description:
Library name: M8_batchA_Cervical_Bulbar
Tissue from Cervical sampled from Patient MALS1008 with Onset: Bulbar (batchA)

value: spinal cord


title: casprib2 biol rep3

status: Public on Mar 10 2026

type: SRA

city: Seattle

first: Alexander

last: Meeske

state: WA

address: 750 Republican St

country: USA

institute: University of Washington

description:
Library name: casprib2 biol rep3
raw reads and RPKM files

library_source: transcriptomic

data_processing:
Reads were mapped to the L. seeligeri LS1 genome using Bowtie2 with settings -I 10 and -X 700. Reads per kilobase values for each gene in the LS1 genome were calculated from the resulting BAM files using Subread featureCounts35, dividing gene read count values by gene length and applying a scale factor to normalize each library to 10M total mapped reads.
Assembly: NC_013891.1
Supplementary files format and content: rpkm files (.txt)

molecule: total RNA

organism: Listeria seeligeri

source_name: bacterial culture

growth_protocol: BHI + kanamycin 50µg/mL + atc 500ng/mL 30°C to OD 0.3-0.8

extract_protocol:
Cultures were pelleted via centrifugation and pellets were resuspended in 100 μL of 1X PBS with 2 mg/mL lysozyme and incubated for 5 min in a 37°C water bath. After incubation, 10 μL of 10% sarkosyl was added to each sample and resuspended before adding 300μL TRI reagent. Samples were vortexed and 400μL of 100% ethanol was added. Each sample solution was then purified using the Zymo Direct-zol RNA Miniprep Kit to the manufacturer’s instructions. RNA-extraction samples were treated with DNAse and column purified using the Zymo RNA Clean & Concentrator kit per the manufacturer’s instructions.
Performed by Azenta (Strand-specific RNA-seq) including rRNA depletion

tag: cell type

value: bacterial culture

tag: genotype

value: pBH228 (inducible type I-B CRISPR system) + casPRIB2 expression plasmid

tag: treatment

value: aTc 500ng/mL (to induce CRISPR)


title: Mll4_AID_MyoD_ER_4OHT_T7

city: Bethesda

first: Kai

last: Ge

state: MD

address: 50 South Dr Rm 4154

department: NIDDK

institute: NIH

description: Library name: Sample 58

library_source: genomic

data_processing:
Basecalls performed using Bcl2fastq.
CUT&RUN and ATAC-Seq reads were aligned to the mm10 genome assembly using bowtie2.
CUT&RUN peaks were called using SICER (v2) with following configuration: For transcription factors and co-factors, a window size of 50 bp, gap size of 50 bp, and a false discovery rate of 1E-10 were used. For histone modifications, a window size of 200 bp, gap size of 200 bp, and a false discovery rate of 1E-3 were used. The effective genome fraction of 0.8 was applied to all CUT&RUN data.
Assembly: mm10
Supplementary files format and content: wig files were generated using in-house script and the scores represent RPKM
Library strategy: CUT&Run

molecule: genomic DNA

organism: Mus musculus

source_name: Kmt2c-/-;Kmt2dAID/AID preadipocytes

growth_protocol: Preadipocytes were maintained in growth media (DMEM plus 10% Foundation FBS).

extract_protocol:
CUT&RUN was performed using CUTANA™ ChIC/CUT&RUN Kit (#14-1048, Epicypher), following the manufacturer’s instructions with the following modifications. 0.5 million cells were used for each CUT&RUN reaction. Cells were gently fixed with 0.1% formaldehyde for 1 min. Cells were resuspended in cold Nuclei Extract Buffer (20 mM HEPES pH 7.9, 10 mM KCl, 0.1% Triton X-100, 20% Glycerol, 1 mM MgCl2 supplemented with protease inhibitor and 0.5 mM Spermidine), lysed on ice for 10 mins, and centrifuged at 600 g for 3 min 4°C. Nuclei were resuspended in 100 uL cold Nuclei Extract Buffer and incubated with activated 10 uL/reaction Concanavalin A beads for 10 min at RT. The following steps are performed according to the manufacturer’s instructions.
Libraries were prepared following the instruction of CUTANA™ CUT&RUN Library Prep Kit for Illumina sequencing. CUT&RUN enriched DNA were used to construct libraries, using NEBNext® Ultra™ II DNA Library Prep kit (#E7645L, NEB) with AMPure XP magnetic beads (Beckman Coulter).

treatment_protocol: MyoD-ER-T7 preadipocytes were grown in growth media (DMEM plus 10% Foundation FBS) for 2 days. On the day of experiments, growth medium was replaced with differentiation medium (DMEM plus 2% horse serum), and cells were rested for 3h. Cells were then pretreated with 1 µM BRM014 or 3 µM A-485 for 1h; 1 µM 5Ph-IAA for 2h; or 250 nM dCBP-1 for 3h, before 400 nM 4-OHT was added for 1h to induced MyoD nuclear translocation.

tag: cell line

value: Kmt2c-/-;Kmt2dAID/AID preadipocytes

value: Preadipocyte

value: 4OHT (1h)

tag: chip antibody


title: PS_M9gluc_H2O2_rep1

city: La Jolla

first: Heera

last: Bajpe

state: CA

address: 9500 Gilman Dr.

institute: UC San Diego

description: Library name: PS_M9gluc_H2O2_1

data_processing:
Raw sequencing reads were mapped to the reference genome using bowtie v1.1.2 with parameters -X 1000 -3 3 -n 2 -S
Read counts were generated using featureCounts
Assembly: NC_004578.1, NC_004633.1, NC_004632.1
Supplementary files format and content: Supplementary files format and content: Comma-delimited text file containing logTPM values for each sample.

organism: Pseudomonas syringae pv. tomato str. DC3000

source_name: bacterial cell

growth_protocol: Cells were grown overnight at 30 °C with shaking in 4 mL of the base medium (M9 or CAMHB). The following day, fresh cultures were inoculated in duplicate to an initial OD600 of 0.05. For nitrogen starvation (nitrogen-free M9 media with addition of eighteen amino acids, with the exception of cysteine and methionine) conditions, cultures were grown in medium supplemented as specified. Cultures were harvested at mid-log phase (OD₆₀₀ ≈ 0.5) and 3 mL of culture was immediately mixed with 6 mL of RNAprotect reagent for the following conditions: growth in M9 glucose media (with and without trace element solution), CAMHB media, and nitrogen starvation. Cells were pelleted and stored at −80 °C until RNA extraction and RNA-seq library preparation.

extract_protocol:
Total RNA was isolated and prepared from cell pellets using reagents from Zymo Research and silica-coated magnetic beads (Luna Nanotech) accroding to the vendor's manual. Concentrations and quality of the RNA were assessed using an Agilent TapeStation and Nanodrop, respectively. For each sample, 1 ug was used as input for the standard RiboRid procedure.
RNA-seq libraries were prepared from 5 uL of the rRNA-subtracted RNA using KAPA RNA HyperPrep Kit (Roche) according to the manufacturer’s protocol with volumes reduced by half. The final PCR amplification step was modified by incorporating SYBR and using a qPCR instrument. Samples were removed from the thermal cycler just before entering the plateau phase of amplification to avoid biases being potentially introduced by reagent depletion. The quality and concentrations of the libraries were checked with an Agilent TapeStation and Qubit instrument, respectively. Pooled libraries were then run on an AVITI instrument (Element Biosciences).

treatment_protocol: For ceftazidime treatment conditions, ceftazidime (¼ of minimum inhibitory concentration) was added at OD₆₀₀ ≈ 0.2, and cultures were incubated for 1 h prior to collection. For osmotic stress (NaCl treatment), oxidative stress (H2O2 treatment), and metal ion stress (CuSO4 and ZnSO4 treatment) conditions, cultures were grown to OD₆₀₀ ≈ 0.5 before addition of the corresponding supplement to the final concentration indicated. This was followed by incubation for an additional 30 min (osmotic and oxidative stress conditions) or 1 h (metal ion stress conditions) before mixing 3 mL of culture with 6 mL of RNAprotect reagent.

tag: strain

value: bacterial cell

value: wild-type

value: glucose (2 g/L) and H2O2 (5 mM)

tag: medium

value: M9 media


title: Mll4_AID_MyoD_ER_4OHT_IgG

description: Library name: Sample 56

value: Anti-IgG (13-0042K, Epicypher)


title: PS_M9gluc_NaCl_rep2

description: Library name: PS_M9gluc_NaCl_2

value: glucose (2 g/L) and NaCl (0.35 M)


title: Mll4_AID_MyoD_ER_4OHT_p300_CST

description: Library name: Sample 59


city: Chicago

first: Fabien

last: Dachet

state: IL

address: 1200 W Harrison St

institute: University of Illinois Chicago

description:
Library name: UIC8_batchA_Medulla_Arm
Tissue from Medulla sampled from Patient UIC-ALS8 with Onset: Arm (batchA)

data_processing:
FASTQ files obtained from Illumina BaseSpace were processed using Trim Galore (v0.6.10) for adapter trimming and quality filtering, then aligned to the human reference genome GRCh38.p14 using the STAR aligner (v2.7.11b) with GENCODE v47 annotation. Gene-level read counts were generated with featureCounts from the Subread package (v2.0.4). Differential expression analysis was performed in R (v4.4.1) using DESeq2 (v1.44.0), with comparisons conducted in the medulla (bulbar vs non-bulbar onset) and cervical spinal cord (arm vs non-arm onset). Gene ontology enrichment analysis was carried out using the DAVID database, considering pathways with a gene count ≥5 and a Benjamini–Hochberg false discovery rate <0.01
Assembly: Sequencing reads were aligned to the human reference genome assembly GRCh38.p14 using gene annotations from GENCODE release 47
Supplementary files format and content: Raw count from FeatureCount using same strand counting

organism: Homo sapiens

source_name: medulla

extract_protocol:
Human postmortem tissues were collected through a rapid autopsy protocol performed within 3 hours of death, with samples obtained from the medulla, cervical spinal cord (C4–6), and lumbar spinal cord (L3–5); tissues were rapidly frozen and stored at −80 °C. For RNA extraction, multiple thin frozen sections from each region were pooled, and total RNA was isolated using the Qiagen RNeasy Lipid Tissue Kit (Cat#74804, Qiagen, Venlo, Netherlands) according to the manufacturer’s protocol, yielding approximately 3 µg of RNA per sample. RNA samples were treated with DNase (RCC-5, Zymo Research, Irvine, CA) to remove genomic DNA, and RNA quality was assessed using a Qubit 4.0 instrument, with only samples having a RNA integrity number (RIN) ≥ 7 included for downstream analyses
RNA library construction was performed by the UIC Genome Research Core facility using total RNA extracted from postmortem human tissues. Following quality control, samples meeting inclusion criteria were used for library preparation, after which sequencing was carried out on an Illumina NovaSeq 6000 platform at the UIC-Urbana Core Facility. Sequencing was performed in paired-end mode, generating an average depth of approximately 30–40 million paired reads per sample, and resulting FASTQ files were retrieved from Illumina BaseSpace for downstream analyses.

tag: tissue

value: medulla

value: Multiple type of cells present in human medulla

tag: batch

value: Batch A


description:
Library name: UIC7_batchA_Lumbar_Bulbar
Tissue from Lumbar sampled from Patient UIC-ALS7 with Onset: Bulbar (batchA)

source_name: spinal cord

value: Multiple type of cells present in human spinal cord


title: casprib2 biol rep1

description:
Library name: casprib2 biol rep1
raw reads and RPKM files


title: casprib2 biol rep2

description:
Library name: casprib2 biol rep2
raw reads and RPKM files


title: PS_M9gluc_NaCl_rep1

description: Library name: PS_M9gluc_NaCl_1


description:
Library name: UIC7_batchA_Cervical_Bulbar
Tissue from Cervical sampled from Patient UIC-ALS7 with Onset: Bulbar (batchA)

value: spinal cord


title: Mll4_AID_MyoD_ER_4OHT_ARID1A

description: Library name: Sample 55

value: Anti-ARID1A/BAF250A (D2A8U) (12354, CST)


title: PS_CAMHB_ceftazidime_rep2

description: Library name: PS_CAMHB_ceftazidime_2

value: ceftazidime (1 ug/mL)

value: CAMHB meda


title: capsprib1 biol rep3

description:
Library name: capsprib1 biol rep3
raw reads and RPKM files

value: bacterial culture

value: pBH228 (inducible type I-B CRISPR system) + casPRIB1 expression plasmid

value: aTc 500ng/mL (to induce CRISPR)


title: PS_CAMHB_rep2

description: Library name: PS_CAMHB_2

value: bacterial cell


title: casprib1 biol rep2

description:
Library name: casprib1 biol rep2
raw reads and RPKM files


description:
Library name: UIC6_batchA_Lumbar_Bulbar
Tissue from Lumbar sampled from Patient UIC-ALS6 with Onset: Bulbar (batchA)


description:
Library name: UIC6_batchA_Medulla_Bulbar
Tissue from Medulla sampled from Patient UIC-ALS6 with Onset: Bulbar (batchA)


title: Mll4_AID_MyoD_ER_unt_T7

description: Library name: Sample 53

value: Kmt2c-/-;Kmt2dAID/AID preadipocytes

value: Preadipocyte

value: untreated


title: Mll4_AID_MyoD_ER_unt_p300_CST

description: Library name: Sample 54


title: Mll4_AID_MyoD_ER_unt_MLL4

description: Library name: Sample 52

value: Anti-MLL4 #3 (home-made, Kai Ge Lab)


title: empty vector biol rep3

description:
Library name: empty vector biol rep3
raw reads and RPKM files

value: pBH228 (inducible type I-B CRISPR system) + empty vector


description:
Library name: UIC7_batchA_Medulla_Bulbar
Tissue from Medulla sampled from Patient UIC-ALS7 with Onset: Bulbar (batchA)

value: medulla


description:
Library name: UIC6_batchA_Cervical_Bulbar
Tissue from Cervical sampled from Patient UIC-ALS6 with Onset: Bulbar (batchA)


title: PS_M9gluc_rep2

description: Library name: PS_M9gluc_2

value: wild-type

value: glucose (2 g/L) and trace element solution

value: M9 media


title: Mll4_AID_MyoD_ER_4OHT_MLL4

description: Library name: Sample 57

value: 4OHT (1h)


title: empty vector biol rep2

description:
Library name: empty vector biol rep2
raw reads and RPKM files


title: casprib1 biol rep1

description:
Library name: casprib1 biol rep1
raw reads and RPKM files


title: PS_CAMHB_rep1

description: Library name: PS_CAMHB_1


title: Mll4_AID_MyoD_ER_unt_ARID1A

description: Library name: Sample 50

value: Anti-ARID1A/BAF250A (D2A8U) (12354, CST)


title: PS_M9gluc_rep1

description: Library name: PS_M9gluc_1


title: Mll4_AID_MyoD_ER_unt_IgG

description: Library name: Sample 51

value: Anti-IgG (13-0042K, Epicypher)


title: empty vector biol rep1

description:
Library name: empty vector biol rep1
raw reads and RPKM files


title: Mll4-AID_MyoD-ER_A485_4OHT_T7

description: Library name: Sample 49


description:
Library name: UIC5_batchA_Lumbar_Leg
Tissue from Lumbar sampled from Patient UIC-ALS5 with Onset: Leg (batchA)

value: spinal cord

value: Multiple type of cells present in human spinal cord

value: Batch A


title: MUT_3

city: Xiamen

first: Tiansheng

last: Chen

address: No. 43, Yindou Road, Jimei District, Fisheries College of Jimei University

country: China

institute: Jimei University

description:
Library name: Sample 6
MUT_3

data_processing:
Illumina Casava 1.8 software used for basecalling
Raw data (raw reads) of fastp format were firstly processed through in-house perl scripts..In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. All the downstream analyses were based on the clean data with high quality.
Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.0.5.
featureCounts v1.5.0-p3 was used to count the reads numbers mapped to each gene.
Differential expression analysis of two conditions/groups (two biological replicates per condition) was performed using the DESeq2 R package (1.20.0).
Assembly: http://ftp.ensembl.org/pub/release-94/fasta/oryzias_latipes/dna/Oryzias_latipes.ASM223467v1.dna.toplevel.fa.gz http://ftp.ensembl.org/pub/release-94/gtf/oryzias_latipes/Oryzias_latipes.ASM223467v1.94.gtf.gz
Supplementary files format and content: tab  files include FPKM values  for all Sample

organism: Oryzias latipes

source_name: Testis

extract_protocol:
Total RNA was isolated using TRIzol according to the manufacturer’s instructions.
RNA libraries for RNA-seq were prepared using Fast RNA-seq Lib Prep Kit V2 following manufacturer's protocols.

value: Testis


title: 48hpf

city: bad nauheim

first: Stefan

last: Günther

address: ludwigtr. 43

country: Germany

institute: MPI for heart and lung research

description: Library name: sample9

library_source: transcriptomic single cell

data_processing:
Sequencing was done on Nextseq2000 and raw reads were aligned against the zebrafish genome (DanRer11) and counted by StarSolo
Preprocessed counts were further analysed using Scanpy ollowed by secondary analysis in Annotated Data Format.
Final data visualization was done by scanpy and cellxgene packages.
Assembly: danRer11
Supplementary files format and content: starsolo outputs

organism: Danio rerio

source_name: Whole heart

extract_protocol:
For all datasets, the cell suspensions were depleted for dead cells using LeviCell 1.0 device (Levitas Bio), counted again with MOX Z cell counter and diluted according to manufacturer’s protocol to obtain 10.000 single-cell data points per sample.  Each sample was run separately on a lane in Chromium controller with Chromium Next GEM Single-cell 3ʹ Reagent Kits v3.1 (10xGenomics).  Single-cell RNAseq library preparation was done using standard protocol.
Library preparation was conducted according to the manufacturer indications.

value: Whole heart

tag: developmental stage

value: 48 hours post fertilization

value: Heart cells


title: PS_CAMHB_ceftazidime_rep1

description: Library name: PS_CAMHB_ceftazidime_1

value: bacterial cell

value: ceftazidime (1 ug/mL)

value: CAMHB meda


title: Mll4-AID_MyoD-ER_A485_4OHT_P300

description: Library name: Sample 48

value: Kmt2c-/-;Kmt2dAID/AID preadipocytes

value: Preadipocyte

value: Anti-p300 (C-20)  (sc-585, Santa Cruz, gift from Bing Ren Lab)


description:
Library name: UIC5_batchA_Medulla_Leg
Tissue from Medulla sampled from Patient UIC-ALS5 with Onset: Leg (batchA)

value: medulla

value: Multiple type of cells present in human medulla


title: 25hpf

description: Library name: sample8

value: 25 hours post fertilization


title: MUT_2

description:
Library name: Sample 5
MUT_2


description:
Library name: UIC3_batchA_Lumbar_Leg
Tissue from Lumbar sampled from Patient UIC-ALS3 with Onset: Leg (batchA)


description:
Library name: UIC5_batchA_Cervical_Leg
Tissue from Cervical sampled from Patient UIC-ALS5 with Onset: Leg (batchA)


title: Mll4-AID_MyoD-ER_A485_4OHT_MLL4

description: Library name: Sample 47

value: Anti-MLL4 #3 (home-made, Kai Ge Lab)


title: MUT_1

description:
Library name: Sample 4
MUT_1


title: 60dpf

description: Library name: sample7

value: 60 days post fertilization


description:
Library name: UIC3_batchA_Cervical_Leg
Tissue from Cervical sampled from Patient UIC-ALS3 with Onset: Leg (batchA)

value: spinal cord

value: Multiple type of cells present in human spinal cord


title: WT_3

description:
Library name: Sample 3
WT_3

value: Testis


description:
Library name: UIC2_batchA_Lumbar_Arm
Tissue from Lumbar sampled from Patient UIC-ALS2 with Onset: Arm (batchA)

value: Batch A


title: WT_2

description:
Library name: Sample 2
WT_2


title: Mll4-AID_MyoD-ER_BRM_4OHT_T7

description: Library name: Sample 44


title: Mll4-AID_MyoD-ER_A485_4OHT_ARID1A

description: Library name: Sample 45

value: Anti-ARID1A/BAF250A (D2A8U) (12354, CST)


title: Mll4-AID_MyoD-ER_A485_4OHT_IgG

description: Library name: Sample 46

value: Anti-IgG (13-0042K, Epicypher)


title: 14dpf

description: Library name: sample4

value: Whole heart

value: 14 days post fertilization

value: Heart cells


title: 21dpf

description: Library name: sample5

value: 21 days post fertilization


title: 7dpf

description: Library name: sample3

value: 7 days post fertilization


title: WT_1

description:
Library name: Sample 1
WT_1


description:
Library name: UIC2_batchA_Cervical_Arm
Tissue from Cervical sampled from Patient UIC-ALS2 with Onset: Arm (batchA)


title: Mll4-AID_MyoD-ER_BRM_4OHT_MLL4

description: Library name: Sample 42

value: Kmt2c-/-;Kmt2dAID/AID preadipocytes

value: Preadipocyte


title: 30dpf

description: Library name: sample6

value: 30 days post fertilization


title: Mll4-AID_MyoD-ER_BRM_4OHT_P300

description: Library name: Sample 43

value: Anti-p300 (C-20)  (sc-585, Santa Cruz, gift from Bing Ren Lab)


description:
Library name: UIC1_batchA_Lumbar_Arm
Tissue from Lumbar sampled from Patient UIC-ALS1 with Onset: Arm (batchA)


title: Mll4-AID_MyoD-ER_BRM_4OHT_IgG

description: Library name: Sample 41


description:
Library name: UIC2_batchA_Medulla_Arm
Tissue from Medulla sampled from Patient UIC-ALS2 with Onset: Arm (batchA)

value: medulla

value: Multiple type of cells present in human medulla


title: 5dpf

description: Library name: sample2

value: 5 days post fertilization


description:
Library name: UIC1_batchA_Cervical_Arm
Tissue from Cervical sampled from Patient UIC-ALS1 with Onset: Arm (batchA)

value: spinal cord

value: Multiple type of cells present in human spinal cord


title: 36hpf

description: Library name: sample1

value: 36 hours post fertilization


title: Mll4-AID_MyoD-ER_BRM_4OHT_ARID1A

description: Library name: Sample 40

value: Anti-ARID1A/BAF250A (D2A8U) (12354, CST)


description:
Library name: UIC3_batchA_Medulla_Leg
Tissue from Medulla sampled from Patient UIC-ALS3 with Onset: Leg (batchA)

value: Batch A


title: RNA-seq of single blastomere (8cell_E05_B7)

city: Fuchu

first: Satoshi

last: Sugimura

state: Tokyo

address: 3-5-8 Saiwai-cho

country: Japan

institute: Tokyo Uiniversity of Agriculture and Technology

description: Library name: 8cell_E05_B7

data_processing:
Adapter trimming was performed using Trim Galore.
Sequence quality was assessed with FastQC.
Reads were aligned to the bovine reference genome (ARS-UCD1.2/bosTau9) using STAR.
Transcript quantification was performed using RSEM.
Assembly: ARS-UCD1.2/bosTau9
Supplementary files format and content: Gene-level count matrix generated by RSEM, provided as tab-delimited text files. Each file contains Ensembl gene IDs and corresponding TPM and expected count values for each sample.

organism: Bos taurus

source_name: blastomerer derived from IVF

growth_protocol: Bovine embryos were produced by in vitro fertilization (IVF) using cumulus-oocyte complexes (COCs) collected from slaughterhouse-derived ovaries. COCs were matured and fertilized in vitro, and then cultured in a time-lapse incubator at 38.5°C under 5% CO₂, 5% O₂, and 90% N₂ until the desired developmental stage.

extract_protocol:
Total RNA was extracted from single blastomeres using the SMART-Seq HT PLUS Kit (Takara Bio, Shiga, Japan), with modifications for single-cell analysis.
Reverse transcription and cDNA amplification were performed using the SMART-Seq HT PLUS Kit (Takara Bio, Shiga, Japan). cDNA quality was evaluated using an Agilent 2200 TapeStation with High Sensitivity D5000 ScreenTape (Agilent Technologies). Libraries were sequenced on the NovaSeq 6000 platform (Illumina) with 150 bp paired-end reads.

treatment_protocol: Not applicable. No experimental treatment was performed.

value: blastomerer derived from IVF

tag: Stage

tag: embryo id

tag: blastomere id


title: RNA-seq of single blastomere (8cell_E05_B8)

description: Library name: 8cell_E05_B8


title: RNA-seq of single blastomere (8cell_E05_B6)

description: Library name: 8cell_E05_B6


title: RNA-seq of single blastomere (8cell_E05_B5)

description: Library name: 8cell_E05_B5


title: 17_5B4-HepG2-10-uM-Lexi-24-hrs_2

city: Philadelphia

first: Priyankara

middle: J

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

description: Library name: 17_5B4-HepG2-10-uM-Lexi-24-hrs_S17

data_processing:
Sequenced reads were aligned using RSEM along with Bowtie2 against the human genome (version: hg19)
DESeq2 was used to normalize the reads and obtain the differentially expressed genes
Assembly: Hg19
Supplementary files format and content: Text RSEM raw counts

molecule: polyA RNA

source_name: Liver Cancer Cells

growth_protocol: Cells were cultured in MEM + 10% FBS + 1% Pen-Strep

extract_protocol:
1x10e6 cells were lysed in Trizol (Sigma) and RNA was extracted according to protocol using Direct-zol RNA mini prep kit (Zymo Research), in-column DNAseI treatment was performed during the extraction. Quantity was assessed using Qubit (Thermofisher) and quality was assessed using RNA Tapestation (Agilent).
500ng of input total RNA was used to make Quantseq libraries for Differential gene expression analysis (Lexogen) according to protocol. 13 PCR cycles were used to amplfy and index cDNA into final library. Quantity and library size was assessed using Qubit and DNA tapestation, respectively. Equimolar quantities of each library were pooled into one sample and sequenced on the Nextseq 2000 (Illumina), single read length 76bp and at least 10M reads/sample.

treatment_protocol: Cells were treated with vehicle (DMSO) or 10 µM Lexibulin for 8 or 24 hours in normal culture media

value: Liver Cancer Cells

value: Liver Cancer

value: 10 µM Lexibulin 24 hours


title: 18_5B4-HepG2-10-uM-Lexi-24-hrs_3

description: Library name: 18_5B4-HepG2-10-uM-Lexi-24-hrs_S18


title: WERI-Rb1 sgELF2, Topotecan, rep3

city: Guangzhou

first: Jing

last: Zhuang

address: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, No. 54 South Xianlie Road

description:
Library name: WERI_sgELF2_TPT_3
ELF2 knockout, topotecan treatment, biological replicate 3

data_processing:
Raw reads were filtered using fastp to remove adapters, low-quality reads, and undetermined bases. Clean reads were mapped to the Homo sapiens reference genome (GRCh38) using HISAT2. Transcript assembly was performed using StringTie. Transcriptomes from all samples were merged with gffcompare to generate a comprehensive reference transcriptome. Expression levels of mRNAs were quantified by StringTie in FPKM units. Differential expression analysis was performed using edgeR, with thresholds of fold change >2 or <0.5 and p < 0.05.
Assembly: GRCh38 (GCF_000001405.39)
Supplementary files format and content: gene_fpkm_matrix.txt: A tab-delimited text file containing FPKM values for all protein-coding genes across all samples. Columns are labeled with the corresponding library name.

growth_protocol: WERI-Rb1 cells were cultured in RPMI-1640 medium supplemented with 10% fetal bovine serum in a 5% CO2 atmosphere.

extract_protocol:
Total RNA was extracted using TRIzol reagent (Invitrogen, Carlsbad, CA, USA) following the manufacturer’s instructions. RNA purity was assessed by NanoDrop ND-1000 spectrophotometer, and integrity was evaluated with Agilent Bioanalyzer 2100 (RIN > 7.0).
Poly(A) RNA was purified from 1 µg total RNA using Dynabeads Oligo(dT)25 (Thermo Fisher), fragmented with Magnesium RNA Fragmentation Module (NEB) at 94 °C for 5–7 min, and reverse-transcribed with SuperScript II (Invitrogen). Second-strand cDNA synthesis incorporated dUTP, followed by adapter ligation and size selection (~300 ± 50 bp) with AMPure XP beads. Libraries were amplified by PCR (8 cycles) and sequenced on an Illumina NovaSeq 6000 platform (LC-Bio Technology Co., Ltd., Hangzhou, China) with 2×150 bp paired-end reads.

treatment_protocol: Cells were treated with either vehicle (DMSO) or 30 nM topotecan for 96 hours prior to RNA extraction.

value: 30 nM topotecan for 96 hours


title: Mll4-AID_MyoD-ER_IAA_4OHT_T7

description: Library name: Sample 39

value: Kmt2c-/-;Kmt2dAID/AID preadipocytes

value: 5-Ph-IAA (2h) 4OHT (1h)


title: Mll4-AID_MyoD-ER_IAA_4OHT_P300

description: Library name: Sample 38

value: Preadipocyte


title: 16_5B4-HepG2-10-uM-Lexi-24-hrs_1

description: Library name: 16_5B4-HepG2-10-uM-Lexi-24-hrs_S16


title: RNA-seq of single blastomere (8cell_E05_B3)

description: Library name: 8cell_E05_B3


title: WERI-Rb1 sgELF2, Topotecan, rep1

institute: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University

description:
Library name: WERI_sgELF2_TPT_1
ELF2 knockout, topotecan treatment, biological replicate 1


title: WERI-Rb1 sgELF2, Topotecan, rep2

description:
Library name: WERI_sgELF2_TPT_2
ELF2 knockout, topotecan treatment, biological replicate 2


title: RNA-seq of single blastomere (8cell_E05_B4)

description: Library name: 8cell_E05_B4


title: 15_WT-HepG2-10-uM-Lexi-24-hrs_3

description: Library name: 15_WT-HepG2-10-uM-Lexi-24-hrs_S15

value: WT p53


title: RNA-seq of single blastomere (8cell_E05_B2)

description: Library name: 8cell_E05_B2


title: Mll4-AID_MyoD-ER_IAA_4OHT_MLL4

description: Library name: Sample 37

value: Anti-MLL4 #3 (home-made, Kai Ge Lab)


title: 14_WT-HepG2-10-uM-Lexi-24-hrs_2

description: Library name: 14_WT-HepG2-10-uM-Lexi-24-hrs_S14


title: WERI-Rb1 sgELF2, Vehicle, rep3

description:
Library name: WERI_sgELF2_Veh_3
ELF2 knockout, vehicle treatment, biological replicate 3

value: vehicle


title: Mll4-AID_MyoD-ER_IAA_4OHT_IgG

description: Library name: Sample 36

value: Anti-IgG (13-0042K, Epicypher)


title: RNA-seq of single blastomere (8cell_E04_B8)

description: Library name: 8cell_E04_B8


title: RNA-seq of single blastomere (8cell_E04_B7)

description: Library name: 8cell_E04_B7


title: Mll4-AID_MyoD-ER_4OHT_P300

description: Library name: Sample 33

value: 4OHT (1h)

value: Anti-p300 (C-20)  (sc-585, Santa Cruz, gift from Bing Ren Lab)


title: Mll4-AID_MyoD-ER_IAA_4OHT_ARID1A

description: Library name: Sample 35


title: Mll4-AID_MyoD-ER_4OHT_T7

description: Library name: Sample 34


title: RNA-seq of single blastomere (8cell_E04_B6)

description: Library name: 8cell_E04_B6


title: WERI-Rb1 NTC, Topotecan, rep2

description:
Library name: WERI_NTC_TPT_2
Non-targeting control, Topotecan treatment, biological replicate 2

value: NTC


title: 11_5B4-HepG2-10-uM-Lexi-8-hrs_2

description: Library name: 11_5B4-HepG2-10-uM-Lexi-8-hrs_S11

value: 10 µM Lexibulin 8 hours


title: 13_WT-HepG2-10-uM-Lexi-24-hrs_1

description: Library name: 13_WT-HepG2-10-uM-Lexi-24-hrs_S13


title: WERI-Rb1 sgELF2, Vehicle, rep1

description:
Library name: WERI_sgELF2_Veh_1
ELF2 knockout, vehicle treatment, biological replicate 1


title: Mll4-AID_MyoD-ER_4OHT_MLL4

description: Library name: Sample 32


title: RNA-seq of single blastomere (8cell_E04_B5)

description: Library name: 8cell_E04_B5


title: 12_5B4-HepG2-10-uM-Lexi-8-hrs_3

description: Library name: 12_5B4-HepG2-10-uM-Lexi-8-hrs_S12


title: 10_5B4-HepG2-10-uM-Lexi-8-hrs_1

description: Library name: 10_5B4-HepG2-10-uM-Lexi-8-hrs_S10


title: RNA-seq of single blastomere (8cell_E05_B1)

description: Library name: 8cell_E05_B1


title: WERI-Rb1 NTC, Topotecan, rep1

description:
Library name: WERI_NTC_TPT_1
Non-targeting control, Topotecan treatment, biological replicate 1


title: 9_WT-HepG2-10-uM-Lexi-8-hrs_3

description: Library name: 9_WT-HepG2-10-uM-Lexi-8-hrs_S9


title: RNA-seq of single blastomere (8cell_E04_B4)

description: Library name: 8cell_E04_B4


title: WERI-Rb1 sgELF2, Vehicle, rep2

description:
Library name: WERI_sgELF2_Veh_2
ELF2 knockout, vehicle treatment, biological replicate 2


title: WERI-Rb1 NTC, Topotecan, rep3

description:
Library name: WERI_NTC_TPT_3
Non-targeting control, Topotecan treatment, biological replicate 3


title: Mll4-AID_MyoD-ER_4OHT_IgG

description: Library name: Sample 31


title: 8_WT-HepG2-10-uM-Lexi-8-hrs_2

description: Library name: 8_WT-HepG2-10-uM-Lexi-8-hrs_S8


title: WERI-Rb1 NTC, Vehicle, rep3

description:
Library name: WERI_NTC_Veh_3
Non-targeting control, vehicle treatment, biological replicate 3


title: Mll4-AID_MyoD-ER_4OHT_ARID1A

description: Library name: Sample 30

value: Anti-ARID1A/BAF250A (D2A8U) (12354, CST)


title: WERI-Rb1 NTC, Vehicle, rep2

description:
Library name: WERI_NTC_Veh_2
Non-targeting control, vehicle treatment, biological replicate 2


title: Mll4-AID_MyoD-ER_unt_T7

description: Library name: Sample 29

value: untreated


title: 7_WT-HepG2-10-uM-Lexi-8-hrs_1

description: Library name: 7_WT-HepG2-10-uM-Lexi-8-hrs_S7

value: Liver Cancer Cells


title: RNA-seq of single blastomere (8cell_E04_B2)

description: Library name: 8cell_E04_B2

value: blastomerer derived from IVF


title: Mll4-AID_MyoD-ER_unt_P300

description: Library name: Sample 28


title: WERI-Rb1 NTC, Vehicle, rep1

description:
Library name: WERI_NTC_Veh_1
Non-targeting control, vehicle treatment, biological replicate 1


title: 6_5B4-HepG2--DMSO-control_3

description: Library name: 6_5B4-HepG2--DMSO-control_S6

value: Liver Cancer

value: Vehicle (DMSO)


title: Mll4-AID_MyoD-ER_unt_MLL4

description: Library name: Sample 27

value: Kmt2c-/-;Kmt2dAID/AID preadipocytes


title: RNA-seq of single blastomere (8cell_E04_B1)

description: Library name: 8cell_E04_B1


title: RNA-seq of single blastomere (8cell_E04_B3)

description: Library name: 8cell_E04_B3


title: USP28 knockout_Initial_T0

city: New York

first: Bic

last: MSKCC

state: NY

address: 1275 York Ave.

institute: Memorial SLoan-Kettering Cancer Center

description: Library name: s_USP28_T0

data_processing:
fastx-toolkit (0.0.13) is used to find the sequence that appears in the vector 5′ to all sgRNA sequences, as well as grab the 20nt sgRNA sequence, and count how many occurrences that sequence was found in each sample. (fastx_reverse_complement -Q33, fastx_clipper -M 20 -c -a CGGTGTTTCGTCCTTTCCACAAG -Q33, fastx_reverse_complement, fastx_trimmer -p 1 -q 20, fastx_collapser, fasta_formatter -t )
R was then used to map back to a reference file of all possible sgRNAs present in the library. Git repo with code used for data processing: https://github.com/soccin/ScreenSEQ/tree/proj/11458
Assembly: GRCh38.p14
Supplementary files format and content: counts files are excels with columns containing sgRNA sequence, gene name, probe ID, what library reaction the sgRNA is from and counts for the samples.
Library strategy: CRISPR Screen

growth_protocol: hTERT-RPE1 cells were cultured in DME/F12 (1:1) +15mM HEPES + 2.4mM Glutamate media, supplemented with 10% FBS and 1% penicillin/streptomycin. Puromycin selection was done at 10mg/mL for 3 days. Throughout the screen cells were split every 4-5 days depending on reaching confluency, re-seeding a minimum of 10 million cells. Multilayer flasks were used to streamline culture work.

extract_protocol:
Genomic DNA was isloated using QIAGEN DNA Blood Maxi kit protocol, from the BRAOD institute
gRNAs were amplified with barcoded primers following the “PCR of sgRNAs, shRNAs, and ORFs from genomic DNA for Illumina sequencing”  protocol, with the following P7 barcode primers for the indicated libraries. s_USP28_T0 : A01 (P7_MT01) s_USP28_T4 : A02 (P7_MT02) s_WT-T4 : A01 (P7_MT01) s_P53-T4 : A02 (P7_MT02) s_WT-T0 : A07 (P7_MT07) s_P53-T0 : A08 (P7_MT08)


title: 5_5B4-HepG2--DMSO-control_2

description: Library name: 5_5B4-HepG2--DMSO-control_S5


title: RNA-seq of single blastomere (8cell_E03_B8)

description: Library name: 8cell_E03_B8


title: USP28 knockout_End_T4

description: Library name: s_USP28_T4


title: 4_5B4-HepG2--DMSO-control_1

description: Library name: 4_5B4-HepG2--DMSO-control_S4


title: RNA-seq of single blastomere (8cell_E03_B7)

description: Library name: 8cell_E03_B7


title: Mll4-AID_MyoD-ER_unt_ARID1A

description: Library name: Sample 25


title: 3_WT-HepG2-DMSO-control_3

description: Library name: 3_WT-HepG2-DMSO-control_S3


title: RNA-seq of single blastomere (8cell_E03_B6)

description: Library name: 8cell_E03_B6


title: 2_WT-HepG2-DMSO-control_2

description: Library name: 2_WT-HepG2-DMSO-control_S2


title: PMCs, DNP, 6 hours, biol rep7

city: Heidelberg

first: Qihua

last: Liang

state: Baden-Württemberg

address: Im Neuenheimer Feld 366

department: Institute of Pharmacology

institute: Universitat Heidelberg

description:
Library name: Sample 2.4.5
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_D_6_7

data_processing:
Raw RNA-Seq reads were processed using the nf-core/rnaseq pipeline (version 3.14.0). The pipeline was executed using Nextflow with the Singularity profile.
Adapter and quality trimming was performed using Trim Galore! with nf-core/rnaseq pipeline default settings.
Read alignment was performed against the Mus musculus GRCm39 reference genome (GRCm39.primary_assembly.genome.fa) using STAR (version 2.7.11a).
Gene-level quantification was carried out using RSEM (version 1.3.1) with annotations from GENCODE release M35 (gencode.vM35.primary_assembly.annotation.gtf).
Technical replicates were aggregated using the DESeq2 collapseReplicates function to produce a raw gene count matrix (raw_counts_All_Samples.txt). To account for technical variation, batch effects were corrected using ComBat-seq. Finally, library size normalization was applied via the DESeq2 median-of-ratios method, with the final output saved as normalized_counts_ComBatSeq_DESeq2_All_Samples.txt.
Assembly: mm39
Supplementary files format and content: tab-delimited text file includes raw counts for all samples
Supplementary files format and content: tab-delimited text file includes batch adjusted and DESeq2 normalized counts for all samples

source_name: connective tissue

growth_protocol: The isolated cell suspension was cultured at 37∘C  and 5% CO2. Two days after the isolation, the culture medium was changed, and all non-adherent cells were removed by aspiration. On day nine, the cells were split and cultivated further at a concentration 1×10^6 of cells/mL. The PMCs were used for the experiments 12-16 days after isolation.

extract_protocol:
For RNA isolation, each sample (0.2 – 2 x 105 PMCs) was lysed in 400 µL RLT Buffer supplemented with 4 μL-Mercaptoethanol. RNA extraction was performed using the RNeasy Micro Kit (Qiagen, Cat. No. 74004). RNA was eluted in 14 µL of buffer, and 10 µL of the eluate was used for library preparation.
For samples 1.1–1.3, transcript libraries were constructed using the Ovation SoLo RNA-Seq Library Preparation Kit (NuGEN/Tecan), utilizing 25 ng of cDNA per sample as input. Sequencing was performed on an Illumina NextSeq 2000 platform (EMBL Heidelberg Genomics Core Facility). For samples 2.1–2.4, libraries were prepared using the MGIEasy Fast RNA Library Prep Set following ribosomal RNA removal with the MGIEasy rRNA Depletion Kit V1.3. Sequencing was conducted on the DNBSEQ-G400 platform (MGI Tech) using the DNBSEQ-G400RS High-throughput Sequencing Reagent Set (FCL PE100).

treatment_protocol: Each PMC preparation was stimulated with four experimental conditions: adenosine (ADO, 10 µM) for 2 hours, compound 48/80 (C48/80, 50µg/mL) for 6 hours, antigen (dinitrophenyl (DNP)-human serum albumin 100 ng/mL) for 2 hours, and antigen (dinitrophenyl (DNP)-human serum albumin 100 ng/mL) for 6 hours.

value: connective tissue

value: primary peritoneal mast cells

value: WT

value: DNP

tag: time

value: 6 hours


title: RNA-seq of single blastomere (8cell_E03_B5)

description: Library name: 8cell_E03_B5


title: hlsec, shPML_TNFa, rep1

city: Chengdu

first: Enjiang

last: Lai

address: 37 Guoxue Alley

institute: Sichuan University

description:
Library name: shPML_TNF_1
column in prcessed data: shPML.TNF_R1

data_processing:
RNA-seq reads were aligned to the hg38 revision of the human genome using STAR (v2.7.3a). Mapped reads were counted using HTseq-count (v0.11.3) Gene expression was quantified as reads per kilobase per million mapped reads (RPKM).
Assembly: hg38
Supplementary files format and content: tab-delimited txt file of RPKM

source_name: Human liver sinusoidal endothelial cells

extract_protocol:
The cells used for RNA-seq were collected and washed twice with cold PBS, and total RNA was extracted. Then mRNA was purified from total RNA using poly-T oligo-attached magnetic beads.
Strand cDNA was synthesized using random hexamer primers, followed by second-strand cDNA synthesis. The libraries then underwent PCR amplification and purification. After the libraries passed quality control, they were pooled and sequenced on the Illunima sequencing platform.

treatment_protocol: Sequences of BRD4 and PML genes were inserted into the backbone vector GV513 and GV492, respectively, according to the manufacturer’s protocols. Lentivector overexpressing BRD4 (BRD4OE) or PML (PMLOE) genes were transfected to HEK293T cells, and supernatants containing lentivirus were collected. hLSECs at 30% confluence were transduced with BRD4OE or PMLOE lentivirus using the transfection reagent (Genechem) according to the manufacture’s protocols. Then puromycin was treated for 24 hours to select positive cells.


title: hlsec, shCON_TNFa, rep3

description:
Library name: shCON_TNF_3
column in prcessed data: shCON.TNF_R3

value: Human liver sinusoidal endothelial cells


title: RNA-seq of single blastomere (8cell_E03_B4)

description: Library name: 8cell_E03_B4


title: 1_WT-HepG2-DMSO-control_1

description: Library name: 1_WT-HepG2-DMSO-control_S1

value: WT p53


title: Mll4-AID_MyoD-ER_unt_IgG

description: Library name: Sample 26

value: Preadipocyte

value: Anti-IgG (13-0042K, Epicypher)


title: hlsec, shPML_TNFa, rep3

description:
Library name: shPML_TNF_3
column in prcessed data: shPML.TNF_R3


title: PMCs, C48/80, 6 hours, biol rep7

description:
Library name: Sample 2.4.3
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_C_6_7


title: hlsec, shCON_TNFa, rep1

description:
Library name: shCON_TNF_1
column in prcessed data: shCON.TNF_R1


title: RNA-seq of single blastomere (8cell_E03_B2)

description: Library name: 8cell_E03_B2


title: Gpnmb+_Pool 2

city: Durham

first: Paloma

middle: B

last: Liton

state: NC

address: DUMC Box 3802

institute: Duke University

description:
Library name: 526_523_TM
Column name in Normalized counts All Samples DBA and Gpnmb_Iridocornea: 5C_4a_1101_TM in

data_processing:
RNA-seq data was processed using the fastp toolkit trimming low-quality bases and sequencing adapters from the 3’ end of reads, then mapped to GRCm39 (downloaded from Ensembl, version 110) using the STAR RNA-seq alignment tool. Reads aligning to a single genomic location were summed across genes. Genes with at least 10 mapped reads in at least a single sample, were normalized and differential expression was carried out using the DESeq2 Bioconductor package implemented for the R programming environment. Consistent with the recommendation of the DESeq authors, independent filtering was utilized prior to calculating adjusted p-values and moderated log2 fold-changes were derived using the ashr package.
Assembly: GRCm39 (downloaded from Ensembl, version 110) using the STAR RNA-seq alignment tool
Supplementary files format and content: CVS file includes raw counts for each sample

source_name: Iridocorneal angle region tissue

growth_protocol: 12 month old, male and female mice housed under a 12-hour light/dark cycle, received a standard mouse diet, and had free access to water

extract_protocol:
Mouse eyes were enucleated and immediately placed in individual 1.5 ml tubes containing 0.5 mL of RNAprotect® Tissue Reagent (Qiagen, 76104) and stored at 4 °C until dissection. For dissection, the anterior and posterior chambers were separated. The anterior segment was divided into four quadrants, and the iris and ciliary body were carefully removed under a dissection microscope to avoid damaging the trabecular meshwork (TM). The iridocorneal angle was carefully dissected, with trimming of non-target tissues to minimize contamination. Iridocorneal angles from three individual mice were pooled in a 1.5 mL tube containing 100 µL of RNAprotect and stored at 4 °C until RNA extraction. RNA isolation was performed using the RNeasy® Plus Micro Kit (Qiagen,74034) according to the manufacturer’s protocol, with some modifications. Briefly, RNAprotect was removed, and tissues were lysed in 350 µL Buffer RLT Plus supplemented with 1% β-mercaptoethanol and Molecular Grinding Resin (Gbiosciences, 786038). Tissues were homogenized using motorized pestles for 1 min and passed through a Qiashredder column (Qiagen, 79656) by centrifugation at 13,000 rpm for 1 min. The lysate was transferred to a gDNA Eliminator spin column and centrifuged at ≥8000 × g for 30 s. An equal volume of 70% ethanol was added to the flow-through, mixed well, and the mixture was loaded onto an RNeasy MinElute spin column. After centrifugation at ≥8000 × g for 15 s, the column was sequentially washed with 700 µL Buffer RW1, 500 µL Buffer RPE, and 500 µL of 80% ethanol (repeated three times), each followed by centrifugation at ≥8000 × g. The column was then centrifuged at full speed for 5 min with the lid open to remove residual ethanol. RNA was eluted with 14–22 µL RNase-free water and collected by centrifugation at full speed for 1 min. RNA concentration and purity were measured using a NanoDrop spectrophotometer (DS-11+ spectrophotometer, DeNovix).
The Sequencing and Genomics Technologies Core Facility at Duke University performed RNA-sequencing. The SGT enriched mRNA from total RNA and reversed transcribed into cDNA to build sequencing libraries using the Kapa mRNA HyperPrep Kit from Roche (Code: KK8581). Libraries were pooled to equimolar concentration and sequenced on the NovaSeq X Plus 1.5B lane to produce 100 bp paired-end reads targeting 50M reads/sample

treatment_protocol: No treatment

value: Iridocorneal angle region tissue


title: hlsec, shPML_TNFa, rep2

description:
Library name: shPML_TNF_2
column in prcessed data: shPML.TNF_R2


title: PMCs, CONTROL, 0 hour, biol rep7

description:
Library name: Sample 2.4.1
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_0_7

value: CONTROL

value: 0 hour


title: hlsec, shCON_veh, rep3

description:
Library name: shCON_veh_3
column in prcessed data: shCON.veh_R3


title: PMCs, DNP, 2 hours, biol rep7

description:
Library name: Sample 2.4.4
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_D_2_7

value: 2 hours


title: PMCs, ADO, 2 hours, biol rep7

description:
Library name: Sample 2.4.2
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_2_7

value: ADO


title: PMCs, DNP, 6 hours, biol rep6

description:
Library name: Sample 2.3.5
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_D_6_6


title: DBA/2J_Pool 3

description:
Library name: 1020_1d_963TM
Column name in Normalized counts All Samples DBA and Gpnmb_Iridocornea: 5C_4a_1101_TM in


title: RNA-seq of single blastomere (8cell_E02_B8)

description: Library name: 8cell_E02_B8

value: blastomerer derived from IVF


title: RNA-seq of single blastomere (8cell_E03_B1)

description: Library name: 8cell_E03_B1


title: hlsec, shCON_veh, rep2

description:
Library name: shCON_veh_2
column in prcessed data: shCON.veh_R2


title: hlsec, shCON_TNFa, rep2

description:
Library name: shCON_TNF_2
column in prcessed data: shCON.TNF_R2


title: DBA/2J_Pool 1

description:
Library name: 5C_4a_1101_TM
Column name in Normalized counts All Samples DBA and Gpnmb_Iridocornea: 5C_4a_1101_TM in


title: PMCs, DNP, 2 hours, biol rep6

description:
Library name: Sample 2.3.4
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_D_2_6


title: RNA-seq of single blastomere (8cell_E03_B3)

description: Library name: 8cell_E03_B3


title: RNA-seq of single blastomere (8cell_E02_B7)

description: Library name: 8cell_E02_B7


title: DBA/2J_Pool 2

description:
Library name: 824_749_750_TM
Column name in Normalized counts All Samples DBA and Gpnmb_Iridocornea: 5C_4a_1101_TM in


title: PMCs, C48/80, 6 hours, biol rep6

description:
Library name: Sample 2.3.3
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_C_6_6


title: hlsec, shCON_veh, rep1

description:
Library name: shCON_veh_1
column in prcessed data: shCON.veh_R1


title: Gpnmb+_Pool 1

description:
Library name: 525_522TM
Column name in Normalized counts All Samples DBA and Gpnmb_Iridocornea: 5C_4a_1101_TM in


title: PMCs, ADO, 2 hours, biol rep6

description:
Library name: Sample 2.3.2
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_2_6


title: hlsec, iBET151, rep3

description: Library name: iBET151_R3

treatment_protocol: Low-passage cells were starved with basal endothelial medium for 2 hours followed by treatment with BRD4 inhibitor (iBET151, 2.5 mol/L, Selleck, S2780) for 2 hours. Then, human recombinant TNFα (50 ng/mL; PeproTech, 300-01A) was treated to hLSECs for 6 hours before cells were collected for further analysis.


title: RNA-seq of single blastomere (8cell_E02_B6)

description: Library name: 8cell_E02_B6


title: RiboSeq_CtrlR4

city: Schlieren

first: Umesh

last: Ghoshdastider

address: Wagistrasse 12

country: Switzerland

institute: University of Zurich

description:
Library name: RiboSeq_CtrlR4
Ribosome Profiling from nontargeting control keratinocytes Replicate4
Ribo-seq_counts.csv

library_source: other

data_processing:
Bioinformatic analysis of single-cell RNA sequencing data: Raw sequencing data, consisting of BCL files, were demultiplexed using Illumina bcl2fastq v2.20 with default settings. One mismatch in the sample barcode sequences was allowed. The resulting fastq files were processed using the BD Rhapsody Pipeline (version 1.9.1) hosted at the Seven Brigdge’s cloud platform (https://www.sevenbridges.com/). A custom mouse reference genome, based on Gencode vM25 (GRCm38.p6), was generated and combined with the 720 sgRNAs sequences. Based on STAR aligner, a custom STAR reference genome was created and the Refined Putative Cell Calling option was disabled during the pipeline execution. For downstream analysis, unique molecular identifier (UMI) count data was corrected using BD Genomics Recursive Substitution Error Correction (RSEC). Processing PCR sgRNA dial-out data PCR dial-out data was processed using an in-house custom Python script as previously described (15). Three 9-nucleotide cell barcodes were extracted from the 97 possible barcodes from Read 1. One mismatch was allowed per barcode (Hamming distance of 1). The script also retrieved the 8-nucleotide UMI sequences from Read 1, as specified in the BD manual. For valid barcodes, sgRNAs were identified in Readd 2 by performing an exact match search for the 20-nucleotide sgRNA sequence. UMI counts per cell were deduplicated and sgRNA UMI counts per cell were computed. If multiple sgRNAs were detected for one cell, the cell was assigned to a specific sgRNA only if its UMI count exceeded the 99th percentile. Processing of single-cell RNA sequencing data The Scrublet Python package was used to remove doublets from each dataset separately. Genes expressed in a minimum of 5 cells were retained and an expected double rate of 20% (as estimated by BD Rhapsody) was applied. After filtering, seven P4 single-cell RNA-seq datasets and eight P60 datasets were merged using Seurat. Only cells containing detected sgRNAs and classified as singlets were selected for further analysis. Cells were filtered based on the following criteria: UMI counts > 500, UMI counts lower than the 0.99 quantile (to remove outliers) and mitochondrial gene content < 20%. Filtered datasets were then processed using Seurat’s standard pipeline. Gene expression was normalized using the NormalizeData function, which scales counts to 10,000 per cell followed by a natural log transformation (log1p). The FindVariableGenes function identified 2,000 highly variable genes, and data scaling was performed using ScaleData. Principal Component Analysis (PCA) was conducted with RunPCA and the ElbowPlot function was used to determine the top principal components that contributed to explaining the variance of the data for each dataset at P4 and P60. The FindNeighbors function was applied to identify nearest neighbors based in the selected principal components. Clustering was performed using FindClusters with Louvain modularity optimization. To visualize the data in two dimensions, UMAP (Uniform Manifold Approximation and Projection) was used. No batch effects were observed in the P4 and P60 datasets, so batch effect correction was not performed.
Assembly: Gencode Mouse vM25 (GRCm38.p6)
Supplementary files format and content: Comma-separated value files and matrix files with count numbers, *_counts.csv.gz files contain Cell Barcode, Gene Symbol and RSEC adjusted counts.
Library strategy: Ribo-Seq

molecule: other

growth_protocol: All mice were housed at the Laboratory Animal Services Center (LASC) of the University of Zurich in individually-ventilated cages in a humidity- and light-controlled environment (22˚C, 45-50%, 12 h light/dark cycle), and had access to food and water ad libitum.

extract_protocol:
P4 back skin was processed as previously described (15). Fat was scraped off and the back skin was washed once in cold PBS and then placed in dispase (Corning #354235) dermis-side facing down, incubated for 35 min at 37 °C on an orbital shaker. Next, epidermis was separated from dermis with fine forceps and placed in 4 ml 0.25% Trypsin-EDTA (1×, Gibco; 25200056) for 15 min at 37 °C with orbital shaking. Epidermis was torn into smaller pieces and washed with cold PBS, pipetted vigorously up and down to achieve a single-cell suspension. Suspension was filtered through a 70-µm and subsequently a 40-µm strainer (Corning; #431750, #431751). Suspension was then centrifuged for 10 min at 400 g and resuspended in FACS buffer (PBS + 2% chelexed FBS (FBS(-)) + DAPI). P60 back skin was processed as previously described (15). Back skin was harvested with surgical scissors and placed dermis-facing up on a styrofoam rack with pins. Fat was scraped off with a scalpel. Fat-free skin was then washed, dermis-side facing down in 1× PBS. Skin was then placed in the same orientation in 0.5% Trypsin-EDTA (10×, Gibco; 15400054) and incubated at 37 °C on an orbital shaker for 25 min (females) or 50 min (males). Using a glass microscopy slide, skin was secured to the bottom of the dish and scraped again with a scalpel until it started to diverge. Excess cold PBS with 2% FBS(-) was added to neutralize trypsin. Cell suspension was strained on ice first through a 70-µm strainer, washed with an additional 15 ml of cold 1× PBS with 2% FBS (-) and then filtered through a 40-µm strainer. Cell suspension was spun down at 400 g for 10 min and resuspended in FACS buffer. P4: a total of 18 injected P4 mice were collected and assessed in 7 independent single-cell RNA sequencing runs. P60: a total of 7 injected mice were collected and assessed in 8 independent single-cell RNA sequencing runs.
Library construction protocol for scRNA-seq: EGFP-positive cells were sorted on a BD FACSAria III using a 70-µm nozzle and processed for single-cell capture with a BD Rhapsody Single-Cell Analysis System using BD Rhapsody Cartridge Kit (#633733). For P4, 7 separate cartridges with an average of 76,613 cells were loaded. For P60, 8 cartridges with an average of 68,171 cells were loaded. Reverse transcription from beads and sequencing library production was carried out according to the manufacturer’s instructions (BD Biosciences, doc ID: 210967 rev. 1.0). Library construction protocol for RNA-seq: RNA was isolated from lysates from nontargeting control and Gm10076 C7 keratinocytes as described in the Ribosome profiling section using TRIzol®  LS Reagent (Invitrogen, 10296028) and column purified with a Direct-zol RNA MiniPrep kit (Zymo Research, R2052). Libraries were prepared and sequenced at Novogene. Library construction protocol for Other:  As previously described, the sgRNA-containing region was amplified from the BD Rhapsody beads using a nested KAPA Polymerase reaction. In the first polymerase reaction (PCR1), the Rhapsody beads from each separate cartridges were resuspended and distributed into 4 separate 50 µL reactions  (100 µl KAPA HiFi HotStart ReadyMix (Roche 07958935001), 6 µl forward primer (5′-ACACGACGCTCTTCCGATCT-3′, 10 μM), 6 µl reverse primer (5′-TCTTGTGGAAAGGACGA-3′, 10 μM), 12 μl Bead RT/PCR Enhancer reagent from BD Biosciences and 72 μl nuclease-free water for a total volume of 200 μl). PCR1 conditions were as follows: initial denaturation at 95 °C for 5 min, followed by 25 cycles of denaturation 95 °C for 30 s, annealing 53 °C for 30 s, extension 72 °C for 20 s and a final extension of 10 min at 72 °C. The 4 x 50 µl PCR1 product was pooled, and beads were removed using a magnet. Amplicons were cleaned up using Agencourt AMPure XP beads (Beckman Coulter, A63881) following manufacturer’s instructions. 3 μl of cleaned up PCR1 was used as template for PCR 2 (25 µL KAPA HiFi HotStart Ready Mix, 2 μl forward primer (5′-ACACGACGCTCTTCCGATCT-3′, 10 μM), 2 μl reverse primer (CAGACGTGTGCTCTTCCGATCTCTTGTGGAAAGGACGAAACA*C*C*G-3′, 10 μM), 18 μl nuclease-free water) for a total volume of 50 μl. PCR2 conditions were as follows: initial denaturation at 95 °C for 3 min, followed by 10 cycles of denaturation 95 °C for 30 s, annealing 60 °C for 3 min, extension 72 °C for 60 s, and a final extension of 5 min at 72 °C. Agencourt AMPure XP beads were used to clean up PCR2 product according to the manufacturer’s instructions  before the indexing PCR. Indexing PCR was performed according to BD Biosciences ‘mRNA Targeted Library Preparation’ protocol (doc ID: 210968 rev. 3.0). Library construction protocol for Ribo-seq: Ribosome profiling was conducted with some adjustments of the original method (64, 65). For RNA digestion, cell lysates were incubated with 1 U/µg of RNase 1 (Epicentre/Lucigen, 10U/µL) for 45 minutes at room temperature. Reaction was stopped by adding 4 µL of SUPERase Inhibitor (20 U/µL, SUPERase-InTM RNase Inhibitor Thermo Scientific AM2694). To eliminate free nucleotides and isolate ribosome-protected fragments, the RNase I-treated samples were processed using Amersham MicroSpin S-400 HR columns (Cytiva, 27514001). RNA extraction was performed utilizing TRIzol® LS Reagent (Invitrogen 10296028) in combination with the Direct-zolTM RNA MiniPrep kit (Zymo Research, R2052). Total RNA concentration was measured using QubitTM RNA broad range kit (Invitrogen, Q10210). Ribosome footprints were excised and extracted from a 15% TBE-UREA gel (Invitrogen, EC68852BOX) using the 20nt, 22nt, 27nt, 30nt and 60nt oligonucleotides mix as markers (1 µL of 10 µM dilution in 19µL RNase-free H2O). Extraction was performed overnight with RNA extraction buffer (300mM NaOAc pH=5.5, 1mM EDTA, 0.25% v/v SDS). T4 PNK (NEB, M0201S) end-healing reaction was performed on the ribosome footprints for 1 hour at 37˚C. Linker ligation reaction followed using T4 RnI truncated K227Q (NEB, M0351L) in combination with 20 µM of preadenylated linker for 3 hours at 22˚C (50% w/v PEG-8000, 10X T4 RNA ligase, preadenylated linker, T4 RnI truncated K227Q). Ribosomal RNA was depleted using biotinylated rRNA depletion oligos (2 µM) by mixing 10 µL of sample with 10 µL of oligo mix and incubated for 2 minutes at 80˚C, followed by a gradual decrease in temperature to 25˚C. MyOne Streptavidin dynabeads (Life Technologies, 65001) were used to recover the samples. Reverse transcription was performed on the Ribosomal RNA-free samples using SuperScriptTM IV Reverse Transcriptase (Invitrogen, 18090050). Footprints were excised and extracted from a 15% TBE-UREA gel. Extraction was performed overnight with DNA extraction buffer (10mM Tris-HCl pH=8, 300mM NaCl, 1mM EDTA). cDNA was circularized using CircLigase II (100 U/µL, Biosearch, CL4115K) for 1 hour at 60˚C. cDNA was amplified using Phusion High-Fidelity DNA polymerase (Thermo Scientific, F530S) using an individual index per sample. 6-12 cycle amplicons were visualized using an 8% TBE non-denaturing polyacrylamide gel to determine optimal cycle number. PCR was carried out with 7-9 cycles and analyzed on an 8% TBE non-denaturing polyacrylamide gel (Invitrogen, EC61252BOX). DNA was excised from gel and extracted using DNA extraction buffer by incubating overnight with gentle shaking. DNA was recovered and precipitated in 10 mM Tris-HCl pH=8. DNA was quantified using Qubit TM 1X dsDNA High Sensitivity Assay (Invitrogen, Q33230) and quality was verified using the Bioanalyzer High Sensitivity DNA kit (Agilent, 5067-4626). Samples were pooled and prepared for sequencing.

value: Keratinocytes


title: hlsec, iBET151, rep2

description: Library name: iBET151_R2


title: RiboSeq_KOR4

description:
Library name: RiboSeq_KOR4
Ribosome Profiling from Gm10076 C7 knockout keratinocytes Replicate4
Ribo-seq_counts.csv


title: PMCs, CONTROL, 0 hour, biol rep6

description:
Library name: Sample 2.3.1
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_0_6


title: RNA-seq of single blastomere (8cell_E02_B5)

description: Library name: 8cell_E02_B5


title: RiboSeq_CtrlR3

description:
Library name: RiboSeq_CtrlR3
Ribosome Profiling from nontargeting control keratinocytes Replicate3
Ribo-seq_counts.csv


title: RiboSeq_KOR3

description:
Library name: RiboSeq_KOR3
Ribosome Profiling from Gm10076 C7 knockout keratinocytes Replicate3
Ribo-seq_counts.csv


title: RNA-seq of single blastomere (8cell_E02_B3)

description: Library name: 8cell_E02_B3


title: RiboSeq_KOR2

description:
Library name: RiboSeq_KOR2
Ribosome Profiling from Gm10076 C7 knockout keratinocytes Replicate2
Ribo-seq_counts.csv


title: hlsec, Tnfa, rep3

description: Library name: Tnfa_R3

value: Tnfa


title: PMCs, CONTROL, 0 hour, biol rep5

description:
Library name: Sample 2.2.1
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_0_5

value: connective tissue


title: RiboSeq_CtrlR2

description:
Library name: RiboSeq_CtrlR2
Ribosome Profiling from nontargeting control keratinocytes Replicate2
Ribo-seq_counts.csv


title: hlsec, iBET151, rep1

description: Library name: iBET151_R1


title: RNA-seq of single blastomere (8cell_E02_B4)

description: Library name: 8cell_E02_B4


title: hlsec, Tnfa, rep2

description: Library name: Tnfa_R2


title: RNA-seq of single blastomere (8cell_E02_B2)

description: Library name: 8cell_E02_B2


title: hlsec, Tnfa, rep1

description: Library name: Tnfa_R1


title: PMCs, C48/80, 6 hours, biol rep5

description:
Library name: Sample 2.2.3
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_C_6_5

value: primary peritoneal mast cells


title: RNA-seq of single blastomere (8cell_E02_B1)

description: Library name: 8cell_E02_B1


title: RiboSeq_KOR1

description:
Library name: RiboSeq_KOR1
Ribosome Profiling from Gm10076 C7 knockout keratinocytes Replicate1
Ribo-seq_counts.csv


title: PMCs, ADO, 2 hours, biol rep5

description:
Library name: Sample 2.2.2
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_2_5

value: WT


title: RNA-seq of single blastomere (8cell_E01_B8)

description: Library name: 8cell_E01_B8


title: RNA-seq of single blastomere (8cell_E01_B7)

description: Library name: 8cell_E01_B7


title: RiboSeq_CtrlR1

description:
Library name: RiboSeq_CtrlR1
Ribosome Profiling from nontargeting control keratinocytes Replicate1
Ribo-seq_counts.csv


title: PMCs, DNP, 2 hours, biol rep5

description:
Library name: Sample 2.2.4
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_D_2_5

value: DNP


title: hlsec, control, rep1

description: Library name: Vehicle_R1

value: DMSO


title: PMCs, DNP, 2 hours, biol rep4

description:
Library name: Sample 2.1.4
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_D_2_4


title: RNA8_KOR4

description:
Library name: RNA8_KOR4
RNA sequencing from Gm10076 C7 knockout keratinocytes Replicate4
RNA-seq_counts.csv

data_processing:
Bioinformatic analysis of single-cell RNA sequencing data: Raw sequencing data, consisting of BCL files, were demultiplexed using Illumina bcl2fastq v2.20 with default settings. One mismatch in the sample barcode sequences was allowed. The resulting fastq files were processed using the BD Rhapsody Pipeline (version 1.9.1) hosted at the Seven Brigdge’s cloud platform (https://www.sevenbridges.com/). A custom mouse reference genome, based on Gencode vM25 (GRCm38.p6), was generated and combined with the 720 sgRNAs sequences. Based on STAR aligner, a custom STAR reference genome was created and the Refined Putative Cell Calling option was disabled during the pipeline execution. For downstream analysis, unique molecular identifier (UMI) count data was corrected using BD Genomics Recursive Substitution Error Correction (RSEC). Processing PCR sgRNA dial-out data PCR dial-out data was processed using an in-house custom Python script as previously described (15). Three 9-nucleotide cell barcodes were extracted from the 97 possible barcodes from Read 1. One mismatch was allowed per barcode (Hamming distance of 1). The script also retrieved the 8-nucleotide UMI sequences from Read 1, as specified in the BD manual. For valid barcodes, sgRNAs were identified in Readd 2 by performing an exact match search for the 20-nucleotide sgRNA sequence. UMI counts per cell were deduplicated and sgRNA UMI counts per cell were computed. If multiple sgRNAs were detected for one cell, the cell was assigned to a specific sgRNA only if its UMI count exceeded the 99th percentile. Processing of single-cell RNA sequencing data The Scrublet Python package was used to remove doublets from each dataset separately. Genes expressed in a minimum of 5 cells were retained and an expected double rate of 20% (as estimated by BD Rhapsody) was applied. After filtering, seven P4 single-cell RNA-seq datasets and eight P60 datasets were merged using Seurat. Only cells containing detected sgRNAs and classified as singlets were selected for further analysis. Cells were filtered based on the following criteria: UMI counts > 500, UMI counts lower than the 0.99 quantile (to remove outliers) and mitochondrial gene content < 20%. Filtered datasets were then processed using Seurat’s standard pipeline. Gene expression was normalized using the NormalizeData function, which scales counts to 10,000 per cell followed by a natural log transformation (log1p). The FindVariableGenes function identified 2,000 highly variable genes, and data scaling was performed using ScaleData. Principal Component Analysis (PCA) was conducted with RunPCA and the ElbowPlot function was used to determine the top principal components that contributed to explaining the variance of the data for each dataset at P4 and P60. The FindNeighbors function was applied to identify nearest neighbors based in the selected principal components. Clustering was performed using FindClusters with Louvain modularity optimization. To visualize the data in two dimensions, UMAP (Uniform Manifold Approximation and Projection) was used. No batch effects were observed in the P4 and P60 datasets, so batch effect correction was not performed.
Assembly: Gencode Mouse vM25 (GRCm38.p6)
Supplementary files format and content: Comma-separated value files and matrix files with count numbers, *_counts.csv.gz files contain Cell Barcode, Gene Symbol and RSEC adjusted counts.


title: RNA-seq of single blastomere (8cell_E01_B6)

description: Library name: 8cell_E01_B6


title: RNA7_CtrlR4

description:
Library name: RNA7_CtrlR4
RNA sequencing from nontargeting control keratinocytes Replicate4
RNA-seq_counts.csv


title: PMCs, DNP, 6 hours, biol rep4

description:
Library name: Sample 2.1.5
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_D_6_4

value: 6 hours


title: hlsec, control, rep2

description: Library name: Vehicle_R2

value: Human liver sinusoidal endothelial cells


title: PMCs, ADO, 2 hours, biol rep4

description:
Library name: Sample 2.1.2
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_2_4


title: hlsec, shPML, Vehicle, Input

description: Library name: shPML_Tnfa_Input

data_processing:
ChIP-seq data was first quality assessed using the ChIPQC package (v1.21.0). The reads were then mapped to the human reference genome hg38 using bowtie2 (v2.3.1). Duplicated mapped reads were discarded using the MarkDuplicates function from Picard (v2.25.6). Narrow peaks were identified utilizing MACS2.
Assembly: hg38
Supplementary files format and content: BigWig files of genome-wide PML binding sites

extract_protocol:
Cells used for ChIP-seq were harvested and crosslinked with 1% formaldehyde in PBS, followed by quenching with 125 mM glycine on ice. The samples were washed with cold PBS and collected. The cells were then pelleted for 10 minutes at 4 °C, resuspended in lysis buffer I (50 mM HEPES-KOH, pH 7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100, protease inhibitors), rotated for 10 minutes at 4 °C, then resuspended in lysis buffer II (10 mM Tris-HCl, pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, protease inhibitors). The cells were rotated once more for 10 minutes at 4 °C, and resuspended in sonication buffer (20 mM Tris-HCl pH 8.0, 150 mM NaCl, 2 mM EDTA pH 8.0, 0.1% SDS, and 1% Triton X-100, protease inhibitors) for sonication. Sonicated lysates were centrifuged for 10 minutes at 4 °C. The sonicated chromatin was then incubated with magnetic beads bound with PML antibody overnight at 4 °C to enrich for DNA. The beads were then washed with wash buffer (50 mM HEPES-KOH pH 7.5, 500 mM LiCl, 1mM EDTA pH 8.0, 0.7% Na-Deoxycholate, 1% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 50 mM NaCl) in order. Chromatin elution was performed by incubation in elution buffer (50 mM Tris-HCl pH 8.0, 10 mM EDTA, 1% SDS) at 65°C for 30 minutes. Cross-links were reversed overnight at 65 °C.
To remove protein, proteinase K was added and incubated at 37 °C for 2 hours. DNA was then purified, amplified via PCR, and sequenced on the Illumina platform.

treatment_protocol: The lentivector expressing shPML or negative control shRNA (shCon) was transfected into HEK293T cells using Lipofectamine 2000 (Thermo Fisher). Supernatants containing lentivirus were collected, centrifugated, and filtered. hLSECs at 30% confluence were transduced with shPML or shCon lentivirus using the transfection reagent (Genechem) according to manufacture protocols. Then geneticin was treated for 24 hours to select positive cells. Human recombinant TNFα (50 ng/mL; PeproTech, 300-01A) was treated to hLSECs for 6 hours before cells were collected for further analysis.

value: shPML+Tnfa


title: PMCs, C48/80, 6 hours, biol rep4

description:
Library name: Sample 2.1.3
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_C_6_4


title: RNA-seq of single blastomere (8cell_E01_B5)

description: Library name: 8cell_E01_B5


title: hlsec, control, rep3

description: Library name: Vehicle_R3


title: hlsec, shPML, Vehicle, PML(ChIP), rep2

description: Library name: shPML_Tnfa_PML_R2

value: Santa Cruz Biotechology sc-71911


title: RNA-seq of single blastomere (8cell_E01_B4)

description: Library name: 8cell_E01_B4


title: RNA5_CtrlR3

description:
Library name: RNA5_CtrlR3
RNA sequencing from nontargeting control keratinocytes Replicate3
RNA-seq_counts.csv


title: RNA4_KOR2

description:
Library name: RNA4_KOR2
RNA sequencing  from Gm10076 C7 knockout keratinocytes Replicate2
RNA-seq_counts.csv


title: RNA-seq of single blastomere (8cell_E01_B2)

description: Library name: 8cell_E01_B2


title: ipsilateral dorsal striatum, M1144 & F1149, Late learning

city: Stanford

first: Yue

last: Sun

address: 290 Jane Stanford Way

institute: Stanford University

description: Library name: YS13-2

data_processing:
A custom reference was build using Cell Ranger pipeline (v3.1.0; https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger; RRID:SCR_017344). Briefly, a WPRE sequence (589 bp) located at 3’ end of Ai9 allele (Rosa-CAG-LSL-tdTomato-WPRE) was added at the end of mouse reference genome FASTA file and annotated in corresponding GTF file (Mus musculus, GRCm38/mm10, Ensemble database) . The genome and anonation were compiled to a reference using "cellranger mkref".
Base call files were demultiplexed into FASTAQ files (cellranger mkfastq) and aligned to the custom reference (cellranger count) following standard workflow of Cell Ranger.
Assembly: Custom mouse mm10 with reporter gene added (See data processing steps)
Supplementary files format and content: Matrix

source_name: dorsal striatum

extract_protocol:
Mice were anesthetized and perfused with cold, oxygenated ACSF. Coronal brain slices (300 µm) were prepared using a vibratome and incubated in ACSF at 3 °C for 30 min. Motor cortex (Fos-TRAP) and dorsal striatum (Arc-TRAP) were dissected, transferred to oxygenated HBSS/s solution, and pooled by hemisphere from two mice per condition. Tissue was digested with papain (20 U/ml, 37 °C, 1 h), triturated, and filtered through 40 µm strainers. Dead cells were removed using a Dead Cell Removal kit, and single cells were resuspended in cold PBS at 700–1200 cells/ml for downstream processing.
cDNA libraries were prepared using Chromium Next GEM Single Cell 3’ Library Construction Kit v2 or v3 (10x Genomics). Single-cell suspensions were loaded into the Chromium Chip for target recovery of 10,000 cells. cells were barcoded and lysed in droplets. Single-stranded cDNAs were then released from droplets, PCR amplified (11 cycles), fragmented, size-selected, ligated with adapters, and amplified by index PCR (11~12 cycles) to construct the final cDNA libraries.

value: dorsal striatum

tag: learning stage

value: Late


title: contralateral dorsal striatum, M1144 & F1149, Late learning

description: Library name: YS13-1


title: hlsec, shPML, Vehicle, PML(ChIP), rep1

description: Library name: shPML_Tnfa_PML_R1

value: Santa Cruz Biotechology sc-71910


title: hlsec, shCon, Tnfa, Input

description: Library name: shCon_Tnfa_Input

value: shCon+TNFa


title: hlsec, shCon, Tnfa, PML(ChIP), rep2

description: Library name: shCon_Tnfa_PML_R2


title: RNA6_KOR3

description:
Library name: RNA6_KOR3
RNA sequencing from Gm10076 C7 knockout keratinocytes Replicate3
RNA-seq_counts.csv


title: PMCs, CONTROL, 0 hour, biol rep4

description:
Library name: Sample 2.1.1
Column name in raw_counts_All_Samples.txt and normalized_counts_ComBatSeq_DESeq2_All_Samples.txt: WT_A_0_4

value: CONTROL

value: 0 hour


title: RNA1_CtrlR1

description:
Library name: RNA1_CtrlR1
RNA sequencing from nontargeting control keratinocytes Replicate1
RNA-seq_counts.csv

value: Keratinocytes


title: RNA-seq of single blastomere (8cell_E01_B3)

description: Library name: 8cell_E01_B3

value: blastomerer derived from IVF


title: RNA-seq of single blastomere (8cell_E01_B1)

description: Library name: 8cell_E01_B1


title: RNA-seq of single blastomere (4cell_E05_B4)

description: Library name: 4cell_E05_B4


title: hlsec, shCon, Tnfa, PML(ChIP), rep1

description: Library name: shCon_Tnfa_PML_R1


title: ipsilateral primary motor cortex, M1124 & F1128, Late learning

description: Library name: YS12-2

source_name: primary motor cortex

value: primary motor cortex


title: ipsilateral dorsal striatum, M1142 & F1148, Early learning

description: Library name: YS11-2

value: Early


title: hlsec, shCon, Vehicle, PML(ChIP), rep2

description: Library name: shCon_Vehicle_PML_R2

value: shCon+DMSO


title: contralateral primary motor cortex, M1124 & F1128, Late learning

description: Library name: YS12-1


title: RNA-seq of single blastomere (4cell_E05_B2)

description: Library name: 4cell_E05_B2


title: RNA-seq of single blastomere (4cell_E05_B3)

description: Library name: 4cell_E05_B3


title: hlsec, shCon, Vehicle, Input

description: Library name: shCon_Vehicle_Input


title: PLLA_B

city: Qujing

first: Dayuan

last: Tang

state: Yunnan

address: No. 1 , Qilin Road, Qujing, Yunnan

institute: Qujing Hospital Affiliated of Kunming Medical University

description: Single cell sequencing

data_processing:
After obtaining Sequenced Reads, bioinformatics was analyzed through the following procedure. The core of single-cell transcriptome analysis is the identification of subgroup and subgroup differential genes. 10X official analysis software Cell Ranger is used for cell typing and differential analysis, and these differential genes are subsequently enriched to identify the functional characteristics of subgroups.
Assembly: Through the enrichment analysis of differentially expressed genes in each cluster, we can find out which biological functions or pathways are significantly correlated with differentially expressed genes in each cluster. The software clusterProfiler was used to perform GO functional enrichment analysis and KEGG pathway enrichment analysis for the differentially expressed gene sets in each cluster
Supplementary files format and content: For the species included in the database, the interaction relationship of the target gene set (such as the list of differentially highly expressed genes in each cluster) is extracted directly from the database to build the network

source_name: BMSCs

extract_protocol:
The qualified cells were washed and re-suspended to prepare a suitable concentration of single-cell suspension
Illumina NovaSeq PE150

value: BMSCs

value: Primary cell

value: PLLA


title: RNA2_KOR1

description:
Library name: RNA2_KOR1
RNA sequencing from Gm10076 C7 knockout keratinocytes Replicate1
RNA-seq_counts.csv


title: RNA-seq of single blastomere (4cell_E05_B1)

description: Library name: 4cell_E05_B1


title: ipsilateral primary motor cortex, M1125 & F1130, Early learning

description: Library name: YS10-2


title: contralateral dorsal striatum, M1142 & F1148, Early learning

description: Library name: YS11-1


title: RNA-seq of single blastomere (4cell_E04_B4)

description: Library name: 4cell_E04_B4


title: RNA3_CtrlR2

description:
Library name: RNA3_CtrlR2
RNA sequencing from nontargeting control keratinocytes Replicate2
RNA-seq_counts.csv


title: hlsec, shCon, Vehicle, PML(ChIP), rep1

description: Library name: shCon_Vehicle_PML_R1

value: Human liver sinusoidal endothelial cells


title: RNA-seq of single blastomere (4cell_E04_B3)

description: Library name: 4cell_E04_B3


title: contralateral primary motor cortex, M1125 & F1130, Early learning

description: Library name: YS10-1


title: Model_C

description: Single cell sequencing

value: Model


title: hlsec, iBET151, BRD4(ChIP), rep2

description: Library name: iBET151_BRD4_R2

data_processing:
ChIP-seq data was first quality assessed using the ChIPQC package (v1.21.0). The reads were then mapped to the human reference genome hg38 using bowtie2 (v2.3.1). Duplicated mapped reads were discarded using the MarkDuplicates function from Picard (v2.25.6). Narrow peaks were identified utilizing MACS2.
Assembly: hg38
Supplementary files format and content: BigWig files of genome-wide BRD4 binding sites

extract_protocol:
Cells used for ChIP-seq were harvested and crosslinked with 1% formaldehyde in PBS, followed by quenching with 125 mM glycine on ice. The samples were washed with cold PBS and collected. The cells were then pelleted for 10 minutes at 4 °C, resuspended in lysis buffer I (50 mM HEPES-KOH, pH 7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100, protease inhibitors), rotated for 10 minutes at 4 °C, then resuspended in lysis buffer II (10 mM Tris-HCl, pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, protease inhibitors). The cells were rotated once more for 10 minutes at 4 °C, and resuspended in sonication buffer (20 mM Tris-HCl pH 8.0, 150 mM NaCl, 2 mM EDTA pH 8.0, 0.1% SDS, and 1% Triton X-100, protease inhibitors) for sonication. Sonicated lysates were centrifuged for 10 minutes at 4 °C. The sonicated chromatin was then incubated with magnetic beads bound with BRD4 antibody overnight at 4 °C to enrich for DNA. The beads were then washed with wash buffer (50 mM HEPES-KOH pH 7.5, 500 mM LiCl, 1mM EDTA pH 8.0, 0.7% Na-Deoxycholate, 1% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 50 mM NaCl) in order. Chromatin elution was performed by incubation in elution buffer (50 mM Tris-HCl pH 8.0, 10 mM EDTA, 1% SDS) at 65°C for 30 minutes. Cross-links were reversed overnight at 65 °C.
To remove protein, proteinase K was added and incubated at 37 °C for 2 hours. DNA was then purified, amplified via PCR, and sequenced on the Illumina platform.

treatment_protocol: Low-passage cells were starved with basal endothelial medium for 2 hours followed by treatment with BRD4 inhibitor (iBET151, 2.5 umol/L, Selleck, S2780) for 2 hours. Then, human recombinant TNFα (50 ng/mL; PeproTech, 300-01A) was treated to hLSECs for 6 hours before cells were collected for further analysis.

value: Bethyl Labs #A301-985A50


title: hlsec, iBET151, Input.

description: Library name: iBET151_Input


title: Model_B

description: Single cell sequencing


title: Model_A

description: Single cell sequencing


title: PLLA_A

description: Single cell sequencing


title: ipsilateral dorsal striatum, M697 & F698, Early learning

description: Library name: YS09-2

value: dorsal striatum


title: spleen CD8+ T cells, KLF2 control, D7, Rep 4

city: Minneapolis

first: Christine

middle: H

last: O'Connor

address: 117 Pleasant St. SE

department: Minnesota Supercomputer Institute

institute: University of Minnesota

description: Library name: D7_KLF2_Ctrl_4

data_processing:
Reads were trimmed using trimmomatic
Samples were aligned using HISAT2 v2.1.0
featureCounts v2.0.6 (Liao et al., 2014) was used to count mapped reads to genes
DE analysis was performed using EdgeR in R
Assembly: GRCm39.109
Supplementary files format and content: raw_counts.txt: raw gene counts for all samples
Supplementary files format and content: D25_Klf2_fullanno.txt: noramlized counts, DE results and gene annotation information for D25 KLF2 KO and matched controls
Supplementary files format and content: D25_Klf3_fullanno.txt: noramlized counts, DE results and gene annotation information for D25 KLF3 KO and matched controls
Supplementary files format and content: D7_Klf2_fullanno.txt: noramlized counts, DE results and gene annotation information for D7 KLF2 KO and matched controls

source_name: spleen

extract_protocol:
RNA was extracted using Qiagen Rneasy Plus Micro Kit. >500ng RNA was used to generate sequencing library.
RNA libraries for RNA-sequencing were prepared using TakaraBio PicoMammalian V2 kit following manufacturer's protocol. TakaraBio PicoMammalian V2 Library preparation

treatment_protocol: Congenically distinct WT or KLF2-GFP P14 CD8+ T cells were negatively enriched from spleen and activated with plate bound α-CD3 and α-CD28 with rhIL-2 for 48 hours, and then electroporated with sgRNAs/Cas9 complexes for target (Klf2 or Klf3) or control (Cd19 or Thy1) genes through a Lonza 4DNucleofector X Unit, using published conditions (Seki et al. 2018). Following culture in rhIL-2 for 24-48h, 5x10e4 CRISPR-modified cells were co-adoptively transferred into congenically distinct host mice, which received LCMV-Arm infection (2x10e5 PFU) within 2h of transfer. CRISPR modified donor P14 CD8+ T cells were FACS sorted from the spleen, 8- or 25-days post LCMV infection.

value: spleen


title: RNA-seq of single blastomere (4cell_E04_B2)

description: Library name: 4cell_E04_B2


title: RNA-seq of single blastomere (4cell_E04_B1)

description: Library name: 4cell_E04_B1


title: PLLA_C

description: Single cell sequencing


title: hlsec, iBET151, BRD4(ChIP), rep1

description: Library name: iBET151_BRD4_R1


title: contralateral dorsal striatum, M697 & F698, Early learning

description: Library name: YS09-1


title: RNA-seq of single blastomere (4cell_E03_B3)

description: Library name: 4cell_E03_B3


title: RNA-seq of single blastomere (4cell_E03_B4)

description: Library name: 4cell_E03_B4


title: left dorsal striatum, M457 & F461, Ctrl

description: Library name: YS08-2

value: Ctrl


title: hlsec, Tnfa, BRD4(ChIP), rep2

description: Library name: Tnfa_BRD4_R2

value: Tnfa


title: hlsec, Tnfa, Input.

description: Library name: Tnfa_Input


title: hlsec, Tnfa, BRD4(ChIP), rep1

description: Library name: Tnfa_BRD4_R1


title: spleen CD8+ T cells, KLF2 control, D7, Rep 3

description: Library name: D7_KLF2_Ctrl_3


title: spleen CD8+ T cells, KLF2 control, D7, Rep 2

description: Library name: D7_KLF2_Ctrl_2


title: spleen CD8+ T cells, KLF2 control, D7, Rep 1

description: Library name: D7_KLF2_Ctrl_1


title: right primary motor cortex, M4252 & F4266, Ctrl

description: Library name: YS07-1


title: contralateral primary motor cortex, M4255 & F4267, Early learning

description: Library name: YS06-2


title: RNA-seq of single blastomere (4cell_E03_B2)

description: Library name: 4cell_E03_B2


title: hlsec, Vehicle, BRD4(ChIP), rep1

description: Library name: Vehicle_BRD4_R1

value: DMSO


title: hlsec, Vehicle, Input.

description: Library name: Vehicle_Input


title: right dorsal striatum, M457 & F461, Ctrl

description: Library name: YS08-1


title: left primary motor cortex, M4252 & F4266, Ctrl

description: Library name: YS07-2


title: spleen CD8+ T cells, KLF2 KO, D7, Rep 3

description: Library name: D7_KLF2_KO_3


title: hlsec, Vehicle, BRD4(ChIP), rep2

description: Library name: Vehicle_BRD4_R2


title: hlsec, iBET151, Input

description: Library name: iBET151_Input

data_processing:
ChIP-seq data was first quality assessed using the ChIPQC package (v1.21.0). The reads were then mapped to the human reference genome hg38 using bowtie2 (v2.3.1). Duplicated mapped reads were discarded using the MarkDuplicates function from Picard (v2.25.6). Narrow peaks were identified utilizing MACS2.
Assembly: hg38
Supplementary files format and content: BigWig files of genome-wide H3K27ac modification

extract_protocol:
Cells used for ChIP-seq were harvested and crosslinked with 1% formaldehyde in PBS, followed by quenching with 125 mM glycine on ice. The samples were washed with cold PBS and collected. The cells were then pelleted for 10 minutes at 4 °C, resuspended in lysis buffer I (50 mM HEPES-KOH, pH 7.5, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100, protease inhibitors), rotated for 10 minutes at 4 °C, then resuspended in lysis buffer II (10 mM Tris-HCl, pH 8.0, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, protease inhibitors). The cells were rotated once more for 10 minutes at 4 °C, and resuspended in sonication buffer (20 mM Tris-HCl pH 8.0, 150 mM NaCl, 2 mM EDTA pH 8.0, 0.1% SDS, and 1% Triton X-100, protease inhibitors) for sonication. Sonicated lysates were centrifuged for 10 minutes at 4 °C. The sonicated chromatin was then incubated with magnetic beads bound with H3K27ac antibody overnight at 4 °C to enrich for DNA. The beads were then washed with wash buffer (50 mM HEPES-KOH pH 7.5, 500 mM LiCl, 1mM EDTA pH 8.0, 0.7% Na-Deoxycholate, 1% NP-40) and TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA, 50 mM NaCl) in order. Chromatin elution was performed by incubation in elution buffer (50 mM Tris-HCl pH 8.0, 10 mM EDTA, 1% SDS) at 65°C for 30 minutes. Cross-links were reversed overnight at 65 °C.
To remove protein, proteinase K was added and incubated at 37 °C for 2 hours. DNA was then purified, amplified via PCR, and sequenced on the Illumina platform.


title: RNA-seq of single blastomere (4cell_E03_B1)

description: Library name: 4cell_E03_B1


title: ipsilateral primary motor cortex, M4255 & F4267, Early learning

description: Library name: YS06-1


title: RNA-seq of single blastomere (4cell_E02_B4)

description: Library name: 4cell_E02_B4


title: spleen CD8+ T cells, KLF2 KO, D7, Rep 2

description: Library name: D7_KLF2_KO_2


title: hlsec, iBET151, H3K27ac(ChIP), rep2

description: Library name: iBET151_H3K27ac_R2

value: Abcam #4729


title: right dorsal striatum, M3320 & F3374, Ctrl

description: Library name: YS05-2


title: RNA-seq of single blastomere (4cell_E02_B1)

description: Library name: 4cell_E02_B1


title: ipsilateral dorsal striatum, M3367 & F3325, Late learning

description: Library name: YS04-2

value: Late


title: left dorsal striatum, M3320 & F3374, Ctrl

description: Library name: YS05-1


title: spleen CD8+ T cells, KLF2 KO, D7, Rep 1

description: Library name: D7_KLF2_KO_1


title: spleen CD8+ T cells, KLF2 KO, D7, Rep 4

description: Library name: D7_KLF2_KO_4


title: spleen CD8+ T cells, KLF3 control, D25, Rep 3

description: Library name: D25_KLF3_Ctrl_6


title: hlsec, iBET151, H3K27ac(ChIP), rep1

description: Library name: iBET151_H3K27ac_R1


title: RNA-seq of single blastomere (4cell_E01_B4)

description: Library name: 4cell_E01_B4

value: blastomerer derived from IVF


title: RNA-seq of single blastomere (4cell_E02_B3)

description: Library name: 4cell_E02_B3


title: RNA-seq of single blastomere (4cell_E01_B3)

description: Library name: 4cell_E01_B3


title: spleen CD8+ T cells, KLF3 control, D25, Rep 2

description: Library name: D25_KLF3_Ctrl_5


title: contralateral dorsal striatum, M3367 & F3325, Late learning

description: Library name: YS04-1


title: hlsec, Tnfa, Input

description: Library name: Tnfa_Input


title: RNA-seq of single blastomere (4cell_E01_B2)

description: Library name: 4cell_E01_B2


title: left primary motor cortex, M3259 & F3252, Ctrl

description: Library name: YS03-2

value: primary motor cortex


title: hlsec, Tnfa, H3K27ac(ChIP), rep1

description: Library name: Tnfa_H3K27ac_R1


title: RNA-seq of single blastomere (4cell_E02_B2)

description: Library name: 4cell_E02_B2


title: spleen CD8+ T cells, KLF2 control, D25, Rep 2

description: Library name: D25_KLF2_Ctrl_2


title: RNA-seq of single blastomere (4cell_E01_B1)

description: Library name: 4cell_E01_B1


title: spleen CD8+ T cells, KLF3 control, D25, Rep 1

description: Library name: D25_KLF3_Ctrl_4


title: spleen CD8+ T cells, KLF2 control, D25, Rep 3

description: Library name: D25_KLF2_Ctrl_3


title: hlsec, Tnfa, H3K27ac(ChIP), rep2

description: Library name: Tnfa_H3K27ac_R2


title: ipsilateral primary motor cortex, M3263 & F3269, Late learning

description: Library name: YS02-2


title: right primary motor cortex, M3259 & F3252, Ctrl

description: Library name: YS03-1


title: spleen CD8+ T cells, KLF2 control, D25, Rep 1

description: Library name: D25_KLF2_Ctrl_1


title: hlsec, Vehicle, Input

description: Library name: Vehicle_Input


title: RNA-seq of single blastomere (2cell_E10_B1)

description: Library name: 2cell_E10_B1


title: hlsec, Vehicle, H3K27ac(ChIP), rep1

description: Library name: Vehicle_H3K27ac_R1


title: RNA-seq of single blastomere (2cell_E10_B2)

description: Library name: 2cell_E10_B2


title: RNA-seq of single blastomere (2cell_E09_B2)

description: Library name: 2cell_E09_B2


title: contralateral primary motor cortex, M3263 & F3269, Late learning

description: Library name: YS02-1


title: hlsec, Vehicle, H3K27ac(ChIP), rep2

description: Library name: Vehicle_H3K27ac_R2


title: TBM, blood,  NSH012, D14

city: London

first: James

middle: Royce

last: Barnacle

state: London

address: Francis Crick Institute, 1 Midland Road

department: Infectious Disease

country: United Kingdom

institute: Imperial College London

description: Library name: DAV1202A134

data_processing:
Raw data were processed using nf-core/rnaseq (v3.10.1) of the nf-core collection of workflows. This workflow used FASTQ files as input, performed quality control (QC), trimming and (pseudo-)alignment to the GRCh38 release 95 human reference genome, and produced a gene expression matrix and QC report. The pipeline was executed with Nextflow (v22.10.3). The output matrix was read into R (v4.2.0) in which subsequent analysis was done.
Assembly: GRCh38, Homo sapiens
Supplementary files format and content: .csv file with samples as colnames and genes as rownames

source_name: Blood

extract_protocol:
Whole blood was collected directly into PAXgene RNA tubes (QIAGEN/BD Biosciences) from 40 patients enrolled into the LASER-TBM trial. Total RNA was extracted using the PAXgene Blood RNA Kit (QIAGEN, Venlo, Netherlands). Samples were stored at -80˚C before and after RNA extraction. Quantity and quality of RNA was assessed using a Qubit fluorimeter (ThermoFisher) and Agilent 2100 Bioanalyzer. Samples with RNA integrity number (RIN) <7 were excluded.
RNA libraries were constructed using Tecan’s Universal Human Blood RNA-Seq kit. Ribosomal and globin RNA were removed using Tecan’s AnyDeplete system. DNA library size distribution was assessed using the D1000 TapeStation kit processed on the 4200 TapeStation (Agilent Technologies). The data were analysed by Agilent’s 4200 TapeStation Analysis Software. Concentration was measured using the QuantiFluor dsDNA System (Promega Corporation). Libraries were sequenced on an Illumina HiSeq4000 with a single-end 100 cycle read configuration.

value: Blood

tag: patient id

tag: study

value: laser

tag: timepoint

tag: age

tag: Sex

value: male

tag: antibiotics

value: soc

tag: aspirin

value: no

tag: diagnosis

value: possible

tag: gxpu

value: negative

tag: mtb culture

tag: hiv vl

tag: blood wcc

tag: blood neutrophils


title: spleen CD8+ T cells, KLF3 KO, D25, Rep 3

description: Library name: D25_KLF3_KO_6

tag: tissue


title: hlsec, iBET151, Hi-C, rep2

description: Library name: iBET151_R2

data_processing:
Hi-C reads were processed using the Hi-C Pro pipeline (v3.1.0). Reads were first aligned to the hg38 reference genome using bowtie2, the aligned reads were assigned to MboI restriction fragments, and the interaction pairs were reconstructed. Then quality assessment was performed, singletons and multi-hit pairs and failed ligation products were filtered. The remaining valid pairs were then used to build contact matrices, and then normalized by iterative correction and eigenvector decomposition.
Assembly: hg38
Supplementary files format and content: hic files of Hi-C juicebox contact map

extract_protocol:
Hi-C library preparation was carried out following existing protocols. Briefly, approximately 100,000 to 1,000,000 cell pellets were used as input per sample. Nuclei were digested overnight with MboI at 37 °C, followed by end-labeling of the DNA with biotin using 25 μl of fill-in master mix at 37 °C for 2 hours. The DNA was then ligated using 450 μl of ligation master mix at room temperature for 4 hours. DNA was purified using phenol-chloroform extraction. To remove linear DNA, the sample was treated with Lambda exonuclease (NEB) and of Exonuclease I (NEB) at 37 °C for 30 minutes. Fragmentation of 100–500 ng of DNA was performed using the TruePrep DNA Library Prep Kit V2 (Vazyme).
Biotin-labeled DNA was enriched by pull-down with Dynabeads MyOne Streptavidin T1 beads (Life Technologies). Library amplification and size selection were conducted using TruePrep Amplify Enzyme (Vazyme) and SPRIselect Beads (Beckman Coulter). Libraries were sequenced on Illumina platform.

tag: cell type

value: Human liver sinusoidal endothelial cells

tag: treatment


title: spleen CD8+ T cells, KLF3 KO, D25, Rep 2

description: Library name: D25_KLF3_KO_5


title: RNA-seq of single blastomere (2cell_E09_B1)

description: Library name: 2cell_E09_B1

data_processing:
Adapter trimming was performed using Trim Galore.
Sequence quality was assessed with FastQC.
Reads were aligned to the bovine reference genome (ARS-UCD1.2/bosTau9) using STAR.
Transcript quantification was performed using RSEM.
Assembly: ARS-UCD1.2/bosTau9
Supplementary files format and content: Gene-level count matrix generated by RSEM, provided as tab-delimited text files. Each file contains Ensembl gene IDs and corresponding TPM and expected count values for each sample.

tag: Stage

tag: embryo id

tag: blastomere id


title: TBM, blood,  MPH011, D0

description: Library name: DAV1202A132

tag: patient id

tag: study

tag: timepoint

value: female

value: intensified

value: yes

value: probable


title: hlsec, iBET151, Hi-C, rep1

description: Library name: iBET151_R1


title: spleen CD8+ T cells, KLF3 KO, D25, Rep 1

description: Library name: D25_KLF3_KO_4

value: spleen


title: RNA-seq of single blastomere (2cell_E08_B2)

description: Library name: 2cell_E08_B2


title: RNA-seq of single blastomere (2cell_E08_B1)

description: Library name: 2cell_E08_B1


title: TBM, blood,  MPH041, D7

description: Library name: DAV1202A131

tag: age

tag: Sex

tag: antibiotics


title: hlsec, Tnfa, Hi-C, rep1

description: Library name: Tnfa_R1

value: Tnfa


title: TBM, blood,  LH007, D7

description: Library name: DAV1202A130

tag: aspirin

tag: diagnosis

value: definite

tag: gxpu

value: positive


title: spleen CD8+ T cells, KLF2 KO, D25, Rep 3

description: Library name: D25_KLF2_KO_3


title: TBM, blood,  NSH016, D14

description: Library name: DAV1202A135

tag: mtb culture

tag: hiv vl

tag: blood wcc


title: hlsec, Tnfa, Hi-C, rep2

description: Library name: Tnfa_R2

tag: cell type


title: RNA-seq of single blastomere (2cell_E07_B2)

description: Library name: 2cell_E07_B2

value: blastomerer derived from IVF


title: spleen CD8+ T cells, KLF2 KO, D25, Rep 2

description: Library name: D25_KLF2_KO_2

tag: tissue


title: TBM, blood,  NSH011, D14

description: Library name: DAV1202A129

value: Blood

tag: blood neutrophils


title: RNA-seq of single blastomere (2cell_E07_B1)

description: Library name: 2cell_E07_B1

tag: Stage

tag: embryo id


title: TBM, blood,  GSH004, D56

description: Library name: DAV1202A127

tag: patient id

value: laser


title: RNA-seq of single blastomere (2cell_E06_B2)

description: Library name: 2cell_E06_B2

tag: blastomere id


title: RNA-seq of single blastomere (2cell_E06_B1)

description: Library name: 2cell_E06_B1