title: Female_03

city: Kuala Lumpur

first: Amir Feisal

last: Merican

state: Kuala Lumpur

address: University of Malaya

department: Institute of Biological Science

country: Malaysia

institute: University of Malaya

data_processing:
The RNA libraries were sequenced on Illumina HiSeq 2000 Platform
The qualities of all sequences in fastq format were assessed using FASTQC . The adaptors and low quality bases were trimmed from the sequence using Trimmomatic. Readwith a phred quality (Q > 20) were used for further alignment and assembly.
Quality trimmed raw reads from all 6 samples were aligned to the human reference genome sequence (GRCh38.79 assembly) separately using HISAT (v 0.1.4) with GENCODE junctions as a guided reference annotation
The aligned reads for each sample was assembled separately into transcripts by StringTie (v1.3.3) using a GENCODE reference annotation (version 22). into transcripts by StringTie (v1.3.3) [51] using a GENCODE reference annotation (version 22). The transcripts abundance was estimated as fragments per kilobases of exon per million fragments mapped (FPKM)
Genome_build: GRCh38.79
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

source_name: Monocyte Cells

extract_protocol:
CD14-Positive Monocytes cells were isolated from PBMC by negative selection method using Miltenyi Monocyte Isolation kit II (Miltenyi biotec). Total RNA was extracted from monocytes cells using the RNesasy mini kit (QIAGEN).
Messenger RNA isolation and sequencing libraries preparation was performed using TruSeq RNA Sample preparation Kit and SuperScript II Reverse Transcriptase.

tag: cell type

value: CD14-Positive Monocytes cells

tag: gender

value: female

tag: disease status

value: Healthy individuals


title: Male_03

status: Public on Dec 22 2016

type: SRA

library_source: transcriptomic

molecule: total RNA

organism: Homo sapiens

value: male

tag: age


title: T05

city: Beijing

first: Mingming

last: Cui

state: Beijing

address: No. 35 Tsinghua East Road

department: College of Forestry

country: China

institute: Beijing Forestry University

description: mixed sample of 5 degree Celsius treatment and -5 degree Celsius treatment

data_processing:
Trinity are used for de novo assembling.
Cufflinks are used for assemble transcripts and remap and quanlity gene-expression level, which is estimate by FPKM value.
Genome_build: GFBH00000000, Holcocerus.Unigene.fa
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample ...

organism: Eogystia hippophaecolus

source_name: larva body

extract_protocol:
Total RNA was extracted using TRIzol reagent (Ambion) and the RNeasy Plus Mini Kit (No. 74134; Qiagen, Hilden, Germany) following the manufacturer’s instructions. A NanoDrop 8000 (Thermo, Waltham, MA, USA) was used to quantify the RNA. The mRNA samples were purified and fragmented using the TruSeq RNA Sample Preparation Kit v2-Set A (No. RS-122-2001; Illumina, San Diego, CA, USA) to remove rRNA.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: There is one moth in T1 and T2 sample, while two individuals in T4, T5 and T6 sample. Prior to treatment, we cleaned the moths with sterile water and starved them for three days. And all of the instruments touched the moths were sterilized. After housing for 48h at room temperature, three samples of each development stage were treated differently: one stay at room temperature as a control and the other two were kept in 5℃ and -5℃, respectively, for 10h, and then snap-frozen in liquid nitrogen and stored at -80°C until the RNA extraction step.

tag: tissue

value: larva whole body

value: 10th instar

tag: treatment

value: low temperature


title: T01

description: CK sample of 25 degree Celsius

value: 6th instar

value: control


title: Anaerobic, fumarate, rep1

city: Seoul

first: kucsbl

last: submitter

state: Seoul

address: Anamdong 5-ga, Seongbuk-gu

department: Department of Biotechnology

country: South Korea

institute: Korea University

description: AsuRPKM.csv

data_processing:
Low quality (Q<30) reads were trimmed at the 5' and 3' ends using the ShortRead package.
Quality controlled read data were aligned on the reference genome sequence using Bowtie2.
DESeq and edgeR package in the Bioconductor/R packages were used for making RPKM values.
Genome_build: Actinobacillus succinogenes strain 130Z (NCBI RefSeq ID: NC_009655.1)
Supplementary_files_format_and_content: CSV file of normalized RPKM value from edgeR for every samples

organism: Actinobacillus succinogenes 130Z

source_name: Mid-exponential growth phase

extract_protocol:
TruSeq RNA Sample Preparation Kit (Illumina) was used.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: oxygen

value: Anaerobic

tag: substrate

value: Fumarate with glycerol


title: T06

value: 13th instar


title: Male_02


title: T04


title: Input

city: Almada

first: Ana

middle: Rita

last: Grosso

address: Campus de Caparica

department: Departamento Ciências da Vida

country: Portugal

institute: Unidade de Ciências Biomoleculares Aplicadas (UCIBIO)/Faculdade de Ciências e Tecnologia - Universidade Nova de Lisboa

description: Input DNA

library_source: genomic

data_processing:
Read quality was assessed with the FastQC quality-control tool for high throughput sequence data (Babraham Bioinformatics).
High-throughput sequencing reads were aligned to the reference human genome (hg19 assembly) with Bowtie (Langmead et al. 2009) and filtering for uniquely aligned reads.
The detection of enriched regions were detected by MACS (Zhang et al. 2008), with a false-discovery rate of 0.05 and absolute change in density >1.5-fold relative to the control sample (input DNA).
For tag density profiles (bedgraphs), no-duplicated and uniquely mapped reads were extended in the 3’ direction to reach 150 nt with the Pyicos (Althammer et al. 2011). Only read counts that overlapped enriched regions identified above were considered.
Genome_build: hg19
Supplementary_files_format_and_content: Tag density profiles for enriched regions in bedgraph format.

molecule: genomic DNA

source_name: Input, MCF10A

growth_protocol: MCF10A cells were grown on monolayers as described in DOI:10.1016/S1046-2023(03)00032-X

extract_protocol:
Fast-ChIP protocol as mentioned in DOI: 10.1038/nsmb.2123, 10.1093/nar/gks1472
The samples were sequencing  by the Core Genomic Faciity - EMBL using Illumina standard protocols.
ChIP-seq libraries were sequenced as single-end 51-bp sequence tags using the standard Illumina pipeline

treatment_protocol: No special treatments

tag: cell line

value: MCF10A cells

tag: chip antibody


title: SRPK1 ChIP-seq - replicate 1

description: SRPK1 immunoprecipitated DNA

source_name: SRPK1, MCF10A

value: anti-SRPK1 (Abcam, catalog# ab90527)


title: T02

description: CK sample of 25 degree Celsius


title: HIV-1 virion (produced in CEM-SS) total small RNA replicate 2

city: Durham

first: Edward

middle: Matthew

last: Kennedy

state: NC

address: Department of Molecular Genetics and Microbiology, Box 3025

department: Molecular Genetics and Microbiology

country: USA

institute: Duke University

description: Total small RNA

data_processing:
Initial reads were quality filtered with Cassava 1.8.2.
Reads>15 nt were collapsed into FASTA format with the FASTX-Toolkit (http://hannonlab.cshl.edu/fastx_toolkit/index.html) using the following pipeline: fastq_quality_filter –Q 33|  fastx_clipper -a TruSeq-Index# -l 15 –c | fastq_to_fasta –Q 33 | fastx_collapser.
All reads were then subject to alignment using Bowtie v.0.12.7 (Langmead et al., 2009) with the following options: –a --best --strata –m 25.  The reads were sequentially aligned and filtered to adapter sequences, miRBase v. 20 annotations of miRNA hairpins or mature miRNA sequences, Ensembl noncoding RNA database release 70, human functional RNA database version 3.4, the viral sequence of interest, and finally human genome 19.  Reverse complement alignments were only allowed for the human and viral genomes.  No mismatches or were allowed except for the human genome for which two mismatches were allowed.
miRNA sequences were given a -5p or -3p designation if they aligned to the 5′ or 3′ stem region, respectively, of the miRNA precursor, as annotated in miRbase.  miRNAs that represented ≥0.1% of the total miRNA population in both cellular and virion libraries were considered significant.
Genome_build: human genome 19
Supplementary_files_format_and_content: Processed data are in a tab-delineated BED format presented as chromosome/gene, starting location for aligned read relative to the start of the gene beginning at 1, ending position of the aligned read, aligned reads per million of total library reads, and the strand (+ or -) that the read aligns to.

source_name: HIV-1 virions produced in the CEM-SS cell line (#776, AIDS Reagent Program)

growth_protocol: Cells were maintained in RPMI supplemented with 10% FBS and Gentamicin.

extract_protocol:
Total RNA was isolated using TRIzol for cells and purified virons.
All small RNA input was processed with the TruSeq Small RNA Sample Preparation Kit (Illumina). Adapter-ligated RNA was reverse transcribed using SuperScript III (Life Technologies) and amplified using GoTaq green PCR master mix (Promega) with TruSeq 3′ indices.  DNA bands corresponding to ~145 bp were isolated by gel electrophoresis on 10% polyacrylamide gels.

tag: illumina index number

value: CEM-SS


title: BLCL-10

city: Montreal

first: Diana

middle: Paola

last: Granados

state: Quebec

address: C.P. 6128, succursale Centre-ville

department: Medecine

country: Canada

institute: IRIC

data_processing:
Basecalls performed using CASAVA 1.8.2
Sequenced reads were mapped to the reference genome using Tophat version 2.0.10
Cufflinks 2.2.1 was then run on the output sorted BAM file in addition to the Ensembl gtf file in order to obtain FPKM (Fragment Per Kilobase of transcript per Million mapped reads) values for all known transcripts
Genome_build: GRCh37.75
Supplementary_files_format_and_content: Tab-delimited text files include FPKM values of transcript isoforms for each sample in the Cufflinks FPKM_tracking file format.

source_name: EBV-infected B lymphoblasts

growth_protocol: Peripheral blood mononuclear cells (PBMCs) were isolated from blood samples of 2 HLA-identical female siblings. B lymphoblastoid cell lines (B-LCLs) were derived from PBMCs with Ficoll-Paque Plus (Amersham )and infected with Epstein-Barr virus and as described by Tosato et al (Tosato,G. & Cohen,J.I. Generation of Epstein-Barr Virus (EBV)-immortalized B cell lines. Curr. Protoc. Immunol. Chapter 7, Unit (2007)). Established B-LCLs were maintained in RPMI 1640 medium supplemented with 10% fetal bovine serum, 25 mM of HEPES, 2mM L-glutamine and penicillin-streptomycin. Cells were seeded at a density of 0.5 million cells per ml 18 hours before RNA extraction.

extract_protocol:
Total RNA was isolated from 5 million B-LCLs using  RNeasy mini kit (Qiagen) (samples 1-2) or TRizol RNA reagent (Life Technologies®)(samples 3 - 4) including DNase I treatment (Qiagen®) according to the manufacturer’s instructions. Total RNA was quantified using the NanoDrop™ 2000 (Thermo Scientific®) and RNA quality was assessed with the 2100 Bioanalyzer™ (Agilent Technologies®).
TruSeq RNA Sample Preparation (Illumina)

treatment_protocol: Cells were seeded at a density of 0.5 million cells per ml, 20 ml in total in a T75 flask, 18 hours before RNA extraction. Cells were not treated.

value: EBV-infected human B lymphoblastoid cell line

tag: source tissue

value: peripheral blood mononuclear cells

tag: passages

tag: host sex


title: HIV-1 virion (produced in CEM-SS) total small RNA replicate 1

description: Total small RNA


title: Female_02

value: CD14-Positive Monocytes cells


title: L4 larval P-granule(-); UNC-119::GFP(-) gonads #3

city: Santa Cruz

first: Susan

last: Strome

state: CA

address: 1156 High St

department: MCD Biology

institute: UC Santa Cruz

data_processing:
Raw sequences were mapped to transcriptome version WS220 using Tophat2.  Only reads with one unique mapping were allowed; otherwise default options were used.  Reads mapping to ribosomal RNAs were removed from further analysis.
The Bioconductor package DESeq2 was used for differential expression analysis. A Benjamini-Hochberg multiple hypothesis corrected p-value cutoff of 0.05 was used as a significance cutoff.
Normalized read counts per gene were obtained with DESeq2, size factor normalization was used.
Genome_build: WS220
Supplementary_files_format_and_content: Wormbase ID, WS220 Gene names, gene sequence, transcript type, gene category designation, and Differential expression comparison data (Mean expression, log2(Fold Change), BH adjusted p-value) and read counts per gene averaged over replicates for control samples, RNAi samples and mutant samples.

organism: Caenorhabditis elegans

source_name: dissected gonads

growth_protocol: C. elegans strains were maintained at 15° or 20° on OP50 bacteria as described (Brenner 1974).

extract_protocol:
Total RNA was extracted using Trizol reagent and ribosomal RNA was depleted using the NEBNext rRNA Depletion Kit (Human/Mouse/Rat) (Cat. #E6310).
Libraries were constructed using the NEBNext Ultra RNA Library Prep Kit for Illumina sequencing (Cat. #E7530). Libraries were sequenced at the Vincent J. Coates Genomics Sequencing Laboratory at UC Berkeley using the Illumina HiSeq 2000, 2500, or 4000 platforms.

treatment_protocol: For experiments using RNAi, synchronized unc-119::gfp L1 worms (P0) were placed on HT115 bacteria containing empty RNAi vector (control) or expressing double strand RNA that targets P granules (i.e., pgl-1, pgl-3, glh-1, and glh-4) or targets pgl-1 or pgl-1; pgl-3 (Updike et al. 2014; Campbell and Updike 2015) at 24° (Day 0).  Three days later, F1 larvae were transferred to new control or P-granule RNAi plates and returned to 24°.  The majority of these F1 larvae were at the L1 or L2 stage at the time of transfer.  The next day (Day 4), L4 worms were visually identified by their size and the presence of a white crescent at their vulva.  Gonads were dissected from L4 control worms and L4 P-granule RNAi worms and cut at the gonad bend with 30½ gauge needles in egg buffer (27.5 mM HEPES, 130 mM NaCl, 2.2 mM MgCl2, 2.2 mM CaCl2, and 0.528 mM KCl) containing 0.5% Tween-20 and 1 mM levamisole.  Sterility could not be assessed in L4 worms.  However, greater than 80% sterility was observed in P-granule RNAi adults that were aged from the population of worms from which L4 gonads were collected.  For Day 1 adults, worms were allowed to age one day past the L4 stage and then dissected.  For Day 2 adults, worms were allowed to age two days past the L4 stage and then dissected.  In Day 2 adults, gonads that visually expressed the unc-119::gfp transgene were collected as a separate sample from those that did not detectably express unc-119::gfp.  For adult control gonads, gonads were dissected from fully fertile worms and cut at the gonad bend.  For adult P-granule RNAi worms, gonads were collected only from sterile worms, as judged by the lack of embryos in the uterus.  Large P-granule RNAi gonads were cut at the gonad bend; stunted P-granule RNAi gonads were collected in their entirety.  Once dissected, gonad arms were collected in Trizol, and total RNA was extracted.  For control samples, 75 to 100 gonads were collected per replicate.  For P-granule RNAi samples, 125 to 150 gonads were collected per replicate. Three biological replicates were performed for each treatment and time point. For experiments using P-granule mutants, wild-type (control), pgl-1 single mutants, or glh-1 single mutant adults (isolated from glh-1(gk100)/hT2g balanced parents) were allowed to lay eggs overnight on OP50 bacteria at 24°.  The day after the egg lay, the adults were removed and the F1 progeny were returned to 24° for two days.  Day 1 adult gonads were dissected, collected into Trizol, and total RNA was extracted.  Only distal gonad arms were collected from fertile wild-type worms or sterile P-granule mutant worms, as judged by the lack of embryos in the uterus.  Three biological replicates were performed for the wild-type and pgl-1 samples, and two biological replicates were performed for the glh-1 sample.

tag: strain

tag: genotype

value: edIs6[unc-119p::unc-119::gfp + rol-6] IV

tag: RNAi treatment

value: P-granule RNAi

tag: temperature


title: Female_01

tag: cell type

tag: gender

value: female

value: Healthy individuals


title: Male_01

value: male

tag: age

tag: disease status


title: Day 2 adult control gonads #1

value: empty vector RNAi


title: SRPK2 ChIP-seq - replicate 1

description: SRPK2 immunoprecipitated DNA

source_name: SRPK2, MCF10A

value: anti-SRPK2 (Bethyl Labs, catalog# A302-467A)


title: Tcf4-sh1-RNA-seq-rep1

city: Rotterdam

first: Raymond

middle: A

last: Poot

address: Wytemaweg 80

department: Cell Biology

country: Netherlands

institute: Erasmus MC

description: RNA-seq NS5 Tcf4 KD shRNA1 replicate1

data_processing:
RNA-seq was mapped against mouse reference NCBIM37.67 (mm9) using Tophat50 v2.0.11 with default settings and a segment length of 20
The aligned exon reads were normalized and differential expression was calculated using Bioconductor DESeq2 package in R
Genome_build: mm9
Supplementary_files_format_and_content: 150814_RNAseq_all_GEO.txt: Processed data is a tabular text file that contains all genes according to ensembl  NCBIM37.67 (mm9) with the RPKM values based on the RNA-seq NS5 wt RNA-seq, corrosponding standard deviation of RPKMs, FC upon shRNA mediated KD of Tcf4 compared to mock KD, to FC corresponding adjusted p.values and gene symbol

molecule: polyA RNA

organism: Mus musculus

source_name: NS5 Neural Stem Cell

growth_protocol: Mouse neural stem cell line NS-5 was grown on gelatin coated dishes. Cells were cultured in N2 medium.

extract_protocol:
NSCs were harvested after 44 hrs after electroporation using 1mL of Trizol. This was mixed thoroughly with 200µl chloroform and centrifuged at max. speed for 15 minutes. The water phase was collected and subsequently the poly(A) RNA was isolated from this sample using the RNeasy kit (Qiagen). The RNA was retrieved from the column in 30 µl RNase-free water (Qiagen)
RNA was isolated using the RNeasy kit (Qiagen), tested for quality with the Bioanalyzer and prepared using the TruSeq RNA sample prep kit v2

treatment_protocol: Both the Tcf4 knockdown and the control knockdown were performed in triplo: 3x Tcf4-sh1 construct and 3x control shRNA construct (Dharmacon). For each sample 6x10^6 NSCs were used. Cells were grown to 80% confluency and washed with PBS. Next, they were detached using Accutase, PBS was added and cells were centrifuged for 5min at 1000rpm. The PBS was aspirated and 10ml of N2 medium was used to resuspend the cell pellet. After measuring the concentration, 6x10^6 cells were divided per tube per condition (max. 2 conditions each time). Then the cells were pelleted again and 200µl Nucleofection buffer was added to each tube. Per condition, 6µg (1µg/1*10^6) of vector DNA was added and mixed with the pellet and buffer, to create a cell suspension. This was next divided over 2 amaxa cuvettes (100µl each). Electroporation was performed using the Amaxa program T-30. Medium was added after the shock as soon as possible, and the cells were subsequently seeded on a gelatin-coated 60mm culture dish. This sequence was repeated for the remaining samples. The medium was refreshed after 18 hrs and puromycin (2ug/mL) was added.

value: NSC

tag: genotype/variation

value: Tcf4 knockdown


title: Tcf4-sh1-RNA-seq-rep3

description: RNA-seq NS5 Tcf4 KD shRNA1 replicate3


title: SRPK2 ChIP-seq - replicate 2

description: SRPK2 immunoprecipitated DNA


title: Day 1 adult P-granule(-); UNC-119::GFP(-) gonads #1


title: B-LCL-2.2 [re-analysis]

description:
reanalysis of GSM1186812
This is duplicated sample record of GSM1186812 for the convenient retrieval of the complete raw data from SRA.


title: L4 larval P-granule(-); UNC-119::GFP(-) gonads #1


title: B-LCL-2.1 [re-analysis]

description:
reanalysis of GSM1186811
This is duplicated sample record of GSM1186811 for the convenient retrieval of the complete raw data from SRA.


title: L4 larval control gonads #3


title: L4 larval control gonads #2


title: V5 ChIP-seq in NS5 Neural Stem Cells

description: ChIP against V5 in NS5 cells expressing no V5 tag (control for NS5 NSCs expressing Tcf4-V5)

data_processing:
Filtering against low complexity reads using prinseq-lite using options -lc_method dust and -lc_treshhold 7
mapping reads using bowtie 0.12.7 the options -n 2 -l 36 -e 70 -k 1 --best
Picard-tools-1.81 MarkDuplicates was used to remove the duplicates from the sam/bam file
MACS 1.4.2 was used for generating wig files using default settings. The corresponding IgG-mock was used as control input file and was scaled to the treatment file by MACS. The .wig files from MACS were converted to .bw files using UCSC WigToBigWig
Genome_build: mm9
Supplementary_files_format_and_content: bigwig files

growth_protocol: Mouse neural stem cell line NS-5 was grown on gelatin coated dishes. Cells were cultured in N2 medium (Euromed (Clontech) with 1x N2 supplement (Gibco/Life Technologies), 1mM glutamine, BSA (50g/mL), Insulin (20g/mL), EGF and FGF2 both at 10 ng/mL)

extract_protocol: Extraction protocol For each ChIP 1.5*10^8 NSCs were used. NS-5 NSCs expressing no V5-tag were used for the control experiment. NSCs were crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Fisher Scientific) for 45 min at RT, washed three times (PBS + 1mM PMSF) and then crosslinked for 10min with 1% formaldehyde. This reaction was quenched by addition of 1/20 volume of 2.5M glycine for 10min at RT. Cells were washed twice with cold 1x PBS+1mM PMSF, collected by centrifugation, and cell pellets frozen with N2(l). Cells were resuspended in LB1 (50mM Hepes-KOH pH7.5, 140mM NaCl, 1mM EDTA, 10% Glycerol, 0.5% NP40, 0.25% Triton-X-100 . After 10 minutes incubation, cells were pelleted by centrifugation and resuspended in LB2 (10 mM Tris-HCl, pH8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA. After 10 minutes incubation, cells were pelleted and resuspended in freshly prepared LB3 (10mM Tris-HCl pH8.0, 100mM NaCl, 1mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified that the DNA fragment size was between 100 and 1000bp. V5-Agarose beads were blocked 1 hr with 0.5 mg/ml BSA. The chromatin was incubated overnight with the blocked V5-Agarose beads. The following day the beads were washed 5 times with RIPA buffer and eluted with elution buffer (50mM Tris-HCl pH8, 10mM EDTA and 1% SDS) for 30 minutes. Samples were de-cross-linked overnight at 65C and treated the following day with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol and resuspended in water. The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC. Cluster generation was performed using the Illumina HiSeq 2000 platform.

treatment_protocol: Untreated

value: NS5 neural stem cell

value: mouse anti-V5 Agarose affinity gel antibody (sigma A7345)


title: NS5-untreated-RNA-seq-rep2

description: RNA-seq NS5 wt RNA-seq replicate2

data_processing:
RNA-seq was mapped against mouse reference NCBIM37.67 (mm9) using Tophat50 v2.0.11 with default settings and a segment length of 20
The aligned exon reads were normalized and differential expression was calculated using Bioconductor DESeq2 package in R
Genome_build: mm9
Supplementary_files_format_and_content: 150814_RNAseq_all_GEO.txt: Processed data is a tabular text file that contains all genes according to ensembl  NCBIM37.67 (mm9) with the RPKM values based on the RNA-seq NS5 wt RNA-seq, corrosponding standard deviation of RPKMs, FC upon shRNA mediated KD of Tcf4 compared to mock KD, to FC corresponding adjusted p.values and gene symbol
Supplementary_files_format_and_content: wt_NSC_RNAseq_RPKM_protein_coding.txt

treatment_protocol: untreated

value: wild-type


title: Npas3 ChIP-seq in NS5 Neural Stem Cells

description: ChIP against Npas3 in NS5 neural stem cells

extract_protocol: For each ChIP 1.5*10^8 NSCs were used. As a control, the same experiment was performed in parallel using rabbit IgG. NSCs were crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Fisher Scientific) for 45 min at RT, washed three times (PBS + 1mM PMSF) and then crosslinked for 10min with 1% formaldehyde. This reaction was quenched by addition of 1/20 volume of 2.5M glycine for 10min at RT. Cells were washed twice with cold 1x PBS+1mM PMSF, collected by centrifugation, and cell pellets frozen with N2(l). Cells were resuspended in LB1 (50mM Hepes-KOH pH7.5, 140mM NaCl, 1mM EDTA, 10% Glycerol, 0.5% NP40, 0.25% Triton-X-100 . After 10 minutes incubation, cells were pelleted by centrifugation and resuspended in LB2 (10 mM Tris-HCl, pH8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA. After 10 minutes incubation, cells were pelleted and resuspended in freshly prepared LB3 (10mM Tris-HCl pH8.0, 100mM NaCl, 1mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified whether the DNA fragment size was between 100 and 1000bp. The DNA concentration was measured in order to take a volume of chromatin solution containing 300µg DNA per ChIP, that was next diluted 1 to 5 with ChIP dilution buffer (20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100 and 1x CEF protease inhibitor). Equilibration of preblocked protein G beads (Millipore) was performed by washing them 3x with 1 ml of ChIP dilution buffer. To lower unspecific binding, the diluted chromatin was incubated with beads without the presence of antibody, rotating for 30 min at 4 C. The beads were discarded and the ‘precleared’ chromatin was incubated overnight at 4C with 15µg of antibody. The next day, 60µl of beads was added to this mixture and rotated for 3hrs at 4 C. The supernatant was discarded and the beads were washed subsequently by rotation for 5min for each wash: 1x ChIP dilution buffer, 1x high salt buffer (20mM Tris-HCl pH8, 500mM NaCl, 2mM EDTA, 1%Triton X-100, 0.1% SDS),1x LiCl buffer (10mM Tris-HCl pH8, 250mM LiCl, 1mM EDTA, 0.5% NP-40, 0.5% deoxycholate) and twice in TE (10 mM Tris-HCl pH8, 1mM EDTA). Elution was performed two times by incubation in 250 l of pre-warmed (37C) freshly prepared elution buffer (0.1M NaHCO3, 1% SDS)  for 15 minutes at RT. Supernatants were combined and centrifuged to get rid of beads. The crosslinks were reversed by incubation overnight at 65C. The following day, the samples were treated with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol. 	The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC.  Cluster generation was performed using the Illumina HiSeq 2000 platform.

value: NS5 Neural Stem cell

value: Rabbit Anti-Npas3 (Sigma:HPA002892)


title: NS5-untreated-RNA-seq-rep1

description: RNA-seq NS5 wt RNA-seq replicate1


title: SRPK1 ChIP-seq - replicate 2

description: SRPK1 immunoprecipitated DNA

value: MCF10A cells

value: anti-SRPK1 (Abcam, catalog# ab90527)


title: NS5-untreated-RNA-seq-rep3

description: RNA-seq NS5 wt RNA-seq replicate3


title: Tcf4-sh1-RNA-seq-rep2

description: RNA-seq NS5 Tcf4 KD shRNA1 replicate2


title: KO-HSC

city: Shanghai

first: Qiumin

last: Le

address: 130 Dong'an Road

institute: Fudan University

description: PCR amplified cDNA library

data_processing:
Base calling was performed by Wuxi Apptec.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm9 by TopHat v1.4.0 with  -p 8  -a 8 -m 2 -i 30 -I 500000 -g 20 -G /galaxy-repl/main/files/006/992/dataset_6992006.dat --library-type fr-unstranded --max-insertion-length 3 --max-deletion-length 3 --coverage-search --min-coverage-intron 10 --max-coverage-intr.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each Sample

source_name: hematopoietic stem cells

extract_protocol:
Sorted cells were individually transferred into lysis buffer with a mouth pipette and reverse transcription reaction was directly on the whole-cell lysate. Poly (dT) primer was used to specifically reverse transcribe mRNA from the lysate. Terminal deoxynucleotidyl transferase was used to add a poly(A) tail to the 3’ end of the first-strand cDNA. The unincorporated primer was digested by exonuclease and 20 + 10 cycles of PCR was applied to amplify the cDNA.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: WT and GRK6-/- mice were sacrificed. 13-15 sorted bone marrow cells of HSC (Lin-c-kit+sca-1+)  as well as CLP (Lin-c-kit+sca-1+Il-7Ra+) from 3-4 mice were picked and transferred into lysis buffer with a mouth pipette.

value: Lin-c-kit+sca-1+ cells

tag: tissue

value: bone marrow

value: GRK6 knockout


title: mock-RNA-seq-rep3

description: RNA-seq NS5 mock shRNA replicate3

value: control knockdown


title: HSF_FPLD-p4_expression

city: Oslo

first: Philippe

last: Collas

address: PO Box 1112 Blindern

department: Institute of Basic Medical Sciences

country: Norway

institute: University of Oslo

data_processing:
ChIP-seq reads were aligned to the hg19 reference genome using bowtie2 version 2.1.0
Duplicate reads were removed using picard MarkDuplicates
ChIP and Input experiments were ensured to have equal read depth by downsampling the deeper sequenced experiment using picard DownsampleSam
Peak calling was done using EDD version 1.1.7
RNA-seq was processed using the tuxedo protocol
Genome_build: hg19
Supplementary_files_format_and_content: peaks.bed files with four columns: chromosome, start position, end position, score; fpkm_tracking files with 13 columns tracking_id, class_code, nearest_ref_id, gene_id, gene_short_name, tss_id, locus, length, coverage, FPKM, FPKM_conf_lo, FPKM_conf_hi, FPKM_status

source_name: primary dermal fibroblasts

growth_protocol: Cells were cultured in DMEM/F12 containing 13% FCS, 2 ng/ml basic fibroblast growth factor and antibiotics. Cells were exponentially growing and used at passage 5-7. Cells were obtained with Norwegian Ethics Committee Approval REK2617A.

extract_protocol:
Cells (107 per ChIP) were cross-linked in suspension for 10 min in PBS containing 1% formaldehyde before quenching with 1.25 mM glycine. Cells were lysed for 30 min at 4oC on a rotator in RIPA buffer (140 mM NaCl, 10 mM Tris-HCl, pH 8.0, 1 mM EDTA, 0.5 mM EGTA, 1% Triton X-100, 0.1% SDS, 0.1% Na-deoxycholate, 1 mM PMSF, 1x protease inhibitor cocktail) adjusted to 1% SDS, and sonicated for 3 times 15 min in a Bioruptor (Diagenode) with 30 sec ON/OFF at high power to generate chromatin fragments of ~200-400 base pairs (bp). After sedimentation, chromatin (supernatant from 107 cell-equivalents) was diluted 10 times in RIPA buffer without SDS, and incubated on a rotator overnight at 4oC with 50 µg anti-lamin A/C antibody (Santa Cruz sc-7292) pre-coupled to magnetic Dynabeads Protein G (Invitrogen). An irrelevant mouse IgG was used as control. ChIP material was collected and washed 3 times in 1 ml of ice-cold RIPA buffer without protease inhibitors. The crosslink was reversed and DNA eluted for 6 h on a shaker at 37oC in elution buffer (50 mM NaCl, 20 mM Tris-HCl, pH 7.5, 5 mM EDTA, 1% SDS) containing 0.5 µg/ml RNase A and 2 µg/ml Proteinase K). DNA was purified as described, processed for library preparation.
The sequencing library was prepared according to the Illumina protocol for the HSeq2500 at the Norwegian Sequencing Center.

treatment_protocol: Cells were cultured as described under Growth protocol. Cells were cultured to confluency before harvesting to ensure consistency of cell cycle stages between the two cell types.

value: lipodystrophy patient with LMNA p.R482W mutation

value: Fibroblasts


title: mock-RNA-seq-rep2

description: RNA-seq NS5 mock shRNA replicate2


title: BLCL-3

value: EBV-infected human B lymphoblastoid cell line

value: peripheral blood mononuclear cells

value: male


title: mock-RNA-seq-rep1

description: RNA-seq NS5 mock shRNA replicate1

value: NSC


title: IgG control ChIP-seq in NS5 Neural Stem Cells for Smad4

description: Goat-IgG mock control for Smad4 ChIP in NS5 cells

extract_protocol: For each ChIP 1.5*10^8 NSCs were used. As a control, the same experiment was performed in parallel using goat IgG. NSCs were crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Fisher Scientific) for 45 min at RT, washed three times (PBS + 1mM PMSF) and then crosslinked for 10min with 1% formaldehyde. This reaction was quenched by addition of 1/20 volume of 2.5M glycine for 10min at RT. Cells were washed twice with cold 1x PBS+1mM PMSF, collected by centrifugation, and cell pellets frozen with N2(l). Cells were resuspended in LB1 (50mM Hepes-KOH pH7.5, 140mM NaCl, 1mM EDTA, 10% Glycerol, 0.5% NP40, 0.25% Triton-X-100 . After 10 minutes incubation, cells were pelleted by centrifugation and resuspended in LB2 (10 mM Tris-HCl, pH8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA. After 10 minutes incubation, cells were pelleted and resuspended in freshly prepared LB3 (10mM Tris-HCl pH8.0, 100mM NaCl, 1mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified whether the DNA fragment size was between 100 and 1000bp. The DNA concentration was measured in order to take a volume of chromatin solution containing 300µg DNA per ChIP. Per sample, 150µl prot-G Dynabead solution (Life Technologies) was used. Beads were blocked for 1hr at RT with 0.5mg/ml BSA and 0.2mg/ml sonicated salmon sperm DNA in IP buffer (20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100) After blocking, the beads were washed 3 times in IP buffer. To lower unspecific binding, the chromatin was incubated with beads without the presence of antibody, rotating for 30 min at 4 C. The beads were discarded and the ‘precleared’ chromatin was incubated overnight at 4 C with 15µg of antibody. The next day, 150µl of blocked beads was added to this mixture and rotated for 3hrs at 4 C. The supernatant was discarded and the beads were washed subsequently by rotation for 10min for each wash: 2x low salt buffer(20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100), 2x high salt buffer (20mM Tris-HCl pH8, 500mM NaCl, 2mM EDTA, 1%Triton X-100, 0.1% SDS), 2x LiCl buffer (10mM Tris-HCl pH8, 250mM LiCl, 1mM EDTA, 0.5% NP-40, 0.5% deoxycholate) and finally twice in TE (10 mM Tris-HCl pH8, 1mM EDTA). Immune complexes were eluted in 210ul elution buffer (1% SDS, 0.1 M NaHCO3) for 1 h at 65 °C while softly shaking. The crosslinks were reversed by incubation overnight at 65C. The following day, the samples were treated with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol. The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC. Cluster generation was performed using the Illumina HiSeq 2000 platform.

value: NS5 Neural Stem Cell

value: Goat IgG (Santa Cruz:sc2028)


title: HSF_FPLD-p2_expression


title: HSF_FPLD-p1_Input


title: Smad4 ChIP-seq in NS5 Neural Stem Cells

description: ChIP against Smad4 in NS5 cells

extract_protocol:
For each ChIP 1.5*10^8 NSCs were used. As a control, the same experiment was performed in parallel using goat IgG. NSCs were crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Fisher Scientific) for 45 min at RT, washed three times (PBS + 1mM PMSF) and then crosslinked for 10min with 1% formaldehyde. This reaction was quenched by addition of 1/20 volume of 2.5M glycine for 10min at RT. Cells were washed twice with cold 1x PBS+1mM PMSF, collected by centrifugation, and cell pellets frozen with N2(l). Cells were resuspended in LB1 (50mM Hepes-KOH pH7.5, 140mM NaCl, 1mM EDTA, 10% Glycerol, 0.5% NP40, 0.25% Triton-X-100 . After 10 minutes incubation, cells were pelleted by centrifugation and resuspended in LB2 (10 mM Tris-HCl, pH8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA. After 10 minutes incubation, cells were pelleted and resuspended in freshly prepared LB3 (10mM Tris-HCl pH8.0, 100mM NaCl, 1mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified whether the DNA fragment size was between 100 and 1000bp. The DNA concentration was measured in order to take a volume of chromatin solution containing 300µg DNA per ChIP. Per sample, 150µl prot-G Dynabead solution (Life Technologies) was used. Beads were blocked for 1hr at RT with 0.5mg/ml BSA and 0.2mg/ml sonicated salmon sperm DNA in IP buffer (20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100) After blocking, the beads were washed 3 times in IP buffer. To lower unspecific binding, the chromatin was incubated with beads without the presence of antibody, rotating for 30 min at 4 C. The beads were discarded and the ‘precleared’ chromatin was incubated overnight at 4 C with 15µg of antibody. The next day, 150µl of blocked beads was added to this mixture and rotated for 3hrs at 4 C. The supernatant was discarded and the beads were washed subsequently by rotation for 10min for each wash: 2x low salt buffer(20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100), 2x high salt buffer (20mM Tris-HCl pH8, 500mM NaCl, 2mM EDTA, 1%Triton X-100, 0.1% SDS), 2x LiCl buffer (10mM Tris-HCl pH8, 250mM LiCl, 1mM EDTA, 0.5% NP-40, 0.5% deoxycholate) and finally twice in TE (10 mM Tris-HCl pH8, 1mM EDTA). Immune complexes were eluted in 210ul elution buffer (1% SDS, 0.1 M NaHCO3)  for 1 h at 65 °C while softly shaking. The crosslinks were reversed by incubation overnight at 65C. The following day, the samples were treated with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol.
The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC.  Cluster generation was performed using the Illumina HiSeq 2000 platform.

value: Goat Smad4 antibody (R&D Systems:AF2097)


title: HSF_FPLD-p3_expression


title: IgG control ChIP-seq in NS5 Neural Stem Cells for Olig2

description: rabbit-IgG mock control for Olig2 ChIP in NS5 cella

extract_protocol: For each ChIP 1.5*10^8 NSCs were used. As a control, the same experiment was performed in parallel using rabbit IgG. NSCs were washed 3 times with PBS, crosslinked with 1/10 volume of fresh 11% buffered formaldehyde solution for 12 min., quenched with 1/20 volume of 2.5M Glycine for 5 min, washed with ice-cold PBS and cell pellets frozen with N2(l) and resuspended and washed 2 times in ice-cold cell lysis buffer (10mM Tris-Cl pH 7.5, 10mM NaCl, 3 mM MgCl2, 0.5% NP40) and cell pellet resuspended in lysis buffer with 1mM CaCl2 and 4% NP40 and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified whether the DNA fragment size was between 100 and 1000bp. The DNA concentration was measured in order to take a volume of chromatin solution containing 300µg DNA per ChIP, that was next diluted 1 to 5 with ChIP dilution buffer (20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100 and 1x CEF protease inhibitor). Equilibration of preblocked protein G beads (Millipore) was performed by washing them 3x with 1 ml of ChIP dilution buffer. To lower unspecific binding, the diluted chromatin was incubated with beads without the presence of antibody, rotating for 30 min at 4 C. The beads were discarded and the ‘precleared’ chromatin was incubated overnight at 4 C with 15µg of antibody. The next day, 60µl of beads was added to this mixture and rotated for 3hrs at 4 C. The supernatant was discarded and the beads were washed subsequently by rotation, 5min for each wash: 1x ChIP dilution buffer, 1x high salt buffer (20mM Tris-HCl pH8, 500mM NaCl, 2mM EDTA, 1%Triton X-100, 0.1% SDS),1x LiCl buffer (10mM Tris-HCl pH8, 250mM LiCl, 1mM EDTA, 0.5% NP-40, 0.5% deoxycholate) and twice in TE (10 mM Tris-HCl pH8, 1mM EDTA). Elution was performed two times by incubation in 250 l of pre-warmed (37C) freshly prepared elution buffer (0.1M NaHCO3, 1% SDS) for 15 minutes at RT. Supernatants were combined and centrifuged to get rid of beads. The crosslinks were reversed by incubation overnight at 65C. The following day, the samples were treated with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol. The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC. Cluster generation was performed using the Illumina HiSeq 2000 platform.

value: NS5 Neural Stem Cells

value: Rabbit IgG  (Santa Cruz:sc2027)


title: HeLa_L647R_LMNA

source_name: cervical cancer (HeLa cells)

value: anti-Flag, Sigma F-1804

value: HeLa


title: Olig2 ChIP-seq in NS5 Neural Stem Cells

description: ChIP against Olig2  in NS5 cells

extract_protocol:
For each ChIP 1.5*10^8 NSCs were used. As a control, the same experiment was performed in parallel using rabbit IgG. NSCs were washed 3 times with PBS, crosslinked with 1/10 volume of fresh 11% buffered formaldehyde solution for 12 min., quenched with 1/20 volume of 2.5M Glycine for 5 min, washed with ice-cold PBS and cell pellets frozen with N2(l) and resuspended and washed 2 times in ice-cold cell lysis buffer (10mM Tris-Cl pH 7.5, 10mM NaCl, 3 mM MgCl2, 0.5% NP40) and cell pellet resuspended in lysis buffer with 1mM CaCl2 and 4% NP40 and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified whether the DNA fragment size was between 100 and 1000bp. The DNA concentration was measured in order to take a volume of chromatin solution containing 300µg DNA per ChIP, that was next diluted 1 to 5 with ChIP dilution buffer (20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100 and 1x CEF protease inhibitor). Equilibration of preblocked protein G beads (Millipore) was performed by washing them 3x with 1 ml of ChIP dilution buffer. To lower unspecific binding, the diluted chromatin was incubated with beads without the presence of antibody, rotating for 30 min at 4 C. The beads were discarded and the ‘precleared’ chromatin was incubated overnight at 4 C with 15µg of antibody. The next day, 60µl of beads was added to this mixture and rotated for 3hrs at 4 C. The supernatant was discarded and the beads were washed subsequently by rotation, 5min for each wash: 1x ChIP dilution buffer, 1x high salt buffer (20mM Tris-HCl pH8, 500mM NaCl, 2mM EDTA, 1%Triton X-100, 0.1% SDS),1x LiCl buffer (10mM Tris-HCl pH8, 250mM LiCl, 1mM EDTA, 0.5% NP-40, 0.5% deoxycholate) and twice in TE (10 mM Tris-HCl pH8, 1mM EDTA). Elution was performed two times by incubation in 250 l of pre-warmed (37C) freshly prepared elution buffer (0.1M NaHCO3, 1% SDS) for 15 minutes at RT. Supernatants were combined and centrifuged to get rid of beads. The crosslinks were reversed by incubation overnight at 65C. The following day, the samples were treated with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol.
The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC.  Cluster generation was performed using the Illumina HiSeq 2000 platform.

value: Rabbit Anti-Olig2 (Milipore:AB9610)


title: IgG control ChIP-seq in NS5 Neural Stem Cells for Npas3

description: Mock control-IgG-control for Npas3 ChIP in NS5 cells

growth_protocol: Mouse neural stem cell line NS-5 was grown on gelatin coated dishes. Cells were cultured in N2 medium (Euromed (Clontech) with 1x N2 supplement (Gibco/Life Technologies), 1mM glutamine, BSA (50g/mL), Insulin (20g/mL), EGF and FGF2 both at 10 ng/mL).

extract_protocol: For each ChIP 1.5*10^8 NSCs were used. As a control, the same experiment was performed in parallel using rabbit IgG. NSCs were crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Fisher Scientific) for 45 min at RT, washed three times (PBS + 1mM PMSF) and then crosslinked for 10min with 1% formaldehyde. This reaction was quenched by addition of 1/20 volume of 2.5M glycine for 10min at RT. Cells were washed twice with cold 1x PBS+1mM PMSF, collected by centrifugation, and cell pellets frozen with N2(l). Cells were resuspended in LB1 (50mM Hepes-KOH pH7.5, 140mM NaCl, 1mM EDTA, 10% Glycerol, 0.5% NP40, 0.25% Triton-X-100 . After 10 minutes incubation, cells were pelleted by centrifugation and resuspended in LB2 (10 mM Tris-HCl, pH8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA. After 10 minutes incubation, cells were pelleted and resuspended in freshly prepared LB3 (10mM Tris-HCl pH8.0, 100mM NaCl, 1mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified whether the DNA fragment size was between 100 and 1000bp. The DNA concentration was measured in order to take a volume of chromatin solution containing 300µg DNA per ChIP, that was next diluted 1 to 5 with ChIP dilution buffer (20mM Tris-HCl pH8, 150mM NaCl, 2mM EDTA, 1% Triton X-100 and 1x CEF protease inhibitor). Equilibration of preblocked protein G beads (Millipore) was performed by washing them 3x with 1 ml of ChIP dilution buffer. To lower unspecific binding, the diluted chromatin was incubated with beads without the presence of antibody, rotating for 30 min at 4 C. The beads were discarded and the ‘precleared’ chromatin was incubated overnight at 4C with 15µg of antibody. The next day, 60µl of beads was added to this mixture and rotated for 3hrs at 4 C. The supernatant was discarded and the beads were washed subsequently by rotation for 5min for each wash: 1x ChIP dilution buffer, 1x high salt buffer (20mM Tris-HCl pH8, 500mM NaCl, 2mM EDTA, 1%Triton X-100, 0.1% SDS),1x LiCl buffer (10mM Tris-HCl pH8, 250mM LiCl, 1mM EDTA, 0.5% NP-40, 0.5% deoxycholate) and twice in TE (10 mM Tris-HCl pH8, 1mM EDTA). Elution was performed two times by incubation in 250 l of pre-warmed (37C) freshly prepared elution buffer (0.1M NaHCO3, 1% SDS) for 15 minutes at RT. Supernatants were combined and centrifuged to get rid of beads. The crosslinks were reversed by incubation overnight at 65C. The following day, the samples were treated with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol. The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC. Cluster generation was performed using the Illumina HiSeq 2000 platform.


title: H3.3 knockdown

first: Jianwei

last: Jiao

address: 1 Beichen West Road

department: State Key Laboratory of Reproductive Biology

institute: Institute of Zoology, Chinese Academy of Sciences

data_processing:
Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mouse whole genome using TopHat v2.0.12 and bowtie v1.0.1 with default parameters.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using HTSeq v0.6.0
Genome_build: reference sequences were download from http://www.ensembl.org/index.html
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each Sample

source_name: neural stem cells

growth_protocol: Neural stem cell isolated from E12 mice and cultured in proliferated medium for 5 days

extract_protocol:
Total RNA was extracted by using Trizol reagent. Total 2ug RNA for the construction of sequencing libraries
mRNA were enriched by Oligo(dT) bread and then fragmented by fragmentation buffer. cDNA was synthesized by reverse transcription using 6bp random primers and purified by QIAQuick PCR kit. After adapter ligation, DNA was PCR amplified sequences and band isolated from an agarose gel.

treatment_protocol: Cultured neural stem cell were infected with lentivirus packaged with Control or H3.3 knockdown plasmid

value: mouse embryonic brain

tag: developmental stage

value: H3.3 knockdown


title: V5-Tcf4 ChIP-seq in NS5 Neural Stem Cells

description:
ChIP against V5 in
NS5 NSCs expressing Tcf4-V5

extract_protocol:
Extraction protocol	For each ChIP 1.5*10^8 NSCs were used. NS-5 NSCs expressing no V5-tag were used for the control experiment. NSCs were crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Fisher Scientific) for 45 min at RT, washed three times (PBS + 1mM PMSF) and then crosslinked for 10min with 1% formaldehyde. This reaction was quenched by addition of 1/20 volume of 2.5M glycine for 10min at RT. Cells were washed twice with cold 1x PBS+1mM PMSF, collected by centrifugation, and cell pellets frozen with N2(l). Cells were resuspended in LB1 (50mM Hepes-KOH pH7.5, 140mM NaCl, 1mM EDTA, 10% Glycerol, 0.5% NP40, 0.25% Triton-X-100 . After 10 minutes incubation, cells were pelleted by centrifugation and resuspended in LB2 (10 mM Tris-HCl, pH8.0, 200mM NaCl, 1mM EDTA, 0.5mM EGTA. After 10 minutes incubation, cells were pelleted and resuspended in freshly prepared LB3 (10mM Tris-HCl pH8.0, 100mM NaCl, 1mM EDTA, 0.5 mM EGTA, 0.1% Na-Deoxycholate, 0.5% N-lauroylsarcosine and sonicated on a Soniprep 150(MSE). For quality control of the sonication, it was verified that the DNA fragment size was between 100 and 1000bp. V5-Agarose beads were blocked 1 hr with 0.5 mg/ml BSA. The chromatin was incubated overnight with the blocked V5-Agarose beads. The following day the beads were washed 5 times with RIPA buffer and eluted with elution buffer (50mM Tris-HCl pH8, 10mM EDTA and 1% SDS) for 30 minutes. Samples were de-cross-linked overnight at 65C and treated the following day with proteinase K for one hour at 45C. DNA was extracted by phenol/chloroform/isoamyl extraction and precipitated with ethanol and resuspended in water.
The library was prepared according to the illumina protocol. Briefly 10 ng of DNA ChIP material was end-repaired, ligated to adapters, size selected on gel (200 bp +-25 bp range) and PCR amplified using Phusion polymerase with 30 seconds incubation at 98 ᴼC, 18 cycles of 10 second 98 ᴼC , 30 seconds 65 ᴼC, 30 seconds 72ᴼC. Followed with 5 minutes of final extension at 72 ᴼC.  Cluster generation was performed using the Illumina HiSeq 2000 platform.

treatment_protocol: The coding sequence of Tcf4 was amplified from mouse NSC cDNA. The cDNA was cloned including a N-terminal V5-tag into a pPyCAG driven expression vector. NS-5 cells were transfected with the Amaxa nucleofector I (Lonza kit V, program T30) and clones were selected using puromycin. Expression of stable clones was tested by Westernblot.

value: mouse anti-V5 Agarose affinity gel antibody (sigma A7345)


title: KO-CLP

description: PCR amplified cDNA library

source_name: common lymphoid progenitor cells

value: Lin-c-kit+sca-1+Il-7Ra+ cells

value: bone marrow

value: GRK6 knockout


title: WT-CLP

description: PCR amplified cDNA library

value: wildtype


title: Day 1 adult glh-1(gk100) gonad #2


title: Day 1 adult glh-1(gk100) gonad #1


title: HSF_FPLD-p4_LMNA

value: Santa Cruz sc-7292

value: lipodystrophy patient with LMNA p.R482W mutation

value: Fibroblasts


title: Slc7a5-KO replicate 2

city: Wien

first: Christoph

middle: P

last: Dotter

address: Zimmermannplatz 10

country: Austria

institute: St. Anna Children’s Cancer Research Institute (St. Anna CCRI)

data_processing:
Raw reads were trimmed as suggested by the library preparation kit, i.e. the first 9 bases of each read were removed using the FASTX toolkit (parameter: -f 10)
Reads were aligned using STAR version 2.5.4 (genome: GrCm38/mm10, gene annotation: Gencode release M8). Reads that didn’t align uniquely were discarded. Parameters: --outSAMtype BAM SortedByCoordinate --alignIntronMax 1000000 --outFilterIntronMotifs RemoveNoncanonical --outFilterMultimapNmax 1 --quantMode GeneCounts
Read counts per gene were also calculated using STAR (option - quantMode GeneCounts).
Differential expression analysis was performed in R 3.2.4 employing the Bioconductor package DESeq2 (Love et al., 2014) version 1.10.1 using an FDR threshold of 0.05, a local fit for the dispersion estimation and default parameters otherwise.
Genome_build: GRCm38/mm10
Supplementary_files_format_and_content: Read count table STAR format

extract_protocol:
Mice were euthanized under anesthesia and the brains were quickly dissected on ice. The cerebellum was used for RNA extraction by using 700 μL Trizol (Invitrogen) for homogenization and 140 μL chloroform (Sigma), followed by centrifugation at 12000g for 15 min at 4°C. The upper aqueous phase was transferred to a new tube and 1.5 volumes of 100% ethanol were added. Total RNA was extracted by using the RNA Clean&Concentrator-5 prep Kit (Zymo Research). DNAse I (Promega) treatment was also applied as described in the kit instructions manual. RNA sample quality was checked by using the NanoDrop spectrophotometer (Thermo Scientific) and the Bioanalyzer 2100 with the RNA 6000 Nano kit (Agilent).
1.5 μg total RNA were used to generate cDNA libraries by following the manufacturer’s instructions in the SENSE mRNA-Seq Library Prep Kit V2 (Lexogen). Quality check of the generated libraries was assessed by using the Bioanalyzer 2100 with the High Sensitivity DNA Analysis Kit (Agilent).

tag: strain background

value: Tie2-Cre Slc7a5 fl/fl; Slc7a5-KO

value: post natal day 60

value: cerebellum


title: HSF_FPLD-p1_expression

tag: chip antibody


title: HSF_CTL-2_expression

value: normal


title: HSF_CTL-3_LMNA


title: HSF_CTL-3_expression


title: Slc7a5-KO replicate 1


title: HeLa_Input

tag: cell line


title: HSF_CTL-1_expression


title: WT-HSC

description: PCR amplified cDNA library

value: Lin-c-kit+sca-1+ cells


title: HeLa_flagLMNA_LMNA

value: anti-Flag, Sigma F-1804

value: HeLa


title: HSF_FPLD-p2_Input


title: HSF_FPLD-p2_LMNA


title: WT replicate 2

value: Tie2-Cre Slc7a5 fl/+; wild type


title: Day 1 adult P-granule(-); UNC-119::GFP(-) gonads #3

value: edIs6[unc-119p::unc-119::gfp + rol-6] IV

value: P-granule RNAi


title: HSF_CTL-1_Input


title: HSF_CTL-2_Input


title: Day 1 adult pgl-1(ct131) gonad #3


title: Day 2 adult control gonads #3

value: empty vector RNAi


title: HSF_CTL-1_LMNA


title: Nap1l1-shRNA1

status: Public on Dec 21 2016

source_name: neural stem cell

growth_protocol: Neural stem cell isolated from E12 mice and cultured in proliferated medium for one day

treatment_protocol: Cultured neural stem cell were infected with lentivirus packaged with Piscor or Nap1l1-shRNA1 plasmid

value: ICR

value: brain

tag: lentivirus


title: Day 1 adult wild type gonad #2

value: wild-type


title: Day 1 adult pgl-1(ct131) gonad #2


title: HSF_FPLD-p1_LMNA

value: Santa Cruz sc-7292

tag: passages


title: HSF_FPLD-p4_Input

value: lipodystrophy patient with LMNA p.R482W mutation


title: Control for Fto knockout fto_wt3

city: Heidelberg

first: Jan

last: Mauer

state: BW

address: Im Neuenheimer Feld, 583

country: Germany

institute: BioMed X Innovation Center

data_processing:
To generate m6Am_regions.bed, raw miCLIP reads were first trimmed of their 3' linker (sequence: AGATCGGAAGAGCGGTTCAG) using flexbar (version 2.5).
Reads belonging to each of the 6 biological replicates, as well as other nonspecific reads, were demultiplexed using the pyCRAC tool suite (version 1.2.2.3) and the following 5' barcodes: NNNGGTT, NNNTTGT, NNNCAAT, NNNACCT, NNNGGCG, NNNCCGG.
Reads belonging to each of the 6 biological replicates, but not other nonspecific reads, were concatenated together.
PCR duplicates were removed from the concatenated file using the pyCRAC tool suite (version 1.2.2.3).
Unique reads were aligned to the human genome (hg19) using bowtie (version 1.1.1).
m6Am-containing regions were identified using custom bash and perl scripts.
Genome_build: hg19
Supplementary_files_format_and_content: The *.results files contain normalized gene counts derived from the standard RNA-Seq by Expectation-Maximization (RSEM) pipeline. m6Am_regions.bed is a .bed format file and contains the coordinates of m6Am-containing regions in HEK293T cells. hek293t_rnaseq.bed is a .bed format file that contains the RNA-Seq for normalization of m6Am site relative stoichiometry. The *counts* files contain raw STAR aligner counts.

growth_protocol: Cells were grown as per ATCC guidelines.

extract_protocol:
For RNA-Seq, total RNA was collected using the TRIzol reagent, and was DNase-treated prior to library construction. For miCLIP, total RNA was collected using the TRIzol reagent and treated with DNase. Then, poly(A)+ RNA was selected using oligo(dT) magnetic beads prior to subjecting the samples to the miCLIP protocol.
For RNA-Seq, libraries were prepared using the Illumina TruSeq Stranded mRNA Library Prep Kit. For miCLIP, libraries were constructed using the iCLIP cloning process, as previously described (see GSE63753).

treatment_protocol: FTO and ALKBH5 knockdown experiments were carried out in HEK293T cells using either Pepmute transfection reagent (Signagen) or Lipofectamine RNAiMAX (ThermoFisher Scientific) with 20 nM dsiRNA duplex directed against FTO (HSC.RNAI.N001080432.12.1, Integrated DNA Technologies) or 50 nM Silencer Select siRNA duplex pool targeting AlkBH5 (s29686, s29687, s29688, ThermoFisher Scientific), respectively.  Scrambled siRNA was used as non-targeting control. DCP2 CRISPR knockout HEK293T cells and wild type control HEK293T cells were treated with actinomycin D for 2 hours at a concentration of 5µg/ml.

value: liver biopsy

tag: treatment

value: wild type mouse liver

tag: number of replicates


title: Control for Fto knockout fto_wt2


title: Slc7a5-KO replicate 3

value: Tie2-Cre Slc7a5 fl/fl; Slc7a5-KO

value: post natal day 60


title: HSF_CTL-3_Input

tag: genotype

value: normal

value: Fibroblasts


title: WT replicate 1

value: cerebellum


title: Day 1 adult wild type gonad #3

tag: strain


title: HSF_CTL-2_LMNA


title: Control

value: mouse embryonic brain

value: control


title: FL_HSC_86

city: Baltimore

first: Patrick

last: Cahan

state: MD

address: 733 North Broadway, MRB 653

department: ICE and Biomedical Engineering

institute: Johns Hopkins University School of Medicine

data_processing:
Alignment to transcriptome with HISAT 0.1.6.
Transcript quantification using Salmon 0.3.2.
Gene-level summarization.
Natural log transform (plus one pseudocount)
Genome_build: GRCm38

growth_protocol: Described in paper

extract_protocol:
Described in paper
Described in paper

treatment_protocol: Described in paper

value: fetal liver hematopoietic stem cell


title: FL_HSC_88


title: FL_HSC_89


title: HSF_FPLD-p3_Input


title: HeLa_R388P_LMNA

value: anti-Flag, Sigma F-1804

value: HeLa


title: FL_HSC_90


title: WT replicate 3

value: Tie2-Cre Slc7a5 fl/+; wild type


title: FL_HSC_82


title: Day 1 adult pgl-1(ct131) gonad #1

tag: temperature


title: FL_HSC_83

tag: cell type


title: HSF_FPLD-p3_LMNA


title: FL_HSC_79


title: Control for Fto knockout fto_wt1


title: Piscor

value: ICR

value: brain

value: Piscor


title: Fto knockout fto_ko_3

value: fto knockout mouse liver


title: Day 1 adult wild type gonad #1

value: wild-type


title: Fto knockout fto_ko_2

value: liver biopsy


title: FL_HSC_73


title: FL_HSC_92


title: FL_HSC_93


title: Fto knockout fto_ko_1


title: FL_HSC_87


title: FL_HSC_84


title: FL_HSC_81


title: FL_HSC_94


title: FL_HSC_80


title: FL_HSC_78


title: FL_HSC_62


title: FL_HSC_91


title: FL_HSC_77


title: FL_HSC_85


title: FL_HSC_74


title: FL_HSC_76


title: FL_HSC_72


title: FL_HSC_75


title: FL_HSC_58


title: FL_HSC_66


title: FL_HSC_57


title: FL_HSC_55


title: FL_HSC_71


title: FL_HSC_70


title: FL_HSC_69


title: FL_HSC_53


title: FL_HSC_52


title: FL_HSC_67


title: FL_HSC_65


title: FL_HSC_54


title: FL_HSC_59


title: FL_HSC_60


title: FL_HSC_56


title: FL_HSC_68


title: FL_HSC_50


title: FL_HSC_48


title: FL_HSC_47


title: FL_HSC_64


title: FL_HSC_61


title: FL_HSC_44


title: FL_HSC_51


title: FL_HSC_42


title: FL_HSC_49


title: FL_HSC_63


title: FL_HSC_46


title: FL_HSC_45


title: FL_HSC_39


title: FL_HSC_38


title: FL_HSC_37


title: FL_HSC_43


title: FL_HSC_35


title: FL_HSC_36


title: FL_HSC_30


title: FL_HSC_32


title: FL_HSC_34


title: FL_HSC_31


title: FL_HSC_28


title: FL_HSC_26


title: FL_HSC_29


title: FL_HSC_27


title: FL_HSC_24


title: FL_HSC_25


title: FL_HSC_21


title: FL_HSC_41


title: FL_HSC_22


title: FL_HSC_40


title: FL_HSC_19


title: FL_HSC_20


title: FL_HSC_18


title: FL_HSC_15


title: FL_HSC_14


title: FL_HSC_16


title: FL_HSC_13


title: FL_HSC_12


title: FL_HSC_11


title: FL_HSC_10


title: FL_HSC_9


title: FL_HSC_33


title: FL_HSC_8


title: FL_HSC_7


title: FL_HSC_6


title: FL_HSC_4


title: FL_HSC_23


title: FL_HSC_3


title: FL_HSC_2


title: FL_HSC_1


title: ESC_77

source_name: ESC

value: embryonic stem cell


title: ESC_78


title: ESC_75


title: FL_HSC_17


title: ESC_73


title: ESC_72


title: ESC_74


title: ESC_70


title: ESC_71


title: ESC_68


title: ESC_66


title: ESC_64


title: ESC_67


title: ESC_62


title: ESC_69


title: ESC_63


title: ESC_58


title: ESC_59


title: ESC_60


title: FL_HSC_5


title: ESC_54


title: ESC_57


title: ESC_55


title: ESC_56


title: ESC_52


title: ESC_49


title: ESC_53


title: ESC_50


title: ESC_76


title: ESC_51


title: ESC_47


title: ESC_44


title: ESC_46


title: ESC_45


title: ESC_65


title: ESC_48


title: ESC_43


title: ESC_42


title: ESC_61


title: ESC_37


title: ESC_31


title: ESC_36


title: ESC_32


title: ESC_39


title: ESC_34


title: ESC_28


title: ESC_30


title: ESC_40


title: ESC_29


title: ESC_38


title: ESC_24


title: ESC_33


title: ESC_23


title: ESC_27


title: ESC_22


title: ESC_41


title: ESC_20


title: ESC_25


title: ESC_21


title: ESC_18


title: ESC_19


title: ESC_16


title: ESC_17


title: ESC_15


title: ESC_14


title: ESC_35


title: ESC_12


title: ESC_13


title: ESC_26


title: ESC_11


title: ESC_10


title: ESC_8


title: ESC_5


title: ESC_7


title: ESC_3


title: ESC_4


title: C1_gDNA_rep1

city: Charlottesville

first: Michael

last: Guertin

state: Virginia

address: 1340 Jefferson Park Ave

institute: University Of Virginia

data_processing:
Align with bowtie2
Merge bam files for all replicates
Peak calling with hotspot (standard paramters)
Genome_build: hg38
Supplementary_files_format_and_content: bigWig file for merged replicates. Generated using seqOutBias software to correct for enzymatic bias.

source_name: T lymphoblast

growth_protocol: CEM-C1 cells were cultured in RPMI medium 1640 (Invitrogen) with 1% penicillin/streptomycin (Invitrogen) and 10% FBS . Cells were grown in a humidified incubator at 37°C with 5% CO2.

extract_protocol:
gDNA was extracted using DNeasy blood and tissue kit (Qiagen)
nextera kit

treatment_protocol: baseline

value: T lymphoblast

tag: description

value: Healthy cells with passage number < 30


title: ESC_2


title: MCF 10A MRTF dN Y305A/dPY #1

city: Cambridge

first: Tackhoon

last: Kim

state: Massachusetts

address: 500 Technology Square, NE47-221

institute: MIT

description:
MRTF YAdPY1
MCF-10A cells expressing G-actin, YAP and SRF binding deficient (dPY/Y305A) MRTFB mutant sample #1

data_processing:
Raw data is processed with Tuxedo suite
Aligned to genome with Bowtie and Tophat
Transcripts were assembled with Cufflinks, and fpkm values are normalized by Cuffnorm
Genome_build: hg19
Supplementary_files_format_and_content: Data is processed as a table with normalized transcript count for each gene using cuffnorm

source_name: MCF-10A MRTF dN Y305A/dPY

growth_protocol: MCF 10A cells are maintained as previously described (Debnath J. et. al. Methods, 2003). Cells are harvested two days after retroviral infection of indicated genes

extract_protocol:
RNA was isolated by EasyBlue (Intron Biotechnology, Korea)
For library construction, we used the TruSeq Stranded mRNA Library Prep Kit (Illumina, San Diego, CA). Briefly, the steps of strand-specific protocol are: first strand cDNA synthesis; second strand synthesized using dUTPs instead of dTTPs; end reapair, A-tailing and adaptor ligation; PCR amplification. Then, each library was diluted to 8 pM for 76 cycles of paired-read sequencing (2 X 75bp) on the Illumina NextSeq 500 per the manufacturer’s recommended protocol.

value: mammary epithelial cells


title: ESC_1


title: MCF 10A MRTF dN Y305A #1

description:
MRTF YA1
MCF-10A cells expressing G-actin and SRF binding deficient (Y305A) MRTFB mutant sample #1

source_name: MCF-10A MRTF dN Y305A


title: MCF 10A MRTF dN Y305A/dPY #2

description:
MRTF YAdPY2
MCF-10A cells expressing G-actin, YAP and SRF binding deficient (dPY/Y305A) MRTFB mutant sample #2


title: MCF 10A MRTF dN dPY #2

description:
MRTF dPY2
MCF-10A cells expressing G-actin  and YAP binding deficient (dPY) MRTFB mutant sample #2

source_name: MCF-10A MRTF dN dPY


title: MCF 10A MRTF dN WT #2

description:
MRTF WT2
MCF-10A cells expressing G-actin binding deficient MRTFB mutant sample #2

source_name: MCF-10A MRTF dN WT


title: MCF 10A MRTF dN dPY #1

description:
MRTF dPY1
MCF-10A cells expressing G-actin  and YAP binding deficient (dPY) MRTFB mutant sample #1


title: MCF 10A Puro v. #1

description:
Puro1
MCF-10A cells expressing empty pMSCV Puro vector sample #1

source_name: MCF-10A pMSCV Puro vector

value: expressing empty pMSCV Puro vector


title: MCF 10A Hygro v. #2

description:
Hyg2
MCF-10A cells expressing empty pMSCV Hygro vector sample #2

source_name: MCF-10A Hygro vector

value: expressing empty pMSCV Hygro vector


title: MCF 10A Puro v. #2

description:
Puro2
MCF-10A cells expressing empty pMSCV Puro vector sample #2


title: MCF 10A YAP 5SA #2

description:
YAP2
MCF-10A cells expressing hyperactive YAP #2

source_name: MCF-10A YAP 5SA

value: expressing hyperactive YAP


title: MCF 10A YAP 5SA #1

description:
YAP1
MCF-10A cells expressing hyperactive YAP #1


title: MCF 10A Hygro v. #1

description:
Hyg1
MCF-10A cells expressing empty pMSCV Hygro vector sample #1


title: ESC_6


title: Eng_ES-HSC_48

value: engrafted embryonic stem cell-derived hematopoietic stem cell


title: Eng_ES-HSC_49


title: Eng_ES-HSC_47


title: Eng_ES-HSC_46


title: Eng_ES-HSC_45


title: Eng_ES-HSC_44


title: Eng_ES-HSC_41


title: Eng_ES-HSC_42


title: Eng_ES-HSC_40


title: ESC_9


title: Eng_ES-HSC_37


title: Eng_ES-HSC_39


title: Eng_ES-HSC_35


title: Eng_ES-HSC_34


title: Eng_ES-HSC_36


title: Eng_ES-HSC_30


title: C1_gDNA_rep2


title: Eng_ES-HSC_31


title: Eng_ES-HSC_33


title: MCF 10A MRTF dN Y305A #2

description:
MRTF YA2
MCF-10A cells expressing G-actin and SRF binding deficient (Y305A) MRTFB mutant sample #2


title: Eng_ES-HSC_27


title: Eng_ES-HSC_29


title: Eng_ES-HSC_28


title: Eng_ES-HSC_26


title: Eng_ES-HSC_24


title: Eng_ES-HSC_25


title: Eng_ES-HSC_23


title: Eng_ES-HSC_21


title: Eng_ES-HSC_22


title: Eng_ES-HSC_20


title: Eng_ES-HSC_38


title: Eng_ES-HSC_17


title: Eng_ES-HSC_16


title: Eng_ES-HSC_18


title: Eng_ES-HSC_19


title: Eng_ES-HSC_12


title: Eng_ES-HSC_13


title: Eng_ES-HSC_32


title: Eng_ES-HSC_14


title: Eng_ES-HSC_11


title: Eng_ES-HSC_10


title: Eng_ES-HSC_15


title: Eng_ES-HSC_8


title: Eng_ES-HSC_5


title: Eng_ES-HSC_6


title: Eng_ES-HSC_9


title: Eng_ES-HSC_7


title: Eng_ES-HSC_4


title: Eng_ES-HSC_3


title: Eng_ES-HSC_2


title: Eng_ES-HSC_1


title: Eng_BM-HSC_95

value: engrafted bone marrow-derived hematopoietic stem cell


title: Eng_BM-HSC_93


title: Eng_BM-HSC_92


title: Eng_BM-HSC_94


title: Eng_BM-HSC_91


title: Eng_BM-HSC_90


title: Eng_BM-HSC_88


title: Eng_BM-HSC_86


title: Eng_BM-HSC_85


title: Eng_BM-HSC_84


title: Eng_BM-HSC_81


title: Nrf1_ChIP-seq_MEF_replicate3

city: Sendai

first: Eri

middle: H

last: Kobayashi

state: Miyagi

address: 2-1 Seiryo-machi, Aobaku

department: Medical Biochemistry

country: Japan

institute: Tohoku University Graduate School of Medicine

description: ChIP samples using antibody to Nrf1 obtained from MG132-treated wild-type MEFs, replicate 3

data_processing:
ChIP-seq reads were aligned to the mm9 genome assembly using bwa version 0.6.2-r126.
Data were filtered using samtools version 0.1.18
Peaks were called using macs2 version 2.0.10
Genome_build: mm9
Supplementary_files_format_and_content: BigWig files were generated using BEDtools version 2.17.0 and UCSC Genome Bioinformatics

source_name: Wild-type MEFs immortalized with SV40 large T antigen and treated with MG132.

growth_protocol: MEFs were maintained in DMEM supplemented with 10% FBS and antibiotics.

extract_protocol:
Lysates were clarified from sonicated nuclei, and protein-DNA complexes were isolated using antibody to Nrf1 and Dynabeads-proteinA and -proteinG
Libraries were prepared according to NuGEN's instructions accompanying the Ovation Ultralow DR Multiplex System 1-8 (NuGEN, 0330-32).

treatment_protocol: MEFs were treated with 10 μM of MG132 for 6 hr.

tag: chip-antibody

value: Cell Signaling Technology, anti-TCF11 (D5B10)

value: Mouse enbryonic fibroblasts


title: Eng_BM-HSC_83


title: Eng_BM-HSC_80


title: Eng_BM-HSC_79


title: Nrf1_ChIP-seq_MEF_replicate1

description: ChIP samples using antibody to Nrf1 obtained from MG132-treated wild-type MEFs, replicate 1


title: Eng_BM-HSC_78


title: Eng_BM-HSC_77


title: Eng_BM-HSC_76


title: Input_replicate2

description: Input DNA obtained from MG132-treated wild-type MEFs, replicate 2


title: Input_replicate3

description: Input DNA obtained from MG132-treated wild-type MEFs, replicate 3


title: Input_replicate1

description: Input DNA obtained from MG132-treated wild-type MEFs, replicate 1


title: Eng_BM-HSC_73


title: Eng_BM-HSC_74


title: Eng_BM-HSC_75


title: Eng_BM-HSC_72


title: Eng_BM-HSC_69


title: Eng_BM-HSC_82


title: Eng_BM-HSC_68


title: Eng_BM-HSC_70


title: Eng_BM-HSC_71


title: Eng_BM-HSC_67


title: Eng_BM-HSC_87


title: Nrf1_ChIP-seq_MEF_replicate2

description: ChIP samples using antibody to Nrf1 obtained from MG132-treated wild-type MEFs, replicate 2


title: Eng_BM-HSC_89


title: Eng_BM-HSC_66


title: Eng_BM-HSC_62


title: Eng_BM-HSC_61


title: Eng_BM-HSC_63


title: Eng_BM-HSC_64


title: Eng_BM-HSC_58


title: Eng_BM-HSC_65


title: Eng_BM-HSC_60


title: Eng_BM-HSC_56


title: Eng_BM-HSC_54


title: Eng_BM-HSC_59


title: Eng_BM-HSC_57


title: Eng_BM-HSC_55


title: Eng_BM-HSC_52


title: C10_Control6h_Chase6d_Cebpa13h_RNAseq_Bio2

city: Paris

first: samuel

last: Collombet

address: 46 rue d'Ulm

department: Biology

country: France

institute: Ecole Normale Superieure

data_processing:
reads were mapped using STAR with the Ensembl gene annotation gtf (GRCm38.75) (parameters: -sjdbOverhang 100 -outFilterType BySJout -alignSJoverhangMin 8 -alignSJDBoverhangMin 1 -outFilterMultimapNmax 1 -outFilterMismatchNmax 999 -outFilterMismatchNoverLmax 0.06 -alignIntronMin 20 -alignIntronMax 1000000 -alignMatesGapMax 1000000)
bigwig tracks were made using DeepTools BamCoverage (parameters: -normalizeUsingRPKM -fragmentLength 200 -binSize 1)
For quantitative analysis, reads were counted on genes exon using HtseqCount in unin mode
Samples scaling and statistical analysis were performed using the Rpackage DESeq2 (1.6.3)
Genome_build: mm10
Supplementary_files_format_and_content: bigwig, track of read signal

growth_protocol: C10 (HAFTL cells with a stably integrated ER-C/EBPa transgene) cells were seeded at a concentration of 2e6 cells/mL in B cell media (RPMI no Phenol red, 10% charcoal-stripped FBS, L-glutamine, 50 uM b-mercaptoethanol, pen-strep).

extract_protocol:
RNA was extracted using TRIzol (15596-026) following the manufacturers instructions.
5 ug of RNA was used as starting material for rRNA depletion using the Ambion Ribominus kit (A10837-08). 100 ng of rRNA-depleted RNA was then used in the NEBNext Ultra-directional RNA-seq kit (E7420L) to generate sequencing libraries, following the manufacturers instructions.

treatment_protocol: To initiate trans-differentiation (Td), a 2X Td mix was made by adding the following components to B cell media: b-estradiol at 200nM, IL-3 (peprotech) at 20 nM, and M-CSF (peprotech) at 20 nM. The 2X mixture was added 1:1 to cell containing-media to start the Td process. After the pulse period, cells were washed 3 times in warm B cell media and re-plated in fresh B cell media. Media was refreshed every two days. Td was initiated as above at day 6 to start the re-stimulation period.

value: C10 (HFATL-1 + ER-C/EBPa)

tag: cell status


title: C10_Control6h_Chase6d_Cebpa13h_RNAseq_Bio3


title: Eng_BM-HSC_50


title: Eng_BM-HSC_49


title: C10_Control6h_Chase6d_Cebpa13h_RNAseq_Bio1


title: C10_Control6h_Chase6d_RNAseq_Bio2


title: Eng_BM-HSC_48


title: C10_Control6h_Chase6d_RNAseq_Bio3


title: Eng_BM-HSC_47


title: Eng_BM-HSC_46


title: C10_Control6h_RNAseq_Bio3


title: C10_Control6h_Chase6d_RNAseq_Bio1


title: Eng_BM-HSC_45


title: Eng_BM-HSC_53


title: Eng_BM-HSC_41


title: C10_Control6h_RNAseq_Bio2


title: Eng_BM-HSC_44


title: C10_Cebpa6h_Chase6d_RNAseq_Bio3


title: C10_Control6h_RNAseq_Bio1


title: Eng_BM-HSC_43


title: C10_Cebpa6h_Chase6d_Cebpa13h_RNAseq_Bio3


title: C10_Cebpa6h_Chase6d_Cebpa13h_RNAseq_Bio2


title: Eng_BM-HSC_51


title: Eng_BM-HSC_40


title: Eng_BM-HSC_37


title: Eng_BM-HSC_39


title: Eng_BM-HSC_36


title: Eng_BM-HSC_34


title: C10_Cebpa6h_RNAseq_Bio3


title: C10_Cebpa6h_Chase6d_RNAseq_Bio1


title: Eng_BM-HSC_42


title: Eng_BM-HSC_35


title: C10_Cebpa6h_RNAseq_Bio2


title: C10_Cebpa6h_RNAseq_Bio1


title: C10_Cebpa6h_Chase6d_Cebpa13h_RNAseq_Bio1


title: Eng_BM-HSC_33


title: Pulse_ChIPseq_Pu1

description: ChIP Pu1

data_processing:
reads were mapped using STAR (parameters: -outFilterMultimapNmax 1 -outFilterMismatchNmax 999 -outFilterMismatchNoverLmax 0.1 -alignIntronMax 1 -alignEndsType EndToEnd).
Duplicates reads were removed using picard
bigwig tracks were made using DeepTools BamCoverage (parameters: -normalizeUsingRPKM -fragmentLength 200 -binSize 1)
Genome_build: mm10
Supplementary_files_format_and_content: bigwig, track of fragment signal (elongated to 200b)

extract_protocol:
Cells at 10e6 cells/mL were fixed in 1% formaldehyde for 10 minutes at room temperature. Fixation was quenched with .125M glycine for 5 minutes. Cells were then spun down at 1200 rpm for 4 minutes, and the pellet washed 3 times in cold PBS. The cells were then re-suspended in swelling buffer (25 mM HEPES pH 7.9, 1.5 mM MgCl2, 10 mM KCl, 0.1% NP-40) containing protease and phosphatase inhibitors, and incubated for 10 minutes in ice. At this point, the cell suspension was transferred to a glass Dounce homogenizer with a ‘tight’ pestle and the cell membrane broken with 25 strokes of the pestle. Nuclei were spun down at 3000xg for 5 minutes at 4 degrees. Nuclei were re-suspended in sonication buffer (50 nM HEPES pH 7.9, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS) at a concentration of ~10e6 cell nuclei per mL sonication buffer. Nuclei were sonicated with a Branson probe sonicator for 3 minutes total, 10 seconds on and 45 seconds off, at 30 mA amplitute. Following sonication, the lysate was spun down at 16,000xg for 15 minutes to remove the insoluble material, and this step repeated and the supernatant retained (the ‘chromatin’) for ChIP.  5% of the chromatin lysate was frozen to serve as input material. The rest was used in immunoprecipitation with the following antibodies. 1 mL (about 10e6 cells of starting material) of chromatin was used for each IP. Antibodies and amount used were as follows: PU.1 (sc-352x, 6 ul), p300 (sc-585x, 10 ul), H3K27me3 (07-449, 3 ul), H3K4me1 (Diagenode C15410037, 5 ug) H3K4me2 (04-790, 2 ul), H3K4me3 (05-1339, 5ul). 50 ul of protein-G Dynabeads were added with the chromatin and antibody mixture, and tubes were places on a 360 rotor at 4 degrees overnight. The following morning, the IP was washed with the following buffers (all at 4 degrees, except for TE which is at room temperature): 1X in sonication buffer, 1X in wash A (50 mM HEPES pH 7.9, 500 mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, 0.1% SDS), 1X in wash B (20mM Tris pH 8.0, 1mM EDTA, 250 mM LiCl, 0.5% NP-40, 0.5% Na-deoxycholate), 2X in TE (10mM Tris pH 8.0, 1mM EDTA). Chromatin was then eluted from the washed beads by incubating with 250ul elution buffer (1% SDS, 0.1M NaHCO3) for 15 minutes at room temperature. This step was repeated, and the eluted fractions combined for a total of 500 ul. To this, 20 ul 5M NaCl was added, and the samples incubated at 65 degrees for 4 hours to reverse the crosslinks. Afterwards, to remove the protein in the sample, 10 ul of 0.5M EDTA, 20 ul of 1M Tris pH 6.5, and 2 ul of 10mg/ml Proteinase K was added and incubated for 1 hr at 45 degrees. DNA was recovered by Phenol/chloroform extraction and quantified on a Qubit® 3.0 Fluorometer.
10 ng of ChIP DNA was used for each library. The NEBNext DNA library Prep Reagent set for Illumina (E6000L) was used to generate the libraries, along with the standard Illumina barcoding primers, following the manufacturers instructions.

tag: chip antibody info

value: SantaCruz Sc-352X, lot B0314


title: Pulse_ChIPseq_H3K27me3

description: ChIP H3K27me3

value: EMD Millipore 07-449, lot 1959680


title: Eng_BM-HSC_30


title: Eng_BM-HSC_31


title: Eng_BM-HSC_38


title: Eng_BM-HSC_29


title: Chase_ChIPseq_Pu1

description: ChIP Pu1


title: Eng_BM-HSC_28


title: Control_ChIPseq_Pu1

description: ChIP Pu1

source_name: Bcell

value: Bcell


title: Eng_BM-HSC_26


title: Eng_BM-HSC_27


title: C10_Cebpa6h_Chase6d_RNAseq_Bio2


title: Control_ChIPseq_H3K27me3

description: ChIP H3K27me3


title: Control_H3K4me1

description: ChIP H3K4me1

value: EMB Millipore 04-790, clone AW30


title: Control_H3K4me3

description: ChIP H3K4me3


title: Eng_BM-HSC_32


title: Eng_BM-HSC_23


title: Control_Chase_input

description: ChIP input

value: none (input)


title: Chase_ChIPseq_H3K27me3

description: ChIP H3K27me3


title: Eng_BM-HSC_24


title: Eng_BM-HSC_25


title: Pulse_input

description: ChIP input


title: Eng_BM-HSC_22


title: Eng_BM-HSC_20


title: Eng_BM-HSC_21


title: Eng_BM-HSC_19


title: Chase_input

description: ChIP input


title: Pulse_H3K4me2

description: ChIP H3K4me2


title: Chase_H3K4me1

description: ChIP H3K4me1

value: Diagenode C15410037, lot A1657D


title: Eng_BM-HSC_16


title: Eng_BM-HSC_18


title: Eng_BM-HSC_15


title: Control_input

description: ChIP input


title: Chase_H3K4me3

description: ChIP H3K4me3

value: EMB Millipore 05-1339, clone CMA304


title: Eng_BM-HSC_14


title: Eng_BM-HSC_13


title: Eng_BM-HSC_12


title: MCF 10A MRTF dN WT #1

description:
MRTF WT1
MCF-10A cells expressing G-actin binding deficient MRTFB mutant sample #1

tag: genotype/variation


title: Control_Chase_H3K4me2

description: ChIP H3K4me2


title: Eng_BM-HSC_17


title: Eng_BM-HSC_9


title: Eng_BM-HSC_11


title: Eng_BM-HSC_7


title: Eng_BM-HSC_6


title: Eng_BM-HSC_8


title: Eng_BM-HSC_3


title: Eng_BM-HSC_5


title: Eng_BM-HSC_10


title: Eng_BM-HSC_4


title: Eng_BM-HSC_2


title: SH colon RNA-seq

city: Tai’an

first: Chao

last: Tianle

state: Shandong

address: Daizong street, no. 61, 6/5000 Shandong Agricultural University

institute: 6/5000 Shandong Agricultural University

data_processing:
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to Oar_v4.0 whole genome using BWA, and mapped to reference transcripts using bowtie v0.12.2
RPKM value calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009.
Genome_build: Oar_v4.0
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for exons of transcripts from each sample

organism: Ovis aries

source_name: intestinal mucosal epithelial tissue

extract_protocol:
flash frozen on dry ice, and RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Small tailed Han sheep

tag: tissue

value: colon

value: intestinal mucosal epithelial


title: half-life measurement by transcriptional inhibition nes-fto_actD_1

source_name: HEK 293T (ATCC CRL-11268)

value: human embryonic kidney cells

value: nes-fto vector transfected, 6h actinomycin D


title: half-life measurement by transcriptional inhibition nes-fto_actD_2


title: SH duodenum RNA-seq

value: duodenum


title: Chase_H3K4me2

description: ChIP H3K4me2


title: half-life measurement by transcriptional inhibition nes-fto_nt_2

value: nes-fto vector transfected, non-treated


title: half-life measurement by transcriptional inhibition ctrl_actD_2

value: ctrl vector transfected, 6h actinomycin D


title: Eng_ES-HSC_43

value: engrafted embryonic stem cell-derived hematopoietic stem cell


title: half-life measurement by transcriptional inhibition ctrl_nt_1

value: ctrl vector transfected, non-treated


title: half-life measurement by transcriptional inhibition ctrl_actD_1


title: half-life measurement by transcriptional inhibition nes-fto_nt_1


title: half-life measurement by transcriptional inhibition ctrl_nt_2


title: miCLIP-Seq of HEK293T cells hek293t_rnaseq

value: wild type


title: SLKK_hypoxia_microRNAseq_rep3 [microRNA]

city: Bethesda

first: Coralie

last: Viollet

address: 10 Center Dr

institute: NIH/NCI

description: Total RNA containing short RNA fraction

data_processing:
For small RNA sequencing: Reads were aligned against known miRNAs from miRbase (version 19.0) using the software package SHRIMP (Rumble et al., 2009) (http://compbio.cs.toronto.edu/shrimp/README) with miRNA specific settings (-o 1 -n 2  -r 30% -h 50% --local -Q --qv-offset 32 --sam). As paired-end sequencing, reads were aligned separately covering the mature miRNA both on the forward and reverse read and the obtained number of matches were averaged. The match counts were normalized and tested for differential expression using the edgeR package (McCarthy, Chen, & Smyth, 2012) with default settings.
Genome_build: Human genome (hg19)
Supplementary_files_format_and_content: bedGraph files were generated using BEDtools
Supplementary_files_format_and_content: matrix '_1' and '_2' data columns are paired-end read_1 and paired-end read_2, respectively

source_name: SLKK cells

growth_protocol: Human KS-derived SLK and SLKK cells (also known as SLK+rKSHV.219) were a gift from Dr. Don Ganem (UCSF, CA). They were expanded on receipt, frozen in liquid nitrogen, and stored in a cryogenic tank until used in the experiments described hereafter. Cells were thawed and maintained in Dulbecco’s Modified Eagle medium supplemented with 10% v/v fetal bovine serum and 1% Penicillin/streptomycin/glutamine solution. Additionally, KSHV-positive SLKK cells were periodically grown under selection with 10 μg/mL puromycin to maintain the viral episome.

extract_protocol:
Total RNA was extracted from cells using miRVana miRNA isolation kit according to manufacturer’s instructions. RNA abundance and integrity were determined after isolation using a Nanodrop-ND-1000 spectrophotometer and an Agilent 2100 Bioanalyzer, respectively. Only samples of total RNA with an RNA integrity number (RIN) >9 were further used for RNA-sequencing and small RNA-sequencing. All samples were stored at -80°C.
For microRNA-Seq: Small RNA libraries were constructed using the Illumina TruSeq RNA Sample Prep kit. Briefly, after running 2μg of total RNA in a 15% urea-TBE gel for 1hr at 200V, the 20 to 30 nucleotide RNA fraction was excised and eluted in 0.3M NaCl.  Following separation of the elute from the gel debris using a Spin-X-column, the small RNA samples were precipitated in 100% ethanol and 1mg.mL-1 glycogen, incubated at -80°C for 30 minutes, centrifuged at 14,000rpm for 25 minutes, washed with 75% ethanol, air dried and resuspended in RNAse-free water. Illumina TruSeq libraries were then prepared according to the manufacturer’s protocol and the final RNA library concentration was measured by a Qubit 2.0 fluorometer using Qubit dcDNA HS Assay kit. We verified the size of the products contained in the libraries using a high sensitivity DNA chip and an Agilent 2100 Bioanalyzer. Finally, a total of 6 miRNA libraries (3 SLK and 3 SLKK samples in hypoxia) were sequenced using the Illumina HiSeq platform.

treatment_protocol: Exposure of cell cultures to 1% oxygen was undertaken in an InVivo2 Hypoxia Work Station (Ruskinn Technology, UK). This was undertaken in parallel with cells maintained in normoxic conditions (21% O2). All cells were harvested for protein and RNA 24hrs post-treatment. All experiments were done at least in triplicate from independent cell cultures.

tag: infection status

value: Chronically KSHV-infected (Human herpesvirus 8)

tag: oxygen concentration

value: Hypoxia (1% O2 for 24hrs)

value: SLKK cells


title: Control for FTO knockdown 2

data_processing:
To generate m6Am_regions.bed, raw miCLIP reads were first trimmed of their 3' linker (sequence: AGATCGGAAGAGCGGTTCAG) using flexbar (version 2.5).
Reads belonging to each of the 6 biological replicates, as well as other nonspecific reads, were demultiplexed using the pyCRAC tool suite (version 1.2.2.3) and the following 5' barcodes: NNNGGTT, NNNTTGT, NNNCAAT, NNNACCT, NNNGGCG, NNNCCGG.
Reads belonging to each of the 6 biological replicates, but not other nonspecific reads, were concatenated together.
PCR duplicates were removed from the concatenated file using the pyCRAC tool suite (version 1.2.2.3).
Unique reads were aligned to the human genome (hg19) using bowtie (version 1.1.1).
m6Am-containing regions were identified using custom bash and perl scripts.
Genome_build: hg19
Supplementary_files_format_and_content: The *.results files contain normalized gene counts derived from the standard RNA-Seq by Expectation-Maximization (RSEM) pipeline. m6Am_regions.bed is a .bed format file and contains the coordinates of m6Am-containing regions in HEK293T cells.

value: scrambled siRNA transfected

tag: cell line


title: miCLIP of HEK293T cells


title: FTO knockdown 2

value: FTO siRNA transfected


title: Control for DCP2 knockout 2


title: FTO knockdown 1


title: SH cecum RNA-seq

value: cecum


title: Day6_rep1_RiboSeq

city: La Jolla

first: Nathan

last: Lewis

address: 9500 Gilman Drive

department: Pediatrics

institute: UCSD

description:
Ribosome protected RNA
S010

data_processing:
Sequence data for both RNA-Seq and Ribo-Seq were quality controlled using FastQC
Trimmomatic was used to trim adapter sequences and low quality bases from the reads.
Sequence alignment was accomplished using Bowtie2 (Langmead and Salzberg, 2012) and Tophat2 (Kim et al., 2013). Specifically, rRNA sequences were eliminated by aligning reads to CHO-K1 rRNA sequences using Bowtie2. Reads that did not align to rRNA were then aligned to CHO-K1 reference genome using Tophat2. If Tophat2 reports multiple alignments with the same best quality score, we randomly chose one hit by setting the parameter -g as 1
read counts at each base pair of a gene were based on the 5' end of aligned reads for sense mRNA and based on the 3' end of aligned reads for antisense mRNAs.
To recalibrate the distance between the 5' end and the ribosomal A site, we analyzed the sequence coverage around the translation start site and the translation stop site for every alignment length.
Genome_build: GCF_000223135.1
Supplementary_files_format_and_content: Riboseq and RNAseq quantification are stored in text files. Each row represent rawCount.

organism: Cricetulus griseus

source_name: Chinese hamster ovary cells

growth_protocol: CS13-1.0 CHO cells expressing chimeric IgG against S surface antigen of hepatitis B virus (Kim et al., 1998) were maintained in PowerCHO-2 serum free media (Lonza #12-771Q) supplied with 4mM glutamine (Lonza #17-605F) in vented Erlenmeyer shake flasks (Corning, NY) in a shaking incubator operated at 37°C, 5% CO2 and 120rpm. Additionally, cells were supplemented with 0.2% anti-clumping agent (Life Technologies #0010057AE) and, during maintenance, 1µM Methotrexate hydrate (Sigma #M8407-100MG). Due to high degree of clumping, cells were passed through a 40µm Corning cell strainer (Sigma #CLS431750-50EA) at each passage.   CS13-1.0 cells were cultivated in 1L (SR0700) bioreactors (Eppendorf DASGIP, Jülich, Germany) with a working volume of 600mL. Temperature was maintained at 37°C with an agitation rate of 200 rpm using one three-way segmented impellers. Dissolved oxygen was maintained at 50% of air saturation using air, N2, O2 and CO2 operated at a constant flow rate of 0.6 L/h. Culture pH was maintained at 7.15 with a dead-band of 0.15 using intermittent CO2 addition to the gas mix and 2M sodium carbonate. Culture pH and pO2 was measured on-line. Cell number and viability were measured using a hemocytometer with tryphan blue staining (Sigma-Aldrich). Batch cultures were seeded with 0.3x106 cells/mL in triplicates and samples were drawn on a daily basis. On day 3 and day 6 samples were taken for ribosome profiling. The culture was terminated after 170 hours.

extract_protocol:
RNA was extracted from a minimum of 1x106 cells using Trizol (Life Technologies #15596-026) following manufacturer’s protocol. For qPCR analysis, cDNA was made from 500ng TURBO-DNase (Life Technologies #AM1907) treated RNA, using the qScript Flex cDNA kit (Quanta Bioscience # 95049-100) with random priming. qPCR was run in an Mx3005P (Agilent Technologies) using Brilliant III Ultra-Fast SYBR® Green master mix (Agilent Technologies # 600882). Oligos for qPCR are listed in Supplementary table S1. Fold changes was calculated using the ∆∆CT method. GAPDH was used as reference gene in all calculations.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: antibody

value: IgG

tag: phase

value: Stationary phase


title: Day6_rep2_RiboSeq

description:
Ribosome protected RNA
S011


title: Day6_rep3_RiboSeq

description:
Ribosome protected RNA
S012


title: Eng_BM-HSC_1

value: engrafted bone marrow-derived hematopoietic stem cell


title: Control for DCP2 knockout 1

value: human embryonic kidney cells

tag: treatment


title: SLKK_hypoxia_microRNAseq_rep1 [microRNA]

description: Total RNA containing short RNA fraction


title: SLKK_hypoxia_microRNAseq_rep2 [microRNA]

description: Total RNA containing short RNA fraction


title: ALKBH5 knockdown 2

value: ALKBH5 siRNA transfected

tag: number of replicates


title: SLK_hypoxia_microRNAseq_rep3 [microRNA]

description: Total RNA containing short RNA fraction

source_name: SLK cells

value: Not infected

value: SLK cells


title: ALKBH5 knockdown 1


title: SLK_hypoxia_microRNAseq_rep2 [microRNA]

description: Total RNA containing short RNA fraction


title: Day6_rep2_RNAseq


title: Day6_rep1_RNAseq


title: SLK_hypoxia_microRNAseq_rep1 [microRNA]

description: Total RNA containing short RNA fraction


title: Day6_rep3_RNAseq


title: Day3_rep3_RiboSeq

description:
Ribosome protected RNA
S06

value: Exponential phase


title: uzcR_Zn_RNA-seq

city: Livermore

first: Dan

last: Park

address: 7000 East Ave

department: BBTD

institute: Lawrence Livermore National Lab

data_processing:
Reads were mapped using Bowtie v1.1.1
HTSeq-0.6.1p1 was used to quantify the number of counts per genomic feature
HTSeq-0.6.1p1 output was converted to a .bam file, sorted, and indexed using samtools-0.1.19
The DESeq package in R was used to determine differential expresssion
Genome_build: GenBank accession number NC_011916.1
Supplementary_files_format_and_content: RSeQC-2.3.9 was used to normalize the counts data and generate wig files for visualization. Triplicate samples were merged using the median value at each coordinate. Each condition has a forward and reverse wig file associated with it
Supplementary_files_format_and_content: The wig files were generated using the genome browser mochiview and contain the following header:
track
variableStep chrom=NC_011916.1 span=1
which would be broadly compatible with other genome browser

organism: Caulobacter vibrioides

source_name: ΔCCNA_02845 (uzcR)

growth_protocol: NA1000 and ΔuzcR strains were grown in PYE (peptone, yeast extract) at 30°C to an OD600 of 0.2

extract_protocol:
Cells were harvested and immediately added to ice-cold ethanol containing 5% (vol/vol) water-saturated phenol (pH 6.6). Total RNA was isolated using hot phenol and phenol/chloroform extractions.
5 ug of total RNA was treated with the ScriptSeq Complete Kit - Bacteria from Epicentre, an Illumina company (Cat# BB1224) for ribosomal RNA depletion and library generation with ScriptSeq Index Primers.

treatment_protocol: with or without one hour exposure to 40 µM ZnSO4

tag: genotype background

value: deltaCCNA_02845 (uzcR)

tag: exposed to

value: 40 µM ZnSO4 for 1 hr


title: DCP2 knockout 2

value: DCP2 CRISPR knockout


title: Control for ALKBH5 knockdown 2


title: DCP2 knockout 1


title: SLKK_hypoxia_polyA_mRNAseq_rep3 [mRNA]

description: Total RNA containing short RNA fraction; DNase treated; polyA enriched fraction

data_processing:
For mRNA sequencing: Adapter sequences were trimmed using the Fastx toolkit. Reads were aligned against two target genomes, human (hg19) and KSHV (Genbank accession number NC_009333.1) using TopHat (Trapnell, Pachter, & Salzberg, 2009) to generate spliced alignments. Transcripts were assembled using Cufflinks and Cuffdiff (Trapnell et al., 2010) in order to reveal differentially expressed genes. Significant mRNA fold change was determined by an adjusted p-value lower than 0.05 based on the Benjamini and Hochberg multiple testing correction.
Genome_build: Human genome (hg19) and KSHV genome (Genbank accession number NC_009333.1)
Supplementary_files_format_and_content: bedGraph files were generated using BEDtools
Supplementary_files_format_and_content: Cufflinks ouput
Supplementary_files_format_and_content: Cuffdiff output

extract_protocol:
Total RNA was extracted from cells using miRVana miRNA isolation kit according to manufacturer’s instructions. RNA abundance and integrity were determined after isolation using a Nanodrop-ND-1000 spectrophotometer and an Agilent 2100 Bioanalyzer, respectively. Only samples of total RNA with an RNA integrity number (RIN) >9 were further used for RNA-sequencing and small RNA-sequencing. All samples were stored at -80°C.
For mRNA-Seq: Total RNA was treated with Turbo DNA-free DNase I and Dynabeads to deplete samples from the residual DNA and to isolate the polyadenylated mRNA transcriptome, respectively. PolyA+ RNA libraries were then prepared with the ScriptSeq v2 RNA-Seq kit (Epicentre). The final concentration and size distribution of the RNA libraries were measured by using a Nanodrop-ND-1000 spectrophotometer, and by running a DNA 100 chip on an Agilent 2100 Bioanalyzer. Finally, a total of 6 polyadenylated mRNA libraries (3 SLK and 3 SLKK samples in hypoxia) were sequenced using the Illumina HiSeq platform.

value: Chronically KSHV-infected (Human herpesvirus 8)


title: Control for FTO knockdown 1


title: WT_Zn_RNA-seq

source_name: wild type

value: wild type


title: uzcR_noZn_RNA-seq


title: Day3_rep2_RiboSeq

description:
Ribosome protected RNA
S05

tag: strain