read2 fastq>
Extracted barcode sequences were aligned to the library using bowtie2 (v2.3.5): bowtie2 --very-sensitive -p 8 --no-mixed -x <barcode index> -1 <trimmed Read1 fastq> -2 <trimmed Read2 fastq> -S <alignment SAM file>
Barcode alignments were sorted using Samtools (v1.9): samtools view -bS <alignment SAM file> > <alignment BAM file>; samtools sort <alignment BAM file> > <sorted BAM file>
Reads per barcode were counted using featureCounts (subread v2.0.0): featureCounts -p -B -P -d 10 -D 10 -T 4 -t BC -g barcode -a <.gtf file with barcodes> -R BAM <sorted BAM file> -o <featureCounts outfile>
The alignment output by featureCounts was resorted and indexed: samtools sort <featureCounts sorted BAM file> > <resorted BAM file>; samtools index <resorted BAM file>
Reads were de-duplicated to obtain UMI counts per barcode using umi_tools (v1.0.0): umi_tools count --paired --per-gene --gene-tag=XT -I <resorted BAM file> -S <UMI counts per barcode>
Assembly: NA
Supplementary files format and content: hek_combined_umi_counts.csv: comma-separated values format; each sequencing library from the HEK experiment as columns, with UMI counts for each barcode as rows
Supplementary files format and content: vglut_combined_umi_counts.csv: comma-separated values format; each sequencing library from the in vivo experiment as columns, with UMI counts for each barcode as rows
Library strategy: MPRA

source_name: Brain, cortex

growth_protocol: For HEK experiments, cells were maintained at 5% CO2, 37°C, 95% relative humidity in 1:1 Dulbecco Modified Eagle Medium/Nutrient Mixture F-12 (DMEM/F12) supplemented with 10% fetal bovine serum and 1% Penicillin-streptomycin. Cells were passaged every 2-3 days or once 80% confluent using 0.25% Trypsin-EDTA. All procedures involving animals were approved by the Washington University Institutional Animal Care and Use Committee (protocol no. 20-0097). Mice were group housed and maintained in the vivarium at Washington University in St. Louis. The colony room lighting was a 12–12 h light–dark cycle, and room temperature (~20–22 °C) and relative humidity (50%) were controlled automatically. Standard laboratory diet and water were freely available.

extract_protocol:
For HEK experiments, cells were lifted into plating medium, spun down, and washed in ice-cold PBS containing 100 µg/mL cycloheximide. Cells were resuspended in lysis buffer (20 mM HEPES, 150 mM KCl, 10 mM MgCl2, 0.5 mM DTT, 1% NP-40, 100 µg/mL cycloheximide, 0.02 U/µL rRNasin, 0.02 U/µL Superasin, EDTA-free protease inhibitor) containing 1% NP-40 and triturated by passing ten times through a 27 gauge needle. Lysate was clarified by centrifugation at 20,000 xg. For in vivo experiments, cortex tissue was dissected and washed in ice-cold lysis buffer and homogenized using a hand-operated tissue grinder. Crude lysates were clarified and then incubated on ice following the addition of  1% NP-40 and 30 mM DHPC. Lysates were clarified again by spinning at 20,000 xg. In both experiments, a fraction of the clarified lysate was fractionated using a 10-50% sucrose gradient. Fractions containing 40S subunits, monosomes, and polysomes were harvested for HEK experiments; only monosome and polysome fractions were taken for in vivo experiments. For the HEK experiment, a fraction of lysate was used for Translating Ribosome Affinity Purification (TRAP). Lysates for TRAP were further incubated with 30 mM DHPC before spinning at 20,000xg. Streptavidin-coated magnetic beads (Invitrogen) conjugated to anti-eGFP IgG antibodies (clones 19C8 and 19F7) via biotinylated Protein L were added to lysate supernatants and incubated for 6 hours at 4°C. Beads were washed four times in wash buffer (20 mM HEPES, 350 mM KCl, 10 mM MgCl2, 0.5 mM DTT, 1% NP-40, 100 µg/mL cycloheximide, 0.02 U/µL rRNasin, 0.02 U/µL Superasin, EDTA-free protease inhibitor) before collecting in lysis buffer. RNA from lysates, TRAP immunoprecipitation, 40S fractions, and monosome fractions was extracted using Trizol LS and RNA clean up kit (Zymo). RNA from polysome fractions was extracted by ammonium acetate precipitation and RNA clean up kit (Zymo). DNA from lysate pellets for both HEK and in vivo experiments was extracted using Trizol LS.
For HEK and "CreOFF" (from Cre-ngative cells) RNA-seq libraries, cDNA was prepared by reverse transcription with a "CreOFF RT" primer. For "Cre-ON" (from Cre-positive glutatmetergic neurons) RNA-seq libraries, a "Cre ON RT" reverse transcription primer was used instead. For all RNA libraries, cDNA was amplified using "FivePrime_Sense" and "FivePrime_Antisense" primers followed by an indexing PCR using Illumina TruSeq adapters. DNA-seq for HEK cells and "CreOFF" samples were prepared by first amplifying with "CreOFF RT" and "FivePrime_Antisense" primers followed by an indexing PCR. DNA-seq libraries from "CreON" samples were prepared by first amplifying with "CreON RT"  and "FivePrime_Antisense" primers followed by an indexing PCR.

treatment_protocol: For HEK experiments, cells were transiently transfected with a pooled reporter plasmid library using Lipofectamine 2000. Media was changed 6 hrs post-transfection, and cells were harvested 48 hrs post-transfection. For in vivo experiments, sub-pools of reporters packaged in AAV9 were delivered transcranially at day P1, and cortical tissue was harvested at day P21.

value: Brain, cortex

value: Ef1a:LSL-eGFP-RPl10 +/-,  Vglut1-IRES2-Cre-D +/-


title: CNT_F_S10_06_S6

description: Gene Expression in hippocampus


title: CNT_M_S09_05_S3

description: Gene Expression in hippocampus


title: Vglut, Sublibrary 3, Replicate 6, polysome RNA, CreON

description:
Prepared from polysome RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv

library_source: other

molecule: other


title: OPFR_M_S32-03_S16

description: Gene Expression in hippocampus


title: Vglut, Sublibrary 3, Replicate 5, polysome RNA, CreON


title: Vglut, Sublibrary 3, Replicate 6, 80S RNA, CreON

description:
Prepared from 80S RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 4, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 6, polysome RNA, CreON

description:
Prepared from polysome RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 5, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 6, 80S RNA, CreON

description:
Prepared from 80S RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 6, total RNA, CreON

description:
Prepared from total RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 2, Replicate 5, polysome RNA, CreON


title: Vglut, Sublibrary 2, Replicate 5, total RNA, CreON

description:
Prepared from total RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 4, DNA, CreON


title: Vglut, Sublibrary 3, Replicate 5, total RNA, CreON


title: CNT_M_S08_01_S1

description: Gene Expression in hippocampus


title: Vglut, Sublibrary 3, Replicate 4, polysome RNA, CreON


title: Vglut, Sublibrary 2, Replicate 4, DNA, CreON

description:
Prepared from DNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, amplified with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 4, total RNA, CreON


title: Vglut, Sublibrary 3, Replicate 5, DNA, CreON


title: Vglut, Sublibrary 1, Replicate 6, DNA, CreON

description:
Prepared from DNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, amplified with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 6, total RNA, CreON

description:
Prepared from total RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 5, 80S RNA, CreON

description:
Prepared from 80S RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 5, DNA, CreON


title: Vglut, Sublibrary 3, Replicate 3, polysome RNA, CreON


title: Vglut, Sublibrary 2, Replicate 6, DNA, CreON


title: Vglut, Sublibrary 1, Replicate 5, polysome RNA, CreON

description:
Prepared from polysome RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, reverse transcription primed with CreON primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 5, total RNA, CreON


title: Vglut, Sublibrary 2, Replicate 6, total RNA, CreON


title: Vglut, Sublibrary 2, Replicate 5, DNA, CreON


title: CNT_F_S08_02_S2

description: Gene Expression in hippocampus


title: Vglut, Sublibrary 3, Replicate 3, DNA, CreON


title: Vglut, Sublibrary 2, Replicate 5, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 4, 80S RNA, CreON


title: Vglut, Sublibrary 3, Replicate 2, 80S RNA, CreON


title: Vglut, Sublibrary 3, Replicate 1, 80S RNA, CreON


title: Vglut, Sublibrary 3, Replicate 1, total RNA, CreON


title: Vglut, Sublibrary 2, Replicate 4, polysome RNA, CreON


title: Vglut, Sublibrary 2, Replicate 3, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 4, total RNA, CreON


title: Vglut, Sublibrary 1, Replicate 6, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 3, polysome RNA, CreON


title: Vglut, Sublibrary 1, Replicate 6, polysome RNA, CreON


title: Vglut, Sublibrary 2, Replicate 2, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 1, total RNA, CreON


title: Vglut, Sublibrary 3, Replicate 3, total RNA, CreON


title: Vglut, Sublibrary 1, Replicate 4, 80S RNA, CreON


title: Vglut, Sublibrary 3, Replicate 3, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 1, polysome RNA, CreON


title: Vglut, Sublibrary 3, Replicate 2, DNA, CreON


title: Vglut, Sublibrary 3, Replicate 2, total RNA, CreON


title: Vglut, Sublibrary 3, Replicate 1, DNA, CreON


title: Vglut, Sublibrary 1, Replicate 4, DNA, CreON


title: Vglut, Sublibrary 1, Replicate 4, total RNA, CreON


title: Vglut, Sublibrary 3, Replicate 2, polysome RNA, CreON


title: Vglut, Sublibrary 3, Replicate 1, polysome RNA, CreON


title: Vglut, Sublibrary 1, Replicate 3, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 3, total RNA, CreON


title: Vglut, Sublibrary 2, Replicate 3, DNA, CreON


title: Vglut, Sublibrary 2, Replicate 2, DNA, CreON


title: Vglut, Sublibrary 2, Replicate 2, polysome RNA, CreON


title: Vglut, Sublibrary 1, Replicate 2, 80S RNA, CreON


title: Vglut, Sublibrary 1, Replicate 2, polysome RNA, CreON


title: Vglut, Sublibrary 1, Replicate 2, total RNA, CreON


title: Vglut, Sublibrary 2, Replicate 1, 80S RNA, CreON


title: Vglut, Sublibrary 2, Replicate 2, total RNA, CreON


title: Vglut, Sublibrary 2, Replicate 1, DNA, CreON


title: Vglut, Sublibrary 1, Replicate 1, total RNA, CreON


title: Vglut, Sublibrary 1, Replicate 3, total RNA, CreON


title: Vglut, Sublibrary 1, Replicate 3, polysome RNA, CreON


title: Vglut, Sublibrary 1, Replicate 4, polysome RNA, CreON


title: Vglut, Sublibrary 1, Replicate 1, polysome RNA, CreON


title: Vglut, Sublibrary 1, Replicate 3, DNA, CreON


title: Vglut, Sublibrary 3, Replicate 6, 80S RNA, CreOFF

description:
Prepared from 80S RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 5, DNA, CreOFF

description:
Prepared from DNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, amplified with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 1, DNA, CreON


title: Vglut, Sublibrary 3, Replicate 5, polysome RNA, CreOFF

description:
Prepared from polysome RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 5, total RNA, CreOFF

description:
Prepared from total RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 3, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 4, DNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 6, polysome RNA, CreOFF

description:
Prepared from polysome RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 1, 80S RNA, CreON


title: Vglut, Sublibrary 3, Replicate 6, DNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 6, total RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 6, polysome RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 5, 80S RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 4, 80S RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 6, 80S RNA, CreOFF

description:
Prepared from 80S RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 3, Replicate 4, total RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 2, DNA, CreON


title: Vglut, Sublibrary 2, Replicate 5, total RNA, CreOFF

description:
Prepared from total RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 2, Replicate 5, polysome RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 5, DNA, CreOFF

description:
Prepared from DNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 2, amplified with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 2, Replicate 4, DNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 5, 80S RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 4, polysome RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 4, total RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 6, total RNA, CreOFF

description:
Prepared from total RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 6, polysome RNA, CreOFF

description:
Prepared from polysome RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 5, 80S RNA, CreOFF

description:
Prepared from 80S RNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, reverse transcription primed with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 5, polysome RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 6, DNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 6, 80S RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 6, DNA, CreOFF

description:
Prepared from DNA from Vglut-Cre animal transduced with AAV-packaged  sublibrary 1, amplified with CreOFF primer.
vglut_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 5, total RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 4, polysome RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 5, DNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 3, DNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 3, total RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 3, polysome RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 3, 80S RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 6, total RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 2, DNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 2, total RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 2, 80S RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 1, DNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 1, 80S RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 4, 80S RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 3, polysome RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 3, 80S RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 1, polysome RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 3, DNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 1, total RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 2, total RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 2, DNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 1, 80S RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 1, DNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 2, polysome RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 1, polysome RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 2, 80S RNA, CreOFF


title: Vglut, Sublibrary 2, Replicate 3, total RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 4, 80S RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 4, DNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 3, DNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 4, polysome RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 2, total RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 1, total RNA, CreOFF


title: HEK, Replicate 6, Plasmid_Library

description:
Prepared from plasmid DNA of the complete library
hek_combined_umi_counts.csv

source_name: HEK

value: HEK


title: Vglut, Sublibrary 1, Replicate 3, polysome RNA, CreOFF


title: HEK, Replicate 4, Plasmid_Library


title: Vglut, Sublibrary 1, Replicate 4, total RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 1, polysome RNA, CreOFF


title: Vglut, Sublibrary 3, Replicate 2, polysome RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 3, 80S RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 3, total RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 2, DNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 2, polysome RNA, CreOFF


title: Vglut, Sublibrary 1, Replicate 1, DNA, CreOFF


title: HEK, Replicate 6, TRAP_RNA

description:
Prepared from TRAP RNA from HEK cells transiently transfected with the complete library
hek_combined_umi_counts.csv


title: Vglut, Sublibrary 1, Replicate 1, 80S RNA, CreOFF


title: HEK, Replicate 4, TRAP_RNA


title: HEK, Replicate 1, TRAP_RNA


title: HEK, Replicate 3, TRAP_RNA


title: HEK, Replicate 5, Plasmid_Library


title: HEK, Replicate 2, Plasmid_Library


title: HEK, Replicate 5, TRAP_RNA


title: Vglut, Sublibrary 1, Replicate 2, 80S RNA, CreOFF


title: HEK, Replicate 5, Polysome_RNA

description:
Prepared from Polysome RNA from HEK cells transiently transfected with the complete library
hek_combined_umi_counts.csv


title: HEK, Replicate 3, Polysome_RNA


title: HEK, Replicate 1, Polysome_RNA


title: HEK, Replicate 6, Polysome_RNA


title: HEK, Replicate 4, 40S_RNA

description:
Prepared from 40S RNA from HEK cells transiently transfected with the complete library
hek_combined_umi_counts.csv


title: HEK, Replicate 6, 40S_RNA


title: HEK, Replicate 2, 40S_RNA


title: HEK, Replicate 3, 40S_RNA


title: HEK, Replicate 1, 40S_RNA


title: HEK, Replicate 3, Plasmid_Library


title: HEK, Replicate 4, 80S_RNA

description:
Prepared from 80S RNA from HEK cells transiently transfected with the complete library
hek_combined_umi_counts.csv


title: HEK, Replicate 5, 40S_RNA


title: HEK, Replicate 6, 80S_RNA


title: HEK, Replicate 1, Plasmid_Library


title: HEK, Replicate 6, TotalRNA

description:
Prepared from TotalRNA from HEK cells transiently transfected with the complete library
hek_combined_umi_counts.csv


title: HEK, Replicate 5, 80S_RNA


title: HEK, Replicate 2, 80S_RNA


title: HEK, Replicate 2, TRAP_RNA


title: HEK, Replicate 4, Polysome_RNA


title: HEK, Replicate 4, TotalRNA


title: HEK, Replicate 3, TotalRNA


title: HEK, Replicate 1, TotalRNA


title: HEK, Replicate 6, DNA

description:
Prepared from DNA from HEK cells transiently transfected with the complete library
hek_combined_umi_counts.csv


title: HEK, Replicate 5, DNA


title: HEK, Replicate 4, DNA


title: HEK, Replicate 3, DNA


title: HEK, Replicate 2, DNA


title: HEK, Replicate 1, DNA


title: HEK, Replicate 3, 80S_RNA


title: LOUCY-DMSO, H3K27ac [HiChIP]

status: Public on Sep 25 2023

city: New York

first: Javier

last: Rodriguez Hernaez

state: New York

address: East 30th Street. 8th Floor

institute: NYU Langone Health

data_processing:
library strategy: HiChIP
Read alignment: The Hi-C reads were aligned by the BWA aligner (v0.7.17). Params: "-A1 -B4 -E50 -L0"
Filtering: Mapped reads were filtered by the GenomicTools tools-hic filter command integrated in HiC-bench for known artifacts of the Hi-C protocol.
Valid pairs: We used the accepted intra-chromosomal read-pairs (‘ds.accepted intra’) and filtered out multi-mapped reads (‘multihit’), read-pairs with only one mappable read (‘single sided’), duplicated read-pairs (‘ds.duplicate’), low mapping quality reads (MAPQ < 20), read-pairs resulting from self-ligated fragments.
Normalization: Contact matrices were generated using the HiC-Bench pipeline and ICE correction was performed using the algorithm detailed in Imakaev et. al (2012) publication. For this, we used the matrix-ic functionality built into HiC-bench which uses the ‘mirnylib.numutils’ python library.
Loop calling: We used the HiC valid pairs with the FitHiC tool (v2.0.7) to identify significant interactions. Parameters: resolution=10kb ; q-value <= 0.01; Min. distance=30kb; Max. distance=1Mb.
Virtual4C: HiC-Bench ‘virtual4C’ pipeline was used to compute the interactions of each selected viewpoint in a roll-window fashion. We summed the valid read pairs in a 5 kb area centered at 200 bp bins that covered the area of +-2.5 Mb from the viewpoint (25k bins per viewpoint). The interactions were normalized by the total number of valid pairs of the sample.
Assembly: hg19
Supplementary files format and content: The ‘*_filtered.reg.txt.gz’ files contains all the HiChIP valid intra and inter-chromosomal read pairs.

source_name: LOUCY

extract_protocol:
Chromatin was extracted and processed by Arima HiC+ kit following manufacturer's instructions
HiChIP libraries were prepared using Swift Biosciences Accel-NGS 2S Plus DNA Library Kit

value: LOUCY

tag: cell type

value: ETP-ALL

tag: chip antibody


title: wingDisc-yw-aGFP-CnR-rep2

status: Public on Oct 03 2023

city: Chapel Hill

first: Matthew

last: Niederhuber

address: 250 Bell Tower Dr.

institute: UNC-Chapel Hill

data_processing:
Adapters were trimmed using Bbmap BBDuk (v38.71)
Reads were aligned to the dm6 reference genome with Bowtie2 (v2.3.4.1) and the settings: "--local --very-sensitive-local --no-unal --no-mixed --no-discordant --phred33 -I 10 -X 700"
Aligned reads were filtered on a custom CUT&RUN negative control exclusion list
Aligned reads were quality filtered (q > 5) with Samtools (v1.10)
Duplicate reads were filtered out with Picard (v2.2.4)
Coverage files were generated with deepTools (v2.4.1) and normalized to 1x genomic coverage (RPGC)
Peaks were called with MACS2 (v2.1.2) without a control and using the settings: "--nomodel --nolambda"
Coverage tracks were Z-normalized with a custom R script
Assembly: dm6
Supplementary files format and content: MACS2 narrowPeak files from individual replicates
Supplementary files format and content: Z-normalized bigwig files from individual replicates
Library strategy: Cut & Run

organism: Drosophila melanogaster

source_name: wing

growth_protocol: Animals were raised at 25C.

extract_protocol:
20-22 female third larval wandering wing discs were dissected in 1x PBS per replicate
Libraries were prepared from the “supernatant” fraction using the Takara ThruPLEX DNA-seq kit with unique dual-indexes and following the manufacturer’s specifications but with a modified amplification step (10s annealing, 68C extension)

value: wing

tag: developmental stage

value: third larval wandering

tag: antibody

value: Rockland rabbit anti-GFP 600-401-2156

value: yw


title: CUTLL1_sgMYB, H3K27Ac [HiChIP]

value: T-ALL

value: sgMYB


title: wingDisc-OsaGFP-aGFP-CnR-rep1

value: female

value: Osa-GFP


title: wingDisc-OsaGFP-aGFP-CnR-rep2


title: CUTLL1_sgCTRL, H3K27ac [HiChIP]


title: CUTLL3-untreated, H3K27ac [HiChIP]


title: CUTLL1_DMSO, H3K27ac [HiChIP]


title: HEK, Replicate 1, 80S_RNA


title: HEK, Replicate 5, TotalRNA


title: Thy_2124, H3K27ac [HiChIP]

source_name: thymus

value: thymus

value: Thymocytes


title: CD34_2583, H3K27ac [HiChIP]

source_name: bone marrow

value: bone marrow


title: CD34_2190, H3K27ac [HiChIP]


title: HEK, Replicate 2, TotalRNA


title: ETP-ALL_p15, H3K27ac [HiChIP]

source_name: peripheral blood

value: peripheral blood


title: T-ALL_p11, H3K27ac [HiChIP]


title: ETP-ALL_p16, H3K27ac [HiChIP]

source_name: bone marrow aspirate

value: bone marrow aspirate


title: T-ALL_p06, H3K27ac [HiChIP]


title: T-ALL_p03, H3K27ac [HiChIP]


title: HEK, Replicate 2, Polysome_RNA


title: T-ALL_p01, H3K27ac [HiChIP]


title: KO urea4C-seq BUR-2 bait

status: Public on Aug 01 2023

data_processing:
The 4C libraries were sequenced by Illumina-HiSeq4000, using BUR-1-bait sequence or  BUR-2-bait sequence as custom sequence oligomers. Reads were mapped with Bowtie2 (version 2.3.1), then assigned to HindIII sites with the same utility tools used in Ben Zouari et al., 2019 (4See: A Flexible Browser to Explore 4C Data. Front Genet 10, 1372). These same tools also normalized the counts to per million intrachromosomal reads and removed reads to restriction fragments comprising more than 2% of the total reads (namely the contiguous adjacent restriction fragment corresponding to undigested chromatin during the urea 4C, and a rare (<5) numbers of fragments with extreme PCR duplications). The generated bedGraph files and these were converted to bigWig format using the bedGraphToBigWig program for direct visualization of the peaks on the UCSC browser.
assembly: mm9

extract_protocol:
Single cell suspension of primary thymocyte samples from Satb1 -/- mice (P13) were cross-linked with 1% formaldehyde for 10 min at room temperature. After quenching cross-linking reaction (0.125M Gly at final), these cells were lysed in buffer (4% SDS, 50 mM Tris-HCl [pH 8.0], 10 mM EDTA, 100 mM NaCl) with Protease inhibitor (Roche) and RNase inhibitor and loaded on top of 8 M urea followed by ultracentrifugation in a Beckman TL-100 Ultracentrifuge for 4-5 hrs at 45K rpm (181,000g, g=relative centrifugation force or RCF; TLS55 rotor). Urea-purified crosslinked chromatin from this method was processed for urea-4C-seq basically as described (Kohwi-Shigematsu et al., 2012, Methods, 58, 243-254)  with some modifications. The isolated crosslinked chromatin was very brief sonicated using Branson sonifier (15% power, 10 sec once) and then it was digested with HindIII (10-20 units/mg DNA) overnight at 37oC.  Briefly, after HindIII digestion, HindIII was heat inactivated at 80oC for 20 min, ligation was performed in 1.0 ml with 1X ligation buffer with ligase (600 units) at 16oC for overnight, followed by reverse-crosslinking at 65oC for overnight.  The chromatin DNAs were purified by Phenol-CHCl3 extraction (or spin column, Qiagen), and the purified DNAs were 1-4 mg and were further digested with HaeIII (100-125 units) overnight.  The HindIII-HaeIII digested DNA fragments were treated with Klenow.  The resulting blunt-end HindIII-HaeIII DNA fragments were ligated with our custom double-stranded (ds) Linker DNA with one strand-Biotin modified. The Linker-ligated dsDNA fragments were purified by spin column to remove free Linker.  The Linker-ligated dsDNA fragments were further digested with XhoI at the linker site (next site from HindIII).  Single stranded DNAs were isolated from the XhoI digested DNA fragments by coupling to StreptAvidin Dynabeads (Invitrogen, Dynabeads M-280 streptavidin) followed by incubation in 40ml of 0.15M NaOH for 10 min at 25oC, followed by neutralization. Only single-stranded DNA, containing biotin coupled with streptavidin, were retained on the Dynabeads, releasing the non-biotin modified single-stranded DNA (single strand DNA capture).  The released non-biotin modified single strand DNAs were amplified by PCR (AmpliTaq Gold system, AppliedBiosystems); 10 cycles at 94C for 30”, 55oC for 60” and 72oC for 60”, followed by 15-20 cycles at 94oC for 30”, 65oC for 60”, 72oC for 60”, using the custom Illumina-bait oligomers and Illumina-linker oligomer for library preparations as described below. The PCR amplified DNAs were purified by high and low size selection by beads (SPRIselect, Beckman Coulter).  The purified DNAs were analyzed by Bioanalyzer to confirm their average size of ~300-500 bp with mostly uniform distribution, followed by the Illumina sequencing using custom bait-specific sequence oligomers and Linker specific barcode oligomer.  We used two bait sequences (BUR-1 or BUR-2), both including a HindIII site.   To validate successful experiments, we confirmed that no PCR amplified product was generated by using either the bait oligomer or linker oligomer alone.
The library was constructed by amplification using either Illumina-BUR-1-bait or Illumina-BUR-2 bait primer with Illumina-Linker oligomer.  Taq-Gold polymerase (NEB) was used for PCR by the following condition: 94oC for 5-10min (one cycle), 94C for 30 seconds, 55C for 30 seconds, 72C for 60 seconds (for 5-10 cycles), 94oC for 30 second, 65oC for 30 seconds, 72oC for 60 seconds (for 20 cycles), followed by 72oC for 10 min.  PCR products were purified by Phenol-CHCl3 extraction twice, CHCl3 extraction once, followed by EtOH precipitation (Optional; PCR purification by spin column, Qiagen).  The 4C libraries were sequenced by Illumina-HiSeq4000, using BUR-1-bait sequence and BUR-2-bait sequence as custom sequence oligomers.  For sequencing, the custom barcode sequence oligomer is required instead of the common Illumina barcode sequencing oligomer.
urea 4C-seq

value: thymocytes

value: male /female mix

value: formaldehyde fixation


title: KO urea4C-seq BUR-1 bait


title: wingDisc-yw-aGFP-CnR-rep1


title: WT urea4C-seq BUR-1 bait

extract_protocol:
Single cell suspension of primary thymocyte samples from Satb1+/+  (P13) were cross-linked with 1% formaldehyde for 10 min at room temperature. After quenching cross-linking reaction (0.125M Gly at final), these cells were lysed in buffer (4% SDS, 50 mM Tris-HCl [pH 8.0], 10 mM EDTA, 100 mM NaCl) with Protease inhibitor (Roche) and RNase inhibitor and loaded on top of 8 M urea followed by ultracentrifugation in a Beckman TL-100 Ultracentrifuge for 4-5 hrs at 45K rpm (181,000g, g=relative centrifugation force or RCF; TLS55 rotor). Urea-purified crosslinked chromatin from this method was processed for urea-4C-seq basically as described (Kohwi-Shigematsu et al., 2012, Methods, 58, 243-254)  with some modifications. The isolated crosslinked chromatin was very brief sonicated using Branson sonifier (15% power, 10 sec once) and then it was digested with HindIII (10-20 units/mg DNA) overnight at 37oC.  Briefly, after HindIII digestion, HindIII was heat inactivated at 80oC for 20 min, ligation was performed in 1.0 ml with 1X ligation buffer with ligase (600 units) at 16oC for overnight, followed by reverse-crosslinking at 65oC for overnight.  The chromatin DNAs were purified by Phenol-CHCl3 extraction (or spin column, Qiagen), and the purified DNAs were 1-4 mg and were further digested with HaeIII (100-125 units) overnight.  The HindIII-HaeIII digested DNA fragments were treated with Klenow.  The resulting blunt-end HindIII-HaeIII DNA fragments were ligated with our custom double-stranded (ds) Linker DNA with one strand-Biotin modified. The Linker-ligated dsDNA fragments were purified by spin column to remove free Linker.  The Linker-ligated dsDNA fragments were further digested with XhoI at the linker site (next site from HindIII).  Single stranded DNAs were isolated from the XhoI digested DNA fragments by coupling to StreptAvidin Dynabeads (Invitrogen, Dynabeads M-280 streptavidin) followed by incubation in 40ml of 0.15M NaOH for 10 min at 25oC, followed by neutralization. Only single-stranded DNA, containing biotin coupled with streptavidin, were retained on the Dynabeads, releasing the non-biotin modified single-stranded DNA (single strand DNA capture).  The released non-biotin modified single strand DNAs were amplified by PCR (AmpliTaq Gold system, AppliedBiosystems); 10 cycles at 94C for 30”, 55oC for 60” and 72oC for 60”, followed by 15-20 cycles at 94oC for 30”, 65oC for 60”, 72oC for 60”, using the custom Illumina-bait oligomers and Illumina-linker oligomer for library preparations as described below. The PCR amplified DNAs were purified by high and low size selection by beads (SPRIselect, Beckman Coulter).  The purified DNAs were analyzed by Bioanalyzer to confirm their average size of ~300-500 bp with mostly uniform distribution, followed by the Illumina sequencing using custom bait-specific sequence oligomers and Linker specific barcode oligomer.  We used two bait sequences (BUR-1 or BUR-2), both including a HindIII site.   To validate successful experiments, we confirmed that no PCR amplified product was generated by using either the bait oligomer or linker oligomer alone.
The library was constructed by amplification using either Illumina-BUR-1-bait or Illumina-BUR-2 bait primer with Illumina-Linker oligomer.  Taq-Gold polymerase (NEB) was used for PCR by the following condition: 94oC for 5-10min (one cycle), 94C for 30 seconds, 55C for 30 seconds, 72C for 60 seconds (for 5-10 cycles), 94oC for 30 second, 65oC for 30 seconds, 72oC for 60 seconds (for 20 cycles), followed by 72oC for 10 min.  PCR products were purified by Phenol-CHCl3 extraction twice, CHCl3 extraction once, followed by EtOH precipitation (Optional; PCR purification by spin column, Qiagen).  The 4C libraries were sequenced by Illumina-HiSeq4000, using BUR-1-bait sequence and BUR-2-bait sequence as custom sequence oligomers.  For sequencing, the custom barcode sequence oligomer is required instead of the common Illumina barcode sequencing oligomer.
urea 4C-seq


title: WT urea4C-seq BUR-2 bait

extract_protocol:
Single cell suspension of primary thymocyte samples from Satb1+/+ (P13) were cross-linked with 1% formaldehyde for 10 min at room temperature. After quenching cross-linking reaction (0.125M Gly at final), these cells were lysed in buffer (4% SDS, 50 mM Tris-HCl [pH 8.0], 10 mM EDTA, 100 mM NaCl) with Protease inhibitor (Roche) and RNase inhibitor and loaded on top of 8 M urea followed by ultracentrifugation in a Beckman TL-100 Ultracentrifuge for 4-5 hrs at 45K rpm (181,000g, g=relative centrifugation force or RCF; TLS55 rotor). Urea-purified crosslinked chromatin from this method was processed for urea-4C-seq basically as described (Kohwi-Shigematsu et al., 2012, Methods, 58, 243-254)  with some modifications. The isolated crosslinked chromatin was very brief sonicated using Branson sonifier (15% power, 10 sec once) and then it was digested with HindIII (10-20 units/mg DNA) overnight at 37oC.  Briefly, after HindIII digestion, HindIII was heat inactivated at 80oC for 20 min, ligation was performed in 1.0 ml with 1X ligation buffer with ligase (600 units) at 16oC for overnight, followed by reverse-crosslinking at 65oC for overnight.  The chromatin DNAs were purified by Phenol-CHCl3 extraction (or spin column, Qiagen), and the purified DNAs were 1-4 mg and were further digested with HaeIII (100-125 units) overnight.  The HindIII-HaeIII digested DNA fragments were treated with Klenow.  The resulting blunt-end HindIII-HaeIII DNA fragments were ligated with our custom double-stranded (ds) Linker DNA with one strand-Biotin modified. The Linker-ligated dsDNA fragments were purified by spin column to remove free Linker.  The Linker-ligated dsDNA fragments were further digested with XhoI at the linker site (next site from HindIII).  Single stranded DNAs were isolated from the XhoI digested DNA fragments by coupling to StreptAvidin Dynabeads (Invitrogen, Dynabeads M-280 streptavidin) followed by incubation in 40ml of 0.15M NaOH for 10 min at 25oC, followed by neutralization. Only single-stranded DNA, containing biotin coupled with streptavidin, were retained on the Dynabeads, releasing the non-biotin modified single-stranded DNA (single strand DNA capture).  The released non-biotin modified single strand DNAs were amplified by PCR (AmpliTaq Gold system, AppliedBiosystems); 10 cycles at 94C for 30”, 55oC for 60” and 72oC for 60”, followed by 15-20 cycles at 94oC for 30”, 65oC for 60”, 72oC for 60”, using the custom Illumina-bait oligomers and Illumina-linker oligomer for library preparations as described below. The PCR amplified DNAs were purified by high and low size selection by beads (SPRIselect, Beckman Coulter).  The purified DNAs were analyzed by Bioanalyzer to confirm their average size of ~300-500 bp with mostly uniform distribution, followed by the Illumina sequencing using custom bait-specific sequence oligomers and Linker specific barcode oligomer.  We used two bait sequences (BUR-1 or BUR-2), both including a HindIII site.   To validate successful experiments, we confirmed that no PCR amplified product was generated by using either the bait oligomer or linker oligomer alone.
The library was constructed by amplification using either Illumina-BUR-1-bait or Illumina-BUR-2 bait primer with Illumina-Linker oligomer.  Taq-Gold polymerase (NEB) was used for PCR by the following condition: 94oC for 5-10min (one cycle), 94C for 30 seconds, 55C for 30 seconds, 72C for 60 seconds (for 5-10 cycles), 94oC for 30 second, 65oC for 30 seconds, 72oC for 60 seconds (for 20 cycles), followed by 72oC for 10 min.  PCR products were purified by Phenol-CHCl3 extraction twice, CHCl3 extraction once, followed by EtOH precipitation (Optional; PCR purification by spin column, Qiagen).  The 4C libraries were sequenced by Illumina-HiSeq4000, using BUR-1-bait sequence and BUR-2-bait sequence as custom sequence oligomers.  For sequencing, the custom barcode sequence oligomer is required instead of the common Illumina barcode sequencing oligomer.
urea 4C-seq


title: LS180-gfp-3

status: Public on Jul 01 2023

city: Amsterdam

first: Nicolas

last: Léveillé

address: Meibergdreef 9

country: Netherlands

institute: Amsterdam UMC

description: 201215_CRL1kdCTNkd_GencBroad_featureCounts_genes.txt

data_processing:
library strategy: GRO-seq
GROseq data was analysed by mapping reads to the human genome and quantifying the expression of annotated coding genes and lncR NAs as well as performing de novo detection of enhancer RNAs (eRNAs)
Assembly: hg19
Supplementary files format and content: For RNA-seq data we used Hisat2 (2.0.4) for alignment and Subread (1.6.4) for identifying gene counts (Liao et al., 2014; Pertea et al., 2016). For the GRO-seq datawe used NRSA (Wang et al., 2018)

extract_protocol:
For RNA sequencing, total RNA was isolated using Trizol-Chloroform extraction (see section RNA isolation). RNA integrity was assessed with the Tapestation RNA ScreenTape Assay (Agilent Technologies).
libraries were prepared using the KAPA mRNA Hyperprep with RiboErase (Roche) according to manufacturer’s recommendation from 1 ug total RNA. Barcoded samples were multiplexed sequenced using single-end 50 bp reads on the Illumina HiSeq4000 platform.

value: knockdown GFP


title: LS180-ctn-3

value: knockdown CTNNB1


title: LS180-gfp-2


title: LS180-sh40-3

value: knockdown CRL1-sh2


title: LS180-sh40-2


title: LS180-sh12-1

value: knockdown CRL1-sh1


title: LS180-sh12-3


title: LS180-sh40-1


title: Thy_2123, H3K27ac [HiChIP]


title: LS180-ctn-2


title: LS180-sh12-2


title: LS180-gfp-1


title: T84-GFP2

description: count.txt


title: LS180-ctn-1


title: T84-CTN1

description: count.txt

value: β-catenin knockdown


title: T84-CTN2

description: count.txt


title: T84-GFP1

description: count.txt


title: HCT116_GFP2

description: count.txt


title: HCT116_CTN2

description: count.txt


title: HCT116_GFP1

description: count.txt


title: ETP-ALL_p04, H3K27ac [HiChIP]

value: ETP-ALL


title: LS411N-GFP2

description: count.txt


title: LS411N-GFP1

description: count.txt


title: LS411N-CTN1

description: count.txt


title: LS180-GFP2

description: count.txt


title: LS180-GFP1

description: count.txt


title: LS180-CTN2

description: count.txt


title: LS180-CTN1

description: count.txt


title: CL40-CTN2

description: count.txt


title: CACO2-CTN1

description: count.txt


title: CL40-GFP2

description: count.txt


title: CACO2-GFP2

description: count.txt


title: CL40-GFP1

description: count.txt


title: CACO2-CTN2

description: count.txt


title: RC511-GFP2

description: count.txt


title: CACO2-GFP1

description: count.txt


title: RC511-GFP1

description: count.txt


title: RC511-CTN1

description: count.txt


title: SW1463-GFP2P

description: count.txt


title: SW1463-CTN2P

description: count.txt


title: CL40-CTN1

description: count.txt


title: RC511-CTN2

description: count.txt


title: SW1463-CTN1P

description: count.txt


title: SW1463-GFP1P

description: count.txt


title: SW48-CTN2P

description: count.txt


title: SW48-CTN1P

description: count.txt


title: HCT116_CTN1

description: count.txt


title: LS411N-CTN2

description: count.txt


title: SW48-GFP2P

description: count.txt


title: SW48-GFP1P

description: count.txt


title: MDST8-GFP1P

description: count.txt


title: MDST8-GFP2P

description: count.txt


title: MDST8-CTN1P

description: count.txt


title: LOVO-GFP2P

description: count.txt

source_name: LOVO

value: LOVO


title: LOVO-GFP1P

description: count.txt


title: MDST8-CTN2P

description: count.txt


title: LOVO-CTN1P

description: count.txt


title: LOVO-CTN2P

description: count.txt


title: HUTU80-GFP2P

description: count.txt


title: HT55-CTN1P

description: count.txt


title: HUTU80-GFP1P

description: count.txt


title: HT55-CTN2P

description: count.txt


title: HT55-GFP1P

description: count.txt


title: GTG7d-GFP2P

description: count.txt


title: GTG7d-CTN2P

description: count.txt


title: GTG7d-CTN1P

description: count.txt


title: Vglut, Sublibrary 2, Replicate 1, total RNA, CreOFF

value: Brain, cortex

value: Ef1a:LSL-eGFP-RPl10 +/-,  Vglut1-IRES2-Cre-D +/-


title: GTG7d-GFP1P

description: count.txt


title: COLO320-GFP2P

description: count.txt


title: COLO320-CTN1P

description: count.txt


title: COLO320-GFP1P

description: count.txt


title: COLO320-CTN2P

description: count.txt