title: R1_Control liver_scRNAseq

description: 10x Genomics

data_processing:
The sequencing reads were mapped using Cell Ranger (6.1.2) to genome build GRCm38 (mm10).
All downstream processing and analysis were performed using the R package Seurat (4.1.1). The counts were loaded into R and to remove low quality cells first, only features (genes) detected in a minimum of three cells and cells expressing a minimum of 200 features (genes) were considered for the further quality control. To systematically filter cells with high percentage of mitochondrial reads and low number of gene expressed depending on each sample, we used the median absolute deviation (MADs) higher (for mitochondrial percentage) or lower (for the number of genes detected) than the median value for these metrics to remove cells.
The remaining cells were SCT normalized and integrated using the IntegrateData() function. The data was reduced and clustered (Louvain algorithm, 0.1 resolution) which were then annotated as Granulocytes, Neutrophils, Monocytic cells, cDC1s, pDCs, B cell, T cell, ILCs and NK cells.
Assembly: mm10
Supplementary files format and content: h5:binary format containing a matrix with columns containing barcode sequences, UMI-counts, indexes

source_name: immune and non immune cells

extract_protocol:
For each sample the Chromium Controller platform of 10X Genomics was used for single cell partitioning and barcoding.
Each cell’s transcriptome was barcoded during reverse transcription, pooled cDNA was amplified and Single Cell 3’ Gene Expression were prepared according to the manufacturer’s protocol “Chromium NextGEM Single Cell 3’ Reagent Kits v3.1” (CG000315, 10X Genomics). Both Single Cell 3’ Gene Expression libraries were quantified on a 2100 Bioanalyzer Instrument following the manufacturer’s protocol “Agilent DNA 7500 kit” (G2938-90024, Agilent Technologies). These Single Cell 3’ Gene Expression libraries were combined to create one sequence library pool which was quantified by qPCR, according to manufacturer’s protocol “KAPA Library Quantification Kit Illumina® Platforms” (KR0405, KAPA Biosystems). A NovaSeq 6000 Illumina sequencing system was used for paired end sequencing of the Single Cell 3’ Gene Expression libraries at a sequencing depth of approximately 17.000 mean reads per cell for Control liver  and 35.000 mean reads per cell for NrasG12DPtenKO.

value: immune and non immune cells

value: 4 weeks

value: normal


title: whole lung_stress_bio rep1

status: Public on Feb 22 2024

type: SRA

city: CLEVELAND

first: Yuan

last: Gao

state: OH

address: 2109 Adelbert Rd., Wood Building Research Tower 300-7, Wood Building Research Tower 300-7

department: Pharmacology

country: USA

institute: Case Western Reserve University

library_source: transcriptomic

data_processing:
Raw reads from RNA-seq were mapped to reference genome mm10 using RNA STAR with default parameters. Read count tables were created using FeatureCounts with a custom GTF file containing protein coding genes only. Differentially expressed genes were analyzed using DESeq2 with two independent replicates using default parameters.
Raw reads from ChIP-seq were aligned to the reference genome hg38 using Bowtie2 with sensitive end-to-end setting. After removal of PCR duplication reads by SAMtools, peaks were identified using MACS2, with narrow peak option for TFs and broad peak option for H3K27Ac. Genome-wide read coverage was calculated by deepTools with a bin size of 50 bp.
Assembly: mm10
Supplementary files format and content: bigwig

molecule: total RNA

organism: Mus musculus

source_name: lung

growth_protocol: Mouse neutrophils were isolated from the bone marrow of 8-week-old female BL/6 mice. Briefly, bone marrow was flushed from both femurs and tibias with 1x Hanks’ Balanced Salt Solution (HBSS) using a syringe with a 26G needle. After washing the bone marrow cells once with 1x HBSS, the cells were resuspended in ammonium-chloride-potassium (ACK) lysis buffer for 3 minutes on ice. The cells were then washed twice with 1x HBSS, resuspended in 2 ml of HBSS. Neutrophils were isolated by density gradient separation. The density gradient was made by layering 2 mL of bone marrow cells on top of 3 mL of 62% Percoll (#17-0891-02, GE Healthcare) on top of 3 mL of 81% Percoll in a 15 ml falcon tube, followed by centrifugation at 2,500 x g for 20 minutes at 4°C. Neutrophils were then taken from the middle interface, washed in HBSS, and resuspended in serum-free DMEM before use.

extract_protocol:
For RNA-seq experiments, total RNA was isolated using TRIzol (Thermo Fisher, 15596026) according to the manufacturer’s instructions. For ChIP_seq experiments, lysates were clarified from sonicated nuclei and protein-DNA complexes were isolated with antibody.
was constructed using the Illumina TruSeq RNA library prep kit v2 (Illumina, RS-122-2001) or  Illumina TruSeq ChIP library prep kit (Illumina, IP-202-1012) based on the manufacturer’s instructions.

tag: tissue

value: lung

tag: cell type

value: whole lung

tag: genotype

value: WT

tag: treatment

value: stress


title: D5-BMYFP-09-01

city: Bronx

first: David

middle: Robert

last: Fooksman

state: NY

address: 1300 Morris Park

department: Pathology

institute: Albert Einstein College of Medicine

data_processing:
RNA-seq reads were aligned to the mouse genome (mm39/GRCm39) using STAR aligner (v2.6.1b). Differential expression was computed using the DESeq2 (v 1.26.0).Alignment was done using STAR
Counts for individual genes were quantified using the RSEM program (v1.3.1)
immunoglobulin genes were removed
Differential expression was computed using the DESeq2 (v 1.26.0).
Assembly: mm10/GRCm38
Supplementary files format and content: immunoglobulin genes removed for total processing normalized counts
Supplementary files format and content: assembled BCR clones using mixcr analysis

source_name: Bone marrow

extract_protocol:
SMART-Seq v4 Ultra Low Input RNA Kit for Sequencing
Nextera XT DNA Library Preparation Kit

value: Bone marrow

tag: cell line

value: primary FACS-purified cells

value: LLPC

value: BEC-YFP

value: 24 week old mouse, D5 post Tamoxifen


title: M14 3D CD20- replicate 2

city: Cardiff

first: Girish

middle: K

last: Patel

address: Maindy Road

country: United Kingdom

institute: Cardiff University

data_processing:
Trimmed and quality-checked reads were aligned to the GRCh38 human reference genome using HISAT2
normalised reads were generated using featureCounts (v1.5.0)
differentially expressed genes were then generated using the standard DESeq2 pipeline
Assembly: GRCh38/hg38
Supplementary files format and content: .txt data file normalised reads for each sample.

molecule: polyA RNA

organism: Homo sapiens

extract_protocol:
M14 cells were grown under non-adherent culture conditions to generate melanospheres. Melanospheres were mechanically dissociated, antibody labelled for CD20 and flow sorted to isolate CD20+ and CD20- populations. RNA was extracted from samples using an RNeasy mini kit as per manufacturers instructions. RNA was then quantified and quality assessed before being sent to Novogene (Cambridge, UK).
total RNA was extracted using Rneasy mini kit (QIAGEN).
RNA was sent to Novogene (Cambridge, UK). Libraries were generated using Illumina TruSeq RNA Library Prep Kit (v2) and were subsequently sequenced on a Illumina NovaSeq 6000


title: DM8-2FTSP2HS57_L002

city: Duluth

first: Daniel

middle: L.

last: Villeneuve

state: MN

address: 6201 Congdon Blvd

department: Mid-Continent Ecology Division

institute: US EPA

description:
DM_8-2FTS_P2_H
DM_8-2FTS_counts.filtered.+1.normFinal.txt

data_processing:
RNA sequencing was performed at the Michigan State University Research Technology Support Facility Genomics Core. Libraries were quality control checked and quantified using a combination of Qubit dsDNA HS, Agilent 4200 TapeStation HS DNA1000, and Invitrogen Collibri Library Quantification qPCR assays. Each pool (corresponding to ≈ 96 samples from assays with two PFAS) was loaded onto one lane of a NovaSeq S4 flow cell. Sequencing was performed in a 2x150bp paired end format using a NovaSeq 6000 v1.5 300 cycle reagent kit (target read depth of ≈28,000 MB/sample). Base calling was done by Illumina Real Time Analysis v3.4.4, and the output was demultiplexed and converted to FASTQ format with Illumina Bcl2fastq v2.20.0.
The Daphnia magna reference genome (assembly ASM2063170v1.1, RefSeq GCF_020631705.1) was indexed with STAR version 2.7.10a using the following sample specific parameters: --genomeSAindexNbases 12 --sjdbOverhang 149.
After performing quality control on the raw FASTQ data using FastQC and MultiQC version 1.12, raw reads were trimmed using Trim Galore! version 0.6.6, with the parameters --illumina --stringency 3 --quality 20 –e 0.1 --length 20 --paired, to remove sequencing adapters and low-quality bases and filter out trimmed reads shorter than 20 bp.
Trimmed reads were then mapped with STAR version 2.7.10a to the indexed genome using the default parameters except for the use of: --outSAMattributes NH HI AS NM XS. Samtools version 1.15.1 was used to sort and index the alignment files.
HTSeq version 2.0.1 was used to assign counts to each gene with the following parameters: -f bam -r pos -s no. FastQC and MultiQC version 1.12 were used to assess quality control metrics of the processed reads. Count data for treatment and plate-specific control samples for each PFAS were written to a single tab-delimited matrix file, where each column represented a sample and each row represented a gene.
Additional quality control and filter steps were applied to each count matrix using the R statistical computing software (R Core Team, 2022) prior to concentration response modeling. Lowly expressed genes were removed from the raw count matrices using edgeR’s (Robinson et al. 2010) built-in filtering function, filterByExpr(), in version 3.40.2; namely, genes with < 15 reads across all samples and < 10 reads in the smallest group (treatment or control) sample size were filtered out. Normalization factors for the filtered count matrices were calculated using the trimmed mean of M-values method via the edgeR function calcNormFactors() which was used for counts per million mapped reads (CPM) normalization. The filtered CPM normalized count matrix was log2 transformed with a pseudo-count of 1 added (to prevent occurrence of any negative count values for downstream analyses; i.e., log2(CPM+1)).
Assembly: Daphnia magna reference genome (assembly ASM2063170v1.1, RefSeq GCF_020631705.1)
Supplementary files format and content: Count matrices provided as tab delimited text files. Top row = sample names. Second row = treatment concentration (ug/L); all subsequent rows provide filtered counts per million (CPM) log2 transformed with a pseudo-count of 1 added (to prevent occurrence of any negative count values for downstream analyses; i.e., log2(CPM+1), with each row corresponding to a separate transcript and each column corresponding to a sample. One count matrix file per chemical exposure experiment.

organism: Daphnia magna

source_name: RNA-Seq of daphnia magna whole body after 24 h exposure to 8:2FTS

growth_protocol: Daphnia magna were obtained from an onsite culture unit at the U.S. EPA Great Lakes Toxicology and Ecology Division, Duluth, MN. For use in the assays, Daphnia magna were reared in mass culture in amended LSW and held at 20° C until they were 72 hours old. They were fed a diet of yeast-Cerophyll-trout chow and algae (USEPA 2016d) until ready for use in the assays. All procedures involving animals were approved by a local animal care and use committee.

extract_protocol:
RNA was extracted from pooled homogenates using Zymo Research Direct-zol-96 RNA kit (Zymo Research) protocol adapted to the Biomek i5 to yield 40 µl of RNA. Previously extracted control RNA was added to empty wells on the final RNA plate. Concentrations of RNA for each well were measured on a NanoDrop spectrophotometer (Thermo Scientific, NanoDrop Eight) and diluted if necessary.
RNA extracts (10 µl) were used to prepare cDNA libraries with the Lexogen Poly(A) RNA Selection Kit V1.5 (catalog #157; Lexogen GmbH, Austria) and Lexogen CORALL mRNA-Seq Library Prep Kit with UDIs (catalog #158-163) both adapted to the Biomek i5. Resulting cDNA library samples were evaluated on a 4200 TapeStation (Agilent Technologies, CA, USA) using the D1000 ScreenTape (Agilent) analysis protocol to assess cDNA quality. Samples were quantified using the Quant-iT Picogreen dsDNA assay kit (catalog # P11495; Thermo Fischer Scientific), diluted if necessary, and equal portions of each sample pooled for use in RNA sequencing by Michigan State University.

treatment_protocol: All assays were conducted in 96-well plate format using 1 ml volume deep-well plates (Catalog # 502162, NEST Scientific). Daphnia magna, one individual per well, were exposed to eight different concentrations of PFAS (two PFAS tested per plate) plus controls. Nominal PFAS concentrations ranged from 0.03 to 100 uM with 1/2 log spacing. Five replicate individuals were exposed to each treatment per plate, except control treatments (n=8). Uniform age (72 ± 6 h) Daphnia magna were manually loaded to each well of three replicate 96-well plates using a P1000 pipet with the end of the tip cut off to a one cm bore diameter. The volume in each well was reduced to a uniform volume of 50 µl using the Biomek i5 liquid handler. Assays were initiated by adding 550 µl of exposure solution from a dosing plate to the three replicate exposure plates for each assay. Immediately after dosing, each plate was covered with a silicone sealing mat (catalog #506065; NEST Scientific) and then placed in a 20°C incubator for 24 h. At test termination, each well was inspected and all mortalities (immobile with gentle prodding) were recorded and removed. Live daphnia were also examined for phenotypic abnormalities, although none were noted at sublethal concentrations. The Biomek i5 was then used to transfer 100 µl of exposure solution to an “exposure verification plate”, where the n=5 samples for each treatment concentration were subsequently pooled together into a total volume of 500 on a per-plate basis. The pooled exposure solution for each treatment was collected for quantitification of PFAS concentrations, with measured concentrations ultimately used for data analysis. The remaining volume of exposure medium in each well was replaced with 60 µl of Trizol (Zymo Research, CA, USA). A 2.3 mm stainless steel bead was added to each well, then the plate was homogenized at 30 Hz for two minutes using an Omni-Bead Ruptor bead mill. Homogenates of individual Daphnia magna from the same treatment group were pooled together, on a per plate basis  to generate one pooled sample per treatment, per plate. Only live daphnia were included in the homogenate pools.

value: whole body

value: GLTED-wt

tag: conc.

tag: conc. units

value: ug/L

tag: conc code

value: H

tag: exposure duration

tag: age

tag: replicate plate_id


title: R1_NrasG12DPtenKO_scRNAseq

city: Amsterdam

first: Masami

last: Ando-Kuri

address: Plesmanlaan 121

country: Netherlands

institute: Netherlands Cancer Institute (NKI)

description: 10x Genomics

library_source: transcriptomic single cell

value: NrasG12D overexpression Pten knockout

tag: time

value: End-stage

tag: disease state

value: cancer


title: whole lung_stress_bio rep2


title: DM8-2FTSP2GS58_L002

description:
DM_8-2FTS_P2_G
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: G


title: DM8-2FTSP2FS59_L002

description:
DM_8-2FTS_P2_F
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: F


title: whole lung_control_bio rep2

value: lung

value: whole lung

value: control


title: R2_Cancer_cell_lines_NrasG12VPtenKO

description:
6794_5_716_C_2_CATAAGCGAC-CCTCTTCAAG_S6
Read_Counts_perGene_CancerCellLines.txt

data_processing:
The reads that are of low complexity or repetitive sequences were filtered.
Paired-end sequencing reads were trimmed with seqpurge and aligned with Hisat2 using as reference GRCm38 (mm10) (--min-intronlen 20 --max-intronlen 500000 -k 5 --minins 0 --maxins 500 --fr --new-summary --threads 16).
The read counts were generated using the function gensum (https://github.com/NKI-GCF/gensum) with gtf (v. 100) annotation.
Assembly: mm10
Supplementary files format and content: Read_Counts_perGene_Bulktumor.txt: tab-separated gene counts and gene annotation for all samples
Supplementary files format and content: Read_Counts_perGene_CD45pos.txt: tab-separated gene counts and gene annotation for all samples
Supplementary files format and content: Read_Counts_perGene_CancerCellLines.txt: tab-separated gene counts and gene annotation for all samples

extract_protocol:
Tissue samples were homogenized in 600ul of buffer RLT (79216, Qiagen) with the addition of 1% B-mercaptoethanol using the TissueLyserII (85300, Qiagen) for 6 minutes  with frequency setting 30 (1/s) in combination with 5mm stainless steel beads (69989, Qiagen). The total RNA was isolated  using the RNeasy Mini Kit (74106, Qiagen), including an on column DNase digestion (79254, Qiagen),  according to the manufactures instructions,.  Quality and quantity of the total RNA was assessed by the 2100 Bioanalyzer using a Nano chip (Agilent, Santa Clara, CA).  Total RNA samples having RIN>8 were subjected to library generation. CD45+ sorted cells (30.000-75.000 cells) from somatic HCC mouse model were homogenized in RLT buffer (79216, Qiagen), total RNA was isolated using the RNeasy Micro Kit (74004, Qiagen), including an on column DNase digestion (79254, Qiagen), according to the manufacturer’s instructions. Quality and quantity of the total RNA was assessed on the 2100 Bioanalyzer instrument following manufacturer’s instructions “Agilent RNA 6000 Pico” (G2938-90046, Agilent Technologies). In general RNA yields of 0.5-45 ng total RNA and RNA integrity numbers (RIN) above 7 were obtained, with only 1 sample had a RIN value of 5.7. RNA samples were concentrated with Agencourt RNAClean XP beads (A63987), according to manufacturer’s instructions (Protocol 001298v001, Agencourt) with modifications.
The library preparation for bulk tumors and cell lines was generated using the TruSeq Stranded mRNA sample preparation kit (Illumina Inc., San Diego, RS-122-2101/2) according to the manufacturer's instructions (Illumina, Document # 1000000040498 v00). The libraries were sequenced with  paired-end reads on a NovaSeq-6000 using a Reagent Kit v1.5 (Illumina Inc., San Diego). For CD45+ RNA-seq, the libraries were generated using Smart-Seq2 RNA (Illumina) according to the manufacturer’s instructions and paired-end sequenced using the Nextseq-550 High Output Kit v2.5 (150 Cycles).

value: HCC-derived-cell-line

value: NrasG12V overexpression Pten knockout


title: M14 3D CD20- replicate 1


title: R3_Cancer_cell_lines_NrasG12VPtenKO

description:
6794_6_716_C_3_AACTCTCTAC-ACGGCTTCTC_S7
Read_Counts_perGene_CancerCellLines.txt


title: M14 3D CD20+ replicate 2


title: M14 3D CD20+ replicate 1


title: M14 3D CD20+ replicate 3


title: DM8-2FTSP2ES60_L002

description:
DM_8-2FTS_P2_E
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: E


title: 15090-CD8_S27

city: Syracuse

first: Frank

middle: A

last: Middleton

address: 505 Irving Avenue

department: Neuroscience and Physiology

institute: SUNY Upstate Medical Univ

description: Sample 27

data_processing:
Raw BCL files were converted to FASTQ files in BaseSpace (Illumina)
Sequence reads were trimmed for adaptor sequence with CUTADAPT 1.12
Trimmed reads were mapped to mm10 of the mouse genome with STAR 2.7.3a
Alignments were quantified to mm10 - Ensembl Transcripts release 100
Raw gene counts were normalized using counts per million (CPM) method
Assembly: mm10
Supplementary files format and content: tab-delimited text files includes matrix of raw gene counts and normalized gene counts for all samples

source_name: spleen

extract_protocol:
Total RNA was extracted from splenocytes using the Qiagen RNeasy kit (Qiagen). Sequencing libraries were quantified using the KAPA Library Quantification Complete Kit Universal (KAPA Biosystems). The pooled library (1.4pM) was loaded onto the NextSeq 500 instrument, using the NextSeq 500/550 High Output v2 Kit for 75 cycles of paired-end reads (Illumina).
Sequencing library preparation was done with the Illumina TruSeq Stranded Total RNA with RiboZero Gold kit following manufacturer's instructions.

value: spleen

value: primary splenocyte

value: SLE1.2.3. triple-congenic (B6.TC)

value: female


title: DM8-2FTSP2DS61_L002

description:
DM_8-2FTS_P2_D
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: ug/L

value: D


title: DM8-2FTSP2CS62_L002

description:
DM_8-2FTS_P2_C
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: C


title: R1_Cancer_cell_lines_NrasG12VPtenKO

description:
6794_4_716_C_1_ACGAGACGTC-CGGTCTCTCT_S5
Read_Counts_perGene_CancerCellLines.txt

value: End-stage

value: cancer


title: R3_Cancer_cell_lines_NrasG12DPtenKO

description:
6794_3_580_C_3_TTACTTCGTG-AAGATCGGAT_S4
Read_Counts_perGene_CancerCellLines.txt

value: NrasG12D overexpression Pten knockout


title: 15090-CD4_S26

description: Sample 26


title: M14 3D replicate 3

description:
3D-3
Using the M14 melanoma cell line, we define culture conditions for reproducible 3D growth of melanospheres to investigate putative cancer stem cell populations. We undertook RNA sequencing to characterise cell populations between adherent and non-adherent cultures.

source_name: cell line

extract_protocol:
total RNA was extracted using Rneasy mini kit (QIAGEN).
RNA was sent to Novogene (Cambridge, UK). Libraries were generated using Illumina TruSeq RNA Library Prep Kit (v2) and were subsequently sequenced on a Illumina NovaSeq 6000

value: cell line


title: M14 3D replicate 2

description:
3D-2
Using the M14 melanoma cell line, we define culture conditions for reproducible 3D growth of melanospheres to investigate putative cancer stem cell populations. We undertook RNA sequencing to characterise cell populations between adherent and non-adherent cultures.


title: M14 3D replicate 1

description:
3D-1
Using the M14 melanoma cell line, we define culture conditions for reproducible 3D growth of melanospheres to investigate putative cancer stem cell populations. We undertook RNA sequencing to characterise cell populations between adherent and non-adherent cultures.


title: R2_Cancer_cell_lines_NrasG12DPtenKO

description:
6794_2_580_C_2_TCGAACACGA-GCGAGGTTCT_S3
Read_Counts_perGene_CancerCellLines.txt


title: DM8-2FTSP2AS64_L002

description:
DM_8-2FTS_P2_A
DM_8-2FTS_counts.filtered.+1.normFinal.txt
DM_PFTDoDA_counts.filtered.+1.normFinal.txt

value: A


title: 15086-CD8_S24

description: Sample 24


title: R1_Cancer_cell_lines_NrasG12DPtenKO

description:
6794_1_580_C_1_TAACTTGGTC-GTCGTGAATC_S2
Read_Counts_perGene_CancerCellLines.txt

value: HCC-derived-cell-line


title: DM8-2FTSP1HS49_L002

description:
DM_8-2FTS_P1_H
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: H


title: whole lung_control_bio rep1

value: lung

value: whole lung

value: WT

value: control


title: R3_Bulk tumor NrasG12VPtenKO

description:
6577_13_Ras12V_PTEN_3_-_619_CCTAAGCTGA-CGTCGCATCT_S13
Read_Counts_perGene_Bulktumor.txt

source_name: liver

value: liver

value: NrasG12V overexpression Pten knockout


title: M14 2D replicate 3

description:
2D-3
Using the M14 melanoma cell line, we define culture conditions for reproducible 3D growth of melanospheres to investigate putative cancer stem cell populations. We undertook RNA sequencing to characterise cell populations between adherent and non-adherent cultures.


title: M14 2D replicate 2

description:
2D-2
Using the M14 melanoma cell line, we define culture conditions for reproducible 3D growth of melanospheres to investigate putative cancer stem cell populations. We undertook RNA sequencing to characterise cell populations between adherent and non-adherent cultures.


title: R2_Bulk tumor NrasG12VPtenKO

description:
6577_12_Ras12V_PTEN_2_-_616_TATGGCAAGC-AACAGCAATG_S12
Read_Counts_perGene_Bulktumor.txt


title: DM8-2FTSP1FS51_L002

description:
DM_8-2FTS_P1_F
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: F


title: 15086-CD19_S22

description: Sample 22

value: spleen

value: primary splenocyte

value: female


title: 15086-CD4_S23

description: Sample 23


title: 15085-CD8_S21

description: Sample 21

value: SLE1.2.3. triple-congenic (B6.TC)/Rab4AQ72L -KO


title: DM8-2FTSP1ES52_L002

description:
DM_8-2FTS_P1_E
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L

value: E


title: M14 2D replicate 1

description:
2D-1
Using the M14 melanoma cell line, we define culture conditions for reproducible 3D growth of melanospheres to investigate putative cancer stem cell populations. We undertook RNA sequencing to characterise cell populations between adherent and non-adherent cultures.

value: cell line


title: ThalAndCtxOrganoids_Batch4_Lane3

city: San Francisco

last: Shin

state: CA

address: 1651 4th Street Room 521A

department: Neurosurgery

institute: University of California, San Francisco

description: 10X Genomics v3 chemistry, the processed data files associated with this library are found in GSMnnnn

data_processing:
Read alignment and generation of feature-barcode matrices were generated using CellRanger. Pooled samples were demultiplexed using SNP genotype data following reference genotyping using the Virio package. Feature-barcode matrices were processed using the Seurat package in R for clustering. Prior to clustering, we first removed cells that contained fewer than 1000 genes or greater than 10% of reads that corresponded to mitochondrial genes. Datasets were normalized using SCTransform. We performed principal compontents analysis to identify the major sources of variance, and performed batch correction using the Harmony package. We defined a batch as scRNAseq experiments that were run on the same 10X Chromium chip. We performed nearest neighbor embedding in the space of harmony embeddings. To determine molecularly distinct clusters, we performed Louvain clustering for community detection based on Jaccard similarities between pairs of cells in the space of the top 30 principal components. Alternatively, dataset projection mapping to primary tissue reference dataset was performed using the Symphony package.
Assembly: hg38
Supplementary files format and content: Processed data files are in RDS format, and contain Seurat Objects after preprocessing and clustering (CorticalOrganoids_WK10.rds, ThalamusOrganoids_WK10.rds) or reference mapping to a primary tissue dataset (ThalamusOrganoid_ReferenceMappedtoPrimary.rds).

source_name: Thalamic and Cortical Organoids

growth_protocol: Cortical and thalamic organoids were generated from iPS cells using recently published protocols (Bhaduri et al., 2020; Xiang et al., 2019). To generate cortical organoids, iPS cells were dissociated with PBS-EDTA and plated in an ultra-low attachment 6-well plate containing a DMEM/F12-based induction media containing 15% KSR, 1% MEM-NEAA, 1% Glutamax, 1% Pen-Strep, 100 μM β-Mercaptoethanol, 5 μM Dorsomorphin, 5 μM SB-431542, 3 μM IWR1-endo, and 1X CEPT apoptosis inhibitor cocktail (50 nM chroman 1, 5 µM emricasan, 0.1% polyamine supplement, and 0.7 µM trans-ISRIB). Neural induction media was replaced every two days for eight days, and Y27632 was removed from the media on the fourth day. After neural induction, plates containing cortical organoids were transferred to a plate shaker rotating at 80 rpm. From Days 8-18, cortical organoids were cultured in an expansion media comprised of a 1:1 mixture of DMEM/F12 and Neurobasal medium containing 1% N2, 2% B27 without vitamin A, 1% Glutamax, 1% MEM-NEAA, 55 mM β-Mercaptoethanol, 1% Antibiotic-Antimycotic, 10 ng/mL FGF2, and 10 ng/mL EGF, with a media exchange every two days. After Day 18, cortical organoids were moved into a maintenance media comprised of a 1:1 mixture of DMEM/F12 and Neurobasal medium containing 1% N2, 2% B27 with vitamin A, 1% Glutamax, 1% MEM-NEAA, 55 mM β-Mercaptoethanol, 1% Antibiotic-Antimycotic, and 200 μM ascorbic acid.   To generate thalamic organoids, iPS cells were dissociated with PBS-EDTA and resuspended in a DMEM/F12-based induction media containing 15% KSR, 1% MEM-NEAA, 1% Glutamax, 1% Pen-Strep, 100 μM β-Mercaptoethanol, 100 nM LDN-193189, 10 μM SB-431542, 4 μg/mL Insulin, and 20 μM Y27632. Induction media was replaced every two days, and Y27632 was removed on the fourth day. After eight days of induction, plates containing thalamic organoids were transferred to a plate shaker rotating at 80 rpm. From Days 8-16, thalamic organoids were cultured in a patterning media comprised of a 1:1 mixture of DMEM/F12 and Neurobasal medium containing 1% N2, 2% B27 without vitamin A, 1% Glutamax, 1% MEM-NEAA, 55 mM β-Mercaptoethanol, 1% Antibiotic-Antimycotic, 30 ng/mL BMP7, and 1 μM PD325901, with a media exchange every two days. After Day 16, thalamic organoids were moved into the maintenance media described above for cortical organoids.

extract_protocol:
Organoids were dissociated with papain for 30 minutes and then triturated with a fire-polished glass pipette. The cell suspension was washed three times in PBS and then strained through a 35um strainer mesh, and then run through 10X v2 or v3 chemistry protocol.
Libraries were constructed according to the 10X Genomics v2 or v3 chemistry protocols

value: Thalamic and Cortical Organoids

value: week 10

value: neural cells

value: 22q11 deletion and control lines


title: 15085-CD4_S20

description: Sample 20


title: R1_Bulk tumor NrasG12VPtenKO

description:
6577_11_Ras12V_PTEN_1_-_614_ATTCGGCTTA-TTAACGTCCG_S6
Read_Counts_perGene_Bulktumor.txt

value: End-stage

value: cancer


title: DM8-2FTSP1DS53_L002

description:
DM_8-2FTS_P1_D
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: D


title: R4_Bulk tumor NrasG12DPtenKO

description:
6577_10_Ras12D_PTEN_4_-_049_TCCAACTGAA-TGTCCGTAGG_S3
Read_Counts_perGene_Bulktumor.txt

value: liver

value: NrasG12D overexpression Pten knockout


title: 15085-CD19_S19

description: Sample 19


title: R3_Bulk tumor NrasG12DPtenKO

description:
6577_9_Ras12D_PTEN_3_-_594_ATAAGCGGAG-CTGTTCTACG_S1
Read_Counts_perGene_Bulktumor.txt


title: DM8-2FTSP1CS54_L002

description:
DM_8-2FTS_P1_C
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: C


title: DM8-2FTSP2BS63_L002

description:
DM_8-2FTS_P2_B
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: B


title: ThalAndCtxOrganoids_Batch4_Lane2

description: 10X Genomics v3 chemistry, the processed data files associated with this library are contained in the Batch5_Ctx_WTC11* files found at the foot of this record and in the files found on GSMnnnn


title: 15090-CD19_S25

description: Sample 25

value: spleen

value: primary splenocyte

value: SLE1.2.3. triple-congenic (B6.TC)

value: female


title: 15041-CD8_S18

description: Sample 18


title: DM8-2FTSP1BS55_L002

description:
DM_8-2FTS_P1_B
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L


title: DM8-2FTSP1GS50_L002

description:
DM_8-2FTS_P1_G
DM_8-2FTS_counts.filtered.+1.normFinal.txt

value: G


title: DM8-2FTSP1AS56_L002

description:
DM_8-2FTS_P1_A
DM_8-2FTS_counts.filtered.+1.normFinal.txt
DM_PFTDoDA_counts.filtered.+1.normFinal.txt

value: A


title: ThalAndCtxOrganoids_Batch4_Lane1

description: 10X Genomics v3 chemistry

value: Thalamic and Cortical Organoids

value: week 10

value: neural cells

value: 22q11 deletion and control lines


title: 15041-CD4_S17

description: Sample 17


title: 15041-CD19_S16

description: Sample 16


title: ThalamicOrganoids_Batch3

description: 10X Genomics v3 chemistry

source_name: Thalamic Organoid

value: Thalamic Organoid


title: R2_Bulk tumor MycOEPtenKO

description:
6577_5_Myc_PTEN_2_-_575_CATAAGCGAC-CCTCTTCAAG_S9
Read_Counts_perGene_Bulktumor.txt

value: liver

value: Myc overexpression Pten knockout

value: End-stage

value: cancer


title: 15016-CD8_S15

description: Sample 15


title: ThalAndCtxOrganoids_Batch1_L7


title: DM6-2FTSP3GS124_L002

description:
DM_6-2FTS_P3_G
DM_6-2FTS_counts.filtered.+1.normFinal.txt

source_name: RNA-Seq of daphnia magna whole body after 24 h exposure to 6:2FTS

value: whole body

value: GLTED-wt


title: ThalAndCtxOrganoids_Batch1_L6


title: ThalOrg_Batch2

description: 10X Genomics v3 chemistry


title: 15016-CD4_S14

description: Sample 14

value: spleen

value: primary splenocyte


title: DM6-2FTSP3FS125_L002

description:
DM_6-2FTS_P3_F
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: F


title: 15016-CD19_S13

description: Sample 13

value: female


title: ThalAndCtxOrganoids_Batch1_L5


title: R1_Bulk tumor MycOEPtenKO

description:
6577_4_Myc_PTEN_1_-_574_ACGAGACGTC-CGGTCTCTCT_S8
Read_Counts_perGene_Bulktumor.txt


title: R3_Bulk tumor MycOETrp53KO

description:
6577_3_Myc_p53_3_-_687_TTACTTCGTG-AAGATCGGAT_S4
Read_Counts_perGene_Bulktumor.txt

value: Myc overexpression Trp53 knockout


title: 15015-CD8_S12

description: Sample 12


title: R2_Bulk tumor MycOETrp53KO

description:
6577_2_Myc_p53_2_-_387_TCGAACACGA-GCGAGGTTCT_S5
Read_Counts_perGene_Bulktumor.txt


title: DM6-2FTSP3DS127_L002

description:
DM_6-2FTS_P3_D
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L

value: D


title: R2_Bulk tumor NrasG12DPtenKO

description:
6577_8_Ras12D_PTEN_2_-_580_GCCAACAGAC-CGTTCACGTT_S11
Read_Counts_perGene_Bulktumor.txt

value: NrasG12D overexpression Pten knockout


title: R1_Bulk tumor NrasG12DPtenKO

description:
6577_7_Ras12D_PTEN_1_-_579_GTATGCCACA-CCGAGTCTTC_S10
Read_Counts_perGene_Bulktumor.txt


title: ThalAndCtxOrganoids_Batch1_L4

description: 10X Genomics v3 chemistry

value: Thalamic and Cortical Organoids

value: week 10

value: neural cells


title: CtxOrgOnly_Batch3

description: 10X Genomics v3 chemistry

source_name: Cortical Organoid

value: Cortical Organoid

value: 22q11 deletion and control lines


title: R1_Bulk tumor MycOETrp53KO

description:
6577_1_Myc_p53_1_-_374_TAACTTGGTC-GTCGTGAATC_S2
Read_Counts_perGene_Bulktumor.txt

value: liver

value: End-stage


title: 15015-CD4_S11

description: Sample 11


title: DM6-2FTSP3CS128_L002

description:
DM_6-2FTS_P3_C
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: C


title: DM6-2FTSP3HS123_L002

description:
DM_6-2FTS_P3_H
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: GLTED-wt

value: H


title: R3_Bulk tumor MycOEPtenKO

description:
6577_6_Myc_PTEN_3_-_597_AACTCTCTAC-ACGGCTTCTC_S7
Read_Counts_perGene_Bulktumor.txt

value: Myc overexpression Pten knockout

value: cancer


title: R3_Control liver

description:
6577_16_Ctrl_E_V_3_-_610_CCTCGTTGGA-GACTGCGTTA_S16
Read_Counts_perGene_Bulktumor.txt

value: WT

value: 15 weeks

value: normal


title: DM6-2FTSP3BS129_L002

description:
DM_6-2FTS_P3_B
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L

value: B


title: CtxOrgOnly_Batch2

description: 10X Genomics v3 chemistry

value: week 10


title: DM6-2FTSP2GS117_L002

description:
DM_6-2FTS_P2_G
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: G


title: R2_Control liver

description:
6577_15_Ctrl_E_V_2_-_609_AGTATCCGAT-TATTGCGCTA_S15
Read_Counts_perGene_Bulktumor.txt


title: R1_Control liver

description:
6577_14_Ctrl_E_V_1_-_379_GCCACCTAGT-TGCGATGTAT_S14
Read_Counts_perGene_Bulktumor.txt

value: 4 weeks


title: 15007-CD8_S9

description: Sample 9

value: spleen

value: primary splenocyte

value: SLE1.2.3. triple-congenic (B6.TC)/Rab4AQ72L -KO

value: female


title: CtxOrgOnly_Batch1_replicate2

description: 10X Genomics v2 chemistry, the processed data files associated with this library are identical to those in org1

value: Cortical Organoid

value: neural cells

value: 22q11 deletion and control lines


title: DM6-2FTSP2FS118_L002

description:
DM_6-2FTS_P2_F
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: F


title: 15007-CD4_S8

description: Sample 8


title: CtxOrgOnly_Batch1

description: 10X Genomics v2 chemistry, the processed data files associated with this library are identical to those in org2


title: HGSOC Patient EOC227, pre-chemotherapy

city: Helsinki

first: Anna

last: Perkiö

address: Haartmaninkatu 8

country: Finland

institute: University of Helsinki

data_processing:
Reads originating from poor-quality cells were excluded (quality control steps done and explained by Zhang et al., 2022, doi:10.1126/sciadv.abm1831 )
STAR 2.7.8a was used to perform alignment. Genome indices were generated based on the GRCh38.d1.vd1 reference genome and GENCODE v25 annotation. A maximum of two mismatches per read was allowed.
For the L1 expression matrix only: Reads mapping more than 1 percent or more to exons were excluded. In addition, reads mapping 90 percent or more to repeats were excluded. The annotations were derived from RepeatMasker (<http://www.repeatmasker.org>). The described steps were performed with Intersect from Pybedtools (v0.8.1), a wrapper of Bedtools
For the L1 expression matrix only: To extract reads aligned in 1kb downstream windows of L1s, Intersect was again used to only retain reads with a minimum overlap of 60 percent with the reference regions. The 1kb reference windows were based on the list from Deininger et al. (2017, doi:10.1093/nar/gkw1067), consisting of L1 loci with potentially functional promoters.
For the L1 expression matrix only: To exclude reads that mapped ambiguously to polyA regions, Pysam 0.16.0.1, a python wrapper of SAMtools, was used to exclude reads for which the A or T base composition was 60 percent or higher.
The expression of both L1s and the rest of the genome was quantified with UMI-tools 1.1.1. Before the alignment, cell barcodes and UMIs were extracted from the reads. Subsequently, the unfiltered reads were assigned straight to the genome (featureCounts from Subread v2.0.1), and the filtered reads were assigned to the L1 1kb downstream windows. Only reads that mapped uniquely and to the same strand as the reference were considered, deduplicated and counted per feature per cell.
For the gene expression matrix only: The genes with a total of less than 95 UMIs (from all samples combined) were excluded.
Assembly: GRCh38.d1.vd1, GENCODE v25
Supplementary files format and content: comma-separated files for gene expression (tumor samples) and L1 expression (all samples)

source_name: High-grade serous ovarian cancer

extract_protocol:
The samples were incubated overnight in a mixture of collagenase and hyaluronidase to obtain single cell suspensions.
Standard Chromium Single Cell 3′ Reagent Kit v. 2.0 (10x Genomics)

value: High-grade serous ovarian cancer

value: clinical patient sample

value: tumor + microenvironment

value: no treatment


title: DM6-2FTSP3ES126_L002

description:
DM_6-2FTS_P3_E
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L

value: E


title: DM6-2FTSP2ES119_L002

description:
DM_6-2FTS_P2_E
DM_6-2FTS_counts.filtered.+1.normFinal.txt


title: HGSOC Patient EOC349, post-chemotherapy

value: neoadjuvant chemotherapy


title: DM6-2FTSP2DS120_L002

description:
DM_6-2FTS_P2_D
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: D


title: 15006-CD8_S6

description: Sample 6

value: spleen

value: primary splenocyte


title: ce_mRNA_ebax_TC_mut_30h

city: Basel

first: Dimos

last: Gaidatzis

address: Maulbeerstrasse 66

department: Computational Biology

country: Switzerland

institute: Friedrich Miescher Institute for Biomedical Research

data_processing:
The RNA-seq samples were mapped to the c.elegans genome (ce10) with the R package QuasR (version 1.26) using the spliced alignment algorithm HISAT2. The command used to perform the alignments was "proj <- qAlign("samples.txt","BSgenome.Celegans.UCSC.ce10",splicedAlignment=TRUE,aligner="Rhisat2")". For gene quantification, gene annotation from WormBase was used (WS220). The command used to create the expression count table was qCount(proj,exons,orientation="opposite", useRead="first"). To normalize for sequencing depth, each sample was divided by the total number of reads and multiplied by the average library size. Finally the count table was log2 transformed after the addition of a pseudocount of 8 in order to minimize large changes in expression caused by low count numbers.
Supplementary files format and content: tab-delimited text file with raw counts for each sample
Supplementary files format and content: tab-delimited text file with normalized counts for each sample

organism: Caenorhabditis elegans

growth_protocol: Synchronized L1's of N2 and HW3527 (ebax-1(tm2321) IV) were plated on 10 cm plates (2000 worms/plate) and kept at 25°C for the time-course. Every hour samples were collected from 18h to 31h

extract_protocol:
We harvested worms from 18h to 31h in hourly interval. Worms were collected from plates and washed 3 times and resuspended in Tri Reagent (MRC, TR118) and RNA was extracted using  Direct-zol™ RNA MicroPrep kit (Zymo Research; R2062) as per manufacturere's instructions. Total RNA was treated with DNAse on column
Sequencing libraries were generated using Illumina Stranded mRNA Prep Kit according to the manufacturer’s instructions. Subsequently, library was sequenced on NovaSeq6000, as 56 cycles paired-end reads.

treatment_protocol: no treatment

tag: strain

tag: time (h)


title: 15007-CD19_S7

description: Sample 7

value: SLE1.2.3. triple-congenic (B6.TC)/Rab4AQ72L -KO

value: female


title: ce_mRNA_ebax_TC_mut_31h


title: DM6-2FTSP2CS121_L002

description:
DM_6-2FTS_P2_C
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: C


title: 15006-CD4_S5

description: Sample 5


title: HGSOC Patient EOC349, pre-chemotherapy


title: 15015-CD19_S10

description: Sample 10


title: 15006-CD19_S4

description: Sample 4


title: ce_mRNA_ebax_TC_mut_29h


title: DM6-2FTSP2BS122_L002

description:
DM_6-2FTS_P2_B
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: B


title: 14959-CD8_S3

description: Sample 3

value: SLE1.2.3. triple-congenic (B6.TC)


title: Day0

city: Portland

last: Bottomly

address: 3181 S.W. Sam Jackson Park Rd.

institute: Oregon Health and Science University

description: PCR library sequencing

library_source: genomic

data_processing:
Read Trimming: cutadapt -j 2 -n 2 -g ACGAAACACCG -a GTTTAAGAGC  (Version 2.3)
Alignment: bowtie2 -p 1 -x yusa_library --norc -U {file}.trimmed.fastq {file}.tmp.sam (Version 2.3.5.1)
Conversion to BAM: samtools view -bS {file}.tmp.sam > {file}.bam (Version 1.9)
Filter to unique matches allowing no mismatches using bamtools (Version 2.5.1)
Counting reads for each sgRNA using MAGeCK: mageck count --norm-method none (Version: v0.5.8)
Assembly: Custom reference created from CRISPR library
Supplementary files format and content: Read counts per sgRNA for each sample

molecule: genomic DNA

source_name: Trametinib Resistant MOLM13 cells infected with library

growth_protocol: Trametinib resistant MOLM13 cells were grown in RPMI-1640, supplemented with 20% serum, pen/strep, fungizone.

extract_protocol:
30 million cells were collected for Trametinib day 14. DNA was extracted by desalting method
50 microgramms of DNA were used to PCR amplify library using Herculase II enzyme following published protocols and instructions supplied by authors on Addgene  (Shalem, et al Science 2014. 343(6166): p. 84-87, Tzelepis et al, Cell Rep, 2016. 17(4): p. 1193-1205)
Shalem et al Science 2014. 343(6166): p. 84-87

treatment_protocol: Puromycin was applied at 2mg/ml concentration for 14 days. Trametinib (100 nanoMolar) was applied to 30 million cells in single flask

value: Trametinib Resistant MOLM13

tag: genotype/variation

value: infected with CRISPR/Cas9 library

value: day 0 post puromycin selection


title: DM6-2FTSP1HS111_L002

description:
DM_6-2FTS_P1_H
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: H


title: ce_mRNA_ebax_TC_mut_27h


title: ce_mRNA_ebax_TC_mut_28h


title: Plamid

description: PCR library sequencing

source_name: Plasmid from Y. Kosuke library

value: Plasmid from Y. Kosuke library


title: HGSOC Patient EOC227, post-chemotherapy

value: High-grade serous ovarian cancer

value: clinical patient sample

value: tumor + microenvironment


title: ce_mRNA_ebax_TC_mut_26h


title: DM6-2FTSP1GS112_L002

description:
DM_6-2FTS_P1_G
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L

value: G


title: 14959-CD4_S2

description: Sample 2

value: spleen

value: primary splenocyte


title: DM6-2FTSP1FS113_L002

description:
DM_6-2FTS_P1_F
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: F


title: Trametinib day 14

description: PCR library sequencing

value: day 14 post puromycin selection, Trametinib


title: Trametinib day 14 replicate

description: PCR library sequencing


title: ce_mRNA_ebax_TC_mut_25h


title: DMSO day 14 replicate

description: PCR library sequencing

value: day 14 post puromycin selection, DMSO


title: ce_mRNA_ebax_TC_mut_24h


title: DM6-2FTSP1DS114_L002

description:
DM_6-2FTS_P1_D
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: D


title: ce_mRNA_ebax_TC_mut_23h


title: 5xFAD with daily APC injections

city: Tampa

first: Ji

last: Li

state: FL

address: 12901 Bruce B. Downs Blvd

department: Surgery

institute: University of South Florida

description: 10x Genomics

data_processing:
Demultiplexing, barcoded processing, and gene counting  were done by the 10x Genomics Cell Ranger software v6.0.2
Assembly: Mm10 Mouse Reference Genome supplied from 10x Genomics: refdata-gex-mm10-2020-A
Supplementary files format and content: Sample separate barcode, matrix, and genes

source_name: Brain

extract_protocol:
Whole brain tissue was collected at 7 months of age after 5 months of daily i.p. injection.  Brain tissue disassociated following the 10x Genomics Mouse Neural Tissue protocol.
Library constructed follwing manufacturer protocol for the 10x Genomics Chromium Next GEM Single Cell 3’ GEM, Library & Gel Bead Kit

value: Brain

value: Astrocyte, Endothelial, Microglia, Neuron, Oligodendrocyte, Oligodendrocyte Precursor Cell

value: Male and Female, 7 Months old

value: 100 μg/kg APC i.p. injection for 5 months


title: DM6-2FTSP1CS115_L002

description:
DM_6-2FTS_P1_C
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: C


title: DMSO day 14

description: PCR library sequencing

value: Trametinib Resistant MOLM13

value: infected with CRISPR/Cas9 library


title: 5xFAD with daily saline injections

description: 10x Genomics

value: Daily 100 μg/kg saline i.p. injection for 5 months


title: Young Wild-Type with daily APC injections

description: 10x Genomics


title: Polychromatophil, SF3B1 WT rep 4

city: Paris

last: Rombaut

address: 22 rue Méchain

country: France

institute: Institut Cochin

data_processing:
FASTQ files were mapped using STAR (v2.7.9) to align the reads against the human reference genome GRCh37 (UCSC version hg19). Read count normalizations and groups comparisons were performed using DESeq2 (v1.30.1), using the Wald test for significance testing. Genes with low counts were filtered, if at least half the samples have less than 20 normalized reads.
For differential expression study, the results obtained after DESeq2 comparison were selected for further analysis and filtered at Benjamini-Hochberg (BH)- adjusted P-value < 0.05 and log2(FC) > |1|.
To identify differentially expressed splicing events, we used KisSplice (v2.6.2), KisSplice2refgenome (v2.0.7) and KissDE (v1.15.3), and filtered results at BH adjusted P-value < 0.05 and ΔPSI > |0.10|.
Assembly: GRCh37
Supplementary files format and content: rawCOUNT_RNAseq_erythro.txt : raw count table (tabulated txt file). event_k2ref_erythro.tsv.gz : kissplice2refgenome output (splicing raw count, txt file), input_order_kissplice (txt file, order necessary to run splicing differential analysis with kissDE).

extract_protocol:
The RNA integrity (RNA Integrity Score ≥7.0) was checked on the Agilent Fragment Analyzer (Agilent) and quantity was determined using Qubit (Invitrogen). SureSelect Automated Strand Specific RNA Library Preparation Kit was used according to the manufacturer’s instructions with the Bravo Platform. Briefly, 50–100 ng of total RNA sample was used for poly-A mRNA selection using oligo(dT) beads and subjected to thermal mRNA fragmentation.
The fragmented mRNA samples were subjected to cDNA synthesis and were further converted into double stranded DNA using the reagents supplied in the kit, and the resulting dsDNA was used for library preparation. The final libraries were bar-coded, purified, pooled together in equal concentrations, and subjected to 100bp paired-end sequencing on Novaseq-6000 sequencer (Illumina) at Gustave Roussy.

value: Bone marrow

value: CD34+-derived polyE

value: SF3B1 WT


title: DM4-2FTSP3HS103_L002

description:
DM_4-2FTS_P3_H
DM_4-2FTS_counts.filtered.+1.normFinal.txt

source_name: RNA-Seq of daphnia magna whole body after 24 h exposure to 4:2FTS

value: ug/L

value: H


title: Young Wild-Type with daily saline injections

description: 10x Genomics


title: ce_mRNA_ebax_TC_mut_22h


title: DM4-2FTSP3GS104_L002

description:
DM_4-2FTS_P3_G
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: G


title: SF3B1 WT t24, rep 3

data_processing:
FASTQ files were mapped using STAR (v2.7.9) to align the reads against the mouse reference genome GRCm39 (GENCODE release M31). Read count normalizations and groups comparisons were performed using DESeq2 (v1.30.1), using the Wald test for significance testing. Genes with low counts were filtered, if at least half the samples have less than 20 normalized reads.
For differential expression study, the results obtained after DESeq2 comparison were selected for further analysis and filtered at Benjamini-Hochberg (BH)- adjusted P-value < 0.05 and log2(FC) > |1|.
To identify differentially expressed splicing events, we used KisSplice (v2.6.2), KisSplice2refgenome (v2.0.7) and KissDE (v1.15.3), and filtered results at BH adjusted P-value < 0.05 and ΔPSI > |0.10|.
Assembly: GRCm39
Supplementary files format and content: rawCOUNT_RNAseq_G1EER.txt : raw count table (tabulated txt file). event_k2ref_G1EER.tsv.gz: kissplice2refgenome output (splicing raw count, txt file), input_order_kissplice (txt file, order necessary to run splicing differential analysis with kissDE).

extract_protocol:
Samples were controled with an Agilent 2100 Bioanalyzer. RNA polyA was realized with the NEBNext polyA kit + Ultra II < 150ng.
Libraries were constructed using the TruSeq Stranded mRNA Sample Preparation Kit (Illumina, San Diego, CA), > 150ng BA-mRNA. Bank were normalized and controled with a Bioanalyzer.

treatment_protocol:
The murine G1E-ER4 cell line was used to generate isogenic Sf3b1K700E and Sf3b1WT cell lines using CRISPR-Cas9-stimulated homology-mediated repair. Cells were transfected using Amaxa Nucleofector™ 2b Device program A-024 and Nucleofector kit L (Lonza, Basel, Switzerland) with Cas9 (Addgene, Cambridge, MA), a mSf3b1-specific gRNA built from gRNA cloning vector and a donor template encoding a hygromycin selection cassette at a 1:1:1 ratio (Addgene).
After two days, hygromycin (Merck) was added to the culture at 800µg/mL for 72h, and maintained at 400µg/mL for another 72h. Hygromycin-resistant cells were then single cell-sorted using the FACS Aria III (Becton Dickinson Biosciences, Franklin Lakes, NJ) and clones were analysed 3 weeks later by DNA sequencing to assess the presence of the Sf3b1K700E mutation. The selection cassette was removed by flippase-mediated excision.
Sf3b1K700E and Sf3b1WT hygromycin-resistant clones were transfected with a flippase-encoding vector and single cell-sorted.
G1E-ER4 cells were treated or not with β-estradiol at 10-7 M (Merck).

value: G1E-ER4 isogenic SF3B1 WT

value: estradiol


title: Polychromatophil, SF3B1 WT rep 3


title: ce_mRNA_ebax_TC_mut_20h


title: Polychromatophil, SF3B1 WT rep 2


title: SF3B1 WT t24, rep 2


title: 14959-CD19_S1

description: Sample 1

value: spleen

value: primary splenocyte

value: SLE1.2.3. triple-congenic (B6.TC)

value: female


title: DM4-2FTSP3FS105_L002

description:
DM_4-2FTS_P3_F
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: F


title: DM4-2FTSP3ES106_L002

description:
DM_4-2FTS_P3_E
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: E


title: ce_mRNA_ebax_TC_mut_19h


title: SF3B1 WT t24, rep 1


title: ce_mRNA_ebax_TC_mut_18h


title: DM4-2FTSP3DS107_L002

description:
DM_4-2FTS_P3_D
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: D


title: Polychromatophil, SF3B1 MUT rep 5

value: Bone marrow

value: SF3B1 MUT


title: Polychromatophil, SF3B1 WT rep 1

value: CD34+-derived polyE

value: SF3B1 WT


title: SF3B1 WT t0, rep 3

value: no


title: ce_mRNA_ebax_TC_N2_31h

value: WT


title: DM4-2FTSP3CS108_L002

description:
DM_4-2FTS_P3_C
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L

value: C


title: Polychromatophil, SF3B1 MUT rep 4


title: Polychromatophil, SF3B1 MUT rep 3


title: DM4-2FTSP3BS109_L002

description:
DM_4-2FTS_P3_B
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: B


title: SF3B1 WT t0, rep 1

value: G1E-ER4 isogenic SF3B1 WT


title: ce_mRNA_ebax_TC_N2_30h


title: ce_mRNA_ebax_TC_mut_21h


title: ce_mRNA_ebax_TC_N2_29h


title: DM4-2FTSP3AS110_L002

description:
DM_4-2FTS_P3_A
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: A


title: Polychromatophil, SF3B1 MUT rep 2


title: ce_mRNA_ebax_TC_N2_28h


title: SF3B1 MUT t24, rep 3

value: G1E-ER4 isogenic SF3B1 K700E

value: estradiol


title: SF3B1 WT t0, rep 2


title: DM4-2FTSP2HS95_L002

description:
DM_4-2FTS_P2_H
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: H


title: Polychromatophil, SF3B1 MUT rep 1


title: SF3B1 MUT t24, rep 2


title: ce_mRNA_ebax_TC_N2_27h


title: DM4-2FTSP2GS96_L002

description:
DM_4-2FTS_P2_G
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: G


title: Basophil, SF3B1 WT, rep 3

value: Bone marrow

value: CD34+-derived basoE


title: SF3B1 MUT t24, rep 1

value: SF3B1 MUT


title: ce_mRNA_ebax_TC_N2_26h


title: Basophil, SF3B1 WT, rep 2

value: SF3B1 WT


title: DM4-2FTSP2ES98_L002

description:
DM_4-2FTS_P2_E
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: E


title: Basophil, SF3B1 WT, rep 1


title: DM4-2FTSP2DS99_L002

description:
DM_4-2FTS_P2_D
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: D


title: SF3B1 MUT t0, rep 1

value: no


title: SF3B1 MUT t0, rep 3


title: Basophil, SF3B1 MUT, rep 4


title: ce_mRNA_ebax_TC_N2_23h

value: WT


title: ce_mRNA_ebax_TC_N2_22h


title: Basophil, SF3B1 MUT, rep 3


title: DM4-2FTSP2BS101_L002

description:
DM_4-2FTS_P2_B
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: ug/L

value: B


title: ce_mRNA_ebax_TC_N2_21h


title: Basophil, SF3B1 MUT, rep 1


title: DM4-2FTSP2AS102_L002

description:
DM_4-2FTS_P2_A
DM_4-2FTS_counts.filtered.+1.normFinal.txt
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: A


title: Basophil, SF3B1 MUT, rep 2


title: DM4-2FTSP1HS89_L002

description:
DM_4-2FTS_P1_H
DM_4-2FTS_counts.filtered.+1.normFinal.txt


title: ce_mRNA_ebax_TC_N2_20h


title: DM4-2FTSP2FS97_L002

description:
DM_4-2FTS_P2_F
DM_4-2FTS_counts.filtered.+1.normFinal.txt

value: F


title: DM4-2FTSP1GS90_L002

description:
DM_4-2FTS_P1_G
DM_4-2FTS_counts.filtered.+1.normFinal.txt


title: CD34+-derived proE/basoE cells, ctrl input, NT

data_processing:
Reads were trimmed and filtered using fastp (v0.23.4) to remove low quality reads, low complexity sequences, as well as polyG tails. Reads were mapped to the human reference genome (GENCODE, GRCh37) with Bowtie2 (v2.3.5.1), using parameters --no-discordant and --no-mixed. Samblaster (v0.1.24) was used to remove duplicated reads, then Samtools (v1.10) to generate sorted (by coordinates) BAM files, with a cut-off for MAPQ score of 10.
Peaks were called for each replicate with the MACS algorithm (MACS3) in broad mode, with the input DNA as control as well as the same sample treated with RNH1, at q-value < 0.1. Resulting peaks were analysed by groups of biological replicates with MSPC (v5.5.0) with default cut-off.
To identify differentially expressed R-loops, we intersected the peaks from MACS3 calling with the 5.5 million of restriction fragments generated before S9.6 immunoprecipitation, using the resulting matrix as a reference frame for featureCounts (v2.0.0). Batch effect removal is necessary as samples has been sequenced at different times. Normalization and differential R-loop expression analysis between wild-type, mutant and control samples was performed with DESeq2, using at Benjamini-Hochberg (BH)- adjusted P-value < 0.05 and log2 (FC) > |1|.
Assembly: GRCh37
Supplementary files format and content: Compressed bedGraph file. Considering exclusion of certain files, depending of the needed comparison (HDACi treatment, genotype), bedgraph are not normalized.

source_name: CD34+-derived proE/basoE

extract_protocol:
8x10^6 cells were harvested and processed for genomic DNA extraction by SDS/ Proteinase K treatment at 37°C followed by phenol-chloroform extraction and ethanol precipitation. 8 µg of genomic DNA was fragmented using HindIII, EcoRI, BsrGI, XbaI, and SspI. Then DNA pre-treated or not with RNase H1 (16UI/mL) overnight (New Engl Biolabs, Ipswitch, MA) was immunoprecipitated using S9.6 antibody overnight at 4°C.
For DRIP-seq, good quality DRIPed DNA samples (with and without RNase H1-treatment) were sonicated to get an average length 200-300 bp. Then samples were subjected to end-repair, dATP tailing, adaptor ligation and library indexing. Lastly, the libraries were cleaned up using AMPure beads, and amplified. After checking library quality on Agilent Bioanalyzer using Agilent High Sensitivity DNA 1000 kit, libraries were sequenced on Illumina NovaSeq instruments.

value: CD34+-derived proE/basoE

value: ctrl

tag: antibody


title: ce_mRNA_ebax_TC_N2_25h


title: CD34+-derived proE/basoE cells, ctrl input, RH

value: Rnase H1


title: DM4-2FTSP2CS100_L002

description:
DM_4-2FTS_P2_C
DM_4-2FTS_counts.filtered.+1.normFinal.txt

tag: tissue

value: whole body

value: GLTED-wt

value: C


title: Primary Lung fibroblast, Positive control, biol rep 3

city: Seodaemun-gu

first: Chanho

last: Lee

state: Seoul

address: 50-1 Yonsei-ro

department: Department of Internal Medicine

country: South Korea

institute: Yonsei University College of Medicine

description: processed data file's column name: ./sample9_transcript.bam

data_processing:
Base call files from mRNA sequencing were converted to FASTQ format using BCL Convert v3.10 (Illumina). The adapter sequences were trimmed during this process.
After trimming the raw reads using Trimmomatic v0.39, the paired reads were aligned to the GRCh38 human reference transcriptome (Ensembl release 106) by HISAT2 v.2.2.1.
Alignment counts were summarized using the featureCounts program of the Subread package v.2.0.3.
Data normalization and differential expression analysis were performed using DESeq2 in R.
Assembly: GRCh38 (Ensembl release 106)
Supplementary files format and content: tab-delimited text file includes alignment counts without normalization.

source_name: Primary Lung fibroblast

growth_protocol: Primary human lung fibroblasts were isolated and cultured in EMEM supplemented with 15% fetal bovine serum and antibiotics in humified atmosphere with 5% CO2 at 37℃.

extract_protocol:
RNA was harvested using Rneasy mini kit (QIAGEN).
RNA libraries for mRNA (polyA RNA) sequencing were prepared using Illumina stranded mRNA prep kit following the manufacturer's protocols.

treatment_protocol: At passage 2, the cells were seeded into 60mm cell dishes and incubated overnight. After 1.5 hours of serum starvation with 0.1% fetal bovine serum-supplemented media, 2 ng/ml TGFβ1 and/or 20μM ezetimibe was treated and incubated 24 hours before harvest.

tag: cell type

value: Primary Lung fibroblast

value: TGFβ1 + / ezetimibe -


title: ce_mRNA_ebax_TC_N2_24h

tag: genotype


title: ce_mRNA_ebax_TC_N2_18h

tag: treatment


title: QGP-1_PSC conditional media replicate 2

type: RNA

city: Taipei

first: Yu-Wen

last: Tien

address: 7 Chung-Shan S.Rd.

country: Taiwan

institute: NTUH

hyb_protocol: Phalanx OneArray Plus Protocol

scan_protocol: Agilent 0.1 XDR Protocol

data_processing: GPR files were loaded into Rosetta Resolver ®  System (Rosetta Biosoftware) to process data analysis.   1.  Rosetta profile error model calculation: the error due to random factors and systematic biases are estimated   by an error model.     2.  Squeeze replicated probes: the repeated probes within one chip are averaged.   3.  Normalize intensities: Median scaling performed on data set without flagged and control data.

label: NHS-Cy5 Dye

source_name: pancreatic neuroendocrine tumor cell line

label_protocol: Target preparation was performed using an Eberwine-based amplification method with Amino Allyl   MessageAmp II aRNA Amplification Kit (Ambion, AM1753) to generate amino-allyl antisense RNA(aa-aRNA).   Labeled aRNA coupled with NHS-CyDye was prepared and purified prior to hybridization. Purified coupled aRNA   was quantified using NanoDrop ND-1000; pass criteria for CyDye incorporation efficiency at > 15 dye   molecular/1000nt.

growth_protocol: QGP-1 were cultured in 10% FBS(Gibco, #10437-028) and Roswell Park Memorial Institute (RPMI)-1640 medium (Gibco, #11875-085) according to the manufacturer’s instructions.

extract_protocol: Total RNA was extracted using NucleoSpin RNA (Macherey-Nagel).

treatment_protocol: QGP-1 were treated in 0.1%FBS in RPMI-1640 with or without conditional media of PSCs for 24 hr.

tag: cell line

value: PanNET cell line QGP-1

value: with PSC conditional media


title: Primary Lung fibroblast, Positive control, biol rep 2

description: processed data file's column name: ./sample8_transcript.bam


title: DM4-2FTSP1CS92_L002

description:
DM_4-2FTS_P1_C
DM_4-2FTS_counts.filtered.+1.normFinal.txt

tag: conc.


title: Primary Lung fibroblast, Positive control, biol rep 1

description: processed data file's column name: ./sample7_transcript.bam


title: QGP-1_PSC conditional media replicate 1


title: QGP-1_Mock_replicate 2

value: Mock


title: F5 CD8 T cells, GEX

city: Lyon

first: Simon

last: de Bernard

address: 30 rue Pré Gaudry

institute: AltraBio

description: scRNA-seq with feature barcode technology

data_processing:
Quality control of raw data was performed using FastQC
The TSO, poly-A sequences and low-quality bases were trimmed using cutadapt
Transcript expression quantification was performed using the alevin-fry pipeline and version M31 ofGENCODE mouse genome and annotations
The gene/count matrix was generated with fishpond.
Assembly: mm10
Supplementary files format and content: fishpond output files for rna and hto(directories "quant/crlike")

source_name: Spleen

growth_protocol: Briefly, 5x10^4 naive F5 CD8 T cells were activated in vitro for 3, 4 and 5 days in the presence or absence of 5% IL-2.

extract_protocol:
Cells were barcoded with hashtag oligonucleotides (HTO) antibodies specific for each condition (TotalSeq™-B – Biolegend) for 30 minutes on ice. After washing with FACS buffer, cells were sorted on a FACS Aria III cytometer (Becton, Dickinson and Company). Sorted cells were immediately loaded into the Chromium Controller (10X genomics, Pleasanton, California) to generate single-cell gel beads-in-emulsion (GEM).
The scRNA-seq library was generated using Chromium Next GEM Single Cell 3′ reagent v.3.1 with feature barcode technology (10X Genomics), according to the manufacturer’s protocol.After reverse transcription, gel-beads in emulsion were disrupted, barcoded cDNA were amplified by PCR and then size-selected to separate 3’ gene expression and cell surface protein (HTO) library construction according to the manufacturer's instructions. On one hand, the gene expression library was fragmented, end repaired and sample indexes were added by PCR. On the other hand, sample indexes were added by PCR for the HTO library. The purified libraries were quantified using the Library Quantification Sample Kit Kapa (Illumina-Uni San Diego, California). Gene expression and HTO libraries were pooled in a 4/1 ratio and paired-end (2x150pb) sequenced (375M reads) on a HiSeq X platform (Illumina) with 1% of PhiX.

treatment_protocol: Cells of each experimental conditions were FACS stained using the following antibodies for surface staining : CD8 (53.6.7), CD8 (), CD45.1 (A20), CD45.2 (104), CD11b (M1/70), CD11c (HL3), CD19 (1D3) from BD Biosciences and CD44 (IM7.8.1) from Biolegend.

value: Spleen

value: CD8 T lymphocytes

value: mixed sample (Day 0, Day 3 without ex-IL-2, Day 3 with ex-IL-2, Day 4 without ex-IL-2, Day 4 with ex-IL-2, Day 5 without ex-IL-2, Day 5 with ex-IL-2)

tag: library type

value: mRNA


title: CD34+-derived proE/basoE cells, SF3B1 WT, rep RH 1

value: SF3B1 WT

value: S9.6 antibody


title: QGP-1_Mock_replicate 1


title: Primary Lung fibroblast, ezetimibe treated, biol rep 3

description: processed data file's column name: ./sample6_transcript.bam

value: TGFβ1 + / ezetimibe +


title: DM4-2FTSP1BS93_L002

description:
DM_4-2FTS_P1_B
DM_4-2FTS_counts.filtered.+1.normFinal.txt

tag: conc. units

value: ug/L

tag: conc code

value: B


title: F5 CD8 T cells, HTO

library_source: other

molecule: protein

value: HTO

tag: hto

value: TotalSeq™-B0301 anti-mouse Hashtag 1, TotalSeq™-B0302 anti-mouse Hashtag 1, TotalSeq™-B0303 anti-mouse Hashtag 1, TotalSeq™-B0304 anti-mouse Hashtag 1, TotalSeq™-B0305 anti-mouse Hashtag 1, TotalSeq™-B0306 anti-mouse Hashtag 1, TotalSeq™-B0310 anti-mouse Hashtag 1


title: DM4-2FTSP1AS94_L002

description:
DM_4-2FTS_P1_A
DM_4-2FTS_counts.filtered.+1.normFinal.txt
DM_6-2FTS_counts.filtered.+1.normFinal.txt

value: whole body

value: GLTED-wt

value: A

tag: exposure duration

tag: age

tag: replicate plate_id


title: ce_mRNA_ebax_TC_N2_19h

tag: strain

value: WT


title: CD34+-derived proE/basoE cells, SF3B1 WT, rep NT 1

value: CD34+-derived proE/basoE

value: no


title: Primary Lung fibroblast, ezetimibe treated, biol rep 2

description: processed data file's column name: ./sample5_transcript.bam

value: Primary Lung fibroblast


title: CD34+-derived proE/basoE cells, SF3B1 WT, rep RH 2

value: Rnase H1


title: Primary Lung fibroblast, ezetimibe treated, biol rep 1

description: processed data file's column name: ./sample4_transcript.bam


title: CD34+-derived proE/basoE cells, SF3B1 MUT, rep RH 2

value: SF3B1 MUT


title: TH-_2

city: Pasadena

first: Brittany

middle: Megan

last: Edens

address: 1200 E California Blvd

institute: Caltech

data_processing:
Reads were mapped to the Stowers Institute SIMRBASE Petromyzon marinus reference genome, kPetMar1.0 (https://simrbase.stowers.org/sealamprey?pane=resource-1) using Kallisto
Differential expression was determined using the DEseq2 package in R
Assembly: kPetmar1
Supplementary files format and content: tab-separated values file includes counts for each sample

organism: Petromyzon marinus

source_name: ammocete trunk

extract_protocol:
RNA was extracted using the NucleoSpin RNA SX kit (Takara), and cDNAs were synthesized using the SMARTseq v4 ultra low input kit (Takara)
RNA libraries for RNA-seq were prepared using NEBext Ultra II Kit (NEB) following manufacturer's protocols

treatment_protocol: Tissues were dissociated and fixed in paraformaldehyde followed by antibody-based selection for TH+ and TH- cells. Samples underwent reverse crosslinking prior to RNA extraction

value: ammocete trunk

value: TH- control


title: StereoSeq_MouseBrain_Rep1

city: Leuven

first: Kristofer

middle: James

last: Davie

address: Herestraat 49

department: Center for Brain and Disease Research

country: Belgium

institute: Vlaams Instituut voor Biotechnologie

description: Fresh frozen Stereo-seq spatial transcriptome sequencing

data_processing:
BGI's SAW pipeline (v7.0.0) was used to process fastq files into resulting gene expression matrices.
Assembly: mm10
Supplementary files format and content: ipr and tar.gz: BGI formatted image files
Supplementary files format and content: barcodeToPos.h5: Stereo-seq mask file
Supplementary files format and content: counts.gef: GEF formatted counts file including X/Y coordinates
Library strategy: Spatial Transcriptomics (Stereo-seq)

growth_protocol: Six to eight weeks old mice (C57BL/6J) used in the study were kept on a 14 h light, 10 h dark–light cycle from 7:00 to 21:00

extract_protocol:
Brains were dissected and immediately snap-frozen in isopentane for 10 min. Afterwards, brains were embedded in Tissue-Tek OCT cryo embedding compound, 10μm sections were cut using a cryostat and placed on the Stereo-seq chip
Tissue optimization was performed using the Stereo-seq Permeabilization kit according to the manufacturer’s protocol. Following this, the spatial transcriptomics analysis was performed using the Stereo-seq Transcriptomics kit according to the manufacturer’s protocol.

value: Brain


title: TH-_1


title: CD34+-derived proE/basoE cells, SF3B1 WT, rep NT 2

value: SF3B1 WT

value: S9.6 antibody


title: Primary Lung fibroblast, Negative control, biol rep 3

description: processed data file's column name: ./sample3_transcript.bam

value: TGFβ1 - / ezetimibe -


title: GDKO, LPS [DNBSEQ]

city: Beijing

first: Minmin

last: Luo

address: 7 Science Park Road, ZGC Life Science Park

country: China

institute: National Institute of Biological Sciences, Beijing

description: 10x Genomics

data_processing:
Demultiplexing was performed using MGI.Lite.BaseCall_1.2.0.99. Barcode processing, gene counting and aggregation were performed using the Cell Ranger software v6.0.1, with the "--include-introns" flag added, according to manufacturer's suggestion. For GDKO PBS, GDKO LPS, the digital expression matrix generated using default parameters contained large fraction of empty droplets. Therefore, fastq files from these two experiments were re-analyzed using the parameter --force-cells=8000 (GDKO PBS) and --force-cells=9000 (GDKO LPS), respectively.
Assembly: mm10
Supplementary files format and content: Preprocessed gene-by-cell matrix in the CellRanger h5 output format.
Supplementary files format and content: Tab-separated values files and matrix files (barcodes.tsv, features.tsv, matrix.mtx).

source_name: brain, motor cortex

extract_protocol:
For single cell dissociation, artificial cerebral spinal fluid (ACSF) was prepared using the “protective cutting” formula. Before the experiment, ACSF was bubbled with carbogen gas (95% O2 and 5% CO2) for ~30 mins. Mice were anesthetized and perfused transcardially with ACSF. The brain was quickly removed and submerged in carbogen bubbled ice-cold ACSF. 8 brain slices (200μm) were sectioned using vibratome and the motor cortex was micro-dissected. The dissected tissue pieces were then digested using the Papain dissociation system, supplemented with DNaseI. After 30 mins, the digestion was quenched by exchanging the solution with quenching (Q) buffer (ACSF with 1% FBS and 0.2% BSA) 3 times. The quenched solution was placed on ice for 5 mins and then proceeded to trituration. The tissue pieces were gently triturated through Pasteur pipettes with fire polished tip openings at 3 different diameters (600μm, 300μm, 150μm). The cell suspension was then centrifuged at 100g for 10 mins and resuspended in Q buffer for FACS sorting. Single cells were sorted (BD Aria III) using a 130μm nozzle and a sheath pressure of 10 psi. 15mL conical tube containing ~12mL collection (C) buffer (ACSF with 13.2mM trehalose, 2% FBS and 1% BSA) was used for collecting the target population.
Library construction was performed according to the manufacturer’s instructions (single cell 3’ v3.1 protocol, 10x Genomics). Briefly, dissociated brain cells were resuspended in the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscribed to cDNA. For each library, cDNA were amplified with polymerase chain reaction (PCR). The purified product were then converted to DNA nanoballs (MGIEasy Universal Library Conversion Kit) for sequencing with the MGISEQ-2000 or DNBSEQ-T7 model.

treatment_protocol: PBS or LPS (40mg/kg) were administered ~8 hours prior to single cell dissociation.

value: brain, motor cortex

value: Gsdmd-/-

value: LPS

tag: dose

tag: time

tag: Sex

value: female

value: Adult (6-8 weeks)


title: SF3B1 MUT t0, rep 2

value: G1E-ER4 isogenic SF3B1 K700E


title: DM4-2FTSP1FS91_L002

description:
DM_4-2FTS_P1_F
DM_4-2FTS_counts.filtered.+1.normFinal.txt

data_processing:
RNA sequencing was performed at the Michigan State University Research Technology Support Facility Genomics Core. Libraries were quality control checked and quantified using a combination of Qubit dsDNA HS, Agilent 4200 TapeStation HS DNA1000, and Invitrogen Collibri Library Quantification qPCR assays. Each pool (corresponding to ≈ 96 samples from assays with two PFAS) was loaded onto one lane of a NovaSeq S4 flow cell. Sequencing was performed in a 2x150bp paired end format using a NovaSeq 6000 v1.5 300 cycle reagent kit (target read depth of ≈28,000 MB/sample). Base calling was done by Illumina Real Time Analysis v3.4.4, and the output was demultiplexed and converted to FASTQ format with Illumina Bcl2fastq v2.20.0.
The Daphnia magna reference genome (assembly ASM2063170v1.1, RefSeq GCF_020631705.1) was indexed with STAR version 2.7.10a using the following sample specific parameters: --genomeSAindexNbases 12 --sjdbOverhang 149.
After performing quality control on the raw FASTQ data using FastQC and MultiQC version 1.12, raw reads were trimmed using Trim Galore! version 0.6.6, with the parameters --illumina --stringency 3 --quality 20 –e 0.1 --length 20 --paired, to remove sequencing adapters and low-quality bases and filter out trimmed reads shorter than 20 bp.
Trimmed reads were then mapped with STAR version 2.7.10a to the indexed genome using the default parameters except for the use of: --outSAMattributes NH HI AS NM XS. Samtools version 1.15.1 was used to sort and index the alignment files.
HTSeq version 2.0.1 was used to assign counts to each gene with the following parameters: -f bam -r pos -s no. FastQC and MultiQC version 1.12 were used to assess quality control metrics of the processed reads. Count data for treatment and plate-specific control samples for each PFAS were written to a single tab-delimited matrix file, where each column represented a sample and each row represented a gene.
Additional quality control and filter steps were applied to each count matrix using the R statistical computing software (R Core Team, 2022) prior to concentration response modeling. Lowly expressed genes were removed from the raw count matrices using edgeR’s (Robinson et al. 2010) built-in filtering function, filterByExpr(), in version 3.40.2; namely, genes with < 15 reads across all samples and < 10 reads in the smallest group (treatment or control) sample size were filtered out. Normalization factors for the filtered count matrices were calculated using the trimmed mean of M-values method via the edgeR function calcNormFactors() which was used for counts per million mapped reads (CPM) normalization. The filtered CPM normalized count matrix was log2 transformed with a pseudo-count of 1 added (to prevent occurrence of any negative count values for downstream analyses; i.e., log2(CPM+1)).
Assembly: Daphnia magna reference genome (assembly ASM2063170v1.1, RefSeq GCF_020631705.1)
Supplementary files format and content: Count matrices provided as tab delimited text files. Top row = sample names. Second row = treatment concentration (ug/L); all subsequent rows provide filtered counts per million (CPM) log2 transformed with a pseudo-count of 1 added (to prevent occurrence of any negative count values for downstream analyses; i.e., log2(CPM+1), with each row corresponding to a separate transcript and each column corresponding to a sample. One count matrix file per chemical exposure experiment.

extract_protocol:
RNA was extracted from pooled homogenates using Zymo Research Direct-zol-96 RNA kit (Zymo Research) protocol adapted to the Biomek i5 to yield 40 µl of RNA. Previously extracted control RNA was added to empty wells on the final RNA plate. Concentrations of RNA for each well were measured on a NanoDrop spectrophotometer (Thermo Scientific, NanoDrop Eight) and diluted if necessary.
RNA extracts (10 µl) were used to prepare cDNA libraries with the Lexogen Poly(A) RNA Selection Kit V1.5 (catalog #157; Lexogen GmbH, Austria) and Lexogen CORALL mRNA-Seq Library Prep Kit with UDIs (catalog #158-163) both adapted to the Biomek i5. Resulting cDNA library samples were evaluated on a 4200 TapeStation (Agilent Technologies, CA, USA) using the D1000 ScreenTape (Agilent) analysis protocol to assess cDNA quality. Samples were quantified using the Quant-iT Picogreen dsDNA assay kit (catalog # P11495; Thermo Fischer Scientific), diluted if necessary, and equal portions of each sample pooled for use in RNA sequencing by Michigan State University.

tag: tissue

value: whole body

value: GLTED-wt

value: ug/L

value: F


title: CD34+-derived proE/basoE cells, SF3B1 MUT, rep NT 2

tag: cell type

value: CD34+-derived proE/basoE

value: SF3B1 MUT

value: no

tag: antibody


title: CD34+-derived proE/basoE cells, SF3B1 MUT, rep RH 1

tag: genotype

tag: treatment

value: Rnase H1


title: CD34+-derived proE/basoE cells, SF3B1 MUT, rep NT 1


title: TH-_3

value: ammocete trunk

value: TH- control


title: GDKO, PBS [DNBSEQ]

description: 10x Genomics

value: PBS


title: WT, LPS [DNBSEQ]

description: 10x Genomics

value: brain, motor cortex

value: wild type


title: WT, PBS [DNBSEQ]

description: 10x Genomics


title: TH+_1

value: TH+ sympathetic


title: TH+_2


title: Primary Lung fibroblast, Negative control, biol rep 1

description: processed data file's column name: ./sample1_transcript.bam

value: Primary Lung fibroblast

value: TGFβ1 - / ezetimibe -


title: Primary Lung fibroblast, Negative control, biol rep 2

description: processed data file's column name: ./sample2_transcript.bam


title: TH+_3


title: KI1_preg_3.F

status: Public on Feb 21 2024

city: Vienna

first: Mihaela

last: Pavlicev

address: Djerassiplatz 1

department: Evolutionary Biology

country: Austria

institute: University of Vienna

data_processing:
Sequences were aligned to the mouse genome with STAR to generate raw count data
Count data was analyzed, including normalization and differential expression, in R (version 4.1.1) with the program DEseq2
Assembly: GRCm38
Supplementary files format and content: tab-delimited text file includes normalized expression for all samples and differential expression analysis

source_name: Decidua

extract_protocol:
Tissues were homogonized in lysis buffer and mechanical disruption with Precellys Evolution homogenizer and RNA was isolated with the RNeasy mini kit
Library preparation was conducted by the CCHMC sequencing facility for uterine proestrus samples and Novogene for the decidua 17.5dpc pregnant samples

value: Decidua

value: Transgenic


title: KI1_preg_2.F


title: P280_27_L9_14_log2 Signal

city: Gießen

first: Robert

last: Ringseis

address: Heinrich-Buff-Ring 26-32

department: Institute of Animal Nutrition and Nutrition Physiology

country: Germany

institute: JLU Gießen

description: P280_27_L9_14_log2 Signal

hyb_protocol: 3.8 µg of fragmented and labeled ss cDNA were hybridized to Applied Biosystems GeneChip Clariom S rat arrays for 16 h at 45°C and 60 rpm in an Applied Biosystems GeneChip hybridization oven 640.

scan_protocol: Hybridized arrays were washed and stained in an Applied Biosystems GeneChip Fluidics Station FS450, and the fluorescent signals were measured with an Applied Biosystems GeneChip Scanner 3000 7G System. Fluidics and scan functions were controlled by the Applied Biosystems GeneChip Command Console v4.3.3 software.

data_processing: Summarized probe set signals in log2 scale were calculated by using the GCCN-SST-RMA algorithm with the Applied Biosystems GeneChip Expression Console v1.4 Software. After exporting into Microsoft Excel, average signal values, comparison fold changes (FC) and significance P-values were calculated.

label: Biotin

organism: Gallus gallus

source_name: Liver

label_protocol: 200 ng of total RNA was used to generate double-stranded cDNA. 12 µg of subsequently synthesized cRNA were purified and reverse transcribed into single-stranded (ss) cDNA, whereat unnatural dUTP residues were incorporated. Purified ss cDNA was fragmented using a combination of uracil DNA glycosylase (UDG) and apurinic/apyrimidinic endonuclease 1 (APE 1) followed by a terminal labeling with biotin.

growth_protocol: The feeding trial was approved by the Animal Welfare Office of the Justus-Liebig-University Giessen (approval no. JLU-Nr. 806_M). All experimental procedures described followed established guidelines for the care and handling of laboratory animals.

extract_protocol: Total RNA was isolated using TRIzol reagent (Invitrogen, Karlsruhe) according to the manufacturer’s protocol. RNA quantity and quality were assessed spectrophotometrically using an Infinite 200M microplate reader equipped with a NanoQuant plate (both from Tecan, Mainz). The average RNA concentration and the A260/A280 ratio of all total RNA samples (n = 20, means ± SD) were 424 ± 78 ng/μL and 1.89 ± 0.05, respectively.

treatment_protocol: The experiment included 72 male, one-day-old broiler chickens (Cobb 500, Cobb, Wiedemar, Germany) which were assigned to three treatment groups with similar initial body weights (BW). Broilers were kept in groups of 4 broilers each in 2.1 m2 cages which were equipped with feed automates and nipple drinkers providing free access to feed and water. The floor of the cages was lined with a cardboard, which was covered with litter to allow pecking, scratching and dustbathing. The cardboards and litter were changed twice per week in the first 2 wk and every 2 d for the last 3 wk. The light regime and the room temperature followed the recommendation of the breeder (Cobb-Vantress, 2018a). The light intensity was 40 Lux and the light regime was (light:dark) 24 h:0 , 23 :1 , 22 :2 , 21 :3 , 20 :4 , 19 :5  for d 1, 2, 3, 4, 5, 6 and 18 :6  from d 7 to 35. The room temperature was 33°C on d 1, measured at pen height, and decreased to 19°C on d 35. The temperature at the cage floor was adjusted with infrared lamps (Albert Kerbl GmbH, Buchbach, Germany) during the first 6 d. The three groups (HI0, HI7.5, HI15) were fed three different diets, which contained different inclusion levels of a commercially available partially defatted HI larvae meal (Madebymade, Pegau, Germany) as protein source, in a three-phase feeding system (starter period: d 1-10, grower period: d 11-21, finisher period: d 22-35). The concentration of the HI larvae meal in the diets HI0, HI7.5 and HI15 was 0, 75 and 150 g/kg diet, respectively. HI larvae meal was included in the diets at the expense of soybean extraction meal, soybean oil and wheat. The reduction of the amounts of wheat and soybean oil was necessary due the lower protein content and the higher fat content of the HI larvae meal in comparison with soybean meal. The amino acid concentrations of the diets were adjusted with synthetic amino acids to reach the recommended concentrations of digestible essential amino acids (Cobb-Vantress, 2018b). The composition of the diets is shown in Table 1. The finisher diets contained 0.5% TiO2 in order to determine the apparent ileal digestibility (AID) coefficient for amino acids by the indicator method (Short et al., 1996). Individual BW of the broilers were recorded on d 1, 10, 21 and 35. The feed intake of each cage was recorded daily.

tag: sample group

value: 7.5% defatted HI meal

value: Liver

value: male

tag: breed

value: Cobb 500 broiler


title: P280_25_L7_2_log2 Signal

description: P280_25_L7_2_log2 Signal


title: P280_29_L11_29_log2 Signal

description: P280_29_L11_29_log2 Signal


title: P280_28_L10_23_log2 Signal

description: P280_28_L10_23_log2 Signal


title: KI1_preg_1.F


title: P280_31_L13_6_log2 Signal

description: P280_31_L13_6_log2 Signal

value: 15% defatted HI meal


title: P280_36_L18_33_log2 Signal

description: P280_36_L18_33_log2 Signal


title: P280_26_L8_11_log2 Signal

description: P280_26_L8_11_log2 Signal


title: P280_24_L6_31_log2 Signal

description: P280_24_L6_31_log2 Signal

value: 0% defatted HI meal


title: P280_35_L17_30_log2 Signal

description: P280_35_L17_30_log2 Signal


title: P280_34_L16_24_log2 Signal

description: P280_34_L16_24_log2 Signal


title: P280_32_L14_12_log2 Signal

description: P280_32_L14_12_log2 Signal


title: P280_33_L15_18_log2 Signal

description: P280_33_L15_18_log2 Signal


title: P280_23_L5_28_log2 Signal

description: P280_23_L5_28_log2 Signal


title: P280_30_L12_32_log2 Signal

description: P280_30_L12_32_log2 Signal


title: P280_22_L4_22_log2 Signal

description: P280_22_L4_22_log2 Signal


title: WT.PE.926.F

source_name: Uterus

value: Uterus

value: Wildtype

value: Proestrus


title: P280_20_L2_7_log2 Signal

description: P280_20_L2_7_log2 Signal


title: TG.PE.857.F


title: WT.PE.933.F


title: WT.PE.307.F


title: TG.PE.931.F


title: Barcode sequencing

first: Hani

last: Goodarzi

department: Department of Biochemistry and Biophysics

institute: UCSF

data_processing:
Cell Ranger (version 3.0.1, 10X Genomics) with default parameters was used to align reads and generate digital expression matrices from single-cell sequencing data. To assign cell genotypes, a bwa reference (H. Li and Durbin 2009) database was created containing all guide sequences present in the library using bwa index command. The barcode-enrichment libraries were mapped to this database to establish the guide identities; to detect the cell barcodes, the barcode correction scheme used in Cell Ranger was used (the mapping of uncorrected to corrected barcodes was extracted from Cell Ranger analysis run of the whole transcriptome libraries; this mapping was then applied to the reads of barcode-enrichment libraries). UMI correction was performed by merging the UMIs within the hamming distance of 1 from each other. For each UMI, the guide assignment was done by choosing the guide sequence most represented among the reads containing the given UMI. To make the final assignment of a guide to cell barcodes, we only considered the barcodes that were represented by at least 5 different UMIs, with >80% UMIs representing the same guide.  Data filtering was performed using scanpy package (F. A. Wolf, Angerer, and Theis 2018). Data were denoised using a modification of scvi autoencoder (Gayoso et al. 2022) with loss function penalizing for the similarity between cells having different RBPs knocked down. The distance between transcriptome profiles of individual RBP knockdowns was calculated by applying the t-test to individual gene counts across the cells that were assigned the respective guide sequence.
Assembly: hg38
Supplementary files format and content: Tab-separated values files and matrix files.
Supplementary files format and content: Cell barcode to sgRNA comma-separated mapping file

source_name: sgR

growth_protocol: All cells were cultured in a 37°C 5% CO2 humidified incubator. The 293T cells (ATCC CRL-3216) were cultured in DMEM high-glucose medium supplemented with 10% FBS, glucose (4.5 g/L), L-glutamine (4 mM), sodium pyruvate (1 mM), penicillin (100 units/mL), streptomycin (100 μg/mL) and amphotericin B (1 μg/mL) (Gibco). The K562 cell line was cultured in RPMI-1640 medium supplemented with 10% FBS, glucose (2 g/L), L-glutamine (2 mM), 25 mM HEPES, penicillin (100 units/mL), streptomycin (100 μg/mL) and amphotericin B (1 μg/mL) (Gibco). All cell lines were routinely screened for mycoplasma with a PCR-based assay.

extract_protocol: 68 RBPs were chosen for Perturb-seq analysis based on the clustering analysis of ENCODE eCLIP dataset and DeepBind dataset (Alipanahi et al. 2015). Perturb-seq experiment was performed as previously described (Datlinger et al. 2017). Briefly, a library of 205 sgRNAs (5 non-targeting sgRNAs and 200 sgRNAs targeting 100 genes, 2 sgRNAs per gene) was ordered as a pooled oligonucleotide library from Twist Bioscience with the following design:  [ATCTTGTGGAAAGGACGAAACACCG]-[Protospacer Sequence]-[GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGC]  The library was PCR-amplified using Q5 Hot Start High-Fidelity 2X Master Mix (NEB) with the primers with the following sequences: 5’-ATCTTGTGGAAAGGAC-3’ and 5’-GCCTTATTTTAACTTGCTA-3’. To clone libraries into CROPseq-Guide-Puro vector (Addgene 86708), the starting vector was digested with BsmBI following the protocol outlined in (Sanjana, Shalem, and Zhang 2014). The library was cloned into the digested backbone using Gibson Assembly method (Thomas, Maynard, and Gill 2015). The reaction product was transformed into Takara Stellar competent cells according to manufacturer recommendations, grown overnight in 100 mL LB with ampicilin and purified using ZymoPURE II Plasmid Midiprep Kit. K562 cells were infected with the plasmid library at a low multiplicity of infection to minimize double infection. The infected cells were selected with 2 µg/mL puromycin for 3 days. Live cells were isolated on a flow cytometer (FACSAria II) by propidium iodide staining. Approximately 5000 live cells were captured by 10X Chromium Controller using Chromium Single Cell 3’ Reagent Kits v2. Sample preparation was performed according to the manufacturer’s protocol. Samples were sequenced on a NovaSeq 6000 using the following configuration: Read 1: 28, i7 index: 8, i5 index: 0, Read 2: 98.  To facilitate sgRNA assignment, sgRNA-containing transcripts were additionally amplified by PCR reactions by modifying a previously published approach (Hill et al. 2018). The following primers were used for amplification: 5'-AATGATACGGCGACCACCGAGATCTACAC-3' and 5'-CAAGCAGAAGACGGCATACGAGATTACGACAGGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTggactatcatatgcttaccgtaacttgaaag-3'. PCR product was cleaned up by 1.0x SPRI beads (SPRIselect; BECKMAN COULTER; Cat. No. B23317). Samples were sequenced using paired-end 150 bp sequencing on an Illumina MiSeq sequencer.

treatment_protocol: K562 cells expressing dCas9-KRAB fusion protein were constructed by lentiviral delivery of pMH0006 (Addgene #135448) and FACS isolation of BFP-positive cells.  The lentiviral constructs were co-transfected with pCMV-dR8.91 and pMD2.D plasmids using TransIT-Lenti (Mirus) into 293T cells, following the manufacturer’s protocol. The virus was harvested 48 hours post-transfection and passed through a 0.45 µm filter. Target cells were then transduced overnight with the filtered virus in the presence of 8 µg/mL polybrene (Millipore).  Guide RNA sequences for CRISPRi-mediated gene knockdown were cloned into pCRISPRia-v2 (Addgene #84832) via BstXI-BlpI sites. After transduction with sgRNA lentivirus, K562 cells were selected with 2 µg/mL puromycin (Gibco). Knockdown of target genes was assessed by RT-qPCR using PerfeCTa SYBR Green SuperMix (QuantaBio) per the manufacturer’s instructions. HPRT1 was used as endogenous control.

tag: cell line

value: sgR


title: melanocyte, KB3

city: Hangzhou

first: Yingzhe

last: Cui

address: Jiefang road

institute: Second Affiliated Hospital, Zhejiang University School of Medicine

data_processing:
The count tables of Smart-seq data were subjected to the variance stabilizing transformations (VST) function from DESeq2
Assembly: GRCM38
Supplementary files format and content: VST readcount

source_name: back epidermis

extract_protocol:
Total RNA was extracted usingTrizol reagent following the manufacturer's procedure. The total RNA quantity and purity were analysis of Bioanalyzer 2100 and RNA 6000 Nano LabChip Kit, high-quality RNA samples with RIN number > 7.0 were used to construct sequencing library.
RNA from each sample was used to generate RNA-seq libraries using a SMART-Seq v4 Ultra Low Input RNA Kit (Clontech, Japan). After quality inspection of the library, Illumina Novaseq™ 6000 was used for sequencing with a read length of 2 * 150 bp (PE150) for paired-end sequencing.

value: back epidermis

value: melanocyte

value: Spry1flox/flox k14-Cre-ERT2


title: P280_21_L3_13_log2 Signal

description: P280_21_L3_13_log2 Signal


title: melanocyte, KB2


title: Vehicle_A

city: Belfast

first: Francesca

last: Amoroso

address: 97 Lisburn Road

institute: Queen's University Belfast

data_processing:
Sequencing was performed on Illumina MiSeq (150) 75 P.E at 2M reads/ sample.
Fastq files were generated using bcl2fastq version 2.19 using the default thresholds.
Reads were aligned to the human genome and analysed using the miRNA Quant tool on the QIAGEN portal using default thresholds (IPA Target Filters are listed among the processed data).
Raw counts were used as input for differential expression analysis using DESeq2, grouping samples in the same ARM compared to the Vehicles.
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: Gene sets were filtered to contain genes upregulated after treatment (FC >1) and downregulated (FC <0.5).

source_name: Prostate Cancer cells

growth_protocol: Cells were injected subcute in nude-nude mice (2x106 in matrigel). When the tumours were visible, the mice were treated as described above. The tumours were harvasted at maximum size of 1000 mm3 or after 21 days of daily treatment.

extract_protocol:
The RNA was extracted from the cells and collected utilizing the column-based method through the miRNeasy Mini Kit (QIAGEN, Hilden, Germany).
The miRNA library preparations were performed with the QIAseq miRNA library prep kit (QIAGEN, Hilden, Germany) according to the manufacturer’s instructions.

treatment_protocol: To obtain single cells, the tumours were dissociated  through mechanical and enzymatic digestion with Collagenase IV (1%) at 37˚C.

tag: treatments

value: 0.1 ml 5% benzyl alcohol and 95% safflower oil solution via IP every day

tag: experiment

value: in vivo (xenograft)

value: nude-nude

value: tumour explant


title: Untr_B

value: Untreated


title: Vehicle_B


title: P280_19_L1_4_log2 Signal

description: P280_19_L1_4_log2 Signal

value: Liver


title: ARM5_D

value: Mycophenolate mofetil (100mg/kg), Abiraterone (0.5 mmol/kg/d in vehicle) and prednisolone phosphate (20mg/kg).


title: Vehicle_C


title: TG.PE.965.F

value: Transgenic


title: melanocyte, KO3

source_name: tail epidermis

value: tail epidermis


title: melanocyte, KB1


title: EOCRC13-P3

type: genomic

city: Scottsdale

first: Michael

middle: Thomas

last: Barrett

state: AZ

address: 13400 East Shea Boulevard

department: Molecular Pharmacology and Experimental Therapeutics

institute: Mayo Clinic Arizona

hyb_protocol: Labeled DNA was hybridized without ozone scavenger and chips washed according to manufacture's protocol for 40 hours in a rotating 65°C oven.

scan_protocol: All microarray slides were scanned using an Agilent 2565C DNA scanner and the images were analyzed with Agilent Feature Extraction version 11.0 using default settings.

data_processing: Data was extracted from the TIFF files using Agilent FE 11.  The aCGH data was assessed with a series of QC metrics then analyzed using an aberration detection algorithm (ADM2)[23]. The latter identifies all aberrant intervals in a given sample with consistently high or low log ratios based on the statistical score derived from the average normalized log ratios of all probes in the genomic interval multiplied by the square root of the number of these probes. This score represents the deviation of the average of the normalized log ratios from its expected value of zero and is proportional to the height h (absolute average log ratio) of the genomic interval, and to the square root of the number of probes in the interval.

source_name: Colon carcinoma

label_protocol: DNAs from frozen tissue and FFPE samples were treated with DNAse 1 prior to Klenow-based labeling. High molecular weight templates were digested for 30 minutes while DNAs from FFPE samples were digested for only 1 minute. In each case 1 µl of 10x DNase 1 reaction buffer and 2 μl of DNase 1 dilution buffer were added to 7 μl of DNA sample and incubated at room temperature then transferred to 70°C for 30 minutes to deactivate DNase 1. Sample and reference templates were then labeled with Cy-5 dUTP and Cy3 dUTP respectively using a BioPrime labeling kit (Invitrogen, Carlsbad, CA) according to our published protocols (22). All labeling reactions were assessed using a Nanodrop assay (Nanodrop, Wilmington, DE) prior to mixing and hybridization to 400k CGH arrays (Agilent Technologies, Santa Clara, CA)

growth_protocol: Fresh tumor samples were flash-frozen and maintained at -80 degrees C.  Formalin fixed paraffin embedded (FFPE) tissues were stored at room temperature

extract_protocol: Excess paraffin was removed from each FFPE sample with a scalpel from either side of scrolls then processed according to our published methods. We used one to three 50 µm scroll(s) from each FFPE tissue block to obtain sufficient numbers of intact nuclei for sorting and molecular assays. Frozen tissue samples were minced in the presence of NST buffer and DAPI according to published protocols. Nuclei from each sample, FFPE or frozen tissue, were disaggregated then filtered through a 40 μm mesh prior to flow sorting with an Influx or Aria III cytometer (Becton-Dickinson, San Jose, CA) with ultraviolet excitation and DAPI emission collected at >450 nm.

tag: disease state

value: Early Onset Colorectal Cancer

value: colorectal carcinoma

tag: tissue source

value: Resection

tag: tissue type

value: formalin fixed paraffin embedded (FFPE) tissue

tag: flow sort content

value: Human tumor

source_name: Reference pooled 46XX

tag: reference

value: Normal female genome


title: EOCRC1MCA-P3

value: frozen tissue


title: ARM5_C

tag: strain


title: Untr_A


title: melanocyte, KO2

value: melanocyte

value: Spry1flox/flox k14-Cre-ERT2


title: melanocyte, KO1


title: ARM5_B

value: in vivo (xenograft)


title: melanocyte, Ctrl3


title: EOCRC17-P3


title: EOCRC12-P4


title: melanocyte, Ctrl2


title: TG.PE.728.F

value: Uterus

value: Proestrus


title: ARM3_D

value: ARN-509 at 30mg/kg- delivered daily by OG in a formulation solution consisting of 15% Vitamin E-TPGS and 65% of a 0.5% w/v CMC solution in 20 mM citrate buffer (pH 4.0)

value: nude-nude

value: tumour explant


title: EOCRC3-P4


title: melanocyte, Ctrl1


title: EOCRC7-P2


title: ARM4_A

value: Mycophenolate mofetil (100mg/kg) and ARN-509 (30mg/kg)


title: ARM3_B


title: ARM4_D


title: EOCRC1MCA-P4


title: ARM2_D

value: Abiraterone at 0.5mmol/kg/d in vehicle - delivered by IP in 5% benzyl alcohol, 95% safflower oil together with


title: ARM5_A

value: Mycophenolate mofetil (100mg/kg), Abiraterone (0.5 mmol/kg/d in vehicle) and prednisolone phosphate (20mg/kg).


title: ARM1_D

value: Mycophenolate mofetil (100 mg/kg) – delivered daily by OG in a 0.9% saline solution


title: ARM2_C


title: ARM3_C


title: ARM2_B