title: V5 pol II ip

data_processing:
Base Calls were performed using CASAVA version 1.8.2
Reads were aligned to Drosophila genome dm3 with bowtie using the following parameters: -v 2 --best --strata -k 1 -m 3
Peaks were called from the aligned reads using MACS2 with the option --nomodel, and a q-value cutoff of 0.001 for yki and RNA polII, and 0.01 for sd.
Genome_build: dm3
Supplementary_files_format_and_content: BED files of called peaks were generated using MACS2, scores represent the -log10 of the q-value.

source_name: eye disks from 3rd instar larvae

extract_protocol:
Chromatin immunoprecipitation from eye discs was performed using a modified protocol from (Gaertner et al., 2012). First, third instar larvae were dissected in PBS (pH 7.4) such that only eye discs and brain remained attached to the mouth hooks. The dissected material was subsequently fixed in 1ml fixation buffer (50 mM HEPES, pH 7.5; 1 mM EDTA; 0.5 mM EGTA; 100 mM NaCl; 2% formaldehyde) for 30 minutes at room temperature. After four washes (PBS, pH 7.4; 0.1% Triton X-100; 0.1% Tween-20), eye discs were hand-dissected and combined into pools of 200 discs in 300 µL buffer A2 (15 mM HEPES, pH 7.5; 140 mM NaCl; 1 mM EDTA; 0.5 mM EGTA; 1% Triton X-100; 0.1% sodium deoxycholate; 0.1 % SDS; 0.5 % N-lauroylsarcosine; 1x Roche complete protease inhibitor cocktail, cat# 5056489001). Sonication was performed in a Bioruptor sonicator for 30 minutes (30 sec on/off cycle at the "high" setting). Following centrifugation (16,000 x g; 10 minutes at 4°C), the supernatant containing soluble chromatin was transferred to fresh tubes, and 50 µL was set aside as whole cell extract (WCE; input). Per ChIP, 10 ug antibodies were added to 450 µl chromatin (corresponding to approximately 300 discs) and incubated overnight at 4 °C with rotation. We used the following antibodies: anti-Pol II (Covance 8WG16, cat# MMS-126R; mouse monoclonal antibody), anti-Sd, and anti-Yki. Immunocomplexes were purified by adding 50 µL pre-washed Dynabeads coated with protein A/protein G (Life Technologies, cat#10002D and 10004D) for four hours, rotating at 4 °C. The beads were washed three times in RIPA buffer (50 mM HEPES, pH 7.5; 1 mM EDTA; 0.7% sodium deoxycholate; 1% NP-40; 500 mM LiCl) and once in TE. Immunoprecipitated DNA was eluted twice in 75 µL elution buffer (50 mM Tris, pH 8.0; 10 mM EDTA; 1% SDS) at 65°C to maximize yields. Crosslinks of ChIP and WCE DNA were reversed over night at 65°C. DNA was purified by RNAse A (Sigma, cat# R6513; [0.2 µg/µl]; 1 h at 37 °C) and proteinase K (Life Technologies, cat# AM2546; [0.2 µg/µl]; 2 h at 55 °C) treatment followed by phenol/phenol-chloroform-isoamylalcohol extractions and ethanol precipitation. The precipitated DNA was resuspended in 30 ul 10 mM Tris buffer (pH 8).
For ChIP-seq library preparation, 30 ng ChIP DNA and 100ng WCE DNA were used to construct ChIP-Seq libraries with the NEBNext ChIP-seq Library Prep Master Mix Set (cat# E6200L) and the NEBNext Multiplex Oligos (cat# E7335S and E7500S) for Illumina, following the manufacturer's instructions. Concentration and size distribution of the libraries were assessed on an Agilent 2100 Bioanalyzer (High Sensitivity DNA assay chip).

value: GMR-Gal4; UAS-Drosophila Yki-V5

value: Ectopic cell proliferation

tag: antidody

value: Anti-Pol II


title: V5 input DNA

status: Public on Mar 01 2014

type: SRA

city: Kansas City

first: Chris

middle: W

last: Seidel

state: MO

department: Genomics

country: USA

institute: Stowers Institute

library_source: genomic

molecule: genomic DNA

organism: Drosophila melanogaster

tag: genotype

tag: phenotype


title: HD3 input control [ChIP-Seq]

city: Vienna

first: Artem

middle: V

last: Artemov

address: Spitalgasse 4

department: Center for Brain Research

country: Austria

institute: Medical University of Vienna

description: HD3 input

data_processing:
Basecalling with CASAVA 1.8.2 software
Mapping to galGal4 genome with bowtie2 aligner
ChIP peak calling with MACS software with P-value threshold of 10^-5. One sample was used as input DNA control.
Genome_build: Gallus_gallus-4.0
Supplementary_files_format_and_content: Bed file reports MACS peaks.

organism: Gallus gallus

source_name: erythroblast cell line

growth_protocol: The avian erythroblastosis virus-transformed chicken erythroblast cell line HD3 (clone A6 of the line LSCC (Beug, 1979;Beug, 1979) and the DT40 lymphoid cell line (CRL-2111, ATCC) were grown in Dulbecco’s Modified Eagle Medium supplemented with 2% chicken serum and 8% fetal bovine serum at 37ºC with 5% CO2. In the case of DT40 cells, the medium additionally contained 50 μM β-mercaptoethanol.

extract_protocol:
Chromatin immunoprecipitation was performed essentially as described prevoiusly (Orlando, 2000) with modifications presented in the Life Technologies protocol (http://tools.lifetechnologies.com/content/sfs/manuals/DynabeadsProteinG_man.pdf). About 108 logarithmically growing cells were fixed with 1% v/v formaldehyde in DMEM/F-12 (1:1) medium (Invitrogen, 42400-010) at room temperature for 8 min. The fixed cells were pelleted at 700xg for 4 min at 4oC, washed with PBS containing 1 mM AEBSF and 2 ul/ml of a protease inhibitor cocktail (Sigma, P8340), pelleted again and resuspended in 500 μl of lysis buffer (50 mM Tris-HCl pH 8.0, 1% SDS, 10 mM EDTA). The lysate was incubated for 10 min on ice, and sonicated with a Cole-Parmer CP750 ultrasonic processor (30% amplitude, 40 cycles for 3 s each with 10 s intervals). Cell debris was removed in a microcentrifuge (10 min, 13000 rpm, 4°C), and the supernatant was diluted 10-fold with 16.7 mM Tris-HCl, pH 8.0, 16.7 mM NaCl, 1.2 mM EDTA, 1% Triton X-100, 0.01% SDS, 1 mM AEBSF, and 1 μl/ml of the protease inhibitor cocktail. Aliquots were taken at this stage for use as an input control. The cell lysates were precleared by incubation with the Dynabeads Protein G magnetic beads (Life Technologies) and then incubated with 30 μg of anti-CTCF antibodies overnight at 4°C with rotation. Rabbit custom anti-CTCF antibodies were prepared against the part of the chicken CTCF gene encoding the N-terminal region of the protein comprising amino acids 86-233. After incubation with antibodies, DNA-protein complexes were collected with the Dynabeads Protein G magnetic beads and washed as described by the manufacturer (Life Technologies) and eluted by two incubations for 15 min in elution buffer (1% SDS, 0.1 M NaHCO3) at room temperature.
The part of the chicken CTCF gene encoding the N-terminal region of the protein comprising amino acids 86–233 was cloned in the pET-28a(+) vector (Novagen). Protein expression was induced with IPTG and purification was carried out through a Ni(+) charged column. Once the protein was obtained, antibodies were generated by immunizing male New Zealand white rabbits. Antibody titers were estimated by ELISA.
DNA samples were purified using the QIAquick Gel Extraction Kit as recommended by the manufacturer (Qiagen).

treatment_protocol: To induce erythroid differentiation, HD3 cells at a density of 8*10^5 cells/ml were incubated in the above medium additionally containing 10mM HEPES (pH 8.0) and 20 mM iso-H-7 (1-(5-Isoquinolinylsulfonyl)-3-methylpiperazine dihydrochloride, Fluka, Seelze, Germany) at 428C in 100% air atmosphere (Nicolas,R.H, 1991). To estimate the percentage of dead cells, seven parts of the cell suspension in growth medium were mixed with one part of 0.4% trypan-blue solution (Gibco, Carlsbad, CA, USA) and the percentage of stained blue cells was determined under a light microscope.  To estimate the percentage of cells containing hemoglobin, 25 ml of 0.4% (w/v) solution of benzidine (Sigma, St.Louis, MO, USA) in 4% (v/v) acetic acid was mixed with 1 ml of 30% H2O2 and then 25 ml of the cell suspension in growth medium was added. After 10 min staining, the percentage of stained dark blue cells was determined under a light microscope (Rowley,P.T.,1985).

tag: cell line

value: HD3 erythroblast cell line

tag: cell line details

value: clone A6 of the line LSCC

tag: chip antibody


title: NPRL3 DT40 rep 1 [4C-Seq]

data_processing:
Library strategy: 4C-Seq
Basecalling with CASAVA 1.8.2 software
Merging the reads from L and S fractions where both fractions were sequenced (see library construction protocol)
Trimming read subsequences originating from the bait region. Leaving only one read from the pair, the subsequence after HindIII restriction site
Mapping to galGal4 genome with bowtie aligner
For every HindIII restriction fragment, summing up the number of reads mapped to the left and the right end of the fragment
Genome_build: Gallus_gallus-4.0
Supplementary_files_format_and_content: bedGraph files containing the number of 4C reads (score column) per HindIII restriction fragment

source_name: lymphoid cell line

extract_protocol:
PCR reactions were performed using the Expand Long Template PCR system (Roche). The 50 µl PCR reaction contained 120 ng of the DNA template, 1× PCR buffer 1, 0.3 µM of each of the primers, 0.35 mM of each of dNTPs, and 3.75 units of Hot start Taq DNA polymerase (SibEnzyme)Expand Long enzyme mix (Roche). The sequences of HindIII / DpnII 4C primers are as follows (5’-3’): TSR3 bait CACTCATCTCCCCGTACTTTG / AAGTTTCTTTTAATTTGGAGACTTTC; GENE-Des bait AATTTGTGAAGCAGTTGTATGTAGTC / TCTTCTCCACATAATCCCACACT; NPRL3 bait GCCAGGATATAGATTCTGCTTT / CCTCTGACATAATTGCCGATAG; TRAP1 bait CCAGAGATTCTCAAATCACAGCA / CTATGGGGACAAGTGAGGAACAG; PPL bait AAAGCATCTCCTCTCCCTGAAG / GTCTCCCACAGTCACTCCTCCT. PCR amplification was performed in the linear range as follows: initial denaturation for 2 min at 94°C; 30 cycles of 15 s at 94°C, 1 min at 57°C, and 3 min at 68°C; final elongation for 4 min at 68°C. PCR products were purified and concentrated using QIAquick PCR purification kit (Qiagen). After separation according to sizes in 1.5% agarose gel, two zones - 70-400 bp (S fraction) and 400-1500bp (L fraction) - were cut. After extraction with QIAquick gel extraction kit (Qiagen), the fragments of L-fraction were sonicated to reduce the fragment size to 100-500 nucleotides using Covaris S220 with following parameters: time 300 seconds, duty cycle 10%, peak power 23W. Then S and L fraction were processed using TruSeq DNA sample prep kit v.2 (Illumina) with post-PCR size selection on agarose gel (fragments with length 200-600 nucleotides were selected). After purification library concentrations were measured using Qubit fluorimeter (Invitrogen) and real-time PCR with primers complementary to distal regions of Illumina adapters (I-qPCR-1.1: AATGATACGGCGACCACCGAGAT and I-qPCR-2.1: CAAGCAGAAGACGGCATACGA). Then libraries from S and L fractions were combined in equal amounts, diluted to 2nM, denatured using 0.1M NaOH and diluted to final concentration 10 pM using HT1 buffer (Illumina).
Cluster generation was performed using cBot instrument and TruSeq PE Cluster Kit v3 reagents and sequencing - using HiSeq2000 instrument and TruSeq SBS Kit v3 reagents (Illumina) with read length 101 nucleotides from each end.

treatment_protocol: 1×107 cells were fixed with 2% formaldehyde in DMEM medium supplemented with 10% fetal bovine serum (FBS) for 10 min at room temperature, and then the reaction was stopped by the addition of glycine to 0.125 M. After washing with PBS/10% FBS, the fixed cells were incubated for 10 min in an ice-cold lysis solution (10 mM Tris pH 8.0, 10 mM NaCl, 0.2% Nonidet P40, and a protease inhibitor cocktail (Fermentas)) at a concentration of 2×107 cells/ml to release the nuclei. The nuclei were harvested and suspended in 0.5 ml of 1.2× restriction buffer 2 (New England Biolabs). SDS was added to a final concentration of 0.3%, and the solution was incubated for 1 h at 37°C with shaking. Triton X-100 was added to 1.8%, and the solution was further incubated for 1 h at 37°C to sequester the SDS. The DNA was digested by overnight incubation with 800 units of HindIII (New England Biolabs) at 37°C with shaking. The restriction endonuclease was inactivated by the addition of SDS to a final concentration of 1.6% and incubation for 20 min at 65°C. The solution was diluted by adding 7 ml of 1× ligation buffer (Fermentas). Triton X-100 was added to 1%, and the solution was incubated at 37°C for 1 h while shaking. Next, 100 U of T4 DNA Ligase (Fermentas) was added, and the DNA was ligated for 4.5  h at 16°C and then for 30 min at room temperature with slow agitation. Cross-links were reversed by incubation at 65°C for 16 h in the presence of Proteinase K (40 μg/ml). After cross-link reversion, RNase A was added to a final concentration of 40 μg/ml, and RNA was digested for 45 min at 37°C. The DNA was purified by extraction with phenol followed by precipitation with ethanol. To further purify DNA, the samples were processed with QIAEX II Gel Extraction Kit (Qiagen). The concentration of DNA was determined using a fluorometric assay (Qubit, Invitrogen). 50 µg of a ligated 3C DNA template was digested overnight at a concentration of 100 ng/µl with 200 units of DpnII (New England Biolabs). The restriction endonuclease was inactivated by incubation for 20 min at 65°C, and the DNA was purified by phenol-chloroform extraction and ethanol precipitation. Next, the DNA was ligated with 100 units of T4 DNA Ligase (Fermentas) in 14 ml of 1× ligation buffer (Fermentas) for 4 h at 16°C. Ligation products were precipitated with ethanol and further purified using QIAquick PCR purification kit (Qiagen). To linearize the circles of interest, the DNA was treated with a tertiary restriction enzyme of choice: EcoNI for TSR3 bait; NcoI for GENE-Des desert bait, TRAP1 bait and PPL bait; and EcoRV for NPRL3 bait. Digested products were purified using QIAquick PCR purification kit (Qiagen).

value: DT40 lymphoid cell line

value: CRL-2111, ATCC

tag: anchor(s)


title: CTCF HD3 non-induced [ChIP-Seq]

description: HD3 CTCF

tag: erythroid differentiation

value: not induced

value: anti-CTCF (Rabbit custom anti-CTCF antibodies against the part of the chicken CTCF gene encoding the N-terminal region of the protein comprising amino acids 86-233)


title: NPRL3 HD3 rep 2 [4C-Seq]


title: NPRL3 HD3 rep 1 [4C-Seq]


title: MT1

description: Monosiga YAP with human TEAD

library_source: transcriptomic

data_processing:
Data was processed with CASAVA version 1.7.0
Sequence reads in fastq format were mapped to dm3 using tophat-1.0.14. Flybase transcripts (v5.29) were quantified and compared between control and each overexpression condition using cuffdiff v1.0.2.
Genome_build: dm3
Supplementary_files_format_and_content: The file FPKM.txt includes the FPKM values for each gene from each sample as determined by cufflinks. The file cuffdiff_ratio_data.txt includes the log2 ratios, and p-values for each gene for each replicate set of overexpressed variants relative to control as determined by cuffdiff. Also listed are the BH adjusted p-values, gene annotations. There is also a value called Amean, a very general abundance ranking factor, which is the average of the “A” values from each comparison where A is equal to the log2 of the geometric mean of the averaged FPKM values for both treatment and control.

molecule: polyA RNA

source_name: eye imaginal disk

extract_protocol:
Total RNA was recovered from surgically-isolated GMR expression domains (GFP+) from 25 to 30 third instar eye discs using a RNeasy kit (Qiagen). For each genotype, total RNA extraction was conducted in triplicate.
TruSeq RNA Sample Preparation (Low Throughput (LT) Protocol) Part# 15015050, Rev A, November 2010

tag: treatment

value: Overexpression

value: Differentiation defects + Strong ectopic proliferation


title: input DNA

value: GMR-Gal4; UAS-Drosophila Yki S168A


title: sd_ip

value: Anti-Sd


title: M1

description: Monosiga YAP

value: Differentiation defects

value: GMR-Gal4; UAS-eGFP; UAS-MonosigaYAP


title: yki_ip

value: Anti-Yki


title: H3K27me3_ChIPSeq in brm rep1

city: shenzhen

first: Lei

last: Gao

state: guangdong

address: Nanhai Ave 3688

department: College of Life Sciences

country: China

institute: shenzhen university

data_processing:
Basecalls and quality control and pre-aligned (eland) were performed using Illumina Genome analyzer pipeline.
ChIP-seq reads were aligned to the Arabidopsis genome (TAIR10) using BOWTIE version 0.12.7. Only perfect match was allowed. The reads mapped to more than one positions were removed.
The alignments were converted to WIG files using MACS.
The program SICER v1.1 (Zang et al. 2009) was used to identify ChIP-enriched domains (peaks) in histone modification signals.
The quantitative comparisons between WT and brm were performed using the ChIPDiff program.
Genome_build: TAIR10
Supplementary_files_format_and_content: Bed files were generated using SICER.

organism: Arabidopsis thaliana

source_name: Young seedlings, brm mutant, H3K27me3 ChIP

extract_protocol:
Lysates were clarified from sonicated nuclei, and histone-DNA complexes were isolated with H3K27me3 antibody (07-449, Millipore).
Ten ng of ChIP DNA immuoprecipitated by anti-H3K27me3 antibody was used for ChIP-seq library construction. Blunting end, adapter ligation and amplification were carried out using Illumina Genomic DNA Sample Prep Kits according to the manufacturer's protocol. Illumina Genome Analyser IIx was used for high-throughput sequencing of the ChIP-seq library.

value: brm mutant

tag: tissue

value: seedling

tag: age

value: 14 days


title: Rat DRG Control 5

city: Malaga

first: James

middle: R

last: Perkins

address: Pl. Hosp. Civil

department: Hospital Civil

country: Spain

institute: Hospital Carlos Haya

data_processing:
Low quality reads removed (those that did not pass the Illumina quality filter)
Normalisation – the DESeq method was used for library size normalisation (estimateSizeFactors(), estimateDispersions()), as described in pmid:20979621 and the DESeq vignette (http://www.bioconductor.org/packages/devel/bioc/vignettes/DESeq/inst/doc/DESeq.pdf)
Gene Counting – expression was calculated by counting all reads aligning to the known exons of a given gene, according to Ensembl. Exons for a given gene that overlap with exons from another gene (on the same strand or on opposite strand) were removed
Alignment – fastq files (with low quality reads removed) were aligned to the human genome (version Hg19) or the rat genome (version Rn4) using bowtie, with parameters -m 1 -v 1 -p 8 --best, that is, allowing for 1 mismatch and discarding ambiguous reads
Genome_build: Human: Hg19, downloaded from UCSC. Rat: Rn4, downloaded from UCSC.
Supplementary_files_format_and_content: Tables of normalised counts: matrix with genes (Ensemble gene id) as rows and samples as columns. Separate tables are provided for the different sequencing depths

organism: Rattus norvegicus

source_name: Dorsal root ganglia, L5

extract_protocol:
Skin: total RNA was extracted using a combination of the phenol extraction method and column purification. Briefly, samples were homogenised in Trizol (Invitrogen), mixed with chloroform and spun so that the aqueous liquid phase containing the nucleic acids could be removed. This solution was then placed on Qiagen RNeasy columns and subjected to numerous wash steps to purify the RNA sample. DRG: , total RNA from DRG samples was extracted using the miRNeasy kit (Qiagen). All samples were subjected to on-column DNase digestion (Qiagen) to prevent genomic contamination and subsequently analysed using an RNA 6000 nano chip (Agilent Technologies) to confirm sufficient integrity.
cDNA libraries were prepared using the TruSeqTM RNA Sample Preparation Kit (Illumina), low throughput protocol. 200 ng of total RNA were subjected to poly(A) enrichment using poly(T)-attached magnetic beads. Poly(A)-enriched RNA was subsequently used for reverse transcription and library preparation according to Illumina’s instructions

treatment_protocol: Humans: volunteers received 3 MEDs of UVB irradiation to 1cm2 of volar forearm skin using TL01 florescent bulbs. Rats: Anaesthetised rats received 1000mJ/cm2 of UVB to the the plantar surface of the left hind paw

tag: strain

value: Wistar

value: Control

value: Dorsal root ganglia, L5


title: Rat DRG Control 3


title: Rat DRG Control 1


title: Rat DRG UVB 6

value: UVB


title: Rat DRG Control 2


title: Rat DRG UVB 5


title: CTCF HD3 induced [ChIP-Seq]

description: HD3 induced CTCF

value: HD3 erythroblast cell line

value: clone A6 of the line LSCC

value: induced


title: CTCF DT40 [ChIP-Seq]

description: DT40 CTCF

value: DT40 lymphoid cell line

value: CRL-2111, ATCC

value: anti-CTCF (Rabbit custom anti-CTCF antibodies against the part of the chicken CTCF gene encoding the N-terminal region of the protein comprising amino acids 86-233)


title: Rat DRG UVB 3


title: Rat DRG UVB 2


title: Rat Skin Control 6

source_name: Skin

value: skin


title: Rat DRG UVB 4


title: Rat Skin Control 5


title: Rat DRG UVB 1


title: Rat Skin Control 3


title: Rat Skin UVB 6


title: Rat Skin UVB 4


title: Rat Skin UVB 3


title: Rat Skin Control 1


title: Rat Skin Control 4


title: Rat Skin UVB 5


title: Rat Skin UVB 1


title: Rat DRG Control 6


title: lhy21_ChIPSeq

city: Coventry

first: Nigel

middle: Paul

last: Dyer

address: Gibbet Hill Road

country: United Kingdom

institute: University of Warwick

data_processing:
ChIP-seq reads were aligned to the TAIR genome assembly using bowtie, allowing only unique alignments to be retained.
Peaks were called using MACS2, release 2.0.10.09132012. Window size (-bw) was set to 200 and the q value threshold (-q) was set to 0.01
Genome_build: TAIR9
Supplementary_files_format_and_content: Peak file in BED format

source_name: 2 week-old whole seedlings

growth_protocol: For each chromatin preparation, approximately 500µl of seed were sterilised and sown over two 90mm square plates. Seedlings were grown for 2 weeks and samples collected 2h after dawn

extract_protocol:
Samples were cross-linked by vacuum-infiltration in 1% formaldehyde for 10 min. The cross-linking reaction was quenched by adding 1.3 mls of 2 M glycine and infiltrating for a further 5 min. Each sample was homogenised in 30 mls of Extraction Buffer 1 (0.4 M sucrose, 10 mM Tris-HCl pH 8, 5mM beta-mercaptoethanol, 1 mM PMSF and protease inhibitors (Roche complete protease inhibitor cocktail)) using a Polytron Status X120. Debris was removed by filtration through miracloth and the sample was centrifuged at 4°C. The resulting pellet was gently resuspended in 1ml of Extraction Buffer 2 (0.25 M sucrose, 10 mM Tris-HCl pH 8, 10 mM MgCl2, 1% Triton X-100, 5 mM beta-mercaptoethanol, 1 mM PMSF and protease inhibitors) and the nuclei collected by centrifugation. The nuclei were lysed in 500 µl of Nuclear Lysis Buffer (50 mM Tris-Hcl pH 8, 10 mM EDTA, 1% SDS, 1 mM PMSF and protease inhibitors) and sonicated to give chromatin fragment sizes ranging from 100 to 1000bp. For each immunoprecipitation, 250 µl of chromatin was added to 2 mls of ChIP dilution buffer (167 mM NaCl, 16.7 mM Tris-HCl pH8, 1.2 mM EDTA, Triton X-100, 1 mM PMSF and protease inhibitors) and pre-cleared with protein A Dynabeads (Invitrogen). Samples were incubated overnight at 4°C with either anti-LHY (1:200)  antibody. The immuno-complexes were isolated by incubation with protein A Dynabeads for 2 h at 4°C. The beads were washed as described by Haring et al.,  2007 with the addition of three extra high salt buffer washes. Protein A beads were then resuspended in 100 μl of TE and treated with RNase at 37°C for 20 minutes. SDS was added to a final concentration of 0.5% and the samples digested with proteinase K for 2 h at 50°C. To reverse cross-linking, 8 μl of 5 M NaCl was added and the samples were incubated overnight at 65°C. The DNA was then purified using the MinElute PCR purification kit (Qiagen).
9 independent ChIP samples were pooled for the generation of each library.  Wild-type and lhy-21 mutant libraries were sequenced in multiplex. Library preparation and sequencing was conducted at the University of Utah’s Bioinformatic Core facility using an Illumina HiSeq 2000 sequencer. The libraries were prepared using the Illumina Tru Seq DNA sample prep kit according to the instructions of the manufacturer.  Indexes 6 (GCCAAT) and  12 (CTTGTA) were used for wild-type (Ws) and lhy21 samples, respectively.

tag: antibody

value: antiserum against the full-length LHY protein

tag: ecotype

value: WS

value: whole seedlings


title: LHY_ChIPSeq

value: Wild-type


title: Rat DRG Control 4


title: Rat Skin Control 2


title: Footprint_contDevB_N2_36hr

city: Basel

first: Dimos

last: Gaidatzis

address: Maulbeerstrasse 66

country: Switzerland

institute: Friedrich Miescher Institute

data_processing:
The 3' adaptor (TGGAATTCTCGGGTGCCAAGG) was removed from the reads using the function preprocessReads from within the bioconductor package QuasR (default parameters). The short fragments (about 30bp length) were mapped to the c.elegans genome (ce6) with the R package QuasR (www.bioconductor.org/packages/2.12/bioc/html/QuasR.html) using the included aligner bowtie allowing only for uniquely mapping reads. The command used to perform the alignments was "proj <- qAlign("samples.txt","BSgenome.Celegans.UCSC.ce6")". Gene expression was quantified by counting the number of reads that started within any of the exons beloning to a particular gene (WormBase, WS190). The command used to create the count table was qCount(proj,exons,orientation="same"). To compensate for differences in the read depths of the various libraries, we divided each sample by the total number of reads and multiplied by the average library size. Log2 expression levels were calculated after adding a pseudocount of 8 (y=log2(x+8). This was done to minimize the large differences in expression that would otherwise be caused by genes with small number of counts.
Genome_build: ce6
Supplementary_files_format_and_content: ce_geneExpression_Footprint_rawCounts.txt: non normalized raw counts
Supplementary_files_format_and_content: ce_geneExpression_Footprint_normalized.txt: normalized and log2 transformed values

molecule: total RNA

organism: Caenorhabditis elegans

source_name: Whole-worm

growth_protocol: Continuous: Egg-prepped C. elegans eggs were allowed to hatch overnight, hatched L1s were plated on peptone rich 2% NGM plates with NA22 for the desired period of time before harvesting

extract_protocol:
Worms were lysed and monosomes purified (Hendriks & Gaidatzis et.al) and RNA was isolated using Tri-reagent (MRC)
The RNA from the monosomal fraction was separated using a 15% TBE-Urea Gel (Invitrogen) and the region around 28-30 nucleotides excised to isolate Ribosome protected fragments (RPFs). The gel piece was forced through a pierced small tube inside an eppendorf tube by centrifugation and RNA from the gel debris was eluted by overnight incubation in 600 µl cracking buffer (20 mM Tris-HCl (pH 7.9), 1 mM EDTA, 400 mM NH4Acetate, 0.5 % SDS). RNA was precipitated with isopropanol at -80 °C for at least 4 hours (isopropanol precipitation). RPFs were 3’ dephosphorylated with 10 Units of T4 polynucleotide kinase (NEB) in T4 PNK buffer with 40 Units of RNasin for 1 hour at 37 °C. Following isopropanol precipitation, the RNA samples were ligated to 3’ adapters according to the Illumina® TruSeq™ Small RNA Sample Preparation protocol and using the reagents of the kit, then again precipitated with isopropanol. Ligation products were 5’ phosphorylated for 30 minutes at 37 °C with 15 Units of T4 polynucleotide kinase (NEB) in T4 PNK buffer, 1 mM ATP and 40 Units of RNasin. Following heat-inactivation of the enzyme for 10 minutes at 70 °C, the RNA was precipitated by isopropanol. Ligation to 5’ adapters, reverse transcription, PCR amplification with barcoded primers and gel-purification of the PCR products were performed using the Illumina® TruSeq™ Small RNA Sample Prep kit. Four different barcodes (RPIX 2, 4, 5, 6) were used.

treatment_protocol: Worms were washed of the plates and subsequently washed three additional times with M9 before extraction of RNA

tag: time in development

value: 36 hours


title: Footprint_contDevB_N2_28hr

value: 28 hours


title: Footprint_contDevB_N2_24hr

value: 24 hours


title: Footprint_contDevB_N2_30hr

value: 30 hours


title: Footprint_contDevB_N2_26hr

value: 26 hours


title: Footprint_contDevB_N2_22hr

value: 22 hours


title: Footprint_contDevB_N2_32hr

value: 32 hours


title: Footprint_contDevB_N2_18hr

value: 18 hours


title: Footprint_contDevB_N2_20hr

value: 20 hours


title: RNASeq_riboM_contDevB_N2_32hr

data_processing:
The RNA-seq data (50bp read length) were mapped to the c.elegans genome (ce6) with the R package QuasR (www.bioconductor.org/packages/2.12/bioc/html/QuasR.html) using the included spliced alignment algorithm SpliceMap. The command used to perform the alignments was "proj <- qAlign("samples.txt","BSgenome.Celegans.UCSC.ce6",splicedAlignment=TRUE)". Gene expression was quantified by counting the number of reads that started within any of the exons beloning to a particular gene (WormBase, WS190). The command used to create the count table was qCount(proj,exons,orientation="same"). To compensate for differences in the read depths of the various libraries, we divided each sample by the total number of reads and multiplied by the average library size. Log2 expression levels were calculated after adding a pseudocount of 8 (y=log2(x+8). This was done to minimize the large differences in expression that would otherwise be caused by genes with small number of counts.
Genome_build: ce6
Supplementary_files_format_and_content: ce_geneExpression_riboM_rawCounts.txt: non normalized raw counts
Supplementary_files_format_and_content: ce_geneExpression_riboM_normalized.txt: normalized and log2 transformed values

growth_protocol: Continuous 1: Egg-prepped C. elegans eggs were allowed to hatch overnight, hatched L1s were plated on 2% NGM plates with OP50 for the desired period of time before harvesting ; Continuous 2: Egg-prepped C. elegans eggs were allowed to hatch overnight, hatched L1s were plated on peptone rich 2% NGM plates with NA22 for the desired period of time before harvesting

extract_protocol:
total RNA isolation was performed using Tri-Reagent (MRC)
Library preparation was performed using the ScriptSeq v2 RNA-Seq library preparation kit (Epicentre) according to the manufacturer's protocol

tag: growth protocol

value: Continuous 2


title: RNASeq_riboM_contDevB_N2_36hr


title: RNASeq_riboM_contDevB_N2_30hr


title: RNASeq_riboM_contDevB_N2_26hr


title: RNASeq_riboM_contDevB_N2_28hr


title: RNASeq_riboM_contDevB_N2_24hr


title: Rat Skin UVB 2

value: Wistar

value: UVB


title: RNASeq_riboM_contDevB_N2_34hr

value: 34 hours


title: RNASeq_riboM_contDevA_N2_36hr

description: Please note that the discrepancy between the "time in development" and the name of the sample is caused by the relatively slow growth during this particular timecourse when compared to other timecourses that were previously collected (data not shown). The time that is mentioned in the sample name more closely resembles the actual developmental time it takes to reach this point in development.

value: 37 hours

value: Continuous 1


title: RNASeq_riboM_contDevA_N2_35hr


title: RNASeq_riboM_contDevA_N2_33hr


title: RNASeq_riboM_contDevA_N2_34hr

value: 35 hours


title: RNASeq_riboM_contDevB_N2_18hr


title: RNASeq_riboM_contDevA_N2_32hr

value: 33 hours


title: RNASeq_riboM_contDevB_N2_22hr


title: RNASeq_riboM_contDevA_N2_30hr

value: 31 hours


title: Footprint_contDevB_N2_34hr


title: RNASeq_riboM_contDevA_N2_29hr


title: RNASeq_riboM_contDevA_N2_31hr


title: RNASeq_riboM_contDevA_N2_28hr

value: 29 hours


title: RNASeq_riboM_contDevA_N2_26hr

value: 27 hours


title: RNASeq_riboM_contDevA_N2_27hr

value: 28 hours


title: RNASeq_riboM_contDevA_N2_22hr

value: 23 hours


title: RNASeq_riboM_contDevA_N2_21hr

value: 22 hours


title: RNASeq_polyA_contDev_glp-4_52hr

data_processing:
The RNA-seq data (50bp read length) were mapped to the c.elegans genome (ce6) with the R package QuasR (www.bioconductor.org/packages/2.12/bioc/html/QuasR.html) using the included spliced alignment algorithm SpliceMap. The command used to perform the alignments was "proj <- qAlign("samples.txt","BSgenome.Celegans.UCSC.ce6",splicedAlignment=TRUE)". Gene expression was quantified by counting the number of reads that started within any of the exons beloning to a particular gene (WormBase, WS190). The command used to create the count table was qCount(proj,exons,orientation="opposite",selectReadPosition="end"). The library preparation protocol for polyA selected RNAs creates reads that correspond to the reverse complement of the original RNA and therefore we had to count the reads on the opposite strand of the actual gene. To compensate for differences in the read depths of the various libraries, we divided each sample by the total number of reads and multiplied by the average library size. Log2 expression levels were calculated after adding a pseudocount of 8 (y=log2(x+8). This was done to minimize the large differences in expression that would otherwise be caused by genes with small number of counts.
Genome_build: ce6
Supplementary_files_format_and_content: ce_geneExpression_polyA_rawCounts.txt: raw counts and ce_geneExpression_polyA_normalized.txt which contains normalized and log2 transformed values

growth_protocol: Continuous: Egg-prepped C. elegans eggs were allowed to hatch overnight, hatched L1s were plated on 2% NGM plates with OP50 for the desired period of time before harvesting ; Dauer-exit: Dauer larvae were grown as described in Hendriks & Gaidatzis et.al, introduction of food to the larvae, by plating them on 2% NGM plates with OP50, corresponds to the 0-hour timepoint.

extract_protocol:
total RNA isolation was performed using Tri-reagent (MRC)
Library preparation was performed using the Truseq stranded mRNA sample preparation kit (Illumina) according to the manufacturer's protocol

value: 52 hours

value: Continuous


title: RNASeq_riboM_contDevA_N2_23hr

value: 24 hours


title: RNASeq_riboM_contDevA_N2_24hr

value: 25 hours


title: RNASeq_riboM_contDevB_N2_20hr

value: 20 hours

value: Continuous 2


title: RNASeq_polyA_contDev_glp-4_46hr

value: 46 hours


title: RNASeq_polyA_contDev_glp-4_48hr

value: 48 hours


title: RNASeq_polyA_contDev_glp-4_44hr

value: 44 hours


title: RNASeq_polyA_contDev_glp-4_42hr

value: 42 hours


title: RNASeq_polyA_contDevA_N2_34hr

value: 35 hours


title: RNASeq_polyA_contDevA_N2_33hr

value: 34 hours


title: RNASeq_polyA_contDevA_N2_36hr

value: 37 hours


title: RNASeq_polyA_contDevA_N2_35hr

value: 36 hours


title: RNASeq_polyA_contDevA_N2_32hr

value: 33 hours


title: RNASeq_polyA_contDevA_N2_28hr

value: 29 hours


title: RNASeq_polyA_contDevA_N2_29hr

value: 30 hours


title: RNASeq_polyA_contDevA_N2_31hr

value: 32 hours


title: RNASeq_polyA_contDev_glp-4_40hr

value: 40 hours

value: Continuous


title: RNASeq_polyA_contDevA_N2_27hr

value: 28 hours


title: RNASeq_polyA_contDevA_N2_26hr

value: 27 hours


title: RNASeq_polyA_contDevA_N2_25hr

value: 26 hours


title: RNASeq_riboM_contDevA_N2_25hr

value: Continuous 1


title: Human Skin Control 3

organism: Homo sapiens

value: Control

value: skin


title: Human Skin Control 4


title: Human Skin Control 2


title: RNASeq_polyA_contDevA_N2_23hr

value: 24 hours


title: RNASeq_polyA_contDev_glp-4_50hr

value: 50 hours


title: RNASeq_polyA_contDevA_N2_22hr

value: 23 hours


title: Human Skin UVB 4

value: UVB


title: Human Skin Control 1


title: RNASeq_polyA_contDevA_N2_21hr

value: 22 hours


title: Human Skin UVB 2


title: Human Skin UVB 1


title: Human Skin UVB 5


title: HCT116-DNaseI_2

status: Public on Feb 28 2014

city: New York

first: Matthew

middle: T

last: Maurano

state: NY

address: 435 East 30th Street

department: Institute for Systems Genetics

institute: NYU School of Medicine

description: This biological replicate have been previously released in GSE30263, but the data has been reprocessed to be consistent wit the new data in this study.

data_processing:
Methylation values: Capture bisulfite reads were mapped to the human genome (hg19) using Bismark (Krueger and Andrews 2011) and bowtie2 beta 6 (Langmead et al. 2009) with the options “-n 1” and excluding duplicate reads and combining reads from the top and bottom strands. Methylation was monitored at CpGs with ≥8 read coverage, and averaged for all CpGs within the 150 bp window at each CTCF site.
Peak: ChIP-seq affinity zones (HotSpots) were identified using the HotSpot algorithm described in Sabo et al. (2004). 1.0% false discovery rate thresholds (FDR 0.01) were computed for each cell type by applying the HotSpot algorithm to an equivalent number of random uniquely mapping 36mers. DNase data used unthresholded hotspots.
Density: Smoothed density tracks were generated using bedmap (http://code.google.com/p/bedops/) to count the number of tags overlapping a sliding 150-bp window, with a resolution of 20 bp (Neph et al. 2012a). Density tracks for display were normalized for sequencing depth by a global linear scaling to 10 million tags.
Alignment: ChIP-seq and DNase data were mapped to the human genome (GRCh37/ hg19) using bowtie (Langmead et al. 2009) with the options ‘‘bowtie -n 2 -m 1 -e 70 --best’’ allowing up to three mismatches. Reads mapping to multiple locations were then excluded, and reads with identical 5’ ends and strand were presumed to be PCR duplicates and were excluded using Picard MarkDuplicates.
Genome_build: hg19
Supplementary_files_format_and_content: .bed: peak locations generated using the hotspot algorithm; .bed proportion of bisulfite tags indicating methylation per CpG (proportion methylation in bed score column, number of total reads in 7th column) .bw: the number of tags overlapping a sliding 150-bp window, with a resolution of 20 bp; .bam: tags mapped to the human genome

source_name: HCT116 colorectal cancer cells

growth_protocol: Cells were grown according to the approved ENCODE cell culture protocols.

extract_protocol:
Digital DNaseI was performed by DNaseI digestion of intact nuclei, isolating DNaseI 'double-hit' fragments as described in Sabo et al. (2006), 20-50 ng of DNA was end-repaired, adenine ligated to Illumina adapters was added, and then a Solexa library was made for sequencing.
20-50 ng of ChIP DNA was end-repaired, adenine ligated to Illumina adapters was added, and then a Solexa library was made for sequencing.

tag: experiment

value: Dnase-Hypersensitivity

value: No special treatment

tag: replicate


title: RNASeq_polyA_contDevA_N2_24hr

value: 25 hours


title: HCT116-DNaseI_1


title: HCT116-H3K4me3_2

extract_protocol:
Cells were crosslinked with 1% formaldehyde, and the reaction was quenched by the addition of glycine. Fixed cells were rinsed with PBS, lysed in nuclei lysis buffer, and the chromatin was sheared to 200-500 bp fragments using Fisher Dismembrator (model 500). Sheared chromatin fragments were immunoprecipitated with specific polyclonal antibodies at 4 degrees C with gentle rotation. Antibody-chromatin complexes were washed and eluted. The cross linking in immunoprecipitated DNA was reversed and treated with RNase-A. Following proteinase K treatment, the DNA fragments were purified by phenol-chloroform-isoamyl alcohol extraction and ethanol precipitation. 20-50 ng of ChIP DNA was end-repaired, adenine ligated to Illumina adapters was added, and then a Solexa library was made for sequencing.
20-50 ng of ChIP DNA was end-repaired, adenine ligated to Illumina adapters was added, and then a Solexa library was made for sequencing.

value: H3K4me3 ChIP-seq


title: HCT116-H3K4me3_1


title: ChIP3_IGG

city: Berlin

first: Michal

middle: Ruth

last: Schweiger

address: Ihnestrasse 63-73

department: Lehrach

country: Germany

institute: Max Planck Institute For Molecular Genetics

description: Sample 7

data_processing:
Illumina Casava1.3 software used for basecalling for the samples 1,2,5,6,7,8. Illumina Casava 1.8.2 was used for the samples 3 & 4
Sequenced reads were mapped to hg19/GRCh37 whole genome using bwa 0.5.9-r16 with default parameters
Exon read coverages were obtained with coverageBed v2.17.0 using exon coordinates of the Ensembl database v63. Transcript counts were generated by summing read counts across all exons of a given transcript and transcript counts were normalized by the number of sequenced reads (sequencing depth) to ensure comparable counts among samples.
Using the bam files produced by bwa as input peaks were called with MACS v1.4 (PMID: 18798982) using an enrichment ratio against the background of 5 to 15 to build the model (m=5,15).
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include read count values for each transcript model of the Ensembl database v63 and peaks called by MACS.

growth_protocol: Hek293T cells were cultivated at 37°C with 5% C0 2 in Dulbecco’s Modiﬁed Eagle’s Medium (DMEM) containing 10% fetal calf serum (FCS), 2 mM L-glutamine and 100 U penicillin/streptomycin.

extract_protocol:
RNA was harvested using Trizol reagent including a DNase digestion step with the RNase-free DNase set from Qiagen.
The Chromatin Immunoprecipitation was done according the protocol Dahl et al PMID: 17569620.
cDNA library preparation, and sequencing was performed by Illumina’s TrueSeq prep kit following the manufacturer’s instructions with 1 ug of total RNA for the construction of mRNA sequencing libraries.
ChIP Seq libraries were prepared according to Illumina's ChIP-Seq Sample Prep Kit instructions.

treatment_protocol: Plasmids (shRNA against BRD4 as described in Schweiger et al PMID: 16611886) were introduced into cells by X-treme gene 9 transfection reagent (Roche). The knockdown efficiency was checked using qPCR and showed a knockdown of more than 75%

value: IgG antibody

tag: chip antibody vendor

value: Jackson ImmunoResearch

tag: chip antibody cat. #


title: ChIP2_BRD4

description: Sample 6

tag: cell line

value: antibody against BRD4 long form

value: Abcam


title: RNASeq_polyA_contDevA_N2_30hr

value: 31 hours

value: Continuous


title: RNA2_KO_BRD4

description: Sample 2

source_name: HEK_BRD4 KO_RNA-seq

tag: transfected with

value: shRNA against BRD4


title: ChIP1_IGG

description: Sample 5

tag: chip antibody


title: Mut_DRH-3_Rep2

first: Germano

last: Cecere

address: 701 West 168th

department: Biochemistry and Molecular Biophysics

institute: Columbia University

description:
GRO-Seq
Replicate 2
processed data file: Mut_DRH-3_rep1+2.neg_strand.bw
processed data file: Mut_DRH-3_rep1+2.pos_strand.bw

data_processing:
Two independent biological replicates were generated for csr-1 hypomorphic strain and the  corresponding N2 wild type (WT) using ~300,000 worms (called Mut_CSR-1 and WT_CSR-1, respectively). Since drh-3 (ne4253) mutant strain has a reduced number of progeny we performed two biological replicates of drh-3 (ne4253) mutant using ~100,000 worms and we compared them with two biological replicates of N2 (WT) generated using an equal number of worms (Mut_DRH-3 and WT_DRH-3, respectively). Sequencing was performed to a length of 50 nucleotides. Since cloned RNA ranged from 16 to 180 nucleotides in length, the sequencing reads were post-processed to eliminate the 3' linker sequence, if present, so that resulting reads ranged from 16 to 50 nucleotides in length.
The reads were then aligned using “bwa 0.6.1” to the C. elegans genome (ce6 assembly), and rRNA reads were removed based on genomic locations: ChrI:15060000-15072421 for Ce_18S_rDNA; ChrV:17115500-17132700 for Ce_5S_rDNA; chrV:17428800-17429040 for Ce_5S_rDNA-singleton; chrX:16929621-16940441 for Ce_repetitiveLocus. After filtering, we obtained 44,155,515 reads from csr-1 hypomorphic mutant [26,584,972 reads from the first replicate (called Mut_CSR-1_Rep1) and 17,570,543 reads from the second replicate (called Mut_CSR-1_Rep2)] and 36,133,711 reads from the corresponding WT [16,052,870 reads from the first replicate (called WT_CSR-1_Rep1) and 20,080,841 reads from the second replicate (called WT_CSR-1_Rep2)]. From drh-3 (ne4253) mutant, we obtained after filtering 34,588,133 reads [17,492,181 reads from the first replicate (called Mut_DRH-3_Rep1) and 17,095,952 reads from the second replicate (called Mut_DRH-3_Rep2)], and 63,000,071 reads from the corresponding WT [21,660,645 reads from the first replicate (called WT_DRH-3_Rep1) and 41,339,426 reads from the second replicate (called WT_DRH-3_Rep2)].
We further filtered the data by strand (+/-) to analyze sense and antisense Pol II transcription. After ascertaining concordant results with the individual replicate pairs (WT vs. mutant) (data not shown), replicate reads for each WT and mutant were pooled, using downsampling on the replicate with higher read count to ensure equal representation of either replicate, and the pooled reads were processed together to increase statistical power.
For each annotated C. elegans transcript, the aligned reads were binned based on the annotated transcriptional start site (TSS) and transcriptional termination site (TTS) into 3 regions as follows: 'promoter' (from 600bp upstream of the annotated TSS to 200bp downstream), 'gene body' (from 200bp downstream of the annotated TSS to 200bp upstream of the annotated TTS), and 'gene end' (from 200bp upstream of the annotated TTS to 400bp downstream).
We then calculated region-specific RPKM values (#total reads / (region length (KB) X # mapped reads (M)) for these regions. We also calculated statistical significant change in RPKM values using using DESeq package  (Anders and Huber, 2010) considering a p value <0.05.
For the enrichment or depletion of gene sets, we first calculated the number of gene expected by chance according to r = (n1xn2)/N, with n1 =  number genes in set 1, n2 = number of genes in set 2, N = total number of C. elegans genes considered, and then the enrichment was calculated by dividing the number of genes in common to the two datasets by r.   The average data profile in Figure 4B and Figures S4A and S4B were calculated using Cistrome/Galaxy (Liu et al., 2011) with the CEAS tool (SitePro: Aggregation plot tool for signal profiling (version 1.0.0)), and the heatmap showing the genome-wide correlation coefficient values in Figure 4C was calculated using Multiple wiggle files correlation tool (version 1.0.0).  The overlaps between gene lists were calculated using the online tool at http://www.nemates.org.  Representation factors “r” to quantify the enrichment or depletion of gene sets were calculated according to r = (n1,2)/[(n1xn2)/N], with n1,2 = number of genes common to set 1 and 2, n1 =  number genes in set 1, n2 = number of genes in set 2, N = total number of genes considered.
Genome_build: WS190
Supplementary_files_format_and_content: bigWig files, strand-specific pooled replicates, rescaled to sample population size. The files are combined replicates, normalized mapped reads separated by strands.

growth_protocol: Synchronous populations of worms were grown for approximately 40 hr posthatching at 20°C on OP-50 E. coli at a density of ~100,000 animals per 15 cm Petri dish.

extract_protocol:
Worms were washed off the plate with cold M9 and washed 3-4 times. The worm pellet was resuspended in 2ml of cold Nuclear Run-On (NRO) lysis buffer (10mM Tris pH 7.5, 2mM MgCl2, 3mM CaCl2, 0.5% IGEPAL and 10% glycerol), transferred to a steel dounce (on ice) and stroked 20 times. The worm lysate was centrifuged for 2min at 40g and the supernatant was centrifuged again for 5min at 1000g to pellet the nuclei. Next, the supernatant was discarded and the nuclei pellet was washed two times with 1ml of cold NRO lysis buffer and one time with 1ml of cold Freezing Buffer (50mM Tris-CL pH 8.3, 40% glycerol, 5mM MgCl2, 0.1 mM EDTA). Nuclei were then resuspended in 100µl of Freezing Buffer and used for the NRO reaction. // NRO reaction and DNase digestion: 100µl of nuclei were mixed with an equal volume of reaction buffer (10mM Tris-Cl pH 8.0, 5mM MgCl2, 1mM DTT, 300mM KCL, 20 units of SUPERase In, 1% sarkosyl, 500µM ATP, GTP, and Br-UTP, 2μM CTP) and incubated for 5 minutes at 32°C. The reaction was stopped with 750μl of TRIzol reagent (Invitrogen) and the RNA was extracted following the manufacturer's instructions. The RNA pellet was resuspended in 85µl water and treated with 5µl of turbo DNase (Ambion) in a total volume of 100µl and incubated at 37°C for 30 minutes. The RNA was then extracted by phenol:chloroform and precipitated with one volume of isopropanol with 1/10 volume of Sodium Acetate 3M and 10µg of Glycoblue (Ambion). // Base Hydrolysis of RNA: The hydrolysis of RNA was performed using fragmentation reagent (Ambion) for 15min at 70°C and the product was purified by p-30 RNase free spin column (BioRad), following the manufacturer’s instruction. // Immunopurification of Br-U RNA and RNA-end repair: The purification of the Br-U RNA and the end repair of the purified Br-U RNA was performed as described in (Core et al., 2008).
Adapter ligation and library preparation: For the ligation of 5' and 3' adapters to Br-U RNA we used reagents contained in the ScriptMiner™ Small RNA-sequencing kit (Epicentre) and we immunopurified the ligated Br-U RNA product after each ligation with anti-deoxy-BrU beads (Santa Cruz Biotech (sc-32323-AC). The ligated Br-U RNA was then reverse-transcribed and PCR-amplified with properly-indexed primers for 18 cycles, following the ScriptMiner™ Small RNA-sequencing kit (Epicentre) manufacturer's instructions. The NRO-cDNA libraries were purified by phenol:chloroform, the DNA was precipitated as above and then ran on a non-denaturing 1XTBE, 8% acrylamide gel and stained with SYBR gold (Invitrogen). DNA fragments ranging in size from 120bp to 300bp was excised from the gel and eluted by incubating in TE + 300mM NaCl overnight with rotation. The DNA libraries were then extracted, precipitated, and sent for sequencing.

tag: genotype/variation

value: drh-3 (ne4253) mutant

tag: Stage

value: late L3/early L4

value: whole animal


title: Mut_DRH-3_Rep1

description:
GRO-Seq
Replicate 1
processed data file: Mut_DRH-3_rep1+2.neg_strand.bw
processed data file: Mut_DRH-3_rep1+2.pos_strand.bw


title: WT_DRH-3_Rep2

description:
GRO-Seq
Replicate 2
processed data file: WT_DRH-3_rep1+2.neg_strand.bw
processed data file: WT_DRH-3_rep1+2.pos_strand.bw

value: wild type


title: WT_DRH-3_Rep1

description:
GRO-Seq
Replicate 1
processed data file: WT_DRH-3_rep1+2.neg_strand.bw
processed data file: WT_DRH-3_rep1+2.pos_strand.bw


title: Mut_CSR-1_Rep2

description:
GRO-Seq
Replicate 2
processed data file: Mut_CSR-1_rep1+2.neg_strand.bw
processed data file: Mut_CSR-1_rep1+2.pos_strand.bw

value: csr-1 hypomorphic mutant


title: Human Skin UVB 3

tag: treatment

value: UVB

value: skin


title: Mut_CSR-1_Rep1

description:
GRO-Seq
Replicate 1
processed data file: Mut_CSR-1_rep1+2.neg_strand.bw
processed data file: Mut_CSR-1_rep1+2.pos_strand.bw

tag: tissue


title: WT_CSR-1_Rep1

description:
GRO-Seq
Replicate 1
processed data file: WT_CSR-1_rep1+2.neg_strand.bw
processed data file: WT_CSR-1_rep1+2.pos_strand.bw


title: Decidua at E11.5

city: Stanford

first: Elizabeth

middle: Havener

last: Finn

state: California

address: 300 Pasteur Drive, Alway M311

institute: Stanford University

data_processing:
Reads were mapped using BWA to a custom artificial B6/Cast F1 transcriptome which was created in silico based on high-quality SNPs defined by Sanger
High-Quality SNPs were assigned maternal/paternal read counts and votes (normalized so each read contributes only once to sums); SNPs were filtered such that only SNPs with a coverage of at least 10 reads across all samples were considered.
Percent Maternal was calculated for each SNP(counts) and transcript(votes)
Genome_build: mm9
Supplementary_files_format_and_content: For placental and yolk sac samples, Bed files include p(mat) at each SNP.
Supplementary_files_format_and_content: For decidual sample, Bed file includes read count at each SNP

organism: Mus musculus

source_name: Midgestation Maternal Decidua

extract_protocol:
Total RNA was isolated from placenta using a phenol-chloroform purification with Trizol (Invitrogen).  mRNA was subsequently purified using Dynabeads (Invitrogen).
RNA was analyzed using 3SEQ; a modificaiton of standard Illumina protocols. Briefly, mRNA was heat sheared before reverse transcription with the P7_oligodT primer.  The P5 linker was then ligated to the free end, sequences are amplified using primers to P5 and P7, and the resulting library was sequenced from P5 using the Illumina Genome Analyzer II.

tag: maternal strain

value: Decidua


title: WT_CSR-1_Rep2

description:
GRO-Seq
Replicate 2
processed data file: WT_CSR-1_rep1+2.neg_strand.bw
processed data file: WT_CSR-1_rep1+2.pos_strand.bw


title: Yolk Sac at E11.5 Sample 1

source_name: Midgestation Yolk Sac

value: Cast

value: Yolk Sac


title: Yolk Sac at E11.5 Sample 2


title: Fetal Placenta at E17.5, Sample 6

source_name: Late Stage Fetal Placenta

value: Placenta


title: Fetal Placenta at E17.5, Sample 2


title: Fetal Placenta at E17.5, Sample 3


title: Fetal Placenta at E17.5, Sample 4


title: Fetal Placenta at E17.5, Sample 1


title: ChIP4_BRD4

description: Sample 8


title: Fetal Placenta at E11.5, Litter C2, Sample 6

source_name: Midgestation Fetal Placenta


title: Fetal Placenta at E11.5, Litter C2, Sample 5


title: Fetal Placenta at E11.5, Litter C2, Sample 2


title: Fetal Placenta at E11.5, Litter C2, Sample 3


title: Fetal Placenta at E11.5, Litter C2, Sample 1


title: RNA1_ctrl

description: Sample 1

value: none (control)


title: Fetal Placenta at E11.5, Litter C1, Sample 4


title: Fetal Placenta at E11.5, Litter C1, Sample 1


title: Fetal Placenta at E11.5, Litter C1, Sample 3


title: Fetal Placenta at E11.5, Litter B4, Sample 7


title: Fetal Placenta at E11.5, Litter B4, Sample 4


title: Fetal Placenta at E11.5, Litter C1, Sample 2


title: Fetal Placenta at E11.5, Litter B3, Sample 4


title: Fetal Placenta at E11.5, Litter B3, Sample 3


title: Fetal Placenta at E11.5, Litter B1, Sample 4


title: Fetal Placenta at E11.5, Litter B1, Sample 3


title: Fetal Placenta at E11.5, Litter B1, Sample 6


title: Fetal Placenta at E11.5, Litter B3, Sample 1


title: Fetal Placenta at E11.5, Litter B1, Sample 5


title: 7104_LIV_post-puberty

city: Guelph

first: Angela

last: Cánovas

state: ON

address: 50 Stone Road East

department: Animal Biosciences

country: Canada

institute: University of Guelph

data_processing:
Filtering for quality by NGS Quality Control tool using CLC Genomics Workbench (CLC Bio, Aarhus, Denmark).
RNA-Seq analysis using CLC Genomics Workbench Ver. 6.0
Genome_build: Annotated UMD3.1 bovine reference genome (http://www.ncbi.nlm.nih.gov/genome/?term=bos+taurus)
Supplementary_files_format_and_content: .csv containing Ensembl ID, Gene name, chromosome, gene start (bp), gene end (bp), average unique reads, RPKM means.

organism: Bos taurus

source_name: Liver

growth_protocol: All tissues were collected within 15 min postmortem, snap-frozen in liquid nitrogen, and then stored at -80° C until processing.

extract_protocol:
Total RNA was purified using a Trizol protocol (Invitrogen, Carlsbad, CA).
mRNA was purified, fragmented, and converted to cDNA. Adapters were ligated to the ends of double-stranded cDNA and PCR amplified to create libraries. These procedures were executed with the TruSeq RNA Sample Preparation kit of Illumina (San Diego, CA).

treatment_protocol: Duplicate 10 g samples of each tissue were ground in liquid nitrogen with mortar and pestle.

tag: developmental stage

value: POST-puberty

value: Liver

tag: breed

value: Brangus (3/8 Brahman x 5/8 Angus)


title: 40_LIV_post-puberty


title: 36_LIV_post-puberty


title: 105_LIV_post-puberty


title: Fetal Placenta at E11.5, Litter C2, Sample 4


title: Fetal Placenta at E11.5, Litter B1, Sample 2


title: Fetal Placenta at E11.5, Litter C1, Sample 5


title: NIH3T3

city: Seoul

first: Hyeshik

last: Chang

state: South Korea

address: Building 203 Room 525, School of Biological Sciences, Seoul National University, 1 Gwanak-ro, Gwanak-gu

department: School of Biological Sciences

country: South Korea

institute: Seoul National University

description: A TAIL-seq sample from NIH3T3

data_processing:
Library strategy: TAIL-seq
The base calls and signal intensities were acquired from HiSeq 2500 after processing by Illumina RTA 1.17.21.3. The read 1 sequences were aligned to the common contaminants set, which is composed of rDNA repeat units (GenBank accession BK000964.1 for NIH3T3 and U13369.1 for HeLa), PhiX genome (GenBank accession J02482.1), Illumina TruSeq primer sequences, and all sequences for 5S and 5.8S rRNAs of respective species (retrieved from Rfam 11.0 of the Wellcome Trust Sanger Institute) using GSNAP 2013-03-31 with maximum 5% mismatches allowed. Clusters with any match to the contaminants were removed from the subsequent analyses. The sequences having completely identical nucleotides in the 21st to 35th cycle in read 1 (representative region of the insert) and the 1st to 15th cycle in read 2 (degenerate bases in 3′ adapter) are deduplicated by leaving only a cluster with the maximum PHRED quality sum of read 1. The degenerate and fixed delimiter sequence in 3′ adapter was clipped out from read 2 by searching perfect match of delimiter sequence (‘GTCAG’ as in the direction of read 2) between the 14th and 16th cycles in read 2. The clusters missing a delimiter sequence or having low diversity in degenerate region (at least two occurrences for all of A, C, G and T) were removed from further analyses.
The fluorescence signal intensities were processed into “Normalized T signal” as described in fig. S1A. The signals from a spike-in sample were purified with an outlier filter based on robust Mahalanobis distance (mvoutlier package 1.9.9; quan=0.5, alpha=0.025). Random 500 clusters per each spike-in were chosen for parameter calculation of a Gaussian mixture hidden Markov model (GMHMM). We trained the model using Baum-Welch algorithm implemented in the GHMM library (http://ghmm.org) with topology and initial parameters shown in fig. S1A and table S7 and S8 (1,000 iterations). The procedure was iterated to maximize likelihood, not using any property (eg. designed length of poly(A) tail) of spike-ins. Normalized T signals outside the range of [-5, 5] were clipped into the range for both training and later calculations.
The length of poly(A) tails were first measured with base call-based “Strategy II” described in fig. S1A. For clusters with the measured length is shorter than 8 nt, the length is called as the final poly(A) tail length. For the others, normalized T signals starting from the first position in T-stretch detected by Strategy II were analyzed with the GMHMM. The hidden states were decoded with the standard Viterbi algorithm implemented in the GHMM library. The number of cycles with state 1 and 2 was called as the length of poly(A) tail. For the estimation of performance, we applied the process to all spike-in samples except the clusters used for the parameter fitting of the model.
The remaining reads after contaminant filter and the first duplication filters were then aligned to the genome sequences (UCSC mm10 for NIH3T3 and UCSC hg19 for HeLa, positions of splicing junctions were processed from the UCSC Genome Browser database for version of Jan 24, 2013) using GSNAP 2013-03-31. Three different versions of alignments to genome were used in this study. (1) R1 alignment: using only the full read 1 sequences which are 51 nt long. This was used for identification of a cluster. (2) R2 short alignment: using only 40 nt right next to the 3′ adapter of read 2. This was used in searching for the poly(A)-free 3′ hydroxyl ends. (3) paired alignment: using the full read 1 sequences and part of read 2 sequences trimmed of degenerate bases and delimiter. We filtered out poly(A) stretches encoded from genome using this alignment set. All the alignments were performed with maximum mismatches of 5%, minimum mapping quality of 3. All multi-mapped reads were removed. The remaining PCR artifacts with few mismatches were removed again using the R1 alignment with 15 degenerate bases inside the 3′ adapter region. To detect that kind of artifacts, we clustered the R1 alignments with maximum distance between mapped positions of 10 bp, they were then clustered again within the first cluster using degenerate bases from read 2 of respective reads with CD-HIT-EST 4.5.4 (word size=6, sequence identity=0.85). For a set of detected duplicates, we chose a read with maximum sum of PHRED quality in read 1 to leave.
For classification and transcript-level analyses, we compiled reference annotations for human and mouse using NCBI RefSeq, RepeatMasker, gtRNAdb, Rfam and miRBase databases (the first three were downloaded from the UCSC Genome Browser on Apr 25, 2013; Rfam version 11; miRBase version 19). The R1 alignments were annotated with intersection with the compiled annotations using BEDTools {Quinlan, 2010 #66}. When multiple annotations were overlapped to an alignment, we chose a class for the statistics requiring exclusive assignment of a genomic source type by the following priority: miRNA, rRNA, tRNA, Mt-tRNA, snoRNA, scRNA, srpRNA, snRNA, lncRNA, RNA, ncRNA, misc_RNA, Cis-reg, ribozyme, RC, IRES, frameshift_element, LINE, SINE, Simple_repeat, Low_complexity, Satellite, DNA, LTR, CDS, 3′ UTR, 5′ UTR, intron, Other, Unknown (higher priority first). The transcript-level analyses were performed using our custom non-redundant RefSeq (nrRefSeq) transcript set, which is a reduced set retaining only the longest isoform or transcript when regions overlap with each other. The positions of read 1 in nrRefSeq transcripts were positioned with BEDTools intersection between alignments to genome sequences and nrRefSeq annotation set, and then translated to the transcript-level coordination with in-house software.
As poly(A) tails were initially detected with a constraint that it must begin within the first 30 cycles, so the maximum detectable 3′ end modification of poly(A) tails was limited to the last 30 nucleotides of insert. To exclude A stretches obviously encoded from genomic sequence (with or without 3′ end modifications), we masked detected poly(A) tail ranges with read 2 alignments so that the 3′-most position of alignable (not clipped) is eliminated from poly(A) tail or its 3′ end modifications. All statistics regarding transcript-level modification rates were calculated for transcripts having more than 200 tags with poly(A) tails longer than 8 nt.
Genome_build: hg19 for HeLa, mm10 for NIH3T3
Supplementary_files_format_and_content: The .csv files contain the poly(A) tail length distribution and 3' end modification frequencies next to poly(A) tails for all detected transcripts with >= 30 poly(A)+ tags

extract_protocol:
Total RNAs were extracted from NIH3T3 or HeLa cells by TRIzol reagent (Invitrogen, 15596-018) and purified by RNeasy MinElute column (Qiagen, 74204) according to manufacturer’s instruction.
The RNA was depleted of rRNA by using Ribo-zero kit (Epicentre, MRZH11124), ligated to 3′ adapter and partially digested by RNase T1 (Ambion, AM2283). The fragmented RNAs were pull-downed with streptavidin beads (Invitrogen, 11206D), and phosphorylated by PNK reaction (Takara, 2021B) and gel purified in the range of 500 – 1000 nucleotides. The purified RNAs were ligated to 5′ adapter, subjected to reverse-transcription (Invitrogen, 18080085) and amplified by PCR using Phusion DNA polymerase (Thermo, F-530L). PCR products were gel purified again. All adapters and primers are synthesized by IDT.

tag: cell type

value: mouse fibroblast cell line


title: 89_LIV_pre-puberty

value: PRE-puberty


title: 105_FAT_post-puberty

source_name: Fat

value: Fat


title: 7104_FAT_post-puberty


title: 40_FAT_post-puberty


title: Fetal Placenta at E11.5, Litter B4, Sample 5


title: 117_LIV_pre-puberty


title: 87_LIV_pre-puberty


title: 7102_FAT_pre-puberty


title: 89_FAT_pre-puberty


title: 117_FAT_pre-puberty


title: 87_FAT_pre-puberty


title: 7104_LDM_post-puberty

source_name: Longissimus dorsi muscle

value: Longissimus dorsi muscle


title: Fetal Placenta at E11.5, Litter B3, Sample 5


title: 36_FAT_post-puberty


title: 105_LDM_post-puberty


title: 7102_LDM_pre-puberty


title: 40_LDM_post-puberty


title: 117_LDM_pre-puberty


title: 7102_LIV_pre-puberty


title: 89_LDM_pre-puberty


title: 105_ENDO_post-puberty

source_name: Endometrium

value: Endometrium


title: 36_ENDO_post-puberty


title: 87_LDM_pre-puberty


title: 40_ENDO_post-puberty


title: 89_ENDO_pre-puberty


title: 87_ENDO_pre-puberty


title: 117_ENDO_pre-puberty


title: 105_UTE_post-puberty

source_name: Uterus

value: Uterus


title: 40_UTE_post-puberty


title: 7102_UTE_pre-puberty


title: 36_UTE_post-puberty


title: 87_UTE_pre-puberty


title: 7104_OVA_post-puberty

source_name: Ovary

value: Ovary


title: 40_OVA_post-puberty


title: CD4 Input donor E

city: Regensburg

first: Michael

last: Rehli

address: F.-J.-Strauss-Allee 11

department: Internal Med III

institute: University Hospital Regensburg

description: Input

data_processing:
Sequence tags were mapped to the current human reference sequence (GRCh27/hg19) using Bowtie and only uniquely mapped tags were used for downstream analyses.
Genome_build: hg19
Supplementary_files_format_and_content: bed

source_name: CD4+CD25-CD45 RA+ naive conventional T cells

growth_protocol: Peripheral blood mononuclear cells were isolated from leukapheresis products of healthy volunteers by density gradient centrifugation over Ficoll/Hypaque (Biochrom AG, Berlin, Germany). Collection of blood cells from healthy donors was performed in compliance with the Helsinki Declaration. All donors signed an informed consent. The leukapheresis procedure and subsequent purification of peripheral blood cells was approved by the local ethical committee (reference number 92-1782 and 09/066c). CD4+ cells were enriched using magnetically labeled human CD4 MicroBeads (Miltenyi Biotec, Bergisch Gladbach, Germany) and the Midi-MACS system (Miltenyi Biotec). The CD4+ fraction was stained with CD4 FITC (BD, Klon SK3, 345768), CD25 PE (BD, Klon 2A3, 341011) and CD45RA APC and CD3+CD4+CD25- T cells were sorted on a FACS-Aria  high-speed cell sorter (BD Biosciences, Heidelberg, Germany). CD8+ cells were enriched using magnetically labeled human CD8 MicroBeads (Miltenyi) . The CD8+ fraction was stained with CD3 FITC (BD, Klon SK7, 345763) and CD8 APC (BD, Klon SK1, 345775) and sorted for CD3+CD8+ T cells. CD19+ and CD56+ cells were enriched from the CD8- fraction using magnetically labeled human CD19 and CD56 MicroBeads (Miltenyi). Enriched cells were stained with CD3 FITC (BD, Klon SK7, 345763), CD19 PE (BD, Klon 4G7, 345777) and CD56 APC (BD, Klon NCAM16.2, 341027) and sorted into CD3+CD19+ B cells and CD3+CD56+ NK cells.

extract_protocol: DNA from chromatin immunoprecipitation (10-30ng) was adapter-ligated and PCR amplified using NEBNext DNA Sample Prep Reagent Set1 (NEB E6000L) and Adapter Oligo Mix and PCR primers from Illumina following the manufacturer’P fragments were sequenced for 36 cycles on Illumina Genome Analyzers.

tag: donor

value: E

tag: donor gender

value: male

tag: donor age (y)

value: Input


title: 105_OVA_post-puberty


title: 36_OVA_post-puberty


title: 89_UTE_pre-puberty


title: CD8 Input donor C

description: Input

source_name: CD8+ T cells

value: CD8+ T cells

value: C


title: 7102_OVA_pre-puberty


title: CD8 Input donor A

description: Input

value: A


title: 117_OVA_pre-puberty


title: NK H3K27ac donor C

description: Chromatin IP against H3K27ac

source_name: CD56+ Natural Killer cells

value: CD56+ Natural Killer cells

tag: chip antibody manufacturer

value: abcam

tag: chip antibody catalog no.

tag: chip antibody lot no.


title: 89_OVA_pre-puberty


title: JAG-GFP replicate 1

status: Public on Mar 05 2014

city: NORWICH

first: Robert

last: Sablowski

state: Norfolk

address: The John Innes Centre

department: CDB

institute: JIC

data_processing:
Basecalls performed using CASAVA version 1.8
Sequenced reads were mapped to TAIR10 whole genome using SOAPv2 -r 0
CSAR (default parameter except backg=20) was used for the peak calling step
Genome_build: TAIR10
Supplementary_files_format_and_content: csv; significant binding sites

source_name: JAG-GFP

growth_protocol: Plants were grown under long day conditions (16 h light and 8 h dark) at 18° C day temperature and at 20° C night temperature at 80% humidity.

extract_protocol:
pJAG:JAG-GFP jag-2 and wild-type L-er control plants were used. 1300-1500 mg  (fresh weight) of inflorescence apices per sample were fixed in 35 ml fixation buffer (0.4 M sucrose, 10mM Tris pH 8, 1 mM EDTA pH 8.5, 1% formaldehyde, 100 µM PMSF) under vacuum for 20 min on ice. Cross-linking was stopped with 100 µM  glycine for 10 min on ice. After two washes with sterile water, the tissue was blotted dry and frozen in liquid nitrogen. Nuclei were purified as described [Gomez-Mena et al., 2005, Development 132: 429-438] and resupended in 1 ml of sonication buffer containing 500 mM Hepes, 150 mM NaCl, 5 mM MgCl2, 10% TRITON X-100 and ½ a tablet of protease inhibitor cocktail complete Mini, EDTA-free (Roche). Sonication was performed in a Bioruptor waterbath sonicator at 4°C (2 x 5 min high power level with 30 sec on/30sec off cycles) resulting in an average fragment size of 500 bp.  After centrifugation, the supernatant was mixed with 500 ul of immunoprecipitation buffer containing 0.5M Hepes, 150 mM NaCl, 5 mM MgCl2 , 10% TRITON X-100, 1 mg/ml BSA and 25 µl of anti-GFP µMACS Microbeads (Milteyi Biotec), incubated on ice for 30 min and loaded on a µ Column (Milteyi Biotec) that had been equilibrated with 200 µl immunoprecipitation buffer and placed into a magnetic µMACS separator (Milteyi Biotec). After washing twice with 400 µl and twice with 200 µl of immunoprecipitation buffer, then twice with 200 µl TE buffer (100 mM Tris pH 8, 10 mM EDTA pH 8), DNA was eluted once with 20 µl and twice with 50 µl of preheated (96° C) elution buffer containing 50 mM Tris pH8, 10 mM EDTA, 50 mM DTT and 1% SDS. 100 µl of TE-buffer and 9 µl of 25 mg/ml Proteinase K (Sigma) were added to the eluted samples and to the input control samples. Crosslinking was reverted in eluted and input samples at 37°C overnight, followed by addition of 9 µl of 25 mg/ml Proteinase K and 8 h incubation at 65° C, phenol-chloroform extraction and precipitation with ethanol overnight at -20° C. After washing in 70% EtOH, the air-dried DNA was resuspended in 100 µl PCR-grade water (Roche), purified using a PCR purification Kit (18104, Qiagen) and stored at -80° C.
Illumina TruSeq ChIP-Seq libraries (three pJAG: JAG-GFP replicates and three wild-type controls) were produced as described [Kaufmann et al, 2009, PLoS Biology 7: e90.]

value: jag-2, JAG:JAG-GFP

value: inflorescence tips (only unopened buds)


title: CD19 Input donor A

description: Input

source_name: CD19+ B cells

value: CD19+ B cells


title: 7104_PIT_post-puberty

source_name: Pituitary gland

value: POST-puberty

value: Pituitary gland

value: Brangus (3/8 Brahman x 5/8 Angus)


title: 117_UTE_pre-puberty

value: PRE-puberty


title: NK H3K4me1 donor A

description: Chromatin IP against H3K4me1


title: NK H3K27ac donor A

description: Chromatin IP against H3K27ac


title: 105_PIT_post-puberty


title: 7104_UTE_post-puberty

value: Uterus


title: NK H3K4me1 donor C

description: Chromatin IP against H3K4me1


title: NK Input donor A

description: Input


title: 117_PIT_pre-puberty


title: CD4 H3K27ac donor F

description: Chromatin IP against H3K27ac

value: CD4+CD25-CD45 RA+ naive conventional T cells

value: F


title: 36_PIT_post-puberty


title: 87_PIT_pre-puberty


title: 105_HYP_post-puberty

source_name: Hypothalamus

value: Hypothalamus


title: 40_HYP_post-puberty


title: 40_PIT_post-puberty


title: CD4 H3K27ac donor E

description: Chromatin IP against H3K27ac

value: E

value: male


title: CD4 H3K4me1 donor F

description: Chromatin IP against H3K4me1


title: CD4 H3K4me1 donor E

description: Chromatin IP against H3K4me1


title: CD8 H3K27ac donor C

description: Chromatin IP against H3K27ac

value: CD8+ T cells

value: C


title: CD8 H3K4me1 donor C

description: Chromatin IP against H3K4me1


title: CD8 H3K27ac donor A

description: Chromatin IP against H3K27ac

value: A


title: CD8 H3K4me1 donor A

description: Chromatin IP against H3K4me1

value: abcam


title: 87_HYP_pre-puberty


title: 36_LDM_post-puberty

value: Longissimus dorsi muscle


title: 117_HYP_pre-puberty


title: 87_OVA_pre-puberty

value: Ovary


title: 36_HYP_post-puberty


title: CD19 H3K27ac donor C

description: Chromatin IP against H3K27ac

value: CD19+ B cells


title: 89_HYP_pre-puberty


title: CD19 H3K27ac donor A

description: Chromatin IP against H3K27ac


title: Monte Prinzera 3

city: Kuopio

first: Pauliina

last: Halimaa

address: Yliopisonranta 1 E

department: Department of Biology

country: Finland

institute: University of Eastern Finland

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
20_MP3_counts.txt: TAIR9

organism: Noccaea caerulescens

source_name: Monte Prinzera

growth_protocol: hydroponic culture as described in Schat, H. et al. 1996. Evolution Int. J. O. Evolution 50, 1888–1895, except for GA and LC 10 µM Zn, for MP 2  µM Zn and 10  µM Ni

extract_protocol: The libraries were constructed following the SOLiD™ Total RNA-Seq Kit's (Whole Transcriptome library) protocol, Applied Biosystems Part Number 4452437 Rev A, 01/2010

tag: ecotype

value: Monte Prinzera

tag: origin

value: Ultramafic soil (serpentine, high Ni)

tag: metal accumulation and tolerance

value: Assunção, A.G.L., et al. 2003. New Phytol. 159, 411-419.

value: pool of 12 root samples


title: Monte Prinzera 6

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
22_MP6_counts.txt: TAIR9


title: Ganges 6

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
19_GA6_counts.txt: TAIR9

source_name: Ganges

value: Ganges

value: Zn/Pb mine spoil (calamine)


title: Ganges 4

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
18_GA4_counts.txt: TAIR9


title: CD19 H3K4me1 donor C

description: Chromatin IP against H3K4me1

tag: chip antibody


title: H3K27ac CD14dimCD16+ donor D

description: Chromatin IP against H3K27ac

data_processing:
Sequence tags were mapped to the current human reference sequence (GRCh27/hg19) using Bowtie and only uniquely mapped tags were used for downstream analyses. All samples (except the inputs) represent data from 2 donors.
Genome_build: hg19
Supplementary_files_format_and_content: bed files

source_name: CD14dimCD16+ "non-classical" blood monocytes

growth_protocol: Peripheral blood monocytes were separated by leukapheresis of healthy donors, followed by density gradient centrifugation over Ficoll/Hypaque and subsequent counter current centrifugal elutriation in a J6M-E centrifuge (Beckman, München, Germany). Collection of blood cells from healthy donors was performed in compliance with the Helsinki Declaration. All donors signed an informed consent. The leukapheresis procedure and subsequent purification of peripheral blood monocytes was approved by the local ethical committee (reference number 92-1782 and 09/066c).

treatment_protocol: The monocyte-enriched cell fraction was further subdivided by FACS (using BDs FACSAria) after staining with CD2-PE (RPA-2.10), CD15-PE (HI98), CD19-PE (4G7), CD56-PE (B159), NKp46-PE (9E2), to exclude remaining lymphocytes and granulocytes, as well as CD14-V450 (MΦP9) and CD16-Alexa647 (3G8) antibodies (all from BD). Monocytes were sorted into CD14++CD16– ("classical" subset), CD14+CD16+("intermediate" subset) as well as CD14dimCD16+ cells ("nonclassica"l subset).

value: D

tag: chip antibody catalog #

tag: chip antibody lot #


title: CD19 H3K4me1 donor A

description: Chromatin IP against H3K4me1


title: Input donor A

description: Input control

value: no antibody

value: CD14dimCD16+ "non-classical" blood monocytes


title: Monte Prinzera 4

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
21_MP4_counts.txt: TAIR9

tag: tissue


title: H3K27ac CD14dimCD16+ donor A

description: Chromatin IP against H3K27ac


title: H3K4me1 CD14dimCD16+ donor D

description: Chromatin IP against H3K4me1


title: H3K4me1 CD14dimCD16+ donor A

description: Chromatin IP against H3K4me1


title: 89_PIT_pre-puberty

value: PRE-puberty

value: Pituitary gland

value: Brangus (3/8 Brahman x 5/8 Angus)


title: Input donor D

description: Input control


title: La Calamine 4

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
15_LC4_counts.txt: TAIR9

source_name: La Calamine

value: La Calamine


title: H3K27ac CD14+CD16- donor D

description: Chromatin IP against H3K27ac

source_name: CD14+CD16- "classical" blood monocytes

value: CD14+CD16- "classical" blood monocytes

value: abcam


title: La Calamine 6

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
16_LC6_counts.txt: TAIR9


title: H3K4me1 CD14+CD16- donor A

description: Chromatin IP against H3K4me1

value: A


title: Ganges 3

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
17_GA3_counts.txt: TAIR9


title: H3K27ac CD14+CD16- donor A

description: Chromatin IP against H3K27ac


title: La Calamine 3

data_processing:
Counts: Sequence reads were obtained and mapped to the Arabidopsis (TAIR9) genome using SOLiD Bioscope software with default settings. Counts included.
Genome Build:
14_LC3_counts.txt: TAIR9


title: Clone 12

city: Newcastle

first: Simon

last: Cockell

address: Framlington Place

institute: Newcastle University

data_processing:
Read Alignment: Bowtie v 1.0.0, default parameters
Unaligned reads filtered: Samtools v0.1.18, options:  view -F 0x04
Mapped reads intersecting with exons & miRNAs found: BEDTools (intersectBed) v2.12.0 - target BED files from UCSC genome browser
Count reads intersecting with features (standard Linux tools - awk, sort, uniq)
Genome_build: hg18
Supplementary_files_format_and_content: BED, reads intersecting with exons and miRNAs, Excel file (read counts)

source_name: HEK293 cells mutagenised with 100 μg/ml ethyl methanesulfonate

growth_protocol: The human embryonic kidney cell line HEK293 was maintained and passaged according to established cell culture conditions

extract_protocol:
RNA and DNA were isolated using Trizol according to standard methodology. The RNA was quality tested and size selected on a denaturing polyacrylamide gel (19-29 bases).
Tagged 3’ adaptors and 5’ adaptors were ligated to the recovered RNA to ensure strand specificity. The material was used for cDNA synthesis.

treatment_protocol: When the cells were about 80% confluent, the medium in one well was replaced and 100 μg/ml ethyl methanesulfonate (EMS; Sigma) was added. Cells were grown in the presence of EMS for 24 hours followed by a 24 hour recovery period in fresh medium.

tag: cell line

value: human embryonic kidney cells

tag: exposed to

value: 100 μg/ml ethyl methanesulfonate

tag: exposure length


title: 7104_HYP_post-puberty

tag: developmental stage

value: POST-puberty

value: Hypothalamus

tag: breed


title: 7102_HYP_pre-puberty


title: K562-aza-CTCF-bisulfite_1

data_processing:
Methylation values: Capture bisulfite reads were mapped to the human genome (hg19) using Bismark (Krueger and Andrews 2011) and bowtie2 beta 6 (Langmead et al. 2009) with the options “-n 1” and excluding duplicate reads and combining reads from the top and bottom strands. Methylation was monitored at CpGs with ≥8 read coverage, and averaged for all CpGs within the 150 bp window at each CTCF site.
Peak: ChIP-seq affinity zones (HotSpots) were identified using the HotSpot algorithm described in Sabo et al. (2004). 1.0% false discovery rate thresholds (FDR 0.01) were computed for each cell type by applying the HotSpot algorithm to an equivalent number of random uniquely mapping 36mers. DNase data used unthresholded hotspots.
Density: Smoothed density tracks were generated using bedmap (http://code.google.com/p/bedops/) to count the number of tags overlapping a sliding 150-bp window, with a resolution of 20 bp (Neph et al. 2012a). Density tracks for display were normalized for sequencing depth by a global linear scaling to 10 million tags.
Alignment: ChIP-seq and DNase data were mapped to the human genome (GRCh37/ hg19) using bowtie (Langmead et al. 2009) with the options ‘‘bowtie -n 2 -m 1 -e 70 --best’’ allowing up to three mismatches. Reads mapping to multiple locations were then excluded, and reads with identical 5’ ends and strand were presumed to be PCR duplicates and were excluded using Picard MarkDuplicates.
Genome_build: hg19
Supplementary_files_format_and_content: .bed: peak locations generated using the hotspot algorithm; .bed proportion of bisulfite tags indicating methylation per CpG (proportion methylation in bed score column, number of total reads in 7th column) .bw: the number of tags overlapping a sliding 150-bp window, with a resolution of 20 bp; .bam: tags mapped to the human genome

source_name: 5-aza-2-deoxycytidine-treated K562 erythroleukemia cells

extract_protocol:
Purified DNA from mock- and 5-aza-CdR-treated K562 cells was fragmented following the Agilent SureSelect Methyl-Seq protocol with slight modifications.  4 ug from each of the samples was fragmented in a Covaris S2 (Covaris) under the following conditions: 10% duty cycle at intensity 5 for 60 seconds with 200 cycles per burst for 6 cycles with mode set to sweeping.  The 250 bp fragmented DNA was then end repaired and adenylated followed by ligation to adapters synthesized with 5’ –methyl-cytosine instead of cytosine. The adaptor-ligated library was purified using AMPure XP beads (Beckman Coulter Genomics). 500 ng of each library was hybridized to Agilent SureSelect Methyl-Seq biotinylated RNA baits (84 Mb) for 24 h at 65 C.  The biotinylated probe/target hybrids were captured on Dynal MyOne Streptavidin T1 (Invitrogen), washed, eluted, and desalted following purification on a MinElute PCR column (Qiagen) as described in the SureSelect protocol. Bisulfite conversion of the purified captured library was performed using the EZ DNA Methylation Gold Kit (Zymo Research) as per manufacturer’s instructions. The bisulfite converted captured library was amplified by PCR with minimal amount of PCR cycles, then purified using AMPure XP beads and quantified by Qubit dsDNA assay (Invitrogen) following SureSelect protocol.
20-50 ng of ChIP DNA was end-repaired, adenine ligated to Illumina adapters was added, and then a Solexa library was made for sequencing.

tag: experiment

value: Bisulfite-Seq

tag: treatment

value: K562 cells were treated with 5-aza-2’-deoxycytidine (decitabine; Sigma, A3656) dissolved in DMSO to 10mM. Drug was administered at 1uM daily for 3 days. Control K562 cells were mock treated with DMSO.

tag: replicate


title: K562-Mock-CTCF-bisulfite_1

source_name: Mock-treated K562 erythroleukemia cells


title: HCT116-DKO-DNaseI_1

source_name: HCT116 colorectal cancer cells DNMT1KO/DNMT3BKO

value: Dnase-Hypersensitivity

value: No special treatment


title: HCT116-H3K27ac_1

source_name: HCT116 colorectal cancer cells

value: H3K27ac ChIP-seq


title: HCT116-DKO-H3K4me3_1

value: H3K4me3 ChIP-seq


title: HCT116-DKO-H3K27ac_1


title: K562-Mock-CTCF_3

value: CTCF ChIP-seq


title: K562-aza-CTCF_1


title: HeLa-miR-1-9h

description: TAIL-seq for HeLa cell line 9h post-transfection of miR-1

data_processing:
The base calls and signal intensities were acquired from HiSeq 2500 after processing by Illumina RTA 1.17.21.3. The read 1 sequences were aligned to the common contaminants set, which is composed of rDNA repeat units (GenBank accession BK000964.1 for NIH3T3 and U13369.1 for HeLa), PhiX genome (GenBank accession J02482.1), Illumina TruSeq primer sequences, and all sequences for 5S and 5.8S rRNAs of respective species (retrieved from Rfam 11.0 of the Wellcome Trust Sanger Institute) using GSNAP 2013-03-31 with maximum 5% mismatches allowed. Clusters with any match to the contaminants were removed from the subsequent analyses. The sequences having completely identical nucleotides in the 21st to 35th cycle in read 1 (representative region of the insert) and the 1st to 15th cycle in read 2 (degenerate bases in 3′ adapter) are deduplicated by leaving only a cluster with the maximum PHRED quality sum of read 1. The degenerate and fixed delimiter sequence in 3′ adapter was clipped out from read 2 by searching perfect match of delimiter sequence (‘GTCAG’ as in the direction of read 2) between the 14th and 16th cycles in read 2. The clusters missing a delimiter sequence or having low diversity in degenerate region (at least two occurrences for all of A, C, G and T) were removed from further analyses.
The fluorescence signal intensities were processed into “Normalized T signal” as described in fig. S1A. The signals from a spike-in sample were purified with an outlier filter based on robust Mahalanobis distance (mvoutlier package 1.9.9; quan=0.5, alpha=0.025). Random 500 clusters per each spike-in were chosen for parameter calculation of a Gaussian mixture hidden Markov model (GMHMM). We trained the model using Baum-Welch algorithm implemented in the GHMM library (http://ghmm.org) with topology and initial parameters shown in fig. S1A and table S7 and S8 (1,000 iterations). The procedure was iterated to maximize likelihood, not using any property (eg. designed length of poly(A) tail) of spike-ins. Normalized T signals outside the range of [-5, 5] were clipped into the range for both training and later calculations.
The length of poly(A) tails were first measured with base call-based “Strategy II” described in fig. S1A. For clusters with the measured length is shorter than 8 nt, the length is called as the final poly(A) tail length. For the others, normalized T signals starting from the first position in T-stretch detected by Strategy II were analyzed with the GMHMM. The hidden states were decoded with the standard Viterbi algorithm implemented in the GHMM library. The number of cycles with state 1 and 2 was called as the length of poly(A) tail. For the estimation of performance, we applied the process to all spike-in samples except the clusters used for the parameter fitting of the model.
The remaining reads after contaminant filter and the first duplication filters were then aligned to the genome sequences (UCSC mm10 for NIH3T3 and UCSC hg19 for HeLa, positions of splicing junctions were processed from the UCSC Genome Browser database for version of Jan 24, 2013) using GSNAP 2013-03-31. Three different versions of alignments to genome were used in this study. (1) R1 alignment: using only the full read 1 sequences which are 51 nt long. This was used for identification of a cluster. (2) R2 short alignment: using only 40 nt right next to the 3′ adapter of read 2. This was used in searching for the poly(A)-free 3′ hydroxyl ends. (3) paired alignment: using the full read 1 sequences and part of read 2 sequences trimmed of degenerate bases and delimiter. We filtered out poly(A) stretches encoded from genome using this alignment set. All the alignments were performed with maximum mismatches of 5%, minimum mapping quality of 3. All multi-mapped reads were removed. The remaining PCR artifacts with few mismatches were removed again using the R1 alignment with 15 degenerate bases inside the 3′ adapter region. To detect that kind of artifacts, we clustered the R1 alignments with maximum distance between mapped positions of 10 bp, they were then clustered again within the first cluster using degenerate bases from read 2 of respective reads with CD-HIT-EST 4.5.4 (word size=6, sequence identity=0.85). For a set of detected duplicates, we chose a read with maximum sum of PHRED quality in read 1 to leave.
For classification and transcript-level analyses, we compiled reference annotations for human and mouse using NCBI RefSeq, RepeatMasker, gtRNAdb, Rfam and miRBase databases (the first three were downloaded from the UCSC Genome Browser on Apr 25, 2013; Rfam version 11; miRBase version 19). The R1 alignments were annotated with intersection with the compiled annotations using BEDTools {Quinlan, 2010 #66}. When multiple annotations were overlapped to an alignment, we chose a class for the statistics requiring exclusive assignment of a genomic source type by the following priority: miRNA, rRNA, tRNA, Mt-tRNA, snoRNA, scRNA, srpRNA, snRNA, lncRNA, RNA, ncRNA, misc_RNA, Cis-reg, ribozyme, RC, IRES, frameshift_element, LINE, SINE, Simple_repeat, Low_complexity, Satellite, DNA, LTR, CDS, 3′ UTR, 5′ UTR, intron, Other, Unknown (higher priority first). The transcript-level analyses were performed using our custom non-redundant RefSeq (nrRefSeq) transcript set, which is a reduced set retaining only the longest isoform or transcript when regions overlap with each other. The positions of read 1 in nrRefSeq transcripts were positioned with BEDTools intersection between alignments to genome sequences and nrRefSeq annotation set, and then translated to the transcript-level coordination with in-house software.
As poly(A) tails were initially detected with a constraint that it must begin within the first 30 cycles, so the maximum detectable 3′ end modification of poly(A) tails was limited to the last 30 nucleotides of insert. To exclude A stretches obviously encoded from genomic sequence (with or without 3′ end modifications), we masked detected poly(A) tail ranges with read 2 alignments so that the 3′-most position of alignable (not clipped) is eliminated from poly(A) tail or its 3′ end modifications. All statistics regarding transcript-level modification rates were calculated for transcripts having more than 200 tags with poly(A) tails longer than 8 nt.
Genome_build: hg19
Supplementary_files_format_and_content: The csv files contain the poly(A) tail length distribution and 3' end modification frequencies next to poly(A) tails for all detected transcripts with >= 30 poly(A)+ tags

growth_protocol: HeLa cells were maintained in DMEM (Welgene) supplemented with 10% fetal bovine serum (Welgene).

extract_protocol:
Total RNAs were extracted from NIH3T3 or HeLa cells by TRIzol reagent (Invitrogen, 15596-018) and purified by RNeasy MinElute column (Qiagen, 74204) according to manufacturer’s instruction.
The RNA was depleted of rRNA by using Ribo-zero kit (Epicentre, MRZH11124), ligated to 3′ adapter and partially digested by RNase T1 (Ambion, AM2283). The fragmented RNAs were pull-downed with streptavidin beads (Invitrogen, 11206D), and phosphorylated by PNK reaction (Takara, 2021B) and gel purified in the range of 500-1000 nucleotides. The purified RNAs were ligated to 5′ adapter, subjected to reverse-transcription (Invitrogen, 18080085) and amplified by PCR using Phusion DNA polymerase (Thermo, F-530L). PCR products were gel purified again. All adapters and primers are synthesized by IDT.

treatment_protocol: HeLa cells were transfected with 50 nM of synthetic miR-1 duplex using Lipofectamine 2000 (Life Technologies). Total RNAs were extracted at 0, 3, 6, and 9 hr after transfection. miR-1 guide 5′- UGGAAUGUAAAGAAGUAUGUA-3′;; miR-1 passenger 5′-CAUACUUCUUUACAUUCCAUU- 3′.

value: HeLa

tag: cell type

value: human cervical cancer cells

tag: transfected with

value: synthetic miR-1 mimic

tag: time point

value: 9hrs post-transfection


title: K562-Mock-CTCF_2


title: K562-Mock-CTCF_1


title: HCT116-DKO-CTCF_1


title: HCT116-DNMT1KO-CTCF_1

source_name: HCT116 colorectal cancer cells DNMT1KO


title: HeLa-miR-1-3h

description: TAIL-seq for HeLa cell line 3h post-transfection of miR-1

value: 3hr post-transfection


title: HCT116-DKO-CTCF_2


title: HCT116-CTCF_3


title: HeLa-miR-1-0h

description: TAIL-seq for HeLa cell line 0h post-transfection of miR-1

value: 0h post-transfection


title: RNA4_KO_BRD4

description: Sample 4

data_processing:
Illumina Casava1.3 software used for basecalling for the samples 1,2,5,6,7,8. Illumina Casava 1.8.2 was used for the samples 3 & 4
Sequenced reads were mapped to hg19/GRCh37 whole genome using bwa 0.5.9-r16 with default parameters
Exon read coverages were obtained with coverageBed v2.17.0 using exon coordinates of the Ensembl database v63. Transcript counts were generated by summing read counts across all exons of a given transcript and transcript counts were normalized by the number of sequenced reads (sequencing depth) to ensure comparable counts among samples.
Using the bam files produced by bwa as input peaks were called with MACS v1.4 (PMID: 18798982) using an enrichment ratio against the background of 5 to 15 to build the model (m=5,15).
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include read count values for each transcript model of the Ensembl database v63 and peaks called by MACS.

source_name: HEK_BRD4 KO_RNA-seq

value: shRNA against BRD4


title: H3K4me1 CD14+CD16- donor D

description: Chromatin IP against H3K4me1

value: CD14+CD16- "classical" blood monocytes

tag: donor

value: D

tag: chip antibody manufacturer

value: abcam


title: HCT116-CTCF_2


title: RNA3_ctrl

description: Sample 3

value: none (control)


title: Clone 5

value: human embryonic kidney cells

value: 100 μg/ml ethyl methanesulfonate


title: RNA-seq; Sample 36_HTS589

status: Public on Feb 27 2014

city: Corvallis

first: Kristina

middle: M

last: Smith

state: OR

department: Biochemisty and Biophysics

institute: Oregon State University

data_processing:
Basecalls were made with RTA1.8_CASAVA1.7
ChIPSeq reads were trimmed of adapter sequence and mapped to the F. graminearum genome with BWA software using default options
RNASeq reads were trimmed of adapter sequence and mapped to the F. graminearum genome with Tophat software with options (-a 5 -m 1 -i 30 -I 2000)
samtools were used to convert BWA or Tophat output to sorted bam files
cufflinks v2.1.1 was used (--max-bundle-length 20000 -I 2000 --min-intron-length 30) to calculate FPKM values for Broad assembly 3 transcripts from sorted bam files of RNA-seq reads
Genome_build: Broad chromosome assembly 3
Supplementary_files_format_and_content: ChIP-seq processed files are tab delimited EpiChIP output containing gene name, NLCS value
Supplementary_files_format_and_content: RNA-seq processed files are tab delimited cufflinks output containing gene name, FPKM value

organism: Fusarium graminearum

source_name: mycelium

extract_protocol: mRNA was isolated using a Poly(A)Purist MAG kit (Ambion). We removed DNA by treatment with RNase-free DNAase (Qiagen), followed by column clean-up according to manufacturers instructions. We used Illumina TruSeq RNA Sample Preparation kits to make RNA-seq libraries

tag: strain number

tag: genotype

tag: nitrogen level

value: low


title: C

city: Lanzhou

first: Ruiqing

last: Zhu

state: Gansu

address: DingxiSouth Road 197

department: Instituye of biology

institute: Gansu Academy of Sciences

data_processing:
Sequencing-received raw image data were transformed by base calling into sequence data. Prior to mapping reads to the reference database, all sequences were filtered to remove 3′ adaptor sequence, low quality sequences (tags with unknown sequences ‘N’), tags with a copy number of 1 (probably sequencing error), and tags which are too long or too short, leaving clean tags of 21 nt. For annotation, all clean tags were mapped to the reference sequences, and a mismatch of only 1 bp was considered.
The clean reads mapped to reference database from multiple genes were filtered.
The number of unambiguous clean tags for each gene was calculated and then normalized to TPM (number of transcripts per million clean tags)

organism: Reaumuria songarica

source_name: leaf

growth_protocol: The three-year-old plants were transferred to a greenhouse under the following conditions: 150 µmol m-2∙s-1 (fluorescent tubes), photoperiod 16 h light/8 h darkness, day/night temperature of 25/20 °C. The plants were washed with deionized water to remove surface contaminants. Soil attached to roots was also rinsed off with deionized water, and then the plants were cultivated in 1/8 Murashige and Skoog medium solution with insufflating air for two weeks to accommodate the liquid environment during treatments.

extract_protocol:
CTAB-method
library construction, and sequencing were carried out at Beijing Genome Institute at Shenzhen, China. An extract of 8 μg of total RNA was obtained and treated with oligo(dT) magnetic bead adsorption to purify mRNA; oligo(dT) was then used as a primer to synthesize the first- and second-strand cDNA. The 5′-ends of tags can be generated by two types of endonuclease: NlaIII or DpnII. Then, the bead-bound cDNA is subsequently digested with restriction enzyme NlaIII, which recognizes and severs the CATG sites. Fragments other than the 3′-cDNA fragments connected to oligo(dT) beads are washed away, and the Illumina adaptor 1 is ligated to the sticky 5′-end of the digested bead-bound cDNA fragments. The junction of the Illumina adaptor 1 and the CATG site constitutes the recognition site of MmeI, which is a type of endonuclease with separate recognition and digestion sites; it breaks 17 bp downstream of the CATG site, producing tags with adaptor 1. After removal of 3′ fragments with magnetic beads precipitation, the Illumina adaptor 2 is ligated to the 3′-ends of tags, acquiring tags with different adaptors of both ends to form a tag library.

treatment_protocol: The experiment was arranged with two treatments (untreated and PEG-treated) and three replications. The untreated samples were maintained with the cultivating conditions as described previously. For drought treatment, 15% PEG6000 was added to the solution for dissolving. All the plants were treated for 12 h, from 6:00 to 18:00.

tag: age


title: RNA-seq; Sample 34_HTS587

value: high


title: RNA-seq; Sample 35_HTS588


title: K562-aza-CTCF_2

value: K562 cells were treated with 5-aza-2’-deoxycytidine (decitabine; Sigma, A3656) dissolved in DMSO to 10mM. Drug was administered at 1uM daily for 3 days. Control K562 cells were mock treated with DMSO.


title: RNA-seq; Sample 31_HTS584

value: FMF1 (FGSC9075)

value: WT


title: RNA-seq; Sample 33_HTS586


title: RNA-seq; Sample 32_HTS585