title: 1782163197_C43_H1


title: 1782163197_C57_B6

status: Public on Jun 29 2020

type: SRA

city: Stanford

first: Wanxin

last: Wang

state: CA

address: James H Clark Center, 318 Campus Drive, room E300

country: USA

institute: Stanford Univeristy

library_source: transcriptomic

data_processing:
bcl2fastq v2.17.1.14 was used for demultiplexing and generating *.fastq.
Raw reads in the *.fastq files were trimmed to 75bp using fastqx,
duplicate-removed using picard MarkDuplicates, and
aligned to reference using STAR with default parameters.
Aligned reads were converted to counts using HTSeq.
Genome_build: Ensembl human reference genome GRCh38.87 (dna.primary_assembly)
Supplementary_files_format_and_content: comma separated file including HTSeq count for each cell

molecule: polyA RNA

organism: Homo sapiens

source_name: endometrium biopsy

extract_protocol:
Single cells were lysed using the reagents (Takara/Clontech) and default protocol for Fluidigm C1 mRNA Seq
RNA libraries were prepared for sequencing using Illumina Nextera-XT protocols for single cells

tag: time in menstrual cycle

tag: tissue

value: endometrium biopsy

tag: donor condition

value: fertile healthy

tag: gender

value: female


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title: AA0028_Neut_Baseline

city: Bethesda

first: Luis

middle: M

last: Franco

state: MD

address: 9000 Rockville Pike

department: National Institute of Allergy and Infectious Diseases

institute: National Institutes of Health

description: RawReadCounts.txt

data_processing:
bcl conversion and de-multiplexing was performed with bcl2fastq2, version 2.20
Adapter trimming was performed with Cutadapt, v. 1.10. The FASTQ files uploaded to GEO are the untrimmed version of the files
Read alignment was performed with STAR, v. 2.6.0. The genome index was generated with a version of the GRCh38 build obtained from GENCODE. The transcript annotation file was also obtained from GENCODE (v28 primary assembly annotation)
Read counting was performed with featureCounts (part of the subread package, v1.6.2)
The ouptut of featureCounts (raw read counts) was converted to a matrix (RawReadCounts.txt) with R v3.6.1
Genome_build: GRCh38
Supplementary_files_format_and_content: RawReadCounts.txt: A tab-delimited text file with 56871 rows and 20 columns. The first row is a header row with sample IDs. The first column has gene symbols. The values are raw (un-normalized) read counts generated by featureCounts.

molecule: total RNA

source_name: Neutrophils

extract_protocol:
Total RNA was isolated by extraction with TRIzol Reagent (Thermo Fisher Scientific, Cat. No. 15596018) followed by column-based purification with the RNA Clean & Concentrator-5 kit (Zymo Research; Cat. No. R1016).
RNA libraries were prepared with the Illumina TruSeq Stranded Total RNA Gold Library Preparation Kit (Illumina, Cat. No. 20020599) and the TruSeq RNA CD Index Plate, 96 indices (Cat. No. 20019792).

tag: donor gender

value: Female

tag: donor type

value: Control

tag: donor age (years)

tag: library prep

value: TruSeq


title: 1772159322_C77_E11


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title: 1772159322_C27_E3


title: 1772159322_C38_G2


title: 1772159322_C23_D8


title: 1772159322_C12_B7


title: AA0022_Neut_Baseline

description: RawReadCounts.txt


title: AA0024_Neut_Baseline

description: RawReadCounts.txt

value: Male


title: AA0025_Neut_Baseline

description: RawReadCounts.txt


title: 1772159322_C06_A7


title: 1772159322_C10_B9


title: 1772159322_C16_C9


title: AA0019_Neut_Baseline

description: RawReadCounts.txt


title: 1772159322_C09_B1


title: AA0023_Neut_Baseline

description: RawReadCounts.txt


title: 1772159322_C07_B3


title: AA0021_Neut_Baseline

description: RawReadCounts.txt


title: 1772159322_C05_A8


title: AA0027_Neut_Baseline

description: RawReadCounts.txt


title: AA0016_Neut_Baseline

description: RawReadCounts.txt


title: AA0014_Neut_Baseline

description: RawReadCounts.txt


title: AA0015_Neut_Baseline

description: RawReadCounts.txt


title: AA0012_Neut_Baseline

description: RawReadCounts.txt


title: AA0011_Neut_Baseline

description: RawReadCounts.txt


title: AA0013_Neut_Baseline

description: RawReadCounts.txt


title: 1772159146_C92_H5


title: 1772159146_C96_H10


title: 1772159146_C93_H4


title: 1772159146_C95_H11


title: 1772159146_C90_G10


title: AA0010_Neut_Baseline

description: RawReadCounts.txt


title: 1772159146_C87_G4


title: AA0007_Neut_Baseline

description: RawReadCounts.txt


title: AA0004_Neut_Baseline

description: RawReadCounts.txt


title: 1772159322_C03_A1


title: AA0006_Neut_Baseline

description: RawReadCounts.txt


title: 1772159146_C82_F12


title: 1772159146_C83_F11


title: AA0003_Neut_Baseline

description: RawReadCounts.txt


title: 1772159322_C01_A3


title: 1772159146_C80_F5


title: AA0001_Neut_Baseline

description: RawReadCounts.txt


title: 1772159146_C79_F6


title: 1772159146_C78_E10


title: 1772159146_C74_E5


title: +CPI colitis C2 sample CD3 TCR sequencing

city: Boston

first: Shengbao

last: Suo

state: MA

address: 360 Longwood Ave

institute: Dana-Farber Cancer Institute

description:
C2-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv

data_processing:
Library strategy: TCR-seq
Raw single-cell RNA-seq data were processed using 10X Genomics Cell Ranger (v3.0.2)
“cellranger count” function is used to align sequencing reads to the human reference genome (GRCh38, v3.0.0, from 10X Genomics), and count the unique molecular identifier (UMI)
All patient samples from each dataset were then aggregated by using the “cellranger aggr” function without depth normalization
After getting the digital cell by gene expression matrix, the cells with poor quality (cells with high percentage of mitochondrial gene reads, cells express less than 200 genes and cells which show high doublet scores determined by DoubletFinder (v2.0.2)) were filtered)
Raw TCR sequencing data were processed by using 10X Genomics Cell Ranger (v3.1.0). Briefly, FASTQ reads were aligned to human GRCh38 V(D)J reference genome (v3.1.0, from 10X Genomics) by using the “cellranger vdj” function
Genome_build: GRCh38
Supplementary_files_format_and_content: *genes-barcodes-matrix.tar.gz (Each RNA sequencing sample contains a tar archive including matrix.mtx (UMI count matrix in a sparse matrix format), genes.tsv (Gene names) and barcodes.tsv (Cell barcodes). )
TCR_filtered_contig_annotations_all.csv (TCR sequencing data for all CD3 samples are saved in a csv file, which is generated by 'cellranger vdj' function and shows the detailed clonotype information.)

source_name: Colon pinch biopsy

growth_protocol: single cell suspensions were prepared from colon biopsies.

extract_protocol: Manufacturer's protocols for 5' barcoded GEX and V(D)J TCR

treatment_protocol: Biopsies were transferred to 6 cm tissue culture dishes placed on ice, and cut into <1 mm pieces with a razor blade while immersed in a small volume in DMEM to preserve tissue hydration. Tissue pieces were centrifuged (400 g for 5 min at 4°C for all centrifuge steps) and resuspended in 2 mL of collagenase-containing enzymatic digestion solution prepared according to manufacturer’s instructions (Miltenyi Human Tumor Dissociation Kit #30-095-929). Biopsies were incubated at 37°C for 25-30 min with mixing using a P1000 pipet tip (10 minute intervals) and intervening gentle vortexing (2-5 minute intervals). Digestion produced a nearly uniform single cell suspension, after which cells were filtered through a 70 um nylon mesh filter. Filtered cell suspensions were centrifuged and resuspended in 3 mL ACK red blood cell lysis buffer (Gibco) for 1 minute at RT, then immediately diluted with 10 mL of PBS buffer containing 0.5% FBS and 1 mM EDTA to halt osmotic stress. Cells were centrifuged and subsequently resuspended in Zombie NIR (Biolegend #423106) viability dye solution (1:500 in PBS), followed by 15 min incubation at RT in the dark. Cells were counted with a hemocytometer, generally yielding >100,000 live cells per biopsy. Following viability dye staining, cells were allocated for sorting/single-cell RNA-seq. For patients with biopsies spanning multiple regions, descending colon and sigmoid-rectum cell suspensions were combined in equal numbers to provide an aggregate analysis of the lower colon. Cells stained with viability dye (0.4-0.6 x 106) were resuspended in 100 uL FACS wash buffer (FWB; PBS, 2%FBS, 1 mM EDTA) with the addition of the following antibodies: TruStain FcX (Biolegend 422302), CD3-PE/Dazzle (Biolegend 300450), CD45-Pacific Blue (Biolegend 304022), CD66b-PE/Cy7 (Biolegend 305116) and CD15-APC (Biolegend 301908). Cells were incubated on ice for 20 minutes, washed with 1 mL FWB, and then resuspended in 500 uL FWB for sorting on a FACS Aria II or III (BD Biosciences). For the majority of samples, 12-15 x 10e3 live CD45+CD66b- mononuclear cells (MNC) cells and CD3+ cells (pre-gated on live MNC) were separately sorted into T cell media (RMPI + 10% FBS) and immediately transferred to ice.

tag: tissue region

value: Ascending, transverse, descending, sigmoidal colon; rectum

tag: checkpoint inhibitor-treated

value: yes

tag: colon status

value: +CPI colitis

tag: cell marker

tag: 10x chemistry

value: scRNA-seq (10X Genomics 5' V1 VDJ)


title: +CPI colitis C2 sample CD45 RNA-seq

data_processing:
Raw single-cell RNA-seq data were processed using 10X Genomics Cell Ranger (v3.0.2)
“cellranger count” function is used to align sequencing reads to the human reference genome (GRCh38, v3.0.0, from 10X Genomics), and count the unique molecular identifier (UMI)
All patient samples from each dataset were then aggregated by using the “cellranger aggr” function without depth normalization
After getting the digital cell by gene expression matrix, the cells with poor quality (cells with high percentage of mitochondrial gene reads, cells express less than 200 genes and cells which show high doublet scores determined by DoubletFinder (v2.0.2)) were filtered)
Raw TCR sequencing data were processed by using 10X Genomics Cell Ranger (v3.1.0). Briefly, FASTQ reads were aligned to human GRCh38 V(D)J reference genome (v3.1.0, from 10X Genomics) by using the “cellranger vdj” function
Genome_build: GRCh38
Supplementary_files_format_and_content: *genes-barcodes-matrix.tar.gz (Each RNA sequencing sample contains a tar archive including matrix.mtx (UMI count matrix in a sparse matrix format), genes.tsv (Gene names) and barcodes.tsv (Cell barcodes). )
TCR_filtered_contig_annotations_all.csv (TCR sequencing data for all CD3 samples are saved in a csv file, which is generated by 'cellranger vdj' function and shows the detailed clonotype information.)

value: scRNA-seq (10X Genomics 5' V1 GEX)


title: +CPI colitis C1 sample CD45 RNA-seq

value: Sigmoidal colon, rectum


title: 1772159322_C22_D9


title: 1772159146_C84_F10


title: 1772159146_C72_D12


title: 1772159146_C85_G6


title: 1772159146_C81_F4


title: 1772159146_C73_E6


title: 1772159146_C71_D11


title: 1772159146_C70_D10


title: 1772159146_C68_D5


title: 1772159146_C67_D4


title: 1772159146_C69_D6


title: 1772159146_C64_C10


title: 1772159146_C65_C11


title: 1772159146_C63_C6


title: 1772159146_C61_C4


title: 1772159146_C59_B11


title: PDX of bone metastasis, patient 142

type: RNA

city: Paris

first: Elisabetta

last: Marangoni

address: 26 rue d'Ulm

country: France

institute: Institut Curie

description: Gene expression data HBCx-142

hyb_protocol: GeneChip Human 1.1 ST arrays were hybridized according to Affymetrix recommendations, using Ambion WT Expression Kit protocol (Life Technologies) and Affymetrix labeling and hybridization kits

scan_protocol: Standard Affymetrix protocol

data_processing: Affymetrix CEL files were imported into the Gene Expression Workflow in Partek® Genomics Suite version 7.0 (Partek Inc., St. Louis, MO, USA, www.partek.com)

label: Biotin

source_name: human breast cancer xenograft 142

label_protocol: Standard Affymetrix protocol

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

tag: disease state

value: Oestrogen receptor positive breast cancer

tag: tissus

value: Human bone metastasis xenograft

tag: treatment protocol

value: Endocrine treatments

tag: growth protocol

value: Engrafted into the interscapular fat pad of female Swiss nude mice


title: Breast tumor, patient 142

description: Gene expression data HBC-142

source_name: human breast cancer 142

value: Primary breast tumor

value: Untreated samples

value: Breast cancer biopsy


title: PDX of bone metastasis, patient 131

description: Gene expression data HBCx-131

source_name: human breast cancer xenograft 131


title: Breast tumor, patient 134

description: Gene expression data HBC-134

source_name: human breast cancer 134


title: 1772159146_C58_B10

tag: time in menstrual cycle

value: endometrium biopsy

value: fertile healthy

value: female


title: 1772159146_C55_B4

tag: tissue


title: 1772159146_C50_A5

tag: donor condition


title: 1772159146_C57_B6

tag: gender


title: PDX of bone metastasis, patient 137

description: Gene expression data HBCx-137

source_name: human breast cancer xenograft 137


title: Breast tumor, patient 118

description: Gene expression data HBC-118

source_name: human breast cancer 118


title: PDX of bone metastasis, patient 134

description: Gene expression data HBCx-134

source_name: human breast cancer xenograft 134


title: 1772159146_C56_B5


title: PDX of bone metastasis, patient 139

description: Gene expression data HBCx-139

source_name: human breast cancer xenograft 139


title: Breast tumor, patient 124

description: Gene expression data HBC-124

source_name: human breast cancer 124


title: 1772159146_C49_A4


title: PDX of bone metastasis, patient 118

description: Gene expression data HBCx-118

source_name: human breast cancer xenograft 118


title: 1772159146_C60_B12


title: 1772159146_C47_H8


title: 10_IC_Stimulated_14hr_rep1

city: Cambridge

first: John

middle: Robert

last: Ferdinand

address: Department of Medicine

country: United Kingdom

institute: University of Cambridge

description: gene expression data

hyb_protocol: GeneTitan Hybridization, Wash and Stain kit (Affymetrix)

scan_protocol: Plate arrays were processed on the GeneTitan instrument (Affymetrix)

data_processing: Bioconductor, limma, oligo

label: biotin

organism: Mus musculus

label_protocol: RNA was assessed for concentration and quality using a SpectroStar (BMG Labtech, Aylesbury, UK) and a Bioanalyser (Agilent Technologies, Cheadle, UK). Microarray experiments were performed at Cambridge Genomic Services, University of Cambridge, using a species specific Gene 2.1 ST Array Plate (Affymetrix, Wooburn Green, UK) in combination with WT PLUS amplification kit (Affymetrix) according to the manufacturer’s instructions. Briefly, 100ng Total RNA was amplified along with inline PolyA spike in control RNA, using the WT PLUS amplification kit (Affymetrix). Successfully amplified samples were labelled using the GeneChip WT terminal labelling kit (Affymetrix) using the in line hybridization controls.

growth_protocol: Murine bone marrow was flushed form femurs and tibias, plated in regular petri dishes and supplemented with 100 ng mL-1 macrophage colony-stimulating factor (M-CSF, Peprotech) in complete medium (10% Hyclone FBS, 1% penicillin/streptomycin in RPMI 1640) twice over 5 days. On day 6, macrophages were transferred to tissue culture plates for stimulation.

extract_protocol: Cells were lysed and RNA extracted using Ambion pure link RNA minikit as per kit instructions.

treatment_protocol: For ovalbumin immune complexes, endotoxin-free ovalbumin (EndoGrade Ovalbumin, Hyglos GmbH) was opsonized with a polyclonal rabbit anti-ovalbumin antibody (Sigma-Aldrich) at 37C for 1h. Subsequently BMDMs were treated with OVA-IC for 4  or 14 hrs or left untreated.

tag: mouse id#

tag: cell type

value: Bone marrow derived cells

tag: treatment

value: OVA immune Complex

tag: timepoint


title: 11_IC_Stimulated_14hr_rep2

description: gene expression data


title: PDX of bone metastasis, patient 124

description: Gene expression data HBCx-124

source_name: human breast cancer xenograft 124


title: 8_Control_14hr_rep2

description: gene expression data

value: Unstimulated


title: 9_Control_14hr_rep3

description: gene expression data


title: 12_IC_Stimulated_14hr_rep3

description: gene expression data


title: 7_Control_14hr_rep1

description: gene expression data


title: 6_IC_Stimulated_4hr_rep3

description: gene expression data


title: 5_IC_Stimulated_4hr_rep2

description: gene expression data


title: 3_Control_4hr_rep3

description: gene expression data


title: 1772159146_C43_H1


title: 1772159146_C44_H2


title: 4_IC_Stimulated_4hr_rep1

description: gene expression data


title: 1772159146_C42_G9


title: 1772159146_C41_G8


title: 1772159146_C39_G3


title: 1772159146_C38_G2


title: 1772159146_C37_G1


title: 2_Control_4hr_rep2

description: gene expression data


title: 1772159146_C36_F9


title: 1_Control_4hr_rep1

description: gene expression data


title: 1772159146_C35_F8


title: 1772159146_C34_F7


title: HC190128-4

city: changchun

first: Feng

last: Hu

state: jilin

address: No. 3302, Jilin Dadu

department: Department of Hepatology and Gastroenterology

country: China

institute: The Second Part of First Hospital of Jilin University

data_processing:
Basecalls performed using CASAVA version 1.8
lncRNA and mRNA-seq reads  were aligned to the human reference genome (UCSC HG19) with hisat2 software.
Then, guided by the Ensembl gtf gene annotation file, cuffdiff software (part of cufflinks) was used to get the FPKM as the expression profiles of LncRNA and mRNA, and fold change and p-value were calculated based on FPKM, differentially expressed LncRNA and mRNA were identified.
Genome_build: hg19
Supplementary_files_format_and_content: excel format:Number of raw junction read

source_name: plasma

extract_protocol:
Collect the plasma of GHGIN and HC, extract plasma exosomes.Total RNA in the exosomes was extracted using TRIzol reagent (Life Technologies, Carlsbad, CA, USA).First, rRNAs were eliminated from total RNA using the NEB Next rRNA depletion kit (New England Biolabs, Inc., Massachusetts, USA). Then, NEB Next UltraTM II Directional RNA Library Prep Kit (New England Biolabs, Inc., Massachusetts, USA) was used to construct RNA libraries from rRNA-depleted RNAs according to the instructions of manufacturer. The BioAnalyzer 2100 system (Agilent Technologies, Inc., USA) was used to evaluate the quality and quantity of the libraries. These libraries were sequenced on an Illumina HiSeq 4000 sequencer (Illumina, USA) and 150 bp paired-end reads were generated according to the instructions of manufacturer.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: patient diagnosis

value: Healthy control

value: plasma

tag: cell fraction

value: exosome


title: pancreatic islets, isolated from β cell-specific Sel1L-deficient mouse, replicate 1

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

country: Netherlands

institute: Wageningen University

hyb_protocol: Hybridization and washing of the Affymetrix GeneChip Mouse Gene 2.1 ST peg arrays were performed on a GeneTitan Instrument according to the manufacturer's recommendations (Affymetrix, Santa Clara, CA, USA).

scan_protocol: Arrays were scanned on an Affymetrix GeneTitan instrument (Affymetrix, Santa Clara, CA, USA).

data_processing: Expression estimates were calculated applying the RMA algorithm in the Bioconductor library 'Oligo' (v1.50.0).

source_name: pancreatic islets, isolated from 5-wk old female β cell-specific Sel1L-deficient mouse

label_protocol: Total RNA (100ng per sample) was labeled with the Whole-Transcript Sense Target Assay (Affymetrix, Santa Clara, CA, USA; P/N 900652).

growth_protocol: Sel1Lf/f mice on the C57BL/6J background were crossed with B6(Cg)-Ins1tm1.1(cre)Thor/J (Ins1Cre, JAX 026801) on the C57BL/6J background to generate β cell-specific Sel1L (Sel1LIns1) deficient mice with respective control littermates (Sel1L f/f). All mice were housed in an ambient temperature room with 12 h light cycle and fed a regular chow diet (13% fat, 57% carbohydrate, and 30% protein; LabDiet 5L0D).

extract_protocol: RNA was extracted using RNeasy micro kit (Qiagen) including a column for elimination of genomic DNA as per manufacturer’s instructions. RNA concentration was determined using NanoDrop 2000 UV-Vis Spectrophotometer. The quality and concentration were determined using the RNA 6000 Nano kit on an Agilent 2100 bioanalyser.

treatment_protocol: To isolate pancreatic islets, mice were euthanized by cervical dislocation and immediately processed for pancreatic perfusion. The pancreas was distended via the intraductal injection of 3 mL of liberase and incubated at 37°C for 12 minutes in additional 1 mL of digestion solution. Liberase solution was prepared by dissolving 5 mg Liberase TL (Roche, 5401020001) in 20 ml of serum free RPMI (Fisher, 11875-085). Digestion was stopped by adding cold media containing 10% FBS. After gentle shaking for complete dissociation and brief centrifugation, the digested suspension was passed through a nylon mesh and islets were isolated by density gradient centrifugation on a Histopaque gradient (1.077 g/mL density; Sigma) for 20 min at 900 x g without brake. Islets were then collected from the interface, washed and hand-picked under a dissecting microscope. Isolated islets were allowed to recover overnight in RPMI 1640 medium in a humidified incubator (95% air, 5% CO2) at 37°C. Islets were collected from 5-week WT and Sel1LIns1 mice and recovered overnight prior to RNA extraction.

tag: genotype

value: Beta cell-specific Sel1L KO

tag: background strain

value: pancreatic islets

tag: age of mice (wks)

tag: Sex


title: HC190128-5


title: HC190128-3


title: pancreatic islets, isolated from β cell-specific Sel1L-deficient mouse, replicate 3


title: pancreatic islets, isolated from β cell-specific Sel1L-deficient mouse, replicate 2


title: pancreatic islets, isolated from wildtype mouse, replicate 2

source_name: pancreatic islets, isolated from 5-wk old female wildtype (C57BL/6J) mouse

value: wildtype


title: pancreatic islets, isolated from wildtype mouse, replicate 1


title: pancreatic islets, isolated from wildtype mouse, replicate 3


title: HC190128-1


title: 1772159146_C31_F1

source_name: endometrium biopsy


title: 1772159146_C32_F2


title: HC190128-2


title: EGC5

value: gastric high-grade intraepithelial neoplasia


title: 1772159146_C30_E9


title: 1772159146_C29_E8


title: EGC4


title: EGC2


title: 1772159146_C25_E1


title: 1772159146_C22_D9


title: 1772159146_C26_E2


title: EGC3


title: EGC1


title: 1772159146_C23_D8


title: Normal control CT6 sample CD3 gd TCR sequencing

description:
T cell receptor
CT6-gdTCR

data_processing:
Library strategy: TCR-seq
Raw single-cell RNA-seq data were processed using 10X Genomics Cell Ranger (v3.0.2)
“cellranger count” function is used to align sequencing reads to the human reference genome (GRCh38, v3.0.0, from 10X Genomics), and count the unique molecular identifier (UMI
All patient samples from each dataset were then aggregated by using the “cellranger aggr” function without depth normalization
After getting the digital cell by gene expression matrix, the cells with poor quality (cells with high percentage of mitochondrial gene reads, cells express less than 200 genes and cells which show high doublet scores determined by DoubletFinder (v2.0.2)) were filted
Raw TCR sequencing data were processed by using 10X Genomics Cell Ranger (abTCR: v3.1.0, gdTCR: v3.0.0). Briefly, FASTQ reads were aligned to human GRCh38 V(D)J reference genome (V3.1.0) by using the “cellranger vdj” function
Genome_build: GRCh38
Supplementary_files_format_and_content: Each RNA sequenting sample contains a compressed tab-delimited text files including matrix.mtx (UMI count matrix in a sparse matrix format), genes.tsv (Gene names) and barcodes.tsv (Cell barcodes). TCR sequencing data for each gdTCR sample are saved in a csv file, which is generated by 'cellranger vdj' function and shows the detailed clonotype information.

extract_protocol:
Manufacturer's protocols for 5' barcoded GEX and V(D)J TCR
Manufacturer's protocols for 5' barcoded GEX and V(D)J TCR

treatment_protocol: Biopsies were transferred to 6 cm tissue culture dishes placed on ice, and cut into <1 mm pieces with a razor blade while immersed in a small volume in DMEM to preserve tissue hydration. Tissue pieces were centrifuged (400 g for 5 min at 4°C for all centrifuge steps) and resuspended in 2 mL of collagenase-containing enzymatic digestion solution prepared according to manufacturer’s instructions (Miltenyi Human Tumor Dissociation Kit #30-095-929). Biopsies were incubated at 37°C for 25-30 min with mixing using a P1000 pipet tip (10 minute intervals) and intervening gentle vortexing (2-5 minute intervals). Digestion produced a nearly uniform single cell suspension, after which cells were filtered through a 70 m nylon mesh filter. Filtered cell suspensions were centrifuged and resuspended in 3 mL ACK red blood cell lysis buffer (Gibco) for 1 minute at RT, then immediately diluted with 10 mL of PBS buffer containing 0.5% FBS and 1 mM EDTA to halt osmotic stress. Cells were centrifuged and subsequently resuspended in Zombie NIR (Biolegend #423106) viability dye solution (1:500 in PBS), followed by 15 min incubation at RT in the dark. Cells were counted with a hemocytometer, generally yielding >100,000 live cells per biopsy. Following viability dye staining, cells were allocated for sorting/single-cell RNA-seq. For patients with biopsies spanning multiple regions, descending colon and sigmoid-rectum cell suspensions were combined in equal numbers to provide an aggregate analysis of the lower colon. Cells stained with viability dye (0.4-0.6 x 106) were resuspended in 100 L FACS wash buffer (FWB; PBS, 2%FBS, 1 mM EDTA) with the addition of the following antibodies: TruStain FcX (Biolegend 422302), CD3-PE/Dazzle (Biolegend 300450), CD45-Pacific Blue (Biolegend 304022), CD66b-PE/Cy7 (Biolegend 305116) and CD15-APC (Biolegend 301908). Cells were incubated on ice for 20 minutes, washed with 1 mL FWB, and then resuspended in 500 L FWB for sorting on a FACS Aria II or III (BD Biosciences). For the majority of samples, 12-15 x 103 live CD45+CD66b- mononuclear cells (MNC) cells and CD3+ cells (pre-gated on live MNC) were separately sorted into T cell media (RMPI + 10% FBS) and immediately transferred to ice.

tag: tissue region

value: Descending, sigmoidal colon; rectum

tag: checkpoint inhibitor (cpi)-treated

value: no

tag: colon status

value: Normal

tag: cell marker

tag: 10x chemistry

value: scRNA-seq (10X Genomics 5' V1 VDJ)


title: 1772159146_C24_D7

tag: time in menstrual cycle

tag: tissue

value: endometrium biopsy

tag: donor condition


title: 1772159146_C20_D2

value: fertile healthy

tag: gender


title: Normal control CT7 sample CD3 gd TCR sequencing

description:
T cell receptor
CT7-gdTCR


title: 1772159146_C17_C8

value: female


title: 1772159146_C16_C9


title: Normal control CT1 sample CD3 gd TCR sequencing

description:
T cell receptor
CT1-gdTCR


title: Normal control CT2 sample CD3 gd TCR sequencing

description:
T cell receptor
CT2-gdTCR


title: +CPI no colitis NC4 sample CD3 gd TCR sequencing

description:
T cell receptor
NC4-gdTCR

value: yes

value: +CPI no colitis


title: Normal control CT5 sample CD3 gd TCR sequencing

description:
T cell receptor
CT5-gdTCR


title: 1772159146_C12_B7


title: 1772159146_C10_B9


title: 1772159146_C07_B3


title: 1772159146_C04_A9


title: +CPI no colitis NC3 sample CD3 gd TCR sequencing

description:
T cell receptor
NC3-gdTCR


title: 1772159146_C06_A7


title: 1772159146_C03_A1


title: +CPI no colitis NC2 sample CD3 gd TCR sequencing

description:
T cell receptor
NC2-gdTCR

value: Ascending, Descending, sigmoidal colon; rectum


title: 1772159146_C02_A2


title: +CPI no colitis NC1 sample CD3 gd TCR sequencing

description:
T cell receptor
NC1-gdTCR


title: Normal control CT3 sample CD3 gd TCR sequencing

description:
T cell receptor
CT3-gdTCR


title: 1772158110_C94_H12


title: +CPI no colitis NC6 sample CD3 gd TCR sequencing

description:
T cell receptor
NC6-gdTCR


title: Normal control CT8 sample CD3 gd TCR sequencing

description:
T cell receptor
CT8-gdTCR


title: 1772159146_C01_A3


title: +CPI colitis C8 sample CD3 gd TCR sequencing

description:
T cell receptor
C8-gdTCR

value: Sigmoidal colon

value: +CPI colitis


title: 1772159146_C19_D3


title: +CPI colitis C7 sample CD3 gd TCR sequencing

description:
T cell receptor
C7-gdTCR


title: +CPI no colitis NC5 sample CD3 gd TCR sequencing

description:
T cell receptor
NC5-gdTCR


title: +CPI colitis C6 sample CD3 gd TCR sequencing

description:
T cell receptor
C6-gdTCR


title: 1772158110_C93_H4


title: 1772158110_C91_H6


title: 1772159146_C11_B8


title: Normal control CT4 sample CD3 gd TCR sequencing

description:
T cell receptor
CT4-gdTCR


title: +CPI colitis C5 sample CD3 gd TCR sequencing

description:
T cell receptor
C5-gdTCR


title: 1772158110_C90_G10


title: +CPI colitis C4 sample CD3 gd TCR sequencing

description:
T cell receptor
C4-gdTCR


title: 1772158110_C89_G11


title: +CPI colitis C3 sample CD3 gd TCR sequencing

description:
T cell receptor
C3-gdTCR


title: Normal control CT8 sample CD3 TCR sequencing

description:
CT8-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv

tag: checkpoint inhibitor-treated


title: Normal control CT7 sample CD3 TCR sequencing

description:
CT7-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C88_G12


title: 1772158110_C86_G5


title: 1772158110_C84_F10


title: 1772158110_C85_G6


title: Normal control CT4 sample CD3 TCR sequencing

description:
CT4-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: Normal control CT6 sample CD3 TCR sequencing

description:
CT6-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: Normal control CT3 sample CD3 TCR sequencing

description:
CT3-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C82_F12


title: 1772158110_C78_E10


title: 1772158110_C73_E6


title: Normal control CT5 sample CD3 TCR sequencing

description:
CT5-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: Normal control CT2 sample CD3 TCR sequencing

description:
CT2-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: Normal control CT1 sample CD3 TCR sequencing

description:
CT1-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C72_D12


title: +CPI no colitis NC6 sample CD3 TCR sequencing

description:
NC6-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C71_D11


title: +CPI no colitis NC5 sample CD3 TCR sequencing

description:
NC5-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C70_D10


title: +CPI no colitis NC3 sample CD3 TCR sequencing

description:
NC3-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C69_D6


title: 1772158110_C67_D4


title: 1772158110_C63_C6


title: +CPI colitis C8 sample CD3 TCR sequencing

description:
C8-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C65_C11


title: 1772158110_C79_F6


title: +CPI no colitis NC2 sample CD3 TCR sequencing

description:
NC2-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C57_B6


title: +CPI colitis C7 sample CD3 TCR sequencing

description:
C7-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C62_C5


title: +CPI colitis C6 sample CD3 TCR sequencing

description:
C6-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: +CPI colitis C3 sample CD3 TCR sequencing

description:
C3-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: Normal control CT6 sample CD45 RNA-seq

value: scRNA-seq (10X Genomics 5' V1 GEX)


title: 1772158110_C56_B5


title: +CPI colitis C5 sample CD3 TCR sequencing

description:
C5-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: +CPI colitis C4 sample CD3 TCR sequencing

description:
C4-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C54_A12


title: 1772158110_C52_A10


title: +CPI no colitis NC4 sample CD3 TCR sequencing

description:
NC4-TCR
T cell receptor
TCR_filtered_contig_annotations_all.csv


title: 1772158110_C50_A5


title: Normal control CT4 sample CD45 RNA-seq


title: 1772158110_C48_H9


title: 1772158110_C43_H1


title: Normal control CT3 sample CD45 RNA-seq


title: 1772158110_C45_H3


title: Normal control CT2 sample CD45 RNA-seq


title: 1772158110_C42_G9


title: Normal control CT1 sample CD45 RNA-seq


title: 1772158110_C41_G8


title: +CPI no colitis NC6 sample CD45 RNA-seq


title: +CPI no colitis NC4 sample CD45 RNA-seq


title: 1772158110_C40_G7


title: 1772158110_C39_G3


title: +CPI no colitis NC3 sample CD45 RNA-seq


title: +CPI no colitis NC2 sample CD45 RNA-seq


title: +CPI colitis C6 sample CD45 RNA-seq


title: 1772158110_C36_F9


title: +CPI no colitis NC1 sample CD45 RNA-seq


title: +CPI colitis C5 sample CD45 RNA-seq


title: 1772158110_C33_F3


title: 1772158110_C38_G2


title: 1772158110_C28_E7


title: 1772158110_C31_F1


title: +CPI colitis C3 sample CD45 RNA-seq


title: 1772158110_C26_E2


title: 1772158110_C25_E1


title: 1772158110_C23_D8


title: Normal control CT6 sample CD3 RNA-seq


title: Normal control CT8 sample CD3 RNA-seq


title: Normal control CT7 sample CD3 RNA-seq


title: Normal control CT5 sample CD3 RNA-seq


title: 1772158110_C20_D2


title: Normal control CT4 sample CD3 RNA-seq


title: 1772158110_C19_D3


title: Normal control CT3 sample CD3 RNA-seq


title: 1772158110_C18_C7


title: Normal control CT2 sample CD3 RNA-seq


title: 1772158110_C17_C8


title: Normal control CT1 sample CD3 RNA-seq


title: 1772158110_C16_C9


title: 1772158110_C14_C2


title: +CPI no colitis NC6 sample CD3 RNA-seq


title: 1772158110_C11_B8


title: +CPI colitis C4 sample CD45 RNA-seq


title: +CPI no colitis NC5 sample CD3 RNA-seq


title: +CPI no colitis NC3 sample CD3 RNA-seq


title: 1772158110_C05_A8


title: +CPI no colitis NC4 sample CD3 RNA-seq


title: 1772158110_C03_A1


title: 1772158110_C04_A9


title: 1772158110_C08_B2


title: +CPI no colitis NC2 sample CD3 RNA-seq


title: 1772158110_C02_A2


title: +CPI colitis C7 sample CD3 RNA-seq


title: +CPI no colitis NC1 sample CD3 RNA-seq


title: 1772158110_C24_D7


title: +CPI colitis C6 sample CD3 RNA-seq


title: 1772158109_C95_H11


title: 1772158109_C96_H10


title: +CPI colitis C5 sample CD3 RNA-seq


title: 1772158109_C94_H12


title: 1772158109_C93_H4


title: 1772158109_C88_G12


title: 1772158109_C90_G10


title: 1772158109_C87_G4


title: +CPI colitis C3 sample CD3 RNA-seq


title: +CPI colitis C4 sample CD3 RNA-seq


title: +CPI colitis C2 sample CD3 RNA-seq

value: Ascending, transverse, descending, sigmoidal colon; rectum


title: 1772158109_C89_G11


title: +CPI colitis C1 sample CD3 RNA-seq

value: Sigmoidal colon, rectum


title: +CPI colitis C8 sample CD3 RNA-seq


title: 1772158109_C86_G5


title: 1772158109_C85_G6


title: 1772158109_C78_E10


title: KIT-purified dSPG Rep3

city: La Jolla

first: Kun

last: Tan

address: 2880 Torrey Pines Scenic Dr. Room 4812

department: Department of Obstetrics, Gynecology, and Reproductive Sciences

institute: UCSD

description: fcounts.txt

data_processing:
For bulk RNA-Seq, Reads were filtered for quality and aligned with STAR (2.5.2b) against Homo sapiens, release-95, Ensembl genome (GRCh38). The exon counts were aggregated for each gene to build a read count table using SubRead function featureCounts. Differentially regulated genes were calculated using DESeq2. For differentially regulated genes a threshold (log2FC ≥ 1 or ≤ –1, q-val < 0.01) was followed. The R package pheatmap was used for clustering and heatmap plotting. The database for annotation, visualization and integrated discovery (DAVID) v6.8 was used for gene ontology analysis. The Ingenuity pathway analysis (IPA) was used for signaling pathway and genetic regulatory network analysis.
For single cell RNA-Seq, Demultiplexed raw sequencing reads were processed and mapped to the human genome (GRCh38) using Cell Ranger software (v3.0.2) with default parameters. Filtered count matrices for each library were tagged with a library batch ID and combined across independent experiments using the Seurat package (version 3.1.0) in R. To identify cell clusters, we employed Pathway and Geneset OverDispersion Analysis (PAGODA2). Single-cell pseudotime trajectories were constructed with the Monocle 2 package (v2.10.1).
Genome_build: GRCh38
Supplementary_files_format_and_content: For bulk RNA-Seq, processed data files are: fcounts.   For single cell RNA-Seq, processed data files are: matrix of genes, counts and cell barcodes.

source_name: human testicular cells

growth_protocol: For single cell RNA-Seq, Integrin-6 (ITGA6)+ SPG were purified from the isolated human testicular cells, and plated in 24-well plate pre-coated with laminin (20 μg/ml). The culture media are IMDM/SFM plus different growth factors or molecular inhibitor. Cells were passaged by incubation in StemPro accutase solution (Thermo Fisher Scientific) for 5 min.

extract_protocol:
For bulk RNA-Seq, total RNAs were extracted using the RNeasy Plus Micro Kit (Qiagen) according to the manufacturer’s protocol.   For single cell RNA-Seq, cells were incubated in StemPro accutase solution for 5 min. Single cell suspension were diluted for loading on Chromium chip for 10X Genomics library preparation and subsequent sequencing.
For bulk RNA-Seq, 10 ng of total RNA was subjected to prep the library using SMARTer® stranded total RNA-Seq kit v2 - pico input system (Takara Bio USA) per manufacturer's instructions.   For single cell RNA-Seq, Viable cells were washed once in PBS and resuspended in 0.04% BSA in PBS for loading on the 10x Chromium chip. Cell capturing, and library preparation was carried as per kit instructions (Chromium Single Cell 3′ Library Construction Kit v3).

treatment_protocol: For single cell RNA-Seq, cells were cultured under different conditions (FGF2 alone vs AKT-I+FGF2). Before loading for library prep, dead cells were removed using the ClioCell Dead Cell Removal kit (Amsbio) following the manufacturers’ instructions.

value: Testis biopsy

tag: age


title: PLPPR3-purified uSPG Rep4

description: fcounts.txt


title: KIT-purified dSPG Rep1

description: fcounts.txt


title: 1772158109_C72_D12


title: 1772158109_C71_D11


title: PLPPR3-purified uSPG Rep3

description: fcounts.txt


title: PLPPR3-purified uSPG Rep2

description: fcounts.txt


title: 1772158109_C79_F6


title: 1772158109_C70_D10


title: PLPPR3-purified uSPG Rep1

description: fcounts.txt


title: cloneIDs_from_V16

city: Solna

first: Michael

last: Ratz

address: Solnavägen 9

department: CMB

country: Sweden

institute: Karolinska Institute

description: amplicon library

data_processing:
Chromium: Sequencing reads were processed with Cell Ranger v2.2.0 (Chromium v2) or Cell Ranger v3.0.1 (Chromium v3); Visium: Sequencing reads were processed with Space Ranger 1.0.0
Genome_build: mm10 with additional sequence representing the H2B-EGFP-N transgene
Supplementary_files_format_and_content: Chromium: filtered gene count matrix; Visium: filtered gene count matrix, aligned images, spot coordinates

source_name: brain section

growth_protocol: mice were kept in standard housing conditions

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V16
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR

treatment_protocol: barcoding of progenitor cells was achieved via in utero microinjection

tag: region

value: Bregma AP -1.65 mm

tag: treatment

value: barcoded

tag: replicate

tag: technology

value: Visium Spatial Transcriptomics


title: cloneIDs_from_V14

description: amplicon library

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V14
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: KIT-purified dSPG Rep4

description: fcounts.txt


title: cloneIDs_from_V15

description: amplicon library

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V15
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: 1772158109_C67_D4


title: 1772158109_C66_C12


title: cloneIDs_from_V13

description: amplicon library

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V13
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: 1772158109_C84_F10


title: cloneIDs_from_V12

description: amplicon library

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V12
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: 1772158109_C64_C10


title: KIT-purified dSPG Rep2

description: fcounts.txt


title: 1772158109_C59_B11


title: cloneIDs_from_V11

description: amplicon library

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V11
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: 1772158109_C63_C6


title: section6

description: gene expression library

extract_protocol:
Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide, labelled using direct immunohistochemical staining and libraries prepared according to the manufacturer's instructions
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: cloneIDs_from_V10

description: amplicon library

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V10
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: cloneIDs_from_V9

description: amplicon library

extract_protocol:
Targeted PCR for enrichment of cloneIDs on full length cDNA obtained from sample V9
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: section4

description: gene expression library

extract_protocol:
Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide and libraries prepared according to the manufacturer's instructions
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR


title: section8

description: gene expression library


title: 1772158109_C75_E4

extract_protocol:
Single cells were lysed using the reagents (Takara/Clontech) and default protocol for Fluidigm C1 mRNA Seq
RNA libraries were prepared for sequencing using Illumina Nextera-XT protocols for single cells


title: section3

description: gene expression library


title: 1772158109_C58_B10


title: 1772158109_C50_A5


title: 1772158109_C51_A6


title: section1

description: gene expression library


title: 1772158109_C41_G8


title: section5

description: gene expression library


title: section2

description: gene expression library


title: 1772158109_C42_G9


title: 1772158109_C62_C5


title: 1772158109_C38_G2


title: 1772158109_C36_F9


title: 1772158109_C39_G3


title: section7

description: gene expression library


title: 1772158109_C32_F2


title: 1772158109_C54_A12


title: R9485 d3_mid_grade_sepsis_preneu

city: Singapore

first: Nicholas

last: Ang

address: 8a Biomedical Grove, Singapore 138648

country: Singapore

institute: Singapore Immunology Network (SIgN)

data_processing:
FASTQ files were mapped using STAR to  GRCm38 genome build
Gene counts were obtained using featureCounts on GENCODE vM20 annotations
Genome_build: GRCm38
Supplementary_files_format_and_content:  genecounts_20200601.txt

source_name: bone marrow

extract_protocol:
Total RNA was extracted using Arcturus PicoPure RNA Isolation kit (Applied Biosystems™ Thermo Fisher Scientific) according to manufacturer's protocol.
All RNAs were analyzed on Agilent Bioanalyser for quality assessment with a median RNA Integrity Number (RIN) of 9.4. cDNA libraries were prepared using 2 ng of total RNA with 1ul of 1:50,000 dilution of ERCC RNA Spike in Controls respectively (Ambion® Thermo Fisher Scientific) using the SMARTSeq v2 protocol (Picelli et al., 2014) with the following modifications: 1. Addition of 20 µM TSO; 2. Use of 200pg cDNA with 1/5 reaction of Illumina Nextera XT kit (Illumina, San Diego, CA, USA). The length distribution of the cDNA libraries was monitored using a DNA High Sensitivity Reagent Kit on the Perkin Elmer Labchip (Perkin Elmer, Waltham, MA, USA). All samples were subjected to an indexed paired-end sequencing run of 2x151 cycles on an Illumina HiSeq 4000 system (Illumina) (26 samples/lane).

tag: cell type


title: R9483 d3_mid_grade_sepsis_preneu


title: R9484 d3_mid_grade_sepsis_preneu


title: 1772158109_C28_E7


title: 1772158109_C43_H1


title: 1772158109_C24_D7


title: brain5_hc

description: gene expression library

source_name: brain cells

extract_protocol:
Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions.
Chromium: Brains were dissected and single cell dissociations prepared using the Papain dissociation system with manual trituration. Purified single cell suspensions were processed according to the manufacturer's instructions. Visium: Brains were fresh frozen, 10 µm cryosections mounted and fixed on a Visium glass slide +/- immunostaining and libraries prepared according to the manufacturer's instructions. Amplicon: full length cDNA obtained during Chromium or Visium library preparation.
10X Genomics Chromium Single Cell 3' v2 and v3; 10X Visium Spatial Gene Expression; Nested PCR

value: hippocampus

value: control

value: Chromium Single Cell 3' v3


title: R9480 d3_mid_grade_sepsis_proneu2


title: R9479 d3_mid_grade_sepsis_proneu1


title: 1772158109_C49_A4

value: endometrium biopsy

value: fertile healthy


title: 1772158109_C23_D8

value: female


title: R9476 d3_mid_grade_sepsis_ly6c_neg_gmp


title: R9475 d3_mid_grade_sepsis_ly6c_neg_gmp


title: R9477 d3_mid_grade_sepsis_proneu1


title: 1772158109_C18_C7


title: 1772158109_C21_D1


title: 1772158109_C17_C8


title: R9495 ly6chi_mo


title: 1772158109_C22_D9


title: R9472 ly6chi_mo


title: R9474 d3_mid_grade_sepsis_ly6c_neg_gmp


title: R9478 d3_mid_grade_sepsis_proneu1


title: 1772158109_C16_C9


title: R9481 d3_mid_grade_sepsis_proneu2


title: R9471 ly6chi_mo


title: 1772158109_C11_B8


title: 1772158109_C10_B9


title: R9470 tpmo

value: tpmo


title: 1772158109_C25_E1


title: 1772158109_C26_E2


title: R9482 d3_mid_grade_sepsis_proneu2


title: 1772158109_C09_B1


title: R9468 tpmo


title: 1772158109_C20_D2


title: R9467 cmop

value: cmop


title: 1772158109_C08_B2


title: 1772158109_C07_B3


title: R9466 cmop


title: R9465 cmop


title: 1772158109_C03_A1


title: 1772158109_C05_A8


title: R9464 preneu

value: preneu


title: R9463 preneu


title: 1772158108_C96_H10


title: R9461 proneu2


title: 1772158108_C87_G4


title: 1772158108_C75_E4


title: 1772158108_C72_D12


title: R9458 proneu1


title: R9459 proneu2


title: 1772158108_C59_B11


title: R9456 proneu1


title: R9469 tpmo


title: R9457 proneu1


title: 1772158108_C45_H3


title: 1772158108_C53_A11


title: R9455 ly6c_neg_gmp


title: 1772158108_C42_G9


title: R9454 ly6c_neg_gmp


title: 1772158108_C38_G2


title: 1772158109_C01_A3


title: R9453 ly6c_neg_gmp


title: R9462 preneu


title: 1772158108_C37_G1


title: 1772158108_C84_F10


title: 1772158108_C28_E7


title: R9460 proneu2


title: 1772158108_C32_F2


title: 1772158108_C26_E2


title: 1772158108_C16_C9


title: 1772158108_C21_D1