title: RNA_RC4_d108h_DMSO

description: processed data file: RNA_RC4_RG9_d108h_counts.txt

source_name: RNAseq, clone C4

value: CAST/Ei x 129/Sv/Jae


title: RNA_RG9_d108h_dTAG13

status: Public on Mar 15 2021

type: SRA

city: Boston

first: Andrea

middle: June

last: Kriz

state: MA

address: 185 Cambridge St

department: Molecular Biology

country: USA

institute: Massachusetts General Hospital, Harvard Medical School

library_source: transcriptomic

data_processing:
ChIP-seq and Calibrated ChIP-seq: For uncalibrated ChIP-seq, input and ChIP-seq reads were aligned to the 129S1/SvJm (mus) and CAST/Eih (cas) genome as previously described (Kung et al., 2015; Minajigi et al., 2015). PCR duplicates were removed and uniquely mapped reads used for further analysis. Fpm-normalized bigwig tracks were generated for all uniquely mapped (comp), mus-specific (mus), and cas-specific (cas) reads. Heatmaps and metaplots were generated using deepTools functions computeMatrix and plotHeatmap with the reference point as the summit of the called ChIP peaks.Calibrated ChIP-seq analysis was performed as previously described (Hu et al., 2015). Briefly, paired-end reads were aligned to a concatenated mouse and human genome (mm9 + hg19) using Bowtie2 with ?--no-mixed? and ?--no-discordant? options. PCR duplicates were filtered out with SAMBAMBA (Tarasov et al., 2015). Uniquely aligned mouse reads were normalized to human spike-in by subsampling mm9 reads so that total number of hg19 reads was matched between samples and bigWig tracks generated from normalized samples. For calibration of RAD21 cChIP between RAD21-FKBPdegron clones, only reads within hg19 peaks (hg19 tracks combined) were counted for normalization purposes. Subsampling factors were further adjusted based on ratio of hg19 to mm9 reads in matched input samples. After normalization factor was determined, paired end reads were aligned to mus and cas genomes as previously described, except using Bowtie2 with the same parameters as above (with ?--no-mixed? and ?--no-discordant?) instead of Novoalign. PCR duplicates were filtered out with SAMBAMBA and allelic reads normalized using the same normalization factor as above.Input subtracted H3K27me3 calibrated ChIP coverage profiles were generated with SPP (Kharchenko et al., 2008) with smoothing using 1kb windows recorded every 500 bp, as previously described (Simon et al., 2013). For input subtraction, inputs from DMSO and dTAG-13 treated samples were combined to ensure equal profiles were subtracted from each calibrated IP.Heatmaps and metaplots were generated using deepTools functions computeMatrix and plotHeatmap with the reference point as the summit of the called ChIP peaks. Plots of calibrated H3K27me3 signal over upregulated or downregulated genes were calculated using ?scale-regions? with the entire gene scaled to 3kb. To compare the allelic densities of H3K27me3 over different classes of X-linked genes, we computed average densities over gene bodies using multiBigwigSummary from deepTools with default parameters. Non-expressed genes were defined as X-linked genes have 0 RPKM in at least one RNA-seq replicate of DMSO control. Genes subject to XCI were identified as the set of genes have allelic information (see RNA-seq analysis above), with escapees, Xist and Tsix excluded. Escapees were as defined previously (Wang et al., 2018). To compare allelic densities of H3K27me3 over S1 and S2 compartments, we defined S1 compartments as regions having positive value in the first Principal Component calculated in the WAPL-FKBPdegron DMSO control days 4-7 sample and S2 compartments as regions have negative value. S1 and S2 compartments were then binned into 100kb windows using bedtools makewindows, excluding windows that overlapped with unmappable regions as defined previously (Wang et al., 2018). We then we computed average densities over these 100kb bins using multiBigwigSummary from deepTools with default parameters.
RNA-seq: RNA-seq reads were aligned allele-specifically to the 129S1/SvJm (mus) and CAST/Eih (cas) genomes using TopHat2 (Kim et al., 2013) as previously described (Kung et al., 2015; Minajigi et al., 2015). PCR duplicates were removed and unique reads mapping to exons of each gene quantified by Homer (Heinz et al., 2010). DESeq 2 was used to call differentially expressed genes with a cut off of adjusted p-value < 0.1. An expression threshold of FKPM > 0.5 was also applied. Expressed genes with a minimum of 13 allelic-specific reads in at least one sample were counted as genes with allelic information (Colognori et al., 2019). For a given gene, percent mus was calculate as the percentage of allelic reads that were mus-specific (mus / (mus+cas) reads).
in situ Hi-C: Reads were trimmed using cutadapt with the options --adapter=GATCGATC (MboI ligation junction) and --minimum-length=20. Reads of each pair were individually mapped to the mus (Xi) and cas (Xa) reference genomes using novoalign and merged into Hi-C summary files and filtered using HOMER as previously described (Minajigi et al., 2015). Hi-C contact maps were generated from reads in the filtered HOMER mus and cas tag directories using the ?pre? command of Juicer tools with the genomeID mm9 (Durand et al., 2016b). The resulting Hi-C contact maps in .hic format were visualized and normalized with the ?Coverage Square Root? option in Juicebox version 1.8.8 (Durand et al., 2016a). Filtered reads from separate RAD21-FKBPdegron clones were combined before generating Hi-C contact maps.
CHART-seq: Input and ChIP-seq reads were aligned to the 129S1/SvJm (mus) and CAST/Eih (cas) genome as previously described (Kung et al., 2015). PCR duplicates were removed and uniquely mapped reads used for further analysis. Fpm-normalized bigwig tracks were generated for all uniquely mapped (comp), mus-specific (mus), and cas-specific (cas) reads. CHART libraries were sampled to equal read depth and input subtracted CHART coverage profiles were generated with SPP (Kharchenko et al., 2008) with smoothing using 1kb windows recorded every 500 bp, as previously described (Simon et al., 2013). After scaling, we computed base 2 logarithm of the ratio of the coverage of DMSO control divided by that of dTAG-13 using bigWigCompare from deepTools with --binSize 1000 and default parameters otherwise.
PRO-seq: Paired-end reads were aligned to a concatenated mouse and fly genome (mm9 + dm6) using Bowtie2 with ?--no-mixed? and ?--no-discordant? options. DMSO and dTAG-13 samples for each clone were normalized to drosophila spike-in by subsampling mm9 reads so that total number of dm6 reads was matched between samples. For RAD21-FKBPdegron clone G9, downsampling factors corresponded to library size (i.e., were equivalent to downsampling based on total number of reads). For RAD21-FKBPdegron clone C4, downsampling factors were 1 for DMSO and 0.73 for dTAG-13 although library sizes were similar, indicating that there were overall lower levels of nascent transcription in the dTAG-13 sample. Duplicates were not removed to preserve reads corresponding to TSS and promoter proximal PolII positions. Reads mapping to genes (full gene bodies) were quantified by Homer (Heinz et al., 2010), removing miRNA, snoRNA and scaRNAs. DESeq 2 was used to call differentially expressed genes with a cut off of adjusted p-value < 0.1. For analyses on all genes, no FPKM threshold was applied, but genes were required to have at least 1 count in one sample. An expression threshold of FKPM > 0.5 was applied for analyses on expressed genes. Log2FoldChange and log2baseMean from DESeq2 were plotted for indicated classes of genes. H3K27me3 genes were defined as genes overlapping H3K27me3 cChIP-seq peaks in their respective samples (DMSO and dTAG-13 peaks combined). H3K27me3 loop genes were defined as H3K27me3 genes overlapping H3K27me3-bound loop anchors called in RAD21-FKBPdegron dTAG-13 Hi-C maps (i.e., cohesin-independent H3K27me3 anchored loops). For overlap with RAD21 loop anchors, loops were called in respective DMSO Hi-C maps.For allelic analysis, PRO-seq reads not mapping to the fly genome (dm6) were aligned to mus and cas genomes using Bowtie2 with the same parameters as above (?--no-mixed? and ?--no-discordant?) and downsampled using the same factors as calculated above. Expressed genes with a minimum of 13 allelic-specific reads in at least one sample were counted as genes with allelic information (Colognori et al., 2019). For CDPs, normalized mus and cas counts divided by gene length were plotted.
Genome_build: mm9
Supplementary_files_format_and_content: ChIP-seq: bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bigWig files are fpm normalized and H3K27me3 bigWig files are additionally input normalized.
Supplementary_files_format_and_content: Calibrated ChIP-seq: bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bigWig files are normalized to spike-in HEK293 control and H3K27me3 bigWig files are additionally input normalized.
Supplementary_files_format_and_content: RNA-seq: Strand specific (cwig, Crick strand, wwig, Watson strand) bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bw files are fpm normalized. Raw exonic reads in each RNA-seq library were quantified and summarized in .txt files.
Supplementary_files_format_and_content: PRO-seq: Strand specific (cwig, Crick strand, wwig, Watson strand) bigWig (bw) files for all reads (.bw), regardless of whether they are allele-specific or not, cas-specific (.cas.bw) and mus-specific (.mus.bw). All bw files are downsampled according to Drosophila spike-in. Gene body counts in each PRO-seq library were quantified and summarized in .txt files.
Supplementary_files_format_and_content: In situ Hi-C: Compressed Hi-C matrix (.hic) files for all contacts genome-wide (.hic), regardless of whether they are allele-specific or not, cas-specific chrX (chrX.cas.his) contacts and mus-specific chrX (chrX.mus.hic) contacts. File can be loaded onto Juicebox to view either raw or normalized Hi-C interaction matrices at 5kb to 2.5Mb resolutions.

molecule: polyA RNA

organism: Mus musculus

source_name: RNAseq, clone G9

growth_protocol: Undifferentiated mESCs were grown on γ-irradiated MEF feeders in medium containing DMEM, high glucose, GlutaMAX supplement, pyruvate, 15% Hyclone FBS (Sigma), 25 mM HEPES pH 7.2-7.5, 1x MEM non-essential amino acids, 1x Pen/Strep, 0.1 mM βME, and 500 U/mL ESGRO recombinant mouse Leukemia Inhibitory Factor (LIF) protein (Sigma, ESG1107) at 37°C with 5% CO2. To induce differentiation in EBs, mESCs were trypsinized and resuspended in the same media without LIF. For the first 3 days of differentiation, EBs were grown in suspension. On day 4 and subsequent days, EBs were grown on gelatinized plates until harvested at day 7 or day 10.

extract_protocol:
ChIP-seq: ChIP-seq was performed as described previously with minor modifications (Del Rosario et al., 2019). Briefly, cells were trypsinized, resuspended in 1 million cells/mL of media and crosslinked with 1% formaldehyde for 10 min at room temperature with rotation. Cells were then quenched with 0.2 M glycine for 5 min. Crosslinked cells were washed once with cold PBS, pelleted, and snap frozen in an EtOH/dry ice bath. Cell pellets were stored at -80C until further processing. 5-10 million cross-linked cells per IP were thawed on ice and resuspended in 300ul Lysis Buffer 1 (25mM HEPES pH 7.2-7.5, 25mM EDTA, 0.5% SDS). Nuclei were sonicated (Qsonica Q800 Sonicator) in polystyrene tubes at 40% power setting, 30s ON/30s OFF for a total sonication (ON) time of 6 min. After sonication, 1.2 mL of Lysis Buffer 2 (25mM HEPES pH 7.2-7.5, 187.5mM NaCl, 1.25% Triton X-100, 0.625% Na-deoxycholate, 6.25% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail) was added. 30ul of Dynabeads Protein G (Thermo Fisher Scientific) per IP were washed 3 times with ChIP Wash Buffer 1 (25mM HEPES pH 7.2-7.5, 150mM NaCl, 5mM EDTA, 1% Triton X-100, 0.5% Na-deoxycholate, 5% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail) and resuspended in 60ul ChIP Wash Buffer 1. Each IP was blocked with 20ul washed beads for 2 hours at 4C with rotation. The other 40ul of washed beads were incubated with antibody (5ul of RAD21 antibody or 2ul of H3K27me3 antibody/IP) for 2 hours at 4C. 5% input was taken from blocked lysates. Beads conjugated to antibody were added to blocked lysates and IP allowed to proceed overnight at 4C with rotation. IPs were then washed 3 times with cold ChIP Wash Buffer 1, 3 times with cold ChIP Wash Buffer 2 (25mM HEPES pH 7.2-7.5, 300mM NaCl, 5mM EDTA, 1% Triton X-100, 0.5% Na-deoxycholate, 5% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail). And 1 time with cold TENG (50mM Tris-HCl pH 8, 1mM EDTA, 50mM NaCl, 5% glycerol, 1x cOmplete EDTA-free protease inhibitor cocktail). All washes were performed by inverting tubes and setting aside for ~2 minutes per wash. IPs were then resuspended in 500ul TENG with 100ug RNAse A (Thermo Fisher Scientific) and incubated for 30 min at 37C. IPs were then washed 2 times with cold ChIP Wash Buffer 2 and 2 times with cold TENG without protease inhibitors. IPs were resuspended in 200ul elution buffer (50mM Tris-HCl pH 7.5, 10mM EDTA, 1% SDS) and incubated 15 min at 65C. Eluate was then transferred to a new tube and 198ul TE buffer (Ambion) and 40ug Proteinase K (Sigma) added. Inputs were thawed, TE buffer added to equalize volume to IPs and 200ul elution buffer and 40ug Proteinase K added. Inputs and IPs were then incubated for 2h at 45C and crosslinks reversed overnight at 65C. DNA was purified with phenol-chloroform extraction and resuspended in 50ul TE. Efficiency of ChIP was assayed via qPCR, with samples displaying over 10-fold enrichment of positive regions over negative control prepared for sequencing. Libraries were prepared from ~500ng of input or IP DNA using the NEBNext® Ultra? II DNA Library Prep Kit for Illumina® (NEB) according to kit instructions. Experiments were performed in two biological replicates. Input and ChIP-seq libraries were sequenced on Illumina HiSeq, generating 20-50 million paired end 50 or 150 nucleotide reads per sample.
Calibrated ChIP-seq: Calibrated ChIP-seq was performed using the same protocol as ChIP-seq except that 50,000 HEK293 cells per 5 million mouse cells were mixed in at the initial lysis step. Cells were counted and equal numbers of cells used per IP.
in situ Hi-C: As described in Rao et al. (2014), 3-5 million cells were crosslinked with 1% formaldehyde for 10min. Crosslinked cell pellets were lysed with 300uL ice-cold HiC lysis buffer (10mM Tris HCl pH 8, 10mM NaCl, 0.2% Igepal CA630) with protease inhibitors (Roche). Cell suspension was incubated for 30min at 4C with rotation, then pelleted at 2500xg for 5min. Pelleted nuclei were then washed with 500ul ice-cold HiC lysis buffer and resuspended in 50uL 0.5% SDS solution and incubated at 62C for 10min. 145ul H2O and 25ul 10% Triton X-100 were added to quench SDS and nuclei incubated at 37C for 15min. 26.1ul of 10X NEB Buffer 2 and 100U of MboI restriction enzyme were added and nuclei digested overnight at 37C with rotation. MboI was then heat activated at 62C for 20min, then nuclei cooled to room temperature. To fill in MboI fragment overhangs and mark DNA ends with biotin, 50ul of fill-in master mix was added (37.5ul 0.4mM biotin-14-dATP, 1.5ul 10mM dCTP, 1.5ul 10mM dGTP, 1.5ul 10mM dTTP, 8ul 5U/ul DNA Polymerase I, Large (Klenow) Fragment) and mixed by pipetting. Nuclei were then incubated at 37C for 1h with rotation. To ligate biotinylated DNA ends together, 900ul ligation master mix was added (649ul H2O, 120ul 10X NEB T4 ligase buffer, 100ul 10% Triton X-100, 6ul of 20mg/ml BSA, 25ul of 400U/ul T4 DNA ligase). Reaction was mixed by inverting and incubated at room temperature for 4h with rotation. Reaction was then centrifuged at 1700xg for 5min at room temperature. Pelleted nuclei were resuspended in 750ul TES buffer (1mM EDTA, 10mM Tris-HCl pH 8, 0.5% SDS), 50ul Proteinase K (20mg/ml) added and incubated at 55C for 1h with rotation. To reverse crosslinks, 88.9ul 3M NaCl was added and crosslinks reversed by incubated overnight at 65C with shaking (Eppendorf thermomixer, 950rpm). Samples were cooled to room temperature and DNA extracted with phase-lock gel tubes (5Prime). RNA was digested from purifying DNA by adding 2ul RNAse A (20mg/ml) and incubating at 37C for 1h. DNA was then extracted with phase-lock gel tubes (5Prime) and resuspended in 130ul 1X Tris buffer (10mM Tris-HCl, pH 8). To ensure complete resuspension, DNA was then incubated at 37C for 15min. DNA was then sonicated to a size of 300-500bp using a Covaris with the following parameters: 140W, 10% duty cycle, 200 bursts/cycle, 100s. Samples were then size selected to 300-500bp using AMPure XP beads and eluted in 300ul 1X Tris buffer. To pulldown biotinylated ligation products, 30ul (per sample) of MyOne Streptavidin C1 beads were washed with 400ul of 1X Tween washing buffer (5mM Tris-HCl pH 7.5, 0.5mM EDTA, 1M NaCl, 0.05% Tween 20) and resuspended in 300ul 2X binding buffer (10mM Tris-HCl pH 7.5, 1mM EDTA, 2M NaCl). Beads were then added to purified DNA and sample incubated at room temperature for 15min with rotation. Beads were washed twice with 600ul 1X Tween Washing buffer. Subsequent library preparation steps were then done directly on beads.
CHART-seq: Xist CHART-seq was performed as previously described (Wang et al., 2018). Briefly, cells were trypsinized, resuspended in 1 million cells/mL of media and crosslinked with 1% formaldehyde for 10 min at room temperature with rotation. Cells were then quenched with 0.2 M glycine for 5 min. Crosslinked cells were washed once with cold PBS, pelleted, and snap frozen in an EtOH/dry ice bath. Cell pellets were stored at -80C until further processing. 25 million cross-linked cells per CHART were thawed on ice and resuspended in 1 mL cold sucrose buffer. (10 mM HEPES pH 7.5, 0.3 M sucrose, 1% Triton X-100, 100 mM potassium acetate, 0.1 mM EGTA) supplemented with 0.5 mM spermidine, 0.15 mM spermine, 1 mM DTT, 1x protease inhibitor cocktail (Sigma, P8340), 10 U/mL RNase inhibitor (Roche). The cell suspension was rotated for 10 min at 4°C, after which it was diluted with additional 2 mL cold sucrose buffer. This was added to a pre-chilled 15-mL glass Wheaton dounce tissue grinder (Thermo Fisher Scientific) that had been pre-rinsed once with RNaseZAP RNase decontamination solution (Thermo Fisher Scientific), three times with DEPC-treated water, and once with 2 mL sucrose buffer. Release of nuclei was accomplished by douncing cells 20 times with a tight pestle, and verified by staining with Trypan Blue Solution (Thermo Fisher Scientific). The nuclear suspension was then layered atop a cushion of 7.5 mL cold glycerol buffer (10 mM HEPES pH 7.5, 25% glycerol, 1 mM EDTA, 0.1 mM EGTA, 100 mM potassium acetate) supplemented with 0.5 mM spermidine, 0.15 mM spermine, 1 mM DTT, 1x cOmplete EDTA-free protease inhibitor cocktail (Roche), and 5 U/mL RNase inhibitor, and centrifuged at 1500 g for 10 min at 4°C. The purified nuclear pellet was resuspended in 6 mL PBS and further cross-linked with 3% formaldehyde for 30 min at room temp with rotation. Afterward, nuclei were pelleted by centrifugation at 1,000 g for 5 min at 4°C and washed three times with cold PBS. Nuclei were resuspended in in 1 mL cold nuclear extraction buffer (50 mM HEPES pH 7.5, 250 mM NaCl, 0.1 mM EGTA, 0.5% sarkosyl, 0.1% sodium deoxycholate, 5 mM DTT, 10 U/mL RNase inhibitor) and incubated for 10 min at 4°C with rotation. Nuclei were then centrifuged at 400 g for 5 min at 4°C and resuspended in 230 uL cold sonication buffer (50 mM HEPES pH 7.5, 75 mM NaCl, 0.1 mM EGTA, 0.5% sarkosyl, 0.1% sodium deoxycholate, 0.1% SDS, 5 mM DTT, 10 U/mL RNase inhibitor) to a final volume of ?270 uL. Nuclei were sonicated for 5 min at 4°C using a Covaris E220e (140W peak incident power, 10% duty factor, 200 cycles/burst). Sonicated chromatin was centrifuged at 16,000 g for 20 min at 4°C to remove debris. The supernatant (?220 uL) was mixed with 110 uL sonication buffer to a final volume of ?330 uL, which was split into two CHART reactions (Xist capture using antisense probes and control capture using sense probes). For two CHART reactions, 600 uL MyOne Streptavidin C1 beads (Thermo Fisher Scientific) were used, which were pre-washed with 1 mL DEPC-treated water, blocked with 600 uL blocking buffer (33% sonication buffer, 67% 2x hybridization buffer, 500 ng/uL yeast RNA [Thermo Fisher Scientific], 1% BSA) at room temp for 1 hr, washed with 1 mL DEPC-treated water again, and resuspended in 610 uL 1x hybridization buffer (33% sonication buffer, 67% 2x hybridization buffer [see below for composition]). For each CHART, 160 uL chromatin was mixed with 320 iL 2x hybridization buffer (50 mM Tris pH 7.0, 750 mM NaCl, 1% SDS, 1 mM EDTA, 15% formamide, 1 mM DTT, 1 mM PMSF, 1x cOmplete EDTA-free protease inhibitor cocktail, 100 U/mL RNase inhibitor) and pre-cleared with 60 uL pre-blocked MyOne Streptavidin C1 beads at room temp for 1 hr with rotation. Pre-cleared chromatin was separated from beads, with 1% saved as ?input? sample. The remaining pre-cleared chromatin (?500 uL) was mixed with 36 pmol of either antisense (Xist-targeting) or sense (control) biotinylated capture probes. We used a pool of 9 capture probes, which were selected from the 11 probes described previously (Simon et al., 2013), excluding probes X.3 and X.8. Hybridization was carried out at 37°C for 4 hr followed by incubation with 240 uL blocked MyOne Streptavidin C1 beads at 37°C for 1 hr. The beads were washed once with 1x hybridization buffer (33% sonication buffer, 67% 2x hybridization buffer) at 37°C for 10 min, five times with wash buffer (10 mM HEPES pH 7.5, 150 mM NaCl, 2% SDS, 2 mM EDTA, 2 mM EGTA, 1 mM DTT) at 37°C for 5 min, and twice with elution buffer (10 mM HEPES pH 7.5, 150 mM NaCl, 0.5% NP-40, 3 mM MgCl2, 10 mM DTT) at 37°C for 5 min. 10% of the final wash was saved as an ?RNA capture? sample to estimate the efficiency of Xist RNA capture afterward by qRT-PCR. CHART-enriched DNA was eluted twice in 200 uL elution buffer supplemented with 5 U/uL RNase H (New England BioLabs) at 37°C for 30 min. An ?input? sample and CHART DNA were treated with 0.5 mg/mL RNase A (Thermo Fisher Scientific) at 37°C for 1 hr and then with 1% SDS, 10 mM EDTA, and 0.5 mg/mL proteinase K (Sigma) at 55°C for 1 hr. Reversal of crosslinks was performed by addition of 150 mM NaCl (final 300 mM) and incubation at 65°C overnight. DNA was purified by phenol-chloroform extraction and further sheared to < 500-bp fragments using Covaris E220e (140W peak incident power, 10% duty factor, 200 cycles/burst). Sonicated DNA was purified using 1.8x Agencourt AMPure XP beads (Beckman Coulter). Libraries were sequenced on Illumina, generating ?50 million 150-nt paired-end reads per sample
RNA-seq: Total RNA was extracted using TRIzol Reagent (Thermo Fisher Scientific) and mRNA isolated using the NEBNext Poly(A) Magnetic Isolation Module (NEB) according to the kit instructions. Libraries were sequenced on the Illumina HiSeq2500 (rapid run), generated 30-50 million 50 nt paired end reads per sample.
PRO-seq: PRO-Seq library construction was performed by the Nascent Transcriptomics Core at Harvard Medical School, Boston, MA. Aliquots of frozen (-80°C) permeabilized cells were thawed on ice and pipetted gently to fully resuspend. An aliquot was removed and diluted 1:10 in 1xPBS for cell counting using an automatic counter (Biorad TC-20) with counts spot checked by manual recount using a hemocytometer as needed. An aliquot of each dilution was also stained with Trypan Blue and counting repeated to determine permeabilization efficiency. For each sample, 1 million permeabilized cells were used for nuclear run-on. For normalization, 50,000 permeabilized Drosophila S2 cells were added to each sample of 1 million cells. Nuclear run on assays and library preparation were then performed essentially as described [Elrod, N.D., et al. (2019) Molecular Cell 76: 738-752.e7] with modifications noted below. The nuclear run-on buffer was prepared as 4X stock (20mM Tris (pH 8), 20mM MgCl2, 2mM DTT, 600mM KCl, 40uM/ea biotin-11-NTPs (Perkin Elmer), 20U SuperaseIN (Thermo)). The 4X stock was mixed 1:1 with 2% sarkosyl (Sigma) to yield 2X complete run-on mix. The 3' adapter (RNA oligo: 5'P-GAUCGUCGGACUGUAGAACUCUGAAC-3'InvdT) was pre-adenylated prior to use (5' DNA adenylation kit, NEB, according to the manufacturer's instructions). Adenylated oligo was purified by ethanol precipitation, resuspended in water and quality was modification confirmed by electrophoresis on a 15% TBE-Urea gel (Novex) next to appropriate controls. The adapter concentration was adjusted to 10uM and 1uL was used for each ligation which used T4 RNA ligase 2, truncated KQ (NEB) with 15% PEG-8000 in the reaction mix. Reactions were incubated at 16°C overnight. Each ligation reaction was then diluted with 180uL of betaine binding buffer (1.42g of betaine brought to 10mL with binding buffer and sterile filtered) containing 1uL of 100uM blocking oligo (TCCGACGATCCCACGTTCCCGTGG/3InvdT/). The blocking oligo is complementary to and overhangs the 3' end of the 3' adapter and reduces 5'/3' adapter dimer formation during the 5' adapter ligation step. The blocking oligo has modifications at its 3? end, to prevent extension by reverse transcriptase. Blocking oligo was also included in final wash solution prior to the 5' adapter ligation and in the ligation reaction itself, both at 3-fold molar excess over the initial amount of 3' adapter (i.e. 1uL of 30uM per reaction). The 5' adapter ligation was performed as in Elrod except PEG-8000 was increased to 15%. Libraries were prepared using the NEBNext® Ultra? II DNA Library Prep Kit for Illumina® (NEB) according to kit instructions. PRO-seq: After reverse transcription, cDNA was immediately amplified for 5-cycles (preCR step). A PCR cocktail consisting of NEBNext Ultra II Q5 master mix (NEB) and Illumina TruSeq PCR primers (RP-1, common; and RPI-X, indexing) was added to each cDNA and amplification was done according to manufacturer's suggested 2-step cycling conditions for NGS applications. To determine optimal library amplification, the preCRs were thawed on ice and an aliquot serially diluted. Each dilution was mixed with a PCR cocktail (Q5 DNA polymerase with optional high GC enhancer) containing 10uM/ea of Illumina universal P5 and P7 primers and amplified 15-cycles. The reactions were electrophoresed on a 2.2% agarose gel (SeaKem) cast with SYBR gold (Thermo) and analyzed to determine the optimal number of cycles for final amplification. preCRs were transferred from ice to a pre-heated thermal cycler block and amplified for the appropriate number of additional cycles to reach the total determined in the test amplification.

treatment_protocol: Degradation of RAD21 or WAPL in FKBPdegron lines during differentiation was induced by adding a 1:20,000 dilution of 10mM dTAG-13 in DMSO to cell culture media (500nM final concentration) for the indicated time frames. The small molecule dTAG-13 was synthesized as previously described (Erb et al., 2017; Nabet et al., 2018). For DMSO vehicle controls, the same volume of DMSO alone was added to cell culture media.

tag: strain

tag: genotype/variation

tag: cell type

value: ES cell (Tsix-stop)

tag: treatment

tag: differentiation day

value: day 10


title: RNA_RG9_d108h_DMSO

value: 8h DMSO


title: Liver lysate 3

city: Quebec

first: Luca

last: Pellegrini

address: 1050 Rue de la Médecine

department: Department of Molecular Biology

country: Canada

institute: Université Laval

data_processing:
Trimming: fastp v0.12.4 with --length_required 30, -qualified_quality_phred 20, --correction, --cut_by_quality3, --cut_window_size 10 and --cut_mean_quality 30
Pseudoalignment: kallisto quant v0.44.0 with default parameters values.
Differential expression: DESeq2 v1.20.0 with R v3.5.0 and Bioc v3.7.
Genome_build: Ensembl release 94
Supplementary_files_format_and_content: Excel files containing raw counts, tpm counts and supplementary table.

molecule: total RNA

source_name: Lysate

extract_protocol:
1 g of liver was homogenized with a modified teflon pestle. The liver lysate was obtained by removing from this homogenate all the tissue debris and cell nuclei through 3 rounds of low speed centrifugation. WAM fractions were prepared from this lysate usi
NA was isolated from mouse liver homogenates using the Dynabeads mRNA Direct Micropurification Kit (Thermo Fisher Scientific); of this, 250 ng were then used to prepare sequencing libraries using the NEBNext Ultra II directional RNA library prep kit for I

tag: tissue

value: Liver

tag: fraction

value: liver postnuclear lysate

tag: age

value: 9-12 weeks old

tag: Sex

value: male

tag: feeding state

value: 3 hours postprandial, after overnight fasting


title: WAM fraction 5

source_name: WrappER-associated mitochondria (WAM) fraction

value: wrappER-associated mitochondria (WAM) fraction


title: WAM fraction 6


title: Liver lysate 6


title: Liver lysate 5


title: WAM fraction 3


title: WAM fraction 2


title: RNA_RC4_d108h_dTAG13


title: ChIP_RAD21_RC4_d102h_dTAG13

library_source: genomic

molecule: genomic DNA

source_name: RAD21 ChIP, clone C4

tag: chip antibody

value: RAD21 (Abcam ab992)


title: ChIP_RAD21_RC4_d102h_DMSO

value: 2h DMSO


title: RNA_WH7_d7-10_dTAG13.rep1

description: processed data file: RNA_WH7_d7-10_rep1.rep2.counts.txt

source_name: RNAseq, clone H7, rep1

value: WAPL-FKBP(degron)

value: 3 days dTAG-13 (500nM)


title: Liver lysate 1


title: WAM fraction 1


title: Liver lysate 2


title: nPOD-6251-2 snATAC (CB)

city: La Jolla

first: Kyle

last: Gaulton

state: CA

address: 9500 Gilman Drive

department: Pediatrics

institute: University of California, San Diego

data_processing:
Note: Barcodes for combinatorial barcoding snATAC-seq experiments are integrated into the original read names.
Alignment: Trimmed Nextera adapter sequences using trim_galore v.0.4.4 and aligned reads to hg19 using bwa mem v.0.7.17-r1188.
Filtering: Low mapping quality (MAPQ<30), mitochondrial reads, duplicate reads on a per-barcode basis with MarkDuplicates from picard.
Clustering: Custom code for processing snATAC-seq datasets is available at https://github.com/kjgaulton/pipelines/tree/master/T1D_snATAC_pipeline.
Peak calling: MACS2 (v.2.1.2) with --nomodel --keep-dup all on aggregate chromatin accessibility profiles for each cluster.
Genome_build: hg19
Supplementary_files_format_and_content: merged peaks
Supplementary_files_format_and_content: single cell cluster labels

organism: Homo sapiens

source_name: whole pancreas

extract_protocol:
Nuclei were isolated from fresh pancreatic islets using a glass dounce homogenizer with a tight-fitting pestle, frozen whole pancreas using a mortar and pestle, and PBMCs using a lysis buffer.
(CB; combinatorial barcoding) Permeabilized nuclei were distributed to 96 wells and tagmentation was carried out to introduce a first barcode combination. After pooling, 20 nuclei were sorted into each well of a 96 well plate and a second barcode was introduced by PCR. Libraries were purified according to the MinElute PCR Purification Kit manual using a vacuum manifold and size selection was performed with SPRI Beads. (10X; 10X Genomics) Libraries were generated using the 10X Genomics Chromium Chip E Single Cell ATAC kit following the manufacturers instructions.

value: pancreas

value: female

tag: bmi

tag: donor id


title: IIDP-AEHU156 snATAC (CB)

source_name: pancreatic islets

value: pancreatic islets


title: nPOD-6251-1 snATAC (CB)


title: nPOD-6007 snATAC (CB)


title: nPOD-6282 snATAC (CB)


title: Cytotrophoblast_male_12-14GW rep5

city: PARIS

first: Severine

middle: Aude

last: Degrelle

address: 4 avenue de l'Observatoire

country: France

institute: INSERM UMRS-1139

data_processing:
STAR alignment with default parameters, on the Ensembl reference GRCh38
STAR --genomeDir /path/to/Index/Genome --sjdbGTFfile /path/to/GTF --runThreadN 14 --readFilesIn /path/to/fq1 /path/to/fq2 --outFileNamePrefix $prefix --twopassMode Basic --sjdbOverhang 74 --outSAMtype BAM SortedByCoordinate --readFilesCommand zcat --outSAMunmapped Within --outFilterType BySJout --quantMode TranscriptomeSAM
Counting has been made with featureCount
Genome_build: Homo sapiens (human) genome assembly GRCh38 (hg38)
Supplementary_files_format_and_content: Normalized Matrix table for every gene and every sample, using DESeq2
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: Cytotrophoblast

growth_protocol: Cytotrophoblasts were isolated from first term placenta by enzyme tissue digestion and purified on percoll gradient. After washes, cell pellets were snap-frozen and stored at -80°C until RNA extraction.

extract_protocol:
Total RNA extration was performed using RNeasy Plus Mini Kit  (Qiagen) following manufacturer's instructions
Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 800 ng of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: gestational age

value: 12-14 gestational week

tag: fetal sex

value: Male

tag: developmental stage

value: Late

tag: oxygen level

value: High


title: Cytotrophoblast_male_12-14GW rep2


title: Cytotrophoblast_male_12-14GW rep1


title: Cytotrophoblast_male_12-14GW rep4


title: Cytotrophoblast_female_12-14GW rep2

value: Female


title: Cytotrophoblast_female_12-14GW rep1


title: Cytotrophoblast_female_12-14GW rep5


title: Cytotrophoblast_male_12-14GW rep3


title: Cytotrophoblast_female_12-14GW rep3


title: Cytotrophoblast_male_8-10GW rep4

value: 8-10 gestational week

value: Early

value: Low


title: Cytotrophoblast_male_8-10GW rep3


title: Cytotrophoblast_male_8-10GW rep1


title: Cytotrophoblast_female_8-10GW rep5


title: Cytotrophoblast_male_8-10GW rep2


title: Cytotrophoblast_female_8-10GW rep4


title: Cytotrophoblast_female_8-10GW rep3


title: Cytotrophoblast_female_8-10GW rep2


title: Cytotrophoblast_female_8-10GW rep1


title: 16H145

city: Montréal

first: Claude

last: Perreault

state: Québec

address: 2950, Chemin de Polytechnique

institute: Institute for Research in Immunology and Cancer

data_processing:
Basecalls performed using Illumina RTA 2.4.11.
Samples demultiplexed using bcl2fastq 2.20.
Sequenced reads were trimmed for sequencing adapters and low quality extrimities using Trimmomatic 0.35 and then mapped to the reference transcriptome using kallisto 0.43.0, taking into account that libraries are stranded.
TPM values were computed by kallisto 0.43.0 using a transcriptome index built on human genome GRCh38 along with Ensembl 88 annotations.
Genome_build: GRCh38
Supplementary_files_format_and_content: Expression files are tab delimited with columns 1) Ensembl transcript identifier, 2) transcript length , 3) effective transcript length, 4) estimated counts, and 5) transcript per million reads sequenced (TPM).

source_name: BCLQ (http://bclq.org)

growth_protocol: Except for 14H124 (expansion in NSG mice), all samples were not expanded.

extract_protocol:
TriZol + RNeasy mini Kit (Qiagen, #74106)
KAPA RNA Hyperprep Stranded PolyA

treatment_protocol: No treatment applied

tag: expansion

value: RNA Extraction from 2 million blasts directly obtained from BCLQ

tag: blast percentage at extraction

tag: tissue of origin at extraction

value: Peripheral blood

value: Acute myeloid leukemia blasts


title: 15H080

tag: cell type


title: 15H063


title: 16H123


title: 15H013


title: 10H005

value: Bone marrow


title: 15H023


title: 12H172


title: 11H008


title: Cytotrophoblast_female_12-14GW rep4


title: Cytotrophoblast_male_8-10GW rep5


title: 11H035


title: 07H141


title: 07H122


title: 08H039


title: 14H124

value: RNA Extraction from 2 million blasts expanded in NSG mice


title: 05H143


title: 07H060


title: 07H063


title: P21 BALB/c Txnip cones 1

first: Yunlu

last: Xue

address: 77 Avenue Louis Pasteur

department: Genetics

institute: Harvard University

data_processing:
Smart-Seq2 libraries were constructed and sequenced by the Broad Institute Genomics Platform on an Illumina NextSeq 500
Reads were aligned using STAR and quantified using featureCounts
Analysis of differential gene expression was performed using DESeq2
Genome_build: GRCm38.p6
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample.

source_name: Retinal cones

extract_protocol:
Retinas were dissected, dissociated with papain, and subjected to cell sorting. Cones were isolated in Buffer TCL by antibody staining.
Libraries were constructed by the Broad Institute Genomics Platform using a modified Smart-Seq2 protocol

treatment_protocol: AAVs were injected to neonatal eyes of mice at P0.

tag: strain

value: BALBC (albino wt)

value: retinal cones

value: post natal day 21

tag: treatment

value: AAV-Txnip

tag: labeling

value: cone-specific H2BGFP for FACS sorting


title: P35 C57BL/6J control cones 1

value: C57BL/6J (pigmented wt)

tag: tissue

tag: age

value: post natal day 35

value: Control


title: P21 BALB/c Txnip cones 5


title: P21 BALB/c control cones 4


title: P21 BALB/c Txnip cones 4


title: P35 C57BL/6J Txnip cones 1


title: P21 BALB/c Txnip cones 6


title: P35 C57BL/6J control cones 3


title: P21 BALB/c control cones 2


title: P21 BALB/c Txnip cones 3


title: P21 BALB/c control cones 6


title: P21 BALB/c Txnip cones 2


title: NESC2.3D.BP10

city: Kunming

first: Tianqing

last: Li

state: Yunnan

address: 727 South Jingming Road, Chenggong District

department: Yunnan Key Laboratory of Primate Biomedical Research & Institute of Primate Translational Medicine

country: China

institute: Kunming University of Science and Technology

data_processing:
The sequencing qualities of 6 RNA-seq profiles were examined with the FASTQC
HISAT2 (v 2.1.0) was used to align the RNA-seq profiles to the human genome.
The alignment results of HISAT2 in the SAM format were converted to BAM format and sorted with SAMTools (v 1.1).
Stringtie (v 1.3.4) was used to calculate the abundances of genes (in FPKM) annotated in GENCODE (v29) using the options of "-G gencode.v29.gtf -B -e –v".
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each sample
cellranger was used to align reads, generate feature-barcode matrices and filtered_feature_bc_matrix.h5 with 10x genomic single cells data
Supplementary_files_format_and_content: Cell-related barcode information and expression data

source_name: NESC

growth_protocol: To extensively expand NESCs in vitro, NESCs were digested into single cells and cultured in ultra-low attachment plates. They were cultured in chemically defined CHbFSB+LIF or CHbFSB culture medium. The CHbFSB culture medium consists of Neurobasal media surplus with 0.25% N2, 0.5% B27, 1% NEAA, 1% Glutmax, 3 μM CHIR99021, 5 μM SB431542, and 10 ng/ml OsrbFGF. TrypLE™ Express Enzyme (Gibco, 12,605,028) was diluted for 2 times with PBS (Sigma, D5652) to digest NESCs for encouraging cell propagation when passaging. NESCs were routinely passaged at 1:3 to 1:4 ratios every 3 days.

extract_protocol:
hESCs were digested into small clumps for suspension culture on ultra-low attachment plates (Corning, 3471) in the NESC-derived medium [26], which is composed of Advance DMEM/F12 (Gibco, 10565-018): Neurobasal media (Gibco, 21103-049) (1:1) supplemented with 1% N2 (Gibco, 17502-048), 2% B27 (Gibco, 17504-044), 1% Glutmax (Gibco, 35050-061), 10 ng/mL OsrbFGF (Oryza sativa recombinant human basic fibroblast growth factor, Wuhan Healthgen, China, HYC005M01), 3 μM CHIR99021 (Selleck, S2924), 5 μM SB431542 (Cellagen technology, C7243), 0.2 μM Compound E (Calbiochem, 565790), 0.1 μM LDN193189 (Selleck, S2618), and 0.1 mM β-mercaptoethanol (Sigma, M3148). After suspension culture for 6 days, neuron bodies (NBs) were digested into single cells and inoculated into ultra-low attachment plates with CHbFSB+LIF culture medium. The CHbFSB+LIF culture medium [26, 28, 31] is composed of Neurobasal medium, 1% N2, 2% B27, 1% NEAA (Gibco, 11140-050), 1% Glutmax, 3 μM CHIR99021, 5 μM SB431542, and 10 ng/ml OsrbFGF surplus with 1000 U/ml hLIF (Millipore, LIF1050).
Total RNA was isolated from NESC  using the TRIzol Reagent (Thermo Fisher Scientific, 15596018). RNA sequencing libraries were constructed using the NEBNext Ultra RNA Library Prep Kit for Illumina (NEB England BioLabs, E7530L). The fragmented and randomly primed 2×150-bp paired end libraries were sequenced using an Illumina HiSeq X Ten.
according to the protocol of Chromium Single Cell 3' Reagent Kits V3
10x genomic single cell gene expression

treatment_protocol: Digested hNESCs (passage 19) were inoculated into a 125 ml suspension bioreactor (Wiggens, BIOMIX Control MS4) with a 100-ml CHbFSB medium at the cell density of 3 × 105 cells/ml. Every 3 days, the NESCspheres were dissociated and passaged using TrypLE™ Express Enzyme: PBS (1:2). The agitation rate of NESCs growing in a stirred suspension bioreactor is 60 rpm. The bioreactor was housed in a humidified incubator with 5% CO2 at 37 °C. The NESCs were fed 2 days after inoculation by replacing 50% of the medium with the fresh medium

value: NESC

tag: condition

value: CHbFSB+LIF

value: passage 10

value: Neuroepithelial stem cells


title: P35 C57BL/6J Txnip cones 2


title: P35 C57BL/6J control cones 2


title: P21 BALB/c control cones 1


title: 08H053

value: RNA Extraction from 2 million blasts directly obtained from BCLQ


title: P21 BALB/c control cones 5


title: NESC2.3D.AP10


title: NESC2.2D.P25

value: passage 25


title: 05H149

value: Peripheral blood

value: Acute myeloid leukemia blasts


title: P35 C57BL/6J Txnip cones 3


title: NESC1.3D.AP15

value: passage 15


title: NESC1.2D.P20

value: passage 20


title: P21 BALB/c control cones 3

value: BALBC (albino wt)

value: retinal cones


title: Control

city: Yiwu

first: Qiang

last: Sun

state: Zhejiang

address: No. N1, Shangcheng Avenue

institute: Zhejiang University

description: ADSCs

data_processing:
Illumina software used for basecalling.
The preprocessed data were compared to the reference genome using HISAT2).Reference genomes and genomic information are downloaded from the Ensembl database.
Using the CuffQuant and CuffNorm components of CuffLinks software, the expression levels of transcripts and genes were quantified by the location information of Mapped Reads on genes.
Input data of gene differential expression were readcount data obtained in gene expression level analysis.As for the samples with biological duplicates, we analyzed them by DESEQ and used the Quantile Normalization method to list the differentially expressed genes with Pvalue less than 0.05 and difference multiple greater than or equal to 2.
The sequencing data were compared to the reference genome using HisAT2, and the lncRNA genes were identified using the comparison results.
Genome_build: hg19
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: Control 1

growth_protocol: cells were cultured in DMEM /f12(10%FBS)

extract_protocol:
RNA extraction method: Trizol  The first step is to build a circular RNA library and remove ribosomes, using RiboMinus Human/Mouse Transcriptome Isolation Kit (invitrogen, thermofisher); the second step is to remove linear RNA, using RNase R enzyme (epicentre, illumina), and the third step is to construct Library, using KAPA Stranded mRNA-Seq Kit (roche)
A RNA library was constructed to remove ribosomes, using RiboMinus Human/Mouse Transcriptome Isolation Kit (invitrogen, thermofisher)

treatment_protocol: treatment with high glucose (25 mM) for 48 h

value: adipose-derived stem cells (ADSCs)

value: untreatment


title: CM of HCT-8/FU rep2

city: Beijing

first: niu

last: huan

address: fengtai district of beijing

department: hereditary system

institute: Capital Medical University

data_processing:
Raw reads were obtained in FASTQ format from Illumina Hiseq X Ten sequencing platform.
The clean reads in compressed FASTQ format were aligned using HISAT2 to the reference of human genome (Homo_sapiens.GRCh38.dna.primary_assembly.fa).
The resulted BAM files were sorted with SAMtools (version 1.18).
The depth counts were called with HTSeq (version 0.11.2. Linux_x86_64, Simon Anders (sanders@fs.tum.de)) with the reference of human genome (Homo_sapiens.GRCh38.94.gtf), European Molecular Biology Laboratory (EMBL)) used to calculate the Fold change (FC) of FPKM and p value among sample groups according to an over-dispersed Poisson model.
The p value was got by DESeq2(version 1.30.0)
The analyses for alternative splicing events of expressed genes were performed using the rMATS software package (version 4.1.0, http://rnaseq-mats.sourceforge.net/, Xing Lab, Children's Hospital of Philadelphia)
Genome_build: Ensembl hg38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: CM of HCT-8/FU

growth_protocol: Cells were maintained at 37°C in a 5% CO2 incubator in Ham’s F12 Nutrient Medium (Ham’s F12) with 10% fetal bovine serum (FBS)  and 1% penicillin-streptomycin.

extract_protocol:
Cells were scraped off from the surface in trypsin-versene solution and collected by 500 g centrifugation. The pellet was washed with PBS to remove residual media. Total RNA extractions were performed with the RNA-Quick Purification Kit following the manufacturer's protocol. The RNA concentrations were determined using a Nanodrop ND1000 spectrophotometer .
The mRNA sequencing libraries were performed with the VAHTS Universal V6 RNA-seq Library Prep Kit for Illumina (NR604-01, Vazyme Biotech, Nanjing, China), VAHTS RNA Multiplex Oligos Set1- Set2 for Illumina (N323, Vazyme Biotech, Nanjing, China), VAHTS DNA Clean Beads (N311-01, Vazyme Biotech, Nanjing, China), VAHTS mRNA Capture Beads (N401-01, Vazyme Biotech, Nanjing, China). The quality assessments were performed in the laboratory of GENEWIZ, lnc (Suzhou, China).

treatment_protocol: 5-FU-sensitive HCT-8 cells and 5-FU-resistant HCT-8/FU cells cultured in 6-well plate.The conditioned medium were collected by cultivate of 48 hours of HCT-8 or HCT-8/FU.

value: colon cancer cell lines

tag: cell line

tag: medium

value: CM with HCT-8/FU


title: High_glucose

description: ADSCs

source_name: Highglucose 1

value: treatment with high glucose (25 mM) for 48 h


title: CM of HCT-8/FU rep1


title: NESC1.3D.BP15

value: NESC


title: CM of HCT-8 rep1

source_name: CM of HCT-8

value: CM with HCT-8


title: HCT-8/FU rep2

value: unconditioned


title: RNAseq_late2C_Rep2

city: Doha

first: Mohamed Nadhir

last: Djekidel

state: Doha

address: Gharafa

department: Translational Medicine Department

country: Qatar

institute: Sidra Medicine

data_processing:
The raw RNA-seq data were first adapter- and quality-trimmed using Trimgalore (v0.6.5) with parameters “‘--nextseq 20 -j 6 --clip_R2 5  --paired ”. The trimmed reads were then mapped to rRNA and tRNA using Hisat2 (v2.1.0) with parameters ‘--rna-strandness RF  --un-conc-gz unmapped.hisat2.gz’. The reads that were not mapped to rRNA/tRNA were then aligned to the mm10 SNP-masked genome using Hisat2 with paramters ‘--rna-strandness RF --sp 1000,1000 --no-mixed --no-discordant’. Known splicing sites were also supplied to improve mapping efficiency. Only uniquely aligned reads (tag NH = 1) were used for subsequent analyses. Gene FPKM values were quantified using StringTie (v1.3.3b) with parameters ‘--rf -v’.
Genome_build: mm10
Supplementary_files_format_and_content: tab

source_name: late 2-cell embryos

extract_protocol:
As all libraries generated in this study involved low-input techniques,  samples collected were directly put into lysis buffer for further library construction.
Low-input RNA-seq libraries were prepared using SMART-Seq Stranded Kit (Illumina) according to the manufacturer’s instructions. Paired-end 76bp sequencing was performed on a NextSeq550/HiSeq2000 sequencer (Illumina).

tag: background

value: B6/DBA x B6/DBA

tag: manipulation

value: WT x WT

tag: embryos/oocytes

value: 2-cell embryos


title: RNAseq_late2C_Rep1


title: HCT-8/FU rep1


title: HCT-8 rep1


title: RNAseq_MII_Rep1

source_name: MII oocytes

value: WT

value: MII oocytes


title: RNAseq_MII_Rep2


title: HCT-8 rep2


title: RNAseq_4cell_Eed_OE_Rep2

source_name: 4-cell embryo

value: B6/129mixed x PWK

value: Eed CKO x WT, (matKO)

value: 4-cell embryos


title: RNAseq_4cell_Eed_OE_Rep1


title: SAMPLE 217_510267FCB1AG01280119_C09

type: RNA

city: Bethesda

first: Sai

last: Kanaparthi

state: MD

address: 7500 Old Georgetown Rd

department: BiG

institute: Immune Tolerance Network

description: Liver biopsy  from Non-tolerant SAMPLE 217

hyb_protocol: Standard Affymetrix protocol

scan_protocol: GeneChips were scanned using the GeneChip Scanner 3000.

data_processing: Data processing cel files were normalized with 'rma' function of the Bioconductor package (v3.1) in R.3.4.3

label: Biotin

source_name: Liver biopsy

label_protocol: Standard Affymetrix protocol

extract_protocol: For total RNA extraction cryopreserved liver tissue samples were homogenized in TRIzol reagent (Invitrogen, San Diego, CA, USA) using pestle and nuclease-free 1,5ml reaction tubes (Ambion). Total RNA was then extracted following the manufacturers guidelines and quality was assessed with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, USA).

value: liver

tag: isw outcome (tolerant/non-tolerant)

value: Non-tolerant

tag: time point

value: For-cause Biopsy Visit

tag: visit code


title: SAMPLE 220_141477MF23281216H01

description: Liver biopsy  from Non-tolerant SAMPLE 220

value: Primary Endpoint Visit


title: SAMPLE 221_810205MF26281216F05

description: Liver biopsy  from Non-tolerant SAMPLE 221


title: RNAseq_4cell_Eed_CTR_Rep1

value: Eed CTR x WT, (CTR)


title: SAMPLE 222_733922MF22281216A02

description: Liver biopsy  from Non-tolerant SAMPLE 222


title: SAMPLE 213_259491MF22281216D02

description: Liver biopsy  from Non-tolerant SAMPLE 213


title: RNAseq_4cell_Eed_CTR_Rep2


title: SAMPLE 218_420006MF22281216D12

description: Liver biopsy  from Non-tolerant SAMPLE 218


title: SAMPLE 219_933569MF23221216A09

description: Liver biopsy  from Non-tolerant SAMPLE 219


title: CM of HCT-8 rep2

value: colon cancer cell lines


title: SAMPLE 214_679850MF22281216A08

description: Liver biopsy  from Non-tolerant SAMPLE 214


title: RNAseq 4cell KDM6B (OE) Rep1

value: B6/DBA x PWK

value: WT x WT, KDM6B (OE)


title: RNAseq 4cell KDM6B (OE) Rep2


title: SAMPLE 216_334078MF22281216F07

description: Liver biopsy  from Non-tolerant SAMPLE 216


title: SAMPLE 212_971198MF22281216E07

description: Liver biopsy  from Non-tolerant SAMPLE 212


title: RNAseq 4cell KDM6B (CTR) Rep2

value: WT x WT, KDM6B (CTR)


title: H3K27me3_Sperm_WT

data_processing:
Library strategy: STAR-ChIP
Low quality reads and adaptor sequences of the STAR-ChIP data were trimmed using Trimgalore (v0.4.5) with the parameter “--paired”. The trimmed reads were then aligned to mm10 genome using Bowtie2 (v2.3.4.3)  with parameters “‘--no-unal --no-mixed --no-discordant -I 0”. Unmapped and low quality reads were removed using sambamba (v0.5.9) with parameters “-f bam -F ‘mapping_quality >= 20 and not (unmapped or mate_is_unmapped)’”. Only uniquely mapped reads were kept for downstream analyses. STAR-ChIP reads within the mm10 black list regions (https://github.com/Boyle-Lab/Blacklist/tree/master/lists) were not used for subsequent analyses.
Genome_build: mm10
Supplementary_files_format_and_content: tab

source_name: Sperm

extract_protocol:
Sperm were collected using the "swim-up" method
Mouse sperm H2Aub ChIP-seq was performed as described previously with some modifications. Briefly, ~2 million mouse spermatozoa (PWK) collected by the swim-up method were treated with 50 mM DTT (Sigma-Aldrich) for 2 hrs at room temperature and quenched by adding N-ethylmaleimide (NEM, Sigma-Aldrich) and incubating for additional 30 min. Pelleted sperm cells were resuspended using 100 µl complete buffer with detergent [15 mM Tris-HCl (pH 7.5), 60 mM KCl, 5 mM MgCl2, 0.1 mM EGTA, 0.3 M sucrose, 0.5 mM DTT, 0.5% NP-40 and 1% sodium deoxycholate] and lysed on ice for 10 min. The same volume of MNase buffer [(85 mM Tris-HCl (pH 7.5), 3 mM MgCl2, 2 mM CaCl2, 0.3 M sucrose] containing 10 units MNase (Sigma-Aldrich) was added and incubated at 37ºC for 5min. MNase treatment was stopped by adding 2 µl of 0.5M EDTA (Thermo Fisher). For STAR-ChIP, 200 µl cold 2×RIPA buffer (1% Triton, 280 mM NaCl, 0.1% SDS, 0.2% DOC, 5mM EGTA, and 1×proteinase inhibitor) was supplemented. After spinning at max speed at 4ºC for 10 min, the supernatant was transferred to a new tube and supplemented with 176 µl of RIPA buffer [10 mM Tris-HCl (pH 8.0), 140 mM NaCl, 1% Triton, 0.1% SDS, 0.1% DOC, 1 mM EDTA, 1×proteinase inhibitor]. The samples were incubated with a rabbit anti-H2Aub antibody (1:200; Cell Signaling, #8240) overnight with rotation at 4ºC. The following day, the samples were incubated with 100 µg protein A dynabeads (Life Technologies) for 2 hrs with rotation at 4 ºC. The beads were than washed five times with 150 µl RIPA buffer and once with LiCl buffer [250 mM LiCl, 10mM Tris-HCl (pH 8.0), 1 mM EDTA, 0.5% NP-40, 0.5% DOC].  To elute DNA, the beads were resuspended with 200 µl TE buffer with 2.5 µl 10% SDS (Sigma-Aldrich) and 2.5 µl mg/ml Protease K (Thermo Fisher) and incubated in a shaker at 55ºC for 90 min. Supernatant was collected to a new tube and DNA were extracted by phenol-chloroform followed by ethanol precipitation. Sequencing libraries were prepared using the NEBNext Ultra II DNA library preparation kit for Illumina (New England Biolabs) according to the manufacturer’s instructions.

value: PWK

value: sperm


title: H2Aub_Sperm_WT_Rep2

data_processing:
Library strategy: STAR-ChIP
Low quality reads and adaptor sequences of the STAR-ChIP data were trimmed using Trimgalore (v0.4.5) with the parameter “--paired”. The trimmed reads were then aligned to mm10 genome using Bowtie2 (v2.3.4.3)  with parameters “‘--no-unal --no-mixed --no-discordant -I 0”. Unmapped and low quality reads were removed using sambamba (v0.5.9) with parameters “-f bam -F ‘mapping_quality >= 20 and not (unmapped or mate_is_unmapped)’”. Only uniquely mapped reads were kept for downstream analyses. PCR duplicates were removed using the “MarkDuplicates” command from Picard tools (v2.8.0) (https://broadinstitute.github.io/picard/). The bigwig files were generated using the “bamCoverage” command from deepTools (v3.0.2) with parameters “--binSize 25 --extendReads 150 --normalizeUsing RPKM --outFileFormat bigwig --scaleFactor 1”. Reproducibility were assessed by calculating Pearson correlation coefficient of RPKM in 5-kb bins between replicates. The biological replicates were pooled using the ‘merge’ command from sambamba (v0.5.9). STAR-ChIP reads within the mm10 black list regions (https://github.com/Boyle-Lab/Blacklist/tree/master/lists) were not used for subsequent analyses.
Genome_build: mm10

value: Sperm


title: SAMPLE 209_944000MF22221216C07

description: Liver biopsy  from Non-tolerant SAMPLE 209


title: SAMPLE 206_324363MF26281216B11

description: Liver biopsy  from Non-tolerant SAMPLE 206


title: SAMPLE 210_768987MF22281216E08

description: Liver biopsy  from Non-tolerant SAMPLE 210


title: H2Aub_Sperm_WT_Rep1

value: WT


title: H2Aub 1cell WT Rep2

data_processing:
Library strategy: CUT&RUN
Low quality reads and adaptor sequences of the CUT&RUN data were trimmed using Trimgalore (v0.4.5) with the parameter “--paired”. The trimmed reads were then aligned to mm10 genome using Bowtie2 (v2.3.4.3)  with parameters “‘--no-unal --no-mixed --no-discordant -I 10 -X 700”. To minimize mapping bias toward reference genome (B6) for the F1 hybrid data, SNPs between PWK and B6/129 or B6/DBA were masked as “N”, depending the maternal genetic background. Unmapped and low quality reads were removed using sambamba (v0.5.9) with parameters “-f bam -F ‘mapping_quality >= 20 and not (unmapped or mate_is_unmapped)’”. Only uniquely mapped reads were kept for downstream analyses. PCR duplicates were removed using the “MarkDuplicates” command from Picard tools (v2.8.0) (https://broadinstitute.github.io/picard/). SNPsplit (v0.3.2) was used to determine the parental origin of the CUT&RUN read pairs. The bigwig files were generated using the “bamCoverage” command from deepTools (v3.0.2) with parameters “--binSize 25 --extendReads 150 --normalizeUsing RPKM --outFileFormat bigwig --scaleFactor 1”. Reproducibility were assessed by calculating Pearson correlation coefficient of RPKM in 5-kb bins between replicates. The biological replicates were pooled using the ‘merge’ command from sambamba (v0.5.9). CUT&RUN reads within the mm10 black list regions (https://github.com/Boyle-Lab/Blacklist/tree/master/lists) were not used for subsequent analyses.
Genome_build: mm10 with SNPs  masked as "N"
Supplementary_files_format_and_content: Bigwig files that can be uploaded on a genome browser to view H3K27me3 and H2Aub at different genomic locations.

source_name: zygote

extract_protocol:
As all libraries generated in this study involved low-input techniques,  samples collected were directly put into lysis buffer for further library construction.
H3K27me3 and H2Aub CUT&RUN libraries were constructed as previously described (Skene and Henikoff, 2017 eLife. 2017; 6: e21856, PMID: 28079019).

value: WT x WT

value: zygote


title: SAMPLE 211_313003MF26281216E06

description: Liver biopsy  from Non-tolerant SAMPLE 211


title: SAMPLE 215_648298FCB1AG01280119_D10

description: Liver biopsy  from Non-tolerant SAMPLE 215


title: SAMPLE 207_417706MF22281216C05

description: Liver biopsy  from Non-tolerant SAMPLE 207


title: H2Aub MII WT Rep2

source_name: metaphase II oocyte

value: metaphase II oocyte


title: SAMPLE 208_332151MF23281216F12

description: Liver biopsy  from Non-tolerant SAMPLE 208


title: RNAseq 2cell KDM6B (OE) Rep2

source_name: 2-cell embryo

extract_protocol:
As all libraries generated in this study involved low-input techniques,  samples collected were directly put into lysis buffer for further library construction.
Low-input RNA-seq libraries were prepared using SMART-Seq Stranded Kit (Illumina) according to the manufacturer’s instructions. Paired-end 76bp sequencing was performed on a NextSeq550 sequencer (Illumina).

value: WT x WT, PR-DUP (OE)

value: 17  2-cell


title: SAMPLE 204_821656MF22281216H07

description: Liver biopsy  from Non-tolerant SAMPLE 204

value: liver

value: Non-tolerant


title: H2Aub GV WT_Rep2

source_name: germinal vesicle oocyte

value: B6/129 mixed

value: Eed (fl/fl)

value: germinal vesicle oocyte


title: RNAseq 2cell KDM6B (OE) Rep1

value: B6/DBA x PWK


title: SAMPLE 202_597774MF23281216B04

description: Liver biopsy  from Non-tolerant SAMPLE 202

value: Primary Endpoint Visit


title: RNAseq 2cell KDM6B (CTR) Rep2

value: WT x WT, KDM6B (CTR)

value: 18  2-cell


title: RNAseq 2cell KDM6B (CTR) Rep1


title: SAMPLE 200_349892MF26281216C01

description: Liver biopsy  from Non-tolerant SAMPLE 200


title: RNAseq 4cell PR-DUB (OE) Rep2

value: 16  4-cell


title: SAMPLE 199_910844MF26281216C03

description: Liver biopsy  from Non-tolerant SAMPLE 199


title: SAMPLE 205_376456MF26281216B01

description: Liver biopsy  from Non-tolerant SAMPLE 205


title: SAMPLE 196_558421MF23221216E09

description: Liver biopsy  from Non-tolerant SAMPLE 196


title: SAMPLE 198_802143MF26221216H07

description: Liver biopsy  from Non-tolerant SAMPLE 198


title: RNAseq 4cell PR-DUB (CTR) Rep2

value: WT x WT, PR-DUP (CTR)

value: 19  4-cell


title: RNAseq 2cell PR-DUB (OE) Rep2

value: 15  2-cell


title: RNAseq 2cell PR-DUB (OE) Rep1


title: SAMPLE 197_214859FCB1221216E07

description: Liver biopsy  from Non-tolerant SAMPLE 197

value: For-cause Biopsy Visit


title: RNAseq 4cell PR-DUB (CTR) Rep1


title: SAMPLE 203_461059MF24281216C12

description: Liver biopsy  from Non-tolerant SAMPLE 203


title: RNAseq 2cell PR-DUB (CTR) Rep2


title: SAMPLE 194_951840FCB1281216D11

description: Liver biopsy  from Non-tolerant SAMPLE 194


title: SAMPLE 201_751760MF22221216F07

description: Liver biopsy  from Non-tolerant SAMPLE 201


title: SAMPLE 193_162449MF26281216E04

description: Liver biopsy  from Non-tolerant SAMPLE 193


title: SAMPLE 195_141477FCB1AG01280119_E01

description: Liver biopsy  from Non-tolerant SAMPLE 195


title: H3K27me3 4cell PR-DUB (OE) Rep1

description:
PR-DUB_OE_4cell_K27me3_rep1_Mat.bw
PR-DUB_OE_4cell_K27me3_rep1_Pat.bw

data_processing:
Low quality reads and adaptor sequences of the CUT&RUN data were trimmed using Trimgalore (v0.4.5) with the parameter “--paired”. The trimmed reads were then aligned to mm10 genome using Bowtie2 (v2.3.4.3)  with parameters “‘--no-unal --no-mixed --no-discordant -I 10 -X 700”. To minimize mapping bias toward reference genome (B6) for the F1 hybrid data, SNPs between PWK and B6/129 or B6/DBA were masked as “N”, depending the maternal genetic background. Unmapped and low quality reads were removed using sambamba (v0.5.9) with parameters “-f bam -F ‘mapping_quality >= 20 and not (unmapped or mate_is_unmapped)’”. Only uniquely mapped reads were kept for downstream analyses. PCR duplicates were removed using the “MarkDuplicates” command from Picard tools (v2.8.0) (https://broadinstitute.github.io/picard/). SNPsplit (v0.3.2) was used to determine the parental origin of the CUT&RUN read pairs. The bigwig files were generated using the “bamCoverage” command from deepTools (v3.0.2) with parameters “--binSize 25 --extendReads 150 --normalizeUsing RPKM --outFileFormat bigwig --scaleFactor 1”. Reproducibility were assessed by calculating Pearson correlation coefficient of RPKM in 5-kb bins between replicates. The biological replicates were pooled using the ‘merge’ command from sambamba (v0.5.9). CUT&RUN reads within the mm10 black list regions (https://github.com/Boyle-Lab/Blacklist/tree/master/lists) were not used for subsequent analyses.
Genome_build: mm10 with SNPs masked as "N"
Supplementary_files_format_and_content: Bigwig files that can be uploaded on a genome browser to view H3K27me3 and H2Aub at different genomic locations.

extract_protocol:
As all libraries generated in this study involved low-input techniques,  samples collected were directly put into lysis buffer for further library construction.
H3K27me3 and H2Aub CUT&RUN libraries were constructed as previously described (Skene and Henikoff, 2017 eLife. 2017; 6: e21856, PMID: 28079019).
CUT&RUN

value: 40 4-cell


title: H3K27me3 4cell PR-DUB (OE) Rep2

description:
PR-DUB_OE_4cell_K27me3_rep2_Mat.bw
PR-DUB_OE_4cell_K27me3_rep2_Pat.bw

value: WT x WT, PR-DUP (OE)


title: SAMPLE 192_233127FCB1281216E03

description: Liver biopsy  from Non-tolerant SAMPLE 192


title: SAMPLE 189_828298MF23281216D09

description: Liver biopsy  from Non-tolerant SAMPLE 189


title: SAMPLE 188_160690MF26281216B07

description: Liver biopsy  from Non-tolerant SAMPLE 188


title: RNAseq 4cell PR-DUB (OE) Rep1


title: SAMPLE 187_907381FCB1AG01280119_E09

description: Liver biopsy  from Non-tolerant SAMPLE 187


title: H3K27me3 4cell PR-DUB (CTR) Rep2

description:
PR-DUB_CTR_4cell_K27me3_rep2_Mat.bw
PR-DUB_CTR_400ng_4cell_K27me3_rep2_Pat.bw


title: H2Aub 4cell PR-DUB (CTR) Rep1


title: H3K27me3 4cell PR-DUB (CTR) Rep1

description:
PR-DUB_CTR_4cell_K27me3_rep1_Mat.bw
PR-DUB_CTR_4cell_K27me3_rep1_Pat.bw


title: H2Aub 4cell Eed MatKO Rep2

description:
H2Aub_4Cell_Eedko_Mat.bw
H2Aub_4Cell_Eedko_Pat.bw

value: B6/129 mixed x PWK

value: Gdf9-Eed (fl/fl) x WT

value: 50 4-cell


title: H2Aub ICM WT Rep1

description:
H2Aub_ICM_WT.bw
H2Aub_ICM_Mat.bw
H2Aub_ICM_Pat.bw

source_name: Inner cell mass

value: WT x WT

value: 20 ICM


title: SAMPLE 186_736429MF22221216B09

description: Liver biopsy  from Non-tolerant SAMPLE 186

value: liver

value: Non-tolerant


title: SAMPLE 185_632724MF26281216D06

description: Liver biopsy  from Non-tolerant SAMPLE 185

value: Primary Endpoint Visit


title: SAMPLE 183_300934FCB1281216C07

description: Liver biopsy  from Non-tolerant SAMPLE 183


title: H2Aub 1cell WT Rep1

description:
H2Aub_1C_WT.bw
H2Aub_1cell_Mat.bw
H2Aub_1cell_Pat.bw

value: B6/DBA x PWK

value: 91  1-cell


title: SAMPLE 191_461082MF22281216D05

description: Liver biopsy  from Non-tolerant SAMPLE 191


title: H2Aub 4cell PR-DUB (OE) Rep1


title: SAMPLE 184_827472MF26281216B12

description: Liver biopsy  from Non-tolerant SAMPLE 184


title: SAMPLE 181_731083MF26281216E05

description: Liver biopsy  from Non-tolerant SAMPLE 181


title: SAMPLE 175_679850SV281015HGU219F01

description: Liver biopsy  from Tolerant SAMPLE 175

value: Tolerant

value: Screening Visit

value: SV


title: SAMPLE 190_907381MF22281216E01

description: Liver biopsy  from Non-tolerant SAMPLE 190


title: SAMPLE 177_843475MF26281216A11

description: Liver biopsy  from Non-tolerant SAMPLE 177


title: SAMPLE 174_996155SV281015HGU219A03

description: Liver biopsy  from Non-tolerant SAMPLE 174


title: SAMPLE 176_516308FCB1281216G02

description: Liver biopsy  from Non-tolerant SAMPLE 176


title: SAMPLE 180_113192SV281015HGU219C06

description: Liver biopsy  from Non-tolerant SAMPLE 180


title: RNAseq 2cell PR-DUB (CTR) Rep1

value: WT x WT, PR-DUP (CTR)

value: 15  2-cell


title: SAMPLE 182_374426MF26281216D07

description: Liver biopsy  from Non-tolerant SAMPLE 182


title: H2Aub 4cell Eed MatKO Rep1

value: 35 4-cell


title: SAMPLE 173_821656SV151015HGU219A12

description: Liver biopsy  from Non-tolerant SAMPLE 173


title: H2Aub Epi E65 WT Rep1

description:
H2Aub_Epi_E65.bw
H2Aub_Epi_E65_Mat.merged.bw
H2Aub_Epi_E65_Pat.merged.bw

source_name: epiblast E6.5

value: 1  Epiblast


title: SAMPLE 170_938137FCB1281216E09

description: Liver biopsy  from Non-tolerant SAMPLE 170

value: For-cause Biopsy Visit


title: SAMPLE 171_907381SV151015HGU219D12

description: Liver biopsy  from Non-tolerant SAMPLE 171


title: SAMPLE 179_419162SV281015HGU219E02

description: Liver biopsy  from Tolerant SAMPLE 179


title: SAMPLE 169_165255FCB1281216B10

description: Liver biopsy  from Non-tolerant SAMPLE 169


title: SAMPLE 178_214859SV281015HGU219E06

description: Liver biopsy  from Non-tolerant SAMPLE 178


title: H2Aub 2cell WT Rep3

description:
H2Aub_2C_WT.bw
H2Aub_2Cell_Mat.merged.bw
H2Aub_2Cell_Pat.merged.bw

value: B6/129 mixed x PWK

value: Eed (fl/fl) x WT

value: 80  2-cell


title: SAMPLE 168_794852FCB1221216A05

description: Liver biopsy  from Non-tolerant SAMPLE 168


title: H2Aub MII WT Rep1

value: WT

value: 300 MII oocytes


title: H2Aub Epi E65 WT Rep2

value: WT x WT


title: SAMPLE 167_540674MF22281216A03

description: Liver biopsy  from Non-tolerant SAMPLE 167

value: liver

value: Non-tolerant

value: Primary Endpoint Visit


title: Ube3am-/p+-3

city: shanghai

first: ronggui

last: hu

state: China

address: Shanghai Institute of Biochemistry and Cell Biology

institute: Center for Excellence in Molecular Cell Science

data_processing:
Illumina Casava1.8 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm10 whole genome using STAR v2.7.4a with parameters --readFilesCommand gunzip -c --readFilesIn --outSAMtype BAM
Reads count were calculated using htseq-count software with default parameters.
Genome_build: mm10
Supplementary_files_format_and_content: raw count text files

source_name: hippocampus

extract_protocol:
hippocampus were removed, and RNA was harvested using Trizol reagent.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: hippocampus

tag: genotype

tag: Sex

value: male


title: SAMPLE 163_648298MF23281216D04

description: Liver biopsy  from Non-tolerant SAMPLE 163


title: SAMPLE 162_148518FCB1281216A09

description: Liver biopsy  from Non-tolerant SAMPLE 162


title: SAMPLE 159_148518SV151015HGU219D11

description: Liver biopsy  from Non-tolerant SAMPLE 159

value: Screening Visit

value: SV


title: SAMPLE 172_143261MF23281216F08

description: Liver biopsy  from Non-tolerant SAMPLE 172


title: SAMPLE 165_990077FCB1281216H11

description: Liver biopsy  from Non-tolerant SAMPLE 165


title: SAMPLE 166_702777FCB1281216A10

description: Liver biopsy  from Non-tolerant SAMPLE 166


title: SAMPLE 158_182516FCB1281216C10

description: Liver biopsy  from Non-tolerant SAMPLE 158


title: SAMPLE 154_334078SV281015HGU219A01

description: Liver biopsy  from Tolerant SAMPLE 154

value: Tolerant


title: SAMPLE 156_772315FCB1AG01280119_D07

description: Liver biopsy  from Non-tolerant SAMPLE 156


title: SAMPLE 161_510267MF22281216A05

description: Liver biopsy  from Non-tolerant SAMPLE 161


title: SAMPLE 153_971198SV281015HGU219C04

description: Liver biopsy  from Tolerant SAMPLE 153


title: SAMPLE 151_233127SV151015HGU219E09

description: Liver biopsy  from Non-tolerant SAMPLE 151


title: SAMPLE 155_772315MF22281216E11

description: Liver biopsy  from Non-tolerant SAMPLE 155


title: SAMPLE 149_702777SV281015HGU219E03

description: Liver biopsy  from Non-tolerant SAMPLE 149


title: SAMPLE 146_141477SV281015HGU219F06

description: Liver biopsy  from Non-tolerant SAMPLE 146


title: SAMPLE 157_455735FCB1281216G05

description: Liver biopsy  from Non-tolerant SAMPLE 157


title: SAMPLE 164_813272FCB1281216H09

description: Liver biopsy  from Non-tolerant SAMPLE 164


title: SAMPLE 150_393758SV151015HGU219E10

description: Liver biopsy  from Non-tolerant SAMPLE 150


title: Ube3am-/p+-2


title: SAMPLE 152_550115SV151015HGU219F09

description: Liver biopsy  from Tolerant SAMPLE 152


title: KO_Kidney_Rep4

city: Piscataway

first: Michael

middle: P

last: Verzi

state: NJ

address: 145 Bevier Road

institute: Rutgers, the State University of New Jersey

data_processing:
Read alignment and mapping done with TopHat2, version 2.1.0
Gene and transcript level computation done with CuffQuant, Version 2.2.1
Normalized abundance measurements generated by CuffNorm, Version 2.2.1, calculate FPKM values using quartile normalization
CuffDiff, Version 2.2.1, geometric normalization, per-condition dispersion
Genome_build: mm9
Supplementary_files_format_and_content: Kidney_HNF4AKOvsCtrl_gene_exp.diff containing significant changes in transcript expression outputted by Cuffdiff

extract_protocol:
RNAs were extracted from kidney tissues of indicated genotypes. The whole kidney samples were washed with PBS, and then quick frozen in liquid nitrogen and stored at -80°C for later processing for RNA extraction. Kidney samples were homogenized in Trizol (Invitrogen) and processed for RNA extraction according to the manufacturer’s instructions.
BGI in-house kit (BGI-Shenzhen, China)

treatment_protocol: Experimental UBC-CreERT2 positive and negative mice (5 weeks old) were all treated with tamoxifen at 100 mg/kg/day by intraperitoneal injection for 4 consecutive days. Kidney tissues were collected 7 days after the first injection

value: Kidney

value: UBC-CreERT2; Hnf4af/f (HNF4AKO)

value: tamoxifen 100mg/kg/day


title: Ube3am-/p+-1


title: Ube3am+/p+-3

value: wild type


title: SAMPLE 148_777934SV151015HGU219F11

description: Liver biopsy  from Non-tolerant SAMPLE 148


title: Ctrl_Kidney_Rep2

value: Hnf4af/f (Control)


title: Ctrl_Kidney_Rep3


title: Ctrl_Kidney_Rep4


title: KO_Kidney_Rep2


title: Ube3am+/p+-1


title: Sample 21_CREBmut-body-fasted1

city: San Diego

first: Ezra

last: Wiater

address: 10010 N Torrey Pines Rd

department: PBL

institute: Salk Institute For Biological Studies

description:
q6
processed data file:
4_Abdo-gene_exp_sample16-21.diff

data_processing:
.diff file from cuffdiff comparing the samples indicated in the file name:
MiSeq Generation of fastq files
tophat2 alignment
cuffdiff Differential Gene Expression
.txt file from DESeq2 comparing thhe samples indicated in the file name:
MiSeq Generation of fastq files
STAR alignment
DESeq2 Differential Gene Expression
Genome_build: dm3 (.diff files); dm6 (DESeq2 output)
Supplementary_files_format_and_content: .diff file from cuffdiff or .txt file from DESeq2 comparing the samples indicated in the file name

organism: Drosophila melanogaster

source_name: CREBmut-body-fasted

growth_protocol: standard fly media

extract_protocol:
total RNA extracted with Qiagen Rneasy kit
polyA RNA isolated and library prepared with NEBnext Ultra kit

treatment_protocol: 16h fasting

value: creb mutant fruit fly

value: 3-5 days old

value: adult fly body

value: 16 hours fasting treatment


title: SAMPLE 144_944000SV151015HGU219A06

description: Liver biopsy  from Tolerant SAMPLE 144

value: liver


title: Ctrl_Kidney_Rep1


title: KO_Kidney_Rep3


title: RNAseq 4cell KDM6B (CTR) Rep1

value: B6/DBA x PWK

value: WT x WT, KDM6B (CTR)

value: 4-cell embryos


title: SAMPLE 145_516308SV151015HGU219C12

description: Liver biopsy  from Non-tolerant SAMPLE 145

value: Non-tolerant


title: SAMPLE 147_526574FCB1281216E12

description: Liver biopsy  from Non-tolerant SAMPLE 147

value: For-cause Biopsy Visit


title: Sample 16_wt-body-fed1

description:
q1
processed data file:
4_Abdo-gene_exp_sample16-21.diff

source_name: wt-body-fed

value: w1118 fruit fly

value: fed


title: Sample 14_CRTCko-head-fed3

description:
q3
processed data file:
3_fed-head-KO_gene_exp_sample13-15.diff
6_VolcanoPlot-DESeq2_crtcko_fastVSfed_sample3-4-9-10-14.txt
8_VolcanoPlot-DESeq2_fed_crtckoVSwt_sample1-3-7-9-13-14.txt

source_name: CRTCko-head-fed

value: crtc ko fruit fly

value: adult fly head


title: SAMPLE 141_983489FCB1281216C06

description: Liver biopsy  from Non-tolerant SAMPLE 141


title: Ube3am+/p+-2

value: hippocampus

value: male


title: KO_Kidney_Rep1

value: Kidney

value: UBC-CreERT2; Hnf4af/f (HNF4AKO)

value: tamoxifen 100mg/kg/day


title: SAMPLE 142_276711SV151015HGU219G06

description: Liver biopsy  from Non-tolerant SAMPLE 142

value: Screening Visit

value: SV


title: Sample 15_SIKmut-head-fed1

description:
q2
processed data file:
3_fed-head-KO_gene_exp_sample13-15.diff

source_name: SIKmut-head-fed

value: sik2 mutant fruit fly

value: 3-5 days old


title: SAMPLE 138_731083SV151015HGU219G05

description: Liver biopsy  from Tolerant SAMPLE 138

value: liver

value: Tolerant


title: NGN3-EGFP_2

city: Aurora

first: Alexandra

last: Theis

state: Colorado

address: 1775 Aurora Ct, Room 4202A

department: Barbara Davis Center

institute: University of Colorado Anschutz Medical Campus

data_processing:
FastQC for quality check
Cutadapt for adapter trimming
Tophat (version 2.0.4) for genome alignment
Cufflinks (version 2.0.2) for gene counts
DESeq for differential gene expression
Genome_build: mm9
Supplementary_files_format_and_content: .txt with column 1 (gene name) and column 2 (read counts)

source_name: e15.5 pancreas sorted NGN3-EGFP

extract_protocol:
Whole pancreata were dissected from e15.5 NGN3-EGFP embryos. Cells were disocciated and GFP+ cells were sorted, lysed in Buffer RLT, and flash frozen. RNA was extracted using Qiagen RNAeasy micro kit, eluted in 30ul RNAse free H20.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: mixed, C57BL6

value: pancreas

tag: Stage


title: NGN3-EGFP_1

tag: strain


title: Sample 20_CREBmut-body-fed1

description:
q5
processed data file:
4_Abdo-gene_exp_sample16-21.diff

source_name: CREBmut-body-fed

value: creb mutant fruit fly

value: adult fly body

tag: treatment


title: Sample 12_CREBmut-head-fasted2

description:
q6
processed data file:
2_DMHead2-gene_exp_sample7-12.diff

source_name: CREBmut-head-fasted

value: 16 hours fasting treatment


title: Sample 19_CRTCko-body-fasted1

description:
q4
processed data file:
4_Abdo-gene_exp_sample16-21.diff

source_name: CRTCko-body-fasted

value: crtc ko fruit fly


title: Sample 18_CRTCko-body-fed1

description:
q3
processed data file:
4_Abdo-gene_exp_sample16-21.diff

source_name: CRTCko-body-fed

value: fed


title: Sample 17_wt-body-fasted1

description:
q2
processed data file:
4_Abdo-gene_exp_sample16-21.diff

source_name: wt-body-fasted

value: w1118 fruit fly


title: SAMPLE 136_374426SV151015HGU219H11

description: Liver biopsy  from Tolerant SAMPLE 136

source_name: Liver biopsy


title: SAMPLE 143_143261SV151015HGU219F06

description: Liver biopsy  from Non-tolerant SAMPLE 143

value: Non-tolerant


title: SAMPLE 140_420006SV151015HGU219A08

description: Liver biopsy  from Tolerant SAMPLE 140


title: SAMPLE 134_632724SV151015HGU219B04

description: Liver biopsy  from Tolerant SAMPLE 134