title: SKNMC STAG2ko

city: Madrid

first: Daniel

last: Gimenez

address: Calle de Melchor Fernández Almagro, 3

institute: CNIO

data_processing:
Fastq files with single-end sequenced reads were quality-checked with FastQC (S. Andrews, http://www.bioinformatics.babraham.ac.uk/projects/fastqc/) and aligned to the human genome (hg38) with Nextpresso executing TopHat-2.0.0 using Bowtie 0.12.7 and Samtools 0.1.16 allowing two mismatches and five multi-hits  . Finally, differential expressed genes were obtained using DESeq2 and filtered by adjusted p-value<0.05.
Assembly: hg38
Supplementary files format and content: csv/txt

source_name: SKNMC

extract_protocol:
Total RNA was extracted with NZY Total RNA Isolation kit (MB13402) following manufacturer’s instructions.
Total RNA samples (500ng) were processed with the "NEBNext Single Cell/Low Input RNA Library Prep" kit (NEB #E6420) by following manufacturer instructions. RNA Quality scores were 9.9 on average (range 9.1-10) when assayed on a PerkinElmer LabChip analyzer. Briefly, an oligo(dT) primed reverse transcription with a template switching reaction was followed by double stranded cDNA production by limited-cycle PCR.  Non-directional sequencing libraries were completed with the "NEBNext Ultra II FS DNA Library Prep Kit for Illumina" (NEB #E7805) and subsequently analyzed on an Illumina NextSeq 550 with v2.5 reagent kits following manufacturer's protocols.

value: SKNMC

tag: genotype


title: SKNMC WT

status: Public on Sep 18 2024

type: SRA

country: Spain

library_source: transcriptomic

molecule: total RNA

organism: Homo sapiens

tag: cell line


title: A673 siSTAG2 Rep3 Exp4


title: A673 siSTAG2 Rep2Exp4


title: A4573 WT


title: A4573 STAG2 KI

value: iCas9 +STAG2KI


title: A673 WT Rep3 Exp4

value: WT


title: Cycling Tg Treated Ribo-seq, 1

city: Baltimore

first: Manolis

last: Maragkakis

address: 251 Bayview Boulevard

country: USA

institute: National Institute on Aging

data_processing:
FASTQ files were demultiplexed using bcl2fastq conversion software (Illumina) and then aligned to the human genome or transcriptome using STAR v2.7.10b.
Differential gene expression and other analysis was performed using DESeq2 v1.36.0.
Assembly: hg38
Supplementary files format and content: proceesed data files are tab-deliminted outputs from STAR v2.7.10b using gene quant mode
Library strategy: Ribo-seq

growth_protocol: Cells were grown in 10% FBS DMEM high glucose media supplemented with non-essential amino acids

extract_protocol:
RNA was extracted using by drop-wise addition of polysome lysis buffer (20 mM Tris pH 7.4, 50 mM NaCl, 5 mM MgCl2, 1 mM DTT, 50 mM KCl, 1% Triton x-100, 0.1 mg/mL cyclohexamide, 1X HALT protease-phosphatase inhibitors). 50 ug of bulk RNA from lysate (measured by nanodrop) was used for Trizol purification of RNA-seq and 50 ug of lysate was prepared for Ribo-seq. Ribo-seq lysates were treated with 1 uL of RNAse I (thermofisher) and incubated at 4C with rotation for 40 min; reaction was then stopped with 100 U of SUPERasein (thermofisher). Lysates were then centrifuged through a sucrose cusion to precipitate ribosome pellets. Pellets weree then suspended in Trizol for RNA preparation. Prepared RNA was DNAse I treated and then gel purified for riboseq footprints (25-40 nts) followed by extraction. Extracted RNA was then depleted of rRNA (thermofisher).
RNA-seq RNA was prepared using Tru-Seq Stranded RNA library kit (Illumina) while Ribo-seq RNA was prepared using NEBNext Multiplex Small RNA Library prep (New England Biolabs)

treatment_protocol: CTg cells were grown to 40-60% confluency and then treated with 25 nM of thapsigargin (Tg) for 3h prior to harvest.

tag: cell type

value: Fibroblast

tag: rna or_ribo

value: Ribosome Protected Fragments


title: A673 siSTAG2 Rep1 Exp4


title: Cycling Tg Treated RNA-seq, 1

data_processing:
FASTQ files were demultiplexed using bcl2fastq conversion software (Illumina) and then aligned to the human genome or transcriptome using STAR v2.7.10b.
Differential gene expression and other analysis was performed using DESeq2 v1.36.0.
Assembly: hg38
Supplementary files format and content: proceesed data files are tab-deliminted outputs from STAR v2.7.10b using gene quant mode


title: Cycling Tg Treated RNA-seq, 2


title: Cycling Tg Treated Ribo-seq, 2


title: A673 WT Rep2 Exp4


title: A_1_stomach_0d

city: Wien

first: Andre

last: Oszwald

address: Währinger Gürtel 18-20

country: Austria

institute: Medical University of Vienna

data_processing:
BCL files were converted to FASTQ files using the Illumina bcl2fastq app (v2.2)
FASTQ files were converted to .DCC files and extracted as a matrix using the Nanostring NGS pipeline and GeoMxTools R package, respectively.
Assembly: n.a. (barcodes only; corresponding to unique probes and regions)
Supplementary files format and content: .dcc: processed counts (used for input in GeoMxTools R package)
Supplementary files format and content: .stats: statistics output of Nanostring GeoMx NGS pipeline

source_name: stomach

extract_protocol:
Sections were deparaffinized and subjected to heat-induced protein epitope retrieval (using retrieval buffer at pH 9, Thermo Fisher Scientific, 00-4956-58) for 15 minutes at 100°C using a steam cooker. Retrieval for mRNA was performed using proteinase K (Thermo Fisher Scientific, 25530049) at 1µg/ml for 15 minutes. Slides were incubated with the human Whole Transcriptome Atlas (WTA) probe panel (Nanostring, NA-GMX-RNA-NGS-HuWTA-4, Lot HWTA21003) at 37°C in a humidified environment overnight. Sections were washed according to standard protocols and direct immunofluorescence was performed with antibodies against CD34 conjugated to Alexa-647 (Novus Biologicals, clone QBEnd-10, NBP2-34713AF647), Pan-cytokeratin conjugated to to Alexa532 (Novus Biologicals, clone AE1/AE3, NBP2-33200), CD45 conjugated to Alexa594 (clone 2B11 + PD7/26, Novus Biologicals, NBP2-34528), and a nuclear counterstain (Syto-13, Nanostring). DSP device run was performed according to the manufacturer's protocol (https://university.nanostring.com/geomx-dsp-ngs-readout-user-manual).
Library preparation and sequencing were performed according to the manufacturers’ protocol (https://university.nanostring.com/geomx-dsp-ngs-readout-user-manual). The library was sequenced using an Illumina NextSeq 550 sequencer with the specifications provided by Nanostring (paired-end reads at length 27, index length 8, PhiX 5%) at a final library concentration of 1.6 pM using a NextSeq 550 High Output 75-cycle kit (Illumina, 20024906).

tag: tissue

value: stomach

tag: case

value: A

tag: specimen

tag: duration edta

tag: roi


title: C.iS_3

city: Wageningen

first: Benthe

last: van der Lugt

address: Akkermaalsbos 2

country: Netherlands

institute: Wageningen University & Research

data_processing:
RNA-seq reads were used to quantify transcript abundances. To this end the tool Cutadapt (version 1.16) was used to trim adapters from the reads and HISAT2 (version 2.2.1) was used for mapping the reads to the GRCh38.p13 human genome assembly-based transcriptome sequences as annotated by the GENCODE consortium. The output from HISAT2 was subsequently converted and sorted by chromosomal position using Samtools (version 1.9).  RSeQC  (v3.0.1) and  PRINSEQ  (v0.20.4) were used for quality control. HTSeq (version 0.11.2) was applied to count reads in transcripts using gene-level quantification. Prior to conducting statistical analyses, a general filtering of the count table was conducted to increase detection power. This filtering was based on the criterion that a gene must exhibit an expression level exceeding approximately 10 counts in a minimum of 3 samples. Differences in library size were adjusted using the trimmed mean of M-values normalization method, which was implemented in the edgeR package (version 3.36.0).
Assembly: GRCh38.p13
Supplementary files format and content: Text file including the normalized counts for each sample.

growth_protocol: Caco-2 cells were maintained in DMEM supplemented with 10% heat-inactivated FBS and 1% penicillin/streptomycin. Cells were kept at in a humidified incubator at 37⁰C and 5% CO2. To start differentiation, Caco-2 cells were seeded at a density of 4 x 105 cells/cm2 in 12-well cell culture inserts with a polyester membrane (0.4µm pore, Corning, New York, NY). The apical and basolateral compartments were filled with 500 µL and 1500 µL cell culture medium, respectively, and medium was refreshed every 2 to 3 days. Cells were differentiated for 21 days. hiPSCs were cultured on human embryonic stem cell qualified Matrigel-coated (Corning #354277) 6-well plates in mTeSR1 medium (Stem Cell Technologies, Vancouver, Canada). hiPSCs were dissociated into single cells, seeded in 24-well plates (220,000 cells/well) and differentiated towards definitive endoderm (DE). Subsequently, cells were differentiated to intestinal stem cells (ISCs), dissociated again and seeded on 12-well Transwell inserts with a polyester membrane (0.4 µm, Corning) coated with Growth Factor Reduced Matrigel (Corning #356231) at a density of 2.5 x 105 cells per insert. The cells were cultured for 12 days in differentiation medium (DMEM/F-12 containing 2% fetal bovine serum, 1% non-essential amino acids (Gibco), 1 × B27 (Gibco), 1 × N2 (Gibco), 1% penicillin/streptomycin (Gibco) and 2 mM L-glutamine (Gibco)), supplemented with 20 ng/ml EGF, 30 µM Forskolin, 5 µM 5-aza-2′-deoxycytidine (Sigma), 20 µM PD98059 (Stem Cell Technologies), and 0.5 µM A-83-01 (Sigma). This medium was used throughout the differentiation period and was refreshed every 2 to 3 days (400 µl apical and 1200 µl basolateral).

extract_protocol:
RNA was extracted from Caco-2 cells and hiPSC-derived IECs using the RNeasy Mini Kit. RNA integrity was analyzed using a total RNA Pico chip in an Agilent 2100 Bioanalyzer.
After poly(A) selection, library preparation was performed using the NEBNext Ultra II RNA Library Preparation kit.

treatment_protocol: IECs and Caco-2 cells were challenged with a pro-inflammatory cytokine cocktail, comprising exposure to 2 ng/mL human interferon gamma (IFN-γ) protein for 24 hours, followed by exposure to 10 ng/ml human tumor necrose factor-alpha (TNFα) protein and 1 ng/ml human interleukin 1 beta (IL-1β) protein for 48 hours. Cells were exposed to these cytokines in both the apical and basolateral compartment.  Exposure to L. plantarum took place 24h prior to the challenge. Cells were exposed to stationary or heat-killed L. plantarum for 96 hours, during which medium (and thus bacteria) was refreshed after 48 hours.

tag: treatment

value: challenged, exposed to stationary L. plantarum


title: A673 WT Rep1 Exp4


title: C.iS_2


title: C.iS_1


title: C.iH_3

value: challenged, exposed to heat-killed L. plantarum


title: C.iH_2


title: A673 STAG1ko clone3 Exp3


title: C.iH_1


title: C.iC_3

value: challenged, unexposed


title: C.iC_2


title: A673 STAG1ko clone1 Exp3


title: C.iC_1


title: A673 WT-Parental Exp3


title: A673 WT Rep1 Exp3


title: C.hH_2

value: unchallenged, exposed to heat-killed L. plantarum


title: C.hH_1


title: A673 SUZ12ko clone3 Exp2


title: A673 SUZ12ko clone2 Exp2


title: A673SUZ12ko clone1 Exp2


title: A673 EZH2ko clone3 Exp2


title: A673 EZH2ko clone2 Exp2


title: C.hS_1

value: unchallenged, exposed to stationary L. plantarum


title: A673 STAG1ko clone2 Exp3


title: C.hS_2


title: A673 EZH2ko clone1 Exp2


title: C.hC_2

value: unchallenged, unexposed


title: C.hC_1


title: C.hC_3


title: A673 WT Rep2 Exp2


title: I.iH_1

source_name: CS83iCTR-33n1 hiPSC

value: CS83iCTR-33n1 hiPSC


title: A673 STAG2ko clone2 Exp2


title: I.iH_3


title: A673 STAG2ko clone5 Exp2


title: A673 STAG2ko clone3 Exp2


title: A673 WT Rep1 Exp2


title: I.iS_3


title: I.iS_2


title: C.hH_3


title: C.hS_3


title: I.iC_3


title: I.iC_2


title: A673 STAG2ko clone1 Exp1


title: A673 STAG2ko clone4 Exp1


title: A673 WT-Parental Exp1


title: I.iS_1


title: A673 WT Rep2 Exp1


title: I.iH_2


title: A673 WT-Parental Exp2


title: A673 STAG2ko clone2 rep2 exp2

library_source: genomic

data_processing:
We performed two independent experiments with two technical replicates for each of the four cell lines used. Each sample was analyzed using the Arima Capture Hi-C pipeline, aligning to the hg38 genome at 3-kb resolution. The resulting BAM files from the technical replicates and the different experiments were merged to create a single file for each cell line (4 in total).
To allow comparisons between the four conditions, we performed downsampling to equalize the number of reads that aligned with the Arima probes (% on-target). Loops were called using ChiCAGO for the individual cell lines with at least 5 reads in the interaction and at least a score >3, resulting in more than 200,000 loops called in each condition .
Assembly: hg38
Supplementary files format and content: ibed

molecule: genomic DNA

extract_protocol:
Promoter Capture Hi-C was performed using the Arima HiC Kit (A510008) and the Arima Human Promoter Capture HiC Kit, which contains probes to 23,711 human promoters  (GRCh38 ensemble database, version 95).
We used 7-8 million cell for each condition (P, WT#2, KO#1 and KO#2) and 10-11 PCR cycles to prepare Hi-C libraries. Post-capture libraries were amplified with 11-12 PCR cycles.

value: iCas9 STAG2ko


title: A673 STAG2ko clone2 rep1 exp2


title: I.hH_2


title: I.iC_1


title: I.hH_1


title: A673 STAG2ko clone1 rep1 exp2


title: Control A673 rep2 exp2


title: Control A673 rep1 exp2


title: I.hC_2


title: Parental A673 rep2 exp2


title: I.hS_1


title: I.hC_1


title: A673 STAG2ko clone2 rep2 exp1


title: A673 STAG2ko clone1 rep2 exp1


title: A673 STAG2ko clone1 rep1 exp1


title: midgut,W22,rep3

city: Shenzhen

first: Mei

last: Luo

state: Gangdong

address: Dapeng

country: China

institute: Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences

data_processing:
Six gigabytes of Illumina PE150 raw reads were obtained for each sample. After filtering out adaptor and low-quality reads, the cleaned reads were mapped to the B73 v4 reference genome using the STAR 2.4.0j software
The original fluorescence image files obtained from Illumina platform are transformed to short reads (Raw data) by base calling and these short reads are recorded in FASTQ format (Cock P. et al, 2010), which contains sequence information and corresponding sequencing quality information.
Sequence artifacts, including reads containing adapter contamination, low-quality nucleotides and unrecognizable nucleotide (N), undoubtedly set the barrier for the subsequent reliable bioinformatics analysis. Hence quality control is an essential step and applied to guarantee the meaningful downstream analysis. we used Fastp (version 0.23.1)(Chen S. et al, 2018) to perform basic statistics on the quality of the raw reads.
The steps of data processing were as follows: (1) Discard a paired reads if either one read contains adapter contamination; (2) Discard a paired reads if more than 10% of bases are uncertain in either one read; (3) Discard a paired reads if the proportion of low quality (Phred quality
Assembly: FAW reference genome: GCA_011064685.2
Supplementary files format and content: Format: txt
Supplementary files format and content: Content: The processed data file contains reads numble for each sample. Rows represent genes, and columns represent individual samples.

organism: Spodoptera frugiperda

source_name: midgut

growth_protocol: S. frugiperda (fall armyworms, FAW) were originally collected from maize fields in Yunnan Province, China, in 2019 and were reared in the laboratory for several generations under controlled experimental conditions. The larvae of these species were reared on an artificial diet in an insect rearing chamber with a 14:10 light/dark photoperiod, a temperature of 25°C ± 2°C, and a relative humidity of 60% ± 5%. Maize seedings were grown in 1-liter pots filled with a growing medium composed of a 5:1 mixture of potting soil and vermiculite. Plants were cultivated in a growth chamber with a 16:8 light/dark photoperiod and temperature set at 24 ± 2°C during the day and 20 ± 2°C at night. The plants were watered regularly and were used for experiments at the two- to three-week-old stage, with the second leaf fully developed and expanded from the whorl.

extract_protocol:
Total RNA was extracted from the midgut samples using the TRIzol reagent kit (Invitrogen, Waltham, MA, USA) according to the manufacturer’s instructions.
Total RNA was used as input material for the RNA sample preparations. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in First Strand Synthesis Reaction Buffer. First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase(RNase H-). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3’ ends of DNA fragments, Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 370~420 bp in length, the library fragments were purified with AMPure XP system. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified.
After the library construction is completed, preliminary quantification is carried out using Qubit, and the library is diluted to 1.5ng/ul. Subsequently, the inserted fragments of the library are detected. Once the inserted fragments meet expectations, the effective concentration of the library is accurately quantified using qRT PCR to ensure the quality of the library. The qualified libraries were pooled and sequenced on sequencing platforms, according to effective library concentration and data amount required.

treatment_protocol: To investigate the gene expression profiles of FAW under different feeding conditions, fourth-instar larvae of each species were fed fresh maize leaves (W22) for 24 hours. Larvae fed with artificial feed (AF) served as controls. After the feeding period, midguts were rapidly dissected and frozen in liquid nitrogen for further analysis. Each treatment consisted of three biological replicates, with five midguts pooled per replicate.

value: midgut


title: I.hH_3


title: Organoid, CDYL KO, Day30, input

city: Beijing

first: Liang-Yu

last: Ni

state: Beijing

address: Peking university Health Science center

institute: Peking University

data_processing:
Raw data (raw reads) of fastq format were firstly processed using fastp (version 0.19.11, Chen et al., 2018)  software. In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content of  the clean data were calculated. All the downstream analyses were based on the clean data with high quality.
Reference genome and gene model annotation files were downloaded from genome website directly. Bowtie 2 were used to align the raw data with the reference genome. The SAM file generated from Bowtie2 was then converted to a BAM file using SAMtools. Next, we generated coverage files in the BigWig (.bw) format for visualization in genome browsers such as IGV or UCSC Genome Browser. This was accomplished using bedtools and deepTools.
Assembly: hg18 & mm19
Supplementary files format and content: .bw

source_name: Organoid, CDYL KO

extract_protocol:
ChIP DNA degradation and contamination was monitored on agarose gels. DNA purity was checked using the NanoPhotometer® spectrophotometer (IMPLEN, CA, USA). DNA concentration was measured using Qubit® DNA Assay Kit in Qubit® 3.0 Flurometer (Life Technologies, CA, USA).
The purified DNA was used for ChIP-seq library preparation. The library was constructed by Novogene Corporation (Beijing, China). Subsequently, pair-end sequencing of sample was performed on Illumina platform (Illumina, CA, USA). Library quality was assessed on the Agilent Bioanalyzer 2100 system.

value: Organoid, CDYL KO

value: Cortical organoids

value: CDYL Knock out

tag: chip antibody

tag: time

value: Day 30


title: Organoid, CDYL KO, Day30, CDYL antibody

value: CDYL (Proteintech, 17763-2-AP)


title: Organoid, Control, Day30, input

source_name: Organoid, Control

value: Organoid, Control

value: Flag-CDYL Konck in


title: Control A673 rep2 exp1


title: A673 WT Rep1 Exp1


title: I.hS_2


title: midgut,W22,rep2


title: A673 STAG2ko clone1 rep2 exp2


title: Control A673 rep1 exp1


title: Organoid, Control, Day30, Flag antibody

value: Flag (Sigma, F1804)


title: I.hS_3


title: Parental A673 rep2 exp1

value: WT


title: midgut,W22,rep1


title: Organoid, Control, Day30, CDYL antibody, Rep2


title: Parental A673 rep1 exp2


title: Parental A673 rep1 exp1


title: A673 STAG2ko, H3K27me3 ChIP-Seq

data_processing:
Alignment of reads to the reference human genome (hg38) was performed using ‘Bowtie2’ (version 2.4.2) under default settings . Duplicates were removed using GATK4 (version 4.1.9.0) and peak calling was carried out using MACS2 (version 2.2.7.1) after setting the q value (FDR) to 0.05 and using the ‘–extsize’ argument with the values obtained in the ‘macs2 predictd’ step (Zhang et al., 2008)
For analysis of calibrated ChIP-seq, profiles for each antibody were normalized by coverage and then multiplied by the occupancy ratio (OR) = (WmIPh)/(WhIPm), where Wh and IPh are the number of reads mapped to the human genome from input (W) and immunoprecipitated (IP) fractions, and Wm and IPm are reads mapped to the mouse genome from the input and IP fractions used for calibrating
Assembly: hg38
Supplementary files format and content: bw

extract_protocol:
A673 cells grown at high confluence were crosslinked with 1% formaldehyde for 15 min at room temperature. After quenching the reaction with 0.125 M Glycine, fixed cells were washed twice with PBS containing 1μM PMSF and protease inhibitors. Cells were lysed in lysis buffer (1% SDS, 10 mM EDTA, 50 mM Tris-HCl pH 8.1) at a concentration of 2x107 cells/ml. Sonication was performed with a Covaris S220 (shearing time 20 min, 20% duty cycle, intensity 6, 200 cycles per burst and 30 sec per cycle) in a minimum volume of 2 ml. Chromatin from 107 cells was incubated with 5 μg of antibody as described . For calibration, 5% of chromatin from mouse ES cells was added to the human chromatin
For library preparation, at least 5 ng of DNA were processed through subsequent enzymatic treatments with "NEBNext Ultra II FS DNA Library Prep Kit for Illumina" from New England BioLabs (cat# E7805). Briefly, a short fragmentation of 10 min was followed by end-repair, dA-tailing, and ligation to adapters. Adapter-ligated libraries were completed by limited-cycle PCR (8-12 cycles). Resulting average fragment size is 300 bp from which 120 bp correspond to adaptor sequences. Libraries were applied to an Illumina flow cell for cluster generation and sequenced on Illumina NextSeq 500 (with v2.5 reagent kits) following manufacturer's recommendation


title: I.hC_3


title: A673 STAG2ko, Input ChIP-Seq


title: midgut,AF,rep3


title: A673 WT, H3K27me3 ChIP-Seq


title: A673 STAG2ko clone2 rep1 exp1


title: midgut,AF,rep2


title: A673 WT, Input ChIP-Seq


title: Untr_HCTaidRPB1_H3K4me3_CnT_rep2

city: Research Triangle Park

first: Jackson

middle: Andrew

last: Hoffman

state: North Carolina

address: 111 TW Alexander Dr, Building 101 Room C432

department: DIR

institute: NIEHS

data_processing:
ChIP-seq analysis. Reads were trimmed using Cutadapt and aligned to both hg19 and dm6 genomes using bowtie2. Aligned reads were deduplicated with picardtools. Scale factors were calculated for each sample by dividing the minimum number of dm6 reads for a given antibody by number of dm6 reads for each sample using the same antibody. Scaled and normalized coverage files were generated using deeptools with calculated scale factors and rpkm normalization.
Reads were trimmed using Cutadapt and aligned to both hg19 and mm10 genomes using bowtie2. Aligned reads were deduplicated with picardtools. Deduplicated read counts were used to calculate the human:mouse ratio, which was then used to calculate scaling factors for samples performed side by side with the same antibody. The scale factor was calculated by dividing the human:mouse read ratio for each sample using a given antibody by the maximum human:mouse read ratio from samples using the same antibody. Scaled and normalized coverage files were generated using deeptools with calculated scale factors and rpkm normalization.
Assembly: hg19
Supplementary files format and content: bigwig coverage files
Library strategy: CUT&TAG

growth_protocol: Human T47D A1-2 (Archer et al., 1994) and MCF-7 cell were grown in MEM (Gibco 10370) supplemented with 10% fetal bovine serum (Gibco 26140-079), GlutaMAX (Gibco 3505), HEPES (Sigma H0887), and Penicillin/Streptomycin (Sigma P0781).  HeLa, SW-13, U2OS, primary human fibroblast and mouse C127 cells were grown DMEM (Gibco 11965-092) supplemented with 10% fetal bovine serum (Gibco 26140-079), GlutaMAX (Gibco 3505), HEPES (Sigma H0887), and Penicillin/Streptomycin (Sigma P0781).  K562 cells were grown in suspension in RPMI Medium 1640 (Gibco 11875-093) supplemented with 10% fetal bovine serum (Gibco 26140-079), GlutaMAX (Gibco 3505), HEPES (Sigma H0887), and Penicillin/Streptomycin (Sigma P0781).  All cells were cultured in a humidified incubator at 37°C with 5% CO2 in the indicated growth medium with media changed every 2-3 days.  Cells were seeded in new tissue culture treated dishes when 80% confluency reached.

extract_protocol:
Cells were fixed in PBS with 1% methanol-free formaldehyde (ThermoFisher Scientific 28906) at 37C for 10 min and quenched with glycine (0.125M) for 5 min at room temperature.  After glycine quench, cells were washed three times and scraped into ice-cold PBS and pelleted by centrifugations at 500g for 5 min.  Compact cell pellets were resuspended in MNase swelling buffer (25 mM HEPES pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40, 0.5 mM PMSF, and 1x cOmplete EDTA-free protease inhibitors (PIC)) (Roche), incubated on ice for 10 min then subjected to Dounce homogenization using 20-strokes with “tight” pestle (Duran Wheaton Kimble, 357542).  Nuclei were sedimented through digestion buffer (15mM Hepes pH 7.9, 60mM KCl, 15mM NaCl, 0.32M Sucrose, 0.5 mM PMSF, and PIC) by centrifugation for 930g for 7 min at 4C. Nuclei pellets were fully resuspended in digestion buffer containing 3.3 mM CaCl2 and digested with MNase (0.75U/1x10ˆ6 cells, Worthington, cat#) at 37C for 15min. MNase fragmentation was stopped by addition of 10 mM EGTA and incubation for 5 min on ice.  Digested nuclei were centrifuged at 930g for 7 min at 4C and compact nuclei pellets resuspended in 20 volumes (v/v) ChIP Shearing buffer (50 mM HEPES pH7.9, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS containing spermidine, PIC and PMSF) and incubated on ice for 10 min.  Samples were subjected to sonication using PIXUL Multi-Sample Sonicator (Active Motif) at 12C using the following parameters: Pulse [N]: 50; PRF [kHz]: 1.00; Process Time: 10 min; Burst Rate [Hz]: 20.00.  After sonication, fragmented chromatin was recovered in supernatant after centrifugation at 18,000g for 10 min with extent of fragmentation determined by agarose gel electrophorese prior to immunoprecipitation.  The concentration of fragmentated chromatin determined by Bradford assay.  Fragmentated chromatin was diluted in two-fold in 2xIP buffer (20 mM Tris-HCl pH 8.0, 300 mM NaCl, 2 mM EDTA, 20% Glycerol, 1% Triton X-100, 0.5 mM PMSF and PIC) and immunoprecipitation performed with the indicated antibodies at a ratio of 1 ug antibody per 200 ug chromatin.  Equal amounts of Spike-IN antibody and Drosophila chromatin (Active Motif 61686 and 53083) added to each immunoprecipitation for normalization.  Immune complexes were captured using protein G dynabeads (Invitrogen 10004D), washed once each with low salt buffer (20 mM Tris-HCl pH 8.0, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 0.1% SDS), high salt buffer (same as low salt buffer, except 500 mM NaCl), and LiCl buffer (Tris-HCl pH 8.0, 250 mM LiCl, 2 mM EDTA, 1 % NP-40, 1% (wt/vol) sodium deoxycholate), and twice with TE.  Immunoprecipitated DNA recovered from beads by incubation in ChIP Elution Buffer (1% SDS, 0.1M Sodium Bicarbonate) for 30 min at room temperature.  Eluted DNA was treated with RNaseA (Roche 10109169001) and Proteinase K (Invitrogen #25530015) and purified using Qiagen PCR purification columns.  ChIP-seq in K562 cells with milder cross-linking and fragmentation performed using a SimpleChIP Plus Enzymatic Chromatin IP Kit with Magnetic Beads (Cell Signaling Technologies #9005S) according to manufacturer’s protocol.
Cells were treated, harvested, and subjected to CUT&TAG analysis using a modified published protocol [Kaya-Okur et al., 2019].  Trypsinized cells were washed once with PBS and incubated with NE nuclear extraction buffer (20 mM HEPES-KOH, pH 7.9, 10 mM KCl, 0.1% Triton X-100, 20% Glycerol, 0.5 mM Spermidine, 100 uM PMSF, and 1x EDTA-free protease inhibitors (Roche cOmplete) for 30 min on ice followed by Dounce homogenization using 20-strokes from Dounce Tissue Grinder (Wheaton 357542) tight pestle.  Following homogenization, samples were centrifuged for 3 min at 600g, nuclei pellet resuspended in NE buffer supplemented with 0.1 % formaldehyde and incubated at room temperature for 2 minutes followed by quench with 0.125 M glycine.  Following formaldehyde cross-link, nuclei were re-centrifuged, resuspended in fresh NE buffer and counted with trypan blue dye using Bio-Rad TC20 Automated Cell Counter.   After isolation, 150,000 total nuclei [135,000 nuclei from experimental sample with 15,000 mouse spike-in nuclei] were incubated with activated concanavalin A-coated (ConA) magnetic beads (EpiCypher) in 0.2 ml thin-walled tubes for 10 min at room temperature.  Nuclei-bound ConA beads were resuspended in antibody incubation buffer (20 mM HEPES, pH 7.5, 150 mM NaCl, 0.01% Digitonin, 2 mM EDTA, 0.5 mM Spermidine, 100 uM PMSF, and 1x EDTA-free protease inhibitor), 0.75 ug indicated primary antibody (see Table S2) added and samples incubated overnight at 4C with gently rotation.  Primary antibody removed from ConA bead solution by placing samples on magnetic stand until clear followed by supernatant removal.  Samples were incubated with 0.75 ug species specific secondary antibody for 30 min at room temperature with gentle rotation followed by two washes with Digitonin150 wash buffer (20 mM HEPES, pH 7.5, 150 mM NaCl, 0.01% Digitonin, 0.5 mM Spermidine, 100 uM PMSF, and 1x EDTA-free protease inhibitor) then incubation with pAG-Tn5 in Digitonin300-wash buffer (20 mM HEPES, pH 7.5, 300 mM NaCl, 0.01% Digitonin, 0.5 mM Spermidine, 100 uM PMSF, and 1x EDTA-free protease inhibitor) for 1h at room temperature.   After pAG-Tn5 incubation, bound ConA beads were washed twice with Digitonin300-wash buffer then resuspended in Tagmentation buffer (Digitonin300 wash buffer containing 10 mM MgCl2) and incubated for 1h at 37C.  Beads were then pelleted, resuspended in TAPS buffer (10 mM TAPS pH 8.5, 0.2 mM EDTA).  TAPS buffer removed and beads carefully resuspended in SDS Release buffer (10 mM TAPS pH 8.5, 0.1% SDS) and incubated for 1 h at 58 C.  Following incubation, SDS Quench buffer (0.67% Triton X-100) was added to each sample along with 2 μl of a universal i5 and a uniquely barcoded i7 primers (from 10 μM stocks).  Equal volume (25 ul) NEBNext High-Fidelity 2x PCR Master mix (M0541, New England Biolabs) was added to each sample and mixed by gentle pipetting.  Samples subjected to DNA amplification using a thermocycler and the CUT&TAG-specific PCR cycling parameters: 58C for 5 min; 72C for 5 min; 98C for 45 sec; with 19 cycles of 98C for 15 sec and 60C for 10 sec; final extension at 72 °C for 1 min; and hold at 4C.  DNA clean-up performed by adding 1.3x AMPure XP beads (A63880, Beckman Coulter) to each sample, allowing libraries to incubated with beads for 10 min at room temperature, followed by two washes with 80% ethanol, then elution in 15 μl of 10 mM Tris pH 8.0.  While on magnetic stand, supernatant was carefully taken and transferred to a fresh tube.  Library size distribution determined by Agilent TapeStation 4200 (Agilent Technologies) and libraries mixed to yield equal representation prior to paired-end (2 x 50 bp) Illumina sequencing by NIEHS The Epigenomics and DNA Sequencing Core Facility.
Two replicates were used for each treatment condition. Cells were collected and subjected to the ATAC-seq protocol described by Buenrostro et al (2013).  Isolated cells were treated in 10 mM PIPES, pH 6.8, 100 mM NaCl, 300 mM sucrose, 3 mM MgCl2, and 0.1% Triton X-100 and then treated with the Illumina Tagment DNA TDE1 Enzyme and Buffer Kits for 30 min followed by mixing every 10 min. Libraries were sequenced at the NIEHS Epigenomics Core Facility on a NovaSeq instrument
ChIP-seq libraries were generated using the xGen ssDNA & Low-Input DNA Library Prep Kit and xGen CDI Primers (IDT 10009817 and 10009815) according to manufacturer’s protocol and sequenced on the Illumina NextSeq platforms.

treatment_protocol: Cells were treated for the indicated time individually, or in combination, with 100 nM Dexamethasone (Sigma D4902) or vehicle (equal volume EtOH or DMSO), 0.5 uM Triptolide (Tocris 3253), 0.5 uM Flavopiridol (Tocris 3094), 0.5 uM A485 (Tocris 6387), 0.1 uM Trichostatin A (Sigma T8552), 10 ug/ml Doxycycline (Sigma D9891), or 1 uM 5-Ph-IAA (AID) ligand (Sigma SML3574).

value: colorectal carcinoma

tag: spike-in control

value: Mouse nuclei

value: H3K4me3 (Diagenode C15410003)

tag: treatment 1

value: DMSO

tag: treatment 2

value: 10 ug/ml Doxycycline


title: midgut,AF,rep1


title: Aux_HCTaidRPB1_H3K4me3_CnT_rep2

value: 1 uM 5-Ph-IAA


title: BlisterFluid91, BURN, rep1

city: Perth

first: Andrew

last: Gibson

state: Western Australia

address: Building 390, Discovery Way, Murdoch University

country: Australia

institute: Murdoch University

library_source: transcriptomic single cell

data_processing:
Single-cell sequencing data were processed using CellRanger v6.1.2 to demultiplex individual cell barcodes, remove PCR duplicates, and align reads to the reference GRCh38 human transcriptome. The individual hashes within each 10x run were further demultiplexed using Souporcell to cluster samples based on SNPs in the RNA-seq reads. The hashtag negatives or doublets, as well as cells with >50% mitochondrial content were removed to filter low quality, dead, or dying populations for initial QC, with a second pass filter excluding apoptotic populations characterized by a low percentage of ribosomal genes and a high percentage of mitochondrial genes, using a defined mitochondrial-ribosomal RNA ratio of >0.47. A standardized mitochondrial DNA threshold of 10% is recommended for human tissue, which was met by 98% of ‘live’ cells identified. The remaining 2% of cells were excluded. Downstream transcriptome-based graph clustering and PCA with cell phenotype calling were performed using R Seurat v4.1.1 package, without input of TCR, BCR, mitochondrial, ribosomal, or sex-linked genes to avoid bias due to known expression variability. Batch correction for 10x run, sample type (skin or blister fluid), and cell cycle phase were performed using harmony. Experimental doublets were identified and removed using a majority consensus benchmark of three independent bioinformatic algorithms (DoubletFinder, scDBLfinder, scDS).
Assembly: GRCh38
Supplementary files format and content: Normalized_Counts.csv : This is the normalized count matrix derived from Seurat's normalize method for all the single-cells included in the project.
Supplementary files format and content: Metadata_CITESeq+TCR.tsv : This tab-delimited text file contains for each cell barcode - sample meta-data  (such as age, sex, etc.) , CITESeq antibody counts and TCR alpha/beta chains.

molecule: polyA RNA

source_name: Blister fluid

extract_protocol:
Cryopreserved skin biopsies were thawed and digested using published methods optimized for lymphoid cell recovery and single-cell sequencing (RPMI 1640 medium with 10% FBS, 1mg/ml collagenase P and 40ug/ml DNAase I with agitation for 90 minutes). Tissue lysates were washed and passed through a 70µm Flowmi™ Cell Strainer. Single-cell suspensions of enzymatically-digested skin or thawed unprocessed blister fluid were then subject to surface antibody staining for a panel of up to 137 functional and surface lineage markers in the TotalSeq™-C Human Universal Cocktail, V1.0 (Biolegend) as per manufacturer’s protocol. 1µl of the appropriate hashtag antibody was also added, with subsequent incubation for 30mins at 4°C. Each cell-antibody cocktail was then resuspended in 3.5ml cell staining buffer and washed three times in PBS before cells were passed through a 70µm Flowmi™ Cell Strainer. Cell count and viability were recorded and cells were diluted as per the manufacturer’s instruction for loading onto the 10X Chromium Next GEM chip K.
Cells were loaded onto the chromium Next GEM chip K chip at the manufacturer (10x Genomics) recommended concentration and single-cell libraries made using a chromium controller and Chromium Next GEM Single Cell 5' Reagent Kit, Version 2.0 (10x Genomics) as per manufacturer instruction (https://www.10xgenomics.com/support/single-cell-immune-profiling/documentation/steps/library-prep/chromium-single-cell-5-reagent-kits-user-guide-v-2-chemistry-dual-index-with-feature-barcoding-technology-for-cell-surface-protein-and- immune-receptor-mapping), with a target of 1,000-20,000 cells. 5’ libraries were then sequenced using the Illumina NovaSeq 6000 platform.

value: Blister fluid

tag: Sex

value: female


title: BlisterFluid90, BURN, rep1


title: Aux_HCTaidRPB1_H3K4me3_CnT_rep1


title: BlisterFluid89, BURN, rep1


title: 8E LiCl rep 3

city: Worcester

first: Jennifer

last: Benanti

state: MA

address: 364 Plantation St., LRB 525

department: Molecular, Cell and Cancer Biology

institute: University of Massachusetts Medical School

data_processing:
Paired-end reads were aligned to Saccharomyces_cerevisiae.R64-1-1, with star_2.5.3a, annotated with Saccharomyces_cerevisiae.R64-1-1.90.gtf.
Aligned exon fragments with mapping quality higher than 20 were counted toward gene expression with featureCounts. Differential expression (DE) analysis was performed with DESeq2. Within DE analysis, 'ashr' was used to create log2 Fold Change (LFC) shrinkage for each comparison. Significant DE genes (DEGs) were filtered with the criteria FDR < 0.05.
Assembly: Saccharomyces_cerevisiae.R64-1-1,
Supplementary files format and content: csv, raw count and TPM

organism: Saccharomyces cerevisiae

source_name: haploid yeast

growth_protocol: All strains are in the BY4741 background and were grown in synthetic media without uracil. Starter cultures were grown in medium contianing 2% galactose, to induce expression of the genomic copy of HCM1 from the GAL1 promoter, which is integrated upstream of the HCM1 coding sequence. At the start of the experiment, samples were diluted into medium contianing 2% dextrose to shut-off expression of the genomic copy of HCM1. Throughout the experiment all Hcm1 is expressed from centromeric plasmids expressing WT or mutant HCM1 from the HCM1 promoter.

extract_protocol:
Total RNA was purified using an acid-phenol extraction protocol.
poylA enrichment and library construction were performed by Innomics/BGI Americas

treatment_protocol: Cells were grown in medium with or without 150mM LiCl. Samples were diluted after 24 hours, and subsequently collected at the 40 hour time point when all cultures were in logarithmic growth.

value: haploid yeast

value: MATa his3delta1 ura3delta0 leu2delta0 lys2delta0 HIS3MX6-GAL1p-HCM1-3HA-KanMX ChrVIdelta181901-182001::Hyg-TEFp-GFP + pRS316 HCM1p-hcm1-8E-3V5

value: 150mM LiCl


title: BlisterFluid88, SCAR, rep1


title: 8E LiCl rep 2


title: Untr_HCTaidRPB1_K27ac_CnT_rep4

value: K27ac (Abcam ab4729)


title: 8E LiCl rep 1


title: Aux_HCTaidRPB1_K27ac_CnT_rep4


title: BlisterFluid86, SCAR, rep1


title: Aux_HCTaidRPB1_K27ac_CnT_rep3


title: BlisterFluid87, SCAR, rep1


title: 3N LiCl rep 3

value: MATa his3delta1 ura3delta0 leu2delta0 lys2delta0 HIS3MX6-GAL1p-HCM1-3HA-KanMX ChrVIdelta181901-182001::Hyg-TEFp-GFP + pRS316-HCM1p-hcm1-3N-3V5


title: Untr_HCTaidRPB1_K27ac_CnT_rep2


title: Untr_HCTaidRPB1_K27ac_CnT_rep3


title: Aux_HCTaidRPB1_K27ac_CnT_rep2


title: Untr_HCTaidRPB1_K27ac_CnT_rep1


title: Organoid, Control, Day30, CDYL antibody, Rep1

value: Organoid, Control

value: Cortical organoids

value: CDYL (Proteintech, 17763-1-AP)

value: Day 30


title: Skin29, SCAR, rep1

source_name: Skin biopsy Unaffected

value: Skin biopsy Unaffected

value: Male


title: Aux_HCTaidRPB1_K27ac_CnT_rep1

value: colorectal carcinoma

value: Mouse nuclei


title: 3N LiCl rep 2


title: Flav_A12_H3K4me3_CnT

value: breast cancer

value: H3K4me3 (Diagenode C15410003)

value: 0.5uM Flavopiridol


title: Flav_A12_K27ac_CnT


title: 3N LiCl rep 1


title: Skin19, SCAR, rep1


title: Untr_hPDF_H2AZ_CnT

source_name: primary dermal fibroblasts

value: primary dermal fibroblasts

value: H2AZ (Diagenode C15410201)

value: DMSO


title: WT LiCl rep 3

value: haploid yeast

value: MATa his3delta1 ura3delta0 leu2delta0 lys2delta0 HIS3MX6-GAL1p-HCM1-3HA-KanMX ChrVIdelta181901-182001::Hyg-TEFp-GFP + pRS316-HCM1p-HCM1-3V5

value: 150mM LiCl


title: Untr_HCTaidRPB1_H3K4me3_CnT_rep1

value: 10 ug/ml Doxycycline


title: WT LiCl rep 2


title: Trip_hPDF_H2AZ_CnT

value: 0.5uM Triptolide


title: Trip_HCT116_H3K4me3_CnT


title: Skin17, SCAR, rep1

source_name: Skin biopsy Affected

value: Skin biopsy Affected


title: Skin18, SCAR, rep1


title: WT LiCl rep 1


title: Untr_hPDF_H3K4me3_CnT


title: BlisterFluid81, SCAR, rep1

value: Blister fluid

value: Female


title: L_13_carcinoma_1d

source_name: carcinoma

value: carcinoma

value: L


title: 8E control rep 3

value: MATa his3delta1 ura3delta0 leu2delta0 lys2delta0 HIS3MX6-GAL1p-HCM1-3HA-KanMX ChrVIdelta181901-182001::Hyg-TEFp-GFP + pRS316 HCM1p-hcm1-8E-3V5


title: Trip_HCT116_K27ac_CnT

value: colorectal carcinoma

value: Mouse nuclei

value: K27ac (Abcam ab4729)


title: BlisterFluid80, SCAR, rep1


title: Trip_SW13_H3K4me3_CnT

value: adrenal carcinoma

value: H3K4me3 (Diagenode C15410003)


title: Trip_SW13_K27ac_CnT


title: Untr_SW13_H3K4me3_CnT

value: DMSO


title: 8E control rep 2

value: haploid yeast


title: 8E control rep 1


title: Skin15, SCAR, rep1

source_name: Skin biopsy

value: Skin biopsy


title: Untr_hPDF_K27ac_CnT

value: primary dermal fibroblasts


title: Skin16, SCAR, rep1


title: 3N control rep 3

value: MATa his3delta1 ura3delta0 leu2delta0 lys2delta0 HIS3MX6-GAL1p-HCM1-3HA-KanMX ChrVIdelta181901-182001::Hyg-TEFp-GFP + pRS316-HCM1p-hcm1-3N-3V5


title: Trip_A12_K27ac_CnT_rep4

value: breast cancer

value: 0.5uM Triptolide


title: Untr_A12_K27ac_CnT_rep4


title: Trip_hPDF_K27ac_CnT


title: Skin14, SCAR, rep1

value: Skin biopsy Affected


title: 3N control rep 2


title: Skin13, SCAR, rep1

value: Female


title: 3N control rep 1


title: Alveolus Lung-on-a-chip, uninfected, 24 hr, biol rep 3

city: Dallas

first: Victoria

middle: Anette

last: Ektnitphong

state: TX

address: 5323 Harry Hines Blvd.

department: Internal Medicine

institute: UTSW Medical Center

data_processing:
Single-end 100 bp read length Fastq files were checked for quality using fastqc (v0.11.2) and fastq_screen (v0.4.4)
Reads from fastq files for each library was mapped to human reference genome using STAR (v2.5.3a)
Read counts were generated using featureCounts (subread) for coding genes corresponding to igenomes gtf file
Normalized read counts were generated using edgeR
Assembly: hg19
Supplementary files format and content: tab delimited txt files that includes normalized read counts for each sample per gene

source_name: human primary alveolar cells

growth_protocol: Primary human alveolar epithelial cells (ATs) were obtained from Cell Biologics (H-6053). Prior to seeding on Alveolus Lung-on-Chips (ALoCs), ATs were expanded in vitro in a T25 flask coated with a 1% gelatin-based coating solution (Cell Biologics, catalog #6950) and complete medium comprising of base media and supplements (Lonza, CC-3118). Medium was prepared according to manufacturer’s instructions using all supplements except GA-1000 where 1% Pen-Strep solution (Gibco 15140-122) was added instead along with 5% FBS and hereby called Small Airway Growth Medium (SAGM). Primary human microvascular endothelial cells were purchased from Lonza (CC-2527). Human Lung Microvascular Endothelial cells (HMVECs) were expanded in a T75 in complete medium comprising of base media and supplements (Lonza, CC-3202). Medium was prepared according to manufacturer’s instructions using all supplements except GA-1000 where 1% Pen-Strep solution was used instead along with 5% FBS and hereby called EGM-2MV. Both primary cell lines were cultured at 37°C in 5% CO2 until ~80% confluency before detachment with TrypLETM Express (Gibco 12604013) and use on the ALoC. One week prior to seeding the macrophages on the ALoC, peripheral blood mononuclear cells (PBMCs) were obtained from buffy coat (Carter BloodCare) taken from anonymous donors. PBMCs were isolated using Ficoll (Cytivia, 17144003) and SepMate50 tubes from StemCell Technologies (85450). CD14+ monocytes were positively selected from the PBMCs using CD14 Microbeads (Miltenyi, 130-050-201), and seeded onto uncoated petri dishes. The monocytes were cultured overnight in RPMI medium (Gibco 11875-093) supplemented with 10% heat-inactivated human serum obtained from the buffy coat of the donor, 1% HEPES buffered solution (Lonza CC-5022), 1% sodium pyruvate (Gibco 11360-070), and 50 ng/mL human granulocyte-macrophage colony-stimulating factor (GM-CSF) (Peprotech 300-03-100UG), and is hereby called Macrophage medium. The next day, the serum in the medium was changed to 10% FBS for the remainder of the culturing. The CD14+ monocytes were differentiated to human monocyte-derived macrophages (HMDMs) for 7 days at 37°C in 5% CO2 until use on the ALoCs. GM-CSF was added to medium for the first 4 days to differentiate the CD14+ monocytes.

extract_protocol:
ALoCs were removed from the Zoe and Pods. Top and bottom channels were washed 3X with 200 µL of ice-cold 1X DPBS (-Ca+/-Mg+). 200 µL of TRIzol reagent (Invitrogen 15596026) was pipetted in each ALoC by rapidly pressing and releasing the plunger 3X before collecting the supernatant in an RNase-free 1.5 mL tube. This process was repeated once more for a total of ~400 µL of supernatant. TRIzol was added to each tube of RNA to a final volume of 1 mL. The RNA was stored at -80°C before extraction using the Qiagen RNeasy Mini Columns (74106). AT and macrophage lysates were thawed on ice and incubated at room temperature for 5 min. 200 µL of chloroform per 1 mL of lysates was added to each tube, shaken vigorously for 15 sec., incubated at room temperature for 2-3 min, and centrifuged for 5 min at 12,000xg at 4°C. The aqueous phase was transferred to fresh RNase-free tubes and the rest of the RNA extraction was performed according to the Qiagen RNeasy Mini Column Kit. Extracted RNA was stored at -80°C until use for library preparation and RNA-sequencing.
Samples were analyzed on an Agilent Tapestation 4200 to determine level of degradation to ensure that only high quality RNA was used (RIN Score 8 or higher). We used a Qubit® 4.0 Fluorimeter (ThermoFisher) to determine RNA concentration prior to starting library prep. One microgram of total DNAse treated RNA was then prepared with the TruSeq Stranded mRNA Library Prep Kit (Illumina). Poly-A RNA was purified and fragmented before strand-specific cDNA synthesis. cDNA was then A-tailed and indexed adapters ligated. After adapter ligation, samples were PCR amplified and purified with AmpureXP beads, then validated again on the Agilent Tapestation 4200. Before being normalized and pooled, samples were quantified by Qubit then sequenced on the Illumina NextSeq 2000 using a P2-100 flowcell.

value: human primary alveolar cells

value: human monocyte-derived macrophages

value: Control, uninfected


title: Trip_A12_K27ac_CnT_rep3


title: BlisterFluid75, SCAR, rep1

value: Blister fluid


title: Alveolus Lung-on-a-chip, uninfected, 24 hr, biol rep 2


title: Untr_SW13_K27ac_CnT

value: Mouse nuclei


title: Trip_hPDF_H3K4me3_CnT


title: WT control rep 3

value: MATa his3delta1 ura3delta0 leu2delta0 lys2delta0 HIS3MX6-GAL1p-HCM1-3HA-KanMX ChrVIdelta181901-182001::Hyg-TEFp-GFP + pRS316-HCM1p-HCM1-3V5


title: Alveolus Lung-on-a-chip uninfected, 24 hr, biol rep 1


title: BlisterFluid74, SCAR, rep1


title: Alveolus Lung-on-a-chip infected with M. fortuitum, 24 hr, biol rep 1

value: M.fortuitum infection


title: Alveolus Lung-on-a-chip infected with M. fortuitum, 24 hr, biol rep 2


title: Alveolus Lung-on-a-chip infected with M. fortuitum, 24 hr, biol rep 3


title: WT control rep 2


title: BlisterFluid73, SCAR, rep1


title: WT control rep 1


title: HMVEC-L, siNegControl 3

city: Los Angeles

first: Jean-Pierre

last: Amoakon

state: California

address: 8700 Beverly Blvd

department: Medicine

institute: Cedars-sinai Medical center

data_processing:
General bioinformatic analysis was performed via BaseSpaceSEQUECNCE HUBappRNA-Seq Alignment v2.0.2followed byRNA-Seq Differential Expression version 1.0.1.
The analysis used STAR for alignment and Salmon for quantification (Transcripts Per Million, TPM), followed by DESeq2 to identify differentially expressed genes.
Assembly: Homo sapiens HG19
Supplementary files format and content: tab-delimited text file includes raw couns for each sample

source_name: Lung microvasculature

growth_protocol: HMVEC-L (Lonza, Cat#CC-2527) were cultured in EGM-2 Bulletkit Culture Medium with supplements (Lonza, Cat#CC-3162) at 37°C and 5% CO2. Culture media bottle was supplemented with 1% penicillin-streptomycin (Gibco, Cat#15-140-163) and 1 mL plasmocin prophylactic (InvivoGen, Cat#ant-mpp).

extract_protocol:
RNA was isolated using Ambion miRNA Isolation Kit (Invitrogen, Cat#AM1561) according to manufacturer’s instructions.
NEBNext Single Cell/Low Input RNA Library Prep kit (non-directional) was used for ultra-low input bulk RNA-seq library preparation.

treatment_protocol: Silencer Select Human CFTR siRNA (ThermoFisher, Cat#4392420/s534180) or Silencer Select Negative Control siRNA (ThermoFisher, Cat#4390843) were transfected (40 nM siRNA final concentration) into cells using Lipofectamine RNAiMax (ThermoFischer, Cat#13778150)

value: Lung microvasculature

value: HMVEC-L

value: Primary cells

value: WT

value: siRNA negative control


title: BlisterFluid71, BURN, rep1

value: Male


title: HMVEC-L, siCFTR 1

value: CFTR knockdwon

value: siRNA CFTR


title: HMVEC-L, siNegControl 1


title: HMVEC-L, siCFTR 3


title: Untr_A12_K27ac_CnT_rep3

value: breast cancer

value: K27ac (Abcam ab4729)

value: DMSO


title: RNA-seq in XY E13.5 Fgf9 Δ93 gonads, biol rep 5

city: Berlin

first: Irene

last: Mota-Gomez

address: Hannoversche strasse 28

country: Germany

institute: Max Delbruck Center

data_processing:
Alignment using STAR
Quantification using Genomic alignments::summarizeOverlaps
Differential analysis using DESeq2
Assembly: mm10
Supplementary files format and content: Counts.csv= Merged matrix containing counts from all sampels in the different columns
Supplementary files format and content: DEseq2.csv= Differential gene expression analysis, direct output from DESeq2

organism: Mus musculus

source_name: gonad

growth_protocol: E13.5 gonads were microdissected in 1XPBS  and separated from mesonephros using srynges and snapped frozen in liquid N

extract_protocol:
RNA was extracted using  Rnase Micro kit (Qiagen).
RNA seq librares were performed using the NEBNext Ultra II Directional RNA library prep kit for Illumina including the Poly(A) magnetic isolation module

value: gonad

value: Fgf9 delta93


title: RNA-seq in XY E13.5 Fgf9 Δ93 gonads, biol rep 6


title: HMVEC-L, siNegControl 2


title: WT, WGBS

city: BeiJing

first: Tian

last: Yuan

address: CAU

institute: CAU

data_processing:
Raw sequence reads were trimmed by trimm_galore
Trimmed sequence reads were mapping to NC10 using bismark
Methylation information were extracted by bismark_methylation_extractor
Assembly: NC10
Supplementary files format and content: bedGraph

organism: Neurospora crassa

growth_protocol: The medium used for plate assays contained 1ⅹVogel’s salts, 3% sucrose, and 1.5% (w/v) agar with or without H2O2. Liquid cultures were grown in minimal medium (1ⅹVogel’s and 2% glucose) under continuous light oscillation at 25°C for 18 hours.

extract_protocol:
phenol-chloroform extraction
MGIEasy Genome-wide methylation library construction

tag: strain

value: Neurospora crassa OR74A

value: mycelium


title: BlisterFluid69, SCAR, rep1

value: Blister fluid

value: Female


title: ckb1ckb2, WGBS

tag: genotype

value: ckb1ckb2 knockout


title: cka_ectopic expression of CKA protein in ckaKO strain, WGBS

tag: cell line


title: CKA_ko, WGBS

tag: cell type

value: cka knockout


title: RNA-seq in XY E13.5 Fgf9 Δ93 gonads, biol rep 3


title: BlisterFluid68, SCAR, rep1


title: BlisterFluid67, SCAR, rep1


title: HMVEC-L, siCFTR 2

value: Lung microvasculature

value: HMVEC-L

value: Primary cells


title: RNA-seq in XY E13.5 Fgf9 Δ93 gonads, biol rep 2


title: MCF-7_Nutlin-3a

city: Jena

first: Martin

last: Fischer

state: Thuringia

address: Beutenbergstraße 11

department: Computational Biology

institute: Leibniz Institute on Aging - Fritz Lipmann Institute (FLI)

data_processing:
Base calling was performed using guppy v6.0.7 with fast model.
After identification, orientation, and trimming of full-length cDNA reads using pychopper v2.7.2, reads were aligned to hg38 by minimap2 v2.24-r1122 in spliced long-read mode and with disabled secondary alignments.
Assembly: hg38
Supplementary files format and content: RPM count table

source_name: breast

growth_protocol: MCF-7 cells were grown in high glucose DMEM with pyruvate. Culture media were supplemented with 10% FBS and penicillin/streptomycin. In addition, DMEM was supplemented with non-essential amino acids.

extract_protocol:
Total RNA was extracted using the innuPREP RNA Mini Kit (Analytik Jena, Jena, Germany) following the manufacturer’s protocol.
Oxford Nanopore genomic library preparation protocol SQK-PCB109 was followed using 50 ng of total RNA as input. The samples were barcoded according to protocol with 16 PCR cycles.
Libraries were run for 72h according to the guidelines of the manufacture using Mk1B-MinION sequencer and R9.4.1 flow cells (FLO-MIN106D). Sequencing run monitoring and real-time data acquisition were performed using the MinKNOW software suite v22.03.6.

treatment_protocol: Cells were treated with Nutlin-3a 10 µM or DMSO control for 24 h

value: breast

value: epithelial cells

value: wild-type

value: Nutlin-3a 10 µM 24 h


title: cka_D149A, WGBS

value: ckaD149A point mutation


title: RNA-seq in XY E13.5 Fgf9 Δ93 gonads, biol rep 1


title: MCF-7_DMSO

value: DMSO 24 h


title: BlisterFluid72, BURN, rep1

value: Male


title: RNA-seq in XY E13.5 Fgf9 Δ93 gonads, biol rep 4

value: gonad

value: Fgf9 delta93


title: H3T11A, WGBS

value: Neurospora crassa OR74A

value: mycelium

value: H3T11A point mutation


title: Skin12, SCAR, rep1

value: Skin biopsy

value: Female


title: Skin11, SCAR, rep1


title: Etoposide-induced senescent cells cells starved for Methionine and Cysteine, 3h, rep 2

first: Emmanouil

state: MD

address: 251 Bayview Blvd, 10B133 Baltimore

department: Laboratory of Genetics and Genomics (LGG)

institute: National Institute on aging

data_processing:
Sequencing data was basecalled using Guppy (v3.4.5) and the adaptors were removed using cutadapt (v2.8).
rRNA sequences were filtered by alignment to sequences obtained from SILVA and resulting non-rRNA reads were mapped against the human genome (hg38) using minimap2 (version 2.17)
Reads were also aligned against the human transcriptome.
The poly(A) tail lengths were extracted from sequenced reads using the nanopolish polya package, with quality control scores as “PASS” selectively used in our analysis.
Assembly: hg38
Supplementary files format and content: Tab-delimited text files include transcript id based raw read counts, respective average transcript length and polyA lengths values for each Sample
Supplementary files format and content: DESeq2 differential gene expression

growth_protocol: Cells were grown in 10% FBS DMEM high glucose media supplemented with non-essential amino acids and antibiotic/antimycotics.

extract_protocol:
Bulk RNA was extracted using Trizol and then poly-A selected.
Poly-A selected RNA was ligated in bulk to a biontinylated linke sequence using T4 RNA ligase 1. Libraries were then prepared using the direct RNA seqneicng kit (Nanopore Technologies) as described by the manufacturer.
direct-RNA long read sequencing

treatment_protocol:
Senescence was induced through 3 treatments with 50 uM etoposide over 10 days with harvest or further treatment on the 10th day.
Cycling cells were harvested at 40-60% confluency before harvest or futher treatment.
Methionine and Cysteine starvation by incubating cells in media described in the growth protocol but missing methoinine and cysteine amino acids.
Thapsigargin treatment was performed by spiking cells with thapsigargin dissolved in DMSO to a final concentration of 25 nM.

value: Fibroblasts

value: WT

value: Etoposide 50 uM, 10d; Methionine and Cysteine starvation, 3h


title: Etoposide-induced senescent cells cells treated with 25 nM thapsigargin, 3h, rep 1

value: Etoposide 50 uM, 10d; Thapsigargin 25 nM, 3h


title: BlisterFluid70, BURN, rep1

value: Blister fluid


title: RNA-seq in XY E13.5 Fgf9 Δ225 gonads, biol rep 4

value: Fgf9 delta225


title: Cycling cells treated with 25 nM thapsigargin, 3h, rep 2

value: Thapsigargin 25 nM, 3h


title: Etoposide-induced senescent cells cells treated with 25 nM thapsigargin, 3h, rep 2


title: Cycling cells treated with 25 nM thapsigargin, 3h, rep 1


title: Etoposide-induced senescent cells cells starved for Methionine and Cysteine, 3h, rep 1


title: Skin10, SCAR, rep1


title: RNA-seq in XY E13.5 Fgf9 Δ225 gonads, biol rep 3


title: Cycling cells starved for Methionine and Cysteine, 3h, rep 2

value: Methionine and Cysteine starvation, 3h


title: Cycling cells starved for Methionine and Cysteine, 3h, rep 1


title: Skin9, SCAR, rep1

value: Skin biopsy Affected


title: RNA-seq in XY E13.5 Fgf9 Δ225 gonads, biol rep 2


title: Skin, Healthy Donor, rep1


title: Etoposide-induced senecent cells, rep 2

description: DESeq2_results_Cycling_versus_Senescent.tab

value: Etoposide 50 uM, 10d


title: Etoposide-induced senecent cells, rep 1


title: Skin8, SCAR, rep1


title: RNA-seq in XY E13.5 Fgf9 Δ225 gonads, biol rep 6

value: gonad


title: Cycling cells, rep 2


title: RNA-seq in XY E13.5 Fgf9 Δ104 gonads, biol rep4

value: Fgf9 delta104


title: RNA-seq in XY E13.5 Fgf9 Δ225 gonads, biol rep 1


title: HMHM

city: Funabashi

first: Shinpei

last: Yamaguchi

state: Chiba

address: Miyama 2-2-1

department: Faculty of Science

country: Japan

institute: Toho University

description:
Re-differentiated CYP3A4G7R-HepaRG cells for 7 days in HepaRG medium + DMSO after de-differentiation in HepaRG medium + DMSO for 7 days. Prior to de-differentiation, cells were grown in HepaRG medium for 14 days and differentiated with HepaRG medium + DMSO for 14+7 days (with passage).
metadata.csv
count.csv
naka_225

data_processing:
Read 1 reads were processed with fastp (version 0.21.0) (S. Chen et al. 2018) using the following command: “ fastp --trim_poly_x -w 20 --adapter_sequence=AGATCGGAAGAGCACACGTCTGAACTCCAGTCA --adapter_sequence_r2=AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT -l 31”. The trimmed reads were then mapped to a D. japonica transcriptome reference (An et al. 2018), using BWA mem (version 0.7.17-r1188) (W. Chen et al. 2013) with the default parameters. The read count for each gene was calculated with salmon using -l IU, which specifies the library type (version v0.12.0) (Patro et al. 2017).
Assembly: Homo_sapiens.GRCh38.cdna.all.fa
Supplementary files format and content: csv-delimited text file includes rawcnt values for each Sample

source_name: HepaRG

growth_protocol: The HepaRG cells at the proliferation, differentiation, de-differentiation, and redifferentiation stages were harvested with 0.05% Trypsin-EDTA (Gibco) treatment.

extract_protocol:
Total RNA of the resulting cell suspension was extracted using RNeasy Plus (QIAGEN).
3’ mRNA-Seq were conducted according to the Lasy-Seq ver. 1.1 protocol (https://sites.google.com/view/lasy-seq/) (Kamitani et al. 2019; Kashima et al. 2021). Briefly, total RNA was reverse transcribed using an RT primer with index and SuperScript IV reverse transcriptase (Thermo Fisher Scientific, Waltham, MA, USA). Then, all RT mixtures of the samples were pooled and purified using an equal volume of AMpure XP beads (Beckman Coulter, Brea, CA, USA) according to the manufacturer’s instructions. Second strand synthesis was conducted on the pooled samples using RnaseH (5 U/μL, Enzymatics, Beverly, MA, USA), and DNA polymerase I (10 U/μL, Enzymatics, Beverly, MA, USA). To avoid the carryover of large amounts of rRNAs, the mixture was subjected to Rnase treatment using Rnase T1 (Thermo Fisher Scientific, Waltham, MA, USA). Then, purification was conducted with a 0.8× volume of Ampure XP beads. Fragmentation, end-repair, and A-tailing were conducted using 5× WGS Fragmentation Mix (Enzymatics, Beverly, MA, USA). The Adapter for Lasy-Seq was ligated using 5× Ligation Mix (Enzymatics, Beverly, MA, USA), and the adapter-ligated DNA was purified twice with a 0.8× volume of Ampure XP beads. After optimisation of PCR cycles for library amplification by qPCR using Evagreen, 20× in water (Biotium, Fremont, CA, USA) and the QuantStudio5 Real-Time PCR System (Applied Biosystems, Waltham, MA, USA), the library was amplified using KAPA HiFi HotStart ReadyMix (KAPA BIOSYSTEMS, Wilmington, MA, USA) on the ProFlex PCR System (Applied Biosystems, Waltham, MA, USA). The amplified library was purified with an equal volume of Ampure XP beads. One microliter of the library was then used for electrophoresis using a Bioanalyzer 2100 with the Agilent High Sensitivity DNA kit (Agilent Technologies, Santa Clara, CA, USA) to check for quality. Then, sequencing of 150-bp paired-end reads was performed using HiSeq X Ten (Illumina, San Diego, CA, USA) by Macrogen, Inc. (Seoul, Korea).

value: HepaRG


title: HMHMWnt

description:
Re-differentiated CYP3A4G7R-HepaRG cells for 7 days in HepaRG medium + DMSO after de-differentiation in HepaRG medium + Wnt3a for 7 days. Prior to de-differentiation, cells were grown in HepaRG medium for 14 days and differentiated with HepaRG medium + DMSO for 14+7 days (with passage).
metadata.csv
count.csv
naka_224


title: Cycling cells, rep 1


title: RNA-seq in XY E13.5 Fgf9 Δ225 gonads, biol rep 5


title: BlisterFluid60, SCAR, rep1

source_name: Hash 1_SCAR patient 3, blister fluid / Hash 2_SCAR patient 3, blister fluid


title: RNA-seq in XY E13.5 Fgf9 Δ104 gonads, biol rep3


title: HM28_2

description:
Differentiated CYP3A4G7R-HepaRG cells for 14 days in HepaRG medium + DMSO conditions after growth in HepaRG medium.
metadata.csv
count.csv
naka_221


title: HM28_3

description:
Differentiated CYP3A4G7R-HepaRG cells for 14 days in HepaRG medium + DMSO conditions after growth in HepaRG medium.
metadata.csv
count.csv
naka_222


title: RNA-seq in XY E13.5 Fgf9 Δ104 gonads, biol rep2


title: SMHM38

description:
Differentiated CYP3A4G7R-HepaRG cells for 14+7 days (with passage) in HepaRG medium + DMSO conditions after growth in HepaRG medium for 7 days followed by small hepatocyte medium for 7 days.
metadata.csv
count.csv
naka_220


title: RNA-seq in XY E13.5 Fgf9 Δ104 gonads, biol rep1


title: HMHMA

description:
Re-differentiated CYP3A4G7R-HepaRG cells for 7 days in HepaRG medium + DMSO after de-differentiation in HepaRG medium + A-83-01 for 7 days. Prior to de-differentiation, cells were grown in HepaRG medium for 14 days and differentiated with HepaRG medium + DMSO for 14+7 days (with passage).
metadata.csv
count.csv
naka_223


title: BlisterFluid59, SCAR, rep1

value: Hash 1_SCAR patient 3, blister fluid / Hash 2_SCAR patient 3, blister fluid


title: BlisterFluid58, SCAR, rep1


title: deSMCTR

description:
De-differentiated HepaRG cells for 5 days in Small hepatocyte medium conditions after differentiation by DMSO for 14+7days (with passage) following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_218


title: SMHM28

description:
Differentiated CYP3A4G7R-HepaRG cells for 14 days in HepaRG medium + DMSO conditions after growth in HepaRG medium for 7 days followed by small hepatocyte medium for 7 days.
metadata.csv
count.csv
naka_219


title: BlisterFluid57, SCAR, rep1


title: RNA-seq in XY E13.5 Meis1 KO gonads, biol rep 4

value: Meis1 KO


title: Skin7, SCAR, rep1


title: BlisterFluid56, SCAR, rep1


title: deSMYAC

description:
De-differentiated HepaRG cells for 5 days in Small hepatocyte medium + YAC conditions after differentiation by DMSO for 14+7days (with passage) following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_217


title: deHMCTR_2

description:
De-differentiated HepaRG cells for 5 days in HepaRG medium conditions after differentiation by DMSO for 14+7days (with passage) following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_216


title: deHMYAC_2

description:
De-differentiated HepaRG cells for 5 days in HepaRG medium + YAC conditions after differentiation by DMSO for 14+7days (with passage) following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_215


title: HM28_1

description:
Differentiated HepaRG cells for 14 days in HepaRG medium + DMSO conditions after growth in HepaRG medium.
metadata.csv
count.csv
naka_214


title: deHMCTR_1

description:
De-differentiated HepaRG cells for 5 days in HepaRG medium conditions after differentiation by DMSO for 14 days following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_213


title: RNA-seq in XY E13.5 Meis1 KO gonads, biol rep 1


title: BlisterFluid55, SCAR, rep1


title: RNA-seq in XY E13.5 Meis1 KO gonads, biol rep 2


title: BlisterFluid53, SCAR, rep1


title: deHMA

description:
De-differentiated HepaRG cells for 5 days in HepaRG medium + A-83-01 conditions after differentiation by DMSO for 14 days following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_212


title: BlisterFluid54, SCAR, rep1


title: HM14_2

description:
Proliferating HepaRG cells cultured in HepaRG medium for 14 days, repliocate #2
metadata.csv
count.csv
naka_209


title: deHMYAC_1

description:
De-differentiated HepaRG cells for 5 days in HepaRG medium + YAC conditions after differentiation by DMSO for 14 days following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_210


title: HMYAC_2

description:
Proliferating HepaRG cells cultured in HepaRG medium for 7 days and then in HepaRG medium with YAC for one day, repliocate #2
metadata.csv
count.csv
naka_207


title: deHMY

description:
De-differentiated HepaRG cells for 5 days in HepaRG medium + Y-27632 conditions after differentiation by DMSO for 14 days following 14 days of growth in HepaRG medium.
metadata.csv
count.csv
naka_211


title: BMDM_CM_4OI_3(biol rep3)

city: Shanghai

first: Chuanyang

last: Wang

state: Shanghai

address: Shanghai Wujin Road 85#

institute: Shanghai Jiaotong University

data_processing:
The raw data was handled by Skewer v0.2.2 (https://sourceforge.net/projects/skewer/files/?source=navbar) and data quality was checked by FastQC v0.11.2 (http:// www.bioinformatics.babraham.ac.uk/projects/fastqc/). The read length was 2×150 bp.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: bone marrow

extract_protocol:
Total RNA was extracted from BMDMs with the Universal RNA Extraction CZ Kit (RNC643, ONREW) according to the manufacturer’s instructions. All RNA-seq experiments were performed in n = 3 independent cultures.
RNA libraries were prepared using the VVAHTS® Universal V8 RNA-seq Library Prep Kit for Illumina (NR605-0, Vazyme)
Illumina NovaSeq 6000 platform with the 150 paired-end sequencing strategy

value: bone marrow

value: primary cell

value: macrophage

value: wildtype

value: CM+4OI 4h


title: HM14_1

description:
Proliferating HepaRG cells cultured in HepaRG medium for 14 days, repliocate #1
metadata.csv
count.csv
naka_208


title: HMYAC_1

description:
Proliferating HepaRG cells cultured in HepaRG medium for 7 days and then in HepaRG medium with YAC for one day, repliocate #1
metadata.csv
count.csv
naka_206


title: BMDM_CM_4OI_2(biol rep2)


title: BlisterFluid51, SCAR, rep1

value: Female


title: BMDM_CM_4OI_1(biol rep1)


title: HM8_2

description:
Proliferating HepaRG cells cultured in HepaRG medium for 8 days, repliocate #2
metadata.csv
count.csv
naka_205


title: BlisterFluid50, SCAR, rep1

value: Blister fluid


title: HM8_1

description:
Proliferating HepaRG cells cultured in HepaRG medium for 8 days, repliocate #1
metadata.csv
count.csv
naka_204


title: HMSHM_2

description:
Proliferating HepaRG cells cultured in HepaRG medium for 7 days and then in Small hepatocyte medium for one day, repliocate #2
metadata.csv
count.csv
naka_203


title: RNA-seq in XY E13.5 Meis1 KO gonads, biol rep 3


title: BMDM_CM_3(biol rep3)

value: CM 4h


title: HAL

description:
RNA from Human adult liver (HAL).
metadata.csv
count.csv
naka_201


title: BlisterFluid49, SCAR, rep1


title: BMDM_CM_2(biol rep2)


title: HMSHM_1

description:
Proliferating HepaRG cells cultured in HepaRG medium for 7 days and then in Small hepatocyte medium for one day, repliocate #1
metadata.csv
count.csv
naka_202


title: BlisterFluid52, SCAR, rep1


title: BMDM_CM_1(biol rep1)


title: BlisterFluid48, SCAR, rep1

value: Male


title: BlisterFluid47, SCAR, rep1


title: BlisterFluid46, SCAR, rep1


title: Mouse, Naïve, E15.5, Input

source_name: Mouse cortex

value: Mouse cortex

value: whole cortex

value: naive

value: CDYL (Proteintech, 17763-3-AP)


title: Mouse, Naïve, E15.5, CDYL antibody, Rep2

value: CDYL (Proteintech, 17763-2-AP)


title: Human_lung_inserts_1_2_RNA

city: Amsterdam

first: Joe

last: Butler

address: Meibergdreef 9

department: Infectious diseases

institute: Amsterdam UMC

description:
AMC-SC-S21_GEX
AMC-SC-S22_GEX

data_processing:
Libraries were sequenced using the Illumina HiSeq2000 (Illumina, San Diego, CA, USA) to generate 150bp paired-end reads.
The sequence read quality was assessed using FastQC methods (version 0.11.5; Babraham Institute, Babraham, Cambridgeshire, UK).
Sequencing fastq output files were processed using CellRanger (10x genomics)_and alignged to human genome as specified below.
The remaining high-quality reads were aligned against the human Genome Reference Consortium Build 38 (GRCh38, Ensembl 84)
Assembly: Human Genome Reference Consortium Build 38 (GRCh38, Ensembl 84)
Supplementary files format and content: Tab-delimited text file includes raw counts for each cell and samplewise metadata

source_name: Lung

extract_protocol:
Samples were processed using 10x genomics chromium single cell v2 protocol.
Libraries were generated according to 10x genomics recommended v2 protocol.

value: Lung

value: Air Liquid Interface Inserts

tag: time point

value: Mixed


title: BlisterFluid45, SCAR, rep1

source_name: Hash 3_SCAR patient 14, blister fluid / Hash 4_SCAR patient 14, blister fluid

tag: tissue


title: Human_lung_inserts_1_2_Hashtags

description:
AMC-SC-S21_HTO
AMC-SC-S22_HTO


title: BlisterFluid44, SCAR, rep1

value: Hash 3_SCAR patient 14, blister fluid / Hash 4_SCAR patient 14, blister fluid


title: Mouse, Naïve, E15.5, CDYL antibody, Rep1

value: CDYL (Proteintech, 17763-1-AP)


title: agshZIKV-h1#2-Input

city: Wuhan

first: Yang

last: Qiu

state: Hubei

address: No. 44 Xiaohongshan Central District, Wuchang District,

institute: Wuhan institue of virology, CAS

data_processing:
Fo small RNAs, libraries of small RNAs were constructed using TruSeq Small RNA Library Preparation Kits (Illumina) according to manufacturer’s protocols, and then sequenced by Illumina HiSeq 2000 at Bioacme (Wuhan, China). Quality control and adapter trimming process were performed using Trim Galore (v0.6.4) with default parameters, and reads length between 17 and 70 were used for subsequent analyses. The sequenced reads were aligned to the human genome (GRCh38) by using Bowtie2 (v2.3.5.1). Reads that cannot be aligned to the human genome were then aligned to agshRNA. The sequences and lengths of reads aligned to agshRNA were then determined using an in-house script. Reads mapped to genome were further categorized as miRNA, tRNA, snoRNA etc. by using htseq-count (v0.11.2).
For total RNAs, libraries of total RNAs were constructed using MGIEasy RNA Library Preparation Kits (BGI) according to manufacturer’s protocols, and then sequenced by MGISEQ2000 (BGI) at Wuhan Institute of Virology, CAS. Quality control and adapter trimming were performed using Trim Galore (v0.6.4) with default parameter. All reads were mapped to the human genome (GRCh38) using Hisat2(v2.1.0) and then annotated using htseq-count(v0.11.2)
Assembly: Human genome (GRCh38)
Supplementary files format and content: small RNA sequencing: bed format
Supplementary files format and content: Transcriptome sequencing: expression matrix counted by htseq-count

source_name: Human  HEK293T cells

extract_protocol:
For small RNA sequencing, Total RNA was extrated using Trizol regent accounding to the manufacturer's instructions. For transcriptome sequencing, RNA was extracted using Total RNA kit according to the manufacturer's instuctions
TruSeq Small RNA Library Preparation Kits (Illumina) (for small RNAs); MGIEasy RNA Library Preparation Kits (BGI) (for total RNAs)
Illumina HiSeq 2000 (for small RNAs); MGISEQ2000 (BGI) (for total RNAs)

value: Human  HEK293T cells


title: agshZIKV-h1#2-IP


title: BlisterFluid43, SCAR, rep1


title: Lung_mTFF2 1d_rep3

type: RNA

first: Wei

middle: hui

last: Fu

address: No. 2901 Callang Road,Jinshan District

institute: Shanghai Public Health Clinical Center

description: Gene expression in lungs of mice infected by H7N9 with TFF2 treatment for 1 day

hyb_protocol: The labeled cRNAs were hybridized onto the microarray. After washing

scan_protocol: the arrays were scanned by the Agilent Scanner G2505C(Agilent Technologies).

data_processing: Feature Extraction software (version10.7.1.1, Agilent Technologies) was used to analyze array images to get raw data. Genespring(version13.1,Agilent Technologies)were employed to finish the basic analysis with the raw data. To begin with, the raw data was normalized with the quantile algorithm.

source_name: Lung, mTFF2 1d, replicate3

label_protocol: total RNA were transcribed to double strand cDNA, then synthesized into cRNA and labeled with Cyanine-3-CTP.

extract_protocol: Total RNA was quantified by the NanoDrop ND-2000 (Thermo Scientific)and the RNA integrity was assessed using Agilent Bioanalyzer 2100 (Agilent Technologies).

treatment_protocol: Fresh mouse lung tissue was washed with PBS, frozen in liquid nitrogen, further crushed into powder, dissolved with RNAzol, and total RNA was extracted.

tag: gender, age

value: female

tag: age

tag: infection


title: Con_3

first: Wanbo

last: Zhu

state: None Selected

address: No600, Yishan road, Shanghai, China

institute: Shanghai Sixth Peoples Hospital Affiliated to Shanghai Jiao Tong University

description:
Con_3
Processed_data.txt

data_processing:
1 Data preprocessing and genomic alignment Raw reads generated during high-throughput sequencing were fastq format sequences. In order to get high-quality reads that could be used for later analysis, raw reads need to be filtered by quality. Trimmomatic [1] software was first used for quality control and linker removal, and then low-quality bases and N-bases or low-quality reads were filtered out. Finally, we got high-quality clean reads. Using Rockhooper2 [2] to align clean reads to the reference genome of the experimental specie, the sample should be assessed by genomic and gene alignment
2 Genetic assembly and structural 2.1 UTR and novel gene prediction Identify the sequence of genes in an annotated gene or an unannotated area. Seeds gene sequence is a sequence area at genome that contains 10 bases at least and each one in this area are covered at least T sequence (T means a function of the2.2 Operator prediction Prokaryotes function on several related genes in are arranged in series with the often constitute manipulation substructure expression as a unit, is regulated by the common upstream area and the downstream transcription termination signal control. When transcribed, multiple structural genes are transcribed onto one mRNA strand and then translated into proteins. Rockhopper2 was used to develop the operon prediction algorithm from pure sequence features to combine experimental data of RNA-seq (ie, the gene expression quantity), that is to use bayesian classifier model to predict operons by combine the inter-genic distance and gene expression quantity, the sensitivity and specificity of the prediction algorithm are as high as 95%. Calculate and visualize 5 the predicted length distribution of operons, the number of structural genes involved, and the operon chain distribution count of genome-wide base average coverage sequence ). Through the bayesian model to judge whether the seed, seed each gene sequences repeatedly extended again after merging adjacent sequence of seeds or overlapping genes ultimately get the genetic map is obtained based on sequence alignment data reference gene annotation to the map and comparison, identification of boundary and the new genes.
3 Gene expression analysis The direct expression of a gene expression level is its abundance. The higher the degree of gene abundance, the higher the gene expression level. In transcriptome sequencing analysis, the expression level of genes was estimated by locating the count of sequencing sequence (reads) in the genome area or the exon region of the gene. The Reads count is positively correlated with the length of the gene and the depth of the sequencing, in addition to the true expression level of the gene. We used the known reference gene sequences as the database, and the expression abundance of each gene in each sample was identified by sequence similarity comparison.Rockhooper2 was used to obtain the number of reads aligned to the gene in each sample and then to calculate RPKM[3].The calculated transcript expression levels can be directly used to compare transcript expression differences between different samples, and this method can eliminate the effect of differences in transcriptional length and sequencing amount on the expression of transcripts. Visualize quantitative data to obtain box plots, density plots, expression plots, etc. to determine differences in expression across all sample
4 Difference analysis and functional analysisUsing the estimateSizeFactors function of the R package of DESeq [4] (2012) to standardize counts, and use the nbinomTest function to calculate the pvalue and foldchange of difference comparison. Pick out the difference transcripts that p-value less than 0.05 and Difference of multiples more than two, and the GO and KEGG enrichment analysis of the differential genes was performed by hypergeometric distribution tests to determine the biological functions or pathways that are mainly affected by differential genes
Assembly: GCF_003052445.1_ASM305244v1
Supplementary files format and content: Processed data.txt

source_name: peri-implant tissue

growth_protocol: Infected mice were executed on the third day and the tissue surrounding the implant was collected.

extract_protocol:
A. Add 600 μL Lysis/Binding Buffer to the cell or tissue lysate. Then add 30 μL miRNA Homogenate Additive to homogenate, and mix well by vortexing or inverting the tube several times.  Leave the mixture on ice for 10 min.  B. Add a volume of Acid-Phenol: Chloroform that is equal to the lysate volume before addition of the miRNA Homogenate Additive. Centrifuge for 5 min at maximum speed (10,000 x g) to separate  the aqueous and organic phases. Carefully remove the aqueous (upper) phase without disturbing the  lower phase, and transfer it to a fresh tube.  C. Add 1.25 volumes of room temperature 100% ethanol to the aqueous phase. D. Pipet the lysate/ethanol mixture (from the previous step) onto the Filter Cartridge (Note: Up to 700 μL can be applied to a Filter Cartridge at a time.). Centrifuge for 30 sec at 13,000 rpm to pass  the mixture through the filter. Discard the flow-through. E. Apply 350 μL miRNA Wash Solution 1 to the Filter Cartridge and centrifuge for 30 sec at 13,000 rpm. Discard the flow-through from the Collection Tube, and replace the Filter Cartridge into  the same Collection Tube.  F. The 10 μL DNase I and 70 μL Buffer RDD QIAGEN (#79254) were mixed. Then the mixture was added to the Filter Cartridge. Leave it at the room temperature for 15 min.  G. Apply 350 μL miRNA Wash Solution 1 to the Filter Cartridge and centrifuge for 30 sec at 13,000 rpm. Discard the flow-through from the Collection Tube, and replace the Filter Cartridge into  the same Collection Tube.  H. Apply 500 μL Wash Solution 2/3 and centrifuge for 30 sec at 13,000 rpm. Draw it through the Filter Cartridge as in the previous step. Repeat with a second 500 μL aliquot of Wash Solution 2/3.  I. Spin the assembly for 1 min to remove residual fluid from the filter. Transfer the Filter Cartridge into a fresh Collection Tube. Apply 100 μL of pre-heated (95°C) Elution Solution to the  center of the filter. Leave it at the room temperature for 2 min. Spin for 20–30 sec at maximum speed  to recover the RNA. Collect the eluate (which contains the RNA) and store it at -70°C.
a. Add 5 μL rRNA Binding Buffer and 5 μL rRNA Removal Mix-Gold to each well with 10 μL total RNA (1 μg). Gently pipette the entire volume up anddown 6 times to mix thoroughly.  b. Incubate the PCR plate at 68°C for 5 min, and then place at room temperature for 1 min. c. Add 35 μL of rRNA Removal Beads to each well of the new PCR plate. Adjust the pipette to 45 μL, then with the tip of the pipette at the bottom of the well, pipette quickly up and down 20 times to mix thoroughly. Incubate at room temperature for 1 min. Place the PCR plate on the magnetic stand at room temperature for 1 min. Transfer all of the supernatant from each well to the corresponding well of the new PCR plate. d. Add 99 μL of RNAClean XP beads (If starting with degraded total RNA, add 193 μL of RNAClean XP beads to each well) to each well of the PCR plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. Incubate the PCR plate at room temperature for 15 min. e. Place the PCR plate on the magnetic stand at room temperature for 5 min. Remove and discard all of the supernatant from each well of the PCR plate. f. Add 200 μL freshly prepared 70% EtOH to each well. Incubate the PCR plate at room temperature for 30 s, and then remove and discard all of the supernatant from each well. Let the PCR plate stand at room temperature for 15 min to dry. g. Add 11 μL Elution Buffer to each well of the PCR plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. Incubate the PCR plate at room temperature for 2 min. Place the PCR plate on the magnetic stand at room temperature for 5 min. Transfer 8.5 μL supernatant from the PCR plate to the new 0.2 ml tube. h. Add 8.5 μL Elute, Prime, Fragment High Mix to each well of the plate. Gently pipette the entire volume up and down 6 times to mix thoroughly. i. Incubate at 94°C for 8 min, hold at 4°C, and then centrifuge shortlya. Add 8 μL First Strand Synthesis Act D Mix and SuperScript II Reverse Transcriptase in each well of the PCR plate. Gently pipette the entire volume up and down 6 times to mix thoroughly.  b. The mixture was incubated in the following progress: 25°C for 10 min, 42°C for 15 min, 70°C for 15 min, hold at 4°C.
2.3 Synthesize Second Strand cDNA  a. Add 5 μL End Repair Control (2 μL End Repair Control + 98 μL Resuspension Buffer) in each well of the PCR plate. b. Add 20 μL Second Strand Marking Master Mix. Gently pipette the entire volume up and down 6 times to mix thoroughly. c. Incubate at 16°C for 60 min. Remove the plate from the thermal cycler2.3.1 Purification  a. Add 90 μL mixed AMPure XP beads to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. Incubate the plate at room temperature for 15 min. b. Place it on the magnetic stand at room temperature for 5 min. c. Remove and discard 135 μL supernatant from each well of the plate. d. With the plate on the magnetic stand, add 200 μL freshly prepared 80% EtOH to each well. Incubate the plate at room temperature for 30 s, and then remove and discard all of the supernatant from each well. e. Repeat step d one time. f. Let the plate stand at room temperature for 15 min to dry, and then remove the plate from the magnetic stand. g. Add 17.5 μL Resuspension Buffer to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. h. Incubate the plate at room temperature for 2 min. Then place the plate on the magnetic stand at room temperature for 5 min. i. Transfer 15 μL supernatant from the plate to the new PCR plate. 2.4 Adenylate 3’ Ends a. Add 2.5 μL of diluted A-Tailing Control (1 μL A-Tailing Control + 99 μL 9  Resuspension Buffer) to each well of the plate.  b. Add 12.5 μL of A-Tailing Mix to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. c. The mixture was incubated in the following progress: 37°C for 30 min, 70°C for 5 min, hold at 4°C. 2.5 Ligate Adapters  a. Add 2.5 μL of diluted Ligation Control (1 μL Ligation Control + 99 μL Resuspension Buffer) to each well of the plate. b. Add 2.5 μL of Ligation Mix to each well of the plate. c. Add 2.5 μL of RNA Adapter Index to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. d. Incubate at 30°C for 10 min. e. Add 5 μL of Stop Ligation Buffer to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. 2.5.1 Purification  a. Add 42 μL of mixed AMPure XP Beads to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. Incubate the plate at room temperature for 15 min. b. Place the plate on the magnetic stand at room temperature for 15 min or until the liquid is clear. c. Remove and discard 79.5 μL supernatant from each well of the plate. d. With the plate on the magnetic stand, add 200 μL freshly prepared 80% EtOH to each well. Incubate the plate at room temperature for 30 s, and then remove and discard all of the supernatant from each well. e. Repeat step d one time for a total of two 80% EtOH washes. f. With the plate on the magnetic stand, let the samples air-dry at room temperature for 15 min. Remove the plate from the magnetic stand. g. Add 52.5 μL Resuspension Buffer to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. 10 h. Incubate the plate at room temperature for 2 min. Place the plate on the magnetic stand at room temperature for 5 minutes or until the liquid is clear. i. Transfer 50 μL supernatant from each well of the plate to the corresponding well of the new PCR plate. j. Add 50 μL of mixed AMPure XP Beads to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. Incubate the plate at room temperature for 15 min. k. Place the plate on the magnetic stand at room temperature for 15 min or until the liquid is clear. l. Remove and discard 95 μL supernatant from each well of the plate. m. With the plate on the magnetic stand, add 200 μL freshly prepared 80% EtOH to each well. Incubate the plate at room temperature for 30 s, and then remove and discard all of the supernatant from each well. n. Repeat step m one time for a total of two 80% EtOH washes. o. With the plate on the magnetic stand, let the samples air-dry at room temperature for 15 min, and then remove the plate from the magnetic stand. p. Add 22.5 μL Resuspension Buffer to each well of the plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. q. Incubate the plate at room temperature for 2 min. Place the plate on the magnetic stand at room temperature for 5 min or until the liquid is clear. r. Transfer 20 μL supernatant from each well of the plate to the corresponding well of the new PCR plate. 2.6 Enrich DNA Fragments  a. Add 5 μL of PCR Primer Cocktail to each well of the PCR plate. b. Add 25 μL of PCR Master Mix to each well of the PCR plate. Gently pipette the entire volume up and down to mix thoroughly. c. The mixture was incubated in the following progress: 1 cycle 98°C for 30 s; 15 cycles 98°C for 10 s, 60°C for 30 s, 72°C for 30 s; 1 cycle 72°C for 5 min; hold at 10°C. 11 2.7.1 Purification  a. Add 50 μL of the mixed AMPure XP Beads to each well of the PCR plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. Incubate the PCR plate at room temperature for 15 min. b. Place the plate on the magnetic stand at room temperature for 15 min or until the liquid is clear. c. Remove and discard 95 μL supernatant from each well of the PCR plate. d. With the PCR plate on the magnetic stand, add 200 μL freshly prepared 80% EtOH to each well. Incubate the PCR plate at room temperature for 30 s, and then remove and discard all of the supernatant from each well. e. Repeat step d one time for a total of two 80% EtOH washes. f. With the PCR plate on the magnetic stand, let the samples air-dry at room temperature for 15 min, and then remove the plate from the magnetic stand. g. Add 32.5 μL Resuspension Buffer to each well of the PCR plate. Gently pipette the entire volume up and down 10 times to mix thoroughly. h. Incubate the PCR plate at room temperature for 2 min. Place the PCR plate on the magnetic stand at room temperature for 5 min or until the liquid is clear. i. Transfer 30 μL supernatant from each well of the PCR plate to the corresponding well of the new PCR plate. 2.8 Validate library  a. Load 1 μL sample on an Agilent Technologies 2100 Bioanalyze. b. Check the size and purity of the sample

treatment_protocol: The collected tissues were sheared and excess red blood cells were removed from the tissues using erythrocyte lysate and digested using collagenase I, II, and IV to obtain single cell suspensions.

value: peri-implant tissue

value: ICR

value: WT

value: Non treatment


title: Con_2

description:
Con_2
Processed_data.txt


title: agshZIKV-h1#1-IP


title: agshZIKV-h1#1-Input


title: Lung_mTFF2 7d_rep3

description: Gene expression in lungs of mice infected by H7N9 with TFF2 treatment for 7 day

source_name: Lung, mTFF2 7d, replicate3


title: agshMHV-h1#2-IP


title: Lung_mTFF2 3d_rep3

description: Gene expression in lungs of mice infected by H7N9 with TFF2 treatment for 3 day

source_name: Lung, mTFF2 3d, replicate3


title: Lung_NC3d_rep3

description: Gene expression in lungs of mice infected by H7N9 with PBS treatment for 3 day

source_name: Lung, NC3d, replicate3

value: NC


title: agshMHV-h1#2-Input


title: Lung_NC7d_rep3

description: Gene expression in lungs of mice infected by H7N9 with PBS treatment for 7 day

source_name: Lung, NC7d, replicate3


title: Lung_NC1d_rep3

description: Gene expression in lungs of mice infected by H7N9 with PBS treatment for 1 day

source_name: Lung, NC1d, replicate3


title: Con_1

description:
Con_1
Processed_data.txt


title: Lung_Blank3_rep3

description: Gene expression in lungs of mice without H7N9 infection or treatment

source_name: Lung, Blank, replicate3

value: Blank


title: BlisterFluid41, SCAR, rep1

source_name: Hash 1_SCAR patient 18, blister fluid / Hash 2_SCAR patient 18, blister fluid

value: Female


title: agshMHV-h1#1-IP


title: agshMHV-h1#1-Input


title: post 10wAIR_F3_RNAseq

city: Minneapolis

first: Natalia

last: Tretyakova

state: Minnesota

address: 2231 6th St SE Room 2-147

department: Medicinal Chemistry

institute: University of Minnesota

description:
Project_017_Raw_Counts.txt.gz
Project_017_14w_FS_DEG_Test.csv.gz
14-f-air-3_S108

data_processing:
Trimmomatic to remove low quality bases and adapter contamination
HISAT2 to map read pairs to the mm10 genome
Samtools to sort mapped reads by query name and filter by mapping quality
featureCounts to quantify gene expression
edgeR to analyze gene expression data
Assembly: mm10
Supplementary files format and content: CSV with the test results for differential gene expression. Contains Ensembl ID, base mean expression, log2(fold change), nominal P-values, and FDR values

extract_protocol:
Alveolar type II epithelial cells (Type II pneumocytes) were isolated according to published procedures. Briefly, the lungs were perfused with 10 mL cold phosphate buffered saline (PBS) before enzymatic digestion with 2 mL of dispase infused into the lung, after which they were removed and incubated in an additional 2 mL of dispase for one hour. The lungs were then manually disintegrated and the resulting cell suspension labeled with antibodies specific for CD11c, CD11b, F4/80, CD19, CD45 and CD16/CD32 (Thermo Fisher Scientific). Samples from 9 mice were pooled to include three sets of lungs per sample for a total of three samples for FACS separation. Type II pneumocytes were isolated by negative selection and thus identified as the unlabeled cell population. Type II pneumocytes were also gated as sideward scatter high (SSChigh) cell population which minimizes contamination with lymphoid cells by selecting cells with a higher granularity. The cells were separated by fluorescence activated cell sorting (FACS) by the University Flow Cytometry Resource at the University of Minnesota using a BD FACS Aria II P07800142 (BSL2) (BD Biosciences, San Jose, CA). Following isolation via FACS, Type II pneumocytes were pelleted by centrifugation. The samples were first centrifuged for 12 min at 200 x g and 4°C and the supernatant was removed, except for the bottom 1 mL, which was transferred to a 1.7 mL Eppendorf tube. To this tube, 500 µL of PBS was added, and the tube was centrifuged for 12 min at 200 x g at 4°C. The supernatant was removed while the bottom 100 μL was saved. One mL of PBS was added to the tube, and the sample was centrifuged for 12 min at 800 x g and 4°C. After this final centrifugation, all supernatant is removed, and the cell pellet was saved for downstream analyses. A portion of the cell pellet (1.25 x 105 – 5 x 105 cells) was set aside to isolate protein. The remaining sample was used to isolate DNA and RNA using the Qiagen AllPrep DNA/RNA Mini Kit (Qiagen, Hilden Germany) according to the man-ufacturer’s instructions.
Total RNA samples were converted to Illumina sequencing libraries using SMARTer Stranded Total RNA-Seq Kit – Pico Mammalian Input (Takara Bio USA, Mountain View CA).
Alveolar type II epithelial cells (Type II pneumocytes) were isolated according to published procedures. Briefly, the lungs were perfused with 10 mL cold phosphate buffered saline (PBS) before enzymatic digestion with 2 mL of dispase infused into the lung, after which they were removed and incubated in an additional 2 mL of dispase for one hour. The lungs were then manually disintegrated and the resulting cell suspension labeled with antibodies specific for CD11c, CD11b, F4/80, CD19, CD45 and CD16/CD32 (Thermo Fisher Scientific). Samples from 9 mice were pooled to include three sets of lungs per sample for a total of three samples for FACS separation. Type II pneumocytes were isolated by negative selection and thus identified as the unlabeled cell population. Type II pneumocytes were also gated as sideward scatter high (SSChigh) cell population which minimizes contamination with lymphoid cells by selecting cells with a higher granularity. The cells were separated by fluorescence activated cell sorting (FACS) by the University Flow Cytometry Resource at the University of Minnesota using a BD FACS Aria II P07800142 (BSL2) (BD Biosciences, San Jose, CA). Following isolation via FACS, Type II pneumocytes were pelleted by centrifugation. The samples were first centrifuged for 12 min at 200 x g and 4°C and the supernatant was removed, except for the bottom 1 mL, which was transferred to a 1.7 mL Eppendorf tube. To this tube, 500 µL of PBS was added, and the tube was centrifuged for 12 min at 200 x g at 4°C. The supernatant was removed while the bottom 100 μL was saved. One mL of PBS was added to the tube, and the sample was centrifuged for 12 min at 800 x g and 4°C. After this final centrifugation, all supernatant is removed, and the cell pellet was saved for downstream analyses. A portion of the cell pellet (1.25 x 105 – 5 x 105 cells) was set aside to isolate protein. The remaining sample was used to isolate DNA and RNA using the Qiagen AllPrep DNA/RNA Mini Kit (Qiagen, Hilden Germany) according to the man-ufacturer’s instructions.

value: Alveolar type II epithelial cells

value: 14 weeks filtered air


title: BlisterFluid40, SCAR, rep1

value: Hash 1_SCAR patient 18, blister fluid / Hash 2_SCAR patient 18, blister fluid


title: post 10wAIR_F2_RNAseq

description:
Project_017_Raw_Counts.txt.gz
Project_017_14w_FS_DEG_Test.csv.gz
14-f-air-2_S119

value: Lung


title: agshEV71-h1#2-IP


title: post 10wAIR_F1_RNAseq

description:
Project_017_Raw_Counts.txt.gz
Project_017_14w_FS_DEG_Test.csv.gz
14-f-air-1_S107


title: agshEV71-h1#2-Input


title: BlisterFluid39, SCAR, rep1

source_name: Blister fluid

value: Blister fluid


title: BlisterFluid38, SCAR, rep1


title: BlisterFluid42, SCAR, rep1


title: post 10wCS_F3_RNAseq

description:
Project_017_Raw_Counts.txt.gz
Project_017_14w_FS_DEG_Test.csv.gz
14-f-cs-3_S111

value: 10 weeks cigarette smoke folowed by 4 weeks filtered air


title: BlisterFluid37, SCAR, rep1


title: 3wAIR_F2_RNAseq

description:
Project_011_Raw_Counts.txt.gz
Project_011_DEG_Test.csv.gz
3S-FC2_S5

value: 3 weeks filtered air