title: MII oocyte, IP, rep1

status: Public on Feb 10 2023

first: Kin Fai

last: Au

state: MI

address: 100 Washtenaw Avenue

department: Department of Computational Medicine and Bioinformatics

data_processing:
Quality assessment of raw sequencing reads (both Input and IP samples) was performed using FastQC (v0.11.8), and sequencing adapters and low-quality bases were trimmed using Cutadapt (v1.8.1) with the parameter “-q 20,20 -m 20 --max-n 0.01 --trim-n”.
Trimmed reads were aligned to the mouse reference genome (mm10) using HISAT2 (v2.1.0) with the parameter “-5 8 --no-mixed --no-discordant”. Only the uniquely-aligned reads (i.e., only one genomic position per read pair was reported by HISAT2) were kept, and PCR duplicates were removed using SAMtools (v1.9) fixmate & markdup. After removing multiply-aligned read pairs and PCR duplicated read pairs, the genome coverage bigWig files (bin size =10 bp, normalized by RPKM/reads per kilobase per million reads) were generated by deepTools (v3.2.0) bamCoverage with the parameter “-bs 10 --normalizeUsing RPKM”.
Further, the read mates which overlap with ribosomal RNA loci (annotated by GENCODE vM23) were removed using BEDTools (v2.28.0) intersect. m6A peaks (IP versus the matched Input) were called using MACS2 (v2.1.2) callpeak with the parameter “-g 242010196 --keep-dup all -B --nomodel --call-summits”, with the cutoff q value <0.05.
Genome_build: mm10
Supplementary_files_format_and_content: The bigWig files are generated by deepTools (v3.2.0) bamCoverage with the bin size=10bp and normalized by RPKM. The bed files are m6A peaks (q value <0.05), generated by MACS2 (v2.1.2) callpeak. Both the bigWig and bed files can be loaded into genome browser, such as UCSC or IGV, for visualizing the read density and m6A peak regions, respectively.

source_name: MII oocyte

growth_protocol: Mice were housed with a 12 h light/dark cycle (7 am to 7 pm), with 55% relative humidity at 22°C, with free access to food and water.

extract_protocol:
Step 1. One-tube rRNA and DNA depletion. To deplete rRNA, we used the NEBNext® rRNA Depletion Kit (NEB) according to the user manual with minor modifications: add 3 µl RNA/probe master mix to a 12 µl sample; place samples in a thermocycler at 95°C for 2 min and then gradually ramped the temperature (−0.1°C/sec) to 22°C, held samples at 22°C for 5 min; add 5 µl RNase H reaction mix to the samples immediately, incubate at 37°C for 30 min and then place on ice. Then, to remove DNA, 30 µl DNase I digestion mix was added to the samples and incubated at 37°C for 30 min. Samples were purified with 2.2X volume of RNAClean XP beads. Freshly prepared ethanol (80%) was used to wash the samples twice. After rRNA and DNA depletion, the RNA was eluted from the beads with 78 μl nuclease-free water (Thermo Fisher Scientific). Then, 2 μl RiboLock RNase inhibitor (40 U/µl) was added to the samples.
Step 2. RNA shearing by sonication. The samples were sonicated for 2.5 × 30 sec using a UP100H Ultrasonic Processor (Hielscher) fitted with a 2-mm probe using pulse settings with 0.5 sec cycles and 27% power. Each sonication cycle was 30 seconds sonication plus 30 seconds on ice. After sonication, taking 7 µl for regular RNA-seq sequencing as Input control, the remaining 73 μl was subjected to immunoprecipitation. After that, 7 µl nuclease-free water and 20 µl 5X IP buffer (50 mM Tris-HCl pH 7.5, 750 mM NaCl, 0.5% (vol/vol) NP-40, 5 U/µl RiboLock RNase inhibitor) were added to the 73 μl samples. The final volume is 100 µl.
Step 3. Immunoprecipitation and washes. Dynabeads (30 μl, Invitrogen) were washed with 1X IP buffer (200 µl 5X IP buffer with 800 µl nuclease-free water) twice. Premixed antibody mix (4 µl m6A antibody (Millipore, ABE572), 16 µl 5X IP buffer, 60 µl nuclease-free water) was added to the washed beads, and the samples were incubated with rotation on a HulaMixer at 4°C overnight. Preincubated antibody-bead complexes were washed in 200 μl 1X IP buffer by vortexing. A second wash was performed by adding 200 μl 1X IP buffer, vortexing briefly, aliquoting 10 μl into PCR tubes for each sample, pulse spinning and placing tubes back to a magnet until the solution was clear. Supernatants were discarded, and sonicated RNA mix (100 μl) was added to the antibody-bead complex. The samples were incubated with rotation on a HulaMixer at 4°C for 2 h. After incubation, the supernatants were removed. The beads were washed 4 times in order with the following buffers: first-round wash with cold medium RIPA buffer (10 mM Tris–HCl, pH 8.0, 300 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% (vol/vol) Triton X-100, 0.2% (vol/vol) SDS, and 0.1% (vol/vol) Na-deoxycholate); second- and third-round washes with cold stringent RIPA buffer (10 mM Tris–HCl, pH 8.0, 350 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% (vol/vol) Triton X-100, 0.23% (vol/vol) SDS, 0.1% (vol/vol) Na-deoxycholate); last-round wash with cold medium RIPA buffer. After four washes, the beads were resuspended in 100 μl 1X IP buffer to recover for 5 min. The samples were placed on a magnet, and the supernatants were removed. The pellet was resuspended in 147.9 μl Elution buffer (5 mM Tris–HCl, pH 7.5, 1 mM EDTA, 0.05% (vol/vol) SDS, and 1 U/µl RiboLock RNase inhibitor). After adding 2.1 μL Proteinase K (NEB), the samples were immediately put in a Thermomixer at 1200 rpm, 55°C for 1.5 h. After incubation, the enzyme was inactivated through briefly spinning the tubes and incubating in a Thermomixer at 80°C for 20 min. After inactivation, the samples were placed on a magnet for 2-3 min. The supernatant (~150 μl) containing the immunoprecipitated RNA was transferred into a new 1.5 ml low-binding tube. We further resuspended the beads in 147.9 μl Elution buffer; added 2.1 μl of Proteinase K and placed immediately in a Thermomixer at 1200 rpm, 55°C for 5 min and then at 80°C for 20 min; placed back on a magnet for 2-3 min, and transferred the supernatant (~150 μl) to the previous 1.5 ml low-binding tube to pool both volumes together, yielding approximately 300 μl.
Step 4. Ethanol precipitation. An appropriate volume of nuclease-free water (~100 μl) was added to the immunoprecipitated RNA to a final volume of 400 μl, followed by adding 40 μl 3 M Sodium Acetate pH 5.2 (Thermo Fisher Scientific), 10 μl Linear acrylamide 5 mg/μl (Thermo Fisher Scientific), 1000 μl ice-cold 100% ethanol in the order. Samples were heavily vortexed and incubated immediately at −80°C for at least 2 h until completely frozen. After centrifugation at 20000 xg at 4°C for 15 min, the supernatant was carefully removed without disrupting the pellet. The pellet was washed twice with 1 ml ice-cold 75% ethanol and resuspended in 7 μl nuclease-free water.
For both Input and immunoprecipitated (IP) samples, the SMART-Seq® Stranded Kit (Takara) was used to prepare a library according to the manufacturer’s protocol with some modifications. The Option 2 protocol was followed without fragmentation for highly fragmented RNA. After PCR1 amplification and AMPure bead purification, 46.5 µl nuclease-free water was added to the beads, the ribosomal cDNA depletion protocol was skipped in Section D, the tubes were removed from the magnetic rack and mixed thoroughly by vortexing to resuspend the beads. The sample was incubated at room temperature for 5 min to rehydrate and recover 46 µl of sample. Then, we followed the protocol until sequencing. Libraries were quantified with KAPA Library Quantification Kits (Roche), and the size distribution was checked using TapeStation D1000 ScreenTape (Agilent Technologies). The equally pooled libraries were sequenced using a NovaSeq system (Illumina) in 100 bp paired-end mode.

treatment_protocol:
For GV oocytes, 8-week-old female mice were injected with 5 units (U) of pregnant mare serum gonadotropin (PMSG), and oocytes were isolated by puncturing the follicles of dissected ovaries at 48 h after PMSG injection. The isolation was performed in at 37°C M2 medium with 0.2 mM cyclic nucleotide phosphodiesterase inhibitor 3-Isobutyl-1-methylxanthine (IBMX, Sigma) added to prevent the oocytes from undergoing germinal vesicle breakdown (GVBD). Cumulus cells were removed by gentle pipetting. Next, the GV oocytes were sorted into non-surrounded nucleolus (NSN) and surrounded nucleolus (SN) stages by looking for the presence of a perivitelline space (gap between the oolemma and the zona pellucida) within 1 h of in vitro culture with IBMX. SN-type oocytes (used in this study) were then exposed to acidic Tyrode’s solution (pH 2.5, Sigma) for a few seconds to remove the zona pellucida followed by three washes in M2 medium. The oocytes (SN stage) were then manually picked and sorted into 12 μl 1X Lysis buffer (Takara).
For MII oocytes, 8-week-old female mice were injected with 5 U of PMSG followed by injecting 5 U of human chorionic gonadotropin (hCG) 45 h later. MII oocytes with cumulus mass were released from the oviduct ampullae 14 h after hCG injection. Cumulus cells were dispersed by 0.3 mg/ml hyaluronidase in HEPES-M2 medium. Oocytes were exposed to acidic Tyrode’s solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. The oocytes were then manually picked and sorted into 12 μl 1X Lysis buffer.
For embryos, 8-week-old female mice were superovulated (5 U PMSG followed by 5 U hCG 45 h later), mated with 8-week-old male mice. Embryos at a particular stage were flushed from the reproductive tract at defined time periods after hCG administration: 27-28 h (zygote), 39-43 h (2-cell), 68-70 h (8-cell), and 92-94 h (blastocyst) in HEPES-buffered CZB medium. Embryos were transferred to acidic Tyrode’s solution for a few seconds to remove the zona pellucida followed by three washes in M2 medium. The embryos were then manually picked and sorted into 12 μl 1X Lysis buffer.

value: oocyte

tag: developmental stage

value: MII oocyte

tag: oocyte/embryo number

tag: fraction

value: rRNA-free RNA

tag: rip antibody

value: m6A  (Millipore, ABE572)


title: Zygote, Input, rep1

type: SRA

city: Ann Arbor

country: USA

institute: University of Michigan

library_source: transcriptomic

molecule: total RNA

organism: Mus musculus

source_name: Zygote

tag: cell type

value: embryo

value: Zygote

tag: strain


title: MII oocyte, IP, rep2


title: Zygote, Input, rep2


title: HCT116 cells, siControl-MG132

status: Public on Mar 18 2023

type: RNA

city: Kyotanabe

first: Akira

last: Kobayashi

address: 1-3 Tatara Miyakodani

country: Japan

institute: Doshisha University

hyb_protocol: Samples were hybridized to the Affymetrix Human Gene 1.0 ST Arrays. GeneChip fluidics station 450 was used for processing of the arrays.

scan_protocol: Array scanning was performed with the GeneChip scanner 3000-7G.

data_processing:
Images were analyzed with the GeneChip operating software (Affymetrix). Expression console and Transcription analysis console (Affymetrix) were used to identify the gene expression.
RMA expression value derived from Expression Console software

label: Biotin

organism: Homo sapiens

source_name: HCT116 cells

label_protocol: Total RNA was processed with the Ambion WT Expression Kit (Affymetrix) accord- ing to the manufacturers’ instructions. cRNA was then fragmented, labelled using the GeneChip WT Terminal Labeling Kit (Affymetrix).

growth_protocol: HCT116 cells were cultured in DMEM/high glucose medium (Wako) supplemented with 10% FBS (Nichirei Biosciences), 40 µg/ml streptomycin, and 40 units/ml penicillin (Wako). Cells were cultured in 37˚C with 5% CO2 incubator.

extract_protocol: RNA was extracted and purification using ISOGEN II (NIPPON GENE) according to the manufacturer's protocol

treatment_protocol: Transfection of siRNA was performed using RNAiMAX (Invitrogen). After 48 h from siRNA transfection, cells were treated with DMSO or 1 µM MG132 (Peptide Institute) for 16 h.

tag: tissue

value: colon

tag: Sex

value: male

tag: cell line

tag: genotype

value: control

tag: treatment


title: GV oocyte, Input, rep2

source_name: GV oocyte

value: GV oocyte


title: MII oocyte, Input, rep1


title: GV oocyte, IP, rep1


title: GV oocyte, IP, rep2


title: 220228_L2 wild-type infected rep3

status: Public on Feb 13 2023

city: Omaha

first: Rey

last: Carabeo

state: Nebraska

address: 985900 Nebraska Medical Center

department: Pathology and Microbiology

institute: University of Nebraska Medical Center

data_processing:
CLC Genomics Workbench version 10.1.1 used for all data processing steps.
Adaptor sequences and low-quality reads were trimmed using default parameters, then aligned to the hg19 reference genome.
Transcripts per million (TPM) were calculated using default parameters.
An initial ANOVA-like pass of differential expression testing was performed. Genes that did not significantly vary between any of the three treatments were culled from downstream analysis.
Differential expression testing was performed using default paramters to identify differentially expressed genes in either L2-wt or L2ΔCTL0480-infected samples relative to the mock-infected control, accounting for variation between replicates.
Assembly: hg19
Supplementary files format and content: CLC file containing raw counts, TPM, and FDRP values for every gene and sample.

molecule: polyA RNA

source_name: Primary endocervical epithelial cells

growth_protocol: Human primary endocervical epithelial cells (PCS-480-011, Lot 80306190) were cultured at 37C with 5% atmospheric CO2 in Cervical Epithelial Cell Basal Medium (CECBM, ATCC PCS-480-032) supplemented with all contents of the Cervical Epithelial Growth Kit (ATCC PCS-080-042), per manufacturer's instructions. Chlamydia trachomatis serovar L2/434/Bu was originally obtained from Dr. Ted Hackstadt (Rocky Mountain National Laboratory, NIAID), and the L2ΔCTL0480 mutant strain was provided by Dr. Derek Fisher and Dr. Erika Lutter.

extract_protocol:
Primary cells were harvested for Total RNA at 24 hpi using TRIzol (Thermo Fisher Scientific 15596026) and the DNA-free DNA removal kit (Thermo Fisher Scientific AM1906), according to manufacturers’ protocols, and were subsequently assayed for fragmentation using an Agilent Bioanalyzer 2100.
cDNA library preparation of intact samples was performed using the NuGEN Universal mRNA-Seq Library Preparation kit, according to the manufacturer's protocol.

treatment_protocol: Primary cells cultured to confluence (6-7 days) seeded on fibronectin-coated 6-well plates (Corning 354402) were washed with pre-warmed Hanks Buffered Saline Solution (HBSS) prior to inoculation with Chlamydia-containing CECBM at a multiplicity of infection (MOI) of 2. Tissue culture plates were centrifuged at 4C and 500 rcf (Eppendorf 5810 R tabletop centrifuge, A-4-81 rotor) for 15 minutes to synchronize infection. Inoculum was then aspirated, and cells were washed with chilled HBSS prior to the addition of pre-warmed KSFM. Infected cultures were then returned to the tissue culture incubator until 24 hours post-infection.

value: Primary endocervical epithelial cells

value: L2 wild-type infected

tag: infection time

value: 24 hpi


title: GV oocyte, Input, rep1


title: MII oocyte, Input, rep2


title: Zygote, IP, rep1


title: Zygote, IP, rep2


title: Blastocyst, IP, rep2

source_name: Blastocyst

value: Blastocyst


title: 8-cell, IP, rep2


title: Blastocyst, Input, rep1


title: HCT116 cells, siNRF1-DMSO

value: NRF1 KD

value: DMSO


title: 8-cell, IP, rep1


title: Blastocyst, Input, rep2


title: 8-cell, Input, rep2


title: 2-cell, Input, rep1


title: 2-cell, Input, rep2


title: 8-cell, Input, rep1


title: 2-cell, IP, rep1


title: Blastocyst, IP, rep1


title: human induced pluripotent stem cell-derived 3D heart organoids D15

status: Public on Feb 04 2023

city: Munich

first: Gianluca

last: Santamaria

state: Germany

address: Ismaninger Strasse 22

department: Department of Medicine I

country: Germany

institute: Klinikum rechts der Isar – Technical University Munich

data_processing:
Cell Ranger workflow using the DropletUtils was used to generate FASTQ and bam files
Genome_build: hg19

source_name: cardiac organoids

growth_protocol: Human embryonic stem cells were differentiated as described in methods of Meier et al., (2021) using a stepwise growth-factor based protocol  to direct hiPSCs towards CMs through defined developmental stages of primitive-like cells and early cardiovascular progenitor cells.

extract_protocol:
Epicardioids were dissociated with papain as described in methods of Meier et al. (2021), filtered through 40 μm filter, washed with 5 mL 1% BSA (Gibco; 15260037) in PBS-/-, centrifuged for 3 min at 200 g, and resuspended in 0.04% BSA in PBS for counting. After counting, 8 000 cells for each sample (targeted cell recovery) were processed using Chromium Single Cell 3’ GEM, Library & Gel Bead Kit v3 (10x Genomics; PN-1000092), the Chromium Chip B Single Cell Kit (10x Genomics; PN- 1000073), and the Chromium i7 Multiplex Kit v2 (10x Genomics; PN-120262) for samples d2-d15 and Chromium Next GEM Single Cell 3’ Library & Gel Bead Kit v3.1 (1000128, 10x Genomics) Chromium Single Cell G Chip Kit (1000127, 10x Genomics) and Single Index Kit T Set A (PN-1000213, 10x Genomics) for sample d30 to generate Gel Bead-In-EMulsions (GEMs) and single cell sequencing libraries.
8 000 cells for each sample (targeted cell recovery) were processed using Chromium Single Cell 3’ GEM, Library & Gel Bead Kit v3 (10x Genomics; PN-1000092), the Chromium Chip B Single Cell Kit (10x Genomics; PN- 1000073), and the Chromium i7 Multiplex Kit v2 (10x Genomics; PN-120262) for samples d2-d15 and Chromium Next GEM Single Cell 3’ Library & Gel Bead Kit v3.1 (1000128, 10x Genomics) Chromium Single Cell G Chip Kit (1000127, 10x Genomics) and Single Index Kit T Set A (PN-1000213, 10x Genomics) for sample d30 to generate Gel Bead-In-EMulsions (GEMs) and single cell sequencing libraries. Libraries were pooled and sequenced using the NovaSeq (Ilumina; S1 flow cell) with 150bp paired end reads with 28 cycles for read1, 91 cycles for read2 8 cycles for i7 and 0 cycles for i5, and with a read depth of at least 25-30 000 pair reads per cell.

value: in vitro differentiated human induced pluriotent stem cells (hiPSC)

value: Human Pluripotent Stem Cell Registry ID:MRIi003-A

tag: time

value: Day 15


title: human induced pluripotent stem cell-derived 3D heart organoids D30

value: Day 30


title: human induced pluripotent stem cell-derived 3D heart organoids D10

value: Day 10


title: 2-cell, IP, rep2

value: embryo

value: rRNA-free RNA

value: m6A  (Millipore, ABE572)


title: Victoria, skin, ripen stage, rep2

status: Public on Jan 31 2023

city: Zhengzhou

first: Yongzhou

last: Li

address: No. 28, Gangwan Road, Guancheng District

country: China

institute: Chinese Academy of Agricultural Sciences Zhengzhou Fruit Research Institute

data_processing:
The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v4-cBot-HS (Illumia) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina platform and paired-end reads were generated.
The raw reads were further processed with a bioinformatic pipeline tool, BMKCloud(www.biocloud.net) online platform.
Raw data (raw reads) of fastq format were firstly processed through in-house perl scripts. In this step, clean data(clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30, GC-content and sequence duplication level of the clean data were calculated. All the downstream analyses were based on clean data with high quality.
The adaptor sequences and low-quality sequence reads were removed from the data sets. Raw sequences were transformed into clean reads after data processing. These clean reads were then mapped to the reference genome sequence. Only reads with a perfect match or one mismatch were further analyzed and annotated based on the reference genome. Hisat2 tools soft were used to map with reference genome.
Quantification of gene expression levelsGene expression levels were estimated by fragments per kilobase of transcript per million fragments mapped.
Assembly: StringTie
Supplementary files format and content: tab-delimited text file includes FPKM values for each Sample

organism: Vitis vinifera

source_name: skin

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen). 1ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols.

tag: cultivar

value: Victoria

value: skin

tag: Stage

value: ripening


title: Victoria, skin, ripen stage, rep1


title: Centennial Seedless, skin, ripen stage, rep2

value: Centennial Seedless


title: Centennial Seedless, skin, ripen stage, rep1


title: WIS HYP_SE4

status: Public on Feb 02 2023

city: Inzai

first: Thu

middle: Van

last: NGUYEN

state: Other

address: Hirakagakuendai 1-1

institute: Juntendo University

description: Gene expression profile in the rat hypothalamus after stress and exercise interventions

hyb_protocol: 0.6ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60ºC for 30 minutes with 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Sureprint G3 Rat GE 8x60k v2 (Agilent) Microarrays for 17 hours at 65ºC in a rotating Agilent hybridization oven. After hybridization, microarrays were washed for 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37ºC GE Wash buffer 2 (Agilent), then dried.

scan_protocol: Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner using one color scan setting for Sureprint G3 unrestricted GE 8x60 (G4858A) slides. Agilent DNA Microarray Scanner (G2600D) using one color scan setting for 8x60k array slides (Scan Area 61x21.6 mm, Profile AgilentHD_GX_1 color, Scan resolution 3um, Dye channel is set to Green and Green PMT is set to 100%)

data_processing: The scanned images were analyzed with Feature Extraction Software 9.1 (Agilent) using default parameters (protocol GE1_107_Sep09 and Grid: 074036_D_F_20180103) to obtain background subtracted and spatially detrended Processed Signal intensities. Processed signal intensities were normalized by the global scaling method. A trimmed mean probe intensity was determined by removing 2% of the lower and the higher end of the probe intensities in order to calculate the scaling factor. Normalized signal intensities were then calculated from the target intensity on each array using the scaling factor, so that the trimmed mean target intensity of each array was arbitrarily set to 2500.

organism: Rattus norvegicus

source_name: Wistar, Hypothalamus, stress+exercise

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng of RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA).  Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

growth_protocol: Stress rats (St and SE, n=12) were submitted to restraint stress (1-1.5h , 5 days per week) for 3 weeks, while control rats (Co, n=6) rats were not. Stress bined with exercise rats (SE, n=6) were allowed to run spontaneously in a wheel for 4 weeks (one week before three-week stress intervention and during it), while stress and control rats (St and Co, n=12) were not.

extract_protocol: The tissues were homogenized in TRIzol reagent and stored at −80ºC before being processed. Total RNAs from each sample were isolated according to the protocol supplied by the manufacturer (Life Technologies) and stored at −80ºC. The purity (260/230 and 260/280 ratio >1.7) and integrity (RIN > 7) for all total RNA samples were verified by using a Nanodrop1000 and a 2100 Bioanalyzer Agilent respectively. 100 ng of total RNA from each rat were used for labeling with Cyanine-3 in microarray experiment.

treatment_protocol: All animals were allowed to eat and drink ad libitum until the day of the tissue collection. Animals were killed by ether inhalation followed by decapitation. The brain was rapidly removed from the cranium, placed on an ice-cold platform, the hypothalamus was dissected under a dissecting microscope, and stored at -80ºC until processing. A single operator performed all dissections.

value: Hypothalamus

tag: gender

value: Male

tag: strains

value: Wistar

tag: age

value: 9 weeks old


title: Summer Black, skin, ripen stage, rep2

value: Summer Black


title: Summer Black, skin, ripen stage, rep1


title: WIS HYP_St1

source_name: Wistar, Hypothalamus, stress


title: Summer Black, skin, ripen stage, rep3


title: WIS HYP_Co1

hyb_protocol: 0.6ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60ºC for 30 minutes with 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Sureprint G3 Rat GE 8x60k v2 (Agilent) Microarrays for 17 hours at 65ºC in a rotating Agilent hybridization oven.  After hybridization, microarrays were washed for 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37ºC GE Wash buffer 2 (Agilent), then dried.

source_name: Wistar, Hypothalamus, control

treatment_protocol: All animals were allowed to eat and drink ad libitum until the day of the tissue collection. Animals were killed by ether inhalation followed by decapitation. The brain was rapidly removed from the cranium, placed on an ice-cold platform, the hypothalamus was dissected under a dissecting microscope, and stored at -80ºC until processing.  A single operator performed all dissections.


title: WIS HYP_SE6


title: WIS HYP_Co5


title: WIS HYP_St3


title: WIS HYP_Co4


title: WIS HYP_St6


title: Midknight Beauty, skin, ripen stage, rep3

value: Midknight Beauty


title: WIS HYP_St2


title: WIS HYP_Co2

growth_protocol: Stress rats (St and SE, n=12) were submitted to restraint stress (1-1.5h , 5 days per week) for 3 weeks, while control rats (Co, n=6) rats were not. Stress combined with exercise rats (SE, n=6) were allowed to run spontaneously in a wheel for 4 weeks (one week before three-week stress intervention and during it), while stress and control rats (St and Co, n=12) were not.


title: WIS HYP_Co6


title: Centennial Seedless, skin, ripen stage, rep3


title: Victoria, skin, ripen stage, rep3


title: WIS HYP_Co3

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 200 ng of RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

treatment_protocol: All animals were allowed to eat and drink ad libitum until the day of the tissue collection. Animals were killed by ether inhalation followed by decapitation. The brain was rapidly removed from the cranium, placed on an ice-cold platform, the hypothalamus was dissected under a dissecting microscope, and stored at -80ºC until processing. A single operator performed all dissections


title: WIS HYP_SE2


title: WIS HYP_St5


title: WIS HYP_SE5


title: WIS HYP_SE3


title: WIS HYP_St4


title: WIS HYP_SE1


title: Midknight Beauty, skin, ripen stage, rep2


title: Midknight Beauty, skin, ripen stage, rep1


title: Shine muscat, skin, ripen stage, rep3

value: Shine muscat


title: Shine muscat, skin, ripen stage, rep2


title: Shine muscat, skin, ripen stage, rep1


title: Smn_ax_5

status: Public on Jan 05 2023

city: Würzburg

first: Silke

last: Appenzeller

address: Schweinfurter Str. 28

institute: University Wuerzburg

description: Gene expression data from axonal compartment of cultured Smn knockdown motoneurons

data_processing:
Quality Assessment (FASTQC version 0.10.1)
Adapter Clipping and Collapsing (inhouse script)
Alignment (Star version 2.4.0d)
Calculation of expression and differential expression (Cufflinks package version 2.2.1)
Genome_build: Mus_musculus.GRCm38.75
Supplementary_files_format_and_content: genes.fpkm_tracking_annotated.xlsx

growth_protocol: Primary mouse motoneurons were cultured in microfluidic chambers as described in Saal et al. (RNA 20: 1789).

extract_protocol:
Total RNA was extracted from the somatodendritic and the axonal compartment using the Arcturus PicoPure RNA Isolation Kit (Applied Biosystems) with 10 μl elution volume.
For whole transcriptome profiling see Briese et al. (Nucleic Acids Res, doi: 10.1093/nar/gkv1027)

tag: strain background

value: Primary mouse motoneuron culture


title: Mouse sagittal brain sample

status: Public on Jan 12 2023

city: Acton

first: Lixinyu

last: Liu

state: ACT

address: 131 Garran road

country: Australia

institute: John Curtin School of Medical Research

data_processing:
The raw data is processed by SAW (https://github.com/BGIResearch/SAW) to generate spatial gene expression matrix.
The spatial gene expression matrix is pre-processed to remove low quality counts using stereo-py (https://github.com/BGIResearch/stereopy)
The filtered matrix undergos Gaussian smoothing based on steroepy(https://github.com/BGIResearch/stereopy)
The smoothed and normalised matrix then is normalised and used for clustering.
Assembly: mm10
Supplementary files format and content: Gaussian smoothed data with clustering is in h5ad format
Supplementary files format and content: The processed spatial expression matrix is in gef format.
Library strategy: Stereo-seq

source_name: brain

extract_protocol: 10um sagittal brain slices

value: brain


title: Smn_sd_3

description: Gene expression data from somatodendritic compartment of cultured Smn knockdown motoneurons


title: Smn_sd_4

source_name: somatodendritic compartment of cultured Smn knockdown motoneurons


title: Smn_ax_6

source_name: axonal compartment of cultured Smn knockdown motoneurons


title: Smn_sd_6


title: Smn_sd_5


title: Smn_ax_4


title: Smn_ax_3


title: Smn_sd_2


title: Smn_ax_2


title: Smn_ax_1


title: In vitro haustoria, untreated RNA, replicate 2

status: Public on Jan 03 2023

city: University Park

first: Michael

middle: J

last: Axtell

state: PA

address: 280 Mueller Lab

department: Biology

institute: Pennsylvania State University

data_processing:
3' adapters were removed with cutadapt, discarding reads that were less than 15 nts after trimming. An example command: cutadapt -j 4 -a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A GATCGTCGGACTGTAGAACTCTGAACGGAAAGAGTGT -o 1d_R1_trimmed.fastq.gz -p 1d_R2_trimmed.fastq.gz -m 15 1d_S1_R1_001.fastq.gz 1d_S1_R2_001.fastq.gz > 1d_cutadapt.txt
Adapter-trimmed reads were aligned to the Cuscuta campestris v0.32 genome assembly (Vogel et al.) using bowtie2, and converted to sorted bam format. An example command: bowtie2 -p 6 -x cuscuta_campestris_assembly_r0-32.fasta -1 3d_R1_trimmed.fastq.gz -2 3d_R2_trimmed.fastq.gz 2> 3d_bowtie2_genome.txt | samtools view -b - | samtools sort -T 3d - > 3d_genome.bam
Interface-induced microRNA primary transcripts were discovered from the aligned data. Quantifications of the primary transcripts were done using a samtools view / bedtools bamtobed pipeline which counted transcripts as full-length concordant read pairs at the annotated coordinates. Transcript numbers were then converted in transcripts per million (TPM). Only transcripts exactly matching the annotated locations were used for analysis.
Capped snRNA (U1, U2, U4, and U5) loci were annotated in the C. campestris genome by homology to Arabidopsis thaliana snRNAs. Observed transcripts were then tallied with a samtools view / bedtools bamtobed pipeline. Full-length concordant read-pairs whose start and end were within +/- 3nts of the expected locations were tallied. Tallies were then converted to transcripts per million (TPM).
tRNA loci were annotated in the C. campestris genome using tRNAScan-SE 2.0. Pre-tRNAs were considered to be any observed transcripts with a 5' end between 5 and 15 nts upstream of the predicted mature tRNA's 5' end. The 3'-ends of valid pre-tRNAs were required to be anywhere between -5 and +15 relative to the predicted mature tRNA 3 end. Qualifying transcripts (supported by conccordant read-pairs) were tallied, and the resulting tallies converted in transcripts per million (TPM).
Assembly: Version 0.32, as described by Vogel et al. (10.1038/s41467-018-04344-z) and retrieved from https://www.plabipd.de/project_cuscuta2/start.ep
Supplementary files format and content: A single file, 'TPMs.tsv', is a tab-delimited file containing all results. Format is similar to bed for the first 6 columns, with column 5 (normally the 'Score') containing the number of observed transcripts. Additional columns are self-explanatory. Data are organized in "tidy" format, with one observation per row. The last column, "LibraryName" contains the library name which links back to one of the 9 samples.

organism: Cuscuta campestris

source_name: In vitro haustoria (host-free)

growth_protocol: Cuscuta campestris were cultivated on beets. Shoot tips 4-6cm in length were cut and stimulated to form haustoria by treatment with far-red light and physical pressure (stack of microscope slides pressing the tips against 3% agar) for 4 days. The regions with visible haustoria were dissected with a scalpel and collected for RNA extractions. 3 RNA samples were collected (numbered 1, 2, and 3).

extract_protocol:
Total RNA was extracted using the Zymo Quick-RNA Plant kit per the manufacturer's instructions.
Each RNA was treated in 3 different ways: Dcp ("Decapping Enzyme" from NEB), RppH (NEB), or no treatment, thus producing the nine samples. After treatments, libraries were constructed using a splint-ligation protocol as described for microRNAs by Maguire et al. (10.1093/nar/gkaa480) except that no size-selection was performed. The resulting libraries include full-length, stranded RNAs between ~10 and ~400 nts in length. The libraries require a 5' monophosphate, so that the abundance in the Dcp or RppH treatments can give an indication as to the identity of the in vivo 5'-ends. Specifically, RNAs enriched in both RppH and Dcp are likely capped, while RNAs enriched only in RppH are likely 5'-ppp.

value: In vitro haustoria (host-free)

value: wild-type strain "Dotti"


title: In vitro haustoria, RppH-treated RNA, replicate 3

value: RppH


title: In vitro haustoria, Dcp-treated RNA, replicate 3

value: Dcp


title: Rapal_H3K79me3

status: Public on Dec 10 2022

city: wuhan

first: Yu

last: Qi

state: Hubei

address: xujiapeng

department: College of life sciences

institute: Hubei university

library_source: genomic

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to sacCer3 whole genome using bowtie v0.12.2 with parameters -q -p 5 -e 100 -y -a -m 10 --best --strata
Assembly: sacCer3
Supplementary files format and content: bigWig files

molecule: genomic DNA

organism: Saccharomyces cerevisiae

growth_protocol: Cells were grown in YPD until mid-log phase.

extract_protocol:
Lysates were clarified from sonicated yeast cell pellets and histone-DNA complexes were isolated with indicated antibodies.
Libraries were prepared using the HTP KAPA kit.

treatment_protocol: No treatment.

tag: chip antibody

value: H3K79me3 (Abclonal, A2369)


title: In vitro haustoria, Dcp-treated RNA, replicate 2


title: In vitro haustoria, Dcp-treated RNA, replicate 1


title: Rapa_H3

value: H3 (Abcam, ab1791)


title: control_H3


title: In vitro haustoria, RppH-treated RNA, replicate 1


title: In vitro haustoria, RppH-treated RNA, replicate 2


title: Rapa_WTH3


title: Smn_sd_1


title: Rapa_H3T11p

value: H3Tp11 (Abcam, ab5168)


title: control_H3K79me3


title: control_H3T11p


title: Patient 20, 25, 34 lesional atopic dermatitis, lichen planus, rep 1 [21L008970]

status: Public on Nov 14 2022

first: Christina

last: Hillig

address: Ingolstaedter Landstraße 1

department: ICB

institute: Helmholtz Center Munich

description: 10x Genomics

data_processing:
Spots were partially manually aligned and data was processed using the Space Ranger Software v1.0.0 from 10x Genomics
Assembly: hg38_ensrel99
Supplementary files format and content: .h5 files

growth_protocol: Skin biopsies were obtained and immediately snap frozen in liquid nitrogen. Samples were then stored at -80°C until further preparation.

extract_protocol:
Upon cryosectioning, samples were equilibrated to cryostat chamber temperature and covered in optimal cutting temperature compound. Sections were taken at 10 µm thickness and directly placed onto the Visium Spatial Gene Expression slide (10x Genomics). Slides were processed using the Visium Spatial Gene Expression Kit (10x Genomics) following the CG000239 Visium Spatial Gene Expression Reagent Kits - User Guide RevA.
Libraries were constructed following the the CG000239 Visium Spatial Gene Expression Reagent Kits - User Guide RevA by 10x Genomics.

value: skin

tag: disease

value: atopic dermatitis, lichen planus

tag: tissue type

value: lesional

tag: patient


title: Patient 35, 36 lesional atopic dermatitis, rep 1 [21L008972]

description: 10x Genomics

tag: tissue

value: atopic dermatitis


title: Patient 35, 36 lesional atopic dermatitis, rep 2 [21L008973]

description: 10x Genomics


title: Patient 27, 37 lesional lichen planus, rep 1 [21L008968]

description: 10x Genomics

value: lichen planus


title: Patient 26, 30 lesional lichen planus, rep 1 [21L008966]

description: 10x Genomics


title: Patient 20, 25, 34 lesional atopic dermatitis, lichen planus, rep 2 [21L008971]

description: 10x Genomics


title: Patient 26, 30 lesional lichen planus, rep 2 [21L008967]

description: 10x Genomics


title: In vitro haustoria, untreated RNA, replicate 3


title: Patient 27, 37 lesional lichen planus, rep 2 [21L008969]

description: 10x Genomics


title: Patient 28, 33 lesional psoriasis, lichen planus, rep 2 [21L008965]

description: 10x Genomics

value: psoriasis, lichen planus


title: In vitro haustoria, untreated RNA, replicate 1


title: Patient 28, 33 lesional psoriasis, lichen planus, rep 1 [21L008964]

description: 10x Genomics


title: Patient 22, 19 lesional psoriasis, rep 2 [21L008959]

description: 10x Genomics

value: psoriasis


title: Patient 29 lesional pityriasis rubra pilaris / psoriasis, rep 2 [21L008961]

description: 10x Genomics

value: pityriasis rubra pilaris / psoriasis


title: Patient 31, 32 lesional psoriasis, rep 2 [21L008963]

description: 10x Genomics


title: Patient 15 lesional atopic dermatitis, rep 1 [P16357_1053]

description: 10x Genomics


title: Patient 29 lesional pityriasis rubra pilaris / psoriasis, rep 1 [21L008960]

description: 10x Genomics


title: Patient 15 lesional atopic dermatitis, rep 2 [P16357_1054]

description: 10x Genomics


title: Patient 14 non lesional lichen planus, rep 2 [P16357_1050]

description: 10x Genomics

value: non lesional


title: HCT116, input

status: Public on Mar 19 2023

data_processing:
Basecalls were performed using BCL2FASTQ Conversion Software 1.8.4 in the CASAVA 1.8.2 pipeline.
Mapping was performed by Bowtie2 (version 2.2.6) using the reference human genome, NCBI build 37 (hg19) with default settings.
ChIP-peaks were called by MACS (version 1.4.2) with default settings.
Assembly: hg19
Supplementary files format and content: BED, Wiggle (except for input sample)

extract_protocol:
Cells were fixed in 1% formaldehyde for 10 min, followed by quenching with 125 mM glycine and two washes with phosphate-buffered saline (PBS). The cells were lysed in cell lysis buffer (5 mM Tris-HCl [pH 8.0], 85 mM KCl, 0.5% NP-40 and protease inhibitor cocktail (Nacalai Tesque)), and then centrifuged at 2,000 rpm for 3 min at 4˚C . The pellets were further lysed with nuclei lysis buffer (50 mM Tris-HCl[pH8.0], 10 mM EDTA, 1% SDS and protease inhibitor cocktail) followed by sonication using Bioruptor (Tosho Electric Co., Ltd.).
ChIP-seq libraries were prepared from 500 pg of immunoprecipitated DNA fragments using KAPA Hyper Prep Kit (KAPA Biosystems).

treatment_protocol: Cells were treated with 1 µM MG132 (Peptide Institute) for 16 h.

tag: cell type

value: Human colon cancer


title: Patient 14 non lesional lichen planus, rep 1 [P16357_1049]

description: 10x Genomics


title: Patient 13 lesional psoriasis, rep 2 [P16357_1048]

description: 10x Genomics


title: Patient 14 lesional lichen planus, rep 1 [P16357_1051]

description: 10x Genomics


title: control_WTH3

tag: strain


title: Patient 13 lesional psoriasis, rep 1 [P16357_1047]

description: 10x Genomics


title: HCT116, NRF1

tag: cell line


title: Patient 13 non lesional psoriasis, rep 2 [P16357_1046]

description: 10x Genomics


title: Patient 13 non lesional psoriasis, rep 1 [P16357_1045]

description: 10x Genomics


title: Patient 22, 19 lesional psoriasis, rep 1 [21L008958]

description: 10x Genomics


title: Patient 12 lesional lichen planus, rep 2 [P16357_1044]

description: 10x Genomics


title: Patient 12 lesional lichen planus, rep 1 [P16357_1043]

description: 10x Genomics


title: Patient 31, 32 lesional psoriasis, rep 1 [21L008962]

description: 10x Genomics


title: Patient 11 lesional atopic dermatitis, rep 2 [P16357_1040]

description: 10x Genomics


title: Patient 12 non lesional lichen planus, rep 1 [P16357_1041]

description: 10x Genomics


title: Patient 10 lesional psoriasis, rep 2 [P16357_1036]

description: 10x Genomics


title: Patient 11 lesional atopic dermatitis, rep 1 [P16357_1039]

description: 10x Genomics


title: Patient 11 non lesional atopic dermatitis, rep 2 [P16357_1038]

description: 10x Genomics


title: Patient 11 non lesional atopic dermatitis, rep 1 [P16357_1037]

description: 10x Genomics


title: Patient 14 lesional lichen planus, rep 2 [P16357_1052]

description: 10x Genomics


title: Patient 10 lesional psoriasis, rep 1 [P16357_1035]

description: 10x Genomics


title: Patient 10 non lesional psoriasis, rep 2 [P16357_1034]

description: 10x Genomics


title: Patient 10 non lesional psoriasis, rep 1 [P16357_1033]

description: 10x Genomics


title: Patient 9 lesional lichen planus, rep 1 [P16357_1031]

description: 10x Genomics


title: Patient 9 lesional lichen planus, rep 2 [P16357_1032]

description: 10x Genomics


title: Patient 9 non lesional lichen planus, rep 2 [P16357_1030]

description: 10x Genomics


title: Patient 8 lesional atopic dermatitis, rep 1 [P16357_1027]

description: 10x Genomics


title: Patient 8 lesional atopic dermatitis, rep 2 [P16357_1028]

description: 10x Genomics


title: Patient 8 non lesional atopic dermatitis, rep 2 [P16357_1026]

description: 10x Genomics


title: Patient 8 non lesional atopic dermatitis, rep 1 [P16357_1025]

description: 10x Genomics


title: Patient 7 lesional psoriasis, rep 1 [P16357_1023]

description: 10x Genomics


title: Patient 7 non lesional psoriasis, rep 2 [P16357_1022]

description: 10x Genomics


title: Patient 6 lesional lichen planus, rep 1 [P16357_1019]

description: 10x Genomics


title: Patient 6 lesional lichen planus, rep 2 [P16357_1020]

description: 10x Genomics


title: Patient 7 lesional psoriasis, rep 2 [P16357_1024]

description: 10x Genomics


title: Patient 12 non lesional lichen planus, rep 2 [P16357_1042]

description: 10x Genomics


title: Patient 5 lesional atopic dermatitis, rep 2 [P16357_1016]

description: 10x Genomics


title: Patient 6 non lesional lichen planus, rep 1 [P16357_1017]

description: 10x Genomics


title: Patient 5 non lesional atopic dermatitis, rep 1 [P16357_1013]

description: 10x Genomics


title: Patient 4 lesional psoriasis, rep 2 [P16357_1012]

description: 10x Genomics


title: Patient 4 non lesional psoriasis, rep 1 [P16357_1009]

description: 10x Genomics


title: Patient 4 lesional psoriasis, rep 1 [P16357_1011]

description: 10x Genomics


title: Patient 4 non lesional psoriasis, rep 2 [P16357_1010]

description: 10x Genomics


title: Patient 3 lesional lichen planus, rep 2 [P16357_1008]

description: 10x Genomics


title: Patient 5 non lesional atopic dermatitis, rep 2 [P16357_1014]

description: 10x Genomics


title: Patient 3 lesional lichen planus, rep 1 [P16357_1007]

description: 10x Genomics


title: Patient 3 non lesional lichen planus, rep 2 [P16357_1006]

description: 10x Genomics


title: Patient 3 non lesional lichen planus, rep 1 [P16357_1005]

description: 10x Genomics


title: Patient 9 non lesional lichen planus, rep 1 [P16357_1029]

description: 10x Genomics


title: Patient 2 lesional atopic dermatitis, rep 1 [P16357_1003]

description: 10x Genomics


title: Patient 2 non lesional atopic dermatitis, rep 1 [P16357_1001]

description: 10x Genomics


title: Patient 2 lesional atopic dermatitis, rep 2 [P16357_1004]

description: 10x Genomics


title: Patient 2 non lesional atopic dermatitis, rep 2 [P16357_1002]

description: 10x Genomics


title: Patient 1 lesional psoriasis, rep 1 [P15509_1003]

description: 10x Genomics


title: Patient 1 non lesional psoriasis, rep 2 [P15509_1002]

description: 10x Genomics


title: Patient 1 lesional psoriasis, rep 2 [P15509_1004]

description: 10x Genomics


title: Patient 1 non lesional psoriasis, rep 1 [P15509_1001]

description: 10x Genomics


title: Patient 6 non lesional lichen planus, rep 2 [P16357_1018]

description: 10x Genomics


title: Cl13D30+aPDL1

status: Public on Oct 31 2022

city: Philadelphia

first: E

middle: JOHN

last: WHERRY

address: 421 Curie Blvd,354 BRB II/III

department: Systems Pharmacology       and Translational Therapeutics,Institute for Immunology

institute: University of Pennsylvania

data_processing:
scATAC-seq sample demultiplexing, barcode processing, alignment, filtering, unique molecular identifier (UMI) counting were performed using 10x Cell Ranger ARC pipeline (2.0.0) and mm10 genome.
Downstream scATAC-seq analysis was performed in R (version 4.0.2) using Seurat (version 4.0.4), Signac (1.3.0), and ArchR (1.0.1)
Union peak list was generated using peaks called in ArchR and using macs2 with sample bam files.
Assembly: GRCm38/mm10
Supplementary files format and content: Filtered peak-barcode matrices containing only cellular barcodes

source_name: P14 CD8 T cells from LCMV Clone13 infection+anti PDL1 treatment

extract_protocol:
Cells were loaded onto the Chromium Controller (10x Genomics) for a target recovery of 10,000 nuclei.
The scATACseq libraries were generated using 10x Genomics Chromium Cell ATAC Reagent Kit (v1) according to the manufacturer’s protocol.
scATACseq

treatment_protocol: PD1 blockade was performed with 5 treatments of 200ug aPD-L1 antibody (10F.9G2, BioXCell) i.p. every 3 days starting 16 days after infection with LCMV Cl13. Analysis was performed one day after final treatment. For control treatments, phosphate-buffered saline (PBS) was administered i.p.

value: P14 CD8 T cells

tag: background strain

tag: treatment


title: Patient 7 non lesional psoriasis, rep 1 [P16357_1021]

description: 10x Genomics


title: Patient 5 lesional atopic dermatitis, rep 1 [P16357_1015]

description: 10x Genomics


title: Cl13D8

source_name: P14 CD8 T cells from LCMV Clone13 infection 8 days post infection

value: No


title: ArmD30

source_name: P14 CD8 T cells from LCMV Armstrong infection 30 days post infection


title: ArmD15

source_name: P14 CD8 T cells from LCMV Armstrong infection 15 days post infection


title: ArmD8

source_name: P14 CD8 T cells from LCMV Armstrong infection 8 days post infection


title: Arm_d60

data_processing:
scATAC-seq sample demultiplexing, barcode processing, alignment, filtering, unique molecular identifier (UMI) counting were performed using 10x Cell Ranger ARC pipeline (2.0.0) and mm10 genome.
Downstream scATAC-seq analysis was performed in R (version 4.0.2) using Seurat (version 4.0.4), Signac (1.3.0), and ArchR (1.0.1)
Union peak list was generated using peaks called in ArchR and using macs2 with sample bam files.
Assembly: GRCm38/mm10
Supplementary files format and content: Filtered gene-barcode matrices containing only cellular barcodes

source_name: P14 CD8 T cells from LCMV Armstrong infection 60 days post infection


title: Cl13D30

source_name: P14 CD8 T cells from LCMV Clone13 infection 30 days post infection


title: Arm_d200

source_name: P14 CD8 T cells from LCMV Armstrong infection 200 days post infection


title: PDX_HCI_006_ESR1_L536P_miRNA_expression

status: Public on Oct 05 2022

city: Salt Lake City

first: Jason

last: Gertz

state: UT

address: 1950 Cir of Hope Dr

institute: University of Utah

hyb_protocol: Hybridization of nCounter Reporter Probes was performed per manufacturer's instructions using the NanoString nCounter platform with the human v3 miRNA panel.

scan_protocol: Sample scans were performed on the nCounterTM Digital Analyzer (NanoString Technologies).

data_processing: NanoString miRNA expression assay results in the form of raw counts were analyzed using NanoString’s nSolver software. Raw counts were normalized using two approaches. The first approach used the geometric mean for each sample of the top 100 highest expressed miRNAs across all samples to determine a normalization factor for each sample. The second approach used the geometric mean for each sample of the five spike-in miRNAs profiled for the assay to generate the normalization factors. Normalized counts were then log-scaled and used to perform t test to identify significantly differentially expressed miRNAs.

label: NanoString

source_name: mammary gland ductal carcinoma

label_protocol: Samples were prepared per manufacturer's instructions using the NanoString human v3 miRNA panel (Catalog # CSO-MIR3-12).

growth_protocol: Cells were cultured in RPMI1640 Media (ThermoFisher Scientific) supplemented with 10% FBS (Sigma-Aldrich) and 1% penicillin-streptomycin (ThermoFisher Scientific).

extract_protocol: Cells were washed with phosphate-buffered saline (PBS; ThermoFisher Scientific) and treated with RLT plus (Qiagen) lysis buffer with 1% beta-mercaptoethanol (BME; Sigma-Aldrich) for cell lysis. Cells were passed through a 21-gauge needle (Sigma-Aldrich) to facilitate genomic DNA lysis. For PDX lines, tissue samples were used from seven PDX lines (WT: HCI-003 and HCI-011; L536P: HCI-005, HCI-006, HCI-007; Y537S: HCI-013, HCI-013EI). Approximately 20mg of tissue per sample was homogenized in RLT plus lysis buffer with 1% beta-mercaptoethanol using gentleMACS M-tubes and a gentleMACS Octo Dissociator (Miltenyi Biotec). RNA extraction for both T-47D and PDX samples was performed using a Quick-RNA RNA Miniprep kit (Zymo Research), and RNA was measured using a Qubit fluorometer.

treatment_protocol: ER mutant or WT T-47D cells were plated in 100mm dishes and grown in estrogen-deprived media for four days prior to treatment with DMSO (0.1%) for 8 hours. For ESR1 knock-down experiments, cells were treated with DMSO (0.1%) and transfected with siRNAs targeting ESR1 or a non-targeting siRNA control for 48 hours prior to RNA harvest. PDX samples were surgically removed from mice and flash frozen. PDX samples were not treated.

tag: model

value: HCI-006 PDX

tag: esr1 genotype


title: PDX_HCI_007_ESR1_L536P_miRNA_expression

value: HCI-007 PDX


title: PDX_HCI_005_ESR1_L536P_miRNA_expression

value: HCI-005 PDX


title: PDX_HCI_013_ESR1_Y537S_miRNA_expression

value: HCI-013 PDX


title: PDX_HCI_013EI_ESR1_Y537S_miRNA_expression

value: HCI-013EI PDX


title: PDX_HCI_011_rep2_ESR1_WT_miRNA_expression

value: HCI-011 PDX

value: WT


title: PDX_HCI_011_rep1_ESR1_WT_miRNA_expression


title: PDX_HCI_003_rep1_ESR1_WT_miRNA_expression

value: HCI-003 PDX


title: PDX_HCI_003_rep2_ESR1_WT_miRNA_expression


title: HeLa, Diagenode 20ug K27ac, 12.5M reads

status: Public on Sep 03 2022

city: Grand Rapids

first: Ariana

last: Kupai

address: 333 Bostwick Ave NE

institute: Van Andel Institute

description:
params4.in
input is inputDMSO from GSE207783

data_processing:
Final processed data obtained from sans spike-in quantitative ChIP-seq (siQ-ChIP) pipeline. Full codes at github via github.com/BradleyDickson/siQ-ChIP
IP and input data were aligned to genome, quantitative scale was computed from sample masses, fragment lengths, etc, and the siQ-scaled process track is alpha * IP/input
Peaks were called in each 20ug IP dataset and compared to the same genomic region in the 2.5ug IP for the same antibody. Processing code is in frechet.f90 on https://github.com/BradleyDickson/siQ-ChIP.
Assembly: hg38
Supplementary files format and content: bigwig files are quantitative siQ-ChIP tracks
Supplementary files format and content: peak files: The format of that data is as follows: peak_number chr start end frechet 1.0 response area_numerator area_denominator max_numerator max_denominator left right where 'left' and 'right' are the location of the maximum for that interval

source_name: cervical

growth_protocol: DMEM, 10% FBS, 1% penicillin-streptomycin, grown in 37°C with 5% CO2

extract_protocol:
Cells were fixed with 0.75% formaldehyde for 5 minutes. Cells were quenched with 750mM Tris for 5 minutes. After cell lysis, chromatin was fragmented using micrococcal nuclease (worthington).
Library preparation was done using KAPA HyperPrep Kit (Roche, KK8504) with 4 uL of Illumina adapters (IDT, UDI) and sequenced on Illumina NextSeq 500 with a Mid-Output (paired-end 75 bp reads) flow cell. Input libraries had between 83-102M reads, and each IP library had 27-46M reads that passed QC, with 88% of the bases having quality scores >= 30.

value: cervical

value: HeLa


title: Cl13D15

source_name: P14 CD8 T cells from LCMV Clone13 infection 15 days post infection


title: T47D_D538G_clone2_miRNA_expression

value: T-47D cell line


title: T47D_Y537S_clone1_miRNA_expression


title: Naïve - scA101

source_name: P14 naïve CD8 T cells


title: T47D_D538G_clone1_miRNA_expression


title: T47D_Y537S_clone2_miRNA_expression


title: T47D_WT_clone2_miRNA_expression


title: HeLa, Diagenode 2.5ug K27ac, 12.5M reads

description:
params3.in
input is inputDMSO from GSE207783


title: HeLa, CST 2.5ug H3K9me3

description:
9CSTloparams.in
input is inputDMSO from GSE207783

value: CST#13969S H3K9me3 lot:5


title: T47D_WT_clone1_miRNA_expression


title: HeLa, 10ug K9me2, rep 2


title: HeLa, 10ug K9me2, rep 1


title: HeLa, 2.5ug K9me2, rep 2


title: HeLa, Active Motif 20ug K27ac, 12.5M reads

description:
params6.in
input is inputDMSO from GSE207783


title: HeLa, input, rep 2


title: HeLa, input, rep 1


title: MO5_Met-aPD1_3

status: Public on Jul 01 2022

city: Suita

first: TAISAKU

last: OGAWA

state: Osaka

address: 6-2-3 Furuedai

institute: RIKEN

description: rep1-3: independent library preparations;

data_processing:
Basecalls performed by Illumina MiSeq
Sequenced reads were trimmed for adaptor sequence, and were sorted by indexes in Illumina MiSeq
Reads were aligned to the mm10 genome assebly using STAR
Mapped reads seperated by genes, and the reads mapped for each gene were clusterd by molecular barcode
The number of cluteres for each was counted, which provides the number of molecules detected for each gene
The copy number of RNA moelcules per cell for each gene is calculated based on spike-in RNA detection efficiency
Genome_build: mm10
Supplementary_files_format_and_content: The calculated copy number of RNA moelcules per cell for each gene is listed.

source_name: MO5 cell

extract_protocol:
RNAs from cell lysate obtained by heat elevation is directly used for the next RT reaction
Libraries were prepared by using molecular barcoding technique. First, for normalization, the same amount of spike-in RNAs, a known number of External RNA Controls Consortium (ERCC) RNA molecules, was added into every tube which contains 100 cells. Then, the cells were lysed and RNA molecules were fragmentated by temperature elevation. cDNAs were generated by reverse transcription using a reverse transcription primer, in which the first common primer sequence, molecular barcode, fourteen T bases, and one V (A or G or C) base are tandemly arranged from 5' end. In the reverse transcription, the second common primer sequence was also attached to 3' end of each generated cDNA molecule by template switching. Subsequently, cDNAs were amplified by PCR using two primers; one primer contains an illumina adapter sequence, sample index, and the first common primer sequence, and the other primer has another illumina adapter sequence, sample index, and the second common primer sequence.

treatment_protocol: CD45+CD8+ cells and CD45- tumor cells were isolated from B16 melanoma using FACS cell sorter.

tag: genotype/variation

value: Cbf{beta}2 knockout

value: MO5 cell

tag: markers


title: MO5_Met-aPD1_2


title: MO5_aPD1_2


title: MO5_aPD1_1


title: MO5_Met-aPD1_1


title: MO5_Met_2

value: wild type


title: MO5_none_2


title: HeLa, Active Motif 2.5ug K27ac, 12.5M reads

description:
params5.in
input is inputDMSO from GSE207783


title: MO5_none_3


title: MO5_none_1


title: MO5_Met_1


title: MO5_aPD1_3


title: HeLa, 2.5ug K9me2, rep 1


title: CD8TIC_Met-aPD1_1

source_name: CD8 Tumor infiltrating cell

value: CD8 Tumor infiltrating cell


title: CD8TIC_aPD1_1


title: CD8TIC_Met_2


title: CD8TIC_Met-aPD1_3


title: CD8TIC_Met_3


title: CD8TIC_Met_1


title: CD8TIC_none_2


title: CD8TIC_aPD1_3


title: CD8TIC_aPD1_2


title: MO5_Met_3


title: 1231A3, NCC expansion PN2, rep1

status: Public on Jun 19 2022

city: Fujisawa

first: Daisuke

last: Kamiya

state: Kanagawa

address: 26-1 Muraoka-Higashi 2-chome

department: CiRA

institute: Kyoto University

data_processing:
Base calling was performed using standard parameters in build-in software provided with the Ion Torrent S5 XL sequencer (Torrent suite v5.16.1).
Sequenced reads were mapped to hg19_AmpliSeq_Transcriptome_ERCC_v1 using standard parameters and build-in software provided with the Ion Torrent S5 XL sequencer (Torrent suite v5.16.1).
RPM were calculated using plug-in software provided with the Ion Torrent S5 XL sequencer (ampliSeqRNA v5.6.0.3)
Assembly: hg19
Supplementary files format and content: Tab-delimited text file include gene symbol and gene expression in RPM for each sample

growth_protocol: 1231A3 human iPSC were cultured on an iMatrix-511 (Nippi, Tokyo, Japan)-coated cell culture plate or dish in StemFit AK03N (Ajinomoto, Tokyo, Japan). The medium was changed every 2 days

extract_protocol:
Total RNA was purified using an RNeasy Mini Kit (Qiagen, Valencia, CA, USA)
10 ng of total RNA were reverse transcribed  to obtain single-stranded cDNA using the SuperScript VILO cDNA Synthesis Kit (Thermo Fisher Scientific). cDNA libraries were  synthesised using the Ion AmpliSeq Transcriptome Human Gene Expression Core Panel (Thermo Fisher Scientific) and the Ion Ampliseq Library Kit Plus (Thermo Fisher Scientific), according to the manufacturer’s protocol. Barcode-labeled cDNA libraries were analyzed using the Ion S5 XL System (Thermo Fisher Scientific) and the Ion 540 Chip Kit (Thermo Fisher Scientific).

treatment_protocol: For NCC induction, iPS cells were cultured in StemFit Basic03 (equivalent to AK03N without bFGF, Ajinomoto, Tokyo, Japan) with 10 μM SB431542 (FUJIFILM Wako) and 1 μM CHIR99021 (Axon Medchem, Reston, VA, USA) for 10 days.CD271high sorted NCCs were seeded onto fibronectin-coated plates in Basic03 supplemented with 10 μM SB431542, 20 ng/mL EGF (FUJIFILM Wako), and FGF2 (FUJIFILM Wako). MSCs were induced from NCCs by cultured in PRIME-XV MSC Expansion XSFM medium (IrvineScientific).

value: NCCs from iPSCs

tag: genotype

value: NCC expansion

tag: time


title: 1231A3 iPSC, rep2

value: iPSCs derived from PBMC

value: iPSC maintenance

value: maintenance


title: 1231A3 iPSC, rep1


title: 1231A3 iPSC, rep3


title: 1231A3, NCC expansion PN2, rep2


title: B6WT+Mgl2DTR_eGFP_Thymic_cells - HTO

status: Public on May 09 2022

city: Minneapolis

first: Kristin

last: Hogquist

state: MN

address: 2101 6th St SE

institute: University of Minnesota

data_processing:
10X Genomics Cell Ranger (scRNAseq) software was used to generate fastq files.
Each gel bead in emulsion (GEM) was identified and all gene specific reads were mapped via cellranger count (ver. 6.0.0) using the mouse reference genome (mm10) or antibody specific tags (HTOs).
Gene level quantification was completed using cellranger count software (ver. 6.0.0) using a custom GTF (derived from GENCODE, version M23, Ensembl 98). HTO counts were generated by cellranger count using the specific antibody-oligo tags sequences used in this experiment. B6WT and Mgl2DTR (strain)  were classified using HTO counts respectivley via Seurat's HTOdemux function.
Raw counts data was imported into R (ver 4.1.1) for downstream processing, including normalization, principal components analysis, differential gene expression testing, single cell clustering, etc. primarily using the Seurat R package (ver. 4.0.4. Although ADT data were generated they were not used in this analysis.
Seurat (v 4.0.4 ) was used to process and cluster cells, and to create UMAP images and gene expression feature plots
Assembly: mm10 (ver. 3.0.0 provided by 10X Genomics)
Supplementary files format and content: text files that contain raw GEX and HTO counts for every tagged GEM

source_name: murine thymocytes of 7-week-old wildtype (WT) and Mgl2DTR_eGFP treated with diptheria toxin for 48 hours

extract_protocol:
An RNeasy micro kit (Qiagen) was used to isolate RNA obtained from each sample per the manufacturer’s instructions.
For 10X Genomics scRNA-seq, we generated libraries that measure: (1) mRNA transcript expression (RNA) and (2) tissue-specific hashtag oligos (HTO). Reverse transcription PCR and library preparation were carried out under the Chromium Single Cell 3’ v3 protocol (10X Genomics) per manufacturer’s recommendations and sequenced in a NovaSeq.

treatment_protocol: Mgl2DTR_eGFP mice were treated with 500ng of diptheria toxin. Thymus cells were obtained from organs digested with collagenase D (1 mg/ml). CD90.2-expressing thymocytes were depleted with magnetic-activated sorting. Enriched cells were stained with anti-CD11c and anti-CD11b antibodies, and live cells were FACS-sorted with a FACSAria (BD Biosciences).


title: B6WT+Mgl2DTR_eGFP_Thymic_cells - GEX

value: murine thymocytes of 7-week-old wildtype (WT) and Mgl2DTR_eGFP treated with diptheria toxin for 48 hours


title: Lv2, scRNAseq

status: Public on Apr 07 2022

city: Fribourg

first: Jules

last: Duruz

address: Chemin du musée 10

country: Switzerland

institute: University of Fribourg

description:
10x Genomics
Lv_barcodes.tsv.gz (barcode-1 represents Sample Lv1, barcode-2 represents Sample Lv2)
Lv_features.tsv.gz
Lv_matrix.mtx.gz

data_processing:
10x Genomics Cell Ranger version 3.0.2 (Zheng et al., 2017) was used to map the single cell RNAseq reads from 10x Genomics Chromium to the above assembled coding genome and count the number of reads overlapping with each gene to produce a count matrix for each sample. Afterwards, the R package scater v. 1.14.0 (McCarthy et al., 2017) was used to automatically filter low-quality cells (cells with low library size, low number of expressed genes and high proportion of mitochondrial reads).
Assembly: Lv_hg_transcripts.fasta
Supplementary files format and content: Lv_barcodes.tsv.gz, Lv_features.tsv.gz, Lv_matrix.mtx.gz (aggregated datasets).

organism: Loligo vulgaris

source_name: Whole heads

extract_protocol:
Single-cell RNAseq libraries were prepared using a Chromium Single Cell 3’ Library & Gel Bead Kit v2 or v3 (10X Genomics), according to the manufacturer’s protocol (User Guide). Two chips were loaded with the accurate volumes calculated based on the “Cell Suspension Volume Calculator Table”. The initial single-cell suspension being estimated at >600 cells/μl, we targeted to recover a maximum 10000 cells. Once GEMs were obtained, reverse transcription and cDNA amplification steps were performed.
scRNA-seq

value: All cells

value: Whole heads

tag: developmental stage

value: pre-hatchlings

tag: age

value: stage 28


title: Lv1, scRNAseq


title: CD8TIC_none_1


title: DL2: Dexamethason and lithium treated sample replicate 2

status: Public on Jan 01 2022

city: Prague

first: Martin

last: Bezdicka

state: Czech Republic

address: V Uvalu 84

department: Department of Pediatrics

country: Czech Republic

institute: Second Faculty of Medicine and Motol University Hospital

data_processing:
Illumina BaseSpace Sequence Hub was used for basecalling.
The transcriptome profiles were analyzed using the BRB-SeqTools framework, reads were aligned using Subread, duplicated reads eliminated by Picard, and genes counted by HTSeq-count.
The gene counts tables and the sample metadata were assembled into a DESeq2 DESeqDataSet object.
Global differences in gene expression between the four conditions were assessed on ordination plots, and particular differentially expressed genes sought for using a negative binomial regression model in DESeq2.
A minimum of 2-fold increase or decrease in expression, and a corrected P value of <0.1 was needed for a gene to be considered. The DESeq test P values were corrected for multiple testing using Benjamini-Hochberg method.
Genome_build: Alignment was initially to the transcriptome (hg19, GRCh37.72), and then to the genome (hg19, GRCh37).
Supplementary_files_format_and_content: Matrix table and separate sample files with raw gene counts for every gene and every sample.

source_name: Primary cultured fetal rat metatarsal bones

growth_protocol: The standard cultivation medium consisted of DMEM/F12 (1:1) with 15 mM Hepes and L-Glutamin + Gentamycine 40 µM + Bovine serum albumin 0.2% + β-Glycerolphosphate 1 mM + Ascorbic acid 285 µM.

extract_protocol:
Metatarsal bones were crushed by using the Minilys MK 28-R homogenization tubes with beads. The RNA was harvested using Trizol reagent and the PureLink RNA Mini Kit.
The 300 ng of total RNA was subjected to library synthesis using the SureSelect strand-specific mRNA library preparation kit (protocol version E0, Agilent Technologies).

treatment_protocol: Approximately 20 (+-2) metatarsals were incubated at 37°C in 5% CO2 for 72 hours in 8 ml of standard cultivation medium + 8 µL of 99% ethanol/ + 8 µL of 1 mM Dexamethasone/ + 8 µL of 99% ethanol and 8 µL of 10 M LiCl/ + 8 µL of 1 mM Dexamethasone and 8 µL of 10 M LiCl for the control/dexamethasone/lithium chloride/ dexamethasone + lithium chloride treatment groups, respectively. There were three biological replicates performed for all four treatment groups.

value: Sprague Dawley

value: Time-mated rat E19

value: standard cultivation medium + dexamethason and lithium


title: DL3: Dexamethason and lithium treated sample replicate 3

value: Primary cultured fetal rat metatarsal bones


title: CD8TIC_none_3


title: DL1: Dexamethason and lithium treated sample replicate 1


title: Li3: Lithium treated sample replicate 3

value: standard cultivation medium + lithium


title: Acinetobacter baumannii ATCC 17978 (WT)

status: Public on Jan 31 2022

city: Seongbuk-gu

first: Mingyeong

last: Kang

state: Seoul

address: Anam-ro 145

country: South Korea

institute: Korea university

data_processing:
RNA sequencing was performed on an Illumina Novaseq 6000 platform.
The reference genome sequence was retrieved from the NCBI database.
Quality-filtered reads were aligned to the reference genome sequence using Bowtie2.
The relative abundance of transcripts is presented as FPKM (Fragments Per Kilobase of transcript per Million).

organism: Acinetobacter baumannii ATCC 17978

source_name: Bacteria cells

growth_protocol: Exponential phase on LB (Luria-Berani) media

extract_protocol:
Using an RNeasy Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions
Illumina sequencing library was constructed by the TruSeq Stranded Total RNA with Ribo-Zero Plus (Illumina, San Diego, CA, USA) following the manufacturer's protocol

value: Mid-exponential phase (OD=0.5)


title: Li2: Lithium treated sample replicate 2


title: Dex2: Dexamethason treated sample replicate 2

value: standard cultivation medium + dexamethason


title: Dex1: Dexamethason treated sample replicate 1


title: Li1: Lithium treated sample replicate 1


title: C2: Untreated control replicate 2

value: standard cultivation medium


title: Dex3: Dexamethason treated sample replicate 3


title: C3: Untreated control replicate 3


title: 116.STAT5_cKO_4T1.CM

status: Public on Nov 17 2021

first: Kaylee

last: Schwertfeger

address: 420 Delaware St. SE MMC 609

department: Lab Medicine and Pathology

data_processing:
CHRUP pipeline (https://dl.acm.org/doi/10.1145/3332186.3333156) used to process raw sequence data
trimmomatic version 0.33 was used to clean reads for adapter contamination and low-quality sequence
HISAT2 version 2.1.0 was used to align samples
featureCounts v1.6.2 was used to count mapped reads to genes
Genome_build: Mus musculus GRCm38.99
Supplementary_files_format_and_content: count table with raw count data for all genes and samples

source_name: Bone marrow

extract_protocol:
Total RNA was collected using TriPure reagent (Roche) from STAT5fl/fl and STAT5cKO BMDMs treated with DMEM, 20 ng/mL rmGM-CSF (Fisher Scientific), or 4T1 tumor cell conditioned media
Stranded RNAseq libraries were prepared using the TruSeq RNA library kit

value: BALB/c

value: macrophages

value: 6 wk


title: 116.STAT5_cKO_DMEM


title: 116.STAT5_cKO_GM.CSF


title: EpCAM_Prox_1

status: Public on Dec 11 2021

city: Copenhagen N

first: Marianne

middle: Terndrup

last: Pedersen

address: Ole Maaloes Vej 5

department: University of Copenhagen

country: Denmark

institute: BRIC

data_processing:
Mapping and quantification of fastq files were done via the Rsubread package.
Reads were mapped to the mm10/GRch38 genome assembly using the align-function with default settings, except nBestLocations=16 and nTrim5=6.
Mapped reads were quantified across RefSeq gene models using the featureCounts-function with default settings, except strandSpecific=2. This approach retains multi-mapping reads for gene-level quantification, as long as only one of the multiple locations maps to an annotated gene.
DE analysis was carried out using edgeR and limma with default settings unless otherwise noted.
RPKM-values were extracted using the rpkm and rpkmByGroup functions from the edgeR package.
Genome_build: mm10
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample.

source_name: EpCAM+/DAPI-/CD31-/CD45- cells isolated from the proximal region of E16.5 intestine

growth_protocol: Intestines were isolated from E16.5 or E13.5 mouse fetuses. Where indicated, the pregnant dams had been treated twice daily from E12.5 to E16.5 with vehicle or LGK-974, a PORCN inhibitor preventing secretion of Wnt ligands.

extract_protocol:
Cells were sorted into RLT buffer and RNA extracted using the RNeasy micro kit (Qiagen).
RNA-seq libraries were prepared using the SMARTer Stranded Total RNA-Seq v2 Pico Input Mammalian kit (Takara).

treatment_protocol: Intestines dissected from E16.5 mouse fetuses were divided into five sections of equal size of which the most proximal, the mid and the most distal segments were processed. Intestines isolated from E13.5 mouse fetuses were divided into three regions and the proximal and distal regions selected. Intestinal regions were incubated with collagenase to obtain single cells. After antibody staining, the indicated cell populations were isolated using a FACSAria III.

tag: intestinal region

value: Proximal E16.5 intestine


title: 115.STAT5_cKO_GM.CSF


title: 115.STAT5_cKO_DMEM


title: EpCAM_Prox_3


title: dam KO strain in Acinetobacter baumannii ATCC 17978 (dam KO)


title: 115.STAT5_cKO_4T1.CM


title: 114.STAT5_cKO_GM.CSF


title: 114.STAT5_cKO_4T1.CM


title: 114.STAT5_cKO_DMEM


title: 113.STAT5_flfl_4T1.CM


title: 113.STAT5_flfl_DMEM


title: 113.STAT5_flfl_GM.CSF


title: 112.STAT5_flfl_GM.CSF


title: 112.STAT5_flfl_DMEM


title: EpCAM_Prox_2


title: 965-E3_KO_FD1

status: Public on Oct 25 2021

city: Boston

first: Susan

last: Slaugenhaupt

state: MA

address: 185 Cambridge St

department: Center for Genomic Medicine

institute: Massachusetts General Hospital

data_processing:
RNASeq reads were mapped by STAR (version 2.4.2a) allowing 5% mismatch
Raw gene counts were generated by HTSeq-count (version 0.13.5)
Size factors for normalization were estimated by R package DESeq2 (version 1.28.1)
Surrogated variables (SVs) were estimated by R package sva (version 3.36.0), with full model as "~Genotype" and null model as "~1"
The uploaded processed file contains normalized counts with effects from SVs removed, meaning it only counts for expression from "pure" genotype effects
Genome_build: Ensembl GRCm38 version 83, with human ELP1 gene sequence inserted as an independent chromosome
Supplementary_files_format_and_content: Fastq format for RNASeq reads; Tab-delimited txt format for normalized expression counts

source_name: mouse embryo

extract_protocol:
RNA was extracted from 8 Elp1-/-, 7 TgFD1; Elp1-/-, 6 TgFD9; Elp1-/- and 8 Elp1+/+ individual embryos at the embryonic stage of E8.5 using the QIAzol Reagent and following the manufacturer’s instructions.
RNAseq Libraries were prepared using TruSeq® Stranded mRNA Library Prep kit (Illumina 20020594), using 100 ng total RNA as input. Libraries were evaluated for final concentration and size distribution by Agilent 2200 TapeStation and/or qPCR, using Library Quantification Kit (KK4854, Kapa Biosystems), and multiplexed by pooling equimolar amounts of each library prior to sequencing.

value: embryo


title: 965-E5_KO_FD1


title: 965-E6_WT

value: WT


title: 775-E2_KO_FD9


title: 965-E1_WT


title: C1: Untreated control replicate 1


title: 112.STAT5_flfl_4T1.CM


title: 751-E10_KO_FD1


title: 683-E7_KO

value: KO


title: 775-E1_KO_FD9


title: CD8TIC_Met-aPD1_2

value: Cbf{beta}2 knockout

value: CD8 Tumor infiltrating cell


title: 683-E1_WT


title: 682-E9_WT


title: 682-E5_KO


title: 682-E6_WT


title: 682-E7_KO


title: 682-E4_KO


title: 682-E2_WT


title: 682-E3_WT


title: 682-E1_KO


title: 452-E8_KO_FD9


title: 604-E4_KO_FD9


title: 365-E9_KO_FD9


title: 351-E5_KO


title: 351-E3_KO


title: 351-E4_KO_FD1


title: 282-E6_KO_FD1


title: 244-E3_KO_FD9


title: 245-E9_KO_FD1


title: 351-E1_KO


title: K27ac_PDX_93

status: Public on Aug 12 2021

first: Yingtian

last: Xie

address: 450 BROOKLINE AVE

institute: DANA FARBER CANCER INSTITUTE INC

data_processing:
Illumina Casava1.7 software used for basecalling.
Reads were aligned to mm9 using BWA
For peak calling MACS2 was used.
Genome_build: hg19
Supplementary_files_format_and_content: bw files were generated using MACS2 using the chilin pipeline

source_name: PDX

extract_protocol:
Cells was fixed using 1% formaldehyde and chromatin sheared to 300-500 bp in size using the Covaris E220 ultrasonicator. Resulting chromatin was incubated overnight with indicated antibodies. Purified immunoprecipitates were isolated and quantified by Qubit fluorometer.
DNA sequencing libraries were prepared using the ThruPLEX-FD Prep kit (Rubicon Genomics). Libraries were sequenced using 75-bp reads on the Illumina platform at the Dana-Farber Cancer Institute.

tag: chip antibody

value: lucap


title: K27ac_PDX_145_1


title: Bil.no2 [19H5_2D_Bil]

status: Public on Oct 20 2021

first: Takeshi

last: Katsuda

address: 421 Curie Blvd, BRB II/III Rm531

department: Perelman School of Medicine

data_processing:
Alignment independent transcript quantification using Salmon
Differential gene expression analysis using DESeq2
Genome_build: mm10
Supplementary_files_format_and_content: txt file containing Transcript per million

source_name: in vitro cultured hepatocyte-derived biliary cells

growth_protocol: Isolated mouse hepatocytes and hepatocyte-derived biliary cells were cultured in vitro

extract_protocol:
RNA extraction using Qiagen RNeasy Kit
RNA-seq library was constructed by Novogene company following standard protocol

treatment_protocol: No treatment was applied

tag: strain background

tag: cell type source

value: hepatocytes

tag: cell subtype

value: hepatocyte-derived biliary cells

tag: clone number

tag: culture

value: 2D culture


title: Bil.no1 [15c7_2D_Bil]


title: Hep.no2 [19H5_3D_Hep]

source_name: in vitro cultured hepatocytes

value: 3D culture


title: K27ac_PDX_49

tag: cell line


title: K27ac_PDX_145_2


title: Hep.no1 [15c7_3D_Hep]


title: T17-11572_NEC

data_processing:
Illumina Casava1.7 software used for basecalling.
Reads were aligned to hg19 using BWA.
For peak calling MACS2 was used.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: bigWig files were generated using MACS2 using the chilin pipeline.

source_name: Primary tissue

extract_protocol:
Fragment of frozen tissues (PDX models) or 50 um sections (liver metastases) were cut and resuspended in 300 ul of cold 3-detergent-ATAC-Resuspension Buffer (RSB) containing 0.1% NP40, 0.1% Tween-20, and 0.01% Digitonin. Tissues were dounced 10 times with a loose and a tight pestle each until complete homogenization. The homogenate was then transferred to a 1.5 ml pre-chilled microfuge tube and incubated on ice for 10 minutes to lyse. After lysis, 300 ul of ATAC-RSB containing 0.1% Tween-20 was added and the tubes were inverted to mix. Lysates were filtered through a 40 um cell strainer and nuclei were centrifuged for 10 minutes at 1500 RCF in a pre-chilled (4 ºC) fixed-angle centrifuge. Nuclei were resuspended with 300 ul of ATAC-RSB containing 0.1% Tween-20 and counted with a hemocytometer using Trypan blue stain.
ATAC-seq libraries were amplified as described previously (Buenrostro et al.).

tag: tissue

value: NEGI


title: T18-05426_I_NEC


title: T18-01479_NEC


title: LuCaP_93_1X

source_name: Patient derived xenograft, prostate cancer

value: NEPC


title: LuCaP_96_1

value: CRPC


title: LuCaP_49_2X


title: LuCaP_49_1X


title: LuCaP_77


title: LuCaP_23_1


title: LuCaP_173_1_S6


title: LuCaP_208_2


title: LuCaP_145_2


title: LuCaP_173_1_S17


title: LuCaP_93_2X


title: LuCaP_145_1_1X


title: EF1_cfce

source_name: Cell line prostate cancer


title: DFMC_33043-R1_2D10

value: MCC


title: DFMC_96712-R1_2C3


title: DFMC_48396-R0_2I30


title: DFMC_87346-Q1_1A3


title: 351-E7_KO_FD1


title: LuCaP_77CR


title: LuCaP_78


title: lucap173_1_ND1

extract_protocol:
Nuclei isolated as previously described were crosslinked with 1% Formaldehyde for 10 minutes for H3K27Ac ChIP-seq and crosslinked in 2 steps with 2 mM of DSG (Pierce) for 45 minutes at room temperature followed by 1 ml of 1% Formaldehyde for 10 minutes whereas for ASCL1 and NEUROD1. Crosslinked nuclei were then quenched with 0.125 M glycine for 5 minutes at room temperature and washed with PBS. After fixation, pellets were resuspended in 500 ul of 1% SDS (50 mM Tris-HCl pH 8, 10 mM EDTA) and sonicated in 1 ml AFA fiber millitubes for 5 minutes (H3K27ac) or 10 (ASCL1 and NEUROD1) minutes using a Covaris E220 instrument (setting: 140 peak incident power, 5% duty factor and 200 cycles per burst). Chromatin was immunoprecipitated with 10 ug of H3K27Ac antibody (Diagenode cat# C15410196), 10 ug of ASCL1 antibody (abcam ab74065) or 10 ug of NEUROD1 antibody (Cell Signalling mAb #4373). 5 ug of chromatin was used for H3K27Ac ChIPs, and 40 ug of chromatin was used for ASCL1 or NEUROD1 ChIPs.
ChIP-seq libraries were made using Rubicon kit (reference) and purified. 75-bp single-end reads were sequenced on a Nextseq instrument (Illumina).


title: K27ac_PDX_173_1


title: K27ac_ef1

source_name: Cell line


title: NCI-H660


title: DFMC_11112-R1_2F0


title: 1231A3, NCC expansion PN2, rep3

tag: cell type

value: NCCs from iPSCs

tag: treatment

value: NCC expansion


title: MSX2_hTSC_ChIPseq_r1x

status: Public on Aug 28 2021

city: Vienna

first: Andreas

last: Lackner

address: Schwarzspanierstrasse 17

department: Center for Anatomy and Cell Biology

country: Austria

institute: Medical University of Vienna

description:
processed data file:
M2_128246_129321_130126.bw
MSX2_WT_SR_all_IDR_CPA.bed
MSX2_hTSC_128246x2_ACAGTG

data_processing:
RNA-seq
read trimming: TrimGalore! (0.6.5)
alignment: STAR (2.7.3a)
alignment processing: samtools (1.10)
read counting: Rsubread (2.0.0)
ChIP-seq:
read trimming: CUTADAPT (2.8)
alignment: bowtie 2 (2.3.5.1)
alignment processing: samtools (1.10)
peak calling: MACS2 (2.2.7.1)
idr analysis: idr (1.2)
bigwig files: bamCoverage (3.5.0)
Genome_build: hg38
Supplementary_files_format_and_content: read counts, bigwig

growth_protocol: hTSC line CT30 were cultured in the basal media (DMEM/F12 (11320074, Thermo Fisher), 1x ITS-X (51500056, Thermo Fisher), 0.2% FBS (PAA), 1.5 µg/mL L-ascorbic acid (A4403, Sigma) supplemented with 2 µM CHIR99021, 50ng/mL EGF, 0.5 µM A83-01 and 5µM Y27632. Cells were passaged every 3-4 days using TrypLE (12605036, Thermo Scientific).

extract_protocol:
RNAseq. RNA was extracted using RNeasy mini kit (Qiagen) and treated with DNaseI (Qiagen). Indexed libraries were prepared with 500 ng RNA using QuantSEQ 3’ mRNA-Seq Library Prep Kit FWD for Illumina (015.96, Lexogen) according to manufacturer’s recommendations. The libraries were pooled and sequenced with a 100-base-pair single-end (MSX2_iOX) and 50-base-pair single-end protocol on Illumina HiSeq 2500 sequencer. ChIPseq. Cells (1-2x108) were fixed in 2mM Di(N-succinimidyl) glutarate (DSG) (80424, Sigma) in PBS at room temperature (RT) for 45 min. After washing in PBS, cells were fixed again in 1% formaldehyde (28908, Sigma) in TS base media at RT for 12 min. Fixation was stopped by adding glycine to a final concentration of 0.125 M. Cells were washed twice in PBS and resuspended in wash buffer 1 (10mM Hepes pH7.5, 10mM EDTA, 0.5mM EGTA and 0.75% Triton X-100) and incubated at 4C for 10min. After pelleting, cells were resuspended in wash buffer 2 (10mM Hepes pH7.5, 200mM NaCl, 1mM EDTA and 0.5mM EGTA) and incubated at 4C for 10min. After pelleting, cells were lysed in the lysis/sonication buffer (150mM NaCl, 25mM Tris pH 7.5, 5mM EDTA, 0.1%Triton, 1% SDS and 0.5% sodium deoxycholate) with complete protease inhibitors (11873580001, Sigma) on ice for 30min. Chromatin was sonicated 15s on /30s off for 23-25 cycles using the UW2070 sonicator (Bandelin) to the average 300-bp fragments. Chromatin was diluted 1/10 with the dilution buffer (150mM NaCl, 25mM Tris pH 7.5, 5mM EDTA, 1% Triton X-100, 0.1% SDS and 0.5 % sodium deoxycholate) containing complete protease inhibitors. Protein G magnetic Dynabeads (10004D, Invitrogen) were blocked with 1mg/ml BSA and tRNA at 4C for 1h and washed with buffer A (150mM NaCl, 25mM Tris pH 8.0, 1mM EDTA, 1% NP-40, 0.1 % SDS and 0.5 % sodium deoxycholate). Chromatin was pre-cleared with pre-blocked beads at 4C for 1h. Four hundred micrograms of chromatin and seven micrograms of antibody (rabbit anti-MSX2 (HPA005652, Sigma) and rabbit normal IgG (NI01, Sigma) were used per each IP. IP was performed overnight at 4C with rotation. Pre-blocked magnetic beads were added next morning for 7-8 h. Beads were washed at 4C with buffer A three times, buffer B (50mM Tris pH 8.0, 500mM NaCl, 0.1% SDS, 0.5 % sodium deoxycholate, 1% NP-40 and 1mM EDTA), buffer C (50mM Tris pH 8.0, 250mM LiCl, 0.5% sodium deoxycholate, 1% NP-40 and 1mM EDTA) and rinsed with TE buffer. DNA was eluted from beads in the elution buffer (1% SDS, 0.1M NaHCO3). Samples were treated with RNAse A and Proteinase K and reverse-crosslinked overnight by incubation for 2h at 37C, followed by 12h at 65C. DNA was purified on the PCR purification columns (Qiagen).
Quantseq libraries were generated using the Quantseq 3' FWD kit according to the maunfacturers recommendations. ChIPseq libraries were generated using the NEBnext ultra II DNA library prep kit according to the standard protocol.

treatment_protocol: To deplete MSX2, cells were transduced with pLKO.1-based lentiviral particles containing either shRNA against MSX2 (TRCN0000234848 and TRCN0000234849) or a shRNA against GFP as a control for 16h and selected for integrations by treatment with 300µg/ml G418

value: human trophoblast stem cells

tag: differentiation state

value: self-renewal (SR)


title: lucap145_1_AS1


title: h660_AS1


title: lucap49_AS1


title: lucap93_AS1


title: 683-E4_WT

value: embryo

value: WT


title: T18-05426

data_processing:
Illumina Casava1.7 software used for basecalling.
Reads were aligned to hg19 using STAR.
Differential analysis was performed by DESeq2.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: bigWig files include RPKM values for each Sample.

extract_protocol:
A fragment of frozen tissues (PDX models) or 50 um sections (liver metastases) were cut and homogenized in 1 ml of AllPrep DNA/RNA Mini Kit (Qiagen) using a plastic pestle (Cole-Palmer #44468-23). DNA and RNA were simultaneously isolated.
500 ng RNA was used to prepare libraries using the NEBNext Ultra™ RNA Library Prep Kit for Illumina. RNA quantity and quality were assessed on an Agilent 2100 Bioanalyzer. For all RNA-seq, reads were sequenced on a NextSeq 500 instrument (Illumina).


title: T18-01479


title: LuCaP_81


title: T17-11572

source_name: primary tissue


title: prostate_NCI-H660_RNA-se


title: DFMC_63632-R1_2F10


title: LuCaP_145_1_2X


title: prostate_MSKCC-EF1_RNA-se


title: MET_H9


title: MET_H7


title: MET_H10


title: MET_H1


title: MET_H2


title: LuCaP_96_p30s25n5

source_name: patient derived xenograft, prostate cancer


title: LuCaP_81_s23n4


title: LuCaP_78_p12s23n6


title: LuCaP_77_p34s27n7


title: LuCaP_23_1_p58s29n4


title: LUCAP_173_1_S9


title: LUCAP_173_1_S5


title: LUCAP_173_1_S3


title: LuCaP_49_p70s16n16


title: LUCAP_173_1_S2


title: LUCAP_173_1_s1_3


title: LuCAP_173_1_p17


title: LUCAP_173_1_S11


title: LuCaP_145_1_p2s18n5


title: LuCaP_145_2_p9s26n3


title: DFMC-87346


title: DMFC-63632


title: DFMC-48396


title: LuCaP_93_p39s22n6


title: DFMC-11112


title: SB+ANI treated sample rep 4

status: Public on Aug 06 2021

city: Göttingen

first: Hsu-Min

last: Sung

state: Niedersachsen

address: Humboldtallee 23

institute: Heart Centre Göttingen

data_processing:
Reads were aligned using STAR v.2.4.2a to the Human RefSeq genome GRCh38.p10 and the ERCC spike-in sequence.
Reads were counted using the DEXSeq read counting script.
Splicing analysis was performed with DEXSeq using the GENCODE 27 gene annotation.
Differential gene expression analysis was performed using DESeq2.
Genome_build: GRCh38.p10
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each Sample

source_name: HeLa TREx_YFP-TIARr

growth_protocol: HeLa TREx cells were maintained in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% tetracyclin-free FBS, 2 mM L-glutamine, 100 U/ml penicillin and 0.1 mg/ml streptomycin , 2 mM L-glutamine, 100 U/ml penicillin and 0.1 mg/ml streptomycin, 5 µg/ml blasticidin and 100 µg/ml zeocin. Cells were cultured at 37°C in 5% CO2.

extract_protocol:
Total RNA was purified with the GeneMATRIX universal RNA purification kit (EURx), including a DNase (Promega) on-column digestion step. 350 ng of total RNA as quantified by Qubit (Invitrogen) was used for subsequent RNA-Seq. Ribosomal RNA was depleted with the Ribo Zero Human Gold Kit (Illumina)
Libraries were prepared using the TruSeq Directional RNA LT Preparation Kit (Illumina) using standard Illumina protocols.  Before library preparation RNA was prepared without poly(A)-mRNA enrichment step for acquiring pre-mRNA

treatment_protocol: For pretreatment with the p38 inhibitor (SB202190), final concentration of 10 µM SB202190 was added to cell culture medium 1 hour prior to DMSO or Anisomycin treatment. For Anisomycin treatment, final concentration of 0.1 µg/ml anisomycin was added to cell culture medium and incubated for 90 minutes prior to RNA extraction,

value: epithelial cell

value: Cervical cancer

value: HeLa TREx_YFP-TIARr

tag: stable expression

value: Tet repressor

tag: inducible expression

value: YFP-fused TIARb resistant to siRNA

value: SB202190 10 uM, 60 minutes, Anisomycin 0.1 ug/ml, 90 minutes


title: LUCAP_173_1_s6


title: LUCAP_173_1_S4


title: DMSO treated control rep 2

value: DMSO 90 minutes


title: ANI treated sample rep 1

value: Anisomycin 0.1 ug/ml, 90 minutes


title: SB+ANI treated sample rep 3


title: ANI treated sample rep 3


title: ANI treated sample rep 4


title: SB+ANI treated sample rep 2


title: Mouse whole brain

status: Public on May 28 2021

city: Portland

first: Ryan

middle: M

last: Mulqueen

state: OR

address: 3181 SW Sam Jackson Park Rd

department: Molecular and Medical Genetics

institute: Oregon Health and Science

description: single-cell ATAC seq data

data_processing:
FastQ files were generated from BCL files using bcl2fastq (v 2.19.0, Illumina Inc.) with the following options with-failed-reads, no-lane-splitting, fastq-compression-level=9, create-fastq-for-index-reads
FastQ reads were assigned to their expected primer index sequence allowing for sequencing error (Hamming distance ≤2) and indexes were concatenated to form a “cellID”. cellIDs are stored in the FastQ header
Reads that could be assigned unambiguously to a cellID were then aligned to reference genomes. For GM12878, paired reads were aligned with bwa mem (v0.7.15-r1140) to hg38. For s3-ATAC libraries, reads were first aligned to a concatenated hybrid genome of hg38 and GRCm38 (“mm10”, Genome Reference Consortium Mouse Build 38 (GCA_000001635.2)).
Reads were then de-duplicated to remove PCR and optical duplicates by a perl (v5.16.3) script aware of cellID, chromosome and read start, read end and strand. From there putative single-cells were distinguished from debris and error-generated cellIDs by both unique reads and percentage of unique reads.
For s3-ATAC libraries, cellIDs with ≥90% of reads aligning to a single reference genome were considered bona fide single-cells and fastQ files were filtered prior to realigning to the correct genome.
Pseudo-bulked data (agnostic of cellID) was then used to call read pile-ups or “peaks” via macs2 (v.2.2.7.1) with option –keep-dup all
A scitools perl script was then used to generate a sparse matrix of peaks × cellID to count occurrence of reads within peak regions per cell.
The peaks x cellID counts matrix was then used to generate Seurat Objects for Signac processing (see Online Methods for more downstream analyses).
Genome_build: GRCm38 (“mm10”, Genome Reference Consortium Mouse Build 38 (GCA_000001635.2) and GRCh38 (“hg38”,Genome Reference Consortium Human Reference 38 (GCA_000001405.2).
Supplementary_files_format_and_content: SeuratObject.Rds files are RDS files used in Signac processing of data (see https://satijalab.org/signac/ and our Online Methods for more detail). Metadata.csv files are csv files containing metadata information per cell (each row), with relevant cell information (each column). Counts.csv files contain the CSV formatted peaks (rows) by cellIDs (columns) matrix, where each value denotes read counts overlapping peaks per cell. Bam files are standard bams with cellIDs filtered to those used in downstream analyses.
Supplementary_files_format_and_content: s3atac.mm10.fragments.tsv: Tab-separated fragment files in bed format with fields <chr> <start> <end> <cell identifier> <count> for the s3ATAC mouse sample. Generated with custom scripts for input into Signac R library.

source_name: Whole brain

extract_protocol:
Tissue samples were minced on dry ice and resuspended in NIB (10 mM HEPES-KOH (Fisher Scientific, BP310-500 and Sigma Aldrich 1050121000, respectively), pH 7.2, 10 mM NaCl, 3mM MgCl2 (Fisher Scientific AC223210010), 0.1 % (v/v) IGEPAL CA-630 (Sigma Aldrich I3021), 0.1 % (v/v) Tween (Sigma-Aldrich P-7949) and diluted in PCR-grade Ultrapure distilled water (Thermo Fisher Scientific 10977015)). After a 5 minute incubation in ice, tissue was dounce homogenized to isolate nuclei. For GM12878, cells were spun down and resuspended in NIB. See https://dx.doi.org/10.17504/protocols.io.bd6wi9fe and https://dx.doi.org/10.17504/protocols.io.beb3jaqn for more details.
Tagmentation plates were prepared by the combination of 420 uL of 1400 nuclei/uL solution with 540 uL 2X TD Buffer (Nextera XT Kit, Illumina Inc. FC-131-1024). From this mixture, 8uL (~5000 nuclei total) was pipetted into each well of a 96 well plate dependent on well schema (Figure 2a). 1uL of 2.5uM uniquely indexed transposase was then pipetted into each well. Tagmentation was performed at 55°C for 10 minutes on a 300 rcf Eppendorf ThermoMixer. Following this incubation, plate temperature was brought down with a brief incubation on ice to stop the reaction. Dependent on experimental schema pools of tagmented nuclei were combined and 2uL 5mg/mL DAPI (Thermo Fisher Scientific D1306) was added.  Nuclei were then flow sorted via a Sony SH800 to remove debris and attain an accurate count per well prior to PCR. A receptacle 96 well plate was prepared with 9uL 1X TD buffer (Nextera XT Kit, Illumina Inc. FC-131-1024,diluted with ultrapure water), and held in a sample chamber kept at 4°C. Fluorescent nuclei were then flow sorted gating by size, internal complexity and DAPI fluorescence for single nuclei following the same gating strategy as previously described40. Immediately following sorting completion, the plate was sealed and spun down for 5 minutes at 500 rcf and 4°C to ensure nuclei were within the buffer.  Nucleosomes and remaining transposases were then denatured with the addition 1uL of 0.1% SDS (~0.01% f.c.) per well. 4uL of NPM (Nextera XT Kit, Illumina Inc) per well was subsequently added to perform gap-fill on tagmented genomic DNA, with an incubation at 72°C for 10 minutes. 1.5 uL of 1uM A14-LNA-ME oligo was then added to supply the template for adapter switching (Supplementary Table 3). The polymerase based adapter switching was then performed with the following conditions: initial denaturation at 98°C for 30 seconds, 10 cycles of 98°C for 10 seconds, 59°C for 20 seconds and 72°C for 10 seconds. The plate was then held at 10°C. After adapter switching 1% (v/v) Triton-X 100 in ultrapure H2O (Sigma 93426) was added to quench persisting SDS. At this point, some plates were stored at -20°C for several weeks while others were immediately processed.   The following was then combined per well for PCR: 16.5 ul sample, 2.5uL indexed i7 primer at 10 uM, 2.5uL indexed i5 primer at 10 uM, 3 uL of ultrapure H2O, and 25 uL of NEBNext Q5U 2X Master mix (New England Biolabs M0597S), and 0.5uL 100X SYBR Green I (Thermo Scientific S7563) for a 50 uL reaction per well (Supplementary Table 4-5). A real time PCR was performed on a BioRad CFX with the following conditions, measuring SYBR fluorescence every cycle: 98°C for 30 seconds; 16-18 cycles of 98°C for 10 seconds, 55°C for 20 seconds, 72°C for 30 seconds, fluorescent reading, 72°C for 10 seconds. After fluorescence passes an exponential growth and begins to inflect, the samples were held at 72°C for another 30 seconds then stored at 4°C.

tag: condition

tag: strain


title: LUCAP_173_1_S7

value: NEPC


title: SB+ANI treated sample rep 1


title: Control and CTE from Run5.2

status: Public on May 24 2021

first: Kevin

middle: Blake

last: Chancellor

address: 77 Avenue Louis Pasteur

department: Genetics

institute: Harvard Medical School

data_processing:
Data processing, from binary base call files produced from next-generation sequencing to generation of counts matrices, was carried out using the inDrops processing pipeline (https://github.com/indrops/indrops). Project specific details can be found at https://github.com/blakechancellor/Altered-oligodendroglia-and-astroglia-in-chronic-traumatic-encephalopathy.
Step 1. Produciont of fastq files. Fastq files were produced using bcl2fastq with default settings.
Step 2. Filter. This step filters reads and only keeps reads that match the expected inDrops read structure. Default settings were used.
Step 3. Identify abundant barcodes. Reads are organized by abundant barcodes (nuclei) and separated by library (sample) of origin. Default settings were used.
Step 4. Sort reads. Reads within samples were sorted by nucleus of origin. Barcodes with fewer than 250 reads were ignored. Default setting were used.
Step 5. Quantify expression. Reads were mapped to the reference genome using bowtie1.1.1. Alignment created BAM files using samtools. Aligned counts were quantified to create unique molecular identifier filter mapped counts matrices.
Step 6 Aggregate. The pipeline can divide the data and run each step in parallel to expedite the process. This step combines data from several parallelized steps.
Genome_build: GRCh38.97
Supplementary_files_format_and_content: Processed files contain the gene expression counts for each sample. Each file is a counts matrix with cellular barcodes as columns and gene names as rows. Values indicate raw counts for each time a gene was detected in a specific cellular barcode.

source_name: Dorsolateral frontal white matter

extract_protocol:
All tissue was macrodissected into 30mg sections from dorsolateral frontal white matter (Brodmann area 8/9) with a scalpel blade and stored at -80° Celsius. On the day of single-nucleus encapsulation, tissue was placed in a 7mL dounce with ice cold buffer (0.5M sucrose, 2M KCl, 1M MgCl2, 1M Tricine-KOH pH 7.8, spermine, spermidine, DTT, RNasin, H2O) and dounced with a tight pestle ten times to mechanically dissociate and homogenize the tissue. To further dissociate the tissue, 5% IGEPAL-CA630 was added to the homogenate and dounced 5 times. The homogenate was then strained through a 40μm cell strainer (VWR) on ice. 5mL of 50% iodixanol was added to the homogenate and mixed. In a 13mL ultraclear ultracentrifuge tube, 1mL of 40% iodixanol was added with 1mL of 30% iodixanol carefully layered on top, followed by 10mL of the tissue homogenate. The tissue was then ultracentrifuged in a swing-bucket rotor at 10,000xg for 18 minutes. Following ultracentrifugation, the 35% iodixanol layer was collected and the nuclei were counted. Nuclei were diluted with 30% iodixanol to 90,000 nuclei/mL for inDrops chip loading. Nuclei were isolated for inDrops using a previously described method.
Libraries were constructed following the inDrops single-nucleus RNA sequencing pipeline: https://singlecellcore.hms.harvard.edu/ and Klein, et al. Cell. 2015. Briefly, nuclei were encapsulated in a hydrogel via a microfluidic chip. Once encapsulated, nuclei were lysed and free-floating mRNA molecules were reverse transcribed to cDNA and ligated with cellular barcodes, library indices, unique molecular identifiers, and next-generation sequencing primers. Prepared cDNA molecules were pooled, sequenced on a NextSeq500 and de-multiplexed to cell and sample of origin bioinformatically.

tag: brodmann  area

tag: brain region

value: Dorsolateral frontal sulcus


title: DMSO treated control rep 4