title: Beef steer 7, subcutaneous adipose tissue (SCAT)

city: Lubbock

first: Clarissa

last: Strieder-Barboza

state: Texas

address: ESB II, 1006 Canton Ave.

department: Veterinary Sciences

institute: Texas Tech University

data_processing:
Quality control was performed by removing 1) reads containing adapters; 2) reads when uncertain nucleotides constitute more than 10% of either read (N > 10%); and 3) low-quality nucleotides (Base Quality less than 5) constitute more than 50% of the read. Clean reads were mapped to Bos taurus genome ARS-UCD1.3 using HISAT2 and annotated to NCBI RefSeq assembly accession: GCF_002263795.2. Gene expression analysis was performed using the DESeq2 package in R (v4.2.3) with differentially expressed genes determined as genes with false discovery rate (FDR) <0.05 and log2 Fold Change > |0.5|.
Assembly: ARS-UCD1.3
Supplementary files format and content: .csv file includes raw counts for each sample

organism: Bos taurus

source_name: Adpose tissue

extract_protocol:
RNA was extracted from adipose tissue samples using a Qiagen Lipid Minikit (Qiagen, Germantown Road, Germantown, MD; Cat# 74804). In short, 50-75mg of frozen adipose tissue was placed into a polystyrene tube containing two 3mm zirconium beads and 1 mL of Trizol reagent. The tissue was thenhomogenized at 400 m/s in 30s intervals using a bead blaster (Benchmark Scientific Inc, Sayreville, NJ, USA). After homogenization, sample was centrifuged at 12,000 × g for 10 min at 4°C, and a pink middle layer fluid was collected into a new 2-mL RNase-free microtube. Then, 200 µL of Phenol:chloroform (Invitrogen, Carlsbad, CA) at 4°C were added to the sample in order to isolate the RNA from the organic phase. After centrifugation at 12,000 × g for 15 min at 4°C, the clear upper phase was transferred into a new 2-mL RNase-free microtube containing 700 µL of 70% ethanol. The total RNA was purified using RNeasy Plus Mini Kit and eluted in 30 µL of RNase-free water.
RNA integirty was assessed using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system (Agilent Technologies, Santa Clara, CA, USA). First strand cDNA was synthesized using random hexamer primers and M-MuLV Reverse Transcriptase (Rnase H-) using the RNA samples. Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. PCR was performed with Phusion High-Fidelity DNA polymerase, Uni-versal PCR primers, and Index (X) Primer. PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system. The clustering of the index-coded samples was performed on a cBot Cluster Generation System using TruSeq PE Cluster Kit v3-cBot-HS (Illumina, Inc., San Diego, CA, USA) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina Novaseq 6000 S4 platform to generate 30 million stranded paired-end reads (2 x 150 nt) per sample.

value: Adpose tissue


title: oxWGBS-Seq of Human Cs+ HCC T1

status: Public on Dec 20 2024

type: SRA

city: Nanning

first: Junxian

last: Chen

address: 71, Hedi Road, Qingxiu District

department: Department of Clinical Laboratory

country: China

institute: Guangxi Medical University Cancer Hospital

description: Library name: oxWGBS_Cs+_T_1

library_source: genomic

data_processing:
We used the Trim Galore  to remove sequencing adapters and low-quality data from the sequencing reads.
Assembly: hg38
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample

molecule: genomic DNA

organism: Homo sapiens

source_name: HCC tumor tissue

extract_protocol:
After quality control, lambda DNA was added as a bisulfite conversion efficiency control. Genomic DNA was fragmented to 200-400 bp using a Covaris M220 Focused-ultrasonicator (Covaris, USA). The fragmented DNA was then end-repaired, 3' adenylated, and ligated with methylation-modified sequencing adapters.
5hmC was oxidized to 5fC, followed by bisulfite treatment using the EZ DNA Methylation Gold Kit (Zymo Research). Unmethylated C and oxidized 5fC were converted to U (T after PCR amplification), while methylated C remained unchanged. PCR amplification was performed to generate the final oxWGBS library.

tag: tissue

value: HCC tumor tissue


title: WT, 5DAC, Replicate 4

city: Rosario

first: Julia

middle: Lidia

last: Baulies

state: Santa Fe

address: Ocampo y Esmeralda s/n

country: Argentina

institute: Instituto de Biología Molecular y Celular de Rosario

description:
JB14
10x Genomics

library_source: transcriptomic single cell

data_processing:
Yielded reads were pseudo-aligned to the Arabidopsis genome (TAIR10) using the kallisto bustools software.
Seurat matrices were constructed with the Seurat R package.
Assembly: TAIR10
Supplementary files format and content: Seurat object

molecule: total RNA

organism: Arabidopsis thaliana

source_name: Root tips

growth_protocol: Sterilized seeds were sown in Petri dishes containing solid medium (Murashige and Skoog salts, 1% sucrose, 0.5g/L MES, 0.8% w/v agar, pH = 5,7) and stratified at 4°C in the dark for at least one day. The plates were placed vertically under continuous light at 22°C.

extract_protocol:
Approximately 1000 roots per sample (wild type and p35S:MIR396, uncut controls and 5 days after cut) were collected per quadruplicate for protoplasting. The seeds were sown on nylon mesh, roots were cut at approximately 1cm from the tip, and placed in digesting solution (20mM MES, 400mM mannitol, 20mM KCl, 1.5% cellulase, 0.4% macerozyme, 10mM CaCl2) covered from the light. To enrich the samples with meristematic tissue, the roots were incubated for 20 minutes until the transition zone (between the proliferation and elongation zones) was digested and the tips were released, which was confirmed under microscope. Then, the excess tissue was filtered out using a 1 mm-pore mesh. Digestion continued for 90 minutes and, after filtering the solution twice with a 40µm pore cell strainer, cells were centrifuged down for two minutes at 300rcf and 4°C, washed with washing buffer (enzyme-free digestion solution) and centrifuged again with the same conditions. Buffer was discarded, leaving a small amount at the bottom to resuspend protoplasts, which were then counted in a Neubauer chamber under a light field microscope, and diluted to a concentration of 1000cells/µl.
12000 cells per replicate were sorted on a 10x Chromium platform and libraries were prepared using a 3’ Gene Expression v3.1 kit, following the provider’s instructions.

treatment_protocol: Control samples were collected ad 3 days old without treatment. For 5DAC samples, 3-day-old seedlings were cut with surgical microblades under a stereoscopic microscope, on agar plates with a nylon mesh of 100µm pore and 70µm fiber. Pores and fibers were used as reference for cut positions. Te samples were collected 5 days after cutting (8 days old).

tag: ecotype

value: Root tips

tag: age

value: 8 days old (cut at 3 days old)

tag: genotype

value: Empty vector

tag: treatment

value: 5 days after cut


title: oxWGBS-Seq of Human Cs+ HCC P4

description: Library name: oxWGBS_Cs+_P_4

source_name: HCC adjacent tissue

value: HCC adjacent tissue


title: Beef steer 7, omental adipose tissue (OM)

country: USA

library_source: transcriptomic


title: p35S:MIR396, 5DAC, Replicate 3

description:
JB12
10x Genomics


title: Beef steer 7, intramuscular adipose tissue (IMAT)


title: HCT116 cells, 0uM CHIR99021, biol rep 1

city: Sheffield

first: Jack

last: Riley

address: Firth Court, Western Bank

department: School of Biosciences

country: United Kingdom

institute: The University of Sheffield

data_processing:
RNAseq reads were aligned to hg38 using STAR aligner.
Transcript counts were quantified against our custom TCGA transcriptome assembly using Salmon.
Assembly: hg38
Supplementary files format and content: Tar archived file (.tar.gz) containing salmon quantification file (.sf) alongside corresponding metadata.

molecule: polyA RNA

growth_protocol: HCT116 cells were maintained in DMEM supplemented with 10% fetal bovine serum at 37 degrees celsius and 5% CO2.

extract_protocol:
RNA was extracted using Total RNA Purification Plus Kit (Norgen Biotek) as per manufacturers instructions.
mRNA was enriched by PolyA capture. Libraries were produced using standard Illumina protocols

treatment_protocol: Media was changed to RPMI-1640 media supplemented with either 0uM or 20uM CHIR99021 for 24 hours.

tag: cell line

tag: cell type

value: colorectal carcinoma cells

value: 0uM CHIR99021 (DMSO)


title: WGBS-Seq of Human Cs+ HCC T4

description: Library name: WGBS_Cs+_T_4


title: CD14- PR-cre/YFP+, ML, Rep2

city: Parkville

first: Yunshun

state: VIC

address: 1G Royal Parade

department: ACRF Cancer Biology and Stem Cells

country: Australia

institute: Walter and Eliza Hall Institute of Medical Research

data_processing:
Libraries were sequenced on an Illumina Nextseq 500
Reads were aligned to the mouse genome mm10 by the align function of the Rsubread software tool version 1.29.6.
Read counts were summarized by Entrez Gene ID using the Rsubread featureCounts function and Rsubread's inbuilt RefSeq annotation.
Read counts from the two separate runs of the same sample were aggragated together to form the final gene count for that sample.
Assembly: mm10
Supplementary files format and content: tab-delimited text files; columns represent Entrez Gene ID, gene length (total base count of all exons) and the number of reads mapped to that gene.

organism: Mus musculus

source_name: Mammary gland

extract_protocol:
Total RNA was extracted using Qiagen's miRNeasy kit and DNase treated.
A minimum of 50 ng of total RNA was used to construct libraries following Ilumina's recommended protocol for TruSeq RNA Library Prep Kit v2.

value: Mammary gland

tag: cell population

value: luminal

value: CD14- PR-cre/YFP+, ML


title: p35S:MIR396, Control, Replicate 3

description:
JB11
10x Genomics

value: 3 days old

value: Control


title: CO_17_H1cJunKO2_Day24_rp2_R1.fq.gz

city: Macau

first: Jiantao

last: Shi

address: niversity of Macau Avenida da Universidade Taipa, Macau, China

department: ICMS

institute: University of Macau

data_processing:
bcl2fastq conversion software v2.17 used for basecalling.
RNAseq reads were aligned to the mm10 genome assembly using rsem with options --bowtie2 --bowtie2-sensitivity-level very_sensitive
expected_count were extract from the sample_name.genes.results and merged together to get a big table
RNAseq data was normalized by EDAseq
Assembly: mm 10
Supplementary files format and content: tab-delimited text files include count values for each Sample

source_name: Embryonic Stem Cells

growth_protocol: We mainly referred to the technical guidelines for Cerebral Organoids culture published by Lancaster group (Madeline A. Lancaster, 2014; Stefano L. Giandomenico, 2021).

extract_protocol:
Total RNA was prepared with TRIzol (Invitrogen,15596026).1 μg total RNA was used for cDNA synthesis using HiScript®II Q RT SuperMix for qPCR (+gDNA wiper) (Vazyme,R223-01) according to the manufacturer’s instructions.
VAHTS® Total RNA-seq (H/M/R) Library Prep Kit for Illumina (Vazyme,NR603), VAHTS®RNA Adapters set3 for Illumina(Vazyme,N809) were used to make the libraries for sequencing according to the manufacturer’s instructions.

treatment_protocol: 3~4 pooled Cerebral Organoids samples were incubated with TRIzol and lysed through pipetting on ice and next extracted according to the manufacturer's instructions. Purified RNA was analyzed to determine the RNA concentration and integrity. rganoid

value: Embryonic Stem Cells

value: c-Jun Knockout

value: neural differentiation and cerebral organoids induction


title: HCT116 cells, 0uM CHIR99021, biol rep 4


title: Beef steer 6, subcutaneous adipose tissue (SCAT)

value: Adpose tissue


title: WT, 5DAC, Replicate 3

description:
JB10
10x Genomics


title: Beef steer 6, omental adipose tissue (OM)


title: WGBS-Seq of Human Cs+ HCC T3

description: Library name: WGBS_Cs+_T_3

value: HCC tumor tissue


title: WT, Control, Replicate 3

description:
JB9
10x Genomics

value: Root tips


title: WT, Control, Replicate 4

description:
JB13
10x Genomics


title: Beef steer 6, intramuscular adipose tissue (IMAT)


title: p35S:MIR396, 5DAC, Replicate 2

description:
JB8
10x Genomics

value: 8 days old (cut at 3 days old)


title: CO_18_H1cJunKO10_Day24_rp2_R1.fq.gz


title: Beef heifer 5, subcutaneous adipose tissue (SCAT)


title: CO_14_H1cJunKO2_Day12_rp2_R1.fq.gz


title: WGBS-Seq of Human Cs+ HCC T2

description: Library name: WGBS_Cs+_T_2


title: WT, 5DAC, Replicate 2

description:
JB6
10x Genomics

value: Empty vector


title: Beef heifer 5, intramuscular adipose tissue (IMAT)


title: CO_06_H1cJunKO10_Day24_rp1_R1.fq.gz


title: WGBS-Seq of Human Cs+ HCC P2

description: Library name: WGBS_Cs+_P_2


title: WT, Control, Replicate 2

description:
JB5
10x Genomics


title: WGBS-Seq of Human Cs+ HCC T1

description: Library name: WGBS_Cs+_T_1


title: WGBS-Seq of Human Cs+ HCC P4

description: Library name: WGBS_Cs+_P_4


title: CO_13_H1WT_Day12_rp2_R1.fq.gz

value: WT


title: Beef steer 4, subcutaneous adipose tissue (SCAT)


title: CO_16_H1WT_Day24_rp2_R1.fq.gz


title: WGBS-Seq of Human Cs+ HCC P1

description: Library name: WGBS_Cs+_P_1

value: HCC adjacent tissue


title: p35S:MIR396, 5DAC, Replicate 1

description:
JB4
10x Genomics

value: 5 days after cut


title: CO_04_H1WT_Day24_rp1_R1.fq.gz


title: WGBS-Seq of Human Cs+ HCC P3

description: Library name: WGBS_Cs+_P_3


title: CO_15_H1cJunKO10_Day12_rp2_R1.fq.gz


title: HCT116 cells, 0uM CHIR99021, biol rep 2

value: colorectal carcinoma cells

value: 0uM CHIR99021 (DMSO)


title: p35S:MIR396, Control, Replicate 2

description:
JB7
10x Genomics


title: Beef steer 4, omental adipose tissue (OM)


title: ent-1KO mutant feed with ent-2 RNAi HT115 bacteria

city: Ashburn

first: Shihong

middle: Max

last: Gao

address: 19700 Helix Dr

institute: Janelia Research Campus

description: 10x Genomics

data_processing:
We used Cell ranger (6.0, 10x Genomics) to align Raw base call (BCL) sequence files or FASTQ files to the C. elegans genome (WS282, Wormbase), and generated feature-barcode matrices. Doublets were removed based on the recommended ratio provided by the 10X Genomics Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 user guide (CG000204 Rev D), with a score calculated by Doubletfinder (Github/chris-mcginnis-ucsf). The feature-barcode matrices for each sample were constructed into a Seurat (R package, 4.0.5) object for downstream analysis. Cells were filtered by requiring a minimum of 100 genes to be expressed, and genes were filtered by being expressed in at least 3 cells. Integration of samples was performed with Seurat canonical correlation analysis (CCA) method to remove the batch effects. We tested CCA, reciprocal principal component analysis (PCA) and Harmony (0.1) for integration and chose the method with the best performance. The top 2,000 variable features were used for integration anchor identification. UMAP and tSNE dimension reduction were performed with the first 50 dimensions from PCA. Clustering was performed at multiple Leiden resolutions.
Assembly: WS282
Supplementary files format and content: Tab-separated values files and matrix files

molecule: nuclear RNA

organism: Caenorhabditis elegans

source_name: whole body

extract_protocol:
Worms were washed three times with PBS and collected in a 1.5 mL tube. We then added 100 μl of homogenization buffer  and grind the worms with a pestle motor for 30 seconds on ice. To minimize nuclei adhesion on the surface, all the pestles, tubes and filters were pre-coated with a homogenization buffer or 1x PBS. 900 μl of homogenization buffer was added to wash the pestle, and the total 1mL homogenized sample was transferred into a 1mL Dounce tissue grinder (Wheaton 357538). The grinder was autoclaved overnight in a 220℃ oven to deactivate ribonuclease. After placing the grinder on ice, 20 strokes were applied using a loose pestle, followed by another 20 strokes using a tight pestle, while avoiding generating foams. The 1000μl samples were filtered through a cell strainer (35 μm) and then through a Flowmi cell strainer (BelArt, H13680-0040) into a new 1.5 mL tube. The tubes were centrifuged for 10 minutes at 1000 g at 4°C, and the supernatant was removed while not disturbing the pellet, which was hardly visible. The pellet was resuspended with 500 μl 1x PBS with 0.5% BSA and RNAase inhibitor and filtered with a Flowmi cell strainer (BelArt, H13680-0040) again into a 5 mL flow cytometer tube. A total of 20 μl was transferred into another flow cytometer tube and diluted with 180 μl 1x PBS with 0.5% BSA and RNAase inhibitor as an unstained control. Hoechst (Invitrogen 33342) was used to stain the nuclei in 1:1000 working concentration. We coated the 1.5 mL sorting collection tube with 1x PBS with 0.5% BSA and RNAase inhibitor and ~300,000 nuclei were sorted with Hoechst 33342 positive gating, which indicates DNA content, and forward scatter area (FSC-A) gating, representing the particle size above threshold. The intestinal nuclei, which are polypoid (32N), usually form an obvious cluster separating from the other 2N somatic nuclei, and these were also included in our collection. The collection tube was centrifuged at 800g for 8 minutes at 4℃ and the sheath buffer supernatant was carefully removed before resuspending the sorted nuclei with 40-50 μl 1x PBS with 0.5% BSA and RNAase inhibitor. The concentration and morphology of the nuclei were checked under a microscope to ensure high-quality nuclei isolation. If the results were desirable, we proceeded to generate gel emulsion with 10X Chromium Controller.For sorting the nuclei, we used Sony MA800 sorter. To obtain the best nuclei morphology and the least RNA degradation, it is recommended to minimize the sorting time. Two researchers could simultaneously set up the sorter and perform nuclei processing.
Library preparation was carried out using the published 10X Chromium Single Cell 3’ v2/v3 Solution protocol (Single/dual index). The resulting libraries were sequenced on the Novaseq 6000 platform, with the recommended cycle numbers: 26 cycles for Read 1, 8 cycles for i7 index, and 98 cycles for Read 2.

value: whole body

tag: strain

value: ent-1KO mutant


title: CO_03_H1cJunKO10_Day12_rp1_R1.fq.gz


title: p35S:MIR396, Control, Replicate 1

description:
JB3
10x Genomics

value: 3 days old


title: Beef heifer 5, omental adipose tissue (OM)


title: WT, 5DAC, Replicate 1

description:
JB2
10x Genomics


title: s0630-1

city: Wuhan

first: qiwen

last: man

state: Hubei

address: Luyu road

institute: wuhan university

data_processing:
Basecalls performed using bcl2fastq V2.19.
equenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence.
Quantify gene expression using read2 by featureCounts software
Genome_build: GRCh38
Supplementary_files_format_and_content: fastq file was rawdata file

source_name: salivary gland pleomorphic adenoma  tissue

extract_protocol:
Single-cell suspensions were then loaded onto microfluidic devices and scRNA-seq libraries were constructed according to Singleron GEXSCOPE® protocol by GEXSCOPE® Single-Cell RNA Library Kit
Single cell suspension was loaded onto a microfluidic chip and single cell RNA-seq libraries were constructed according to the manufacturer’s instructions (Singleron GEXSCOPE Single Cell RNAseq Library Kit, Singleron Biotechnologies).

treatment_protocol: The tissue pieces were digested with 2 ml GEXSCOPE® Tissue Dissociation Solution at 37℃ for 15 min

value: Untreated

tag: gender

value: Female


title: CO_02_H1cJunKO2_Day12_rp1_R1.fq.gz

value: Embryonic Stem Cells

value: c-Jun Knockout


title: ent-2KO mutant feed with L4440 HT115 control bacteria

description: 10x Genomics

value: ent-2KO mutant


title: oxWGBS-Seq of Human Cs+ HCC P1

description: Library name: oxWGBS_Cs+_P_1


title: CO_01_H1WT_Day12_rp1_R1.fq.gz

value: neural differentiation and cerebral organoids induction


title: CO_05_H1cJunKO2_Day24_rp1_R1.fq.gz


title: s0623-01

value: salivary gland pleomorphic adenoma  tissue


title: Wild-type(N2) feed with L4440 HT115 control bacteria

description: 10x Genomics


title: WT, Control, Replicate 1

description:
JB1
10x Genomics

value: Root tips

value: Control


title: Beef steer 4, intramuscular adipose tissue (IMAT)

value: Adpose tissue


title: ent-1KO mutant feed with L4440 HT115 control bacteria

description: 10x Genomics


title: MUSCLE (E17 embryo) ADCK2 wild-type, prenatally treated with CoQ10, biological rep 2

status: Public on Dec 19 2024

type: RNA

city: Sevilla

first: Daniel Jose

last: Moreno Fernandez-Ayala

address: Carretera de Utrera, Km. 1

country: Spain

institute: Universidad Pablo de Olavide

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo, prenatal CoQ10 administration (maternal route)
WT+Q_MUSCLE-2

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on Clariom™ D assays, mouse, ThermoFisher (previously known as GeneChip™ Mouse Transcriptome Array 1.0 - MTA 1.0.) in a Hibridation Oven 645. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: GeneChips were scanned using the Hewlett-Packard GeneArray Scanner 3000 7G.

data_processing: The data were analysed with the Transcriptome Analysis Console 4.0 software using Affymetrix default analysis settings and global scaling as normalization method.

label: biotin

source_name: muscle of E17-embryo (E17 dev. stage)

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 6 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).

growth_protocol: Both Adck2 (+/-) mutant and Adck2(+/+) wild-type E17-embryos from the same litter were dissected. For each ambryo, muscle from the four limbs, liver and brain were carefully dissected.

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

treatment_protocol: Reduced Coenzyme Q10 (CoQ10) (Kaneka QH stabilized powder type P30) (Kaneka Pharma Europe, Brussels, Belgium) (CAS Registry No. 303-98-0) was dissolved in the water bottles to fed reproductive animals. CoQ10 was dissolved in water (0.5 mg/ml) to achieve a final mean ingestion of 33.33-50 mg/kg/day per mouse, in accordance with the doses normally used in CoQ deficient patients (humans).

tag: development

value: E17 embryo (embryo 17 days post coitus)

value: Muscle (E17 embryo)

value: ADCK2(+/+), wild-type


title: MUSCLE (E17 embryo) ADCK2 wild-type, biological rep 1

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo
WT-MUSCLE-1

value: non-treated (control)


title: MUSCLE (E17 embryo) ADCK2 mutant, prenatally treated with CoQ10, biological rep 2

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo, prenatal CoQ10 administration (maternal route)
ADCK2+Q_MUSCLE-2

source_name: muscle of E17-embryo (E17 dev. stage), CoQ10-prenatally treated

value: ADCK2(+/-), mutant


title: MUSCLE (E17 embryo) ADCK2 wild-type, biological rep 2

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo
WT-MUSCLE-2


title: MUSCLE (E17 embryo) ADCK2 wild-type, prenatally treated with CoQ10, biological rep 1

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo, prenatal CoQ10 administration (maternal route)
WT+Q_MUSCLE-1


title: MUSCLE (E17 embryo) ADCK2 mutant, prenatally treated with CoQ10, biological rep 1

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo, prenatal CoQ10 administration (maternal route)
ADCK2+Q_MUSCLE-1


title: MUSCLE (E17 embryo) ADCK2 mutant, prenatally treated with CoQ10, biological rep 3

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo, prenatal CoQ10 administration (maternal route)
ADCK2+Q_MUSCLE-3


title: MUSCLE (E17 embryo) ADCK2 mutant, biological rep 1

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo
ADCK2-MUSCLE-1


title: LIVER (E17 embryo) ADCK2 wild-type, biological rep 1

description:
Gene expression data from LIVER of ADCK2 wild-type E17 embryo
WT-LIVER-1

source_name: liver of E17-embryo (E17 dev. stage)

value: Liver (E17 embryo)

value: non-treated


title: DMSMapseq, VCE

city: New Haven

first: Carson

last: Thoreen

state: Connecticut

address: 333 Cedar Street

department: Cellular and Molecular Physiology

institute: Yale University

description:
DMSMapseq with RNA
DMS VCE Sample
MutationRate.tsv
VCE_plusDMS_MutRates.tsv

library_source: other

data_processing:
*library strategy: dimethyl sulfate-mutational profiling and sequencing (DMS-MaPseq)
adapter trimming using bbduk.shk trim=r k=23 mink=11 hdist=1 tpe tbo
PCR duplicate/UMI collapsing using dedupe.sh
UMI trimming using bbduk.sh ftl=10
alignment using STAR soloStrand = Forward alignSJoverhangMin = 999 alignIntronMax = 1  alignIntronMin = 999
Reads quantification by pileup.sh
Assembly: artifitial genome
Supplementary files format and content: tsv, CPM
Supplementary files format and content: tsv, pairing parobability

molecule: other

organism: synthetic construct

extract_protocol:
RNA is extracted with hot phenol.
RNA is reverse transcribed with common primer binding to the 3′ end. 5′ Adaptosr with random UMI were ligated to the reverse transcript products. The library was then amplified with primers containing Illumina indice sequences.

value: DMS


title: LIVER (E17 embryo) ADCK2 wild-type, biological rep 3

description:
Gene expression data from LIVER of ADCK2 wild-type E17 embryo
WT-LIVER-3


title: W40_S_12_2.

city: yangzhou

first: xvwen

last: shen

state: JiangSu

address: wenhui

institute: Yangzhou University

description: Library name: Sample23

data_processing:
Sequence reads were trimmed for adaptor sequence /low-qualty sequence using fastp. Trimmed sequence reads were mapped to reference genome（http://www.genoscope.cns.fr/brassicanapus/data/）using hisat2 Read count extraction and normalization were performed using RSEM Differential expression analysis was performed using DESeq2
Assembly: v5（http://www.genoscope.cns.fr/brassicanapus/data/）processed data files format and content：csv file  include TPM values for each sample
Supplementary files format and content: tab-delimited text file includes raw counts for each sample csv files include FPKM values for each Sample

organism: Brassica napus

source_name: leaf

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen). 1.3 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: leaf


title: DMSMapseq, VCE, noDMS

description:
DMSMapseq with RNA
DMS VCE noDMS
MutationRate.tsv
VCE_minusDMS_MutRates.tsv

value: no DMS


title: LIVER (E17 embryo) ADCK2 wild-type, biological rep 2

description:
Gene expression data from LIVER of ADCK2 wild-type E17 embryo
WT-LIVER-2


title: DMSMapseq, VCE, denatured

description:
DMSMapseq with RNA
DMS VCE denatured
MutationRate.tsv
VCE_denatured_MutRates.tsv

value: DMS, denature RNA


title: W40_S_6_2.

description: Library name: Sample19


title: LIVER (E17 embryo) ADCK2 mutant, biological rep 1

description:
Gene expression data from LIVER of ADCK2 mutant E17 embryo
ADCK2-LIVER-1


title: W40_S_12_1.

description: Library name: Sample21


title: LIVER (E17 embryo) ADCK2 mutant, biological rep 2

description:
Gene expression data from LIVER of ADCK2 mutant E17 embryo
ADCK2-LIVER-2


title: LIVER (E17 embryo) ADCK2 mutant, biological rep 3

description:
Gene expression data from LIVER of ADCK2 mutant E17 embryo
ADCK2-LIVER-3


title: BRAIN (E17 embryo) ADCK2 mutant, biological rep 1

description:
Gene expression data from BRAIN of ADCK2 mutant E17 embryo
ADCK2-BRAIN-1

source_name: brain of E17-embryo (E17 dev. stage), ADCK2 mutant

value: Brain (E17 embryo)


title: MUSCLE (E17 embryo) ADCK2 mutant, biological rep 2

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo
ADCK2-MUSCLE-2


title: BRAIN (E17 embryo) ADCK2 wild-type, biological rep 3

description:
Gene expression data from BRAIN of ADCK2 wild-type E17 embryo
WT-BRAIN-3

source_name: brain of E17-embryo (E17 dev. stage), wild-type


title: W40_S_6_1.

description: Library name: Sample17


title: BRAIN (E17 embryo) ADCK2 wild-type, biological rep 1

description:
Gene expression data from BRAIN of ADCK2 wild-type E17 embryo
WT-BRAIN-1


title: W40_S_3_2.

description: Library name: Sample15


title: BRAIN (E17 embryo) ADCK2 mutant, biological rep 2

description:
Gene expression data from BRAIN of ADCK2 mutant E17 embryo
ADCK2-BRAIN-2


title: BRAIN (E17 embryo) ADCK2 wild-type, biological rep 2

description:
Gene expression data from BRAIN of ADCK2 wild-type E17 embryo
WT-BRAIN-2


title: W40_12_2.

description: Library name: Sample11

value: WT


title: MUSCLE (E17 embryo) ADCK2 wild-type, biological rep 3

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo
WT-MUSCLE-3


title: W40_6_1.

description: Library name: Sample5


title: W40_6_2.

description: Library name: Sample7


title: BRAIN (E17 embryo) ADCK2 mutant, biological rep 3

description:
Gene expression data from BRAIN of ADCK2 mutant E17 embryo
ADCK2-BRAIN-3


title: W40_3_2.

description: Library name: Sample3


title: W40_3_1.

description: Library name: Sample1


title: W40_S_3_1.

description: Library name: Sample13


title: RNA-seq of fat body DD condition w1118 CT24_biorep2

city: Geneva

first: Blanca

middle: Lago

last: Solis

address: 30 Quai Ernest-Ansermet

department: Genetics and Evolution

country: Switzerland

institute: University of Geneva

description: Library name: W_CT24_2

data_processing:
For RNA-seq data analysis, raw reads were aligned with the Dm6 Drosophila genome using STAR software (v2.7.9a). The raw counts of reads associated with each of the exons were determined using featureCounts function of the Rsubread package (v1.22.2). All libraries generated on average ~18 million single-end 100 bp reads, from which approximately 80% (~14 million reads) were mapped to the genome. Genes presenting no expression in at least one of the conditions in one of the replicates, as well as genes whose expression was below a count threshold were excluded from further analysis. This threshold was set up based on the distribution of the gene counts, established with edgeR function. After that, the DESeq2 package (v.1.30.1) was used to normalize the gene expression levels. The normalized counts were used for downstream analysis.
Assembly: Dm6 Drosophila genome
Supplementary files format and content: A Comma-delimited files with the raw data for the nine timepoints samples of Per0 (CT.counts_P) and  another file for w1118 (CT.counts_W) larvae.  A Comma-delimited files with the normalized data for per0 samples (per0_fatbody_normalizedcounts_2) and another one for w1118 samples (W_fatbody_normalizedcounts_2)

organism: Drosophila melanogaster

source_name: Fat body

growth_protocol: Drosophila melanogaster strains were maintained on standard cornmeal-agar food at 18 °C or 25 °C in a 12 h light-dark cycle and controlled humidity. To obtain third-instar larvae, flies of each genotype were transferred to tubes with food for 3 days and entrained in 12h light-dark cycle. Then, they were flipped twice every day in 12h:12h LD cycles or in DD. After 3-4 days, the food of the flipped tube was scraped and rinsed with distilled water to expose larvae. A mixed population of male and female larvae was used in all experiments. The developmental stage was confirmed by the characteristic shape of the larval mouth hooks.

extract_protocol:
Full fat body of 5 larvae were dissected and collected into 350 µL Buffer RLT Plus (QIAGEN RNeasy Plus Micro Kit, Cat. # 74034). Then, the tissue was dissociated using an electric Pellet Pestle Cordless Motor, 5. Total RNA extraction was performed according to the manufacturer’s protocol (QIAGEN RNeasy Plus Micro Kit, Cat. # 74034) and eluted in 14 µL of nuclease-free water.
cDNA library preparations were conducted using the SMART-Seq v4 Ultra Low Input kit from TaKaRa (Cat. # 634894) according to the manufacturer’s instructions.

value: Fat body


title: RNA-seq of fat body DD condition w1118 CT28_biorep1

description: Library name: W_CT28_1


title: RNA-seq of fat body DD condition w1118 CT28_biorep2

description: Library name: W_CT28_2


title: RNA-seq of fat body DD condition w1118 CT32_biorep1

description: Library name: W_CT32_1


title: RNA-seq of fat body DD condition w1118 CT32_biorep2

description: Library name: W_CT32_2


title: RNA-seq of fat body DD condition w1118 CT20_biorep2

description: Library name: W_CT20_2


title: MUSCLE (E17 embryo) ADCK2 wild-type, prenatally treated with CoQ10, biological rep 3

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo, prenatal CoQ10 administration (maternal route)
WT+Q_MUSCLE-3


title: RNA-seq of fat body DD condition w1118 CT16_biorep2

description: Library name: W_CT16_2


title: RNA-seq of fat body DD condition w1118 CT24_biorep1

description: Library name: W_CT24_1


title: RNA-seq of fat body DD condition w1118 CT20_biorep1

description: Library name: W_CT20_1


title: RNA-seq of fat body DD condition w1118 CT12_biorep2

description: Library name: W_CT12_2


title: RNA-seq of fat body DD condition w1118 CT16_biorep1

description: Library name: W_CT16_1


title: RNA-seq of fat body DD condition w1118 CT04_biorep2

description: Library name: W_CT04_2


title: RNA-seq of fat body DD condition w1118 CT08_biorep2

description: Library name: W_CT08_2


title: W40_12_1.

description: Library name: Sample9


title: RNA-seq of fat body DD condition w1118 CT08_biorep1

description: Library name: W_CT08_1


title: CD14+ PR-cre/YFP+, HR+ LP, Rep1

value: Mammary gland

value: luminal

value: CD14+ PR-cre/YFP+, HR+ LP


title: RNA-seq of fat body DD condition w1118 CT12_biorep1

description: Library name: W_CT12_1


title: RNA-seq of fat body DD condition w1118 CT04_biorep1

description: Library name: W_CT04_1


title: RNA-seq of fat body DD condition w1118 CT00_biorep2

description: Library name: W_CT00_2


title: RNA-seq of fat body DD condition per0 CT32_biorep1

description: Library name: P_CT32_1


title: RNA-seq of fat body DD condition w1118 CT00_biorep1

description: Library name: W_CT00_1


title: RNA-seq of fat body DD condition per0 CT32_biorep2

description: Library name: P_CT32_2


title: RNA-seq of fat body DD condition per0 CT28_biorep1

description: Library name: P_CT28_1


title: RNA-seq of fat body DD condition per0 CT28_biorep2

description: Library name: P_CT28_2


title: RNA-seq of fat body DD condition per0 CT24_biorep2

description: Library name: P_CT24_2


title: RNA-seq of fat body DD condition per0 CT20_biorep2

description: Library name: P_CT20_2


title: RNA-seq of fat body DD condition per0 CT24_biorep1

description: Library name: P_CT24_1


title: RNA-seq of fat body DD condition per0 CT20_biorep1

description: Library name: P_CT20_1


title: RNA-seq of fat body DD condition per0 CT16_biorep1

description: Library name: P_CT16_1


title: RNA-seq of fat body DD condition per0 CT12_biorep1

description: Library name: P_CT12_1


title: RNA-seq of fat body DD condition per0 CT04_biorep2

description: Library name: P_CT04_2


title: RNA-seq of fat body DD condition per0 CT16_biorep2

description: Library name: P_CT16_2


title: RNA-seq of fat body DD condition per0 CT12_biorep2

description: Library name: P_CT12_2


title: RNA-seq of fat body DD condition per0 CT04_biorep1

description: Library name: P_CT04_1


title: RNA-seq of fat body DD condition per0 CT00_biorep2

description: Library name: P_CT00_2


title: RNA-seq of fat body DD condition per0 CT00_biorep1

description: Library name: P_CT00_1


title: Oxalipaltin-resistant PANC1, sample 2

city: Boston

first: Jonathan

middle: P

last: Celli

state: MA

address: 100 Morrissey Blvd

institute: University of Massachusetts Boston

data_processing:
The quality of the raw fastq files were assessed using FastQC (v.0.11.5)24. Adaptor sequences, “AGATCGGAAGAGCACACGTCTGAACTCCAGTCA”, and “AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT" were trimmed from the 3’ end of the reads using Cutadapt commandline tool form the Trime Galore package (v.0.4.2). The trimmed reads were mapped against the human reference genome (Ensemble, GRCh38) using STAR/2.5 using default parameters25. The average alignment rate was 96%. The sorted BAM files generated by STAR were used to estimate the transcript abundance per sample using featureCount from the Subread package (v.1.6.2)26. Gene expression analysis was performed using the edgR Bioconductor R package (v.3.24.3)27. The edgeR TMM method (trimmed mean of M values) was applied to the filtered genes utilizing the DGElist(), calcNormFactors(), estimateGLMCommonDisp(), estimateGLMTrendedDisp(), estimateGLMTagwiseDisp() functions. The glmFit and glmLRT functions from edgeR were used to fit a negative binomial generalized log-linear model to the read counts.
Assembly: The trimmed reads were mapped against the human reference genome (Ensemble, GRCh38) using STAR/2.5 using default parameters
Supplementary files format and content: The read count file is in CSV format and contains raw read counts for each mapped gene for each sample

source_name: Oxalipaltin-resistant

growth_protocol: Cultures were grown in DMEM with 10% FBS, 5% CO2 at 37C as per ATCC growth guidelines

extract_protocol:
Trizol
Library preparation was performed using Kapa mRNA Hyper Prep kit with Bioo single index barcodes

treatment_protocol: Drug-resistant cultures were grown in media containing increasing concentration of oxaliplatin (Tocris) over successive passages


title: Oxalipaltin-resistant PANC1, sample 3


title: MUSCLE (E17 embryo) ADCK2 mutant, biological rep 3

description:
Gene expression data from MUSCLE of ADCK2 mutant E17 embryo
ADCK2-MUSCLE-3

value: E17 embryo (embryo 17 days post coitus)


title: RNA-seq of fat body DD condition per0 CT08_biorep1

description: Library name: P_CT08_1


title: Oxalipaltin-resistant PANC1, sample 1


title: PANC1 cells, untreated, sample 2

source_name: untreated


title: PANC1 cells, untreated, sample 3


title: OV90, SY-5609 400nm, 4hr 2

city: Boulder

first: Taylor

middle: K

last: Jones

state: CO

address: 3415 Colorado Ave

institute: University of Colorado Boulder

data_processing:
Mapping of RNA-seq data was performed using the RNAFlow Pipeline (https://github.com/Dowell-Lab/RNAseq-Flow)
Assembly: hg38
Supplementary files format and content: tdfs for visualization of the data

growth_protocol: OV-90 (CRL-11732) cells were purchased from American Tissue Culture Collection (ATCC) and cultured according to manufacturer's instructions at 37°C in 5% CO 2 in a humidified incubator.500,000 cells per well were seeded in 6-well tissue culture treated plates the day before treatment.

extract_protocol:
Total RNA was harvested from cell lysates using reagents from RNeasy Mini Kit (Qiagen 74104) and the Qiacube (Qiagen).Total RNA was eluted in water and quantitated using a Nanodrop Spectrophotometer(Thermo Fisher).
The sample with the highest concentration of RNA was 1000 ng, so we used this sample as the standard for submitting equal volumes of all other samples to control for cell number. ERCC spike-in was also added to the samples to control for cell number.

treatment_protocol: Cells were treated with 0.025% DMSO or SY-5609 at 40 and 400nM for 4hrs before total RNA was harvested

value: Ovarian

tag: time


title: OV90, SY-5609 40nm, 4hr 2


title: OV90, DMSO 4hr 3

value: DMSO


title: OV90, DMSO 4hr 2


title: OV90, SY-5609 40nm, 4hr 3


title: OV90, DMSO 4hr 1


title: OV90, SY-5609 400nm, 4hr 1


title: RNA-seq of fat body DD condition per0 CT08_biorep2

description: Library name: P_CT08_2


title: T3M4_plusY318A_S19

city: Lebanon

first: Christine

last: Alewine

state: NH

address: One Medical Center Drive, Williamson 651,

institute: Dartmouth Cancer Center

data_processing:
Sequencing quality was first examined by FastQC (version 0.11.8), Preseq(version 3.1.2), Picard tools (version 2.23.7), and RSeQC (version 4.0.4).
The Illumina reads in FASTQ format were then trimmed by Cutadapt (version 4.0) to remove the adaptor sequences and low quality reads.
STAR (version 2.7.0f) two-pass mode was used to map reads to human reference genome hg38 and RSEM (version 1.3.3) was used to quantify the gene and transcript expression levels.
Assembly: hg38
Supplementary files format and content: T3M4_4CM.RawCountFile_RSEM_genes.txt raw count from RSEM
Supplementary files format and content: KLM1_4CM.RawCountFile_RSEM_genes.txt raw count from RSEM

source_name: Pancreatic ductal adenocarcinoma

growth_protocol: Human pancreatic cancer cell lines KLM1 and T3M4 were the gifts of Udo Rudloff and Mitchell Ho (NCI, Bethesda, MD), respectively. Identity of the cell lines was confirmed by STR testing. Cells were cultured in RPMI 1640 medium (Gibco, Thermo Scientific) supplemented with L-Glutamine (2 mmol/L), penicillin (100 U), streptomycin (100 ug), and 10% FBS (Hyclone, Thermo Scientific). All cells were maintained in a 5% CO2 adjusted incubator at 37°C and tested free of mycoplasma.

extract_protocol:
CM-treated cells were lysed and RNA extracted using RNeasy Kit (Qiagen).
Total RNA from quintuple samples was sequenced by the Center for Cancer Research Sequencing Facility (Bethesda, MD) using an Illumina HiSeq 2500 instrument.

treatment_protocol: KLM1 parent and derivative cells were plated at equal density, then old medium was replaced with equivolume of fresh RPMI after 48 hours. CM was collected 24 hours later and stored in aliquots at -80°C for later use. KLM1 or T3M4 KO cells were grown for 48 hours after plating, before old growth medium was replaced with designated CM and treated for 4 hours.

value: Pancreatic ductal adenocarcinoma

value: MSLN knockout and then over-expressing Y318A mutant MSLN


title: T3M4_plusY318A_S4


title: T3M4_plusWT_S7

value: MSLN knockout and then over-expressing full length MSLN


title: T3M4_plusY318A_S8


title: T3M4_Mock_S5

value: Wild Type


title: T3M4_plusWT_S18


title: T3M4_plusWT_S3


title: T3M4_plusY318A_S15


title: T3M4_KO_S9

value: MSLN knockout


title: T3M4_plusWT_S10


title: OV90, SY-5609 400nm, 4hr 3


title: T3M4_KO_S6


title: T3M4_Mock_S16


title: T3M4_plusY318A_S11


title: T3M4_Mock_S1


title: PANC1 cells, untreated, sample 1


title: T3M4_KO_S2


title: OV90, SY-5609 40nm, 4hr 1


title: T3M4_Mock_S12


title: T3M4_KO_S17


title: T3M4_KO_S13


title: KLM1_Y318A5


title: KLM1_Y318A3


title: KLM1_WT5


title: T3M4_plusWT_S14


title: KLM1_WT4


title: KLM1_WT3


title: KLM1_WT1


title: KLM1_Mock5


title: KLM1_KO5


title: KLM1_Mock2


title: KLM1_Mock1


title: KLM1_KO3


title: KLM1_KO4


title: KLM1_KO1


title: KLM1_KO2


title: VCE, ShortPool, total RNA input

description:
VCE ShortPool IVT
CPM.tsv
CPMandRRS_VCE_Short.tsv

data_processing:
*library strategy: DART-seq
adapter trimming using bbduk.shk trim=r k=23 mink=11 hdist=1 tpe tbo
PCR duplicate/UMI collapsing using dedupe.sh
UMI trimming using bbduk.sh ftl=10
alignment using STAR soloStrand = Forward alignSJoverhangMin = 999 alignIntronMax = 1  alignIntronMin = 999
Reads quantification by pileup.sh
Assembly: artifitial genome
Supplementary files format and content: tsv, CPM
Supplementary files format and content: tsv, pairing parobability

source_name: HeLa

value: HeLa


title: VCE, LongPool, total RNA input

description:
VCE LongPool IVT
CPM.tsv
CPMandRRS_VCE_Long.tsv


title: VCE, ShortPool, monosome fraction

description:
VCE ShortPool monosome
CPM.tsv
CPMandRRS_VCE_Short.tsv


title: VCE, LongPool, monosome fraction

description:
VCE LongPool monosome
CPM.tsv
CPMandRRS_VCE_Long.tsv


title: VCE, VarientPool, monosome fraction

description:
VCE VarientPool monosome
CPM.tsv
CPMandRRS_VCE_Var.tsv


title: VCE, VarientPool, total RNA input

description:
VCE VarientPool IVT
CPM.tsv
CPMandRRS_VCE_Var.tsv


title: KLM1_Y318A4


title: Engineered heart issue_E54K_3

first: Stuart

middle: G

last: Campbell

state: CT

address: 55 Prospect Street, Malone 301

department: Biomedical Engineering

data_processing:
Data analysis was performed on PartekFlow. Alignment was performed using STAR 2.7.8a and hg38 as the reference genome.
Assembly: hg19
Supplementary files format and content: .csv file. Rows are features/genes and columns L through T are samples

source_name: engineered heart tissue

extract_protocol:
For RNA extraction, tissues were flash frozen and crushed with plastic pestles. The aqueous phase was collected after TRIzol (Ambion) phase separation, and the RNA pellets were precipitated for total RNA extraction. Samples were treated with DNAse and cleaned with Qiagen RNeasy columns. Samples were sequenced by Yale Center for Genomic Analysis
mRNA is purified from approximately 200ng of total RNA with oligo-dT beads and sheared by incubation at 94C in the presence of Mg (Roche Kapa mRNA Hyper Prep Cat# KR1352). Following first-strand synthesis with random primers, second strand synthesis and A-tailing are performed with dUTP for generating strand-specific sequencing libraries. Adapter ligation with 3’ dTMP overhangs are ligated to library insert fragments. Library amplification amplifies fragments carrying the appropriate adapter sequences at both ends. Strands marked with dUTP are not amplified. Indexed libraries are quantified by qRT- PCR using a commercially available kit (Roch KAPA Biosystems Cat# KK4854) and insert size distribution determined by either the Agilent Bioanalyzer. Samples with a yield of ≥ 0.5 ng/ul and a size distribution of 150-300bp are used for sequencing.

value: engineered heart tissue

value: iPSC derived cardiomyocyte


title: Engineered heart issue_E54K_2


title: Engineered heart issue_E62Q_3


title: Engineered heart issue_E62Q_2


title: KLM1_Y318A1


title: Engineered heart issue_E54K_1


title: Engineered heart issue_E62Q_1


title: KLM1_WT2


title: KLM1_Mock4


title: Engineered heart issue_WT_3


title: Engineered heart issue_WT_1


title: isolated hepatocytes, ALAS1/2-KO, Rep4

city: New York

first: Seungjae

last: Lee

state: NY

address: 430 East 67th St.

department: Department of Developmental Biology

institute: Sloan-Kettering Institute

data_processing:
After removing 5’ and 3’end Illumina TruSeq adaptor sequences using Cutadapt, the reads were mapped to the mm10 genome assembly using HISAT2
Assembly: mm10
Supplementary files format and content: csv file includes cpm values for each sample

source_name: Liver

extract_protocol:
RNA was harvested using Trizol (Invitrogen). 1ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using TruSeq RNA Library Prep Kit v2 (Illumina)

value: Liver

value: Hepatocytes

value: tamoxifen


title: isolated hepatocytes, ALAS1/2-KO, Rep3


title: isolated hepatocytes, ALAS1/2-KO, Rep2


title: isolated hepatocytes, ALAS1/2-KO, Rep1


title: isolated hepatocytes, Wild-type, Rep2

value: WT

value: non


title: isolated hepatocytes, Wild-type, Rep1


title: hepatocyte, ALAS12-KO, rep2 (miRNA-seq)

data_processing:
After removing adaptor sequences of the small RNA sequencing libraries using Cutadapt, the reads were collapsed subsequently to remove PCR artifacts using fastx_collapser, and 5' and 3' end linker sequences were clipped using fastx_clipper. The remaining small RNA reads were mapped to the hg38 genome assembly, and small RNA counts are normalized by spike-in reads in each library. Differential small RNA expression analysis was performed by Bioconductor R package Limma.
Supplementary files format and content: mm10
Supplementary files format and content: tab-delimited file includes raw miRNA counts

source_name: liver

extract_protocol:
Mice were deeply anesthetized with ketamine/xylazine (100/10 mg/kg, i.p.) and given a toe pinch test to ensure sufficient anesthetization. For whole liver experiments, mice were perfused with 1x phosphate-buffered saline (PBS) via the left cardiac ventricle, and liver tissues were harvested Hepatocyte isolations were performed using the two-step collagenase perfusion method (PMID: 6273298). In brief, a lateral incision was made through the integument and abdominal wall up to the ribcage. The diaphragm and pleural cavities were incised, a 25G i.v. catheter was inserted into the hepatic portal vein, and the inferior vena cava was cut to create an outlet in the circulatory system for exsanguination. Liver Perfusion medium (Gibco, 30 mL), heated to 37℃, was pumped through the catheter line using a Gilson Minipuls 2 peristaltic pump at a rate of 6.5 mL/min, followed by 30 mL of 37℃ Hepatocyte Wash medium containing Collagenase B (0.05% Collagenase B solution). Liver tissue was excised, transferred to a petri dish containing 5 mL of liver digestion buffer [0.05% Collagenase B solution diluted 10X in Hepatocyte Wash medium (Gibco)], and incubated for 10-13 min at 37℃. Digested liver tissue was filtered through a 70 µm cell strainer and then centrifuged at 50 x g for 5 min at 4℃ to isolate a hepatocyte-enriched pellet. The hepatocytes were washed three times using Hepatocyte Wash medium and centrifugation at 50 x g for 5 min at 4℃.
One milligram of total RNA was used to generate small RNA libraries for each sample. After adding 1 μL of QIAseq TM miRNA Library QC Spike-Ins (QIAGEN) to the total RNA, a 3' linker was ligated to it in a reaction containing 100 mM 3' linker, T4 RNA Ligase 2, truncated KQ (NEB), 10% PEG8000, 1x RNA ligation buffer, and 20 U RNase OUT overnight at 17°C. The 48-57 nucleotide sized 3' linker-ligated small RNAs were purified using 12% Urea-PAGE gel with radio-labeled oligonucleotides serving as size-markers. Subsequently, the small RNA-linker hybrid was subjected to a 5’ ligation reaction overnight at 17°C containing 100 mM 5' adaptor, 20% PEG8000, 1x RNA ligase buffer, 1 mM ATP, 20 U RNaseOUT, and 5 U T4 RNA ligase 1. The linker-ligated RNAs were then reverse transcribed, PCR amplified using 12 cycles of PCR with forward and Illumina index reverse primers, and purified by 8% non-denaturing acrylamide gel. Sequencing was performed with HiSeq 4000 in SE50 mode.

treatment_protocol: Mice were housed in standard cages in a barrier facility at the Icahn School of Medicine at Mount Sinai (ISMMS) Center for Comparative Medicine and Surgery, maintained at 22℃ on a 12-12 hour light-dark cycle.

value: liver

value: hepatocyte

value: TMX


title: hepatocyte, ALAS12-KO, rep4 (miRNA-seq)


title: hepatocyte, ALAS12-KO, rep1 (miRNA-seq)


title: hepatocyte, ALAS12-KO, rep3 (miRNA-seq)


title: hepatocyte, Wild-type, rep2 (miRNA-seq)


title: hepatocyte, Wild-type, rep1 (miRNA-seq)


title: hepatocyte, Wild-type, rep3 (miRNA-seq)


title: s23_3

city: Rancagua

first: Nicolas

last: Santander

address: Libertador Bernardo O'Higgins 611

country: Chile

institute: Universidad de O´Higgins

data_processing:
Align to mm10 with Rsubread
Quantification with FeatureCounts
Normalization and diffierential expression with DESeq2
Assembly: mm10
Supplementary files format and content: tab delimited file with raw counts for each sample

source_name: Brain

extract_protocol:
Rneasy Plus micro kit Qiagen
NEBNext Ultra II RNA Library Prep Kit

value: Brain


title: nescon2

value: NestinCre


title: s23_1


title: nescon1


title: nescon3


title: s23_2


title: MGtgfb1_2

source_name: Microglia

value: Microglia


title: MGtgfb1_1


title: MGcx3con2


title: KLM1_Y318A2

value: Pancreatic ductal adenocarcinoma

value: MSLN knockout and then over-expressing Y318A mutant MSLN


title: MGcx3con1


title: Engineered heart issue_WT_2


title: tgfbr2_3


title: tgfbr2_2


title: isolated hepatocytes, Wild-type, Rep3


title: tgfbr2_1


title: tgfb1_2


title: tgfb1_1


title: cx3con1


title: cx3con3


title: cx3con4


title: cx3con2


title: b82


title: HEK293T, ALAS1-KO, rep3 (miRNA-seq)

data_processing:
After removing adaptor sequences of the small RNA sequencing libraries using Cutadapt, the reads were collapsed subsequently to remove PCR artifacts using fastx_collapser, and 5' and 3' end linker sequences were clipped using fastx_clipper. The remaining small RNA reads were mapped to the hg38 genome assembly, and small RNA counts are normalized by spike-in reads in each library. Differential small RNA expression analysis was performed by Bioconductor R package Limma.
Supplementary files format and content: hg38
Supplementary files format and content: tab-deliminted file includes raw miRNA counts

extract_protocol:
To analyze small RNAs, we isolated biologically independent total RNA samples from HEK293T, ALAS1-KO cells using Trizol (Invitrogen)
One milligram of total RNA was used to generate small RNA libraries for each sample. After adding 1 μL of QIAseq TM miRNA Library QC Spike-Ins (QIAGEN) to the total RNA, a 3' linker was ligated to it in a reaction containing 100 mM 3' linker, T4 RNA Ligase 2, truncated KQ (NEB), 10% PEG8000, 1x RNA ligation buffer, and 20 U RNase OUT overnight at 17°C. The 48-57 nucleotide sized 3' linker-ligated small RNAs were purified using 12% Urea-PAGE gel with radio-labeled oligonucleotides serving as size-markers. Subsequently, the small RNA-linker hybrid was subjected to a 5’ ligation reaction overnight at 17°C containing 100 mM 5' adaptor, 20% PEG8000, 1x RNA ligase buffer, 1 mM ATP, 20 U RNaseOUT, and 5 U T4 RNA ligase 1. The linker-ligated RNAs were then reverse transcribed, PCR amplified using 12 cycles of PCR with forward and Illumina index reverse primers, and purified by 8% non-denaturing acrylamide gel. Sequencing was performed with HiSeq 4000 in SE50 mode.

treatment_protocol: HEK293T cells were grown in a humidified incubator at 37°C, supplemented with 5% CO2. We cultured cells in DME-high glucose media containing 10% fetal bovine serum (FBS), and 100 IU/ml penicillin/streptomycin.

value: Human embryonic kidney 293T


title: HEK293T, ALAS1-KO, rep1 (miRNA-seq)


title: HEK293T, Wild-type, rep1 (miRNA-seq)


title: HEK293T, ALAS1-KO, rep2 (miRNA-seq)


title: b81


title: HEK293T, Wild-type, rep3 (miRNA-seq)


title: HEK293T, ALAS1-KO, rep1 (Ago2 CLEAR-CLIP)

description: Ago2 CLEAR-CLIP

data_processing:
After removing adaptor sequences of the small RNA sequencing libraries using Cutadapt, the reads were collapsed subsequently to remove PCR artifacts using fastx_collapser, and 5' and 3' end linker sequences were clipped using fastx_clipper. miRNA-target chimeric reads were analyzed by ChiRA tool (PMID: 33511995)
Supplementary files format and content: hg38
Supplementary files format and content: tab-delimited file includes miRNA-chimeric read counts

extract_protocol:
One 25 cm dish of confluent HEK293T and ALAS1-/- cells were used per sample. The cells were UV irradiated with 150mJ/cm2 UVC (254nM wavelength) two times. The cells were then lysed on ice with iCLIP lysis buffer (50 mM Tris-HCl pH7.4, 100 mM NaCl, 1% NP-40, 0.1% SDS, 0.5% SDC, protease inhibitor cocktail (Roche), and 40 U/ml RNaseOUT (Sigma)) for 20 min on ice. TurboDNase (Invitrogen #AM2238, 10U), RNase I (0.05 U/mL) were added subsequently, and samples were incubated at 37°C for 5 min. Treated lysates were immunoprecipitated with anti-Ago2 antibodies (Abnova) or control IgG (MilliporeSigma) coupled with Dynabeads protein G (Thermo Fisher Scientific) for 2 hrs at 4°C with gentle agitation. The immunoprecipitate was washed sequentially with high-stringency buffer, high-salt buffer (50 mM Tris-HCl pH 7.4, 1 M NaCl, 1 mM EDTA, 1% Nonidet P-40, 0.5% sodium deoxycholate, and 0.1% SDS), and PNK wash buffer (20 mM Tris-HCl pH 7.4, 10 mM MgCl2, and 0.2% Tween-20).
Samples were then phosphorylated at 37°C for 20 minutes in 50 μl of PNK mixture: 41.8ul H2O, 5 μl 10X PNK buffer (NEB), 1 μl RNaseOUT, 1.67 μl ATP (30 mM), 0.5 μl T4 PNK - 3' phosphatase minus (NEB M0236L). The immunoprecipitated materials were washed with PNK wash buffer, and then miRNA-mRNA ligation was carried out overnight at 17°C in 100 μl of mixture: 49.25 μl H2O, 30 μl 50% PEG-8000 (NEB #B1004), 10 μl 10x T4 RNA ligation buffer (NEB), 2.5 μl RNaseOUT, 1 μl ATP (100mM), 1 μl BSA (10 mg/ml), 6.25 μl T4 RNA ligase 1 (10U/μl – NEB M0204). The RNAs in the immunoprecipitate were dephosphorylated on the beads with FastAP Thermosensitive alkaline phosphatase (Thermo Fisher Scientific) and ligated to preadenylated adaptors in 40 μL of mixture: 17 μl H2O, 4 μl 10x T4 RNA ligase buffer (NEB), 1 μl of 3'-linker, 16 μl 50% PEG-8000, 1 μl RNaseOUT and 1μl T4 RNA ligase 2 truncated K227Q (NEB M0351) at room temperature for 5 hrs. Samples were then radiolabeled in 50 μl mixture: 5 μl 10X PNK buffer (NEB), 1 μl RNaseOUT, 1.5 μl γ-32P-ATP (15 μCi), 1 μl PNK enzyme (NEB M0201) and 41.5 μl H2O for 5 min at 37°C. Protein–RNA complexes were eluted from the beads, electrophoresed on 4–20% Bis-Tris NuPAGE gels (Thermo Fisher Scientific), transferred to nitrocellulose membranes (Whatman), and visualized by autoradiography. The area of the membrane corresponding with proteins that were ~100 kDa (the MW of Ago2) to ~150 kDa in size was excised from each lane of the blot. Excised lanes were treated with proteinase K (MilliporeSigma) and 7M urea (MilliporeSigma). The RNA-adaptor products were purified by acid phenol:chloroform: isoamyl alcohol extraction and ethanol precipitation. Purified RNAs were then subjected to reverse transcription using Superscript III (Thermo Fisher) with adapter-specific RT primers according to the manufacturer’s instruction. Resulted single-stranded cDNA products were purified using ExoSAP-IT (Thermo Fisher Scientific), followed by circularization using CircLigase (Illumina), and PCR-amplified. The resulting libraries were gel-purified and sequenced with the Illumina HiSeq platform.
CLEAR-CLIP

tag: rip antibody

value: anti-Ago2 antibodies (Abnova)


title: HEK293T, Wild-type, rep2 (Ago2 CLEAR-CLIP)

description: Ago2 CLEAR-CLIP


title: HEK293T, ALAS1-KO, rep2 (Ago2 CLEAR-CLIP)

description: Ago2 CLEAR-CLIP


title: HEK293T, ALAS1-KO, rep4 (miRNA-seq)


title: b83


title: Parathyroid gland, Control, ZT2, rep7

city: Copenhagen

first: Søren

last: Egstrand

address: Blegdamsvej 9

department: Depart. of Nephrology

country: Denmark

institute: Rigshospitalet

data_processing:
NovaSeq System Software generated base calls
Sequenced reads were alligned to the rn6 genome using STAR (version 2.7.8a+galaxy0)
Unambigously alligned reads were counted by featureCounts (version 2.0.1+galaxy1)
Genes with less then 10 counts in total were excluded
Counts were then normalized for sequencing depth and RNA composition using the median of ratios method employed in DESeq2 (version 1.30.1).
Counts were transformed using the rlog transformation function employed in DESeq2
Assembly: rn6
Supplementary files format and content: count_table.xlsx contains counts from featureCounts for all samples

organism: Rattus norvegicus

source_name: Parathyroid gland

extract_protocol:
For each sample one parathyroid gland was harvested, flash frozen in liquid nitrogen and store at -80 degrees. Total RNA was extracted using TRIzol (Sigma-Aldrich, St Louis, MO, USA) according to the manufacturer’s instruction and purified using Qiagen RNase-Free DNase Set (Qiagen, Hilden, Germany) on a QIAcube Connect (Qiagen, Hilden, Germany). RNA quality was examined using an Agilent Technologies 2100 Bioanalyzer. Library preparation was done using a TruSeq Stranded Total RNA Gold kit with dual adaptor indexing strategy; IDT for Illumina – TruSeq RNA UD Indexes (Illumina, San Diego, USA). The library was diluted and pooled to a concentration of 2 nM according to the procedure of “Denature and Dilute Libraries Guide of Illumina Novaseq 6000” and final loading concentration was 400 pM.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Parathyroid gland

tag: Sex

value: male

tag: time of_investigation


title: HEK293T, Wild-type, rep2 (miRNA-seq)


title: Parathyroid gland, Control, ZT2, rep6


title: Parathyroid gland, Control, ZT14, rep4


title: Parathyroid gland, Control, ZT2, rep5


title: Parathyroid gland, Control, ZT2, rep8


title: Parathyroid gland, Control, ZT2, rep3


title: Parathyroid gland, Control, ZT14, rep3


title: HEK293T, Wild-type, rep4 (miRNA-seq)


title: Parathyroid gland, Control, ZT2, rep4


title: Parathyroid gland, Control, ZT2, rep2


title: Parathyroid gland, Control, ZT14, rep2


title: HEK293T, Wild-type, rep1 (Ago2 CLEAR-CLIP)

description: Ago2 CLEAR-CLIP


title: Parathyroid gland, CKD+Cinacalcet, ZT2, rep7

value: Cinacalcet ZT2


title: Parathyroid gland, Control, ZT2, rep1


title: Parathyroid gland, Control, ZT14, rep1


title: Parathyroid gland, CKD+Cinacalcet, ZT2, rep8


title: Parathyroid gland, CKD, ZT2, rep7


title: Parathyroid gland, CKD, ZT14, rep7


title: Parathyroid gland, CKD+Cinacalcet, ZT2, rep6


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep10

value: Cinacalcet ZT14


title: Parathyroid gland, CKD+Cinacalcet, ZT2, rep5


title: Parathyroid gland, CKD+Cinacalcet, ZT2, rep4


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep9


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep8


title: Parathyroid gland, CKD+Cinacalcet, ZT2, rep2


title: Parathyroid gland, CKD+Cinacalcet, ZT2, rep3


title: Parathyroid gland, CKD, ZT14, rep5


title: Parathyroid gland, CKD, ZT14, rep6


title: Parathyroid gland, CKD, ZT2, rep6


title: Parathyroid gland, CKD, ZT2, rep5


title: Parathyroid gland, CKD, ZT2, rep4


title: Parathyroid gland, CKD, ZT14, rep4


title: Parathyroid gland, CKD, ZT14, rep3


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep4


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep5


title: Parathyroid gland, CKD, ZT2, rep3


title: Parathyroid gland, CKD, ZT14, rep1


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep6


title: KLM1_Mock3

value: Wild Type


title: Parathyroid gland, CKD, ZT14, rep2


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep2


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep1


title: Parathyroid gland, CKD, ZT2, rep2


title: Parathyroid gland, CKD, ZT2, rep1


title: DMSMapseq, CleanCap, denatured

description:
DMSMapseq with RNA
DMS CleanCap denatured
MutationRate.tsv
CC_denatured_MutRates.tsv

value: DMS, denature RNA


title: DMSMapseq, CleanCap, noDMS

description:
DMSMapseq with RNA
DMS CleanCap noDMS
MutationRate.tsv
CC_minusDMS_MutRates.tsv

value: no DMS


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep7


title: DMSMapseq, CleanCap

description:
DMSMapseq with RNA
DMS CleanCap Sample
MutationRate.tsv
CC_plusDMS_MutRates.tsv

value: DMS


title: KOXII5,IgG,biol rep 1

city: wuhan

first: xue

last: lilan

state: 请选择...

address: shizishan

institute: huazhong agriculture university

description: Library name: XII5_control_1

data_processing:
raw sequencing data were initially filtered using cutadapt (version 2.5) to remove low-quality reads and trim adaptor-contaminated sequences
Clean reads were mapped to the T. gondii reference genome (TGME49) using Bowtie2 (version 2.3.4.1) with default parameters
Library quality was assessed by calculating insert sizes using samtools (version 1.12) and analyzing read distribution around transcription start sites (TSS) with deepTools (version 3.5.1)
Peak calling was performed using MACS2 (version 2.1.1, -p 0.01 --shift 75 –extsize 150 --nomodel -B –SPMR --keep-dup all --call-summits) and IgG controls for thresholdingBigwigs were generated using bigWigMerge and bedGraphToBigWig (version 4.11.1). Peaks were annotated using the ChIPseeker (version 1.32.1)
Bigwigs were generated using bigWigMerge and bedGraphToBigWig (version 4.11.1). Peaks were annotated using the ChIPseeker (version 1.32.1)
Assembly: T. gondii reference genome (TGME49)
Supplementary files format and content: bigwig
Library strategy: CUT&Tag

organism: Toxoplasma gondii

source_name: toxoplasma

growth_protocol: ME49 and ME49ΔXII-5  were cultured in human foreskin fibroblasts (HFF), in Dulbecco’s Modified Eagle Medium (DMEM) (Sigma-Aldrich, USA), supplemented with 2% heat-inactivated fetal bovine serum (FBS) (Gibco, USA), 1% penicillin-streptomycin, and 5 mM L-glutamine.

extract_protocol:
Intracellular parasites were released by needle passage and purified through filtration with 3 μm pore-size membranes. Approximately 5 x 10⁶ tachyzoites were collected and processed using the Hyperactive Universal CUT&Tag Assay Kit for Illumina (Vazyme Biotech, China), following the manufacturer’s instructions.
The resulting DNA was amplified for library construction using the TD202 TruePrep Index Kit V2 for Illumina (Vazyme Biotech, China). The libraries were enriched, quantified, and sequenced on a NovaSeq 6000 sequencer (Illumina) using the PE150 model.

value: toxoplasma

value: ME49 deltaXII-5

tag: chip antibody

value: rabbit anti IgG


title: ME49,IgG,biol rep 2

description: Library name: ME49_control_2


title: KOXII5,HDAC3,biol rep 2

description: Library name: XII5_IP_2

value: rabbit anti-TgHDAC3(generated by immunizing rabbit with the synthetic peptide sequence 'CPYRIRRKDYANDFE' conjugated to KLH)


title: KOXII5,HDAC3,biol rep 1

description: Library name: XII5_IP_1


title: ME49,HDAC3,biol rep 2

description: Library name: ME49_IP_2


title: ME49,HDAC3,biol rep 1

description: Library name: ME49_IP_1


title: Parathyroid gland, CKD+Cinacalcet, ZT14, rep3


title: IPEC-J2 cellls, 0 μM OAA, 24h, rep2 IP

city: Hangzhou

first: Luoyi

last: Zhu

state: Zhejiang

address: 866 Yuhang Tang Road

institute: Zhejiang University

description:
Library name: Sample 3
C2_1

data_processing:
reads containing adapters were removed; reads containing more than 10% N were removed; reads with all A-bases were removed; remove low quality reads (the number of bases with quality value Q ≤ 20 accounts for more than 50% of the whole read).
Ribosome alignment: The short reads alignment tool bowtie2 was used to align high quality clean reads to the ribosome database of this species, and the reads on the aligned ribosomal RNA were removed.
Genome alignment: HISAT2 uses global and local search methods to effectively align spliced reads in RNA-Seq sequencing data, which is currently the most accurate alignment software with the highest alignment rate.
Next using Integrative Genomics Viewer (IGV) for visualization. Peak statistics for grouped samples: The R package exomePeak2 was used to perform genome-wide peak calling on a group basis, with a default threshold of p value<1e-5.
Assembly: Sus scrofa Ensembl_release106
Supplementary files format and content: raw_counts_all_samples.txt file includes raw counts for each Sample
Supplementary files format and content: CON_OAA_FPKM.txt files include FPKM values for each Sample

organism: Sus scrofa

source_name: cell line

growth_protocol: The resuscitated IPEC-J2 cells were cultured in DMEM/F12 complete medium (Gibco, 1320033) containing 10% FBS (Gibco, A5670701), 100 IU/mL penicillin, and 100 μg/mL streptomycin at 37°C, 5% CO2 concentration, and 100% humidity. IPEC-J2 cells were digested with trypsin and passaged when the cells reached 80-90% growth, and the medium was changed once a day.

extract_protocol:
RNA was extracted utilizing Trizol reagent.
After total RNA was extracted, the enriched RNA was fragmented into short fragments (about 100nt) by using fragmentation buffer. RNA was divided into two parts, one of which was used as the input (no IP experiment was performed). The other RNA was enriched with m6A specific antibody. The enriched RNA was reverse transcripted into cDNA with random primers. Next, the cDNA fragments was end repaired, and ligated to Illumina sequencing adapters. Finally we got qualified library for sequencing, sequenced using Illumina HiSeqTM 4000 (or other platforms) by Gene Denovo Biotechnology Co. (Guangzhou, China)

treatment_protocol: MeRIP-seq analysis of IPEC-J2 cells after treatment with 0 μM and 25 μM OAA for 24h

value: cell line

value: Intestinal Porcine Epithelial Cell

value: WT

value: 0 μM OAA

tag: batch


title: IPEC-J2 cellls, 0 μM OAA, 24h, rep2 input

description:
Library name: Sample 4
CON2_1

tag: tissue


title: IPEC-J2 cellls, 25 μM OAA, 24h, rep2 IP

description:
Library name: Sample 9
O2_1

value: 25 μM OAA


title: KOXII5,IgG,biol rep 2

description: Library name: XII5_control_2


title: IPEC-J2 cellls, 25 μM OAA, 24h, rep2 input

description:
Library name: Sample 10
OAA2_1


title: IPEC-J2 cellls, 25 μM OAA, 24h, rep1 IP

description:
Library name: Sample 7
O1_1


title: IPEC-J2 cellls, 25 μM OAA, 24h, rep3 input

description:
Library name: Sample 12
OAA3_1


title: IPEC-J2 cellls, 0 μM OAA, 24h, rep3 IP

description:
Library name: Sample 5
C3_1


title: ME49,IgG,biol rep 1

description: Library name: ME49_control_1


title: IPEC-J2 cellls, 25 μM OAA, 24h, rep3 IP

description:
Library name: Sample 11
O3_1


title: IPEC-J2 cellls, 0 μM OAA, 24h, rep3 input

description:
Library name: Sample 6
CON3_1


title: Smartseq2_764

first: Tiago

last: Castro

state: New York

address: 1230 York Avenue

institute: The Rockefeller University

data_processing:
SmartSeq2 Analysis: Fastq files from SmartSeq2 libraries were aligned to the mouse genome (mm39) with Gencode M29 annotations using STAR (v2.7.10a). Genome-mapped BAM files were quantified with RSEM (v1.3.1), and the resulting gene/UMI count matrix was processed in Seurat (v4.1.2). Data were normalized using the LogNormalize method, with mitochondrial gene expression and library size bias regressed out via the ScaleData function. Cells with >5% mitochondrial reads were excluded. Clustering, differential expression, and gene signature scoring were performed in Seurat. Differentially expressed genes were identified using the Wilcoxon Rank Sum test, with Bonferroni-adjusted P-values < 0.05 and log2 fold changes > 0.5 considered significant.
Bulk RNA-Seq raw Fastq files were aligned to the mouse genome (mm39) with Gencode M29 annotations using STAR (v2.7.10a). The count matrix was processed in R with the DESeq2 package (v1.34). Genes with fewer than ten reads were filtered out, and sample normalization was applied using a negative binomial distribution model. Differential gene expression between conditions was assessed with the Wald test. Genes with adjusted P-values (FDR) < 0.1 were retained, and log2 fold changes were shrunk using the apeglm algorithm.
10x Genomics Analysis: Fastq files from cDNA and hashtag libraries generated on the Chromium platform (10x Genomics) were processed using Cell Ranger (v7.0.1). The resulting quantification matrix was processed in Seurat (v5.0) with normalization using the SCTransform function. Non-myeloid cell contaminants were removed based on the PanglaoDB_Augmented_2021 database and EnrichR package for R. Cells were classified by biological condition using the HTODemux function. Differentially expressed genes were identified with the Wilcoxon rank test, with Bonferroni-adjusted P-values < 0.05 and log2 fold changes > 0.3 considered significant. Signature scores were calculated using Seurat’s AddModuleScore function.
Assembly: mm10
Supplementary files format and content: Tab files from bulk RNA-Seq tab files containing the STAR gene quantification.
Supplementary files format and content: gene.results from RSEM quantification of Smartseq2 data
Supplementary files format and content: mtx, cellranger output from 10x genomics libraries.
Supplementary files format and content: rds, images of Seurat objects used in the analysis

source_name: D-LN

extract_protocol:
Lymph nodes and gut tissue were dissected, finely chopped, and digested in RPMI medium with Collagenase D to obtain single-cell suspensions. For gut-associated lymph nodes, duodenal lymph nodes were identified using anatomical markers. Dendritic cells were isolated from the duodenal lamina propria by separating intestines, removing mucus with PBS and DTT, and stripping epithelium with EDTA. The tissue was further digested with DNaseI and collagenase, followed by Percoll gradient centrifugation to isolate DCs. Single-cell sorting of DCs was conducted into 96-well plates with TCL buffer and stored at –80°C.
Library Preparation for Single-cell RNA-seq (SmartSeq2). Single cells were sorted onto plates, covering all conditions in each replicate. RNA was extracted using RNAClean XP SPRI beads, then reverse-transcribed into cDNA with RT and TSO primers. The cDNA was amplified, cleaned with SPRI beads, and tagmented using the Nextera XT DNA Library Preparation Kit (Illumina). Barcoded libraries were pooled and sequenced on the Illumina Nextseq 550 platform. Library Preparation for Single-cell RNA-Seq (10x Genomics). Cells were labeled with hashtag oligonucleotide (HTO)-tagged antibodies, sorted, and counted for viability. scRNA-seq libraries were generated using the 10x Genomics Chromium system, following the manufacturer’s protocol. Libraries were sequenced on an Illumina NovaSeq SP flowcell at a minimum depth of 30,000 reads per cell. Library Preparation for Bulk RNA-seq. RNA was isolated using RNAClean XP beads. Reverse transcription was performed with specific primers, followed by cDNA amplification with KAPA HiFi HS Ready Mix. The amplified product was cleaned with RNAClean XP beads and tagmented using the Nextera XT DNA Library Preparation Kit (Illumina). Libraries were pooled and sequenced as 75-base single-end reads on an Illumina NextSeq platform.

treatment_protocol: Mice were treated with dietary ovalbumin (OVA) to induce tolerance. For infection conditions, mice were infected with Heligmosomoides polygyrus (H.p.) or Strongyloides venezuelensis (S.v.) to model chronic and acute type 2 immune responses, respectively.

value: D-LN

value: Filtered

tag: number of_cells_sorted


title: Smartseq2_762


title: Smartseq2_763

value: HP


title: Smartseq2_766


title: Smartseq2_767


title: Smartseq2_760


title: IPEC-J2 cellls, 25 μM OAA, 24h, rep1 input

description:
Library name: Sample 8
OAA1_1


title: IPEC-J2 cellls, 0 μM OAA, 24h, rep1 IP

description:
Library name: Sample 1
C1_1


title: IPEC-J2 cellls, 0 μM OAA, 24h, rep1 input

description:
Library name: Sample 2
CON1_1


title: Smartseq2_765


title: Smartseq2_759


title: Smartseq2_756


title: Smartseq2_757


title: Smartseq2_758


title: Smartseq2_755


title: Smartseq2_754


title: Smartseq2_761


title: Smartseq2_753


title: Smartseq2_740


title: Smartseq2_748


title: Smartseq2_751


title: Smartseq2_732


title: Smartseq2_730


title: Smartseq2_746


title: Smartseq2_743


title: Smartseq2_747


title: Smartseq2_750


title: Smartseq2_739


title: Smartseq2_752


title: Smartseq2_742


title: Smartseq2_735


title: Smartseq2_749


title: Smartseq2_733


title: Smartseq2_731


title: Smartseq2_727


title: Smartseq2_726


title: Smartseq2_745


title: Smartseq2_744


title: Smartseq2_738


title: Smartseq2_741


title: Smartseq2_734


title: Smartseq2_737


title: Smartseq2_724


title: Smartseq2_736


title: Smartseq2_723


title: Smartseq2_721


title: Smartseq2_720


title: Smartseq2_719

value: OV


title: Smartseq2_725


title: Smartseq2_722


title: Smartseq2_728


title: Smartseq2_729


title: Smartseq2_708


title: Smartseq2_713


title: Smartseq2_715


title: Smartseq2_717


title: Smartseq2_712


title: Smartseq2_710


title: Smartseq2_706


title: Smartseq2_714


title: Smartseq2_711


title: Smartseq2_718


title: Smartseq2_703


title: Smartseq2_702


title: Smartseq2_707


title: Smartseq2_709


title: Smartseq2_704


title: Smartseq2_700


title: Smartseq2_698


title: Smartseq2_699


title: Smartseq2_705


title: Smartseq2_697


title: Smartseq2_701


title: Smartseq2_696


title: Smartseq2_695


title: Smartseq2_694


title: Smartseq2_690


title: Smartseq2_692


title: Smartseq2_685


title: Smartseq2_686


title: Smartseq2_688


title: Smartseq2_682


title: Smartseq2_681


title: Smartseq2_684


title: Smartseq2_680


title: Smartseq2_689


title: Smartseq2_683


title: Smartseq2_687


title: Smartseq2_691


title: Smartseq2_677


title: Smartseq2_678


title: Smartseq2_676


title: Smartseq2_675


title: Smartseq2_670


title: Smartseq2_668


title: Smartseq2_674


title: Smartseq2_671


title: Smartseq2_672


title: Smartseq2_669


title: Smartseq2_667


title: Smartseq2_661


title: Smartseq2_673


title: Smartseq2_660


title: Smartseq2_657


title: Smartseq2_652


title: Smartseq2_679


title: Smartseq2_654


title: Smartseq2_664


title: Smartseq2_655


title: Smartseq2_663


title: Smartseq2_653


title: Smartseq2_665


title: Smartseq2_649


title: Smartseq2_659


title: Smartseq2_648


title: Smartseq2_693


title: Smartseq2_643


title: Smartseq2_645


title: Smartseq2_644


title: Smartseq2_640


title: Smartseq2_666


title: Smartseq2_641


title: Smartseq2_646


title: Smartseq2_637


title: Smartseq2_642


title: Smartseq2_658


title: Smartseq2_647


title: Smartseq2_636


title: Smartseq2_630


title: Smartseq2_639


title: Smartseq2_634


title: Smartseq2_629


title: Smartseq2_631


title: Smartseq2_632


title: Smartseq2_628


title: Smartseq2_627


title: Smartseq2_625


title: Smartseq2_623


title: Smartseq2_635


title: Smartseq2_626


title: Smartseq2_622


title: Smartseq2_621


title: Smartseq2_620


title: Smartseq2_633


title: Smartseq2_619


title: Smartseq2_618


title: Smartseq2_616


title: Smartseq2_613


title: Smartseq2_614


title: Smartseq2_611


title: Smartseq2_615


title: Smartseq2_656


title: Smartseq2_610


title: Smartseq2_609


title: Smartseq2_617


title: Smartseq2_608


title: Smartseq2_601


title: Smartseq2_604


title: Smartseq2_651


title: Smartseq2_605


title: Smartseq2_603


title: Smartseq2_600


title: Smartseq2_716


title: Smartseq2_624


title: Smartseq2_599


title: Smartseq2_596


title: Smartseq2_595


title: Smartseq2_597


title: Smartseq2_612


title: Smartseq2_598


title: Smartseq2_594


title: Smartseq2_593


title: Smartseq2_592


title: Smartseq2_588


title: Smartseq2_589


title: Smartseq2_606


title: Smartseq2_591


title: Smartseq2_607


title: Smartseq2_581


title: Smartseq2_585


title: Smartseq2_582


title: Smartseq2_586


title: Smartseq2_580


title: Smartseq2_583


title: Smartseq2_584


title: Smartseq2_578


title: Smartseq2_579


title: Smartseq2_574


title: Smartseq2_573


title: Smartseq2_577


title: Smartseq2_572


title: Smartseq2_576


title: Smartseq2_575


title: Smartseq2_571


title: Smartseq2_568


title: Smartseq2_570


title: Smartseq2_569


title: Smartseq2_567


title: Smartseq2_562


title: Smartseq2_566


title: Smartseq2_564


title: Smartseq2_563


title: Smartseq2_554


title: Smartseq2_556


title: Smartseq2_565


title: Smartseq2_550


title: Smartseq2_561


title: Smartseq2_560


title: Smartseq2_558


title: Smartseq2_552


title: Smartseq2_559


title: Smartseq2_557


title: Smartseq2_547


title: Smartseq2_545


title: Smartseq2_551


title: Smartseq2_549


title: Smartseq2_542


title: Smartseq2_555


title: Smartseq2_548


title: Smartseq2_538


title: Smartseq2_535


title: Smartseq2_546


title: Smartseq2_537


title: Smartseq2_544


title: Smartseq2_534


title: Smartseq2_543


title: Smartseq2_532


title: Smartseq2_531


title: Smartseq2_536


title: Smartseq2_529


title: Smartseq2_533


title: Smartseq2_553


title: Smartseq2_528


title: Smartseq2_526


title: Smartseq2_527


title: Smartseq2_541


title: Smartseq2_530


title: Smartseq2_524


title: Smartseq2_521


title: Smartseq2_519


title: Smartseq2_523


title: Smartseq2_540


title: Smartseq2_525


title: Smartseq2_522


title: Smartseq2_517


title: Smartseq2_520


title: Smartseq2_514


title: Smartseq2_511


title: Smartseq2_515


title: Smartseq2_516


title: Smartseq2_518


title: Smartseq2_512


title: Smartseq2_513


title: Smartseq2_509


title: Smartseq2_507


title: Smartseq2_508


title: Smartseq2_506


title: Smartseq2_510


title: Smartseq2_503


title: Smartseq2_501


title: Smartseq2_499


title: Smartseq2_498


title: Smartseq2_497


title: Smartseq2_495


title: Smartseq2_494


title: Smartseq2_539


title: Smartseq2_505


title: Smartseq2_504


title: Smartseq2_502


title: Smartseq2_496


title: Smartseq2_500


title: Smartseq2_489


title: Smartseq2_487


title: Smartseq2_483


title: Smartseq2_482


title: Smartseq2_481


title: Smartseq2_480


title: Smartseq2_478


title: Smartseq2_475


title: Smartseq2_493


title: Smartseq2_491


title: Smartseq2_492


title: Smartseq2_486


title: Smartseq2_490


title: Smartseq2_470


title: Smartseq2_485


title: Smartseq2_479


title: Smartseq2_484


title: Smartseq2_466


title: Smartseq2_467


title: Smartseq2_464


title: Smartseq2_472


title: Smartseq2_477


title: Smartseq2_488


title: Smartseq2_476


title: Smartseq2_474


title: Smartseq2_462


title: Smartseq2_471


title: Smartseq2_460


title: Smartseq2_473


title: Smartseq2_469


title: Smartseq2_456


title: Smartseq2_453


title: Smartseq2_455


title: Smartseq2_468


title: Smartseq2_451


title: Smartseq2_465


title: Smartseq2_463


title: Smartseq2_461


title: Smartseq2_459


title: Smartseq2_457


title: Smartseq2_454


title: Smartseq2_452


title: Smartseq2_449


title: Smartseq2_450


title: Smartseq2_447


title: Smartseq2_442


title: Smartseq2_445


title: Smartseq2_443


title: Smartseq2_444


title: Smartseq2_441


title: Smartseq2_435


title: Smartseq2_587


title: Smartseq2_436


title: Smartseq2_437


title: Smartseq2_434


title: Smartseq2_440


title: Smartseq2_433


title: Smartseq2_431


title: Smartseq2_430


title: Smartseq2_429


title: Smartseq2_446


title: Smartseq2_425


title: Smartseq2_428


title: Smartseq2_426


title: Smartseq2_432


title: Smartseq2_448


title: Smartseq2_427


title: Smartseq2_420


title: Smartseq2_439


title: Smartseq2_424


title: Smartseq2_422


title: Smartseq2_438


title: Smartseq2_419


title: Smartseq2_421


title: Smartseq2_418


title: Smartseq2_416


title: Smartseq2_415


title: Smartseq2_412


title: Smartseq2_411


title: Smartseq2_410


title: Smartseq2_423


title: Smartseq2_409


title: Smartseq2_408


title: Smartseq2_405


title: Smartseq2_404


title: Smartseq2_417


title: Smartseq2_401


title: BMDM, KO_Unst_5

city: Chicago

first: Rogan

middle: A

last: Grant

state: IL

address: 303 E Superior St, Simpson Querrey Biomedical Research Center 5-522

department: Pulmonary and Critical Care Medicine

institute: Northwestern University

data_processing:
FASTQ files were generated using bcl-convert 4.0.3 using default parameters. To facilitate reproducible analysis, samples were processed using the publicly available nf-core/RNA-seq pipeline version 3.9 implemented in Nextflow 22.04.5.5708 using Singularity 3.8.1 with the minimal command nextflow run nf-core/rnaseq \ -r ‘3.9’ \ -profile nu_genomics \--additional_fasta ‘transgenes.fa’ \ --star_index false \ --genome ‘GRCm38’ for SMARTer-seq samples.
Assembly: GRCm38
Supplementary files format and content: Raw counts were imported directly from Salmon using tx2gene within nf-core/rnaseq 3.9 and exported with relevant metadata directly to CSV

source_name: Bone-Marrow-Derived Macrophage (BMDM)

growth_protocol: Bone marrow was isolated from mice and plated in 10 cm Primaria tissue culture plates (ThermoFisher 25382-701). To induce differentiation into macrophages, cells were cultured in RPMI medium containing 11 mM glucose, 10% fetal+ serum (Atlas Biologics P16E19A1), 1 mM methyl pyruvate (Sigma 371173), 400 μM uridine (Sigma U3003), 1% antibiotic/mycotic (ThermoFisher 15-240-062) 1% Hepes (ThermoFisher MT25060CI) and 4 mM glutamine (Gibco, catalog no.11965-126) supplemented with 20ng ml–1 M-CSF (Peprotech 315-02) at 37 °C with 5% CO2. The medium was changed every 3 days, and BMDMs were harvested by scraping on day 6 and plated in 12-well plates (2 million cells per well), 24-well plates (940,000 cells per well), 48-well plates (300,000 cells per well) or 96-well plates (150,000 cells per well).

extract_protocol:
RNA was isolated using the Qiagen Allprep Mini kit according to manufacturer's instructions
RNA quality and quantity were assessed using TapeStation 4200 High Sensitivity RNA tapes (Agilent), and RNA-seq libraries were prepared from 100ng of total RNA using the NEBNext® Ultra™ DNA Library Prep Kit for Illumina® (NEB E7370L). Library QC was then performed using TapeStation 4200 High Sensitivity DNA tapes (Agilent). Dual-indexed libraries were pooled and sequenced on a NextSeq2000 instrument (Illumina) for 100 cycles, single-end, to an average sequencing depth of 14.99 million reads per sample.

treatment_protocol: BMDMs were stimulated with 0.1µg/mL ultrapure O5:B55 LPS (Invivogen tlrl-pb5lps) or IL-4 10ng/mL for 18h prior to collection.

value: Bone-Marrow-Derived Macrophage (BMDM)

value: KO

value: Unst


title: BMDM, KO_LPS_5

value: LPS


title: BMDM, WT_Unst_5


title: BMDM, WT_LPS_5


title: Smartseq2_399


title: BMDM, KO+AOX_IL-4_3

value: KO+AOX


title: Smartseq2_413


title: Smartseq2_414


title: TIRE-seq Murine DC library

city: Melbourne

first: Daniel

middle: Victor

last: Brown

address: 1G Royal Parade

institute: Walter and Eliza Hall Institute

description:
Library name: TIRE_DCs
Reverse transcription based sequencing
See feature_README.txt for sample metadata and barcodes.

data_processing:
Fastq data was processed into count matrices with v2.9.7 zUMIs.
Count matrices were converted into SingleCellExperiment objects with Bioconductor package SingleCellExperiment v1.29.1
Assembly: Reads were mapped against 10x Genomics pre-built references (July 7, 2020 version); GRCh38 v2-7-3a for human experiments mm10-2020-A for mouse experiments or hg19 and mm10 for mixed species experiments.
Supplementary files format and content: R Data Serialization (.rds) object that can be read into R with the readRDS() function

source_name: Dendritic cells

extract_protocol:
UHRR was a commerical source of RNA (ThermoFisher QS0639). Prime-seq was SPRI based RNA extraction. No extraction for TIRE-seq.
Prime-seq and TIRE-seq as described in protocols.io dx.doi.org/10.17504/protocols.io.j8nlk8rqdl5r/v1

value: Dendritic cells

value: Flt3L stimulated


title: Smartseq2_398


title: BMDM, WT_IL-4_5


title: Smartseq2_396


title: BMDM, KO+AOX_LPS_4


title: Smartseq2_395


title: Smartseq2_407


title: Smartseq2_394


title: BMDM, KO_LPS_4


title: Smartseq2_406


title: BMDM, WT_LPS_4


title: BMDM, KO+AOX_Unst_3


title: Smartseq2_403


title: Smartseq2_402


title: BMDM, KO_Unst_4


title: Smartseq2_400


title: Smartseq2_392


title: Smartseq2_391


title: BMDM, WT_Unst_4


title: BMDM, KO_IL-4_5


title: Smartseq2_397


title: Smartseq2_390


title: Smartseq2_387


title: BMDM, WT_IL-4_4


title: BMDM, KO+AOX_LPS_3


title: Smartseq2_388


title: Smartseq2_389


title: BMDM, KO+AOX_IL-4_2


title: BMDM, KO_IL-4_4


title: Smartseq2_384


title: Smartseq2_386


title: BMDM, KO_LPS_3


title: BMDM, WT_LPS_3


title: Smartseq2_385


title: BMDM, KO_Unst_3


title: BMDM, WT_Unst_3


title: Smartseq2_393


title: BMDM, KO+AOX_IL-4_1


title: BMDM, KO_IL-4_3


title: Smartseq2_382

value: SV


title: Smartseq2_380


title: Smartseq2_381


title: Smartseq2_383


title: Smartseq2_379


title: BMDM, KO_LPS_2


title: BMDM, WT_IL-4_3


title: Smartseq2_378


title: BMDM, WT_LPS_2


title: Smartseq2_377


title: BMDM, KO+AOX_LPS_2


title: BMDM, KO+AOX_Unst_2


title: Smartseq2_375


title: Smartseq2_373


title: Smartseq2_376


title: BMDM, KO_IL-4_2


title: Smartseq2_374


title: Smartseq2_372


title: BMDM, KO_Unst_2


title: BMDM, WT_Unst_2


title: Control_3

city: Atlanta

first: Teele

last: Palumaa

state: Georgia

address: 1365 Clifton Rd NE Building B

department: Ophthalmology

institute: Emory University

description:
Library name: SL33274
Column name in processed files: Control_3

data_processing:
FASTQ files were uploaded to Galaxy platform (version 24.1.4)
Sequenced reads were trimmed for low-quality bases using Trimmomatic (version 0.39), parameters: SLIDINGWINDOW:4:20.
Transcript abundances were quantified using Salmon (version 1.10.1) with quasi-alignment mode and a k-mer size of 31. Paired-end reads were mapped to the reference transcriptome index built from Mus musculus GRCm38 cDNA sequences. Effective length correction and fragment length distribution modeling were applied. Quantification estimates were aggregated to the gene level for downstream analysis.
Gene-level counts were generated from transcript abundances using the tximport package (version 1.28.0) in R (version 4.3.1).
Assembly: mm10
Supplementary files format and content: raw_counts: The CSV file was generated using the data processing steps listed above. The table contains raw counts per sample.
Supplementary files format and content: normalized_counts: The CSV file was generated using the data processing steps listed above. The table contains normalized counts per sample.

source_name: retina

growth_protocol: C57BL/6J mice were housed in a standard 12-12 hour light-dark cycle. Food was provided ad libitum.

extract_protocol:
Total RNA was extracted using TRIzol (Invitrogen) and RNeasy Micro kit (QIAGEN) according to the manufacturers’ protocols.
RNA-seq libraries were prepared using poly-A enrichment with the TruSeq RNA Library Prep Kit v2.

treatment_protocol: From postnatal day (P) 28, mice were exposed to form-deprivation myopia (FDM) by surgically attaching a head-mounted diffuser goggle over the right eye. The left eye remained uncovered and served as an internal control.

value: retina

value: male

value: wild-type

value: Contralateral control

tag: age

tag: refractive error_at_p42


title: Smartseq2_370


title: Smartseq2_371

tag: treatment


title: Control_1

description:
Library name: SL33272
Column name in processed files: Control_1

tag: genotype


title: Smartseq2_369


title: Control_2

description:
Library name: SL33273
Column name in processed files: Control_2


title: BMDM, KO_LPS_1

tag: cell type


title: BMDM, KO+AOX_LPS_1


title: FDM_2

description:
Library name: SL33271
Column name in processed files: FDM_2

value: FDM for 2 weeks


title: BMDM, WT_LPS_1

value: WT


title: Testis, western diet, Tas1r3 KO 2

city: Seoul

first: Hobin

last: Seong

state: KOREA, REPUBLIC OF

address: Gwanak-ro 1, Seoul National University

country: South Korea

institute: Seoul National University

data_processing:
Partek® Flow® v7.0
Raw sequence reads were trimmed and filtered by removing adaptor sequences and low-quality sequence reads.
Trimmed high-quality sequence reads were mapped and aligned using STAR method.
Aligned reads in gene transcripts were quantified to RefSeq transcripts using the Partek E/M method and normalized as FPKM using the mixed-model approach in Partek® Flow® v7.0
Assembly: mm39
Supplementary files format and content: Excel worksheet files include FPKM values, gene location, transcript id and gene symbol for each Sample

source_name: Testis

extract_protocol:
RNA was harvested using RNAqueous 4PCR Kits (Ambion). 2 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using Ion Total RNA-Seq Kit v2 following manufacturer's protocols.

value: Testis

value: Male

value: Tas1r3 KO

value: western diet (18 weeks)


title: Smartseq2_368

value: D-LN


title: BMDM, KO+AOX_Unst_1

value: Bone-Marrow-Derived Macrophage (BMDM)

value: Unst


title: Smartseq2_367


title: Testis, western diet, Tas1r3 KO 1


title: Testis, western diet, WT 2


title: NR+DNA_3

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

hyb_protocol: Once the hybridization reactions have been removed from the thermal cycler, proceed immediately to an nCounter Prep Station. This step can process up to 12 samples per run in approximately 2.5 to 3 hours.

scan_protocol: Next, the nCounter Digital Analyzer which is a multi-channel epifluorescence scanner collects raw data by taking images of the immobilized fluorescent reporters in the sample cartridge with a CCD camera through a microscope objective lens. Images are processed internally and the results are exported as a.csv file.

data_processing: RRC format files containing the raw data  were analyzed via the NanoString nCounter nSolver 4.0 software (MAN-C0019-08) with the NanoString Advanced Analysis 2.0 plugin (MAN-10,030-03) following the NanoString Gene Expression Data Analysis Guidelines (MAN-C0011-04). Both positive control and housekeeping normalization were used to normalize all sources of variation associated with the platform. Data was analyzed by ROSALIND® (https://rosalind.bio/), with a HyperScale architecture developed by ROSALIND, Inc. (San Diego, CA). Read Distribution percentages, violin plots, identity heatmaps, and sample MDS plots were generated as part of the QC step. Normalization, fold changes and p-values were calculated using criteria provided by Nanostring. ROSALIND® follows the nCounter® Advanced Analysis protocol of dividing counts within a lane by the geometric mean of the normalizer probes from the same lane. Housekeeping probes to be used for normalization are selected based on the geNorm algorithm as implemented in the NormqPCR R library1. Abundance of various cell populations is calculated on ROSALIND using the Nanostring Cell Type Profiling Module. ROSALIND performs a filtering of Cell Type Profiling results to include results that have scores with a p-Value greater than or equal to 0.05. Fold changes and pValues are calculated using the fast method as described in the nCounter® Advanced Analysis 2.0 User Manual. P-value adjustment is performed using the Benjamini-Hochberg method of estimating false discovery rates (FDR).

label: NanoString barcodes

source_name: HMC3 cells

label_protocol: Label a 12-tube PCR hybridization strip. The purified RNA is diluted in nuclease-free water to 20 ng/uL. Recommended total RNA samples are 20 ng to 100 ng for hybridization assay. The prepared samples are used as input into nCounter hybridization reactions containing NanoString Reporter and Capture probes. First, remove Reporter CodeSet and Capture ProbeSet tubes from the freezer and thaw at room temperature. Second, after it has thawed, inspect the tube of Reporter CodeSet to make sure no colored precipitate is present. Third, If you see a colored precipitate, heat the entire tube to 75°C for 10 minutes and cool at room temperature before using .The purified RNA was hybridized at thermal cycler at 65°C for 16 to 24 hours into nCounter 15 µL of hybridization buffer containing NanoString Reporter and Capture probes. Add 8 μL of Master Mix to each well of a strip tube. Add 5 µL of sample to each tube containing Master Mix. Add 2 µL of Capture ProbeSet to each tube. Maximum hybridization time should not exceed 48 hours.

extract_protocol: Total RNA was purified from cells using RNeasy Mini Kit (Qiagen) as per the manufacturer’s protocol. Purified RNA was quantified on a NanoDrop ND-1000 spectrophotometer and diluted in nuclease-free water to 20 ng/uL.

treatment_protocol: HMC3 cells were treated with 10 µM UA or 3 mM NR for 1 week. For gene expression analysis, we used the human neuroinflammation panel (XT-CSO-MNROI1-12).

tag: cell line

value: HMC3 cells

value: NR+DNA


title: BMDM, KO_Unst_1

value: KO


title: Testis, western diet, WT 1

tag: strain


title: BMDM, WT_IL-4_2


title: NR_3

value: NR


title: BMDM, WT_Unst_1


title: Smartseq2_365

value: SV


title: UA_3

value: UA


title: UA+DNA_3

value: UA+DNA


title: DNA_3

value: DNA


title: FDM_1

description:
Library name: SL33269
Column name in processed files: FDM_1

value: retina

value: male

value: wild-type

value: FDM for 2 weeks


title: Smartseq2_364


title: NR+DNA_2


title: BMDM, WT_IL-4_1

value: WT


title: ctr_3

value: control


title: NR_2


title: shLUC_7

city: Moscow

first: Anton

last: Zhiyanov

address: 20 Myasnitskaya ulitsa

department: Faculty of Biology and Biotechnology

country: Russia

institute: HSE University

description: Library name: shLUC_7

data_processing:
Quality of RNA-seq FASTQ files was assessed with FastQC v0.11.9 (Babraham Bioinformatics, Cambridge, UK).
Adapter sequences were removed with the use of cutadapt v2.10.
miRNA read counts were obtained using miRDeep2 v2.0.1.2. miRbase v22.1 database was a source of reference miRNA sequences.
Assembly: GENCODE GRCh38.p13
Supplementary files format and content: counts.tsv – tab-delimited miRNA read counts table

source_name: colon

growth_protocol: The human colon adenocarcinoma cell line HT-29 was purchased from the European Cell Culture Collection (Porton Down, Wiltshire, UK). Cells were grown in RPMI 1640 + L-Glutamine (Gibco, Thermo Fisher Scientific, MA, USA) supplemented with 10% fetal calf serum (Gibco), 1% penicillin and streptomycin (Gibco) and cultured under standard cell culture conditions (37°C, saturated humidity, 5% CO2). The HT-29 cells were additionally tested for mycoplasma contamination. CD44 knockdown in HT-29 cells was achieved by a shRNA-mediated approach. All animals used were pathogen-free Balb/c severe combined immunodeficient (SCID) mice aged 9-14 weeks with a weight of 25-30 g at the beginning of the experiment. Each animal was inoculated subcutaneously (s. c.) above the right scapula with 1 x 106 shLUC or shCD44 HT-29 cells in medium without supplements. The mice were euthanized when the tumors ulcerated or reached a size of ~1.5 cm³.

extract_protocol:
Approximately 50 mg of fresh-frozen xenograft PT tissue was crushed in liquid nitrogen in the presence of QIAzol Lysis Reagent (Qiagen, Hilden, Germany). The total RNA was isolated using miRNeasy Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions. All RNA samples were treated with DNase I during the isolation procedure. The RNA yield was determined by UV absorbance using a NanoDrop 1000 spectrophotometer (Peqlab, Erlangen, Germany). The RNA quality was assessed by analyzing the ribosomal RNA integrity number (RIN) on an Agilent 2100 Bioanalyzer using the RNA 6000 Nano kit (Agilent Technologies, Palo Alto, CA, USA). The RIN values of the isolated RNA samples ranged from 6.7 to 9.6.
Libraries for miRNA sequencing were prepared from total RNA samples using NEBNext Multiplex Small RNA Library Prep Kit for Illumina. Each sample was sequenced on the Illumina NextSeq 550 to generate single-end 50-nucleotide reads: 7 control samples (shLUC) and 8 samples with CD44 knockdown (shCD44).

value: colon

value: human colorectal adenocarcinoma

tag: origin

value: Xenograft

value: SCID

value: shLUC


title: UA+DNA_2

value: HMC3 cells


title: Smartseq2_363

value: D-LN


title: UA_2


title: DNA_2


title: shLUC_6

description: Library name: shLUC_6


title: ctr_2


title: Smartseq2_361

value: SV


title: NR+DNA_1

value: NR+DNA


title: shLUC_5

description: Library name: shLUC_5


title: Smartseq2_366


title: NR_1

value: NR


title: Smartseq2_360


title: shLUC_4

description: Library name: shLUC_4


title: UA+DNA_1

value: UA+DNA


title: BMDM, KO_IL-4_1

value: Bone-Marrow-Derived Macrophage (BMDM)

value: KO


title: UA_1

value: UA


title: Smartseq2_359


title: ctr_1


title: shLUC_3

description: Library name: shLUC_3


title: Smartseq2_358