title: I738

city: Erlangen

first: David

last: Voehringer

address: Wasserturmstrasse 3-5

department: Department of Infection Biology

institute: University Clinic Erlangen

data_processing:
Raw data (raw reads) of FASTQ format were firstly processed through in-house scripts. In this step, clean data (clean reads) were obtained by removing reads containing adapter and poly-N sequences and reads with low quality from raw data.
Reference genome and gene model annotation files were downloaded from genome website browser (NCBI/UCSC/Ensembl) directly. Paired-end clean reads were mapped to the reference genome using HISAT2 software. HISAT2 uses a large set of small GFM indexes that collectively cover the whole genome. These small indexes (called local indexes), combined with several alignment strategies, enable rapid and accurate alignment of sequencing reads.
HTSeq was used to count the read numbers mapped of each gene, including known and novel genes. And then FPKM (Fragments Per Kilobase Million) of each gene were generated.
Genome_build: Mus_musculus GRCm38
Supplementary_files_format_and_content: Table of FPKM values with gene annotations in xls format

source_name: Splenic B cells

extract_protocol:
Total RNA was isolated using the Rneasy kit from Qiagen
A total amount of 1 μg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer (5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase (RNase H-). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3’ ends of DNA fragments, NEBNext Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 150~200 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). Then 3 μl USER Enzyme (NEB, USA) was used with size-selected, adaptorligated cDNA at 37 °C for 15 min followed by 5 min at 95 °C before PCR. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using PE Cluster Kit cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina platform and paired-end reads were generated.

treatment_protocol: Sort of CD19+ CD3- Propidium Iodid- Cells from total splenocytes

tag: cell type

value: Splenic B cells

tag: genotype

value: CD19Cre; STAT6vt

tag: strain

tag: gender

value: male


title: 18B

status: Public on Dec 14 2020

type: SRA

city: New York

first: Kiley

last: Graim

state: NY

address: 162 Fifth Ave

country: USA

institute: Flatiron Institute

library_source: transcriptomic

data_processing:
Reads mapped using HISAT2
Assemblies filtered using FastQC
Read counts constructed using Dexseq-Count
Count matrix normalized using TMM
Genome_build: canfam3.1
Supplementary_files_format_and_content: comma-delimited text file includes TMM values for each Sample

molecule: polyA RNA

organism: Canis lupus familiaris

source_name: mammary gland_Adenoma

extract_protocol:
Qiagen Rneasy kit
PrepX mRNA Library protocol for the Apolloa324 System (Wafergen)

treatment_protocol: cryo-pulverized then homogenized using a rotor-stator in Trizol (Cat. No.15596-026, Invitrogen), lystate further homogenized using a Quashredder spin column

tag: patient number

tag: tumor diameter

tag: goldschmidt

value: adenoma with sclerosis, duct ectasia with papillomatosis

tag: histology

value: B

tag: simhist

value: Adenoma

tag: dethist

tag: library prep

tag: sequencing run

tag: tumor location

tag: repselect

tag: rin


title: 18F

value: complex adenoma

value: Complex Adenoma


title: 18A

source_name: mammary gland_Normal

value: duct ectasia with periductal inflammation

value: N

value: Normal

value: Duct Ectasia


title: I737

country: Germany

molecule: total RNA

organism: Mus musculus


title: 17D

value: Benign adenoma with lobular hyperplasia


title: human WBC DNase [GB4029008_S8]

city: Bethesda

first: Keji

last: Zhao

state: MD

address: Building 10 Room 7B06A

department: National Heart, Lung, and Blood Institute

institute: national Institute of Health

library_source: genomic

data_processing:
library strategy: indexing single-cell DNase sequencing (iscDNase-seq)
Basecalls performed
Each raw fastq file are reads corresponding to 96 cell barcodes . The 96 pairs are shown in barcode_96.txt. Read 1 is considered  to belong to one of the barcodes if  it corresponding read 2 contains  that barcode.  After separting each set of fastq files into 96 files, read 1 is mapped to the human genome hg18 using bowtie2.
The top 30 barcodes are selected from each well based on read count, so 30 nuclei are sorted for each well with doublets removed, where the number removed was calculated based on the collision rate like in (Science. 2015 May 22; 348(6237): 910–914). These are denoted as single cells because of the low likelihood of two cells in the same well having the same barcode. We required that qualifying cells have more than 100 reads and a FRiP greater than 0.15. With these requirements, 20621 cells were obtained . From the pooled single cell data, we obtaind a set of DHS that are shown in "combined_20621sc_noDup2-W100-G0-E100.scoreisland". There are 132926 DHSs. For each cell, we calculated the reads that were located within the DHSs and stored the results in a matrix, which can be found from the file 'scDNase_KZ1839_hg18_rc20621_mat.txt.gz'
Genome_build: hg18
Supplementary_files_format_and_content: barcode_96.txt: one column  refers to a DNA barcode starting from the 3' .
Supplementary_files_format_and_content: combined_20621sc_noDup2-W100-G0-E100.scoreisland: first column is for chromsome, second column is for DHS peak start site,  third column is for DHS peak end site, fourth column is the -log(E-value)
Supplementary_files_format_and_content: scDNase_KZ1839_hg18_rc20621_mat.txt.gz: This file contains read count data for single cells in text format. The rows refer to DHS while the column refer to single cells. Reading the matrix results in a 132926 x 20621 matrix.

molecule: genomic DNA

organism: Homo sapiens

source_name: WBC

extract_protocol:
DNase-seq human 1.25M 2-step crosslink human healthy T/B/NK/monocyte, final 0.1U/μl DNase digestion 37°C 5 minutes, T cell split #1-15 wells, B cell split #16-30 wells, NK cell split #31-45 wells, monocyte split #46-61 wells, TdT&T4 methods adding P7 barcode adaptor (12.5μl volumn/well 37°C 60 minutes), pool, dilution, counting and redistribute into 96-well plate, total 20 nuclei per well seal completely, 65°C overnight; direct PCR1 (12 cycles), pool all the DNA, EXO I 30 minutes, A-tailing and P5 ligation(rt overnight) , column purification and PCR2(16 cycles), run-e-gel and cut from 200-500bp rc#1 i5-01
The isolated 50M of PBMC suspended by 50ml PBS /MgCl2 are firstly fixed by adding 400μl freshly prepared 0.25M Disuccinimidyl glutarate (DSG, ThermoFisher Scientific, catalog no.20593) and incubate at room temperature for 45 minutes with rotation. After protein-protein fixation by DSG and washing three times by PBS, cells suspended by culture medium DMEM supplemented with 10% FBS are further fixed by the addition of 1:15 volume of 16% (w/v) methanol-free formaldehyde solution (Thermo Fisher Scientific) and incubate at room temperature for 10 minutes. The fixing reaction is terminated by a 1:10 volume of 1.25 M glycine and incubate at room temperature for 5 minutes. The fixed cells are collected by centrifugation at 1320rpm for 7 minutes and washed with PBS. The cells are stored in aliquots (1 × 106 cells per tube) at −80°C until use.

tag: cell lines

value: WBC


title: 16F

source_name: mammary gland_Carcinoma

value: intraductal carcinoma

value: M

value: Carcinoma


title: 17B

source_name: mammary gland_Carcinoma-In Situ

value: Duct papillomatosis, epitheliosis and carcinoma-in-situ

value: Carcinoma-In Situ

value: In Situ Carcinoma


title: 16C

value: simple adenoma


title: 16D

value: B


title: 17A

value: Lobular hyperplasia

value: Lobular Hyperplasia


title: 16B

value: complex adenoma

value: Adenoma

value: Complex Adenoma


title: 17C

value: N

value: Normal


title: 15C

value: Ductal adenoma


title: 18D

value: tubular carcinoma, low grade


title: 16E

value: lobular hyperplasia


title: 15B

value: Tubular carcinoma, low grade with desmoplasia

value: M


title: I739

value: Splenic B cells

value: CD19Cre; STAT6vt

value: female


title: 15A

value: Mild duct ectasia and lobular hyperplasia


title: I734

value: male


title: I735


title: 14F

value: Complex adenoma

value: B


title: I736


title: 14E


title: 14D


title: 16A

value: intraductal papillary adenoma


title: 14A

value: Mild lobular hyperplasia and duct ectasia

value: Lobular Hyperplasia


title: 14C

value: Adenoma

value: Complex Adenoma


title: 14B

value: Carcinoma


title: 13D

value: Ductal adenoma


title: 13F

value: Adenoma with sclerosis


title: 13C

value: Tubular carcinoma


title: 13G


title: 13B


title: 13E


title: 13A

value: Duct ectasia, lactation

value: N

value: Normal

value: Duct Ectasia


title: 12D

value: Adenomas


title: 11J

value: Adenocarcinoma with focal lymphatic invasion

value: M


title: 14G

value: Lobular hyperplasia and duct ectasia


title: 12C

value: Severe duct ectasia and lobular hyperplasia


title: 12A


title: 11G

value: Complex adenoma

value: B


title: 12B

value: Severe duct ectasia with papillomatosis, lobular hyperplasia and multifocal periductal carcinoma with sclerosis


title: 10F

value: Lobular hyperplasia

value: Lobular Hyperplasia


title: 10B

value: Carcinoma


title: 11F

source_name: mammary gland_Atypia-Normal

value: Duct ectasia, lobular hyperplasia, duct papillomatosis with atypia and adenoma

value: Atypia-Normal

value: Atypia


title: 11E

value: Intraductal papillary adenoma

value: Adenoma


title: 10D

value: Adenoma with focal mineralization


title: 11A

value: Duct ectasia, lobular hyperplasia, duct papillomatosis with atypia

value: N

value: Normal


title: 11C


title: 10C


title: 10A

value: Adenoma and lobular hyperplasia


title: 11I

value: Adenocarcinoma, lobular hyperplasia, papillary adenoma

value: M


title: 9G

value: Duct ectasia and lobular hyperplasia


title: 9F

value: Low grade tubular carcinoma with necrosis


title: 11H

value: Duct ectasia, lobular hyperplasia, duct papillomatosis with atypia, carcinoma-in-situ

value: Carcinoma-In Situ

value: In Situ Carcinoma


title: 11B

value: Carcinoma


title: 8D

value: Duct ectasia

value: Duct Ectasia


title: 8B

value: Carcinoma (low grade based on discussion w/Dr. G)


title: 6C

value: Carcinoma with multifocal necrosis


title: 6E

value: Lobular hyperplasia with atypia

value: Atypia-Normal

value: Atypia


title: 9C

value: Complex adenoma

value: B

value: Adenoma

value: Complex Adenoma


title: 6B


title: 6D


title: 9B


title: 9D

value: N

value: Normal


title: 9A


title: 9E


title: 6A

value: Normal ductal tissue


title: 8C


title: 4F

value: lobular hyperplasia

value: Lobular Hyperplasia


title: 4D

value: lobular hyperplasia, duct ectasia with atypia


title: 4E

value: complex adenoma


title: 3D

value: adenoma (large duct type) and epitheliosis


title: 4C

value: duct ectasia

value: Duct Ectasia


title: 4A

value: carcinoma arising in mixed mammary tumor

value: M

value: Carcinoma

value: Mixed Carcinoma


title: 4B

value: adenoma

value: B


title: 3C

value: mild lobular hyperplasia


title: 3B

value: low grade, focal carcinoma arising in a complex adenoma


title: 8E

value: Tubular carcinoma (low grade based on discussion w/Dr. G)


title: 3A

value: lobular hypoplasia, adenoma, mastitis

value: Adenoma


title: 8A

value: Complex adenoma

value: Complex Adenoma


title: 2E

value: normal mammary tissue

value: N

value: Normal


title: 2F

value: carcinoma in-situ

value: Carcinoma-In Situ

value: In Situ Carcinoma


title: 2D

value: complex adenoma


title: 2C

value: lobular hyperplasia

value: Lobular Hyperplasia


title: 2B

value: adenoma

value: B


title: 2A


title: 1C

value: carcinoma arising in benign mixed tumor

value: M

value: Carcinoma

value: Mixed Carcinoma


title: 1A

value: severe duct ectasia/cyst formation

value: Duct Ectasia


title: 1E

value: mammary adenoma

value: Adenoma


title: RNA_Bach2KO_gp33tetramer_Cl13_D7_StemLike_3

first: Hong-wei

last: Sun

address: 9000 Rockville Pile

department: Office of Science & Technology

institute: NIAMS

description: RNAseq_WT_Bach2KO_GP33tetramer_Cl13_D7_StemLike_RPKM.txt

data_processing:
Raw sequencing data were processed with CASAVA 1.8.2 to generate FastQ files. Reads of sequentially varying length were mapped to the mouse genome mm10 using TopHat 2.1.0.
RPKM calculated using Partek 6.6
Genome_build: mm10
Supplementary_files_format_and_content: Text files, RPKM values

source_name: CD8 gp33 tetramer

extract_protocol:
Cells from spleen were obtained by mechanical disruption. Isolated cells were further sorted based on their surface markers.
50,000 P14 cells were sorted and resuspended in Qiazol (Qiagen). Total RNA was extracted with miRNeasy Mini kit (Qiagen). mRNA was selected using NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB). RNAseq libraries were generated using NEBNext Ultra RNA Library Prep Kit for Illumina (NEB) and NEBNext Multiplex Oligos for (NEB) according to manufacturer’s instruction.
The purified libraries were multiplexed and captured on an Illumina flow cell for cluster generation. Libraries were sequenced for 50 single read cycles on a HiSeq 3000  according to the manufacturer's protocols.

treatment_protocol: Mice were intravenously (i.v.) injected with 2×106 PFU LCMV clone 13.

value: Bach2 KO

tag: tissue

value: Spleen

tag: sorting marker

tag: culture condition

value: ex vivo

tag: infection

value: 7 days after LCMV Cl13 infection


title: RNA_WT_Cl13_gp33tetramer_D7_StemLike_2

tag: strain


title: RNA_Bach2KO_gp33tetramer_Cl13_D7_StemLike_2


title: RNA_Bach2KO_gp33tetramer_Cl13_D7_StemLike_1


title: RNA_Bach2gRNA_P14_Cl13_D7_StemLike_3

data_processing:
scRNA-seq: Raw sequencing data were processed with CASAVA 1.8.2 to generate FastQ files. Alignment, filtering, barcode counting, and unique molecular identifier (UMI) counting were performed using Cell Ranger v2.0.0
alignment, ATAC-seq: Raw sequencing data were processed with CASAVA 1.8.2 to generate FastQ files. ATAC-seq reads were mapped to the mouse genome (build mm10) with Bowtie 1.1.1.
peaks, ATAC-seq: Peaks were identified using MACS (v 1.4.3; default p-value threshold of 1E-5) .
alignment, RNA-seq: Raw sequencing data were processed with CASAVA 1.8.2 to generate FastQ files. Reads of sequentially varying length were mapped to the mouse genome mm10 using TopHat 2.1.0.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig files for ATAC-seq, UMI versus gene counting matrix for scRNA-Seq, RPKM txt tables for RNAseq.

source_name: CD8 P14

extract_protocol:
Cells from spleen were obtained by mechanical disruption. Isolated cells were further sorted based on their surface markers.
scRNA-seq: The scRNA-seq libraries were generated using Chromium Single Cell 3’ Library & Gel Bead Kit v2 (10X Genomic) according to manufacturer’s protocol. Briefly, 15,000 live cells were FACS-sorted and used to generate single-cell gel-bead in emulsion (GEM). After reverse transcription, GEMs were disrupted. Barcoded cDNA was isolated and amplified by PCR (12 cycles). Following fragmentation, end repair, and A-tailing, sample indexes were added during index PCR (8 cycles).
scRNA-seq: Indexed libraries were multiplexed and sequenced on an Illumina HiSeq 3000 instruments according to the manufacturer's instructions (26 cycles of Read 1, 8 cycles of i7 Index, and 98 cycles of Read2).
ATAC-seq: ATAC-Seq libraries for stem-like and terminally exhausted CD8 T cells were generated using the exact protocol as previously described (Shih et al. Cell. 2016). A Fast-ATAC protocol was used to generate ATAC-Seq libraries for pMIG, BACH2OE, control and Bach2 gRNA samples. Briefly, 10,000 FACS-sorted cells were pelleted and resuspended in 50 ul transposase mixture containing 25 ul of 2x TD buffer, 2.5 ul of TDE1 (Illumina), 0.5 ul of 1% digitonin (Promega), 22 ul of nuclease-free water. After 30 minutes incubation at 37 C, transposed DNA was purified and eluted in 10 ul elution buffer with MinElute PCR purification kit (QIAGEN).
ATAC-seq: The purified libraries were multiplexed and captured on an Illumina flow cell for cluster generation. Libraries were sequenced for 50 paired read cycles on a HiSeq 3000 or a NextSeq 550 according to the manufacturer's protocols.
RNA-seq: 50,000 P14 cells were sorted and resuspended in Qiazol (Qiagen). Total RNA was extracted with miRNeasy Mini kit (Qiagen). mRNA was selected using NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB). RNAseq libraries were generated using NEBNext Ultra RNA Library Prep Kit for Illumina (NEB) and NEBNext Multiplex Oligos for (NEB) according to manufacturer’s instruction.
RNA-seq: The purified libraries were multiplexed and captured on an Illumina flow cell for cluster generation. Libraries were sequenced for 50 single read cycles on a HiSeq 3000  according to the manufacturer's protocols.

value: transduced with Bach2 gRNA construct


title: RNA_Bach2gRNA_P14_Cl13_D7_StemLike_2


title: RNA_WT_Cl13_gp33tetramer_D7_StemLike_1


title: RNA_pMK9_P14_Cl13_D7_StemLike_3

value: transduced with pMK9


title: RNA_pMK9_P14_Cl13_D7_StemLike_2


title: RNA_BACH2OE_P14_Cl13_D7_StemLike_2

value: transduced with BACH2 over expression construct


title: RNA_BACH2OE_P14_Cl13_D7_StemLike_3


title: RNA_pMIG_P14_Cl13_D7_StemLike_3

value: transduced with pMIG


title: RNA_pMIG_P14_Cl13_D7_StemLike_2


title: RNA_pMIG_P14_Cl13_D7_StemLike_1


title: 1F

tag: patient number


title: RNA_WT_Cl13_gp33tetramer_D7_StemLike_3


title: RNA_Bach2gRNA_P14_Cl13_D7_StemLike_1


title: scRNA_Bach2gRNA_P14_Cl13_D7


title: RNA_pMK9_P14_Cl13_D7_StemLike_1


title: melan_a c-kit(K641E)hi exp_rep2

type: RNA

city: Boston

first: Hensin

last: Tsao

address: 55 Fruit St

institute: Massachusetts General Hospital

description: Gene expression data from  c-kit(K641E) transformed melanoma

hyb_protocol: Samples were submitted to the Boston Children’s Hospital BCH IDDRC Molecular Genetics Core (Boston, MA). Affymetrix Clariom S chip was used.

data_processing: CEL files were then processed with the TAC Console software

label: biotin

growth_protocol: mVec, mKitK641E(PT) and mKitK641E(T) cell lines were cultured in RPMI and 10%FBS.

extract_protocol: Total RNA was extracted from mVec, mKitK641E(PT) and mKitK641E(T) cells using the RNeasy isolation kit (Qiagen, Germany)

treatment_protocol: No treatment

value: melanocyte

tag: genotype

tag: age

value: immortalized


title: RNA_BACH2OE_P14_Cl13_D7_StemLike_1


title: melan_a CD516B2_rep2

description: Gene expression data from  control vector transducd melanocyte

scan_protocol: Samples were submitted to the Boston Children’s Hospital BCH IDDRC Molecular Genetics Core (Boston, MA). Affymetrix Clariom S chip was used.

source_name: mVec


title: scRNA_BACH2OE_P14_Cl13_D7


title: melan_a c-kit(K641E)hi exp_rep1

label_protocol: Samples were submitted to the Boston Children’s Hospital BCH IDDRC Molecular Genetics Core (Boston, MA). Affymetrix Clariom S chip was used.


title: scRNA_pMK9_P14_Cl13_D7


title: scRNA_pMIG_P14_Cl13_D7


title: ATAC_P14_Cl13_D7_TerminallyExhaused_2


title: ATAC_P14_Cl13_D7_TerminallyExhaused_1


title: melan_a c-kit(K641E)C3#_rep2


title: ATAC_P14_Cl13_D7_StemLike_2


title: melan_a CD516B2_rep1


title: 6_4.3F_VPS39ko

city: Malmö

first: Charlotte

last: Ling

address: Jan Waldenströms gata 35, CRC 91:12

department: Clinical Sciences

country: Sweden

institute: Lund University

hyb_protocol: according to the manufacturer’s recommendations

data_processing: Expression Console Software v1.4.1.46 was used for quality analyses and data were processed using Robust Multi-array Analysis (RMA) and log2 transformed

source_name: KO skeletal muscle

growth_protocol: Wild-type (WT) and Vps39+/- mice (KO), on a C57BL/6J background, were maintained under standard housing conditions with a 12-hour light/dark cycle at the animal facility, Sahlgrenska Academy, University of Gothenburg

extract_protocol: Whole mouse tibialis anterior muscle was homogenized in TissueLyser II and RNA extracted using RNeasy Fibrous Tissue mini kit (Qiagen #74704).

tag: genotype/variation

value: Vps39+/- mice (KO)

tag: Sex

value: female


title: 15_2.6M_VPS39ko

scan_protocol: according to the manufacturer’s recommendations

value: male


title: 16_4.1M_VPS39ko

label_protocol: according to the manufacturer’s recommendations


title: 3_1.3F_VPS39ko


title: melan_a c-kit(K641E)C3#_rep1


title: 2_1.2F_VPS39ko


title: 18_7.5M_VPS39ko


title: 13_1.1M_VPS39ko


title: 17_7.4M_VPS39ko


title: 5_2.7F_VPS39ko


title: 4_2.6F_VPS39ko


title: 14_1.2M_VPS39ko


title: ATAC_P14_Cl13_D7_StemLike_1


title: Spleen, Survivor, Day 0 [V40S]

city: Frederiksberg

first: Asma

middle: M

last: Karami

state: Denmark

address: Stigbojlen 7

country: Denmark

institute: University of Copenhagen

description: Replica 6

data_processing: All q-PCR assays exhibited efficiencies within 100% ± 5% so the simplified 2-ΔΔCq method was used for analysing data (Livak and Schmittgen, 2001). For normalization and as internal calibrator the average of three genes (ARP, ELF1α and β-actin) was chosen by using NormFinder (Andersen et al., 2004). For gene expression analyses, all challenged groups were compared to non-exposed time point controls using Student’s t-test. We considered differences to be significant when regulations were at least two-fold and p < 0.05. For four of the genes investigated in the non-exposed timepoint control groups less than three fish showed Cq values (excluding a t-test) and therefore we applied a qualitative assessment (presence/absence of Cq values) and analyzed data with the nonparametric Mann-Whitney test using a probability level of 5%.

label: FAM

organism: Oncorhynchus mykiss

source_name: Survivor

label_protocol: cDNA synthesis was performed in T100 thermocycler, Biorad, Denmark, using Oligo d(T)16 primer and TaqMan® Reverse Transcription Reagents (cat.no. N8080234, Thermo Fischer Scientific, Denmark). Finally, cDNA was stored at −20°C until further use. Quantitative PCR assays were performed using an AriaMx Real-Time PCR machine (cat.no.G8830A-04R-010, AH diagnostics AS, Denmark). The cycling conditions were one cycle of predenaturation at 95°C for 3 min. This was followed by 40 cycles of denaturation at 94°C for 5 sec with a combined annealing/elongation process at 60°C for 15 sec with endpoint measurement. Primers and Taq-Man probes targeting immune-relevant rainbow trout genes were synthesized at TAG Copenhagen AS, Denmark. Reaction volumes were 12.5 μl (2.5 μl cDNA, 6.25 μl Brilliant III Ultra-Fast QPCR Master Mix (600881, AH Diagnostics AS, Denmark), 1.0 μl primer-probe mixture (forward primer, 10 μM and reverse primer, 10 μM), Taq-Man probe (5 μM), and 2.75 μl RNase-free water. Reverse transcriptase minus and negative controls were used for every plate setup. . NormFinder (Andersen et al., 2004) was applied to evaluate all combinations of three reference genes (Elongation factor (ELF) 1-α, β-actin and acidic ribosomal phosphoprotein P0 (ARP)) as endogenous control; the average of the three genes was chosen and stability values were 0.002, 0.005 and 0.002 for liver, spleen and gills, respectively.

growth_protocol: Fish were maintain in plastic tanks with low density. Water was recycled (20 L/min) by internal biofilters (EHEIM, Germany) applying 30% water replenishment daily. Water quality was monitored and kept constant at pH 7.6, nitrite < 0.01 mg/l, nitrate < 50 mg/l (TetraTest 6in1, cat.no. T704154, Eldorado A/S, Denmark) and ammonia < 0.5 mg/l (API Ammonia Test Strips, VioVet Ltd, UK). During the 14 days acclimation and 14 days experimental infection period, fish received commercial pelleted feed (1.5% biomass per day, INICIO 917, BioMar A/S, Denmark). After the pathogen exposure fish with clinical signs were euthanized with overdose (300 mg/L) of ethyl-3-aminobenzoate methanesulfonate (MS222) (cat.no. A5040, Sigma-Aldrich, Denmark)

extract_protocol: RNAlater fixed samples from fish (gills, liver and spleen) were homogenized (Tissue-lyser II, Qiagen, Denmark) using a homogenization buffer with 2-mercaptoethanol (Sigma-Aldrich). RNA was extracted using the GenEluteTM mammalian RNA kit (RTN350, Sigma-Aldrich, Denmark). Liver samples were pre-treated by Proteinase K (cat.no.P4850, Sigma-Aldrich, Denmark). DNase I (AMPD1, Sigma-Aldrich, Denmark) treatment was applied to remove genomic DNA. The concentration of RNA was determined in a NanoDrop 2000 spectrophotometer (Saveen & Werner, Denmark). Quality of RNA was evaluated by agarose (cat.no. 16500100, ThermoFisher Scientific, Denmark) gel electrophoresis. RNA was kept at −80°C until cDNA synthesis.

treatment_protocol: Tissue (liver, spleen, gills) samples were fixed in RNAlater (cat.no R0901, Sigma-Aldrich, Denmark), placed at 4°C for 24 h and subsequently stored at -20°C until further processing.

tag: organ

tag: pathogen

value: Vibrio anguillarum

tag: time

value: Day 11


title: Spleen, Survivor, Day 0 [V38S]

description: Replica 6


title: Spleen, Survivor, Day 0 [V37S]

description: Replica 6


title: Spleen, Survivor, Day 0 [V36S]

description: Replica 6


title: RUNX1EVI1-NoDox-Pu1-ChIP

city: Birmingham

first: Peter

last: Keane

address: Vincent Drive

department: Institute for Cancer and Genomic Sciences

country: United Kingdom

institute: University of Birmingham

data_processing:
Raw single-end reads from DNaseI-seq experiments were processed with Trimmomatic v0.32.
The processed reads were aligned to the mouse genome (version mm10) using Bowtie2 v2.2.3 with the option --very-sensitive-local
Peaks were identified using MACS v2.1.1 with the options -g mm --keep-dup all -B --trackline
Genome_build: mm10
Supplementary_files_format_and_content: bedGraph files of sequence read pileups generated by MACS2

source_name: A2lox.cre mESC

growth_protocol: Generation of RUNX1-ETO and RUNX1-EVI1 containing ESCs was previously described (Kellaway et al., 2020; Regha et al., 2015). R201Q and R204X plasmids were generated by site-directed mutagenesis on wild type human RUNX1c, and N-terminal HA tags added using the following primers: R201Q forward 5’-CAGTGGATGGGCCCCAAGAACCTCGAAGAC-3’, reverse 5’-GTCTTCGAGGTTCTTGGGGCCCATCCACTG-3’ and R204X forward 5’-CCCCCTCGAGCCACCATG-3’, reverse 5’-GCCGATGATATCTCAAGGTTCTCG-3’. A2lox ESCs (a gift from Michael Kyba) were transduced with 20 µg of each plasmid using the 4D-Nucleofector (Lonza), mouse ES program and P3 primary cell kit. Note, R201Q, R204X and RUNX1-ETO all included N-terminal HA-tags.Individual colonies were expanded and maintained on mouse embryonic feeder cells in ES cell medium, comprising DMEM (Sigma D6546), 15% FCS (Sigma ES-009), 100 units/ml penicillin, 100 µg/ml streptomycin, 1 mM sodium pyruvate, 1mM L-glutamine, 0.15 mM monothioglycerol, 1x non-essential amino acids and 103 U/ml leukaemia inhibitory factor (ESGRO, Millipore) following 7 days of 300 µg/ml neomycin selection.

extract_protocol:
ChIP was performed as previously described (Obier et al, 2016) with the following modifications. For H3K27ac only KIT+ Progs were crosslinked only in 1% formaldehyde (single crosslinking, as opposed to double crosslinking for RUNX1). For single crosslinked cells nuclei were sonicated for 4 cycles of 30s on/30s off using a Picoruptor (Diagenode). Immunoprecipitation was carried out overnight at 4°C using 2 µg of RUNX1 antibody (ab23980, Abcam), or for four hours at 4°C using 2 µg of H3K27ac antibody (??) coupled to 15 µg Dynabeads Protein G (Invitrogen) per 2 x 106 cells. DNA from 2-3 immunoprecipitations was pooled for RUNX1, but just 1 immunoprecipitation for H3K27ac, and extracted using Ampure beads (Beckman Coulter).
ChIP libraries were generated using the KAPA hyper prep kit, libraries were size selected to obtain fragments between 150 and 450 bp

treatment_protocol: ESCs were differentiated as previously described (Gilmour et al, 2014; Regha et al., 2015) with the following modifications. FLK1+ cells were purified by magnetic cells sorting, using biotin-conjugated CD309 antibody (eBioscience), anti-biotin microbeads (Miltenyi Biotec) and LS columns (Miltenyi Biotec) following culture of embryoid bodies for between 3.25 and 3.75 days (cell line dependent). These FLK1+ cells were then cultured in gelatin-coated flasks – 1.2-1.4x106 cells in a T150 flask to form the blast culture. After 1-2 days (cell line dependent), 0.1-0.5 µg/ml doxycycline was added as appropriate and cells were cultured in the same media for a further 18 hours prior to sorting for HE and Progs. Cell populations were identified and sorted on day 2-3 of blast culture based on surface markers. Cells were stained with KIT-APC (BD pharmingen), Tie2-PE (eBioscience) and CD41-PE-Cy7 (eBioscience) and analyzed on a Cyan ADP flow cytometer (Beckman Coulter) with data analysis using FlowJo, or sorted on a FACS Aria cell sorter (BD Biosciences).

tag: cell type

value: A2lox.cre mESC

tag: target

tag: antibody

value: Santa Cruz (T-21) sc-352


title: R201Q_PlusDox_Gata1

value: Abcam ab11852


title: 10_6.5F_WT

source_name: WT skeletal muscle

value: wild type


title: Spleen, Survivor, Day 0 [V33S]

description: Replica 6

value: Spleen


title: 24_4.3M_WT


title: 9_6.4F_WT


title: 21_2.8M_WT


title: 23_4.2M_WT


title: 22_2.9M_WT


title: R201Q_NoDox_Gata1


title: Spleen, Survivor, Day 0 [V34S]

description: Replica 6


title: RUNX1EVI1-PlusDox-Pu1-ChIP


title: 7_1.4F_WT


title: 19_1.3M_WT


title: 12_7.8F_WT


title: TNF2B41: microglia TNF_rep10

city: Cambridge

first: Alex

last: Delbridge

state: MASSACHUSETTS

address: 225 binney street

institute: Biogen

data_processing:
Reads were aligned to the reference with STAR 2.7.0a and parameters --alignSJDBoverhangMin 1 --outFilterMismatchNoverLmax 0.05 --outFilterScoreMinOverLread 0.90 --outFilterMatchNmin  OverLread 0.90 --alignIntronMax 1000000
Reads were assigned to genes using "featureCounts" in the subread v1.6.4 package with parameters "--minOverlap 25"
Genome_build: mm10
Supplementary_files_format_and_content: tab-separated gene counts table

source_name: brain slice microglia, 2 weeks in culture

extract_protocol:
Microglia were isolated under ice cold conditions. Pure populations of microglia were obtained by cell sorting for live CD11b positive, CD45 intermediate cells. Sorted cells were collected directly into ice cold lysis buffer and RNA was isolated using the Qiagen RNeasy Micro RNA Kit. Flash frozen brain tissue samples were homogenized and RNA was isolated using the Qiagen RNeasy Mini RNA kit.
Takara SMART-Seq v4 Ultra Low Input RNA Kit + Illumina Nextera XT Library Prep Kit

value: brain slice microglia

tag: treatment

value: TNF 50 ng/mL, 24 hours

tag: immunophenotype

value: CD11b high; CD45 intermediate

tag: study id

tag: rna-seq protocol


title: Spleen, Survivor, Day 0 [V32S]

description: Replica 6


title: TNF2A31: microglia TNF_rep8

tag: tissue

value: Takara SMART-Seq v4 Ultra Low Input RNA Kit + Illumina Nextera XT Library Prep Kit


title: Spleen, Survivor, Day 0 [V31S]

description: Replica 6


title: 1_1.1F_VPS39ko

tag: strain


title: 8_1.5F_WT


title: 11_7.7F_WT


title: TNF2A26: microglia TNF_rep7


title: TNF2B46: microglia TNF_rep11


title: Spleen, Survivor, Day 0 [V29S]

description: Replica 6


title: TNF2B36: microglia TNF_rep9


title: TNF1B23: microglia TNF_rep6


title: TNF1B14: microglia TNF_rep4


title: TNF1A2: microglia TNF_rep2


title: media2B45: microglia medium control_rep9

value: medium only control, 24 hours


title: TNF1A12: microglia TNF_rep1


title: media2A25: microglia medium control_rep5


title: Spleen, Survivor, Day 0 [V26S]

description: Replica 6


title: media2B40: microglia medium control_rep8


title: Spleen, Survivor, Day 0 [V25S]

description: Replica 6


title: Spleen, Survivor, Day 0 [V24S]

description: Replica 6


title: Spleen, Survivor, Day 0 [V28S]

description: Replica 6


title: media1A1: microglia medium control_rep2


title: media1B13: microglia medium control_rep4


title: media1A6: microglia medium control_rep3


title: Spleen, Survivor, Day 0 [V22S]

description: Replica 6


title: media2A30: microglia medium control_rep6


title: Spleen, Survivor, Day 0 [V21S]

description: Replica 6


title: GMCSF2B42: microglia GMCSF_rep8

value: GMCSF 20 ng/mL, 24 hours


title: TNF1B19: microglia TNF_rep5


title: media1A11: microglia medium control_rep1


title: Spleen, Control, Day 11 [VC57S]

description: Replica 5

source_name: Time point Control

value: Uninfected


title: GMCSF2B47: microglia GMCSF_rep9


title: Spleen, Control, Day 11 [VC58S]

description: Replica 5


title: Spleen, Control, Day 11 [VC60S]

description: Replica 5


title: GMCSF2A32: microglia GMCSF_rep6


title: GMCSF2B37: microglia GMCSF_rep7


title: TNF1A7: microglia TNF_rep3


title: Spleen, Control, Day 11 [VC56S]

description: Replica 5


title: GMCSF1B15: microglia GMCSF_rep3


title: GMCSF1B20: microglia GMCSF_rep4


title: GMCSF1A8: microglia GMCSF_rep2


title: GMCSF2A27: microglia GMCSF_rep5


title: Spleen, Control, Day 11 [VC55S]

description: Replica 5


title: GMCSF1A3: microglia GMCSF_rep1


title: Runx1Eto_PlusDox_HE2_Rep2

data_processing:
Raw paired-end sequencing data were processed to remove low quality sequences and adaptors with Trimmomatic v0.32
The processed reads were then aligned to the mouse genome (version mm10) using Hisat2 v2.1.0 with default parameters
Gene expression was measured as raw counts  with FeatureCounts using gene models from RefSeq as the reference transcriptome
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited file of read counts

extract_protocol:
RNA was isolated from sorted cells using either the NucleoSpin RNA kit (Macherey-Nagel) or Trizol reagent (Thermo Fisher Scientific).
RNA-seq libraries were prepared from two biological replicates using the True-Seq stranded total RNA kit (Illumina) and sequenced paired-end in a pool of 12 indexed libraries using a Next-Seq 500/550 high output kit v2 150 cycles (Illumina)

treatment_protocol: ESCs were differentiated as previously described (Gilmour et al, 2014; Regha et al., 2015) with the following modifications. FLK1+ cells were purified by magnetic cells sorting, using biotin-conjugated CD309 antibody (eBioscience), anti-biotin microbeads (Miltenyi Biotec) and LS columns (Miltenyi Biotec) following culture of embryoid bodies for between 3.25 and 3.75 days (cell line dependent). These FLK1+ cells were then cultured in gelatin-coated flasks – 1.2-1.4x10e6 cells in a T150 flask to form the blast culture. After 1-2 days (cell line dependent), 0.1-0.5 µg/ml doxycycline was added as appropriate and cells were cultured in the same media for a further 18 hours prior to sorting for HE and Progs. Cell populations were identified and sorted on day 2-3 of blast culture based on surface markers. Cells were stained with KIT-APC (BD pharmingen), Tie2-PE (eBioscience) and CD41-PE-Cy7 (eBioscience) and analyzed on a Cyan ADP flow cytometer (Beckman Coulter) with data analysis using FlowJo, or sorted on a FACS Aria cell sorter (BD Biosciences).

value: A2lox ESCs

tag: sorted cells


title: Runx1Eto_PlusDox_HP_Rep2

value: HP


title: Runx1Eto_PlusDox_HE2_Rep1


title: Spleen, Control, Day 11 [VC51S]

description: Replica 5


title: Runx1Eto_NoDox_HP_Rep1


title: Runx1Eto_PlusDox_HP_Rep1


title: Runx1Eto_NoDox_HP_Rep2


title: R204X_PlusDox_HP_rep2


title: Spleen, Control, Day 11 [VC52S]

description: Replica 5


title: R204X_PlusDox_HP_rep1


title: Runx1Eto_NoDox_HE2_Rep2


title: Runx1Eto_NoDox_HE2_Rep1


title: Spleen, Control, Day 11 [VC49S]

description: Replica 5


title: Spleen, Control, Day 11 [VC53S]

description: Replica 5


title: R204X_PlusDox_HE2_rep1


title: Spleen, Control, Day 11 [VC46S]

description: Replica 5


title: R204X_PlusDox_HE2_rep2


title: R204X_NoDox_HP_rep2


title: Spleen, Control, Day 11 [VC45S]

description: Replica 5


title: R204X_NoDox_HE2_rep1


title: R204X_NoDox_HE2_rep2


title: R201Q_PlusDox_Prog_Rep1

value: Prog


title: R201Q_PlusDox_HE2_Rep2


title: Spleen, Control, Day 11 [VC43S]

description: Replica 5


title: Spleen, Control, Day 11 [VC41S]

description: Replica 5


title: R201Q_NoDox_Prog_Rep2


title: R201Q_PlusDox_HE2_Rep1


title: R201Q_NoDox_Prog_Rep1


title: Runx1Evi1_plusDox_H3K27ac_976

data_processing:
Raw single-end reads were processed with Trimmomatic v0.32.
The processed reads were aligned to the mouse genome (version mm10) using Bowtie2 v2.2.3 with the option --very-sensitive-local
Peaks were identified using MACS v2.1.1 with the options -g mm --keep-dup all -B --trackline
Genome_build: mm10
Supplementary_files_format_and_content: bedGraph files of sequence read pileups generated by MACS2

tag: chip antibody

value: Abcam ab4729


title: R201Q_NoDox_HE2_Rep1


title: Spleen,  Clinical Sign, Day 3 [V19S]

description: Replica 4

source_name: Clinical Sign

value: Vibrio anguillarum

value: Day 3


title: Spleen,  Clinical Sign, Day 3 [V20S]

description: Replica 4


title: R204X_NoDox_HP_rep1


title: Spleen,  Clinical Sign, Day 3 [V17S]

description: Replica 4


title: R201Q_PlusDox_Prog_Rep2


title: Spleen, Control, Day 11 [VC42S]

description: Replica 5

value: Day 11


title: Runx1Evi1_noDox_H3K27ac_974


title: Spleen,  Clinical Sign, Day 3 [V15S]

description: Replica 4


title: R204X_plusDox_H3K27ac_1070_1316


title: Runx1Eto_noDox_H3K27ac_1317_1352


title: Spleen,  Clinical Sign, Day 3 [V14S]

description: Replica 4


title: R204X_noDox_H3K27ac_1069_1317


title: R201Q_plusDox_H3K27ac_1072


title: R201Q_noDox_H3K27ac_1071


title: Spleen,  Clinical Sign, Day 3 [V13S]

description: Replica 4


title: R201Q_NoDox_HE2_Rep2


title: R204X_plusDox_Runx1_ChIP_1026_1794_GB140819

value: Abcam ab23980


title: R204X_noDox_Runx1_ChIPseq_1025


title: Spleen,  Clinical Sign, Day 3 [V18S]

description: Replica 4


title: Spleen,  Clinical Sign, Day 3 [V11S]

description: Replica 4


title: Runx1Eto_plusDox_H3K27ac_1318_1351


title: R201Q_noDox_Runx1_ChIPseq_998


title: Runx1-Evi1_PlusDox

data_processing:
Raw single-end reads were processed with Trimmomatic v0.32
The processed reads were aligned to the mouse genome (version mm10) using Bowtie2 v2.2.3 with the option --very-sensitive-local.
The resulting alignments were filtered to remove potentially PCR duplicated reads using the MarkDuplicates function in Picard v2.10.5
Peaks corresponding to regions of open chromatin were identified using MACS2 v2.1.1 with the options -g mm --keep-dup all -B --trackline --nomodel --shift -100 --extsize 200.
Genome_build: mm10
Supplementary_files_format_and_content: bedGraph files of sequence read pileups generated by MACS2

extract_protocol:
ATAC-seq was performed essentially as described (Buenrostro et al, 2015), briefly, 50,000 KIT+ CD41+ Tie2- Progs were transposed in 1x Tagment DNA buffer (Illumina), Tn5 transposase (Illumina) and 0.01% Digitonin (Promega) for 30 minutes at 37°C with agitation.
DNA was amplified by PCR using Nextera primers and libraries were sequenced using a Next-Seq 500 high output kit v2 75 cycles (Illumina).


title: Runx1-Evi1_NoDox


title: Spleen,  Clinical Sign, Day 3 [V9S]

description: Replica 4


title: Spleen,  Clinical Sign, Day 3 [V10S]

description: Replica 4


title: Runx1-Eto_PlusDox


title: Spleen,  Clinical Sign, Day 3 [V8S]

description: Replica 4


title: Runx1-Eto_NoDox


title: R204X_PlusDox


title: R204X_NoDox


title: R201Q_PlusDox


title: Spleen,  Clinical Sign, Day 3 [V6S]

description: Replica 4


title: R201Q_NoDox


title: Spleen,  Clinical Sign, Day 3 [V5S]

description: Replica 4


title: media2B35: microglia medium control_rep7


title: Time_course_day4

first: Elena

last: Torlai Triglia

state: MA

address: 415 Main Street

institute: Broad Institute

description:
Single-cell RNA-Seq (10X).
I1 fastq file contains sample barcode.
R1 fastq file contains cell barcodes.
R2 fastq file contains cDNA.
processed data file: RNA_timecourse_unadjusted_barcodes.tsv
processed data file: RNA_timecourse_unadjusted_genes.tsv
processed data file: RNA_timecourse_unadjusted_matrix.mtx.gz
processed data file: RNA_timecourse_sctransform_barcodes.tsv
processed data file: RNA_timecourse_sctransform_genes.tsv
processed data file: RNA_timecourse_sctransform_count_data_matrix.mtx.gz
processed data file: RNA_timecourse_sctransform_norm_data_matrix.mtx.gz
processed data file: RNA_timecourse_sctransform_scale_data_matrix.mtx.gz
processed data file: RNA_timecourse_sctransform_variable_genes.tsv
fluorescent_proteins.fa

data_processing:
scRNA-seq: To incorporate the fluorescent reporters into the reference genome for alignment, we added the sequences of Yfp, tdTomato, Gfp, and Bfp (reporting history of Il5 expression, and current expression of Il5, Il17a, and Il22, respectively) to the mm10 reference genome provided by CellRanger (v1.3). Yfp sequence was obtained by Sanger sequencing as described above. We did check that Yfp counts appeared relatively independent of whether Gfp and Bfp were in the reference. TdTomato was not positioned close enough to the 3' end of the transcript to be mapped robustly, and Il5 has relatively low expression; hence, we used dial-out experiments with Cre to assess expression in the scRNA-seq libraries. Exact Gfp and Bfp transcript sequences were unknown, so Gfp was represented in the reference by the "eGFP" and "bGH" terminator in the pBigT-IRES-GFP plasmid (https://www.addgene.org/browse/sequence/229542/), and the Bfp sequence was obtained from Gfp by substituting the sequence of BFP (https://www.addgene.org/browse/sequence_vdb/6363/) for eGFP. We could not confidently map reads to Gfp or Bfp, likely due, at least partly, to the large similarity in those sequences; hence, Il17a and Il22 mapped transcripts were used to quantify transcriptional expression of those genes, without any reporter transcripts. We did not use counts from tdTomato, Gfp, or Bfp fluorescent proteins in the downstream scRNA-seq analysis.   Gene counts were obtained by aligning reads to the modified mm10 genome using CellRanger software (v1.3) (10x Genomics). Counts for Cre were obtained by dial-out, using the dedup tool from UMI-tools v1.0.086 to remove PCR duplicates, with the --per-cell flag and the default method. Deduplicated reads aligned to Cre were then tallied for each cell.  To remove doublets and poor-quality cells, cells were excluded from subsequent analysis if they were outliers in their sample of origin in terms of number of genes, number of Unique Molecular Identifiers (UMIs), or their joint distribution, retaining 94% (6,059), 96.6% (6,200), 91.2% (4,272), 96.7% (6,645), and 92.8% (5,051) of cells per time points 0–4, respectively. Cut-offs for number of genes detected per cell were 500–3,000 for time points 0 and 1, and 700–3,300 for time points 2–4. Cut-offs for number of UMIs detected per cell were 1,000–10,000 for time points 0 and 1, and 1,500–11,000 for time points 2–4. Cells were further excluded if they were outliers across remaining cells in all samples in terms of percentage of mitochondrial genes (>15%), eliminating another 3.7% of the remaining cells, resulting in 27,185 retained high-quality cell profiles.   To normalize gene counts, we used the “SCTransform” function from Seurat v3, setting the “vars.to.regress” parameter to the percentage of mitochondrial genes and defaults for other parameters.  Variable gene selection was performed automatically within the call to SCTransform via selection of the top 3,000 genes, ranked by residual variance.  To cluster and remove cells unlikely to be ILCs, we computed a Principal Components Analysis (PCA) on scaled variable genes, as determined above, using Seurat’s “RunPCA” function, and visualized it by computing a Uniform Manifold Approximation and Projection (UMAP) using Seurat’s “RunUMAP” function on the top 30 PCs. We also computed the k-nearest neighbor graph on the top 30 PCs, using Seurat’s “FindNeighbors” function with default parameters (i.e., “k.param” for the number of nearest neighbors set to 20), and in turn used Seurat’s “FindClusters” function with “algorithm” set to “leiden” to compute clusters based on the graph.  Based on expression of marker genes across clusters, we determined that two very distinct clusters were unlikely to be ILCs. Cells from one small, distinct cluster of 109 cells had unique, very high expression of genes associated with the extracellular matrix and collagen, including Sparc, Dcn, Gcn, Bgn, Col3a1, and Col1a2. Another, even smaller distinct cluster of 8 cells had very specific, high expression of Lyz2. Cells from these putative non-ILC clusters were removed, and the QC and normalization steps were repeated.  The result of this filtering step can be found in the following files: those prefixed with RNA_timecourse_unadjusted_ contain the count data that is not processed by scTrainsform (mtx matrix with genes in rows, barcodes in columns); those prefixed with RNA_timecourse_sctransform_ contain scTransform data in the layers "scale_data" (Pearson residuals), "count_data" (adjusted count data), and "norm_data" (log1p transformed adjusted count data) as mtx matrices with genes in rows, barcodes in columns. RNA_timecourse_sctransform_variable_genes.tsv contains a list of genes selected as variable by scTransform.
scATAC-seq: Fastq files were demultiplexed from the sequencer base call files using CellRanger ATAC mkfastq (v1.1.0) from 10x Genomics. Peak-barcode matrices were obtained by aligning reads to mm10 (CR v1.1.0 pre-built reference) using CellRanger ATAC count.
Genome_build: mm10, with modification described above.
Supplementary_files_format_and_content: RNA_timecourse_unadjusted_barcodes.tsv: RNA tsv with timepoint and barcode sequences of counts (columns).
Supplementary_files_format_and_content: RNA_timecourse_unadjusted_genes.tsv: RNA tsv with gene symbols of counts (rows).
Supplementary_files_format_and_content: RNA_timecourse_unadjusted_matrix.mtx.gz: RNA mtx file with count matrix, genes in rows, barcodes in columns, gzip compressed.
Supplementary_files_format_and_content: RNA_timecourse_sctransform_barcodes.tsv: RNA tsv with timepoint and barcode sequences for sctransform data (columns).
Supplementary_files_format_and_content: RNA_timecourse_sctransform_genes.tsv: RNA tsv with gene symbols (rows) of counts for sctransform data.
Supplementary_files_format_and_content: RNA_timecourse_sctransform_count_data_matrix.mtx.gz: RNA mtx file with count matrix, adjusted by sctransform, genes in rows, barcodes in columns, gzip compressed.
Supplementary_files_format_and_content: RNA_timecourse_sctransform_norm_data_matrix.mtx.gz: RNA mtx file with log1p transformed count matrix, adjusted by sctransform, genes in rows, barcodes in columns, gzip compressed.
Supplementary_files_format_and_content: RNA_timecourse_sctransform_scale_data_matrix.mtx.gz: RNA mtx file with Pearson residual matrix from sctransform, genes in rows, barcodes in columns, gzip compressed.
Supplementary_files_format_and_content: RNA_timecourse_sctransform_variable_genes.tsv: RNA tsv with gene symbols of 3000 most variable genes.
Supplementary_files_format_and_content: ATAC_naive_barcodes.tsv: CellRanger ATAC tsv with barcode sequences.
Supplementary_files_format_and_content: ATAC_naive_fragments.tsv.gz: CellRanger ATAC bed-like tabular file with ATAC-seq fragments.
Supplementary_files_format_and_content: ATAC_naive_fragments.tsv.gz.tbi: CellRanger ATAC tabix index of the fragment interval.
Supplementary_files_format_and_content: ATAC_naive_matrix.mtx: CellRanger ATAC raw peak-barcode matrix.
Supplementary_files_format_and_content: ATAC_naive_peaks.bed: CellRanger ATAC peak bed file.
Supplementary_files_format_and_content: ATAC_naive_peak_annotation.tsv: CellRanger ATAC peak annotation file.
Supplementary_files_format_and_content: ATAC_IL23_barcodes.tsv: CellRanger ATAC tsv with barcode sequences.
Supplementary_files_format_and_content: ATAC_IL23_fragments.tsv.gz: CellRanger ATAC bed-like tabular file with ATAC-seq fragments.
Supplementary_files_format_and_content: ATAC_IL23_fragments.tsv.gz.tbi: CellRanger ATAC tabix index of the fragment interval.
Supplementary_files_format_and_content: ATAC_IL23_matrix.mtx: CellRanger ATAC raw peak-barcode matrix.
Supplementary_files_format_and_content: ATAC_IL23_peaks.bed: CellRanger ATAC peak bed file.
Supplementary_files_format_and_content: ATAC_IL23_peak_annotation.tsv: CellRanger ATAC peak annotation file.
Supp

source_name: FACS-purified total mouse skin ILC

extract_protocol:
Skin ILCs were isolated from ears of IL-5 Fate IL-17AGFP and IL-22BFP mouse reporters from naïve mice (day 0) and at daily intervals thereafter along a four-day time course of IL-23 treated mice (1,2,3, and 4 days post injection; n=5 mice per group). Total skin ILCs were sorted as CD45.2, CD90.2 positive, and lineage, TCRgamma and CD3epsilon negative. Sorted cells were washed with PBS/0.04% BSA and processed for droplet-based 3′ end massively parallel scRNA-seq: sorted ILCs were encapsulated into droplets, and libraries were prepared using Chromium Single Cell 3′ Reagent Kits V3 according to the manufacturer’s protocol (10X Genomics). scRNA-seq libraries were sequenced using a 75 cycle Nextseq 500 High Output kit.   To quantify Cre, as a proxy for Il5 expression, a Cre-specific primer was selected ~200bp upstream from the polyA signal motif (AAUAAA) for the Cre that was used to generate the mouse in this study. Primers were ordered from Genewiz. Cre primer 5’- GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTGGTGTGTCCATCCCTGAAAT -3’, P7-index primer: 5’- CAAGCAGAAGACGGCATACGAGAT[index]GTCTCGTGGGCTCGGAGATGTG -3’, P5 primer: 5'- AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTC -3'  PCR was conducted with 20μl reaction consisting of 2μl dilute template, 2μl of 10μM P5 primer, 2μl of 1μM of Cre-primer, 2μl of 10μM P7 index primer, 2μl H2O and 10μl of Q5® High-Fidelity 2X Master Mix (New England Biolabs) using the following PCR program:: 98°C for 30sec, 22 cycles of 98°C for 10seq, 68°C for 30sec, and 72°C for 30sec followed by 72°C for 50sec and hold at 12°C. Libraries were then pooled equally by volume, purified by MinElute (Qiagen), and eluted in 35μl double distilled water. Purified libraries were loaded on Blue Pippin for size selection of 400-520nt. Tapestation D1000 was used for quality control after size selection. Qubit HS DNA was used to quantify the library. The library was sequenced on an Illumina MiSeq with a V3 150 cycle kit with the following setting: Read1: 28; Read2: 125; I7 Index: 8; I5 Index: 0  YFP was amplified from 10x WTA mixture with P5 primer and Yfp-primer, followed by Sanger Sequencing with P5 and YPF primer. The Sanger sequencing result was then compared to the vector sequence used in the generation of the mice.  Yfp-primer: GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTCCTCGTGACCACCTTCGG
10x RNAseq (3', V3)

value: skin

value: innate lymphoid cells

tag: time point


title: Time_course_day2


title: Time_course_day3


title: Spleen,  Clinical Sign, Day 3 [V4S]

description: Replica 4

value: Spleen


title: Spleen,  Clinical Sign, Day 3 [V3S]

description: Replica 4


title: R201Q_plusDox_Runx1_ChIPseq_1728_1790


title: LH_9_KO

city: Dallas

first: Maria

last: Chahrour

state: Texas

address: 6000 Harry Hines Blvd

institute: University of Texas Southwestern Medical Center

data_processing:
Raw data from the machine were de-multiplexed and converted to fastq files using bcl2fastq (v2.17, Illumina).
The fastq files were checked for quality using fastqc (v0.11.2) and fastq_screen (v0.4.4).
Reads were mapped to mm10 reference Mouse genome using STAR. Read counts were then generated using featureCounts.
TMM normalization and differential expression analysis was performed using edgeR.
Genome_build: mm10
Supplementary_files_format_and_content: differential gene expression analysis

source_name: Kdm5a -/-_hippocampus

extract_protocol:
RNA was extracted from hippocampal tissue of 10 animals (5 WT and 5 Kdm5a -/-). Samples were run on the Agilent Tapestation 4200 to assess quality and those with a RIN score of ≥ 8 were used for sequencing. Concentrations were measured on a Qubit fluorometer.
For each sample, 1μg of total DNase-treated RNA was used for library preparation using the TruSeq Stranded mRNA Library Prep Kit from Illumina. Poly-A RNA was purified, fragmented, and strand specific samples were amplified and purified with AmpureXP beads.

tag: strain background

tag: genotype/variation

value: Kdm5a -/-

tag: age

value: 6-8 weeks

value: Hippocampus


title: Time_course_day0


title: 20_1.5M_WT

value: wild type

tag: Sex

value: male


title: Spleen, No Clinical Sign, Day 3 [VT14-10S]

description: Replica 3

source_name: No Clinical Sign

tag: organ

tag: pathogen

tag: time


title: LH_8_KO


title: ATAC_Bach2gRNA_P14_Cl13_D7_StemLike_2

tag: sorting marker

tag: culture condition

value: transduced with Bach2 gRNA construct

tag: infection

value: 7 days after LCMV Cl13 infection


title: LH_7_KO


title: ATAC_Bach2gRNA_P14_Cl13_D7_StemLike_1


title: Spleen, No Clinical Sign, Day 3 [VT14-7S]

description: Replica 3


title: Spleen, No Clinical Sign, Day 3 [VT14-8S]

description: Replica 3


title: Spleen, No Clinical Sign, Day 3 [VT14-6S]

description: Replica 3


title: ATAC_pMK9_P14_Cl13_D7_StemLike_2

value: transduced with pMK9


title: LH_6_KO


title: Time_course_day1

tag: cell type

tag: treatment


title: LH_5_WT

value: Wild type

tag: tissue


title: Spleen, No Clinical Sign, Day 3 [VT14-4S]

description: Replica 3


title: LH_10_KO


title: LH_4_WT


title: LH_3_WT


title: Spleen, No Clinical Sign, Day 3 [VT14-3S]

description: Replica 3


title: ATAC_pMK9_P14_Cl13_D7_StemLike_1

tag: strain


title: ATAC_BACH2OE_P14_Cl13_D7_StemLike_2

value: transduced with BACH2 over expression construct


title: ATAC_pMIG_P14_Cl13_D7_StemLike_2

value: transduced with pMIG


title: ATAC_BACH2OE_P14_Cl13_D7_StemLike_1


title: Spleen, No Clinical Sign, Day 3 [VT14-1S]

description: Replica 3

value: Vibrio anguillarum

value: Day 3


title: Spleen, No Clinical Sign, Day 3 [VT14-2S]

description: Replica 3


title: Spleen, No Clinical Sign, Day 3 [VT13-10S]

description: Replica 3


title: ATAC_pMIG_P14_Cl13_D7_StemLike_1


title: Spleen, No Clinical Sign, Day 3 [VT13-9S]

description: Replica 3


title: P30_3

city: boston

first: Yuxuan

last: Guo

address: 320 Longwood Ave, Enders 1250,

department: Department of Cardiology

institute: Boston childrens hospital

data_processing:
Basecalls performed using CASAVA version 1.8
Sequenced reads were aligned to mm10 whole genome by STAR 2.5.3a default setting
Read counts were calculated by featureCounts of Subread 1.5.2
Genome_build: mm10
Supplementary_files_format_and_content: Gene reads counts matrix generated by FeatureCounts
Supplementary_files_format_and_content: Differential expression analysis between P6 and P30 data by DESeq2

source_name: cardiomyocyte

extract_protocol:
total RNA was extracted from FACS sorted cells using PureLink™ RNA Micro Scale Kit (ThermoFisher)
10ng total RNA was reverse transcribed, cDNA further amplified by 8 PCR cycles using SMART-Seq v4 Ultra Low Input RNA Kit (Clontech). 500pg cDNA was next used to generate RNA-seq libraries using Nextera XT DNA Library Preparation Kit (Illumina)

treatment_protocol: 5×10^8 vg/g AAV9-cTNT-Cre virus was injected into P1. Cells were collected at P14 through retrograde collagenase perfusion followed by fluorescence activated cell sorting

tag: genotype

value: postnatal day 30

tag: aav injection time

value: postnatal day 1


title: LH_1_WT


title: P30_2


title: ATAC_IL23

description: Single-cell ATAC-Seq (10X).

extract_protocol:
Skin ILCs were isolated from ears of WT naïve mice (day 0) and rIL-23 treated mice (day 4) with n=10 mice per group. Total skin ILCs were sorted (approximately 60,000 cells per group) as CD45.2, CD90.2 positive, lineage, TCRgamma and CD3epsilon negative. After sorting, cells were immediately washed and resuspended in 1ml of CryoStore CS10 freezing media (StemCell Technologies) followed by cryopreservation at -80°C with cooling rate of 1°C/min using Mr. Frosty (Nalgene). For scATAC-seq cells were thawed, washed and lysed for 3 min on ice according to the low input protocol recommendations by 10x Genomics (CG000169-Rev C). For single cell library preparation, the 10x Genomics Chromium Controller and the 10x Genomics Chromium Single Cell ATAC Library & Gel Bead Kit (#1000111) were used, according to the manufacturer’s instructions (CG000168-Rev B).
10x ATACseq v1.0

value: skin

value: innate lymphoid cells

tag: time point


title: P30_1


title: Spleen, No Clinical Sign, Day 3 [VT13-6S]

description: Replica 3

value: Spleen


title: ATAC_naive

description: Single-cell ATAC-Seq (10X).


title: GFP_5

data_processing:
Basecalls performed using CASAVA version 1.8
Sequenced reads were aligned to mm10 whole genome by STAR 2.5.3a default setting
Read counts were calculated by featureCounts of Subread 1.5.2
Genome_build: mm10
Supplementary_files_format_and_content: Gene reads counts matrix generated by FeatureCounts

treatment_protocol: 5×10^8 vg/g AAV9-cTNT-dnMKL1-GFP or AAV9-cTNT-GFP virus was injected into P1. Cells were collected at P14 through retrograde collagenase perfusion followed by fluorescence activated cell sorting

value: Wildtype

value: postnatal day 14

value: cardiomyocyte


title: Spleen, No Clinical Sign, Day 3 [VT13-4S]

description: Replica 3


title: dnMKL1_5


title: P6_4

value: postnatal day 6


title: Spleen, No Clinical Sign, Day 3 [VT13-3S]

description: Replica 3


title: dnMKL1_4


title: P6_3


title: LH_2_WT

tag: strain background

value: 6-8 weeks

value: Hippocampus


title: Spleen, No Clinical Sign, Day 3 [VT13-2S]

description: Replica 3


title: P6_2


title: Spleen, No Clinical Sign, Day 3 [VT13-1S]

description: Replica 3


title: GFP_4


title: P6_1


title: dnMKL1_3


title: Hmga2-/- Ctrl SSRP1

city: Nancy Cedex

first: Guillermo

last: Barreto

address: 9, Avenue de la Forêt de Haye ‐ BP 20199

department: Campus Biologie-Santé. Faculté de Médecine

country: France

institute: CNRS Director of Research

data_processing:
Visualization of raw reads was done using FastQC, trimmed of the raw reads using Trimmomatic (Bolger et al. 2014), using the following parameters LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 HEADCROP:4 MINLEN:15
High quality reads were aligned to the mm10 genome by using bowtie2 with default parameters.
Sam files were converted to bam files using samtools view and sort.
ChIPs were compared to their coresponding inputs  using deeptool's bamCompare. A Bed-File containing UCSC known genes were downloaded using the Table Browser from UCSC Genome Browser and used as region-file. ChIP and Input reads were scaled using the scaleFactorsMethod command and Input reads were substracted from ChIP reads.
Genome_build: mm10
Supplementary_files_format_and_content: BigWig files were generated using deeptool's bamCompare; Scores represent compared values.

source_name: Mouse embryonic fibroblasts

growth_protocol: Immortalized MEFs were cultured at 37 °C in 5 % CO2 in DMEM medium with 4.5 g/l glucose, 10 % FCS 4 mM L-Glutamine, 1 mM Pyruvate, 100 U/ml penicillin and 100 U/ml streptomycin.

extract_protocol:
Cells were cross-linked with 1% formaldehyde for 10 min at room temperature. The reaction was quenched for 5 min on ice by adding 0.125 M glycine. After three washes with 1x PBS, cells were subjected to hypotonic buffer L1 (50 mM Tris-HCl pH 8.0, 2 mM EDTA pH 8.0, 0.1% IGEPAL CA-630 (Sigma), 10% glycerol, 2 mM DTT, 10 mM NaF, 2 mM Na3VO4, 0.2 M imidazole, 1 mM PMSF and protease inhibitor cocktail). Cells were lysed, and sonicated with Diagenode Bioruptor to an average DNA length of 100-300 bp. After centrifugation, the soluble chromatin was immunoprecipitated using antibodies specific against p H2A.X (Millipore), SUPT16H (CellSignaling) and SSRP1 (BioLegend). Precipitated chromatin complexes were removed from the beads and decrosslinked by incubating with 100 μl of 10 mM Tris-HCl pH 8.0, 1 mM EDTA pH 8.0, 0.1 M NaHCO3, 1% SDS and Proteinase K at 65 °C overnight. Reverse cross-linked immunoprecipitated chromatin was purified using the QIAquick PCR purification kit (Qiagen).
Precipitated DNA samples were purified by QIAquick PCR purification kit (Qiagen) and quantified by Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific). 2ng of DNA was used as input for TruSeq ChIP Library Preparation Kit (Illumina) with following modifications. Instead of gel-based size selection before final PCR step, libraries were size selected by SPRI-bead based approach after final PCR with 18 cycles. In detail, samples were 1st cleaned up by 1x bead:DNA ratio to eliminate residuals from PCR reaction, followed by 2-sided-bead cleanup step with initially 0.6x bead:DNA ratio to exclude larger fragments. Supernatant was transferred to new tube and incubated with additional beads in 0.2x bead:DNA ratio for eliminating smaller fragments, like adapter and primer dimers. Bound DNA samples were washed with 80% ethanol, dried and resupended in TE buffer. Library integrity was verified with LabChip Gx Touch 24 (Perkin Elmer).

treatment_protocol: MEFs were starved overnight in DMEM medium containing 1 % FCS and then treated with 10 ng/ml human recombinant TGFB1 (Sigma Aldrich) for 3 h.

value: Immortalized Mouse embryonic fibroblasts

value: whole embryo

value: embryonic day 15.5

value: Hmga-/-

tag: cell extraction

value: Timed-pregnant mice were sacrificed on post coitum day 15.5  according to standard methods. The embryos were collected in sterile PBS complemented with 100 U\ml penicillin and 100 U\ml streptomycin. After removing the head, limbs, tail, viscera, liver and heart, the remaining body was trypsinized with 1x trypsin-EDTA solution (0.05 %) for 20 minutes at 37°C and single cells were obtained via physical disruption by sterile Pasteur pipet. The cells from each embryo were diluted with growth medium containing FCS and plated separately.

tag: antibody

value: SSRP1 (Biolegend, 609710)


title: Spleen, Control, Day 3 [VC39S]

description: Replica 2

value: Uninfected

value: Day 3


title: Spleen, Control, Day 3 [VC40S]

description: Replica 2


title: GFP_2

tag: age

value: postnatal day 1


title: dnMKL1_2


title: Hmga2+/+ Ctrl SSRP1

value: wild type


title: Hmga2-/- Ctrl SUPT16H

value: SPT16 (Cell Siganling, 12191)


title: Spleen, Control, Day 3 [VC38S]

description: Replica 2

tag: organ

value: Spleen


title: Spleen, Control, Day 3 [VC37S]

description: Replica 2

tag: pathogen


title: Hmga2+/+ Ctrl SUPT16H


title: dnMKL1_1

value: Wildtype

value: postnatal day 14


title: Hmga2-/- TGFB1 pH2AX

value: pH2A.XSer139(Millipore, 05-636, clone JBW301)


title: GFP_1

value: cardiomyocyte


title: Spleen, Control, Day 3 [VC35S]

description: Replica 2

tag: time


title: Hmga2-/- Ctrl pH2AX


title: Actn2KO5

treatment_protocol: 3.6×10^8 vg/g AAV9-cTNT-Cre virus was injected into P1. Cells were collected at P14 through retrograde collagenase perfusion followed by fluorescence activated cell sorting


title: Hmga2+/+ TGFB1 pH2AX


title: Hmga2+/+ Ctrl pH2AX


title: Actn2KO4


title: Spleen, Control, Day 11 [VC50S]

description: Replica 5

value: Day 11


title: Hmga2-/- TGFB1 pH2AX input

value: Immortalized Mouse embryonic fibroblasts


title: GFP_3


title: Spleen, Control, Day 3 [VC30S]

description: Replica 2


title: Actn2HET4


title: Hmga2-/- Ctrl pH2AX input

value: whole embryo


title: Spleen, Control, Day 3 [VC29S]

description: Replica 2


title: Actn2KO3


title: Spleen, Control, Day 3 [VC28S]

description: Replica 2


title: Hmga2+/+ TGFB1 pH2AX input

value: embryonic day 15.5


title: Hmga2+/+ Ctrl pH2AX input


title: Actn2HET3


title: Spleen, Control, Day 3 [VC36S]

description: Replica 2


title: Actn2KO2


title: Spleen, Control, Day 3 [VC25S]

description: Replica 2


title: Spleen, Control, Day 3 [VC27S]

description: Replica 2


title: Actn2HET2


title: Actn2HET5


title: Hmga2-/- Ctrl

data_processing:
Visualization of raw reads was done using FastQC, trimmed of the raw reads using Trimmomatic (Bolger et al. 2014), using the following parameters LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 HEADCROP:5 MINLEN:15
High quality reads were mapped by using with HISAT2 v2.1.0 (Kim et al. 2015) with reads corresponding to the transcript with default parameters. RNA-seq reads were mapped to mouse genome mm10.
Sam files were converted to bam files using samtools view and sort.
analyzeRepeats.pl (http://homer.salk.edu/homer/ngs/analyzeRNA.html) with the parameters (analyzeRepeats.pl database.gtf hg19 -count genes -strand + and -rpkm; reads per kilobase per millions mapped). A gtf file for UCSC known genes was downloaded using the Table Browser from UCSC Genome Browser and was used to quantify RNA.
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited text files include RPKM values for each sample.

source_name: Immortalized Mouse embryonic fibroblasts

extract_protocol:
Total RNA was isolated using Trizol (Invitrogen). RNA was treated with DNase (DNase-Free DNase Set, Qiagen) and repurified using the miRNeasy micro plus Kit (Qiagen). Total RNA was quantified using a Nanodrop Spectrophotometer (ThermoFisher Scientific, Germany).
Total RNA and library integrity were verified on LabChip Gx Touch 24 (Perkin Elmer). 1µg of total RNA was used as input for SMARTer Stranded Total RNA Sample Prep Kit - HI Mammalian (Clontech). Sequencing was performed on the NextSeq500 instrument (Illumina) using v2 chemistry with 1x75bp single end setup.

treatment_protocol: MEFs were starved overnight in DMEM medium containing 1 % FCS and then treated with 10 ng/ml human recombinant TGFB1 for 24 h.

value: Mouse embryonic fibroblasts

value: Hmga-/-


title: Spleen, Control, Day 3 [VC33S]

description: Replica 2


title: Spleen, Control, Day 3 [VC24S]

description: Replica 2


title: Spleen, Control, Day 3 [VC32S]

description: Replica 2


title: Actn2KO1


title: Spleen, Control, Day 3 [VC23S]

description: Replica 2


title: Actn2HET1


title: Spleen, Control, Day 0 [VC20S]

description: Replica 1

value: Day 0


title: Hmga2+/+ Ctrl

value: Timed-pregnant mice were sacrificed on post coitum day 15.5  according to standard methods. The embryos were collected in sterile PBS complemented with 100 U\ml penicillin and 100 U\ml streptomycin. After removing the head, limbs, tail, viscera, liver and heart, the remaining body was trypsinized with 1x trypsin-EDTA solution (0.05 %) for 20 minutes at 37°C and single cells were obtained via physical disruption by sterile Pasteur pipet. The cells from each embryo were diluted with growth medium containing FCS and plated separately.


title: Fraction 4, Hmga2-/-

data_processing:
Visualization of raw reads was done using FastQC, trimmed of the raw reads using Trimmomatic (Bolger et al. 2014), using the following parameters LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 HEADCROP:5 MINLEN:15
High quality reads were aligned to the mm10 genome by using bowtie2 with default parameters.
Sam files were converted to bam files using samtools view and sort.
Bigwig files were generated with fragment sizes between 100 and 200 bp using deeptool's bamCompare (--operation log2 --pseudocount 1 --effectiveGenomeSize 2652783500 --normalizeUsing RPKM --ignoreDuplicates --centerReads --scaleFactorsMethod None --minFragmentLength 100 --maxFragmentLength 200). A Bed-File containing UCSC known genes were downloaded using the Table Browser from UCSC Genome Browser and used as region-file.  Data are represented as log2 enrichment over the corresponing inputs.
Genome_build: mm10
Supplementary_files_format_and_content: BigWig files were generated using deeptool's bamCompare; Scores represent compared values.

extract_protocol:
The nuclei were resuspended in Low Salt Buffer (10 mM Tris-HCl pH 7.4, 0.2 mM MgCl2 supplemented with protease and phosphatase inhibitors) including 1% Triton-X100. After 15 min incubation on ice, cells were spun down and the chromatin was resuspended in MNase digestion buffer with 1,250 Units MNase (NEB Biolabs). Chromatin-MNase mix was incubated at 37°C for 30 min. MNase reaction was stopped by adding 50 mM EDTA. Samples were sonicated for 30 sec on / 30 sec off using the Bioruptor with high amplitude. Chromatin was spun down at 14,000 rpm at 4°C for 10 min and the supernatant was used for ultracentrifugation. Sucrose gradients (5%-40%) were prepared in 1,800 μl low salt buffer (10 mM NaCl, 10 mM Tris-HCl pH7.4, 0.2 mM EDTA, 0.2 mM DTT, 20 mM NaF, 20 mM Na3VO4, 40 μg/ml phenylmethylsulfonyl fluoride and protease inhibitor) in polyallomer centrifuge tube (Beckman). Fragmented native chromatin was loaded on top of the 9 ml 5%-40% sucrose gradients and centrifuged for 16 h and 30 min at 37,500 rpm in an SW50.1 ultracentrifuge rotor (Beckman Coulter). Following centrifugation, 11 fractions (1000 μl each) were collected manually from the bottom of the tubes. From fractions 3 and 4, 200 µl  were treated with RNase A and purified using Phenol:Chloroform.
Purified DNA was quantified by Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific). 10 ng DNA was used as input for TruSeq ChIP Library Preparation Kit (Illumina) with following modifications. Instead of gel-based size selection before final PCR step, libraries were size selected by SPRI-bead based approach after final PCR with 18 cycles. In detail, samples were first cleaned up by 1x bead:DNA ratio to eliminate residuals from PCR reaction, followed by 2-sided-bead cleanup step with initially 0.6x bead:DNA ratio to exclude larger fragments. Supernatant was transferred to new tube and incubated with additional beads in 0.2x bead:DNA ratio for eliminating smaller fragments, like adapter and primer dimers. Bound DNA samples were washed with 80% ethanol, dried and resupended in TE buffer. Library integrity was verified with LabChip Gx Touch 24 (Perkin Elmer).


title: Fraction 3, Hmga2-/-


title: Spleen, Control, Day 0 [VC17S]

description: Replica 1

value: Uninfected


title: Spleen, Control, Day 0 [VC19S]

description: Replica 1


title: Spleen, Control, Day 0 [VC15S]

description: Replica 1


title: Hmga2-/- TGFB1


title: Spleen, Control, Day 0 [VC14S]

description: Replica 1


title: Spleen, Control, Day 0 [VC12S]

description: Replica 1


title: Hmga2+/+ TGFB1


title: Spleen, Control, Day 0 [VC10S]

description: Replica 1


title: Spleen, Control, Day 0 [VC11S]

description: Replica 1


title: Input, Hmga2+/+


title: Fraction 3, Hmga2+/+


title: Spleen, Control, Day 0 [VC7S]

description: Replica 1


title: Spleen, Control, Day 0 [VC9S]

description: Replica 1


title: TLSMG 120h

city: Berlin

first: Helene

last: Kretzmer

address: Ihnestraße 63

department: Genome Regulation

institute: Max Planck Institute for Molecular Genetics

description: Single-cell RNA-sequencing of 3 pooled TLSMG (at 120h)

data_processing:
Raw base call (BCL) files generated by Illumina sequencers were demultiplexed into FASTQ files using 10X Genomics' Singel-Cell Gene Expression Software Cell Ranger ("mkfastq" pipeline, version 1.3.1, default parameters).
For each experiment FASTQ files were aligned, filtered, barcode and UMI counted using 10X Genomics' Singel-Cell Gene Expression Software Cell Ranger ("count pipeline", version 1.3.1, default parameters). The count pipeline was run jointly on multiple sequencing runs of the same experiment. Genomic reference: "refdata-cellranger-mm10-1.2.0", prepared by 10x Genomics.
The output of Cell Ranger "count" was aggregated and normalized to the same sequencing depth and then the gene-barcode matrices were recomputed on the combined data. Sample aggregation was evaluated with normalizing for sequencing depth differences in the libraries. Normalized UMI counts used the 'mapped' normalization mode. Analysis of the data was based on the normalized UMI counts.
genome build: mm10
For all samples there is one tar gz archive containing the gene-barcode matrices as produced by the Cell Ranger pipeline. They contain the MEX-formatted UMI count matrices of each sample, or if multiple libraries where sequence, the aggregated gene-barcode matrix. 10x Genomics explains their MEX-formatted UMI counts as follows: "The cellranger pipeline generates a gene-barcode matrix... Each matrix is stored in Market Exchange Format (MEX). It also contains TSV files with genes and barcode sequences corresponding to row and column indices, respectively." More information on the formatting can be found at https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/output/matrices.

source_name: TLSMG 120h aa (3x pooled)

growth_protocol:
mESCs culture: Mouse embryonic stem cells (mESCs)  were cultured on 6 cm plates gelatinized with 0.1% gelatin and coated with feeder cells (3-4x10^4cells/cm²)  in ES+LIF medium at 37 °C and 5%  or 7.5%  CO2. mESCs were split every second day with a dilution suitable to the proliferation velocity (between 1/5 and 1/9). ES+LIF medium was refreshed daily. For splitting, media was aspirated and cells were washed once with PBS and trypsinized for 5-10’ at 37 °C. Trypsin was neutralized by 3 ml ES+LIF and centrifuged for 5’ at 1000 rpm, after which the pellet was resuspended in ES+LIF. For freezing of mESCs, cell pellets were resuspended in ES medium with 20% FCS, and mixed in a 1:1 ratio with ES freezing medium. Cells were frozen down o/n in the –80 °C and transferred to liquid nitrogen the next day.
Generation of gastruloids: Gastruloids were generated as described previously (Beccari et al., Nature 2018), with some minor modifications. First, mESCs were feeder freed. To this end, mESCs were trypsinized on the feeder plate as described above, washed with ES+LIF and resuspended in 2ml ES+LIF. On three gelatinized (0.1% gelatin) wells of a 6-well plate, cells were sequentially plated for 25’, 20’ and 15’. With each transfer, cells were triturated to maintain a single cell suspension. Feeder-freed mESCs were then washed once in 5 ml PBS containing MgCl2 and NaCl and once in 5 ml NDiff227. mESCs were then pelleted by centrifugation for 5’ at 1000rpm and resuspended in 500 µl of N2B27. 10 µl of the cell suspension was mixed with 10 µl of Trypanblue (Bio-Rad) for automated cell counting with Luna Automated Cell Counter. 200-250 live cells were then  plated in a volume of 30 to 40 µl NDiff227 into each well of a 96-well round bottom, low attachment plate  (Cellstar 96 well suspension culture plate or Costar 7007 ultra-low attachment 96 well plate). Cells were then allowed to aggregate for 48 h. After these 48h cells were pulsed with 3 µM CHIR99021 (CHI) in 150 µl of  NDiff227. Between 72 and 120h aa, medium was refreshed every 24 h by removing 150 µl of the old media and adding the same volume of new, pre-incubated NDiff227. For the "96h" samples cells harvested at 96h aa and for the "Gastruloid" at 120h aa and subjected to bulk RNA sequencing and for 96h also for single cell RNAseq ("TLS precursor").
Generation of TLS: The gastruloid protocol described above was followed until 96h aa. Gastruloids were then embedded in 5% Growth-Factor-Reduced Matrigel (MG) (Corning). To this end, fresh NDiff227 medium was pre-incubated for at least 20’ at at 37 °C and 5%  or 7.5%  CO2. Pre-incubated medium was then put on ice for 5’, after which MG was added to achieve a final concentration of 5%. Medium was then put on room temperature for 5’, during which 150µl of old medium was removed from the aggregates. New medium with 5%MG (150µl) was then added, and the cultures were returned to the incubator and further cultured at 37 °C and 5%  or 7.5%. Cultured were allowed to settle for at least 30’ before proceeding to further experimentation (e.g. live imaging). For TLS treated with CHI ("TLSMGC") and CHI+LDN ("TLSMGCL"), 5 µM CHI with or without 600 nM LDN was added from 96h to 120h aa prior to adding the MG. For controls ("TLSMG"), an equal volume of diluent (DMSO) was added. All conditions were subjected to bulk RNAseq and the TLSMG were subjected to single cell RNAseq at 108h and 120h aa.

extract_protocol:
Structures were serially washed through pipette transferring (cut 200ul tip) in wells filled with 200ul of 1xPBS/0.4%BSA (5 transfers) to get rid of the Matrigel embedding. Then structures of the same time point have been pooled together and dissociated in a drop of 200 µl TrypLE Express (Gibco) for 25 minutes with pipetting every 5 minute intervals. The cell suspension was filtered using Scienceware Flowmi Cell Strainers, 40 µM. Cells were washed twice with 1 ml 1xPBS/0.4%BSA with centrifugation steps performed for 5 minutes at 1200 rpm in low DNAbind Eppendorf tubes. The cell concentration was determined using a hemocytometer and cells were subjected to single-cell RNA sequencing (10x Genomic, Chromium™ Single Cell 3’ v3; one reaction per timepoint has been used) aiming for a target cell recovery of up to 10,000 sequenced cells per sequencing library (timepoint).
Single-cell libraries were generated following the manual instructions of 10x Genomic (Chromium™ Single Cell 3’ v3), with the exception of fewer PCR cycles in cDNA amplification and sample index PCR than recommended in order to increase complexity of libraries. Libraries were sequenced with a minimum of 230 Mio paired end reads and parameters as described in the manual.

value: mESC

value: 3 pooled TLSMG (at 120h)


title: Fraction 4, Hmga2+/+

value: wild type


title: 5 GyExo

city: Kamakura

first: Satoshi

last: Kondo

state: Kanagawa

address: 10-1 tebiro 6-chome

country: Japan

institute: Toray Industries,Inc.

hyb_protocol: Hybridized for 16 h at 32 C with rotary shake (250 rpm). Hybridization buffer and washing protocol was followed by the protocol supplied by TORAY Industries, Inc..

scan_protocol: 3D-Gene Scanner ((Toray Industries Inc., Tokyo, Japan) was used for scanning. Images were quantified using Extraction(Toray Industries Inc., Tokyo, Japan).

data_processing: The raw data of each spot was normalized by substitution with a mean intensity of the background signal determined by all blank spots’ signal intensities of 95% confidence intervals.

source_name: pancreatic cancer, 5 Gy, 48 h

label_protocol: Extracted total RNA was labeled with Cy5 using the 3D-Gene miRNA labeling kit (Toray, Kamakura, Japan)

growth_protocol: The MIAPaCa-2 human pancreatic cancer cell line was obtained from Japanese Collection of Research Bioresources Cell Bank (JCRB) and maintained in minimal essential medium (MEM; Sigma Aldrich, Merck KGaA) supplemented with 10% (vol / vol) fetal bovine serum (FBS; Sigma Aldrich, Merck KGaA), 1% penicillin-streptomycin mix solution (Nacalai Tesque) and 1% MEM nonessential amino acids solution (100×) (Nacalai Tesque) in a humidified atmosphere containing 5% CO2 at 37 °C.

extract_protocol: RNA samples were extracted from cells using Toray's 3D-Gene RNA extraction regent from a liquid sample kit (Toray Industries Inc., Tokyo, Japan).

treatment_protocol: To isolate exosomes, cells were seeded at 1.5 × 10^6 cells per T75 cm^2 flask and irradiated after substituting the media with exosome-depleted 10% FBS cell culture media. The cell culture media was centrifuged at 2,000 × g for 10 min at 4 °C, and the supernatants were filtered through a 0.22-µm minisart syringe filter (Sartorius, Gottingen, Germany). The supernatant was centrifuged at 150,000 × g for 90 min at 4 °C. The pellet was washed with phosphate-buffered saline (PBS) and centrifuged at 150,000 × g for 90 min at 4 °C and resuspended in 50 μl PBS. After then, exomal RNA samples were extracted.

value: pancreatic cancer

tag: cell line

tag: gender

value: Male

value: 65 years

value: irradiated (5 Gy)


title: 8 GyExo

source_name: pancreatic cancer, 8 Gy, 48 h

value: MIAPaCa-2 human pancreatic cancer cell line

value: irradiated (8 Gy)


title: Spleen, Control, Day 0 [VC6S]

description: Replica 1

value: Spleen


title: Spleen, Control, Day 0 [VC4S]

description: Replica 1


title: 293T/17 con cpt rep 2

city: Bonn

first: Moritz

last: Kebschull

address: Welschnonnenstr. 17

department: Department of Periodontology

institute: University of Bonn

description: replicate 2

hyb_protocol: standard Illumina hybridization protocol

scan_protocol: standard Illumina scan protocol

data_processing: Data was exported from Illumina BeadStudio, quality controlled, quantile normalized and analyzed using R/Bioconductor and the beadarray package.

source_name: 293T/17 cells

label_protocol: 100 ng of total RNA was labeled using the the Ambion Total Prep RNA Amplification and Labeling kit.

extract_protocol: Total RNA was extracted using Trizol reagent, cleaned up using a Quiagen RNeasy mini kit, quantified on a Nanodrop 2000C, quality controlled using a Agilent Bioanalyzer. All samples showed RIN numbers ≥ 9.

tag: spag4 status

value: wildtype (control)

value: camptothecin 2uM, 48h


title: Spleen, Control, Day 0 [VC2S]

description: Replica 1


title: 293T/17 Spag4 knockdown vehicle rep 3

description: replicate 3

value: shRNA knockdown

value: vehicle (DMSO)


title: Bmi-1 shRNA rep3

city: Jinan

first: Xin

last: Zhang

state: Shandong

address: 107 Wenhua Xi Road

country: China

institute: Qilu Hospital, Shandong University

description: Small RNA

data_processing:
Subsequently, unique sequences with length in 18~25 nucleotide were mapped to specific species precursors in miRBase 22.0 by BLAST search to identify known miRNAs and novel 3p- and 5p- derived miRNAs.
Length variation at both 3' and 5' ends and one mismatch inside of the sequence were allowed in the alignment.
The unique sequences mapping to specific species mature miRNAs in hairpin arms were identified as known miRNAs. The unique sequences mapping to the other arm of known specific species precursor hairpin opposite to the annotated mature miRNA-containing arm were considered to be novel 5p- or 3p-derived miRNA candidates.
The remaining sequences were mapped to other selected species precursors (with the exclusion of specific species) in miRBase 22.0 by BLAST search, and the mapped pre-miRNAs were further BLASTed against the specific species genomes to determine their genomic locations.
The above two we defined as known miRNAs. The unmapped sequences were BLASTed against the genomes, and the hairpin RNA structures containing sequences were predicated from the flank 120 nt sequences using RNAfold software (http://rna.tbi.univie.ac. at/cgi-bin/RNAfold.cgi). The criteria for secondary structure prediction were: (1) number of nucleotides in one bulge in stem (≤12) (2) number of base pairs in the stem region of the predicted hairpin (≥16) (3) cutoff of free energy (kCal/mol ≤-15) (4) length of hairpin (up and down stems + terminal loop ≥50) (5) length of hairpin loop (≤200). (6) number of nucleotides in one bulge in mature region (≤4) (7) number of biased errors in one bulge in mature region (≤2) (8) number of biased bulges in mature region (≤2) (9) number of errors in mature region (≤4) (10) number of base pairs in the mature region of the predicted hairpin (≥12) (11) percent of mature in stem (≥80).
Supplementary_files_format_and_content: excel,expression profiles

source_name: Cholangiocarcinoma cells

growth_protocol: QBC-939 was maintained in DMEM medium supplemented with 10% fetal bovine serum in 5% CO2 resuscitation.

extract_protocol:
Total RNA was extracted using Trizol reagent (Invitrogen, CA, USA) following the manufacturer's procedure
Approximately 5 µg of total RNA were used to prepare nine small RNA libraries according to the protocol of TruSeq Small RNA Sample Prep Kits (Illumina, San Diego, CA, USA). And then the libraries were sequenced by Illumina Hiseq 2500 at the LC-BIO following the vendor’s recommended protocol. Raw reads were subjected to an in-house program, ACGT101-miR (LC Sciences, Houston, Texas, USA) to remove adapter dimers, junk, low complexity, common RNA families (rRNA, tRNA, snRNA, snoRNA) and repeats.

treatment_protocol: QBC-939 was treated with Bmi1 shRNA or scrambled shRNA for 48h.

tag: cell type

value: Cholangiocarcinoma

tag: knockdown

value: Bmi1 shRNA


title: 293T/17 Spag4 knockdown cpt rep 2

description: replicate 2

tag: treatment


title: Input, Hmga2-/-

value: Mouse embryonic fibroblasts

tag: strain

tag: tissue

value: whole embryo

value: embryonic day 15.5

value: Hmga-/-

value: Timed-pregnant mice were sacrificed on post coitum day 15.5  according to standard methods. The embryos were collected in sterile PBS complemented with 100 U\ml penicillin and 100 U\ml streptomycin. After removing the head, limbs, tail, viscera, liver and heart, the remaining body was trypsinized with 1x trypsin-EDTA solution (0.05 %) for 20 minutes at 37°C and single cells were obtained via physical disruption by sterile Pasteur pipet. The cells from each embryo were diluted with growth medium containing FCS and plated separately.


title: 293T/17 con cpt rep 1

description: replicate 1


title: Bmi-1 shRNA rep1

description: Small RNA


title: 293T/17 Spag4 knockdown cpt rep 3

description: replicate 3


title: Spleen, Control, Day 0 [VC1S]

description: Replica 1

data_processing: All q-PCR assays exhibited efficiencies within 100% ± 5% so the simplified 2-ΔΔCq method was used for analysing data (Livak and Schmittgen, 2001). For normalization and as internal calibrator the average of three genes (ARP, ELF1α and β-actin) was chosen by using NormFinder (Andersen et al., 2004). For gene expression analyses, all challenged groups were compared to non-exposed time point controls using Student’s t-test. We considered differences to be significant when regulations were at least two-fold and p < 0.05. For four of the genes investigated in the non-exposed timepoint control groups less than three fish showed Cq values (excluding a t-test) and therefore we applied a qualitative assessment (presence/absence of Cq values) and analyzed data with the nonparametric Mann-Whitney test using a probability level of 5%.

value: Uninfected

value: Day 0


title: Liver, Survivor, Day 0 [V36L]

description: Replica 6

source_name: Survivor

value: Liver

value: Vibrio anguillarum

value: Day 11


title: 293T/17 con cpt rep 3

description: replicate 3


title: Liver, Survivor, Day 0 [V35L]

description: Replica 6


title: Spleen, Control, Day 0 [VC5S]

description: Replica 1

value: Spleen


title: control rep2

description: Small RNA

value: control


title: 293T/17 Spag4 knockdown vehicle rep 2

description: replicate 2


title: 293T/17 Spag4 knockdown cpt rep 1

description: replicate 1

value: camptothecin 2uM, 48h


title: control rep1

description: Small RNA


title: 293T/17 con vehicle rep 1

description: replicate 1

value: wildtype (control)


title: 0 GyExo

description: HCA-control

source_name: pancreatic cancer, 0 Gy, 48 h

value: pancreatic cancer

value: MIAPaCa-2 human pancreatic cancer cell line

value: Male

value: 65 years

value: non-irradiated


title: 293T/17 Spag4 knockdown vehicle rep 1

description: replicate 1

value: shRNA knockdown

value: vehicle (DMSO)


title: Bmi-1 shRNA rep2

description: Small RNA

value: Cholangiocarcinoma

value: Bmi1 shRNA


title: Liver, Survivor, Day 0 [V34L]

description: Replica 6


title: Liver, Survivor, Day 0 [V39L]

description: Replica 6


title: sgUSP22_rep2

city: Toronto

first: Musaddeque

last: Ahmed

state: ON

address: 101 College St

country: Canada

institute: University health Network

description: total RNA

data_processing:
The raw sequencing data were mapped to human genome assembly GRCh38 using STAR with quantMode method against refGene gene annotation
The differential expression analyses were performed using DESeq2 package in R
Genome_build: GRCh38
Supplementary_files_format_and_content: Raw count matrix of refGene genes in all samples

growth_protocol: 24hrs post-transduction, cells were selected with completed medium containing puromycin for 72hrs.

extract_protocol:
Total RNA was extracted from AML3 cells using the RNeasy Plus Mini Kit (Qiagen Cat. 74314) following manufacture’s guidelines
RNA-seq libraries were prepared using the SMARTer Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian (634418, Takara)

treatment_protocol: AML cells were transduced with sgRNAs targeting the genes or control

value: Blood

tag: sgrna


title: sgATXN7L3_rep1

description: total RNA


title: sgUSP22_rep1

description: total RNA


title: sgENY2_rep2

description: total RNA


title: Liver, Survivor, Day 0 [V29L]

description: Replica 6

extract_protocol: RNAlater fixed samples from fish (gills, liver and spleen) were homogenized (Tissue-lyser II, Qiagen, Denmark) using a homogenization buffer with 2-mercaptoethanol (Sigma-Aldrich). RNA was extracted using the GenEluteTM mammalian RNA kit (RTN350, Sigma-Aldrich, Denmark). Liver samples were pre-treated by Proteinase K (cat.no.P4850, Sigma-Aldrich, Denmark). DNase I (AMPD1, Sigma-Aldrich, Denmark) treatment was applied to remove genomic DNA. The concentration of RNA was determined in a NanoDrop 2000 spectrophotometer (Saveen & Werner, Denmark). Quality of RNA was evaluated by agarose (cat.no. 16500100, ThermoFisher Scientific, Denmark) gel electrophoresis. RNA was kept at −80°C until cDNA synthesis.

value: Liver


title: Liver, Survivor, Day 0 [V32L]

description: Replica 6

value: Vibrio anguillarum


title: Liver, Survivor, Day 0 [V30L]

description: Replica 6

value: Day 11


title: 293T/17 con vehicle rep 3

description: replicate 3


title: 293T/17 con vehicle rep 2

description: replicate 2


title: Liver, Survivor, Day 0 [V33L]

description: Replica 6


title: sgAAVS_rep2

description: total RNA

value: sgAAVS


title: Liver, Survivor, Day 0 [V26L]

description: Replica 6


title: OXO-PCR Patient 1031 sample 3E1

city: Oxford

first: Zhiyuan

last: Hu

address: John Radcliffe Hospital/Headley Way

department: MRC Weatherall Institute of Molecular Medicine

institute: University of Oxford

data_processing:
FastQ files were trimmed for adapter sequences and quality.
Trimmed reads were then mapped to the human hg19 reference genome using STAR (v2.4.2a).
Read counts were obtained using subread FeatureCounts (v1.4.5-p1).
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include raw counts for each sample

source_name: High-grade serous ovarian cancer

extract_protocol:
Frozen tumor samples were embedded in OCT (NEG-50, Richard-Allan Scientific) and 10 μm sections were taken using MB DynaSharp microtome blades (ThermoFisher Scientific) in a CryoStar cryostat microtome (ThermoFisher Scientific). Scrolls were also collected to be used for bulk RNA-seq. Bulk scrolls were immediately processed for RNA extraction using the RNeasy Mini kit (Qiagen). The extraction procedure was performed according to the manufacturer’s instructions, including DNase digestion, after which RNA integrity was evaluated using the 2200 TapeStation (Agilent).
The KAPA mRNA HyperPrep Kits (Roche) were use for the library preparation by following the manufacturer's instructions.

tag: patient

value: Pre-chemotherapy high grade serous ovarian cancer

tag: tissue preparation

value: sections


title: OXO-PCR Patient 1027 sample 3D2


title: OXO-PCR Patient 1031 sample 3E2


title: sgAAVS_rep1

description: total RNA


title: AML3_day7_rep1

description:
guide RNA
AML3_D7pre_vs_D0.gene_summary.txt

data_processing:
The fastq files were first converted to fasta file using custom shell script. For each sample, a custom bowtie database was generated by the command bowtie-build in bowtie suite (version 1.1.2). The library sgRNAs were mapped against the database for each sample using bowtie with the parameter v=0 and default values for other parameters.
The differential sgRNA abundance was estimated using the tool MAGeCK (Li et al., 2014).
Genome_build: Hg19
Supplementary_files_format_and_content: MAGeCK output for all cell lines. Please refer to MAGeCK documentation for output details.

growth_protocol: AML2 and AML3 were cultured in alpha-MEM, KG1a and U937 in RPMI-1640, MV4-11 in IMDM, and TEX, all supplemented with 10% fetal bovine serum (FBS).

extract_protocol:
Genomic DNA was extracted from replicate samples (QIAamp Blood Midi kit) and sgRNA inserts were amplified by PCR
The sgRNAs were designed using the CRISPR-DO tool that accounted for sgRNA specificity and cutting efficiency. sgRNAs were synthesized as 73-mer oligonucleotides (CustomArray, USA), GAAAGGACGAAACACCGNNNNNNNNNNNNNNNNNNNNGTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC (N’s denote sgRNA 20 nucleotide target sequence; sense orientation) with a total of 12,472 sequences and amplified by PCR as a pool using the following primers: TAACTTGAAAGTATTTCGATTTCTTGGCTTTATATATCTT GTGGAAAGGACGAAACACCG (Forward) and ACTTTTTCAAGTTGATAACG GACTAGCCTTATTTTAACTTGCTATTTCTAGCTCTAAAAC (Reverse). The PCR product was purified and cloned in the lentiGuide-Puro vector using BsmBI (NEB). Ligation was performed using the NEBuilder® HiFi DNA Assembly Cloning Kit and transformed into an electrocompetent strain (Stbl4; ThermoFisher, USA) to achieve ~300x coverage. Colonies were scraped off Agar plates using LB medium. Plasmid DNA was extracted using NA0310 Sigma GenElute™ HP Plasmid Maxiprep Kit and adequate library representation of each sgRNA was confirmed by NGS.
single end unstranded

treatment_protocol: we co-incubated the transduced AML3 cells with DNTs at a 1:1 E:T ratio for 2h to induce apoptosis in ~40-50% of the targets, depleted DNTs by magnetic separation (EasySep Human CD3 Positive Selection Kit), and performed FACS to separate Annexin V+ (live) and Annexin V- (dead or dying) target cells.

tag: source tissue

tag: expression

value: Cas9 expressing

tag: source type

value: Cell line


title: AML3_day7_rep2


title: OXO-PCR Patient 1027 sample 3D1


title: AML3_day0_rep2


title: Liver, Survivor, Day 0 [V22L]

description: Replica 6

tag: organ


title: Liver, Survivor, Day 0 [V23L]

description: Replica 6

tag: pathogen


title: Liver, Survivor, Day 0 [V28L]

description: Replica 6

tag: time


title: Liver, Survivor, Day 0 [V21L]

description: Replica 6


title: AML3_day0_rep1


title: OXO-PCR Patient 1027 sample 3C9


title: OXO-PCR Patient 1015 sample 1F3


title: AML3_dead_rep2

description:
guide RNA
live_vs_dead.gene_summary.txt


title: Liver, Control, Day 11 [VC60L]

description: Replica 5

value: Uninfected


title: AML3_live_rep2

description:
guide RNA
live_vs_dead.gene_summary.txt


title: Liver, Survivor, Day 0 [V25L]

description: Replica 6


title: Liver, Control, Day 11 [VC58L]

description: Replica 5


title: OXO-PCR Patient 1008 sample 1C7


title: Liver, Control, Day 11 [VC57L]

description: Replica 5


title: AML3_dead_rep1

description:
guide RNA
live_vs_dead.gene_summary.txt


title: OXO-PCR Patient 1002 sample 3F8


title: AML3_live_rep1

description:
guide RNA
live_vs_dead.gene_summary.txt


title: OXO-PCR Patient 1002 sample 3F6


title: Liver, Control, Day 11 [VC56L]

description: Replica 5


title: Liver, Control, Day 11 [VC55L]

description: Replica 5


title: HT29 YB-1471 10uM #2

city: Ottawa

first: Yannick

middle: D.

last: Benoit

state: Ontario

address: 451 Smyth road

institute: University of Ottawa

data_processing:
Quantified transcripts from GENCODE v27 with salmon v0.9.1 using --gcBias flag.
Performed fold change analysis using DESeq2 as follows
Read transcript quantification data with tximport library and summarized to gene level quantification.
Filtered to retain genes with at least 5 reads assigned in at least two samples.
Constructed DESeqDataSet object with fourteen samples (two replicates of seven conditions), using the model ~condition (cell type/treatment).
Calculated fold changes using the lfcShrink() function with the apeglm method.
Genome_build: GRCh38
Supplementary_files_format_and_content: *_quant.sf.txt: Salmon transcript quantification: Quantification of GENCODE v27 transcripts using salmon v0.9.1.
Supplementary_files_format_and_content: *_all.txt: DESeq2 fold change results: Results from the DESeq2 lfcShrink() function comparing two conditions, including results for all genes retained after filtering.

source_name: Colorectal cancer cell line

extract_protocol:
RNA was extracted using the total RNA purification kit by Norgen Biotech Corp, following the manufacturer's guidelines. Quantification of total RNA samples was performed with a Qubit HS RNA assay (ThermoFisher Scientific) and fragment size was evaluated with a Fragment Analyzer HS NGS assay (AATI). A RNA Quality Number (RQN) of 8.0 or higher was considered satisfactory for library construction. Library construction was performed with a TruSeq RNA v2 (Illumina) with unique barcodes compatible with the Illumina NextSeq 500 platform.
RNA libraries were prepared for sequencing using standard Illumina protocols.

tag: cell line

value: Colorectal cancer cell line

tag: origin

value: Colorectal adenocarcinoma

value: 48 hours 10uM YB-1471


title: Liver, Control, Day 11 [VC54L]

description: Replica 5

source_name: Time point Control


title: AML2_postDNT_rep2

description:
guide RNA
AML2_postDNT_vs_noDNT.gene_summary.txt


title: AML2_postDNT_rep1


title: Liver, Control, Day 11 [VC53L]

description: Replica 5


title: AML2_noDNT_rep2


title: AML2_noDNT_rep1


title: OXO-PCR Patient 1008 sample 1C9


title: AML3_postDNT_rep2

description:
guide RNA
AML3_postDNT_vs_noDNT.gene_summary.txt


title: AML3_postDNT_rep1


title: Liver, Control, Day 11 [VC52L]

description: Replica 5


title: Liver, Control, Day 11 [VC49L]

description: Replica 5


title: HIEC YB-1471 10uM #2

source_name: Intestinal epithelial crypt cells

value: Intestinal epithelial crypt cells

value: 16-week fetal ileum


title: Liver, Control, Day 11 [VC48L]

description: Replica 5


title: HIEC YB-1471 10uM #1


title: AML3_noDNT_rep2


title: HIEC DMSO #2

description: processed data file: HT29_vs_HIEC.DMSO_all_salmon.txt

value: 48 hours DMSO


title: Liver, Control, Day 11 [VC47L]

description: Replica 5