title: NaraCOVID_TP10_CD4_035_TCR


title: NaraCOVID_TP10_CD4_033_TCR

status: Public on Apr 01 2024

type: SRA

city: Noda

first: Hiroyasu

last: Aoki

state: Chiba

address: 2669 Yamazaki

department: Research Institute for Biomedical Sciences

country: Japan

institute: Tokyo University of Science

library_source: transcriptomic

data_processing:
Data processing of Bulk TCRseq was performed as described in Aoki et al. (Aoki et al., 2021).
Adapter trimming and quality filtering of sequencing data were performed using Cutadapt-3.2 and PRINSEQ-0.20.4. Sequencing data were processed by MiXCR-3.0.5. In MiXCR, filtered reads were aligned to reference mouse TCR V/D/J sequences registered in the international ImMunoGeneTics (IMGT) information system. The Variable (V) and Joining (J) segments of TCRs were represented in IMGT gene nomenclature. T-cell clones were determined as TCR reads with the same TCR V segment, J segment, and CDR3 nucleotide sequence.
Inter-sample contamination within samples using same UDIs were excluded by "Decontaminate" function of vdjtools with the parameter "-r 10".
For bulk TCRseq data, we regarded the CD4+-CD8+ overlapping clones in the same participant as contamination during T-cell isolation from PBMC because CD4+ and CD8+ T-cell repertoire are mutually exclusive (Aoki 2021). Thus, these overlapping clones were categorized into either CD4+ or CD8+ according to their dominance in CD4+ and CD8+ repertoire using VDJtools-1.2.1. The sequencing reads of bulk TCR-seq samples were normalized to 100,000 reads by the “DownSample” command of VDJtools-1.2.1. To preserve the minor clonotypes, we performed the normalization ten times and pooled these down-sampled repertoire data for each sample, by “JoinSamples” command of VDJtools-1.2.1.
Assembly: IMGT reference sequences of human TRA and TRB
Supplementary files format and content: Text files in VDJtools format include read count and frequency, complementarity determining region 3 (CDR3) nucleotide and amino acid sequences, Variable(V)/ Diversity(D)/ Joining(J) segment names, and V, D and J segment boundaries within CDR3 nucleotide sequence of individual clones
Library strategy: Bulk TCR-seq

molecule: total RNA

organism: Homo sapiens

source_name: Peripheral blood T cells

extract_protocol:
PBMCs were isolated from whole blood with BD vacutainer CPT tubes (BD), resuspended in CELLBANKER 1 (Takara), and stored in -80°C.
CD4+ and CD8+ T cells were magnetically isolated from PBMC. PBMC were thawed by warming frozen cryovials in a 37°C water bath. Cells were stained for 20 minutes at 4°C with 1st antibody mix (Antibody Mix 1 in Supplementary Table S1). After washing with MACS buffer [0.5% BSA (Nacalai), 2 mmol/L EDTA (NIPPON GENE) in PBS (Nacalai)], Cells were stained for 20 minutes at 4°C with 2nd antibody mix; BD IMag anti-PE Magnetic Particles-DM (Clone E31-1459, BD) and anti-APC-Biotin (clone APC003, BioLegend). Then, the cell suspension was diluted with MACS buffer and set on a magnet plate (DynaMag96SideSkirted, Veritas). The positive fraction was collected as CD4+ T cells. The negative fraction was stained with BD IMag Streptavidin Particles Plus-DM (BD) for 20 minutes at 4°C. Then, the cell suspension was diluted with MACS buffer and set on a magnet plate. The positive fraction was collected as CD8+ T cells and B cells. The counts of CD4+ T cells, CD8+ T cells, and B cells were determined using Flow-Count fluorospheres (Beckman) and a Cytoflex flow cytometer (Beckman) and FlowJo software (version 10.8.1; BD Biosciences).
Bulk TCR sequencing library was prepared according to the previous report (Aoki et al., 2023).  In brief, mRNA in T-cell lysates were captured by incubating with 10 μL of Dynabeads M270 streptavidin (Thermo Fisher Scientific) bound to 2.5 pmol of BioEcoP-dT25-adapter primers for 30 minutes at room temperature under gentle rotation. mRNA-trapped beads were suspended in 10 µL of RT mix and incubated for 60 min at 42ºC to perform reverse transcription (RT) and template-switching. The composition of the RT mix was as follows; 1× First Strand buffer (Thermo Fisher Scientific), 1 mM dNTP (Roche), 2.5 mM DTT (Thermo Fisher Scientific), 1 M betaine (Sigma-Aldrich), 9 mM MgCl2 (NIPPON GENE), 1 U/µL RNaseIn Plus RNase Inhibitor (Promega), 10 U/µL Superscript II (Thermo Fisher Scientific), and 1 µM of i5-TSO. To amplify the TCR cDNA containing complementarity determining region 3 (CDR3), nested PCR of the TCR locus was performed using KAPA Hifi Hotstart ReadyMix (KAPA Biosystems), followed by the purification of PCR product using an AM Pure XP kit (Beckman Coulter) at a 0.7:1 beads-to-sample ratio. The third PCR was performed to amplify TCR libraries and add adaptor sequences for the next-generation sequencer. The third-PCR products were purified as second PCR. Final TCR libraries, whose lengths were about 600 base pairs, were quantified using a KAPA SYBR Fast qPCR Kit (KAPA Biosystems), and size distribution was analyzed by Microchip Electrophoresis System MultiNA (Shimadzu). TCR libraries were sequenced using an Illumina Novaseq 6000 S4 flowcell (67 bp read 1 and 140 bp read 2) (Illumina) using NovaSeq 6000 S4 Reagent Kit v1.5 (200 cycles, Illumina). Only read2 contained the sequence regarding the definition of T-cell clones. Oligonucleotides and reaction conditions for library preparation were summarized in Table S2 and S3, respectively.

treatment_protocol: 38 healthcare workers (over 20 years old) at Nara Medical University in Japan received two doses of Pfizer-BNT162b2 vaccine between 10 March and 30 March 2021.

tag: treatment

value: Three shots of Pfizer-BNT162b2 vaccine

tag: cell type

value: CD4+ T cells

tag: age

tag: Sex

value: female

tag: date

value: 3 month after receiving the third shot


title: VUMC0809

type: RNA

city: Nashville

first: Cong

last: Wang

state: TN

address: 2525 West End Ave

country: USA

institute: Vanderbilt University

hyb_protocol: NanoString nCounter miRNA Expression Assay manufacturer's protocol

data_processing: miRNAs were normalized in the following steps: 1) background correction to remove non-specific binding. The background level was calculated as an aggregate summary of seven negative control probes with no target transcript. Mean of the calculated background level for each sample was then subtracted from the adjusted probe value to remove background noise. 2) Normalization of miRNA counts using mean counts of three spike-in synthetic control miRNAs (ath-miR159a, cel-miR-248, and osa-miR414) to control for variances on the starting material and the miRNA purification efficiency. 3) Sample content normalization using geometric mean counts of the top 50 miRNAs expressed excluding hsa-miR-320e, hsa-miR-16-5p and hsa-miR-451a because these miRNAs may be related to red blood cell contamination or hemolysis. After these steps, the expression level of each miRNA was calculated.

label: NanoString

source_name: plasma

extract_protocol: Qiagen’s miRNeasy Plasma Kit manufacturer's protocol

tag: study site (plco=1 swmhs=2 sccs=3 mec=4)

tag: case control status (control=0 case=1)

tag: sets

tag: sex (female=0 male=1)

tag: age at diagnosis

tag: race (white=1 black=2 other=3)


title: VUMC0807


title: VUMC0817


title: VUMC0806


title: NaraCOVID_TP10_CD4_032_TCR

value: male


title: VUMC0805


title: NaraCOVID_TP10_CD4_031_TCR


title: VUMC0804


title: VUMC0814


title: NaraCOVID_TP10_CD4_030_TCR


title: NaraCOVID_TP10_CD4_026_TCR


title: NaraCOVID_TP10_CD4_025_TCR


title: NaraCOVID_TP10_CD4_029_TCR


title: NaraCOVID_TP10_CD4_027_TCR


title: NaraCOVID_TP10_CD4_037_TCR


title: VUMC0811


title: VUMC0816


title: VUMC0803


title: NaraCOVID_TP10_CD4_024_TCR


title: VUMC0810


title: VUMC0802


title: VUMC0801


title: VUMC0798


title: VUMC0799


title: NaraCOVID_TP10_CD4_023_TCR


title: NaraCOVID_TP10_CD4_022_TCR


title: VUMC0800


title: VUMC0795


title: VUMC0797


title: NaraCOVID_TP10_CD4_021_TCR


title: VUMC0794


title: NaraCOVID_TP10_CD4_020_TCR


title: NaraCOVID_TP10_CD4_019_TCR


title: VUMC0792


title: NaraCOVID_TP10_CD4_018_TCR


title: VUMC0793


title: NaraCOVID_TP10_CD4_017_TCR


title: NaraCOVID_TP10_CD4_016_TCR


title: NaraCOVID_TP10_CD4_015_TCR


title: NaraCOVID_TP10_CD4_014_TCR


title: NaraCOVID_TP10_CD4_012_TCR


title: NaraCOVID_TP10_CD4_013_TCR


title: VUMC0789


title: NaraCOVID_TP10_CD4_011_TCR


title: VUMC0785


title: VUMC0787


title: VUMC0786


title: VUMC0791


title: NaraCOVID_TP10_CD4_010_TCR


title: NaraCOVID_TP10_CD4_007_TCR


title: NaraCOVID_TP10_CD4_006_TCR


title: VUMC0784


title: NaraCOVID_TP10_CD4_005_TCR


title: NaraCOVID_TP10_CD4_008_TCR


title: VUMC0779


title: VUMC0783


title: VUMC0782


title: VUMC0781


title: VUMC0788


title: VUMC0777


title: VUMC0776


title: VUMC0778


title: NaraCOVID_TP10_CD4_004_TCR


title: NaraCOVID_TP10_CD4_003_TCR


title: NaraCOVID_TP10_CD4_002_TCR


title: NaraCOVID_TP10_CD4_001_TCR


title: VUMC0775


title: VUMC0773


title: NaraCOVID_TP9_CD8_040_TCR

value: CD8+ T cells and CD19+ B cells

value: 1 month after receiving the third shot


title: NaraCOVID_TP9_CD8_039_TCR


title: VUMC0772


title: NaraCOVID_TP9_CD8_038_TCR


title: VUMC0770


title: VUMC0780


title: NaraCOVID_TP9_CD8_037_TCR


title: NaraCOVID_TP9_CD8_036_TCR


title: VUMC0769


title: VUMC0768


title: VUMC0767


title: VUMC0774


title: NaraCOVID_TP9_CD8_035_TCR


title: NaraCOVID_TP9_CD8_032_TCR


title: NaraCOVID_TP9_CD8_030_TCR


title: NaraCOVID_TP9_CD8_041_TCR


title: NaraCOVID_TP9_CD8_029_TCR


title: VUMC0763


title: NaraCOVID_TP9_CD8_031_TCR


title: VUMC0761


title: VUMC0762


title: NaraCOVID_TP9_CD8_027_TCR


title: VUMC0758


title: VUMC0760


title: VUMC0757


title: NaraCOVID_TP9_CD8_026_TCR


title: VUMC0756


title: NaraCOVID_TP9_CD8_024_TCR


title: NaraCOVID_TP9_CD8_023_TCR


title: VUMC0764


title: NaraCOVID_TP9_CD8_022_TCR


title: NaraCOVID_TP9_CD8_033_TCR


title: NaraCOVID_TP9_CD8_021_TCR


title: NaraCOVID_TP9_CD8_025_TCR


title: NaraCOVID_TP9_CD8_020_TCR


title: VUMC0754


title: VUMC0751


title: VUMC0753


title: VUMC0759


title: VUMC0752


title: VUMC0755


title: NaraCOVID_TP9_CD8_018_TCR


title: NaraCOVID_TP9_CD8_019_TCR


title: VUMC0748


title: NaraCOVID_TP9_CD8_013_TCR


title: NaraCOVID_TP9_CD8_016_TCR


title: NaraCOVID_TP9_CD8_012_TCR


title: VUMC0747


title: NaraCOVID_TP9_CD8_017_TCR


title: VUMC0746


title: VUMC0745


title: NaraCOVID_TP9_CD8_015_TCR


title: VUMC0742


title: VUMC0744


title: NaraCOVID_TP9_CD8_014_TCR


title: VUMC0750


title: NaraCOVID_TP9_CD8_011_TCR


title: NaraCOVID_TP9_CD8_008_TCR


title: NaraCOVID_TP9_CD8_010_TCR


title: VUMC0743


title: NaraCOVID_TP9_CD8_007_TCR


title: NaraCOVID_TP9_CD8_006_TCR


title: NaraCOVID_TP9_CD8_004_TCR


title: NaraCOVID_TP9_CD8_005_TCR


title: VUMC0739


title: VUMC0738


title: VUMC0737


title: VUMC0736


title: NaraCOVID_TP9_CD8_003_TCR


title: NaraCOVID_TP9_CD8_002_TCR


title: NaraCOVID_TP9_CD8_001_TCR


title: NaraCOVID_TP9_CD4_041_TCR


title: VUMC0732


title: VUMC0729


title: VUMC0735


title: VUMC0734


title: NaraCOVID_TP9_CD4_040_TCR


title: NaraCOVID_TP9_CD4_039_TCR


title: NaraCOVID_TP9_CD4_038_TCR


title: VUMC0731


title: VUMC0740


title: VUMC0727


title: VUMC0730


title: NaraCOVID_TP9_CD4_037_TCR


title: NaraCOVID_TP9_CD4_036_TCR


title: VUMC0726


title: VUMC0728


title: VUMC0724


title: VUMC0725


title: VUMC0723


title: NaraCOVID_TP9_CD4_035_TCR


title: NaraCOVID_TP9_CD4_033_TCR


title: NaraCOVID_TP9_CD4_030_TCR


title: VUMC0720


title: NaraCOVID_TP9_CD4_029_TCR


title: VUMC0722


title: VUMC0718


title: VUMC0721


title: NaraCOVID_TP9_CD4_023_TCR


title: VUMC0717


title: VUMC0716


title: NaraCOVID_TP9_CD4_025_TCR


title: VUMC0715


title: NaraCOVID_TP9_CD4_022_TCR


title: NaraCOVID_TP9_CD4_026_TCR


title: NaraCOVID_TP9_CD4_024_TCR


title: NaraCOVID_TP9_CD4_021_TCR


title: VUMC0712


title: VUMC0711


title: VUMC0709


title: VUMC0714


title: NaraCOVID_TP9_CD4_032_TCR


title: NaraCOVID_TP9_CD4_020_TCR


title: VUMC0710


title: NaraCOVID_TP9_CD4_019_TCR


title: VUMC0707


title: NaraCOVID_TP9_CD4_031_TCR


title: NaraCOVID_TP9_CD4_018_TCR


title: VUMC0719


title: NaraCOVID_TP9_CD4_027_TCR


title: NaraCOVID_TP9_CD4_016_TCR


title: VUMC0706


title: VUMC0704


title: VUMC0705


title: NaraCOVID_TP9_CD4_014_TCR


title: VUMC0708


title: VUMC0703


title: NaraCOVID_TP9_CD4_012_TCR


title: NaraCOVID_TP9_CD4_013_TCR


title: VUMC0700


title: VUMC0699


title: NaraCOVID_TP9_CD4_011_TCR


title: NaraCOVID_TP9_CD4_010_TCR


title: VUMC0697


title: NaraCOVID_TP9_CD4_008_TCR


title: VUMC0701


title: NaraCOVID_TP9_CD4_004_TCR


title: NaraCOVID_TP9_CD4_005_TCR


title: NaraCOVID_TP9_CD4_017_TCR


title: NaraCOVID_TP9_CD4_006_TCR


title: VUMC0691


title: NaraCOVID_TP9_CD4_015_TCR


title: VUMC0696


title: VUMC0692


title: VUMC0695


title: VUMC0690


title: NaraCOVID_TP9_CD4_003_TCR


title: VUMC0694


title: NaraCOVID_TP9_CD4_002_TCR


title: VUMC0689


title: VUMC0698


title: VUMC0688


title: NaraCOVID_TP8_CD8_041_TCR

value: 1 week after receiving the third shot


title: NaraCOVID_TP8_CD8_040_TCR


title: NaraCOVID_TP9_CD4_007_TCR


title: VUMC0686


title: NaraCOVID_TP8_CD8_037_TCR


title: NaraCOVID_TP8_CD8_038_TCR


title: NaraCOVID_TP8_CD8_036_TCR


title: VUMC0684


title: NaraCOVID_TP9_CD4_001_TCR


title: NaraCOVID_TP8_CD8_033_TCR


title: VUMC0679


title: VUMC0682


title: OE Spores Rep 4 [OE4_Spo]

city: Madison

first: Nancy

last: Keller

state: WI

address: 1550 Linden Drive

department: Medical Microbiology and Immunology

institute: University of Wisconsin-Madison

data_processing:
Given the raw reads (i.e. the .fq.gz files), we clipped the adapter sequences with Trimmomatic (version 0.32, settings 2:30:10:2:keepBothReads LEADING:5 TRAILING:5 MINLEN:36) (Bolger et al. 2014). Subsequently, we counted the transcripts with RSEM (Li & Dewey 2011) using A. fumigatus strain Af293’s reference transcriptome (ftp://ftp.ensemblgenomes.org/pub/fungi/release-49/gff3/aspergillus_fumigatus/Aspergillus_fumigatus.ASM265v1.49.gff3.gz). RSEM produced a transcripts-per-million (TPM) matrix. In this TPM matrix, the rows represent the genes, and the columns represent the samples in that data set. Finally, we log-transformed and quantile normalized the TPM matrix.
Assembly: A. fumigatus strain Af293 reference transcriptome was downloaded from   ftp://ftp.ensemblgenomes.org/pub/fungi/release-49/gff3/aspergillus_fumigatus/Aspergillus_fumigatus.ASM265v1.49.gff3.gz .
Supplementary files format and content: The processed data file "ZfpaMutant_TPM_normalized.tsv" is a tab-separated file where the rows represent the genes and the columns represent the samples.
Supplementary files format and content: The processed data file "readcount.csv" is a comma-separated text file that includes the raw counts for each sample.

organism: Aspergillus fumigatus

source_name: Resting Conidia

growth_protocol: Glucose minimal media plate overlayed with six mL top media containing one million conidia per mL and grown at 37 degrees celcius for 3 days

extract_protocol:
Conidia were collected into water with 0.01% Tween 80 and filtered thoroughly to remove hyphal fragments. Spores were pelleted at 1000xg and resuspended in a volume of 200 uL which was added to 1.5mL screwcap tubes containing 1mL QIAzol Lysis Reagent (Qiagen) and 250uL of 0.5 mm zirconia/silica beads and bead beaten for five minutes. RNA was extracted according to manufacture's instruction with an additional phenol:chloroform:isoamylalchohol (24:1:1) extraction step before RNA precipitation. Total RNA was further cleaned up using RNeasy Mini Kit with on-column DNase digestion (Qiagen) Library was prepared using TruSeq library preparation protocol with poly-A mRNA enrichment
Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers, followed by the second strand cDNA synthesis using dTTP. For the non-directional library, it was ready after end repair, A-tailing, adapter ligation, size selection, amplification, and purification. The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection. Quantified libraries will be pooled and sequenced on  Illumina platforms, according to effective library concentration and data amount

value: Resting Conidia

tag: genotype

value: ZfpA overexpression


title: NaraCOVID_TP8_CD8_039_TCR


title: NaraCOVID_TP8_CD8_032_TCR


title: NaraCOVID_TP8_CD8_035_TCR


title: OE Spores Rep 2 [OE2_Spo]


title: VUMC0677


title: VUMC0676


title: NaraCOVID_TP8_CD8_031_TCR


title: VUMC0678


title: VUMC0675


title: OE Spores Rep 3 [OE3_Spo]


title: VUMC0674


title: VUMC0673


title: NaraCOVID_TP8_CD8_030_TCR


title: VUMC0680


title: VUMC0683


title: OE Spores Rep 1 [OE1_Spo]


title: NaraCOVID_TP8_CD8_027_TCR


title: VUMC0672


title: NaraCOVID_TP8_CD8_026_TCR


title: VUMC0671


title: NaraCOVID_TP8_CD8_029_TCR


title: NaraCOVID_TP8_CD8_025_TCR


title: VUMC0681


title: KO Spores Rep 2 [KO2_Spo]

value: ZfpA knockout


title: VUMC0669


title: VUMC0667


title: VUMC0670


title: KO Spores Rep 1 [KO1_Spo]


title: VUMC0668


title: VUMC0664


title: NaraCOVID_TP8_CD8_024_TCR


title: VUMC0666


title: WT Spores Rep 4 [WT4_Spo]

value: WT


title: VUMC0662


title: VUMC0661


title: VUMC0663


title: NaraCOVID_TP8_CD8_023_TCR


title: WT Spores Rep 3 [WT3_Spo]


title: NaraCOVID_TP8_CD8_022_TCR


title: WT Spores Rep 2 [WT2_Spo]


title: KO Spores Rep 3 [KO3_Spo]


title: VUMC0656


title: KO Spores Rep 4 [KO4_Spo]


title: VUMC0658


title: VUMC0657


title: NaraCOVID_TP8_CD8_021_TCR


title: VUMC0660


title: VUMC0651


title: VUMC0653


title: VUMC0652


title: VUMC0655


title: WT Spores Rep 1 [WT1_Spo]

extract_protocol:
Conidia were collected into water with 0.01% Tween 80 and filtered thoroughly to remove hyphal fragments. Spores were pelleted at 1000xg and resuspended in a volume of 200 uL which was added to 1.5 mL screwcap tubes containing one mL QIAzol Lysis Reagent (Qiagen) and 250 uL of 0.5 mm zirconia/silica beads. Conidia were bead beaten for five minutes to disrupt cell wall, and RNA was extracted according to manufacture's instruction with an additional phenol:chloroform:isoamylalchohol (24:1:1) extraction step before RNA precipitation. Total RNA was further cleaned up using RNeasy Mini Kit with on-column DNase digestion (Qiagen) Library was prepared using TruSeq library preparation protocol with poly-A mRNA enrichment
Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers, followed by the second strand cDNA synthesis using dTTP. For the non-directional library, it was ready after end repair, A-tailing, adapter ligation, size selection, amplification, and purification. The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection. Quantified libraries will be pooled and sequenced on  Illumina platforms, according to effective library concentration and data amount


title: OE Mycelia Rep 2 [OE2_Myc]

source_name: Mycelia

growth_protocol: Two flasks of 50mL glucose minimal media each inoculated with 50 million spores and incubated at 37 degrees celcius and 250 rpm for 16hrs

extract_protocol:
Fungal biomas was harvested and combined from two flasks, flash-frozen in liquid nitrogen, and lyphilized completely. Total RNA was extracted with QIAzol Lysis Reagent (Qiagen) according to manufacture's instruction with additional phenol:chloroform:isoamylalchohol (24:1:1) extraction step before RNA precipitation. Total RNA was further cleaned up using RNeasy Mini Kit with on-column DNase digestion (Qiagen) Library was prepared using TruSeq library preparation protocol with poly-A mRNA enrichment
Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers, followed by the second strand cDNA synthesis using dTTP. For the non-directional library, it was ready after end repair, A-tailing, adapter ligation, size selection, amplification, and purification. The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection. Quantified libraries will be pooled and sequenced on  Illumina platforms, according to effective library concentration and data amount

value: Mycelia


title: VUMC0654


title: NaraCOVID_TP8_CD8_020_TCR


title: VUMC0650


title: VUMC0644


title: NaraCOVID_TP8_CD8_018_TCR


title: OE Mycelia Rep 4 [OE4_Myc]


title: OE Mycelia Rep 1 [OE1_Myc]


title: NaraCOVID_TP8_CD8_017_TCR


title: NaraCOVID_TP8_CD8_019_TCR


title: NaraCOVID_TP8_CD8_016_TCR


title: NaraCOVID_TP8_CD8_015_TCR


title: VUMC0642


title: KO Mycelia Rep 4 [KO4_Myc]


title: KO Mycelia Rep 1 [KO1_Myc]


title: VUMC0645


title: VUMC0648


title: VUMC0637


title: VUMC0647


title: NaraCOVID_TP8_CD8_012_TCR


title: VUMC0643


title: VUMC0636


title: VUMC0639


title: VUMC0641


title: NaraCOVID_TP8_CD8_014_TCR


title: VUMC0649


title: KO Mycelia Rep 2 [KO2_Myc]


title: OE Mycelia Rep 3 [OE3_Myc]


title: VUMC0640


title: NaraCOVID_TP8_CD8_013_TCR


title: VUMC0632


title: WT Mycelia Rep 4 [WT4_Myc]


title: VUMC0633


title: NaraCOVID_TP8_CD8_008_TCR


title: WT Mycelia Rep 1 [WT1_Myc]


title: NaraCOVID_TP8_CD8_007_TCR


title: WT Mycelia Rep 3 [WT3_Myc]


title: NaraCOVID_TP8_CD8_011_TCR


title: VUMC0634


title: KO Mycelia Rep 3 [KO3_Myc]


title: VUMC0626


title: VUMC0629


title: iPS cell line 10

city: Kagoshima

first: Sumito

last: Matsuya

address: 1-21-24 Korimoto

department: Joint Faculty of Veterinary Medicine

institute: Kagoshima university

data_processing:
CLC Genomics Workbench 22.0.1
Sequence reads were trimmed using CLC Genomics Workbench 22.0.1 (Quality limit = 0.05).
Trimmed sequence reads were mapped to Mus musculus GRCm38 release-94 using CLC Genomics Workbench 22.0.1.
Read count extacrion and normalization were performed using CLC Genomics Workbench 22.0.1.

organism: Mus minutoides

extract_protocol:
RNA was extracted using the Maxwell RSC simply RNA Cell Kit (Promega).
Library preparation for RNA-seq was performed using the NEBNext Ultra Ⅱ RNA Library Prep Kit for Illumina (NEB) following manifacturer's protocols.

tag: cell line

value: iPS cell

value: reprograming


title: VUMC0631


title: iPS cell line 6


title: VUMC0628


title: NaraCOVID_TP8_CD8_010_TCR


title: VUMC0625


title: NaraCOVID_TP8_CD8_004_TCR


title: NaraCOVID_TP8_CD8_005_TCR


title: WT Mycelia Rep 2 [WT2_Myc]


title: fibroblast line 2

value: fibroblast

value: no reprograming


title: VUMC0624


title: VUMC0635


title: VUMC0630


title: VUMC0621


title: VUMC0622


title: fibroblast line 1


title: VUMC0620


title: VUMC0627


title: VUMC0623


title: NaraCOVID_TP8_CD8_003_TCR

value: Three shots of Pfizer-BNT162b2 vaccine


title: MybM_OE_30hrs_Shield_rep_3

city: Philadelphia

first: Katelyn

middle: Ann

last: Walzer

state: PA

address: 380 South University Avenue

department: Pathobiology

institute: University of Pennsylvania

description: RNeasy minikit, Illumina stranded mRNA prep ligation kit (20040534)

data_processing:
Fastq files were pseudoaligned to the Cryptosporidium parvum Iowa II reference transcriptome (version 46, VEuPathDB) using Kallisto v.0.45.0
Kallisto transcript counts were imported into R using the Bioconductor package tximport
Expression data was normalized using the TMM method in the function calcNormFactors from the package edgeR
Differentially expressed genes (log2 fold-change ≥ 1 and p-value ≤ 0.05) were identified by linear modeling and Bayesian statistics using the limma package
Assembly: Version 46 of the Cryptosporidium parvum Iowa II reference from VEuPathDB
Supplementary files format and content: Tab-delimited text files include log2 counts per million (CPM) values for each sample

organism: Cryptosporidium parvum

growth_protocol: HCT-8 cells grown in a 24-well plate were infected with 250,000 oocysts of wild-type or Myb-M-HA overexpression parasites for 30 hours at 5% CO2 and 37º C

extract_protocol:
RNA was isolated using the RNeasy Mini Kit (Qiagen) and processed as per manufacturer’s instructions
Libraries were prepared using the Illumina stranded mRNA prep ligation kit (20040534), according to the manufacturer’s instructions

treatment_protocol: Cultures were treated with vehicle (0.1% ethanol) or Shield-1 (0.5 µM) for 6 hours starting at 24 hours (three biological replicates per strain and condition)

tag: source

value: Cryptosporidium parvum infected HCT-8 cells

tag: background strain

value: Cryptosporidium parvum Iowa II (Bunchgrass Farms)

value: cgd6_2250 (Myb-M) OE in TK locus

value: Shield-1 (0.5 µM) for 6 hours

tag: cell preparation

value: bulk


title: NaraCOVID_TP8_CD8_002_TCR

value: CD8+ T cells and CD19+ B cells

value: male


title: NaraCOVID_TP8_CD8_006_TCR

value: 1 week after receiving the third shot


title: VUMC0619


title: VUMC0617


title: MybM_OE_30hrs_Shield_rep_2


title: VUMC0618


title: VUMC0615


title: VUMC0616


title: NaraCOVID_TP8_CD8_001_TCR


title: MybM_OE_30hrs_Shield_rep_1


title: NaraCOVID_TP8_CD4_041_TCR

value: CD4+ T cells


title: MybM_OE_30hrs_vehicle_rep_3

value: vehicle (0.1% ethanol) for 6 hours


title: VUMC0613


title: VUMC0614


title: MybM_OE_30hrs_vehicle_rep_2


title: NaraCOVID_TP8_CD4_039_TCR


title: VUMC0612


title: NaraCOVID_TP8_CD4_038_TCR


title: VUMC0611


title: NaraCOVID_TP8_CD4_040_TCR


title: WT_30hrs_Shield_rep_3

value: wildtype


title: NaraCOVID_TP8_CD4_037_TCR

value: female


title: MybM_OE_30hrs_vehicle_rep_1


title: VUMC0608


title: VUMC0607


title: WT_30hrs_Shield_rep_2


title: VUMC0609


title: WT_30hrs_Shield_rep_1


title: NaraCOVID_TP8_CD4_036_TCR


title: VUMC0604


title: NaraCOVID_TP8_CD4_035_TCR


title: VUMC0606


title: VUMC0602


title: VUMC0597


title: VUMC0603


title: VUMC0600


title: VUMC0596


title: WT_30hrs_vehicle_rep_3


title: NaraCOVID_TP8_CD4_033_TCR


title: VUMC0595


title: WT_30hrs_vehicle_rep_2


title: VUMC0594


title: NaraCOVID_TP8_CD4_032_TCR


title: WT_30hrs_vehicle_rep_1


title: VUMC0605


title: NaraCOVID_TP8_CD4_031_TCR


title: NaraCOVID_TP8_CD4_030_TCR


title: NaraCOVID_TP8_CD4_029_TCR


title: VUMC0593


title: VUMC0592


title: VUMC0591


title: NaraCOVID_TP8_CD4_027_TCR


title: NaraCOVID_TP8_CD4_026_TCR


title: VUMC0588


title: VUMC0589


title: NaraCOVID_TP8_CD4_025_TCR


title: VUMC0590


title: VUMC0587


title: VUMC0583


title: VUMC0585


title: VUMC0584


title: VUMC0580


title: VUMC0582


title: VUMC0578


title: VUMC0581


title: NaraCOVID_TP8_CD4_024_TCR


title: NaraCOVID_TP8_CD4_023_TCR


title: VUMC0577


title: NaraCOVID_TP8_CD4_022_TCR


title: VUMC0574


title: NaraCOVID_TP8_CD4_021_TCR


title: VUMC0575


title: VUMC0573


title: VUMC0572


title: VUMC0570


title: VUMC0586


title: NaraCOVID_TP8_CD4_018_TCR


title: NaraCOVID_TP8_CD4_019_TCR


title: VUMC0567


title: VUMC0568


title: VUMC0566


title: VUMC0576


title: NaraCOVID_TP8_CD4_015_TCR


title: VUMC0562


title: NaraCOVID_TP8_CD4_017_TCR


title: VUMC0560


title: VUMC0559


title: VUMC0565


title: VUMC0558


title: VUMC0557


title: VUMC0556


title: NaraCOVID_TP8_CD4_020_TCR


title: NaraCOVID_TP8_CD4_014_TCR


title: VUMC0571


title: VUMC0555


title: NaraCOVID_TP8_CD4_013_TCR


title: VUMC0554


title: VUMC0569


title: NaraCOVID_TP8_CD4_012_TCR


title: VUMC0553


title: VUMC0552


title: NaraCOVID_TP8_CD4_016_TCR


title: VUMC0551


title: I_R_2

city: Changchun

first: Tongjian

last: Zhao

address: 1266 Fujin Road

country: China

institute: School of Pharmaceutical Sciences, Jilin University

data_processing:
Hisat2 for sequence alignment.
Stringtie tools for transcript assembly, transcript abundance estimation, gene structure optimization.
Samtools tools for related file processing.
Assembly: mm9
Supplementary files format and content: mRNA_FPKM.txt: Tab-delimited text file includes FPKM values for each sample.

organism: Mus musculus

source_name: Heart

extract_protocol:
TRIzol (Thermo Fisher, 15596018) was used to separate and purify total RNA from the samples, following the manufacturer's instructions. The quantity and purity of the total RNA were assessed using NanoDrop ND-1000 (NanoDrop, Wilmington, DE, USA), and the integrity of RNA was evaluated using Bioanalyzer 2100 (Agilent, CA, USA).
Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina following the manufacturer’s recommendations and index codes were added to attribute sequences to each sample.

tag: tissue

value: Heart

value: MIRI

tag: strain

value: Male


title: I_R_3

tag: treatment


title: NaraCOVID_TP8_CD4_010_TCR

tag: cell type

tag: age


title: I_R_1

tag: Sex


title: NaraCOVID_TP8_CD4_008_TCR

tag: date


title: VUMC0549

tag: study site (plco=1 swmhs=2 sccs=3 mec=4)

tag: case control status (control=0 case=1)


title: NaraCOVID_TP8_CD4_007_TCR


title: VUMC0550

tag: sets

tag: sex (female=0 male=1)


title: NaraCOVID_TP8_CD4_006_TCR


title: C_3

value: Sham


title: NaraCOVID_TP8_CD4_011_TCR

value: Three shots of Pfizer-BNT162b2 vaccine


title: C_2


title: VUMC0548

tag: age at diagnosis

tag: race (white=1 black=2 other=3)


title: VUMC0547


title: VUMC0561


title: VUMC0546


title: VUMC0545


title: Arabidopsis thaliana 6d seedling total RNA-seq r3

city: Pullman

first: Sascha

last: Duttke

state: Washington

address: BLS 202, 100 Dairy Road

department: School of Molecular Biosciences

institute: Washington State University

description:
total RNA-seq (Ribo0), steady state transcriptome profiling in A.thaliana 6d seedling; Experiment MS320
A.thaliana-6dseedling-10.tss.txt

data_processing:
csRNA-seq (small capped RNAs, ~20-60 nt) and sRNA-seq reads (“input”) were trimmed of their adapter sequences using HOMER (“homerTools trim -3 AGATCGGAAGAGCACACGTCT -mis 2 -minMatchLength 4 -min 20" none -f {csRNA_fastq_path}/*fastq.gz”)(Heinz et al. 2010). Reads were aligned to the appropriate genome using Hisat2 version 2.2.1 (Kim et. al. 2019) with parameters “hisat2 -p 30 --rna-strandness RF --dta -x {hisat2_genome_index} -U {path_rimmed_csRNA or sRNA} -S {output_sam} 2> {mapping_stats}”. Hisat2 indices were then created for each genome used ("hisat2-build -p 40 genome.dna.toplevel.fa {Hisat2_indexfolder}") with exception for barely which required the addition of "--large-index" due to genome size. Tag directory folders were then created using HOMER makeTagDirectories ("makeTagDirectories {outputName} -omitSN -checkGC -fragLength 150 -single")(Heinz et. al. 2010). Only reads with a single, unique alignment (MAPQ >=10) were considered in the downstream analysis. This consolidates the aligned read information into a read/chromosome format along with other important alignment information. Peaks were called and annotated using HOMER ("findcsRNATSS.pl {csRNA} -o {output} -i {sRNA} -genome hg38 -size 150 -ntagThreshold 10") (Duttke et al. 2019) and annotatePeaks.pl ("annotatePeaks.pl {tss/tsr.txt} hg38 -strand + -fragLength 1 -rlog -d {listOfTagDirs} > {output}") (Heinz et. al. 2010) respectively (additional details available in the manuscript).
Transcription start sites (TSSs) and Transcription start regions (TSRs) were determined in this study. TSRs  were called using findcsRNATSS.pl (detailed above) while TSSs were called using getTSSfromReads.pl ("getTSSfromReads.pl -d {csRNA_tagdir} -dinput {sRNA_tagdir} -min 7 > {output_file}") (Heinz et. al. 2010). To ensure high quality TSSs, at least 7 per 10^7 aligned reads were required and TSS were required to be within called TSR regions.
Paired end total ribosomal RNA-depeleted RNA-seq libraries were trimmed using skewer (“time -p skewer -m mp {read1} {read2} -t 40 -o {trimmed_fastq_output}”) (Jiang et al. 2014) and aligned using Hisat2 (Kim et al. 2019) to ensure all data were processed as similar as possible (“hisat2 -p 30 --rna-strandness RF --dta -x {hisat2_index} -1 {trimmed_RNAseq_R1} -2 {trimmed_RNAseq_R2} -S {output_sam} 2> {mapping_file}”). In this manuscript, total RNA-seq was exclusively used to determine RNA-stability as described in the csRNA-seq analysis.
Assembly: TAIR10, Phypa V3
Supplementary files format and content: bed files: Locations indicate TSS positions. TSS.txt files: text files containing tab delimited  information about found transcription start site culsters (TSRs). Motif file containing new line delimited motifs. Motif files: files containing motifs used with a ‘>’ indicating motif information followed by rows with nucleotide probability.

organism: Arabidopsis thaliana

source_name: 6d seedling

growth_protocol: Physcomitrium (Physcomitrella) patens (Gransden) was grown on plates with BCDA medium in a growth cabinet at 21°C under 16h light.

extract_protocol:
A. thaliana seedlings tissue was ground using the Qiagen TissueLyser II, at 1/30s frequency for 1.5 min twice. RNA was purified using the Zymo Direct-zol RNA MiniPrep kit.
csRNA-seq was performed as described in (S. H. Duttke et al. 2019). Small RNAs of ~20-60 nt were size selected from 0.4-3 µg of total RNA by denaturing gel electrophoresis. A 10% input sample was taken aside and the remainder enriched for 5’-capped RNAs. Monophosphorylated RNAs were selectively degraded by 1 hour incubation with Terminator 5´-Phosphate-Dependent Exonuclease (Lucigen). Subsequently, RNAs were 5’dephosporylated through 90 minutes incubation in total with thermostable QuickCIP (NEB) in which the samples were briefly heated to 75°C and quickly chilled on ice at the 60 minutes mark. Input (sRNA) and csRNA-seq libraries were prepared as described in (Hetzel et al. 2016) using RppH (NEB) and the NEBNext Small RNA Library Prep kit, amplified for 11-14 cycles. Total RNA-Seq Library Preparation Strand-specific, paired-end libraries were prepared from total RNA by ribosomal depletion using the Ribo-Zero Gold plant rRNA removal kit (Illumina, San Diego, CA). Samples were processed following the manufacturer’s instructions.

value: 6d seedling

tag: genotype


title: C_1


title: NaraCOVID_TP8_CD4_004_TCR


title: NaraCOVID_TP8_CD4_002_TCR


title: VUMC0544


title: NaraCOVID_TP8_CD4_003_TCR


title: Arabidopsis thaliana 6d seedling total RNA-seq r1

description:
total RNA-seq (Ribo0), steady state transcriptome profiling in A.thaliana 6d seedling; Experiment MS316
A.thaliana-6dseedling-10.tss.txt


title: VUMC0541


title: VUMC0542


title: High temperature

city: Tianjin

first: Yangyang

last: Zheng

address: 135 Yaguan Road, Jinnan District, Tianjin

institute: Tianjin University

description: DNA fragment

library_source: genomic

data_processing:
Illumina protocol for amplicon sequencing
The editing cassette was used to quantify fitness associated with different mutations in the library. Fitness associated with all mutations in the library was calculated using log2-fold enrichment. Synonymous mutations of every homology arm were regard as a part of each corresponding library. Synonymous mutations were used to determine the standard deviation and mean value of the log2-fitness change of wild-type cells. Then, a mean ± 2σ cutoff (i.e., p=0.05 assuming a normal distribution) was used to estimate strongly enriched mutations in the selections. The confidence interval of 95% was used to calculate statistically significant differences between samples. Differences with p ≤ 0.05 were considered statistically significant.
Assembly: Escherichia coli str. K-12 substr. MG1655, complete genome.
Supplementary files format and content: 91 potential positive mutations selected by CREATE deep sequencing and fitness calculation

molecule: genomic DNA

source_name: Escherichia coli

growth_protocol: Acetate, NaCl, furfural, isobutanol and high temperature were selected as stress screening conditions to verify the library tolerance in mineral medium (M9 medium) . M9 medium contained (per liter of deionized water) 1 g of NH4Cl, 0.5 g of NaCl, 4 g of Na2HPO4, and 3 g of KH2PO4. The following components were sterilized separately and then added to M9 medium (per liter [final volume] of medium): 2 mL of 1 M MgSO4, 1 mL of 0.1 M CaCl2, glucose or xylose, 10 mL of a trace element solution containing (per liter) 1 g of FeCl3, 0.18 g of ZnSO4· 7 H2O, 0.12 g of CuCl2· 2 H2O, 0.12 g of MnSO4· H2O, and 0.18 g of CoCl2· 6 H2O

extract_protocol:
Mutations on chromosomes were consistent with those carried by the CREATE plasmids. Firstly, the CREATE plasmids of tolerant strains were extracted separately. Following extraction, the CREATE plasmids were used as templates for PCR using Illumina-compatible primers designed with Golay barcodes . CREATE library fragments were amplified in 16 cycles of PCR. At the first PCR step, each experimental sample was ligated with a unique barcode used to track the mutations. After that, PCR products were confirmed by electrophoresis and purified using an Agarose Gel DNA Extraction Kit. Deep sequencing was performed according to the Illumina protocol for amplicon sequencing.
using global regulator libraries based on the CRISPR-enabled trackable genome engineering (CREATE) method to engineer tolerance against multiple inhibitors in Escherichia coli. Deep mutagenesis libraries were rationally designed, constructed, and screened to target 34,340 mutations across 23 global regulators.These libraries were divided into G1, G2, G3, G4 and G5 sub-groups according to different gene functions (such as active sites, DNA binding sites, and predicted functional sites) .
Illumina protocol for amplicon sequencing

tag: type

value: plasmids

value: PCR


title: NaraCOVID_TP8_CD4_001_TCR


title: VUMC0540


title: Acetate

description: DNA fragment

organism: Escherichia coli str. K-12 substr. MG1655


title: NaraCOVID_TP6_CD8_041_TCR

value: 6 month after receiving the second shot


title: VUMC0539


title: 3T3_CDX2, RNA, Dox, rep2

city: Lausanne

first: David

middle: Michael

last: Suter

address: EPFL SV-IN Station 19

department: Institute of Bioengineering

country: Switzerland

institute: École polytechnique fédérale de Lausanne

data_processing:
RNA-seq reads were aligned to the mm10 genome using STAR 2.7.1a, allowing only uniquely mapped reads. Reads not aligning to chr1-19, chrX, or, chrY were removed using SAMTools 1.10 and duplicates were removed using Picard. Reads in refGene exons were counted using HOMER 4.10 analyzeRepeats.pl. Counts were normalised using edgeR TMM normalisation and differentially expressed transcripts determined using limma with design matrix ~0+Condition+Replicate
Assembly: mm10
Supplementary files format and content: Differential expression analysis result table

molecule: nuclear RNA

source_name: 3T3 iCDX2

growth_protocol: NIH3T3 cell lines stably expressing H2B-mCherry, the rtTA3G transactivator, and TRE3G-controlled (doxycycline (dox)-inducible) CDX2were maintained in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with Glutamax, 2 μg/ml Puromycin, and 1% Penicillin-Streptomycin, 10% Fetal bovine serum , and 1 mM sodium pyruvate  at 37°C with 5% CO2

extract_protocol:
500,000 NIH-3T3 iCDX2 cells were treated for 6 hours with 500ng/ml Doxycycline or solvent alone prior to cell collection. Cell pellets were resuspended in 500 μlL Nuclear RNA lysis buffer (10 mM Tris pH 7.4, 10 mM NaCl, 3 mMMgCl2, 1% BSA (Merck, B9000S), 0.1% Tween (Fisher Scientific, 10113103), 1 mM DTT (Merck, 646563), 0.5U/μl Protector RNase Inhibitor (Merck, 3335399001), 0.1% NP-40, 0.01% Digitonin (Invitrogen, BN2006)) and incubated 5 min on ice. Then 500 μl Nuclear RNA wash buffer (10 mM Tris pH 7.4, 10 mM NaCl, 3 mM MgCl2, 1% BSA (Merck, B9000S), 0.1% Tween (Fisher Scientific, 10113103), 1 mM DTT (Merck, 646563), 0.5U/μl Protector RNase Inhibitor (Merck, 3335402001) were added before spinning down cells at 300g 4°C for 5 min. Two more washes with Nuclear RNA wash buffer were performed before proceeding to RNA extraction using the Rneasy Plus Micro Kit (Qiagen,74034) with the addition of an on-column DNAse digestion (Qiagen, 79254)MgCl2, 1% BSA (Merck, B9000S), 0.1% Tween (Fisher Scientific, 10113103), 1 mM DTT (Merck, 646563), 0.5U/μl Protector RNase Inhibitor (Merck, 3335399001), 0.1% NP-40, 0.01% Digitonin (Invitrogen, BN2006)) and incubated 5 min on ice. Then 500 μl Nuclear RNA wash buffer (10 mM Tris pH 7.4, 10 mM NaCl, 3 mM MgCl2, 1% BSA (Merck, B9000S), 0.1% Tween (Fisher Scientific, 10113103), 1 mM DTT (Merck, 646563), 0.5U/μl Protector RNase Inhibitor (Merck, 3335402001) were added before spinning down cells at 300g 4°C for 5 min. Two more washes with Nuclear RNA wash buffer were performed before proceeding to RNA extraction using the Rneasy Plus Micro Kit (Qiagen,74034) with the addition of an on-column DNAse digestion (Qiagen, 79254)
NEBNext Ultra II Directional RNA Library Prep Kit for Illumina

treatment_protocol: CDX2 expression was induced by 6-hour treatment with 500ng/µL Doxycycline

tag: cell line

value: 3T3 iCDX2

value: +Dox (6h)


title: NaraCOVID_TP6_CD8_040_TCR

value: CD8+ T cells and CD19+ B cells

value: male


title: NaraCOVID_TP6_CD8_039_TCR


title: VUMC0538


title: Furfural

description: DNA fragment


title: 3T3_CDX2, RNA, noDox, rep2

value: -Dox (6h)


title: NaraCOVID_TP6_CD8_038_TCR


title: M5brain,Model

city: Harbin

first: Bin

last: Li

state: Heilongjiang

address: No. 23, Postal Street, Nangang District

department: Department of Cardiology

institute: First Affiliated Hospital of Harbin Medical University

data_processing:
Samples are sequenced on the platform to get image files, which are transformed by the software of the sequencing platform, and the original data in FASTQ format (Raw Data) is generated. Sequencing data contains a number of connectors, low-quality Reads, so we use fastp(0.22.0) software to filter the sequencing data to get high quality sequence (Clean Data) for further analysis
The reference genome and gene annotation files were downloaded from genome website. The filtered reads were mapping to the reference genome using HISAT2 (v2.1.0).
We used HTSeq (v0.9.1) statistics to compare the Read Count values on each gene as the original expression of the gene, and then used FPKM to standardize the expression.
Assembly: Assembly Rnor6.0
Supplementary files format and content: gene_fpkm.txt

organism: Rattus norvegicus

source_name: Paraventricular hypothalamic nucleus

extract_protocol:
Total RNA was isolated using the Trizol Reagent (Invitrogen Life Technologies), after which the concentration, quality and integrity were determined using a NanoDrop spectrophotometer (Thermo Scientific). Three micrograms of RNA were used as input material for the RNA sample preparations.
Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (#E7530L, NEB, USA) following the manufacturer’s recommendations and index codes were added to attribute sequences to each sample.

value: Paraventricular hypothalamic nucleus

value: WT

value: Immobilized stress


title: 3T3_CDX2, RNA, Dox, rep1


title: Isobutanol

description: DNA fragment


title: M4brain,Model


title: NaraCOVID_TP6_CD8_036_TCR


title: 3T3_CDX2, RNA, noDox, rep1


title: NaraCOVID_TP6_CD8_037_TCR

value: female


title: VUMC0536


title: 3T3_CDX2, HiC, Dox24h, rep2

data_processing:
distiller-nf 0.3.3 was used to align Hi-C data to the mm10 genome
Assembly: mm10
Supplementary files format and content: Multi-resolution cooler (.mcool) file of merged replicates

extract_protocol:
Two million cells were fixed in 2% formaldehyde solution (AppliChem, catalog no. UN2209), and the reaction was quenched with 2 µM glycine (VWR Chemicals, catalog no. 101196X). Cells were incubated in lysis buffer (10 mM Tris-HCl, pH 8.0 (Invitrogen, Thermo Fisher Scientific, catalog no. 15568025), 10 mM NaCl (Sigma-Aldrich, catalog no. 59222C), 0.2% IGEPAL CA-630 (Sigma-Aldrich, catalog no. I8896), 1× cOmplete Protease Inhibitor Cocktail (Sigma-Aldrich, catalog no. 11697498001)) for 30 min at 4 °C. The nuclei were spun down, resuspended in NEB3.1 buffer (New England Biolabs, catalog no. B7203S) and 0.1% sodium dodecylsulfate (SDS; Carl Roth, catalog no. CN30) final concentration, and incubated at 65 °C for 10 min. The SDS was quenched by the addition of 1% Triton X-100 (AppliChem, catalog no. A1388). The chromatin was then digested with 100 U of restriction endonuclease MboI (New England Biolabs, catalog no. R0147) overnight at 37 °C. Digested fragments were filled in with 0.4 mM biotin (Invitrogen, Thermo Fisher Scientific, catalog no. 19524016), 10 mM dCTP, 10 mM dGTP, 10 mM TTP (Promega, catalog no. U1330) using 50 U µl−1 of DNA polymerase I, large Klenow fragment (New England Biolabs, catalog no. M0210) and incubated at room temperature for 4 h. Proximity ligation of the biotin-filled ends was performed by the addition of 5 U µl−1 of T4 DNA Ligase (Thermo Fisher Scientific, catalog no. EL0011), 0.5× bovine serum albumin (New England Biolabs, catalog no. B9000S) and 1% Triton X-100 to the samples, which were incubated at room temperature for an additional 4 h. Crosslinks were reversed by treatment with 300 mM NaCl and 1% SDS overnight at 68 °C. Samples were treated with 50 µg ml−1 of RNase A (Thermo Fisher Scientific, catalog no. EN0531) for 30 min at 37 °C, followed by incubation with 400 µg ml−1 of proteinase K (Promega, catalog no. V3021) at 65 °C for 1 h. DNA was precipitated with 1.6× volumes of 100% ethanol and 0.1× volume of 3 M sodium acetate, pH 5.2 (Thermo Fisher Scientific, catalog no. R1181) and incubation at −80 °C for 1 h. DNA was purified with 70% ethanol and fragmented by sonication (Covaris E220). DNA was double size selected with 0.575×, followed by 1.2×, volumes of Ampure XP beads (Beckman Coulter, catalog no. A6388) to obtain fragments sizes of 300–700 bp. Biotin-bound fragments were isolated on Dynabeads MyOne Streptavidin T1 beads (Thermo Fisher Scientific, catalog no. 65602) by incubation in binding buffer (10 mM Tris-HCl, pH 7.5 (Thermo Fisher Scientific, catalog no. 15567027), 1 mM ethylenediaminetetraacetic acid (Invitrogen, Thermo Fisher Scientific, catalog no. AM9260G), 2 M NaCl) for 30 min at room temperature.
Bead-bound DNA is subjected to end polishing though the addition of 25 mM each of a dNTP mix, 3 U µl−1 of T4 DNA polymerase, 5 U µl−1 of DNA polymerase I, large Klenow fragment, 10 U µl−1 of T4 polynucleotide kinase in 1× T4 DNA ligase reaction buffer, and incubation for 30 min at room temperature. The dA tail was added to the fragments by incubation with 10 mM dATP and 5 U µl−1 of Klenow fragment 3′→5′ exonuclease for 30 min at 37 °C. Illumina Indexed TruSeq adapters were ligated to the DNA fragments with T4 DNA ligase and incubated at room temperature for 2 h. Finally, the bead-bound libraries were amplified using KAPA HiFi HotStart ReadyMix PCR Kit, and universal Illumina forward and reverse primers.

treatment_protocol: CDX2 expression was induced by 6-hour or 24-hour treatment with 500ng/µL Doxycycline


title: VUMC0535


title: VUMC0534


title: M3brain,Model


title: M2brain,Model


title: 3T3_CDX2, HiC, Dox24h, rep1


title: NaraCOVID_TP6_CD8_035_TCR


title: VUMC0533


title: M1brain,Model


title: 3T3_CDX2, HiC, Dox6h, rep2


title: D5brain,Minocycline

value: Minocycline treatment and immobilized stress


title: VUMC0532


title: NaraCOVID_TP6_CD8_032_TCR


title: NaCl

description: DNA fragment


title: 3T3_CDX2, HiC, Dox6h, rep1


title: VUMC0537


title: NaraCOVID_TP6_CD8_031_TCR


title: 3T3_CDX2, HiC, noDox, rep2

value: -Dox


title: D4brain,Minocycline


title: VUMC0530


title: VUMC0531


title: D3brain,Minocycline


title: D2brain,Minocycline


title: NaraCOVID_TP6_CD8_029_TCR


title: FEL4-MT4,scRNAseq

city: Cincinnati

first: Guolun

state: OH

address: 3333 Burnet Avenue

institute: Cincinnati Children's Hospital Medical Center

description: 10X Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcode processing and gene counting were achieved by using the cell ranger software v7.0.
Assembly: mm10
Supplementary files format and content: H5 files

source_name: lung cells

extract_protocol:
Freshly prepared lung cells are FACS purified from mouse chimeric lung generated from blastocyst complementation. The tissues were digested with liberate to prepare single cell suspension.
Library was constructed according to the manufacter’s instructions (single cell 3’ v3.1 protocol, 10X genomics). The cells prepared were resuspended in the the Chromium Single Cell 3’ master mix solution and processed by microfluidic partitioning to capture single cells and create next-generation sequencing (NGS) cDNA libraries in Gel Beads in emulsion.The single cells were lysed, and the Gel Bead is dissolved to release barcoded RT oligonucleotides into solution, and reverse transcription of polyadenylated mRNA occurs. This results in all cDNAs from a single cell having the same barcode for mapping back to their original cell of origin. Next, Illumina primer and adapter sequences were incorporated to make paired-ended constructs with P5 and P7 to assist next generation sequencing.

value: lung cells

value: chimera

value: FACS


title: NaraCOVID_TP6_CD8_027_TCR

value: Three shots of Pfizer-BNT162b2 vaccine


title: VUMC0528


title: NaraCOVID_TP6_CD8_033_TCR


title: D1brain,Minocycline


title: VUMC0527


title: FEL1-MT1,scRNAseq

description: 10X Genomics


title: NaraCOVID_TP6_CD8_026_TCR


title: FEL0-WT,scRNAseq

description: 10X Genomics


title: NaraCOVID_TP6_CD8_025_TCR


title: C4brain, Control

value: No treatment


title: VUMC0526


title: NaraCOVID_TP6_CD8_024_TCR

value: 6 month after receiving the second shot


title: NaraCOVID_TP6_CD8_030_TCR


title: C3brain, Control


title: VUMC0525


title: 4T1, RI, 5

city: Guangzhou

first: Yanpei

last: Zhang

state: Guangdong

address: No.1023,South Shatai Road, Baiyun District, Guangzhou, Guangdong, China

institute: Southern Medical University

data_processing:
Illumina Casava1.7 software used for basecalling.
HISAT2 software was used to conduct genome mapping analysis of preprocessed sequencing sequences. Usually we compare clean No RNA reads to the genome, which is the basis of subsequent analysis. HISAT2 uses global and local search methods, which can perform efficient mapping and effectively compare splICED reads in RNA Seq sequencing data. The default parameters are set, and the reference genome version is GRCm38.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Assembly: mm10 (GRCm38)
Supplementary files format and content: gene_fpkm_matrix.csv: The CSV file includes the FPKM values for each sample.

source_name: tumor

extract_protocol:
Tumors were harvested, and total RNA was isolated and reverse transcribed into cDNA to construct an indexed Illumina library.
RNA libraries were prepared for sequencing using standard Illumina protocols.

value: tumor

value: Murine breast cancer cells

value: radiation+immune checkpoint inhibitor


title: 3T3_CDX2, HiC, noDox, rep1

value: 3T3 iCDX2


title: 4T1, RI, 2


title: VUMC0529


title: VUMC0524


title: 4T1, RI, 4


title: NaraCOVID_TP6_CD8_023_TCR

value: CD8+ T cells and CD19+ B cells

value: male


title: C2brain, Control

value: Paraventricular hypothalamic nucleus

value: WT


title: VUMC0523


title: 4T1, RI, 1


title: VUMC0522


title: NaraCOVID_TP6_CD8_021_TCR


title: 4T1, ICI, 4

value: immune checkpoint inhibitor


title: C1brain, Control


title: C5brain, Control


title: MSC42_UTD_Baseline

city: Rochester

first: Saad

last: Kenderian

state: MN

address: 200 First Street SW

department: Hematology Research

institute: Mayo Clinic

data_processing:
Binary base call data converted to fastq using Illumina bcl2fastq software
Check quality with FastQC, trim adapters with trimmomatic, and re-check quality with FastQC
Latest human reference genome GRCh38 was downloaded from NCBI and indexed using STAR
Paired end reads were mapped to the genome, and HTSeq was used to generate expression counts for each gene
DeSeq2 was used to calculate differential expression
Assembly: GRCh38
Supplementary files format and content: tab-delimited text files contain the gene expression counts for each sample or DeSeq2 dataset

source_name: MSC

extract_protocol:
RNA was isolated using miRNeasy Micro kit (Qiagen, Gaithersburg, MD, USA) and treated with RNase-Free DNase Set (Qiagen, Gaithersburg, MD, USA).
TruSeq RNA Library Prep Kit v2
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Untransduced and EcCAR-MSCs were produced on 3 biological replicates of normal donor MSCs and stimulated or not stimulated with soluble E-cadherin fc chimera and total RNA was isolated.

tag: donor

value: MSC

value: untransduced


title: NaraCOVID_TP6_CD8_020_TCR

value: female


title: VUMC0520


title: 4T1, ICI, 3

tag: tissue


title: NaraCOVID_TP6_CD8_018_TCR


title: MSC42_K129_Activated

value: transduced with EcCAR


title: NaraCOVID_TP6_CD8_019_TCR


title: 4T1, ICI, 1


title: 4T1, ICI, 2


title: VUMC0518


title: NaraCOVID_TP6_CD8_022_TCR


title: VUMC0517


title: VUMC0519


title: NaraCOVID_TP6_CD8_017_TCR


title: H3K9me3_ExE_Zfp57-WT_input

city: Shanghai

first: Liping

address: 1239 Siping Road

institute: Tongji University

data_processing:
Sequencing reads were first trimmed to remove adapters and then mapped to mouse genome (UCSC mm10) using Bowtie2 (v2.3.4.3) with the parameters: -t --no-mixed --no-discordant --no-unal(Langmead and Salzberg, 2012).
Mapped reads with high confidence were kept for further analysis using SAMtools (v1.7) with the parameters: -bf 0x2 -q 20. Duplicated reads were removed using sambamba (v0.6.8)(Tarasov et al., 2015).
Genome-wide signals were calculated using a 25-bp window and normalized to the uniquely mapped fragments using ‘bamCoverage’ from deepTools (v2.5.7)(Ramirez et al., 2014).
Promoters were [-2k, 0.5k] relative intervals of TSSs. 5’ most TSS selected if multiple TSSs found in one gene. Peaks were identified by macs2 (v2.1.1)(Zhang et al., 2008) with the parameters: -q 0.05 --keep-dup 1 and peaks with signal FPKM > 15 were finally reserved.
The programs ‘computeMatrix’ and ‘multiBigwigSummary’ of deepTools (v2.5.7) were used to compute the CUT&RUN-seq and ChIP-seq signals over the region of interest. Genomic features annotation and prediction of transcription factor motifs were performed using Homer (4.11.1)(Heinz et al., 2010).
Assembly: mm10
Supplementary files format and content: TF binding signal (FPKM) bigwig file
Library strategy: CUT&RUN

source_name: extraembryonic lineage

extract_protocol:
Pre-implantation embryos were obtained by in vitro fertilization (IVF). MII-stage oocytes were collected from 7-week-old superovulated C57BL/6N female mice by intraperitoneally injecting with 6 IU of pregnant mare’s serum gonadotropin (PMSG) and 6 IU of human chorionic gonadotrophin (hCG) (San-Sheng Pharmaceutical). MII oocytes were then transferred into G-IVF PLUS medium (Vitrolife) and inseminated with activated spermatozoa obtained from the caudal epididymides of adult PWK/PhJ male mice. The zona pellucida of each MII oocyte was punched to make a small hole using a Piezo-drill micromanipulator, and the spermatozoa capacitation was attained by 20-30 minutes incubation in G-IVF PLUS medium. At 4 hours post-insemination (hpi), fertilized oocytes were transferred and cultured in G-1 PLUS medium (Vitrolife) at 37°C under 5% CO2. The morula and blastocyst stage IVF embryos were harvested at 78 and 96 hpi, respectively. To obtain post-implantation embryos, female mice at 6.5 days post-coitum (dpc) were executed and the uteri were dissected and transferred into Dulbecco's phosphate-buffered saline (DPBS). Next, each decidua was carefully freed from the uterine muscle layers using properly sharpened forceps. 6.5 dpc embryos were carefully separated from deciduas. Reichart's membrane and the ectoplacental cone were also removed from the embryos.
uliCUT&RUN was conducted following the published protocols(Patty and Hainer, 2021; Skene et al., 2018) with a few modifications. Briefly, all isolated fresh blastomeres or cells were washed three times with CUT&RUN wash buffer to avoid possible contamination and then transferred into a 1.5mL low-binding PCR tube (Eppendorf) containing 600µL of the wash buffer. The samples were then bound to Concanavalin A-coated magnetic beads (Bangs Laboratoris), which had been activated and resuspended in CUT&RUN binding buffer. After cell immobilization, bead-bound samples were successively incubated with appropriate amount of primary antibodies against protein-of-interest (TFAP2C antibody: Santa Cruz, sc-12762; CDX2 antibody: BioGenex, MU392A-5UC; NANOG antibody: Cosmo Bio, RCAB002P-F; SOX2 antibody: Active Motif, 39843) in 250µL of CUT&RUN antibody buffer and a secondary antibody if necessary (rabbit anti-mouse IgG: Abcam, ab46540) in 250µL of CUT&RUN digitonin buffer for overnight at 4°C. Cell membranes were permeabilized with 0.01% digitonin to allow the specific antibody to find its target. After unbound antibodies were washed away, 700 ng/mL protein A-MNase (pA-MN; a gift from Steven Henikoff lab or purchased from Vazyme Biotech) was added and incubated for 1 hour at 4°C. After washing, CaCl2 was added to a final concentration of 2 mM to activate pA-MN, and the digestion reaction was carried out for 30 min at 0°C and then stopped by adding 150μL 2× CUT&RUN stop buffer. The protein–DNA complex fragments were then released by 20-min incubation at 37°C. After transferring the supernatant to a new tube, 3µL of 10% SDS and 2.5µL of Proteinase K (Thermo) were added and incubated for 30 min at 55°C. DNA was then precipitated by phenol/chloroform/isoamylalcohol followed by ethanol precipitation with glycogen and then dissolved in nuclease-free water. Sequencing libraries were prepared using KAPA Hyper Prep Kit (KAPA biosystems) following the manufacturer’s instructions with slight modifications. Briefly, end repair was conducted for 30 min at 20°C followed by end-repair/A-tailing for 30 min at 50°C. After adaptor ligation for 30 min at 20°C, the DNA fragments were purified by 1.4× volume of AMPure beads (Beckman Coulter) followed by 18 cycles of PCR amplification with 2× KAPA HiFi HotStart Ready Mix. The final libraries were cleaned up with 1.2× volume of AMPure beads, and all CUT&RUN libraries were sequenced on a NovaSeq6000 (Illumina) platform at Novogene Co., Ltd. with paired-ended 150-bp reads.

value: extraembryonic lineage

tag: antibody

value: wild type


title: MSC17_UTD_Activated

tag: treatment


title: H3K9me3_ExE_Zfp57-WT


title: MSC17_UTD_Baseline

tag: cell type


title: NaraCOVID_TP6_CD8_016_TCR

extract_protocol:
PBMCs were isolated from whole blood with BD vacutainer CPT tubes (BD), resuspended in CELLBANKER 1 (Takara), and stored in -80°C.
CD4+ and CD8+ T cells were magnetically isolated from PBMC. PBMC were thawed by warming frozen cryovials in a 37°C water bath. Cells were stained for 20 minutes at 4°C with 1st antibody mix (Antibody Mix 1 in Supplementary Table S1). After washing with MACS buffer [0.5% BSA (Nacalai), 2 mmol/L EDTA (NIPPON GENE) in PBS (Nacalai)], Cells were stained for 20 minutes at 4°C with 2nd antibody mix; BD IMag anti-PE Magnetic Particles-DM (Clone E31-1459, BD) and anti-APC-Biotin (clone APC003, BioLegend). Then, the cell suspension was diluted with MACS buffer and set on a magnet plate (DynaMag96SideSkirted, Veritas). The positive fraction was collected as CD4+ T cells. The negative fraction was stained with BD IMag Streptavidin Particles Plus-DM (BD) for 20 minutes at 4°C. Then, the cell suspension was diluted with MACS buffer and set on a magnet plate. The positive fraction was collected as CD8+ T cells and B cells. The counts of CD4+ T cells, CD8+ T cells, and B cells were determined using Flow-Count fluorospheres (Beckman) and a Cytoflex flow cytometer (Beckman) and FlowJo software (version 10.8.1; BD Biosciences).
Bulk TCR sequencing library was prepared according to the previous report (Aoki et al., 2023).  In brief, mRNA in T-cell lysates were captured by incubating with 10 μL of Dynabeads M270 streptavidin (Thermo Fisher Scientific) bound to 2.5 pmol of BioEcoP-dT25-adapter primers for 30 minutes at room temperature under gentle rotation. mRNA-trapped beads were suspended in 10 µL of RT mix and incubated for 60 min at 42ºC to perform reverse transcription (RT) and template-switching. The composition of the RT mix was as follows; 1× First Strand buffer (Thermo Fisher Scientific), 1 mM dNTP (Roche), 2.5 mM DTT (Thermo Fisher Scientific), 1 M betaine (Sigma-Aldrich), 9 mM MgCl2 (NIPPON GENE), 1 U/µL RNaseIn Plus RNase Inhibitor (Promega), 10 U/µL Superscript II (Thermo Fisher Scientific), and 1 µM of i5-TSO. To amplify the TCR cDNA containing complementarity determining region 3 (CDR3), nested PCR of the TCR locus was performed using KAPA Hifi Hotstart ReadyMix (KAPA Biosystems), followed by the purification of PCR product using an AM Pure XP kit (Beckman Coulter) at a 0.7:1 beads-to-sample ratio. The third PCR was performed to amplify TCR libraries and add adaptor sequences for the next-generation sequencer. The third-PCR products were purified as second PCR. Final TCR libraries, whose lengths were about 600 base pairs, were quantified using a KAPA SYBR Fast qPCR Kit (KAPA Biosystems), and size distribution was analyzed by Microchip Electrophoresis System MultiNA (Shimadzu). TCR libraries were sequenced using an Illumina Novaseq 6000 S4 flowcell (67 bp read 1 and 140 bp read 2) (Illumina) using NovaSeq 6000 S4 Reagent Kit v1.5 (200 cycles, Illumina). Only read2 contained the sequence regarding the definition of T-cell clones. Oligonucleotides and reaction conditions for library preparation were summarized in Table S2 and S3, respectively.

value: Three shots of Pfizer-BNT162b2 vaccine

tag: age


title: VUMC0516

extract_protocol: Qiagen’s miRNeasy Plasma Kit manufacturer's protocol


title: NaraCOVID_TP6_CD8_015_TCR

tag: Sex

value: 6 month after receiving the second shot


title: H3K9me3_Epi_Zfp57-WT_input

source_name: epiblast

value: epiblast


title: MSC42_K129_Baseline


title: MSC17_K129_Baseline


title: MSC42_UTD_Activated


title: NaraCOVID_TP6_CD8_014_TCR

tag: date


title: VUMC0521

tag: study site (plco=1 swmhs=2 sccs=3 mec=4)


title: VUMC0515

tag: case control status (control=0 case=1)


title: MSC17_K129_Activated


title: H3K9me3_ExE_Zfp57-KO_input


title: MSC16_UTD_Baseline


title: VUMC0513

tag: sets


title: NaraCOVID_TP6_CD8_012_TCR


title: NaraCOVID_TP6_CD8_011_TCR


title: H3K9me3_ExE_Zfp57-KO


title: H3K9me3_Epi_Zfp57-KO_input


title: MSC16_UTD_Activated


title: VUMC0511

tag: sex (female=0 male=1)


title: VUMC0514

tag: age at diagnosis


title: VUMC0512

tag: race (white=1 black=2 other=3)


title: MSC16_K129_Activated


title: MSC16_K129_Baseline


title: COPD 1, day 5

city: Biberach

first: Meshal

last: Ansari

address: Birkendorfer Straße 65

country: Germany

institute: Boehringer Ingelheim

description:
Drop-seq
muc14439

data_processing:
The Drop-Seq core computational pipeline was used for processing next generation sequencing reads of the scRNA-seq data, as previously described (Pubmed ID: 26000488). After quality control and pre-processing, the final data set encompasses 64062 cells and 32989 genes. The cells were divided into time series (basal_cells_nonCLD_trajectory.h5ad, basal_cells_COPD_trajectory.h5ad), starting point (basal_cells_nonCLD_COPD_day0.h5ad) and end point objects (basal_cells_nonCLD_COPD_day28.h5ad) for further analysis.
Assembly: hg38
Supplementary files format and content: Tab-separated values files and h5ad files

source_name: Airway liquid interface

growth_protocol: Isolated pHBECs seeded on CollagenIV-coated transwell inserts were cultured in PneumaCultTM - Ex Plus Media (STEMCELL Technologies, REF:05041) until confluent before air-lift (Air-Liquid-Interface day 0 (ALId0); basal medium changed to supplemented PneumaCultTM (100x PneumaCultTM ALI Media, STEMCELL Technologies, REF:05006). Basal medium changed 3x/week, apical side washed 1x/week (HBSS Ca2+/Mg2+, Gibco Life Technologies, REF:14065-049) (Schamberger et al, 2014).

extract_protocol: Dropseq experiments were performed according to the original Dropseq protocol (Macosko et al., 2015;). Single cell suspensions were processed through Drop-Seq to generate single-cell cDNA libraries attached to microbeads. Microbeads were counted, and amplified by PCR, and the 3' end of the cDNA prepared for sequencing. For each sample, 1 ng of pre-amplified cDNA from an estimated 2000 cells was tagmented by Nextera XT (Illumina) with a custom P5 primer (Integrated DNA Technologies). Single cell libraries were sequenced in a 100 bp paired-end run on the Illumina HiSeq4000 using 0.2 nM denatured sample and 5% PhiX spike-in. For priming of read 1, 0.5 µM Read1CustSeqB was used.

value: Airway epithelium

tag: patient id

value: Airway liquid interface

tag: time point

value: day 5

tag: health state

value: COPD


title: TFAP2C_ExE_dKO_rep2

tag: cell line

value: Prdm2/Suv39h2 dKO


title: COPD 2, day 28

description:
Drop-seq
muc14438

tag: tissue

value: day 28


title: H3K9me3_Epi_Zfp57-WT

tag: antibody


title: NaraCOVID_TP6_CD8_010_TCR

tag: treatment

value: CD8+ T cells and CD19+ B cells

value: male


title: NaraCOVID_TP6_CD8_007_TCR

tag: cell type


title: NaraCOVID_TP6_CD8_008_TCR

tag: age


title: NaraCOVID_TP6_CD8_013_TCR

tag: Sex


title: VUMC0510


title: NaraCOVID_TP6_CD8_006_TCR

tag: date


title: VUMC0509

tag: study site (plco=1 swmhs=2 sccs=3 mec=4)


title: COPD 2, day 21

description:
Drop-seq
muc14437

value: day 21


title: TFAP2C_ExE_dKO_rep1


title: H3K9me3_Epi_Zfp57-KO


title: NaraCOVID_TP8_CD4_005_TCR

value: CD4+ T cells

value: 1 week after receiving the third shot


title: NaraCOVID_TP6_CD8_005_TCR


title: VUMC0508

tag: case control status (control=0 case=1)


title: H3K9me3_ExE_WT_input


title: VUMC0504

tag: sets


title: NaraCOVID_TP6_CD8_004_TCR


title: TFAP2C_ExE_WT


title: COPD 2, day 7

description:
Drop-seq
muc14435

value: day 7


title: H3K9me3_ExE_dKO_input

value: extraembryonic lineage


title: COPD 2, day 3

description:
Drop-seq
muc14433

value: day 3


title: COPD 2, day 0

description:
Drop-seq
muc14432

value: day 0


title: COPD 2, day 5

description:
Drop-seq
muc14434


title: VUMC0503

tag: sex (female=0 male=1)


title: H3K9me3_ExE_dKO_rep2


title: VUMC0507

tag: age at diagnosis


title: NaraCOVID_TP6_CD8_003_TCR

value: Three shots of Pfizer-BNT162b2 vaccine

value: 6 month after receiving the second shot


title: VUMC0500

tag: race (white=1 black=2 other=3)


title: H3K9me3_ExE_WT_rep1

value: wild type


title: VUMC0502


title: NaraCOVID_TP6_CD8_002_TCR


title: NaraCOVID_TP6_CD8_001_TCR


title: COPD 1, day 14

description:
Drop-seq
muc13949

value: Airway epithelium

tag: patient id

value: Airway liquid interface

value: day 14


title: VUMC0501


title: NaraCOVID_TP6_CD4_039_TCR


title: NaraCOVID_TP6_CD4_041_TCR


title: NaraCOVID_TP6_CD4_037_TCR

value: female


title: NaraCOVID_TP6_CD4_038_TCR


title: COPD 1, day 7

description:
Drop-seq
muc13948

tag: time point

value: COPD


title: H3K9me3_ExE_dKO_rep1

value: Prdm2/Suv39h2 dKO


title: Tat+METH 4, Female

city: Riverside

first: Marcus

last: Kaul

state: California

address: 900 University Ave, SOM RB Room 311

department: Biomedical Sciences

institute: University of California, Riverside

description: instrument model: Illumina NovaSeq X Plus

data_processing:
FASTQ files were processed using the reference cDNA sequence for Mus musculus (GRCm39), from Ensembl database (release 110). Kallisto [PMID: 27043002] was utilized as the software to generate an index facilitating transcript quantification.
Assembly: GRCm39
Supplementary files format and content: Excel File (.xlsx) containing: target_id, lengths of transcripts (length), effective lengths (eff_length), estimated counts (est_counts), transcripts per million (tpm).

source_name: Hippocampus

extract_protocol:
RNA was extracted from the brain tissue (snap frozen hippocampus) using the RNeasy mini plus kit (Qiagen). RNA samples were first bioanalyzed using an Agilent Tapestation through the University of California Riverside (UCR) Genomics Core. Samples with RIN > 7.0 were sent to the University of California San Diego (UCSD) genomics core (IGM), for RNAseq (rRNA depletion, PE100, 25M).
RNA libraries for RNA-seq were prepared by the UCSD genomics core (IGM): Illumina Total RNA Prep (Ribodepletion).

treatment_protocol: The study employed transgenic mice that express a tetracycline-inducible viral protein Tat in the brain (iTat-tg mice - GFAP-rtTA + TRE-Tat). The mouse cohort included rtTA-positive TRE-Tat-negative control animals, which cannot express Tat upon Dox injection, and comprised each 50 % females and males. The transgenic animals were subjected at 4 months of age to a 12-week METH regimen ('chronic METH regimen'): Starting week 1 at 0.5 mg/kg s.c., 1 x day, step-wise increase by 0.5 mg/kg with each injection over 5 days (Mon–Fri), followed by 11 weeks of 1 x 2.5 mg/kg/day (Mon – Fri = 20+ days per month). During week 4, the mice received Doxycyclin (Dox, 100 mg/kg, i.p., for 7 days) for induction of Tat expression.  Four months after the last METH exposure, at 11-12 months of age, all iTat-tg mice underwent behavioral testing before the animals were euthanized and brain tissue collected for analysis of gene expression at 12-13 months of age.

value: Hippocampus

tag: gender

value: Female

tag: genotype

value: GFAP-rtTA+/pTRE-Tat+

value: Doxycycline+METH


title: COPD 1, day 28

description:
Drop-seq
muc13951

tag: tissue

value: day 28

tag: health state


title: VUMC0496


title: VUMC0499


title: NaraCOVID_TP6_CD4_036_TCR

data_processing:
Data processing of Bulk TCRseq was performed as described in Aoki et al. (Aoki et al., 2021).
Adapter trimming and quality filtering of sequencing data were performed using Cutadapt-3.2 and PRINSEQ-0.20.4. Sequencing data were processed by MiXCR-3.0.5. In MiXCR, filtered reads were aligned to reference mouse TCR V/D/J sequences registered in the international ImMunoGeneTics (IMGT) information system. The Variable (V) and Joining (J) segments of TCRs were represented in IMGT gene nomenclature. T-cell clones were determined as TCR reads with the same TCR V segment, J segment, and CDR3 nucleotide sequence.
Inter-sample contamination within samples using same UDIs were excluded by "Decontaminate" function of vdjtools with the parameter "-r 10".
For bulk TCRseq data, we regarded the CD4+-CD8+ overlapping clones in the same participant as contamination during T-cell isolation from PBMC because CD4+ and CD8+ T-cell repertoire are mutually exclusive (Aoki 2021). Thus, these overlapping clones were categorized into either CD4+ or CD8+ according to their dominance in CD4+ and CD8+ repertoire using VDJtools-1.2.1. The sequencing reads of bulk TCR-seq samples were normalized to 100,000 reads by the “DownSample” command of VDJtools-1.2.1. To preserve the minor clonotypes, we performed the normalization ten times and pooled these down-sampled repertoire data for each sample, by “JoinSamples” command of VDJtools-1.2.1.
Assembly: IMGT reference sequences of human TRA and TRB
Supplementary files format and content: Text files in VDJtools format include read count and frequency, complementarity determining region 3 (CDR3) nucleotide and amino acid sequences, Variable(V)/ Diversity(D)/ Joining(J) segment names, and V, D and J segment boundaries within CDR3 nucleotide sequence of individual clones
Library strategy: Bulk TCR-seq

tag: treatment

value: CD4+ T cells

value: male


title: VUMC0495

data_processing: miRNAs were normalized in the following steps: 1) background correction to remove non-specific binding. The background level was calculated as an aggregate summary of seven negative control probes with no target transcript. Mean of the calculated background level for each sample was then subtracted from the adjusted probe value to remove background noise. 2) Normalization of miRNA counts using mean counts of three spike-in synthetic control miRNAs (ath-miR159a, cel-miR-248, and osa-miR414) to control for variances on the starting material and the miRNA purification efficiency. 3) Sample content normalization using geometric mean counts of the top 50 miRNAs expressed excluding hsa-miR-320e, hsa-miR-16-5p and hsa-miR-451a because these miRNAs may be related to red blood cell contamination or hemolysis. After these steps, the expression level of each miRNA was calculated.


title: NaraCOVID_TP6_CD4_040_TCR

tag: cell type


title: VUMC0498


title: H3K9me3_ExE_WT_rep2

tag: cell line

value: extraembryonic lineage

tag: age

tag: antibody


title: COPD 1, day 21

description:
Drop-seq
muc13950

value: day 21


title: VUMC0497

tag: study site (plco=1 swmhs=2 sccs=3 mec=4)


title: NaraCOVID_TP6_CD4_033_TCR

value: Three shots of Pfizer-BNT162b2 vaccine

tag: Sex

tag: date

value: 6 month after receiving the second shot


title: Tat+METH 6, Female


title: Donor 2, day 21

description:
Drop-seq
muc7512

value: Airway epithelium

value: Donor


title: NaraCOVID_TP6_CD4_032_TCR


title: Donor 1, day 28

description:
Drop-seq
muc7510

value: Airway liquid interface


title: Tat+METH 2, Male

value: Male


title: NaraCOVID_TP6_CD4_031_TCR


title: COPD 1, day 3

description:
Drop-seq
muc13947

value: day 3

value: COPD


title: Tat+METH 3, Male

value: Hippocampus


title: NaraCOVID_TP6_CD4_030_TCR

value: female


title: Donor 1, day 21

description:
Drop-seq
muc7509


title: VUMC0491

tag: case control status (control=0 case=1)

tag: sets

tag: sex (female=0 male=1)


title: VUMC0490

tag: age at diagnosis

tag: race (white=1 black=2 other=3)


title: VUMC0488


title: Donor 2, day 3 (2)

description:
Drop-seq
muc7508

tag: patient id

tag: time point


title: COPD 2, day 14

description:
Drop-seq
muc14436

tag: tissue

value: day 14


title: VUMC0494


title: VUMC0493


title: Donor 1, day 14

description:
Drop-seq
muc7507

tag: health state


title: NaraCOVID_TP6_CD4_027_TCR

tag: treatment


title: METH 1, Male

tag: gender

tag: genotype

value: GFAP-rtTA+

value: Doxycycline+METH


title: Donor 2, day 28

description:
Drop-seq
muc7513

tag: cell type

value: day 28


title: NaraCOVID_TP6_CD4_026_TCR

value: CD4+ T cells

tag: age

value: male


title: Tat 6, Female

value: Female

value: GFAP-rtTA+/pTRE-Tat+

value: Doxycycline+Saline


title: NaraCOVID_TP6_CD4_035_TCR

value: Three shots of Pfizer-BNT162b2 vaccine

value: 6 month after receiving the second shot


title: Donor 1, day 7

description:
Drop-seq
muc7503

value: Airway epithelium

value: Airway liquid interface

value: day 7

value: Donor


title: Tat+METH 5, Female

value: Hippocampus


title: Donor 2, day 0

description:
Drop-seq
muc7501

value: day 0


title: NaraCOVID_TP6_CD4_025_TCR


title: NaraCOVID_TP6_CD4_024_TCR

value: female


title: Tat+METH 1, Male

value: Male


title: Donor 1, day 5

description:
Drop-seq
muc7499

value: day 5


title: VUMC0492

tag: study site (plco=1 swmhs=2 sccs=3 mec=4)


title: COPD 1, day 0

description:
Drop-seq
muc13946

value: COPD


title: METH 6, Female

value: GFAP-rtTA+

value: Doxycycline+METH


title: Donor 2, day 14

description:
Drop-seq
muc7511

value: day 14


title: VUMC0485


title: METH 5, Female

value: Female


title: NaraCOVID_TP6_CD4_023_TCR

value: CD4+ T cells

tag: Sex

value: male


title: Tat 4, Female

value: GFAP-rtTA+/pTRE-Tat+

value: Doxycycline+Saline


title: NaraCOVID_TP6_CD4_022_TCR

value: Three shots of Pfizer-BNT162b2 vaccine

tag: date

value: 6 month after receiving the second shot


title: Donor 1, day 3

description:
Drop-seq
muc7495

value: Airway epithelium

value: Airway liquid interface

value: day 3

value: Donor


title: METH 3, Male

value: Hippocampus

value: Male


title: METH 4, Female


title: NaraCOVID_TP6_CD4_029_TCR

value: female


title: VUMC0482

tag: case control status (control=0 case=1)


title: NaraCOVID_TP6_CD4_021_TCR


title: VUMC0487

tag: sets


title: NaraCOVID_TP6_CD4_020_TCR


title: VUMC0486

tag: sex (female=0 male=1)


title: VUMC0489

tag: age at diagnosis


title: METH 2, Male

value: GFAP-rtTA+


title: Donor 2, day 7

description:
Drop-seq
muc7489

value: day 7


title: Ctl 6, Female


title: Donor 2, day 5

description:
Drop-seq
muc7488

value: day 5


title: Tat 5, Female

value: Female


title: Ctl 5, Female


title: Donor 2, day 3

description:
Drop-seq
muc7497


title: VUMC0484

tag: race (white=1 black=2 other=3)


title: NaraCOVID_TP6_CD4_018_TCR

value: CD4+ T cells


title: VUMC0479


title: Ctl 4, Female


title: Tat 2, Male


title: PDX anti-OPN-treated 5

city: Chicago

first: Ronen

last: Sumagin

state: IL

address: 303 E Chicago Ave

department: Pathology

institute: Northwestern University

data_processing:
Read alignment with STAR. Raw reads were de-multiplexed into FASTQ format using bcl2fastq software (Illumina) The FASTQ files were quality checked for called bases of each read. Pre-processing of raw data was performed by Trimmomatic option in Chipster where reads of quality below 20 were eliminated. The Illumina adapters were removed from the 3’ ends by cutadapt command-line program that operated under two adapter trimming algorithms 71.
Trimmed reads were aligned to the Mus musculus genome mm10 and human genome hg38 using STAR RNA-seq aligner (version STAR_2.5.1 72). Of note, pre-processing of the reads was performed in conjunction with the mm10 mouse gene annotation file obtained from Ensembl (http://useast.ensembl.org/index. html). Reads with read lengths <35 were discarded. The read counts for individual genes were generated by htseq-count script derived from the HTseq framework v.0.6.1p1 that counts and calculates the read overlaps of genes within the mouse and human genome.
To identify DEGs, normalization for sequencing depth followed by variance-stabilizing transformation as well as differential expression calculations were performed via DESeq2 R Bioconductor package 1.16.1, which employed the Wald test to shrink Log-Fold-Changes (LFC) and their standard errors.  The cut-off for determining significantly differentially expressed genes was determined with a stringency level where an FDR-adjusted p<0.05 and |log2FC| <1 using the Benjamini-Hochberg method. In addition, genes with normalized counts < 100 were removed and not considered for future analysis. To execute principal-component analysis (PCA) and heatmap visualization, read counts were transformed to log2(count per million) and corrected for batch effects using limma package. In these analyses, PCA plots were made using the pca() function and hierarchical clustering with the dist() function.
Assembly: hg18 (PDX data) and mm3 (neutrophil AOM/DSS data)
Supplementary files format and content: count matrices
Supplementary files format and content: GSEA, GO, and DEG outputs

source_name: PDX tumor tissues

extract_protocol:
For sorting of PMNs or other myeloid cells, we utilized Lyz2EGFP reporter mice with additional staining for Ly6G, Ly6C, CD11b, CD45, and CD64. FACS was performed on a 3-laser sorter FACS Melody and a 6-color laser sorter FACS Aria at the Northwestern Flow Cytometry core. EpCAM+ tumor or epithelial cells as well as Ly6Gneg CD64hi¬¬ macrophages were gated out to avoid RNA contamination from other cell types. CD45+ CD11b+ Ly6Cint Lyz2EGFP Ly6Ghi/int were defined as PMNs/ TANs and sorted directly into RNA Lysis Buffer (RLT plus buffer, Qiagen), supplemented with excess amount of RNaseOUT (10 U/μL, Invitrogen). RNA from PMNs was immediately extracted using the RNeasy Mini Plus Kit (Qiagen) and subjected to RNA integrity (RIN) analysis. For quality control of PMN cell sorting, additional replicates were performed with identical gating strategy and subjected to either more rigorous flow cytometric analyses using additional PMN markers, or using Cytospin and immunofluorescence for imaging. Enrichment of PMN gene sets was performed following RNA sequencing to confirm the transcriptomic identity of the sorted cells as well as validity of downstream bioinformatic analyses
The NEBNext Ultra II RNA Library Prep Kit for Illumina (New England Biolabs, Ipswich, MA) was used for library preparation per manufacturer’s protocol with the addition of NEBNext rRNA depletion option. Following fragmentation to 200-500 base pairs (bp) by Bioruptor Sonicator (Diagenode), reverse transcription (RT) was performed to convert RNA to cDNA, followed by end-repair, adaptor ligation, and PCR amplification of cDNA libraries per manufacturer’s instruction (New England Biolabs, Ipswich, MA). Prior to sequencing, prepared libraries were assessed for fragment sizing on a Bioanalyzer using High Sensitivity DNA chip and re-quantified with Qubit. Library sequencing was performed on an Illumina HiSeq 4000 NGS System (Illumina, San Diego, CA) to generate 50 bp reads with 300 million reads and 30x read depth.

treatment_protocol: To initiate carcinogenesis, mice were given a single dose of azoxymethane (AOM) by intraperitoneal (i.p.) injection (12.5 mg/kg body weight). A week after, mice were treated with up to 5 cycles of DSS 3% in drinking water (1 cycle = 5 days of DSS + 14 days recovery). Immediately after sacrifice, peripheral blood was drawn from the vena cava; bone marrows from both hind-legs (femur and tibia) were collected; spleen and tumor-bearing colons were extracted for phenotypic evaluations and downstream molecular/functional assays. For PDX experiments, a total of 15 PDX-bearing mice were purchased from Jackson Laboratory (Bar Harbor). PDXs were established by the Jackson Labs as follows; metastatic stage IV colon adenocarcinoma with BRAFV600E and PI3KCAG1049R mutations derived from a 30-year-old male patient (Colon Cancer, PDX Model TM00170) were subcutaneously grafted (2x106 tumor cells/mouse in 40μL PBS) into flanks of NOD.Cg-Prkdcscid Il2rgtm-1Wjl/SzJ NSG mice aged 7-8 weeks (strain 005557 NSG). Through the duration of the experiments, tumor diameters were measured bi-weekly by calipers. Once tumors reached 500mm3, mice were randomly divided into 3 treatment groups (Control, Anti-OPN, and NSC405020. Anti-OPN (clone 103D6) was administered at 5.0mg/kg/mice, daily, i.p.) while NSC 405020 was administered 1.0mg/kg/mice (3 times per week, i.p.) for 21 days (endpoint).

value: PDX tumor tissues

value: PDX primary tumor cells

value: BRAFV600E/PI3KCAG1049R mutations

value: anti-OPN (103D6, 5.0 mg/kg)


title: VUMC0483


title: NaraCOVID_TP6_CD4_019_TCR

city: Noda

first: Hiroyasu

address: 2669 Yamazaki

institute: Tokyo University of Science

source_name: Peripheral blood T cells

value: Three shots of Pfizer-BNT162b2 vaccine

value: 6 month after receiving the second shot


title: VUMC0481

city: Nashville

first: Cong

address: 2525 West End Ave

institute: Vanderbilt University

source_name: plasma


title: NaraCOVID_TP6_CD4_016_TCR


title: Tat 1, Male

value: GFAP-rtTA+/pTRE-Tat+

value: Doxycycline+Saline


title: Donor 1, day 0

description:
Drop-seq
muc7490

value: Airway epithelium

tag: patient id

value: Airway liquid interface

value: day 0

value: Donor


title: VUMC0480


title: VUMC0476


title: NaraCOVID_TP6_CD4_015_TCR

value: male


title: NaraCOVID_TP6_CD4_017_TCR


title: PDX anti-OPN-treated 4


title: Ctl 3, Male

value: Hippocampus

value: Male

value: GFAP-rtTA+


title: VUMC0474


title: Ctl 2, Male


title: VUMC0477


title: Pia_mmu, scRNAseq

city: Bentley

first: Paul

middle: James

last: Whatmore

state: Western Australia

address: Kent St

department: eResearch

country: Australia

institute: Curtin University

description: 10x Genomics

data_processing:
Demultiplexing, barcoding processing, gene counting and aggregation were completed using Cell Ranger software v7.0.0 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/using/tutorial_ov)
Downstream analysis (see below) was completed in R v4.0.5
QC, dimensionality reduction, clustering, and differential expression analysis was completed using the R package Seurat (https://satijalab.org/seurat/)
Functional annotation of GE genes to KEGG pathways and GO terms was completed using the R package clusterProfiler (https://bioconductor.org/packages/release/bioc/html/clusterProfiler.html)
Assembly: mm10, hg38
Supplementary files format and content: Quality-filtered barcodes (cell IDs), features (genes) and count matrix, generated by Cell Ranger.

source_name: Pia

extract_protocol:
Mouse samples: Immune cells (CD45+ cells) were isolated from the choroid and pia mater of n=20 C57Bl/6J female mice (7 weeks of age) by FACS. Freshly dissected choroid and pia mater were pooled to obtain sufficient numbers of cells. The dissected tissues were transferred to 50 mL tubes containing dissection buffer (1x HBSS, no calcium, no magnesium, no phenol red; 5 mM glucose; 15 mM HEPES) and all processing was performed on ice. The tissues were passed through a 70 μm nylon cell strainer (Falcon 352350) using the plunger of a 5 mL syringe to obtain single cell suspensions, and then pelleted by centrifugation at 400 g for 5 min at 4 °C.  Pia mater-enriched tissue was subsequently resuspended in 30% (v/v) Percoll (GE Healthcare 17089102) in 1 x PBS and centrifuged at 700 g for 10 min at 4 °C without a brake being applied. The myelinated (i.e., hydrophobic) layer formed at the top of the tube was discarded prior to resuspending the cell pellet. The number of viable cells was determined by exclusion of trypan blue stain as viewed by microscopy, assisted by use of a hemocytometer. Cells were subsequently resuspended in FACS buffer (3 mM EDTA, 0.1% (w/v) BSA, 1 x PBS, 100 μg/mL DNase I) and stained with fluorescent-conjugated antibodies (rat anti- mouse CD45-BV605, BD Biosciences 563890) for 30 min on ice, then centrifuged and resuspended in FACS buffer. Immune cells (CD45+) were isolated from stained cell suspensions using a BD FACS Aria IIIu (100 µm nozzle) at QIMR Berghofer Flow Cytometry Facility. Dead cells were identified and excluded from analysis by staining with propidium iodide (PI, 1 μg/ mL, BD Biosciences, Franklin Lakes, NJ, 556463).  Human samples: Fresh (unfixed) human eye cups from a 45-year-old male donor with no history of eye disease or diabetes was obtained within 17 hours post-mortem from the Queensland Eye Bank. The vitreous and retinas were removed from the eye cups and choroid-RPE was subsequently dissected from the sclera. Choroid-RPE tissue was transferred into a 12-well plate and subjected to enzymatic digestion with collagenase type II (200 U/mL in HBSS (calcium, magnesium, no phenol red)) for 60 minutes at 37 °C, with tissue triturated every 15 minutes. After 60 minutes, choroid tissue was triturated with a pipette and then passed through a 70 µm cell strainer to generate a single cell suspension. The cell suspension was pelleted by centrifugation at 400 x g for 5 mins, resuspended in freezing media (DMEM/F12 containing 10% (v/v) DMSO and 10% (w/v) bovine serum albumin) and transferred to a Cool Cell freezing chamber at -80 °C for controlled freezing. Cells then were transferred to liquid nitrogen storage. For scRNA-seq studies, cells were thawed and pre-warmed DMEM/F12 containing 10% (w/v) BSA was added in a dropwise manner. The viability and yield of thawed cells were determined by trypan blue staining using a haemocytometer. Human choroidal immune cells were isolated from thawed cell suspensions by FACS. Briefly, thawed cell suspensions were incubated with 2.5 µg anti-human Fc block (BD Pharmingen 564220) for 15 mins, then centrifuged at 400 x g for 5 mins. The pellet was resuspended in FACS buffer containing BV421 Mouse Anti-Human CD45 antibody (BD Biosciences 563879) and incubated on ice for 30 mins. Cells were centrifuged at 400 x g for 5 mins then resuspended in FACS buffer for sorting. Live (propidium iodide negative) immune cells (CD45+) were sorted using a BD FACS Aria IIIu with a 100 µm nozzle.
scRNA-seq libraries were prepared using a Chromium single cell reaction 3’ v3.1 kit, following the manufacturer's instructions.

value: Pia


title: PDX anti-OPN-treated 3

value: PDX tumor tissues


title: PDX anti-OPN-treated 2

value: PDX primary tumor cells


title: NaraCOVID_TP6_CD4_013_TCR

value: CD4+ T cells

value: female


title: Ctl 1, Male


title: PDX MMP14i-treated 5

value: BRAFV600E/PI3KCAG1049R mutations


title: VUMC0475


title: Tat 3, Male


title: PDX anti-OPN-treated 1

value: anti-OPN (103D6, 5.0 mg/kg)


title: NaraCOVID_TP6_CD4_014_TCR

value: Three shots of Pfizer-BNT162b2 vaccine

value: 6 month after receiving the second shot


title: PDX MMP14i-treated 3


title: Aged mouse HSCs, control

city: West Hollywood

first: AGCT

last: Core

state: CA

address: 8687 Melrose Ave, PDC B230

department: BMS

institute: Cedars Sinai Medical Center

data_processing:
Sequence reads were aligned and analyzed using the Cell Ranger Count pipeline (v7.0.1) (10X Genomics) to generate cell-associated gene expression matrices. Data was then normalized via Scran deconvolution and differential expression among samples was measured with DESeq 2 algorithm in Partek Flow
UMAP plots were generated in R v4.2.2 using the Seurat v4.2.1 package following dimensionality reduction using the first ten principal components while density plots were generated using ggplot2 v3.4.0 and the stat_density_2d function.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and metric files

source_name: Bone marrow CD34-KSL cells

extract_protocol:
RNA was extracted using the Chromium Single Cell 3’ V2 Reagents Kits
Single-cell RNA-Seq libraries were prepared as per the 10x Genomics Single Cell 3′ v3.1 Reagent Kit User Guide using the Chromium X (10x Genomics, Pleasanton, California). Fragment analyses of the cDNA and indexed libraries were performed on the 4200 TapeStation (Agilent Technologies, Santa Clara, California). Barcoded sequencing libraries were quantified by quantitative PCR using the Collibri Library Quantification Kit (Thermo Fisher Scientific, Waltham, MA). Libraries were sequenced on a NovaSeq 6000 (Illumina, San Diego, CA) with the sequencing parameters 28x10x10x90 and a sequencing depth of ~40,000 reads/cell.

treatment_protocol: BM CD34-KSL cells were sorted from mice after subcutaneous injection of 1 µg/g EGF or saline in 200 µL daily for 3 days

value: Bone marrow CD34-KSL cells

value: CD34- KSL

value: 200ul PBS for 3 days


title: VUMC0472


title: VUMC0470


title: VUMC0471


title: PDX MMP14i-treated 4


title: NaraCOVID_TP6_CD4_010_TCR

last: Aoki


title: NaraCOVID_TP6_CD4_008_TCR


title: Choroid_mmu, scRNAseq

description: 10x Genomics

source_name: Choroid

value: Choroid


title: VUMC0468

last: Wang


title: Young mouse HSCs, EGF treated

value: 1ug/g EGF for 3 days


title: VUMC0469


title: PDX MMP14i-treated 2


title: PDX MMP14i-treated 1


title: Young mouse HSCs, control


title: PDX vehicle-treated 4

value: IgG isotype


title: Aged mouse HSCs, EGF treated


title: NaraCOVID_TP6_CD4_007_TCR

value: male


title: VUMC0466


title: NaraCOVID_TP6_CD4_012_TCR


title: VUMC0467


title: VUMC0465


title: NaraCOVID_TP6_CD4_005_TCR


title: PDX vehicle-treated 2


title: NaraCOVID_TP6_CD4_011_TCR


title: VUMC0464


title: SOX2_ESC_rep1

source_name: ESC

value: ESC


title: SOX2_E6.5_Epi_rep2


title: PDX vehicle-treated 1


title: SOX2_E6.5_Epi_rep1


title: VUMC0463


title: CARD14WT_9hr_3

city: London

first: Tolar

last: Pavel

address: Gower St

country: United Kingdom

institute: UCL

data_processing:
The RSEM package (version 1.3.0) in conjunction with the STAR alignment algorithm (version 2.5.2)  was used for the mapping and subsequent gene-level counting of the sequenced reads with respect to Ensembl GRCm38-release-89 transcriptome
Differential expression analysis was performed with the DESeq2 package (version 1.10.1) within the R programming environment (version 3.3.2)
Assembly: GRCm38
Supplementary files format and content: Tab delimited text files containing TMM normalised data. Rows represent ensembl gene identifiers and columns represent individual samples.

source_name: CARD14WT-3XFLAG HaCaT-TR

growth_protocol: Cells are at least two doublings from liquid nitrogen thaw and never allowed to be in prolonged culture with high confluency

extract_protocol:
RNA was extracted with and RNEasy kit (Qiagen). A fraction of each sample was taken and assessed for RNA quality. mRNA was then purified using a poly-A KAPA mRNA Hyper Prep kit (Roche)
Samples were processed with the KAPA mRNA HyperPrep kit, according to manufacturer’s instructions, which includes: End repair and A-tailing, which produces end-repaired, 5′-phosphorylated, 3′-dA-tailed, dsDNA fragments. Adapter ligation, during which dsDNA adapters with 3′-dTMP overhangs are ligated to 3′-dA-tailed library fragments. Library amplification, which employs high-fidelity, low-bias PCR to amplify library fragments carrying appropriate adapter sequences on both ends.

treatment_protocol: Cells were induced with tetracycline as part of the Tet-on (Trex) system to cause gene induction of CARD14 WT or mutant

value: Skin cell line

value: keratinocyte

value: +Tetracycline for indicated timepoint

tag: environmental condition

value: 37C, 5% CO2 gas


title: NaraCOVID_TP6_CD4_003_TCR

value: CD4+ T cells


title: VUMC0461


title: VUMC0460


title: PDX vehicle-treated 3

tag: tissue

value: PDX tumor tissues

value: PDX primary tumor cells

value: BRAFV600E/PI3KCAG1049R mutations


title: CARD14WT_9hr_2


title: SOX2_TE_rep2

source_name: TE

value: TE


title: CARD14WT_9hr_1


title: NaraCOVID_TP6_CD4_006_TCR

value: Three shots of Pfizer-BNT162b2 vaccine

value: 6 month after receiving the second shot


title: SOX2_ICM_rep1

source_name: ICM

value: ICM


title: NaraCOVID_TP6_CD4_001_TCR


title: CARD14WT_6hr_3


title: SOX2_TE_rep1


title: VUMC0459


title: NANOG_ESC_rep2


title: NaraCOVID_TP5_CD8_041_TCR

value: CD8+ T cells and CD19+ B cells

value: 3 month after receiving the second shot


title: NaraCOVID_TP5_CD8_039_TCR


title: NANOG_ESC_rep1


title: NaraCOVID_TP5_CD8_040_TCR


title: VUMC0456


title: VUMC0457


title: NANOG_E6.5_Epi_rep3


title: NaraCOVID_TP6_CD4_002_TCR


title: CARD14WT_6hr_1


title: NaraCOVID_TP5_CD8_038_TCR


title: CARD14WT_3hr_3


title: NANOG_E6.5_Epi_rep2


title: VUMC0454


title: NANOG_E6.5_Epi_rep1


title: NaraCOVID_TP5_CD8_037_TCR

value: female


title: NaraCOVID_TP5_CD8_036_TCR

value: male


title: CARD14WT_3hr_2


title: CARD14WT_0hr_3


title: VUMC0453


title: CARD14WT_3hr_1


title: NANOG_ICM_rep2


title: NaraCOVID_TP5_CD8_035_TCR


title: VUMC0451


title: NaraCOVID_TP6_CD4_004_TCR


title: CARD14WT_0hr_2


title: NaraCOVID_TP5_CD8_032_TCR


title: CARD14WT_0hr_1


title: VUMC0450


title: SOX2_ESC_rep2

value: ESC


title: NaraCOVID_TP5_CD8_033_TCR


title: NANOG_ICM_rep1


title: VUMC0455


title: NANOG_Morula_rep2

source_name: Morula

value: Morula


title: VUMC0449


title: NANOG_Morula_rep1


title: NaraCOVID_TP5_CD8_031_TCR


title: VUMC0448


title: CARD14E138A_9hr_3

source_name: CARD14E138A-3xFLAG HaCaT-TR

value: Skin cell line

value: keratinocyte


title: NaraCOVID_TP5_CD8_029_TCR


title: CARD14E138A_9hr_1

value: +Tetracycline for indicated timepoint

value: 37C, 5% CO2 gas


title: CARD14E138A_9hr_2


title: NaraCOVID_TP5_CD8_027_TCR


title: VUMC0447


title: CDX2_E6.5_ExE_rep2


title: VUMC0446


title: CARD14E138A_6hr_2


title: CDX2_E6.5_ExE_rep1


title: VUMC0445


title: VUMC0452


title: NaraCOVID_TP5_CD8_025_TCR


title: NaraCOVID_TP5_CD8_026_TCR


title: CDX2_TE_rep2


title: VUMC0443


title: CARD14E138A_6hr_1


title: CDX2_TSC_rep2

source_name: TSC

value: TSC


title: CARD14E138A_3hr_3


title: CDX2_TE_rep1


title: VUMC0442


title: NaraCOVID_TP5_CD8_024_TCR


title: NaraCOVID_TP5_CD8_030_TCR


title: CARD14E138A_3hr_2


title: NaraCOVID_TP5_CD8_023_TCR


title: CDX2_TSC_rep1


title: VUMC0441


title: CDX2_E6.5_ExE_rep3


title: CARD14E138A_6hr_3


title: NaraCOVID_TP5_CD8_022_TCR


title: CARD14WT_6hr_2


title: CDX2_Morula_rep2


title: CDX2_Morula_rep1


title: NaraCOVID_TP5_CD8_021_TCR


title: VUMC0440


title: CARD14E138A_0hr_3


title: NaraCOVID_TP5_CD8_020_TCR


title: Inguinal White adipose tissue, Day21, scRNA-seq

first: Yeung

last: Sivannn

address: Dongan Road

institute: Fudan University

description: SAMPLES 4

data_processing:
Raw sequencing reads of the cDNA library were mapped to the reference genome by 10X Genomics Cell Ranger (v3.1.0). Genome Reference Consortium Mouse Build 38 (GRCm38) was used as reference for 10X Cell Ranger.
The cell barcodes in the file are designated by the suffixes -1, -2, -3, -4, and -5, representing samples 1, 2, 3, 4, and 5, respectively.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Inguinal White adipose tissue

extract_protocol:
Stromal vascular fraction of mouse iWAT were prepared by enzymatic dissociation with collagenase Ⅷ (sigma c2139).
Cells were loaded into Chromium microfluidic chips and barcoded within a 10× Chromium Controller (10X Genomics), Single cell transcriptome were reverse-transcribed into cDNA library containing 10X cell barcodes and unique molecular identifiers (UMI) information. All procedures were performed with reagents from Chromium Next GEM Single Cell 3ʹ GEM, Library & Gel Bead Kit v3.1 (10X Genomics, Cat. No. 1000121), following the manufacturers’ protocol. All the libraries were sequenced in a PE150 mode (Pair-End for 150bp read) on the NovaSeq platform (Illumina).

value: Inguinal White adipose tissue

value: 21 days old


title: CARD14E138A_0hr_2


title: Inguinal White adipose tissue, Day28, scRNA-seq

description: SAMPLES 5

value: 28 days old


title: VUMC0438


title: VUMC0439


title: Inguinal White adipose tissue, Day18, scRNA-seq

description: SAMPLES 3

value: 18 days old