title: Erythroid_CTCF_2

value: CTCF

value: Active Motif, DMABT-H19813


title: HSPC_rep3

status: Public on Jun 27 2016

type: SRA

city: New Haven

first: Vince

last: Schulz

state: CT

address: 333 Cedar St. LMP 4085

department: Department of Pediatrics

country: USA

institute: Yale University

library_source: transcriptomic

data_processing:
Illumina Casava 1.8.2 software was used for basecalling.
FASTQ format sequencing reads were aligned to the hg19 genome using TopHat Version 2.0.4 software with default parameters
Reads in every gene were counted using htseq-count (http://www.ncbi.nlm.nih.gov/pubmed/25260700). Read counts per gene and log2 reads per kilobase per million reads were obtained using the edgeR package.
Genome_build: hg19
Supplementary_files_format_and_content: Counts per gene and log2 RPKM

molecule: total RNA

organism: Homo sapiens

source_name: Human Stem and Progenitor Cell

growth_protocol: To obtain primary human erythroid cells, CD34+ cells were cultured and selected as described.(Panzenbock, B., Bartunek, P., Mapara, M. Y., and Zenke, M. (1998) Blood 92, 3658-3668.  Migliaccio, A. R., Migliaccio, G., Di Baldassarre, A., and Eddleman, K. (2002) Bone Marrow Transplant 30, 75-80) Briefly, Human CD34-selected stem and progenitor cells were cultured in StemSpan SF expansion medium (StemSpan 09650) with estradiol (100 ng/ml), dexamethasone (10 ng/ml), human transferrin (200 ng/ml), insulin (10 ng/ml), Flt3 ligand (100 ng/ml), stem cell factor (100 ng/ml), interleukin-3 (50 ng/ml), interleukin-6 (20 ng/ml), insulin-like growth factor 1 (50 ng/ml), and erythropoietin (3 U/ml) for 9 to 14 days. FACS analysis was used to analyze cellular expression of CD71 (transferrin receptor) and CD235a (glycophorin A). These cells represent the R3/R4 cell population of nucleated erythroid cells defined by Zhang et al.(Zhang, J., Socolovsky, M., Gross, A. W., and Lodish, H. F. (2003) Blood 102, 3938-3946)

extract_protocol:
RNA was prepared using the Qiagen RNeasy kit (Qiagen, Valencia, CA) according to the manufacturer's instructions.
Single end RNA libraries were prepared for sequencing using standard Illumina protocols at the Yale Center for Genome Analysis.

tag: cell type

value: HSPC


title: Erythroid_rep1

source_name: Erythroid Cell

value: Erythroid


title: HSPC_CTCF_2

library_source: genomic

data_processing:
Illumina Casava 1.8.2 software was used for basecalling.
FASTQ format sequencing reads were mapped to the human genome (hg19 NCBI Build 37) using the BWA version 0.6.1 alignment program with default parameters.
The Model-based Alignment of ChIP-Seq (MACS) program version 1.4.0rc2 was used to identify CTCF and SA1 peaks using default parameters.
The SICER program version 1.1 was used to identify broad regions bound by H3K27me3
Genome_build: hg19
Supplementary_files_format_and_content: The CTCF and SA1 macs1.4 peaks bed output files have a fifth field that is -10*log10(pvalue).  The H3K27me3 files have additional SICER fields ChIP_island_read_count, CONTROL_island_read_count, p_value, fold_change, FDR_threshold

molecule: genomic DNA

extract_protocol:
ChIP assays were performed as previously described with minor variations (Steiner, L. A., Maksimova, Y., Schulz, V., Wong, C., Raha, D., Mahajan, M. C., Weissman, S. M., and Gallagher, P. G. (2009) Mol Cell Biol). 1 × 107 cells were cross-linked with 1% formaldehyde for 10 min at room temperature, followed by Dounce homogenization. Cross-linked nuclei were isolated, followed by sonication with a Covaris S2 shearing device (duty cycle 10%, intensity 5, 15 minutes), to obtain chromatin-containing DNA fragments with an average size of ~200-400bp. For each ChIP, 20 μg of antibody was used. Samples were immunoprecipitated with antibodies against antibody against CTCF (Millipore,07-729),  the SA-1 subunit of cohesin (Abcam, ab4457), trimethyl histone 3 lysine 27 (Abcam ab6002) or nonspecific rabbit IgG (sc-2091, Santa Cruz). The antigen-antibody complex was captured on protein G beads, washed four times with radioimmunoprecipitation assay buffer, and then washed with phosphate-buffered saline. The DNA-protein complex was eluted from the protein G beads with 1% sodium dodecyl sulfate at 65°C, and cross-linking of DNA-protein adducts was reversed by overnight incubation at 65°C.   After proteinase K and RNase digestion of the reverse-cross-linked sample, DNA was extracted with phenol-chloroform, precipitated with ethanol, then cleaned with the QIAquick PCR Purification Kit (Qiagen) according to manufacturer instructions.
ChIP libraries were prepared using the KAPA Library Preparation Kit (KAPA Biosystems) per the manufacturer's protocol. Briefly, ChIP DNA was end repaired, followed by addition of an adenine base at the 3' and NEXTflex ChIP-Seq Barcodes (BiooScientific) adaptor ligation. The modified DNA was then size selected on a 2% agarose and purified by using a gel extraction kit (Qiagen).  One microliter of size-selected adaptor ligated DNA was initially analyzed by SYBR Green Q-PCR to determine the number of cycles required to reach 50% of complete amplification.  Adaptor ligated DNA was PCR amplified with one initial heating step of 98°C for 45 second, followed by the pre-determined number of cycles with a melting temperature of 98°C for 15 s, an annealing temperature of 65°C for 30 s, and a product extension at 72°C for 30 s. At the end of the amplification, a final extension at 72°C for 1 min was performed. Amplified ChIP DNA was subjected to deep sequencing using standard Yale Center for Genome analysis protocols.

tag: chip antibody

tag: antibody source


title: HSPC_rep2


title: Jurkat_Pol2_JQ1_500nM_4h

city: Cambridge

first: Richard

middle: A

last: Young

state: MA

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

data_processing:
Aligned using /usr/local/bin/bowtie with configuration -p 4 --best -k 2 -m 2 --sam -l 40
hg19
Supplementary_files_format_and_content: WIG files(s) represent counts of aligned reads within 50 bp bins with each read being extended 200 in the direction of alignment. Counts are in reads-per-million and floored at 0.1

source_name: T-ALL

growth_protocol: Human Jurkat cells were purchased from the American Type Culture Collection (ATCC), Manassas, VA. Cells were maintained under typical conditions in RPMI media with 10% Bovine Calf Serum. For location analysis, cells were grown to a density of 1 million per ml and 99% viability prior to cross-linked with formaldehyde for 20 min.

extract_protocol:
Purified immunoprecipitated DNA were prepared for sequencing according to a modified version of the Solexa Genomic DNA protocol. Fragmented DNA was end repaired and subjected to 18 cycles of LM-PCR using oligos provided by Illumina. Amplified fragments between 150 and 300bp (representing shear fragments between 50 and 200nt in length and ~100bp of primer sequence) were isolated by agarose gel electrophoresis and purified.
Whole cell extracts were sonicated to solubilize the chromatin. The chromatin extracts containing DNA fragments with an average size of 500 bp were immunoprecipitated using different antibodies.

treatment_protocol: Jurkat cells treated with JQ1 (500 nM) for four hours. RNA Pol2 chip-seq

value: Jurkat T-ALL cells

tag: treated with

value: JQ1 (500 nM) for 4hrs

value: RNA Pol II (Rpb1 N-terminus)

tag: chip antibody vendor

value: Santa Cruz

tag: chip antibody cat. #


title: Erythroid_rep2


title: HSPC_SA1_2

value: Abcam, ab4457


title: Erythroid_SA1_1

data_processing:
Illumina Casava 1.8.2 software was used for basecalling.
FASTQ format sequencing reads were mapped to the human genome (hg19 NCBI Build 37) using the BWA version 0.6.1 alignment program with default parameters.
The Model-based Alignment of ChIP-Seq (MACS) program version 1.4.0rc2 was used to identify CTCF and SA1 peaks using default parameters.
The SICER program version 1.1 was used to identify broad regions bound by H3K27me3
Genome_build: hg19
Supplementary_files_format_and_content: The CTCF and SA1 macs1.4 peaks bed output files have a fifth field that is -10*log10(pvalue).  The H3K27me3 files have additional SICER fields ChIP_island_read_count, CONTROL_island_read_count,p_value, fold_change, FDR_threshold


title: DMSO_WCE

treatment_protocol: DMSO 4 hours

value: DMSO 4 hrs


title: HSPC_H3K27me3

value: Abcam, ab6002


title: Erythroid_CTCF_1


title: HSPC_SA1_1


title: HSPC_CTCF_1


title: HSPC_rep1


title: Stem_Butyrate_Input_Rep1

city: St Louis

first: Gerard

last: Kaiko

state: Missouri

address: 660 S Euclid avenue

institute: Washington University in St Louis

description: Input DNA

data_processing:
Sequencing tags from FAIRE- and ChIP-Seq were aligned to the reference genome (mouse MM10) with Novoalign
MACS was used to identify FAIRE or ChIP peaks by comparison of matched treatment to input samples using default settings.
Genome_build: MM10
Supplementary_files_format_and_content: wig files were generated from the matched treatment to input aligned sequence files (bam) using MACS

organism: Mus musculus

source_name: Colon

growth_protocol: Intestinal epithelial spheroid lines were generated from the colon and maintained as described in our previous publication (Miyoshi and Stappenbeck. In vitro expansion and genetic modification of gastrointestinal stem cells in spheroid culture. Nature Protcols. 2013. PMID: 24232249).

extract_protocol:
FAIRE-Seq:  Cells were fixed with 1% formaldehyde in culture medium for 10 min at room temperature followed by quenching with 0.125 M glycine for 5 min.  Crosslinked chromatin was lysed for 10 minutes sequentially in L1 buffer (50mM Hepes KOH pH 7.5, 140mM NaCl, 1mM EDTA pH 8.0, 10% glycerol, 5% NP40 and 0.25% Triton X-100) followed by L2 buffer (200mM NaCl, 1mM EDTA pH 8.0, 0.5mM EGTA pH 8.0, 1M and Tris pH 8.0). The pellets were resuspended in 300ul of buffer L3 (1mM EDTA pH 8.0, 0.5mM EGTA pH 8.0, 1M Tris pH 8.0, 100mM NaCl, 0.1% Na-deoxycholate and 5mg/ml N-lauroyl sarcosine) and subjected to sonication using a Bioruptor to an average size of 500bp. The soluble DNA fraction was isolated by performing two consecutive phenol:chloroform:isoamylalcohol (25:24:1, pH 8) extractions followed by a chloroform-isoamyl alcohol (24:1) extraction. Precipitated DNA was resuspended in 10mM Tris-HCl (pH 7.4) and incubated with 10 ug of RNase A for 1hr. Genomic DNA was isolated using QIAquick PCR purification column.  ChIP-Seq: Cells were fixed with 1% formaldehyde in culture medium for 10 min at room temperature followed by quenching with 0.125 M glycine for 5 min. The cells were washed with ice-cold PBS and lysed in SDS lysis buffer (1% SDS, 10 mM EDTA, 50 mM Tris pH8.0) for 20 min on ice. The crosslinked chromatin was sheared to an average size of less than 300 bp using a BiorupterTM sonicator (Diagenode). 5 µg of antibody was incubated with Dynabeads ProteinA beads (Invitrogen at 4C for 2-4 hours .  Sonicated lysates were subjected to immunopreciptation overnight at 4°C with acetylated H3K27 antibody conjugated to Dynabeads (Invitrogen, USA) coated with Protein-A.  After incubation, the immune complexes were collected by centrifugation and washed with the following buffers each for 3 min at 4C;low salt (150 mM NaCl, 50 mM Tris pH8.0, 0.1% SDS), high-salt buffer (500 mM NaCl, 50 mM Tris pH8.0, 0.1% SDS), LiCl buffer (250 mM LiCl, 50 mM Tris pH8.0, 0.5% Na deoxycholate, 1% NP40) and TE (10 mM Tris pH8.0, 1 mM EDTA). The protein-DNA complexes were eluted from the beads in 250 µl elution buffer (1% SDS, 100 mM NaHCO3) at 65C overnight followed by the addition of proteinaseK to 500 µg/ml and incubaton at 56C for 1 hour. Genomic DNA was isolated using QIAquick PCR purification column.
At least 5 ng of ChIP or input DNA was used for library preparation according to the Illumina ChIP-seq protocol. After end-repair and indexed adapter ligation, fragments were size-selected using Agencourt AMPure XP beads (Beckman Coulter, Brea, CA, USA) prior to amplification. The size-selected and amplified fragment libraries were verified on a 2100 Bioanalyzer (Agilent) prior to being pooled (7-10 per lane) and sequenced

treatment_protocol: After trypsinization, colonic epithelial spheroid cells were cultured in conditioned medium from L cells expressing Wnt3a, R-spondin3, and Noggin (L-WRN cells) (for stem cells). Colonic epithelial stem cells were then treated with either 1mM Butyrate or 1mM NaCl (mock) as a control.

tag: strain

value: colonic epithelial stem cell

tag: treatment

value: 1mM Butyrate


title: Stem_Mock_Input_Rep1

description: Input DNA

value: 1mM NaCl


title: Stem_Mock_Input_Rep2

description: Input DNA


title: Stem_Butyrate_H3K27ac_Rep1


title: Stem_Mock_H3K27ac_Rep1


title: Stem_Mock_H3K27ac_Rep2


title: Stem_Mock_Faire_Rep1


title: Stem_Butyrate_H3K27ac_Rep2


title: HSPC_Total_Input

value: TI


title: Stem_Butyrate_Input_Rep2

description: Input DNA


title: Stem_Butyrate_Faire_Rep2


title: Stem_Butyrate_Faire_Rep1


title: WTFL4_count.txt

status: Public on Jun 26 2016

city: Boston

first: Areum

last: Han

address: Karp Family Building, 7th Floor. 300 Longwood Ave.

institute: Boston Children's Hospital

data_processing:
From the raw reads, we removed adaptor sequences using a Cutadapt tool, allowing some mismatches (The specific parameters we used were -n 2 -e 0.02 –o 15).
Then, the cleaned reads were aligned to the mouse genome/transcriptome (mm10 and corresponding UCSC gene model) using a TopHat2 software with the following parameters (--library-type=fr-unstranded --min-intron-length=10 --coverage-search --microexon-search --min-isoform-fraction=0).
To estimate expression levels for each gene, we counted aligned reads per gene using a htseq-count tool with the following parameters (--stranded=no --idattr=gene_id -t exon -i transcript_id).
Genome_build: mm10
Supplementary_files_format_and_content: Text files generated from the htseq-count tool. The text files have information about counts of aligned reads for each gene.

source_name: wild-type_fetal liver

extract_protocol:
Total RNA was collected using Trizol (Invitrogen) according to the manufacturer's instructions.
For the reverse transcription and library preparation, we used 'SMARTer ultra low RNA kit (Sequencing V3)' and the 'Low Input Library Prep Kit' from Clontech. We followed the manufacturer's protocols.

treatment_protocol: Lin28b overexpression was induced by doxycycline exposure.

tag: genotype/variation

value: wild-type

tag: tissue

value: fetal liver

value: FACS sorted common myloid progenitor cells


title: WTBM5_count.txt

source_name: wild-type_adult bone marrow

value: adult bone marrow


title: WTFL5_count.txt


title: WTFL1_count.txt


title: WTBM6_count.txt


title: Stem_Mock_Faire_Rep2


title: iLin28_4_vavcre_count.txt

source_name: inducible LIN28B under vavcre_adult bone marrow

value: inducible LIN28B under vavcre


title: FLD3_count.txt

source_name: Let-7b/c knock-out_adult bone marrow

value: albino black 6

value: Let-7b/c knock-out


title: 9b_p7m_YFP_pos

city: Davis

first: Mark

middle: O.

last: Huising

state: CA

address: One Shields Avenue

department: Neurobiology, Physiology and Behavior

institute: University of California, Davis

description: index: TAGCTT

data_processing:
reads were mapped to GenCode M5 (GRCm38.p3) using STAR v2.4
FeatureCounts was used to create count tables of the sorted bam files using reads aligning to RefSeq defined exons.
EdgeR v3.12.0 was used for statistical comparison.
Genome_build: GenCode M5 (GRCm38.p3)

source_name: FACS-purified alpha cells

growth_protocol: Isolated were isolated by collagenase injection through the common bile duct per common protocols. Isolated islets were cultured overnight in RPMI supplemented with 5.5 mM glucose and 10% FBS and pen/strep and dissociated the following morning by trypsin (0.25%) EDTA and gentle titturation for FACS purification.

extract_protocol:
Cells were collected in Trizol post FACS purification and isolated by a chloroform extraction, assisted by phase lock tubes. RNA was precipitated by isopropanol and cleaned up over an RNEasy microcolumn per the Qiagen's taking great care to avoid carry over of ethanol following the column washes. Elution volume was 30 microliter.
Indexed sequencing libraries were constructed using the TruSeq RNA sample prep Kit v2 per Illumina instructions.

value: islets

value: FACS-purified alpha cells

tag: Stage

value: adult

tag: gender

value: male


title: 9_p7m_YFP_pos

description: index: GATCAG


title: 12b_p6f_Chy_pos

description: index: GAGTGG

source_name: FACS-purified beta cells

value: FACS-purified beta cells

value: female


title: 13b_p7f_YFP_pos

description: index: ACTGAT


title: iLin28_6_vavcre_count.txt


title: iLin28_1_count.txt

source_name: inducible LIN28B_adult bone marrow

value: inducible LIN28B


title: 8b_p6m_Chy_pos

description: index: TTAGGC


title: 20_p6f_Chy_pos

description: index: GTTTCG


title: 12_p6f_Chy_pos

description: index: CGTACG


title: FLD2_count.txt


title: BKO8_count.txt

source_name: Lin28b KO_fetal liver

value: Lin28b KO


title: 16_p6m_Chy_pos

description: index: ACTTGA


title: 21_p7f_YFP_pos

description: index: CGTACG

source_name: FACS-purified delta cells

value: FACS-purified delta cells


title: BKO6_count.txt


title: FLD1_count.txt


title: CD34- HSC cultured under medium depleted of valine

city: Yokohama

first: Takaho

middle: A.

last: Endo

state: Kanagawa

address: 1-7-22 Suehiro, Tsurumi

department: IMS

country: Japan

institute: RIKEN

description: ValDepleted

data_processing:
Basecalls performed using Illumina offline basecaller (OLB) version 1.9.4
Sequences were aligned to mm9 genome using tophat2 (version 2.8.0) with default parameters.
Aligned seqeunces were converted relative expression using cufflinks (version 2.1.1).
Genome_build: mm9
Supplementary_files_format_and_content: plain text containig FPKM (fragments per kilobase exon per million reads) values.

source_name: HSC

growth_protocol: Ten thousand CD34-KSL cells were obtained from 9-week old B6 mouse.

extract_protocol:
Total RNA was isolated from cultured CD34-KSL cells by using RNeasy mini kit (Qiagen)
Library was prepared according to Illumina TruSeq protocol.

treatment_protocol: Cells were cultured with complete, cysteine deficient, or valine deficient media with SCF and TPO for 24 hours.

tag: strain background

tag: age

value: 9-week old

value: CD34-KSL hematopoietic stem cells (HSCs)

tag: cultured in

value: valine-deficient medium for 24 h


title: CD34- HSC cultured under medium depleted of cystein

description: CysDepleted

value: cysteine-deficient medium for 24 h


title: BKO3_count.txt

value: fetal liver

value: FACS sorted common myloid progenitor cells


title: WTBM2_count.txt

value: wild-type

value: adult bone marrow


title: iLet7_1_count.txt

source_name: inducible Let-7_fetal liver

value: inducible Let-7


title: G3

status: Public on Jun 25 2016

city: Pittsburgh

first: Varsha

last: Shridhar

state: PA

address: 204 Craft Ave

institute: Magee-Womens Research Institute

description: Sample 12

data_processing:
Illumina CASAVA 1.8.2 was used for basecalling and demultiplexing
TrimmomaticPE with the following settings: -phred 33 LEADING:20 TRAILING:20 MINLEN:30
PE Alignment with Bismark v0.7.6, default settings
removed deplicates with script deduplicate_bismark_alignment_output.pl
methylation calls made using Bismark (v.0.7.6) methylation extractor
Used bismark2bedGraph.pl to generate methylation counts, settings: --counts
Used the methylKit package in R for statistical analysis
Genome_build: GRCh37
Supplementary_files_format_and_content: tab-delimited text files for each sample contain columns: chr, coordinate, coordinate, total counts, methylated calls, unmethylated calls (bismark_to_bedgraph output)
Supplementary_files_format_and_content: fetal_maternal_methdiff.txt contains the fetal average and maternal average at a specific site as well as statistical significance of the difference calculated by methylKit

source_name: blood

extract_protocol:
CVS: Qiagen DNeasy Blood and Tissue Kit, MBC: Qiagen QIAmp DNA Blood Midi Kit
Agilent SureSelectXT  Methyl-Seq Target Enrichment Workflow

value: maternal blood cell


title: G2_r1

description: Sample 10


title: CVS_6

description: Sample 8

source_name: chorionic villus DNA

tag: subject age

value: fetus (1st trimester)

value: chorionic villus sample (CVS)


title: G2_r2

description: Sample 11


title: G1

description: Sample 9


title: CD34- HSC cultured under complete medium

description: Complete

value: complete medium


title: iLin28_5_count.txt


title: CVS_5

description: Sample 7

value: male


title: CVS_4_r1

description: Sample 5

value: female


title: CVS_3 r1

description: Sample 3


title: CVS_2

description: Sample 2


title: CVS_3_r2

description: Sample 4


title: Rat-CeA 2-1 PARP1 Chip-seq

city: Montreal

first: Moshe

last: Szyf

state: Quebec

address: McIntyre Medical Building, 3655 Promenade Sir William Osler, Room 1309

department: Pharmacology

country: Canada

institute: McGill University

data_processing:
 Base-calling by the Illumina HiSeq 2000 pipeline
Read alignment to assembly RN4 with Bowtie v0.12.7 with "-m 1", only unique alignments permitted.
Duplicate alignments removed using MarkDuplicates from Picard Tools v 1.37
Tag density and peak calling performed using FP4 with "-dist_type" set to 3.
Genome_build: rn4
Supplementary_files_format_and_content: [.wig] files provide tag density
Supplementary_files_format_and_content: [.peaks] tab-delimited files provide peak locations identified by FP4 (columns: id, chrom, start, end, max_coord, score, fdr)

organism: Rattus norvegicus

source_name: CeA 2-1 PARP1 Chip-seq

growth_protocol: Male Sprague-Dawley rats were maintained under conditions of constant temperature (23°C), humidity (50%), reversed 12:12h light/dark cycle, ad libitum food and water.

extract_protocol:
On completion of the test, brains were immediately removed (within <1 min) and flash frozen. The bi-lateral central amygdale were isolated on dry ice, were homogenized in 1 X PBS including 1% formaldehyde, and the homogenates were kept for   10 min at 25°C. Cross-linking reactions were stopped by the addition of glycine (125 mM) for 10 min at 25°C. Fixed chromatin samples were then homogenized in cell lysis solution (PIPES 5 mM (pH 8), KCl 85 mM, NP40 0.5%) and centrifuge for 5 min at 3000 rpm, 4°C. Pellets were resuspended in RIPA-light solution (NaCl 150 mM, SDS 0.3%, Tris-HCl 50 mM (pH 8)) and sonicated using a covaris E220. Sonicated chromatin samples were then centrifuged for 15 min at 14000 rpm, 4°C. Pellets were resuspended in 1 ml of RIPA-light solution. Chromatin samples were pre-cleared with 50 µl of dynabeads protein G (Life Technologies) pre-blocked with BSA and incubated overnight at 4°C with an anti-PARP1 (AbD serotec, MCA1522G) antibody. Antibodies and chromatin were then mixed with   100 µl of dynabeads protein G for 3 hours at 4°C. The beads were then washed with RIPA-light solution, washing-solution (Tris-HCl 100 mM (pH 8), LiCl 500 mM). Protein–DNA complexes were eluted from the beads, de-cross-linked, treated with proteinase K and purified. The DNA concentration was determined by fluorometry on the Qubit system (Invitrogen). DNA were sheared a second time with the Covaris E220 instrument in 53 µl reaction volume (duty factor 10%, Pic Incident Power 175, Cycles per burst 200, time 360 sec) to obtain fragments in the size range of 150 bp followed by purification with AMPure XP beads   (×1.8v/v) (Beckman Coulter A63881). Purified DNA was resuspended in 45 µl elution buffer. A library of the chromatin immunoprecipitated DNA fragments was prepared using the Tru Seq DNA Low Throughput Protocol (Illumina). PCR enrichment of ligation products was performed using the Illumina Primer Cocktail; 15 cycles of PCR were performed for ChIP libraries and 10 cycles for the input. The libraries were purified using AMPure XP beads ×1.0 v/v. Quality of libraries was validated by 260 nm absorbance measurement, quality control on HSdna chip (Agilent Bioanalyzer : size of libraries around 275bp bp) and quantification  by Q-PCR with Kappa Library Quantification kit for Illumina Sequencing Platforms (KAPPA Biosystems). The DNA concentration of the different sequencing libraries was from 40 to 500 nM. Clusters (13.5 pM) were generated using TruSeq PE Cluster Kit v3, for cBot protocol, which was followed by Pair End Sequencing, 50 cycles, on an Illumina HiSeq 2000, according to the manufacturer recommendations.
DNA samples from 2 rats per group were pooled, a total of 10–12 ng DNA were used for the preparation of the library. The immunoprecipitated DNA and input DNA were sheared a second time with the Covaris E220 instrument in 53 µl reaction volume (duty factor 10%, Pic Incident Power 175, Cycles per burst 200, time 360 sec) to obtain fragments in the size range of 150 bp followed by purification with AMPure XP beads   (×1.8v/v) (Beckman Coulter A63881). Purified DNA was resuspended in 45 µl elution buffer. A library of the chromatin immunoprecipitated DNA fragments was prepared using the Tru Seq DNA Low Throughput Protocol (Illumina). PCR enrichment of ligation products was performed using the Illumina Primer Cocktail; 15 cycles of PCR were performed for ChIP libraries and 10 cycles for the input. The libraries were purified using AMPure XP beads ×1.0 v/v. Quality of libraries was validated by 260 nm absorbance measurement, quality control on HSdna chip (Agilent Bioanalyzer : size of libraries around 275bp bp) and quantification  by Q-PCR with Kappa Library Quantification kit for Illumina Sequencing Platforms (KAPPA Biosystems). The DNA concentration of the different sequencing libraries was from 40 to 500 nM. Clusters (13.5 pM) were generated using TruSeq PE Cluster Kit v3, for cBot protocol, which was followed by Pair End Sequencing, 50 cycles, on an Illumina HiSeq 2000, according to the manufacturer recommendations.

treatment_protocol: Rats were trained in cocaine-CPP paradigm for 10 consecutive days. Cocaine (15 mg/kg; IP) or saline were injected in alternating days. Following injection, rats were correspondingly placed in the cocaine-associated compartment or in the saline-associated compartment, for 20 min. 24 hours after the last injections, rats were tested for their preference of the cocaine-associated compartment during 15 minutes.

value: saline

tag: group

value: control saline only

value: Sprague Dawley

value: central amygdala

value: anti-PARP1 (AbD serotec, MCA1522G)


title: 17_p7m_YFP_pos

description: index: TAGCTT

value: islets

tag: cell type

value: FACS-purified delta cells

value: adult


title: MP

city: Milan

first: Francesco

last: Blasi

address: Via Adamello 16

country: Italy

institute: IFOM

data_processing:
mRNAseq data sets were aligned to mouse (mm9) genome using default parameters of TopHat.
Cufflinks was used to assemble mapped reads into iso-forms.
Biological replicates were merged using Cuffmerge.
Differentially expression analysis was performed using Cuffdiff, and differentially expressed genes were visualized using CummeRbund.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include differentially expressed genes values for each Sample versus ev sample.

source_name: Meis1-Prep1 overexpressing cells (MP)

growth_protocol: Mouse embronic fibroblasts (MEFs) were taken from E14.5 Prep1 hypomorphic embros and cultured in DMEM supplemented with 10% FBS and 2mM L-glutamin. Mefs were passed every 2-3 days.

extract_protocol:
Total RNA was extracted from various cell-types using the Qiagen easy RNA kit.
Illumina multiplexed sequencing libraries were prepared according to the manufacturer’s instructions.

value: Prep1 hypomorphic

value: Mouse embronic fibroblasts (MEFs)

tag: cell genotype/variation

value: Meis1-Prep1 overexpressing cells (MP)

tag: passages


title: M

source_name: Meis1 overexpressing cells (M)

value: Meis1 overexpressing cells (M)


title: P

source_name: Prep1 overexpressing cells (P)

value: Prep1 overexpressing cells (P)


title: Rat-CeA 5-11

source_name: CeA 5-11 input

value: cocaine

value: low cocaine CPP

tag: chip antibody


title: ev

source_name: empty vector infected cells (ev)

value: empty vector infected cells (ev)


title: Rat-CeA 5-11 PARP1 Chip-seq

source_name: CeA 5-11 PARP1 Chip-seq

value: male


title: Rat-CeA 8-6 PARP1 Chip-seq

source_name: CeA 8-6 PARP1 Chip-seq

value: high cocaine CPP


title: outCtr_II

status: Public on Jun 24 2016

city: Marburg

first: Marie

last: Tollot

state: Hessen

address: Karl-von-Frisch-Strasse 10

department: Organismic Interactions

country: Germany

institute: MPI Marburg

data_processing:
mapping of paired end reads to U. maydis 521 genome using the CLC Genomics software 7.5  (CLC Bio-Qiagen)
peak-calling based on peak shape score and p-value using the ChIP-seq analysis tool of the CLC genomics Workbench software 7.5 (CLC Bio-Qiagen) combining all three replicates from Ros1HA sample and Ros1 control sample.
Genome_build: The reference file used for mapping is the list of U. maydis open reading frames (file p3_t237631_Ust_maydi_v2GB.orf found on MIPS database (ftp://ftpmips.gsf.de/fungi/Ustilaginaceae/Ustilago_maydis_521/))
Supplementary_files_format_and_content: List of ChIP peaks identified in output DNA from Ros1HA sample compared to input DNA. The table gives the localization, the length, the peak shape score and the p-value of each ChIP peak sequence.

organism: Mycosarcoma maydis

source_name: infected plant material

growth_protocol: Ustilago maydis cells were grown to OD=1.0 in YEPSL, pelleted and resuspended in water. Compatible strains were mixed in equal quantities and injected into 7-day old maize seedlings

extract_protocol:
Samples were ground to a fine powder in liquid nitrogen. For chromatin preparation, powder was resuspended in lysis buffer (50mM HEPES  pH 7.5, 150 mM NaCl, 1 mM EDTA, 1% Triton X-100, 5 mM benzamidine, 2 mM PMSF, 1 X Roche complete EDTA free protease inhibitors) and further treated by sonication to lyse the remaining cells using a microtip sonifier (Branson). Chromatin was then sheared in a bioruptor sonication bath (Diagenode) for 10 cycles (30s on / 30s off) at high power setting. An aliquot was saved to serve as input control and the rest of the chromatin solution was incubated with ChIP grade Protein A/G magnetic beads (Life technologies) coupled to a monoclonal anti-HA antibody (Sigma) for 10h at 4°C. Beads were washed three times in lysis buffer, two times in high salt buffer (50 mM HEPES pH 7.5, 300 mM NaCl, 1 mM EDTA, 1% Triton X-100, 1 X Roche complete EDTA free protease inhibitors) and once in TE buffer (10 mM Tris-HCl, 1 mM EDTA, pH 7.5). DNA / Ros1HA complexes were eluted from the beads in TE SDS buffer (10 mM Tris-HCl, 1 mM EDTA, 1% SDS, pH 7.5) by incubation for15 min at 65°C. Samples and input controls were de-crosslinked for 8-10h at 65°C in TE SDS buffer containing 200 mM NaCl and 0.65 µg/µL proteinase K. DNA was purified using the ChIP DNA clean and concentrator kit (Zymo research). The experiment was done in three biological replicates which were sequenced separately.
DNA libraries were prepared using the Illumina TruSeq RNA sample preparation Kit v2 starting from the end repair step of the protocol. Up to 100 ng ChIP DNA was used as starting material. After end repair and A-tailing, indexing adapters were ligated. The products were then purified and amplified for 18 PCR cycles to create the final libraries. After validation (Agilent 2200 TapeStation) and quantification using the KAPA Library Quantification Kit (VWR, Erlangen, Germany) and the Applied Biosystems 7900HT Sequence Detection System, equimolar amounts of library were pooled. The pool was sequenced using the Illumina TruSeq PE Cluster Kit v3 and the Illumina TruSeq SBS Kit v3-HS (101x7x101 Cycles) on an Illumina HiSeq 2000 sequencer with a paired-end (101x7x101 cycles) protocol.

treatment_protocol: Leaf sections from 5 different plants infected with Dros1 complemented strains expressing an HA tagged version of Ros1 or with complementation strains expressing the native Ros1 protein were collected and frozen in liquid nitrogen.

tag: fungal strain

tag: host plant

value: Zea Mays (Gaspé Flint)

value: 8 days post infection


title: outCtr_III


title: Rat-CeA 10-9 PARP1 Chip-seq

source_name: CeA 10-9 PARP1 Chip-seq

value: saline

value: control saline only

tag: strain

value: Sprague Dawley

value: central amygdala


title: CVS_1

description: Sample 1

value: fetus (1st trimester)

value: chorionic villus sample (CVS)


title: inCtr_I


title: inCtr_III


title: outHA_II


title: WT

source_name: wild-type cells (WT)

value: Prep1 wild-type

value: Mouse embronic fibroblasts (MEFs)

value: wild-type cells (WT)


title: outHA_III


title: outHA_I


title: inCtr_II


title: outCtr_I


title: CVS_4_r2

description: Sample 6

value: female


title: inHA_III


title: inHA_I


title: Dros1_II

data_processing:
mapping of paired end reads to U. maydis 521 genome using the CLC Genomics Workbench software 7.5  (CLC Bio-Qiagen)
Differential gene expression analysis was carried out using the R software framework for statistical analyses (version 3.2.2), using the package edgeR (Robinson et al., 2010) (version 3.8.6).
Read count data from replicates were merged, given that corresponding genes were expressed under both conditions. Read count data was subsequently normalized using the method
introduced by Robinson and Oshlack (2010). Differentially expressed genes were identified by genewise exact tests for differences in the means of the negative-binomially distributed
read counts between the two treatments. A significance threshold of 0.01 was applied.
Genome_build: The reference file used for mapping is the list of chromosome sequences (file p3_t237631_Ust_maydi_v2GB.scaf available at  ftp://ftpmips.gsf.de/fungi/Ustilaginaceae/Ustilago_maydis_521/)
Supplementary_files_format_and_content: Processed data are presented in two files one is the list of differentially expressed U.maydis genes in wild type vs ros1 mutant (plant infected with strains FB1 x FB2 or FB1Dros1 x FB2Dros1) the other is a table presenting the number of reads mapping to each U. maydis open reading frame in each sample and each replicate.

extract_protocol:
Samples were ground to a fine powder in liquid nitrogen and total RNAs were extracted using the TRIzolâ method (Invitrogen)
Sequencing libraries were prepared with the Illumina TruSeq® RNA sample preparation Kit starting from 2µg total RNA. After poly-A selection (using poly-T oligo-attached magnetic beads), mRNA was purified and fragmented using divalent cations under elevated temperature. The RNA fragments were reverse transcribed using random primers. A second strand cDNA synthesis was carried out with DNA Polymerase I and RNase H. After end repair and A-tailing, indexing adapters were ligated to the cDNA. The products were then purified and amplified (14 PCR cycles) to create the final cDNA libraries. After library validation and quantification (Agilent 2100 Bioanalyzer), equimolar amounts of library were pooled. The pool was quantified using the Peqlab KAPA Library Quantification Kit and the Applied Biosystems 7900HT Sequence Detection System. The pool was sequenced using an Illumina TruSeq PE Cluster Kit v3 and an Illumina TruSeq SBS Kit v3-HS on an Illumina HiSeq 2000 sequencer with a paired-end (101x7x101 cycles) protocol.

treatment_protocol: leaf sections from 10 different plants infected with FB1 x FB2 U. maydis wild type strains or the corresponding Dros1 strains were collected and frozen in liquid nitrogen.

value: FB1{delta}ros1 x FB2{delta}ros1


title: Dros1_III


title: WT_II

value: FB1 x FB2


title: Dros1_I


title: WT_III


title: WT_I


title: 100µM_AZA-treated_3

city: Athens

first: Robert

middle: J

last: Schmitz

state: GA

address: B416 Davison Life Sciences

department: Genetics

institute: University of Georgia

description: RNA Sequencing on 100µM AZA-treated 3

data_processing:
Data were mapped to the TAIR 10
MethylC-seq analysis were performed as described in Schmtiz et al. Epigenome-wide inheritance of DNA methylation variants in a recombinant inbred population. Genome Research (2013).
Genome_build: TAIR 10
Supplementary_files_format_and_content: MethylC-seq data (marked as “allc_sampel.tsv”) column 1 = chr = chromosome; column 2 = pos = coordinate for the cytosine position on the chromosome; column 3 = strand = + or - strand; column 4 = mc_class = context of the cytosine and the two following bases from the same strand; column 5 = mc_count = number of reads supporting a methylated cytosine; column 6 = total = total number of reads at that position; column 7 = methylated = cytosine is considered methylated if there is a 1 or unmethylated if there is a 0
Supplementary_files_format_and_content: RNA-seq data (marked as "gene_exp.diff") column 1 = Tested id = A unique identifier describing the transcipt, gene, primary transcript, or CDS being tested; column 2 = gene = The gene_name(s) or gene_id(s) being tested; column 3 = locus = Genomic coordinates for easy browsing to the genes or transcripts being tested.; column 4 = sample 1 = Label (or number if no labels provided) of the first sample being tested; column 5 = sample 2 = Label (or number if no labels provided) of the second sample being tested; column 6 = Test status = Can be one of OK (test successful), NOTEST (not enough alignments for testing), LOWDATA (too complex or shallowly sequenced), HIDATA (too many fragments in locus), or FAIL, when an ill-conditioned covariance matrix or other numerical exception prevents testing.; column 7 = FPKMx = FPKM of the gene in sample x; column 8 = FPKMy = FPKM of the gene in sample y; column 9 = log2(FPKMy/FPKMx) = The (base 2) log of the fold change y/x; column 10 = test stat = The value of the test statistic used to compute significance of the observed change in FPKM; column 11 = p = The uncorrected p-value of the test statistic; column 12 = q = The FDR-adjusted p-value of the test statistic; column 13 = significant = Can be either “yes” or “no”, depending on whether p is greater then the FDR after Benjamini-Hochberg correction for multiple-testing

organism: Arabidopsis thaliana

source_name: 100µM AZA-treated 3

growth_protocol: Agarose gel (Ameresco) with added half-strength Linsmaier and Skoog nutrients (Caisson Laboratories, Inc) was prepared and autoclaved. 5-azacytidine (Sigma) and zebularine (APExBIO) were dissolved in dimethyl-sulfoxide (DMSO) and water, respectively, before being added to the liquefied cooling agar at final concentrations of 25 µM, 50 µM and 100 µM. Columbia-0 (col-0) background Arabidopsis thaliana seeds were then subjected to a ethanol-based seed sterilization and approximately 30 seeds were plated on the solid agar. As a control, seeds were plated on agar containing DMSO with no chemical demethylating agent (AZA mock-treated control) and agar containing no DMSO or chemical demethylating agent (untreated control). Three replicates of each treatment type were simultaneously plated and grown in parallel. After a two-day stratification period at 4ºC, the seeds were removed to room temperature and allowed to grow for 8 days under constant light.

extract_protocol:
Tissue was flash frozen in liquid nitrogen for all experiments including RNA-seq and MethylC-seq. DNA was isolated using a Qiagen Plant DNeasy kit (Qiagen, Valencia, CA) following the manufacturer’s recommendations. RNA was isolated using TRIzol (Thermo Scientific, Waltham, MA) following the manufacturer’s instructions.
RNA-seq: RNA-seq libraries were constructed using Illumina TruSeq Stranded RNA LT Kit (Illumina, San Diego, CA) following the manufacturer’s instructions with limited modifications. The starting quantity of total RNA was adjusted to 1.3 µg, and all volumes were reduced to a third of the described quantity.
MethylC-seq libraries were constructed using the MethylC-seq protocol (Urich et al. 2015).

tag: ecotype

value: whole seedling

tag: treatment

value: 100 uM 5-azacytidine


title: 100µM_AZA-treated_2

description: RNA Sequencing on 100µM AZA-treated 2

source_name: 100µM AZA-treated 2

tag: tissue


title: inHA_II

tag: Stage


title: Mock_treated_1

description: RNA Sequencing on Mock treated 1

source_name: Mock treated 1

value: 0 uM


title: Mock_treated_2

description: RNA Sequencing on Mock treated 2

source_name: Mock treated 2


title: sorted hematopoietic stem cells [JC48P4]

city: Utrecht

first: Jean-Charles

last: Boisset

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

data_processing:
Paired end reads were aligned to the transcriptome using bwa. Paired end reads obtained by CEL-seq were aligned to the transcriptome using bwa (version 0.6.2-r126) with default parameters. The transcriptome contained all RefSeq gene models based on the mouse genome release mm10 downloaded from the UCSC genome browser and contained 31,109 isoforms derived from 23,480 gene loci. All isoforms of the same gene were merged to a single gene locus. The right mate of each read pair was mapped to the ensemble of all gene loci and to the set of 92 ERCC spike-ins in sense direction. Reads mapping to multiple loci were discarded. The left read contains the barcode information: the first eight bases correspond to the cell specific barcode followed by 4 bases representing the unique molecular identifier. The remainder of the left read contains a polyT stretch followed by few (<15 transcript derived bases). The left read was not used for quantification. For each cell barcode we counted the number of unique molecular identifiers for every transcript and aggregated this number across all transcripts derived from the same gene locus. Based on binomial statistics we converted the number of observed unique molecular identifiers into transcript counts.
Genome_build: mm10
Supplementary_files_format_and_content: Tab separated data file for each sequencing library, listing all genes (rows) and the number of sequenced transcripts for all samples. Column name indicate the library, followed by micro-structure number, and cell number within the micro-structure. Column names containing "HSC" are single sorted hematopoietic stem cells. The rest are micro-dissected bone-marrow cells. The first column lists the official gene symbol followed by the chromosome name, separated by a double underscore.

source_name: Bone marrow

extract_protocol:
Single hematopoietic stem cells were sorted (Lineage- Kit+ Sca1+ CD150+ CD48-) into Trizol (Life Technologies) and total RNA was isolated according to the manufacturer’s instructions, with the following alterations. RNA was precipitated overnight at -20 °C, with 2.5µg GlycoBlue (Life Technologies).
RNA was processed using the previously described CEL-seq technique, with the indicated modifications. Libraries were sequenced on an Illumina HiSeq2500 using 50bp paired end sequencing or Illumina NextSeq500 using 75bp paired end sequencing.

value: bone marrow cells

tag: cell subtype

value: hematopoietic stem cells


title: ZEB_100

description: Whole Genome Bisulfite Sequencing 100 µM ZEB-treated

source_name: ZEB 100

value: 100 uM zebularine


title: ZEB_50

description: Whole Genome Bisulfite Sequencing 50 µM ZEB-treated

source_name: ZEB 50

value: 50 uM zebularine


title: ZEB_25

description: Whole Genome Bisulfite Sequencing 25 µM ZEB-treated

source_name: ZEB 25

value: 25 uM zebularine


title: sorted hematopoietic stem cells [JC48P7]


title: Leaf BGM-infested 24hr rep4

city: Salt Lake City

middle: M

last: Clark

state: Utah

department: Department of Biology

institute: University of Utah

data_processing:
Following transfer of the flowcell to an Illumina HiSeq 2500 instrument (HCS v2.2.38 and RTA v1.18.61), a 125 cycle paired-end sequence run was performed using HiSeq SBS Kit v4 sequencing reagents (FC-401-4003), and the resulting reads were aligned to the H. vulgare reference genome (082214) with STAR 2.5.1b in two-pass alignment mode (alignments were independent of the existing reference annotation).
Read counts for genes in the ASM32608v1 annotation were obtained using HTSeq 0.6.0 with --stranded reverse and --feature transcript.
Genome_build: 082214 (genome sequence) and ASM32608v1 (genome annotation) were downloaded from the barley genome site at ensembl.org.
Supplementary_files_format_and_content: Tab-delimited text file with counts of reads per gene from alignments as assessed with HTSeq (expression across all samples is included in the single processed data file).

molecule: polyA RNA

organism: Hordeum vulgare

source_name: Leaf tissue

growth_protocol: The barley (Hordeum vulgare, Morex variety) plants used for the experiments were germinated and grown in a growth chamber with 16h-light/8h-dark photoperiod at a temperature of approximately 20 degrees C. They were cultivated using Sun Gro® Metro-Mix® 900 Grower Mix, watered as needed, and fertilized weekly with 200ppm NutriCulture 18N-6P-18K.

extract_protocol:
Leaf tissue was ground to fine powder in liquid nitrogen. Total RNA was prepared using the DirectZol RNA extraction kit (Zymo Research). Quality of the RNA extractions was assessed using a Bioanalyzer (Aligent).
The libraries were prepared at the University of Utah High Throughput Genomics facility using the Illumina TruSeq Stranded mRNA Library Preparation Kit with poly(A) selection.

treatment_protocol: The 5th leaves of 30-day-old barley plants were used for the experiment. Mite movement was restrained by barriers made of a non-toxic wax (TangleFoot). Mites were allowed to feed on both sides of a leaf section defined by TangleFoot barriers. The barley leaf sections were infested with healthy adult females, 400 mites per leaf section for 2hr-timepoints and 200 mites per leaf section for 24hr-timepoints. Wounding treatment was applied by rubbing 60 grit sand paper on both sides of the leaf. The control leaves received the TangleFoot barriers but none of the treatments. Mite infestation and wounding treatments were applied at 24hr and 2hr before sample collection. All samples were collected at the same time to limit the effects of the circadian cycle and any minor environmental differences on gene expression.

value: V5-30 days old

tag: leaf number

value: leaf number 5

tag: mite strain

value: Oligonychus pratensis, BGM


title: Leaf BGM-infested 24hr rep2


title: sorted hematopoietic stem cells [JC48P2]


title: Leaf BGM-infested 24hr rep3


title: micro-dissected cells [JC46]

data_processing:
Paired end reads obtained by CEL-seq were aligned to the transcriptome using bwa (version 0.6.2-r126) with default parameters. The transcriptome contained all RefSeq gene models based on the mouse genome release mm10 downloaded from the UCSC genome browser and contained 31,109 isoforms derived from 23,480 gene loci. All isoforms of the same gene were merged to a single gene locus. The right mate of each read pair was mapped to the ensemble of all gene loci. Reads mapping to multiple loci were discarded. The left read contains the barcode information: the first eight bases correspond to a sample specific barcode. The remainder of the left read contains a polyT stretch followed by few (<15 transcript derived bases).
The 'cel-seq_barcodes.csv'  file contains the 96 different barcodes that we used. The description of the reads structure, and how to process the fastq files, was already described in a recent paper: Grün et al. 2015, Nature, Sep 10;525(7568):251-5, in methods: quantification of transcript abundance.
Genome_build: mm10
Supplementary_files_format_and_content: Tab separated data file for each sequencing library, listing all genes (rows)  and the number of sequenced transcripts for all samples. Column name indicate  the cell number within the experiment, separated by a space. The first column lists the official gene symbol followed by the chromosome name,  separated by a double underscore.

extract_protocol:
Cells were directly placed into Trizol (Life Technologies) and total RNA was isolated according to the manufacturer’s instructions, with the following alterations. RNA was precipitated overnight at -20 °C, with 2.5µg GlycoBlue (Life Technologies).
RNA was processed using the previously described CEL-seq technique, with the indicated modifications. Libraries were sequenced on an Illumina NextSeq500 using 75bp paired end sequencing.

value: micro-dissected cells


title: micro-dissected cells [JC41]


title: micro-dissected cells [JC47]


title: micro-dissected cells [JC44]


title: micro-dissected cells [JC45]


title: micro-dissected cells [JC40]


title: micro-dissected cells [JC39]


title: Mock_treated_3

description: RNA Sequencing on Mock treated 3

source_name: Mock treated 3


title: Mock_treated_4

description: RNA Sequencing on Mock treated 4

source_name: Mock treated 4


title: Leaf BGM-infested 2hr rep4


title: Leaf BGM-infested 2hr rep3


title: sorted hematopoietic stem cells [JC48P6]


title: 100µM_AZA-treated_1

description: RNA Sequencing on 100µM AZA-treated 1

source_name: 100µM AZA-treated 1


title: micro-dissected cells [JC36]


title: micro-dissected cells [JC35]


title: Leaf BGM-infested 2hr rep2


title: micro-dissected cells [JC30]


title: Leaf BGM-infested 2hr rep1


title: Leaf TSSM-infested 24hr rep4

value: Tetranychus urticae, TSSM


title: micro-dissected cells [JC32]


title: micro-dissected cells [JC27]


title: micro-dissected cells [JC26]


title: Leaf BGM-infested 24hr rep1


title: micro-dissected cells [JC43]


title: micro-dissected cells [JC20]


title: Leaf TSSM-infested 24hr rep3


title: micro-dissected cells [JC21]


title: Donor number 7 live sorted cells,  library 4

first: Mauro

last: Muraro

state: Utrecht

address: Uppsalalaan, 8

institute: Single Cell Discoveries

data_processing:
Paired end reads obtained by CEL-seq were aligned to the transcriptome using bwa (version 0.6.2-r126) with default parameters. The transcriptome contained all RefSeq gene models based on the mouse genome release mm10 downloaded from the UCSC genome browser and contained 31,109 isoforms derived from 23,480 gene loci. All isoforms of the same gene were merged to a single gene locus. The right mate of each read pair was mapped to the ensemble of all gene loci. Reads mapping to multiple loci were discarded. The left read contains the barcode information: the first eight bases correspond to a sample specific barcode. The remainder of the left read contains a polyT stretch followed by few (<15 transcript derived bases).
Genome_build: hg19
Supplementary_files_format_and_content: D2_3_7_10_17.csv: Tab separated data file for all sequencing libraries combined, listing all genes (rows) and the number of sequenced unique transcripts for all samples. Column name indicates the experiment (D*) and the cell number of the experiment, separated by an underscore. The first column lists the official gene symbol followed by the chromosome name, separated by a double underscore.

source_name: human pancreas

extract_protocol:
Cells were sorted into Trizol (Life Technologies) and total RNA was isolated according to the manufacturer’s instructions, with the following alterations. RNA was precipitated overnight at -20 °C, with 2.5ug GlycoBlue (Life Technologies).
RNA was processed using the previously described CEL-seq technique, with the indicated modifications. Libraries were sequenced on an Illumina HiSeq2500 using 50bp paired end sequencing and Illumina NextSeq500 using 75bp paired end sequencing.

value: live sorted cells

tag: library


title: micro-dissected cells [JC37]


title: Leaf TSSM-infested 24hr rep1


title: Leaf TSSM-infested 24hr rep2


title: Donor number 7 live sorted cells,  library 1


title: Donor number 17, TGFBR3+ sorted cells

value: TGFBR3+ sorted cells


title: Leaf TSSM-infested 2hr rep4


title: Donor number 7 live sorted cells,  library 2


title: Donor number 17, live sorted cells,  library 2


title: Donor number 17, live sorted cells,  library 1


title: Donor number 17, CD24+ CD44+ live sorted cells

value: CD24+ CD44+ live sorted cells


title: Leaf TSSM-infested 2hr rep3


title: micro-dissected cells [JC28]


title: Donor number 17, CD13+ sorted cells

value: CD13+ sorted cells


title: Leaf TSSM-infested 2hr rep2


title: Donor number 10, CD63+ sorted cells

value: CD63+ sorted cells


title: Leaf TSSM-infested 2hr rep1


title: SVE (mNKX3.1 + UTY_sh) + UGM, CAD120

city: Newark

first: Antonina

last: Mitrofanova

state: NJ

address: 65 Bergen Street, Rm 923B

institute: Rutgers University

description:
CAD120
Processed data file: RNAseq_mouse_recomb.txt

data_processing:
Libraries were sequenced using Illumina HiSeq 2000 at Columbia Genome Center. We multiplexed samples in each lane, which yielded targeted number of single-end/paired-end 100bp reads for each sample, as a fraction of 180 million reads for the whole lane.
We used RTA (Illumina) for base calling and bcl2fastq (version 1.8.4) for converting BCL to fastq format, coupled with adaptor trimming. We mapped the reads to a reference genome (Mouse: UCSC/mm9) using TopHat (version 2.0.4) with 4 mismatches (--read-mismatches = 4) and 10 maximum multiple hits (--max-multihits = 10). To tackle the mapping issue of reads that are from exon-exon junctions, TopHat infers novel exon-exon junctions ab initio, and combines them with junctions from known mRNA sequences (refgenes) as the reference annotation.  We estimated the relative abundance (aka expression level) of genes and splice isoforms using Cufflinks (version 2.0.2) with default settings. We tested for differentially expressed genes under various conditions using DESeq. It is an R package based on a negative binomial distribution that models the number of reads from RNA-seq experiments and tests for differential expression.
Raw RNA-seq counts were normalized (variance stabilization) using DESeq package in R-studio Version 0.99.484.
Genome_build: Mouse: UCSC/mm9
Supplementary_files_format_and_content: Tab-delimited text file includes DESeq normalized values for each Sample.

source_name: tissue recombinants, SVE (mNKX3.1 + UTY_sh) + UGM

growth_protocol: Tissue recombinants generated from combining engineered mouse epithelial cells (seminal vesicle epithelial [SVE] cells or prostate epithelial [PE] cells from 2-month-old C57Bl/6J mice) and rat urogenital sinus mesenchymal (UGM) cells were harvested, and snap frozen for subsequent molecular analysis.

extract_protocol:
RNA was extracted using the RNeasy kit (Qiagen). Quality control was performed with Agilent Bioanalyser 2100. We used poly-A pull-down to enrich mRNAs from total RNA samples (200ng-1ug per sample, RIN>8 required).
Library preparation was performed using the Illumina TruSeq RNA prep kit. Libraries were then sequenced using Illumina HiSeq 2000 at Columbia Genome Center.

treatment_protocol: Mice were administered doxycycline in drinking water for 3 months to induce expression of NKX3.1 wild type and NKX3.1 T164A mutant from altered pTRIPZ inducible vector.

tag: mouse strain/background

value: hybrid 129/SvImJ and C57Bl/6J

tag: rat strain/background

value: Sprague Dawley

value: tissue recombinants (mouse seminal vesicle epithelial cells + rat urogenital sinus mesenchymal cells)

tag: mouse genotype/variation

value: mNKX3.1 + UTY_sh (SVE)

tag: age

value: 3 months post implantation


title: SVE (mNKX3.1 + UTY_sh) + UGM, CAD119

description:
CAD119
Processed data file: RNAseq_mouse_recomb.txt


title: Donor number 10, live sorted cells,  library 1

tag: cell type


title: SVE (mNKX3.1 + UTY_sh) + UGM, CAD118

description:
CAD118
Processed data file: RNAseq_mouse_recomb.txt


title: Leaf wounded 24hr rep4


title: Leaf wounded 24hr rep3


title: Leaf wounded 24hr rep2


title: SVE (UTY_sh) + UGM, CAD116

description:
CAD116
Processed data file: RNAseq_mouse_recomb.txt

source_name: tissue recombinants, SVE (UTY_sh) + UGM

value: UTY_sh (SVE)


title: SVE (UTY_sh) + UGM, CAD114

description:
CAD114
Processed data file: RNAseq_mouse_recomb.txt


title: Leaf wounded 2hr rep4


title: Leaf wounded 24hr rep1


title: SVE (UTY_sh) + UGM, CAD115

description:
CAD115
Processed data file: RNAseq_mouse_recomb.txt


title: Leaf wounded 2hr rep3


title: SVE (mNKX3.1) + UGM, CAD112

description:
CAD112
Processed data file: RNAseq_mouse_recomb.txt

source_name: tissue recombinants, SVE (mNKX3.1) + UGM

value: mNKX3.1 (SVE)


title: SVE (mNKX3.1) + UGM, CAD111

description:
CAD111
Processed data file: RNAseq_mouse_recomb.txt


title: Leaf wounded 2hr rep1

value: V5-30 days old


title: SVE (mNKX3.1 + UTY_sh) + UGM, CAD117

description:
CAD117
Processed data file: RNAseq_mouse_recomb.txt


title: Donor number 7 live sorted cells,  library 3


title: Leaf non-infested rep4

value: leaf number 5


title: Donor number 10, live sorted cells,  library 2


title: SVE (mNKX3.1) + UGM, CAD109

description:
CAD109
Processed data file: RNAseq_mouse_recomb.txt


title: SVE (mNKX3.1) + UGM, CAD110

description:
CAD110
Processed data file: RNAseq_mouse_recomb.txt


title: Leaf non-infested rep3


title: PE + UGM, CAD107

description:
CAD107
Processed data file: RNAseq_mouse_recomb.txt

source_name: tissue recombinants, PE + UGM

value: tissue recombinants (mouse prostate epithelial cells + rat urogenital sinus mesenchymal cells)

value: wild-type NKX3.1 (PE)


title: PE + UGM, CAD108

description:
CAD108
Processed data file: RNAseq_mouse_recomb.txt


title: PE + UGM, CAD106

description:
CAD106
Processed data file: RNAseq_mouse_recomb.txt


title: PE + UGM, CAD105

description:
CAD105
Processed data file: RNAseq_mouse_recomb.txt


title: SVE + UGM, CAD104

description:
CAD104
Processed data file: RNAseq_mouse_recomb.txt

source_name: tissue recombinants, SVE + UGM

value: wild-type NKX3.1 (SVE)


title: Leaf non-infested rep2


title: SVE + UGM, CAD103

description:
CAD103
Processed data file: RNAseq_mouse_recomb.txt


title: Nkx3.1 -/- Prostate, CAD66

description:
CAD66
Processed data file: RNAseq_mouse_4_month_prostate.txt

data_processing:
Libraries were sequenced using Illumina HiSeq 2000 at Columbia Genome Center. We multiplexed samples in each lane, which yielded targeted number of single-end/paired-end 100bp reads for each sample, as a fraction of 180 million reads for the whole lane.
We used RTA (Illumina) for base calling and CASAVA (version 1.8.2) for converting BCL to fastq format, coupled with adaptor trimming. We mapped the reads to a reference genome (Mouse: UCSC/mm9) using TopHat (version 2.0.4) with 4 mismatches (--read-mismatches = 4) and 10 maximum multiple hits (--max-multihits = 10). To tackle the mapping issue of reads that are from exon-exon junctions, TopHat infers novel exon-exon junctions ab initio, and combines them with junctions from known mRNA sequences (refgenes) as the reference annotation. We estimated the relative abundance (aka expression level) of genes and splice isoforms using Cufflinks (version 2.0.2) with default settings. We tested for differentially expressed genes under various conditions using DESeq. It is an R package based on a negative binomial distribution that models the number of reads from RNA-seq experiments and tests for differential expression.
Raw RNA-seq counts were normalized (variance stabilization) using DESeq package in R-studio Version 0.99.484.
Genome_build: mm9 (MGSCv37)
Supplementary_files_format_and_content: Tab-delimited text file includes DESeq normalized values for each Sample.

source_name: Prostate tissue, Nkx3.1 -/-, 4 months

extract_protocol:
RNA was extracted using the MagMAX-96 RNA isolation kit (Ambion). Quality control was performed with Agilent Bioanalyser 2100. We used poly-A pull-down to enrich mRNAs from total RNA samples (200ng-1ug per sample, RIN>8 required).
Library preparation was performed using the Illumina TruSeq RNA prep kit. Libraries were then sequenced using Illumina HiSeq 2000 at Columbia Genome Center.

treatment_protocol: Prostate tissue from 4-month-old Nkx3.1+/+ and Nkx3.1-/- mice was harvested, and snap frozen for subsequent molecular analysis.

tag: strain/background

tag: genotype/variation

value: 4 months

value: prostate


title: SVE (UTY_sh) + UGM, CAD113

description:
CAD113
Processed data file: RNAseq_mouse_recomb.txt


title: Leaf non-infested rep1


title: SVE + UGM, CAD101

description:
CAD101
Processed data file: RNAseq_mouse_recomb.txt


title: SVE + UGM, CAD102

description:
CAD102
Processed data file: RNAseq_mouse_recomb.txt


title: RWPE1- NKX3.1 (T164A), CAD94

description:
CAD94
Processed data file: RNAseq_human_RWPE1.txt

data_processing:
Libraries were sequenced using Illumina HiSeq 2000 at Columbia Genome Center. We multiplexed samples in each lane, which yielded targeted number of single-end/paired-end 100bp reads for each sample, as a fraction of 180 million reads for the whole lane.
We used RTA (Illumina) for base calling and CASAVA (version 1.8.2) for converting BCL to fastq format, coupled with adaptor trimming. We mapped the reads to a reference genome (Human: NCBI/build37.2) using TopHat (version 2.0.4) with 4 mismatches (--read-mismatches = 4) and 10 maximum multiple hits (--max-multihits = 10). To tackle the mapping issue of reads that are from exon-exon junctions, TopHat infers novel exon-exon junctions ab initio, and combines them with junctions from known mRNA sequences (refgenes) as the reference annotation. We estimated the relative abundance (aka expression level) of genes and splice isoforms using Cufflinks (version 2.0.2) with default settings. We tested for differentially expressed genes under various conditions using DESeq. It is an R package based on a negative binomial distribution that models the number of reads from RNA-seq experiments and tests for differential expression.
Raw RNA-seq counts were normalized (variance stabilization) using DESeq package in R-studio Version 0.99.484.
Genome_build: GRCh37
Supplementary_files_format_and_content: Tab-delimited text file includes DESeq normalized values for each Sample.

extract_protocol:
RNA was extracted using the MagMAX-96 RNA isolation kit (Ambion). Quality control was performed with Agilent Bioanalyser 2100. We used poly-A pull-down to enrich mRNAs from total RNA samples (200ng-1ug per sample, RIN>8 required).
Library preparation was performed using the Illumina TruSeq RNA prep kit. Libraries were sequenced using Illumina HiSeq 2000 at Columbia Genome Center.

treatment_protocol: Human RWPE1 cells were engineered with empty vector (altered pTRIPZ), NKX3.1 wild type over-expression, and NKX3.1 (T164A) mutant over-expression. Cells were snap frozen for subsequent molecular analysis.

tag: cell line

tag: expression


title: RWPE1- NKX3.1 (T164A), CAD93

description:
CAD93
Processed data file: RNAseq_human_RWPE1.txt


title: Leaf wounded 2hr rep2


title: RWPE1- NKX3.1 (T164A), CAD91

description:
CAD91
Processed data file: RNAseq_human_RWPE1.txt


title: RWPE1- NKX3.1 (T164A), CAD92

description:
CAD92
Processed data file: RNAseq_human_RWPE1.txt


title: Nkx3.1 -/- Prostate, CAD65

description:
CAD65
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: RWPE1 - NKX3.1, CAD17

description:
CAD17
Processed data file: RNAseq_human_RWPE1.txt

source_name: RWPE1, NKX3.1 (WT)


title: RWPE1 - NKX3.1, CAD18

description:
CAD18
Processed data file: RNAseq_human_RWPE1.txt


title: Nkx3.1 -/- Prostate, CAD63

description:
CAD63
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: RWPE1 - NKX3.1, CAD15

description:
CAD15
Processed data file: RNAseq_human_RWPE1.txt


title: Nkx3.1 -/- Prostate, CAD62

description:
CAD62
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: RWPE1 - NKX3.1, CAD14

description:
CAD14
Processed data file: RNAseq_human_RWPE1.txt


title: RWPE1 - NKX3.1, CAD16

description:
CAD16
Processed data file: RNAseq_human_RWPE1.txt


title: RWPE1 - Control Vector, CAD5

description:
CAD5
Processed data file: RNAseq_human_RWPE1.txt

source_name: RWPE1, control vector

value: Control vector


title: RWPE1 - NKX3.1, CAD13

description:
CAD13
Processed data file: RNAseq_human_RWPE1.txt


title: Nkx3.1 +/+ Prostate, CAD42

description:
CAD42
Processed data file: RNAseq_mouse_4_month_prostate.txt

source_name: Prostate tissue, Nkx3.1 +/+, 4 months


title: RWPE1 - Control Vector, CAD1

description:
CAD1
Processed data file: RNAseq_human_RWPE1.txt


title: Nkx3.1 -/- Prostate, CAD61

description:
CAD61
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: RWPE1 - Control Vector, CAD4

description:
CAD4
Processed data file: RNAseq_human_RWPE1.txt


title: RWPE1 - Control Vector, CAD6

description:
CAD6
Processed data file: RNAseq_human_RWPE1.txt


title: RWPE1 - Control Vector, CAD2

description:
CAD2
Processed data file: RNAseq_human_RWPE1.txt


title: Nkx3.1 +/+ Prostate, CAD40

description:
CAD40
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: Nkx3.1 +/+ Prostate, CAD39

description:
CAD39
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: Nkx3.1 +/+ Prostate, CAD41

description:
CAD41
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: Nkx3.1 -/- Prostate, CAD64

description:
CAD64
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: Nkx3.1 +/+ Prostate, CAD38

description:
CAD38
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: GM12878 rep1 input

city: Los Angeles

first: Jinkai

last: Wang

address: 650 Charles E. Young Drive South, CHS 33-228

institute: University of California, Los Angeles

description: without antibody treatment

data_processing:
We mapped strand-specific paired-end RNA-seq reads first to Ensembl genes and then to hg19 human genome incorporated with all spike-in sequences using tophat (v 1.4.1).
Only the properly paired and uniquely mapped reads were used in future analyses.
Genome_build: hg19
Supplementary_files_format_and_content: BigWig files were generated using BEDTools, the socres represent coverage of raw reads.

growth_protocol: cell culture: cultured on complete RPMI media supplemented with 15% FBS and 10% Pen/Strep antibiotic

extract_protocol:
Total RNA was isolated from cells according to manufacturer’s instructions using TRIzol LS reagent (Ambion). Total RNA was treated using DNAse I (Promega) for 20 minutes at 37ºC. The treated RNA was then acid phenol/chloroform extracted and chloroform extracted. The RNA was precipitated using 300mM final concentration of NaCl2 spiked with 1µl of 50mg/ml of Ultra Pure Glycogen (Promega) and 2.5 volume of 100% ethanol at -20ºC either for 2 hours or overnight. The precipitated RNA was then centrifuged using a refrigerated table-top at maximum speed (>13,000g) at 4ºC for 20 minutes. The precipitated RNA was then washed with 70ºC ethanol and centrifuged at maximum speed for an additional 10 minutes. The final pellet was then re-suspended in ultra pure H2O. PolyA RNA selection was performed twice using Dynabeads mRNA Purification Kit (Invitrogen Cat. #610.06) according to the manufacturer's protocol. The second polyA RNA selection was performed using the eluate of the first polyaA RNA selection as starting material according to the manufacture’s instruction.
100ng RNA were used for library construction and m6A-LAIC-seq using TrueSeq Stranded mRNA Sample Preparation Guide, entering the protocol by adding the Fragment, Prime, Finish Mix, skipping the elution step and proceeding immediately to the synthesis of the First Strand cDNA. From that point on, the exact steps of the Illumina TruSeq Stranded mRNA sample Preparation Guide were followed to the end.

value: without antibody treatment


title: Nkx3.1 +/+ Prostate, CAD37

description:
CAD37
Processed data file: RNAseq_mouse_4_month_prostate.txt


title: GM12878 rep1 m6A neg

description: supernatant (m6A negative) fraction of m6A-LAIC-seq


title: H1-ESC rep2 m6A pos

description: eluate (m6A positive) fraction of m6A-LAIC-seq

growth_protocol: cell culture: cultured in feeder-free condition using mTESR1 media on 6-well plates coated with matrigel


title: GM12878 rep2 m6A neg

value: supernatant (m6A negative) fraction of m6A-LAIC-seq


title: GM12878 rep2 m6A pos

value: eluate (m6A positive) fraction of m6A-LAIC-seq


title: GM12878 rep2 input

description: without antibody treatment


title: H1-ESC rep2 m6A neg


title: GM12878 rep1 m6A pos


title: Myelin 24m R2

city: MAinz

first: Vijay

last: Tiwari

address: Ackermannweg 4

institute: IMB Mainz

data_processing:
The FASTQ files were aligned mouse genome (mm9) using TopHat v2.0.8
Samtools v0.1.19 was used for File format conversions (bam to sam)
Reads at transcripts were counted using HTSeq (v0.5.4p1)
Normalization (for library size) was done using DESeq
Differential expression analysis was done using DESeq
Genome_build: mm9
Supplementary_files_format_and_content: Table containing normalized read counts

source_name: myelin extracted from mouse

growth_protocol: A light-weight membrane fraction enriched for myelin was purified from mouse brains as described previously (Jahn et al., Neuromethods, 2013).

extract_protocol:
RNA was isolated using column purification according to manufacture instruction
RNA libraries were prepared for sequencing using standard Illumina protocols

value: myelin

tag: tissue type

value: Primary tissue


title: Myelin 24m R3


title: H1-ESC rep2 input

description: without antibody treatment

tag: treatment


title: H1-ESC rep1 m6A neg


title: Myelin 24m R1

tag: tissue


title: RWPE1 - Control Vector, CAD3

description:
CAD3
Processed data file: RNAseq_human_RWPE1.txt


title: H1-ESC rep1 m6A pos


title: H1-ESC rep1 input

description: without antibody treatment


title: Myelin 6mo R2


title: Myelin 6mo R3


title: Myelin P75 R2


title: Myelin 6mo R1


title: Myelin P75 R3


title: Myelin P75 R1


title: Myelin P18 R3


title: Myelin P18 R1


title: Donor number 3 live sorted cells,  library 4

value: live sorted cells


title: Donor number 3, exocrine fraction, live sorted cells

value: exocrine fraction, live sorted cells


title: Myelin P18 R2


title: NC1+ Cranial Neural Crest B

city: Ithaca

first: Marcos

last: Simoes-Costa

state: New York

address: 526 Campus Rd

department: Molecular Biology and Genetics

institute: Cornell University

data_processing:
Sequence reads were aligned to the Gallus Gallus genome with TopHat.
Cufflinks was used to calculate gene expression levels.
Differentially expressed transcripts were identified with Cuffdiff 2.1.
Genome_build: galGal4
Supplementary_files_format_and_content: Tab-delimited text files include RPKM values and enrichment for each gene.

organism: Gallus gallus

source_name: Cranial neural crest

growth_protocol: Embryos were electroporated with either cranial or trunk neural crest enahncers, and cultured until desired stages.

extract_protocol:
Total RNA was extracted from the sorted RNA with the RNAqueous micro kit (Ambion). cDNA was synthesized and amplified with the Ovation RNA-Seq System V2 (NuGEN).
The sequencing libraries were built according to the Illumina Standard Protocols. SR50 sequencing was performed in a HiSeq2000 Illumina machine.

treatment_protocol: Dissected cranial and trunks were dissociated with Accumax and FACS sorted for GFP+ cells on BD FACSARIA Cell Sorter,  (BD Biosciences), with 7AAD exclusion.

tag: developmental stage

tag: eletroporated with

value: FoxD3 NC1 enhancer

value: cranial neural crest

tag: library id


title: NC2+ Trunk Neural Crest C

source_name: Trunk neural crest

value: FoxD3 NC2 enhancer

value: trunk neural crest


title: Donor number 3 live sorted cells,  library 2


title: NC1+ Cranial Neural Crest A

value: Cranial neural crest


title: Donor number 3 live sorted cells,  library 3


title: Donor number 2, exocrine fraction, live sorted cells


title: Donor number 3 live sorted cells,  library 1


title: sorted hematopoietic stem cells [JC4]

source_name: Bone marrow

value: bone marrow cells

tag: cell subtype

value: hematopoietic stem cells


title: TET #3

status: Public on Jun 23 2016

city: Ann Arbor

first: Stefan

middle: W.

last: Stoll

state: MI

address: 1301 Catherine Street

department: Dermatology

institute: University of Michigan

data_processing:
Sequenced reads were aligned to the reference genome NCBI build 37 using TopHat version 1.3.3. We only used mapped reads with less than 3 mismatches.
We used the genes annotated in RefSeq to quantify gene expression levels. The expression level of each exon was obtained by counting the number of reads mapped to that exon and the expression of a gene is the sum of read counts of all exons in a gene.  Reads mapped to multiple exons in a gene were counted only once.
The gene or exon level expression was normalized to the number of reads per kilobase per million mapped reads (RPKM).
Genome_build: genome NCBI build 37
Supplementary_files_format_and_content: RPKM

source_name: N/TERT-2G_skin keratinocyte_TET

growth_protocol: Keratinocytes were grown in keratinocyte serum-free medium (KFSM, Invitrogen) with 0.4 mM CaCl2.

extract_protocol:
Total RNA was isolated from subconfluent N/TERT keratinocytes treated for 60 hours with and without 1 µg/ml Tet with and without 20 ng/ml EGF (Qiagen RNeasy Minikits, Valencia, CA) and RNA quality and quantity were assessed using an Agilent Bioanalyzer.
Complementary DNA libraries were prepared from polyadenylated RNA, and libraries for high throughput sequencing were prepared using the Illumina mRNA-Seq kit according to the manufacturer’s protocol (Illumina, San Diego, CA). The prepared libraries were sequenced one sample per lane in an 8-lane flow cell on the Illumina Genome Analyzer IIx. One sample per sequencing lane sequencing generated on average 31 million reads, resulting in approximately 2.4 gigabases per sample.

treatment_protocol: Subconfluent N/TERT keratinocytes with tetracycline-inducible AREG shRNA were treated for 60 hours with and without 1 µg/ml Tet with or with Tet and 20 ng/ml EGF.

value: keratinocyte

value: Tet-inducible AREG shRNA

tag: treated with

value: 1 µg/ml Tet for 60hrs


title: TET #1


title: TET #2


title: Control #3

source_name: N/TERT-2G_skin keratinocyte_control

value: none (control)


title: TET-EGF #1

source_name: N/TERT-2G_skin keratinocyte_TET_EGF

value: 1 µg/ml Tet and 20 ng/ml EGF for 60hrs


title: Control #2


title: Control #1


title: p1C-1

city: Shanghai

first: Ligang

last: Wu

state: Shanghai

address: 320 Yueyang Road

department: Shanghai Institutes for biological Sciences

country: China

institute: Chinese Academy of Sciences

data_processing: The raw reads were quality filtered and collapsed with FASTX Toolkit (http://hannonlab.cshl.edu/fastx_toolkit/). Read sequences longer than 17 nt after adapter trimming were aligned to the reference genome (mm10 for mouse, hg38 for human) with no mismatch allowed. The genome mapped reads were aligned to the libraries of miRNAs (miRBase v. 21), tRNAs, rRNAs, sn/snoRNAs (small nuclear/small nucleolar RNAs), endo-siRNA, piRNAs, and repetitive elements sequentially. Normalization was performed by dividing the read counts of the individual small RNA by the total number of reads mapped to the genome and presented as RPM.

source_name: parthenogenetic one-cell embryos

extract_protocol: cDNA libraries of small RNAs were constructed using HEK293 total  RNA, with threefold serial dilutions ranging from 100 to 1 ng. For  mouse oocyte and embryo samples, total RNA was extracted from  111 MII oocytes, 167 1C embryos, 137 2C embryos, 57 4C embryos,  102 8C embryos, 200 p1C embryos, or ~3 × 106 sperms using TRIzol  Reagent (Ambion) containing 45 mM MgCl2 (72), respectively. The  quantity of isolated RNA was measured with Qubit 2.0 Fluorometer  (Invitrogen). Each cDNA library of small RNAs was constructed using  10 ng of total RNA with two technical replicates.

tag: replicate

value: technical replicate 1

tag: strain

value: FVB/N

value: 8 to 12 weeks old

tag: Sex

value: female

value: Parthenogenetic embryos in 1-cell stage


title: 4C-2

source_name: 4-cell embryos

value: technical replicate 2

tag: age

value: Embryos in 4-cell stage


title: 4C-1


title: 2C-1

source_name: 2-cell embryos

value: Embryos in 2-cell stage


title: 2C-2


title: 8C-2

source_name: 8-cell embryos

value: Embryos in 8-cell stage


title: 1C-3

source_name: zygotes

value: technical replicate 1 of the second biological replicate sample

value: Embryos in zygote stage


title: Oo-1

source_name: oocytes

value: Oocyte in metaphase II


title: Oo-2


title: HOXA9-MEIS1/ H3K4me1

city: Birmingham

first: Rui

last: Lu

state: AL

address: 1824 6th Ave S

institute: Univ of Alabama at Birmingham

data_processing:
All reads were mapped to the mouse genome (mm9) using the BWA alignment software. Unique reads mapped to a single best-matching location with no more than two mismatches were kept, which were then subject to removal of reads generated by PCR- caused duplicates using the Picard and Samtools.
Profiles of ChIP-Seq read densities were displayed in the Integrative Genomics Viewer (IGV, Broad Institute) and  the Integrative Genomics Browser (IGB, Affymetrix).
The MACS2 software was used for peak identification with data from input as controls and default parameters.
Genome_build: mm9
Supplementary_files_format_and_content: The bedGraph files, which were generated by MACS2 software with default parameters, report the number of ChIP-Seq sequencing reads in a genomic interval (normalized to a read depth of 1 million reads).

source_name: HOXA9-MEIS1 LSCs

growth_protocol: RH-RAS and other Leukemia stem cell (LSC) lines were cultured in Opti-MEM base medium supplemented with 10% FBS, 1% antibiotics, 50 μM mercaptoethanol and stem cell factor

extract_protocol:
Lysates were clarified from sonicated nuclei and DNMT3A R882H- or histone-DNA complexes were isolated with indicated antibodies.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced following the manufacturer's protocols.

value: HOXA9-MEIS1 LSC

tag: chip antibody


title: 1C-2

value: technical replicate 2 of the first biological replicate sample


title: 1C-4

value: technical replicate 2 of the second biological replicate sample


title: LSC RH-RAS/ H3K4me3

source_name: RH-RAS LSCs

value: RH-RAS LSC


title: LSC RH-RAS/ H3K27me3


title: HOXA9-MEIS1/ Input


title: LSC RH-RAS/ Input


title: LSC RH-RAS/ Myc-DNMT3AR882H (9E10)

value: myc tag, 9e10


title: p1C-2

value: technical replicate 1


title: TET-EGF #2

tag: cell type

value: keratinocyte

value: Tet-inducible AREG shRNA

value: 1 µg/ml Tet and 20 ng/ml EGF for 60hrs


title: EV_RAS_eRRBS

data_processing:
Sequence reads from bisulfite-treated libraries were identified using standard Illumina base-calling software.
Residual cytosines (Cs) in each read were first converted to thymines (Ts), with each such conversion noted for subsequent analysis. A reference sequence database was constructed from the 50-bp ends of each computationally predicted MspI-TaqI fragment in the 40–350 bp size range.
All Cs in each fragment end were then converted to Ts; the converted reads were aligned to the converted reference by Bowtie. The number of mismatches in the induced alignment was then counted between the unconverted read and reference, ignoring cases in which a T in the unconverted read is matched to a C in the unconverted reference. For a given read, the best alignment was kept. If there was more than one best alignment, the read was discarded as non-unique.
The methylation level of each sampled cytosine was estimated as the number of reads reporting a C, divided by the total number of reads reporting a C or T.
Genome_build: mm9
Supplementary_files_format_and_content: bigBed files contain all mapped CpGs with the number of methylated reads and number of total reads listed.

source_name: Primary HSPC Cells

growth_protocol: Lin- enriched murine hematopoietic stem/progenitor cells leukemia progenitor cell lines were cultured in Opti-MEM base medium supplemented with 10% FBS, 1% antibiotics, 50 μM mercaptoethanol and stem cell factor

extract_protocol:
Genomic DNA was extracted using DNeasy Blood & Tissue Kit (Qiagen).
eRRBS was carried out with a previously described protocol with slight modification (Garrett-Bakelman et al., 2015). Briefly, ~300 ng of genomic DNA were digested with three different enzymes (~80 units of MspI, 40 units of BfaI and 40 units of MseI) to enhance genomic fragmentation and coverage. The produced fragments were ligated to pre-annealed adapters containing 5’-methyl-cytosine instead of cytosine, followed by overhang fill-in, 3’-terminal-A extension and purification. Bisulfite treatment of the fragments was done using the EZ DNA Methylation–Lightning kit (Zymo Research). Amplified eRRBS libraries was quality checked with Agilent 2200 TapeStation, followed by deep sequencing on the Illumina HiSeq-2000 genome analyzer with 50 bp SE parameters.

treatment_protocol: Primary HSPC Cells were retrovirally infected with NRAS G12D alone (EV-RAS), DNMT3A R882H with NRAS G12D (RH-RAS) or DNMT3A WT with NRAS G12D (WT-RAS)

value: Lin- enriched hematopoietic stem/progenitor cells

tag: genotype

value: NRAS G12D alone (EV)

tag: time point

value: day 16 post-transduction


title: 8C-1

value: FVB/N

value: 8 to 12 weeks old

value: female


title: HSPC_EVRAS_INPUT_ForH3K79me2

tag: chip batch

value: batch 2


title: HSPC_RHRAS_H3K79me2

value: DNMT3A R882H with NRAS G12D

value: H3K79me2 (Abcam, catalog# ab3594, lot# GR201271-1)


title: HSPC_EVRAS_H3K79me2


title: HSPC_RHRAS_INPUT_ForH3K79me2


title: HSPC_WTRAS_INPUT

value: DNMT3A WT with NRAS G12D

value: batch 1


title: NC2+ Trunk Neural Crest A

value: FoxD3 NC2 enhancer

value: trunk neural crest


title: HSPC_RHRAS_INPUT


title: HSPC_WTRAS_H3K4me1

value: H3K4me1 (Abcam, catalog# ab8895, lot# GR114265-2)


title: HSPC_WTRAS_H3K27ac

value: H3K27ac (Abcam, catalog# ab4729, lot# GR183919-1)


title: HSPC_RHRAS_H3K27ac


title: HSPC_RHRAS_H3K4me1


title: 1C-1

value: technical replicate 1 of the first biological replicate sample

value: Embryos in zygote stage


title: HSPC_EVRAS_INPUT

value: Lin- enriched hematopoietic stem/progenitor cells


title: HSPC_EVRAS_H3K27ac

value: NRAS G12D alone (EV)


title: LSC RH-RAS/ H3K4me1

tag: cell line

value: RH-RAS LSC


title: Yorkshire pig

city: Changsha City

first: Hu

last: Yang

address: Furong District

institute: Hunan Agricultural University

data_processing:
Basecalls performed using CASAVA version 1.8
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence using FASTX-Toolkit  version 0.0.13 and Perl version 5.8.8
mRNA-seq reads were aligned to the mm10 genome assembly using Tophat version 2.0.6 with parameters: -r 100 -I 50000  --max-segment-intron 50000 -a 10 -g 10
Gene abundance measurements were generationed and normalized  using Cufflink version 2.0.2.Linux_x86_64 ; The normalized standard were Fragments Per Kilobase of exon per Megabase of library size (FPKM)
Differential expression were analysed  using DEGseq function package in R program version 2.10.0
Genome_build: UCSC mm10
Supplementary_files_format_and_content: tab-delimited text files include normalized RPKM values and raw fragment counts for each Sample

organism: Sus scrofa

source_name: longissimus dorsi muscle

extract_protocol:
Total RNA was extracted with Trizol Reagent (Invitrogen, Carlsbad, CA, USA).
Following "TruSeq RNA Sample Preparation Guide".

treatment_protocol: longissimus dorsi muscle were dissected and stored in liquid nitrogen until RNA extraction

value: longissimus dorsi muscle

tag: Stage

value: Chinese indigenous pig breeds


title: Shaziling pig

tag: tissue

value: lean-type pig breeds


title: RH_RAS_eRRBS

value: day 16 post-transduction


title: WT_RAS_eRRBS


title: HSPC_EVRAS_H3K4me1


title: Grass_222 (muscle)

city: College Park

first: Jose

middle: Adrian

last: Carrillo

state: MD

address: 1413 Animal Sciences Center

department: Department of Animal and Avian Sciences

institute: University of Maryland

data_processing:
Illumina HiSeq 2000 sequencer for basecalling
Alignment to the reference genome (Bos_taurus_UMD3.1/bosTau6) obtained from the UCSC (http://genome.ucsc.edu/) was performed employing Bowtie (Ultrafast, memory-efficient short read aligner). During this step, the first 15 bases of each read (50 bp) were trimmed to avoid low Phred quality scores, resulting in 35 bp tags
Counting of reads per gene was executed by the summarizeOverlaps function implemented in R
The identification of differentially expressed genes was achieved employing the edgeR software package and the included generalized linear model (GLM) approach, which requires a design matrix to describe the treatment conditions (grass-fed and grain-fed).
Genome_build: Bos_taurus_UMD3.1/bosTau6
Supplementary_files_format_and_content: BovMuscleCountRaw_2.xls: Raw read counts per gene
Supplementary_files_format_and_content: MuscleCountNormalizededgeR_2.xls: Normalized read counst per gene using edgeR
Supplementary_files_format_and_content: BovMuscleSigensembl_edgeRGLM_2.xls: Differentially expressed genes, with statistical measurement obtained from edgeR using GLM approach

organism: Bos taurus

source_name: longissimius dorsi muscle

growth_protocol: The animals were born, raised and maintained at the Wye Angus farm. All animals received the same diet until weaning

extract_protocol:
Total RNA was extracted individually (two animals per group) using Trizol (Invitrogen, Carlsbad, CA, USA) followed by DNase digestion and Qiagen RNeasy column purification (Invitrogen, Carlsbad, CA, USA). The RNA sample was dissolved in RNAse-free H2O; the integrity and quality of RNA were then checked by a NanoDrop 1000 spectrophotometer and by resolution on a 1.5% non-denaturing agarose gel
Each library was identified by adding 6-bp adaptors and sequenced at 50 bp/sequence read using an Illumina HiSeq 2000 sequencer

treatment_protocol: After weaning, the calves were randomly assigned to one diet and exclusively received that regimen until termination. The grain group received conventional diet consisting of corn silage, shelled corn, soy bean and trace minerals. The grass fed steers consumed normally grazed alfalfa; during wintertime, bailage was utilized. The alfalfa has been harvested from land without any fertilizers, pesticides or other chemicals. The steers ate no animal, agricultural or industrial byproducts and never receive any type of grain

tag: breed

value: Aberdeen Angus

tag: treatment

value: Grass-fed

value: Muscle


title: Grain_224 (muscle)

value: Grain-fed


title: Grain_184 (muscle)


title: Grass_104 (muscle)


title: Mouse brain tissue_rep2

city: La Jolla

first: Sheng

last: Zhong

state: California

address: 9500 Gilman Drive, Mail Code 0412

department: Bioengineering Department

institute: University of California, San Diego

description:
Sample name: Brain_2
crosslinked RNA

data_processing:
Library strategy: MARIO
MARIO-tools is a package of command-line tools for analyses of MARIO data. It is written   in Python and R and is version controlled by GitHub. The full documentation is at http://mariotools.ucsd.edu/. The pipeline takes pair-end sequencing reads as input. The oligonucleotide sequences of the RNA linker and the sample barcodes used for multiplexed sequencing should also be provided to the pipeline. Script files mentioned below are all components from the pipeline.
Removing PCR duplicates. The forward read contains a 4nt sample barcode and a 6nt random barcode at the 5' end. A read pair was classified as a PCR duplicate of another read pair and is therefore discarded if the two read pairs had identical sequences and contained identical barcodes (10nt). The tool ‘remove_dup_PE.py’ provides this function, and generates a fastq/fasta file containing the non-duplicated reads, and reports the number of duplicates removed.
Assigning multiplexed sequencing reads into corresponding experimental samples. The tool ‘split_library_pairend.py’ assigns each pair-end read into a sample by matching the sample barcode in each read with those in the list of sample barcodes (a user input text file), generates a fastq/fasta file for the reads assigned to each sample, as well as a fastq/fasta file for the unassigned reads.
Recovering the cDNAs in the sequencing library. This step identifies the overlapping regions of the two ends of every read pair, if any. It also recovers the entire sequences of the cDNAs in the sequencing library, whenever possible. This function is achieved by ‘recoverFragment.py’, which uses local alignment to identify the overlapping regions. When the overlap was small (15bp or less) compared to read length (100bp on each end), local alignment could be insensitive. To overcome this insensitivity, ‘recoverFragment.py’ collects the read pairs without identifiable overlaps after the first alignment, truncates each read into one third of its length (retaining 33bp at the 3’ of each read), and repeats local alignment.
Parsing the chimeric cDNAs. This step categorizes the cDNAs based on their configurations. This takes the completely and partially recovered cDNA sequences, as well as the linker sequence as inputs. It identifies the location of the linker in the cDNA, and generates five categories of cDNAs based the locations of the linker sequence.
Mapping to the genome. Hereafter, all analyses were based on the RNA1-Linker-RNA2 type of read pairs. First, any cDNA containing less than 15bp on either the RNA1 or RNA2 side of linker was discarded, because it is unlikely to uniquely map a 15bp or less sequence to the genome in the mapping step. Then the two Mapping to the genome. RNA fragments on each side of the linker (RNA1 and RNA2) were separately mapped to the mouse genome mm9/NCBI37 using Tophat version 2.1.1. This step, implemented in ‘Stitch-seq_Aligner.py’ outputs the read pairs where both RNA1 and RNA2 were uniquely mapped to the genome. The aligned tab-delimited text files are results from this step.
The annotations were retrieved from Ensembl (release 67, mouse NCBIM37), including the genes of mRNAs, lincRNAs, rRNAs, snRNAs, snoRNAs, miRNAs, misc_RNAs, tRNAs, and transposons. The different genomic copies of the same transposon were considered as different genes in this analysis. The reads mapped to rRNAs were removed from further analysis. The number of uniquely aligned reads (from either RNA1 or RNA2 of the RNA1-Linker-RNA2 type) were counted on every gene. Any gene with a read count less than 5 was filtered out. Next, the association between any two genes was tested with Fisher’s exact test. The null hypothesis was that gene A and gene B independently contributed to the sequencing reads. The alternative hypothesis was that their contributions to read counts were associated. We denoted c_A, c_B as the read counts for gene A and gene B, respectively, and I_{A,B} as the read counts of co-appearance, where the two genes co-appeared on the same read pair. A Fisher’s exact test was carried out on each gene pair, with c_A, c_B, I_{A,B}, and the read counts on other genes besides gene A (gene B) as the test statistics. Both p-values and FDRs (Benjamini-Hochberg procedure) were calculated for every gene pair. This step outputs gene pairs with FDR < 0.05 and fold change (FC) >= 3. The FC was calculated as (I_{A,B} + 0.5) / (I'_{A,B} + 0.5), where I'_{A,B} was the co-appearing read counts in the control sample (ES-indirect). This step was implemented in ‘Select_strongInteraction_RNA.py’ which outputs strong interacting RNA pairs with information of their interaction regions, number of supporting pairs, p-value of significance, FDR and fold changes. The strong interaction clusters tab-delimited text files are from this step.
For further analysis steps to identify RNA interaction sites, calculating binding energies between the interaction sites, network analysis, intra-molecule cutting/ligation detection and RNA secondary structure prediction, please refer to corresponding manuscript.
Genome_build: mm9
Supplementary_files_format_and_content: The *fragment_paired_align.txt.gz file column headers: Chromosome_RNA_1, Start_RNA_1, End_RNA_1, Strand_RNA_1, Fragment_in_read_RNA_1, Reference_RNA_1, Type_RNA_1, Name_RNA_1, Sub_type_RNA_1, Strand_Properness_RNA_1, Read_ID, Chromosome_RNA_2, Start_RNA_2, END_RNA_2, Strand_RNA_2, Fragment_in_read_RNA_2, Reference_RNA_2, Type_RNA_2, Name_RNA_2, Sub_type_RNA_2, Strand_Properness_RNA_2.

source_name: Brain tissue_technical replicate

extract_protocol:
Adult female was sacrificed by cervical dislocation and the whole brain was immediately collected, rinsed with ice-cold PBS three times and snap frozen. Frozen whole mouse brain tissue was ground into fine powder in liquid nitrogen using a mortar and pestle. The tissue powder was quickly transferred into a Petri dish on a bed of dry ice and irradiated on dry ice three times at 400 mJ/cm2 in a UV crosslinker (254 nm) with gentle swirling between each irradiation. Crosslinked powdered tissue was immediately lysed and subjected to MARIO procedure as described.
Protein-RNA complex was eluted from streptavidin beads and RNA is recovered by digesting the bound protein. Eluted RNA was subjected to rigorous DNase treatment to eliminate DNA contamination. Purified RNAs were hybridized with a DNA probe that is complementary to the biotin-tagged RNA linker, and treated with T7 exonuclease to remove the non-ligated biotinylated RNA linkers. As a result, mainly the successfully ligated chimeric RNAs retained a biotin-tagged linker at the junction.
This chimeric RNA library was fragmented again to an average of 150 nucleotides, and the ligation junctions were pulled-down with streptavidin-coated magnetic beads. The end product was a library of ~150nt chimeric RNAs. This library was enriched with chimeras of in the form of 5'-R1-linker-R2, where R1 and R2 were fragments of interacting RNAs. This library was converted into cDNAs and sequenced with paired-end next-generation sequencing.

value: brain

tag: gender

value: female


title: Mouse brain tissue_rep1

description:
Sample name: Brain_1
crosslinked RNA


title: Tfrc RIA sample

description: Tfrc antisense oligonucleotides used to pull down RNA from cell samples

data_processing:
The mapping method is the same as described in GSE61489 and is shown below.
Removing PCR duplicates. The forward read contains a 4nt sample barcode and a 6nt random barcode at the 5' end. A read pair was classified as a PCR duplicate of another read pair and is therefore discarded if the two read pairs had identical sequences and contained identical barcodes (10nt). The tool ‘remove_dup_PE.py’ provides this function, and generates a fastq/fasta file containing the non-duplicated reads, and reports the number of duplicates removed.
RNA were separately mapped to the mouse genome transcriptome and mm9/NCBI37 using Bowtie version 0.12.7 (24), and parameters -f -n 1 -l 15 -e 200 -p 9 –S. This step, implemented in ‘Stitch-seq_Aligner.py’ outputs the reads uniquely mapped to the genome. The aligned tab-delimited text files are results from this step.
The annotations were retrieved from Ensembl (release 67, mouse NCBIM37), including the genes of mRNAs, lincRNAs, rRNAs, snRNAs, snoRNAs, miRNAs, misc_RNAs, tRNAs, and transposons. The different genomic copies of the same transposon were considered as different genes in this analysis. The reads mapped to rRNAs were removed from further analysis. The number of uniquely aligned reads were counted on every gene. The reads mapped to transcriptome and genome is marked separately to indicate potential pre-mRNA alignment.
Genome_build: mm9

source_name: ES cells

growth_protocol: Undifferentiated mouse E14 ES cells were cultured under feeder-free conditions. ES cells were seeded on gelatin-coated dishes and were cultured in Dulbecco's modified Eagle medium (DMEM; GIBCO) supplemented with 15% fetal bovine serum (FBS; Gemini Gemcell), 0.055 mM 2-mercaptoethanol (Sigma), 2 mM Glutamax (GIBCO), 0.1 mM MEM nonessential amino acid (GIBCO), 5,000 U/ml penicillin/streptomycin (GIBCO) and 1,000 U/ml of LIF (Millipore). The cells were maintained in an incubator at 37 °C and 5 % CO2.

extract_protocol:
Cells were rinsed with PBS for 3 times and scraped off plates and centrifuged at 1500g for 5 minutes. Then pellets were lysed in lysis buffer (50mM Tris-HCl, pH7.0, 10mM EDTA, 1%SDS and freshly added DTT, phenylmethylsulphonyl fluoride (PMSF), Protease Inhibitor Cocktail Tablets). Lysate was incubated on ice for 20 minutes and sonicated by Covaris, and then diluted by two folds of hybridization buffer (500mM NaCl, 1%SDS, 100mM Tris-HCl, pH7.0, 10mMEDTA, 15%formamide, and freshly added DTT, PMSF, protease inhibitor, RNase-in). Antisense oligonucleotides for the target RNA were incubated with the lysate, rotating at 37 C overnight. Streptavidin beads were added to the hybridization buffer and incubated for 30 minutes at 37 C. Beads were accumulated by magnet rack and washed five times with wash buffer (2XSSC, 0.5%SDS, PMSF).RNA were eluted by 200ul RNA proteinase K buffer (100mMNaCl, 10mM TrisHCl, pH7.0, 1mM EDTA, 0.5%SDS, 1mg/ml proteinase K) and incubated at 50 C for 45 minutes, followed by 10 minutes boiling.
RNA was purified by Trizol. Eluted RNA was treated by DNase and converted into cDNA by Ovation V2 kit (NuGEN), and subsequently converted into sequencing library using KAPA Library Preparation Kit.

treatment_protocol: RNA pulldown based on antisense nucleotides is done following the method described in Kretz M. et al. (Nature. 2013 Jan 10;493(7431):231-5). Mouse ES cells were fixed by 1% glutaraldehyde for 10 minutes at room temperature, followed by 0.125 M glycine for 5 minutes to quench the reaction.

value: Tfrc antisense