title: B14/15_late-1


title: BT2-2_rep3

status: Public on Oct 02 2024

type: SRA

city: Montreal

first: William

last: Pastor

state: Quebec

address: 3655 Promenade Sir-William-Osler rm900

department: Biochemistry

country: Canada

institute: McGill University

library_source: transcriptomic

data_processing:
For RNA and ChIP/ATAC sequencing analysis, Genpipes 4.3.2 (https://bitbucket.org/mugqic/genpipes) provided the pipeline for basic sequencing processing.  In brief for RNAseq, sequencing quality and adaptor removal was trimmed with trimmomatic (v0.36). Trimmed fastq files alignment was performed with STAR aligner (v2.7.8a) using hg38/GRCh38 (ensembl v104). Picard (v2.9.0) was then used merged, mark duplicates, identify unique read, and sort .bam files proceeding alignment. Read counts were collected using HTseq-count and StringTie (v1.3.5). Differentially expressed gene (DEG) comparison was performed with DESeq2 package on RStudio (R v4.3.1). Correlation matrix, hierarchal gene cluster analysis, and PCA were generated in R. Gene pathway analysis were performed on ConsensusPathwayDB and EnrichR. For ChIPSeq and ATACseq, in brief, raw fastq files trimmed using trimmomatic (v.0.36). Then qualified fastq reads were mapped with BWA (v.0.7.17) with post processing with sambamda (v0.8.1) to merge replicates, mark and filter duplicates, and remove blacklist regions. Peak calling and differential-bind was performed with MACS2 (v2.2.7.1). Gene annotations and motif analysis used with Homer (v4.11). Bam to bigwig bamCoverage from deepTools (v3.5.1) was used to generate the tracks for viewing on IGV (v2.9.4).
Assembly: hg38
Supplementary files format and content: tab-delimited text includes raw counts for each sample

molecule: total RNA

organism: Homo sapiens

source_name: Trophoblast Stem Cell

growth_protocol: TSC were cultured in TSC basal media containing DMEM F-12 (GIBCO), 1x ITS-X (GIBCO), 0.3% BSA (WISENT), 1% Penicillin/Streptomyocin (GIBCO), 1% ESC qualified Fetal Bovine Serum (GIBCO), 0.1mM β-mercaptoethanol (GIBCO), 15ug/ml L-ascorbic acid (SIGMA), and 50ng/ml recombinant hEGF (GIBCO). To this, made fresh, 0.75mM valporic acid, 2uM CHIR99021 (CaymenChem), 0.5uM A8301 (CaymenChem), 1uM SB431542 (CaymenChem), and 5μM Y27632 (CaymenChem) was added to make TSCMedia. TSC were dissociated using TRYPLE (GIBCO) diluted with PBS to 30% and incubated at 37C for 10 min. Deactivation of TRYPLE was done by using 1:1 vol of 0.5mg/ml Soybean trypsin inhibitor (GIBCO) diluted in PBS. For experiments performed in figure 1-2, TSC were passaged on to 5ug/ml of Collagen IV (Corning) coated plates, but due to product availability, TSC from experiment in figure 3-6 were passaged onto Laminin 511(SIGMA) coated plates.

extract_protocol:
Total RNA isolation used manufacturing protocol indicated by Sigma-Aldrich RNAzolRT R4533. Qubit RNA BR Assay kit (Q10211) was used to accurately measure RNA concentration.
For RNA library synthesis, in brief, purified mRNA was cleaned using the NEBNext Poly(A) mRNA Magnetic Isolation Module kit (NEB E7490) and final library generation was created with the NEBNextÒ Ultra RNA Library Prep Kit for IlluminaÒ (NEB E7530) following manufacturing instructions. Barcoding came from TruSeq Unique Dual Indexes (Illumina, San Diego, CA) QubitÔ 1x dsDNA HS Assay kit (Q33231) was used to measure synthesis of the library.

treatment_protocol: StemCell Technologies Hypoxia Incubator Chamber (cat:27310) and mixed O2 and CO2 Praxiar tanks were injected into control the O2 levels.

tag: cell line

value: Trophoblast Stem Cell

tag: genotype

value: WT

tag: treatment


title: RP_0-2

city: Wuhan

first: Liang

last: Hu

address: 1 Nanhu Yaoyuante, Hongshan District

country: China

institute: Hubei Academy of Agricultural Sciences

data_processing:
Data were filtered by abandoning adapter sequences and low-quality reads.
The obtained clean reads, ranging from 18 nt to 36 nt in length, underwent deduplication to generate unique reads for further exploration. These reads were matched to the Rfam (ver. 13.0) database using BLAST to identify rRNA, tRNA, snRNA, and snoRNA, adhering to a criterion of no more than two mismatches.
Unclassified reads were matched to the miRBase database (ver. 22.0) using BLAST with the same criterion to identify mature miRNA sequences.
The identified known miRNAs were classified into families by aligning them with reference miRNAs from the miRBase database.
Unmapped reads were analyzed against the genome using the Mireap online program to predict novel miRNAs.
Assembly: Rfam (ver. 13.0) database and miRBase database (ver. 22.0)
Supplementary files format and content: TXT file contain counts for each sample

organism: Oryza sativa

source_name: Leaf sheaths of rice

growth_protocol: The pyramiding line containing both BPH14/BPH15 genes (B1415) was created using the indica cartivar ‘Wushansimiao’ as the recurrent parent (RP) [12]. Rice seeds were sown in plastic contains (15 cm tall and 9 cm wide) with 15 per container and maintained in a controlled environment with 14 h light (32 ± 2°C)/10 h dark (26 ± 2°C) cycle. BPHs were reared on the susceptible cultivar ‘Taichung Native1’ (TN1, IRRI Acc. No.00105) under the same environment.

extract_protocol:
Total RNA was extracted using TRIzol from leaf sheaths of rice.
Eighteen small RNA libraries were produced using the NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England Biolabs, Inc.), encompassing samples from B1415 and RP at non-infestation (B1415_0, RP_0), early infestation (B1415_early, RP_early), and late infestation (B1415_late, RP_late) stages, each with three biological replicates. The small RNA libraries underwent quantification using a high-sensitivity DNA Kit on a Bioanalyzer 2100 system (Agilent) prior to sequencing on the Illumina NovaSeq 6000 platform.

treatment_protocol: Sample collection and BPH treatments were executed following the endpoint method [33]. Despite varying start times, all treatments were concluded concurrently. Four-leaf stage seedlings of B1415 and RP were infested with eight 3rd instar BPH nymphs per seedling at 0, 3, 6, 12, 24, 48, and 72 h with 15 seedlings each in three biological replicates. Leaf sheath samples were categorized as non-infested (0 h), early infestation stage (3, 6, 12 h), or late infestation stage (24, 48, 72 h), following prior methodology [33] and designated as B1415_0, B1415_early, B1415_late, RP_0, RP_early, and RP_late, respectively. All samples were snap frozen, ground, and preserved in TRIzol (Invitrogen Life Technologies) at -80°C.

tag: tissue

value: Leaf sheaths of rice

value: The recurrent parent (RP) 'Wushansimiao', an indica inbred rice cultivar

value: Non-infested by BPH


title: 34156

type: genomic

city: Bethesda

first: Alexander

middle: Christian

last: Harris

state: Maryland

address: 50 South Drive

department: National Human Genome Research Institute

country: USA

institute: National Institutes of Health

description: BELS

hyb_protocol: Resuspended DNA was loaded onto BeadChips arrays and hybridized overnight at at 48oC.  Single-base extension and allele-specific staining was performed by hand.

scan_protocol: After allele-specific staining BeadChip arrays were coated with XC4/ethanol , dried for 1 hour and scanned on a BeadArray Reader (Illumina).

data_processing:
Image data was analyzed using GenomeStudio with GT module (Illumina).
genotypes AA, AB, BB, NC=no call

molecule: genomic DNA

organism: Canis lupus familiaris

source_name: Whole Blood

label_protocol: 200ng of genomic DNA was amplified in an overnight reaction at 37oC. The amplified DNA was enzymatically fragmented, precipitated with isopropanol, and resuspended in hybridization buffer.

extract_protocol: Genomic DNA was extracted from whole blood or Buccal cells using Proteinase K and phenol/chloroform with standard protocols. Genomic DNA extracted from saliva using standard Performagene genotek DNA extraction protocols.

tag: breed

value: Belgian Sheepdog


title: CT3-5_rep3


title: 1151 CA1-

city: Minneapolis

first: Maria

last: Razzoli

state: Minnesota

department: Integrative Biology and Physiology

institute: University of Minnesota

data_processing:
FASTQ files were filtered and demultiplexed using DND2.0 with the following parameters: quality trim score = 20, adaptor trim match length = 10, adaptor trim max mismatch = 3, barcode max mismatch = 1, stitching max mismatch = 2, dedup-hd = 1
DND was used to convert the raw FASTQ files to Digital Count Conversion (DCC) file format and uploaded to the Nanostring GeoMx DSP Anaysis Suite
Limit of Quantification was derived and used to exclude genes that were not expressed at a level of LOQ raised to the power of 2 in <20% of AOIs
Individual probe counts were normalized to the median probe count for each AOI
Assembly: mm10
Supplementary files format and content: Lyons_et_al_geomix_median_norm_q_datasets2.0.csv
Library strategy: spatial transcriptomics

organism: Mus musculus

source_name: brain

extract_protocol:
Formalin fixed paraffin embedded tissue was sectioned at 5um and prepared using the Nanostring GeoMx workflow according to the manufacturer's instructions
Spatial transcriptomic sequencing libraries were prepared using the Nanostring GeoMx workflow according to the manufacturer's instructions

value: brain

tag: group

value: ctrl

value: low

tag: region

value: hippocampus

tag: subregion

tag: mouse id


title: CT1-2_rep2


title: BT2-5_rep2


title: CT1-20_rep3


title: RP_early-1

value: Early feeding stage


title: 34310

description: AFFN

value: Affenpinscher


title: CT3-5_rep2


title: B14/15_early-2

value: The near isogenic lines (NILs) contain both the BPH14 and BPH15


title: BT2-2_rep2


title: 1175 SS-

value: cortex

value: SS


title: 34322

description: BELS


title: 1151 CA1+

value: high


title: 33500

description: AFFN

source_name: Saliva


title: 34153

description: SEAL

value: Sealyham Terrier


title: B14/15_early-1


title: CT3-2_rep2

value: Trophoblast Stem Cell


title: 34142

description: SHWD

value: Spanish Water Dog


title: 1175 MC+

value: MC


title: 33501

description: AFFN


title: B14/15_early-3

value: Leaf sheaths of rice


title: 33512

description: LANC

value: Lancashire Heeler


title: 1175 CP-

value: choroid plexus

value: CP


title: 1175 CP+

value: brain

value: ctrl


title: 33431

description: CESK

value: Cesky Terrier


title: B14/15_0-2

value: Non-infested by BPH


title: 1175 CA1+

value: hippocampus


title: B14/15_0-1

value: The near isogenic lines (NILs) contain both the BPH14 and BPH15


title: CT1-5_rep2

value: WT


title: BT2-2_rep1


title: 33463

description: CESK


title: 1175 MC-

value: low

value: cortex


title: B14/15_0-3


title: CT3-20_rep2


title: 33334

description: SEAL

value: Sealyham Terrier


title: 33398

description: SEAL


title: 1175 CA3-


title: 1168 SS+

value: high

value: SS


title: CT1-20_rep2

value: Trophoblast Stem Cell


title: 1168 SS-


title: 1168 MC+

value: MC


title: 33281

description: SEAL


title: lhp1-14d-R3

city: Guangzhou

last: Zhenwei

state: Guangdong

address: Xingang West Road

department: School of Life Science

institute: Sun Yat-sen University

data_processing:
Reads were mapped to the TAIR10 Arabidopsis genome using TopHat (Galaxy V2.1.1 in usegalaxy.org) with default settings, except that a minimum intron length of 20 bp and a maximum intron length of 5,000 bp were required (Paired-end). Then, mapped reads were assembled according to TAIR10 version of genome annotation using cufflinks (version 2.1.1) with default settings. To analyze differential expression, the assembled transcripts from three independent biological replicates in Col and other mutants were included and compared using Cuffdiff (version 2.1.1) with default settings.
Assembly: TAIR10
Supplementary files format and content: tab-delimited text files include RPKM values for wt, pkl, clf and lhp1

organism: Arabidopsis thaliana

source_name: whole seedlings

growth_protocol: Plants were grown in 1/2MS media with 22 degree in 16h light/8h dark

extract_protocol:
RNA was harvested using the HiPure Universal RNA Mini Kit (MAGEN, Cat. No. R4130). 1 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: whole seedlings


title: 1175 CA3+


title: clf-14d-R3


title: 33254

description: NGSD

value: New Guinea Singing Dog


title: CT3-5_rep1


title: CT3-2_rep1


title: 33312

description: MCUR

value: Mountain Cur


title: BT2-5_rep1


title: lhp1-14d-R2


title: lhp1-14d-R1


title: 1168 CP+

value: brain

value: choroid plexus

value: CP


title: 1168 MC-

value: ctrl


title: CT1-5_rep1


title: clf-14d-R2


title: 33252

description: BGPC

value: Berger Picard


title: 33251

description: BGPC


title: 1168 CP-


title: clf-14d-R1


title: 33235

description: SEAL


title: 1168 CA3+

value: hippocampus


title: BT2-20_rep1

value: WT


title: CT1-2_rep1


title: 33229

description: POPO

value: Portuguese Podengo Pequeno


title: CT3-20_rep1


title: 1168 CA1+


title: 33193

description: SBYH

value: Stabyhoun


title: CT1-20_rep1


title: pkl-14d-R2


title: 33148

description: BOER

value: Boerboel


title: 1168 CA1-

value: low


title: CT3-TSC-non-target2_c5

treatment_protocol: Cells were grown in either steady-state TSC or differentiated to EVT or STB3D. TSC were converted to EVT using a modified TSC basal media containing DMEM F-12 (GIBCO), 1x ITS-X (GIBCO), 0.3% BSA (WISENT), 1% Penicillin/Streptomyocin, and 0.1mM β-mercaptoethanol (GIBCO). For day 1-2, TSC were passage onto 1ug/ml Collagen IV coated plates in mod. TSC basal with added 5% Knock-serum replacement (GIBCO), 5uM Y27632 (CaymenChem), 3uM A8301 (CaymenChem), 100ng/ml NRG1 (NEB) and 2% GFR-Matrigel (Corning) while the media is still cold. Day 3-5, the media is change to the previously mention ETVM but this time without NRG1 and reduction to 0.5% GFR-Matrigel. Day6-8, the media is change to the previously mention ETVM but this time without NRG1, KSR, and reduction to 0.5% GFR-Matrigel. At the end of day 8, differentiated EVT can be assess by Flowcytometry. TSC were converted to STB3D using a modified TSC basal media containing DMEM F-12 (GIBCO), 1x ITS-X (GIBCO), 0.3% BSA (WISENT), 1% Penicillin/Streptomyocin, and 0.1mM β-mercaptoethanol (GIBCO). For day 1-2, TSC were passage on to suspension plate (Starsted) in TSC basal media containing 5uM Y27632, 2uM Forskolin (CaymenChem), 5% KSR (GIBCO), and 50ng/ml recombinant hEGF. On day 3, cell clustered were collect and pulse-spin for 30sec to separate single cell from STB fusion-clusters and replate in STBMedia mentioned above. At day 5, STB-3D are ready to be assessed by immunofluorescence and/or hCG- ELISA.

value: TSC


title: pkl-14d-R1


title: CT3-STB3D-non-target2_c5

value: Trophoblast Stem Cell


title: 33067

description: DARG

value: Dogo Argentino


title: CT3-EVT-non-target2_c4

value: EVT


title: 1162 SS-

value: CSS

value: cortex

value: SS


title: 32908

description: IRSE

value: Irish Setter


title: WT-14d-R2

value: whole seedlings


title: 32896

description: SAAR

value: Saarloos Wolfdog


title: 1162 MC+

value: brain

value: high

value: MC


title: 1168 CA3-

value: ctrl

value: hippocampus


title: CT3-STB3D-non-target2_c3

value: WT


title: pkl-MNase-R2

library_source: genomic

data_processing:
To normalize data among samples, we applied quantile normalization methods to normalizeoccupancy with Wiq function in DANPOS. To identify high-quality nucleosome peaks, the Dpos algorithm was used. To analyze the distributions of chromatin features flanking each group of genomic sites, we calculated nucleosome read coverage by bedmap from BEDPOS with option -wmean
Assembly: TAIR10
Supplementary files format and content: bigwig

extract_protocol:
For nuclear extraction, 2-week-old Arabidopsis seedlings without roots were ground in liquid nitrogen and resuspended in lysis buffer [50 mM HEPES, pH7.5, 150 mM NaCl, 1 mM ethylenediaminetetraacetic acid (EDTA), 1% Triton X-100, 10% glycerol, 5 mM b-mercaptoethanol and protease inhibitor cocktail. After lysis for about half an hour, the lysis mixture was filtered through a 40-lm cell strainer. The filtered plant extracts were centrifuged for 20 min at 3200 g to precipitate nuclei. Isolated nuclei were washed once with HBB buffer (25 mM Tris–HCl, pH 7.5, 0.44 M sucrose, 10 mM MgCl2, 0.1% Triton-X and 10 mM b-mercaptoethanol) and centrifuge at 1500g for 10 min at 4°C. Once with HBC buffer (20 mM Tris–HCl, pH 7.5, 352 mM sucrose, 8 mM MgCl2, 0.08% Triton-X, 8 mM b-mercaptoethanol and 20% glycerol) and centrifuge at 1500g for 10 min at 4°C. The obtained nuclei were resuspended with 100 ul of HBC buffer and flash frozen in liquid nitrogen.
Two ng of DNA was used for library construction.  Libraries were constructed by the VAHTSTM Universal DNA Library Prep Kit for Illumina® V3 (ND607), VAHTSTM DNA Adapters set3 - set6 for Illumina® (N805) and VAHTS® DNA Clean Beads (N411-02, Vazyme) according to the manufacturer’s protocol. protocol. Illumina Novaseq was used for high-throughput sequencing of the ChIP-seq library.


title: CT3-TSC-non-target2_c3


title: 32885

description: CZEC

value: Czechoslovakian Wolfdog


title: 1162 MC-

value: low


title: pkl-MNase-R1


title: 1162 CP+

value: choroid plexus

value: CP


title: CT3-TSC-non-target2_c2

value: Trophoblast Stem Cell

value: TSC


title: 31969

description: NGSD

value: New Guinea Singing Dog


title: 31968

description: NELK

value: Norwegian Elkhound


title: CT3-STB3D-non-target2_c2


title: WT-MNase-R2


title: 1162 CP-

value: CSS


title: WT-MNase-R1

value: whole seedlings


title: 1162 CA3+


title: 1162 SS+

value: cortex

value: SS


title: CT3-EVT-non-target2_c2

value: EVT


title: WT-14d-R1


title: pkl-14d-R3


title: CT3-EVT-non-target2_c1


title: 31787

description: BEDT

value: Bedlington Terrier


title: 31730

description: RSTR

value: Russell Terrier


title: 1162 CA3-

value: brain

value: hippocampus


title: CLF-GFP pkl R2

data_processing:
The raw reads were mapped to the reference genome (Arabidopsis thaliana genome, TAIR10) with Bowtie for Illumina (Galaxy Version 1.1.2) for DNA sequencing on web https://usegalaxy.org/ with default setting, except that the paired-end was required.
MACS2 (Galaxy Version 2.1.1) was used for peak calling using the following parameters “gsize=119,667,750, bw=300, q=0.01, nomodel, extsize=200”.
The alignments were first converted to Wiggle (WIG) files and the bigwig files were generated using bamCoverage with “-bs 10”, “--normalizeUsing RPKM (reads per kilobase per million)” in DeepTools and then the data were imported into the Integrative Genomics Viewer (IGV) for visualization.
Assembly: TAIR10
Supplementary files format and content: bigwig

source_name: whole seedling

extract_protocol:
Lysates were clarified from sonicated protein-DNA complexes were isolated with GFP antibody.
Two ng of ChIP DNA immuoprecipitated by anti-GFP antibody was used for ChIP-seq library construction.  Libraries were constructed by the VAHTSTM Universal DNA Library Prep Kit for Illumina® V3 (ND607), VAHTSTM DNA Adapters set3 - set6 for Illumina® (N805) and VAHTS® DNA Clean Beads (N411-02, Vazyme) according to the manufacturer’s protocol. protocol. Illumina Novaseq was used for high-throughput sequencing of the ChIP-seq library.

value: whole seedling


title: CLF-GFP pkl R1


title: WT-14d-R3


title: CT3-EVT-GCM1KO2_c4


title: 31729

description: RSTR


title: CLF-GFP R2


title: 1162 CA1+

value: high


title: 31614

description: KMUN

value: Kleine Munsterlander


title: 1162 CA1-

value: low


title: CT3-TSC-GCM1KO2_c1

value: Trophoblast Stem Cell


title: 1157 SS+


title: CT3-STB3D-GCM1KO2_c1


title: CLF-GFP R1


title: CT3-TSC-GCM1KO2_c2

value: TSC


title: 31612

description: KMUN


title: PKLdSNF GFP-IP-R2


title: 31519

description: SILA

value: Pastore della Sila


title: 1157 SS-


title: CT3-STB3D-GCM1KO2_c2


title: 31378

description: CAUC

value: Caucasian Ovcharka


title: PKLdCHROMO GFP-IP-R2


title: 31375

description: CAUC


title: 1157 MC+

value: CSS

value: MC


title: CT3-EVT-GCM1KO2_c2


title: CT1-STB3D-GCM1KO1_c2


title: 31284

description: CIRN

value: Cirneco dellEtna


title: PKLdCHROMO GFP-IP-R1


title: PG GFP-IP-R2


title: 1157 CP+

value: choroid plexus

value: CP


title: CT1-STB3D-non-target_c2

value: WT


title: 31283

description: CIRN


title: CT1-STB3D-GCM1KO1_c1


title: PG GFP-IP-R1


title: 1157 CP-


title: 31121

description: CESK

value: Cesky Terrier


title: AMIRMSI1-anti-PKL-R2


title: 1157CA3+

value: brain


title: AMIRMSI1-anti-PKL-R1


title: 1157 CA3-

value: hippocampus


title: CT1-STB3D-non-target_c1


title: PKLdSNF GFP-IP-R1

value: whole seedling


title: 1157 CA1+

value: high


title: 30861

description: IWSP

value: Irish Water Spaniel


title: CT3-STB3D-GCM1KO1_c3


title: CT3-STB3D-GCM1KO1_c2


title: 31228

description: BRUS

value: Brussels Griffon


title: 1157 CA1-

value: low


title: WT-anti-PKL-R1


title: CT3-STB3D-non-target_c4

value: Trophoblast Stem Cell


title: 1157 MC-

value: cortex


title: 1152 SS+

value: SS


title: 30359

description: ESHP

value: English Shepherd


title: CT3-STB3D-non-target_c3


title: 30504

description: WSSP

value: Welsh Springer Spaniel


title: 1152 SS-

value: CSS


title: 30183

description: PTWD

value: Portuguese Water Dog


title: CT1-EVT-GCM1KO1_c3

value: EVT


title: MSI1-GFP-R1


title: pkl-K27-spike-in-R2


title: 1152 MC-

value: MC


title: 30108

description: ALKK

value: Alaskan Klee Kai


title: 30054

description: PTWD


title: CT1-EVT-GCM1KO1_c1_rep1


title: pkl-K27-spike-in-R1


title: WT-anti-PKL-R2


title: WT-K27-spike-in-R2


title: CT1-EVT-non-target_c1_rep2

value: WT


title: 30034

description: PTWD


title: 1152 CP+

value: brain

value: choroid plexus

value: CP


title: WT-K27-spike-in-R1

value: whole seedling


title: CT3-TSC-GCM1KO1_c3

value: TSC


title: 1152 CA3+

value: high

value: hippocampus


title: MSI1-GFP-R2


title: 30003

description: PTWD


title: CT3-TSC-non-target_c4

value: Trophoblast Stem Cell


title: CT3-TSC-GCM1KO1_c2


title: 29896

description: PTWD


title: 1152 CA3-

value: low


title: 1152 MC+

value: cortex


title: CT3-TSC-non-target_c2


title: 29629

description: CARD

value: Cardigan Welsh Corgi


title: 1175 CA1-

value: ctrl


title: 29612

description: TERV

value: Belgian Tervuren


title: 29609

description: IWSP

value: Irish Water Spaniel


title: 1152 CA1-

value: CSS


title: pkl pkr2-K27me3 R2


title: 63-3

city: Wilmington

first: Qi

last: Zheng

state: DE

address: 1701 Rockland Road

institute: Nemours Children's Health

description: 10x Genomics

library_source: transcriptomic single cell

data_processing:
All data processing steps used the cellranger v7.1.0 pipeline, incluidng fastq calling, demultiplexing, counting shown below.
Call: FASTQ files were called using the cellranger 'mkfastq' program
Combine: raw reads sequenced over 2 NextSeq runs were combined for every sample with Linux 'cat' cmd
Count: cell level counts were generated using the cellranger  'count' program based on the pre-built 'mm10-2020-A' database.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

extract_protocol:
Single cell RNA sequencing (scRNAseq) was performed by creation of a single cell solution using the GentleMACS brain dissociator (Miltenyi Biotec) and Adult Brain Dissociation Kit (Miltenyi Biotec). Cell viability was confirmed via trypan blue staining. Live and dead cells were counted using a hemocytometer.
The 10x Genomics Chromium Next GEM Single Cell 3’ Kit v3.1, Dual Index Kit TT Set A, Chromium Next GEM Chip G Single Cell Kit, and SPRIselect Reagent Kit were used for library creation. Sample and library quality control was achieved using Agilent High Sensitivity D5000 ScreenTape, D5000 Reagents, D5000 Ladder and KAPA Universal Library Quantification Kit.

treatment_protocol: Adapted from Aravamuthan et al., 2020, Postnatal day 6 (P6) was chosen for hypoxia administration due to poor animal survival at later time points. Mice were placed in a hypoxia chamber (Biospheryx) on a heated pad (37.2 °C) and subjected to 8 minutes of either progressive hypoxia (HIE) from 21% to 0% oxygen or normoxia (CTRL) (21% oxygen). After 8 minutes of treatment, the chamber door was opened to allow rapid recovery to 21% oxygen. Surviving mice were returned to their mother for further recovery.

value: CF-1 wildtype

value: CTRL


title: pkl pkr2-K27me3 R1


title: PKL-dSANTdSLIDE-GFP IP-R1


title: PKL-GFP IP-R2


title: 1152 CA1+

value: brain


title: 29597

description: GOLD

value: Golden Retriever


title: PKL-dSANTdSLIDE-GFP IP-R2


title: 63-2

description: 10x Genomics


title: CT1-TSC-non-target_c1_rep1

value: WT


title: 29584

description: NEWF

value: Newfoundland


title: CT1-TSC-GCM1KO1_c1_rep2

value: TSC


title: CT1-EVT-GCM1KO1_c1_rep2

value: EVT


title: pkr2-K27me3 R1

value: whole seedling


title: 29583

description: AMST

value: American Staffordshire Terrier


title: pkr2-K27me3 R2


title: CT1-EVT-non-target_c1_rep1

value: Trophoblast Stem Cell


title: 29595

description: GOLD


title: 63-1

description: 10x Genomics


title: pkl-14d-H3K4me3-R2


title: PKL-GFP IP-R1


title: CT1-TSC-GCM1KO1_c3


title: 60-7

description: 10x Genomics

value: HIE


title: 29317

description: PTWD

source_name: Unknown Source

value: Portuguese Water Dog


title: CT1-TSC-GCM1KO1_c1_rep1


title: 63-4

description: 10x Genomics


title: pkl-14d-H3K4me3-R1


title: 29268

description: PTWD


title: CT1-TSC-non-target_c1_rep2


title: WT-14d-H3K4me3-R2


title: 29216

description: PTWD


title: 29065

description: PTWD


title: 60-5

description: 10x Genomics


title: CT3-EVT-GCM1KO1_c2


title: 28908

description: BEDT

value: Bedlington Terrier


title: WT-14d-H3K4me3-R1


title: 60-4

description: 10x Genomics


title: CT3-EVT-GCM1KO1_c1_rep2


title: CT3-EVT-non-target_c3


title: WT-H3


title: 28513

description: SKYE

value: Skye Terrier


title: pkl-5d-H3K27me3-R2


title: CT3-EVT-non-target_c1_rep2


title: 59-9

description: 10x Genomics


title: 59-2

description: 10x Genomics


title: 28510

description: SKYE


title: CT3-EVT-GCM1KO1_c1_rep1


title: pkl-5d-H3K27me3-R1


title: 28502

description: BOX-

value: Boxer


title: 30171

description: PTWD


title: 59-1

description: 10x Genomics


title: 60-6

description: 10x Genomics


title: pkl-H3


title: CT3-EVT-non-target_c1_rep1


title: WT-5d-H3K27me3-R2


title: 59-10

description: 10x Genomics


title: 28483

description: PARS

value: Parson Russell Terrier


title: 56-4

description: 10x Genomics

value: CF-1 wildtype


title: 28467

description: GOLD


title: CT3-EVTd3_INPUT_rep2

data_processing:
For RNA and ChIP/ATAC sequencing analysis, Genpipes 4.3.2 (https://bitbucket.org/mugqic/genpipes) provided the pipeline for basic sequencing processing.  In brief for RNAseq, sequencing quality and adaptor removal was trimmed with trimmomatic (v0.36). Trimmed fastq files alignment was performed with STAR aligner (v2.7.8a) using hg38/GRCh38 (ensembl v104). Picard (v2.9.0) was then used merged, mark duplicates, identify unique read, and sort .bam files proceeding alignment. Read counts were collected using HTseq-count and StringTie (v1.3.5). Differentially expressed gene (DEG) comparison was performed with DESeq2 package on RStudio (R v4.3.1). Correlation matrix, hierarchal gene cluster analysis, and PCA were generated in R. Gene pathway analysis were performed on ConsensusPathwayDB and EnrichR. For ChIPSeq and ATACseq, in brief, raw fastq files trimmed using trimmomatic (v.0.36). Then qualified fastq reads were mapped with BWA (v.0.7.17) with post processing with sambamda (v0.8.1) to merge replicates, mark and filter duplicates, and remove blacklist regions. Peak calling and differential-bind was performed with MACS2 (v2.2.7.1). Gene annotations and motif analysis used with Homer (v4.11). Bam to bigwig bamCoverage from deepTools (v3.5.1) was used to generate the tracks for viewing on IGV (v2.9.4).
Assembly: hg38
Supplementary files format and content: tab-delimited text includes peakCalls over INPUT of the merged replicates

extract_protocol:
Cell samples are fixed with 0.66% paraformaldehyde (FisherSci) diluted with PBS and incubated on a rotator for 10 minutes at room temperature. Samples are quenched by adding glycine to a final concentration of 0.125M with continued rotation for another 10 minutes at room temperature then wash with cold PBS 2x and replaced with PBS + 1mM EDTA + protease inhibitor (Roche)before collection with a cell scrapper. Cell pellets were centrifuged at 13200 rpm for 5 mins. Nuclear lysis extraction begins with 10 min incubation on a rotator in nuclear lysis buffer (50mM HEPES pH 7.8, 0.5% Triton X-100, 1mM EDTA, 0.5mM EGTA, 140 mM NaCl, 10% glycerol and 1% NP-40). Nuclear pellets are collected by centrifugation at 13200 rpm for 5 minutes at 4°C. Nuclei are resuspended in nuclear wash buffer (10mM Tris-HCl pH 8.0, 200mM NaCl, 1mM EDTA, and 0.5mM EGTA) and then rotated 5 minutes at 4°C the repeated twice. Nuclear pellets are then resuspended in SDS lysis buffer (50mM Tris-HCl pH 8, 10mM EDTA, 1% SDS, and 1% Triton X-100). Nuclei samples are transferred to a 1ml tube (Bioruptor) and keep cold before sonication. Sonication was performed using the Diagenode Bioruptor sonicator (settings: 30sec on, 30 sec off, 30-35 cycles). Dilute the sonicate by 10-fold with Dilution buffer (25mM Tris-HCl pH 8, 150 mM NaCl, 3mM EDTA, and 1% Triton X-100). Pre-washed Protein G Sepharose beads (SIGMA, P3296-5ml) are prepared by washing them three times with dilution buffer. A pre-clearing step is performed by using 40μl of pre-washed beads combined sonicated sample. This rotated for 1hrs at 4°C then centrifuged at 1000xg for 1 min. Sticky-chromatin is remove by the beads and the supernatant is retained for the IP. 2ug of GCM1(HPA001343) antibody is added to each pre-cleared sample and then rotated overnight at 4°C. The next day, pre-washed Protein G Sepharose beads are added to the ChIP samples and rotated 2 hours at 4°C. The beads washed with 3 buffer: DB150 (25mM Tris-HCl pH 8.0, 150mM NaCl, 3mM EDTA, 1% Triton X-100, and 0.05% SDS), DB500 (25mM Tris-HCl pH 8.0, 500mM NaCl, 3mM EDTA, 1% Triton X-100, and 0.05% SDS), Buffer III (10mM Tris-HCl pH 8.0, 250mM LiCl, 1% Sodium Deoxycholate, 1% NP-40, and 1mM EDTA, and lastly TE buffer (10mM Tris-HCl pH 8.0, 1mM EDTA). DNA is eluted for 15 min on shaker rotating at 1000rpm with 200ul elution buffer (100 mM NaHCO3, and 1% SDS). 16ul of 5M NaCl is added to each sample and further incubated at 65C overnight for decrosslinking. Ethanol precipitation is performed to collect ChIP material, and further purification was performed with Geneaid Gel/PCR cleanup protocol. Purified DNA fragments passed to sequencing library preparation.
Up to 5ng of isolated genomic DNA were re-sonicated to generate fragment ~100bp. Libraries were prepared using and following the NEBNext Ultra II DNA Library Prep with Sample Purification Beads protocol.

treatment_protocol: Cells were differenitated to EVT for 3 days before collection. Protocol: TSC were converted to EVT using a modified TSC basal media containing DMEM F-12 (GIBCO), 1x ITS-X (GIBCO), 0.3% BSA (WISENT), 1% Penicillin/Streptomyocin, and 0.1mM β-mercaptoethanol (GIBCO). For day 1-2, TSC were passage onto 1ug/ml Collagen IV coated plates in mod. TSC basal with added 5% Knock-serum replacement (GIBCO), 5uM Y27632 (CaymenChem), 3uM A8301 (CaymenChem), 100ng/ml NRG1 (NEB) and 2% GFR-Matrigel (Corning) while the media is still cold. Day 3-5, the media is change to the previously mention ETVM but this time without NRG1 and reduction to 0.5% GFR-Matrigel. Day6-8, the media is change to the previously mention ETVM but this time without NRG1, KSR, and reduction to 0.5% GFR-Matrigel. At the end of day 8, differentiated EVT can be assess by Flowcytometry.

tag: cell type

tag: chip antibody


title: WT-5d-H3K27me3-R1


title: 56-3

description: 10x Genomics


title: pkl tfl2-2-14d-K27-R2


title: CT3-EVTd3_INPUT_rep1


title: 56-2

description: 10x Genomics


title: pkl tfl2-2-14d-K27-R1


title: 28310

description: TERV

value: Belgian Tervuren


title: 56-1

description: 10x Genomics

value: brain


title: CT3-EVTd3_GCM1_rep2


title: pkl swn-14d-K27-R2


title: CT3-EVTd3_GCM1_rep1


title: 28309

description: BELS

value: Belgian Sheepdog


title: 54-6

description: 10x Genomics


title: pkl swn-14d-K27-R1


title: 28308

description: TERV


title: 54-5

description: 10x Genomics


title: pkl clf-14d-K27-R2


title: 54-3

description: 10x Genomics


title: pkl clf-14d-K27-R1


title: 54-2

description: 10x Genomics


title: 28277

description: BELS


title: 28316

description: TERV


title: 44-6

description: 10x Genomics

value: CTRL


title: tfl2-2-14d-K27-R1


title: 28301

description: TERV


title: tfl2-2-14d-K27-R2


title: STB-rep2

data_processing:
For RNA and ChIP/ATAC sequencing analysis, Genpipes 4.3.2 (https://bitbucket.org/mugqic/genpipes) provided the pipeline for basic sequencing processing.  In brief for RNAseq, sequencing quality and adaptor removal was trimmed with trimmomatic (v0.36). Trimmed fastq files alignment was performed with STAR aligner (v2.7.8a) using hg38/GRCh38 (ensembl v104). Picard (v2.9.0) was then used merged, mark duplicates, identify unique read, and sort .bam files proceeding alignment. Read counts were collected using HTseq-count and StringTie (v1.3.5). Differentially expressed gene (DEG) comparison was performed with DESeq2 package on RStudio (R v4.3.1). Correlation matrix, hierarchal gene cluster analysis, and PCA were generated in R. Gene pathway analysis were performed on ConsensusPathwayDB and EnrichR. For ChIPSeq and ATACseq, in brief, raw fastq files trimmed using trimmomatic (v.0.36). Then qualified fastq reads were mapped with BWA (v.0.7.17) with post processing with sambamda (v0.8.1) to merge replicates, mark and filter duplicates, and remove blacklist regions. Peak calling and differential-bind was performed with MACS2 (v2.2.7.1). Gene annotations and motif analysis used with Homer (v4.11). Bam to bigwig bamCoverage from deepTools (v3.5.1) was used to generate the tracks for viewing on IGV (v2.9.4).
Assembly: hg38
Supplementary files format and content: tab-delimited text includes peakCalls for each sample of the merged replicates

extract_protocol:
ATAC-seq library preparation was performed on 1 × 106freshly cultured cells using a commercially available ATAC-Seq kit from Active Motif (#53150, Carlsbad, CA). We followed manufacturers protocol with some minor modification. Each sample was lysed in the ATAC-seq lysis buffer. Next, the samples were processed for the transposase reaction. After cleanup of the transposed DNA, samples were stored at −20 °C until library amplification.
Samples were subsequently thawed at room temperature and library construction completed according to manufacturer protocol. Libraries were quantified using a Qubit dsDNA BR Assay Kit (Q32853, Thermo-Fisher) and the size was determined with a High Sensitivity DNA Bioanalyzer Kit (5067-4626, Agilent, Santa Clara, CA) and sequenced on a NovaSeq 6000 (Illumina, San Diego, CA) using Nextera Sequencing primers.

value: WT

value: STB


title: EVT-rep3

value: EVT


title: 28072

description: FCR-

value: Flat-Coated Retriever


title: CT1-EVT-non-target_c2


title: 28040

description: FCR-


title: 44-2

description: 10x Genomics


title: EVT-rep1


title: swn-14d-K27-R2

value: whole seedling


title: 44-9

description: 10x Genomics


title: EVT-rep2

value: Trophoblast Stem Cell


title: 27923

description: NOWT

value: Norwich Terrier


title: STB-rep3


title: swn-14d-K27-R1


title: 27912

description: BMD-

value: Bernese Mountain Dog


title: 44-1

description: 10x Genomics


title: hTSC-rep2

value: TSC


title: clf-14d-K27-R2


title: STB-rep1


title: hCMECD3_WSE50_S48_Batch3

city: Livermore

first: Aimy

last: Sebastian

state: CA

address: 7000 East Ave

department: BBTD

institute: LLNL

data_processing:
Sequence reads were aligned against human reference genome (hg38) using STAR.  After read alignment, the number of reads per gene locus was summarized with ‘featureCounts’ from Rsubread package, using hg38 gene annotation.
Assembly: hg38
Supplementary files format and content: Tab-delimited text file includes read counts for each sample .

source_name: immortalized human cerebral microvascular endothelial cell line (hCMEC/D3)

extract_protocol:
Co-cultures were treated with with eucalyptus wood smoke extract (10,30,50 mg/mL or 20,70, 100 mg/mL), lipopolysaccharide (LPS) (1 mg/mL), or LiCl (10 mM) for 24 hours. Cells were fixed with 4% paraformaldehyde (PFA) for ICC, lysed to collect mRNA for bulk-RNA sequencing, or supernatants were collected for ELISA and cytotoxicity assays. Total RNA was extracted from cell lysed in Qiagen Buffer RLT containing β-mercaptoethanol followed by RNA isolation using RNAeasy mini spin columns (Qiagen)
Sequencing libraries were preparedusingIlluminaStranded mRNA prep, Ligation kit (Illumina)

value: eucalyptus wood smoke extract 50 ug/mL


title: 27853

description: GSD-

value: German Shepherd Dog


title: 27843

description: GSD-


title: clf-14d-K27-R1


title: pkl-14d-K27-R2


title: hCMECD3_WSE50_S47_Batch3

treatment_protocol: Co-cultures were treated with with eucalyptus wood smoke extract (10,30,50 mg/mL or 20,70, 100 mg/mL)

value: immortalized human cerebral microvascular endothelial cell line (hCMEC/D3)


title: 27836

description: GSD-


title: hCMECD3_WSE30_S46_Batch3

value: eucalyptus wood smoke extract 30 ug/mL


title: hTSC-rep1


title: Senescent IMR90 shATG7 dox+ rep2

city: Seoul

first: Jaejin

last: Kim

address: Gwanak-ro 1

country: South Korea

institute: Seoul National University

data_processing:
The sequence reads were trimmed using Trimmomatic version 0.36 and mapped against the human reference genome (hg38) using Hisat2 version 2.1.0.
Read counts per gene were calculated by using featureCounts version 1.6.3.
Assembly: hg38
Supplementary files format and content: tab-delimited text files include raw counts for each sample

source_name: Lung

growth_protocol: IMR90 cells were maintained in 3% O2 and cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 15% fetal bovine serum (FBS), penicillin/streptomycin, and 0.1 mM nonessential amino acids.

extract_protocol:
Total RNAs were isolated using the acid phenol extraction method.
One microgram of high-quality RNA samples (RIN > 7.0) was used to construct RNA-seq libraries using the TruSeq Stranded mRNA LT Sample Prep Kit (Illumina). Total RNA integrity was checked using an Agilent 2200 TapeStation. RNA-seq was performed on an Illumina NovaSeq 6000 sequencer at Macrogen.

treatment_protocol: Senescent cells were assessed at 10 days after exposure to bleomycin (10 ug/ml for 24 h). Doxycycline (1ug/ml) were treated for 10 days to express shRNA targeting ATG7 or ATG13.

value: Lung

value: Fibroblast

value: hTERT pinducer10 RFP X shATG7

value: 10 days after exposure to bleomycin (10 ug/ml for 24 h), Doxycycline for the last 10 days (1ug/ml)


title: pkl-14d-K27-R1


title: 27834

description: GSD-


title: Senescent IMR90 shATG7 dox+ rep1


title: hCMECD3_WSE30_S45_Batch3


title: Senescent IMR90 shATG7 dox- rep1

value: 10 days after exposure to bleomycin (10 ug/ml for 24 h)


title: hCMECD3_WSE10_S43_Batch3

value: eucalyptus wood smoke extract 10 ug/mL


title: hTSC-rep3

value: WT


title: 27789

description: PHAR

value: Pharaoh Hound


title: PG-R2

value: whole seedling


title: hCMECD3_Control_S42_Batch3

value: eucalyptus wood smoke extract 0 ug/mL


title: WT-14d-K27-R1


title: Senescent IMR90 shATG7 dox- rep2


title: 27788

description: PHAR


title: WT-14d-K27-R2


title: Proliferating IMR90 shATG7 dox+ rep2

value: Doxycycline for 10 days (1ug/ml)


title: PG-R1


title: Proliferating IMR90 shATG7 dox+ rep1


title: hCMECD3_Control_S41_Batch3


title: Proliferating IMR90 shATG7 dox- rep2

value: untreated


title: 27575

description: SKYE

value: Skye Terrier


title: 27795

description: GSD-


title: Proliferating IMR90 shATG7 dox- rep1


title: hCMECD3_WSE50_S40_Batch2

value: eucalyptus wood smoke extract 50 ug/mL


title: PG swn-R2


title: hCMECD3_WSE10_S44_Batch3


title: Senescent IMR90 shATG13 dox+ rep2

value: hTERT pinducer10 RFP X shATG13


title: PG swn-R1


title: 27193

description: CNCS

value: Cane Corso


title: hCMECD3_WSE30_S38_Batch2

value: immortalized human cerebral microvascular endothelial cell line (hCMEC/D3)

value: eucalyptus wood smoke extract 30 ug/mL


title: 27482

description: IWSP

value: Irish Water Spaniel


title: Senescent IMR90 shATG13 dox+ rep1

value: Lung

value: Fibroblast


title: PG clf-R2


title: 27119

description: EFOX

value: English Foxhound


title: hCMECD3_WSE30_S37_Batch2


title: PG clf-R1


title: Senescent IMR90 shATG13 dox- rep2

value: 10 days after exposure to bleomycin (10 ug/ml for 24 h)


title: ATAC-pkl clf-#3

extract_protocol:
Lysates were clarified from sonicated protein-DNA complexes were isolated with GFP antibody.
The purified nuclei were incubated with Tn5 transposome and tagmentation buffer at 37℃ for 30 min (Vazyme Biotech, Cat No./ID: 501-02). After the tagmentation, the DNA was purified using a Qiagen MinElute PCR Purification Kit (Qiagen, Cat No./ID: 28004) and then amplified for 8-10 cycles. Amplified libraries were purified with VAHTS DNA Clean Beads (Vazyme, cat. no. N411-02) VAHTSTM DNA Adapters set3 - set6 for Illumina® (N805) and VAHTS® DNA Clean Beads (N411-02, Vazyme) according to the manufacturer’s protocol. protocol. Illumina Novaseq was used for high-throughput sequencing of the ChIP-seq library.

value: pkl clf

tag: developmental stage

tag: treatment of_tn-5

value: Vazyme Cat# TD501-02


title: hCMECD3_WSE10_D36_Batch2

value: eucalyptus wood smoke extract 10 ug/mL


title: Senescent IMR90 shATG13 dox- rep1


title: 27072

description: CNCS


title: 27778

description: PHAR

value: Pharaoh Hound


title: ATAC-tfl2-#3

value: whole seedling


title: hCMECD3_WSE10_S35_Batch2


title: 27011

description: WSSP

value: Welsh Springer Spaniel


title: CT1-TSC-non-target_c2

value: Trophoblast Stem Cell

value: WT

value: TSC


title: ATAC-clf-#3

value: clf


title: hCMECD3_WSE50_S39_Batch2

value: immortalized human cerebral microvascular endothelial cell line (hCMEC/D3)

value: eucalyptus wood smoke extract 50 ug/mL


title: hCMECD3_Control_S34_Batch2

value: eucalyptus wood smoke extract 0 ug/mL


title: Proliferating IMR90 shATG13 dox+ rep2

value: Lung

value: hTERT pinducer10 RFP X shATG13

value: Doxycycline for 10 days (1ug/ml)


title: ATAC-pkl-#3

value: pkl


title: 26987

description: TREE

value: Treeing Walker Coonhound


title: Proliferating IMR90 shATG13 dox+ rep1

value: Fibroblast


title: hCMECD3_Control_S33_Batch2


title: ATAC-WT-#3

value: Vazyme Cat# TD501-02


title: 26902

description: IWSP

value: Irish Water Spaniel


title: ATAC-PG-dCHROMO-R2-#2

value: whole seedling

value: PG dCHROMO


title: hCMECD3_WSE50_S32_Batch1


title: 1152 CP-

value: brain

value: CSS

value: low

value: choroid plexus

value: CP


title: hCMECD3_WSE50_S31_Batch1

value: immortalized human cerebral microvascular endothelial cell line (hCMEC/D3)

value: eucalyptus wood smoke extract 50 ug/mL


title: Proliferating IMR90 shATG13 dox- rep2

value: Lung

value: untreated


title: Proliferating IMR90 shATG13 dox- rep1

value: hTERT pinducer10 RFP X shATG13


title: ATAC-PG-dCHROMO-R1-#2


title: 25632

description: FCR-

value: Flat-Coated Retriever


title: 26830

description: IWSP


title: OCI-AML22_ATAC

city: Toronto

middle: James

last: Murison

state: Ontario

address: 101 College Street, TMDT room 11-706

institute: University Health Network

description:
filtered_feature_bc_matrix.h5
atac_fragments_post_doublets.tsv

library_source: genomic single cell

data_processing:
Cellranger-Arc used to align raw data with default settings
Amulet and Doublet Finder used to filter likely doublets
Cells filtered for QC as described in Boutzen et al, 2024
Assembly: hg38
Supplementary files format and content: gex_molecule_info.h5: Per molecule information from cell-ranger
Supplementary files format and content: per_barcode_metrics.csv: metrics for every observed barcode

growth_protocol: cells were expanded in culture in the following medium, with final concentrations as indicated : X-VIVO 10 (Lonza, BE04-380Q) supplemented with 20% BIT 9500 Serum Substitute (StemCell technologies, 09500), 1x Glutamax Supplement (Thermo Fisher Scientific, 35050061), Primocin 0.1 mg/ml (invivogen), SCF (200 ng/ml;Miltenyi Biotech, 130-096-696), IL3 (20 ng/ml;Miltenyi Biotech, 130-095-069), TPO (20 ng/ml; Peprotech, 300-18), FLT3L (40 ng/ml; Peprotech, 300-19 B), IL6 (10 ng/ml; Miltenyi Biotech, 130-093-934), G-CSF (10 ng/ml; Miltenyi Biotech, 130-093-861). Cells were maintained at a density of 0.8x106 cells/ml and passaged every 3-5 days, in a 96 well flat bottom plate. The CD34+CD38- fraction or CD34+ fraction was regularly sorted to serially expand the cells.

extract_protocol:
CD34+CD38- OCI-AML22 flow sorter cells were washed in PBS + %0.04 BSA and then prepared for the 10x multiome assay with a 4 min incubation in lysis buffer (10 mM Tris-HCl (pH 7.4), 10 mM NaCl, 3 mM MgCl2, 1% BSA, 0.1% Tween-20, 1 mM DTT, 1 U/µl RNase inhibitor) as perthe Nuclei Isolation for Single Cell Multiome ATAC + Gene Expression Sequencingprotocol from 10x genomics (protocol 1).
Lysed cells were then transposed and partitioned into single nuclei droplets usingthe 10x chromium Next GEM (protocol 2).Chip J before undergoing library preparation as outlined in theChromium Next GEM Single Cell Multiome ATAC + Gene Expressionprotocol from 10x genomics
10x Multiome. The molarity of each library was calculated based on library size as measured bioanalyzer (Agilent Technologies) and qPCR amplification data (Roche, cat. no. 07960140001). ATAC and Gene Expression libraries were sequenced separately on Illumina’s NovaSeq 6000. The RNA portion was sequenced on a S1 flow cell with the following configuration: 28-10-10-90 (Read 1-Index 1-Index 2- Read 2) and the ATAC portion was sequenced on a SP-XP flow cell with the following configuration: 50-8-24-50. Sequencing was performed at the Princess Margaret Genomics Centre, Toronto, Canada (www.pmgenomics.ca).

treatment_protocol: no treatment performed

value: human primary AML cell derived OCI-AML22


title: ATAC-PG-dSNF-R2-#2

value: PG dSNF


title: 25382

description: ESHP

value: English Shepherd


title: OCI-AML22_RNA

molecule: polyA RNA


title: hCMECD3_WSE30_S29_Batch1

value: eucalyptus wood smoke extract 30 ug/mL


title: hCMECD3_WSE30_S30_Batch1


title: hCMECD3_WSE10_S28_Batch1

value: eucalyptus wood smoke extract 10 ug/mL


title: 24956

description: CHIN

value: Japanese Chin


title: ATAC-PG-dSNF-R1-#2

value: whole seedling

value: Vazyme Cat# TD501-02


title: ATAC-PG-R2-#2

value: PG


title: Antibody

city: Leiden

first: Jessica

last: Roelands

address: Albinusdreef 2

country: Netherlands

institute: Leiden University Medical Center

library_source: other

data_processing:
Fastq files were generated using Cell Ranger mkfastq (10x Genomics)
These fastq files were analyzed using the 10x Genomics Cell Ranger software version 7.0.0 and a custom reference with GRCh38-2020 containing the tdTomato gene.
Assembly: Custom reference with GRCh38-2020 containing the tdTomato gene
Supplementary files format and content: Tab-separated values files and matrix files

molecule: other

source_name: Bone marrow

extract_protocol:
Bone marrow was harvested from euthanized mice on day 2 (n=4/group), day 5 (n=3/group), and day 8 (n=3/group). Femurs were cleaned of surrounding tissue and cut open on the knee-side. The open femurs were placed into a 1.5ml Eppendorf tube containing 100ul of T cell medium and spun down at 2500g at room temperature. Bone marrow suspension was filtered through a cell strainer (70μM) (sysmex) into a sterile tube. Subsequently, bone marrow was spun down (500g) and resuspended in 500 μL red blood cell lysis buffer (XX mM ammonium chloride) for 10 minutes on ice. Lysed samples were spun and supernatant was removed. For each mouse, bone marrow from both paws was combined. Negative MACS enrichment of human cells from the bone marrow was conducted, utilizing anti-mCD45 (APC) antibodies and APC-beads. The enriched human cell population from each mouse was then labeled with unique hashtags (totalSeq-A human hashtags). Subsequently, to enhance sample purity, positive selection of human cells was performed using anti-hCD45 antibodies through sorting. Following these procedures, a single run of 10x Genomics 3’ v3.1 chemistry was executed for downstream analysis.
Single-cell gene expression libraries were prepared using the 10x Genomics Chromium X platform, specifically employing the Chromium Next GEM Single Cell 3’ Library & Gel Bead Kit v3.1 and Chromium Next GEM Chip G Single Cell Kit (10x Genomics) according to manufacturer's instructions.

value: Bone marrow


title: hCMECD3_Control_S26_Batch1

value: eucalyptus wood smoke extract 0 ug/mL


title: Sorted human cells from mouse bone marrow

value: CD45-positive cells


title: 26832

description: IWSP

value: Irish Water Spaniel


title: 24776

description: ROOS

value: Teddy Roosevelt Terrier


title: hsa-miR-487b-3p mimic, replicate 3

city: Columbus

first: Cankun

last: Wang

state: OH

address: 281 W Lane Ave, Columbus, OH 43210

institute: The Ohio State University Wexner Medical Center

data_processing:
Raw FASTQ reads derived from 24 samples underwent quality control and filtering using the Fastp package. Subsequently, these high-quality reads were aligned to the GENCODE GRCh38 reference genome. Quantification of gene expression was accomplished using the featureCounts software, which provided the count data for each gene.
Assembly: GRCh38
Supplementary files format and content: csv formatted files including the sample name, counts, and related metadata.

source_name: Cardiac

growth_protocol: Ye, et al., "Generation and Expansion of Human Cardiomyocytes from Patient Peripheral Blood Mononuclear Cells", JOVE, 2021

extract_protocol:
Qiagen miRNeasy Micro Kit
Novogene Library Construction

treatment_protocol: Lipofectamine RNAiMAX Trasfection Reagent Protocol

value: Cardiac

value: 273 Control (iPSC)

value: iPSC-CM

value: Normal

value: miR-487b mimic


title: ATAC-pkl-R2-#2

value: pkl


title: 24723

description: BULT

value: Bull Terrier


title: hsa-miR-487b-3p mimic, replicate 2


title: hCMECD3_WSE10_S27_Batch1

value: immortalized human cerebral microvascular endothelial cell line (hCMEC/D3)

value: eucalyptus wood smoke extract 10 ug/mL


title: hsa-miR-487b-3p mimic, replicate 1


title: pEC_WSE50_S24_Batch3

source_name: primary human brain microvascular endothelial cells (HBMEC)

value: eucalyptus wood smoke extract 50 ug/mL


title: hCMECD3_Control_S25_Batch1


title: 24712

description: BULT


title: hsa-miR-487b-3p inhibitor, replicate 3

value: miR-487b inhibitor


title: ATAC-WT-R2-#2

value: whole seedling

value: WT

value: Vazyme Cat# TD501-02


title: hsa-miR-487b-3p inhibitor, replicate 2


title: ATAC-pkl-R1-#2


title: 24709

description: BULT


title: pEC_WSE50_S23_Batch3

value: primary human brain microvascular endothelial cells (HBMEC)


title: ATAC-WT-R1-#2


title: pEC_WSE30_S22_Batch3

value: eucalyptus wood smoke extract 30 ug/mL


title: 24639

description: TERV

value: Belgian Tervuren


title: pEC_WSE30_S21_Batch3


title: 24777

description: ROOS

value: Teddy Roosevelt Terrier


title: hsa-miR-487b-3p inhibitor, replicate 1

value: Cardiac

value: 273 Control (iPSC)


title: 24325

description: BSJI

value: Basenji


title: ATAC-pkl-R1-#1


title: ATAC-PG-R1-#2

value: PG


title: pEC_WSE10_S20_Batch3


title: 24211

description: WIFX

value: Wire Fox Terrier


title: hsa-miR-433-3p mimic, replicate 3

value: iPSC-CM

value: Normal

value: miR-433 mimic


title: ATAC-WT-R2-#1


title: hsa-miR-433-3p mimic, replicate 2


title: pEC_WSE10_S19_Batch3

value: eucalyptus wood smoke extract 10 ug/mL


title: 24017

description: WHWT

value: West Highland White Terrier


title: ATAC-WT-R1-#1

value: whole seedling


title: hsa-miR-433-3p mimic, replicate 1


title: pEC_Control_S18_Batch3

value: eucalyptus wood smoke extract 0 ug/mL


title: PG lhp1-IP R2

tag: antibody

value: GFP (ab290, abcam)


title: hsa-miR-433-3p inhibitor, replicate 3

value: miR-433 inhibitor


title: 23691

description: BULT

value: Bull Terrier


title: 23731

description: BULT


title: pEC_Control_S17_Batch3

value: primary human brain microvascular endothelial cells (HBMEC)


title: hsa-miR-433-3p inhibitor, replicate 2


title: ATAC-pkl-R2-#1

value: pkl

value: Vazyme Cat# TD501-02


title: PG lhp1 INPUT


title: hsa-miR-433-3p inhibitor, replicate 1

value: Cardiac

value: 273 Control (iPSC)


title: pEC_WSE50_S16_Batch2

value: eucalyptus wood smoke extract 50 ug/mL


title: PG lhp1-IP R1


title: 23604

description: BULT


title: pEC_WSE50_S15_Batch2


title: pEC_WSE30_S14_Batch2

value: eucalyptus wood smoke extract 30 ug/mL


title: 23592

description: BULT


title: hsa-miR-134-5p mimic, replicate 2

value: iPSC-CM

value: miR-134 mimic


title: 23578

description: TERV

value: Belgian Tervuren


title: PG-IP R2


title: pEC_WSE30_S13_Batch2


title: PG-IP R1


title: PG INPUT


title: hsa-miR-134-5p mimic, replicate 3

value: Normal


title: pEC_WSE10_S12_Batch2

value: eucalyptus wood smoke extract 10 ug/mL


title: 23377

description: CHOW

value: Chow Chow


title: hsa-miR-134-5p inhibitor, replicate 3

value: miR-134 inhibitor


title: 23572

description: BELS

value: Belgian Sheepdog


title: hsa-miR-134-5p mimic, replicate 1


title: PKL-GFP R1

value: whole seedling

value: GFP (ab290, abcam)


title: 23671

description: BULT

value: Bull Terrier


title: pEC_WSE10_S11_Batch2

value: primary human brain microvascular endothelial cells (HBMEC)


title: pEC_Control_S10_Batch2

value: eucalyptus wood smoke extract 0 ug/mL


title: PKL-GFP R2


title: hsa-miR-134-5p inhibitor, replicate 1

value: Cardiac


title: PKL-GFP INPUT


title: Mock, Replicate 3

value: 273 Control (iPSC)

value: Buffer


title: 23115

description: TREE

value: Treeing Walker Coonhound


title: 22900

description: DARG

value: Dogo Argentino


title: PKLdSANTdSLIDE-K27 R1


title: PKLdSANTdSLIDE-K27 R2


title: pEC_Control_S9_Batch2


title: pEC_WSE50_S7_Batch1

value: eucalyptus wood smoke extract 50 ug/mL


title: Mock, Replicate 2

value: iPSC-CM


title: PKLdSNF-K27 R2


title: 22415

description: BMD-

value: Bernese Mountain Dog


title: pEC_WSE30_S6_Batch1

value: eucalyptus wood smoke extract 30 ug/mL


title: 22437

description: BMD-


title: Mock, Replicate 1

value: Normal


title: PKLdSNF-K27 R1


title: Scramble Control, Replicate 3

value: Scramble


title: PKLdCHROMO-K27 R2


title: Scramble Control, Replicate 2


title: pEC_WSE30_S5_Batch1


title: hsa-miR-134-5p inhibitor, replicate 2

value: miR-134 inhibitor


title: 22244

description: WELT

value: Welsh Terrier


title: PKLdCHROMO-K27 R1

value: whole seedling


title: pEC_WSE10_S4_Batch1

value: eucalyptus wood smoke extract 10 ug/mL


title: 22235

description: WELT


title: 22112

description: SCOT

value: Scottish Terrier


title: pEC_WSE50_S8_Batch1

value: primary human brain microvascular endothelial cells (HBMEC)


title: 22810

description: CHOW

value: Chow Chow


title: pEC_WSE10_S3_Batch1


title: PKLdPHD-K27 R2


title: Scramble Control, Replicate 1

value: Cardiac

value: 273 Control (iPSC)


title: 21511

description: BRUS

value: Brussels Griffon


title: KOLF2 cells, SETBP1_PATH3, Day 24, biol rep 3

city: Nedlands

first: Kevin

middle: Guangxing

last: Chen

address: Hospital Avenue

country: Australia

institute: The Kids Research Institute Australia

description: RawCountsMatrix.tsv

data_processing:
Raw sequencing reads were processed using the ENCODE/DCC pipeline (https://github.com/ENCODE-DCC/rna-seq-pipeline)
Reads were aligned to GRCh38 and quantified using kallisto.
Kallisto quantificiation files were read into edgeR/limma using the tximport package in R to generate counts matrices
Amplicon sequencing: reads were aligned to either WT or HDR amplicon using CRISPResso2
Assembly: GRCh38
Supplementary files format and content: RawCountsMatrix.tsv contains the Kallisto-quantified counts of all genes for each sample

source_name: KOLF2 derived NPCs

growth_protocol: KOLF2 cells were grown on Matrigel-coated plates in TeSR-E8 media. The STEMdiff SMADi Neural Induction Kit (STEMCELL technologies) were used to induce differentiation into NPCs.

extract_protocol:
Cells were collected on day 0 and 24 for RNA extraction using RNeasy (Thermo Fisher). gDNA for amplicon sequencing was extracted using PureLinkTM Genomic DNA Mini Kit (Life Technologies)
RNA integrity was determined using the Agilent TapeStation system (Genomics WA). RNA sequencing was performed according to SureSelect Strand-Specific RNA Library preparation for Illumina Multiplexed sequencing, paired-end, 100 bp, 30 M read RNA sequencing on the NOVAseq 6000 platform (Illumina, USA) at Genomics WA (Perth, Australia). PCR amplicon products corresponding to the SNV sites in SETBP1, PATH2, PATH3 and VUS2, were prepared from extracted gDNA with forward and reverse amplicon primers and Phusion High-Fidelity PCR Master Mix with HF Buffer (Thermo Scientific). Purified samples were barcoded with TruSeq primers (IDT) and SETBP1 sequences determined with paired-end, 250bp DNA amplicon sequencing with a MiniSeq Mid-Output Kit, 300 cycles.

value: KOLF2 derived NPCs

value: neural progenitor cells


title: 21455

description: FCR-

value: Flat-Coated Retriever


title: KOLF2 cells, SETBP1_PATH3, Day 24, biol rep 2

description: RawCountsMatrix.tsv


title: pEC_Control_S2_Batch1

value: eucalyptus wood smoke extract 0 ug/mL


title: PKLdPHD-K27 R1


title: 21453

description: FCR-


title: NCI-H226-HC31278-A-6

city: Stony Brook

first: Jyoti

last: Misra

address: 101 Nicolls Rd

institute: Stony Brook University

data_processing:
CLC Genomics Workbench (CLC Bio,v23.0.3)
Sequence reads were trimmed for adapter sequences using CLC Genomics work bench
Sequence reads were Mapped to HG38 using CLC genomics Work bench
Read count extraction and normalization were perfromed using CLC Genomics workbench
Assembly: ENSEMBL v99 hg38
Supplementary files format and content: Total counts, RPKM, TPM, CPM

source_name: Pleura

growth_protocol: NCI-H226 cells were maintained in DMEM supplemented with 10% FBS.

extract_protocol:
RNA was extracted using trizol reagent
RNA sequencing library was prepared using Illumina Stranded mRNA prep kit (20040532) according to Manufacturer’s protocol.

treatment_protocol: NCI-H226 cells were treated with either dmso or 500nM HC31278A  for 24 hours

value: Pleura

value: MESOTHELIOMA


title: PG-K27 R1


title: KOLF2 cells, SETBP1_PATH3, Day 0, biol rep 3

description: RawCountsMatrix.tsv

source_name: KOLF2, Day 0

value: KOLF2, Day 0

value: induced pluripotent stem cells


title: NCI-H226-HC31278-A-2


title: 21437

description: SCOT


title: pkl-K27 R2


title: NCI-H226-HC31278-A-1


title: KOLF2 cells, SETBP1_PATH3, Day 0, biol rep 1

description: RawCountsMatrix.tsv


title: pkl-K27 R1


title: NCI-H226-DMSO-3

value: DMSO


title: 20637

description: WHWT

value: West Highland White Terrier


title: NCI-H226-DMSO-2


title: WT-K27 R1

value: whole seedling


title: KOLF2 cells, SETBP1_PATH2, Day 24, biol rep 2

description: RawCountsMatrix.tsv


title: pEC_Control_S1_Batch1


title: KOLF2 cells, SETBP1_PATH3, Day 24, biol rep 1

description: RawCountsMatrix.tsv


title: NCI-H226-DMSO-1


title: 20136

description: GOLD

value: Golden Retriever


title: KOLF2 cells, SETBP1_WT, Day 24, biol rep 6

description: RawCountsMatrix.tsv

value: WT


title: patient I, post NACT

city: Birmingham

first: Paul

last: Moss

state: West Midlands

address: Vincent Drive

department: Immunology and Immunotherapy

country: United Kingdom

institute: University of Birmingham

description: Nextera DNA library preparation

data_processing:
Reads were processed into FASTQ format and peak-barcode counts using Cell Ranger ATAC v2.0.0, aligning to the prebuilt Cell Ranger human reference genome GRCh38-2020-A-2.0.0.
The counts matrix for each sample was read into R and stored as Seurat objects for downstream analysis with Seurat v4.1 and Signac v1.6.0 R packages.
Assembly: GRCh38
Supplementary files format and content: read fragments tsv file

source_name: HGSOC Omentum

growth_protocol: Tumour biopsies were obtained following the surgical resection of omental tissue and were immediately snap frozen in liquid nitrogen

extract_protocol:
Each tissue specimen was minced using a scalpel into 1-2mm sections. These were transferred with 1ml of the 0.1X lysis buffer solution into a 7ml Dounce tissue homogeniser on ice. The tissue was pushed to the base of the homogeniser and compressed with the larger pestle A for 50 strokes. This was then repeated with pestle B and the sample was transferred into a sterile 15ml Conical tube. Following incubation on ice for 20 minutes, the sample was centrifuged at 4°C for 5 minutes at 500xG. The pellet was resuspended in 2ml of new 0.1X lysis buffer solution and incubated on ice for a further 20 minutes. The sample was filtured using a 40-micron cell strainer and was centigured again at 4°C for 5 minutes at 500xG. A wash buffer was formulated by combining 3.5ml of nuclease-free water, 40µl of Tris-HCl, 8µl of NaCl, 12µl of MgCl2, 400µl of BSA and 40µl of Tween-20. The cell pellet was resuspended in 1ml of this wash buffer, before being filtering with a 40-micron cell strainer and transferred into a sterile 1.5ml Eppendorf tube. Individual nuclei within the solution were counted and viability assessed using the Countess II FL automated cell counter (ThermoFisher Scientific). Diluted nuclei buffer (10X Genomics Single Cell ATAC Library and Gel Bead Kit; 1000175) was added to achieve a concentration of 2000 nuclei per µl, with a targeted nuclei recovery of 4000 nuclei
Following nuclei extraction, single-nuclei solutions were processed by the 10X Genomics Sequencing laboratory at Genomics Birmingham. The Chromium Single Cell ATAC Solution and the Chromium Next GEM Single Cell ATAC Library and Gel Bead Kit (10X Genomics) were utilised according to the manufacturer’s instructions. Briefly, diluted nuclei were added to a transposition solution, where transposase entered nuclei and fragmented the DNA in open chromatin regions, while adapter sequences were merged onto the DNA fragments. Individual transposed nuclei were joined with a barcoded gel bead and partitioning oil to create a GEM. By dissolving gel beads, primers and 10x barcodes were added to single-stranded DNA, and the GEM’s fragmented to collect the DNA. All transposed DNA fractions from the same nucleus were tagged with individual barcodes to identify the nucleus of origin after DNA pooling.

value: HGSOC Omentum

value: mixed

value: NACT


title: PG-K27 R2


title: pkl-K27me3 R2


title: 16133

description: WHWT

extract_protocol: Genomic DNA was extracted from whole blood or Buccal cells using Proteinase K and phenol/chloroform with standard protocols. Genomic DNA extracted from saliva using standard Performagene genotek DNA extraction protocols.


title: patient H, post NACT

description: Nextera DNA library preparation


title: pkl-K27me3 INPUT


title: 16129

description: SSHP

value: Shetland Sheepdog


title: KOLF2 cells, SETBP1_PATH2, Day 24, biol rep 1

description: RawCountsMatrix.tsv

value: KOLF2 derived NPCs


title: KOLF2 cells, SETBP1_PATH3, Day 0, biol rep 2

description: RawCountsMatrix.tsv


title: KOLF2 cells, SETBP1_PATH2, Day 0, biol rep 3

description: RawCountsMatrix.tsv


title: patient G, post NACT

description: Nextera DNA library preparation


title: 21193

description: SCOT

value: Scottish Terrier


title: 15625

description: CHOW

value: Chow Chow


title: WT-K27me3 R2


title: WT-K27 R2


title: KOLF2 cells, SETBP1_PATH2, Day 24, biol rep 3

description: RawCountsMatrix.tsv

value: neural progenitor cells


title: KOLF2 cells, SETBP1_PATH2, Day 0, biol rep 2

description: RawCountsMatrix.tsv


title: patient F, post NACT

description: Nextera DNA library preparation


title: 15087

description: SCOT


title: WT-K27me3 R1


title: 20104

description: LHSA

value: Lhasa Apso


title: 14566

description: BULD

value: Bulldog


title: KOLF2 cells, SETBP1_WT, Day 0, biol rep 6

description: RawCountsMatrix.tsv

value: KOLF2, Day 0


title: pkl-K27me3 R1


title: KOLF2 cells, SETBP1_PATH2, Day 0, biol rep 1

description: RawCountsMatrix.tsv

value: induced pluripotent stem cells


title: WT-K27me3-INPUT


title: patient E, treatment-naïve

description: Nextera DNA library preparation

value: treatment-naive


title: KOLF2 cells, SETBP1_WT, Day 24, biol rep 5

description: RawCountsMatrix.tsv


title: 13819

description: CHOW


title: 13811

description: CHOW


title: patient C, treatment-naïve

description: Nextera DNA library preparation


title: patient D, treatment-naïve

description: Nextera DNA library preparation


title: 13797

description: CHOW


title: KOLF2 cells, SETBP1_WT, Day 24, biol rep 4

description: RawCountsMatrix.tsv


title: patient B, treatment-naïve

description: Nextera DNA library preparation


title: KOLF2 cells, SETBP1_WT, Day 0, biol rep 5

description: RawCountsMatrix.tsv


title: KOLF2 cells, SETBP1_WT, Day 0, biol rep 4

description: RawCountsMatrix.tsv


title: KOLF2 cells, SETBP1_WT, Day 24, biol rep 3

description: RawCountsMatrix.tsv


title: patient A, treatment-naïve

description: Nextera DNA library preparation


title: 13767

description: CHOW


title: OL_KO_2_FPKM

city: Cincinnati

first: Richard

last: Lu

state: Ohio

institute: CINCINNATI CHILDREN'S HOSPITAL MEDICAL CENTER

data_processing:
RNA-Seq data were aligned to mouse genome using TopHat with default settings.
Assembly: mm10
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

extract_protocol:
Total RNA from mouse OLs were extracted using RNeasy Micro Kit (Qiagen)
RNA-seq libraries were prepared using Illumina RNA-Seq Preparation Kit

value: brain

value: mouse OL

value: Zfp488 KO


title: 13682

description: CHOW

data_processing:
Image data was analyzed using GenomeStudio with GT module (Illumina).
genotypes AA, AB, BB, NC=no call


title: KOLF2 cells, SETBP1_WT, Day 24, biol rep 2

description: RawCountsMatrix.tsv

value: WT


title: OL_KO_1_FPKM


title: patient A, post NACT

description: Nextera DNA library preparation

value: HGSOC Omentum


title: KOLF2 cells, SETBP1_WT, Day 24, biol rep 1

description: RawCountsMatrix.tsv


title: OL_2_FPKM


title: KOLF2 cells, SETBP1_WT, Day 0, biol rep 3

description: RawCountsMatrix.tsv


title: OL_1_FPKM


title: 12999

description: LAB-

value: Labrador Retriever


title: 12994

description: LAB-


title: 12995

description: LAB-


title: SmartSeq2 scRNA-seq, KRN

city: Barcelona

first: Josep

last: Garnica

address: Rosselló, 149

country: Spain

institute: IDIBAPS

data_processing:
Aligment - STAR, Bowtie2, Cellranger v6. TCR reconstriuction - TraCeR
PeakCalling - MACS2
Downstream - DiffBind, Seurat, Signac, GenomicRanges, Trackviewer…
Assembly: GRCm38.p6
Supplementary files format and content: Peakset; differential peaks called
Supplementary files format and content: feature count matrices

source_name: Liver

extract_protocol:
Cells were lysed in cold lysis buffer (10 mM Tris-HCl, pH 7.4,10 mM NaCl, 3 mM MgCl2, and 0.1% IGEPAL CA-630), washed, and right after resuspended in transposase reaction mix.  10X single-cell RNA-seq: Alive cells were collected from untreated and aGalCer/CD1d-NP-treated 13-15 wk-old NOD mice in DMEM media (Sigma-Aldrich) supplemented with 10% FBS (Hyclone) at 4 ºC, separated into nanoscale gel beads emulsions with a 10X barcode. Cell numbers and viability were assessed using a TC20™ Automated Cell Counter (Bio-Rad Laboratories, Hercules, CA, USA), with a minimum target of 4000 cells. Later, cDNA sequencing libraries were produced using the NextGEM Single-cell 3’ mRNA kit (v3.1; 10X Genomics) following the manufacturer's instructions. These steps involved GEM-RT clean-up, cDNA Amplification for 13 cycles, and cDNA quality control and quantification using the Agilent Bioanalyzer High Sensitivity chip (Agilent Technologies). Libraries were indexed by PCR using the PN-220103 Chromium i7 Sample Index Plate. Finally, sequencing was carried out on a NovaSeq 6000 sequencer (Illumina). 10X single-cell multiome (scRNAseq+scATACseq). For 10X multiome RNA-seq+ATAC-seq, LiNKT cells were sorted from untreated (n=5; 13-15wk-old) and αGalCer/CD1d-NP-treated male NOD mice (n=4; 13-15wk-old), to obtain a total of 300.000 and 250.000 live LiNKT cells in DMEM media (Sigma-Aldrich) supplemented with 10% FBS (Hyclone). Cells were lysed at 4 ºC, and nuclei isolated. Nuclei were used for transposition of adapter sequences and processed for single-cell barcoding and library generation following the manufacturer’s instructions (CG000338; 10X Genomics). Briefly, isolated nuclei were partitioned into Gel Bead-In-Emulsions to produce barcoded cDNA from poly-adenylated mRNA as described above, as well as barcoded DNA fragments, and processed for library amplification and sequencing on a NovaSeq 6000 sequencer (Illumina), also as described above.  Smartseq2 scRNAseq: One-cell sorting of LiNKTs from untreated and αGalCer/CD1d-NP-treated mice (n=3 male mice/group; 13-15 wk-old) was performed in SMARTseq2 96-well plates (PerkinElmer, Waltham, MA, USA) containing 100 µl lysis buffer/well, consisting in 0.2% v/v Triton-100 with RNase inhibitor. Full-length single-cell RNA sequencing libraries were prepared using a modified Smart-seq protocol 64. A reverse transcription reaction was done in presence of oligo-dT30VN, betaine and template-switching oligonucleotides. Complementary DNA was then amplified with KAPA Hifi Hotsart RadyMix (Kappa Biosystems) and ISPCR primer in 25 cycles. The PCR product was purified with Agencourt Ampure XP beads (Beckman Coulter) and analyzed with a High Sensitivity DNA kit (Agilent Technologies). 200 pg of the cDNA product were fragmented with Nextera® XT (Illumina) and amplified with indexed Nextera® PCR primers. Products were purified twice with Agencourt AMPure XP beads and quantified again using a Bioanalyzer High Sensitivity DNA Kit. Sequencing of Nextera® libraries from 384 cells was carried out using one sequencing lane on an Illumina HiSeq2500 v4 or HiSeq4000 to 500K reads/cell.  ATACseq:  ATAC-seq was done on LiNKT cells isolated from untreated and αGalCer/CD1d-NP-treated mice (n=3 male mice/group; 13-15 wk-old). 7x104 cells were sorted in PBS and processed for library preparation as described 27. Briefly, cells were lysed in cold lysis buffer (10 mM Tris-HCl, pH 7.4,10 mM NaCl, 3 mM MgCl2, and 0.1% IGEPAL CA-630) to isolate nuclei. The purified nuclei were washed, resuspended in the transposase reaction mix (25 μl 2x TD buffer, 2.5 μl transposase (Illumina) and 22.5 μl nuclease-free water) and incubated for 30 min at 37°C. DNA was purified with MinElute Reaction Cleanup kit (Qiagen). Libraries were generated with NEBNext® High Fidelity PCR kit (New England Biolabs, Ipswich, MA, USA) using 1x NEB Next PCR master mix (New England BioLabs) and 1.25 μM of custom Nextera PCR primers. Libraries were rendered using the barcoded primers Ad1_noMX as forward and Ad2.1-6 as reverse and purified using a PCR cleanup kit (Qiagen), yielding a final concentration of about 30 nM in 20 μl. Libraries were then analyzed on Bioanalyzer using an Agilent DNA High Sensitivity chip (Agilent Technologies, Santa Clara, CA, USA) to estimate the quantity and size distribution. Next, they were quantified by qPCR using the KAPA Library Quantification Kit before amplification with Illumina’s cBot. Libraries were finally loaded at 3.33 pM onto the flowcell and sequenced 1 x 50 on Illumina’s HiSeq 2500 to obtain 30-40 million reads/sample.  ChIP-seq: Chromatin immunoprecipitation (ChIP) and sequencing was performed on LiNKT cell samples from untreated and aGalCer/CD1d-NP-treated NOD mice (n=3/group; 13-15 wk-old) for H3K4me3, H3K27me, and H3K27Ac bound DNA via ChIP-seq following van Arensbergen’s protocols 65. LiNKT cells were sorted, fixed with 16% paraformaldehyde (PFA; Electron Microscopy Sciences, Hatfield, PA, USA) in DMEM (Sigma-Aldrich) supplemented with 10% FBS (Hyclone) and frozen at -80ºC until processed. We used 1-1.5x106 LiNKTs/sample. Cells were lysed, sheared, and sonicated using an S220 Focused-ultrasonicator (Covaris, Woburn, MA, USA) (13 min, 105 W, 2% Duty Factor, 200 cycles). This was followed by overnight incubation with the precipitating antibody: 0.5 µL of H3K4me3 (Sigma), 0.5 µL of H3K27me3 (Cell Signaling, Danvers, Massachusetts, USA), and 2 µL of H3K27Ac (Abcam, Cambridge, United Carlsbad, CA, USA) and precipitated using Protein-A-Dynabeads (Abcam). RNA was cleared using RNAse A (Qiagen) (1 hour at 65 ºC), and decrosslinking was performed overnight with proteinase K at 65 ºC. DNA was finally purified with Phenol-Chloroform and EtOH-precipitation. After quality control validation on a Bioanalyzer (Agilent Technologies), samples were sent for sequencing. Libraries were prepared using the NEB Next Ultra DNA Library Prep kit (Illumina) following the manufacturer’s protocol, analyzed with DNA High Sensitivity kit (Agilent Technologies), and quantified via qPCR using the KAPA Library Quantification Kit (Kapa Biosystems). Libraries were loaded at a concentration of 2.75 pM onto flowcells and were sequenced 1 x 50 on Illumina’s HiSeq 2500 to get 30-40 million reads/sample.  WGBS: Six replicates of LiNKTs (4-5x105 cells/sample) were obtained from untreated female NOD mice (n=7, 17wk-old) and αGalCer/CD1d-NP-treated NOD mice (n=4, 16wk-old) by cell sorting. Genomic DNA was extracted using the DNeasy Blood and Tissue kit (Qiagen) following the manufacturer’s instructions, and frozen at -20ºC. DNA was processed by Whole Genome Bisulfite Sequencing (WGBS). Briefly, DNA was sonicated to a mean size of 250 bp using a Bioruptor (Diagenode, Seraing (Ougrée), Belgium) followed by blunt-ending, dA addition to the 3'-end and ligation of methylated adaptors to protect from bisulfite conversion. Ligated DNA was bisulfite converted using the EZ DNA Methylation-Gold kit (ZYMO, Irvine, CA, USA). After treatment with sodium bisulfite, unmethylated cytosine residues were converted to uracil, leaving 5-methylcytosine (5mC) unaffected. Different insert size fragments were excised from the same lane of a 2% TAE agarose gel. Products were purified using a QIAquick PCR; after PCR amplification, uracil residues were converted to thymine. Sequencing was performed 2 x 150 bp using the NovaSeq 6000 System (Illumina).
Library fragments were amplified using 1x NEB Next PCR master mix (New England BioLabs) and 1.25 μM of custom Nextera PCR primers forward and reverse. Libraries were rendered using the barcoded primers Ad1_noMX as forward and Ad2.1-6 as reverse and purified using a PCR cleanup kit (Qiagen), yielding a final concentration of about 30 nM in 20 μL. Libraries were then analyzed on Bioanalyzer using an Agilent DNA High Sensitivity chip (Agilent Technologies, Santa Clara, CA, USA) to estimate the quantity and size distribution. Next, they were quantified by qPCR using the KAPA Library Quantification Kit before amplification with Illumina’s cBot.
Libraries were loaded at 3.33 pM onto the flowcell and sequenced 1 x 50 on Illumina’s HiSeq 2500

value: Liver

value: Liver iNKT (aGalCer/CD1d tetramer+)

tag: strain

value: NOD mouse


title: KOLF2 cells, SETBP1_WT, Day 0, biol rep 2

description: RawCountsMatrix.tsv


title: 13785

description: CHOW


title: SmartSeq2 scRNA-seq, CTL

value: untreated


title: KOLF2 cells, SETBP1_VUS2, Day 24, biol rep 4

description: RawCountsMatrix.tsv

value: KOLF2 derived NPCs


title: KOLF2 cells, SETBP1_VUS2, Day 24, biol rep 3

description: RawCountsMatrix.tsv

value: neural progenitor cells


title: KOLF2 cells, SETBP1_WT, Day 0, biol rep 1

description: RawCountsMatrix.tsv


title: 12744

description: GOLD

value: Golden Retriever


title: ChIPseq H3K4me3, KRN

value: H3K4me3 (Sigma)


title: 12777

description: CHIN

value: Japanese Chin


title: KOLF2 cells, SETBP1_VUS2, Day 0, biol rep 3

description: RawCountsMatrix.tsv

value: KOLF2, Day 0

value: induced pluripotent stem cells


title: 12393

description: WHWT

value: West Highland White Terrier


title: ChIPseq H3K27Ac, CTL

description: ChIP-seqGSA_K27ac_Treated_Untreated.xlsx, Peaks_ChIP_K27ac_CTL.xlsx

value: H3K27Ac (Abcam)


title: KOLF2 cells, SETBP1_VUS2, Day 24, biol rep 1

description: RawCountsMatrix.tsv


title: ChIPseq H3K27Ac, KRN


title: 13000

description: LAB-


title: KOLF2 cells, SETBP1_VUS2, Day 0, biol rep 4

description: RawCountsMatrix.tsv


title: KOLF2 cells, SETBP1_VUS2, Day 0, biol rep 2

description: RawCountsMatrix.tsv


title: 11110

description: WHWT


title: 12679

description: SSHP

value: Shetland Sheepdog


title: 12738

description: GOLD


title: ChIPseq H3K27me3, Input CTL

description: No significant peaks (MACS2) were found for H3K27me3 in any experimental condition, hence no processed data was obtained.

value: Input

value: H3K27me3 (Cell Signaling)


title: ChIPseq H3K4me3, CTL

description: GSA_K4me3_Treated_Untreated.xlsx,Peaks_chip_K4me3_CTL.xlsx


title: ChIPseq H3K27me3, Input KRN


title: KOLF2 cells, SETBP1_VUS2, Day 24, biol rep 2

description: RawCountsMatrix.tsv


title: ChIPseq H3K27Ac, Input


title: 11673

description: GOLD


title: KOLF2 cells, SETBP1_VUS2, Day 0, biol rep 1

description: RawCountsMatrix.tsv


title: ChIPseq H3K27me3, KRN


title: 11385

description: SCOT

value: Scottish Terrier


title: ChIPseq H3K27me3, CTL

value: Liver


title: N102_2

city: Nanchang

first: Wenxuan

last: Fu

address: china

institute: Nanchang university

data_processing:
Sequenced reads were trimmed for adaptor sequence and low-quality sequence.
The trimmed reads were alignmented to the human reference genome GRCh38 with HISAT2 software.
The expression level of each gene was calculated by kilobase of transcript per million mapped reads (FPKM), and then the differentially expressed genes (DEGs) were identified by DESeq2 software.
Assembly: GRCh38
Supplementary files format and content: Tab-delimited text file includes readcount values for each sample.

source_name: CAL 27

growth_protocol: CAL 27 cells were cultured in Dulbecco’s modified Eagle medium (DMEM) supplied with 10% fetal bovine serum.

extract_protocol:
RNA was collected by using Trizol regeant.
RNA library was prepared by using  VAHTS Universal V8 RNA-seq Library Prep Kit for Illumina.

treatment_protocol: CAL 27 cells were transfected with 20nM anti-MORC3 siRNA or non-specific siRNA twice in an interval of 48 hours. Total RNAs were collected at 48 hours after first transfection.

value: CAL 27

value: Oral squamous cell carcinoma

value: MORC3 knockdown

value: siRNA against MORC3


title: 10811

description: AESK

source_name: Whole Blood

value: American Eskimo Dog


title: N102_3


title: 11104

description: FCR-

value: Flat-Coated Retriever


title: ChIPseq H3K4me3, Input

value: Liver iNKT (aGalCer/CD1d tetramer+)

value: NOD mouse

value: H3K4me3 (Sigma)


title: 10810

description: AESK


title: ATACseq KRN, replicate 3

description: Diff_ATAC_CTL_peaks_annotated.xlsx, Shared_ATAC_peaks_annotated.xlsx


title: 12739

description: GOLD

value: Golden Retriever


title: ATACseq KRN, replicate 2


title: 10804

description: AESK


title: 10807

description: AESK


title: Cohort 2-RSD-PLX

first: Ethan

last: Goodman

address: 460 Medical Center Drive

institute: Ohio State University

data_processing:
Sequencing reads were aligned to the mm10 mouse genome and count matrices were generated using CellRanger (10X Genomics). Low-quality cells (doublets, low counts, high counts) were filtered using the Seurat package in R. Cells expressing >20% mitochondria RNA were excluded from analysis.
Assembly: mm10
Supplementary files format and content: fastq, mtx

source_name: hippocampus

extract_protocol:
Mice mice underwent CO2 asphyxiation and were perfused with phosphate buffered saline (PBS). The hippocampus was dissected and dissociated using the Miltenyi Adult Mouse Brain Dissociation Kit per manufacturer’s instructions with minor modifications. Tissue was immediately placed into Enzyme P solution (37°C), Enzyme A solution was added, and tissue was dissociated in C-tubes using a gentleMACS dissociator. Density centrifugation was used to remove myelin debris, remaining red blood cells were lysed, and cell viability and concentration was determined using a hemocytometer. Hippocampi from three mice per group were pooled into each sample. This experiment was performed in two replicates, with six mice per group total, and approximately 3,000-5,000 cells were recovered from each sample.
Single-cell suspension was loaded onto a 10X chip and run on a Chromium Controller to generate gel-bead emulsions. Protocols for UMI barcoding, cDNA amplification, and library construction followed the Chromium Next GEM Single Cell 3' GEM, Library & Gel Bead Kit v3.1 kit protocol (10x Genomics; 1000121). Library quality was determined by Agilent High Sensitivity DNA BioAnalyzer chip and was sequenced using an Illumina HiSeq.

value: hippocampus

value: PLX Diet


title: NC_2

value: WT

value: non-specific siRNA


title: ATACseq CTL, replicate 3

value: untreated


title: N102_1


title: NC_1


title: 10802

description: AESK


title: 10805

description: AESK


title: Donor2_HTO

city: Huddinge

first: Johan

middle: K.

last: Sandberg

state: Stockholms

address: Alfred Nobels Allé 8b

department: Department of Medicine Huddinge

country: Sweden

institute: Karolinska Institutet

description: HTO

data_processing:
BCL to FASTQ conversion was performed using CASAVA software's bcl2fastq_v2.20.0.422.
Sequence alignment was performed following the cellranger multi pipeline (v6.1.2).
Singlets were identified using HTODemux (Stoeckius et al., DOI: 10.1186/s13059-018-1603-1) and scDblFinder (Germain et al., DOI: 10.12688/f1000research.73600.2).
Cells were quality controlled (nFeature_RNA>200, nCount_RNA>1000, nCount_RNA<8000, percent_mito<10, percent_ribo>10).
Cells annotated as T cells or NK cells by sc-type algorithm were retained for downstream analysis.
Assembly: GRCh38-2020-A
Library strategy: CITE/TCR-seq

molecule: protein

source_name: Peripheral blood

extract_protocol:
Peripheral blood was obtained from four healthy human donors. After lymphocyte isolation, 5-OP-RU-hMR1-tetramer-PE-positive cells were magnetically enriched and marked (HTO#1). 5-OP-RU-hMR1-tetramer-PE-negative cells were counter-stained (HTO#3). Both populations were stained with CITE-seq antibodies and sorted as live CD14- CD19- CD45+ MAITcells  (5OPRU-hMR1-tetramer-positive) and non-MAIT cells (5-OPRU-hMR1-tetramer-negative).
Single cell libraries were created following the 10X Chromium Next GEM Single Cell 5' v2 protocols with feature barcode technology for cell surface protein and immune receptor mapping. Libraries were pooled and sequenced with a 26-10-10-90 nucleotide read setup on a Illumina NovaSeq6000 v1.5 S1-100 at the National Genomics Infrastructure at the Science for Life Laborartory in Stockholm, Sweden.

value: Peripheral blood

value: CD14- CD19- CD45+ 5-OP-RU-hMR1-tetramer+/-


title: 10808

last: Harris

institute: National Institutes of Health

description: AESK


title: Cohort 2-Control-PLX


title: ATACseq KRN, replicate 1

value: Liver


title: ATACseq CTL, replicate 1


title: ATACseq CTL, replicate 2


title: Podocytes, NPR1 KO, rep6

city: Regensburg

first: Uwe

last: Schwartz

address: Universitätstraße 32

department: NGS Analysis Center

country: Germany

institute: University of Regensburg

data_processing:
Reads were mapped to the reference genome (GRCm38) and corresponding gene annotation (Ensembl version 98) using the Spliced Transcripts Alignment to a Reference (STAR) software (version v2.7.8a)(Dobin et al., 2013). The following options were used to optimize the alignment process: --outFilterType BySJout, --outFilterMultimapNmax 20, --alignSJoverhangMin 8, --alignSJDBoverhangMin 1, --outFilterMismatchNmax 999, --alignIntronMin 10, --alignIntronMax 1000000, --outFilterMismatchNoverReadLmax 0.04, --runThreadN 12, --outSAMtype BAM SortedByCoordinate, --outSAMmultNmax 1, and --outMultimapperOrder Random
The level of PCR duplication was assessed using Picard MarkDuplicates (v2.21.8) (Broad Institute, 2019), and dupRadar (version v1.15.0) (Sayols et al., 2016).
Gene expression quantification was carried out using featureCounts (v1.6.3) (Liao et al., 2014).
Assembly: GRCm38
Supplementary files format and content: tab-delimited text file includes raw counts for each sample
Supplementary files format and content: tab-delimited text file includes TPM values for each sample

source_name: Kidney

growth_protocol: Eight weeks old mice with a podocyte-specific ablation of GC-A/NPR-A were included in the study. The mice expressed Cre recombinase under the control of the podocin (NPHS2) promoter and carried floxed alleles of the gene Npr1 encoding GC-A/NPR-A (NPHS2-Cre x GC-Aflox/flox), designated as Podo GC-A KO. Control mice (NPHS2-Cre x GC-AWT/WT) had a functional GC-A in podocytes and were designated as Podo GC-A control. For separation of podocytes from other glomerular cells by fluorescence-activated cell sorting (FACS) the mouse line was crossed with a double fluorescent Cre-reporter mouse (Gt(ROSA)26Sortm4(ACTB-tdTomato,-EGFP)Luo/J,mG/mT) .

extract_protocol:
Total RNA was extracted from FACS sorted mouse podocytes according to the “Purification of total RNA from animal and human cells” protocol of the RNeasy Plus Micro Kit (QIAGEN, Hilden, Germany)
Library preparation were carried out as described in the Illumina TruSeq Stranded mRNA.

treatment_protocol: In the first step, glomeruli were isolated by the magnetic bead method from six Podo GC-A KO and six Podo GC-A control mice, all with GFP-expressing podocytes, (20, 24).  The day before perfusion the magnetic beads (Dynabeads M-450 Epoxy, Invitrogen, 4.5 microns) were cleaned and blocked in 5 mg/mL BSA in HBS for 16-20 hours at room temperature with orbital shaking at 300 rpm. Before kidney perfusion the beads were rinsed in HBSS and added to an enzyme solution containing Collagenase type 2 (Worthington, LS004176, 230 U/g), PronaseE (Merck, 1074330001), DNase I (Applichem A3778) and HBSS and prewarmed to 37 °C. The mice were sacrificed by cervical dislocation, a perfusion cannula was inserted into the abdominal aorta, the aorta was clamped superior to the renal arteries together with the superior mesenteric artery and the kidneys were perfused with 10 mL HBSS followed by perfusion with 8 mL beads suspension (5 μl beads/ml HBSS) and 2 mL enzyme beads solution (5 μl beads/ml enzyme solution). Subsequently, kidneys were removed, weighted, decapsulated, minced by a scalpel, and digested in 1 mL enzyme solution (37 °C, 30 minutes at 300 rpm). The resulting suspension was pressed through a 100 µm cell strainer twice into 50 mL falcons with HBSS. The falcons were placed next to a magnet for seven minutes and the HBSS was discarded. The remaining magnetic glomeruli were resuspended in HBSS, and the procedure was repeated three times.  Finally, the glomerulus suspension was centrifuged (five minutes, 4 °C, 1500 rpm) and the supernatant was removed. To isolate podocytes, the glomeruli were resuspended in 2 mL enzyme solution (50 minutes, 37 °C, 1400 rpm). The incubation was stopped after every five minutes to release the podocytes by pipetting or shaking. After the end incubation, a magnet was used to remove unwanted beads and tissue. The supernatant was transferred to a 40 μm cell strainer and washed with HBSS. The samples were centrifuged at 1500 rpm for five minutes at 4 °C and the supernatant was removed. The cells were resuspended in 475 μl FACS-buffer (1xPBS + 0,2% Fetal Calf Serum (FCS) in 50 ml 1x PBS + 100 μl FCS) and a 25 μl Propidium iodide solution (1 mg/ml in HBSS, Sigma, P4170) and sorted by FACS. Podocytes (green fluorescence, GFP) and other glomerular cells (red fluorescence, tomato) were sorted in separate tubes containing 450 μl RNAprotect cell reagent (Qiagen, 76526) to stabilize the RNA.

value: Kidney

value: Podocytes

value: NPR1 KO


title: Donor4_HTO

description: HTO


title: 10801

city: Bethesda

description: AESK

value: American Eskimo Dog


title: Cohort 2-RSD-Veh

value: Control Diet


title: 10800

description: AESK


title: Cohort 1-Control-PLX


title: Podocytes, NPR1 KO, rep4


title: Cohort 2-Control-Veh


title: Donor1_HTO

description: HTO


title: Cohort 1-RSD-PLX


title: Donor3_HTO

description: HTO


title: NC_3

value: CAL 27

value: Oral squamous cell carcinoma


title: 10705

description: FCR-

value: Flat-Coated Retriever


title: Donor3_ADT

description: ADT


title: 10048

description: FCR-


title: Podocytes, NPR1 KO, rep3


title: 10023

description: WHWT

value: West Highland White Terrier


title: Donor2_ADT

description: ADT


title: Donor1_ADT

description: ADT


title: 9321

description: MCUR

value: Mountain Cur


title: Donor4_ADT

description: ADT


title: Podocytes, NPR1 KO, rep5


title: input, Rosi, biological replicate3

city: Los Angeles

first: Xia

last: Yang

state: California

address: 2000 Terasaki Life Sciences Bldg

department: Department of Integrative Biology and Physiology

institute: University of California, Los Angeles

data_processing:
Image analysis and base calling were conducted by the HiSeq Control Software (HCS). Raw sequence data (.bcl files) generated from Illumina HiSeq was converted into fastq files and de-multiplexed using Illumina's bcl2fastq 2.17 software. One mismatch was allowed for index sequence identification. Quality control of raw sequencing data was conducted using FastQC, employing default parameters. Subsequently, sequencing reads were mapped to the mouse genome mm10 using Salmon. Differential gene expression analysis was then undertaken with DESeq2.
Assembly: mm10
Supplementary files format and content: Plain-text, tab-separated file that includes Salmon's quantification output  for each sample
Library strategy: Polysome-Seq

source_name: epididymal adipose tissue

growth_protocol: DMEM (GIBCO) 10% FBS (Omega FB#11), 1% PenStrep (GIBCO), and 50ng/mL Primocin (Invivogen) - 37C incubator 5% CO2

extract_protocol:
Stromal vascular fractions were isolated as previously described. Briefly, entire WAT depots were cut into small pieces, suspended in DMEM (GIBCO) + 50 mM HEPES (GIBCO) + 1mg/mL type II collagenase (Sigma), + 1% BSA - low fatty acid (Gemini), and digested for 30 minutes at 37°C with agitation (120 rpm). The resulting dissociated tissue was passed through 100μm strainers, and adipocytes were removed from the supernatant by centrifugation (750g, 10min, 4C). After a second filtration with 40μm strainers, red blood cells were lysed using ACK lysis buffer (GIBCO). Cells were then treated to differentiate with complete adipogenic cocktail (0.5mM 3-isobutyl-1-methylxanthine (Sigma), 1μM dexamethasone (Sigma), 5μg/mL insulin (GIBCO) plus either 1μM rosiglitazone (Novo Nordisk) or vehicle (DMSO) for 2 days. After that, media was replaced for minimal maintenance media (Insulin/Rosi or Insulin/DMSO) for 2 days. At day 4 of differentiation cells were boosted with fresh media for one hour and for the last 5 minutes cycloheximide (Sigma) was added to the media (final concentration 100μg/mL). Cells were lysed and collected in hypotonic lysis buffer (5mM Tris HCl pH 7.5, 2.5 mM MgCl2, 1.5 mM KCl, 100 μg/mL cycloheximide, 2 mM DTT), and 0.5% Triton, 0.5% Sodium Deoxycholate were then added to the cell lysates after collection. The cytoplasmic extracts were resolved on 5-50% sucrose gradients by centrifugation in an SW40 rotor at 150,000 x g for 2 hours. The absorbance at 260 nm was measured using a Piston Gradient Fractionator Model 153 instrument (BioComp, Canada). RNA from polysomes or total RNA pool was resuspended in Trizol reagent (ThermoFisher), and RNA was isolated using Direct-zol RNA Microprep kits as described by the manufacture (Zymo research).
The RNA sample received was quantified using Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA) and RNA integrity was checked using TapeStation (Agilent Technologies, Palo Alto, CA, USA). The RNA sequencing library was prepared using the NEBNext Ultra RNA Library Prep Kit for Illumina using manufacturer’s instructions (NEB, Ipswich, MA, USA). Briefly, mRNAs were initially enriched with Oligod(T) beads. Enriched mRNAs were fragmented for 15 minutes at 94 °C. First-strand and second-strand cDNA were subsequently synthesized. cDNA fragments were end-repaired and adenylated at 3’ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by PCR with limited cycles. The sequencing library was validated on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and quantified by using Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA) as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA). The sequencing library was clustered on three lanes of a flowcell. After clustering, the flowcell was loaded on the Illumina HiSeq instrument (4000 or equivalent) according to the manufacturer’s instructions. The sample was sequenced using a 2x150bp Paired End (PE) configuration.

value: epididymal adipose tissue

tag: Sex

value: male

value: Lepob/Lepob (The Jackson Laboratory #000632)

value: stromal vascular fraction

tag: age

value: Rosiglitazone


title: Podocytes, NPR1 KO, rep2


title: Donor3_GEX

description: GEX

value: Peripheral blood

value: CD14- CD19- CD45+ 5-OP-RU-hMR1-tetramer+/-


title: Cohort 1-Control-Veh

value: hippocampus


title: input, Rosi, biological replicate1

treatment_protocol: Cells were then treated to differentiate with complete adipogenic cocktail (0.5mM 3-isobutyl-1-methylxanthine (Sigma), 1μM dexamethasone (Sigma), 5μg/mL insulin (GIBCO) plus either 1μM rosiglitazone (Novo Nordisk) or vehicle (DMSO) for 2 days. After that, media was replaced for minimal maintenance media (Insulin/Rosi or Insulin/DMSO) for 2 days. At day 4 of differentiation cells were boosted with fresh media for one hour and for the last 5 minutes cycloheximide (Sigma) was added to the media (final concentration 100μg/mL).


title: Donor4_GEX

description: GEX


title: 7789

first: Alexander

address: 50 South Drive

description: SCOT

value: Scottish Terrier


title: input, Veh, biological replicate3

value: Vehicle


title: input, Rosi, biological replicate2


title: Podocytes, NPR1 KO, rep1

value: Kidney

value: Podocytes

value: NPR1 KO


title: Podocytes, WT, rep6

value: WT


title: Podocytes, WT, rep4


title: Donor1_GEX

description: GEX


title: 7733

description: DDMT

value: Dandie Dinmont Terrier


title: Donor2_GEX

description: GEX


title: Podocytes, WT, rep3


title: Cohort 1-RSD-Veh

value: Control Diet


title: 7979

description: FCR-

value: Flat-Coated Retriever


title: 7731

description: DDMT


title: Podocytes, WT, rep5


title: 7555

description: IWSP

value: Irish Water Spaniel


title: input, Veh, biological replicate1

value: epididymal adipose tissue

value: male


title: Podocytes, WT, rep2


title: MIA PaCa-2, 24hr, sotorasib (1μM), rep5

city: Palo Alto

first: Jin Hyun

last: Ju

address: 3160 Porter Dr, Suite 250

institute: BridgeBio

data_processing:
Sequence reads were trimmed to remove possible adapter sequences and nucleotides with poor quality using Trimmomatic v.0.36. The trimmed reads were mapped to the Homo sapiens GRCh38 reference genome available on ENSEMBL using the STAR aligner v.2.5.2b to generate BAM files.
Unique gene hit counts were calculated by using featureCounts from the Subread package v.1.5.2. The hit counts were summarized and reported using the gene_id feature in the annotation file. Only unique reads that fell within exon regions were counted. If a strand-specific library preparation was performed, the reads were strand-specifically counted.
After extraction of gene hit counts, the gene hit counts table was used for downstream differential expression analysis. Using DESeq2, a comparison of gene expression between the customer-defined groups of samples was performed.
Assembly: GRCh38
Supplementary files format and content: comma separated raw counts for all samples

source_name: MIA PaCa-2

growth_protocol: Mia PaCa-2 cells pancretic adenocarcinoma cells were maintined in DMEM supplemented with 10% fetal bovine serum (FBS) and penicillin/streptomycin (Pen/Strep) in humidified atmosphere at 37C

extract_protocol:
RNA was extracted by Azenta life sciences' standard protocol.
cDNA libraries were constructed by Azenta life sciences' standard protocol

treatment_protocol: Cells were treated for 24 hours with DMSO,  30nM BBO-8520, 300nM adagrasib (MRTX-849) or 1μM sotorasib (AMG-510) with 5 replicates/condition. All had a final DMSO concentration of 0.1%.

tag: cell line

value: MIA PaCa-2

value: Pancreatic Adenocarcinoma

tag: time

value: 24 hrs

value: sotorasib (1μM)


title: 7333

description: GOLD

value: Golden Retriever


title: eWAT, Rosi, biological replicate2

value: Lepob/Lepob (The Jackson Laboratory #000632)

value: stromal vascular fraction

tag: treatment

value: Rosiglitazone


title: Podocytes, WT, rep1

tag: tissue

value: Kidney

value: Podocytes

tag: genotype


title: eWAT, Rosi, biological replicate3


title: 6220

description: LHSA

tag: breed

value: Lhasa Apso


title: 6033

description: BRUS

value: Brussels Griffon


title: 7285

description: LAB-

value: Labrador Retriever


title: eWAT, Veh, biological replicate3

value: Vehicle


title: eWAT, Veh, biological replicate2


title: MIA PaCa-2, 24hr, sotorasib (1μM), rep4


title: eWAT, Rosi, biological replicate1


title: MIA PaCa-2, 24hr, sotorasib (1μM), rep1


title: MIA PaCa-2, 24hr, sotorasib (1μM), rep3


title: MIA PaCa-2, 24hr, sotorasib (1μM), rep2


title: 5905

description: SUSX

value: Sussex Spaniel


title: input, Veh, biological replicate2

value: epididymal adipose tissue


title: MIA PaCa-2, 24hr, adagrasib (300nM), rep5

value: adagrasib (300nM)


title: 5378

description: LANC

source_name: Unknown Source

value: Lancashire Heeler