title: Dies_Ant_B2

city: Austin

first: Anna

last: Battenhouse

address: 100 East 24th St., Stop A5000

department: Center for Biomedical Research Support

institute: The University of Texas at Austin

data_processing:
Fastq datasets were initially processed to collapse duplicates based on TagSeq molecular barcodes. Sequencing data quality, both before and after TagSeq pre-processing, was evaluated using the FastQC tool.
Paired-end pseudo alignment was performed using kallisto (ver. 0.45.0) with the options --single-overhang -l 200 -s 50, against the mouse transcriptome (GENCODE M23 transcript sequences).
Downstream analysis of kallisto estimated counts was performed in R (ver. 3.4.4).
The tximport package (v1.6.0) was used to roll up transcript-level counts into gene-level counts provided to the DESeq2 package (ver. 1.18.1).
Before DESeq2 analysis, count data matrices were filtered to remove genes with fewer than 1 read across all included samples.
Genome_build: GENCODE M23 transcriptome (transcript sequences and main gene annotation set)
Supplementary_files_format_and_content: Tab-delimited kallisto_counts.tsv text files provide kallisto per-transcript estimated counts and transcripts per million values for each sample. Full DESeq2 DEG and GO analysis details can be found in the Supplementary zip file.

source_name: Anterior oviduct, Diestrus stage

extract_protocol:
Dissected oviduct samples were collected in duplicate for each of the four stages across both anterior and posterior regions. Oviduct tissue was manually disrupted and the lysate spun through a QIAshredder column (Qiagen, cat#: 79656) to fully homogenize. Total RNA was isolated from each sample by addition of Trizol (Thermo Fisher) and the sample was transferred to a Phasemaker tube (Thermo Fisher). Total RNA was extracted following the protocol supplied by the manufacturer and further cleaned up using a RNeasy MinElute Cleanup Kit (Qiagen).
RNA integrity number (RIN) was measured using an Agilent Bioanalyzer and 100 ng of RNA was used for the TagSeq protocol. The fragmentation/RT mix was prepared and added to each RNA sample, then heated to 95°C for 2.5 minutes on a Thermocycler and immediately put on ice for 2 minutes. After cooling and addition of the template switching oligo and SmartScribe RT, the fragmented RNA reaction was incubated at 42°C for 1hr, 65°C for 15 min. Next an AmPure bead clean-up was completed for the cDNA before it was amplified to incorporate the Illumina sequencing primer site, followed by another cleanup. The remaining portions of the Illumina adapter (the i5 and i7 indices) were then added through an additional 4 cycles of PCR. Final libraries were quantified with PicoGreen then pooled equally for size selection using the Blue Pippin from 355-550 bp.

value: Swiss Webster

tag: Sex

value: female

value: oviduct

tag: location

value: anterior

tag: estrous cycle stage

value: diestrus


title: Dies_Post_B1

status: Public on Jan 15 2021

type: SRA

state: TX

country: USA

library_source: transcriptomic

molecule: polyA RNA

organism: Mus musculus

source_name: Posterior oviduct, Diestrus stage

tag: strain

tag: tissue

value: posterior


title: Dies_Ant_B1


title: Dies_Post_B2


title: Metes_Post_B1

source_name: Posterior oviduct, Metestrus stage

value: metestrus


title: Metes_Post_B2


title: Metes_Ant_B2

source_name: Anterior oviduct, Metestrus stage


title: Vanillin, rep2

type: RNA

city: Taichung

first: Chien-Yun

last: Hsiang

address: 91 Hsueh-Shih Road

country: Taiwan

institute: China Medical University

description: Mice were treated with AOM/DSS as sham group and orally with 100 mg/kg vanillin for 91 consecutive days.

hyb_protocol: Fluorescent targets were hybridized to Mouse Whole Genome OneArrayTM (Phalanx Biotech Group, Hsinchu, Taiwan).

scan_protocol: The slides were scanned by an Axon 4000 scanner (Molecular Devices, Sunnyvale, CA, USA).

data_processing: The Cy5 fluorescent intensity of each spot was analyzed by GenePix 4.1 software (Molecular Devices, Sunnyvale, CA, USA). The signal intensity of each spot was corrected by subtracting background signals in the surrounding. We filtered out spots that signal-to-noise ratio was less than 0 or control probes. Spots that pass aforementioned criteria were normalized by R program in the limma package.

molecule: total RNA

source_name: Colon, vanillin

label_protocol: Fluorescence-labeled RNA targets were prepared from 5 μg of total RNA using MessageAmpTM aRNA kit (Ambion, Austin, TX, USA) and Cy5 dye (Amersham Pharmacia, Piscataway, NJ, USA).

extract_protocol: Total RNAs were extracted from colon tissues using RNeasy Mini kit (Qiagen, Valencia, CA, USA).

treatment_protocol: On Day 1, mice were injected intraperitoneally with 12.5 mg/kg AOM. On Day 8, mice receiving the AOM injection were treated with 2.5% DSS in supplemented drinking water for 5 consecutive days, followed by normal drinking water for 16 days. DSS treatment was repeated for three additional cycles. In vanillin group, mice were treated with AOM/DSS as sham group and orally with 100 mg/kg vanillin for 91 consecutive days.

tag: strain/background

value: BALB/c

tag: gender

value: Female

value: Colon

tag: treatment

value: Vanillin


title: Mock, rep3

description: No treatment.

source_name: Colon, untreated


title: Mock, rep1

description: No treatment.


title: Sham, rep2

description: On Day 1, mice were injected intraperitoneally with 12.5 mg/kg AOM. On Day 8, mice receiving the AOM injection were treated with 2.5% DSS in supplemented drinking water for 5 consecutive days, followed by normal drinking water for 16 days. DSS treatment was repeated for three additional cycles.

source_name: Colon, sham

value: Sham


title: Metes_Ant_B1


title: Es_Post_B2

source_name: Posterior oviduct, Estrus stage

value: estrus


title: Sham, rep3


title: Sham, rep1


title: Vanillin, rep3


title: Vanillin, rep1


title: Mock, rep2

description: No treatment.


title: NT D0 4

city: Singapore

first: Henry

last: Yang

address: 28 Medical Drive

country: Singapore

institute: Cancer Science Institute of Singapore

description: Replicate 4

hyb_protocol: 750 ng of cRNA was hybridized to HumanHT-12 v4 Expression BeadChip (Illumina CA, USA) for 16h at 58°C. Arrays were then washed and stained with Streptavidin-Cy3 (GE Healthcare, CSP, UK).

scan_protocol: Arrays were scanned using Illumina BeadArray Reader (Illumina, CA, USA) at 25 scan factor 1

data_processing:
Raw expression data were obtained using the GenomeStudio Software. Background was subtracted and lower or negative expression was set to be 40 as p value for this intensity was at 0.05.
Normalization between samples was performed based the Cross-correlation method (Chua et al 2006, NAR) and  values were represent by expression intensities

label: biotin-labelled

organism: Homo sapiens

source_name: N/TERT1 undifferentiated keratinocytes

label_protocol: 500ng total RNA was reverse transcribed using ArrayScript™ Reverse Transcriptase (Illumina CA, USA) with T7oligo(dT) primer for 2h at 42°C. This was followed by an in vitro transcription of cDNA to biotin-labelled cRNA by T7 RNA polymerase in the presence of biotinylated ribonucleotides (Enzo Diagnostics, USA) at 37°C for 4h using the Illumina TotalPrep RNA Amplification kit (Applied Biosystems, CA, USA).

growth_protocol: The N/TERT-1 human immortalized keratinocyte cell line was obtained from James Rheinwald (Dickson et al., 2000). N/TERT-1 cells were maintained in keratinocyte serum-free media (K-SFM; Gibco®, Life Technologies, CA, USA), supplemented with 25µg/mL bovine pituitary extract (BPE), 0.2ng/mL epidermal growth factor (EGF) and 0.3mM CaCl2 at ~50-60% confluence. Differentiation was induced at high confluence by switching to high calcium (1.2mM CaCl2) media, as previously described (Rheinwald et al., 2002).

extract_protocol: RNA was harvested from cultured keratinocytes with TRIzol (Life Technologies, CA, USA) and purified on RNeasy columns (Qiagen, Germany)

treatment_protocol: 50µM EGBG was added at D3 of keratinocyte differentiation. Spd, Spm and Put (Sigma) were used at 1mM and added at day 3.

tag: cell line

tag: cell type/treatment

value: undifferentiated keratinocytes


title: lea13 T-DNA plants vs. Wild Type Col-0

city: Celaya

first: Gerardo

last: Acosta-Garcia

state: Guanajuato

address: Av. García Cubas Pte. # 600, Esquina Av. Tecnológico, Colonia Alfredo V. Bonfil

department: Ingeniería Bioquímica

country: Mexico

institute: Instituto Tecnológico de Celaya

hyb_protocol: For pre-hybridization, the slide was re-hydrated with water vapor at 60ºC and fixed with two cycles of UV ligth. Then, the slide was submerged in boiling water for two minutes (92ºC) and washed with 95% ethanol for one minute. After that, the oligo array was prepared for hybridization in a buffer solution 5X SSC, 0.1% SDS and 1% BSA for one hour at 41ºC. Finally, the array was washed and dried for hybridization. Equal quantities of labeled cDNA were hybridized to the Arabidopsis array using hybridization solution UniHyb (TeleChem International INC) for 14 hr at 42ºC.

scan_protocol: Scanned on an ScanArray 4000 scanner with ScanArray 4000 software from Packard BioChips.

data_processing:
The analysis of the microarray data: background correction, Lowess normalization, intensity filter and selection of differentially expressed genes, was performed with the software genArise (http://www.ifc.unam.mx/genarise/). The identification of differentially expressed genes was done by the calculation of a Z score using a sliding-window approach, where the algorithm calculates the mean and standard deviation (SD) within a window surrounding each data point to define a Z score, where Z is the number of SD a data point is from the window mean. To obtain the Z score, the next formula was applied, zi = (Ri - mean(R))/sd(R), where zi is the Z score for a given data point, Ri is the log ratio of signal intensity for each element, mean(R) and sd(R) are the mean and SD of the log ration of signal intensity of the surrounding window. Only the elements with an absolute Z score greater than 1.5 were considered differentially expressed (|Zscore| > 1.5)
The file processed_data_Cy3andCy5.txt includes the individual cy3 and cy5 values in addition to the zscore for each probe id.

organism: Arabidopsis thaliana

source_name: Cotyledons, 15 days old Wt plants

label_protocol: Total RNA were used for cDNA synthesis incorporating Alexa555 or Alexa647 making use of First-Strand cDNA labeling kit (Invitrogen). Incorporation of fluorophore was tested by measuring the absorbance at 555 nm for Alexa555 and 650 nm for Alexa647.

growth_protocol: Plants were initially grown on Murashige and Skoog (MS) agar plates in an incubator with 16 h light/8 h dark cycles for 7 a 10 days at 23°C and then transferred to soil for growth in a greenhouse at 25°C

extract_protocol: Total RNA was extracted from 15 days old frozen cotyledon tissue with the Trizol reagent (Invitrogen, Carlsbad, CA, USA), according to the manufacturer’s instructions. All RNA samples concentration were checked using a NanoDrop 2000 spectrophotometer. Approximately 500 ng was used for the later cDNA synthesis

treatment_protocol: Plants were subjected to normal growing conditions

tag: ecotype

tag: genotype

value: Wild Type

value: Cotyledon

source_name: Cotyledons, 15 days old lea13 plants

value: T-DNA insertion alleles for LEA13 (At2g18340; lea13)


title: Es_Post_B1

value: Swiss Webster


title: NT D6 + SDSM 4

description: Replicate 4

source_name: N/TERT1 day 6 differentiated keratinocytes + 1mM spermidine/spermine


title: NT D6 PUT 4

description: Replicate 4

source_name: N/TERT1 day 6 differentiated keratinocytes + 1mM putrescine


title: NT D6 + EGBG + SDSM 4

description: Replicate 4

source_name: N/TERT1 day 6 differentiated keratinocytes + EGBG  + 1mM spermidine/spermine


title: Multiplexed_Subcellular_Samples_File16

city: Frankfurt

first: Julio

middle: David

last: Perez

state: Hessen

department: Synaptic plasticity

country: Germany

institute: Max Planck Institute for Brain Research

data_processing:
bcl2fastq v2.20.0.422 to generate fastq files
Drop-seq tools v2.3.0 and Fastp for removal of adapter sequences and low quality reads
Reads were aligned against the RGSC 6.0/rn6 rat genome using STAR v2.5.4b
Gene assigment, and digital expression quantification per samples as done using Drop-seq tools v2.3.0
Genome_build: Rn6
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

organism: Rattus norvegicus

growth_protocol: The hippocampus of P1 rat pups or cortex and hippocampus of P0/P1 mice of either sex was dissected out, dissociated with papain and plated on poly-D- lysine coated coverslips of MatTek glass bottom dishes at a density of 20,000.

extract_protocol:
Dendrites and somata were dissected using LCM and collected in a buffer containing detergent and proteases to lyse the cell and release RNAs. RNAs were reverse transcribed, pre-amplified and prepared for Illumina sequencing.
Nextera XT DNA Library prep kit (Illumina)

value: Sprague Dawley

tag: days in vitro

value: wild type


title: NT D6 PUT 3

description: Replicate 3

value: day 6 differentiated keratinocytes + 1mM putrescine


title: Es_Ant_B2

source_name: Anterior oviduct, Estrus stage

value: female

value: oviduct

value: anterior


title: NT D6 + EGBG 4

description: Replicate 4

source_name: N/TERT1 day 6 differentiated keratinocytes + EGBG


title: NT D6 4

description: Replicate 4

source_name: N/TERT1 day 6 differentiated keratinocytes

value: day 6 differentiated keratinocytes


title: NT D6 + SDSM 3

description: Replicate 3

value: day 6 differentiated keratinocytes + 1mM spermidine/spermine


title: NT D6 + EGBG + SDSM 3

description: Replicate 3

value: day 6 differentiated keratinocytes + EGBG  + 1mM spermidine/spermine


title: NT D6 PUT 2

description: Replicate 2


title: NT D0 3

description: Replicate 3


title: NT D6 + EGBG 3

description: Replicate 3

value: day 6 differentiated keratinocytes + EGBG


title: NT D6 3

description: Replicate 3


title: Proes_Post_B2

source_name: Posterior oviduct, Proestrus stage

value: posterior

value: proestrus


title: Proes_Post_B1


title: liver_WD

city: Suita

first: Daisuke

last: Okuzaki

state: Osaka

address: Yamadaoka 3-1

department: Immunology Frontier Research Center

country: Japan

institute: Osaka univ.

description: liver of ob/ob mice fed western type diet for 4 weeks without 0.01% 7KC

data_processing:
Sequencing was performed on an Illumina HiSeq 2500 platform in a 75-base single-end mode.
Illumina Casava1.8.2 software was used for base calling.
The sequenced reads were mapped to the mouse reference genome sequences (mm10) using TopHat version 2.0.13 in combination with Bowtie2 version 2.2.3 and SAMtools version 0.1.19.
The fragments per kilobase of exon per million mapped fragments (FPKMs) were calculated using Cufflinks version 2.2.1.
Genome_build: mm10
Supplementary_files_format_and_content: FPKM processed value in csv file

extract_protocol:
Total RNA was isolated from the liver tissues using the Rneasy® Mini Kit (QIAGEN, Hilden, Germany)。
Library preparation was performed using a TruSeq stranded mRNA sample prep kit (Illumina, San Diego, CA) according to the manufacturer’s instructions.

value: liver

value: ob/ob mice

tag: diet

value: western


title: Es_Ant_B1

value: estrus


title: Proes_Ant_B2

source_name: Anterior oviduct, Proestrus stage

value: Swiss Webster


title: liver_CD_7KC

description: liver of ob/ob mice fed normal chow diet for 4 weeks with 0.01% 7KC

value: normal with 7-KC for 4 weeks


title: Proes_Ant_B1


title: liver_CD

description: liver of ob/ob mice fed normal chow diet for 4 weeks without 0.01% 7KC

value: normal


title: 575518_LM9

city: Basel

first: Mohamed

last: Bentires-Alj

address: Hebelstrasse 20

department: DBM

country: Switzerland

institute: University Hospital Basel

library_source: genomic

data_processing:
Library strategy: piggyBac-Seq
Standard Illumina software was used for base calling.
Mapping of insertion sequences to the mouse genome and identification of common integration sites (CIS). Pre-processing and alignment: Paired-End reads were first pre-processed by removing the expected transposon-derived sequence (5’-TAGGGTTAA-3’) from the beginning of the first read (read-pairs with non-matching first reads were discarded; typically around 1%) using the preprocessReads function from QuasR (version 1.12.0,33). Read pairs were then aligned to the mouse genome (BSgenome.Mmusculus.UCSC.mm10) using the QuasR qAlign function and parameters “-m 1 --best --strata --maxins 1000”; only uniquely mapping pairs with up to 1000 bp between-pair distance were reported. Mapping rates were recorded and non-mapped read pairs were further aligned against the non-mobilized transposon sequence to estimate the probable fraction of read pairs that originate from non-mobilized copies of the transposon. For each aligned read pair, the piggyBac insertion coordinate was identified as the coordinate of the first (most 5’) base of the first read.  Integration site identification and quantification: For each unique integration site, the number of distinct supporting alignments (distinct read pairs), and the genomic sequence from the four base pairs on the same strand as the first read directly upstream of the insertion site were recorded. Only integration sites with the expected upstream TTAA sequence were used for the downstream analysis.  Association of integration sites with genes: Coordinates of known genes (exons/introns, 5’-untranslated region [UTR], coding sequence [CDS], 3’UTR) were obtained from the TxDb.Mmusculus.UCSC.mm10.knownGene Bioconductor package (version 3.2.2). Promoter regions were defined as regions 2,000 bp upstream of known transcript start sites. Integration sites were matched against these genomic regions to identify overlaps on any strand and orientation, selecting the first overlap in the case of multiple overlaps. Integration sites were classified hierarchically as follows: Sites without overlaps to any transcript were labeled as promoter sites (in the case of an overlap with a promoter region) or intergenic sites. All other sites were labeled with the first region type that they overlapped, in the following order: 5’UTR, CDS, 3’UTR, intron, or ncRNA (defined as an overlap with a transcript without annotated CDS). Sites were further labeled according to their orientation with respect to the associated gene (same or opposite). Finally, sites were grouped according to the gene they overlapped (including promoter sites) or, for intergenic sites, according to the pair of flanking genes.  Selection of the enriched integration sites: For each CIS (all insertions associated to a single gene or genomic locus), the diversity (Div) was calculated by summing up the Div values of the individual insertions. Finally, this value was divided by the total number of different insertions for a given sample and converted to percentage. In order to find putative metastatic genes, we looked at genes with a higher number of insertions in the metastatic than in the tumour samples.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: processed_data_insertion_wise.csv
Supplementary_files_format_and_content: processed_data_gene_wise.csv

molecule: genomic DNA

source_name: breast cancer cell line

growth_protocol: HR1.Ctrl and HR1.PB cells (1,000,000) were resuspended in 50 μl Matrigel:PBS (1:1) and injected into the mammary fat pads of four- to eight-week-old female NOD/SCID mice. Tumours were resected and processed before they reached 1500 mm3. After tumour removal metastases were assessed allowing isolation of metastatic lungs and lines: 9 lung metastases (LM) cell lines and 3 lung-metastases were processed. LM1 and LM9 (250,000) were resuspended in 50 μl Matrigel:PBS (1:1) and injected into the mammary fat pads of four- to eight-week-old female NOD/SCID mice. 6 lung macro.metastases were isolated and processed.

extract_protocol:
Genomic DNA was isolated using DNeasy Blood and Tissue Kit (Qiagen).
Extracted genomic DNA was sheared to a fragment length of 250 bp with a Covaris sonicator. After end repair and A-tailing, purified DNA fragments were ligated to a splinkerette adaptor. Transposon-containing fragments were enriched by 18 cycles of transposon-specific PCR1 for the 5′ transposon ends in a unique library. Bar coding of individual samples and completion of Illumina adaptor sequences were achieved by an additional 12 cycles (primary tumours and LM cell lines) and 18 cycles (lung-metastases and lung macro-metastases) of transposon-specific PCR and a custom array of 35 unique bar-coding primers. After magnetic bead purification (Beckman Ampure XP), libraries were assembled in two pools and sequenced.

tag: breast cancer model

tag: mouse strain

value: NOD.CB17-Prkcd<scid>/J (NOD/SCID)

tag: sample type

value: Lung metastasis cell line


title: NT D6 + EGBG + SDSM 2

description: Replicate 2


title: 598079_LM9.Macromets

value: Lung macro-metastasis


title: Allograft tumor 3

city: Dallas

first: Thomas

last: Carroll

address: 6000 Harry Hines Blvd.

institute: UT Southwestern

data_processing:
Base calling was performed using bcl2fastq version 2.17.1.14.
Transcript abundance was estimated without aligning reads using Salmon against an index of coding sequences from the Ensembl GRCm38 assembly.
Transcript-level abundance was imported and count and offset matrices generated using the tximport R/Bioconductor package.
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited text file including FPKM values for each sample.

source_name: Allograft tumor

extract_protocol:
Dissected kidneys stored in RNA later solution (Invitrogen, Cat. AM7020). RNA was extracted with the QIAGEN's RNeasy Micro Kit (QIAGEN Cat. 74004).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: 1. A series of three injectons of tamoxifen at a dose of 5 mg/40 mg body weight was administered to one-month old Slc34a1CreERT2;Lats1/2flox/flox mice. 2. Lats1-/-;Lats2-/- cells were implanted by subcapsular injection of 6-8 week old NOD/SCID mice.

tag: strain background

value: mixed source;NOD/scid host

value: allograft tumor


title: Allograft tumor 2


title: Slc34a1CreERT2;Lats1c/c;Lats2c/+;RosaTomato proximal tubule 3

value: mixed

value: proximal tubule cells


title: Allograft tumor 1


title: 575515_LM8


title: 597079_LM9.Macromets2


title: 597079_LM9.Macromets1


title: 575518_LM9.Macromets


title: NT D6 + EGBG 2

description: Replicate 2

value: day 6 differentiated keratinocytes + EGBG


title: Slc34a1CreERT2;Lats1c/c;Lats2c/+;RosaTomato proximal tubule 2


title: Slc34a1CreERT2;Lats1c/c;Lats2c/+;RosaTomato proximal tubule 1


title: liver_WD_7KC

description: liver of ob/ob mice fed western type diet for 4 weeks with 0.01% 7KC

value: western with 7-KC for 4 weeks


title: NT D6 2

description: Replicate 2

value: day 6 differentiated keratinocytes


title: Slc34a1CreERT2;Lats1/2flox/flox lung metastasis 2

value: lung metasis


title: NT D6 PUT 1

description: Replicate 1

value: day 6 differentiated keratinocytes + 1mM putrescine


title: Slc34a1CreERT2;Lats1/2flox/flox lung metastasis 3


title: NT D6 + SDSM 2

description: Replicate 2

value: day 6 differentiated keratinocytes + 1mM spermidine/spermine


title: 571941_LM1.Macromets


title: 586941_LM1.Macromets


title: NT D6 + SDSM 1

description: Replicate 1


title: Slc34a1CreERT2;Lats1/2flox/flox lung metastasis 1


title: 575518_Tumor

value: Primary tumor


title: LM9_2

data_processing:
Standard Illumina software was used for base calling
Reads were aligned on the mm10 mouse genome using the R/Bioconductor package QuasR (with the unspliced option, using the TxDb.Mmusculus.UCSC.mm10.knownGene package for raw gene counts).
Genome_build: mm10

growth_protocol: HR1, LM1, LM9 (250,000 cells) were resuspended in 50 µl Matrigel:PBS (1:1) and injected into the mammary fat pads of four- to eight-week-old female NOD/SCID mice. Tumours were resected and processed before they reached 1500 mm3. Mice were monitored regularly for signs of metastatic outgrowth and distress. Sorted GFP+ cells from tumors were collected.

extract_protocol:
Total RNA was extracted using a Qiagene, RNeasy Plus Mini kit (cat. number 74136)
The library was prepared using Illumina TruSeq stranded mRNA-seq (according to recommendations from the manufacturer).


title: Slc34a1CreERT2;Lats1/2flox/flox primary tumor 2

value: kidney tumor


title: LM9_3


title: Slc34a1CreERT2;Lats1/2flox/flox primary tumor 3


title: Slc34a1CreERT2;Lats1/2flox/flox primary tumor 1


title: NT D6 + EGBG + SDSM 1

description: Replicate 1

value: day 6 differentiated keratinocytes + EGBG  + 1mM spermidine/spermine


title: NT D0 2

description: Replicate 2

value: undifferentiated keratinocytes


title: 575515_Tumor

value: NOD.CB17-Prkcd<scid>/J (NOD/SCID)


title: NT D0 1

description: Replicate 1


title: NT D6 + EGBG 1

description: Replicate 1


title: LM9_1


title: LM1_3


title: LM1_2


title: HF/TZD KO 2

city: Chicago

first: Jose

last: Cordoba-Chacon

state: IL

address: 835 S Wolcott Ave

department: Medicine

institute: University of Illinois at Chicago

data_processing:
Sequencing Ilumina HiSeq PE150 Platform 6G/sample Q3>95%
Mapping with STAR v 2.6.1
Analsyis of gene expression Htseq v0.6.1
Differential expression analysis DESeq2 R package v2_1.6.3
Enrichment analysis clusterProfiler R package
Genome_build: Mouse reference genome sequence (GRCm38/mm10)
Supplementary_files_format_and_content: These files are the differential expression analysis. The columns are: Gene ID; Values of each mouse; log2FoldChange; p-value; p adjusted; Gene Name; Chromosome; Strand; Start; End; Gene Length; Gene Description; Values of each mouse (reads counts); Values of each  mouse (FPKM)

source_name: Liver

growth_protocol: Adult mice were fed a LFCF diet (LF, D091000304, Research Diets) at libitum, or a HFCF diet (HF, D16010101, Research Diets). Half of the mice in a HFCF diet were switched to a HFCF diet containing rosiglitazone maleate (HF/TZD, D18080301, Research Diets) in the last 8 weeks of the experiment.

extract_protocol:
Whole liver RNA was extracted using Trizol reagent. RIN>7.5 to 9.3
250-300bp insert cDNA libraries non strand specific.

treatment_protocol: Half of the mice in the HFCF diet or HFCF diet with rosiglitazone maleate (HF/TZD) were hepatocyte-specific PPARgamma knockout. The KO of this gene in hepatocytes was induced after 24 weeks of HFCF diet.

value: Hepatocyte-specfic KO of PPARgamma in hepatocytes after NASH

value: TZD (Rosiglitazone maleate)

tag: disease

value: Non-alcoholic steatohepatitis

value: high fat

tag: ID


title: HR1_3


title: HF/TZD C 5

value: Control mouse


title: HR1_1


title: HF/TZD KO 1


title: HF/TZD KO 4


title: 052_CD8_67_LP

city: San Diego

first: Joseph

middle: S

last: Dolina

state: CA

address: 9420 Athena Cir

institute: La Jolla Institute of Immunology

data_processing:
Reads were mapped against the mouse mm10 reference using STAR v2.4.1c
Sequencing read coverage per gene was counted using HTSeq-count v0.7.1 (-m union -s no -t exon -i gene_name -r name)
Genome_build: mm10

source_name: splenocytes and tumor draining lymph node lymphocytes

extract_protocol:
This assay was adapted from the Smart-seq2 protocol (7).  Single-cells were sorted into 96-well V-bottom PCR plates containing 4 μL cell lysis buffer composed of 0.2% Triton X-100, 1 U/μL recombinant RNase inhibitor (Takara Bio, Kusatsu, Shiga, Japan), 2.5 mM dNTP mix (Life Technologies, Carlsbad, CA).  FACS-sorting was performed with a calibrated sort stream to ensure single-cells landed on the liquid interface.
RNA loss was minimized by performing on-plate RNA reverse-transcription and whole-transcriptome amplification to generate ~1-10 ng cDNA.  Briefly, mRNA was captured using poly-dT oligos and directly reverse-transcribed into full-length cDNA using the described template-switching LNA oligo (7).  Whole-transcriptome cDNA was amplified by PCR for 24 cycles and purified using Agencourt AMPure XP magnetic beads at 0.9:1 (v:v) 2× (Beckman Coulter).  The number of PCR cycles for optimal cDNA amplification was determined by Q-PCR.  Quality and quantity of cDNA amplification were assessed by Fragment Analyzer Automated CE System (Advanced Analytical, Ankeny, IA) and PicoGreen dsDNA intercalating-dye based assay (Invitrogen).  Samples that failed quality control steps as described (8) were eliminated from downstream steps.  0.4 ng of cDNA was used to prepare a sequencing library with the Nextera XT DNA Library Prep and Index Kit (Illumina), using a Biomek FXP automated platform (Beckman Coulter).  Before sequencing, all libraries were purified using Agencourt AMPure XP beads and quality control steps were performed (8).  Samples that passed strict quality controls were pooled at equimolar concentration, loaded on flow cells and sequenced on the Illumina HiSeq 2500 in Rapid Run mode (Illumina) to obtain more than 0.3×106 125×125 bp paired-end reads, generating a mean/median of ~2.0×106 filtered uniquely mapped reads per sample.

tag: tissue type

value: lymphocytes


title: HF/TZD C 4


title: HF/TZD C 3


title: HR1_2


title: HF/TZD KO 3


title: HF/TZD C 2


title: HF/TZD C 1


title: HF KO 5

value: no treatment


title: HF/TZD KO 5


title: HF KO 4


title: 042_CD4_65_LP


title: HF KO 3


title: 038_CD4_67_LP


title: LM1_1


title: HF KO 2


title: 068_CD8_67_LP


title: 001_CD4_65_LP


title: data_Input4_Total  (WT 9J8h)

city: Leiden

first: Martijn

middle: S.

last: Luijsterburg

state: zuid-holland

address: Einthovenweg 20

department: Human Genetics

country: Netherlands

institute: LUMC

data_processing:
ChIP-seq: Reads were aligned to the Human Genome 38 (Hg38) using bwa-mem tools (Li, 2013) during which only high-quality reads (> q30) were included in the analyses and duplicates were removed. Bam files were converted into TagDirectories using HOMER tools (Heinz et al., 2010). Binding profiles within selected areas of individual genes (e.g. around TSS or TTS), were defined using the AnnotatePeaks.pl tool of HOMER using the default normalization to 10mln reads. Metagene profiles were defined using the makeMetaGeneProfile.pl tool of HOMER, using default settings. Individual datasets were subsequently processed in R (Team, 2019). BrU-seq: Data was processed as previously described (Paulsen et al., 2014).  Briefly, reads were pre-filtered by alignment to the human ribosomal repeating subunit (GenBank U13369.1) and human mitochondrial genome (chrM) from the hg38 reference genome using bowtie2 v.2.3.3.1. The remaining reads were mapped to the hg38 reference genome using STAR v2.7.0f. Base coverages were used to compute read counts for features, such as genes and bins, which were then normalized to feature length and number of uniquely-mapped reads (RPKM method). Gene selection was based on the following criteria: length as indicated per figure, TSS is at least 10kb apart, expression is at least 0.05 RPKM. The median expression was calculated for each 500 bp bins from 5kb upstream until 100kb downstream of the selected genes for each time-point.
Genome_build: hg38
Supplementary_files_format_and_content: excel tables providing avearged read coverage around the TSS or normalized traverling ratios for the top 3000 genes for each individual sample, and the BrU-seq counts versus distance from the TSS for genes of 25-50Kb, 50-100Kb and >100Kb

source_name: U2OS WT

growth_protocol: DMEM GlutaMAX +10% Fetal Calf Serum + antibiotics

extract_protocol:
ChIP-seq: Cells were plated and grown to ~90% confluency and crosslinked with 0.5 mg/mL disuccinimidyl glutarate (DSG; Thermo Fisher) in PBS for 45 min at room temperature. Cells were washed with PBS and crosslinked with 1 % PFA for 20 min at room temperature. Fixation was stopped by adding 1.25 M Glycin in PBS to a final concentration of 0.1 M for 3 minutes at room temperature. Cells were washed with cold PBS and lysed and collected in a buffer containing 0.25% Triton X-100, 10 mM EDTA (pH 8.0), 0.5 mM EGTA (pH 8.0) and 20 mM Hepes (pH 7.6). Chromatin was pelleted in 5 min at 400 g and incubated in a buffer containing 150 mM NaCl, 1 mM EDTA (pH 8.0), 0.5 mM EGTA (pH 8.0) and 50 mM Hepes (pH 7.6) for 10 minutes at 4°C. Chromatin was again pelleted for 5 min at 400 g and resuspended in ChIP-buffer (0.15 % SDS, 1 % Triton X-100, 150 mM NaCl, 1 mM EDTA (pH 8.0), 0.5 mM EGTA (pH 8.0) and 20 mM Hepes (pH 7.6)) to a final concentration of 15x106 cells/ml. BrU-seq: U2OS osTIR1 or PAF1-AID cells were induced with doxycycline for 24 hrs, and subsequently with auxin for 5 hrs. After this treatment, cells were either mock-treated or irradiated with UV-C light (7 J/m2). Cells were then incubated in conditioned media for different periods of time (0, 3, 8, 24h) before being incubated with 2 mM bromouridine (Bru) at 37°C for a 30 min. The cells were then lysed in TRIzol reagent (Invitrogen) and Bru-containing RNA isolated as previously described (Andrade-Lima et al., 2015).
ChIP-seq: Chromatin was sonicated to approximately 1 nucleosome using a Bioruptor waterbath sonicator (Diagenode). Chromatin of ~5x106 cells was incubated with 3ug antibody (RNAPII, rabbit polyclonal, Bethyl laboratories, A304-405A; PAF1, rabbit polyclonal, Bethyl laboratories, A300-172A) over night at 4°C, followed by a 1.5 hrs protein-chromatin pull-down with a 1:1 mix of protein A and protein G Dynabeads (Thermo Fisher; 10001D and 10003D). ChIP samples were washed extensively, followed by decrosslinking for 4 hrs at 65°C in the presence of proteinase K. DNA was purified using a Qiagen MinElute kit. Sample libraries were prepared using Hifi Kapa sample prep kit and A-T mediated ligation of Nextflex adapters. Samples were sequenced using an Illumina NextSeq500, using paired-end sequencing with 42 bp from each end. BrU-seq: cDNA libraries were made from the Bru-labeled RNA using the Illumina TruSeq library kit and paired-end 151 bp sequenced using the Illumina NovaSeq platform at the University of Michigan DNA Sequencing Core. Only single-end sequencing data was used for downstream analyses

treatment_protocol: DNA damage was induced by irradiating cells with varying doses of UV-C as indicated per sample; samples were assayed at different time-periods after UV treatment as indicated per sample; to deplete PAF1 using the auxin-inducible degron system cells were induced with 2ug/ml doxycycline for 24 hrs and pre-treated with 500uM auxin for 5hours prior to UV-C irradiation or harversting.

tag: cell type

tag: genotype/variation

value: WT

tag: uv-c dose

tag: time after uv-c

value: 8 hours

tag: chip-antibody

value: not applicable


title: 065_CD8_65_LP


title: HF C 4

tag: treatment


title: 004_CD4_67_LP


title: 057_CD8_65_LP


title: 022_CD4_67_LP


title: HF C 3


title: HF KO 1


title: HF C 5


title: HF C 2


title: 036_CD4_67_LP


title: 033_CD4_65_LP


title: HF C 1


title: 026_CD4_65_LP


title: 089_CD8_65_LP


title: 010_CD4_65_LP


title: 073_CD8_65_LP


title: LF C 2

value: Healthy mouse

value: low fat


title: LF C 3


title: NT D6 1

description: Replicate 1

tag: cell line

tag: cell type/treatment

value: day 6 differentiated keratinocytes


title: 028_CD4_67_LP


title: LF C 1

tag: genotype


title: 009_CD4_65_LP


title: 045_CD4_67_LP


title: HF-KO-5

data_processing:
The expression level analysis was done with HTSeq v0.6.1 to calculate the FPKM of each gene  based on the length of the gene and reads count mapped to this gene. To allow for log adjustment, genes with 0 FPKM are assigned a value of 0.001. Correlation were determined using the cor.test function in R with options set alternative ="greater" and method = "Spearman".
The differential expression analysis between two different groups was done with DESeq2 R package (v2_1.6.3). DESeq2 provide statistical routines for determining differential expression in digital gene expression data using a model based on the negative binomial distribution. The resulting P-values were adjusted using the Benjamini-Hochberg's approach for controlling the False Discovery Rate (FDR). TFCat and Cosmic database were used to annotate the differential expressed gene.
The enrichment analysis was performed with ClusterProfiler v2.4.3.

growth_protocol: Adult mice were fed a LFCF diet (LF, D091000304, Research Diets) at libitum, or a HFCF diet (HF, D16010101, Research Diets).

extract_protocol:
Whole liver RNA was extracted using Trizol reagent. RNA with RIN>7 was selected for RNAseq
The 250-300bp insert cDNA libraries, non strand-specific, were prepared with NEBNext(r) UltraTM RNA Library Prep Kit. Libraries were sequenced in Ilumina HiSeq PE150 Platform ~6G/sample Q30>90%. RNAseq reads were mapped to the mouse reference genome sequence using STAR (v2.5) with a total mapping rate >90%/sample.

treatment_protocol: Half of the mice in the LFCF or HFCF diet were hepatocyte-specific PPARgamma knockout. The KO of this gene in hepatocytes was induced before the 24 weeks of HFCF diet.

tag: control or ko

value: Hepatocyte-specfic KO of PPARgamma

tag: diet


title: 041_CD4_65_LP


title: HF-KO-6


title: HF-KO-4


title: HF-KO-3


title: 014_CD4_67_LP


title: 029_CD4_67_LP


title: HF-KO-1


title: HF-C-5


title: HF-KO-2


title: 070_CD3_contrl_


title: 005_CD4_67_LP


title: 081_CD8_65_LP


title: 013_CD4_67_LP


title: 093_CD8_67_LP


title: HF-C-2


title: 084_CD8_67_LP


title: 076_CD8_67_LP


title: 025_CD4_65_LP


title: LF-KO-4


title: HF-C-1


title: 002_CD4_65_LP


title: LF-KO-3


title: 092_CD8_67_LP


title: LF-KO-2


title: 066_CD8_65_LP


title: LF-C-4


title: 062_CD3_contrl_


title: LF-C-3


title: 020_CD4_67_LP


title: LF-C-2


title: HF-C-4


title: 090_CD8_65_LP


title: HF-C-3


title: LF-C-1


title: 074_CD8_65_LP


title: 082_CD8_65_LP


title: 058_CD8_65_LP


title: RGC that have been transduced with SOX11_MUTANT, replicate2

city: Palo Alto

first: Jeffrey

middle: L

last: Goldberg

address: 1651 Page Mill Road

department: Opthalmology

institute: Stanford University

data_processing:
The RNA-seq reads were aligned to the mouse genome mm10 with the RNA computational pipeline on the DNAnexus cloud computing platform comprised of the STAR alignment package and RSEM counting algorithm. Each sample (control, SOX11 wildtype, and SOX11 K91A ) had 2 replicates.
The differentially expressed genes were called by using limma-voom in R/BioC on the RNA-seq replicates, and the genes with a FDR < 0.05 and an absolute fold change (FC) > 1.2 were considered for down-stream analysis.
The functional enrichment of differentially expressed genes was computed in enrichR, and volcano plots were displayed in R.
Genome_build: mm10
Supplementary_files_format_and_content: rsem (counts, fpkm, tpm)

source_name: RGC

growth_protocol: Mouse RGCs from postnatal day 2 (P2) pups were purified by immunopanning using CD90.1 as described previously (Barres et al., 1988). RGCs were plated onto PDL/laminin-coated tissue culture plates in Full Sato (FS) medium including forskolin (5 mM), BDNF (50 ng/ml), and CNTF (10 ng/ml) as described previously (Meyer-Franke et al., 1995). Plating densities were 2.5x10 3 RGCs/well in 24-well plates for neurite outgrowth assay, or 2.5x10 5 RGCs/well in 6-well plates for RNA sequencing and Western blot assays.

extract_protocol:
For viral transduction of E18 RGCs for RNA sequencing, 2.5x10^5 RGC/well were plated on PDL/laminin-coated 6-well plates in quadruplicates. Four hours after plating, 1.5 μl AAV (2x10^13 vector genome (vg)/ml) was added to cultured cells at a MOI of about 10^5 vg/cell. Full medium changes were performed 5.5 and 24 hours after virus exposure. Total RNA was extracted using the RNEasy kit (QIAgen) 72 hours after plating.
Library construction and next generation sequencing was performed at Genewiz on 3 RNA samples (500 ng/sample) in duplicates.

treatment_protocol: For viral transduction in neurite outgrowth assay, Sox11 and Sox11 K91A AAV2 viruses were diluted to 2x10 11 vector genomes (vg)/ml in medium and added at a multiple of infection (MOI) of about 10^5 vg/cell overnight, and then changed into fresh culture medium for another 2 days. RGCs were immunstained with antibodies against β-III-tubulin. Briefly,equal amount 37°C 4% paraformaldehyde (PFA) was added into the medium for 20 min for fixation. The neurites were stained with mouse anti-β-III-tubulin antibody E7 (1:500, hybridoma from Developmental Studies Hybridoma Bank) overnight at 4°C and probed with Alexa Fluor 647-tagged anti-mouse antibody (1:500; Life technologies) for 2 hours at room temperature. GFP+ and DAPI+ RGCs were imaged on a Zeiss Axio Observer inverted microscope using a 10× objective to measure neurite outgrowth and survival, respectively. Total length of neurites per cell (30~60 cells average per condition per experiment) was measured using ImageJ Neurite Tracer.

tag: transduction


title: 012_CD4_67_LP


title: RGC that have been transduced with SOX11_WT, replicate2


title: RGC that have been transduced with SOX11_MUTANT, replicate1


title: 044_CD4_67_LP


title: 037_CD4_67_LP


title: RGC that have been transduced with CONTROL, replicate1

value: CONTROL


title: RGC that have been transduced with CONTROL, replicate2


title: RGC that have been transduced with SOX11_WT, replicate1


title: 006_CD4_67_LP


title: LF-KO-1


title: 030_CD4_67_LP


title: M. albugensis + SynCom Peeled Rep3

city: Cologne

first: Gunther

last: Doehlemann

address: Zülpicher Straße 47a

department: CEPLAS / Institute of Botany

institute: University of Cologne

data_processing:
Sequencing read quality control was performed using FastQC version 0.11.5
Sequenced reads were mapped to the genome of M. albugensis using tophat v2.0.10
Read counts per gene were extracted by using HTSeq version 0.9.0
Genome_build: Moesziomyces albugensis WT v.2 (currently in submission).
Supplementary_files_format_and_content: tab delimited .txt file with raw read counts for each predicted M. albugensis transcript, samples in columns

organism:
Arabidopsis thaliana
Moesziomyces sp. GD-2020a

source_name: Latex-peeled microbes from plant leaves, 4dpi

growth_protocol: For axenic culture samples and prior to inoculation M. albugensis was grown in liquid YEPSlight medium at 22°C and 200rpm. Bacterial SynCom member were grown in KingsB media under the same conditions. A. thaliana seedlings were grown under short day conditions (22°C/18°C 8h day/16h night).

extract_protocol:
Four weeks old A. thaliana plants were fixed and liquid latex was applied to the leaf surface. The latex was dried using the cold air option of a hair dryer, carefully peeled off  and immediately frozen in liquid nitrogen. Afterwards the frozen latex pieces were grinded with liquid nitrogen and the RNA was isolated by using Trizol® Reagent (Invitrogen) according to the manufacturer’s instructions.DNA contaminations were removed by using Turbo DNA-Free™ Kit (Ambion).
RNA libraries were prepared by the CCG (Cologne genome center) using standard Illumina protocols

treatment_protocol: 3-weeks old seedlings were spray inoculated, each with 500µl of M. albugensis/ SynCom members (OD600 0.6) solved in MgCl2. After two days plants are challenged with the oomycete pathogen A. laibachii. Therefore zoospores were harvested from infected plants, washed and set to a spore number of 16*10^4 spores/ml. 500µl are evenly sprayed to each plant. Plants that are not infected with the pathogen were sprayed with MgCl2. All samples were incubated at 4°C over night with 100% humidity and moved to the growth chamber on the next day. 24h after moving samples were harvested.

tag: host plant cultivar

tag: biotic interaction

value: M. albugensis, A. thaliana, bacterial SynCom

tag: tissue

value: Latex-peeled microbes from plant leaves


title: 034_CD4_65_LP


title: 054_CD8_67_LP


title: M. albugensis + SynCom Peeled Rep1


title: LF C 4

tag: disease

tag: ID


title: 021_CD4_67_LP

tag: tissue type


title: 017_CD4_65_LP


title: 020_CD4_44_LP


title: M. albugensis Peeled Rep3

value: M. albugensis, A. thaliana


title: M. albugensis Peeled Rep1


title: 090_CD8_61_SP


title: 046_CD4_67_LP


title: 081_CD8_48_SP


title: 062_CD8_61_LP


title: E– 24 hpf sample 4 [sample_24_lane8]

city: Corvallis

first: Maret

last: Traber

state: OR

address: 2900 SW Campus Way

department: Linus Pauling Institute

institute: Oregon State University

description: lane8-s030-index-AGACCA-Sample-24_S30_L008

data_processing:
Sequence read quality was first assessed using FastQC v0.11.5. Adapters and poly(A) tails were trimmed using bbduk v35.92.
Trimmed reads were then aligned to the current reference genome (GRCz11, D. rerio) with current annotations (Ensembl 100) using STAR aligner, v2.7.3a.
Genome_build: GRCz11
Supplementary_files_format_and_content: tab-delimited text file includes CPM

organism: Danio rerio

source_name: E– 24 hpf_whole embryo lysate

growth_protocol: Tropical 5D strain zebrafish were reared in Sinnhuber Aquatic Research Laboratory at Oregon State University under standard laboratory conditions of 28°C on a 14 h light/10h dark photoperiod according to standard zebrafish breeding protocols.

extract_protocol:
RNA was extracted from embryos staged developmentally at 12, 18 and 24 hpf, pooled (n=10 embryos/sample) and homogenized; 4 sample pools per VitE status. RNA integrity (9.02  0.15, n=16) was assessed using the 2100 BioAnalyzer Instrument (Agilent) at the Center for Genome Research and Biocomputing at Oregon State University.
RNA libraries were prepared with the Lexogen QuantSeq 3’mRNA Seq Library Prep Kit-FWD kits (Lexogen, Vienna, Austria). After library preparation, samples were pooled and sequenced using single-end sequencing with 100 bp reads on an Illumina HiSeq3000 instrument (Illumina, San Diego, CA).

treatment_protocol: At 55 days post-fertilization (dpf), adult zebrafish were randomly allocated to two experimental diets, vitamin E deficient (E–) or sufficient (E+)]. E– and E+ embryos were obtained by group spawning of adult fish fed either E– or E+ diets for a minimum of 80 days up to 9 months. Embryos were collected, staged and incubated until use in standard embryo media [15 mM NaCl, 0.5 mM KCl, 1 mM MgSO4, 0.15 mM KH2PO4, 0.05 mM Na2HPO4, 1 mM CaCl2, NaHCO3 in fish system water, (EM)].

tag: strain

value: E–

value: Whole embryo


title: 010_CD4_61_SP


title: E+ 24 hpf sample 4 [sample_20_lane8]

description: lane8-s028-index-AAGTGG-Sample-20_S28_L008

source_name: E+ 24 hpf_whole embryo lysate

value: E+


title: E– 24 hpf sample 3 [sample_23_lane8]

description: lane8-s029-index-GGCCAA-Sample-23_S29_L008


title: E– 24 hpf sample 1 [sample_21_lane7]

description: lane7-s017-index-CTCATA-Sample-21_S17_L007


title: E+ 24 hpf sample 3 [sample_19_lane8]

description: lane8-s027-index-TGCTAT-Sample-19_S27_L008


title: M. albugensis + SynCom Peeled Rep2


title: 095_CD8_61_LP


title: E+ 24 hpf sample 2 [sample_18_lane7]

description: lane7-s016-index-ACTCTT-Sample-18_S16_L007


title: E+ 24 hpf sample 1 [sample_17_lane7]

description: lane7-s015-index-GCAGCC-Sample-17_S15_L007


title: 033_CD4_48_SP


title: 093_CD8_48_LP


title: 068_CD8_44_LP


title: E– 18 hpf sample 3 [sample_15_lane8]

description: lane8-s025-index-CATCTA-Sample-15_S25_L008

source_name: E– 18 hpf_whole embryo lysate


title: 037_CD4_48_LP


title: E– 18 hpf sample 4 [sample_16_lane8]

description: lane8-s026-index-AACAAG-Sample-16_S26_L008


title: 003_CD4_65_SP


title: 005_CD4_48_LP


title: M. albugensis Peeled Rep2


title: E– 18 hpf sample 1 [sample_13_lane7]

description: lane7-s013-index-ATTGGT-Sample-13_S13_L007


title: E+ 18 hpf sample 4 [sample_12_lane8]

description: lane8-s024-index-ATGGCG-Sample-12_S24_L008

source_name: E+ 18 hpf_whole embryo lysate


title: 063_CD8_61_LP


title: E+ 18 hpf sample 3 [sample_11_lane8]

description: lane8-s023-index-CGTCGC-Sample-11_S23_L008


title: E– 18 hpf sample 2 [sample_14_lane7]

description: lane7-s014-index-GCCACA-Sample-14_S14_L007


title: 035_CD4_65_SP


title: 078_CD8_61_LP


title: E+ 18 hpf sample 1 [sample_9_lane7]

description: lane7-s011-index-GACATC-Sample-9_S11_L007


title: E– 24 hpf sample 2 [sample_22_lane7]

description: lane7-s018-index-CCGACC-Sample-22_S18_L007


title: 023_CD4_61_LP


title: 069_CD8_48_LP


title: E– 12 hpf sample 3 [sample_7_lane8]

description: lane8-s021-index-CAGATG-Sample-7_S21_L008

source_name: E– 12 hpf_whole embryo lysate


title: E– 12 hpf sample 2 [sample_6_lane7]

description: lane7-s010-index-AACCGA-Sample-6_S10_L007


title: 074_CD4_61_SP


title: 038_CD4_61_LP


title: E+ 18 hpf sample 2 [sample_10_lane7]

description: lane7-s012-index-CGATCT-Sample-10_S12_L007


title: E+ 12 hpf sample 3 [sample_3_lane8]

description: lane8-s019-index-TCGTTC-Sample-3_S19_L008

source_name: E+ 12 hpf_whole embryo lysate


title: 065_CD8_48_SP


title: E– 12 hpf sample 4 [sample_8_lane8]

description: lane8-s022-index-GTAGAA-Sample-8_S22_L008


title: 027_CD4_65_SP


title: E+ 12 hpf sample 1 [sample_1_lane7]

description: lane7-s007-index-AAGCTC-Sample-1_S7_L007


title: E+ 12 hpf sample 4 [sample_4_lane8]

description: lane8-s020-index-CCAATT-Sample-4_S20_L008


title: 089_CD8_48_SP


title: 007_CD3_control_control


title: 086_CD8_61_LP


title: MSC dervived Adipocyte donor 8002 [Capture C]

city: Philadelphia

first: Matthew

middle: C

last: Pahl

state: PA

address: 3615 Civic Center Blvd

department: Center for Spatial and Functional Genomics

institute: Children's Hospital of Philadelphia

data_processing:
library strategy: Capture C
Bases were called using Illumina bcl2fastq2 conversion v2.29
Paired-end reads from three replicates from MSC-dervived Adipocytes were pre-processed using the HICUP pipeline (v0.5.9) with the default parameters. Reads were aligned to hg19 using bowtie2. We called call significant promoter interactions using the read count from promoters included in our reference bait.
Significant interactions were called CHiCAGO (v1.1.8) with default parameters except for bin-size set to 2,500. In addition to our analysis per individual DpnII fragment (1frag), we also called interactions by binning four fragments, which improves detection of long-distance interactions. Significant interactions at 4-DpnII fragment resolution were also called using CHiCAGO with artificial baitmap and .rmap files where four DpnII fragments were concatenated. Interactions with a CHiCAGO score > 5 in at least one cell type in either 1-fragment or 4-fragment resolution were considered significant.
Genome_build: hg19
Supplementary_files_format_and_content: ibed

source_name: Adipocytes dervived from MSCs from donor 8002

growth_protocol: Primary bone-marrow derived human MSCs isolated from healthy donors (age range: 22 –29 years) were characterized for cell surface expression (CD166+CD90+CD105+/CD36−CD34−CD10−CD11b−CD45−) at the Institute of Regenerative Medicine, Texas A&M University. Expansion and maintenance of the cells were carried out using alpha-MEM supplemented with 16.5% FBS in standard culture conditions by plating cells at a density of 3000 cells/cm^2.

extract_protocol:
Cells were fixed after 2 weeks of adipogenic induction. Cells were collected and single-cell suspension were made with aliquots of 10^7 cells in 10 ml media (e.g. RPMI + 10%FCS). 540 µl 37% formaldehyde was added and incubated for 10 min at RT. The reaction was quenched by adding 1.5 ml 1 M cold glycine (4 °C) for a total volume of 12 ml. Fixed cells were centrifuged at 1000 rpm for 5 min at 4 °C and supernatant removed. The cell pellets were washed in 10 ml cold PBS (4 °C) followed by centrifugation. Supernatant was removed and cell pellets were resuspended in 5 ml of cold lysis buffer (10 mM Tris pH8, 10 mM NaCl, 0.2% NP-40 (Igepal) supplemented with protease inhibitor cocktails). Resuspended cells were incubated for 20 min on ice, centrifuged as above, and the lysis buffer removed. Finally, cell pellets were resuspended in 1 ml fresh lysis buffer, transferred to 1.5 ml Eppendorf tubes and snap frozen (ethanol/dry ice or liquid nitrogen). Cells were stored at −80 °C until they were thawed for 3C library generation.
Samples underwent a pre-digestion incubation with the addition of 0.3% SDS, 1x NEBuffer DpnII, and dH2O for 1hr at 37ºC in a Thermomixer shaking at 1,000rpm. A 1.7% solution of Triton X-100 was added to each tube and shaking was continued for another hour. After the pre-digestion incubation, 10 μL of DpnII (was added only to each sample tube, and continued shaking until the end of the day. An additional 10 µL of DpnII was added to each digestion reaction and digestion continued overnight. The next day, another additional 10 µL of DpnII was added and the incubation continued for another 2-3 hours. 100 μL of each digestion reaction was then removed, pooled into one 1.5 mL tube. The remaining samples were heat inactivated at 65°C for 20 min at 1000 rpm in a Thermomixer, and cooled on ice for 20 additional minutes. Digested samples were ligated with 8 μL of T4 DNA ligase (HC 30U/uL) and 1X ligase buffer at 1,000 rpm overnight at 16°C in a Thermomixer. The next day, an additional 2 µL of T4 DNA ligase was spiked into each sample and incubated for another few hours. The ligated samples were then de-crosslinked overnight at 65°C with Proteinase K along with the pre-digestion and digestion controls. The following morning, both controls and ligated samples were incubated for 30 min at 37°C with RNase A, followed by phenol/chloroform/isoamyl alcohol extraction and ethanol precipitation at -20°C. The 3C libraries were centrifuged at 3000 rpm for 45 min at 4°C 14,000 rpm, to pellet the samples. DNA pellets were resuspended in 70% ethanol and again centrifuged as described above. The 3C library pellets were resuspended in 300 μL dH2O and stored at −20°C. 10 μg of isolated DNA from 3C libraries of each library was sheared in dH2O using a QSonica Q800R to an average fragment size of 350bp.  After shearing, DNA was purified using AMPure XP beads. One microgram of adaptor-ligated library was used as input for the SureSelect XT capture kit using manufacturer protocol and our custom-designed 41K promoter Capture-C probe set.

treatment_protocol: Adipogenic differentiation of the hMSCs was carried out 2 days after seeding using standard procedure for human cells. Briefly, cell monolayers were washed twice with phosphate buffered saline and supplemented with alpha-MEM consisting of 8.25% FBS, 0.5 µM water-soluble Dexamethasone, 0.5 µM isobutylmethylxanthine, and 50 µM Indomethacin. Alpha-MEM consisting of 8.25% FBS with or without supplementation were exchanged every 2–3 days until harvested.

tag: subject status/id

value: healthy donor 8002

tag: cell type

value: Adipocytes dervived from mesenchymal stem cells (MSCs)


title: MSC dervived Adipocyte donor 8004 [Capture C]

source_name: Adipocytes dervived from MSCs from donor 8004

value: healthy donor 8004


title: 002_CD4_61_SP

value: lymphocytes


title: 042_CD4_61_SP


title: E+ 12 hpf sample 2 [sample_2_lane7]

description: lane7-s008-index-GACGAT-Sample-2_S8_L007


title: 006_CD4_61_LP


title: MSC dervived Adipocyte donor 7043 [Capture C]

source_name: Adipocytes dervived from MSCs donor 7043

value: healthy donor 7043


title: 025_CD4_48_SP


title: 091_CD8_48_SP


title: E– 12 hpf sample 1 [sample_5_lane7]

description: lane7-s009-index-AGTTGA-Sample-5_S9_L007


title: MSC dervived Adipocyte donor 7043

data_processing:
Bases were called using Illumina bcl2fastq2 conversion v2.29
Reads were aligned using bowtie2 implimented in the ENCODE pipeline. Reads alignment by offsetting +4bp for all the reads aligned to the forward strand, and -5bp for all the reads aligned to the reverse strand
Open chromatin regions (OCRs) were called using MACS2 implemented in the ENCODE pipeline with the following parameters -p 0.01 --nomodel --shift -75 --extsize 150 -B --SPMR --keep-dup all --call-summits. The encode blacklist regions (wgEncodeDacMapabilityConsensusExcludable.bed.gz) were removed from called peaks.
Genome_build: hg19
Supplementary_files_format_and_content: NarrowPeak, peak calls (IDR conservative peaks + pooled_rep calls)

source_name: Adipocytes dervived from mesenchymal stem cells donor 7043

extract_protocol:
Cells were fixed after 2 weeks of adipogenic induction. Cells were collected and single-cell suspension were made with aliquots of 10^7 cells in 10 ml media (e.g. RPMI + 10%FCS). 540 µl 37% formaldehyde was added and incubated for 10 min at RT. The reaction was quenched by adding 1.5 ml 1 M cold glycine (4 °C) for a total volume of 12 ml. Fixed cells were centrifuged at 1000 rpm for 5 min at 4 °C and supernatant removed. The cell pellets were washed in 10 ml cold PBS (4 °C) followed by centrifugation. Supernatant was removed and cell pellets were resuspended in 5 ml of cold lysis buffer (10 mM Tris pH8, 10 mM NaCl, 0.2% NP-40 (Igepal) supplemented with protease inhibitor cocktail). Resuspended cells were incubated for 20 min on ice, centrifuged as above, and the lysis buffer removed. Finally, cell pellets were resuspended in 1 ml fresh lysis buffer, transferred to 1.5 ml microfuge tubes and snap frozen (ethanol/dry ice or liquid nitrogen). Cells were stored at −80 °C until they were thawed for 3C library generation.
Samples underwent a pre-digestion incubation with the addition of 0.3% SDS, 1x NEBuffer DpnII, and dH2O for 1hr at 37ºC in a Thermomixer shaking at 1,000rpm. A 1.7% solution of Triton X-100 was added to each tube and shaking was continued for another hour. After the pre-digestion incubation, 10 μL of DpnII (was added only to each sample tube, and continued shaking until the end of the day. An additional 10 µL of DpnII was added to each digestion reaction and digestion continued overnight. The next day, another additional 10 µL of DpnII was added and the incubation continued for another 2-3 hours. 100 μL of each digestion reaction was then removed, pooled into one 1.5 mL tube. The remaining samples were heat inactivated at 65°C for 20 min at 1000 rpm in a Thermomixer, and cooled on ice for 20 additional minutes. Digested samples were ligated with 8 μL of T4 DNA ligase (HC 30U/uL) and 1X ligase buffer at 1,000 rpm overnight at 16°C in a Thermomixer. The next day, an additional 2 µL of T4 DNA ligase was spiked into each sample and incubated for another few hours. The ligated samples were then de-crosslinked overnight at 65°C with Proteinase K along with the pre-digestion and digestion controls. The following morning, both controls and ligated samples were incubated for 30 min at 37°C with RNase A, followed by phenol/chloroform/isoamyl alcohol extraction and ethanol precipitation at -20°C. The 3C libraries were centrifuged at 3000 rpm for 45 min at 4°C 14,000 rpm, to pellet the samples. DNA pellets were resuspended in 70% ethanol and again centrifuged as described above. The 3C library pellets were resuspended in 300 μL dH2O and stored at −20°C. 10 μg of isolated DNA from 3C libraries of each library was sheared in dH2O using a QSonica Q800R to an average fragment size of 350bp.  After shearing, DNA was purified using AMPure XP beads. One microgram of adaptor-ligated library was used as input for the SureSelect XT capture kit using manufacturer’s protocol and our custom-designed 41K promoter Capture-C probe set.

tag: subject status

value: healthy donor

tag: gender

value: M

value: Adipocyte


title: MSC dervived Adipocyte donor 8013

source_name: Adipocytes dervived from mesenchymal stem cells from donor 8013


title: MSC dervived Adipocyte donor 8004

source_name: Adipocytes dervived from mesenchymal stem cells from donor 8004


title: 017_CD4_48_SP


title: 046_CD4_61_LP


title: 085_CD8_48_LP


title: 039_CD4_61_LP


title: 094_CD8_61_LP


title: 009_CD4_44_SP


title: 028_CD4_44_LP


title: PBMC ECCITE-seq HTO_5P

city: New York

first: Yuhan

last: Hao

address: 101 6th Ave

institute: New York Genome Center

data_processing:
Library strategy: ECCITE-seq
CITE-seq and ECCITE-seq RNA reads were mapped to the human genome (hg38). Transcripts were quanitfied using CellRanger v3.1.0
ADT, HTO counts were quantified using Alevin (https://salmon.readthedocs.io/en/latest/alevin.html#)
Cells were demultiplexed into their experimental conditions after normalization (centered log ratio transformation, margin = 2) using the HTOdemux function in Seurat.
TCR and BCR clonotypes were identified using the 10x Genomics cellranger vdj pipeline (https://support.10xgenomics.com/single-cell-vdj/software/pipelines/latest/using/vdj)
Genome_build: hg38
Supplementary_files_format_and_content: Processed data contains 3P and 5P single cell expression matrices where the rows correspond to features  (genes, or protein) and the columns to single cells, their corresponding meta data tables, and 5P BCR, TCR analysis results.

source_name: PBMC

extract_protocol: For the CITE/ECCITE-seq experiment: a panel of antibodies (ADTs) was used to stain the cells according to the CITE/ECCITE-seq protocol. To identify cell doublets. and keep track of patients identity and their vaccination time points, we split cells into aliquots and stained each aliquot with a unique hashing (HTOs) antibody.  Samples were washed and recombined and loaded on a 10X genomics chip. 5prime 10X genomics was ran and the manufacturers protocols was followed for cDNA library generation. To recover HTOs, ADTs and TCR/BCRs we followed the CITE/ECCITE-seq protocol found here: https://cite-seq.com/protocol/.

value: PBMC


title: 013_CD4_48_LP

source_name: splenocytes and tumor draining lymph node lymphocytes


title: PBMC ECCITE-seq BCR_5P


title: PBMC ECCITE-seq TCRgd_5P


title: 021_CD4_48_LP


title: PBMC ECCITE-seq RNA_5P


title: 049_CD8_48_SP


title: 004_CD4_44_LP


title: PBMC CITE-seq ADT_3P

data_processing:
Library strategy: CITE-seq
CITE-seq and ECCITE-seq RNA reads were mapped to the human genome (hg38). Transcripts were quanitfied using CellRanger v3.1.0
ADT, HTO counts were quantified using Alevin (https://salmon.readthedocs.io/en/latest/alevin.html#)
Cells were demultiplexed into their experimental conditions after normalization (centered log ratio transformation, margin = 2) using the HTOdemux function in Seurat.
TCR and BCR clonotypes were identified using the 10x Genomics cellranger vdj pipeline (https://support.10xgenomics.com/single-cell-vdj/software/pipelines/latest/using/vdj)
Genome_build: hg38
Supplementary_files_format_and_content: Processed data contains 3P and 5P single cell expression matrices where the rows correspond to features  (genes, or protein) and the columns to single cells, their corresponding meta data tables, and 5P BCR, TCR analysis results.


title: PBMC CITE-seq RNA_3P


title: PBMC_10x3'v3

city: SOUTH SAN FRANCISCO

first: Tracy

last: Yamawaki

state: California

address: 1120 VETERANS BLVD.

institute: AMGEN, INC.

description: PBMC

data_processing:
fastq files were downsampled using seqtk v1.3-r106
fastq files were processed with cellranger v3.0.2 for cell demultiplexing, read alignment and transcript counting
Data was aligned to a “cellRanger_1.2.0” referenced composed of hg19 with Ensembl 82
Genome_build: hg19

extract_protocol:
cell suspensions were encapsulated in cell lysis buffer with the 10X Chromium
Libraries were constructed with the 10X Genomics 3' v2, v3 or 5' v1 kits.


title: PBMC ECCITE-seq ADT_5P


title: PBMC CITE-seq HTO_3P


title: 012_CD4_44_LP


title: 053_CD8_48_LP


title: 022_CD4_61_LP


title: PBMC_10x5'v1

description: PBMC


title: 001_CD4_44_SP


title: 076_CD8_44_LP


title: 2hAux_4h42C_DoubleAID_rep2

first: Ali

last: Shilatifard

address: 320 E Superior St

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

data_processing:
For ChIP-seq and RNA-seq, single-end raw reads were trimmed by Trimmomatic 0.33 (Bolger et al., 2014) and aligned to hg19 using Bowtie (1.1.2) and TopHat (2.1.0)(Langmead et al. 2009; Trapnell et al. 2009), allowing only uniquely mapping reads with up to two mismatches within the 50 bp read. The resulting reads were extended to 150 bp toward the interior of the sequenced fragment, normalized to total reads aligned (reads per million, rpm) and formatted as bigWig-coverage tracks using GenomicRanges.
Genome_build: hg19
Supplementary_files_format_and_content: bigWig files with normalized read counts were provided for all samples.

growth_protocol: DLD-1 or HCT116 cells were cultured in DMEM (Corning) supplemented with 10% FBS (Corning), 1X GlutaMAX (Gibco), 100 U/ml Penicillin and 100 µg/ml Streptomycin (Gibco).

extract_protocol:
For ChIP-seq, 20-50 million cells were crosslinked with 1% paraformaldehyde (Electron Microscopy Sciences) in PBS for 10 min at r.t. Chromatin was sonicated with the Covaris E220 for 4 min with 10% duty cycle, 140 peak intensity power, 200 cycles per burst. Immunoprecipitations were carried out with antibodies and Protein A/G-agarose beads (Santa Cruz) or Dynabeads Protein G (Invitrogen).
For RNA-seq, total RNAs were isolated using the RNeasy Mini Kit from Qiagen. mRNAs were isolated using NEBNext Poly(A) mRNA Magnetic Isolation Module.
For ChIP-seq, DNA libraries were prepared by the HTP Library Preparation Kit for Illimina (KAPA) and sequenced on the NextSeq 500 (Illumina).
RNA libraries were prepared using NEBNext Ultra II Directional RNA Library Prep Kit and sequenced on Illumina NovaSeq 6000.

treatment_protocol:
Cells were treated with 500 µM Auxin (3-indole-acetic acid sodium salt, abcam), 250 nM dBET6 (AOBIOUS), or 20 nM KL-1 (Liang et al. 2018). Heat shock treatment at 42 ˚C for 1 h was carried out as described previously (Mahat et al, 2016)
For RNA-seq, cells were treated with or without auxin for 2 h in 37°C incubator and were directly placed into 42°C incubator for heat shock.

tag: cell line

value: Colorectal adenocarcinoma

value: AFF1/4-AID OsTIR1

tag: growth temperature

value: Heat shock

value: Auxin


title: 2hAux_4h42C_DoubleAID_rep1

tag: genotype


title: Sample1_UE_SingleCell

city: Parkville

first: Sean

last: Wilson

state: Victoria

address: 50 Flemington Road

country: Australia

institute: MCRI

data_processing:
10x library demultiplexing:  CellRanger 3.1.0
Single Cell RNA-Seq:  Quality control, sample integration and analysis was performed in Seurat (v3.1.4)
Bulk RNA-seq:  The raw reads were quality and adapter trimmed using trimmomatic (v0.39)
Bulk RNA-seq:  The trimmed reads were mapped to the human genome (hg38) using the STAR aligner version 2.5.2a in the two pass mapping mode
Bulk RNA-seq:  The mapped reads were quantified at the gene level using featureCounts version 1.5.0-p3 using GENCODE Release 20 comprehensive annotation
Genome_build: GRCh38
Supplementary_files_format_and_content: scRNA-seq samples:  barcode, features and matrix files from CellRanger output, scUE_METADATA.csv table containing the metadata related to single cell dataset (column 1 barcodes are appended with "_#" with being 1 for SH1 and 2 for SH2 barcodes).
bulk RNA-seq samples: counts

source_name: Human iPSC kidney organoid

growth_protocol: Takasato et al, 2015 (PMID: 26444236) kidney organoid protocol, modified to use E6 base media as in Howden et al, 2019 (PMID: 30858339).

extract_protocol:
Single Cell RNA-Seq: Samples were dissociated with a 1:1 TrypLE/Accutase solution (~0.2 ml per organoid) at 37˚C for 15–25 min, with occasional agitation (flicking) until large clumps were no longer clearly visible. The cell suspension was then centrifuged (3 min, 300 g) and the dissociation solution removed. Cells  were resuspended in HBBS supplemented with 2% FBS and passed through a 40 μM FACS tube cell strainer. Following centrifugation at 300 g for 3 minutes, the supernatant was discarded and cells resuspended in 50 μl TeSR-E6 medium. Viability and cell number were assessed and samples were run across separate runs on a Chromium Chip Kit (10x Genomics)
Single Cell RNA-seq: 10x libraries were constructed using 10x Chromium Next GEM Single Cell 3' Reagent kit v3.1. Sequenced using Illumina Novaseq

treatment_protocol: Day 25 organoids were dissociated with a 1:1 TrypLE/Accutase solution (~0.2 ml per organoid) at 37˚C for 15–25 min, with occasional agitation (flicking) until large clumps were no longer clearly visible. The cell suspension was then centrifuged (3 min, 300 g) and the dissociation solution removed. Cells  were resuspended in HBBS supplemented with 2% FBS and the mCherry+ fraction was isolated by fluorescent activated cell sorting (FACS) . The purified mCherry+ cells were centrifuged and resuspended in UB medium (TesR-E6 supplemented with 200 ng/ml FGF2, 3 mM CHIR99021, 2 mM ATRA, 10 mM Y-27632 and 100 ng/ml GDNF), counted using a haemocytometer and transferred to 24-well Transwell inserts (5e4–10e4 cells per well). Cells were mounted in a 1:1 Matrigel:UB medium solution (placed on top of the Transwell) and 0.5 ml UB medium was placed in the basolateral subcompartment.  Media was changed every other day. At 14 days culture the a single culture was isolated and dissociated for single cell RNA sequencing

value: Kidney organoid

tag: batch


title: 057_CD8_48_SP


title: Sample2_UE_SingleCell


title: 2hAux_4h37C_DoubleAID_rep2


title: PBMC ECCITE-seq TCRab_5P


title: 061_CD8_48_LP


title: 2hAux_4h37C_DoubleAID_rep1


title: 070_CD8_61_LP


title: 2hH2O_4h42C_DoubleAID_rep2


title: 2hH2O_4h42C_DoubleAID_rep1


title: 2hH2O_4h37C_DoubleAID_rep2


title: 2hH2O_4h37C_DoubleAID_rep1


title: 2hAux_4h42C_BRD4AID_rep2

value: BRD4-AID OsTIR1


title: 050_CD4_61_SP


title: 2hAux_4h37C_BRD4AID_rep2


title: 026_CD4_61_SP


title: 2hAux_4h37C_BRD4AID_rep1


title: 077_CD8_48_LP


title: 060_CD8_44_LP


title: 055_CD8_61_LP


title: 031_CD4_61_LP


title: 2hH2O_4h42C_BRD4AID_rep1


title: 066_CD4_61_SP


title: 073_CD8_48_SP


title: 036_CD4_44_LP


title: 2hH2O_4h37C_BRD4AID_rep1


title: pSS-5

city: 深圳

first: Shuhui

last: Meng

state: 广东

address: 翠竹街道

country: China

institute: SHENZHEN PEOPLE' HOSPITAL

data_processing:
Using the 10x Genomics official analysis software Cell Ranger (https://support.10xgenomics.com/single-cell-gene expression/software/overview/welcome), the original data was filtered, compared, quantified, identified and recovered cells, and finally the gene expression matrix of each cell was obtained.
Seurat perform quality control, analysis, and exploration of scRNA-seq data.
likelihood-ratio test was used to find differential expression for a single cluster, compared to all other cells.
GO database (http://www.geneontology.org/), differently enriched GO term was calculated by hypergeometric test. The calculated p-value was corrected by FDR, and p-value ≤ 0.05 was considered to be statistically significant. KEGG pathway was calculated in the same way as GO.
Genome_build: GRCh38
Supplementary_files_format_and_content: processed data files format and content is mtx and tsv

extract_protocol:
The PBMCs were used for single-cell RNA sequencing library preparation by Chromium Single Cell 3’ Reagent Kit (version 2) and Chromium Controller (10X Genomics, CA, USA) according to the manufacturer's instructions.
Firstly, the cDNA was digested into fragments of about 200-300 bp, and then library construction process of traditional second-generation sequencing such as sequencing adapter P5 and sequencing primer R1, and finally the DNA library was obtained by PCR amplification.contain the P5 and P7 primers used in Illumina bridge amplification.
paired-end sequencing model

value: blood

tag: sex

value: female

tag: disease state

value: oral and ocular dryness


title: 2hH2O_4h37C_BRD4AID_rep2


title: 087_CD8_61_LP


title: pSS-4


title: 018_CD4_61_SP


title: 052_CD8_44_LP


title: pSS-3


title: 015_CD4_61_LP


title: 084_CD8_44_LP


title: 082_CD4_61_SP


title: pSS-2


title: 2hAux_4h42C_BRD4AID_rep1


title: 014_CD4_61_LP


title: HC-5


title: 019_CD4_65_SP


title: 045_CD4_48_LP


title: HC-3


title: HC-4


title: pSS-1


title: 071_CD8_61_LP


title: 2hH2O_4h42C_BRD4AID_rep2


title: 079_CD8_61_LP


title: HC-2


title: HC-1


title: 054_CD8_61_LP


title: 058_CD4_61_SP


title: 2hH2O_42C1h_AID660_AFF4AID_DLD_rep1

description: AID ChIP-seq

data_processing:
For ChIP-seq, single-end raw reads were trimmed by Trimmomatic 0.33 (Bolger et al., 2014) and aligned to hg19 using Bowtie version 1.1.2, allowing only uniquely mapping reads with up to two mismatches within the 50 bp read. The resulting reads were extended to 150 bp toward the interior of the sequenced fragment, normalized to total reads aligned (reads per million, rpm), and formatted as bigWig-coverage tracks using GenomicRanges.
Genome_build: hg19
Supplementary_files_format_and_content: bigWig files with normalized read counts were provided for all samples.

growth_protocol: DLD-1 cells were cultured in DMEM (Corning) supplemented with 10% FBS (Corning), 1X GlutaMAX (Gibco), 100 U/ml Penicillin and 100 µg/ml Streptomycin (Gibco).

extract_protocol:
For ChIP-seq, 20-50 million cells were crosslinked with 1% paraformaldehyde (Electron Microscopy Sciences) in PBS for 10 min at r.t. Chromatin was sonicated with the Covaris E220 for 4 min with 10% duty cycle, 140 peak intensity power, 200 cycles per burst. Immunoprecipitations were carried out with antibodies and Protein A/G-agarose beads (Santa Cruz) or Dynabeads Protein G (Invitrogen).
For ChIP-seq, DNA libraries were prepared by the HTP Library Preparation Kit for Illimina (KAPA) and sequenced on the NextSeq 500 or NovaSeq 6000 (Illumina).

treatment_protocol: Cells were treated with 500 µM Auxin (3-indole-acetic acid sodium salt, abcam), 250 nM dBET6 (AOBIOUS), or 20 nM KL-1 (Liang et al. 2018). Heat shock treatment at 42 ˚C for 1 h was carried out as described previously (Mahat et al, 2016)

value: Colorectal adenocarcinoma DLD-1

value: AFF4-AID OsTIR1

tag: chip antibody


title: 034_CD4_61_SP

value: lymphocytes


title: 2hAux_42C1h_AID660_AFF4AID_DLD_rep1

description: AID ChIP-seq

tag: treatment


title: 011_CD4_65_SP


title: 2hAux_37C1h_AID660_AFF4AID_DLD_rep1

description: AID ChIP-seq


title: 2hH2O_42C1h_AID660_AFF1AID_DLD_rep1

description: AID ChIP-seq

value: AFF1-AID OsTIR1


title: 2hH2O_37C1h_AID660_AFF4AID_DLD_rep1

description: AID ChIP-seq


title: 041_CD8_48_SP


title: 2hAux_37C1h_AID660_AFF1AID_DLD_rep1

description: AID ChIP-seq


title: Mus_002_TUM_FFT_RNA

source_name: abdominal wall tissue

extract_protocol:
Tumor tissue was collected from mice inoculated with SCC VII to perform Exome-Seq and RNA-Seq, and mouse tail samples were collected and subjected to Exome-Seq to be used as germline controls.  Total DNA and RNA was purified using the AllPrep DNA/RNA Micro Kit (Qiagen, Valencia, CA).
For Exome-Seq, sequencing libraries were prepared and captured using SureSelect All Exon V4 Library Kits (Agilent Technologies, Santa Clara, CA) following the manufacturer’s instructions.  Briefly, 1-200 μg of genomic DNA from each sample was fragmented by Adaptive Focused Acoustics (E220 Focused Ultrasonicator, Covaris, Woburn, MA) to produce an average fragment size of ~175 bp.  Fragmented DNA was purified using Agencourt AMPure XP beads (Beckman Coulter).  The quality of the fragmentation and purification was assessed with an Agilent 2200 Tapestation (Agilent Technologies).  The fragment ends were repaired and adaptors were ligated to the fragments.  The resulting DNA library was amplified by 7 cycles of PCR, and 500 ng of each library was captured by solution hybridization to biotinylated RNA library baits for 24 hrs at 65°C.  Bound genomic DNA was purified with streptavidin coated magnetic Dynabeads (Invitrogen) and further amplified using 12 cycles of PCR to add barcode tags.  For RNA-Seq, total RNA was assessed for quality using an Agilent 2100 Tapestation (Agilent Technologies).  RNA-Seq libraries were generated from 200 ng of total RNA using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold (Illumina, San Diego, CA).  Manufacturer’s instructions were followed with shear time modified to 5 minutes.  Both Exome- and RNA-enrichment libraries were sequenced on the Illumina HiSeq 2500 in Rapid Run mode (Illumina) generating 100×100 bp paired-end reads.  An average Exome-Seq depth of 36.07 and RNA-Seq fold-coverage of 224.75 (~200×106 reads) was achieved.

value: Tumor


title: 029_CD4_48_LP


title: 2hH2O_37C1h_AID660_AFF1AID_DLD_rep1

description: AID ChIP-seq


title: mES KO postHS_2

first: Pierre

last: Klein

state: NY

address: 1300 York Avenue

department: Department of Pharmacology

institute: Weill Medical College, Cornell University

description:
ribosome protected fragment
heat shock 30min at 42°C + 1h recovery at 37°C

data_processing:
After sequencing, fastq files were quality-based trimmed, reads below 18 nt were excluded and adapter sequence were removed using the FLEXBAR tool.
Reads were demultiplexed based on 5’ barcodes for individual replicates using an in-house Linux shell script
The second part of the iCLIP random barcode was then moved to the read header (awk -F “##” '{sub(/..../,”##”$2, $2); getline($3); $4 = substr($3,1,2); $5 = substr($3,3); print $1 $2 $4”\n”$5}')
Ribosomal RNA reads were removed using STAR-genome-aligning software
The STAR-genome-aligning software was used to generate gene-count tables after aligning to the mm10 transcriptome.
Ribosome protected fragments reads were aligned to the transcriptome to avoid any possible contamination coming from the untranslated area of the genome
Gene count tables for each sample were then normalized and processed using DESeq2
Genome_build: mm10

source_name: mouse embryonnic stem cells (mES)

growth_protocol: Mettl14 knockout and wild type mouse ES cells were previously described by Geula et al. and were a generous gift of J. Hanna and S. Geula (Weizmann Institute of Science). mES cells were grown in Knockout DMEM (10829018, Invitrogen) with 15% heat inactivated FBS, 100 U ml−1 penicillin and 100 μg ml−1 of streptomycin, 200 mM L-glutamine, 1% non-essential Amino-Acid (Gibco 11140076), 50 μM β-mercaptoethanol (21985023, Gibco), 1000 U ml-1 mouse LIF (ESG1107, EMD Millipore), 30 μM GSK3 inhibitor CHIR99021 (04-0004-02, Stemgent), 10 μM MEK1 inhibitor PD0325901 (04-0006-02, Stemgent). Media was removed and changed every day. Cells were cultured on 0.1% gelatin-coated (07903, StemCell Technologies Inc.) plates. Cells were passaged every 2-3 days enzymatically using TrypLE Express (Life Technologies) according to the manufacturer’s instructions

extract_protocol:
Ribosome protected fragment  were obtained aw previously described by McGlincy, N.J. and Ingolia N.T. Methods 126, 112-129 (2017).  Briefly, to inhibit ribosome transit post-lysis, cells were rapidly washed twice with ice-cold PBS containing 50 µg/ml of cycloheximide. To generate ribosome-protected fragments, cells were pelleted and immediately lysed in 400 µL of cell lysis buffer (20 mM Tris pH 7.4, 150 mM NaCl, 5 mM MgCl2, 1 mM DTT, 100 µg/mL cycloheximide, 25 U DNase I). 30 µg of RNA were used to isolate ribosome-protected fragments following RNase I digestion. After RNase digestion, lysates were loaded on a sucrose gradient and centrifuged at 100,000 rpm to recover ribosome protected fragments. RNA from the resuspended ribosomal pellet was purified and run on a gel to selectively excise footprinted RNAs (from 17 nt to 34 nt RNA fragments). To avoid any ribosomal contamination in the library preparation steps, we then performed Ribo-Zero Gold depletion of the footprinted RNA. The rRNA depleted RNA fragments were dephosphorylated and the linker was added. To specifically deplete unligated linker, yeast 5’-deadenylase and RecJ exonuclease digestion was performed
library preparation steps were performed essentially as previously described: Linder, B. , Grozhik, A.V. et al. Nat methods 12, 767-772 (2015)

treatment_protocol: mES cells were plated on a 10 cm dish and allowed to reach 70-80% confluence.  Heat shock was performed for 30 min in a water bath at 42°C. For the recovery condition, heat shocked cells were placed back at 37°C for 1 h in a water bath

value: mouse embryonnic stem cells (mES) V6.5

value: Mettl14 -/-


title: 2hAux_42C1h_AID660_DoubleAID_DLD_rep1

description: AID ChIP-seq


title: 2hH2O_37C1h_AID660_DoubleAID_DLD_rep1

description: AID ChIP-seq


title: mES KO HS_2

description:
ribosome protected fragment
heat shock 30min at 42°C

value: heat shock 30min at 42°C


title: mES WT postHS_1

value: wild type

value: heat shock 30min at 42°C + 1h recovery at 37°C


title: 2hH2O_42C1h_AID660_DoubleAID_DLD_rep1

description: AID ChIP-seq

value: AFF1/4-AID OsTIR1

value: Heat shock


title: mES WT postHS_2


title: 044_CD4_44_LP


title: mES KO postHS_1


title: mES KO HS_1


title: mES WT HS_2


title: mES WT HS_1


title: 2hAux_37C1h_AID660_BRD3AID_DLD_rep1

description: AID ChIP-seq

value: BRD3-AID OsTIR1

value: Auxin


title: mES KO preHS_2

description:
ribosome protected fragment
No heat shock

value: No heat shock


title: 2hAux_42C1h_AID660_AFF1AID_DLD_rep1

description: AID ChIP-seq


title: mES KO preHS_1


title: 2hAux_42C1h_AID660_BRD3AID_DLD_rep1

description: AID ChIP-seq


title: 2hAux_42C1h_AID660_BRD4AID_DLD_rep1

description: AID ChIP-seq

value: BRD4-AID OsTIR1


title: 2hH2O_42C1h_AID660_BRD3AID_DLD_rep1

description: AID ChIP-seq


title: mES WT preHS_2


title: 2hH2O_42C1h_AID660_BRD4AID_DLD_rep1

description: AID ChIP-seq


title: mES WT preHS_1


title: 540712_Lung.Mets

data_processing:
Library strategy: piggyBac-Seq
Standard Illumina software was used for base calling.
Mapping of insertion sequences to the mouse genome and identification of common integration sites (CIS). Pre-processing and alignment: Paired-End reads were first pre-processed by removing the expected transposon-derived sequence (5’-TAGGGTTAA-3’) from the beginning of the first read (read-pairs with non-matching first reads were discarded; typically around 1%) using the preprocessReads function from QuasR (version 1.12.0,33). Read pairs were then aligned to the mouse genome (BSgenome.Mmusculus.UCSC.mm10) using the QuasR qAlign function and parameters “-m 1 --best --strata --maxins 1000”; only uniquely mapping pairs with up to 1000 bp between-pair distance were reported. Mapping rates were recorded and non-mapped read pairs were further aligned against the non-mobilized transposon sequence to estimate the probable fraction of read pairs that originate from non-mobilized copies of the transposon. For each aligned read pair, the piggyBac insertion coordinate was identified as the coordinate of the first (most 5’) base of the first read.  Integration site identification and quantification: For each unique integration site, the number of distinct supporting alignments (distinct read pairs), and the genomic sequence from the four base pairs on the same strand as the first read directly upstream of the insertion site were recorded. Only integration sites with the expected upstream TTAA sequence were used for the downstream analysis.  Association of integration sites with genes: Coordinates of known genes (exons/introns, 5’-untranslated region [UTR], coding sequence [CDS], 3’UTR) were obtained from the TxDb.Mmusculus.UCSC.mm10.knownGene Bioconductor package (version 3.2.2). Promoter regions were defined as regions 2,000 bp upstream of known transcript start sites. Integration sites were matched against these genomic regions to identify overlaps on any strand and orientation, selecting the first overlap in the case of multiple overlaps. Integration sites were classified hierarchically as follows: Sites without overlaps to any transcript were labeled as promoter sites (in the case of an overlap with a promoter region) or intergenic sites. All other sites were labeled with the first region type that they overlapped, in the following order: 5’UTR, CDS, 3’UTR, intron, or ncRNA (defined as an overlap with a transcript without annotated CDS). Sites were further labeled according to their orientation with respect to the associated gene (same or opposite). Finally, sites were grouped according to the gene they overlapped (including promoter sites) or, for intergenic sites, according to the pair of flanking genes.  Selection of the enriched integration sites: For each CIS (all insertions associated to a single gene or genomic locus), the diversity (Div) was calculated by summing up the Div values of the individual insertions. Finally, this value was divided by the total number of different insertions for a given sample and converted to percentage. In order to find putative metastatic genes, we looked at genes with a higher number of insertions in the metastatic than in the tumour samples.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: processed_data_insertion_wise.csv
Supplementary_files_format_and_content: processed_data_gene_wise.csv

source_name: breast cancer cell line

extract_protocol:
Genomic DNA was isolated using DNeasy Blood and Tissue Kit (Qiagen).
Extracted genomic DNA was sheared to a fragment length of 250 bp with a Covaris sonicator. After end repair and A-tailing, purified DNA fragments were ligated to a splinkerette adaptor. Transposon-containing fragments were enriched by 18 cycles of transposon-specific PCR1 for the 5′ transposon ends in a unique library. Bar coding of individual samples and completion of Illumina adaptor sequences were achieved by an additional 12 cycles (primary tumours and LM cell lines) and 18 cycles (lung-metastases and lung macro-metastases) of transposon-specific PCR and a custom array of 35 unique bar-coding primers. After magnetic bead purification (Beckman Ampure XP), libraries were assembled in two pools and sequenced.

tag: breast cancer model

tag: mouse strain

value: NOD.CB17-Prkcd<scid>/J (NOD/SCID)

tag: sample type

value: Lung metastasis


title: 2hAux_37C1h_AID660_BRD4AID_DLD_rep1

description: AID ChIP-seq

value: Colorectal adenocarcinoma DLD-1


title: 543630_Lung.Mets


title: 2hAux_37C1h_AID660_DoubleAID_DLD_rep1

description: AID ChIP-seq


title: 2hH2O_37C1h_AID660_BRD4AID_DLD_rep1

description: AID ChIP-seq


title: 543632_Lung.Mets


title: 540700_LM7

value: Lung metastasis cell line


title: 2hAux_42C1h_AID660_BRD2AID_DLD_rep1

description: AID ChIP-seq

value: BRD2-AID OsTIR1


title: 543630_LM4


title: 2hH2O_37C1h_AID660_BRD3AID_DLD_rep1

description: AID ChIP-seq


title: 643609_LM2


title: 2hH2O_42C1h_AID660_BRD2AID_DLD_rep1

description: AID ChIP-seq


title: 2hAux_37C1h_AID660_BRD2AID_DLD_rep1

description: AID ChIP-seq


title: 643636_LM1


title: 542627_Tumor

value: Primary tumor


title: 2hH2O_37C1h_AID660_BRD2AID_DLD_rep1

description: AID ChIP-seq


title: 28 WOA D/DA

city: Houston

first: Peixin

last: Jiang

address: One Baylor Plaza

institute: Baylor College of Medicine

hyb_protocol: Standard Illumina hybridization protocol.

scan_protocol: Standard Illumina scanning protocol.

data_processing: The data was processed and normalized using R package lumi.

label: biotin

source_name: 28 weeks of age D/DA-treated

label_protocol: Biotinylated cRNA were prepared with the Ambion MessageAmp kit for Illumina arrays

extract_protocol: RNA was extracted by using the AllPrep DNA/RNA/Protein Mini kit.

value: TRAMP

tag: age

value: 28 weeks

tag: genotype/treatment

value: D/DA-treated

tag: tissue

value: dorsal-lateral prostate lobes


title: 28 WOA AGN

source_name: 28 weeks of age AGN-treated

tag: strain

value: AGN-treated


title: 16 WOA D/DA

source_name: 16 weeks of age D/DA-treated

value: 16 weeks


title: 28 WOA WT

source_name: 28 weeks of age wild type

value: wild type


title: 540712_Tumor


title: 543630_Tumor


title: 2hAux_2hDMSO_AID660_BRD4AID_DLD_rep1

description: AID ChIP-seq


title: 540712_LM5


title: 543641_LM6


title: 543641_Tumor


title: 543610_Tumor


title: 543631_LM3


title: 543626_Tumor


title: 540700_Tumor


title: 504701_Tumor


title: 28 WOA Vehicle

source_name: 28 weeks of age vehicle-treated

value: vehicle-treated


title: 16 WOA WT

source_name: 16 weeks of age wild type


title: 0hDMSO_AID660_BRD4AID_DLD_rep1

description: AID ChIP-seq


title: 16 WOA Vehicle

source_name: 16 weeks of age vehicle-treated


title: 543639_Tumor


title: 543625_Tumor


title: 2hAux_BRD3Bethyl_BRD3AID_DLD_rep1

description: BRD3 ChIP-seq

value: BRD3-AID OsTIR1

value: Auxin

value: Brd3 Bethyl A302-368A


title: AGN 2

source_name: AGN-treated

value: neuroendocrine carcinomas


title: 2hH2O_BRD3Bethyl_BRD3AID_DLD_rep1

description: BRD3 ChIP-seq


title: 543631_Tumor

value: NOD.CB17-Prkcd<scid>/J (NOD/SCID)


title: 643636_Tumor


title: 643609_Tumor


title: 2hAux_BRD2CST_BRD2AID_DLD_rep1

description: BRD2 ChIP-seq

value: Colorectal adenocarcinoma DLD-1

value: Brd2 (D89B4) CST mAb #5848


title: 2hdBET6_AID660_BRD4AID_DLD_rep1

description: AID ChIP-seq

value: BRD4-AID OsTIR1


title: 16 WOA AGN

source_name: 16 weeks of age AGN-treated


title: AGN 3


title: 2hH2O_BRD2CST_BRD2AID_DLD_rep1

description: BRD2 ChIP-seq

value: BRD2-AID OsTIR1


title: AGN 1

value: TRAMP


title: 543637_Tumor

value: Primary tumor


title: Vehicle 3

source_name: vehicle-treated


title: Vehicle 2


title: 2hAux_42C1h_Pol2CST_AFF4AID_DLD_rep3

description: Pol2 ChIP-seq

value: AFF4-AID OsTIR1

value: Heat shock

value: Rpb1 NTD (D8L4Y)


title: 2hH2O_42C1h_Pol2CST_AFF4AID_DLD_rep3

description: Pol2 ChIP-seq


title: 2hAux_37C1h_Pol2CST_AFF4AID_DLD_rep3

description: Pol2 ChIP-seq


title: HTO libraries

city: Hannover

first: Berislav

last: Bosnjak

address: Carl-Neuberg Straße 1

department: Institute of Immunology

institute: Hannover Medical School

data_processing:
For cDNA libraries, Cell Ranger v3.1 02 was used for alignment to the mouse genome (mm10), filtering, barcode counting and unique molecular identifier counting , resulting in 82% of 270,993,014 or 311,497,445 reads from WT or CD25Y129H CD4+GFP+ Treg cells, respectively, confidently mapped to the genome.
The R package Seurat v2.3 was used under R 3.5 for further data trimming, unsupervised clustering, and visualization (Butler et al., 2018).
Downstream analysis was performed using the Seurat R package (version 3.0, Satija Lab; PMID: 31178118) which enables the integrated processing of multi-modal (RNA & HTO) single-cell datasets (PMID: 29608179). Initially, raw cDNA library datasets were loaded and genes detected in less than 3 barcodes or barcodes with less than 200 genes were filtered out. Barcodes passing this initial filtering served for retrieval of the cell hashing tags.The cell hashing tags (WT mice: ACCCACCAGTAAGAC,GGTCGAGAGCATTCA,CTTGCCGCATGTCAT; CD25Y129H mice: AAAGCATTCTTCACG, CTTTGTCTTTGTGAG, TATGCTGCCACGGTA) were counted using CITE-seq-Count protocol (https://cite-seq.com/protocol/; PMID: 30567574), resulting in an average of 1111 and 967 reads/cell for WT or CD25Y129H CD4+GFP+ Treg cells, respectively. Next, all barcodes designated as “singlets” were extracted and further filtered using Seurat’s standard pre-processing workflow.  Normalization and downstream analysis was performed using the Seurat R package (version 3.0, Satija Lab; PMID: 31178118) which enables the integrated processing of multi-modal (RNA & HTO) single-cell datasets (PMID: 29608179). In the analysis, single cells with 900-3500 detected genes and with a percentage of mitochondrial genes below 10% were included, resulting in total of 3704 WT CD4+GFP+ Treg cells (with average of 1702 genes / 4722 reads) and 2142 CD25Y129H CD4+GFP+ Treg cells (with average of  1725 genes / 4862 reads) passing the filters.
Clustering and UMAP plots were generated using resolution 0.5 and first 30 principal component analysis (PCA) dimensions applying SCTransform function of Seurat package. After inspection, we detected the presence of few contaminating B cells that were excluded from further analyses. After removal of contaminating cells, remaining events were re-clustered with resolution 0.5 and first 30 PCA dimensions using SCTransform function of Seurat package, resulting in 6 clusters. For detection of differently expressed genes (DEGs), data were log normalized and scaled using NormalizeData and ScaleData function of Seurat, before DEGs were found using FindAllMarker or FindMarker function. All neighboring clusters with less than 20 DEGs (adjusted p value < 0.05) were merged, resulting in final UMAP plot with 4 clusters. Heatmaps and Venn diagrams for DEGs were done in R using the ggplot2 package. To find enriched GO biological processes, data were exported from R and analyzed with PANTHER classification system software (v.14.1) using Fisher’s exact test with false discovery rate calculated and significance level adjusted to 0.05 (http://pantherdb.org; PMID 23193289). To identify subset-specific markers, we manually compared the DEGs from each cluster, determined by a threshold of false discovery rate (FDR) < 0.001, to markers previously reported in canonical pathways and Treg cell subsets 25.
The cells were ordered in pseudotime using PCA and Seurat cluster assignments with the cluster 2 as a starting point. Two main trajectories were found, one including cells from cluster 2, 3 and 4 and the other including cells from cluster 2, 3 and 1. To find the temporally expressed genes, generalized additive model (GAM) for top 1000 the most variable genes was fitted with a LOESS term for pseudotime.
Genome_build: mm10
Supplementary_files_format_and_content: Processed data include cellranger raw counts (including barcode, feature, and matrix files) before processing in Seurat package and matrices from cell hashing libraries, as determined by CITE-seq-Count protocol (https://cite-seq.com/protocol/; PMID: 30567574), resulting in an average of 1111 and 967 reads/cell for WT or CD25Y129H CD4+GFP+ Treg cells, respectively.
Supplementary_files_format_and_content: Merged matrix from HTO counts

source_name: cell hashing libraries

extract_protocol:
After euthanasia, spleens from WT and CD25-Y129H Foxp3-GFP mice were collected. Single cell suspensions were obtained by filtering through nylon gaze and re-suspended in FACS buffer. Treg cells were sorted based on CD4 and GFP expression. Cells from three mice per genotype were labelled with TotalSeq cell hashing antibodies (Biolegend), mixed together, and used to generate single-cell gel-beads in emulsion using Chromium Single Cell 3’ Library & Gel Bead Kit v.3 according to the manufacturer´s protocol (10x Genomics).
The scRNA-seq libraries were prepared following the user guide manual (CG000183 Rev A) provided by the 10X Genomics and Chromium™ Single Cell 3' Reagent Kit (v3). Briefly, after labeling with cell hashing antibodies, 9000 cells from each sample were encapsulated in microfluidic droplets (estimated doublet rate ~4.3%). Encapsulated cells were subjected to reverse transcription (RT) reaction at 53ºC for 45 min. After RT step, the emulsion droplets were broken and barcoded-cDNA and -HTO was purified with DynaBeads, followed by 12-cycles of PCR-amplification (98 °C for 180 s; [98 °C for 15 s, 63 °C for 20 s, 72 °C for 60 s] x 12-cycles; 72 °C for 60 s). Afterwards, PCR products were cleaned using SPRI selection and in this step HTO-libraries were separated from cDNA libraries as descibed in cell hashing protocol (version 2019-02-13, https://cite-seq.com/protocol/; PMID: 30567574). For cDNA libraries, 10 µl of PCR-amplified barcoded-cDNA was fragmented with the reagents provided in the kit and purified with SPRI beads to obtain an average fragment size of ~400 bp. Next, the DNA library was ligated to the sequencing adapter followed by indexing PCR (98 °C for 45 s; [98 °C for 20 s, 54 °C for 30 s, 72 °C for 20 s] x 16 cycles; 72 °C for 60 s). The resulting DNA library was double-size purified (0.6-0.8X) with SPRI beads. For HTO libraries, they were amplified and ligated to sequencing adaptors using 2x KAPA Hifi PCR (95 °C for 45 s; [95 °C for 20 s, 64 °C for 30 s, 72 °C for 20 s] x 12 cycles; 72 °C for 300 s) and purified with SPRI beads (1.6x). Both cDNA- and HTO-libraries were sequenced on an Illumina NextSeq platform (R1 – 26 cycles, i7 – 8 cycles, R2 – 56 cycles) resulting in ~300 million reads per cDNA sample (average reads per single-cell being 100,000 a).

value: spleen


title: CD25Y129H CD4+GFP+ Treg cells

source_name: spleen


title: WT CD4+GFP+ Treg cells

value: WT


title: 2hAux_42C1h_Pol2CST_AFF1AID_DLD_rep3

description: Pol2 ChIP-seq

value: AFF1-AID OsTIR1


title: MCF10A_GFP

city: Detroit

first: Zeng-Quan

last: Yang

state: MI

address: 4100 John R HWCRC 815

department: Oncology

institute: Karmanos Cancer Institute

description: MCF10A cells were infected by adenoviruses of the control GFP.

hyb_protocol: Hybridization was performed overnight on a µParaflo microfluidic chip based on the LC Sciences service procedure. On the microfluidic chip, each detection probe consisted of a chemically modified nucleotide coding segment complementary to target microRNA (from miRBase, http://microrna.sanger.ac.uk/sequences/) or other RNA (control or customer defined sequences) and a spacer segment of polyethylene glycol to extend the coding segment away from the substrate. The hybridization melting temperatures were balanced by chemical modifications of the detection probes. Hybridization used 100 mL 6xSSPE buffer (0.90 M NaCl, 60 mM Na2HPO4, 6 mM EDTA, pH 6.8) containing 25% formamide at 34 °C. After RNA hybridization, tag-conjugating Cy3 dye was circulated through the microfluidic chip for dye staining.

scan_protocol: Fluorescence images were collected using a laser scanner and digitized using Array-Pro image analysis software (Media Cybernetics). Microarray images were carefully scanned for a balanced view. Numerical intensities are extracted for control, background, and microRNA probes.

data_processing: Data were analyzed by first subtracting the background and then normalizing the signals using a LOWESS filter (Locally-weighted Regression).

source_name: Nontumorigenic mammary epithelial cell line

label_protocol: Cy3 - LC Sciences service procedure.

extract_protocol: Total RNA was extracted with miRNeasy Mini kits (Qiagen, Hilden, Germany), and miRNA microarray analysis was performed by LC Sciences (Houston,TX).

value: Breast


title: MCF10A_IRE1

description: MCF10A cells were infected by adenoviruses of the wild-type IRE1.


title: 2hH2O_42C1h_Pol2CST_AFF1AID_DLD_rep3

description: Pol2 ChIP-seq


title: 2hAux_37C1h_Pol2CST_AFF1AID_DLD_rep3

description: Pol2 ChIP-seq

value: Auxin


title: SUM52_K599A

description: SUM52 cells were infected by adenoviruses of the IRE1 dominant-negative K599A.

source_name: Breast cancer line


title: SUM52_K907A

description: SUM52 cells were infected by adenoviruses of the IRE1 dominant-negative K907A.