title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL03

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL03_DpGC51_60_S12


title: Day 10 DIO [X62]

status: Public on Sep 01 2023

type: SRA

city: Hershey

first: Yuka

last: Imamura Kawasawa

state: PA

address: 500 University Dr.

department: College of Medicine, Pharmacology

country: USA

institute: Penn State University

library_source: transcriptomic

data_processing:
De-multiplexed and adapter-trimmed sequencing reads were generated using Illumina bcl2fastq (released version 2.20.0.422) allowing no mismatches in the index read. BBDuk was used to trim/filter low quality sequences using “qtrim=lr trimq=10 maq=10” option. Next, alignment of the filtered reads to the mouse reference genome (GRCm38) was done using HISAT2 (version 2.1.0) applying --no-mixed and --no-discordant options. Read counts were calculated using HTSeq by supplementing Ensembl gene annotation (GRCm38.78).
Assembly: GRCm38
Supplementary files format and content: raw read counts are provided in a csv file

molecule: total RNA

organism: Mus musculus

source_name: Bone fracture healing callus (Day 10 post-fracture)

extract_protocol:
Total RNA was isolated from the fracture callus using TRIzol. Optical density values of extracted RNA were measured using NanoDrop (Thermo Scientific) to confirm an A260:A280 ratio above 1.9. RNA integration number (RIN) was measured using BioAnalyzer (Agilent Technologies) RNA 6000 Nano Kit.
The cDNA libraries were prepared using the Illumina® Stranded mRNA Prep, Ligation kit (Illumina) as per the manufacturer’s instructions. Briefly, polyA RNA was purified from 200 ng of total RNA using oligo (dT) beads. The extracted mRNA fraction was subjected to fragmentation, reverse transcription, end repair, 3’– end adenylation, and adaptor ligation, followed by PCR amplification and SPRI bead purification (Beckman Coulter). The unique dual index sequences (IDT® for Illumina® RNA UD Indexes Set A, Ligation, Illumina) were incorporated in the adaptors for multiplexed high-throughput sequencing. The final product was assessed for its size distribution and concentration using BioAnalyzer High Sensitivity DNA Kit (Agilent Technologies). The libraries were pooled and diluted to 3 nM using 10 mM Tris-HCl, pH 8.5 and then denatured using the Illumina protocol. The denatured libraries were loaded onto an S1 flow cell on an Illumina NovaSeq 6000 (Illumina) and run for 2X53 cycles according to the manufacturer's instructions.

tag: tissue

tag: biological repeats

value: Biol rep 3

tag: disease state

value: diet-induced obese (DIO)

tag: condition

value: Day 10 DIO


title: Day 14 DIO [X49]

source_name: Bone fracture healing callus (Day 14 post-fracture)

value: Biol rep 1

value: Day 14 DIO


title: MCF7_NAIL_ΔNFκB_TNFa_90min_H3K4Me3_chip

city: Singapore

first: Vinay

last: Tergaonkar

address: 61 Biopolis Drive,  Proteos

country: Singapore

institute: Institute of Molecular and Cell Biology

library_source: genomic

data_processing:
Raw fastq files were aligned to hg19 human reference genome assembly using Bowtie2 read-aligner to obtain *.bam alignment files.
Peaks were called by Homer (v4.10) using default parameters with narrow (p65, p300) or broad peak (H3K4Me3 and H3K27Ac) options.
Differential peaks of respective time points were identified using Deseq2 in Homer suite.
Bedgraph files were generated using Homer and peaks were visualized by Integrative Genomics Viewer.
Genome_build: hg19
Supplementary_files_format_and_content: BigWig

molecule: genomic DNA

organism: Homo sapiens

growth_protocol: Cells were cultured in DMEM media supplemented with 10% FBS and antibiotics.

extract_protocol:
Cells were crosslinked on the plate with disuccinimidyl glutarate (DSG) (1.5mM) (Thermo Fischer Scientific) for 30 mins at room temperature with gentle shaking. After removing DSG, cells were fixed with 1% formaldehyde for 10 mins and the reaction was quenched with a final concentration of 0.125M glycine. Lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with antibodies.
Chip-seq library preparation was performed by NEB Next Ultra II DNA library preparation kit (New England Biolabs) according to the manufacturer’s instructions.
Sequencing was performed on HiSeq2500 system (Illumina) with high output run mode and paired-end sequencing (2x101bp).

treatment_protocol: MCF7-NAIL-WT and MCF7-NAIL-ΔNFκB cells stimulated with TNF alpha for 0min, 45min and 90.

tag: cell line

tag: genotype

tag: treatment

value: TNFa

tag: time

tag: chip antibody

value: H3K4Me3 (Millipore, 04-745)


title: Day 14 DIO [X50]

value: Bone fracture healing callus (Day 14 post-fracture)

value: Biol rep 2


title: Day 10 DIO [X61]

value: Bone fracture healing callus (Day 10 post-fracture)


title: MCF7_NAIL_WT_TNFa_90min_H3K27Ac_chip

value: H3K27Ac (Millipore, 07-360)


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL03

city: Kashihara

first: Hiroki

last: Ikeda

address: 840 Shijo-Cho, Kashihara, Nara

country: Japan

institute: Nara Medical University

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL03_DpGC31_40_S10

data_processing:
After trimming polyA/T sequences, adapter sequences, and poor quality reads, using fastp (v. 0.20.0) and cutadapt (ver. 1.16), the sequenced reads were aligned to the GRCm38 mouse genome assemblywith ERCC spike-in RNA sequences using the HiSAT2 (v. 2.1.0).
Best-matched reads on annotated genes were counted using HTSeq 0.10.0 with a modified GRCm38 gtf file which is extended up to 10 Kb downstream from transcription termination sites of reference genes.
Mapped reads on exons and exon-exon junctions were counted by featureCounts (v2.0.1) with the GRCm38 gtf file and the option "-J" "-p" "-M" "-O" "-t　exon" "-g　transcript_id".
Assembly: GRCm38
Supplementary files format and content: Matrix table with raw gene, exon and exon-exon junction counts for every sample.

molecule: polyA RNA

source_name: Granulosa cell

growth_protocol: Mouse embryonic stem cells (mESCs) were cultured in a medium containing 2iLIF [GSK inhibitor (CHIR99021; 3 µM), MEK inhibitor (PD0325901; 0.4 µM), and LIF (1 U/µL) (2iLIF)]

extract_protocol:
A Smart-seq v4 3’ DE Kit (Takara Bio) was adapted to DRaqL as follows.  Cells were isolated with LCM in 6.4 μL of cell lysis buffer (0.25 μL of 40 U/μL Takara RNase Inhibitor, 0.4 μL of 100 mM DTT, 0.16 μL of 1:500,000 diluted ERCC RNA Spike-In Mix 1, 0.4 μL of 5% SDc, and 5.19 μL of nuclease-free water) in the caps of PCR tubes, and spun down into the tubes by brief centrifugation.  Cells were then lysed at 72°C for 3 min, followed by the addition of 2.8 μL quenching buffer (0.7 μL of Triton X-100, 0.7 μL of 2% BSA, and 1.4 μL of 5× SuperScript II buffer [Thermo Fisher]) to reduce the denaturing effect of SDc.  Then, 2.8 μL of RT oligo (1 μL of 12 μM Oligo dT In-line Primer and 1.8 uL of nuclease-free water: kit components) was added, followed by incubation at 72°C for 90 sec.  Reverse transcription was performed by adding 7.5 μL of Master Mix (4 ?L of 5× Ultra Low First-Strand Buffer, 1 μL of 48 μM SMART-Seq V4 oligonucleotide, 0.5 μL of 40 U/μL RNase Inhibitor, and 2 μL of SMARTScribe Reverse Transcriptase: kit component) supplemented with 0.25 μL of SuperScript II and 0.25 μL of SuperScript III, and by incubating at 42°C for 90 min and 70°C for 10 min.  Then, cDNA amplification was performed by adding 30 μL of kit component PCR Mix (25 μL of 2× SeqAmp PCR Buffer, 1 μL of 12 μM Blocked PCR Primer II A, 1 μL of SeqAmp DNA polymerase, and 3μL of nuclease-free water) and applying the thermal cycling program for SMART-seq v4.  The amplified cDNA (50 μL) was purified with a 0.8× volume of AxyPrep MAG PCR clean-up reagent according to the manufacturer’s instructions.
The cDNA synthesis and amplification onward were performed as described previously with minor modifications [Kurimoto, K.et.al., Nucleic Acids Research, 34(5), e42.]. In brief, the first-strand cDNA was synthesized by SuperScript III [Thermo Fisher] and incubating at 50℃  for 5 min followed by heat inactivation at 70℃  for 10 min.  The excess RT primers were degraded by exonuclease I and incubating at 37℃ for 30 min followed by heat inactivation at 80℃ for 25 min. The first-strand cDNA product was poly dA-tailed by TdT and incubating at 37℃  for 1 min followed by heat inactivation at 70℃ for 10 min. Second-strand cDNA synthesis were performed by adding 9.5 μL reaction mixture of ExTaq, Hot Start version [Takara Bio] and V3[dT]24 primer in the ExTaq buffer with the following thermal cycling program (95℃  for 3 min, 50℃  for 2 min, and 72℃  for 3 min). cDNA amplification was performed by addingthe mixture of V1[dT]24 and ExTaq, Hot Start version [Takara Bio] in the ExTaq buffer with the following thermal cycling program (95℃  for 1 min; 20 cycles of 95℃  for 30 sec, 67℃  for 1 min and 72℃  for 3 min with a 6 sec elongation per cycle; and 72℃  for 10 min).  The amplified cDNA was purified with a 0.6× volume of AxyPrep MAG PCR clean-up reagent (Axygen) according to the manufacturer's instructions. Preparation of the libraries for SC3-seq was performed essentially as previously described (Nakamura et al., 2015)(Nakamura et al., 2017).  For Y-adapter sequencing, single-cell cDNAs were pre-amplified using the primers (V3[dT]24, V1[dT]24) and ExTaq HS with the thermal cycling program (95℃  for 3 min; 4 cycles of 95°C for 30 sec, 67℃  for 1 min and 72℃  for 5 min; 72℃  for 10 min). Pre-amplified cDNAs were then subjected to the primer-dimer removal, sonication, end polishing, and the size selection as in SC3-seq. Then, purified cDNA was dA-tailed in a 10 μL dA tailing mixture with incubation at 72℃  for 10 min. The dA-tailed product was subjected to the Y-shaped adaptor ligation by adding a mixture of Y-adaptor and NEB Next Quick Ligation Reaction Buffer and incubating at 20℃  for 15 min and at 72℃  for 20 min. The ligation product was purified with a 0.9×volume of AxyPrep reagent, and subjected to additional PCR amplification in 20 μL with the thermal cycling program (95℃  for 3 min, 55℃  for 1 min, and 72℃  for 1 min; 11 cycles of 95℃  for 30 sec, 60℃  for 1 min, and 72℃  for 1 min; and 72℃  for 3 min), followed by DNA purification with a 0.9×volume of AxyPrep reagent.

treatment_protocol: Mouse ovaries were excised from 8-week-old female mice and snap-frozen in liquid N2. Then, the frozen ovaries were sunk in 10% PVA in cryomolds, and again frozen in liquid N2. The embedded ovaries were sectioned using a CM1860UV cryostat at a thickness of 15 μm, dried at room temperature for more than 1 h, fixed and stained with 1% cresyl violet acetate dissolved in 50% isopropanol, washed with 100% isopropanol, and dried for more than 1 h. Cells in the frozen sections were isolated using a laser microdissection system PALM MB4 (Zeiss).

tag: collecting conditions

value: LCM

tag: class

value: Neighboring

tag: analysis target

value: gene expression


title: Day 7 DIO [X28]

source_name: Bone fracture healing callus (Day 7 post-fracture)

value: Day 7 DIO


title: MCF7_NAIL_ΔNFκB_TNFa_0min_H3K4Me3_chip

value: control


title: MCF7_NAIL_WT_TNFa_45min_H3K4Me3_chip


title: Day 10 DIO [X59]


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL03

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL03_DpGC21_30_S9


title: Day 7 DIO [X27]

value: Bone fracture healing callus (Day 7 post-fracture)


title: MCF7_NAIL_WT_TNFa_90min_H3K4Me3_chip


title: Day 7 DIO [X24]


title: Day 5 DIO [X18]

source_name: Bone fracture healing callus (Day 5 post-fracture)

value: Biol rep 3

value: Day 5 DIO


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL03

description:
SMART-Seq v4 3' DE Kit
IL03_DmGC41_50_S5

extract_protocol:
The cDNA amplification using SMART-Seq v4 3’DE Kit was performed according manufacturer’s instruction (Takara Bio), except for the step of cell capture.  20 μL of 10×Reaction buffer was prepared (19 μL of 10×Lysis Buffer, 1 μL of RNase Inhibitor), of which 1 μL was mixed with 10.5 μL DDW, resulting in 11.5 μL of cell lysis buffer.  Then, 6.4 μL of cell lysis buffer was added the flat caps of PCR tube, and cells were isolated with LCM into the reaction buffer, followed by the brief centrifugation.  Then, another 5.1 μL of cell lysis buffer was added to the tubes (11.5 μL in total).  The following steps were performed exactly according to the manufacturer’s instruction.  Briefly, 1 μL of 12 μM Oligo dT In-line Primer was added to the mixture, and incubated at 72°C for 3 min.  Then, reverse transcription was performed by adding 7.5 μL of Master Mix (4 μL of 5x Ultra Low First-Strand Buffer, 1 μL of 48 μM SMART-Seq V4 Oligonucleotide, 0.5 μL of 40 U/μL RNase Inhibitor, 2 μL of SMARTScribe Reverse Transcriptase) and by incubating at 42°C for 90 min and 70 °C for 10 min.  Then, cDNA amplification was performed by adding 30 μL of kit component PCR Mix (25 μL of 2× SeqAmp PCR Buffer, 1μL of 12 μM Blocked PCR Primer II A, 1μL of SeqAmp DNA Polymerase, and 3μL of Nuclease free water) and applying the thermal cycling program for SMART-seq v4.
The cDNA synthesis and amplification onward were performed as described previously with minor modifications [Kurimoto, K.et.al., Nucleic Acids Research, 34(5), e42.]. In brief, the first-strand cDNA was synthesized by SuperScript III [Thermo Fisher] and incubating at 50℃  for 5 min followed by heat inactivation at 70℃  for 10 min.  The excess RT primers were degraded by exonuclease I and incubating at 37℃ for 30 min followed by heat inactivation at 80℃ for 25 min. The first-strand cDNA product was poly dA-tailed by TdT and incubating at 37℃  for 1 min followed by heat inactivation at 70℃ for 10 min. Second-strand cDNA synthesis were performed by adding 9.5 μL reaction mixture of ExTaq, Hot Start version [Takara Bio] and V3[dT]24 primer in the ExTaq buffer with the following thermal cycling program (95℃  for 3 min, 50℃  for 2 min, and 72℃  for 3 min). cDNA amplification was performed by addingthe mixture of V1[dT]24 and ExTaq, Hot Start version [Takara Bio] in the ExTaq buffer with the following thermal cycling program (95℃  for 1 min; 20 cycles of 95℃  for 30 sec, 67℃  for 1 min and 72℃  for 3 min with a 6 sec elongation per cycle; and 72℃  for 10 min).  The amplified cDNA was purified with a 0.6× volume of AxyPrep MAG PCR clean-up reagent (Axygen) according to the manufacturer's instructions. Preparation of the libraries for SC3-seq was performed essentially as previously described (Nakamura et al., 2015)(Nakamura et al., 2017).  For Y-adapter sequencing, single-cell cDNAs were pre-amplified using the primers (V3[dT]24, V1[dT]24) and ExTaq HS with the thermal cycling program (95℃  for 3 min; 4 cycles of 95°C for 30 sec, 67℃  for 1 min and 72℃  for 5 min; 72℃  for 10 min). Pre-amplified cDNAs were then subjected to the primer-dimer removal, sonication, end polishing, and the size selection as in SC3-seq. Then, purified cDNA was dA-tailed in a 10 μL dA tailing mixture with incubation at 72℃  for 10 min. The dA-tailed product was subjected to the Y-shaped adaptor ligation by adding a mixture of Y-adaptor and NEB Next Quick Ligation Reaction Buffer and incubating at 20℃  for 15 min and at 72℃  for 20 min. The ligation product was purified with a 0.9×volume of AxyPrep reagent, and subjected to additional PCR amplification in 20 μL with the thermal cycling program (95℃  for 3 min, 55℃  for 1 min, and 72℃  for 1 min; 11 cycles of 95℃  for 30 sec, 60℃  for 1 min, and 72℃  for 1 min; and 72℃  for 3 min), followed by DNA purification with a 0.9×volume of AxyPrep reagent.


title: Day 5 DIO [X1]

value: Bone fracture healing callus (Day 5 post-fracture)

value: diet-induced obese (DIO)


title: Day 5 DIO [X16]


title: MCF7_NAIL_WT_TNFa_0min_H3K4Me3_chip


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL03

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL03_DpGC41_50_S11


title: Day 3 DIO [X6]

source_name: Bone fracture healing callus (Day 3 post-fracture)

value: Day 3 DIO


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC31_40_S4_IL03

description:
SMART-Seq v4 3' DE Kit
IL03_DmGC31_40_S4


title: MCF7_NAIL_ΔNFκB_TNFa_90min_p65_chip_rep2

value: TNFa

value: p65 (Santa-Cruz, sc-372)


title: MCF7_NAIL_ΔNFκB_TNFa_90min_p65_chip_rep1


title: Day 3 DIO [X5]

value: Bone fracture healing callus (Day 3 post-fracture)


title: Day 3 DIO [X4]

value: Biol rep 1


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL03

description:
SMART-Seq v4 3' DE Kit
IL03_DmGC21_30_S3


title: Day 10 lean [X57]

value: Bone fracture healing callus (Day 10 post-fracture)

value: Lean

value: Day 10 lean


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL02

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL02_DpGC51_60_S12

value: LCM

value: Neighboring


title: MCF7_NAIL_ΔNFκB_TNFa_45min_p65_chip_rep1


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL02

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL02_DpGC41_50_S11

value: gene expression


title: Butyrate 3

city: Los Angeles

first: Andrew

middle: Scott

last: Goldstein

state: CA

address: 610 Charles E Young Dr East

department: Urology

institute: University of California, Los Angeles

data_processing:
The RNAseq reads were mapped by STAR 2.7.9a and read counts per gene were quantified using the mouse genome GRCm39.105. In Partek Flow, read counts were normalized by CPM +1.0E-4.
Assembly: GRCm39.105
Supplementary files format and content: tab-delimited text files include TPM values for each Sample

source_name: prostate

growth_protocol: Organoids were cultured following Drost et al Nature Protocols 2016

extract_protocol: Libraries for RNA-Seq were prepared with KAPA RNA Hyper Kit. The workflow consists of depletion of rRNA by hybridization of complementary DNA oligonucleotides, followed by treatment with RNase H and DNase. Next steps include mRNA enrichment and fragmentation, first strand cDNA synthesis using random priming followed by second strand synthesis converting cDNA:RNA hybrid to double-stranded cDNA (dscDNA), and incorporating dUTP into the second cDNA strand. cDNA generation is followed by end repair to generate blunt ends, A-tailing, adaptor ligation and PCR amplification. Different adaptors were used for multiplexing samples in one lane. Sequencing was performed on Illumina NovaSeq6000 for a paired-end 2x50 run. Data quality check was done on Illumina SAV. Demultiplexing was performed with Illumina Bcl2fastq v2.19.1.403 software. L001 and L002 refer to the two lanes where samples were run.

value: prostate

tag: cell type

value: basal-derived organoid

value: Wildtype

value: Butyrate

tag: strain


title: Day 10 lean [X56]

value: Biol rep 2


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL02

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL02_DpGC31_40_S10


title: Day 10 lean [X54]


title: Butyrate 2


title: MCF7_NAIL_WT_TNFa_90min_p65_chip_rep2


title: UK5099 + Butyrate 3

value: UK5099 + Butyrate


title: Day 7 Lean [X23]

value: Bone fracture healing callus (Day 7 post-fracture)

value: Biol rep 3

value: Day 7 Lean


title: MCF7_NAIL_WT_TNFa_90min_p65_chip_rep1

value: TNFa

value: p65 (Santa-Cruz, sc-372)


title: Butyrate 1


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL02

description:
SMART-Seq v4 3' DE Kit
IL02_DmGC51_60_S6


title: Day 7 Lean [X20]


title: UK5099 3


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL02

description:
SMART-Seq v4 3' DE Kit
IL02_DmGC41_50_S5


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL02

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL02_DpGC21_30_S9


title: MCF7_NAIL_ΔNFκB_TNFa_45min_p65_chip_rep2


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL03

description:
SMART-Seq v4 3' DE Kit
IL03_DmGC51_60_S6


title: UK5099 + Butyrate 2


title: MCF7_NAIL_WT_TNFa_45min_p65_chip_rep2


title: MCF7_NAIL_ΔNFκB_TNFa_0min_p65_chip_rep2

value: control


title: UK5099 2


title: UK5099 + Butyrate 1


title: Day 7 Lean [X19]

value: Biol rep 1

value: Lean


title: Day 5 Lean [X13]

value: Bone fracture healing callus (Day 5 post-fracture)

value: Day 5 Lean


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL02

description:
SMART-Seq v4 3' DE Kit
IL02_DmGC21_30_S3

value: LCM

value: Neighboring

value: gene expression


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL01

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL01_DpGC51_60_S12


title: MCF7_NAIL_WT_TNFa_45min_p65_chip_rep1


title: UK5099 1

value: prostate

value: basal-derived organoid

value: Wildtype


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL01

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL01_DpGC41_50_S11


title: Day 5 Lean [X12]

value: Biol rep 2


title: MCF7_NAIL_ΔNFκB_TNFa_0min_p65_chip_rep1


title: Vehicle 2

value: Vehicle


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL01

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL01_DpGC21_30_S9


title: Vehicle 1


title: ganglioid culture, DIV4_rep4

city: London

first: Vassilis

last: Pachnis

state: London

address: 1 Midland Road

department: Development and Homeostasis of the Nervous System Laboratory

country: United Kingdom

institute: The Francis Crick Institute

description:
DIV4_60.mSm_peaks.annotatePeaks.txt
DIV4_60.mSm_peaks.broadPeak
DIV4_60.mSm_peaks.gappedPeak
DIV4_60.mSm_peaks.txt

data_processing:
Reads from ATAC-Seq fastq files were aligned to the Ensembl GRCm38 genome, peaks mapped and quantified using the nextflow atacseq pipeline version 1.0dev (https://nf-co.re/atacseq).
Assembly: hg38
Supplementary files format and content: NF-core ATAC-seq pipeline MACS output

source_name: Enteric nervous system

growth_protocol: Mice used in this study were maintained according to standard practices and all procedures were approved by The Francis Crick Institute Animal Ethics Committee and conformed to UK Home Office requirements (ASPA 1986) and the ARRIVE guidelines.

extract_protocol:
Crude nuclei were isolated using IGEPAL solution from 50,000 FACS sorted tdTomato cells after 4 days in vitro.
Nuclei were subjected to tagmentation using the Illumina Nextera Tn5 transposase following published protocol with some modifications. Transposition was carried out at 37°C for an hour instead of the recommended 30mins for all samples processed. DNA fragments were purified using Qiagen MinElute PCR kit and subjected to PCR amplifications using the NEBNext High-Fidelity 2x PCR Master Mix (New England Biolabs) and appropriate barcoded primers as described. Amplified fragments were further purified using AMPure XP Beads (Beckman Coulter) according to manufacturer’s recommendation to remove unamplified primers before sequencing. The resulting ATAC libraries were quantified using the GloMax multi-detection system (Promega) and size distribution assayed using a TapeStation D1000 ScreenTape (Agilent). The libraries were then pooled (4 nM) for sequencing and loaded onto the HiSeq 4000 (Illumina) for paired end 100 bp reads sequencing.

treatment_protocol: Neurogenic EGC cultures were established from Sox10Cre|tdT mice (24 hours after a single dose of tamoxifen injection at 100 μg/g of body weight).

value: Enteric nervous system

value: Enteric glial culture


title: MCF7_NAIL_WT_TNFa_0min_p65_chip_rep2


title: ganglioid culture, DIV4_rep1


title: Mpc1-KO 3 [RNA9_Cre_S47]

description: Master File_CPM+0.0001_Normalized_Counts-0120.xlsx

data_processing:
The reads were mapped by STAR 2.7.9a and read counts per gene were quantified using the mouse Ensembl GRCm39.105 GTF file. In Partek Flow, read counts were normalized by CPM +1.0E-4.
Supplementary files format and content: tab-delimited text files include CPM values for each Sample

extract_protocol: Libraries for RNA-Seq were prepared with KAPA Stranded mRNA-Seq Kit. The workflow consists of mRNA enrichment  and fragmentation, first strand cDNA synthesis using random priming followed by second strand synthesis converting  cDNA:RNA hybrid to double-stranded cDNA (dscDNA), and incorporates dUTP into the second cDNA strand. cDNA  generation is followed by end repair to generate blunt ends, A-tailing, adaptor ligation and PCR amplification. Different  adaptors were used for multiplexing samples in one lane. Sequencing was performed on Illumina HiSeq3000 for a SE  1x50 run. Data quality check was done on Illumina SAV. Demultiplexing was performed with Illumina Bcl2fastq  v2.19.1.403 software.

treatment_protocol: Organoids were treated with vehicle or 10uM UK5099 for one week

value: Mpc1 floxed

value: Mpc1 knockout


title: ganglioid culture, DIV4_rep2


title: ganglioid culture, DIV4_rep3


title: Mpc1-KO 2 [RNA8_Cre_S46]


title: therapeutic, saline-OT2-7

city: Chicago

first: Jeffrey

last: Hubbell

state: Illinois

address: 5640 S. Ellis Ave

institute: University of Chicago

description:
sb7
count_protein_coding_kallisto_therapeutic.txt

data_processing:
Fastq files were generated using bcl2fastq
kallisto (v0.46.1) was used to quantify abundances of transcripts from the fastq files.
Count tables were generated using R package tximport to import transcript-level estimates from kallisto output and summarize them at gene level
Genome_build: GRCm38.97
Supplementary_files_format_and_content: tab-delimited text file includes matrix table with raw gene count values for each sample

source_name: CD4 T cells

extract_protocol:
Spleens and lymph nodes (inguinal, axial, brachial, popliteal, cervical) were pooled. OT-I and OT-II were isolated separately using fluorescence activated cell sorting (FACS) into Trizol (prophylactic) or FACS buffer, spun, decanted, and snap frozen (therapeutic) and frozen at -80. RNA was extracted (Qiagen). For prophylactic three mice were pooled per sample, for therapeutic one mouse per sample.
Pooled samples were converted to cDNA using SmartSeq v4 protocol. Libraries were constructed using iDT Nextera, paired end 150.

treatment_protocol: At 1 day post adoptive transfer of OTI and OTII cells, mice received either 5ug plain OVA, the molar antigen equivalent of pGal-OVA, or saline.

value: OT-II

value: Saline

tag: group

value: therapeutic


title: Vehicle 3


title: MCF7_NAIL_WT_TNFa_0min_p65_chip_rep1

value: control

value: p65 (Santa-Cruz, sc-372)


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL01

description:
SMART-Seq v4 3' DE Kit
IL01_DmGC21_30_S3

value: LCM


title: UK5099 3 [RNA6_UK_S44]


title: Day 3 Lean [X29]

value: Bone fracture healing callus (Day 3 post-fracture)

value: Biol rep 1

value: Lean

value: Day 3 Lean


title: BL6 microbiota-colonized ileal Paneth cells

city: Newark

last: Gao

state: NJ

address: 225 University Ave

department: Biological Sciences

institute: Rutgers University

description: Illumina Novaseq 6000 - 10x Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software (v3.0.1)
Supplementary files format and content: mm10
Supplementary files format and content: Matrix file

source_name: Distal 2/3 of ileum

growth_protocol: Paneth cell reporter mice were maintained under gnotobiotic conditions before and after microbiota transfer.

extract_protocol:
Crypt epithelial cells were isolated from GF and exGF+B6M mice and subjected to FACS sorting for isolation of tDT positive Paneth cells
Single Cell 3' v3

treatment_protocol: Germ-free (GF) Paneth cell reporter mice were orally gavaged with sterile PBS to keep as germ-free controls, or orally gavaged with a freshly prepared fecal slurry prepared from the cecum contents of C57B/L6 mice under anaerobic condition. 3 weeks later, mice were intraperitoneally injected with sterile tamoxifen to activate the reporter. Mice were sacrificed 2 days later when ileal Paneth cells were flow sorted for single cell RNA sequencing.

value: Distal 2/3 of ileum

value: Paneth cells

value: Lyz1-3BACTg(IRES-CreER); R26R-TdtL/+

value: BL6 fecal slurry & tamoxifen two days prior to sacrifice


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL01

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL01_DpGC31_40_S10

value: Neighboring

value: gene expression


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL01

description:
SMART-Seq v4 3' DE Kit
IL01_DmGC51_60_S6


title: trunk_e95

city: Vienna

first: Artem

middle: V

last: Artemov

address: Spitalgasse 4

department: Center for Brain Research

country: Austria

institute: Medical University of Vienna

description: Neural crest derivatives from the trunk (cut posterior to the ottic vesicle) of mouse embryos at embryonic day E9.5

data_processing:
Sequenced bcl files were converted to fastq files with cellranger-atac, version 1.2.0
The sequenced 10X libraries were mapped to mm10 mouse genome and assigned to droplets with cellranger-atac, version 1.2.0 with default parameters
Assembly: mm10
Supplementary files format and content: *.filtered_peak_bc_matrix.h5 - peak count matrix (in h5 format); *.fragments.tsv.gz - all mapped ATAC fragments per cell; *.singlecell.csv - metadata (quality metrics) for individual cells (generated by cellranger-atac)

source_name: Neural crest derivatives

growth_protocol: Mouse embryos were collected at embryonic day E9.5. Transgenic embryos contained Wnt1-Cre/R26Tomato construct.

extract_protocol:
After dissociation, cells were FACS-sorted based on the expression of Tomato. Tomato-positive fluorescent cells were sorted for ATAC sequencing. Cell nuclei were isolated according to a standard 10X protocol for single-nucleus ATAC-seq.
Library was prepared according to the standard 10X single cell ATAC-seq protocol

value: Neural crest derivatives


title: Germ-free ileal Paneth cells

description: Illumina Novaseq 6000 - 10x Genomics

value: PBS & tamoxifen 2 days prior to sacrifice


title: UK5099 2 [RNA5_UK_S43]

value: prostate

value: basal-derived organoid

value: Mpc1 floxed


title: Day 3 Lean [X32]

value: Biol rep 3


title: therapeutic, saline-OT2-6

description:
sb6
count_protein_coding_kallisto_therapeutic.txt

value: OT-II

value: Saline

value: therapeutic


title: Mpc1-KO 1 [RNA7_Cre_S45]

value: Mpc1 knockout


title: Lung, LAIR-1 KO sample 6

first: Georges Doumet

last: HELOU

state: California

address: 1450 Biggy St, NRT5505, Keck School of Medicine

department: Molecular Microbiology and Immunology

institute: University of Southern California

description: 21079R-03-24_S92 -cKO

data_processing:
Illumina Casava1.7 software used for basecalling, algorithm
RNAseq data were analyzed using Partek® Flow® software, version 9.0 Copyright ©
Raw reads were first trimmed based on Quality Score (min quality level 20, trim from both ends, min read length 25)
Trimmed reads were aligned to mm10 with Gencode M21 as guidance using STAR - 2.7.3a
Aligned reads were quantified to Gencode M21 genes using Partek E/M. Normalized reads counts were tested for differential expression using Partek's GSA algorithm.
Assembly: mm10
Supplementary files format and content: Counts and normalized counts

source_name: Freshly sorted LAIR-1 KO lung macrophages

extract_protocol:
Total RNA was isolated using MicroRNAeasy (Qiagen). 10ng of input RNA was used to produce cDNA for downstream library preparation.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: RNAseq was performed on freshly sorted macrophages after three intranasal administrations of Poly(I:C).

value: Lungs

value: LAIR-1 KO

tag: age

value: 10 weeks

value: macrophages


title: Day 3 Lean [X31]

value: Bone fracture healing callus (Day 3 post-fracture)

value: Biol rep 2

value: Lean

value: Day 3 Lean


title: Ileum Paneth Cells

description: Illumina Novaseq 6000 - 10x Genomics

source_name: Ileum of small intestine

growth_protocol: Paneth cell reporter mice were maintained under specific pathogen free condition.

extract_protocol:
Crypt epithelial cells were isolated from mouse duodenum and ilea and subjected to FACS sorting for isolation of tDT positive Paneth cells
Single Cell 3' v3

treatment_protocol: Mice were intraperitoneally injected with tamoxifen, and sacrificed 48 hours later. Duodenum and ilea were dissected.

value: Ileum of small intestine

value: Paneth cells

value: Lyz1-3BACTg(IRES-CreER); R26R-TdtL/+

value: tamoxifen


title: therapeutic, saline-OT2-5

description:
sb5
count_protein_coding_kallisto_therapeutic.txt


title: Lung, LAIR-1 KO sample 4

description: 21079R-03-23_S91 -cKO


title: UK5099 1 [RNA4_UK_S42]

value: prostate

value: basal-derived organoid

value: Mpc1 floxed


title: therapeutic, saline-OT2-4

description:
sb4
count_protein_coding_kallisto_therapeutic.txt

value: OT-II

value: Saline

value: therapeutic


title: Duodenum Paneth Cells

description: Illumina Novaseq 6000 - 10x Genomics

source_name: Duodenum of small intestine

value: Duodenum of small intestine


title: Lung, WT sample 3

description: 21079R-03-17_S74 -WT

source_name: Freshly sorted WT lung macrophages

value: Lungs

value: WT

value: 10 weeks


title: Day 5 Lean [X10]

value: Bone fracture healing callus (Day 5 post-fracture)

value: Biol rep 1

value: Day 5 Lean


title: Vehicle 3 [RNA3_RFP_S41]

value: Vehicle


title: Abx SAL Paneth cells

description: Illumina Novaseq 6000 - 10x Genomics

extract_protocol:
Epithelial cells were isolated from the ilea of both mice and subjected to FACS sorting for isolation of tDT positive Paneth cells
Single Cell 3' v3

treatment_protocol: Mice were intraperitoneally injected with tamoxifen. One day after, mice were orally gavaged with Streptomycin. One day after, mice were orally inoculated with PBS (serve as control "Abx") or with Salmonella (experimental group "Abx SAL").

value: Distal 2/3 of ileum

value: Paneth cells

value: Streptomycin +10^8 CFU Salmonella + tamoxifen 6 days prior to sacrifice


title: therapeutic, saline-OT2-3

description:
sb3
count_protein_coding_kallisto_therapeutic.txt


title: Abx Paneth cells

description: Illumina Novaseq 6000 - 10x Genomics

value: Lyz1-3BACTg(IRES-CreER); R26R-TdtL/+

value: Streptomycin (15mg/100g mouse) + tamoxifen 6 days prior to sacrifice


title: therapeutic, saline-OT2-2

description:
sb2
count_protein_coding_kallisto_therapeutic.txt


title: Lung, LAIR-1 KO sample 5

description: 21079R-03-22_S90 -cKO

value: LAIR-1 KO

value: macrophages


title: Lung, WT sample 2

description: 21079R-03-18_S88 -WT


title: Lung, WT sample 1

description: 21079R-03-20_S89 -WT


title: WT, oxygen-glucose deprivation, replicate 3

city: Granada

first: Jose

middle: Luis

last: Ruiz Rodriguez

state: Granada

address: Avda. del Conocimiento 17. P. T. Ciencias de la Salud

country: Spain

institute: IPBLN-CSIC

description:
OGD_table_samples_29775_annotated_genes.txt
OGD_KO_vs_WT_19_annotated_DEGs.txt

data_processing:
We used the miARma-Seq pipeline to analyze and calculate the differential expressed genes (DEGs) (Andres-Leon et al., 2016), with Mus musculus Gencode version M25 genome-build (mmGRCm38.p6) for alignment of reads. Some divergent replicates according to PCAs and clustering analyses were discared and not included in the normalization step in order to preserve biological differences
Differential expression analysis was conducted by using edgeR package (Nikolayeva & Robinson, 2014; RRID:SCR_012802), and genes were normalized by trimmed mean of M-values method (Robinson & Oshlack, 2010)
We calculated reads per kb per million mapped reads and log2-counts per million per gene in each sample (Nikolayeva & Robinson, 2014).
To infer the replicability/similarity of RNA-sequencing samples, we used Principal Component analysis and Hierarchical Clustering of normalized samples (Reeb et al., 2015; Ritchie et al., 2015). Genes giving false discovery rate (FDR) values <0.05 were marked as DEGs, and Log2-FC was used to calculate the fold-change expression of each gene between KO and WT
To identify the effects of DEGs, functional enrichment study was carried out using the clusterProfiler Bioconductor package (Yu et al., 2012; RRID:SCR_016884), in which DEGs were compared against all expressed genes in the RNA-seq assay. Gene Ontology terms (Biological process, Molecular function) were obtained from the Bioconductor Mus musculus database and associated to Entrez gene identifiers in an orgDB R object through the AnnotationForge package (with clusterProfiler). Multiple exetrnal databases, primarly GeneCards and PathCards were integrated
Assembly: mmGRCm38.p6
Supplementary files format and content: RPKM for all the samples and gene annotation, based on multiple external databases (see paper Methods)
Supplementary files format and content: Subset of genes that are significant (FDR < 0.05) for the comparison between KO and WT samples

source_name: brain

extract_protocol:
RNA (1 μg) from primary Cort−/− and Cort+/+ endothelial cell cultures (three independent biological replicated experiments each, with RNA Integrity Number coefficients >9, using Bioanalyzer RNA 6000 Nano-chip, from Agilent, Santa Clara, CA, USA)
RNA was used to prepare mRNA libraries with TruSeq Stranded mRNA Library Prep Kit (Illumina, San Diego, CA, USA, cat#20020595). Quality and size distributions of indexed mRNA libraries were validated by Bioanalyzer High Sensitivity DNA assay (Agilent), and the final libraries were pooled equimolecularly and diluted/denatured as recommended. The 40x2nt paired-end sequencing was conducted on an Illumina NextSeq-500 sequencer (highest output mode), producing 46,850,000 raw paired-reads on average

value: brain

value: endothelial cells

value: ischemic stroke mouse model (MCAO)

value: OGD

tag: replicate


title: DKO UK5099 3 [Cre_UK_3_S12]

data_processing:
The Partek flow  and Ingenuity Pathway Analysis (IPA) were used for the data analysis. The alignment was performed using STAR with mouse reference genome mm10. The mm10 - Ensembl Transcripts release 100 gtf was used for gene feature annotation. For normalization of transcripts counts, the normalization method, adding 1.0E-4 followed by counts per million (CPM), was used. Normalized TPM was also generated for the project.
Assembly: mm10
Supplementary files format and content: tab-delimited text files include TPM values for each Sample

treatment_protocol: DKO organoids were treated with vehicle or 10uM UK5099 for five days

tag: tissue

value: prostate

value: basal-derived organoid

value: PTEN, Rb1 double knockout

tag: strain background

value: mixed C57BL/6:129/Sv:FVB


title: WT, oxygen-glucose deprivation, replicate 2

tag: disease state

value: WT


title: CON, replicate_2, Ventricular tissue, scRNA-seq

city: Chongqing

first: Li

last: Guoxing

state: Chongqing

address: Street of shiyou Road, Yuzhong District, Chongqing, China

country: China

institute: Chongqing Medical University

description: 10x Genomics

data_processing:
scRNA-seq: The demulitiplexing, barcoded processing, gene counting and agregation were  performed by using Cell Ranger software
Assembly: mm10
Supplementary files format and content: scRNA-seq: separated values files and matrix files

source_name: Ventricular tissue

extract_protocol:
The heart was minced and incubated in a mixture of type II and IV collagenase at 37°C, followed by filtration through a 40 μm filter. The cells were then resuspended in red blood cell lysis buffer and dead cells were removed.
scRNA-seq: Use a cell counter to detect the sample. When the sample is qualified, use the standard 10x Genomics library building process combined with illumina library building process to build the library (library building reagent: chromium next gem single cell 3' library and gel bead kit v3.1).

value: Ventricular tissue

tag: Sex

value: male

value: Normal diet, Normal drinking water


title: CON, replicate_3, Ventricular tissue, scRNA-seq

description: 10x Genomics


title: therapeutic, saline-OT1-8

description:
sa8
count_protein_coding_kallisto_therapeutic.txt

source_name: CD8 T cells

value: OT-I

value: Saline

value: therapeutic


title: HFpEF, replicate_3, Ventricular tissue, scRNA-seq

description: 10x Genomics

value: High-fat diet, L-NAME 0.5 g/L in drinking water


title: WT, oxygen-glucose deprivation, replicate 1


title: CON, replicate_1, Ventricular tissue, scRNA-seq

description: 10x Genomics


title: therapeutic, saline-OT1-6

description:
sa6
count_protein_coding_kallisto_therapeutic.txt


title: DKO Vehicle 3 [Cre_Vehicle_3_S9]

value: Vehicle


title: WT, normoxia, replicate 3

description:
NM_table_samples_30269_annotated_genes.txt
NM_KO_vs_WT_613_annotated_DEGs.txt

value: brain

value: NM


title: DKO UK5099 1 [Cre_UK_1_S10]


title: WT, normoxia, replicate 2

value: endothelial cells

value: ischemic stroke mouse model (MCAO)


title: DKO Vehicle 2 [Cre_Vehicle_2_S8]

value: prostate


title: therapeutic, saline-OT2-1

description:
sb1
count_protein_coding_kallisto_therapeutic.txt

value: OT-II


title: WT, normoxia, replicate 1


title: HFpEF, replicate_1, Ventricular tissue, scRNA-seq

description: 10x Genomics


title: therapeutic, saline-OT1-5

description:
sa5
count_protein_coding_kallisto_therapeutic.txt


title: therapeutic, saline-OT1-3

description:
sa3
count_protein_coding_kallisto_therapeutic.txt


title: HFpEF, replicate_2, Ventricular tissue, scRNA-seq

description: 10x Genomics


title: DKO Vehicle 1 [Cre_Vehicle_1_S7]

value: basal-derived organoid

value: PTEN, Rb1 double knockout


title: CON, replicate_6, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
HF-6

data_processing:
bulk RNA-seq: Quality control, Samples are sequenced on the platform to get image files, which are transformed by the software of the sequencing platform, and the original data in FASTQ format (Raw Data) is generated. Sequencing data contains a number of connectors, low-quality Reads, so we use Cutadapt (v1.15) software to filter the sequencing data to get high quality sequence（Clean Data）for further analysis.
Reads mapping：The reference genome and gene annotation files were downloaded from genome website. The filtered reads were mapping to the reference genome using HISAT2 v2.0.5.
Differential expression analysis: we used HTSeq(0.9.1) statistics to compare the Read Count values on each gene as the original expression of the gene, and then used FPKM to standardize the expression.
Assembly: mm10
Supplementary files format and content: bulk RNA-seq: FPKM

extract_protocol:
bulk RNA-seq: Total RNA was isolated using the Trizol Reagent (Invitrogen Life Technologies), after which the concentration, quality and integrity were determined using a NanoDrop spectrophotometer (Thermo Scientific). Three micrograms of RNA were used as input material for the RNA sample preparations.
bulk RNA-seq: Sequencing libraries were generated according to the following steps. Firstly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in an Illumina proprietary fragmentation buffer. First strand cDNA was synthesized using random oligonucleotides and Super Script II. Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities and the enzymes were removed. After adenylation of the 3′ ends of the DNA fragments, Illumina PE adapter oligonucleotides were ligated to prepare for hybridization. To select cDNA fragments of the preferred 400-500 bp in length, the library fragments were purified using the AMPure XP system (Beckman Coulter,Beverly, CA, USA). DNA fragments with ligated adaptor molecules on both ends were selectively enriched usingIllumina PCR Primer Cocktail in a 15 cycle PCR reaction Products were purified (AMPure XP system) and quantified using the Agilent high sensitivity DNA assay on a Bioanalyzer 2100 system (Agilent).

value: WT


title: KO, oxygen-glucose deprivation, replicate 3

value: KO

value: OGD


title: DKO UK5099 2 [Cre_UK_2_S11]

value: mixed C57BL/6:129/Sv:FVB


title: therapeutic, saline-OT1-7

description:
sa7
count_protein_coding_kallisto_therapeutic.txt

value: OT-I


title: therapeutic, saline-OT1-2

description:
sa2
count_protein_coding_kallisto_therapeutic.txt

value: Saline


title: Vehicle 1 [RNA1_RFP_S39]

description: Master File_CPM+0.0001_Normalized_Counts-0120.xlsx

value: Mpc1 floxed


title: CON, replicate_5, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
HF-5

value: Ventricular tissue

value: male

value: Normal diet, Normal drinking water


title: KO, oxygen-glucose deprivation, replicate 2

description: Processed files should not apply to KO replicate 2 in OGD condition.

value: brain


title: DKO_CRE_RFP_2_S8

description:
DKO_CRE_RFP_2_S8
SKO vs DKO organoid RNAseq TPM 2020.xlsx

data_processing:
The Partek flow and Ingenuity Pathway Analysis (IPA) were used for bioinformatics methods and data analysis. The reads were aligned to reference genome version GRCm38, and reads per gene was quantified by  STAR 2.27a using Ensembl GRCm38.97 GTF file. After obtaining gene counts, the counts were normalized by CPM and added 0.001.
Assembly: GRCm38
Supplementary files format and content: TPM values for each Sample

extract_protocol: Libraries for RNA-Seq were prepared with TruSeq Stranded mRNA Library Prep Kit. The workflow consists of mRNA  enrichment and fragmentation. Cleaved RNA fragments are copied into first strand cDNA using reverse transcriptase  and random primers. Strand specificity is achieved by replacing dTTP with dUTP and followed by second strand cDNA  synthesis using DNA Polymerase I and RNase H. cDNA generation is followed by A-tailing, adaptor ligation and PCR  amplification. Different adaptors were used for multiplexing samples in one lane. Sequencing was performed on Illumina  HiSeq 3000 for SE 1x50 run. Data quality check was done on Illumina SAV. Demultiplexing was performed with Illumina  Bcl2fastq v2.19.1.403 software.

value: PTEN, Rb1 double knockout (DKO)


title: therapeutic, saline-OT1-1

description:
sa1
count_protein_coding_kallisto_therapeutic.txt

value: therapeutic


title: Vehicle 2 [RNA2_RFP_S40]

value: prostate

value: Vehicle


title: therapeutic, pGal-OT2-9

description:
pb9
count_protein_coding_kallisto_therapeutic.txt

value: OT-II

value: pGal


title: KO, oxygen-glucose deprivation, replicate 1

value: endothelial cells

value: ischemic stroke mouse model (MCAO)


title: SKO_CRE_RFP_3_S6

description:
SKO_CRE_RFP_3_S6
SKO vs DKO organoid RNAseq TPM 2020.xlsx

value: basal-derived organoid

value: PTEN single knockout (SKO)


title: KO, normoxia, replicate 3

description: Processed files should not apply to KO replicate 3 in normoxia.

value: KO

value: NM


title: CON, replicate_3, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
HF-3

value: WT


title: DKO_CRE_RFP_1_S7

description:
DKO_CRE_RFP_1_S7
SKO vs DKO organoid RNAseq TPM 2020.xlsx

value: mixed C57BL/6:129/Sv:FVB


title: therapeutic, saline-OT1-4

description:
sa4
count_protein_coding_kallisto_therapeutic.txt

value: OT-I

value: Saline


title: KO, normoxia, replicate 2

value: brain


title: CON, replicate_2, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
HF-2

value: Ventricular tissue

value: male

value: Normal diet, Normal drinking water


title: KO, normoxia, replicate 1


title: therapeutic, pGal-OT2-7

description:
pb7
count_protein_coding_kallisto_therapeutic.txt

value: therapeutic


title: SKO_CRE_RFP_1_S4

description:
SKO_CRE_RFP_1_S4
SKO vs DKO organoid RNAseq TPM 2020.xlsx

value: prostate


title: CON, replicate_1, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
HF-1


title: therapeutic, pGal-OT2-5

description:
pb5
count_protein_coding_kallisto_therapeutic.txt

value: OT-II

value: pGal


title: SKO_RFP_3_S3

description:
SKO_RFP_3_S3
SKO vs DKO organoid RNAseq TPM 2020.xlsx

value: Wildtype


title: HFpEF, replicate_5, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
con-5

value: High-fat diet, L-NAME 0.5 g/L in drinking water


title: HFpEF, replicate_6, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
con-6


title: therapeutic, pGal-OT2-6

description:
pb6
count_protein_coding_kallisto_therapeutic.txt


title: DKO_CRE_RFP_3_S9

description:
DKO_CRE_RFP_3_S9
SKO vs DKO organoid RNAseq TPM 2020.xlsx

value: basal-derived organoid

value: PTEN, Rb1 double knockout (DKO)


title: SKO_RFP_2_S2

description:
SKO_RFP_2_S2
SKO vs DKO organoid RNAseq TPM 2020.xlsx


title: Y-adaptor seq. single oocyte Ex_Oocyte_28_S4

data_processing:
After trimming polyA/T sequences, adapter sequences, and poor quality reads, using fastp (v. 0.20.0) and cutadapt (ver. 1.16), the sequenced reads were aligned to the GRCm38 mouse genome assemblywith ERCC spike-in RNA sequences using the HiSAT2 (v. 2.1.0).
Best-matched reads on annotated genes were counted using HTSeq 0.10.0 with a modified GRCm38 gtf file which is extended up to 10 Kb downstream from transcription termination sites of reference genes.
Mapped reads on exons and exon-exon junctions were counted by featureCounts (v2.0.1) with the GRCm38 gtf file and the option "-J" "-p" "-M" "-O" "-t　exon" "-g　transcript_id".
Genome_build: GRCm38
Supplementary_files_format_and_content: Matrix table with raw gene, exon and exon-exon junction counts for every sample.

source_name: Oocyte

extract_protocol:
Cells were isolated with LCM in 6.4 μL of cell lysis buffer (1× GeneAmp PCR Buffer II [Thermo Fisher], 1.5 mM MgCl2 [Thermo Fisher], 0.5% sodium deoxycholate [SDc] [Nacalai Tesque], and 5 mM dithiothreitol [DTT] [Thermo Fisher] dissolved in deionized distilled water [Gibco] containing 4 U RNaseOUT [Thermo Fisher], 0.4 U porcine liver RNase inhibitor [Takara Bio], 0.05 mM dNTPs [Takara Bio], ERCC RNA Spike-In Mix 1 [Thermo Fisher], and 1.6 ng V1[dT]24 primer ) in the caps of PCR tubes, and spun down into the tubes by brief centrifugation.  Cells were then lysed at 70°C for 6 min, followed by the addition of 2.8 µL quenching buffer (25% Triton X-100 [Nacalai Tesque] and 1% bovine serum albumin [BSA] [Takara Bio]) and incubation at 70°C for 90 sec to quench the denaturing activity of SDc.
The cDNA synthesis and amplification onward were performed as described previously with minor modifications [Kurimoto, K.et.al., Nucleic Acids Research, 34(5), e42.]. In brief, the first-strand cDNA was synthesized by SuperScript III [Thermo Fisher] and incubating at 50℃  for 5 min followed by heat inactivation at 70℃  for 10 min.  The excess RT primers were degraded by exonuclease I and incubating at 37℃ for 30 min followed by heat inactivation at 80℃ for 25 min. The first-strand cDNA product was poly dA-tailed by TdT and incubating at 37℃  for 1 min followed by heat inactivation at 70℃ for 10 min. Second-strand cDNA synthesis were performed by adding 9.5 μL reaction mixture of ExTaq, Hot Start version [Takara Bio] and V3[dT]24 primer in the ExTaq buffer with the following thermal cycling program (95℃  for 3 min, 50℃  for 2 min, and 72℃  for 3 min). cDNA amplification was performed by addingthe mixture of V1[dT]24 and ExTaq, Hot Start version [Takara Bio] in the ExTaq buffer with the following thermal cycling program (95℃  for 1 min; 20 cycles of 95℃  for 30 sec, 67℃  for 1 min and 72℃  for 3 min with a 6 sec elongation per cycle; and 72℃  for 10 min).  The amplified cDNA was purified with a 0.6× volume of AxyPrep MAG PCR clean-up reagent (Axygen) according to the manufacturer's instructions. Preparation of the libraries for SC3-seq was performed essentially as previously described (Nakamura et al., 2015)(Nakamura et al., 2017).  For Y-adapter sequencing, single-cell cDNAs were pre-amplified using the primers (V3[dT]24, V1[dT]24) and ExTaq HS with the thermal cycling program (95℃  for 3 min; 4 cycles of 95°C for 30 sec, 67℃  for 1 min and 72℃  for 5 min; 72℃  for 10 min). Pre-amplified cDNAs were then subjected to the primer-dimer removal, sonication, end polishing, and the size selection as in SC3-seq. Then, purified cDNA was dA-tailed in a 10 μL dA tailing mixture with incubation at 72℃  for 10 min. The dA-tailed product was subjected to the Y-shaped adaptor ligation by adding a mixture of Y-adaptor and NEB Next Quick Ligation Reaction Buffer and incubating at 20℃  for 15 min and at 72℃  for 20 min. The ligation product was purified with a 0.9×volume of AxyPrep reagent, and subjected to additional PCR amplification in 20 μL with the thermal cycling program (95℃  for 3 min, 55℃  for 1 min, and 72℃  for 1 min; 11 cycles of 95℃  for 30 sec, 60℃  for 1 min, and 72℃  for 1 min; and 72℃  for 3 min), followed by DNA purification with a 0.9×volume of AxyPrep reagent.

value: LCM

value: Secondary-antral

value: exon-exon junction


title: Y-adaptor seq. single oocyte Ex_Oocyte_30_S5


title: CON, replicate_4, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
HF-4

value: WT


title: therapeutic, pGal-OT2-4

description:
pb4
count_protein_coding_kallisto_therapeutic.txt


title: HFpEF, replicate_4, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
con-4


title: therapeutic, pGal-OT2-3

description:
pb3
count_protein_coding_kallisto_therapeutic.txt


title: SKO_RFP_1_S1

description:
SKO_RFP_1_S1
SKO vs DKO organoid RNAseq TPM 2020.xlsx

value: mixed C57BL/6:129/Sv:FVB


title: therapeutic, pGal-OT2-8

description:
pb8
count_protein_coding_kallisto_therapeutic.txt


title: Y-adaptor seq. single oocyte Ex_Oocyte_21_S3


title: Y-adaptor seq. single oocyte Ex_Oocyte_17_S2


title: Y-adaptor seq. single oocyte Ex_Oocyte_10_S1


title: HFpEF, replicate_3, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
con-3


title: SSCT F0 bio rep 3

city: Amsterdam

first: Joana

middle: B

last: Serrano

address: Meibergdreef 9

country: Netherlands

institute: Amsterdam UMC - Location AMC

data_processing:
RRBS libraries were sequenced generating 50 bp single reads on an Illumina HiSeq 3000 instrument. The sequenced reads were controlled for quality of sequencing with the FastQC tool. Adapter removal was performed using Trim Galore! version 0.4.1.
The cleaned reads were then aligned to the reference genome using Bismark v0.20.0. The cytosine2coverage and bismark_methylation_extractor modules of Bismark were used to infer the methylation state of all cytosines (for every single uniquely mappable read) and their context, and to compute the percentage methylation. The reported cytosines were filtered to get only the CpGs covered in each sample. The spike-in control sequences were used at this step to check the bisulfite conversion rates and to validate the efficiency of the bisulfite treatment.
Methylkit v1.7.0, a R/Bioconductor package, was used to perform the differential methylation analysis  between the two groups of samples from each generation. The CpG data set was filtered for low coverage (CpGs with coverage  less than 10X in all samples per comparative group were discarded) and for extremely high coverage  to discard reads with possible PCR bias (CpGs with coverage higher than the 99.9th percentile were  discarded). The data was then normalized for read coverage distribution between samples.
Assembly: GRCm38/mm10
Supplementary files format and content: Tab delimited methylation file with .outputForMethylKit.tsv extension. Contains the methylation information on all covered Cs in CpG context only. Coordinates are 1-based. Each line has 5 fields: chromosome - start and end position - strand - methylation level (from 0 to 1) - coverage (number of reads covering the position). These files were be used for further statistical analyses  with MethylKit.

source_name: epidydimal sperm

extract_protocol:
Sperm was isolated from the cauda epididymis from control F1, SSCT F1, SSCT F2/M and SSCT F2/P offspring (n=5 for each). For the three SSCT F0 transplanted W/W-v males and DBA2/J controls, donor-derived DBA2/J and control epididymal sperm were isolated at 16-18 months of age, as they were needed to breed the offspring for a prolonged time. with sSperm was collected in an Eppendorf and washed in 500uL Phosphate-buffered saline (PBS). For purification of the sperm, the suspension was carefully pipetted into a falcon with 2mL PureSperm 70% (Nidacon) while having the falcon almost horizontal and centrifuged for 20 minutes at 300 RCF no break at room temperature (RT). The pellet was then washed with 5 mL HBSS or PBS and centrifuge for 10 minutes at 500 RCF after which the pellet was SNAP frozen and stored at -80ᵒC for further analysis.
RRBS libraries were prepared using the Premium Reduced Representation Bisulfite Sequencing (RRBS) Kit (Diagenode Cat# C02030033). 100ng of genomic DNA were used to start library preparation for each sample, with the exception of samples 21, 23, 24 that had lower input. Following library preparation, samples were pooled together by 9 or 8. PCR clean-up after the final library amplification was performed using a 1.45x beads:sample ratio of Agencourt® AMPure® XP (Beckman Coulter). DNA concentration of the pools was measured using the Qubit® dsDNA HS Assay Kit (Thermo Fisher Scientific).  The profile of the pools was checked using the High Sensitivity DNA chip for 2100 Bioanalyzer (Agilent).  In case of too high adapter dimer peaks, the pools were size selected one more time using a 1.45x beads:sample ratio of Agencourt® AMPure® XP (Beckman Coulter) and quality control steps were performed again. RRBS library pools were sequenced on a HiSeq3000 (Illumina) using 50 bp single-read sequencing (SR50).

value: epidydimal sperm

value: sperm

value: 16-18 months

value: SSCT


title: therapeutic, pGal-OT2-2

description:
pb2
count_protein_coding_kallisto_therapeutic.txt

value: therapeutic


title: HFpEF, replicate_1, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
con-1

value: Ventricular tissue

value: male


title: therapeutic, pGal-OT2-10

description:
pb10
count_protein_coding_kallisto_therapeutic.txt

value: OT-II

value: pGal


title: Control F0 bio rep 2

value: Control


title: therapeutic, pGal-OT2-1

description:
pb1
count_protein_coding_kallisto_therapeutic.txt


title: Y-adaptor seq. single mESC(LCM) Ex_SL06_S4

source_name: Embryonic stem cell


title: Y-adaptor seq. single mESC(LCM) Ex_SL09_S5


title: SSCT F0 bio rep 2


title: Control F0 bio rep 1


title: therapeutic, pGal-OT1-9

description:
pa9
count_protein_coding_kallisto_therapeutic.txt

value: OT-I


title: Control 9464D cells, ATAC-seq, Replicate2

city: Uppsala

first: Xingqi

last: Chen

address: BMC, Husargatan 3

department: Department of Immunology, Genetics and Pathology

country: Sweden

institute: Uppsala university

data_processing:
Sequencing adaptor was trimmed by using pyadapter_trim.py (https://github.com/TheJacksonLaboratory/ATAC-seq/blob/master/auyar/pyadapter_trim.py).
The sequencing reads were aligned to the reference genome (mm10) using Bowtie 2  with the ‘-very-sensitive’ parameter.
The aligned BAM files were sorted and filtered using Samtools. PCR duplicates were removed with Picard (http://broadinstitute.github.io/picard/). BigWig files were generated using the ‘bamCoverage’ function in Deeptools
The transcription start site (TSS) enrichment score was analyzed using the ‘computeMatrix’ and ‘plotProfile’ functions in Deeptools.
MACS2  was used for peak calling, with the parameter ‘-q 0.01 -nomodel -shift 0’. Mouse blacklist regions were removed using bedtools intersect.
Assembly: mm10
Supplementary files format and content: bigwig
Supplementary files format and content: narrowPeak

source_name: Brain

growth_protocol: All cell lines were cultured at 37°C with 5% CO2 using IMDM (Thermo Scientific) supplemented with 10% heat-inactivated FBS and 1% Penicillin-Streptomycin (Thermo Scientific).

extract_protocol:
In brief, 50,000 mouse neuroblastoma cells were centrifuged at 500g for 5 min at room temperature for each reaction. The cell pellet was re-suspended in 50 μL lysis buffer. Samples were purified using Qiagen MinElute PCR Purification kit (Qiagen). The transposed DNA was amplified using NEBNext High-Fidelity 2× PCR master mix.
Standard ATAC-seq libraries were prepared following the previously established protocol. qPCR was performed to determine the optimal number of cycles for final PCR amplification. The PCR product was purified using Qiagen MinElute PCR Purification kit (Qiagen), followed by a size selection step with SPRI beads with 1:1.2 ratio (Beckman Coulter). Finally, the purified DNA was eluted in 20 μL of Elution Buffer.

treatment_protocol: Murine NB cell line 9464D was transfected with ribonucleoprotein (RNP) complexes containing gene-specific crRNA, tracrRNA, and a recombinant Cas9 protein (all from IDT) using an optimized electroporation program.

value: Brain

tag: cell line

value: 9464D cell

value: Mouse neuroblastoma cell line


title: Y-adaptor seq. single mESC(LCM) Ex_SL04_S3

value: LCM

value: exon-exon junction


title: H2afy CRISPR KO 9464D cells, ATAC-seq, Replicate2

value: H2afy CRISPR KO


title: Control F0 bio rep 3

value: epidydimal sperm


title: HFpEF, replicate_2, Ventricular tissue, Bulk RNA-seq

description:
bulk RNA-seq
con-2

tag: genotype

value: WT

value: High-fat diet, L-NAME 0.5 g/L in drinking water


title: Y-adaptor seq. single mESC(LCM) Ex_SL01_S1


title: SSCT F0  bio rep 1

value: sperm

value: 16-18 months

tag: treatment

value: SSCT


title: therapeutic, pGal-OT1-8

description:
pa8
count_protein_coding_kallisto_therapeutic.txt

value: therapeutic


title: therapeutic, pGal-OT1-6

description:
pa6
count_protein_coding_kallisto_therapeutic.txt


title: SKO_CRE_RFP_2_S5

description:
SKO_CRE_RFP_2_S5
SKO vs DKO organoid RNAseq TPM 2020.xlsx

value: prostate

value: basal-derived organoid

value: PTEN single knockout (SKO)

value: mixed C57BL/6:129/Sv:FVB


title: Y-adaptor seq. single mESC(LCM) Ex_SL02_S2


title: SSCT F2/M bio rep 5

value: 3 months

value: male


title: H2afy CRISPR KO 9464D cells, ATAC-seq, Replicate1

value: Brain

value: 9464D cell

value: Mouse neuroblastoma cell line


title: Control 9464D cells, ATAC-seq, Replicate1

value: Control


title: WT4, snRNAseq

city: Dallas

first: Maria

middle: H

last: Chahrour

state: Texas

address: 6000 Harry Hines Boulevard, NB10.104

institute: UT Southwestern Medical Center

description: 10x Genomics

data_processing:
Raw sequencing data were converted to fastq format using the CellRanger mkfastq command (10X Genomics, v1.1.0). Sequencing reads were aligned to the mm10 mouse reference genome and quantified using CellRanger count (10X Genomics, v1.1.0). We used the R package Seurat (v4.1.0) for further filtering and analysis.
Assembly: mm10
Supplementary files format and content: filtered feature barcode matrix files

molecule: nuclear RNA

source_name: Hippocampus

extract_protocol:
Hippocampi were isolated and, for each animal, a punch was collected from the middle of the hippocampus using a 1 mm diameter tissue punch. The punches were immediately stored at -80oC. For each animal, tissue punches from the right and left hippocampi were combined and homogenized using a glass Dounce homogenizer in 500 ml ice-cold Nuclei EZ lysis buffer (NLB) (NUC-101, Sigma) 25 times with pestle A and then 25 times with pestle B. The mixture was transferred to a pre-chilled Eppendorf tube and 1000 ml of NLB was added to the tube and incubated on ice for 5 minutes (min). The tubes were centrifuged at 800 x g for 5 min at 4o C. The nuclei were resuspended in 75 ml of nuclei suspension buffer (consisting of 1X PBS, 1% BSA (AM2618, Thermo Fisher Scientific), and 0.2 U/ml (1%) Rnase inhibitor (AM2694, Thermo Fisher Scientific)). The nuclei were then filtered through a 40 mm Flowmi Cell Strainer (H13680-0040, Bel-Art) to remove cellular debris. Concentration of nuclei for each sample was calculated using Trypan Blue (T8154, Sigma) on a hemocytometer, aiming for a final concentration of ~1000-2000 nuclei/ml.
Single nuclei were processed using the 10X Genomics Next GEM Single Cell 3’ Reagent Kit v3.1 following the manufacturer’s user guide. All mRNA samples were examined for quantity and quality by NanoDrop and Bioanalyzer 2100 (Agilent). The libraries were constructed following TruSeq Stranded mRNA Sample preparation guide (Illumina) and paired end sequencing was performed on the Illumina NovaSeq 6000 platform at the McDermott Center Next Generation Sequencing Core at UTSW.

value: Hippocampus


title: Y-adaptor seq. single mESC(fresh) Ex_S10_S10

value: Fresh

value: exon-exon junction


title: SSCT F2/P  bio rep 4

value: epidydimal sperm

value: sperm


title: Y-adaptor seq. single mESC(fresh) Ex_S06_S8


title: WT2, snRNAseq

description: 10x Genomics

value: WT


title: therapeutic, pGal-OT1-4

description:
pa4
count_protein_coding_kallisto_therapeutic.txt

value: OT-I

value: pGal

value: therapeutic


title: WT3, snRNAseq

description: 10x Genomics


title: SSCT F2/M bio rep 4


title: therapeutic, pGal-OT1-2

description:
pa2
count_protein_coding_kallisto_therapeutic.txt


title: WT1, snRNAseq

description: 10x Genomics


title: SSCT F2/M bio rep 3


title: KO4, snRNAseq

description: 10x Genomics


title: SSCT F2/P bio rep 3

value: SSCT


title: Single oocyte (LCM) O45_S44

value: LCM

value: Primary

value: gene expression (sc3-seq.)


title: Y-adaptor seq. single mESC(fresh) Ex_S02_S6


title: KO2, snRNAseq

description: 10x Genomics


title: SSCT F2/P  bio rep 2

value: 3 months

value: male


title: therapeutic, pGal-OT1-10

description:
pa10
count_protein_coding_kallisto_therapeutic.txt


title: therapeutic, OVA-OT2-8

description:
ob8
count_protein_coding_kallisto_therapeutic.txt

value: OT-II

value: OVA


title: KO3, snRNAseq

description: 10x Genomics

value: Hippocampus


title: therapeutic, pGal-OT1-3

description:
pa3
count_protein_coding_kallisto_therapeutic.txt


title: KO1, snRNAseq

description: 10x Genomics


title: Single oocyte (LCM) O43_S42


title: therapeutic, pGal-OT1-5

description:
pa5
count_protein_coding_kallisto_therapeutic.txt


title: Y-adaptor seq. single mESC(fresh) Ex_S05_S7

value: Fresh


title: Single oocyte (LCM) O42_S41


title: Single oocyte (LCM) O44_S43


title: SSCT F2/M  bio rep 1


title: therapeutic, OVA-OT2-7

description:
ob7
count_protein_coding_kallisto_therapeutic.txt


title: Y-adaptor seq. single mESC(fresh) Ex_S08_S9

value: exon-exon junction


title: SSCT F2/P  bio rep 1

value: epidydimal sperm


title: X10SI4 low cys

city: Utrecht

first: Saskia

middle: van

last: Mil

address: Universiteitsweg 100

department: Center for Molecular Medicine

institute: UMC Utrecht

data_processing:
Quality control was doen with FastQC (v0.11.8)
TrimGalore (v0.6.5) was used to trim reads based on quality and adapter presence after which FastQC was used to check the resulting quality.
rRNA reads were filtered out using SortMeRNA (v 4.3.3)
Reads were aligned to the reference genome using the STAR (v. 2.73a) aligner
QC on the mapped files was done using Sambamba (v0.7.0), RSeQC (v3.0.1) and PreSeq (v2.0.3)
Readcounts were generated using the Subread Featurecounts module (v2.0.0) and normalization using edgeR (v 3.28).
Assembly: Mm_GRCm38_gatk_sorted.fasta)
Supplementary files format and content: tab-delimited text file containing the raw data counts

source_name: Small intestine

extract_protocol:
RNA was isolated using Trizol and then following the protocol of the Qiagen Rneasy mini kit.
Libraries were prepared using the TruSeq stranded mRNA polyA kit

treatment_protocol: Mice were acclimatized for 1 week on AIN93G HF control diet, before starting the two week intervention on either a low cystine or normal cystine AIN93G HF diet.

value: Small intestine

value: low cystine diet


title: Single oocyte (LCM) O41_S40


title: therapeutic, OVA-OT2-6

description:
ob6
count_protein_coding_kallisto_therapeutic.txt


title: SSCT F1  bio rep 5

value: sperm


title: therapeutic, pGal-OT1-7

description:
pa7
count_protein_coding_kallisto_therapeutic.txt

value: OT-I

value: pGal


title: X26SI4 low cys


title: Single oocyte (LCM) O40_S39


title: X23SI4 low cys


title: therapeutic, OVA-OT2-5

description:
ob5
count_protein_coding_kallisto_therapeutic.txt

value: therapeutic


title: therapeutic, OVA-OT2-4

description:
ob4
count_protein_coding_kallisto_therapeutic.txt


title: Control F1  bio rep 5

value: WT


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL01

description:
SMART-Seq v4 3' DE Kit
IL01_DmGC41_50_S5

data_processing:
After trimming polyA/T sequences, adapter sequences, and poor quality reads, using fastp (v. 0.20.0) and cutadapt (ver. 1.16), the sequenced reads were aligned to the GRCm38 mouse genome assemblywith ERCC spike-in RNA sequences using the HiSAT2 (v. 2.1.0).
Best-matched reads on annotated genes were counted using HTSeq 0.10.0 with a modified GRCm38 gtf file which is extended up to 10 Kb downstream from transcription termination sites of reference genes.
Mapped reads on exons and exon-exon junctions were counted by featureCounts (v2.0.1) with the GRCm38 gtf file and the option "-J" "-p" "-M" "-O" "-t　exon" "-g　transcript_id".
Assembly: GRCm38
Supplementary files format and content: Matrix table with raw gene, exon and exon-exon junction counts for every sample.

source_name: Granulosa cell

extract_protocol:
The cDNA amplification using SMART-Seq v4 3’DE Kit was performed according manufacturer’s instruction (Takara Bio), except for the step of cell capture.  20 μL of 10×Reaction buffer was prepared (19 μL of 10×Lysis Buffer, 1 μL of RNase Inhibitor), of which 1 μL was mixed with 10.5 μL DDW, resulting in 11.5 μL of cell lysis buffer.  Then, 6.4 μL of cell lysis buffer was added the flat caps of PCR tube, and cells were isolated with LCM into the reaction buffer, followed by the brief centrifugation.  Then, another 5.1 μL of cell lysis buffer was added to the tubes (11.5 μL in total).  The following steps were performed exactly according to the manufacturer’s instruction.  Briefly, 1 μL of 12 μM Oligo dT In-line Primer was added to the mixture, and incubated at 72°C for 3 min.  Then, reverse transcription was performed by adding 7.5 μL of Master Mix (4 μL of 5x Ultra Low First-Strand Buffer, 1 μL of 48 μM SMART-Seq V4 Oligonucleotide, 0.5 μL of 40 U/μL RNase Inhibitor, 2 μL of SMARTScribe Reverse Transcriptase) and by incubating at 42°C for 90 min and 70 °C for 10 min.  Then, cDNA amplification was performed by adding 30 μL of kit component PCR Mix (25 μL of 2× SeqAmp PCR Buffer, 1μL of 12 μM Blocked PCR Primer II A, 1μL of SeqAmp DNA Polymerase, and 3μL of Nuclease free water) and applying the thermal cycling program for SMART-seq v4.
The cDNA synthesis and amplification onward were performed as described previously with minor modifications [Kurimoto, K.et.al., Nucleic Acids Research, 34(5), e42.]. In brief, the first-strand cDNA was synthesized by SuperScript III [Thermo Fisher] and incubating at 50℃  for 5 min followed by heat inactivation at 70℃  for 10 min.  The excess RT primers were degraded by exonuclease I and incubating at 37℃ for 30 min followed by heat inactivation at 80℃ for 25 min. The first-strand cDNA product was poly dA-tailed by TdT and incubating at 37℃  for 1 min followed by heat inactivation at 70℃ for 10 min. Second-strand cDNA synthesis were performed by adding 9.5 μL reaction mixture of ExTaq, Hot Start version [Takara Bio] and V3[dT]24 primer in the ExTaq buffer with the following thermal cycling program (95℃  for 3 min, 50℃  for 2 min, and 72℃  for 3 min). cDNA amplification was performed by addingthe mixture of V1[dT]24 and ExTaq, Hot Start version [Takara Bio] in the ExTaq buffer with the following thermal cycling program (95℃  for 1 min; 20 cycles of 95℃  for 30 sec, 67℃  for 1 min and 72℃  for 3 min with a 6 sec elongation per cycle; and 72℃  for 10 min).  The amplified cDNA was purified with a 0.6× volume of AxyPrep MAG PCR clean-up reagent (Axygen) according to the manufacturer's instructions. Preparation of the libraries for SC3-seq was performed essentially as previously described (Nakamura et al., 2015)(Nakamura et al., 2017).  For Y-adapter sequencing, single-cell cDNAs were pre-amplified using the primers (V3[dT]24, V1[dT]24) and ExTaq HS with the thermal cycling program (95℃  for 3 min; 4 cycles of 95°C for 30 sec, 67℃  for 1 min and 72℃  for 5 min; 72℃  for 10 min). Pre-amplified cDNAs were then subjected to the primer-dimer removal, sonication, end polishing, and the size selection as in SC3-seq. Then, purified cDNA was dA-tailed in a 10 μL dA tailing mixture with incubation at 72℃  for 10 min. The dA-tailed product was subjected to the Y-shaped adaptor ligation by adding a mixture of Y-adaptor and NEB Next Quick Ligation Reaction Buffer and incubating at 20℃  for 15 min and at 72℃  for 20 min. The ligation product was purified with a 0.9×volume of AxyPrep reagent, and subjected to additional PCR amplification in 20 μL with the thermal cycling program (95℃  for 3 min, 55℃  for 1 min, and 72℃  for 1 min; 11 cycles of 95℃  for 30 sec, 60℃  for 1 min, and 72℃  for 1 min; and 72℃  for 3 min), followed by DNA purification with a 0.9×volume of AxyPrep reagent.

value: LCM

value: Neighboring

value: gene expression


title: therapeutic, OVA-OT2-3

description:
ob3
count_protein_coding_kallisto_therapeutic.txt

source_name: CD4 T cells


title: SSCT F1 bio rep 4

value: 3 months

value: male

value: SSCT


title: SSCT F1 bio rep 3


title: Single oocyte (LCM) O39_S38

value: Primary

value: gene expression (sc3-seq.)


title: therapeutic, OVA-OT2-2

description:
ob2
count_protein_coding_kallisto_therapeutic.txt

value: OT-II

value: OVA


title: SSCT F2/M  bio rep 2

value: epidydimal sperm


title: SI9 normal cys

value: Small intestine

value: normal cystine diet


title: Control F1 biol rep 4

value: sperm


title: SSCT F2/P bio rep 5


title: therapeutic, OVA-OT2-1

description:
ob1
count_protein_coding_kallisto_therapeutic.txt


title: therapeutic, OVA-OT1-9

description:
oa9
count_protein_coding_kallisto_therapeutic.txt

value: OT-I


title: X32SI4 normal cys


title: Single oocyte (LCM) O37_S36


title: X17SI4 normal cys


title: Single oocyte (LCM) O36_S35


title: Single oocyte (LCM) O35_S34


title: Control F1 biol rep 3


title: X1SI4 normal cys


title: Single oocyte (LCM) O34_S33


title: therapeutic, OVA-OT1-7

description:
oa7
count_protein_coding_kallisto_therapeutic.txt

value: therapeutic


title: Control F1 biol rep 2

value: WT


title: therapeutic, OVA-OT1-8

description:
oa8
count_protein_coding_kallisto_therapeutic.txt


title: Single oocyte (LCM) O33_S32

value: LCM


title: WJL2346_RLF_N023

city: Stanford

first: Jinglong

last: Wang

state: CA California

address: 1291 Welch Rd, CCSR Room 1240

department: Radiation Oncology

institute: Stanford University

data_processing:
Data processing is described in Hu et al., 2016 Nature Protocols (PMC4895203) and Liang et al., 2021 PNAS (PMC8166026).
Standard basecalling formats for Illumina reads
Illumina reads were de-multiplexed and adapter sequence trimmed using the fastq-multx tool from ea-utils (https://expressionanalysis.github.io/ea-utils/) and the SeqPrep utility (https://github.com/jstjohn/SeqPrep) respectively.
Reads were mapped to the mm9 reference genome using Bowtie2 (http://bowtiebio.sourceforge.net/bowtie2/manual.shtml) to map translocations
Reads were aligned to the bait and the prey region using Bowtie2.
Assembly: The cross library contamination in the co-tagged PCR was filtered by the inner barcodes from I7-nested primers.
Supplementary files format and content: mm9
Supplementary files format and content: tab delimited text files contain filtered unique junctions and include the following information: sequence ID (Qname); prey chromosome (Rname), prey junction coordinate (Junction), chromosome orientation of prey junction (Strand), beginning (Rstart) and end (Rend) nucleotide position of prey junction aligning to the genome build; bait chromosome (B_Rname); beginning (B_Rstart) and end (B_Rend) nucleotide position of the bait junction; chromosome orientation of the bait junction (B_Strand); position on the read where the bait sequence begins (B_Qstart) and ends (B_Qend); position on the read where the prey junction begins (Qstart) and ends (Qend); the length of the combined and stitched paried end read (Qlen); and the entire stitched paired end read sequence (Seq)
Library strategy: LAM-HTGTS

source_name: vAbl-ProB

growth_protocol: vAbl ProB cells were cultured at 37 °C and 5% CO2 in RPMI supplemented with 10% (vol/vol) FBS, 50 U/mL penicillin/streptomycin, 2 mM L-glutamine , 1× MEM-NEAA, 1 mM sodium pyruvate , 50 μM 2-mercap- toethanol and 20 mM Hepes (pH 7.4).

extract_protocol:
Genomic DNA was isolated using protocol described in Hu et al., 2016 Nature Protocols (PMC4895203)
Libraries were generated as described in Hu et al., 2016 Nature Protocols (PMC4895203), with the enzyme blocking step being omitted, using primers associated with the target locus and Illumina NovoSeq  sequencing

treatment_protocol: G1-arrested cells were treated with STI-571 (3µM) for 96h. DNA-PKcs inhitors, DPKi #1 (Nu7441, 0.5 µM) and DPKi #2 (Nu7026, 20 µM)were added to some of the samples the same day as STI-571.

value: vAbl-ProB

value: pro-B cells


title: WJL2345_RLF_N023


title: WJL2347_RLF_N023


title: therapeutic, pGal-OT1-1

description:
pa1
count_protein_coding_kallisto_therapeutic.txt

extract_protocol:
Spleens and lymph nodes (inguinal, axial, brachial, popliteal, cervical) were pooled. OT-I and OT-II were isolated separately using fluorescence activated cell sorting (FACS) into Trizol (prophylactic) or FACS buffer, spun, decanted, and snap frozen (therapeutic) and frozen at -80. RNA was extracted (Qiagen). For prophylactic three mice were pooled per sample, for therapeutic one mouse per sample.
Pooled samples were converted to cDNA using SmartSeq v4 protocol. Libraries were constructed using iDT Nextera, paired end 150.

value: pGal


title: therapeutic, OVA-OT1-6

description:
oa6
count_protein_coding_kallisto_therapeutic.txt


title: WJL2344_RLF_N023


title: Control F1 bio rep 1

value: 3 months

value: male


title: therapeutic, OVA-OT1-5

description:
oa5
count_protein_coding_kallisto_therapeutic.txt


title: Single oocyte (LCM) O32_S31

value: Secondary-antral


title: therapeutic, OVA-OT1-4

description:
oa4
count_protein_coding_kallisto_therapeutic.txt


title: WJL2342_RLF_N023


title: ESM1-WT-tumor 3

first: Riki

last: Kawaguchi

address: 760 Westwood Plaza, Room 37-420

department: Department of Neurology and Department of Psychiatry

institute: University of California Los Angeles

data_processing:
Basecalls/demultiplexing performed using Illumina CASAVA 1.8.2
Aligned to mm10 using STAR2.4.0
Count read using HT-seq0.6.1 using mm10 RefSeq as reference
Normalization (TMM), quantitation (FPKM and adjFPKM) and differential expression analysis using EdgeR.
Genome_build: mm10
Supplementary_files_format_and_content: xlsx

source_name: Tumor tissue

extract_protocol:
Total RNA was extracted from tissues using the TRIzol Plus RNA Purification System ( Invitrogen).
NEBNext Ultra RNA library preparation kit was used for preparation.

treatment_protocol: All animal experiments were performed at the UAB under an Institutional Animal Care and Use Committee (IACUC)-approved protocol according to NIH guidelines (#20295).

tag: genotype/variation

tag: rna sample type

value: WT group


title: ESM1-WT-tumor 2


title: WJL2343_RLF_N023


title: Single oocyte (LCM) O31_S30

tag: collecting conditions

value: gene expression (sc3-seq.)


title: therapeutic, OVA-OT1-2

description:
oa2
count_protein_coding_kallisto_therapeutic.txt

data_processing:
Fastq files were generated using bcl2fastq
kallisto (v0.46.1) was used to quantify abundances of transcripts from the fastq files.
Count tables were generated using R package tximport to import transcript-level estimates from kallisto output and summarize them at gene level
Genome_build: GRCm38.97
Supplementary_files_format_and_content: tab-delimited text file includes matrix table with raw gene count values for each sample

value: OVA


title: WJL2340_RLF_N023


title: Single oocyte (LCM) O30_S29

tag: class


title: ESM1-WT-tumor 1


title: SSCT F1 biol rep 1

value: epidydimal sperm

value: SSCT


title: WJL2341_RLF_N023


title: Single oocyte (LCM) O28_S27

tag: analysis target


title: SSCT F1 biol rep 2

tag: cell type


title: ESM1-KO tumor 2

tag: strain

value: ESM1 KO group


title: therapeutic, OVA-OT1-10

description:
oa10
count_protein_coding_kallisto_therapeutic.txt

tag: group


title: Single oocyte (LCM) O29_S28

value: Primary


title: Single oocyte (LCM) O27_S26


title: WJL2339_RLF_N023


title: therapeutic, OVA-OT1-1

description:
oa1
count_protein_coding_kallisto_therapeutic.txt

value: OT-I


title: ESM1-KO tumor 1


title: Single oocyte (LCM) O26_S25


title: WJL2337_RLF_N023

value: WT + DPKi #2


title: prophylactic batch 2, OT2-pGal-2-batch2

description:
S14
count_protein_coding_kallisto_batch2.txt

value: OT-II

value: prophylactic batch 2


title: Single oocyte (LCM) O25_S24

value: LCM


title: WJL2338_RLF_N023

value: vAbl-ProB

value: pro-B cells

value: WT + DPKi #1


title: Single oocyte (LCM) O38_S37


title: WJL2335_RLF_N023


title: prophylactic batch 2, OT2-pGal-1-batch2

description:
S13
count_protein_coding_kallisto_batch2.txt

value: pGal


title: ESM1-KO tumor 3


title: prophylactic batch 2, OT2-OVA-2-batch2

description:
S12
count_protein_coding_kallisto_batch2.txt


title: prophylactic batch 2, OT2-OVA-1-batch2

description:
S11
count_protein_coding_kallisto_batch2.txt


title: Single oocyte (LCM) O24_S23

value: Secondary-antral


title: Single oocyte (LCM) O23_S22


title: prophylactic batch 2, OT2-Saline-2-batch2

description:
S10
count_protein_coding_kallisto_batch2.txt

value: Saline


title: prophylactic batch 2, OT2-Saline-1-batch2

description:
S09
count_protein_coding_kallisto_batch2.txt


title: Single oocyte (LCM) O22_S21

value: gene expression (sc3-seq.)


title: WJL2333_RLF_N023


title: prophylactic batch 2, OT1-pGal-2-batch2

description:
S06
count_protein_coding_kallisto_batch2.txt


title: Single oocyte (LCM) O20_S19


title: Single oocyte (LCM) O21_S20


title: PBMC_normal_nonfasting_SCR [95]

type: RNA

city: Oslo

first: Kirsten

last: Holven

address: Sognsvannsveien 9

department: Department of Nutrition

country: Norway

institute: University of Oslo

description: Gene expression data from PBMCs isolated from non-fasting blood samples collected at a screening visit for a randomized trial

hyb_protocol: Gene expression measurements were performed by hybridizing the amplified RNA to Illumina HumanHT-12 v4 Expression BeadChip (Illumina Inc., California, USA) according to the manufacturer’s instructions.

scan_protocol: Samples were scanned using Illumina HiScan System (Illumina Inc., California, USA).

data_processing:
The Illumina HumanHT-12 v4 Expression BeadChip provides genome-wide measurements of the expression of over 48,000 probe sets. IlluminaGenome Studio was used to calculate and report a detection P value, which represents the confidence that a given transcript is expressed above the background. A gene was defined as expressed when relevant probes with a P value below 0.01 were found in more than five samples. After hybridization and scanning, a manual quality control was performed investigating density plots and hierarchical clustering of raw probe densities. One probe per gene (max IQR) was selected for further analysis.
After correcting for background noise, using normexp background correction (neqc filtration, Limma), quantile normalization of the data was performed using the Illumina GenomeStudio software, version 1.7.0. Data were log2-transformed and exported to R [R Core Team (2023). R: A language and environment for statistical computing. R Foundation for Statistical Computing, Vienna, Austria. URL https://www.R-project.org/] for biostatistical analysis.
Analyses in Bålsrud et al. (submitted):
For the PBMC gene expression analyses, in an initial untargeted approach, we fitted standard linear regression models for each of the 11925 mRNA transcripts with frailty index (FI) score as the main exposure. The main model was adjusted for age and BMI; we visualized the regression coefficients and P values in a Volcano plot. Two other models were fitted: a crude, unadjusted model, and a model adjusted for age, BMI, and proportion of monocytes and lymphocytes in blood (not shown). In a more targeted approach, we sought to explore cell type-specific genes. To derive an estimate of cell type proportions in the PBMC pool, we performed gene expression decomposition (in silico flow cytometry) using CIBERSORT [Newman AM, Steen CB, Liu CL, Gentles AJ, Chaudhuri AA, Scherer F, et al. Determining cell type abundance and expression from bulk tissues with digital cytometry. Nature Biotechnology. 2019;37(7):773-82]. We fitted regression models for each CIBERSORT-predicted cell type with FI score as the main exposure (both crude models and models adjusted for age and BMI). We then extracted the top 30 genes/mRNA transcripts associated with CIBERSORT-predicted monocytes and associated them with the six serum inflammatory biomarkers in linear regression models adjusting for age and BMI. The resulting β coefficient matrix was visualized as a heatmap (ComplexHeatmap R package).

label: Biotin

source_name: PBMC, non-fasting blood sample, screening visit

label_protocol: Total RNA preparation (incl. biotin labelling) and amplification were performed using the Illumina Total Prep RNA Amplification Kit (Illumina Inc., California, USA).

extract_protocol: Blood samples were collected in non-fasting state in BD Vacutainer® CPTTM cell preparation tubes with sodium heparin (Becton Deckenson, NJ, USA). Within two hours of blood collection PBMCs were collected by density gradient centrifugation of the blood samples (1636g) for 25 min at room temperature. The cells were washed twice (300g, 10 min at RT) in phosphate-buffered saline (PBS) without calcium chloride and magnesium chloride. After the last washing step, excess PBS was discarded. The pellet was dissolved in the remaining liquid and transferred to an Eppendorf tube, centrifuged (13,000g, 3 min at 4 °C) and frozen at −80 °C until total RNA was extracted using RNeasy Mini Kit (Qiagen). RNA quantity was measured using NanoDrop-1000 (NanoDrop Technologies, Inc., Delaware, USA), while RNA quality was checked with Aglient 2100 Bioanalyzer (Agilent Technologies, Inc., California, USA).

tag: fi score (range 0-1)

tag: sex (female/male)

value: Female

tag: age (y)

tag: bmi (kg(m2)

tag: monocytes (absolute values x109/l)

tag: lymphocytes (absolute values x109/l)


title: WJL2332_RLF_N023

tag: cell line


title: PBMC_normal_nonfasting_SCR [93]


title: prophylactic batch 2, OT1-pGal-1-batch2

description:
S05
count_protein_coding_kallisto_batch2.txt


title: WJL2331_RLF_N023

tag: genotype


title: PBMC_normal_nonfasting_SCR [94]


title: WJL2336_RLF_N023

value: WT

tag: treatment


title: Single oocyte (LCM) O19_S18


title: WJL2330_RLF_N023


title: WJL2329_RLF_N023


title: prophylactic batch 2, OT1-OVA-2-batch2

description:
S04
count_protein_coding_kallisto_batch2.txt

value: OVA


title: WJL2334_RLF_N023


title: PBMC_normal_nonfasting_SCR [90]


title: prophylactic batch 2, OT1-OVA-1-batch2

description:
S03
count_protein_coding_kallisto_batch2.txt


title: PBMC_normal_nonfasting_SCR [89]


title: PBMC_normal_nonfasting_SCR [91]


title: PBMC_normal_nonfasting_SCR [92]


title: Single oocyte (LCM) O17_S16


title: Single oocyte (LCM) O16_S15


title: Single oocyte (LCM) O18_S17


title: WJL2327_RLF_N023


title: PBMC_normal_nonfasting_SCR [88]


title: prophylactic batch 2, OT1-Saline-1-batch2

description:
S01
count_protein_coding_kallisto_batch2.txt


title: prophylactic batch 2, OT1-Saline-2-batch2

description:
S02
count_protein_coding_kallisto_batch2.txt


title: WJL2325_RLF_N023


title: WJL2326_RLF_N023


title: Single oocyte (LCM) O14_S13


title: prophylactic batch 1, OT2-OVA5-2

description:
S32
count_protein_coding_kallisto_batch1.txt

value: prophylactic batch 1


title: PBMC_normal_nonfasting_SCR [86]


title: PBMC_normal_nonfasting_SCR [85]


title: Single oocyte (LCM) O12_S12


title: prophylactic batch 1, OT2-OVA5-3

description:
S33
count_protein_coding_kallisto_batch1.txt


title: WJL2324_RLF_N023


title: Single oocyte (LCM) O11_S11


title: PBMC_normal_nonfasting_SCR [84]


title: prophylactic batch 1, OT2-saline-3

description:
S27
count_protein_coding_kallisto_batch1.txt


title: Single oocyte (LCM) O10_S10


title: WJL2323_RLF_N023


title: Single oocyte (LCM) O15_S14


title: prophylactic batch 1, OT2-saline-2

description:
S26
count_protein_coding_kallisto_batch1.txt


title: WJL2328_RLF_N023


title: PBMC_normal_nonfasting_SCR [83]


title: prophylactic batch 1, OT2-saline-1

description:
S25
count_protein_coding_kallisto_batch1.txt


title: PBMC_normal_nonfasting_SCR [82]


title: WJL2321_RLF_N023


title: Single oocyte (LCM) O08_S8


title: PBMC_normal_nonfasting_SCR [81]


title: prophylactic batch 1, OT2-pGal-3

description:
S21
count_protein_coding_kallisto_batch1.txt


title: Single oocyte (LCM) O07_S7


title: Single oocyte (LCM) O06_S6


title: WJL2319_RLF_N023


title: prophylactic batch 1, OT2-pGal-2

description:
S20
count_protein_coding_kallisto_batch1.txt


title: PBMC_normal_nonfasting_SCR [79]


title: prophylactic batch 1, OT2-pGal-1

description:
S19
count_protein_coding_kallisto_batch1.txt


title: Single oocyte (LCM) O05_S5


title: prophylactic batch 1, OT1-OVA5-3

description:
S15
count_protein_coding_kallisto_batch1.txt


title: Single oocyte (LCM) O03_S3


title: PBMC_normal_nonfasting_SCR [78]


title: Single oocyte (LCM) O04_S4


title: prophylactic batch 1, OT1-OVA5-2

description:
S14
count_protein_coding_kallisto_batch1.txt


title: WJL2317_RLF_N023


title: Single oocyte (LCM) O09_S9


title: WJL2316_RLF_N023


title: prophylactic batch 1, OT1-OVA5-1

description:
S13
count_protein_coding_kallisto_batch1.txt


title: PBMC_normal_nonfasting_SCR [87]

description: Gene expression data from PBMCs isolated from non-fasting blood samples collected at a screening visit for a randomized trial


title: PBMC_normal_nonfasting_SCR [77]


title: WJL2315_RLF_N023


title: Single oocyte (LCM) O02_S2


title: PBMC_normal_nonfasting_SCR [80]


title: WJL2318_RLF_N023


title: Single oocyte (LCM) O01_S1


title: PBMC_normal_nonfasting_SCR [74]


title: prophylactic batch 1, OT1-saline-2

description:
S08
count_protein_coding_kallisto_batch1.txt


title: PBMC_normal_nonfasting_SCR [75]


title: WJL2314_RLF_N023


title: WJL2313_RLF_N023


title: Single granulosa cell (LCM) GC100_S63

value: Neighboring


title: prophylactic batch 1, OT1-saline-3

description:
S09
count_protein_coding_kallisto_batch1.txt

value: OT-I


title: PBMC_normal_nonfasting_SCR [72]


title: prophylactic batch 1, OT1-pGal-3

description:
S03
count_protein_coding_kallisto_batch1.txt


title: Single granulosa cell (LCM) GC097_S62

value: Non-neighboring


title: WJL2311_RLF_N023


title: WJL2312_RLF_N023


title: prophylactic batch 1, OT1-saline-1

description:
S07
count_protein_coding_kallisto_batch1.txt


title: Single granulosa cell (LCM) GC093_S60


title: prophylactic batch 1, OT1-pGal-2

description:
S02
count_protein_coding_kallisto_batch1.txt


title: PBMC_normal_nonfasting_SCR [71]


title: PBMC_normal_nonfasting_SCR [70]


title: WJL2310_RLF_N023


title: PBMC_normal_nonfasting_SCR [73]


title: Single granulosa cell (LCM) GC091_S59


title: WJL2322_RLF_N023


title: WJL2308_RLF_N023


title: prophylactic batch 1, OT1-pGal-1

description:
S01
count_protein_coding_kallisto_batch1.txt


title: PBMC_normal_nonfasting_SCR [69]


title: WJL2309_RLF_N023


title: Postnatal day 7 myeloid cells

city: Edinburgh

first: Nizar

middle: N.

last: Batada

address: Crew Road

institute: Institute for Genetics and Molecular Medicine, University of Edinburgh

data_processing:
demultiplexing and alignment with CellRanger
Genome_build: mm10

source_name: Myeloid cells

growth_protocol: Samples were collected at postnatal day 0, 7, or 21

extract_protocol: Brains were extracted and placed on ice in HBSS with HEPES buffer. Corpus callosi were dissected out and homogenized in glass dounces and filtered. Following centrifugation, cells were resuspended in 10% fetal bovine serum and run through a Percoll gradient. Cells were collected in FACS buffer, spun, and resuspended in HBSS. Myeloid cells were sorted based on expression of Cd11b.

treatment_protocol: Mice were anesthetized and perfused with phosphate buffered saline

tag: age

value: postnatal day 7

tag: brain region

value: corpus callosum


title: therapeutic, OVA-OT1-3

description:
oa3
count_protein_coding_kallisto_therapeutic.txt

value: therapeutic


title: Single granulosa cell (LCM) GC090_S58

last: Ikeda

address: 840 Shijo-Cho, Kashihara, Nara

institute: Nara Medical University

tag: collecting conditions

value: LCM


title: Single granulosa cell (LCM) GC089_S57

tag: class


title: WJL2307_RLF_N023

value: vAbl-ProB

value: pro-B cells


title: Postnatal day 21 myeloid cells

value: postnatal day 21


title: Single granulosa cell (LCM) GC088_S56

tag: analysis target


title: Postnatal day 0 myeloid cells

value: postnatal day 0


title: PBMC_normal_nonfasting_SCR [76]


title: WJL2306_RLF_N023

tag: cell type


title: Single granulosa cell (LCM) GC087_S55


title: WJL2305_RLF_N023


title: PBMC_normal_nonfasting_SCR [66]


title: PBMC_normal_nonfasting_SCR [68]


title: PBMC_normal_nonfasting_SCR [67]


title: Con3_170

city: Irvine

first: Devon

last: Lawson

address: 839 Health Science Road, Sprague Hall 140

institute: University of California, Irvine

description:
Single-cell RNA-Seq [10X Genomics]
PolyA RNA

data_processing:
Sequencing reads were aligned utilizing 10x Genomics Cell Ranger Count 3.0.2 to the dual indexed mm10/GRCh38 genome provided by Cell Ranger.
Cells in dataset were subsequently trimmed and analyzed in Seurat v3, with assigned cell type labels and quality control metrics for each cell passing initial quality filtering included in the metadata sheets.
Genome_build: mm10/GRCh38
Supplementary_files_format_and_content: Space-delimited text files contain a matrix of counts; rows are genes and columns are samples.
Supplementary_files_format_and_content: TSV/MTX sets are output from the Cell Ranger pipeline and can be used to generate a matrix using Seurat's Read10X function.
Supplementary_files_format_and_content: Metadata sheets include quality control and cell type label information for the cells analyzed in the associated study.

source_name: Control brain

extract_protocol:
Single cell suspension from mouse brain was prepared using the Adult Brain Dissociation Kit, Mouse and Rat (Miltenyi Biotec) with some modifications. Briefly, whole dissected brains were chopped into 8 pieces of equal size and placed into C tube (Miltenyi Biotec) containing enzyme P and A. Brain tissue was digested using gentleMACS Octo Dissociator with heaters operating the Adult brain dissociation protocol (Miltenyi Biotec). After digestion, the cell suspension was strained over a sterile 70μm strainer (Fisher Scientific) and washed with 5mL FACS buffer containing ice cold DMEM/F12, 50mM HEPES, and 2% BSA. After removal of myelin by density centrifugation, the cell pellet was washed and remaining red blood cells were lysed with red blood cell lysis buffer. Cells were then re-suspended in FACS buffer and blocked with anti-CD16/32 for 15 minutes on ice. Next, cells were stained with fluorescent antibodies on ice for 15 minutes shielded from light. The labeled cells were washed with 500μL of FACS buffer and resuspended in 500μL of FACS buffer, strained through 40μm strainer prior to sorting on BD FACSAria Fusion sorter. For sorting of microglia, astrocytes, and cancer cells, cells were gated for size based on forward and side scatter, single cells, and Sytox Blue viability (Thermofisher, S34857). All myeloid cells (CD45+ CD11b+) and astrocytes (CD45-, ACSA2+) were sorted from control and metastatic mouse brains into 500μL of chilled FACS buffer. GFP+ 231BR cells were sorted from metastatic brains into 500μL of FACS buffer.
FACS isolated mouse microglia cells were centrifuged for 10 minutes at 300g and washed with 0.04% BSA in PBS. Cells were resuspended to achieve approximately 1,000 cells/µL. Final cell suspensions were counted on the Countess II automated cell counter to determine actual concentration for droplet generation. Cells were loaded onto the 10x Genomics Chromium Single Cell Gene Expression 3’ v2 Chemistry kits for GEMs generation. Following the Chromium Single Cell 3′ Reagents Kits version 2 user guide (CG00052 Rev B), cells were loaded to achieve approximately 10,000 cells for capture. Libraries were sequenced on the Illumina HiSeq 4000 platform to achieve an average of read depth of 50,000 mean reads per cell.

tag: mouse host genotype

value: Foxn1 nu/nu

tag: mouse diagnosis

value: Control

tag: tissue

value: Brain

value: Astrocytes (CD45-, ACSA2+) and myeloid cells (CD45+ CD11b+)

tag: batch

value: Group 2


title: WJL2303_RLF_N023


title: Single granulosa cell (LCM) GC083_S52

first: Hiroki

value: gene expression (sc3-seq.)


title: Single granulosa cell (LCM) GC080_S50

value: Neighboring


title: WJL2304_RLF_N023


title: Con1_131

value: Group 1


title: PBMC_normal_nonfasting_SCR [63]

tag: fi score (range 0-1)

tag: sex (female/male)

value: Female

tag: age (y)


title: Single granulosa cell (LCM) GC078_S49


title: PBMC_normal_nonfasting_SCR [62]

tag: bmi (kg(m2)

tag: monocytes (absolute values x109/l)

tag: lymphocytes (absolute values x109/l)


title: PBMC_normal_nonfasting_SCR [64]


title: WJL2301_RLF_N023

tag: cell line

tag: genotype

value: WT + DPKi #2

tag: treatment


title: Single granulosa cell (LCM) GC084_S53

tag: collecting conditions


title: Single granulosa cell (LCM) GC075_S48

tag: class

value: Non-neighboring


title: PBMC_normal_nonfasting_SCR [65]


title: Con2_169


title: ESM1-KO tumor 2

data_processing:
Alignment was carried out using the Burrows-Wheeler Aligner mem
Aligned to mm10 reference genome
peak calling using MACS2 (with parameter setting-nomodel-shift 75)
GO enrichment analysis and peak annotation using HOMER
Genome_build: mm10

extract_protocol:
Sequencing was carried out by the UCLA ICNN at UCLA. Samples were multiplexed in 1 lanes.
PE 2x75 using Illumina HiSeq4000

tag: strain

tag: genotype/variation

tag: rna sample type

value: ESM1 KO group


title: ESM1-KO tumor 1


title: WJL2300_RLF_N023

tag: cell type

value: WT


title: Single granulosa cell (LCM) GC074_S47

value: LCM

tag: analysis target


title: WJL2320_RLF_N023

value: vAbl-ProB

value: pro-B cells


title: ESM1-WT tumor 1

value: WT group


title: Single granulosa cell (LCM) GC073_S46


title: Single granulosa cell (LCM) GC069_S116


title: WJL2298_RLF_N023


title: WJL2302_RLF_N023

value: WT + DPKi #1