title: Ground Control, Leaf Rep. 1

description: LEAFFT.vs.LEAFGC diff exp

data_processing:
The overall quality of demultiplexed RNA-Seq sequence data is evaluated using FastQC before proceeding with downstream analysis. If problems are identified, corrective actions will be recommended before proceeding with further data analysis steps.
While acceptable sequence quality is expected, the poor quality reads are removed and low confidence sequences are trimmed using Trimmomatic. FastQC analysis is repeated after Trimmomatic filtering to ensure only high confidence reads enter downstream processing steps.
The Star Aligner is used for mapping high quality paired-end reads to Arabidopsis thaliana accession Ws-0.
Gene expression values are obtained using RSEM, which uses a generative statistical model accommodating multiple features of the RNA-Seq read generation process and is capable of handling ambiguously aligned reads to efficiently estimate gene expression.
The estimated counts of mapped read-pairs at each gene locus are normalized using an Upper-Quartile method and then differentially expressed (DE) genes are called using edgeR by designing a generalized linear regression model. The unsupervised learning methods, hierarchical clustering and PCA, are used to examine broad patterns of DE genes to uncover the overall pattern of the samples between Ground control and Spaceflight.
Genome_build: Arabidopsis thaliana WS reference genome (Mott et al. Nature 2011)
Supplementary_files_format_and_content: .xls spreadsheets containing lists of genes,  FPKM values, and CPM values. There are two comparisons: Root FT vs. Root GC and Leaf FT vs. Leaf GC.

growth_protocol: Dry, sterilized Arabidopsis seeds of wild-type cultivar Wassilewskija  were planted aseptically on the surface of 10 cm2 solid media plates comprised of 0.5% Phytagel/0.5× MS media, and then immediately wrapped in light-tight black cloth (Duvetyne, SeattleFabrics.com) and transported to KSC (Kennedy Space Center), FL, USA. The wrapped plates were stored at temperatures between 4 and 10°C and remained dormant until removal from cold stowage and exposure to light at the initiation of the APEX03-2 experiment on the ISS (International Space Station). Plates were integrated and grown in the Vegetable Production System (VPS/Veggie) hardware on the Columbus Module of the ISS. The 10 cm2 Petri plates were housed perpendicular (“vertical”) to the overhead LED lighting of the VPS. Plants received between 100–135 μmoles/m2/s PAR inside the VPS. The plants were grown for eleven days after germination, which was initiated by unwrapping the plates and exposing them to light in the VPS. The ground control plates were grown inside the VPS ground unit within the ISSES (International Space Station Environment Simulator) chamber at KSC with a 48-h delay. The ISSES chamber replicated the temperature, CO2 levels and lighting that had been experienced by the spaceflight plants in the previous 48 h. After eleven days, seedlings were harvested by an astronaut into a KFT (Kennedy Fixation Tube) containing RNAlater. Once the plants were placed in the KFTs, the KFT was actuated with RNAlater to chemically fix the sample. At 24 hours post-harvest, KFTs were then transferred to MELFI (Minus Eight Laboratory Freezer for ISS) and were kept at -80°C throughout the delivery process to the Ferl lab at University of Florida.

extract_protocol:
Seedlings stored in RNA later were recovered from -80°C freezer. Each tube containing the seedlings was allowed to thaw in the fridge overnight. After completely thawing the samples to room temperature, the seedlings were observed under the microscope. The preserved seedlings from the spaceflight and ground control harvests were dissected into distinct plant parts: leaf, hypocotyl, root tip and then the rest of the root. The root tip was defined as the last 2mm of the root for the light-grown plants and the last 1mm for the dark-grown plants. All the sections dissected were saved in RNA later and stored back to -80°C freezer.  Three independent roots were used as three biological replicates for the transcriptomic analysis. The root apex was dissected as 2mm root tip section for light grown samples and 1mm section for the dark grown samples. Dissected root tips were carefully transferred into pre-labeled micro-centrifuge tubes, flash frozen in liquid N. Total RNA was extracted from these frozen samples using Picopure RNA isolation kit (Arcturus®, Life technologies). RNA concentration was determined on a Qubit (Thermo Fisher/Life Technologies, Inc) and sample quality was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Inc.). Five ng RNA from each sample was used for low input array and RNA sequencing.
RNASeq Libraries were prepared using SMART-Seq V4 ultra low input RNA kit for sequencing combined with Illumina Nextera DNA Sample Preparation Kit. Ten ng of total RNA was used for cDNA library construction using CloneTech SMART-Seq V4 ultra low input RNA kit for sequencing (Clontech Laboratories, Inc, cat#: 634890) according to manufacturer's protocol. Briefly, 1st strand cDNA was primed by the SMART-Seq v4 oligonucleotide and then base-pairs with these additional nucleotides, creating an extended template. The reverse transcriptase then switches templates and continues transcribing to the end of the oligonucleotide, resulting full-length cDNA that contains an anchor sequence that serves as a universal priming site for second strand synthesis. The resulting cDNA was amplified with primer II A for 8 PCR cycles. Then Illumina sequencing libraries were generated with 150 pg of cDNA using Illumina Nextera DNA Sample Preparation Kit (Cat#: FC-131-1024) according to manufacturer’s instructions. Briefly, 150 pg of cDNA was fragmented by tagmentation reaction and then adapter sequences added onto template cDNA by PCR amplification. Libraries were quantitated by Bioanalyzer and qPCR (Kapa Biosystems, catalog number: KK4824). Finally, the libraries were pooled equal molar concentration and sequenced on an Illumina HiSeq 3000 using 2x101 cycles. Bisulfite-converted libraries were sequenced alongside RNA-seq libraries (60%:40% ratio of RNA-seq vs Bisulfite-converted libraries), without a PhiX spike-in. Library pools were processed according the Illumina protocol (HiSeq3000) for clustering on the cBOT machine. The final pool concentration for clustering was 0.25 nM, following denaturation, neutralization, and mixing with the ExAmp reagent. A typical sequencing run in the HiSeq3000 produced 600-650 million paired-end reads per lane, 60% of which (~360 million paired-end reads) went to RNAseq.


title: Spaceflight, Root Rep. 3

status: Public on Mar 20 2019

type: SRA

city: Gainesville

first: Robert

middle: J.

last: Ferl

state: Florida

address: 1301 Fifield Hall PO Box 110690

department: Horticultural Sciences

country: USA

institute: University of Florida

description: ROOTFT.vs.ROOTGC diff exp

library_source: transcriptomic

molecule: total RNA

organism: Arabidopsis thaliana

source_name: solid 0.5x MS media plates; grown on the ISS, 11 day old plant harvested into KFT containing RNAlater fixative

treatment_protocol: The two treatments involved in this experiment were spaceflight and ground control. Spaceflight plants were grown for 11 days aboard the ISS in the VPS hardware. Ground control plants were grown for 11 days in the VPS hardware in the ISSES chamber at KSC, which replicates the temperature, CO2 levels and lighting that had been experienced by the spaceflight plants in the previous 48h.

tag: cultivar

value: WS

tag: material type

value: Leaves

tag: organism part

tag: age

value: 11 days old plants

tag: biosource type

value: RNAlater fixed_sample


title: Ground Control, Leaf Rep. 2

description: LEAFFT.vs.LEAFGC diff exp

source_name: solid 0.5x MS media plates; grown on the ground, 11 day old plant harvested into KFT containing RNAlater fixative

value: Roots

value: Whole roots


title: Spaceflight, Root Rep. 2

description: ROOTFT.vs.ROOTGC diff exp


title: Spaceflight, Leaf Rep. 1

description: LEAFFT.vs.LEAFGC diff exp


title: Spaceflight, Root Rep. 1

description: ROOTFT.vs.ROOTGC diff exp


title: Spaceflight, Leaf Rep. 2

description: LEAFFT.vs.LEAFGC diff exp


title: Spaceflight, Leaf Rep. 3

description: LEAFFT.vs.LEAFGC diff exp


title: Ground Control, Leaf Rep. 3

description: LEAFFT.vs.LEAFGC diff exp


title: C1

city: HANGZHOU

first: Chenwen

last: Xiao

address: Shiqiao Road 145 JiangGan District

department: Institute of Animal Husbandry and Veterinary Science

country: China

institute: Zhejiang Academy of Agricultural Sciences

data_processing:
Illumina Casava1.8 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence by fastx_toolkit-0.0.13.2, then mapped to whole genome using SOAP with parameters: maximum number of mismatches allowed = 2bp， number of valid alignments per read pair (-k) = 40
Fragments Per Kilobase of exon model per Million mapped reads (FPKM) were calculated.
Genome_build: reference sequence used for read alignment
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

organism: Trichophyton mentagrophytes

source_name: Fungal

growth_protocol: After 6 h incubation (28°C,150 rpm)

extract_protocol:
Tissue samples were stored at -80°C prior to use, and RNA was harvested using Trizol reagent. 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: The BC treated group (three samples) was subject to medicinal stress dose which means 1/2 MIC (minimum inhibitory concentration) dose (1 mg/mL berberine in DMSO), Clotrimazole (CLO) (99 % pure, Lot No. 23593-75-1) was purchased from BaDaTong Medical Company (TaiZhou, Zhejiang Province, China). And clotrimazole treated group (three samples) was subject to medicinal stress dose which means 1/2 MIC (minimum inhibitory concentration) dose (0.5 μg/mL Clotrimazole in DMSO),while the control group was fed with DMSO (Purity>99%)

tag: agent

value: DMSO


title: Ground Control, Root Rep. 1

description: ROOTFT.vs.ROOTGC diff exp


title: K3

value: Clotrimazole


title: K2


title: C3


title: Ground Control, Root Rep. 2

description: ROOTFT.vs.ROOTGC diff exp


title: C2


title: K1


title: Ground Control, Leaf Rep. 3 [BS]

description:
bisulfite converted
LEAFFT.vs.LEAFGC.full CG
LEAFFT.vs.LEAFGC.full CHG
LEAFFT.vs.LEAFGC.full CHH
LEAFGC-winavg CG
LEAFGC-winavg CHG
LEAFGC-winavg CHH
LEAFFT.vs.LEAFGC.genes01.full CG
LEAFFT.vs.LEAFGC.genes01.full CHG
LEAFFT.vs.LEAFGC.genes01.full CHH
LEAFFT.vs.LEAFGC.genes02.full CG
LEAFFT.vs.LEAFGC.genes02.full CHG
LEAFFT.vs.LEAFGC.genes02.full CHH
LEAFFT.vs.LEAFGC.genes03.full CG
LEAFFT.vs.LEAFGC.genes03.full CHG
LEAFFT.vs.LEAFGC.genes03.full CHH
LEAFFT.vs.LEAFGC.genes04.full CG
LEAFFT.vs.LEAFGC.genes04.full CHG
LEAFFT.vs.LEAFGC.genes04.full CHH
LEAFFT.vs.LEAFGC.genes05.full CG
LEAFFT.vs.LEAFGC.genes05.full CHG
LEAFFT.vs.LEAFGC.genes05.full CHH

library_source: genomic

data_processing:
The fastq files containing short reads from whole-genome bisulfite sequencing were trimmed to remove low-quality bases using trimmomatic (version 0.36). Fastq files before and after trimming were analyzed for quality control with FastQC (version 0.11.4). The fraction of reads removed because of low quality was never more than 0.01%.  Total number of reads after trimming for each sample ranged from 22,446,135 to 39,065,215.
A second filtering step was performed to remove reads showing evidence of incomplete conversion, using the cscall tool (http://compbio.ufl.edu/software/cscall/). The fraction of reads removed in this step ranged between 0.05% and 0.08%.
The remaining reads were aligned to the Arabidopsis genome using bsmap (http://bmcbioinformatics.biomedcentral.com/articles/10.1186/1471-2105-10-232) version 2.87. The percentage of aligned reads ranged from 87.57% and 90.52%.
Methylation calling was performed with cscall using the ws_0.v7.allPlusChlMito-CG index. A site was included in the analysis if it reached a coverage of 20 in at least 2 out of the three replicates of each sample. Differential methylation was determined using the mcomp program, part of the MOABS suite (https://genomebiology.biomedcentral.com/articles/10.1186/gb-2014-15-2-r38), version 1.3.4. Sites for which the P-value of the difference between test and control methylation rates was below 0.01 were considered significant. Sites with a methylation difference above 0.3 were classified as Strongly hyper/hypo methylated(++ or  --), otherwise they were classified as hyper/hypo methylated (+ or -).
Genome_build: Arabidopsis thaliana WS reference genome (Mott et al. Nature 2011)
Supplementary_files_format_and_content: .xls spreadsheets containing methylation levels in a 100 bp sliding window across the genome and differential methylation between the spaceflight vs. ground control comparisons.

molecule: genomic DNA

growth_protocol: Dry, sterilized Arabidopsis seeds of wild-type cultivar Wassilewskija  were planted aseptically on the surface of 10 cm2 solid media plates comprised of 0.5% Phytagel/0.5× MS media, and then immediately wrapped in light-tight black cloth (Duvetyne, SeattleFabrics.com) and transported to KSC (Kennedy Space Center), FL, USA. The wrapped plates were stored at temperatures between 4 and 10°C and remained dormant until removal from cold stowage and exposure to light at the initiation of the APEX03-2 experiment on the ISS (International Space Station). Plates were integrated and grown in the Vegetable Production System (VPS/Veggie) hardware on the Columbus Module of the ISS. The 10 cm2 Petri plates were housed perpendicular (“vertical”) to the overhead LED lighting of the VPS. Plants received between 100–135 μmoles/m2/s PAR inside the VPS. The plants were grown for eleven days after germination, which was initiated by unwrapping the plates and exposing them to light in the VPS. The ground control plates were grown inside the VPS ground unit within the ISSES (International Space Station Environment Simulator) chamber at KSC with a 48-h delay. The ISSES chamber replicated the temperature, CO2 levels and lighting that had been experienced by the spaceflight plants in the previous 48 h. After eleven days, seedlings were harvested by an astronaut into a KFT (Kennedy Fixation Tube) containing RNAlater. Once the plants were placed in the KFTs, the KFT was actuated with RNAlater to chemically fix the sample. At 24 hours post-harvest, KFTs were then transferred to MELFI (Minus Eight Laboratory Freezer for ISS) and were kept at -32°C. Following SpaceX-3 splashdown in the Pacific Ocean, the KFTs were transferred to the Cold Stowage charter plane at the Long Beach Airport, placed into an insulated shipper with dry ice, and flown to Johnson Space Center (JSC). The KFTs were then transferred via FedEx ground to the Kennedy Space Center. The KFTs were removed from dry ice and transferred to a -80°C freezer on May 22, 2014. The PIs retrieved the plant samples from the KFTs on May 29, 2014 and transferred the samples back to the University of Florida.

extract_protocol:
Seedlings stored in RNAlater were recovered from the -80°C freezer. Each tube containing the seedlings from a single plate was allowed to thaw at 4°C overnight. After completely thawing the samples to room temperature, the seedlings were observed under the microscope. The preserved seedlings from the spaceflight and ground control harvests were dissected into distinct plant parts: leaf, hypocotyl, and root. Three plants were allocated for RNA analysis and the remaining plants from a plate (between 10 and 12) were allocated for DNA extraction and subsequent whole genome bisulfite sequencing. DNA was extracted using a modified phenol/chloroform protocol as previously described (LeFrois et al. 2016) from three biological replicates of root and leaf samples from spaceflight and ground controls. This DNA material was then submitted to the Interdisciplinary Center for Biotechnology at the University of Florida for bisulfite conversion and Next-Generation sequencing.
Genomic-wide bisulfite sequencing was performed using a similar procedure as described by Wang et al. (2013) with several modifications. In summary, genomic DNA was extracted from leaves and roots of spaceflight and ground control samples using a modified phenol/chloroform extraction method (LeFrois et al. 2016). Between 700-1700 ng of genomic DNA was processed for sequencing library construction. The average size of the genomic DNA for this type of samples was 5 Kb as observed on a DNA12000 BioAnalyzer chip (Agilent). DNA was transferred into 6x16 mm glass microtubes with AFA fiber and pre-slit snap caps (Cat# 520045, Covaris, Inc.). As it is required by the Illumina sequencing procedure, the DNA was sheared into average fragments size of ~400 bp, using the Covaris S220 ultrasonic disruptor, and following the recommended settings by the manufacturer. Short DNA fragments (<100 bp) were removed using AMPure magnetic beads (Cat# A63881, Beckman Coulter) at a 1:1 bead to sample ratio.  At this point we obtained a yield of 300-750 ng of clean, fragmented DNA. Between 100-250 ng of this material was used for Illumina sequencing library construction using the NEBnext Ultra DNA Illumina construction kit (Cat# E7370S, NEB) in conjunction with Illumina-specific methylated and barcoded adaptors (Cat# E7535S, NEB) from the same source. This particular library construction kit was chosen because it contains reagents that have been optimized for relatively low-input DNA in the library construction process. At the end of the end-repair, 3'-adenylation, and adaptor-ligation steps, 75-150 ng of clean, adaptor-ligated DNA fragments (Illumina library) were recovered in Tris-HCl, pH 8.0. It should be noted that the libraries were not PCR-amplified at this stage, as it would normally be done in the most typical Illumina library construction protocol. Libraries were quantitated using a QUBIT fluorometer (Life Technologies).Illumina libraries (containing methylated adaptors) were subjected to sodium bisulfite treatment using the EZ DNA Methylation Direct kit (ZYMO Research, Cat #D5020) according to the manufacturer's instructions. The resulting libraries were enriched by a 13-15 cycle amplification using an uracil-insensitive polymerase (EpiMark hot start Taq polymerase, NEB, Cat#M0490S). The amplified library products were separated on a 2% agarose gel from which library fragments in the 250-500 bp range were excised (QIAquick gel extraction kit, Cat# 28704, QIAGEN) and AMPure purified (Cat# A63881, Beckman Coulter). Gel staining was done with SYBR safe (Life Technologies) and visualized on a blue light transilluminator (Life Technologies) in order to avoid UV damage of DNA. The final libraries were quantitated by the QUBIT fluorometer, and by qPCR with the Kapa SYBR Fast qPCR reagents (Cat# KK4824, Kapa Biosystems) with monitoring on an ABI7900HT real-time PCR system (LifeTechnologies). The average library size was 350 bp. In preparation for sequencing on the HiSeq3000 Illumina sequencer, barcoded libraries were normalized to 2.5 nM and pooled (equimolarly). Up to 6 samples were pooled for sequencing on a single HiSeq3000 flow cell lane, using a 2x101 cycle multiplex, paired-end configuration. Typically, the PhiX library is used for a spike-in (at 20%) in order to correct for the low base diversity of bisulfite-converted genomic libraries. Alternatively, bisulfite-converted libraries were sequenced alongside RNA-seq libraries (60%:40% ratio of RNA-seq vs Bisulfite-converted libraries), without a PhiX spike-in. Library pools were processed according the Illumina protocol (HiSeq3000) for clustering on the cBOT machine. The final pool concentration for clustering was 0.25 nM, following denaturation, neutralization, and mixing with the ExAmp reagent. A typical sequencing run in the HiSeq3000 produced 600-650 million paired-end reads per lane. For genome-wide bisulfite sequencing 50-100 million reads provided sufficient depth for analysis of methylation. In Arabidopsis (and possibly other plants), this output is required in part due to the fact that up to 50% of the reads map to chloroplast DNA. Assuming the lowest end of reads/lane output in the HiSeq3000, a 36-sample project would need 6 lanes.


title: Ground Control, Root Rep. 3

description: ROOTFT.vs.ROOTGC diff exp


title: X2

value: Berberine


title: X1


title: X3


title: Input_of_MYOD_ChIPSeq_IMR90_MYOD_GM

status: Public on Mar 19 2019

city: Cambridge

first: Alessandra

last: Dall'Agnese

state: MA

address: 455 Main Street

institute: Whitehead Institute

description: ChIPSeq lysates were clarified from sonicated crosslinked nuclei and MYOD-DNA complexes were isolated with antibody for ChIP sample or not for input sample. Libraries were prepared as in http://bioinformatics-renlab.ucsd.edu/RenLabLibraryProtocolV1.pdf

data_processing:
ChIP-Seq - Read quality was determined using FASTQC. Reads were mapped using bowtie2-2.0.5/bowtie2 to the female Homo sapiens hg19 genomes with options: --very-sensitive-local. Peaks were called using macs2 2.1.1.20160309 with qvalue=0.01, macs2 2.1.1.20160309 was also used for differential peak calling among samples.
RNA-Seq - Spike-in were added based on number of nuclei but were not used for the analysis. Read quality was determined using FASTQC. Reads were mapped using TopHat2.1.1 to the female Homo sapiens hg19 genomes with options: -p 8 -g 1 --segment-length 17 --library-type fr-firststrand. HTSeq-0.6.1p1173 with --stranded=reverse option was used to assigned mapped reads to Homo Sapiens GRCh37.75 genes. Differential expression analysis was performed using DESeq2.
Genome_build: hg19

organism: Homo sapiens

growth_protocol: IMR90 cells were cultured in EMEM supplemented with 10% FBS

extract_protocol:
For RNASeq: cells were lyzed with Trizol and RNA was ectracted following manufacturer's instruction. For ChIPSeq lysates were clarified from sonicated crosslinked nuclei and MYOD-DNA complexes were isolated with antibody.
RNASeq libraries were prepared using TruSeq Stranded mRNA Library Prep Kit set A RS-122-2101 following Illumina's recomandations . ChIPSeq libraries were prepared as in http://bioinformatics-renlab.ucsd.edu/RenLabLibraryProtocolV1.pdf.

treatment_protocol: IMR90 cells were transfected with vector containing TET-inducible MYOD or EMPTY vector control, cells were terated with 200ng/ml doxycycline for 24hrs in EMEM 10% FBS and collected for GM time point. Cells were terated with 200ng/ml doxycycline for 24hrs in EMEM 10% FBS and for 3days in EMEM 2% HS 1%ITS collected for DM time point.

tag: cell type

value: Lung fibroblast-derived myoblast

tag: antibody

tag: ectopic gene expression

value: mouse MYOD overexpression

tag: passage from isolation

value: passage 21-28


title: Input_of_MYOD_ChIPSeq_IMR90_MYOD_DM

value: Lung fibroblast-derived myotube


title: MYOD_ChIPSeq_IMR90_MYOD_DM

value: MYOD, Santa Cruz, M-318 sc-760


title: MYOD_ChIPSeq_IMR90_MYOD_GM

description:
ChIPSeq lysates were clarified from sonicated crosslinked nuclei and MYOD-DNA complexes were isolated with antibody for ChIP sample or not for input sample. ChIPSeq libraries were prepared as in http://bioinformatics-renlab.ucsd.edu/RenLabLibraryProtocolV1.pdf
diff_M_GM_vs_M_DM_c3.0_common.bed


title: Ube3a_TG_HFD_1

status: Public on Mar 18 2019

city: Gwangju

first: JANGHYUN

last: KIM

state: Gwangju

address: 33, YONGBONG-RO, BUK-GU, GWANGJU

department: Oral pathology

country: South Korea

institute: CNUDH

description:
Ube3a TG
Ube3a TG homo samples overexpress Ube3a.
Processed data file: WT_Ube3a_TG_count_file.txt

data_processing:
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm10 whole genome using subread v1.5.2.
Either the feature or meta-feature levels were counted using featureCounts and DESeq2 was used to identify differentially expressed genes between WT and Ube3a mutants liver.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: *txt: Tab-delimited text files include RPKM values for each Sample.

organism: Mus musculus

source_name: liver, Ube3a TG homo, HFD

growth_protocol: All mouse works were performed under an approved protocol by the Institutional Animal Care and Use Committee of the Oregon Health & Science University. Mice were housed in a pathogen-free animal facility under standard 12h light/12h dark cycle.

extract_protocol:
Livers were removed and RNA was harvested using Trizol reagent.
Illumina TruSeq RNA Sample Prep Kit was used with 1 ug of total RNA for the construction of sequencing libraries.

treatment_protocol: 42% high fat diet feeding for 8 weeks.

tag: strain/background

tag: genotype/variation

value: Ube3a TG homo

tag: tissue

value: liver

tag: treatment

value: high-fat diet


title: Ube3a_TG_HFD_2


title: WT_HFD_3

description:
Wild type (control of Ube3a het).
Processed data file: WT_Ube3a_het_count_file.txt (WT-1)

source_name: liver, WT, HFD

value: WT


title: Ube3a_het_HFD_1

description:
Ube3a het
Ube3a het samples have one knockout and one wt allele, so they express half the amount of Ube3a.
Processed data file: WT_Ube3a_het_count_file.txt

source_name: liver, Ube3a het, HFD

value: Ube3a het


title: Ube3a_het_HFD_2


title: WT_HFD_4

description:
Wild type (control of Ube3a het).
Processed data file: WT_Ube3a_het_count_file.txt (WT-2)


title: PrimaryMyoblast__Cont__polyA.rep3

city: Higashi-Osaka

first: Kei

last: IIDA

state: Osaka

address: Kowakae 3-4-1

country: Japan

institute: Kindai University

description: mRNA-seq (polyA+) data from primary myoblasts obtained from skeletal muscles of 1-month-old control mice under differentiated condition, replicate 3

data_processing:
Basecalls performed using Torrent Suite version 5.0.2
The seuqence data was mapped to the genome sequence with STAR ver. 2.4.1d .
Genome_build: mm10
Supplementary_files_format_and_content: Expression values of refseq gene models were calculated with an in-house perl script.  We counted read counts mapped on exons, then normalized them with library sizes and sum of exon lengths (RPKM; Reads Per Kilobase of exon Model per million mapped reads). In the processed data files, gene ID and RPKM values were described column 1 and 2, respectively.

source_name: primary myoblast

growth_protocol: Primary myoblasts were expanded in F-10 containing 20% fetal bovine serum, 1% penicillin-streptomycin, and 2.5 ng/mL basic fibroblast growth factor.

extract_protocol:
Total RNA was extracted from differentiated primary myoblasts using the RNeasy mini kit (QIAGEN).
Ion Total RNA-Seq Kit (Life Technology)

treatment_protocol: Induced differentiation in 2% horse serum in Dulbecco’s modified Eagle medium and subjected to sample preparation after 4 days.

tag: condition

value: differentiated condition

tag: derived tissue

value: skeletal muscle (1-month-old male mice)


title: PrimaryMyoblast__Cont__polyA.rep2

description: mRNA-seq (polyA+) data from primary myoblasts obtained from skeletal muscles of 1-month-old control mice under differentiated condition, replicate 2


title: PrimaryMyoblast__SFPQ_KO__polyA.rep2

description: mRNA-seq (polyA+) data from primary myoblasts obtained from skeletal muscles of 1-month-old SfpqSM-KO mice under differentiated condition, replicate 2


title: PrimaryMyoblast__SFPQ_KO__polyA.rep1

description: mRNA-seq (polyA+) data from primary myoblasts obtained from skeletal muscles of 1-month-old SfpqSM-KO mice under differentiated condition, replicate 1


title: PrimaryMyoblast__Cont__polyA.rep1

description: mRNA-seq (polyA+) data from primary myoblasts obtained from skeletal muscles of 1-month-old control mice under differentiated condition, replicate 1


title: PrimaryMyoblast__SFPQ_KO__rRNAdep.rep1

description: mRNA-seq (polyA+/-) data from primary myoblasts obtained from skeletal muscles of 1-month-old SfpqSM-KO mice under differentiated condition.


title: PrimaryMyoblast__Cont__rRNAdep.rep1

description: mRNA-seq (polyA+/-) data from primary myoblasts obtained from skeletal muscles of 1-month-old control mice under differentiated condition.


title: NPC-TW01 biological rep3

type: RNA

city: New Taipei City

first: Yu-Lun

last: Kuo

address: 7F-3, No.99, Sec. 1, Xintai 5th Rd., Xizhi Dist.

country: Taiwan

institute: BIOTOOLS

description: control

hyb_protocol: Following fragmentation, 15 ug of cRNA were hybridized for 16 hr at 45C on GeneChip Affymetrix HG U133 plus 2.0 probe arrays. GeneChips were washed and stained in the Affymetrix Fluidics Station.

scan_protocol: GeneChips were scanned using an Affymetrix 7G Gene Chip Scanner

data_processing: The raw fluorescence intensity data within CEL files of these datasets were preprocessed and normalized using the GC-RMA algorithm

label: biotin

source_name: cell line derived from biopsy specimens of nasopharyngeal carcinoma patient

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 1 ug total RNA

growth_protocol: The NPC cell line was maintained in DMEM with 10% FBS at 37°C under 5% CO2.

extract_protocol: Total RNA was extracted using an illustra RNAspin Mini RNA isolation kit (GE Healthcare, Little Chalfont, UK) according to the manufacturer’s instructions.

treatment_protocol: NPC cells treated with or without 200 nM PDPN siRNA for 48 hours at 37°C

value: NPC cells

tag: passage


title: WT_HFD_1

description:
Wild type (control of Ube3a TG).
Processed data file: WT_Ube3a_TG_count_file.txt (WT-1)


title: WT_HFD_2

description:
Wild type (control of Ube3a TG).
Processed data file: WT_Ube3a_TG_count_file.txt (WT-2)


title: NPC-TW01 biological rep2

description: control


title: NPC-TW01 biological rep1

description: control


title: NPC-TW01 PDPN_200nM_48hr, biological rep 2

description: gene expression after 48hours 200nM PDPN siRNA treatment in NPC cell line

source_name: cell line derived from biopsy specimens of nasopharyngeal carcinoma patient and treated with 4 uM thiostrepton 12 hr

value: 200 nM PDPN siRNA


title: NPC-TW01 PDPN_200nM_48hr, biological rep 1


title: NPAI-C2 (57533)

city: Ann Arbor

first: Richard

middle: C

last: McEachin

state: United States

address: 2800 Plymouth Road

department: DCM&B

institute: University of Michigan

description:
NRAS activated; TP53 knockdown; ATR knockdown; IDH1-R132H expression
Low Grade Glioma model

data_processing:
[QC] We checked the quality of the raw reads data for each sample using FastQC (version 0.11.3) to identify features of the data that may indicate quality problems (e.g. low quality scores, over-represented sequences, inappropriate GC content, etc.).
[Aligment] We used the software package Tuxedo Suite for alignment, differential expression analysis, and post-analysis diagnostics. Briefly, we aligned reads to the reference transcriptome including both mRNAs and lncRNSs (UCSC mm10) using TopHat (version 2.0.3) and Bowtie (version 2.2.1). We used default parameter settings for alignment, with the exception of: “--b2-very-sensitive” telling the software to spend extra time searching for valid alignments.
[QC] We used FastQC for a second round of quality control (post-alignment), to ensure that only high quality data would be input to expression quantitation and differential expression analysis.
[Normalization and Quantitation] We used Cufflinks/CuffDiff (version 2.2.1) for expression quantitation, normalization, and differential expression analysis, using UCSC mm10.fa as the reference genome sequence. For this analysis, we used parameter settings: “--multi-read-correct” to adjust expression calculations for reads that map in more than one locus, as well as “--compatible-hits-norm” and “--upper-quartile –norm” for normalization of expression values. We generated diagnostic plots using the CummeRbund package.
[Differential Expression] We used locally developed scripts to format and annotate the differential expression data output from CuffDiff. Briefly, we identified genes and transcripts as being differentially expressed based on three criteria: test status = “OK”, FDR < 0.05 and fold change ≥ ± 1.5.
[Annotation] We annotated genes and isoforms with NCBI Entrez GeneIDs and text descriptions. We further annotated differentially expressed genes with Gene Ontology (GO) terms using NCBI annotation.
[Enrichment Testing] We used DAVID (version 6.7) for enrichment analysis of the set of differentially expressed genes to identify significantly enriched functional categories.
Genome_build: MM10
Supplementary_files_format_and_content: matrix of mapped read counts for each gene for each sample

source_name: Isolated tumor neurospheres

growth_protocol: Tumor neurospheres were grown in Neuro Stem cell media (RPMI media + B27 and N2 supplement, normacin 1X and Anti-Anti 1X)

extract_protocol:
RNA was harvested using the RNEasy-Plus Kit (Qiagen;74136) and assessed for quality using the TapeStation (Agilent). Samples with RNA Integrity Number of 8 or greater were rRNA depleted using
Ribo Minus (Invitrogen;K1550-04).
rRNA-depleted samples were prepared using the TruSeq mRNA SamplePrep v2 kit (Illumina;RS-122-2001, RS-122-2002). The entire fraction of 0.1-3 ug of rRNA depleted total RNA was fragmented and
copied into first-strand cDNA using reverse transcriptase and random primers. cDNA 3‘-ends were adenylated and adapters were ligated. One of the adapters had a 6-nucleotide barcode that was unique
for each sample, allowing us to multiplex in a HiSeq flow cell (Illumina). Products were purified and enriched by PCR to create the final cDNA library. Libraries were checked for quality and quantity by TapeStation
(Agilent) and qPCR using a library quantification kit for Illumina platforms (Kapa Biosystems;KK4835). They were clustered on the cBot (illumina) and all 9 samples were sequenced on each of 2 llanes, then the reads files for each sample were concatenated.
Sequencing was 50-cycle, single end, on the Illumina HiSeq 2500 High-Output sequencer, using V4 chemistry

value: Tumor neurospheres

tag: genotype

value: NRAS-V12; shP53; shATRX; IDH1-R132H


title: NPA-C54B (57531)

description:
NRAS activated; TP53 knockdown; ATR knockdown
Pediatric Glioma model

value: NRAS-V12; shP53; shATRX


title: NPAI-C1 (57532)


title: NPAI-C3 (57534)


title: PrimaryMyoblast__SFPQ_KO__polyA.rep3

description: mRNA-seq (polyA+) data from primary myoblasts obtained from skeletal muscles of 1-month-old SfpqSM-KO mice under differentiated condition, replicate 3


title: NP-C4 (57528)

description:
NRAS activated and TP53 knockdown
Glioma cells model

value: NRAS-V12; shP53


title: NP-C3 (57527)


title: NP-C1 (57526)


title: Sample_deg_4hrs

city: London

first: Tessa

last: Crompton

address: 30 Guilford Street

department: Immunobiology

country: United Kingdom

institute: UCL

description: Deg 4hours

data_processing:
Base Calling was done using the RTA package from Illumina
Alignment was done on the transcriptome using the Burrows-Wheeler Aligner (BWA) package
Expression levels in Reads Per Million Kilobases (RPKM).
Genome_build: GRCm38
Supplementary_files_format_and_content: comma separated file including RPKM values for each Sample.

source_name: Murine Adult Spleen CD4 T Cells

extract_protocol:
Murine CD4 positive cells were isolated from spleen cells by magnetic bead separation using EasySep mouse CD4+ cell negative selection kit (StemCell Technologies) according to the manufacturer’s instructions. The purity of isolated CD4+ cells was routinely exceeded 95%. For activation, CD4+cells were cultured at 5x106 cells/ml coated with 5µg/ml anti-CD3, and added 1µg/ml soluble anti-CD28 in complete RPMI and analysed at 4 and 20h. Th0, Th1 and Th2 skewing cultures, CD4+ cells were cultured on 5ug/ml plate bound anti-CD3 with were culture with 1µg/ml soluble anti-CD28 in complete RPMI. Th1 conditions the solution containing 5µg/ml anti-IL-4 and 10ng/ml rmIL-12. For Th2 cultures, 5µg/ml anti-IFNɣ; 20ng/ml rmIL-4, and 5µg/ml anti-IL-12 were added and, Cells were cultured at 37 °C under 5% CO2. Cells were harvested 72h and T-bet and Gata-3 were analysed through intracellular staining assay as described (Furmanski et al. 2013) (antibodies/proteins: eBioscience).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: CD4 Th cells were purified and set up in culture under three different Th differentiation conditions: Th0 condition was set up by treating the purified CD4 cells with anti-cd3e and anti-cd28; Th1 condition was set up with anti-cd3e, anti-cd28, anti-IL-4 and recombinant IL-12; and Th2 condition was set up with anti-cd3e, anti-cd28, anti-IL-12, anti-IFNG and recombinant IL-4.

tag: strain

tag: developmental stage

value: Murine Adult CD4 T cells cultured in vitro with Actinomysin

tag: culture condition

value: Stimulated for 16 hours and than Actinomysin added

value: Wildtype


title: Sample_deg_2hrs

description: Deg 2hours


title: Sample_Th2_24hrs_0612

value: Murine Adult CD4 T cells cultured in vitro

value: Stimulated under Th2 for 24 hours


title: Sample_deg_0hrs

description: Deg 0hours


title: Sample_Th2_12hrs_0612

value: Stimulated under Th2 for 12 hours


title: Sample_Th1_16hrs_0612

value: Stimulated under Th1 for 16 hours


title: Sample_Th1_12hrs_0612

value: Stimulated under Th1 for 12 hours


title: Sample_Th2_4hrs_0612

value: Stimulated under Th2 for 4 hours


title: Sample_Th1_24hrs_0612

value: Stimulated under Th1 for 24 hours


title: Sample_Th1_20hrs_0612

value: Stimulated under Th1 for 20 hours


title: Sample_Th1_8hrs_0612

value: Stimulated under Th1 for 8 hours


title: Sample_Th2_8hrs_0612

value: Stimulated under Th2 for 8 hours


title: Sample_deg_1hrs

description: Deg 1hour


title: NPC-TW01 PDPN_200nM_48hr, biological rep 3

value: NPC cells

value: 200 nM PDPN siRNA


title: Sample_Th1_4hrs_0612

value: Stimulated under Th1 for 4 hours


title: Sample_Th0_24hrs_0612

value: Stimulated under Th0 for 24 hours


title: NPA-C54A (57530)

value: Tumor neurospheres

value: NRAS-V12; shP53; shATRX


title: NPA-C2 (57529)


title: Sample_Th0_20hrs_0612

value: Stimulated under Th0 for 20 hours

value: Wildtype


title: Sample_Th0_16hrs_0612

value: Murine Adult CD4 T cells cultured in vitro

value: Stimulated under Th0 for 16 hours


title: Sample_Th0_12hrs_0612

value: Stimulated under Th0 for 12 hours


title: NPAI_c2_H3K4me3

first: Maria

middle: G

last: Castro

state: Michigan

address: 1150 W. Medical Center Drive MSRB II, Room 4570C

department: Dept. of Neurosurgery and Dep. of Cell and Developmental Biology

data_processing:
The data quality was accessed using FastQC, and the first 2 bases were trimmed out due to the low read quality. Reads were aligned to the mouse reference genome (mm10) using bowtie2 with default options, and the unique mapped reads were extracted. PePr (v1.1.14) was used to identify the differential H3K4me3 or H3K27me3 peaks between NPA and NPAI groups using p-value < 1e-5 as the cutoff. The H3K4me3 peaks were set as narrow peaks, while H3K27me3 peaks were set as broad peaks.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig

source_name: Mouse glioma neurospheres

growth_protocol: Tumor glioma neurospheres (NS) wt-IDH1 and mIDH1 NS were generated from genetically engineered mouse brain tumors. NS were cultured with DMEM-F12 media supplemented with N2 and B27 and 10 ng/ml of both EGF and FGF (NS expansion medium).

extract_protocol:
Tumor NS were collected and suspended in digestion buffer.  Digestion was performed using microccocal nuclease to obtain 150 pb DNA fragments. Histone-DNA complexes were isolated with the desired H3 me3 specific antibody.
Libraries for ChIP-seq were prepared using the TruSeq ChIP Sample Preparation Kit. Input and IP DNA  was blunt-ended and phosphorylated, followed by the addition of a single adenine nucleotide prior to ligation with an adapter duplex with a T overhang. Ligated products were size selected by agarose gel electrophoresis, purified and PCR amplified for the final library preparation.

value: NRAS activation; shp53 (inactivation); shATRX (inactivation); IDH1-R132H expression

value: glioma neurospheres

value: 5 to 8


title: NPAI_c3_H3K27me3


title: NPAI_c3_input


title: NPAI_c2_input


title: NPAI_c2_H3K27me3


title: NPAI_c3_H3K4me3


title: Sample_Th2_20hrs_0612

value: Stimulated under Th2 for 20 hours


title: Sample_Th2_16hrs_0612

value: Stimulated under Th2 for 16 hours


title: Sample_Th0_8hrs_0612

value: Stimulated under Th0 for 8 hours


title: NPAI_c1_H3K4me3


title: Sample_Th0_4hrs_0612

value: Stimulated under Th0 for 4 hours


title: Sample_0hrs_0611_B

description: 0hours Rep2

value: Murine Adult Unstimulated CD4 T cells

value: Purified Unstimulated CD4 T cells


title: Sample_0hrs_0611_A

description: 0hours Rep1


title: NPA_c54a_input

value: NRAS activation; shp53 (inactivation) and shATRX (inactivation)


title: NPAI_c1_H3K27me3


title: NPA_c54b_input


title: NPA_C2_input


title: NPA_c54a_H3K4me3


title: NPA_c54a_H3K27me3


title: NPA_C2_H3K4me3


title: NPAI_c1_input


title: NPAI-IR (71578)

description:
NRAS activated; TP53 knockdown; ATR knockdown; IDH1-R132H expression
Glioma implanted mouse model

source_name: Implanted glioma brain tumor

growth_protocol: Brain tumors were generated in BL6 mice by implantation of 50.000 tumor neurospheres expression IDH1-R132H or control WT.

extract_protocol:
RNA was harvested using the RNEasy-Plus Kit (Qiagen;74136) and assessed for quality using the TapeStation (Agilent). Samples with RNA Integrity Number of 8 or greater were rRNA depleted using
Ribo Minus (Invitrogen;K1550-04).
rRNA-depleted samples were prepared using the TruSeq mRNA SamplePrep v2 kit (Illumina;RS-122-2001, RS-122-2002). The entire fraction of 0.1-3 ug of rRNA depleted total RNA was fragmented and
copied into first-strand cDNA using reverse transcriptase and random primers. cDNA 3‘-ends were adenylated and adapters were ligated. One of the adapters had a 6-nucleotide barcode that was unique
for each sample, allowing us to multiplex in a HiSeq flow cell (Illumina). Products were purified and enriched by PCR to create the final cDNA library. Libraries were checked for quality and quantity by TapeStation
(Agilent) and qPCR using a library quantification kit for Illumina platforms (Kapa Biosystems;KK4835). They were clustered on the cBot (illumina) and sequenced 24 samples per lane on 6 lanes of a 50-cycle
single end HiSeq 4000. HiSeq Control Software version 3.3.52 was used according to manufacturer’s protocols. Demultiplexing and Fastq file generation was done using bcl2fastq version 2.17.1.14.

treatment_protocol: After 7 days of cell implantation, the animals were treated with 2 Gy of ionizing radiation (IR) per day during e days  for a total of 10 Gy (IR).

value: Glioma brain tumor

value: NRAS-V12; shP53; shATRX; IDH1-R132H

value: IR 10Gy


title: NPA_C2_H3K27me3


title: NPA_c54b_H3K27me3


title: NPAI-IR (71576)


title: NPAI-IR (71577)


title: NPA_c54b_H3K4me3


title: NPAI-IR (71575)


title: NPAI-control (71572)

value: Control


title: NPA-IR (71569)

description:
NRAS activated; TP53 knockdown; ATR knockdown
Glioma implanted mouse model

value: NRAS-V12; shP53; shATRX


title: NPA-IR (71570)


title: NPA-IR (71567)


title: NPA-control (71566)


title: NPA-IR (71568)


title: NPAI-control (71574)


title: NPA-control (71565)


title: NPA-control (71564)


title: NPA-control (71563)


title: siPFKFB3 RNA-seq

city: Stanford

first: Kazuya

last: Miyagawa

state: California

address: 269 Campus Drive

department: Pediatrics

institute: Stanford Universtiy

data_processing:
Trimmed using FASTQ groomer.
Aligned to the human genome using TopHat.
Gene transcripts were quantified using Cuffdiff
Genome_build: hg19
Supplementary_files_format_and_content: txt files were generated using Cuffdiff

source_name: Large pulmonary artery endothelial cells

growth_protocol: PASMCs suspended in Smooth Muscle Growth Medium-2 (Lonza, Indianapolis, IN) were seeded on the opposite side of an 1um pored cell culture insert and 2 h after seeding, the inserts were flipped and PAECs were seeded on the bottom of the inserts and cultivated in Endothelial Cell Medium (Sciencell, Carlsbad, CA). Co-cultures of PAECs with PAECs were established by using only EC medium.

extract_protocol:
Cells were trypsinized 48 h after co-culture, and PAECs were cross-linked with 1% formaldehyde (EMD Millipore), for 10 minutes at room temperature. To neutralize formaldehyde, 2M glycine (ThermoFisher Scientific) was added and incubated for 5 minutes at room temperature. The cells were washed with ice cold PBS twice, snap-frozen and stored at -80 °C. For ChIP-DNA preparation, cells were thawed by adding PBS and incubated at 4 °C by rotating. To extract cell nuclei, cells were treated with hypotonic buffer for 10 minutes on ice and then were homogenized using glass homogenizer. Nuclear pellets were resuspended in RIPA buffer (Millipore) and incubate for 30 minutes on ice. Chromatin was sheared with SFX250 Sonifier (Branson, Dunbury, CT) and was immunoprecipitated with Notch1 antibody, H3K27ac antibody (Cell Signaling Technology) and p300 antibody (Santa Cruz Biotechnology) at 4°C overnight on a nutator. To save input sample, 100 ul of sheared nuclear lysate was removed and store overnight at 4 °C. The second day, protein A/G agarose beads (Millipore) were added to the chromatin-antibody complex and incubate for 1 hr at 4°C on a nutator and then the beads were washed 3 times with RIPA buffer and 1 time with PBS. DNA-protein complex bound to the beads were eluted with SDS buffer (Santa Cruz Biotechnology) and incubated at 65 °C for 10 minutes. Supernatant containing ChIP-DNA and input sample were reverse cross-linked by incubating overnight at 65 °C. The third day, ChIP-DNA was treated with RNase A (Qiagen) and proteinase K (ThermoFisher Scientific) and then was purified.
RNA was extracted with RNeasy Mini Kit (Qiage), and libraries were prepared with ScriptSeq Complete Gold kit ChIP-DNA samples were end repaired and A-tailed. The illumina TruSeq adapters (illumina) were ligated and size-selected from the gel before doing PCR amplification. PCR products were purified and size selected in the gel again.

treatment_protocol: PAECs with or without siPFKFB3 transfection were contact co-cultured with PASMCs for 48 hours.

tag: passages


title: siControl RNA-seq

value: Large pulmonary artery endothelial cells


title: p300_ChIPSeq__input_siPFKFB3_Repl2

data_processing:
Trimmed using FASTQ groomer.
Aligned to the human genome using Bowtie2.
Peaks were called using MACS2.
MACS2 narrowPeak files from two biological replicates were submitted into Irreproducible Discovery Rate (IDR) pipeline.
Genome_build: hg19
Supplementary_files_format_and_content: txt files were generated using IDR pipeline.

extract_protocol:
Cells were trypsinized 48 h after co-culture, and PAECs were cross-linked with 1% formaldehyde (EMD Millipore), for 10 minutes at room temperature. To neutralize formaldehyde, 2M glycine (ThermoFisher Scientific) was added and incubated for 5 minutes at room temperature. The cells were washed with ice cold PBS twice, snap-frozen and stored at -80 °C. For ChIP-DNA preparation, cells were thawed by adding PBS and incubated at 4 °C by rotating. To extract cell nuclei, cells were treated with hypotonic buffer for 10 minutes on ice and then were homogenized using glass homogenizer. Nuclear pellets were resuspended in RIPA buffer (Millipore) and incubate for 30 minutes on ice. Chromatin was sheared with SFX250 Sonifier (Branson, Dunbury, CT) and was immunoprecipitated with Notch1 antibody, H3K27ac antibody (Cell Signaling Technology) and p300 antibody (Santa Cruz Biotechnology) at 4°C overnight on a nutator. To save input sample, 100 ul of sheared nuclear lysate was removed and store overnight at 4 °C. The second day, protein A/G agarose beads (Millipore) were added to the chromatin-antibody complex and incubate for 1 hr at 4°C on a nutator and then the beads were washed 3 times with RIPA buffer and 1 time with PBS. DNA-protein complex bound to the beads were eluted with SDS buffer (Santa Cruz Biotechnology) and incubated at 65 °C for 10 minutes. Supernatant containing ChIP-DNA and input sample were reverse cross-linked by incubating overnight at 65 °C. The third day, ChIP-DNA was treated with RNase A (Qiagen) and proteinase K (ThermoFisher Scientific) and then was purified.
The ChIP-DNA samples were end repaired and A-tailed. The illumina TruSeq adapters (illumina, San Diego, CA) were ligated and size-selected from the gel before doing PCR amplification. PCR products were purified and size selected in the gel again.

tag: chip antibody


title: p300_ChIPSeq__input_siPFKFB3_Repl1


title: p300_ChIPSeq_siControl_Repl1

value: p300 (Santa Cruz Biotechnology, sc-585)


title: p300_ChIPSeq__input_siControl_Repl2


title: p300_ChIPSeq_siPFKFB3_Repl1


title: p300_ChIPSeq_siControl_Repl2


title: p300_ChIPSeq__input_siControl_Repl1


title: p300_ChIPSeq_siPFKFB3_Repl2


title: Input_ChIPSeq_siControl_Repl2


title: Input_ChIPSeq_siPFKFB3_Repl2


title: NPAI-control (71571)


title: Input_ChIPSeq_siControl_Repl1


title: H3K27ac_ChIPSeq_siControl_Repl2

value: H3K27ac (Cell Signaling Technology, 8173S, lot1)


title: NPAI-control (71573)


title: Notch1_ChIPSeq_siPFKFB3_Repl2

value: Notch1 (Cell Signaling Technology, 3608S, lot5)


title: Notch1_ChIPSeq_siPFKFB3_Repl1


title: Notch1_ChIPSeq_siControl_Repl2


title: Notch1_ChIPSeq_siControl_Repl1


title: H3K27ac_ChIPSeq_siPFKFB3_Repl2


title: DPSC_1

status: Public on Mar 17 2019

city: Taichung

first: Shih-Chieh

last: Hung

address: No. 120, Wuyi Street, North District

department: Integrative Stem Cell Center

institute: China Medical University Hospital

data_processing:
Illumina RTA v1.17.21.3 software used for basecalling, and CASAVA V1.8.2 software used for demultiplex and data converstion
Low quality of sequencing bases were trimmed based on the Phred+33 quality score (> 20) from both of the 5'- and 3'-ends of reads. After trimming, reads are discarded, if they are shorter than 75bp, or have one or more ambiguous base.
The alignment, quantification, normalization, and differential expression analysis were performed by STAR 2.5.3a, htseq-count 0.6.0, TMM, and edgeR 3.18.1, respectively.
Genes with count-per-million (CPM) values above 1 in at least three samples were retained, and genes with no or low expression levels were discarded.
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text file includes count-per-million (CPM) values of 12742 genes for each Sample

molecule: polyA RNA

source_name: dental pulp-derived mesenchymal stem cell

growth_protocol: Normal exfoliated human deciduous incisors were collected from 7- to 8-year-old children in the Pediatric Dental Department, while bone marrow aspirates were collected from patients who received an orthopaedic surgery in the orthopaedic department of the of Taipei Veterans General Hospital with approval of Institute of Review Board. The pulp was isolated from a remnant crown and then digested in a solution of 3 mg/ml collagenase type I (Worthington Biochemical Corp., Lakeside, NJ) and 4 mg/ml dispase (Roche Molecular Biochemicals, Mannheim, Germany) at 37°C for 1 h. The cell suspension was filtered through a 40 μm cell strainer (BD Falcon; BD Biosciences, Bedford, MA) and the nucleated cells were plated in 6-well plastic dishes at clonal density and cultured in complete medium [CM: α-MEM (α-minimal essential medium; Gibco-BRL, Gaithersburg, MD), supplemented with 10% fetal bovine serum (FBS), 100 units/mL penicillin, 100 μg/mL streptomycin, and 2 mM L-glutamine (Invitrogen, Carlsbad, CA)] at 37°C under 5% CO2 atmosphere. The isolation of human BMSCs has been described previously which was originally derived from the iliac crest of donors(Tsai et al. 2011). Briefly, mononuclear cells were isolated from heparinized bone marrow by density gradient centrifugation using Ficoll- Hypaque of a density of 1.077 g/L, followed by seeding into 6-well plate with CM at 37°C under 5% CO2 atmosphere. At 9 days after seeding, both of DPSCs and BMSCs were recovered and reseeded in 10-cm plastic dishes at an initial density of 4×10^3 cells/cm2 and the culture medium was changed twice per week. The medium was changed twice a week and a subculture is performed at 1:3 to 1:5 split every week.

extract_protocol:
Total RNA was isolated with the GeneAll Hybrid-R RNA purification kit
RNA libraries were prepared for sequencing using Illumina TruSeq RNA sample Prep kit

tag: cell type

value: DPSC


title: DPSC_2


title: DPSC_3


title: BMSC_T8E

source_name: bone marrow-derived mesenchymal stem cell

value: BMSC


title: BMSC_T9E


title: BMSC_T7E


title: Input_ChIPSeq_siPFKFB3_Repl1


title: Psd1_oe2-8_rep1

status: Public on Mar 16 2019

first: Charles

middle: Arthur

last: Whittaker

institute: Koch Institute

description:
d1679_Psd1_oe2-8_rep1
mRNA
Carrying a 2µ plasmid to express PSD1 from the GAL promoter.
Processed data file: yeast_count_table.txt.gz
Processed data file: yeast_fpkm_table.txt.gz

data_processing:
HiSeq2000 using v3 chemistry. Control software version 2.0.12.0, RTA version 1.17.21.3, BCL files were converted to fastq using bcl2qseq. Indexes were split using custom scripts allowing 1 mismatch.
Map to Ensembl sacCer3 assembly with tophat 2.0.9, bowtie 2.1.0 and samtools 0.1.18.
Generate transcripts with cufflinks 2.2.0.
Merge transcripts with cuffmerge 2.2.0.
Summarize expression with cuffquant 2.2.0.
Prepare counts and FPKM data with cuffdiff 2.2.0.
Parse count and FPKM data from cufflinks genes.read_group_tracking with custom scripts.
Genome_build: sacCer3 (R64)
Supplementary_files_format_and_content: yeast_merged.gtf.gz is the merged gtf from cuffmerge associating XLOC ids from tophat-cufflinks-cuffquant-cuffdiff run with gene and orf names.
Supplementary_files_format_and_content: yeast_count_table.txt.gz is parsed integer counts for samples from cufflinks genes.read_group_tracking.
Supplementary_files_format_and_content: yeast_fpkm_table.txt.gz is parsed fpkm for samples from cufflinks genes.read_group_tracking.

organism: Saccharomyces cerevisiae

source_name: GAL-PSD1 W303

growth_protocol: YPR (1% yeast extract, 2% peptone, 2% raffinose).

extract_protocol:
RNeasy mini-kit (Qiagen).
Illumina TruSeq.

treatment_protocol: Induction of GAL-PSD1 (+/-) with 2% galactose for 4h.


title: Psd1_oe_Pdr3_ko2-9_rep1

description:
d1680_Psd1_oe_Pdr3_ko2-9
mRNA
Carrying a 2µ plasmid to express PSD1 from the GAL promoter.
Processed data file: yeast_count_table.txt.gz
Processed data file: yeast_fpkm_table.txt.gz

source_name: pdr3∆ GAL-PSD1 W303


title: WT2-10_rep2

description:
d1681_WT2-10_rep2
mRNA
Processed data file: yeast_count_table.txt.gz
Processed data file: yeast_fpkm_table.txt.gz

source_name: Wild type W303

value: Wild type control


title: H3K27ac_ChIPSeq_siControl_Repl1


title: Psd1_oe_Pdr3_ko2-12_rep2

description:
d1683_Psd1_oe_Pdr3_ko2-12
mRNA
Carrying a 2µ plasmid to express PSD1 from the GAL promoter.
Processed data file: yeast_count_table.txt.gz
Processed data file: yeast_fpkm_table.txt.gz


title: WT2-7_rep1

description:
d1678_WT2-7_rep1
mRNA
Processed data file: yeast_count_table.txt.gz
Processed data file: yeast_fpkm_table.txt.gz


title: CCCP_rep1

description:
d843_CCCP
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz

data_processing:
HiSeq2000 using v3 chemistry. Software: Control software version 2.0.12.0, RTA version 1.17.21.3. BCL files were converted to fastq using bcl2qseq. Indexes were split using custom scripts allowing 1 mismatch.
Sequence aligned and transcripts quantified using bowtie2.2.3, rsem1.2.30 and samtools 0.1.19.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: weidberg_HELA_counts.txt.gz is parsed integer counts for samples from rsem.
Supplementary_files_format_and_content: weidberg_HELA_fpkm.txt.gz is parsed fpkm for samples from rsem.

source_name: HeLa cells, CCCP

growth_protocol: DMEM supplemented with 10% FBS and 100 U/mL penicillin. rho0 cells were supplemented with 50 μg/ml uridine and 1mM sodium pyruvate.

extract_protocol:
RNeasy mini-kit (Qiagen).
Illumina TruSeq followed by Spri prep.

treatment_protocol: CCCP was added to a final concentration of 10 μM for 1h.

tag: cell line

value: HeLa

value: Wild type+CCCP


title: CCCP_rep3

description:
d849_CCCP_01.19
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz


title: rho0_rep2

description:
d845_rho0_12.25
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz

source_name: HeLa cells, rho0


title: WT_rep3

description:
d847_WT_01.19
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz

source_name: HeLa cells, WT

value: Wild type


title: CCCP_rep2

description:
d846_CCCP_12.25
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz


title: WT_rep2

description:
d844_WT_12.25
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz


title: rho0_rep1

description:
d842_rho0
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz


title: rho0_rep3

description:
d848_rho0_01.19
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz


title: Donor15 N6OU and antagonist

status: Public on Mar 15 2019

city: Oxford

first: Udo

last: Oppermann

state: Oxfordshire

address: Botnar Research Centre, Nuffield Orthopaedic Centre, Windmill Road

department: Nuffield Department of Orthopaedics, Rheumatology and Musculoskeletal Sciences

institute: University of Oxford

description:
D15_UA
PolyA RNA
Processed data file: counts.csv

data_processing:
Following QC and adaptor trimming, samples were aligned to GRCh38.p3 using HISAT2, sorted and duplicates marked using Sambamba and primary alignments reads counted at the exon level and summarized at the gene level using FeatureCounts.
Differential expression analysis was performed using DESeq2, using a model encompassing treatment group and donor.
A corrected p-value of 0.05 and an absolute log2 fold change of 0.7 was used as the criteria for determining significant regulation of a gene.
Genome_build: GRCh38.p3
Supplementary_files_format_and_content: counts.csv: Comma-separated text file contains counts matrix of reads.

source_name: Macrophage, LPS and N6OU and antagonist

growth_protocol: Human monocytes were isolated by Ficoll and Percoll gradient centrifugation from leukocyte preparations from the NHS blood service. Monocytes were seeded at 250000 cells per well in 24 well plates in 500ul X-VIVO 10 medium in 1% autologous serum.

extract_protocol:
Treated samples were lysed in RLT buffer and total RNA was extracted using the RNeasy Kit (Qiagen) according to the manufacturer's instructions. Integrity was assessed on the Agilent Tape Station 2200 requiring a RIN score greater than 8.
200ng total RNA was used as the starting material. PolyA selection was performed using the NEBNext PolyA selection Kit (NEB) followed by stranded library production using the NEBNext Ultra Directional Library Kit. Samples were sequenced on a NextSeq 500 42bp paired-end.

treatment_protocol: Cells were incubated with either DMSO as a non-stimulated vehicle control or 100ng/ml LPS for 2 hours. LPS-treated cells were then either preincubated with the antagonist (5uM BAY229703 ) or vehicle for 1 hour before treatment with GPR84 agonist, either 500nM N6OU or 5uM embelin.

value: Macrophage

value: LPS and N6OU and antagonist

tag: donor

tag: disease status

value: Healthy


title: Donor15 Embelin

description:
D15_E
PolyA RNA
Processed data file: counts.csv

source_name: Macrophage, LPS and Embelin

value: LPS and Embelin


title: H3K27ac_ChIPSeq_siPFKFB3_Repl1


title: Donor14 Vehicle

description:
D14_V
PolyA RNA
Processed data file: counts.csv

source_name: Macrophage, DMSO

value: DMSO


title: Donor14 N6OU

description:
D14_U
PolyA RNA
Processed data file: counts.csv

source_name: Macrophage, LPS and N6OU

value: LPS and N6OU


title: Donor14 N6OU and antagonist

description:
D14_UA
PolyA RNA
Processed data file: counts.csv


title: Donor14 LPS

description:
D14_LPS
PolyA RNA
Processed data file: counts.csv

source_name: Macrophage, LPS

value: LPS


title: Donor14 Embelin

description:
D14_E
PolyA RNA
Processed data file: counts.csv


title: Psd1_oe2-11_rep2

description:
d1682_Psd1_oe2-11_rep2
mRNA
Carrying a 2µ plasmid to express PSD1 from the GAL promoter.
Processed data file: yeast_count_table.txt.gz
Processed data file: yeast_fpkm_table.txt.gz

tag: strain/background


title: Donor14 Embelin and antagonist

description:
D14_EA
PolyA RNA
Processed data file: counts.csv

source_name: Macrophage, LPS and Embelin and antagonist

value: LPS and Embelin and antagonist


title: Donor13 Vehicle

description:
D13_V
PolyA RNA
Processed data file: counts.csv


title: Donor15 LPS

description:
D15_LPS
PolyA RNA
Processed data file: counts.csv


title: Donor13 LPS

description:
D13_LPS
PolyA RNA
Processed data file: counts.csv


title: Donor15 Embelin and antagonist

description:
D15_EA
PolyA RNA
Processed data file: counts.csv


title: Donor15 N6OU

description:
D15_U
PolyA RNA
Processed data file: counts.csv


title: Donor13 Embelin and antagonist

description:
D13_EA
PolyA RNA
Processed data file: counts.csv


title: Donor11 Vehicle

description:
D11_V
PolyA RNA
Processed data file: counts.csv


title: WT_rep1

description:
d841_WT
mRNA
Processed data file: weidberg_HELA_counts.txt.gz
Processed data file: weidberg_HELA_fpkm.txt.gz


title: Donor13 Embelin

description:
D13_E
PolyA RNA
Processed data file: counts.csv


title: Donor15 Vehicle

description:
D15_V
PolyA RNA
Processed data file: counts.csv


title: Donor11 Embelin and antagonist

description:
D11_EA
PolyA RNA
Processed data file: counts.csv


title: Donor10 LPS

description:
D10_LPS
PolyA RNA
Processed data file: counts.csv


title: Donor11 LPS

description:
D11_LPS
PolyA RNA
Processed data file: counts.csv


title: Donor11 N6OU and antagonist

description:
D11_UA
PolyA RNA
Processed data file: counts.csv


title: Donor10 Vehicle

description:
D10_V
PolyA RNA
Processed data file: counts.csv


title: Donor11 N6OU

description:
D11_U
PolyA RNA
Processed data file: counts.csv


title: Donor10 N6OU

description:
D10_U
PolyA RNA
Processed data file: counts.csv


title: Donor10 Embelin

description:
D10_E
PolyA RNA
Processed data file: counts.csv


title: Donor11 Embelin

description:
D11_E
PolyA RNA
Processed data file: counts.csv


title: Donor10 Embelin and antagonist

description:
D10_EA
PolyA RNA
Processed data file: counts.csv


title: Wild Type rep3

city: Chicago

first: Cambrian

middle: Yangshao

last: Liu

state: IL

address: 5841 S Maryland Ave

department: Department of Medicine

institute: The University of Chicago

data_processing:
pseudoalignment and transcript quantification with kallisto 0.43.1
statistical significance calling with sleuth
Genome_build: mm10
Supplementary_files_format_and_content: Comma-separated text file with tpm calculations for each annotated transcript for each sample

source_name: colon

extract_protocol: Extraction protocol was the standard protocol associated with the PureLink RNA Mini Kit (ThermoFisher Scientific). Library construction protocol was the paired-end protocol in the SMARTer Universal Low Input RNA Kit for Sequencing (Clontech).

value: WT

value: colon


title: Wild Type rep1


title: R1ko rep1


title: R1ko rep2


title: P5.5_KIT_negative_SC71

city: Hong Kong

first: Jinyue

last: Liao

address: Room 38016, 1/F Lui Che Woo Clinical Sciences Building

department: Department of Chemical Pathology

country: Hong Kong

institute: The Chinese University of Hong Kong

description: Processed data file: TPM.matrix.txt

data_processing:
Illumina Casava1.7 software used for basecalling.
Raw reads were pre-processed with trim galore for trimming Illumina adaptor sequences (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/).
Expression level for all RefSeq genes was quantified using Salmon.
Genome_build: mm9 (MGSCv37)
Supplementary_files_format_and_content: Salmon.Expected.count.matrix.xlsx: Excel file includes expected count value for each cell as calculated by Salmon.

source_name: Spermatogonial stem and progenitor cell (SSC)

extract_protocol:
Freshly sorted Oct4-GFP+/KIT- cells were submitted to a Fluidigm C1 System machine for single cell capture and cDNA synthesis.
Libraries were prepared using the Nextera XT kit and sequencing was carried out on an Illumina Hiseq 2000 using standard Illumina protocols.

value: Oct4-GFP+ germ cells

tag: gender

value: male

tag: age

value: postnatal day 5.5


title: Wild Type rep2

tag: tissue


title: Donor13 N6OU and antagonist

description:
D13_UA
PolyA RNA
Processed data file: counts.csv

tag: treatment


title: Donor13 N6OU

description:
D13_U
PolyA RNA
Processed data file: counts.csv


title: P5.5_KIT_negative_SC70

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC66

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC67

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC63

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC65

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC62

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC68

description: Processed data file: TPM.matrix.txt


title: Donor10 N6OU and antagonist

description:
D10_UA
PolyA RNA
Processed data file: counts.csv


title: P5.5_KIT_negative_SC64

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC61

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC59

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC57

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC58

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC60

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC43

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC69

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC44

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC49

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC45

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC54

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC46

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC47

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC51

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC53

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC52

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC55

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC50

description: Processed data file: TPM.matrix.txt


title: R1ko rep3


title: P5.5_KIT_negative_SC41

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC40

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC38

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC39

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC36

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC42

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC35

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC34

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC37

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC56

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC33

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC32

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC30

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC31

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC48

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC29

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC28

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC25

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC27

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC24

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC19

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC26

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC15

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC21

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC18

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC14

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC17

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC11

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC23

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC13

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC9

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC20

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC5

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC12

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC10

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC8

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC4

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC6

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC7

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC1

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC3

description: Processed data file: TPM.matrix.txt


title: P5.5_KIT_negative_SC22

description: Processed data file: TPM.matrix.txt


title: Couple Antibiotics-treated Female Cftr -/- ileum

city: Rotterdam

first: Marcel

last: Bijvelds

address: PO Box 2040

department: Gastroenterology and Hepatology

country: Netherlands

institute: Erasmus MC University Medical Center

description: Littermate couple ABX Cftr KO ileum

data_processing:
Basecalls performed using Illumina's Real Time Analysis Software (HiSeq2000 ) or Ion Proton use Proton™ Torrent Server and Torrent Suite Software
Raw reads are filtered by removing the adaptors, reads in which unknown bases are more than 10%, low quality reads (the percentage of low quality bases is over 50% in a read, where the low quality base is defined as  base whose sequencing quality is no more than 5). After filtering, the remaining reads are stored as FASTQ.
Reads are mapped and RPKM values are calculated by CLC Genomics workbench 7.5.1, using default settings
Genome_build: GRCm38.76
Supplementary_files_format_and_content: Tab-delimited text file with normalized RPKM values for all the samples

source_name: Small intestine, <6 cm proximal to the ileocecal valve

growth_protocol: Animals were reared on a low fiber diet (C1013; Altromin) and an osmotic laxative (ad libitum)

extract_protocol:
Sampling from a CF animal and a sex-matched littermate control was performed within a time window of 20 min, and between 12:00-14:00h. Tissue was homogenized with a rotor-stator homogenizer in Trizol reagent (Qiagen), and total RNA was extracted using the Nucleospin RNA kit (Macherey-Nagel). Integrity of the extracted RNA was verified by gel electrophoresis.
Libraries were prepared by BGI. In brief, the mRNA is enriched by using the oligo(dT) magnetic beads. Mixed with the fragmentation buffer, the mRNA is fragmented into short fragments. Then the first strand of cDNA is synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End reparation and 3’-end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The fragments are enriched by PCR amplification. During the QC step, Agilent 2100 Bioanaylzer and ABI StepOnePlus Real-Time PCR System are used to qualify and quantify of the sample library. The library products are ready for sequencing via Illumina HiSeqTM 2000 or Ion Proton System (4476610).

treatment_protocol: Laxative treatment was discontinued >4 days before sampling. Where indicated (ABX) the drinking water was supplemented with ciprofloxacin (0.3 g/L) and metronidazole (0.5 g/L).

value: Ileum - Whole tissue section

value: F

value: FVB

value: Cftr -/-

value: ABX


title: P5.5_KIT_negative_SC16

description: Processed data file: TPM.matrix.txt


title: Couple 2 Male Cftr -/- ileum

description: Littermate couple 2_m Cftr KO ileum

value: M


title: Couple 1 Female Cftr -/- ileum

description: Littermate couple 1_f Cftr KO ileum


title: Couple Antibiotics-treated Female Cftr N/N ileum

description: Littermate couple ABX Cftr WT ileum

value: Cftr N/N


title: Couple 2 Male Cftr N/N ileum

description: Littermate couple 2_m Cftr WT ileum


title: Couple 3 Female Cftr -/- ileum

description: Littermate couple 3_f Cftr KO ileum


title: Couple 3 Female Cftr N/N ileum

description: Littermate couple 3_f Cftr WT ileum


title: P3HR1_DMSO_8hr_4

status: Public on Mar 14 2019

first: Andrew

last: Chen

address: 500 Main St. Room 389

department: Biological Engineering

institute: Massachusetts Institute of Technology

description:
counts_raw_cpms.txt
Merged fastq files contain genomic reads with barcode informations concatenated with read IDs; collapsed reads contain single exemplars of reads, collapsed by UMI and reads sequence, which were used for downstream analyses.

data_processing:
Post-sequencing, quality-control on each of the libraries was performed to assess coverage depth, enrichment for messenger RNA (exon/intron and exon/intergenic density ratios), fraction of rRNA reads and number of detected genes using bespoke scripts. Reads bearing the same Unique Molecular Identifier (UID) and the same sequence were collapsed into unique exemplar for downstream processing. Sequences were aligned against the human genome hg19 and ERCC reference sequences separately using bwa.
Gene expression was estimated based on reads mapping near the 3' end of transcripts using ESAT (Derr A. et al. End Sequence Analysis Toolkit (ESAT) expands the extractable information from single-cell RNA-seq data. Genome Res. 2016 Oct;26(10):1397-1410. Epub 2016 Jul 28.) based on the hg19 Refseq annotation.
Results were summarized as counts per million mapped reads (CPMs), merged across samples, log-transformed and subjected to hierarchical clustering and visualization. For ERCC quantification, bam files from bwa were sorted and indexed with samtools (Li H. et al. 1000 Genome Project Data Processing Subgroup. The Sequence Alignment/Map format and SAMtools. Bioinformatics. 2009 Aug 15;25(16):2078-9. doi: 10.1093/bioinformatics/btp352. Epub 2009 Jun 8.) and counts were retrieved from the indices using idxstats.
Genome_build: hg19
Supplementary_files_format_and_content: raw counts and Counts per M mapped reads (CPMs) are reported for each annotated gene in each sample (tab-delimited file).

growth_protocol: Cells were plated at 3 million cells in 3 mL media per well in 6-well plates.

extract_protocol:
1.5 million cells were washed with 1 mL cold PBS, followed by extraction of RNA using Aurum™ Total RNA Mini Kit (BioRad 732-6820)
RNA samples were quantified and quality assessed using an Advanced Analytical Fragment Analyzer. 20 ng of totalRNA was used for library preparation with ERCC Spike-in control Mix A (Ambion 10-6 final dilution). All steps were performed on a Tecan EVO150. 3’DGE-custom primers 3V6NEXT-bmc#1-24 were added to a final concentration of 1.2 µM. (5'-/5Biosg/ACACTCTTTCCCTACACGACGCTCTTCCGATCT[BC6]N10T30VN-3'  where  5Biosg = 5’  biotin, [BC6] = 6bp barcode specific to each sample/well, N10 = Unique Molecular Identifiers, Integrated DNA technologies).  After addition of the oligonucleotides, samples were denatured at 72 °C for 2 minutes followed by addition of SMARTScribe RT per manufacturer’s recommendations with Template-Switching oligo5V6NEXT (12 µM, [5V6NEXT : 5’-iCiGiCACACTCTTTCCCTACACGACGCrGrGrG-3’  where  iC: iso-dC, iG: iso-dG, rG: RNA G ]) and incubation at 42 °C for 90 minutes followed by inactivation at 72 °C for 10 minutes.   Following the template switching reaction, cDNA from 24 wells containing unique well identifiers were pooled together and cleaned using RNA Ampure beads at 1.0X. cDNA was eluted with 90 µl of water followed by digestion with Exonuclease I at 37 °C for 45 minutes, inactivation at 80 °C for 20 minutes.  Single stranded cDNA was then cleaned using RNA Ampure beads at 1.0X and eluted in 50 µl of water.   Second strand synthesis and PCR amplification was done using the Advantage 2 Polymerase Mix (Clontech) and the SINGV6 primer (10 pmol, Integrated DNA Technologies 5’-/5Biosg/ACACTCTTTCCCTACACGACGC-3’ ). 12 cycles of PCR were performed followed by clean up using regular SPRI beads at 1.0X, and was eluted with 20 µl of EB. Successful amplification of cDNA was confirmed using the Fragment Analyzer. Illumina libraries were then produced using standard Nextera tagmentation substituting P5NEXTPT5-bmc primer (25 μM, Integrated DNA Technologies, (5’-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCG*A*T*C*T*-3’  where  * = phosphorothioate bonds. ) in place of the normal N500 primer.    Final libraries were cleaned using SPRI beads at 0.8X and quantified using the Fragment Analyzer and qPCR before being loaded for paired-end sequencing using the Illumina NextSeq500.
3' Digital Gene Expression (adapted from Soumillon et al. bioRxiv March 5, 2014 doi: https://doi.org/10.1101/003236  ).

treatment_protocol: Cells were treated with either 0.4% DMSO, 1 µM KI-MS2-008, 10 µM KI-MS2-008 or 0.1 µg/mL doxycycline, followed by brief shaking to mix. Cells were treated for 45 minutes or 8 hours.

value: B lymphocyte

tag: inducible expression system

tag: timepoint

value: 8 hour

value: 0.4% DMSO


title: P3HR1_10uM_8hr_3


title: P3HR1_10uM_8hr_1


title: P3HR1_10uM_8hr_4


title: P5.5_KIT_negative_SC2

description: Processed data file: TPM.matrix.txt


title: ST486_10uM_8hr_1


title: P3HR1_DMSO_8hr_1


title: ST486_1uM_8hr_4


title: ST486_10uM_8hr_4


title: ST486_10uM_8hr_3


title: ST486_1uM_8hr_2


title: ST486_1uM_8hr_3


title: P3HR1_DMSO_8hr_3


title: ST486_10uM_8hr_2


title: ST486_10uM_45min_4

value: 45 min


title: ST486_DMSO_8hr_1


title: ST486_DMSO_8hr_2


title: ST486_DMSO_8hr_3


title: Couple 1 Female Cftr N/N ileum

description: Littermate couple 1_f Cftr WT ileum

tag: genotype


title: ST486_10uM_45min_2


title: ST486_10uM_45min_3


title: ST486_1uM_45min_4


title: ST486_10uM_45min_1


title: P3HR1_DMSO_8hr_2


title: ST486_1uM_45min_3


title: ST486_1uM_45min_2


title: P3HR1_10uM_8hr_2


title: ST486_1uM_8hr_1


title: ST486_1uM_45min_1


title: ST486_DMSO_8hr_4


title: ST486_DMSO_45min_3


title: ST486_DMSO_45min_2


title: P4936_10uM_8hr_3

value: Tet-off for c-Myc


title: P4936_Dox_8hr_2

value: 0.1 ug/mL doxycycline


title: ST486_DMSO_45min_1


title: P4936_10uM_8hr_2


title: P4936_Dox_8hr_3


title: P4936_10uM_8hr_4


title: P4936_10uM_8hr_1


title: P4936_Dox_8hr_4


title: P4936_1uM_8hr_1


title: P4936_1uM_8hr_3


title: P4936_1uM_8hr_4


title: P4936_1uM_8hr_2


title: P4936_DMSO_8hr_4


title: P4936_DMSO_8hr_2


title: P4936_DMSO_8hr_3


title: P4936_Dox_45min_4


title: P4936_Dox_45min_1


title: P4936_Dox_45min_3


title: P4936_Dox_45min_2


title: P4936_10uM_45min_3


title: P4936_10uM_45min_4


title: ST486_DMSO_45min_4


title: P4936_10uM_45min_1


title: P4936_10uM_45min_2


title: P4936_Dox_8hr_1


title: P4936_1uM_45min_4


title: P4936_1uM_45min_2


title: P4936_1uM_45min_3


title: P4936_1uM_45min_1


title: P4936_DMSO_45min_4


title: P4936_DMSO_45min_2


title: P4936_DMSO_45min_1


title: WT CD8+ T cells stimulated for 30 minutes #2

city: Los Angeles

first: Ameya

last: Champhekar

address: 700 Tiverton Ave, 11-272 Factor Bldg

department: Medicine, HemOnc Division

institute: University of California, Los Angeles

data_processing:
Adapter sequences were removed and the resulting 50 bp reads were aligned to the mouse genome release mm10/GRCm38
The splice read aligner TopHat (v2.0.12) was used for alignment with the following command: tophat --library-type fr-firststrand --no-coverage-search
Read counts for each gene were obtained with the HTSeq package (v0.6.1) using the command:  htseq-count -s no -a 10 Mus_musculus.GRCm38.76.gtf
RPKM values were calculated by dividing the number of uniquely mapped reads for each gene by the product of transcript length (kb) and the total reads for that sample (in millions)
Genome_build: mm10/GRCm38
Supplementary_files_format_and_content: Tab-delimited text file with RPKM values for all samples. Only genes expressed at 1 RPKM or more in at least 1 sample are included. The cutoff was applied after averaging RPKM values for replicates of each sample.

source_name: CD8+ T cells from WT spleen

growth_protocol: Naïve CD8+ T cells were enriched from 6 week old WT or Ikaros-Zn4-/- spenocytes by negative selection using the mouse naïve CD8+ T cell isolation kit (StemCell Technologies). Naïve CD8+ T cells, defined as CD8+ CD62Lhi CD44 lo/-,  were then sorted from the enriched samples.

extract_protocol:
Naïve or stimulated CD8+ T cells were harvested and lysed using the TRI reagent (MRC, Inc.). Total RNA was extracted using manufacturer's suggested protocol and directly used for library preparation.
TruSeq RNA Library Prep Kit v2 (Illumina, Inc.) was used to construct single-end libraries and sequenced on the Illumina Hiseq 2000 sequencer.

treatment_protocol: Sorted naïve CD8+ T cells were either used directly or stimulated with plate-bound anti-CD3 and anti-CD28 antibodies for up to 2 hours.

tag: strain

value: WT

value: 6 weeks

tag: Sex

value: Female

value: Spleen


title: Ikaros-Zn4-/- CD8+ T cells stimulated for 30 minutes #1

source_name: CD8+ T cells from Ikaros-Zn4-/- spleen


title: WT CD8+ T cells stimulated for  2 hours


title: Ikaros-Zn4-/- CD8+ T cells stimulated for 2 hours #1


title: Naïve Ikaros-Zn4-/- CD8+ T cells #1, 0 min


title: Ikaros-Zn4-/- CD8+ T cells stimulated for 30 minutes #2


title: Naïve Ikaros-Zn4-/- CD8+ T cells #2, 0 min


title: WT CD8+ T cells stimulated for 30 minutes #1


title: P4936_DMSO_8hr_1


title: Naïve WT CD8+ T cells #1, 0 min


title: Ikaros-Zn4-/- CD8+ T cells stimulated for 24 hours #2

data_processing:
Adapter sequences were removed and the resulting 50 bp reads were aligned to the mouse genome release mm10/GRCm38
The splice read aligner TopHat (v2.0.12) was used for alignment with the following command: tophat --library-type fr-firststrand --no-coverage-search
Read counts for each gene were obtained with the HTSeq package (v0.6.1) using the command:  htseq-count -s no -a 10 Mus_musculus.GRCm38.76.gtf
RPKM values were calculated by dividing the number of uniquely mapped reads for each gene by the product of transcript length (kb) and the total reads for that sample (in millions)
Genome_build: mm10/GRCm38
Supplementary_files_format_and_content: Tab-delimited file containing the RPKM values for all samples. Only genes expressed at 1 RPKM or more in at least 1 sample are included. The cutoff was applied after averaging RPKM values for replicates of each sample.

extract_protocol:
Naïve or stimulated CD8+ T cells were harvested and lysed using the TRI reagent (MRC, Inc.). Total RNA was extracted using manufacturer's suggested protocol and directly used for library preparation.
TruSeq Stranded mRNA Library Prep Kit (Illumina, Inc.) was used to construct single-end libraries and sequenced on the Illumina Hiseq 2000 sequencer.

treatment_protocol: Sorted naïve CD8+ T cells were either used directly or stimulated with plate-bound anti-CD3 and anti-CD28 antibodies for up to 24 hours.

tag: background strain


title: Ikaros-Zn4-/- CD8+ T cells stimulated for 24 hours #1


title: P4936_DMSO_45min_3


title: Naïve Ikaros-Zn4-/- CD8+ T cells #2


title: WT CD8+ T cells stimulated for 24 hours #2


title: Ikaros-Zn4-/- CD8+ T cells stimulated for 12 hours #2


title: Naïve Ikaros-Zn4-/- CD8+ T cells #1


title: Ikaros-Zn4-/- CD8+ T cells stimulated for 12 hours #1


title: WT CD8+ T cells stimulated for 24 hours #1


title: WT CD8+ T cells stimulated for 12 hours #2


title: WT CD8+ T cells stimulated for 12 hours #1


title: Naïve WT CD8+ T cells #1


title: Input, VEGF treatment

status: Public on Mar 12 2019

city: Heidelberg

first: Carmen

last: Ruiz de Almodovar

address: Im Neuenheimer Feld 328

country: Germany

institute: Heidelberg University Biochemistry Center (BZH)

description: Input aliquot

data_processing:
The reads were aligned to the hg38 genome assembly using Bowtie2 v2.2.5
Peak calling was done with MACS v1.4.2, using the corresponding Input samples as controls.
Peaks in the centromeres, in repeat regions (from UCSC database, based on RepeatMasker and Tandem Repeats Finder), and in human blacklist regions (from ENCODE project) were excluded.
Peaks were paired with target genes by a strict +-1kb window at TSS.
Peaks were also converted to hg37 coordinates with UCSC liftover for using in EnhancerAtlas to pair them with target genes.
Genome_build: hg38
Supplementary_files_format_and_content: Bed format files with MACS peak coordinates for hg38, and converted for hg37

source_name: HBMECs

growth_protocol: HBMECs were maintained in Endopan 3 Kit. Before experiment, cells were cultured in subconfluent status and starved for overnight.

extract_protocol:
Cells were then fixed with 1% formaldehyde (sigma) in medium for 10min at room temperature, and lysed in 1%SDS lysis buffer. Chromatin was sheared to approximately 200bp fragments by using Covaris S220X.
Libraries were generated by ThruPLEX DNA-seq Kit (Rubicon).

treatment_protocol: Treated with or without 50ng/ml VEGF for 6h.

value: VEGF treatment (6h)

tag: antibody

tag: beads


title: YAP/TAZ ChIP, Control

description: IP sample

value: no treatment

value: YAP/TAZ (D24E4) Rabbit mAb (Cell signalling)

value: Protein A/G magnetic beads (Pierce)


title: YAP/TAZ ChIP, VEGF treatment

description: IP sample


title: MCF10AsiSIER_NT_rna_rep1

city: Milano

first: Pier Giuseppe

last: Pelicci

address: Via Adamello, 16

department: Department of Experimental Oncology

country: Italy

institute: European Institute of Oncology

description: Poly(A)-containing mRNA

data_processing:
Basecalls performed using CASAVA version 1.9
Data were filtered using the Illumina quality control flag
Reads were aligned to the Homo sapiens genome build hg18 using tophat (v2.0.8), which uses Bowtie (2.1.0) and Samtools (v0.1.18) using default parameters and transcriptome Homo sapiens index, UCSC hg18 gtf.
Raw read counts were summarised using HTSeq (v0.5.3p9), then gene expression counts for the i-th gene (n_i) were converted into “transcripts per million” (TPM) of mapped reads, with TPM(i) = 10^6∙(n_i/L_i )/∑_i(n_i/L_i ) , where L_i is the length of the i-th coding sequence. Every variable was converted to base 2 logarithm.
Genome_build: hg18
Supplementary_files_format_and_content: tab-delimited text files include genomic coordinates, names, strand and expression level means (log2TPM) for each gene.

growth_protocol: MCF10A parental cells and MCF10A AsiSI cells were routinely grown by re-plating adherent cultures (typically 8 × 10^5 cells into a tissue-culture dish 100mm) in a medium composed of DMEM (Lonza) and HAM's F12 (GibcoLife Technologies) 1:1, supplemented with horse serum 1%, hydrocortisone 0,5mg/ml, hEGF10ug/ml, insulin 5mg/ml, glutamine 0,5% and 100 ng/ml of cholera toxin.

extract_protocol:
Total RNA was extracted using Quick-RNA Miniprep kit (Zymo Research, Cat#R1055)
RNA libraries were prepared for sequencing using Illumina TruSeq RNA Sample Prep Kit (Cat#RS-122-2002), with 600ng of input RNA.

tag: tissue source

value: normal mammary gland

tag: genotype/variation


title: Input, Control

description: Input aliquot


title: MCF10AsiSIER_NT_gH2AX_chip

data_processing:
Basecalls performed using CASAVA version 1.9
Data were filtered using the Illumina quality control flag
ChIP-seq: reads were aligned to the hg18 genome assembly using bwa version 0.6.2 with the default parameters
ChIP-seq: low-quality and multiple-mapping reads were removed using Samtools with option ‘‘-q 1’
ChIP-seq: duplicates (reads with the same start and end position mapping) that are potential PCR artifacts are removed with samtools rmdup -s
ChIP-seq: peaks were called using MACS 1.4.1 with default setting
BLISS: reads were aligned to the hg18 genome assembly using bwa version 0.6.2 with the default parameters
BLISS: low-quality and multiple-mapping reads were removed using Samtools with option ‘‘-q 1’
BLISS: duplicates (reads with same start-end position) deriving from the same DSB-end independently generated in different cells, were distinguished from potential PCR artifacts due to the presence, within the BLISS adapters, of random 8-nt long sequences serving as Unique Molecular Identifiers (UMIs; details at https://doi.org/10.1101/091629): using a custom filtering script, duplicated reads with different UMIs were maintained, while the others were discarded.
BLISS: raw BLISS signal was analyzed using the findPeaks algorithm from the HOMER analysis suite enclosed inside a customizable Python-based pipeline.
BLISS: calling of BLISS enrichments was independently performed on each strand, to evaluate the correct orientation of BLISS tags (i.e., reverse tag enrichment upstream and forward tag enrichment downstream the true damaged site).
BLISS: calling of raw tag enrichments was performed using both genome-wide settings (fold-enrichment ≥ 1.5, p-value ≤ 1e-5), and local settings using a 10 kb window (fold-enrichment ≥ 2, p-value ≤ 1e-4), to account for local signal variability.
BLISS: significant BLISS enrichments of 4-OHT-treated samples were called using BLISS signals from untreated samples, while for endogenous DSBs, fold-enrichments of BLISS signals were calculated over random background.
BLISS: from an initial pool of 190,160 raw endogenous DSB clusters, 8,132 (Tier1) showed paired (Pj) p-value ≤ 1e-4, w ≥ 75 and ≥2 fold-enrichment of ChIPseq signals of at least one of three repair factors (NBS1, XRCC4, RAD51) within ±1 kb from the center of the BLISS signal, as compared to the input DNA.
Genome_build: hg18
Supplementary_files_format_and_content: ChIP-seq: bigwig files were obtained with a combination of bedtools (v2.16.1) sub-commands: aligned reads (bam) were converted in bed with bamToBed and extended to approximately 200 bp with slopbed; then genomeCoverageBed were used to generate the wig file, which in turn is transformed to bigwig with the UCSC Genome Browser utility wigToBigWig.
Supplementary_files_format_and_content: BLISS: list of Tier1 DSB clusters identified by BLISS in bed format.

growth_protocol:
ChIP-seq: MCF10A-AsiSIER cells were routinely grown by re-plating adherent cultures (typically 8 × 10^5 cells into a tissue-culture dish 100mm) in a medium composed of DMEM (Lonza) and HAM's F12 (GibcoLife Technologies) 1:1, supplemented with horse serum 1%, hydrocortisone 0,5mg/ml, hEGF10ug/ml, insulin 5mg/ml, glutamine 0,5% and 100 ng/ml of cholera toxin.
BLISS: MCF10A-AsiSIER cells were grown directly on 22 x 22 mm coverglasses in the same culture conditons used for ChIP-seq preparations.

extract_protocol:
ChIP-seq: Cells were cross-linked by adding formaldehyde to the culture medium to a final concentration of 1% (8 min at RT). Cross-linking was stopped by addition of glycine to a final concentration of 125 mM. Cells were washed twice with PBS and lysed in SDS buffer: 100 mM NaCl, 50 mM Tris HCl (pH 8.1), 5 mM EDTA (pH 8), 0.5% SDS, and protease inhibitors. Chromatin lysate was then pelleted and resuspended in IP buffer (100 mM NaCl, 100 mM Tris HCl at pH 8.1, 5 mM EDTA at pH 8, 0.3% SDS, 1.7% Triton X-100), where it was directly sonicated prior to overnight incubation with antibodies .
ChIP-seq: Libraries were prepared according to HT-ChIP-Seq Library Preparation Protocol. Briefly, DNA was end-repaired using a combination of T4 DNA polymerase and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (3´→ 5´ exo minus) and dATP to yield a protruding 3- 'A' base for ligation of indexed oligo adaptors which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with amplification primers for 14 cycles. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.
BLISS: Cells were fixed for 10 min in 4% formaldehyde at room temperature (RT). After permeabilization, cells were incubated in a blunting reaction mix (NEB, cat. no. E1201L) for 1 h at RT, followed by in situ DSB ligation in a T4 DNA ligase reaction mix (NEB, cat. no. M0202M) for 16–18 h at 16 °C. The next day, genomic DNA (gDNA) was in situ fragmented by incubating the samples with HaeIII (NEB, cat. no. R0108L) for 3 h at 37 °C. Afterwards, the cells were scraped off the coverglass, and gDNA purified using proteinase K (NEB, cat. no. P8107S). Purified gDNA was linearly amplified using the T7 RNA polymerase (Thermo, cat. no. AM1334).
BLISS: Libraries were prepared using a modified Illumina TruSeq Small RNA Library Prep Kit (RS-200-0012). A detailed, step-by-step BLISS protocol can be found at https://doi.org/10.1101/091629.

treatment_protocol:
ChIP-seq: MCF10A-AsiSIER cells were treated for 2 hours with 4-Hydroxytamoxifen 300nM (final conc), adding the drug directly into the medium.
BLISS: MCF10A-AsiSIER cells were treated for 2 hours with 4-Hydroxytamoxifen 300nM (final conc), adding the drug directly into the medium.

tag: cell line

tag: treated with

tag: chip antibody

value: gammaH2AC (Biolegend, 613401)


title: Naïve WT CD8+ T cells #2, 0 min


title: MCF10AsiSIER_NT_H3K4me1_chip

value: H3K4me1 (Abcam, ab8895)


title: MCF10AsiSIER_NT_POL2_S5_chip

value: POLR2A-phosphoS5 (Abcam, ab5131)


title: MCF10AsiSIER_NT_H3K4m3_chip

value: H3K4me3 (Active Motif, 39159)


title: MCF10AsiSIER_NT_RAD51_chip

value: RAD51 (Santa Cruz sc8349)


title: MCF10AsiSIER_NT_H3K27ac_chip

value: H3K27ac (Abcam, ab4729)


title: MCF10AsiSIER_NT_TOPO2B_chip

value: TOP2B (Abcam, Ab58442)


title: MCF10AsiSIER_4OHT2h_gH2AX_chip

value: 4-Hydroxytamoxifen (4OHT) (300 nM for 2h)


title: MCF10AsiSIER_4OHT2h_NBS1_rep2_chip

value: NBS1 (Abcam, ab32074)


title: MCF10AsiSIER_INPUT_chip


title: MCF10AsiSIER_4OHT2h_RAD51_chip


title: MCF10AsiSIER_4OHT2h_NBS1_rep1_chip


title: MCF10AsiSIER_NT_NBS1_chip


title: MCF10AsiSIER_NT_BLISS_rep1

description: processed data file: Dellino_BLISS_Processed_Data_Tier1_DSBs.txt

data_processing:
library strategy: Breaks Labeling In Situ and Sequencing (BLISS)
Basecalls performed using CASAVA version 1.9
Data were filtered using the Illumina quality control flag
ChIP-seq: reads were aligned to the hg18 genome assembly using bwa version 0.6.2 with the default parameters
ChIP-seq: low-quality and multiple-mapping reads were removed using Samtools with option ‘‘-q 1’
ChIP-seq: duplicates (reads with the same start and end position mapping) that are potential PCR artifacts are removed with samtools rmdup -s
ChIP-seq: peaks were called using MACS 1.4.1 with default setting
BLISS: reads were aligned to the hg18 genome assembly using bwa version 0.6.2 with the default parameters
BLISS: low-quality and multiple-mapping reads were removed using Samtools with option ‘‘-q 1’
BLISS: duplicates (reads with same start-end position) deriving from the same DSB-end independently generated in different cells, were distinguished from potential PCR artifacts due to the presence, within the BLISS adapters, of random 8-nt long sequences serving as Unique Molecular Identifiers (UMIs; details at https://doi.org/10.1101/091629): using a custom filtering script, duplicated reads with different UMIs were maintained, while the others were discarded.
BLISS: raw BLISS signal was analyzed using the findPeaks algorithm from the HOMER analysis suite enclosed inside a customizable Python-based pipeline.
BLISS: calling of BLISS enrichments was independently performed on each strand, to evaluate the correct orientation of BLISS tags (i.e., reverse tag enrichment upstream and forward tag enrichment downstream the true damaged site).
BLISS: calling of raw tag enrichments was performed using both genome-wide settings (fold-enrichment ≥ 1.5, p-value ≤ 1e-5), and local settings using a 10 kb window (fold-enrichment ≥ 2, p-value ≤ 1e-4), to account for local signal variability.
BLISS: significant BLISS enrichments of 4-OHT-treated samples were called using BLISS signals from untreated samples, while for endogenous DSBs, fold-enrichments of BLISS signals were calculated over random background.
BLISS: from an initial pool of 190,160 raw endogenous DSB clusters, 8,132 (Tier1) showed paired (Pj) p-value ≤ 1e-4, w ≥ 75 and ≥2 fold-enrichment of ChIPseq signals of at least one of three repair factors (NBS1, XRCC4, RAD51) within ±1 kb from the center of the BLISS signal, as compared to the input DNA.
Genome_build: hg18
Supplementary_files_format_and_content: ChIP-seq: bigwig files were obtained with a combination of bedtools (v2.16.1) sub-commands: aligned reads (bam) were converted in bed with bamToBed and extended to approximately 200 bp with slopbed; then genomeCoverageBed were used to generate the wig file, which in turn is transformed to bigwig with the UCSC Genome Browser utility wigToBigWig.
Supplementary_files_format_and_content: BLISS: list of Tier1 DSB clusters identified by BLISS in bed format.


title: MCF10AsiSIER_4OHT2h_BLISS_rep1


title: MCF10AsiSIER_4OHT2h_BLISS_rep2


title: MCF10AsiSIER_NT_BLISS_rep2