title: ChIP ESC Smarcad1KD H3.3

city: Solna

first: Simon

middle: J

last: Elsässer

address: Tomtebodavägen 23A

department: Medical Biochemistry and Biophysics

country: Sweden

institute: Karolinska Institutet

description: Wildtype mouse embryonic stem cell

data_processing:
mapping reads to mm9 using bowtie2/v2.3.5.1 with parameters --fast (>97% alignment rate)
remove duplicates using Picard/v2.20.4 with parameters MarkDuplicates REMOVE_DUPLICATES=TRUE
filter against mm9 blacklist using BEDTools/v2.27.1 with parameters intersect -v -b "blacklist.bed"
generate bigwig file using deepTools/v3.3.2 bamCoverage command with parameters --normalizeUsing RPGC --effectiveGenomeSize 2150570000 -bs 5
Genome_build: mm9
Supplementary_files_format_and_content: Genomic tracks in bigwig format normalized to 1x Genome Coverage

source_name: mESC

growth_protocol: Mouse embryonic stem cells (mESCs) were cultured feeder-free in 0.1% gelatin-coated flasks (Sigma, G1890) under standard conditions (10% CO2, 90% humidity, 37 °C) in KnockOut DMEM (LifeTechnologies, 10829018), 2 mM Alanyl-glutamine (Sigma, G8541), 0.1 mM non-essential amino acids (Sigma, M7145) 15% fetal bovine serum (FBS) (Sigma, F7524), 0.1 mM β-mercaptoethanol (Sigma, M3148), ESGRO Leukemia Inhibitory Factor (LIF) (Millipore, ESG1107), Penicillin-Streptomycin (Sigma, P4333) and 2i;1 μM MEK inhibitor PD0325901 (Sigma, PZ0162) and 3 μM GSK3 inhibitor CHIR99021 (Sigma, SML1046). Generation of H3.3 Knock Out has been previously described (Banaszynski et al. 2013). ESCs were routinely tested for mycoplasma.

extract_protocol: ATAC-seq was performed as described with minor modification (Corces et al. 2017).  Tn5 transposase was produced in-house as described (Picelli et al. 2014) . Briefly, 100’000 cells were centrifuged at 600g for 5 min at 4°C. The pellet were resuspended in 200ul lysis buffer (10mM Tris-HCl pH 7.4, 10mM NaCl, 3mM MgCl2, 0.1% NP40, 0.1% Tween-20, 0.01% digitonin) and incubated on ice for 3min. After incubation, cells were centrifuged at 600g for 3min in the pre-chilled swinging-bucket rotor. The nuclei pellets were washed with wash buffer (10mM Tris-HCl pH 7.4, 10mM NaCl, 3mM MgCl2, 0.01% digitonin). After washing, the nuclei were pelleted at 600g for 3min, resuspended in 50ul transposase mixture with final concentration of 100nM in-house Tn5 and incubated at 37C for 30min with shaking. After transposition, the mixtures were digested with proteinase K at 63°C for 1h, followed by phenol-chloroform purification. Illumina sequencing libraries were prepared based as described (Corces et al. 2017). ChIP-Seq was performed as described in the associated publication. Briefly, cells were cross-linked with 1% methanol-free paraformaldehyde (PFA) in medium for 10 min at room temperature, quenched by Tris-HCl (7.8) and washed twice with ice-cold PBS. 2x1e6 cell aliquots were resuspended in 50 uL Sonication Buffer 1 and chromatin fragmented by using a Bioruptor Plus sonication device (Diagenode) for 80 minutes (30 seconds on/30 seconds off, high) at 4°C to approximately 150-300 bp. Samples were diluted further to a final volume 300 uL with Lysis Buffer. 25µl SureBeads Protein A per IP were incubated with 4 ug a-H3.3 (Millipore, 09-838) or 2.5 ug a-H3K9me3 (Abcam, ab8898). 300-450 uL (2-3 Mio cells) sonicated and cleared chromatin were added to the pre-coupled magnetic beads overnight at 4°C with rotation. Next, the beads were washed each time twice, 5 min each with 1 mL RIPA, 1 mL RIPA high, 1 mL LiCl Buffer and a quick wash with 1 mL TE buffer, then resuspended in 100 uL ChIP elution buffer containing 0.25 mg/mL Proteinase K  and eluted overnight at 63°C. The DNA was isolated using 1x SPRI beads. ChIP libraries were prepared using NEBNext Ultra II DNA library preparation kit (New England Biolabs, E7645) and amplified using NEBNext® Multiplex Oligos for Illumina (New England Biolabs, E7335) according to manufacturer’s protocol.

treatment_protocol:
Approximately 5x10e5 ESCs in single cell suspension in knockout medium without antibiotics were transfected with 10 uL Lipofectamine RNAiMax (Invitrogen, 13778030) and 100 nM esiRNA SMARCAD1 (Sigma, EMU209081), and plated in 6 well-plates for 48 h.
Rescue cell lines: H3.3 knockout (H3.3KO) cells were described previously (Banaszynski et al. 2013; Elsässer et al. 2015). Rescue cell lines expressing wildtype H3.3 (H3.3KO+H3.3), H3.2 (H3.3KO+H3.2) or H3.3 L126A I130A (H3.3KO+H3.3LI->AA) under EF1 promoter were generated by PiggyBac transgenesis. The coding sequences of the respective histones were cloned into a general-purpose PiggyBac vector containing a EF1 promoter, multiple cloning site, IRES and Blasticidin resistance gene  (Elsässer 2018) . 100 000 H3.3KO cells in single cell suspension were transfected with 3 μL TransIT-X2 (Mirus, MIR 6004) with 1 μg total Plasmid DNA: PiggyBac targeting vector with histone gene of interest (GOI) and Super PiggyBac Transposase Plasmid pPBase (PBT) (PB200PA, System Bioscience Inc) at a ratio of 4:1 respectively and plated in a 12-well plate. After 48 hours, cells were split, allowed to recover overnight and subsequently selected with 400-800 μg/ml Blasticidin (Invivogen, ant-bl-10p) for seven days. Experiments were performed at early passages.

value: mouse embryonic stem cells

tag: assay

value: a-H3.3 ChIP-Seq (Millipore, 09-838)


title: ChIP ESC Smarcad1KD H3K9me3

status: Public on Aug 28 2020

type: SRA

description: Wildtype mouse embryonic stem cell

library_source: genomic

molecule: genomic DNA

organism: Mus musculus

tag: strain

tag: cell type

value: a-H3K9me3 ChIP-Seq (Abcam, ab8898)


title: ChIP ESC Smarcad1KD Input

description: Wildtype mouse embryonic stem cell

value: ChIP-Seq


title: ChIP ESC WT Input

description: Wildtype mouse embryonic stem cell

treatment_protocol:
none
Rescue cell lines: H3.3 knockout (H3.3KO) cells were described previously (Banaszynski et al. 2013; Elsässer et al. 2015). Rescue cell lines expressing wildtype H3.3 (H3.3KO+H3.3), H3.2 (H3.3KO+H3.2) or H3.3 L126A I130A (H3.3KO+H3.3LI->AA) under EF1 promoter were generated by PiggyBac transgenesis. The coding sequences of the respective histones were cloned into a general-purpose PiggyBac vector containing a EF1 promoter, multiple cloning site, IRES and Blasticidin resistance gene  (Elsässer 2018) . 100 000 H3.3KO cells in single cell suspension were transfected with 3 μL TransIT-X2 (Mirus, MIR 6004) with 1 μg total Plasmid DNA: PiggyBac targeting vector with histone gene of interest (GOI) and Super PiggyBac Transposase Plasmid pPBase (PBT) (PB200PA, System Bioscience Inc) at a ratio of 4:1 respectively and plated in a 12-well plate. After 48 hours, cells were split, allowed to recover overnight and subsequently selected with 400-800 μg/ml Blasticidin (Invivogen, ant-bl-10p) for seven days. Experiments were performed at early passages.

value: ChIP-Seq (Input)


title: ChIP ESC WT H3K9me3

description: Wildtype mouse embryonic stem cell


title: Control primary hepatocytes

type: RNA

city: Fukuoka

first: Lixiang

last: Wang

state: Fukuoka

address: 3-1-1, Maidashi, Higashi-ku

department: Department of Medicine and Bioregulatory Science

country: Japan

institute: Graduate School of Medical Sciences, Kyushu University

hyb_protocol: cRNA was hybridized to a 60K 60-mer oligomicroarray (SurePrint G3 Mouse Gene Expression Microarray 8x60K v2 ; Agilent Technologies) according to the manufacturer's instructions.

scan_protocol: The hybridized microarray slides were scanned using an Agilent scanner. The relative hybridization intensities and background hybridization values were calculated using Feature Extraction Software version 9.5.1.1 (Agilent Technologies).

data_processing: The scanned images were analyzed with Feature Extraction Software 9.5.1.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. The raw signal intensities and flags for each probe were calculated from the hybridization intensities and spot information according to the procedures recommended by Agilent Technologies using the Flag criteria in the GeneSpring Software. The raw signal intensities of all samples were  normalized by the quantile algorithm with the Bioconductor.

molecule: total RNA

source_name: primary hepatocytes, wild-type

label_protocol: cRNA was amplified and labelled using a Low input Quick Amp Labelling Kit (Agilent Technologies).

growth_protocol: Miice were anesthetized with pentobarbital sodium (50 mg/kg body weight), and liver tissues were perfused with pre-warmed Hank’s balanced salt solution (HBSS) supplemented with 0.5 mM EGTA and 10 mM HEPES for 5 minutes, followed with digestion buffer [low glucose Dulbecco’s modified Eagle’s medium (1000mg/L) supplemented with 0.8 mg/ml collagenase type 1 (Worthington Biochemical Corporation, Lakewood, NJ, USA)]. Single cell suspensions were filtered through a 70-μm cell strainer (BD Falcon, Bedford, MA, USA) and centrifuged at 50 g for 1 min, the cell supernatants were collected separately for the preparation of non-parenchymal cells as described below. The hepatocytes in the pellet were washed twice and suspended in high glucose Dulbecco’s modified Eagle’s medium (4500mg/L) supplemented with 1μM insulin, 2mM L-glutamine, 10 IU/mL penicillin, 10 IU/mL streptomycin and 10% fetal bovine serum and seeded at 300,000 cells/well on 6-well plastic plates coated with collagen type I (Cellmatrix type I-P, Nitta Gelatin, Osaka, Japan). After 4 hours incubation in a 5% CO2 incubator at 37 °C, the culture medium was changed to discard floating unattached cells. We conducted  microarray analysis with primary hepatocyte harvested 24 hours after seeding.

extract_protocol: The total RNA was isolated from cells using TRIzol Reagent (nitrogen) and purified using SV Total RNA Isolation System (Promega) according to the manufacturer's instructions.

tag: strain background

tag: genotype/variation

value: wild type control

tag: tissue

value: primary hepatocytes


title: Drp1LiKO primary hepatocytes

source_name: primary hepatocytes, knockout


title: KPC PDA tumour 2 CD105pos CAFs

city: MANCHESTER

first: Alexander

last: Kononov

address: Oglesby Building, 555 Wilmslow Rd,

country: United Kingdom

institute: Cancer Research UK Manchester Institute

library_source: transcriptomic

data_processing:
Sequencing reads (76bp, paired-end) were aligned to the mouse reference genome GRCm38.mm10 using STAR aligner version 2.5.1b
Gene annotation was taken from Ensembl build 92
Read counts were determined by using the featureCounts function from Bioconductor package Rsubread version 1.28.1
Expression of a gene was calculated as Transcipts Per Kilobase Million (TPM) using standard calculation
Genome_build: mm10
Supplementary_files_format_and_content: Matrix table with TPM values for every gene and every sample

source_name: KPC GEMM mouse Pdx1-Cre; KrasLSL-G12D/+; Trp53LSL-R172H/+ (KPC)

extract_protocol:
FACS isolated CAFs were centrifuged at 400g for 5 min, carefully aspirated and cells lysed in RLT buffer. Lysis and subsequent RNA isolation was conudcted as per instructions ffor the Qiagen RNeasy micro kits
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Spontaneous pancreatic tumours


title: ADO.045-PI

type: protein

city: Siena

first: Erika

last: Bartolini

state: Italy

address: Via Fiorentina 1

department: AIMB

country: Italy

institute: GSK Vaccines

description:
NCT00661713
FS00007559 Area1-ADO.045-PI

hyb_protocol: Protein arrays were first blocked 60 min in Blocking Buffer (Block it, Arrayit Sunnyvale, CA 94085, USA). The array was then incubated 1h a room temperature with human sera diluted 1:500 for anti-human IgG-specific analysis. After washing with 0.1% Tween 20 in PBS buffer (PBST), arrays were incubated with an AlexaFluor®647-labelled anti-human IgG, fab specific secondary antibody (1:800 – Jackson Immunoresearch) at RT for 1 h to detect interactions. Slides were washed again as before, then rinsed in sterile water and dried.

scan_protocol: Fluorescence signals were detected by using a Powerscanner (Tecan Trading AG, Switzerland) and the 16-bit images were generated with Powerscanner software at 10 µm per pixel resolution.

data_processing: Images were processed using ImaGene 9.0 software (Biodiscovery Inc, CA, USA). Elaboration and analysis of image raw fluorescence Intensity (FI) data was performed using in-house developed software and R scripts. For each sample, the mean fluorescence intensity (MFI) of replicated spots was determined, after subtraction of the background value surrounding each spot.

label: AlexaFluor®647-labelled anti-human secondary antibody

molecule: protein

organism: Homo sapiens

label_protocol: The secondary antibody used for arrays was from Jackson Immunoresearch, (872 W Baltimore Pike, West Grove, PA 19390 USA). It was conjugated to the Alexafluor dye (Alexa 647) by the company.

extract_protocol: Human sera collected from different clinical trials

tag: subject group

value: Adolescent

value: blood

tag: tissue subtype

value: serum-PI


title: ESC WT Smarcad1KD batch2

description: Wildtype mouse embryonic stem treated with Smarcad1 esiRNA (Sigma, EMU209081) for 48h

value: Omni-ATAC


title: ESC WT batch2

description: Wildtype mouse embryonic stem cell littermate control for H3.3 knockout


title: ESC H3.3KO +H3.3 Smarcad1KD batch2

description: Rescue cell lines expressing wildtype histone H3.3 under EF1 promoter generated from H3.3KO by PiggyBac transgenesis. Treated with Smarcad1 esiRNA (Sigma, EMU209081) for 48h


title: ChIP ESC WT H3.3

description: Wildtype mouse embryonic stem cell


title: ADO.043-PI

description:
NCT00661713
FS00006765-ADO.043-PI


title: ADO.042-PI

description:
NCT00661713
FS00005219-ADO.042-PI


title: ADO.044-PI

description:
NCT00661713
FS00007559 Area2-ADO.044-PI


title: ADO.041-PI

description:
NCT00661713
FS00007558 Area1-ADO.041-PI


title: ADO.039-PI

description:
NCT00661713
FS00007551 Area1-ADO.039-PI


title: ADO.040-PI

description:
NCT00661713
FS00006267-ADO.040-PI


title: ADO.046-PI

description:
NCT00661713
FS00007558 Area2-ADO.046-PI


title: ESC H3.3KO batch2

description: Histone H3.3 knockout (H3.3KO) cells  described previously (Banaszynski et al. 2013; Elsässer et al. 2015).


title: ESC H3.3KO +H3.3 batch2

description: Rescue cell lines expressing wildtype histone H3.3 under EF1 promoter generated from H3.3KO by PiggyBac transgenesis


title: ESC H3.3KO Smarcad1KD batch2

description: Histone H3.3 knockout (H3.3KO) cells treated with Smarcad1 esiRNA (Sigma, EMU209081) for 48h


title: ADO.036-PI

description:
NCT00661713
FS00007551 Area2-ADO.036-PI


title: ADO.035-PI

description:
NCT00661713
FS00007622-ADO.035-PI


title: ADO.037-PI

description:
NCT00661713
FS00007555 Area1-ADO.037-PI


title: ESC WT ATRXKD batch2

description: Wildtype mouse embryonic stem treated with Atrx siRNA (Thermo AM16704) for 48h

treatment_protocol:
Approximately 5x10e5 ESCs in single cell suspension in knockout medium without antibiotics were transfected with 10 uL Lipofectamine RNAiMax (Invitrogen, 13778030) and 100 nM Atrx siRNA (Thermo AM16704), and plated in 6 well-plates for 48 h.
Rescue cell lines: H3.3 knockout (H3.3KO) cells were described previously (Banaszynski et al. 2013; Elsässer et al. 2015). Rescue cell lines expressing wildtype H3.3 (H3.3KO+H3.3), H3.2 (H3.3KO+H3.2) or H3.3 L126A I130A (H3.3KO+H3.3LI->AA) under EF1 promoter were generated by PiggyBac transgenesis. The coding sequences of the respective histones were cloned into a general-purpose PiggyBac vector containing a EF1 promoter, multiple cloning site, IRES and Blasticidin resistance gene  (Elsässer 2018) . 100 000 H3.3KO cells in single cell suspension were transfected with 3 μL TransIT-X2 (Mirus, MIR 6004) with 1 μg total Plasmid DNA: PiggyBac targeting vector with histone gene of interest (GOI) and Super PiggyBac Transposase Plasmid pPBase (PBT) (PB200PA, System Bioscience Inc) at a ratio of 4:1 respectively and plated in a 12-well plate. After 48 hours, cells were split, allowed to recover overnight and subsequently selected with 400-800 μg/ml Blasticidin (Invivogen, ant-bl-10p) for seven days. Experiments were performed at early passages.


title: ADO.032-PI

description:
NCT00661713
FS00007552 Area1-ADO.032-PI


title: ADO.034-PI

description:
NCT00661713
FS00005222-ADO.034-PI


title: ADO.033-PI

description:
NCT00661713
FS00007555 Area2-ADO.033-PI


title: ADO.030-PI

description:
NCT00661713
FS00007556 Area1-ADO.030-PI


title: ADO.031-PI

description:
NCT00661713
FS00007571-ADO.031-PI


title: ESC H3.3KO Smarcad1KD

treatment_protocol: Rescue cell lines: H3.3 knockout (H3.3KO) cells were described previously (Banaszynski et al. 2013; Elsässer et al. 2015). Rescue cell lines expressing wildtype H3.3 (H3.3KO+H3.3), H3.2 (H3.3KO+H3.2) or H3.3 L126A I130A (H3.3KO+H3.3LI->AA) under EF1 promoter were generated by PiggyBac transgenesis. The coding sequences of the respective histones were cloned into a general-purpose PiggyBac vector containing a EF1 promoter, multiple cloning site, IRES and Blasticidin resistance gene  (Elsässer 2018) . 100 000 H3.3KO cells in single cell suspension were transfected with 3 μL TransIT-X2 (Mirus, MIR 6004) with 1 μg total Plasmid DNA: PiggyBac targeting vector with histone gene of interest (GOI) and Super PiggyBac Transposase Plasmid pPBase (PBT) (PB200PA, System Bioscience Inc) at a ratio of 4:1 respectively and plated in a 12-well plate. After 48 hours, cells were split, allowed to recover overnight and subsequently selected with 400-800 μg/ml Blasticidin (Invivogen, ant-bl-10p) for seven days. Experiments were performed at early passages.

tag: treatment

value: Approximately 5x105 ESCs in single cell suspension in knockout medium without antibiotics were transfected with 10 uL Lipofectamine RNAiMax (Invitrogen, 13778030) and 100 nM esiRNA SMARCAD1 (Sigma, EMU209081), and plated in 6 well-plates for 48 h.

value: H3.3KO Smarcad1KD


title: ADO.029-PI

description:
NCT00661713
FS00007556 Area2-ADO.029-PI


title: ADO.028-PI

description:
NCT00661713
FS00007553 Area1-ADO.028-PI


title: ESC H3.3KO +H3.3LI->AA

description: Rescue cell lines expressing histone H3.3 with mutations L126A I130A under EF1 promoter generated from H3.3KO by PiggyBac transgenesis

value: 3KO +H3.3LI->AA


title: ADO.026-PI

description:
NCT00661713
FS00006349-ADO.026-PI


title: ESC H3.3KO


title: Jukat-EZH2-KO2-rep3

city: London

first: Theresa

middle: Elizabeth

last: Leon

address: 72 Huntley St, Camden Town

department: Haematology

institute: UCL Cancer Institute

data_processing:
RNA-Seq Alignment: Basespace RNA-Seq Alignment Application, Version: 1.1.0
STAR Aligner: 2.5.0b
Cufflinks: 2.2.1
Bedtools: 2.7.0
Fragments Per Kilobase of sequence per Million mapped reads (FPKM) files were obtained from Basespace, cufflinks folder.
Genome_build: UCSC hg19
Supplementary_files_format_and_content: TSV

source_name: Jurkat cell line

growth_protocol: Cells were in culture in complete RPMI (10% FBS, 1% Pen/Strep).

extract_protocol:
Exponentially growing cells were washed and RNA was extracted using RNeasy Mini Kit (Qiagen).
Samples were processed using the KAPA mRNA HyperPrep Kit (Cat. KK8580) according to manufacturer’s instructions, using 500ng of total RNA for the library preparation.

value: T lymphocyte

tag: ezh2 status

value: KO


title: ESC H3.3KO +H3.2

description: Rescue cell lines expressing wildtype histone H3.2 under EF1  generated from H3.3KO by PiggyBac transgenesis

value: mouse embryonic stem cells


title: ESC H3.3KO +H3.3


title: Jukat-EZH2-KO2-rep2


title: Jukat-EZH2-KO2-rep1


title: ADO.022-PI

description:
NCT00661713
FS00007575 Area1-ADO.022-PI


title: ADO.023-PI

description:
NCT00661713
FS00007552 Area2-ADO.023-PI


title: ADO.025-PI

description:
NCT00661713
FS00007553 Area2-ADO.025-PI


title: ADO.024-PI

description:
NCT00661713
FS00006275-ADO.024-PI


title: ADO.018-PI

description:
NCT00661713
FS00007614-ADO.018-PI


title: ADO.020-PI

description:
NCT00661713
FS00007575 Area2-ADO.020-PI


title: ADO.019-PI

description:
NCT00661713
FS00007820 Area1-ADO.019-PI


title: Jukat-EZH2-KO1-rep3


title: ADO.021-PI

description:
NCT00661713
FS00007597 Area1-ADO.021-PI


title: ADO.015-PI

description:
NCT00661713
FS00007820 Area2-ADO.015-PI


title: Jukat-EZH2-KO1-rep2


title: ADO.017-PI

description:
NCT00661713
FS00007618-ADO.017-PI


title: ADO.013-PI

description:
NCT00661713
FS00007821 Area1-ADO.013-PI


title: ADO.014-PI

description:
NCT00661713
FS00007597 Area2-ADO.014-PI


title: ADO.012-PI

description:
NCT00661713
FS00007821 Area2-ADO.012-PI


title: ADO.027-PI

description:
NCT00661713
FS00005225-ADO.027-PI


title: Jurkat-EZH2-WT-rep3

value: WT


title: Jukat-EZH2-KO1-rep1


title: ADO.008-PI

description:
NCT00661713
FS00007819 Area2-ADO.008-PI


title: ADO.011-PI

description:
NCT00661713
FS00007818 Area1-ADO.011-PI


title: ADO.010-PI

description:
NCT00661713
FS00007818 Area2-ADO.010-PI


title: ADO.009-PI

description:
NCT00661713
FS00007819 Area1-ADO.009-PI


title: Jurkat-EZH2-WT-rep2


title: ADO.007-PI

description:
NCT00661713
FS00007815 Area1-ADO.007-PI


title: ADO.006-PI

description:
NCT00661713
FS00007606-ADO.006-PI


title: Jurkat-EZH2-WT-rep1


title: ADO.004-PI

description:
NCT00661713
FS00007815 Area2-ADO.004-PI


title: ADO.005-PI

description:
NCT00661713
FS00007602-ADO.005-PI


title: ADO.003-PI

description:
NCT00661713
FS00007561 Area1-ADO.003-PI


title: ADO.002-PI

description:
NCT00661713
FS00007569-ADO.002-PI


title: ADU.028-PI

description:
NCT00560313
FS00005306-ADU.028-PI

value: Adult


title: ADO.001-PI

description:
NCT00661713
FS00007609-ADO.001-PI


title: ADU.030-PI

description:
NCT00560313
FS00005312-ADU.030-PI


title: ADU.027-PI

description:
NCT00560313
FS00007698-ADU.027-PI


title: ADU.029-PI

description:
NCT00560313
FS00005309-ADU.029-PI


title: ADU.023-PI

description:
NCT00560313
FS00007684-ADU.023-PI


title: ADU.024-PI

description:
NCT00560313
FS00007688-ADU.024-PI


title: ratA25U0_ratA25dmso_r2

city: Düsseldorf

first: Wasco

last: Wruck

address: Moorenstr. 5

country: Germany

institute: Universitätsklinikum Düsseldorf

description: always two replica with a dye swap

hyb_protocol: Microarray hybridisation and washing was performed according to the same manual on Slide BoosterTM hybridisation station (Advalytix, Germany).

scan_protocol: Microarray images were obtained on an Agilent G2565AA scanner at 10 µm resolution.

data_processing: Image analysis was carried out using the ImaGene software (BioDiscovery, Inc., El Segundo, USA). For each target miRNA, two independent hybridizations were done by inverting the Alexa3 and Alexa5 fluorochromes in the labelling reactions. Background subtraction and normalisation was done with GeneSite software (BioDiscovery, Inc, El Segundo, USA) by global scaling normalization, the median to set the average to 1 for all arrays.. For statistical analysis BRB-ArrayTools software, developed by Dr. Richard Simon and Development Team was used

organism: Rattus norvegicus

label_protocol: 100 ng of small RNA fraction were labelled using the NCODETM miRNA labeling system (Invitrogen, CA, USA) according to the manufacturer’s instructions.

growth_protocol: The HRASG12V-transformed human embryonal kidney cells (HA1ER) and their immortalized origin cell line HA1EB were described by Hahn et al. (32) . They were cultivated in Minimum Essential Medium Eagle, Alpha Modification (MEM Alpha) from Sigma, supplemented with 10% IFS, 2 mM ultraglutamine, 1%  penicillin/streptomycin,  0,1mg/ml hygromycin and  0,5µg/ml puromycin.  G418 was freshly added to a final concentration of 0,4 mg/ml . ROSE199, rat ovary epithelial cell line and their KRAS transformed derivative cell line ROSEA25 were decribed in Tchernitsa, 2000.

extract_protocol: Total RNA was isolated from cell lines using the mirVanaTM miRNA Isolation Kit from Ambion® (Austin, TX, USA) according to the maufacturer’s instructions. In order to avoid cross-labeling of premature miRNA we used the gel purification option. Sunseqneutly the quality of the small RNA fraction was controlled on a 15% PAAG gel stained with ethidium bromide.

treatment_protocol: Induble RAS studies were done in HEK cell lines transfected with a constuct bearing Ha-RAS under ER promoter (33) Cells were treated with …µM of tamoxifen before RNA was extracted. U0126 MEK inhibitor was dissolved in DMSO and used in 10µM final concentration for 48h before RNA was extracted. LY294002 PI3K inhibitor was dissolved in ethanol and used at final concentration of 50µM.

tag: cell line

value: KRAS transformed ROSEA25

tag: tissue/cell type

value: ovarium surface epithelium cells

value: treated with DMSO for 48h

value: treated with U0126 for 48h


title: ADU.025-PI

description:
NCT00560313
FS00007686-ADU.025-PI


title: ratA25LY_ratA25Eth_r2

description: always two replica with a dye swap

value: treated with ethanol for 48h

value: treated with LY294002 for 48h


title: ADU.021-PI

description:
NCT00560313
FS00005381-ADU.021-PI

value: blood


title: ADU.022-PI

description:
NCT00560313
FS00005385-ADU.022-PI

value: serum-PI


title: ADU.020-PI

description:
NCT00560313
FS00005379-ADU.020-PI


title: huHAEB_huHAER_r2

description: always two replica with a dye swap

source_name: huHAER

value: HRASG12V-transformed human embryonal kidney cells (HA1ER)

value: human embryonic kidney cells

source_name: huHAEB


title: ADU.026-PI

description:
NCT00560313
FS00007690-ADU.026-PI


title: huHAEBEth_huHAEBTam_r2

description: always two replica with a dye swap

value: treated with tamoxifen for 48h


title: huHAERU0_huHAERdmso_r2

description: always two replica with a dye swap


title: ADU.019-PI

description:
NCT00560313
FS00005377-ADU.019-PI


title: huHAERU0_huHAERdmso_r1

description: always two replica with a dye swap


title: huHAEBEth_huHAEBTam_r1

description: always two replica with a dye swap


title: ADU.017-PI

description:
NCT00560313
FS00007538-ADU.017-PI


title: ADU.018-PI

description:
NCT00560313
FS00005378-ADU.018-PI


title: rat199_ratA25_r2

description: always two replica with a dye swap


title: ADU.015-PI

description:
NCT00560313
FS00005367-ADU.015-PI


title: ADU.016-PI

description:
NCT00560313
FS00005373-ADU.016-PI


title: huHAEB_huHAER_r1

description: always two replica with a dye swap


title: ADU.013-PI

description:
NCT00560313
FS00007540-ADU.013-PI


title: ADU.010-PI

description:
NCT00560313
FS00007530-ADU.010-PI


title: ADU.014-PI

description:
NCT00560313
FS00005368-ADU.014-PI


title: ADU.009-PI

description:
NCT00560313
FS00007524-ADU.009-PI


title: ADU.012-PI

description:
NCT00560313
FS00007536-ADU.012-PI


title: ADU.011-PI

description:
NCT00560313
FS00007528-ADU.011-PI


title: TSKI4R cells, biological rep2

city: Memphis

first: Amy

last: Abell

state: TN

address: 3774 Walker Ave

department: Biological Sciences

country: USA

institute: The University of Memphis

hyb_protocol: Following fragmentation, 3.5 ug of cRNA were hybridized according to manufacturer's instruction. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.

scan_protocol: GeneChips were scanned using Affymetrix GeneChiP Scanner 3000 7G

data_processing: GCRMA normalization; data not provided by submitter

label: biotin

source_name: Trophoblast stem cells

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 100 ng total RNA (Manual Target Preparation for GeneChip™ Whole Transcript (WT) Expression Arrays).

growth_protocol: Cells were cultured in the absence of feeders in 30% TS media (RPMI 1640, 20% fetal bovine serum, 1% penicillin and streptomycin, 1% L-glutamine, 1% sodium pyruvate, and 100 µM β-mercaptoethanol) and 70% MEF-conditioned TS cell media. For maintenance of the stem cell state, TS medium was supplemented with FGF4 (37.5 ng/ml) and Heparin (1 mg/ml

extract_protocol: Total cellular RNA isolated using the RNeasy Plus Minikit (Qiagen) according to the manufacturer's instructions.

value: Trophoblast stem cells

tag: genotype

value: MAP3K4 Knockin K1361R

tag: vector

value: REL expressing lentiviral vector


title: ratA25U0_ratA25dmso_r1

description: always two replica with a dye swap


title: TSKI4R cells, biological rep1


title: TSKI4R cells, biological rep3


title: ratA25LY_ratA25Eth_r1

description: always two replica with a dye swap

value: treated with Ethanol for 48h


title: TSKI4 cells, biological rep2

value: pLKO control  lentiviral vector


title: ADU.008-PI

description:
NCT00560313
FS00007704-ADU.008-PI

value: Adult


title: TSKI4 cells, biological rep3


title: ADU.007-PI

description:
NCT00560313
FS00007706-ADU.007-PI


title: INF.067

description:
NCT00944034
FS00006632-INF.067

source_name: vaccinated_Infants_serum-post IV 4CmenB doses

value: Infants

value: serum-post IV 4CmenB doses


title: ADU.003-PI

description:
NCT00560313
FS00007700-ADU.003-PI


title: ADU.002-PI

description:
NCT00560313
FS00005121-ADU.002-PI


title: ADU.006-PI

description:
NCT00560313
FS00007702-ADU.006-PI


title: rat199_ratA25_r1

description: always two replica with a dye swap

value: ovarium surface epithelium cells

value: KRAS transformed ROSEA25


title: INF.068

description:
NCT00944034
FS00006242-INF.068


title: ADU.001-PI

description:
NCT00560313
FS00005118-ADU.001-PI


title: INF.069

description:
NCT00944034
FS00006548-INF.069


title: miRNA sequencing of iBAT 3

city: New York

first: Sean

last: O'Connor

state: NY

address: 1230 York Ave

department: Laboratory of Molecular Metabolism

institute: Rockefeller University

data_processing:
Raw counts per transcript were determined by pseudo alignment tool Kallisto (version: 0.45.0) to mouse index GRCm38 (version: 10)
Per gene abundance measurements were calculated for each sample in R (version: 3.6.1) using package TxImport (version: 1.14.2)
Normalized abundance and log2FoldChange measurements were calculated using DESeq2 output, method "apelgm" (version: 1.26.0)
Genome_build: mm10
Supplementary_files_format_and_content: DEseq normalized counts of miRNA seq.csv
Supplementary_files_format_and_content: DEseq normalized counts of mRNA seq.csv

source_name: Mouse whole adipose tissue depot

growth_protocol: For miRNAseq, total RNA was extracted from fat pads of 13-week old C57BL/6J male mice housed at room temperature and from inguinal fat pads of 13-week old mice housed at 4° C for 1 week prior to sacrifice. For mRNAseq, iBAT and eWAT tissue was dissected from 15-week old C57BL/6J male mice and immediately flash frozen in liquid nitrogen

extract_protocol:
RNA was extracted from tissues and cells using a TRIzol (Invitrogen)/chloroform extraction, followed by purification with RNeasy mini kits (Qiagen). To retain small RNAs during RNA purification, 1.5X volumes of 100% ethanol was used to precipitate RNA prior to column loading and RW1 buffer was substituted with RWT (Qiagen).
For miRNA, The TruSeq Small RNA Library Prep Kit (Illumina) was used to prepare libraries for sequencing, which was performed by LC Sciences. For mRNA, the library was prepared using the Illumina TruSeq Stranded Total RNA Library Prep Kit, and sequencing was performed on an Illumina NextSeq 500 sequencer.

treatment_protocol: No treatment applied to adipose tissue depots

value: iBAT


title: TSKI4 cells, biological rep1


title: INF.065

description:
NCT00944034
FS00006634-INF.065


title: INF.066

description:
NCT00944034
FS00006631-INF.066


title: miRNA sequencing of iBAT 2


title: ADU.004-PI

description:
NCT00560313
FS00007531-ADU.004-PI


title: ADU.005-PI

description:
NCT00560313
FS00007699-ADU.005-PI


title: INF.064

description:
NCT00944034
FS00006638-INF.064


title: miRNA sequencing of iBAT 1


title: miRNA sequencing of cWAT 3

value: cWAT


title: INF.062

description:
NCT00944034
FS00006516-INF.062

value: blood


title: INF.060

description:
NCT00944034
FS00006543-INF.060


title: INF.059

description:
NCT00944034
FS00006006-INF.059


title: INF.057

description:
NCT00944034
FS00006562-INF.057


title: INF.058

description:
NCT00944034
FS00006558-INF.058


title: INF.063

description:
NCT00944034
FS00006637-INF.063


title: INF.061

description:
NCT00944034
FS00006636-INF.061


title: miRNA sequencing of cWAT 2


title: INF.056

description:
NCT00944034
FS00006540-INF.056


title: INF.054

description:
NCT00944034
FS00006555-INF.054


title: H, biological rep4

city: Campinas

first: Cristiane

middle: de Souza

last: Rocha

state: São Paulo

address: Av Tessália Vieira de Camargo, 126 Cidade Universitária "Zeferino Vaz"

department: Medical Genetics

country: Brazil

institute: Unicamp

description: Gene Expression of Healthy subjects, PBMC

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45oC on Gene Chips U133 Plus 2.0 (Affymetrix) arrays. GeneChips were washed and stained in the Affymetrix Fluidics Station.

scan_protocol: The U133 Plus 2.0 arrays were scanned twice using the GeneChip Scanner 3000 7G (Affymetrix Inc., Santa Clara, CA, USA)

data_processing: The raw CEL files were processed using the RMA method, as implemented in the Affy 3 package. The workflow performs background correction, quantile normalization and summarization via median polish. These log2-expression quantities resulted from the Robust Multi-array Average (RMA) method were further processed by the RankProd package.

source_name: Healthy subjects, PBMC

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from Five hundred nanograms of RNA as initial input for each sample in GeneChip 3' IVT Kit protocol  (P/N 702646 Rev. 8).

extract_protocol: Peripheral venous blood collected from each subject was immediately submitted to centrifugation on a Ficoll-Paque PLUS (GE Healthcare Life Sciences, Oslo, Norway) density gradient and consecutive washing with saline (NaCl 0.9%) to isolate peripheral blood mononuclear cells (PBMCs). Total RNA was extracted using the Trizol® reagent (Invitrogen, Rockville, MD, USA) according to manufacturer's instructions. Samples containing total RNA were purified by RNeasy Protection Mini Kit (Qiagen, Hilden, Germany).

value: peripheral blood mononuclear cells (PBMC)

tag: Sex

value: Female

tag: age (years)

tag: fasting plasma glucose

tag: total cholesterol

tag: triglycerides


title: H, biological rep6

value: Male


title: INF.053

description:
NCT00944034
FS00006546-INF.053


title: miRNA sequencing of cWAT 1

tag: strain


title: P, biological rep6

description: Gene Expression of Periodontitis, PBMC

source_name: Periodontitis, PBMC


title: miRNA sequencing of eWAT 2

value: eWAT


title: miRNA sequencing of eWAT 3


title: INF.052

description:
NCT00944034
FS00006545-INF.052


title: INF.051

description:
NCT00944034
FS00006554-INF.051


title: INF.050

description:
NCT00944034
FS00006552-INF.050


title: miRNA sequencing of eWAT 1


title: H, biological rep1


title: INF.055

description:
NCT00944034
FS00006556-INF.055


title: miRNA sequencing of mWAT 3

value: mWAT


title: H, biological rep3


title: P, biological rep5

description: Gene Expression of Periodontitis, PBMC


title: H, biological rep5


title: H, biological rep2


title: P, biological rep4

description: Gene Expression of Periodontitis, PBMC


title: P, biological rep3

description: Gene Expression of Periodontitis, PBMC


title: INF.048

description:
NCT00944034
FS00006692-INF.048


title: INF.046

description:
NCT00944034
FS00006117-INF.046


title: INF.047

description:
NCT00944034
FS00006651-INF.047


title: INF.045

description:
NCT00944034
FS00006007-INF.045


title: miRNA sequencing of mWAT 2


title: P, biological rep1

description: Gene Expression of Periodontitis, PBMC


title: INF.042

description:
NCT00944034
FS00006647-INF.042


title: DL-P, biological rep5

description: Gene Expression of Dyslipidemia and Periodontitis, PBMC

source_name: Dyslipidemia and Periodontitis, PBMC


title: miRNA sequencing of mWAT 1


title: P, biological rep2

description: Gene Expression of Periodontitis, PBMC


title: INF.043

description:
NCT00944034
FS00006652-INF.043


title: INF.049

description:
NCT00944034
FS00006690-INF.049


title: INF.041

description:
NCT00944034
FS00006648-INF.041


title: miRNA sequencing of aWAT 3

value: aWAT


title: INF.040

description:
NCT00944034
FS00006649-INF.040


title: miRNA sequencing of aWAT 1


title: INF.039

description:
NCT00944034
FS00006729-INF.039


title: miRNA sequencing of aWAT 2


title: INF.044

description:
NCT00944034
FS00006009-INF.044


title: INF.035

description:
NCT00944034
FS00006123-INF.035


title: miRNA sequencing of iWAT 3

value: iWAT


title: INF.036

description:
NCT00944034
FS00006726-INF.036


title: INF.037

description:
NCT00944034
FS00006728-INF.037


title: INF.034

description:
NCT00944034
FS00006124-INF.034


title: INF.038

description:
NCT00944034
FS00006005-INF.038


title: miRNA sequencing of iWAT 2


title: INF.033

description:
NCT00944034
FS00006716-INF.033


title: miRNA sequencing of iWAT 1


title: INF.030

description:
NCT00944034
FS00006118-INF.030


title: INF.031

description:
NCT00944034
FS00006122-INF.031


title: INF.029

description:
NCT00944034
FS00006121-INF.029


title: INF.032

description:
NCT00944034
FS00006715-INF.032


title: Res4: D14_N6_CTTGTA

city: Tokyo

first: Hikaru

last: Ito

address: 1-5-45 Yushima, Bunkyo-ku-ku

institute: Tokyo Medical and Dental University

data_processing:
Sequence reads were mapped against the Mus musculus genome assembly (Genome Reference Consortium GRCm38, UCSC mm10) using CLC genomic workbench (QIAGEN).
Read count normalization with trimmed mean of M values normalization, gene expression calculation, and statistical analysis based on generalized linear model were performed by featureCounts and edgeR package.
Genome_build: GRCm38, UCSC mm10

source_name: lateral habenula

extract_protocol:
RNA was purified using NucleoSpin RNA XS (TAKARA Bio)
Sequencing libraries (RNA 100 pg) were generated using SMART-Seq v4 Ultra Low Input RNA Kit and Low Input Library Prep Kit v2 (TAKARA Bio) and validated with the Agilent 2100 bioanalyzer

treatment_protocol: Brains were removed and coronally sliced with a vibratome at 500 µm. Tissue was rapidly punched under the microscope (MVX10, Olympus) and frozen with liquid nitrogen.

tag: age

value: lateral habenula

tag: stressed

value: defeated stress-resilient


title: INF.027

description:
NCT00944034
FS00006714-INF.027

tag: tissue


title: DL-P, biological rep3


title: INF.028

description:
NCT00944034
FS00006119-INF.028

tag: subject group


title: DL-P, biological rep6


title: Res1: D9_N3_GATCAG


title: INF.022

description:
NCT00944034
FS00005955-INF.022

tag: tissue subtype


title: Cont4: C8_H4_TGACCA

value: non-defeated


title: DL-P, biological rep2


title: INF.026

description:
NCT00944034
FS00006720-INF.026

value: Infants


title: DL-P, biological rep1


title: T2DMwell-DL-P, biological rep7

description: Gene Expression of PBMC, T2DMwell-DL-P

source_name: T2DMwell-DL-P, PBMC


title: INF.025

description:
NCT00944034
FS00006719-INF.025

value: serum-post IV 4CmenB doses


title: Res3: D12_N5_GGCTAC


title: Res2: D10_N4_TAGCTT


title: INF.021

description:
NCT00944034
FS00006723-INF.021


title: T2DMwell-DL-P, biological rep6

description: Gene Expression of PBMC, T2DMwell-DL-P


title: INF.020

description:
NCT00944034
FS00005957-INF.020


title: T2DMwell-DL-P, biological rep4

description: Gene Expression of PBMC, T2DMwell-DL-P


title: T2DMwell-DL-P, biological rep2

description: Gene Expression of PBMC, T2DMwell-DL-P


title: INF.019

description:
NCT00944034
FS00006334-INF.019


title: T2DMwell-DL-P, biological rep3

description: Gene Expression of PBMC, T2DMwell-DL-P


title: T2DMwell-DL-P, biological rep5

description: Gene Expression of PBMC, T2DMwell-DL-P


title: T2DMpoorly-DL-P, biological rep5

description: Gene Expression of PBMC, T2DMpoorly-DL-P

source_name: T2DMpoorly-DL-P, PBMC


title: T2DMwell-DL-P, biological rep1

description: Gene Expression of PBMC, T2DMwell-DL-P


title: DL-P, biological rep4


title: T2DMpoorly-DL-P, biological rep4


title: Sus4: D6_N2_ACTTGA

value: defeated stress-suceptible


title: Cont3: C6_H3_TTAGGC


title: INF.016

description:
NCT00944034
FS00006322-INF.016


title: INF.024

description:
NCT00944034
FS00005964-INF.024


title: INF.023

description:
NCT00944034
FS00005959-INF.023


title: T2DMpoorly-DL-P, biological rep3


title: INF.015

description:
NCT00944034
FS00006321-INF.015


title: INF.014

description:
NCT00944034
FS00006320-INF.014


title: INF.011

description:
NCT00944034
FS00006724-INF.011


title: INF.018

description:
NCT00944034
FS00006323-INF.018


title: Sus2: D3_H6_GCCAAT


title: RR936_m6Am_mouse_adult_Testes_m6A_IP_KO

city: Gneveva

first: Ramesh

last: Pillai

address: 30, Quai Ernest-Ansermet

department: Department of Molecular Biology

country: Switzerland

institute: University of Geneva

description:
m6Am-Exo-Seq
m6A_IP
all_genes_m6A_enrichment_KOvsHET.csv.gz

data_processing:
For gene expression profiling the reads were sorted into individual libraries based on the barcodes and mapped to the mouse genome (Ensembl release 95) using salmon v0.12.0 (salmon quant with options -l A  --validateMappings --gcBias)(Patro et al., 2017).  Further analysis was performed using R version 3.6.2  (R Core Team, 2017) and Bioconductor (Huber et al., 2015). The DESeq function of DESeq2_1.25.10 bioconductor package (Love et al., 2014) was used to obtain log2 fold changes of gene expression between wild-type and mutant samples and the adjusted p-values. Adjusted p-value 0.1 was used as a threshold for statistical significance. The individual mouse tissues were analyzed separately.
Analysis of ribosome footprinting data: To analyze the translational differences between the mutant and wild-type brain samples for individual genes, we compared the ratio of footprint vs. the input reads (i.e. translation efficiency) for individual genes.  We tested the differences in translation efficiency for individual genes using DESeq2 design: '~ assay(input/footprint) + condition(KO/WT) + assay:condition' and applying the likelihood ratio test which removes the interaction term in the reduced model.  For each gene we obtained log2 fold changes of gene translation efficiency difference between wild-type and mutant samples and the adjusted p-value.
Analysis of ribosome m6Am-Exo-Seq data: Reads were sorted into individual libraries based on the barcodes and mapped to the mouse genome (Ensembl release 95) using STAR (ref: PMID: 23104886) with parameters ?--outFilterType BySJout --limitOutSJcollapsed 50000000 --limitIObufferSize 1500000000?. The genomic coverages for individual samples were created in R using GenomicAlignments::coverage method.
For individual genes we tested the differences in m6A enrichment of their TSSs using matrix of read counts covering TSS of individual genes and DESeq2 design: '~ assay(input/m6A_IP) + condition(KO/HET) + assay:condition' and applying the likelihood ratio test which removes the interaction term in the reduced model.  For each gene we obtained log2 fold m6A enrichment difference between wild-type and mutant samples and the adjusted p-value. We identified 1370 genes which had significantly lower m6A TSS enrichment in the mutants. "
Genome_build: Ensembl release 95
Supplementary_files_format_and_content:
Gene expression data file contains information about gene expression changes in the KO.
Ribosome profiling file contains information about differences in footprint enrichment in the KO and control.
m6Am-Exo-Seq data files contains information about differences in m6A TSS enrichment in the KO and control.

source_name: Testes

growth_protocol: We targeted the Pcif1 locus in mouse embryos of the B6D2F1/J hybrid line (also called B6D2; The Jackson Laboratory, stock no. 100006). It is a cross between C57BL/6J (B6) and DBA/2J (D2), and heterozygous for all B6 and D2 alleles. Single-cell mouse embryos were injected with two different guide RNAs (gRNAs) that direct the DNA endonuclease Cas9 to delete a region between exon 15 and intron 16 (Figure S1A). Founder mice were identified by genotyping PCR (Figure S1B) and crossed with wildtype C57BL/6J (Janvier) partners to obtain germline transmission. We identified a mouse line with 89 bp deletion in the exon 15/intron 16 that removes the critical catalytic residues (NPPF) within the methyltransferase domain and also creates a premature termination codon.

extract_protocol:
Total RNA from biological triplicates of 2 months-old wildtype (WT) and Pcif1 KO mice were used for the experiment with mouse brain, spleen and testis samples. The tissues were snap-frozen in liquid nitrogen. Total RNA was isolated from tissues using TRIzol (Thermo Fisher Scientific) according to manufacturer instructions.
For ribosomal profiling brains from three adult wildtype (WT) controls and three Pcif1 KO mice were isolated and snap-frozen in liquid nitrogen. Frozen brain (~400 mg) was homogenized in 1300 µL (3x weight of the tissue in miligrams) of ice cold polysome lysis buffer (20 mM Tris?HCl pH 7.4, 150 mM NaCl, 5 mM MgCl2, 5 mM DTT, 100 ?g/ml Cycloheximide, 40 U/ml RNasin plus, (Promega), Complete EDTA-free protease inhibitor (Thermo Scientific), 1% TritonX?100, 0.5% Sodium deoxycholate) using a glass homogenizer (6-8 strokes). The lysate was transferred to a 1.5 mL Eppendorf tube and incubated on ice for 10 minutes with intermittent mixing. The lysate was centrifuged at 3000 rpm for 3 minutes at 4°C. Supernatant was carefully collected in a new 1.5 mL Eppendorf tube. The OD260 of the lysate was measured with a Nanodrop instrument and approximately 15 OD260 (which correspond to 100 µL of lysate), was used for ribosome footprint preparation, and 50 µL was directly transferred to TriZol (Ambion) for total input RNA extraction. The remaining lysate was divided in aliquots of 150 µL and was snap frozen in liquid nitrogen and stored in -80°C freezer for later use. For ribosome footprint preparation, 100 µL of the supernatant was treated with 2.5 µL (5 units) of Turbo-DNase (Ambion) and 6.5 µL (650 units) of RNase I at 23°C for 45 minutes with rotation at 300rmp in a thermoblock. In meantime, the s-400 HR columns (GE healthcare) were washed three times with 700 µL polysome buffer (20 mM Tris?HCl pH 7.4, 150 mM NaCl, 5 mM MgCl2, 5 mM DTT, 100 ?g/mL Cycloheximide, 40U/mL RNasin plus, (Promega), Complete EDTA-free protease inhibitor (Thermo Scientific)) plus 20U/mL SUPERaseIn (Ambion). During each wash columns with polysome buffer were spun at 2000rpm for 1 minute in a cold centrifuge. The columns were stored on ice with the polysome buffer in it (last wash). Once the RNaseI-treatment of lysate completed, 8.7 µL of SUPERaseIn RNase inhibitor was added to the reaction, gently mixed with the help of a pipette and stored on ice. The S-400 HR columns were spun at 2000 rpm for 1 minute to remove the polysome buffer. RNaseI-treated lysate was applied on the top of the column and spun at 2400 rpm for two minutes. The flow through was transferred to a fresh Eppendorf tube and RNA was extracted using 1.0 mL TriZol reagent. The RNA contain ribosome protected fragments (RPF) of mRNA as well as ribosomal RNA.
Approximately 20-25 µg of RNaseI-treated RNAs were separated on 15% UREA-PAGE to purify RPF which are in the range of 26-34 nt. Radiolabelled RNA marker of length 28 and 30 nt were used to size select the ribosome protected RNA fragment (RPF). The urea polyacrylamide gel was run at 16 W for 1 hour with the samples. Once the bromophenol blue reached the bottom of the gel, the power was switched off and the gel was taken out of the running apparatus. One glass plate was removed and the gel was wrapped in saran wrap and exposed to Storage Phosphor Screens (GE Healthcare; BAS IP MS 2025 E, Cat no. 28-9564-75) for 30 minutes, and scanned (GE Healthcare; Typhoon FLA 9500 IP, Cat no. 29-1885-90). Based on the position of markers, the gel piece corresponding to approximately 26-34 nt was excised and transfer to an RNase-free Eppendorf tube. The gel piece was ground to a fine powder and 600 µL of RNA extraction buffer (0.5 M Ammonium Acetate, 1 mM EDTA pH 8.0, and 0.1% SDS) was added and the tubes were incubated at 25°C overnight. Next day, the slurry was transferred to filter tubes (from Art-seq kit) and spun for 5 minutes at 5000 rpm. The flow-through was collected in a new RNase-free Eppendorf tube and precipitated with 1.0 µL of glycogen and 900 µL isopropanol overnight in -20°C freezer. The RNA solution was centrifuged at 13000 rpm for 20 minutes in a cold centrifuge to pellet RNA. RNA pellet was washed with 80% ethanol once, air dried and resuspended in 20 µL of RNase-free water.
m6Am-Exo-Seq (Sendinc et al., 2019) is a variation of m6A-IP seq. In this protocol, first, the m7G-capped RNA fragments (~100-200 nt) from poly(A)+ RNAs are enriched by multiple enzymatic treatments followed by m6A IP-seq to get m6A modification near the 5´  end of the mRNAs.  Approximately 8.0 µg of poly(A)+ RNAs from biological triplicates from Pcif1+/- and Pcif1-/- mouse testes were fragmented with RNA fragmentation buffer (Ambion) in a total volume of 20 µL for 7.30 minutes at 70°C in a thermcycler to obtain RNA fragments of ~100-200nt length. Fragmentation reaction was stopped by transferring the RNAs quickly to ice and adding 2.2 µL 10x stop solution (Ambion). The fragmented RNAs were purified with RNA Clean & Concentrator-5 (Zymo Research, R1014) and eluted in 17 µL water. Fragmented RNAs were 5´ phosphorylated by adding 10x, 2 µL T4 DNA ligase buffer (NEB) and 20 units of T4 PNK (ThermoScientific) in a final reaction volume of 20 µL. The phosphorylation reaction was carried out for 1.30 hour at 37°C. The 5´ phosphorylated RNA fragments were degraded by Terminator? 5´-Phosphate-Dependent Exonuclease (Epibio). To the phosphorylation reaction, 14 µL of water, 4 µL 10x Exonuclease buffer A and 2 µL Terminator exonuclease was added and the reaction mixture was incubated at 30°C for 3 hours. RNAs were purified from above reaction with RNA Clean & Concentrator-5 and eluted in 15 µL water. At this point we get 300-400 ng of m7G-cap-enriched RNA fragments from each sample.
The RNA obtained from above reaction were directly used for m6A-IP (Wojtas et al., 2017) with some modifications. First, 100 µL of protein A dynabeads (Life Technologies; 10002D) were washed twice with PXL buffer (1X PBS, 0.1% SDS, 0.5% sodium deoxycholate, 0.5% NP-40) and blocked with 1 µg/µL BSA in 400 µL PXL buffer for 45 minutes. Blocked beads were then incubated with 5 µg of anti-m6A antibody (Synaptic Systems, Cat. no. 202003) in 200 µL PXL buffer supplemented with 4 µL of RNasin RNase inhibitor (Promega; N2611) for one hour at room temperature. Antibody-conjugated beads were washed twice with PXL buffer and finally resuspended in 400 µL PXL buffer plus 4 µL RNAsin RNase inhibitor. Fragmented m7G-cap enriched RNAs were added and beads were incubated at 4°C for two hours with rotation. 10% of the RNA was kept separately and used to prepare input libraries. After two hours incubation, the beads were washed twice by ice-cold Nelson low-salt buffer (15 mM Tris at pH 7.5, 5 mM EDTA), once by ice-cold Nelson high-salt buffer (15 mM Tris at pH 7.5, 5 mM EDTA, 2.5 mM EGTA, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 1 M NaCl), once by ice-cold Nelson stringent wash buffer (15 mM Tris at pH 7.5, 5 mM EDTA, 2.5 mM EGTA, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 120 mM NaCl, 25 mM KCl), and last by ice-cold NT-2 buffer (50 mM Tris at pH 7.4, 150 mM NaCl, 1 mM MgCl2, 0.05% NP-40). Antibody-bound RNAs were eluted by adding 1mL TRIzol (Invitrogen) directly to the beads. RNAs were extracted and precipitated with Glycogen and ethanol overnight in -20°C, and used directly for the library preparation.
The TruSeq Stranded Total RNA kit with Ribo-Zero Gold (Illumina, Cat. No. 20020612) was used for library preparation with 400 ng of total RNA as input. Library molarity and quality was assessed with the Qubit and Tapestation using a DNA High sensitivity chip (Agilent Technologies). Libraries were diluted at 2 nM and pooled before the clustering process on a HiSeq 4000 Single Read flow cell. Paired-end reads of 100 bases were generated using the TruSeq SBS reagents on the Illumina HiSeq 4000 sequencer (iGE3 Genomics Platform, University of Geneva).
For ribosomal profiling the ribosomal RNAs were depleted from both input RNAs (400 ng) and RPF RNAs with mouse ribosomal RNAs probe from siTOOLs (mouse riboPOOLs) following their protocol. The ribosomal RNA-depleted input and RPF RNAs were used to make next generation sequencing library with TruSeq Ribo Profile (mammalian) kit from illumina. The libraries were sequenced at Genomics platform (University of Geneva).
The m6Am-Exo-Seq libraries were prepared with NEBNext® Ultra? II Directional RNA Library Prep Kit for Illumina® (E7765) according to the protocol 3 of the kit (Protocol for use with FFPE RNA, step 3.5 onwards). The final PCR was performed for 15 cycles (98°C ? 30 s, 12 cycle of 98°C ? 10 s, 65°C ? 75 s and final extension at 65°C ? 5 minutes). The libraries were sequenced on HiSeq4000 machine at genomics platform (University of Geneva)"

value: adult

value: KO


title: T2DMpoorly-DL-P, biological rep1


title: INF.017

description:
NCT00944034
FS00006721-INF.017


title: RR937_m6Am_mouse_adult_Testes_m6A_IP_KO


title: Cont2: C5_H2_CGATGT


title: T2DMpoorly-DL-P, biological rep2


title: INF.013

description:
NCT00944034
FS00006725-INF.013


title: Sus3: D4_N1_CAGATC


title: Cont1: C3_H1_ATCACG


title: INF.010

description:
NCT00944034
FS00006264-INF.010


title: RR935_m6Am_mouse_adult_Testes_m6A_IP_KO


title: Sus1: D2_H5_ACAGTG


title: INF.012

description:
NCT00944034
FS00006329-INF.012


title: INF.009

description:
NCT00944034
FS00006730-INF.009


title: INF.007

description:
NCT00944034
FS00006268-INF.007


title: amyg16

city: Vancouver

first: Mark

middle: Steven

last: Cembrowski

state: BC

address: 2350 Health Sciences Mall

department: Cellular and Physiological Sciences

country: Canada

institute: University of British Columbia

data_processing:
Trim input file and remove adapters.
Tag reads with cell barcode and UMI
Align reads to PhiX, then to mouse.
Aggregate reads and de-duplicate UMIs.
Post-processing in R with custom scripts and Seurat.
Genome_build: NCBIM37
Supplementary_files_format_and_content: Tab delimited file containing count values.

source_name: Basolateral amygdala

extract_protocol:
Microdissect, dissociate, and pool cells across acute brain slices in B6 mice
As in (Cembrowski et al., Cell, 2018)

tag: cell type

value: basolateral amygdala excitatory neuron


title: INF.005

description:
NCT00944034
FS00006234-INF.005

value: blood


title: amyg15


title: RR932_m6Am_mouse_adult_Testes_m6A_IP_HET

value: HET


title: RR933_m6Am_mouse_adult_Testes_m6A_IP_HET


title: RR934_m6Am_mouse_adult_Testes_m6A_IP_HET


title: INF.008

description:
NCT00944034
FS00006137-INF.008


title: amyg14


title: RR931_m6Am_mouse_adult_Testes_input_KO

description:
m6Am-Exo-Seq
input
all_genes_m6A_enrichment_KOvsHET.csv.gz


title: amyg13


title: amyg11


title: INF.006

description:
NCT00944034
FS00006229-INF.006


title: amyg12


title: PBS-2

city: Suita

first: Kiyoharu

last: Fukushima

state: Osaka

department: Host defense

institute: Osaka university

hyb_protocol: standard Affymetrix protocol

scan_protocol: GeneChips were scanned using the Hewlett-Packard GeneArray Scanner

data_processing:
Background-adjusted robust multichip average expression values were calculated using Expression Console Software (Thermo Fisher Scientific) and Transcriptome Analysis Software (Thermo Fisher Scientific)
affymetrix-algorithm-name = sst-rma-gene-full
affymetrix-algorithm-version = 1.0
program-name = Expression Console
program-version = 1.4.1.46

source_name: mouse lung CD45-negative cells 10 days after PBS

label_protocol: standard Affymetrix protocol

extract_protocol: Extraction of total RNAs of lung CD45-negative cells was performed according to the manufacturer's instructions(Roche).

treatment_protocol: PBS or Bleomycin was intra-tracheally administered

value: lung CD45-negative cells

tag: treatment

value: PBS

tag: time point

value: 10 days after treatment


title: INF.003

description:
NCT00944034
FS00006238-INF.003


title: INF.001

description:
NCT00944034
FS00006235-INF.001


title: INF.002

description:
NCT00944034
FS00006236-INF.002


title: ADO.046

description:
NCT00661713
FS00007834-ADO.046

source_name: vaccinated_Adolescent_serum-post III 4CmenB doses

value: Adolescent

value: serum-post III 4CmenB doses


title: RR929_m6Am_mouse_adult_Testes_input_KO


title: amyg10


title: RR927_m6Am_mouse_adult_Testes_input_HET


title: RR928_m6Am_mouse_adult_Testes_input_HET


title: RR926_m6Am_mouse_adult_Testes_input_HET


title: BLM-1

source_name: mouse lung CD45-negative cells 10 days after BLM

value: Bleomycin (BLM)


title: ADO.045

description:
NCT00661713
FS00007840-ADO.045


title: BLM-2


title: amyg9


title: INF.004

description:
NCT00944034
FS00006232-INF.004


title: amyg8


title: amyg7


title: RR891_RF_mouse_adult_Brain_RF_KO

description:
ribosome footprintig
RF
all_genes_RF_KOvsWT.csv.gz

source_name: Brain


title: PVP-AgNP exposed mouse-rep 4

city: Rensselaer

first: Sridar

middle: V

last: Chittur

address: One Discovery Drive, CRC 342G

department: Center for Functional Genomics

institute: SUNY-University at Albany

description: gene expression data from PVP-AgNP exposed mouse liver

hyb_protocol: 16 h to Affymetrix Mouse Transcriptome 1.0 ST arrays

scan_protocol: Affymetrix Scanner 3000 7G using AGCC software.

data_processing: CEL files were summarized using the SST RMA algorithm in Transcriptome Analysis Console v4.0.1.36 (Affymetrix) and the normalized data were subjected to one-way ANOVA with a Benjamin Hochberg False Discovery Rate correction included (p<0.05). A 2-fold change was used to select entities that were statistically and differentially expressed between any two conditions.

source_name: PVP-AgNP exposed mouse liver

label_protocol: Affymetrix Whole Transcript Sense Target labeling protocol

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions.

treatment_protocol: Mice were treated with commercially available AgNPs (20 nm, BioPure ™, nanoComposix, 1 mg Ag/ml) coated with cit or PVP. Aqueos AgNP solutions were administered to 3 months-old male and female mice via oral gavage daily for 7 consequtive days. Livers were harvested 24 h after administration of last dose, flash frozen in liquid nitrogen and stored in -80 C until analysis.

value: liver

value: C57BL/6J -Oca2p-un/J

tag: gender

value: male

tag: age, months


title: PBS-1


title: RR890_RF_mouse_adult_Brain_RF_KO

tag: genotype


title: PVP-AgNP exposed mouse-rep 3


title: ADO.043

description:
NCT00661713
FS00006325-ADO.043


title: ADO.044

description:
NCT00661713
FS00007835-ADO.044


title: amyg6


title: ADO.042

description:
NCT00661713
FS00006279-ADO.042


title: amyg5


title: PVP-AgNP exposed mouse-rep 2

value: female


title: amyg4


title: RR887_RF_mouse_adult_Brain_input_KO

description:
ribosome footprintig
input
all_genes_RF_KOvsWT.csv.gz


title: ADO.041

description:
NCT00661713
FS00007833-ADO.041


title: ADO.040

description:
NCT00661713
FS00006766-ADO.040


title: amyg3


title: RR888_RF_mouse_adult_Brain_input_KO


title: RR889_RF_mouse_adult_Brain_RF_KO


title: cit-AgNP exposed mouse-rep 4

description: gene expression data from cit-AgNP exposed mouse liver

source_name: cit-AgNP exposed mouse liver


title: RR885_RF_mouse_adult_Brain_RF_WT

value: WT


title: PVP-AgNP exposed mouse-rep 1


title: ADO.038

description:
NCT00661713
FS00006341-ADO.038


title: RR884_RF_mouse_adult_Brain_RF_WT


title: cit-AgNP exposed mouse-rep 1


title: RR883_RF_mouse_adult_Brain_RF_WT


title: control mouse-rep 3

description: gene expression data from control mouse liver

source_name: control mouse liver


title: control mouse-rep 4


title: control mouse-rep 2


title: cit-AgNP exposed mouse-rep 3


title: ADO.037

description:
NCT00661713
FS00007831-ADO.037


title: ADO.036

description:
NCT00661713
FS00007869-ADO.036


title: amyg2


title: RR886_RF_mouse_adult_Brain_input_KO


title: RR882_RF_mouse_adult_Brain_input_WT


title: control mouse-rep 1


title: RR881_RF_mouse_adult_Brain_input_WT


title: ADO.035

description:
NCT00661713
FS00006277-ADO.035


title: ADO.039

description:
NCT00661713
FS00007872-ADO.039


title: TSWTCBPsh

description: alligned using mouse reference genome GRCm38 build 84

data_processing:
The Ensembl mouse reference genome GRCm38 build 84 was used to align all reads generated by Illumina HiSeq
The reference genome index was created using Bowtie2  run with default parameters
RNA-seq reads for each sample were aligned to the reference genome using TopHat2  run with default parameters
The aligned reads for each sample were then independently processed using GFOLD and the reference gene annotation to obtain normalized reads per kilobase million (RPKM) values for each gene
Zero RPKM values were converted to “epsilon”, a small number equaling 2.2204e-16 to avoid division-by-zero errors during fold change calculations.
Genome_build: genome GRCm38 build 84
Supplementary_files_format_and_content: Excel Spreadsheet  includes RPKM values for each Sample.

source_name: Trophoblast stem cells

extract_protocol:
Total RNA was extracted from TS cells using the RNeasy minikit (Qiagen)
RNA libraries were prepared for sequencing using standard Illumina protocols at High-Throughput Sequencing Facility (HTSF) at UNC school of medicine

value: Trophoblast stem cells

value: Wild-type

tag: vector

value: CBPshRNA control  lentiviral vector


title: amyg1

value: basolateral amygdala excitatory neuron


title: RR880_RF_mouse_adult_Brain_input_WT

value: adult


title: TSWT

value: pLKO control  lentiviral vector


title: TSKI4

value: MAP3K4 Knockin K1361R


title: ADO.034

description:
NCT00661713
FS00005221-ADO.034

value: blood


title: TSKI4H6sh

value: HDAC6shRNA expressing lentiviral vector


title: ADO.032

description:
NCT00661713
FS00007866-ADO.032

value: Adolescent


title: ADO.033

description:
NCT00661713
FS00007829-ADO.033

value: serum-post III 4CmenB doses


title: OtB5298

first: Francesca

middle: M.

last: Spagnoli

address: Great Maze Pond

department: CGTRM

institute: King's College London

data_processing:
Illumina sequencing libraries from stranded cDNA were sequenced, paired-end, on Illumina HiSeq2500
Data sets had been mapped against mm10 mouse genome with star 2.4.0j, and then analyzed with cufflinks (2.2.1) for expression level measurement on www.DNAnexus.com
Comparison of expression level between samples had been conducted with cufflinks.cuffdiff (2.2.1)
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample and desired comparisons

source_name: Pool of FACS-sorted pancreatic epithelial cells

extract_protocol:
Dorsal pancreatic buds were dissected, dissociated into single-cell suspensions. Tomato- and GFP- positive cells were then sorted at 4°C using a FACS Aria II flow cytometer (BD) using GFP and DsRed filters and by setting the gate on the GFP and Tomato fluorescence intensity. FACS-sorted fractions were then frozen and kept at -80°C. They were subsequently pooled and RNA was purified using a combination of Trizol reagent and RNeasy RNA Mini Kit (Qiagen).
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Embryonic dorsal pancreatic bud

tag: developmental stage


title: RR871_mouse_adult_Brain_KO

description:
gene expression profiling
total RNA
all_genes_mouse_PCIF1.xlsx.gz

value: KO


title: OtB5296

source_name: Pool of FACS-sorted pancreatic mesenchymal cells


title: RR868_mouse_adult_Brain_WT


title: OtB5295

value: mTmG


title: RR869_mouse_adult_Brain_KO


title: RR870_mouse_adult_Brain_KO


title: RR867_mouse_adult_Brain_WT


title: ADO.029

description:
NCT00661713
FS00007827-ADO.029


title: ADO.031

description:
NCT00661713
FS00007567-ADO.031


title: ADO.030

description:
NCT00661713
FS00007830-ADO.030


title: ADO.028

description:
NCT00661713
FS00007828-ADO.028


title: RR866_mouse_adult_Brain_WT


title: OtB5294


title: OtB5293


title: RR861_mouse_adult_Testes_KO


title: RR865_mouse_adult_Testes_KO


title: RR863_mouse_adult_Testes_KO


title: ADO.026

description:
NCT00661713
FS00006336-ADO.026


title: ADO.027

description:
NCT00661713
FS00007568-ADO.027


title: ADO.024

description:
NCT00661713
FS00006274-ADO.024


title: RR859_mouse_adult_Testes_WT


title: ADO.025

description:
NCT00661713
FS00007876-ADO.025


title: Rif_1.5m_DMS

city: Baltimore

first: Yumeng

last: Hao

state: Maryland

address: 3400 N. Charles Street, Jenkins 408

department: Biophysics

institute: Johns Hopkins University

data_processing:
library strategy: 4U-DMS-MaP
adaptor removed with cutadapt
first 3 nt trimmed with FASTX trimmer
low quality bases trimmed (ShapeMapper)
Reads mapped to rRNA with Bowtie2 (ShapeMapper)
Coverage and mutations computed for each read (ShapeMapper)
Genome_build: Escherichia coli strain MRE600 rRNA with leader and trailer
Supplementary_files_format_and_content: tab-seperated text file of mutation rates (#mutations/coverage), a proxy for extent of DMS modification, for each nucleotide in each dataset.

organism: Escherichia coli

source_name: ribosome

extract_protocol:
Total RNA was extracted from E. coli cells. The 4-thio-uridine labeled RNAs were biotinylated and extracted via pulldown.
RNA after pulldown was fragmented with Zinc Chloride for 5min @ 94C. 3' ends were resolved by treatment with T4 PNK, and RNA was size-selected by gel for ~60-70nt fragments. A preadenylated linker was ligated with T4 PNK, and the ligated products were gel purified. RT was performed with TGIRT RT @ 60C for 1 hour, and full-length RT products were gel purified. cDNA was circularized with circ ligase, and amplified by 11-13 cycles of PCR.

treatment_protocol: For mature ribosomes, the E. coli cells were grown to log phase, then treated with DMS for 30 seconds, and quenched with βME buffer. For 4-thio-uridine labeled mature ribosomes, the E. coli cells were grown to log phase, metabolically labeled with 4-thio uridine for 10 minute. The samples were further treated with treated with DMS for 30 seconds, and quenched with βME buffer. For nascent ribosomes, the E. coli cells were grown to log phase, metabolically labeled with 4-thio uridine for 1 minute then treated with rifampicin. The samples were further treated with treated with DMS for 30 seconds, and quenched with βME buffer.

tag: dms concentration

tag: treatment time

value: 30 seconds

tag: 4su concentrations

tag: 4su treatment time

value: 1 minute

tag: rif concentration

value: 200 µg/mL

tag: rif treatment time

value: 1.5 minutes


title: Rif_1m_DMS

tag: strain


title: Rif_2.5m_DMS

value: 2.5 minutes


title: Rif_1.5m_DMS_2

value: 1.5 minute


title: ADO.023

description:
NCT00661713
FS00007868-ADO.023

tag: subject group

tag: tissue


title: RR857_mouse_adult_Testes_WT

tag: age

value: WT


title: Rif_30s_DMS


title: RR855_mouse_adult_Testes_WT

value: adult


title: ADO.022

description:
NCT00661713
FS00007870-ADO.022

tag: tissue subtype


title: PA14 glycans 15 m

city: Cambridge

first: Katharina

last: Ribbeck

state: Massachusetts

address: 21 Ames St

department: Biological Engineering

institute: Massachusetts Institute of Technology

data_processing:
Illumina genome analyzer modules Firecrest and Bustard were used for image analysis and base-calling, respectively.
Sequence reads were mapped onto the P. aeruginosa PA14 reference genome, which is available for download from the Pseudomonas Genome Database (http://www.pseudomonas.com) using the Burrows Wheeler algorithm on the Galaxy server.
Read counts were quantified in R thes files are provided as processed data)
Gene expression values were normalized based on library size and differentially expressed genes were identified using a negative binomial test with a false discovery rate (FDR) less than 0.05.
Genome_build: Pseudomonas aeruginosa PA14
Supplementary_files_format_and_content: comma-delimited text files with raw read counts

organism: Pseudomonas aeruginosa

source_name: Planktonic bacterial cultures

growth_protocol: Batch cultivation of Pseudomonas aeruginosa was carried out shaking at 37 °C in LB. Overnight cultures were diluted to an OD600 of 0.01 medium with or without (0.5%) MUC5AC or (0.1%) MUC5AC glycans and grown for 15 minutes or 5 h at 37 °C in a static 96-well microtiter plate.

extract_protocol:
Total RNA was extracted using the MasterPure RNA Purification kit (Lucigen) and residual DNA was removed using the Turbo DNA-free kit (Ambion). The integrity of the total RNA was assessed with an Agilent 2100 Bioanalyzer (Agilent Technologies). 16S, 23S, and 5S rRNA were removed using biotinylated rRNA probes (as described in Culviner and Laub, 2019).
Gene expression analysis was conducted via Illumina RNA sequencing (RNA-seq). Libraries were produced using the KAPA RNA HyperPrep kit (Kapa Biosystems). Libraries were sequenced using the Illumina HiSeq platform with a single-end protocol and read lengths of 40 nt or 52 nt.

tag: growth medium

value: ABTGC medium

tag: growth time


title: PA14 no glycans  15 m


title: Rif_ND2

value: 0 minute

value: 6.5 minutes


title: Rif_ND3

value: 10.5 minutes


title: ADO.021

description:
NCT00661713
FS00007864-ADO.021

value: Adolescent

value: blood


title: D858A glycans 5 h

value: PA14 RetS(D858A)

value: 5 h


title: ADO.020

description:
NCT00661713
FS00007871-ADO.020

value: serum-post III 4CmenB doses


title: RR862_mouse_adult_Spleen_KO

source_name: Spleen

value: KO


title: RR864_mouse_adult_Spleen_KO


title: ADO.019

description:
NCT00661713
FS00007656-ADO.019


title: Rif_ND1

value: 1 minute

value: 30 seconds


title: D858A no glycans 5 h


title: 4U_10m_DMS_2

value: 31 seconds

value: 0 µg/mL


title: ADO.018

description:
NCT00661713
FS00006335-ADO.018


title: RR860_mouse_adult_Spleen_KO


title: RR858_mouse_adult_Spleen_WT


title: Dismed glycans 5 h

value: PA14 RetS(delta dismed2)


title: 4U_ND


title: 4U_10m_DMS_1


title: Dismed no glycans 5 h


title: RR856_mouse_adult_Spleen_WT

value: WT


title: RR854_mouse_adult_Spleen_WT

value: adult


title: ADO.017

description:
NCT00661713
FS00007617-ADO.017


title: PA14 glycans 5 h 2

value: ABTGC medium


title: DMS_ctrl

value: 0 mM

value: 0 minute


title: PA14 glycans 5 h 1


title: ND_ctrl


title: Untreat.Hep.no3.ATAC

first: Jinyang

last: Li

state: New York

institute: The Rockefeller University

data_processing:
alignment using Bowtie2 to generate sam files
sam and bam files were processed with SAMtools
peak calling using MACS2
peaks were merged with BEDTools
read counts were calculated and normalized in the merged peaks for every sample
Genome_build: mm10
Supplementary_files_format_and_content: TPM as txt file

growth_protocol: Cells were directly sorted from liver of mice without in vitro growth

extract_protocol:
DNA was extracted using Qiagen Kit.
ATAC-seq libraries were build following standard protocols.

treatment_protocol: Cells were not treated following sorting from liver of mice

tag: cell type

value: hepatocyte


title: ADO.016

description:
NCT00661713
FS00006340-ADO.016

value: Adolescent


title: Untreat.Hep.no2.ATAC


title: ADO.015

description:
NCT00661713
FS00007651-ADO.015

value: blood


title: ADO.014

description:
NCT00661713
FS00007865-ADO.014


title: PA14 no glycans 5 h 1


title: PA14 no glycans 5 h 2


title: ADO.013

description:
NCT00661713
FS00007652-ADO.013


title: Untreat.Bil.no3.ATAC

value: biliary cells


title: cit-AgNP exposed mouse-rep 2

value: liver

value: C57BL/6J -Oca2p-un/J

tag: gender

value: female

tag: age, months


title: RNAseq_hMADS_shGPS2_D14_4

city: Huddinge

first: Serena

last: Barilla

state: Huddinge

address: Hälsovägen 7C

institute: Karolinska Institutet

data_processing:
RNA-seq raw files (fastq files) were aligned to GRCh38 version of the human reference genome, using HISAT2 with defalt settings. SAM files were converted to BAM files and then sorted using SAMtools. featureCounts were then used to count tag in all exons.
Genome_build: hg38
Supplementary_files_format_and_content: RNA-seq subreads counts files are in txt format and contain gene id and tag counts  in the indicated samples.

source_name: human adipose-derived stem cells

growth_protocol: hMADS cells were maintained in DMEM (Lonza, low glucose) supplemented with 10% fetal calf serum (Eurobio), 15 mM Hepes (Lonza), 2 mM L-Glutamine (Lonza), 1% penicillin/streptomycin and 2.5 ng/ml hFGF2 (Peprotech).

extract_protocol:
Total RNA was extracted from cells  using Trizol Reagent and RNeasy RNA Mini Kit. RNA quality was assessed by Agilent TapeStation 2200.
PolyA RNA selection was performed using the Illumina TruSeq RNA Sample Preparation Kit according to the manufacturer's protocol. RNA-seq libraries were prepared and sequenced on the Illumina HiSeq 2000.

treatment_protocol: Two days post-confluence (day 0), cells were induced to differentiate in DMEM/Ham’s F12 (Lonza) media supplemented with 1 μM dexamethasone, 500 μM 3-isobutyl-1-methylxanthine (IBMX), 5 μg/ml insulin, 10 μg/ml transferrin and 0.2 nM triiodothyronine (T3). After two days of induction, dexamethasone and IBMX were omitted from the media, and 100 nM of rosiglitazone was added until day 9. The differentiation was continued until day 14 when full white adipocytes were developed.

tag: cell line

tag: genotype

tag: timepoint

value: Day 14


title: RNAseq_hMADS_shGPS2_D14_3


title: Untreat.Hep.no1.ATAC


title: OtB5297

value: Embryonic dorsal pancreatic bud

tag: developmental stage

value: mTmG


title: RNAseq_hMADS_shGPS2_D14_2


title: PA14 mucin 5 h 2

value: 5 h


title: Untreat.Bil.no2.ATAC


title: PA14 mucin 5 h 1


title: ADO.011

description:
NCT00661713
FS00007648-ADO.011

data_processing: Images were processed using ImaGene 9.0 software (Biodiscovery Inc, CA, USA). Elaboration and analysis of image raw fluorescence Intensity (FI) data was performed using in-house developed software and R scripts. For each sample, the mean fluorescence intensity (MFI) of replicated spots was determined, after subtraction of the background value surrounding each spot.

extract_protocol: Human sera collected from different clinical trials

value: serum-post III 4CmenB doses


title: ADO.010

description:
NCT00661713
FS00007649-ADO.010


title: PA14 no mucin 5 h 2


title: RNAseq_hMADS_shGPS2_D14_1


title: Untreat.Bil.no1.ATAC


title: ADO.009

description:
NCT00661713
FS00007645-ADO.009


title: DDC.Hep.no3.ATAC


title: RNAseq_hMADS_shGFP_D14_4

value: shGFP


title: RNAseq_hMADS_shGFP_D14_3


title: ADO.012

description:
NCT00661713
FS00007653-ADO.012


title: PA14 no mucin 5 h 1


title: DDC.Hep.no2.ATAC


title: DDC.Hep.no1.ATAC


title: IMR90_lowip_MLLT3_1

city: Bronx

first: Yizhou

last: Zhu

address: Price469 1301 Morris Park Ave

department: Genetics

institute: Albert Einstein College of Medicine

description: Baseline

data_processing:
Library strategy: Tri-4C
Basecalls performed using CASAVA version 1.8
Reads were filtered using padding sequence indicated (Zhu, Suh)
Viewpoint sequence was trimmed using fastx_trimmer -f N, where N equals to viewpoint length
Processed reads were aligned to the hg19 genome assembly using bwa mem with default setting
Data were filtered with MAPQ>0. Reads aligned -1k to +2k from viewpoint were removed
Duplication reads were filtered using custom awk script (Zhu, Suh)
Genome_build: hg19
Supplementary_files_format_and_content: bedGraph with 100 bp bin size. Score for each bin indicate read count aligned to 500 bp interval (+-250 bp)

source_name: Lung

growth_protocol: IMR90 cells was maintained in EMEM with 10% FBS

extract_protocol:
Cells were fixed with 1% (v/v) formaldehyde in PBS for 10 min at room temperature. Fixation was quenched 0.2M glycene. Cells were washed with cold PBS twice and pelleted.
Cell pellet was resuspended in a premixture of 250 µl cold lysis buffer (10 mM Tris-HCl pH 8.0, 10 mM NaCl, 0.2% Igepal CA630) and 50 µl protease inhibitors. Suspension was incubated on ice for 15 min and centrifuged. The pellet was washed once with 500 µl of cold lysis buffer, and carefully resuspended in 50 µl of 0.5% SDS. The suspension was then incubated in a 62 oC heating block for 7 min, followed by mixing with 145 µl water and 25 µl 10% Triton X-100, and incubated at 37 oC for 15 min. The suspension was mixed with 50 µl of 10x Cutsmart buffer, 120 U MboI (DpnII), 100 U Csp6I (CviQI), and 100 U CviAII, and filled the total volume to 500 µl. Digestions were conducted at 25 oC for 2 hours and then 37 oC overnight with rotation. The restriction enzymes were inactivated by incubating at 65 oC for 20 min. After cooling to room temperature, end blunting was performed by adding 3 µl of 10 mM dNTP, 8 µl of DNA polymerase I Klenow, and 4 µl of T4 DNA polymerase, and incubating at 37 oC for 1 hour with rotation. The suspension was then mixed with 460 µl water, 120 µl T4 ligase buffer, 100 µl 10% Triton X-100, 6 µl 20 mg/ml BSA, and 5 µl of 400 U/µl T4 ligase, and incubated for 4 hours at room temperature with rotation. The processed nuclei were pelleted and resuspended in 500 µl 1x T4 ligation buffer supplied with 50 µl 20 mg/ml proteinase K and 50 µl 10% SDS, and incubated at 55 oC for 30 min. For de-crosslinking, 60 µl 5M sodium chloride was added and the mixture was incubated at 68 oC for 2 hours. DNA was recovered using phenol-cholorform extraction.The re-arranged genomic DNA was sonicated to 300-400 bp fragments using Covaris S2 ultrasonicator. The fragmented DNA was double size-selected using 0.40x/1.0x of SPRI beads to remove reads below 100 bp and above 1000 bp, and eluted with a final volume of 70 µl of 10 mM Tris-HCl. The eluent was mixed with 10 µl 10x T4 ligation buffer , 10 µl 100 mM ATP, 5 µl 10 mM dNTP mix, 4 µl T4 DNA polymerase , 1 µl DNA polymerase Klenow, and 5 µl T4 PNK, and incubated for 30 min at room temperature. The repaired DNA was purified by using 1.0x SPRI beads, and eluted with a master mix of 94.5 µl 1x NEB buffer 2 and 0.5 µl 100 mM dATP. After removing the beads, 5 µl of Klenow exo- was added, and the mixture was incubated at 37 oC for 30 min for dA tailing. The processed DNA was purified by using 1.0x SPRI beads, and eluted with 20 µl 10 mM Tris-HCl. After dA-tailing, a custom Y shape adaptor was ligated to the fragments. For amplification, eight 100 µl reactions, each containing 400 µg preamplification library, 2 µM universal primer, 0.5 µM viewpoint-specific outer primer for each multiplexed viewpoint, and 1x SuperFi PCR master mix with 20% GC enhancer, was performed at following condition: 98 oC for 30s, 28 cycles of 98 oC for 10s, 62 oC for 10s, and 72 oC for 60s, and final extention at 72 oC for 5 min. The product was pooled and purified with 1.0x SPRI beads.

value: Lung Fibroblast

tag: passages

tag: method

value: Triple

tag: viewpoint

value: CTCGGAGCCCGGGTGTCAGGCGCCACGGCG


title: ADO.007

first: Erika

last: Bartolini

institute: GSK Vaccines

description:
NCT00661713
FS00007646-ADO.007


title: ADO.008

description:
NCT00661713
FS00007650-ADO.008


title: RNAseq_hMADS_shGFP_D14_1


title: IMR90_alt_MLLT3_2

description: Baseline


title: DDC.Bil.no3.ATAC


title: IMR90_alt_MLLT3_1

description: Baseline


title: DDC.Bil.no2.ATAC


title: RNAseq_hMADS_shGPS2_D2_4

value: Day 2


title: ADO.005

description:
NCT00661713
FS00007549-ADO.005


title: RNAseq_hMADS_shGPS2_D2_3


title: ADO.004

description:
NCT00661713
FS00007647-ADO.004


title: DDC.Bil.no1.ATAC


title: DDC.Rep.no3.ATAC

value: reprogrammed hepatocyte


title: IMR90_alt_Boundary_1

description: Baseline

value: CAGTCCTCTCCCGCCATCTAGTGGT


title: IMR90_alt_Boundary_2

description: Baseline


title: RNAseq_hMADS_shGFP_D14_2


title: RNAseq_hMADS_shGPS2_D2_1


title: RNAseq_hMADS_shGPS2_D2_2


title: ADO.002

description:
NCT00661713
FS00007839-ADO.002

value: Adolescent

value: blood


title: ADO.003

description:
NCT00661713
FS00007842-ADO.003


title: DDC.Rep.no2.ATAC


title: RNAseq_hMADS_shGFP_D2_4


title: VSMC_Tri4C_ECAD9_2

description: Baseline

data_processing:
Library strategy: Tri-4C
Basecalls performed using CASAVA version 1.8
Reads were filtered using padding sequence indicated (Zhu, Suh)
Heterozygote reads were split according to the allele genoype of the tag SNP
The raw data was processed after splitting to the paternal and maternal allele (annotated as NR for non-risk and R for risk).
Viewpoint sequence was trimmed using fastx_trimmer -f N, where N equals to viewpoint length
Processed reads were aligned to the hg19 genome assembly using bwa mem with default setting
Data were filtered with MAPQ>0. Reads aligned -1k to +2k from viewpoint were removed
Duplication reads were filtered using custom awk script (Zhu, Suh)
Genome_build: hg19
Supplementary_files_format_and_content: bedGraph with 100 bp bin size. Score for each bin indicate read count aligned to 500 bp interval (+-250 bp)

source_name: Embryonic stem cell-derived VSMC

growth_protocol: hESC cells were maintained in feeder-free E8 system. To differentiate to VSMCs, cells were treated with N2B27 medium with 10 μM CHIR99021 and 25 ng/ml BMP4 for three days, followed by N2B27 supplied with 10ng/ml PDGF-BB and 2ng/ml Activin A for 5 days. Cells were maintained in N2B27 medium supplied with 10  ng/ml PDGF-BB

extract_protocol:
Cells were fixed with 1% (v/v) formaldehyde in PBS for 10 min at room temperature. Fixation was quenched 0.2M glycene. Cells were washed with cold PBS twice and pelleted.
Cell pellet was resuspended in a premixture of 250 µl cold lysis buffer (10 mM Tris-HCl pH 8.0, 10 mM NaCl, 0.2% Igepal CA630) and 50 µl protease inhibitors. Suspension was incubated on ice for 15 min and centrifuged. The pellet was washed once with 500 µl of cold lysis buffer, and carefully resuspended in 50 µl of 0.5% SDS. The suspension was then incubated in a 62 oC heating block for 7 min, followed by mixing with 145 µl water and 25 µl 10% Triton X-100, and incubated at 37 oC for 15 min. The suspension was mixed with 50 µl of buffer G, 120 U MboI (DpnII), 120 U Csp6I (CviQI), and 100 U Hin1II, and filled the total volume to 500 µl. For Single RE UMI-4C experiments, the suspension was mixed with 50 µl of buffer R, buffer B, and buffer G, respectively, for digestion with 100 U MboI, 100 U Csp6I, and 100 U Hin1II   in 500 µl final volume. All digestions were conducted at 37 oC overnight with rotation. The restriction enzymes were inactivated by incubating at 65 oC for 20 min. After cooling to room temperature, end blunting was performed by adding 3 µl of 10 mM dNTP, 8 µl of DNA polymerase I Klenow, and 4 µl of T4 DNA polymerase, and incubating at 37 oC for 1 hour with rotation. The suspension was then mixed with 460 µl water, 120 µl T4 ligase buffer, 100 µl 10% Triton X-100, 6 µl 20 mg/ml BSA, and 5 µl of 400 U/µl T4 ligase, and incubated for 4 hours at room temperature with rotation. The processed nuclei were pelleted and resuspended in 500 µl 1x T4 ligation buffer supplied with 50 µl 20 mg/ml proteinase K and 50 µl 10% SDS, and incubated at 55 oC for 30 min. For de-crosslinking, 60 µl 5M sodium chloride was added and the mixture was incubated at 68 oC for 2 hours. DNA was recovered using phenol-cholorform extraction.The re-arranged genomic DNA was sonicated to 300-400 bp fragments using Covaris S2 ultrasonicator. The fragmented DNA was double size-selected using 0.40x/1.0x of SPRI beads to remove reads below 100 bp and above 1000 bp, and eluted with a final volume of 70 µl of 10 mM Tris-HCl. The eluent was mixed with 10 µl 10x T4 ligation buffer , 10 µl 100 mM ATP, 5 µl 10 mM dNTP mix, 4 µl T4 DNA polymerase , 1 µl DNA polymerase Klenow, and 5 µl T4 PNK, and incubated for 30 min at room temperature. The repaired DNA was purified by using 1.0x SPRI beads, and eluted with a master mix of 94.5 µl 1x NEB buffer 2 and 0.5 µl 100 mM dATP. After removing the beads, 5 µl of Klenow exo- was added, and the mixture was incubated at 37 oC for 30 min for dA tailing. The processed DNA was purified by using 1.0x SPRI beads, and eluted with 20 µl 10 mM Tris-HCl. After dA-tailing, a custom Y shape adaptor was ligated to the fragments. For amplification, eight 100 µl reactions, each containing 400 µg preamplification library, 2 µM universal primer, 0.5 µM viewpoint-specific outer primer for each multiplexed viewpoint, and 1x SuperFi PCR master mix with 20% GC enhancer, was performed at following condition: 98 oC for 30s, 28 cycles of 98 oC for 10s, 62 oC for 10s, and 72 oC for 60s, and final extention at 72 oC for 5 min. The product was pooled and purified with 1.0x SPRI beads.

value: H7-derived vascular smooth muscle cell

tag: digestion


title: ADO.006

description:
NCT00661713
FS00007601-ADO.006

tag: subject group


title: RNAseq_hMADS_shGFP_D2_3


title: VSMC_Tri4C_ECAD9_1

description: Baseline


title: ADU.030

description:
NCT00560313
FS00005311-ADU.030

source_name: vaccinated_Adult_serum-post III 4CmenB doses

value: Adult

tag: tissue


title: DDC.Rep.no1.ATAC


title: IMR90C_NlaIII_MLLT3_2

description: Baseline

data_processing:
Basecalls performed using CASAVA version 1.8
Reads were filtered using padding sequence indicated (Zhu, Suh)
Viewpoint sequence was trimmed using fastx_trimmer -f N, where N equals to viewpoint length
Processed reads were aligned to the hg19 genome assembly using bwa mem with default setting
Data were filtered with MAPQ>0. Reads aligned -1k to +2k from viewpoint were removed
Duplication reads were filtered using custom awk script (Zhu, Suh)
Genome_build: hg19
Supplementary_files_format_and_content: bedGraph with 100 bp bin size. Score for each bin indicate read count aligned to 500 bp interval (+-250 bp)

growth_protocol: IMR90 cellswas maintained in EMEM with 10% FBS

extract_protocol:
Cells were fixed with 1% (v/v) formaldehyde in PBS for 10 min at room temperature. Fixation was quenched 0.2M glycene. Cells were washed with cold PBS twice and pelleted.
Cell pellet was resuspended in a premixture of 250 µl cold lysis buffer (10 mM Tris-HCl pH 8.0, 10 mM NaCl, 0.2% Igepal CA630) and 50 µl protease inhibitors. Suspension was incubated on ice for 15 min and centrifuged. The pellet was washed once with 500 µl of cold lysis buffer, and carefully resuspended in 50 µl of 0.5% SDS. The suspension was then incubated in a 62 oC heating block for 7 min, followed by mixing with 145 µl water and 25 µl 10% Triton X-100, and incubated at 37 oC for 15 min. The suspension was mixed with 50 µl of buffer G, 120 U MboI (DpnII), 120 U Csp6I (CviQI), and 100 U Hin1II, and filled the total volume to 500 µl. For Single RE UMI-4C experiments, the suspension was mixed with 50 µl of buffer R, buffer B, and buffer G, respectively, for digestion with 100 U MboI, 100 U Csp6I, and 100 U Hin1II   in 500 µl final volume. All digestions were conducted at 37 oC overnight with rotation. The restriction enzymes were inactivated by incubating at 65 oC for 20 min. After cooling to room temperature, end blunting was performed by adding 3 µl of 10 mM dNTP, 8 µl of DNA polymerase I Klenow, and 4 µl of T4 DNA polymerase, and incubating at 37 oC for 1 hour with rotation. The suspension was then mixed with 460 µl water, 120 µl T4 ligase buffer, 100 µl 10% Triton X-100, 6 µl 20 mg/ml BSA, and 5 µl of 400 U/µl T4 ligase, and incubated for 4 hours at room temperature with rotation. The processed nuclei were pelleted and resuspended in 500 µl 1x T4 ligation buffer supplied with 50 µl 20 mg/ml proteinase K and 50 µl 10% SDS, and incubated at 55 oC for 30 min. For de-crosslinking, 60 µl 5M sodium chloride was added and the mixture was incubated at 68 oC for 2 hours. DNA was recovered using phenol-cholorform extraction.The re-arranged genomic DNA was sonicated to 300-400 bp fragments using Covaris S2 ultrasonicator. The fragmented DNA was double size-selected using 0.40x/1.0x of SPRI beads to remove reads below 100 bp and above 1000 bp, and eluted with a final volume of 70 µl of 10 mM Tris-HCl. The eluent was mixed with 10 µl 10x T4 ligation buffer , 10 µl 100 mM ATP, 5 µl 10 mM dNTP mix, 4 µl T4 DNA polymerase , 1 µl DNA polymerase Klenow, and 5 µl T4 PNK, and incubated for 30 min at room temperature. The repaired DNA was purified by using 1.0x SPRI beads, and eluted with a master mix of 94.5 µl 1x NEB buffer 2 and 0.5 µl 100 mM dATP. After removing the beads, 5 µl of Klenow exo- was added, and the mixture was incubated at 37 oC for 30 min for dA tailing. The processed DNA was purified by using 1.0x SPRI beads, and eluted with 20 µl 10 mM Tris-HCl. After dA-tailing, a custom Y shape adaptor was ligated to the fragments. For amplification, eight 100 µl reactions, each containing 400 µg preamplification library, 2 µM universal primer, 0.5 µM viewpoint-specific outer primer for each multiplexed viewpoint, and 1x SuperFi PCR master mix with 20% GC enhancer, was performed at following condition: 98 oC for 30s, 28 cycles of 98 oC for 10s, 62 oC for 10s, and 72 oC for 60s, and final extention at 72 oC for 5 min. The product was pooled and purified with 1.0x SPRI beads.
4C, Tri-4C

treatment_protocol: To induce IFNB1 expression, cells were treated with 20 µM 2’3’-cyclic GMP-AMP, 100 ng/mL IFN?, and 10 ng/mL TNF? for 24 hours

tag: strain

value: NlaIII


title: IMR90C_NlaIII_Boundary_2

description: Baseline


title: RNAseq_hMADS_shGFP_D2_2


title: IMR90_lowip_Boundary_1

description: Baseline


title: RNAseq_hMADS_shGPS2_h2_3

value: 2 hours


title: RNAseq_hMADS_shGFP_D2_1


title: IMR90C_NlaIII_Boundary_1

description: Baseline


title: IMR90C_NlaIII_MLLT3_1

description: Baseline


title: Day 140

city: Columbus

first: Summer

middle: Rose

last: Fair

state: OH

address: 575 Children's Crossroad

department: Institute for Genomic Medicine

institute: Nationwide Children's Hospital

data_processing:
Basecalls were performed using bcl2fastq (illumina).
Paired-end 150 bp reads were aligned and processed with cellranger count 3.1.0 (10x genomics)
Genome_build: hg38
Supplementary_files_format_and_content: Gene expression matrices are provided from cellranger count

source_name: Cerebral organoid

extract_protocol: none provided by the submitter

tag: model

value: iPSC-derived Cerebral Organoids

value: Thermofisher Human Episomal iPSC Line (A18945)

value: Wildtype


title: ADU.029

description:
NCT00560313
FS00005314-ADU.029

tag: tissue subtype

value: serum-post III 4CmenB doses


title: ADU.028

description:
NCT00560313
FS00005303-ADU.028


title: ribo_QTRT2KO_rep1

city: Hirosawa Wako city, Saitama prefecture, Japan

first: Shintaro

last: Iwasaki

department: Cluster for Pioneering Research

institute: RIKEN

description: Ribosome Profiling

data_processing:
Library strategy: Ribo-seq
Basecalling with illumina CASAVA bcl2fastq.gz (v2.19)
3' adapter trimming with FastX-toolkit
Library sorting based on sample barcode sequence with custom script
Random barcode triming with custom script
rRNA and other non-coding RNA alignment with STAR(2.7.0a)
Alignment with STAR(2.7.0a)
Read quantitation using fp-transcript
Genome_build: Custom mitochondria transcript sequences
Supplementary_files_format_and_content: Text files contain four columns: 1. transcript name and consecutive number;2. Distance from start codon (nt);3. Distance from start codon (codon);4. read counts

extract_protocol: RNA extraction, RNaseI footprinting, ribosome pelletting, 17-34 nt gel excision, preadenylated linker ligation, rRNA depeletion, reverse transcription, cDNA circularization, PCR amplification

tag: genetic background

value: QTRT2 KO


title: ribo_QTRT2KO_rep2

description: Ribosome Profiling


title: Day 93


title: ADU.027

city: Siena

description:
NCT00560313
FS00005301-ADU.027


title: RNAseq_hMADS_shGPS2_h2_2


title: RNAseq_hMADS_shGPS2_h2_4


title: ADO.001

description:
NCT00661713
FS00007610-ADO.001

source_name: vaccinated_Adolescent_serum-post III 4CmenB doses


title: ribo_naive_rep1

description: Ribosome Profiling

value: WT


title: ADU.024

description:
NCT00560313
FS00007687-ADU.024


title: ribo_naive_rep2

description: Ribosome Profiling


title: ADU.026

description:
NCT00560313
FS00007689-ADU.026


title: ADU.025

description:
NCT00560313
FS00007691-ADU.025


title: RNAseq_hMADS_shGPS2_h2_1


title: RNAseq_hMADS_shGFP_h2_3

value: shGFP


title: E1-HEK293

city: Concepción

first: Carlos

last: Farkas

state: Biobío

address: Chacabuco 1140, 1808

department: Bioquímica y Biología Molecular

country: Chile

institute: Universidad de Concepción (UDEC)

data_processing:
Illumina ChIP-seq reads of SALL2 wild type, E1A-knockout cells and SALL2 total KO performed in HEK293 cells were imported to the Galaxy platform (usegalaxy.org) and aligned to the human genome (hg19 assembly) by using bowtie2 aligner with default settings [23]. Correspondent BAM files were sorted (Galaxy Version 1.7) and alignments were filtered with a sliding quality window of Q20 by using SAMtools (Galaxy Version 1.7) [23]. MACS2 program was used to call peaks in these BAM files by using an effective genome size of 2700000000, read extension size of 151, and peak detection based on p-value cutoff of 0.005. Additionally, bigwig files from aligned bam files were obtained by using deeptools bamCoverage tool (Galaxy Version 3.1.2)[24]. Each bigwig file was normalized to 1x method by using the referred effective genome size and a bin length of 50 bp
Genome_build: hg19
Supplementary_files_format_and_content: BigWig

source_name: HEK293 cells

growth_protocol: cells were grown in 100-mm dishes at one million cells per plate

extract_protocol:
Cell nuclei were sonicated to shear DNA in 300 μl of sonication buffer, using a Bioruptor Plus (Diagenode) (18 times, 15 sec on /20 sec off each time, 9 W potency), obtaining lengths between 300 and 600 bp. Immunoprecipitations were carried out overnight at 4 °C using 40 μg of chromatin and 5 μg of SALL2 (anti-SALL2; Bethyl). The SALL2 total knockout was used as a negative control of antibody binding
To prepare each library was used the KAPA Hiper Prep kit and 7 adapters from the KAPA SI Adapters kit, both according to the manufacturer’s protocols. Briefly, the first step in library preparation was to convert any overhangs in the ChIP DNA into phosphorylated blunt ends. The 3′ ends were then adenylated, and adaptors ligated onto the ends of the fragments. The library size (348 bp) and the DNA concentration (14,486 nM) was obtained by using an Agilent 2100 Bioanalyzer

tag: chip antibody

value: Sall2 (Bethyll)


title: ADU.023

address: Via Fiorentina 1

description:
NCT00560313
FS00007683-ADU.023


title: WT-HEK293