title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL10

description:
SMART-Seq v4 3' DE Kit
IL10_DmOo11_20_S2

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: Primary follicle


title: SNU449_wt_plusRego_r3

status: Public on Sep 01 2023

type: SRA

city: Vienna

first: Wolfgang

last: Mikulits

address: Borschkegasse 8A

country: Austria

institute: Medical University of Vienna

description:
Axl_Rego_Quantseq_star253a_GRCh38v40.SS2_DESeq2-220726_AxlWT.minusRego_VS_AxlWT.plusRego_Prep.Date_DEtable.txt
Axl_Rego_Quantseq_star253a_GRCh38v40.SS2_DESeq2-220726_AxlWT.plusRego_VS_AxlKO.plusRego_Prep.Date_DEtable.txt

library_source: transcriptomic

data_processing:
Computational trimming of poly-As from the 3’-ends of the 100 bp long reads was performed with cutadapt (v. 4.0; with options -O 3 -e 0.05 -m 20 --max-n 2), retaining reads with a trimmed length of at least 20 nt.
Reads were aligned to the human genome GRCh38.v40 and the spike-in sequences (Lexogen “SIRV-Set3”) by STAR 2.5.3a (27), with up to 1000 hits, a maximum error rate of 5%, and counts per gene as output.
These count tables were generated on the basis of the GENCODE annotation v40 concatenated with the SIRV and ERCC annotation.
Assembly: human genome GRCh38.v40
Supplementary files format and content: contain information about: "gene_id" "baseMean" "log2FoldChange" "lfcSE" "stat" "pvalue" "padj" "absLog2FC" "SNU449_AxlKO_parental_r1" "SNU449_AxlKO_parental_r2" "SNU449_AxlKO_parental_r3" "SNU449_AxlKO_plusRego_r1" "SNU449_AxlKO_plusRego_r2" "SNU449_AxlKO_plusRego_r3" "AxlKO.minusRego.AVG" "AxlKO.plusRego.AVG" "chrom_name" "source" "type" "start" "end" "score" "strand"

molecule: total RNA

organism: Homo sapiens

source_name: liver

growth_protocol: SNU449 and derived HCC cells were maintained in RPMI supplemented with 10% fetal bovine serum (FBS) and antibiotics in humidified atmosphere with 5% CO2 at 37C. SNU449 and derived HCC cells were additionally treated with Regorafenib every time after splitting.

extract_protocol:
Total RNA was isolated using the Monarch total RNA miniprep kit (New England Biolabs, Ipswich, Massachusetts, USA).
Library preparation and sequencing was performed by Lexogen (Greenland, New Hampshire, USA). In brief, quality controlled and quantified RNA was used to prepare a QuantSeq 3’ mRNA-seq (fwd) library suitable for Illumina sequencing (12 cycles of library amplification, paired-end 100 nt sequencing).

treatment_protocol: To generate resistant cell lines, SNU449 and SNU449-Axl-KO were initially treated with 50% of the IC50 concentration of Rego (TargetMol Chemicals, Boston, USA). The concentrations was increased by 50% of the IC50 every two weeks until 2 x IC50 was reached and cells proliferated under high concentrations

tag: tissue

value: liver

tag: cell line

tag: cell type

value: Hepatocellular carcinoma cell line

tag: genotype

value: WT

tag: treatment

value: Regorafenib long-term


title: ATAC, KO, 1

city: Hangzhou

first: Heping

last: Xu

state: Zhejiang

address: 18 Shilongshan Road

country: China

institute: Westlake University

description:
Assay for Transposase-Accessible Chromatin
DNA-seq

library_source: genomic

data_processing:
For RNA-seq: raw fastq files were trimmed with cutadapt v2.10. Alignment to mm10 genome/transcriptome and quantification of gene expression were done using STAR v2.7.5c and RSEM v1.3.3.
For ATAC-seq/CUTTag-seq: raw fastq files were trimmed with cutadapt v2.10. Alignment to mm10 genome and advanced filtering of bad-quality, blacklist reads and duplicates were done using bowtie2(v2.2.5), samtools(v1.10), bedtools(v2.29.2) and picard(v2.23.8).
Assembly: mm10
Supplementary files format and content: TPM expression matrix files were extracted from RSEM gene-results and initially filtered for expressed protein-coding genes.
Supplementary files format and content: bigwig files contain genome-wide signal which were generated from clean bam files using deeptools(v3.5.0) 'bamCoverage -e250 -bs100 --normalizedUsing RPKM'.

molecule: genomic DNA

organism: Mus musculus

source_name: Small Intestine

growth_protocol:
primary small intestine ILC2s or small intestinal ILC2s were cultured in vitro with IL-2 and IL-7, or IL-2, IL-7 and IL-25, for 4, 48 and 72 hours
small intestinal ILC2s were cultured with IL-2 and IL-7, or IL-2, IL-7 and IL-25, for 4, 48 and 72 hours

extract_protocol:
DNA and RNA were extracted from cells using AMPure XP beads (Backman)
Full-length Smart-seq2, ATAC-seq and CUT&Tag-seq
RNA-seq and DNA-seq

treatment_protocol: The mice were at the steady state or treated with IL-25 or PBS for 2 days

value: Small Intestine

tag: strain

tag: age

value: 8-12 weeks

tag: Sex

value: pooled male and female

value: KO

value: primary small intestine ILC2s

value: steady state

tag: treatment time


title: iILC2, KO, 4

description:
full-length gene expression
Smart-seq2
iKO_4
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: iILC2, KO, 1

description:
full-length gene expression
Smart-seq2
iKO_1
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo11_20_S8_IL09

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
IL09_DpOo11_20_S8

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: Oocyte

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

tag: analysis target

value: gene expression


title: SNU449_wt_plusRego_r1


title: iILC2, CTRL, 4

description:
full-length gene expression
Smart-seq2
iCL_4
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: cuttag, H3K27ac, KO, 2

description:
CUT&Tag to identify DNA binding sites
DNA-seq

data_processing:
For RNA-seq: raw fastq files were trimmed with cutadapt v2.10. Alignment to mm10 genome/transcriptome and quantification of gene expression were done using STAR v2.7.5c and RSEM v1.3.3.
For ATAC-seq/CUTTag-seq: raw fastq files were trimmed with cutadapt v2.10. Alignment to mm10 genome and advanced filtering of bad-quality, blacklist reads and duplicates were done using bowtie2(v2.2.5), samtools(v1.10), bedtools(v2.29.2) and picard(v2.23.8).
Assembly: mm10
Supplementary files format and content: TPM expression matrix files were extracted from RSEM gene-results and initially filtered for expressed protein-coding genes.
Supplementary files format and content: bigwig files contain genome-wide signal which were generated from clean bam files using deeptools(v3.5.0) 'bamCoverage -e250 -bs100 --normalizedUsing RPKM'.
Library strategy: CUT&Tag-seq

tag: antibody

value: Acetyl-Histone H3 (Lys27) Antibody (Cell Signaling Technology, Cat# 4353S); Goat Anti-Rabbit IgG Antibody (H&L) [HRP], pAb (Genscript, Cat# A00098)


title: SNU449_wt_plusRego_r2


title: iILC2, CTRL, 3

description:
full-length gene expression
Smart-seq2
iCL_3
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL09

description:
SMART-Seq v4 3' DE Kit
IL09_DmOo01_10_S1

value: Secondary-antral


title: cuttag, H3K27ac, KO, 1


title: iILC2, CTRL, 2

description:
full-length gene expression
Smart-seq2
iCL_2
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo11_20_S8_IL10

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL10_DpOo11_20_S8

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. Formalin fixation was performed with 10% formalin neutral buffer solution (Wako) at room temperature for 24 h, and the fixed sections were stained with 1% cresyl violet acetate in 50% isopropanol and dried for more than 1 h. Cells in the sections were isolated using a laser microdissection system PALM MB4 (Zeiss).


title: SNU449_AxlKO_plusRego_r2

description:
Axl_Rego_Quantseq_star253a_GRCh38v40.SS2_DESeq2-220726_AxlKO.minusRego_VS_AxlKO.plusRego_Prep.Date_DEtable.txt
Axl_Rego_Quantseq_star253a_GRCh38v40.SS2_DESeq2-220726_minusRego_VS_plusRego_Prep.Date_DEtable.txt

value: Axl KO


title: nILC2, KO, 4

description:
full-length gene expression
Smart-seq2
nKO_4
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: iILC2, CTRL, 1

description:
full-length gene expression
Smart-seq2
iCL_1
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL10

description:
SMART-Seq v4 3' DE Kit
IL10_DmOo01_10_S1


title: iILC2, KO, 2

description:
full-length gene expression
Smart-seq2
iKO_2
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: SNU449_wt_parental_r2

description:
Axl_Rego_Quantseq_star253a_GRCh38v40.SS2_DESeq2-220726_AxlWT.minusRego_VS_AxlKO.minusRego_Prep.Date_DEtable.txt
Axl_Rego_Quantseq_star253a_GRCh38v40.SS2_DESeq2-220726_AxlWT.minusRego_VS_AxlKO.plusRego_Prep.Date_DEtable.txt

value: naive


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL08

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL08_DpOo01_10_S7


title: nILC2, KO, 2

description:
full-length gene expression
Smart-seq2
nKO_2
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL08

description:
SMART-Seq v4 3' DE Kit
IL08_DmOo11_20_S2


title: SNU449_wt_parental_r1


title: nILC2, CTRL, 4

description:
full-length gene expression
Smart-seq2
nCL_4
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: nILC2, CTRL, 2

description:
full-length gene expression
Smart-seq2
nCL_2
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: nILC2, CTRL, 3

description:
full-length gene expression
Smart-seq2
nCL_3
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: Huh7 cells, sghnRNPA2B1-3

city: shanghai

first: Weijie

last: Hao

state: shanghai

address: 1630 Dongfang Road, Shanghai, China

institute: shanghai jiaotong university

data_processing:
hg19 Genomics Workbench v 11.0.1
Sequence reads were trimmed for adaptor sequence/low-quality sequence usinghg19 genomic benchwork (parameter- Quality limit: 0.05)
Trimmed sequence reads were mapped to hg19 using CLC genomic benchwork (parameters- mismath cost: 2 insertion cost: 3 deletion cost: 3 length fraction: 0.8 similarity fraction: 0.8)
Read count extraction and normalization were performed using hg19 genomic benchwork
Assembly: hg19
Supplementary files format and content: tab-delimited text files include Counts values for each Sample

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen). 1.3 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols.

value: Liver cancer cells

value: Cell culture


title: Huh7 cells, sghnRNPA2B1-2


title: nILC2, KO, 1

description:
full-length gene expression
Smart-seq2
nKO_1
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: C72h, IL2+IL7+IL25+ILC2, 4

description:
full-length gene expression
Smart-seq2
C72h8
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: small intestinal ILC2s

value: IL2 IL7 and IL25


title: Huh7 cells, sghnRNPA2B1-1


title: C72h, IL2+IL7+IL25+ILC2, 3

description:
full-length gene expression
Smart-seq2
C72h7
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: WT


title: SNU449_wt_parental_r3

value: liver

value: Hepatocellular carcinoma cell line


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo11_20_S8_IL07

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL07_DpOo11_20_S8

value: Primary follicle


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL07

description:
SMART-Seq v4 3' DE Kit
IL07_DmOo11_20_S2


title: C72h, IL2+IL7+IL25+ILC2, 2

description:
full-length gene expression
Smart-seq2
C72h6
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: Small Intestine

value: 8-12 weeks

value: pooled male and female


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL07

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL07_DpOo01_10_S7

value: LCM


title: Huh7 cells, sgNC-3


title: nILC2, KO, 3

description:
full-length gene expression
Smart-seq2
nKO_3
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv

value: KO

value: primary small intestine ILC2s


title: C72h, IL2+IL7+ILC2, 4

description:
full-length gene expression
Smart-seq2
C72h4
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: IL2 and IL7


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL07

description:
SMART-Seq v4 3' DE Kit
IL07_DmOo01_10_S1

value: Secondary-antral

value: gene expression


title: Huh7 cells, sgNC-2

value: Liver cancer cells


title: C72h, IL2+IL7+ILC2, 3

description:
full-length gene expression
Smart-seq2
C72h3
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL06

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL06_DpOo01_10_S7


title: C72h, IL2+IL7+ILC2, 1

description:
full-length gene expression
Smart-seq2
C72h1
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: Huh7 cells, sgNC-1

value: Cell culture


title: C72h, IL2+IL7+ILC2, 2

description:
full-length gene expression
Smart-seq2
C72h2
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: small intestinal ILC2s


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL06

description:
SMART-Seq v4 3' DE Kit
IL06_DmOo11_20_S2


title: nILC2, CTRL, 1

description:
full-length gene expression
Smart-seq2
nCL_1
SS2.nILC2_iILC2.Ctrl_cKO.tpm_matrix.csv


title: HD3 [scRNA-seq]

city: Lausanne

first: Tania

last: Wyss

address: Rue du Bugnon 25A

country: Switzerland

institute: SIB Swiss Institute of Bioinformatics

library_source: transcriptomic single cell

data_processing:
Illumina bcl2fastq2 Conversion Software 2.20. Quality score is encoded in a +33 offset ASCII (Sanger like)
The raw sequencing reads were filtered, demultiplexed and aligned to the human genome and transcriptome (hg38, GRCh38-3.1.0) using the 10x Genomics Cell Ranger pipeline (version 3.1.0). The cellranger count function was used with default parameters (i.e. --include-introns=FALSE).
Assembly: hg38
Supplementary files format and content: Output of cellranger count: for each sample, a matrix (mtx) file contains the raw counts per gene (in rows) per cell (in columns). The tab-separated values file features.tsv contains the corresponding row names of the mtx file, and the barcodes.tsv file contains the corresponding cell names of the mtx file

source_name: peripheral blood

extract_protocol:
PBMCs were freshly isolated from human venous blood by Lymphoprep centrifugation. Red blood cell lysis was performed using red blood lysis buffer and platelets were removed by centrifugation. Human ILCs were identified as lineage negative and interleukin-7 receptor α (CD127) positive lymphocytes. Total ILCs were sorted as CD45+Lin-CD127+CD7+ to at least 98% purity using FACSAria. Sorted cells were washed with PBS 0.04% Bovin Serum Albumin (BSA). Cell droplet encapsulation, lysis and RNA capture was performed using 10x Chromium Controller (10x Genomics) and the Chromium Single Cell 3′ v3 Reagent Kit according to the manufacturer’s protocol. One cDNA library per healthy donor was generated and further sequenced (paired-end) on a HiSeq2500 device (Ilumina)
10x genomics Single Cell 3' v3 chemistry

value: peripheral blood

value: total innate lymphoid cells

tag: disease state

value: healthy


title: C48h, IL2+IL7+IL25+ILC2, 3

description:
full-length gene expression
Smart-seq2
C48h7
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: WT

value: IL2 IL7 and IL25


title: C48h, IL2+IL7+IL25+ILC2, 2

description:
full-length gene expression
Smart-seq2
C48h6
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: Small Intestine


title: C48h, IL2+IL7+IL25+ILC2, 4

description:
full-length gene expression
Smart-seq2
C48h8
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: 8-12 weeks


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL05

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL05_DpOo01_10_S7


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL06

description:
SMART-Seq v4 3' DE Kit
IL06_DmOo01_10_S1


title: C48h, IL2+IL7+ILC2, 4

description:
full-length gene expression
Smart-seq2
C48h4
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: pooled male and female


title: C48h, IL2+IL7+ILC2, 3

description:
full-length gene expression
Smart-seq2
C48h3
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: HD1 [scRNA-seq]


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL05

description:
SMART-Seq v4 3' DE Kit
IL05_DmOo11_20_S2

value: LCM

value: Primary follicle


title: C48h, IL2+IL7+ILC2, 2

description:
full-length gene expression
Smart-seq2
C48h2
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: C72h, IL2+IL7+IL25+ILC2, 1

description:
full-length gene expression
Smart-seq2
C72h5
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: HD4 [scATAC-seq]

library_source: genomic single cell

data_processing:
Illumina bcl2fastq2 Conversion Software 2.20. Quality score is encoded in a +33 offset ASCII (Sanger like)
The raw sequencing reads were filtered, demultiplexed, aligned to the human genome (refdata-cellranger-atac-GRCh38-1.1.0, gencode.v28.basic annotation), and accessible chromatin regions were detected using the 10x Genomics Cell Ranger-atac pipeline (version1.1.0).
Assembly: hg38
Supplementary files format and content: Output of cellranger-atac count: For each sample, the bed file indicates the accessible chromatin regions, with chromosome, start and stop position of each peak; a tab-separated values fragment file (fragment.tsv) and its associated samtools tabix index, containing the list of fragments; a comma-separated values file (singlecell.csv) containing information about cells called and their associated QC metrics, with column "is__cell_barcode" indicating whether the barcode is a true cell (=1) or not (=0).

extract_protocol:
PBMCs were freshly isolated from human venous blood by Lymphoprep centrifugation. Red blood cell lysis was performed using red blood lysis buffer and platelets were removed by centrifugation. Human ILCs were identified as lineage negative and interleukin-7 receptor α (CD127) positive lymphocytes. Total ILCs were sorted as CD45+Lin-CD127+CD7+ to at least 98% purity using FACSAria. Single nuclei were isolated, transposed and one scATAC-seq library per healthy donor was generated using the 10x Chromium Single Cell ATAC Reagent Kits.
Chromium Single Cell ATAC Reagent Kits


title: C4h, IL2+IL7+IL25+ILC2, 4

description:
full-length gene expression
Smart-seq2
C4h8
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: C4h, IL2+IL7+IL25+ILC2, 3

description:
full-length gene expression
Smart-seq2
C4h7
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo11_20_S8_IL04

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL04_DpOo11_20_S8


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL08

description:
SMART-Seq v4 3' DE Kit
IL08_DmOo01_10_S1


title: C4h, IL2+IL7+IL25+ILC2, 2

description:
full-length gene expression
Smart-seq2
C4h6
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL04

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL04_DpOo01_10_S7


title: HD2 [scRNA-seq]


title: BRC#2-2, scTCR-seq, replicate 2

city: Daejeon

first: Seung Hyuck

last: Jeon

address: 291 Daehak-ro, Yuseong-gu

country: South Korea

institute: Korea Advanced Institute of Science and Technology

description: 10x Genomics

data_processing:
The scRNA-seq data were aligned and quantified using the Cell Ranger Single-Cell Software Suite (v.2.3, 10x Genomics) against the GRCh38 human reference genome.
The TCR sequence data from 10X Genomics were processed using Cell Ranger software (v.3.1) using the human VDJ reference genome.
Assembly: GRCh38
Supplementary files format and content: 10x Genomics output files: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz
Library strategy: scTCR-seq

source_name: Breast cancer

extract_protocol:
The cells were washed 3 times with PBS supplemented with 1% BSA, and the number of cells was adjusted to 700-1,200 cells/μL.
The scRNA-seq and scTCR-seq libraries were prepared using the Chromium Next GEM Single Cell 5’ Kit v1.1 (10x Genomics) and Chromium Single Cell Human TCR Amplification kit (10x Genomics), respectively, according to the standard manufacturer’s instructions

value: Breast cancer

value: CD8+ cells

value: NY-ESO-1 OLPs


title: BRC#2-1, scTCR-seq, replicate 1

description: 10x Genomics


title: BRC#2-2, scRNA-seq, replicate 2

description: 10x Genomics

data_processing:
The scRNA-seq data were aligned and quantified using the Cell Ranger Single-Cell Software Suite (v.2.3, 10x Genomics) against the GRCh38 human reference genome.
The TCR sequence data from 10X Genomics were processed using Cell Ranger software (v.3.1) using the human VDJ reference genome.
Assembly: GRCh38
Supplementary files format and content: 10x Genomics output files: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz


title: C4h, IL2+IL7+ILC2, 4

description:
full-length gene expression
Smart-seq2
C4h4
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: IL2 and IL7


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL04

description:
SMART-Seq v4 3' DE Kit
IL04_DmOo01_10_S1

value: Secondary-antral

value: gene expression


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo11_20_S8_IL03

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL03_DpOo11_20_S8


title: C4h, IL2+IL7+IL25+ILC2, 1

description:
full-length gene expression
Smart-seq2
C4h5
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: small intestinal ILC2s


title: C48h, IL2+IL7+ILC2, 1

description:
full-length gene expression
Smart-seq2
C48h1
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: BRC#1-2, scTCR-seq, replicate 2

description: 10x Genomics

value: autologous tumor cell


title: C4h, IL2+IL7+ILC2, 2

description:
full-length gene expression
Smart-seq2
C4h2
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: C4h, IL2+IL7+ILC2, 3

description:
full-length gene expression
Smart-seq2
C4h3
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL03

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL03_DpOo01_10_S7


title: Nitro_HhaHNS_rep2

city: Madison

first: Robert

last: Landick

state: WI

address: 1550 Linden Dr.

country: USA

institute: University of Wisconsin

data_processing:
Library strategy: Nitro-seq
base calling and de-multiplexing with Illumina Pipeline, version 1.8.2
Sequencing adapter reads and reads shorter than 30 nt were removed with Trimmomatic (version 0.38) and the quality of remaining read were confirmed by FastQC
Remaining paired-end reads were aligned to the MG1655 genome (Accession: NC_000913.3) using Bowtie2 (version 2.3.4.2)   with the parameters (-q --end-to-end -I 100 -X 800 --fr -t). Correctly paired reads were retained and duplicate reads were removed with Samtools (version 1.9)   and bedgraph files for visualization were created with BEDtools (version 2.8)
ChIP- or Nitro-seq patterns, read coverage in each condition was scaled for sequencing depth by reads per million. To correct ChIP-seq patterns for variations in amounts of input DNA and in sequencing, the log2 of control reads from input DNA was subtracted from the log2 of IP reads to give a corrected read coverage.
To determine the enriched DNA segments more rigorously, we used MOSAiCS (Model-based one and two Sample Analysis and Inference for ChIP-Seq Data) (Kuan et al., 2011), specifically the two-sample analysis and MOSAiCS with a hidden Markov Model (MOSAiCS-HMM) (Chung et al., 2014).
Genome_build: MG1655 genome (Accession: NC_000913.3)
Supplementary_files_format_and_content: bedGraph files include normalized read coverage across the E. coli genome (normalized to input and log2 transformed) for both ChIP-seq and Nitro-seq samples. Files were generated with bedtools "coverage" command. For samples Nitro_noprotein_rep1, Nitro_noprotein_rep2, and Nitro_noprotein_rep3, the bedgraph file contains raw read coverage of the genome.
Supplementary_files_format_and_content: BED files are bed files showing enriched regions identified by MOSAiCS analysis.

organism: Escherichia coli

source_name: Escherichia coli MG1655 purified genomic DNA from stationary cells

extract_protocol: Treated filter with elution buffer (20 mM Tris-HCl, pH 7.8, 0.2% SDS, 0.3 M sodium acetate) to elute DNA retained on filter

tag: growth phase

value: stationary

value: Protein and DNA mixture applied to 0.45 µm nitrocellulose filter

tag: concentration of protein added

value: 100 nM Hha + 100 nM H-NS


title: C48h, IL2+IL7+IL25+ILC2, 1

description:
full-length gene expression
Smart-seq2
C48h5
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: WT

value: IL2 IL7 and IL25


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL03

description:
SMART-Seq v4 3' DE Kit
IL03_DmOo11_20_S2


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL05

description:
SMART-Seq v4 3' DE Kit
IL05_DmOo01_10_S1


title: Nitro_HhaHNS_rep3


title: Nitro_100nMStpA_rep2

value: 100 nM StpA


title: Nitro_100nMStpA_rep3


title: BRC#1-1, scTCR-seq, replicate 1

description: 10x Genomics


title: BRC#1-2, scRNA-seq, replicate 2

description: 10x Genomics


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL03

description:
SMART-Seq v4 3' DE Kit
IL03_DmOo01_10_S1


title: Nitro_50nM_StpA_rep3

value: 50 nM StpA


title: BRC#1-1, scRNA-seq, replicate 1

description: 10x Genomics


title: C4h, IL2+IL7+ILC2, 1

description:
full-length gene expression
Smart-seq2
C4h1
SS2.C4h28h72h.IL2IL7IL25.tpm_matrix.csv

value: Small Intestine

value: 8-12 weeks

value: pooled male and female


title: BRC#2-1, scRNA-seq, replicate 1

description: 10x Genomics

value: Breast cancer

value: CD8+ cells


title: Nitro_50nM_StpA_rep2


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo11_20_S8_IL02

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL02_DpOo11_20_S8

value: LCM

value: Primary follicle


title: Nitro_50nM_StpA_rep1


title: Nitro_12nM_Fis_rep2

value: 12 nM Fis


title: RCC#2, scTCR-seq

description: 10x Genomics

source_name: Renal cell carcinoma

value: Renal cell carcinoma

value: CD45+ cells


title: RCC#2, scRNA-seq

description: 10x Genomics


title: Nitro_12nM_Fis_rep3

value: stationary


title: Nitro_12nM_Fis_rep1

value: Protein and DNA mixture applied to 0.45 µm nitrocellulose filter


title: Nitro_5nM_Fis_rep1

value: 5 nM Fis


title: RCC#1, scTCR-seq

description: 10x Genomics


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL02

description:
SMART-Seq v4 3' DE Kit
IL02_DmOo01_10_S1

value: Secondary-antral

value: gene expression


title: Nitro_200nM_HNS_rep3

value: 200 nM H-NS


title: RCC#1, scRNA-seq

description: 10x Genomics


title: Nitro_200nM_HNS_rep1


title: Nitro_200nM_HNS_rep2


title: Nitro_100nMStpA_rep1

value: 100 nM StpA


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL01

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL01_DpOo01_10_S7


title: Nitro_100nM_HNS_rep3

value: 100 nM H-NS


title: Nitro_100nM_HNS_rep2


title: HNC, scRNA-seq

description: 10x Genomics

source_name: Head and neck carcinoma

value: Head and neck carcinoma


title: Nitro_100nM_HNS_rep1


title: Single oocyte (DRaqL - SMART-Seq v4 3' DE Kit) DpOo01_10_S7_IL02

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL02_DpOo01_10_S7


title: Nitro_noprotein_rep3


title: Nitro_noprotein_rep2


title: HCC, scTCR-seq

description: 10x Genomics

source_name: Hepatocellular carcinoma

value: Hepatocellular carcinoma


title: Nitro_noprotein_rep1


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL10

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL10_DpGC51_60_S12

source_name: Granulosa cell

value: Non-neighboring


title: Nitro_HhaHNS_rep1

value: 100 nM Hha + 100 nM H-NS


title: Nitro_5nM_Fis_rep2


title: Nitro_input

extract_protocol: None, sample was purified genomic DNA


title: HCC, scRNA-seq

description: 10x Genomics


title: HNC, scTCR-seq

description: 10x Genomics


title: miRNA-seq NPC WT rep 2

city: Toronto

first: Marat

last: Mufteev

address: 686 Bay St.

department: Molecular Genetics

country: Canada

institute: University of Toronto

data_processing:
Reads are trimmed with cutadapt.
Custom STAR genome index was generated for hybrid genome of human, fly, yeast, ERCC to facilitate spike-ins quantification. Reads are aligned to hybrid genome. Reads with MAPQ < 2 are filtered out.
Finally, usage of poly-A sites, obtained from PolyA_DB version 3, was defined as a sum of reads whose 3`-ends are falling within 20bp upstream and 10bp downstream of the poly-A sites.
Human counts are normalized to the total count of fly counts. Gene counts are obtained as a sum of normalized counts for all pA sites of a gene.
The abundance of mature miRNAs was quantified with mirdeep2 pipeline.
Assembly: hg38

source_name: iPSC derived neuronal progenitors

growth_protocol: iPSC lines #37 (WT) from a female patient were previously described. This cell line was generated and cultured under the approval of the SickKids Research Ethics Board and the Canadian Institutes of Health Research Stem Cell Oversight Committee. iPSC lines were cultured in 5% CO2 on BD hESC-qualified matrigel (BD) in mTeSR medium (STEMCELL Technologies). Cultures were passaged using ReLeSR (STEMCELL Technologies) following the manufacture’s instruction every 6-7 days. For neuronal induction, iPSCs were aggregated as Embryoid Bodies (EBs) in low-attachment dishes in N2 media containing laminin (1 ml/ml) with 10 mM SB431542, 2 mM DSM, and 1x penicillin-streptomycin changed daily. After 7 days, EBs were plated on poly-L-ornithine + laminin-coated dishes and grown in N2 media + laminin (1 ml/ml). After 7 days, neural rosettes were manually picked and transferred to poly-L-ornithine + laminin-coated wells. After 7 days, neural rosettes were picked a second time, digested with Accutase and plated on poly-L-ornithine + laminin-coated wells. Resulting neural precursor cells (NPC) were grown as a monolayer and split every 5-7 days in NPC media containing DMEM/F12, N2, B27, 1x non-essential amino acid (NEAA), 2 mg/ml Heparin, 1 mg/ml laminin.

extract_protocol: not provided

tag: 4su labeling time before harvest, (hours)

value: wild type

value: line 37 (Cheung 2011, HMG)

tag: sequencing batch

value: C

tag: treatment protocol

tag: extraction protocol

value: To calculate relative and absolute differences in the miRNA population in NEUWT and NEURTT, small RNAs were extracted from two replicates of both lines using the same number of cells followed by the addition of a set of spike-in RNAs. Small RNAs were extracted from 500,000 neurons of each line using the SPLIT RNA extraction Kit (Lexogen) according to the manufacturer’s instructions. A set of 52 RNA spike-ins (QIAseq miRNA Library QC Spike-Ins – Qiagen) that spanned a wide range of concentrations were added to the recovered RNAs according to the manufacturer’s instructions.

value: Sequencing libraries were made using the Small RNA library preparation kit NEBNext (NEB) according to the manufacturer’s instructions. Sequencing was performed on the Illumina HiSeq 2500 using the Rapid Run mode.


title: Rif_input_rep2

data_processing:
base calling and de-multiplexing with Illumina Pipeline, version 1.8.2
Sequencing adapter reads and reads shorter than 30 nt were removed with Trimmomatic (version 0.38) and the quality of remaining read were confirmed by FastQC
Remaining paired-end reads were aligned to the MG1655 genome (Accession: NC_000913.3) using Bowtie2 (version 2.3.4.2)   with the parameters (-q --end-to-end -I 100 -X 800 --fr -t). Correctly paired reads were retained and duplicate reads were removed with Samtools (version 1.9)   and bedgraph files for visualization were created with BEDtools (version 2.8)
ChIP- or Nitro-seq patterns, read coverage in each condition was scaled for sequencing depth by reads per million. To correct ChIP-seq patterns for variations in amounts of input DNA and in sequencing, the log2 of control reads from input DNA was subtracted from the log2 of IP reads to give a corrected read coverage.
To determine the enriched DNA segments more rigorously, we used MOSAiCS (Model-based one and two Sample Analysis and Inference for ChIP-Seq Data) (Kuan et al., 2011), specifically the two-sample analysis and MOSAiCS with a hidden Markov Model (MOSAiCS-HMM) (Chung et al., 2014).
Genome_build: MG1655 genome (Accession: NC_000913.3)
Supplementary_files_format_and_content: bedGraph files include normalized read coverage across the E. coli genome (normalized to input and log2 transformed) for both ChIP-seq and Nitro-seq samples. Files were generated with bedtools "coverage" command. For samples Nitro_noprotein_rep1, Nitro_noprotein_rep2, and Nitro_noprotein_rep3, the bedgraph file contains raw read coverage of the genome.
Supplementary_files_format_and_content: BED files are bed files showing enriched regions identified by MOSAiCS analysis.

source_name: Escherichia coli RL3000 culture

extract_protocol: input DNA was purified from the cell lysate using the Qiagen QIAquick PCR purification kit

tag: genotype

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-14

tag: media

value: MOPS defined minimal media with 0.2% glucose

value: mid-exponential

value: 100 µg rifampicin/mL

tag: chip antibody

value: none (input)


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL10

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL10_DpGC31_40_S10

value: LCM

value: gene expression


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL10

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL10_DpGC41_50_S11


title: Rif_BetaIP_rep1

extract_protocol: specific RNAP-DNA complexes were isolated with an antibody specific to the Beta subunit (Neoclone #WP0022) and the resulting DNA was purified with the Qiagen QIAquick PCR purification kit

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-12

tag: treatment

value: anti-RNAP (Beta subunit)


title: Rif_input_rep1

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-11


title: miRNA-seq iPSC WT rep 2

source_name: iPSC

tag: cell line


title: Rif_HNSIP_rep1

extract_protocol: specific H-NS-DNA complexes were isolated with a polyclonal antibody specific to H-NS (from Harlan Bioproducts) and the resulting DNA was purified with the Qiagen QIAquick PCR purification kit

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-13

value: anti-H-NS


title: miRNA-seq NPC WT rep 1

value: wild type


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL01

description:
SMART-Seq v4 3' DE Kit
IL01_DmOo11_20_S2

value: Primary follicle


title: miRNA-seq iPSC WT rep 1

value: line 37 (Cheung 2011, HMG)

value: C


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL10

description:
SMART-Seq v4 3' DE Kit
IL10_DmGC51_60_S6

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: Non-neighboring


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo01_10_S1_IL01

description:
SMART-Seq v4 3' DE Kit
IL01_DmOo01_10_S1

value: Secondary-antral


title: NoRif_HNSIP_rep2

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-10

value: MOPS defined minimal media with 0.2% glucose

value: methanol (solvent)


title: Rif_BetaIP_rep2

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-15

value: mid-exponential

value: 100 µg rifampicin/mL


title: NoRif_BetaIP_rep2

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-9


title: Rif_HNSIP_rep2

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-16


title: Single oocyte (SMART-Seq v4 3' DE Kit) DmOo11_20_S2_IL02

description:
SMART-Seq v4 3' DE Kit
IL02_DmOo11_20_S2

value: LCM

value: gene expression


title: NoRif_HNSIP_rep1

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-7

value: anti-H-NS


title: NoRif_input_rep2

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-8

value: none (input)


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL10

description:
SMART-Seq v4 3' DE Kit
IL10_DmGC41_50_S5


title: NoRif_input_rep1

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-5


title: RATE-seq UHRR rep2

source_name: UHRR

value: wild type

value: human tissue

value: A

value: RNA-seq libraries were prepared for each time-points and seady-state sample using the QuantSeq 3’ mRNA-Seq Library Prep Kit FWD for Illumina (Lexogen) automated on the NGS WorkStation (Agilent) at The Centre for Applied Genomics (TCAG) according to the manufacturer’s instructions. PCR cycle numbers were determined using the PCR Add-on Kit for Illumina (Lexogen). All steady-state samples were processed with 250 ng of total RNA input. To minimize variability between time-points within a batch, RNA samples were processed with the same total RNA input with a minimum of 100 ng of total RNA used. Each sample was spiked-in with ERCC RNA Spike-In Control Mix 1 (Ambion) according to the manufacturer’s instructions prior to the start of library preparation. Library quality and quantity were measured at TCAG prior to sequencing with Bioanalyzer (Agilent) and KAPA qPCR (Roche). Sequencing was also performed at TCAG on the Illumina HiSeq 2500 with single-end 100bp read length yielding 40 to 50 million reads.


title: RATE-seq UHRR rep1


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL10

description:
SMART-Seq v4 3' DE Kit
IL10_DmGC21_30_S3

value: Non-neighboring


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL09

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL09_DpGC51_60_S12


title: S2R+, IgG, NHS, rep2

city: Helsinki

first: Maria

last: Sokolova

address: Viikinkaari 5

department: Institute of Biotechnology

country: Finland

institute: University of Helsinki

data_processing:
ChIP-Seq data sets were aligned using Bowtie2
Assembly: dm6 (BDGP6.87)
Supplementary files format and content: bigwig files were generated using DeepTools with RPKM normalization method

organism: Drosophila melanogaster

source_name: embryo

growth_protocol: Schneider cell line (S2R+) cultured in Schneider’s Drosophila Medium (Thermo Fisher Scientific) supplemented with 10% heat inactivated fetal bovine serum (Thermo Fisher Scientific) and 1 x antibiotic-antimycotic solution (penicillin, streptomycin, Amphotericin B; Thermo Fisher Scientific) at +25°C incubator.

extract_protocol:
samples were  dissected from fly strains were fixed in 1% PFA for 10 min at RT, homogenized using a pestle in 300 ml of RIPA buffer, and sonicated with Bioruptor (Diagenode). Immunoprecipitations were carried out with 5 µg antibody overnight at 4°C in a rotating wheel.
ChIP libraries were prepared for Illumina NextSeq 500 using NEBNext ChIP-Seq DNA Sample Prep Master Mix Set for Illumina (NEB E6240) and NEBNext® Multiplex Oligos for Illumina® (Index Primers Set 1) (NEB E7335) according to the manufacturer’s protocols.

treatment_protocol: Heat shock was performed by placing sealed flasks into +37°C water bath for 20 mins.

tag: tissue

value: embryo

value: normal mouse IgG:sc-2025

value: non-heat shock


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL10

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL10_DpGC21_30_S9

tag: collecting conditions


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL09

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL09_DpGC41_50_S11

tag: class


title: NoRif_BetaIP_rep1

value: MG1655 F-, lambda-, ilvG468+, rfb-50, rph+, ybhJ(L54->I), yebN(G25xxx->D), ycfK::97bp deltaInsB-5 deltaInsA-5 deltaInsAB-6

value: MOPS defined minimal media with 0.2% glucose

value: mid-exponential

value: methanol (solvent)

value: anti-RNAP (Beta subunit)


title: RATE-seq NPC WT SS rep2

description:
counts_genes.csv
SS_and_TR_genes.csv
counts_pA_sites.csv
SS_and_TR_pA_sites.csv

value: line 37 (Cheung 2011, HMG)

value: The culture media of all three cell types was replaced with media supplemented with 100 μM 4sU (Sigma-Aldrich) reconstituted in DMSO. Cells were harvested at 0.5, 1, 4, 8, and 24 h after the addition of 4sU. Metabolic labeling was designed such that all time points were collected together. Cellular viability in the presence of 100 μM 4sU was monitored up to 24 h of treatment on parallel cultures by using Trypan blue staining and live/dead cell counting.

value: After 4sU incorporation, cells were quickly washed twice with ice-cold 1× PBS Total RNA and scraped into ice-cold 1.5 ml Eppendorf tubes.  Cells were collected by spinning at 1000g for 5 min at 4°C and cell pellets were resuspended in 1 mL of Trizol (Thermo Fisher Scientific). Total RNA was extracted according to manufacturer instructions. 50 µg of total RNA was mixed with 5 µg unlabeled yeast RNA and 5 µg 4sU-labeled S2 fly RNA in a total volume of 120 µL. 1 mg/mL HPDP-biotin (ThermoFisher Scientific) was reconstituted in dimethylformamide by shaking at 37°C for 30 min at 300 RPM. 120 µL of 2.5× citrate buffer (25 mM citrate, pH 4.5, 2.5 mL EDTA) and 60 µL of 1 mg/mL HPDP-biotin were added to the premixed RNA sample for each time point. The solution was incubated at 37°C for 2 h at 300 RPM on an Eppendorf ThermoMixer F1.5 in the dark. Samples were extracted twice with acid phenol, pH 4.5, and once with chloroform. RNA was precipitated with 18 µL 5M NaCl, 750 µL 100% ethanol, and 2 µL GlycoBlue (Invitrogen) overnight at −20°C. Precipitated RNA was pelleted for 30 min at 21,000g at 4°C. The RNA pellet was resuspended in 200 µL of 1× wash buffer (10 mM Tris-HCl, pH 7.4, 50 mM NaCl, 1 mM EDTA). Biotinylated RNA was purified using the µMACS Streptavidin microbeads system (Miltenyi Biotec). 50 µL Miltenyi beads per sample were pre-blocked with 48 µL 1× wash buffer and 2 µL yeast tRNA (Invitrogen), rotating for 20 min at room temperature. µMACS microcolumns were washed 1× with 100 µL nucleic acid equilibration buffer (Miltenyi Biotec), followed by 5× washes with 100 µL 1× wash buffer. Beads were applied to microcolumns in 100 µL aliquots and again washed 5× with 100 µL 1× wash buffer. Beads were demagnetized and eluted off the column with 2× 100 µL 1× wash buffer, and columns were placed back on the magnetic stand. A total of 200 µL beads was mixed with each sample of biotinylated RNA and rotated at room temperature for 20 min. Samples were applied to the microcolumns in 100 µL aliquots, washed 3× with 400 µL wash A buffer (10 mM Tris-HCl, pH 7.4, 6 M urea, 10 mM EDTA) prewarmed to 65°C, and washed 3× with 400 µL wash B buffer (10 mM Tris-HCl, pH 7.4, 1 M NaCl, 10 mM EDTA). RNA was eluted with 5× 100 µL of 1× wash buffer supplemented with 0.1 M DTT, and flow-through was collected in a tube. Purified RNA was precipitated with 30 µL 5 M NaCl, 2 µL GlycoBlue, and 1 mL 100% ethanol, incubated at −20°C overnight. Samples were spun at 21,000g at 4°C for 30 min and resuspended in 20 µL water. RNA quality was assessed by running 3 µL of samples on a 1.5% agarose gel.


title: S2R+, input, rep2

value: heat shock


title: S2R+, IgG, HS, rep2


title: RATE-seq NPC WT 24h rep2

value: wild type


title: S2R+, Actin, HS, rep2


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL09

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL09_DpGC21_30_S9

value: LCM

tag: analysis target

value: gene expression


title: S2R+, Pol2S5P, HS, rep2


title: RATE-seq NPC WT 8h rep2

value: A

value: RNA-seq libraries were prepared for each time-points and seady-state sample using the QuantSeq 3’ mRNA-Seq Library Prep Kit FWD for Illumina (Lexogen) automated on the NGS WorkStation (Agilent) at The Centre for Applied Genomics (TCAG) according to the manufacturer’s instructions. PCR cycle numbers were determined using the PCR Add-on Kit for Illumina (Lexogen). All steady-state samples were processed with 250 ng of total RNA input. To minimize variability between time-points within a batch, RNA samples were processed with the same total RNA input with a minimum of 100 ng of total RNA used. Each sample was spiked-in with ERCC RNA Spike-In Control Mix 1 (Ambion) according to the manufacturer’s instructions prior to the start of library preparation. Library quality and quantity were measured at TCAG prior to sequencing with Bioanalyzer (Agilent) and KAPA qPCR (Roche). Sequencing was also performed at TCAG on the Illumina HiSeq 2500 with single-end 100bp read length yielding 40 to 50 million reads.


title: S2R+, Pol2S5P, NHS, rep1

value: embryo


title: RATE-seq NPC WT 4h rep2


title: RATE-seq NPC WT 1h rep2


title: S2R+, Actin, NHS, rep1


title: S2R+, IgG, NHS, rep1

value: normal mouse IgG:sc-2025


title: S2R+, Pol2S5P, HS, rep1


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC31_40_S4_IL09

description:
SMART-Seq v4 3' DE Kit
IL09_DmGC31_40_S4

value: Non-neighboring


title: S2R+, Actin, HS, rep1


title: RATE-seq NPC WT 30min rep2


title: S2R+, input, rep1


title: S2R+, IgG, HS, rep1


title: RATE-seq NPC WT SS rep1


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL08

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL08_DpGC31_40_S10


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL08

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL08_DpGC41_50_S11


title: Library9b DNA-seq

city: Saint Louis

first: Michael

middle: Aaron

last: White

state: MO

address: 660 South Euclid Ave

department: Genetics

institute: Washington University in St. Louis School of Medicine

description: DNA-seq on reporter gene barcode amplicons from plasmid DNA library

data_processing:
All raw reads were processed regardless of quality score. Reads with the expected in-line multiplexing barcode for each sample followed by the spacer sequence were identified using custom python scripts.
Demultiplexed reads were used to determine the read counts for each barcode with custom python scripts.
To obtain barcode read counts, reads were matched to barcoded reporter gene sequences using custom python scripts. Only exact barcoded matches were counted. FASTA files with reporter gene sequences for each library are included as supplementary files.
Assembly: mm10
Supplementary files format and content: Each sample has its own tab-delimited file. Column 1: Barcode sequence. Column 2: cis-regulatory element name. Column 3: Barcode counts.

growth_protocol: P0 mouse retinal explants were electroporated with plasmid library and cultured ex vivo for 8 days.

extract_protocol:
RNA was extracted from retinas with TRIzol and treated with Turbo Dnase. cDNA was prepared using SuperScript IV.
Sequence barcodes from reporter genes were amplified by PCR and purified with Monarch PCR cleanup kit. Custom Illumina adapters were added by PCR.

value: wild-type


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL09

description:
SMART-Seq v4 3' DE Kit
IL09_DmGC51_60_S6


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL09

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL09_DpGC31_40_S10


title: Library9a DNA-seq


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL08

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL08_DpGC21_30_S9


title: S2R+, Actin, NHS, rep2

value: non-heat shock


title: Library4 DNA-seq


title: S2R+, Pol2S5P, NHS, rep2


title: Library5 DNA-seq


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL08

description:
SMART-Seq v4 3' DE Kit
IL08_DmGC41_50_S5


title: RATE-seq NPC WT 4h rep1


title: RATE-seq NPC WT 24h rep1


title: RATE-seq NPC WT 8h rep1


title: Library3 DNA-seq


title: Library7 DNA-seq


title: RATE-seq NPC WT 1h rep1


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC31_40_S4_IL08

description:
SMART-Seq v4 3' DE Kit
IL08_DmGC31_40_S4


title: PS19-E4_Saline_689

city: San Francisco

first: Ayushi

last: Agrawal

state: CA

address: 1650 Owens St

institute: Gladstone Institutes

description: 10x Genomics single nucleus RNA sequencing

data_processing:
Demultiplexed fastq files were aligned to a custom reference genome built from mm10 2020-A that includes introns, using the Cellranger v6.1.1 counts function with default parameters, as described in the Cellranger documentation. UMI counts were also determined using the Cellranger counts function.
Assembly: The Homo sapiens microtubule associated protein tau (MAPT) (NCBI Reference Sequence: NM_001123066.4) and the Homo sapiens APOE are genes of interest for this study. Custom mouse reference genome was made using these genes of interest , the reference mouse genome sequence (GRCm38) from Ensembl (release 98) and the mouse gene annotation file from GENCODE (release M23), similar to those used in 10x Genomics Cell Ranger mouse reference package mm10 2020-A.
Supplementary files format and content: feature-barcode matrix in the Hierarchical Data Format (HDF5 or H5) (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/advanced/h5_matrices)

molecule: nuclear RNA

source_name: hippocampus

growth_protocol: Male and female PS19-E4 and PS19-E3 mice on a pure C57BL/6 genetic background were housed in a pathogen-free barrier facility at the Gladstone Institutes on a 12 h light cycle at 19–23ºC and 30–70% humidity. Mice were genotyped by polymerase chain reaction (PCR) of a tail clipping. All animal experiments were conducted in accordance with the guidelines and regulation of the National Institutes of Health, the University of California, and the Gladstone Institutes.

extract_protocol:
The frozen left hippocampus was placed into a pre-chilled 2 mL Dounce with 1 mL of cold 1X Homogenization Buffer (1X HB) (250 mM Sucrose, 25 mM KCL, 5 mM MgCl2, 20 mM Tricine-KOH pH7.8, 1 mM DTT, 0.5 mM Sermidine, 0.15 mM Sermine, 0.3% NP40, 0.2 units/µL RNase inhibitor, ~0.07 tablets/sample Protease inhibitor). Dounce with “A” loose pestle (~10 strokes)and then with “B” tight pestle (~15 strokes). The homogenate was filtered using a 70 µM Flowmi strainer (Eppendorf) and transferred to a pre-chilled 2 mL LoBind tube (Fischer Scientific). Nuclei were pelleted by spinning for 5 min at 4°C at 350 RCF. The supernatant was removed and the nuclei were resuspended in 400 µL 1X HB. Next, 400 µL of 50% Iodixanol solution was added to the nuclei and then slowly layered with 600 µL of 30% Iodixanol solution under the 25% mixture, then layered with 600 µL of 40% Iodixanol solution under the 30% mixture. The nuclei were then spun for 20 min at 4°C at 3,000g in a pre-chilled swinging bucket centrifuge. 200 µL of the nuclei band at the 30%-40% interface was collected and transferred to a fresh tube. Then, 800 µL of 2.5% BSA in PBS plus 0.2 units/µL of RNase inhibitor was added to the nuclei and then were spun for 10 min at 500 RCF at 4C. The nuclei were resuspended with 2% BSA in PBS plus 0.2 units/µL RNase inhibitor to reach ~500 nuclei/µL. The nuclei were then filtered with a 40 µM Flowmi stainer. The nuclei were counted and then ~13,000 nuclei per sample were loaded onto 10x Genomics Next GEM chip G. The snRNA-seq libraries were prepared using the Chromium Next GEM Single Cell 3ʹ Library and Gel Bead kit v3.1(10x Genomics)according to the manufacturer’s instructions. Libraries were sequenced on an Illumina NovaSeq 6000 sequencer at the UCSF CAT Core.
The snRNA-seq libraries were prepared using the Chromium Next GEM Single Cell 3ʹ Library and Gel Bead kit v3.1 (10x Genomics) according to the manufacturer’s instructions. Libraries were sequenced on an Illumina NovaSeq 6000 sequencer at the UCSF CAT Core.
10X Single-nucleus RNAseq

treatment_protocol: At 6.5 months of age, male and female PS19-E4 and PS19-E3 mice were randomly assigned to the control or treatment group. Mice received intraperitoneal injections with either sterile grade 0.9% saline (Fisher Scientific) or a mixture of HMGB1 inhibitors: ethyl pyruvate (80mg/kg) (Sigma-Aldrich) and glycyrrhizic acid (20mg/kg) (Sigma-Aldrich) dissolved in 0.9% saline. The mice received three injections per week for 12 weeks, starting at 6.5 months of age until they reached 9.5 months of age. All mice were monitored for weight changes, grooming changes, and posture during the experiments and no changes were observed. Following treatment, the animals were perfused and the left hippocampi were dissected and snap frozen on dry ice and stored at -80ºC for snRNA-seq experiment.

value: hippocampus

value: Saline


title: PS19-E4_Saline_480


title: PS19-E4_Saline_477


title: RATE-seq iPSC WT 24h rep2


title: RATE-seq iPSC WT SS rep2


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL07

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL07_DpGC41_50_S11


title: PS19-E4_Saline_476


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL08

description:
SMART-Seq v4 3' DE Kit
IL08_DmGC21_30_S3


title: RATE-seq iPSC WT 8h rep2


title: PS19-E4_HMGB1_Inhibitors_475

value: HMGB1 inhibitors


title: PS19-E4_HMGB1_Inhibitors_525


title: PS19-E3_Saline_580


title: PS19-E3_Saline_577


title: RATE-seq NPC WT 30min rep1

value: line 37 (Cheung 2011, HMG)


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL07

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL07_DpGC51_60_S12


title: Library6 DNA-seq


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL07

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL07_DpGC31_40_S10


title: RATE-seq iPSC WT 4h rep2

value: The culture media of all three cell types was replaced with media supplemented with 100 μM 4sU (Sigma-Aldrich) reconstituted in DMSO. Cells were harvested at 0.5, 1, 4, 8, and 24 h after the addition of 4sU. Metabolic labeling was designed such that all time points were collected together. Cellular viability in the presence of 100 μM 4sU was monitored up to 24 h of treatment on parallel cultures by using Trypan blue staining and live/dead cell counting.


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL07

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL07_DpGC21_30_S9


title: PS19-E3_Saline_562


title: RATE-seq iPSC WT 1h rep2

description:
counts_genes.csv
SS_and_TR_genes.csv
counts_pA_sites.csv
SS_and_TR_pA_sites.csv

value: After 4sU incorporation, cells were quickly washed twice with ice-cold 1× PBS Total RNA and scraped into ice-cold 1.5 ml Eppendorf tubes.  Cells were collected by spinning at 1000g for 5 min at 4°C and cell pellets were resuspended in 1 mL of Trizol (Thermo Fisher Scientific). Total RNA was extracted according to manufacturer instructions. 50 µg of total RNA was mixed with 5 µg unlabeled yeast RNA and 5 µg 4sU-labeled S2 fly RNA in a total volume of 120 µL. 1 mg/mL HPDP-biotin (ThermoFisher Scientific) was reconstituted in dimethylformamide by shaking at 37°C for 30 min at 300 RPM. 120 µL of 2.5× citrate buffer (25 mM citrate, pH 4.5, 2.5 mL EDTA) and 60 µL of 1 mg/mL HPDP-biotin were added to the premixed RNA sample for each time point. The solution was incubated at 37°C for 2 h at 300 RPM on an Eppendorf ThermoMixer F1.5 in the dark. Samples were extracted twice with acid phenol, pH 4.5, and once with chloroform. RNA was precipitated with 18 µL 5M NaCl, 750 µL 100% ethanol, and 2 µL GlycoBlue (Invitrogen) overnight at −20°C. Precipitated RNA was pelleted for 30 min at 21,000g at 4°C. The RNA pellet was resuspended in 200 µL of 1× wash buffer (10 mM Tris-HCl, pH 7.4, 50 mM NaCl, 1 mM EDTA). Biotinylated RNA was purified using the µMACS Streptavidin microbeads system (Miltenyi Biotec). 50 µL Miltenyi beads per sample were pre-blocked with 48 µL 1× wash buffer and 2 µL yeast tRNA (Invitrogen), rotating for 20 min at room temperature. µMACS microcolumns were washed 1× with 100 µL nucleic acid equilibration buffer (Miltenyi Biotec), followed by 5× washes with 100 µL 1× wash buffer. Beads were applied to microcolumns in 100 µL aliquots and again washed 5× with 100 µL 1× wash buffer. Beads were demagnetized and eluted off the column with 2× 100 µL 1× wash buffer, and columns were placed back on the magnetic stand. A total of 200 µL beads was mixed with each sample of biotinylated RNA and rotated at room temperature for 20 min. Samples were applied to the microcolumns in 100 µL aliquots, washed 3× with 400 µL wash A buffer (10 mM Tris-HCl, pH 7.4, 6 M urea, 10 mM EDTA) prewarmed to 65°C, and washed 3× with 400 µL wash B buffer (10 mM Tris-HCl, pH 7.4, 1 M NaCl, 10 mM EDTA). RNA was eluted with 5× 100 µL of 1× wash buffer supplemented with 0.1 M DTT, and flow-through was collected in a tube. Purified RNA was precipitated with 30 µL 5 M NaCl, 2 µL GlycoBlue, and 1 mL 100% ethanol, incubated at −20°C overnight. Samples were spun at 21,000g at 4°C for 30 min and resuspended in 20 µL water. RNA quality was assessed by running 3 µL of samples on a 1.5% agarose gel.


title: PS19-E3_HMGB1_Inhibitors_668


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL07

description:
SMART-Seq v4 3' DE Kit
IL07_DmGC51_60_S6


title: PS19-E3_HMGB1_Inhibitors_664


title: PS19-E3_HMGB1_Inhibitors_621


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL08

description:
SMART-Seq v4 3' DE Kit
IL08_DmGC51_60_S6


title: PS19-E3_HMGB1_Inhibitors_614


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL07

description:
SMART-Seq v4 3' DE Kit
IL07_DmGC41_50_S5


title: dmPGE2_6h_1 [mRNA_Dgaln]

first: CHAO

last: CHEN

address: The First Affiliated Hospital, College of Medicine, Zhejiang University, NO.79 Qingchun Road, Hangzhou, Zhejiang 310003, China

institute: zhejiang university

description:
seq_mRNA_Dgaln_count.txt
seq_mRNA_Dgaln_fpkm.txt

data_processing:
The raw sequencing data encompasses a minor fraction of reads with sequencing joints or suboptimal sequencing quality. Reads with adapters should be discarded. Additionally, reads with an N ratio exceeding 0.002 (where N signifies an undetermined base) should be eliminated. If the proportion of low-quality bases in a single-end read surpasses 50% of the read's length, the corresponding paired reads should be removed.
Trimmed sequence reads were mapped to GrCM38/mm10 using Hisat2.
Read count extraction and normalization were performed using Hisat2.
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

source_name: Liver

growth_protocol: C57 BL6 mice is 8 week old and have a average weight of 20g.

extract_protocol:
Liver tissue of C57 BL6 mice were treated with liquid nitrogen immediately and maintained at -80℃. 1 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using NEBNext Ultra RNA Library Prep Kit for Illumina following manufacturer's protocols.

treatment_protocol: galactosamine/lipopolysaccharide(Dgaln/lps) induced liver injury model was established by intraperitoneal injection of Dgaln 500mg/kg and LPS 5μg/kg. 300μg/kg dmPGE2 is intraperitoneal injected 1 hour before Dgaln/LPS. The sample is collected after 6 hours of injection of Dgaln/LPS.

value: Liver

tag: strain

value: C57 BL6

value: WT

value: dmPGE2 300μg/kg


title: dmPGE2_6h_3 [mRNA_Dgaln]


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC31_40_S4_IL07

description:
SMART-Seq v4 3' DE Kit
IL07_DmGC31_40_S4

value: LCM

value: gene expression


title: dmPGE2+Dgaln/LPS_6h_3 [mRNA_Dgaln]

value: D-GalN 500mg/kg + LPS 5μg/kg + dmPGE2 300μg/kg


title: dmPGE2+Dgaln/LPS_6h_1 [mRNA_Dgaln]


title: Dgaln/LPS_6h_2 [mRNA_Dgaln]

value: D-GalN 500mg/kg + LPS 5μg/kg


title: Dgaln/LPS_6h_3 [mRNA_Dgaln]


title: PS19-E3_Saline_575


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL06

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL06_DpGC41_50_S11

value: Non-neighboring


title: dmPGE2_6h_2 [mRNA_Dgaln]


title: RATE-seq iPSC WT 24h rep1

value: wild type

value: A

value: RNA-seq libraries were prepared for each time-points and seady-state sample using the QuantSeq 3’ mRNA-Seq Library Prep Kit FWD for Illumina (Lexogen) automated on the NGS WorkStation (Agilent) at The Centre for Applied Genomics (TCAG) according to the manufacturer’s instructions. PCR cycle numbers were determined using the PCR Add-on Kit for Illumina (Lexogen). All steady-state samples were processed with 250 ng of total RNA input. To minimize variability between time-points within a batch, RNA samples were processed with the same total RNA input with a minimum of 100 ng of total RNA used. Each sample was spiked-in with ERCC RNA Spike-In Control Mix 1 (Ambion) according to the manufacturer’s instructions prior to the start of library preparation. Library quality and quantity were measured at TCAG prior to sequencing with Bioanalyzer (Agilent) and KAPA qPCR (Roche). Sequencing was also performed at TCAG on the Illumina HiSeq 2500 with single-end 100bp read length yielding 40 to 50 million reads.


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL06

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL06_DpGC31_40_S10


title: Dgaln/LPS_6h_1 [mRNA_Dgaln]


title: PS19-E4_HMGB1_Inhibitors_570

value: hippocampus


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL06

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL06_DpGC51_60_S12


title: Control_6h_3 [mRNA_Dgaln]

value: Saline


title: RATE-seq iPSC WT 30min rep2

value: line 37 (Cheung 2011, HMG)


title: RATE-seq iPSC WT 8h rep1

value: The culture media of all three cell types was replaced with media supplemented with 100 μM 4sU (Sigma-Aldrich) reconstituted in DMSO. Cells were harvested at 0.5, 1, 4, 8, and 24 h after the addition of 4sU. Metabolic labeling was designed such that all time points were collected together. Cellular viability in the presence of 100 μM 4sU was monitored up to 24 h of treatment on parallel cultures by using Trypan blue staining and live/dead cell counting.


title: dmPGE2+Dgaln/LPS_6h_2 [mRNA_Dgaln]


title: Control_6h_2 [mRNA_Dgaln]


title: RATE-seq iPSC WT 30min rep1

value: After 4sU incorporation, cells were quickly washed twice with ice-cold 1× PBS Total RNA and scraped into ice-cold 1.5 ml Eppendorf tubes.  Cells were collected by spinning at 1000g for 5 min at 4°C and cell pellets were resuspended in 1 mL of Trizol (Thermo Fisher Scientific). Total RNA was extracted according to manufacturer instructions. 50 µg of total RNA was mixed with 5 µg unlabeled yeast RNA and 5 µg 4sU-labeled S2 fly RNA in a total volume of 120 µL. 1 mg/mL HPDP-biotin (ThermoFisher Scientific) was reconstituted in dimethylformamide by shaking at 37°C for 30 min at 300 RPM. 120 µL of 2.5× citrate buffer (25 mM citrate, pH 4.5, 2.5 mL EDTA) and 60 µL of 1 mg/mL HPDP-biotin were added to the premixed RNA sample for each time point. The solution was incubated at 37°C for 2 h at 300 RPM on an Eppendorf ThermoMixer F1.5 in the dark. Samples were extracted twice with acid phenol, pH 4.5, and once with chloroform. RNA was precipitated with 18 µL 5M NaCl, 750 µL 100% ethanol, and 2 µL GlycoBlue (Invitrogen) overnight at −20°C. Precipitated RNA was pelleted for 30 min at 21,000g at 4°C. The RNA pellet was resuspended in 200 µL of 1× wash buffer (10 mM Tris-HCl, pH 7.4, 50 mM NaCl, 1 mM EDTA). Biotinylated RNA was purified using the µMACS Streptavidin microbeads system (Miltenyi Biotec). 50 µL Miltenyi beads per sample were pre-blocked with 48 µL 1× wash buffer and 2 µL yeast tRNA (Invitrogen), rotating for 20 min at room temperature. µMACS microcolumns were washed 1× with 100 µL nucleic acid equilibration buffer (Miltenyi Biotec), followed by 5× washes with 100 µL 1× wash buffer. Beads were applied to microcolumns in 100 µL aliquots and again washed 5× with 100 µL 1× wash buffer. Beads were demagnetized and eluted off the column with 2× 100 µL 1× wash buffer, and columns were placed back on the magnetic stand. A total of 200 µL beads was mixed with each sample of biotinylated RNA and rotated at room temperature for 20 min. Samples were applied to the microcolumns in 100 µL aliquots, washed 3× with 400 µL wash A buffer (10 mM Tris-HCl, pH 7.4, 6 M urea, 10 mM EDTA) prewarmed to 65°C, and washed 3× with 400 µL wash B buffer (10 mM Tris-HCl, pH 7.4, 1 M NaCl, 10 mM EDTA). RNA was eluted with 5× 100 µL of 1× wash buffer supplemented with 0.1 M DTT, and flow-through was collected in a tube. Purified RNA was precipitated with 30 µL 5 M NaCl, 2 µL GlycoBlue, and 1 mL 100% ethanol, incubated at −20°C overnight. Samples were spun at 21,000g at 4°C for 30 min and resuspended in 20 µL water. RNA quality was assessed by running 3 µL of samples on a 1.5% agarose gel.


title: RATE-seq iPSC WT 4h rep1


title: RATE-seq iPSC WT 1h rep1


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL06

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL06_DpGC21_30_S9


title: SMMC7721 cells_siSNRPE_rep3

city: Nanjing

first: Qipeng

last: Wu

state: Jiangsu

address: Tongjiaxiang No.24

institute: China Pharmaceutical University

data_processing:
Quality of bases lower than 20, pass filter data of fastq format were processed by Cutadapt (V1.9.1) to be high quality clean data.
Reference genome sequences and gene model annotation files of relative species were downloaded from genome website.Hisat2 (v2.0.1) was used to index  reference genome sequence.
In the beginning transcripts in fasta format are converted from known gff annotation file and indexed  properly. Then, with the file as a reference gene file, HTSeq (v0.6.1) estimated gene and isoform  expression levels from the pair-end clean data.
Genome_build: hg37
Supplementary_files_format_and_content: xls files include FPKM values for each Sample

source_name: SMMC7721 cells

extract_protocol:
Total RNA of each sample was extracted using TRIZOL Reagent (Vazyme). Total RNA of each sample was quantified and qualified by Agilent 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA, USA), NanoDrop (Thermo Fisher Scientific Inc.), and 1% agrose gel. 1 μg total RNA with RIN value above 6.5 was used for library preparation.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: siSNRPE


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL07

description:
SMART-Seq v4 3' DE Kit
IL07_DmGC21_30_S3


title: dmPGE2+APAP_6h_3 [mRNA]

description:
mRNA_FPKM.txt
mRNA_readcount.txt
lncRNA_FPKM.txt
lncRNA_readcount.txt

extract_protocol:
Liver tissue of C57 BL6 mice were treated with liquid nitrogen immediately and maintained at -80℃. 1 ug of total RNA was used for the construction of sequencing libraries.
Ribosomal RNA is extracted from the total RNA, which is subsequently fragmented into short sequences ranging from 250-300 bp. Utilizing this fragmented RNA as a template and employing random oligonucleotides as primers, the first strand of cDNA is synthesized. Following this, the RNA strand is degraded using RNase H. The second strand of cDNA is then synthesized using dNTPs (dUTP, dATP, dGTP, and dCTP) within the DNA polymerase I system. The purified double-stranded cDNA undergoes processes such as end repair, A-tailed addition, and sequencing linkages. AMPure XP beads are employed to isolate cDNA fragments approximately 350-400 bp in length. The USER enzyme is used to degrade the second cDNA strand containing U, culminating in PCR amplification to produce the final library.

treatment_protocol: Acetaminophen(APAP) induced liver injury model was established by intraperitoneal injection of 500mg/kg APAP. 300μg/kg dmPGE2 is intraperitoneal injected 1 hour before APAP. The sample is collected after 6 hours of injection of APAP.

value: Liver

value: C57 BL6

value: 500mg/kg Acetaminophen + 300μg/kg dmPGE2


title: dmPGE2+Dgaln/LPS_6h_4 [mRNA_Dgaln]

value: WT


title: APAP_6h_3 [mRNA]

value: 500mg/kg Acetaminophen


title: dmPGE2+APAP_6h_1 [mRNA]


title: APAP_6h_2 [mRNA]


title: SMMC7721 cells_siSNRPE_rep2


title: RATE-seq iPSC WT SS rep1


title: Control_6h_1 [mRNA]


title: Control_6h_2 [mRNA]


title: SMMC7721 cells_NC_rep2

value: negative control


title: SMMC7721 cells_NC_rep3


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL06

description:
SMART-Seq v4 3' DE Kit
IL06_DmGC21_30_S3

value: LCM

value: gene expression


title: SMMC7721 cells_NC_rep1


title: SMMC7721 cells_siSNRPE_rep1


title: Control_6h_3 [mRNA]


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL06

description:
SMART-Seq v4 3' DE Kit
IL06_DmGC51_60_S6

value: Non-neighboring


title: APAP_6h_1 [mRNA]


title: APAP_6h_3 [miRNA]

data_processing:
Discard low-quality reads, specifically those with a mass value of sQ lower than 20 and those where the alkali base constitutes over 30% of the total reads. Eliminate reads with an undetermined base information, represented by N, exceeding 10% of the read. Exclude reads contaminated with 5' connectors and those lacking 3' connector sequences and insertion fragments. Truncate the 3' connector sequence. Finally, remove reads derived from continuous polyA/T/G/C sequences, as these may result from sequencing errors and possess low information entropy, rendering them unsuitable for analysis.
Analyze the clean reads from each sample, selecting sRNAs that fall within a specific length range for further analysis. Typically, the sRNA length in animals ranges from 18-35nt, and in plants, it spans 18-30nt. The sRNA length distribution peak plot aids in identifying the sRNA type; for instance, miRNAs predominantly fall within the 21-22nt range, while siRNAs are primarily at 24nt.
Employ Bowtie to map the length-filtered sRNAs to the reference sequence, and then analyze the distribution of these small RNAs on the reference. The subsequent sRNA classification pertains to these mapped sRNAs.
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: tab-delimited text files include TPM values for each Sample

extract_protocol:
Liver tissue of C57 BL6 mice were treated with liquid nitrogen immediately and maintained at -80℃. 1 ug of total RNA was used for the construction of sequencing libraries.
Using the Small RNA Sample Pre Kit, we constructed a library by leveraging the distinct structures at the 3' and 5' ends of Small RNA, which possess complete phosphate groups at the 5' end and hydroxyl groups at the 3' end. Starting with total RNA, the ends of Small RNA were directly ligated with connectors, followed by reverse transcription to synthesize cDNA. After PCR amplification, the target DNA fragment was isolated using PAGE gel electrophoresis. The cDNA library was then obtained through gel excision and recovery.


title: dmPGE2+APAP_6h_3 [miRNA]


title: dmPGE2+APAP_6h_2 [miRNA]


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL05

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL05_DpGC41_50_S11

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.

value: Neighboring


title: dmPGE2+APAP_6h_1 [miRNA]


title: Control_6h_1 [mRNA_Dgaln]


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC31_40_S4_IL06

description:
SMART-Seq v4 3' DE Kit
IL06_DmGC31_40_S4


title: MCF7_input

city: Singapore

first: Vinay

last: Tergaonkar

address: 61 Biopolis Drive,  Proteos

country: Singapore

institute: Institute of Molecular and Cell Biology

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

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.


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL05

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL05_DpGC51_60_S12

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.


title: MCF7_NAIL_ΔNFκB_TNFa_90min_p300_chip

value: TNFa

tag: time

value: p300 (Santa-Cruz, sc-585)


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL06

description:
SMART-Seq v4 3' DE Kit
IL06_DmGC41_50_S5


title: dmPGE2+APAP_6h_2 [mRNA]


title: APAP_6h_2 [miRNA]


title: Control_6h_3 [miRNA]

value: Saline


title: APAP_6h_1 [miRNA]


title: MCF7_NAIL_ΔNFκB_TNFa_45min_p300_chip


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL05

description:
SMART-Seq v4 3' DE Kit
IL05_DmGC51_60_S6


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL05

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL05_DpGC21_30_S9


title: Control_6h_2 [miRNA]


title: Control_6h_1 [miRNA]


title: MCF7_NAIL_ΔNFκB_TNFa_0min_p300_chip

value: control


title: MCF7_NAIL_WT_TNFa_45min_p300_chip


title: dmPGE2+APAP_6h_3 [circRNA]

data_processing:
The raw sequencing data encompasses a minor fraction of reads with sequencing joints or suboptimal sequencing quality. Reads with adapters should be discarded. Additionally, reads with an N ratio exceeding 0.002 (where N signifies an undetermined base) should be eliminated. If the proportion of low-quality bases in a single-end read surpasses 50% of the read's length, the corresponding paired reads should be removed.
Trimmed sequence reads were mapped to GrCM38/mm10 using Hisat2.
Read count extraction and normalization were performed using Hisat2.
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: tab-delimited text files include TPM values for each Sample

extract_protocol:
Liver tissue of C57 BL6 mice were treated with liquid nitrogen immediately and maintained at -80C. 1 ug of total RNA was used for the construction of sequencing libraries.
Ribosomal RNA is extracted from the total RNA, which is subsequently fragmented into short sequences ranging from 250-300 bp. Utilizing this fragmented RNA as a template and employing random oligonucleotides as primers, the first strand of cDNA is synthesized. Following this, the RNA strand is degraded using RNase H. The second strand of cDNA is then synthesized using dNTPs (dUTP, dATP, dGTP, and dCTP) within the DNA polymerase I system. The purified double-stranded cDNA undergoes processes such as end repair, A-tailed addition, and sequencing linkages. AMPure XP beads are employed to isolate cDNA fragments approximately 350-400 bp in length. The USER enzyme is used to degrade the second cDNA strand containing U, culminating in PCR amplification to produce the final library.


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL05

description:
SMART-Seq v4 3' DE Kit
IL05_DmGC41_50_S5


title: MCF7_NAIL_WT_TNFa_90min_p300_chip


title: dmPGE2+APAP_6h_1 [circRNA]


title: APAP_6h_3 [circRNA]


title: MCF7_NAIL_WT_TNFa_0min_p300_chip


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC31_40_S4_IL05

description:
SMART-Seq v4 3' DE Kit
IL05_DmGC31_40_S4


title: Control_6h_2 [circRNA]


title: dmPGE2+APAP_6h_2 [circRNA]


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC41_50_S11_IL04

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL04_DpGC41_50_S11


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL05

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL05_DpGC31_40_S10


title: APAP_6h_1 [circRNA]


title: APAP_6h_2 [circRNA]


title: Control_6h_3 [circRNA]


title: Library8c RNA-seq rep3

description: RNA-seq performed on reporter gene barcode amplicons

source_name: whole retina

value: whole retina

value: wild-type


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL05

description:
SMART-Seq v4 3' DE Kit
IL05_DmGC21_30_S3


title: MCF7_NAIL_ΔNFκB_TNFa_45min_H3K27Ac_chip

value: H3K27Ac (Millipore, 07-360)


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC51_60_S12_IL04

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL04_DpGC51_60_S12


title: Control_6h_1 [circRNA]

value: Liver


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC21_30_S9_IL04

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL04_DpGC21_30_S9


title: Library8c RNA-seq rep2


title: Library8b RNA-seq rep1


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC51_60_S6_IL04

description:
SMART-Seq v4 3' DE Kit
IL04_DmGC51_60_S6


title: Library8a RNA-seq rep3


title: Library8c RNA-seq rep1


title: MCF7_NAIL_ΔNFκB_TNFa_0min_H3K27Ac_chip


title: Library8a RNA-seq rep2


title: Library8a RNA-seq rep1


title: Library8b RNA-seq rep3


title: Library8b RNA-seq rep2


title: MCF7_NAIL_ΔNFκB_TNFa_90min_H3K27Ac_chip


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC41_50_S5_IL04

description:
SMART-Seq v4 3' DE Kit
IL04_DmGC41_50_S5


title: Day 21 DIO [X9]

city: Hershey

first: Yuka

last: Imamura Kawasawa

state: PA

address: 500 University Dr.

department: College of Medicine, Pharmacology

institute: Penn State University

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

source_name: Bone fracture healing callus (Day 21 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: biological repeats

value: Biol rep 2

tag: disease state

value: diet-induced obese (DIO)

tag: condition

value: Day 21 DIO


title: Day 21 DIO [X70]

value: Bone fracture healing callus (Day 21 post-fracture)

value: Biol rep 3


title: Single granulosa cell (SMART-Seq v4 3' DE Kit) DmGC21_30_S3_IL04

description:
SMART-Seq v4 3' DE Kit
IL04_DmGC21_30_S3

value: LCM


title: MCF7_NAIL_WT_TNFa_45min_H3K27Ac_chip


title: Day 21 DIO [X8]

value: Biol rep 1


title: MCF7_NAIL_WT_TNFa_0min_H3K27Ac_chip


title: Day 14 DIO [X53]

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

value: Day 14 DIO


title: Single granulosa cell (DRaqL - SMART-Seq v4 3' DE Kit) DpGC31_40_S10_IL04

description:
DRaqL - SMART-Seq v4 3' DE Kit
IL04_DpGC31_40_S10

value: Neighboring

value: gene expression


title: MCF7_NAIL_ΔNFκB_TNFa_45min_H3K4Me3_chip

value: TNFa

value: H3K4Me3 (Millipore, 04-745)