title: normal fetus in twins 7

source_name: healthy placentas

tag: tissue

value: normal


title: normal fetus in twins 6

status: Public on Aug 31 2015

type: SRA

city: Shenzhen

first: Desheng

last: Gong

address: No.7 PengFei road

country: China

institute: Agricultural Genomes Institute at Shenzhen

library_source: genomic

data_processing:
After removing the adapter sequences and filtering out the low quality reads, the bisulfite sequencing reads were directly aligned to the human reference genome (UCSC hg19) using BSMAP 2.73 with default parameters, which combines genome hashing and bitwise masking to achieve fast and accurate bisulfite mapping.
We filtered out the CpG sites with less than 4X sequencing depth in order to guarantee accurate calculation of methylation levels. Then analysed differential promoter DNA methylation between IUGR and normal fetus.
Genome_build: UCSC hg19
Supplementary_files_format_and_content: *.cout.gz files contain 8 columns. The format is that: (1)Chromosome ID, (2)Cytosine position, (3)+/- Watson/Crick chain, (4)Type (CG/CHH/CHG), (5)Base pattern, (6)Methylation Ratio, (7)Methylated reads num, (8)Un-methylated reads num.

molecule: genomic DNA

organism: Homo sapiens

extract_protocol:
Genomic DNA was extracted using the QIAamp DNA Blood Mini Kit (Qiagen). 1µg of genomic DNA was fragmented into approximately 200-300bp using a Covarias sonication system (Covarias). After purification, the fragmented fractions were treated with a mix of T4 DNA polymerase, Klenow Fragment and T4 polynucleotide kinase to repair blunt and phosphorylate ends. Total RNA was extracted using the Invitrogen TRIzol® Reagent and then treated with RNase-free DNase I (Ambion) for 30 min. The integrity of total RNA was checked using an Agilent 2100 Bioanalyser.
After purification, the fragmented fractions were treated with a mix of T4 DNA polymerase, Klenow Fragment and T4 polynucleotide kinase to repair blunt and phosphorylate ends. The blunt DNA fragments were subsequently 3'-adenylated using Klenow Fragment (3-5' exo-) and then ligated by T4 DNA Ligase to adapters that were synthesized with 5'-methylcytosine instead of 5'-cytosine and contain index sequences inside. After each step, the reaction products were purified using QIAquick PCR purification kit (Qiagen).  The constructed libraries were quantified using a Qubit fluorometer with Quant-iT dsDNA HS Assay Kit (Invitrogen).

value: placentas

tag: gender

value: F

tag: gestational weeks

tag: condition


title: IUGR fetus  in twins 5

source_name: IUGR placentas

value: M

value: IUGR


title: IUGR fetus  in twins 6


title: normal fetus in twins 5


title: normal fetus in twins 4


title: IUGR fetus  in twins 4


title: Mucosal_CD103_P_LC_Rep3

city: Rehovot

first: Ido

last: Amit

address: 234 Herzl st.

department: Immunology

country: Israel

institute: Weizmann Institute of Science

description: 3' RNA-seq for digital gene expression quantitation

library_source: transcriptomic

data_processing:
bcl2fastq v2.15.0.4
alignment: tophat --max-intron-length 10000 --max-multihits 1
Expression table done using ESAT http://garberlab.umassmed.edu/software/esat/
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text file includes mRNA molecule count values for each Sample

molecule: polyA RNA

organism: Mus musculus

source_name: Mucosal LCs CD103+

extract_protocol:
10^4 cells from each cell type were sorted in 100 ul of Lysis/Binding Buffer (Life technologies), lysed for 5 min and frozen at -80°C. Cell lysates were thawed and mRNA was captured with 12 ul of Dynabeads oligo(dT) (Life technologies), and washed according to manufacture guidelines. Purified messenger RNA was eluted at 70°C with 10 ul of 10 mM Tris-Cl pH 7.5 and stored at -80°C
cDNA was generated from 1ul of mRNA of each sample. cDNA quantity in each sample was evaluated by qPCR for Actin B gene, and then equivalent amounts of mRNA of each sample were taken for RNAseq library construction. Library construction was performed in a 96-well plate format. First, to open secondary RNA structures and allow annealing of the RT primer, the samples were incubated at 72˚C for 3 min and immediately transferred to 4˚C. Then, RT reaction mix (10 mM DTT, 4 mM dNTP, 2.5 U/µl Superscript III RT enzyme in 50 mM Tris-HCl (pH 8.3), 75 mM KCl, 3 mM MgCl2) was added into each well of the 96-well plate and the reaction was mixed. The 96-well plate was then spun down and moved into a cycler (Eppendorf) for the following incubation: 2 min at 42˚C, 50 min at 50˚C, 5 min at 85˚C. Indexed samples with equivalent amount of cDNA were pooled.  The pooled cDNA was converted to double-stranded DNA with a second strand synthesis kit (NEB) in a 20µl reaction, incubating for 2.5 h at 16˚C. The product was purified with 1.4x volumes of SPRI beads, eluted in 8 µl and in-vitro transcribed (with the beads) at 37˚C overnight for linear amplification using the T7 High Yield RNA polymerase IVT kit (NEB). Following IVT, the DNA template was removed with Turbo DNase I (Ambion) 15 min at 37˚C and the amplified RNA (aRNA) purified with 1.2x volumes of SPRI beads. Library preparation for high-throughput sequencing: The aRNA was chemically fragmented into short molecules (median size ~200 nucleotides) by incubating 3 min at 70˚C in Zn2+ RNA fragmentation solution (Ambion) and purified with two volumes of SPRI beads. The aRNA (5 µl) was preincubated 3 min at 70˚C with 1 µl of 100 µM ligation adapter; then, 14 µl of a mix containing 9.5% DMSO, 1 mM ATP, 20% PEG8000 and 1 U/µl T4 ligase in 50 mM Tris HCl pH7.5, 10 mM MgCl2 and 1mM DTT was added. The reaction was incubated at 22˚C for 2 h. The ligated product was reverse transcribed using Affinity Script RT enzyme (Agilent; reaction mix contains Affinity Script RT buffer, 10 mM DTT, 4 mM dNTP, 2.5 U/µl RT enzyme) and a primer complementary to the ligated adapter. The reaction was incubated for 2 min at 42˚C, 45 min at 50˚C and 5 min at 85˚C.  The cDNA was purified with 1.5x volumes of SPRI beads. The library was completed and amplified through a nested PCR reaction with 0.5 µM of P5_Rd1 and P7_Rd2 primers and PCR ready mix (Kapa Biosystems). The forward primer contains the Illumina P5-Read1 sequences and the reverse primer contains the P7-Read2 sequences. The amplified pooled library was purified with 0.7x volumes of SPRI beads to remove primer leftovers. Library concentration was measured with a Qubit fluorometer (Life Technologies) and mean molecule size was determined with a 2200 TapeStation instrument (Agilent). MARS-Seq libraries were sequenced using an Illumina HiSeq 1500.

tag: strain

tag: cell type

value: mucosal CD103+ LC


title: IUGR fetus  in twins 3


title: normal fetus in twins 3


title: YoungBeta_H3K27me3_ChIP

status: Public on Sep 01 2015

city: Jerusalem

first: Dana

last: Avrahami Tzfati

address: Ein-Kerem

department: Developmental Biology and cancer research

institute: Hadassah Hebrew University School of medicine

data_processing:
Illumina Casava-1.8.2 software was used for basecalling.
RNA-Seq: Low quality reads as well as ribosomal and other repeat sequences were filtered out before alignment. RUM software was used for aligning reads to mouse genome mm9 and Refseq transcriptome and for transcript quantitation. Differential expression analysis was carried out using a custom script implementing EdgeR software.
ChIP-Seq: Reads were aligned to mm9 genome using bowtie version 0.12.7 with parameters -v 3 -k 1 -m 1 --best --strata. Redundant reads were discarded before creating bedgraph profiles.
Peak calling was done using HOMER for H3K4me1 and H3K27Ac marks at FDR=0.01 with respect to corresponding Input. Peak calling using STAR was done for H3K27me3 marks at FDR=0.01.
BIS-Seq: DNA sequencing data was aligned to mouse genome (mm9) with the BS Seeker program. Data was reported as the fraction of reads that were methylated, calculated as the ratio of cytosine bases (indicating methylated CpGs) to total reads for each CpG coordinate.  Unmethylated cytosines are ultimately converted to thymine bases during the bisulfite conversion process.
Genome_build: mm9
Supplementary_files_format_and_content: Tab delimited file of transcript raw read counts.
Supplementary_files_format_and_content: Tab delimited file of transcript FPKM counts.
Supplementary_files_format_and_content: Peak files are tab-delimited listing genomic coordinates and their respective scores and p-values. Scores represent fold-change with respect to Input.
Supplementary_files_format_and_content: Tab-delimited files of loci, fraction of methylated cytosines and total read count.
Supplementary_files_format_and_content: RawReadCounts.txt: Raw transcript counts
Supplementary_files_format_and_content: TrancriptFPKMCounts.txt: Transcript FPKM counts
Supplementary_files_format_and_content: Old_H3K4me1.peaks.txt: peak
Supplementary_files_format_and_content: Old_H3K27me3.peaks.txt: peak
Supplementary_files_format_and_content: Young_H3K4me1.peaks.txt: peak
Supplementary_files_format_and_content: Young_H3K27me3.peaks.txt: peak
Supplementary_files_format_and_content: Young_H3K27Ac.peaks.txt: peak
Supplementary_files_format_and_content: OldBeta_bs_seeker-CG.tab: Percent methylated cytosines
Supplementary_files_format_and_content: YoungBeta_bs_seeker-CG.tab: Percent methylated cytosines

source_name: Pancreatic Beta Cells

extract_protocol:
RNA-Seq: Flow cytometry sorting was performed on live β cells from isolated pancreatic islets of young and old MIP-GFP mice. Total RNA were isolated using the RNeasy kit (Qiagen). ChIP-Seq: ChIP was performed on sonicated chromatin obtained from sorted old and young mouse β cells using antibody against H3K27ac (39685, Active Motif), H3K4me1 (ab8895, Abcam) and H3K27me3 (07-449, upstate). Input and bound DNA fractions were purified using PCR purification kit (Qiagen) for library preparation. BIS-Seq: Genomic DNA was extracted from FACS sorted old and young beta cells using FFPE AllPrep kit (Ambion), sonicated and bisulfite converted by Qiagen Epitect Bisulfite Kit and used for preparation of libraries.
RNA-Seq:Libraries were constructed from 200ng of total RNA using the standard TruSeq RNA sample prep kit (Illumina) using manufacturer's protocols. ChIP-Seq: Libraries were prepared using NEBNext ChIP-seq library prep reagent kit according to Illumina's instructions. BIS-Seq: Libraries were prepared following manufacturer's instructions using NEBNext ChIP-seq library prep reagent kit with methylated adapters (NEB).

tag: age

value: 4-6 weeks

tag: antibody


title: IUGR fetus  in twins 2


title: normal fetus in twins 2


title: IUGR fetus  in twins 1


title: normal fetus in twins 1


title: P2_AICAR_2

city: Nijmegen

first: Rutger

last: Vogel

state: Gelderland

address: Geert Grooteplein 10

department: Pediatrics

country: Netherlands

institute: Radboud university medical center

data_processing:
Raw sequence data was filtered for reads that contain adaptors in their sequence and with more than 50% of low quality base calls (quality value <= 5).
After filtering the reads were mapped to the reference using SOAP2 pipeline allowing for 1 or 2 mismatched bases.
Reads were subsequently mapped onto reference gene sequences to calculate RPKM (Reads Per Kb per Million mapped reads)
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each Sample

molecule: total RNA

source_name: Fibroblasts

growth_protocol: M199 medium with 10% fetal calf serum and penstrep

extract_protocol:
RNA was isolated using the Purelink RNA Mini Kit (Life Technologies). RNA was treated with DNAse and quality control was performed (OD260/280=1.8~2.2; OD260/230≥2.0; RIN≥7.0; 28S: 18S>1.0). Two 2 μg of RNA was analyzed using Illumina HiSeq2000.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: 125 uM chloramphenicol, 75 uM resveratrol, 500 uM AICAR, or 0.01% DMSO (vehicle), all for 72 hours and in duplicate

tag: cell line

value: fibroblast

tag: passages

tag: genotype

value: complex I deficient (patient ND5)

tag: agent

value: AICAR


title: P2_AICAR_1


title: P2_RES_2

value: resveratrol


title: P2_DMSO_1

value: DMSO


title: P1_RES_2

value: complex I deficient (patient NDUFS2)


title: ILK-KO (ILK-K5)-rep3

city: Münster

first: Sara

middle: A

last: Wickström

address: Röntgenstraße 20

department: Cell & Tissue Dynamics

country: Germany

institute: Max Planck Institute for Molecular Biomedicine

description:
TopHat alignment
KO2252

data_processing:
Illumina Casava1.9 software used for basecalling.
Sequenced reads were trimmed for adaptor sequences, ribosomal sequences were removed, then mapped to GRCm38_79 whole genome using bowtie TopHat 2.0.14 and Bowtie 2.2.4 with parameters --library-type fr-firststrand -p 20
DESeq2 (1.6.3) was used to compute the differential gene expressioon according to the manual. Adjusted p-values for 5,10,15, and 20% have been computed
Genome_build: m38
Supplementary_files_format_and_content: tab-delimited text files include gene expression values for differentially expressed genes

source_name: epidermis

growth_protocol: Bulge hair follicle stem cells were FACS purified as a alpha6 integrin high / CD34 + population from 21d old mouse epidermis. Cells were directly sorted into TrizolLS for RNA isolation

extract_protocol:
RNA was isolated according to manufacturers instructions for TrizolLS (Invitrogen), amplified cDNA was prepared using Ovation RNA-Seq System V2 (NuGen)
Libraries were prepared from amplified cDNA using Nextera XT library preparation kit (Illumina)

value: hair follicle bulge stem cell

value: post natal day 21

value: ILK -/-


title: P2_CAP_2

value: chloramphenicol


title: P1_RES_1


title: P1_DMSO_2


title: P1_DMSO_1


title: P2_DMSO_2


title: P1_AICAR_2


title: C2_RES_1

value: control


title: P1_AICAR_1


title: P2_RES_1


title: P1_CAP_2


title: C2_DMSO_1


title: C2_DMSO_2


title: C2_CAP_2


title: C2_CAP_1


title: C2_AICAR_2


title: C1_RES_2


title: P2_CAP_1


title: C1_DMSO_2


title: C1_DMSO_1


title: C1_CAP_2


title: C1_RES_1


title: C1_AICAR_2


title: C1_AICAR_1


title: Input DNA, E2, rep1, MCF7

city: Sherbrooke

first: Nicolas

last: Gevry

state: Québec

address: 2500 boulevard de l'université

department: Biologie

country: Canada

institute: Université de Sherbrooke

description:
CTCAGA
processed data file: Input_q10_nodup-rep1-2_MCF7_hg18_norm.bw

data_processing:
The alignment of sequencing reads on the human genome (Build 36.1, hg18, Mar. 2006) was done using the software Burrows-Wheeler Aligner (version 0.6.1, Li H et al., 2009) with default parameters. Above 96% of the reads per sample were mappable and ~80% had a Phred Mapping Quality Score above 10 (filtered using Samtools, version 0.1.18, Li H et al., 2009). The Picard command-line tools (version 1.64, http://picard.sourceforge.net) was then used to remove duplicates (80% of the filtered data were kept). Since the Pearson correlation coefficient, calculated in 2 kb bins containing the number of reads across the whole genome, was above 0.9 between each biological replicated, both replicates were combined for subsequent analyses.
The Wiggler tool (version 1.0, https://code.google.com/p/align2rawsignal/, Hoffman et al., 2013) was used to uniformly process and normalize the MNase ChIP-seq data using the default parameters except for the fragment length that was set to 147 bp (reads are shifted by 147/2 from 5' to 3'), the smoothing window that was set to 120 and the smoothing kernel that was set to triweight. This tool normalized the genome-wide signal at each position as a fold-change (the observed fragment count over the expected fragment count per position, i.e, if all the uniquely mappable reads are uniformly distributed over all uniquely mappable locations in the genome).
Genome_build: Build 36.1, hg18, Mar. 2006
Supplementary_files_format_and_content: The bigwig files represent the final normalized filtered and merged duplicates for each mark or H2A.Z generated with Wiggler tool (version 1.0, https://code.google.com/p/align2rawsignal/, Hoffman et al., 2013).

source_name: MCF7_Input DNA_E2

growth_protocol: The MCF-7 cells were grown in DMEM (Wisent) supplemented with 10% fetal bovine serum (FBS) and antibiotics. The cells were mycoplasma free (routinely tested).

extract_protocol:
MCF-7 cells were cross-linked 10 min at room temperature (1% formaldehyde in 1X PBS pH 7.4). ChIP assays were performed as described previously (Svotelis A et al., 2009), except for the chromatin preparation. Briefly, permeabilized cells were incubated 2 min at 37oC in digestion buffer (50 mM Tris-HCl pH 8.0, 1 mM CaCl2, 0.2% Triton and protease inhibitors), before the addition of Micrococcal Nuclease (MNase, USB Corporation) 5 min at 37oC. Then the digestion reactions were stopped (5 mM EDTA, 10 mM Tris-HCl pH 8 and 0.5% SDS and protease inhibitors) and chromatin sample were briefly sonicated to fragment insoluble chromatin from the pellet without affecting the fragmentation of mononucleosomal fragments. No visible pellet remained after a centrifugation at 15,000 g at 4oC for 10 min. The digestion and the full extraction of chromatin were verified on gel electrophoresis prior to the immunoprecipitation of the chromatin.
The sequencing libraries were performed according to the Illumina library preparation protocol except for the size-selection of DNA fragments and for DNA recovery that were executed as described previously (Rodrigue S et al., 2010). Quant-iT PicroGreen dsDNA Assay Kit (Invitrogen) was used to quantify ChIP-DNA and 10 ng was used for the preparation of the sequencing libraries. The size-selection of nucleosome fragments, to exclude subnucleosomal and polynucleosomal particules, was executed in parallel with the preparation of libraries and was performed using solid-phase paramagnetic beads technology (AMPure XP PCR Purification systems, Beckman Coulter), since it allows a more precise size-selection of the DNA fragments than conventional gel electrophoresis and minimizes loss of sample, which is particularly relevant for the dilute samples frequently obtained after ChIP experiments. It was thus used for all the DNA-recovery steps of the libraries preparation as well as to eliminate potential primer dimers or primers and linkers in excess (shorter than the libraries of nucleosome fragments) after the final amplification of the libraries. Importantly, this final step was monitored by qPCR and stopped in the exponential-phase to avoid PCR-duplicates. The quantification of the final libraries, the absence of primer-dimers as well as the validity of the size-selection were determined using the 2100 Bioanalyzer instrument (Agilent Technologies). To ensure reproducibility, two biological replicates of every mark or H2A.Z or control input were single-end sequenced with 40 nucleotides reads using an Illumina HiSeq 2000 platform by the MIT MicroBioCenter.

treatment_protocol: Before experiments, the MCF-7 cells were hormone-deprived for 3 days in DMEM without phenol red (Wisent) supplemented with 5% charcoal-stripped FBS. The cells were treated with 100 nM of 17-b-estradiol (Sigma) or with the vehicle (EtOH) for 30 min.

value: human breast adenocarcinoma

tag: treated with

value: 100 nM of 17-b-estradiol for 30min


title: Input DNA, E2, rep2, MCF7

description:
CTTAAG
processed data file: Input_q10_nodup-rep1-2_MCF7_hg18_norm.bw


title: C1_CAP_1


title: H3K27ac, E2, rep1, MCF7

description:
TAGTCG
processed data file: H3K27ac_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw

tag: chip antibody

tag: chip antibody vendor

value: Abcam

tag: chip antibody cat. #

tag: chip antibody lot #


title: H3K27ac, E2, rep2, MCF7

description:
TCAATC
processed data file: H3K27ac_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: C2_RES_2


title: H2A.Z, E2, rep2, MCF7

description:
GTCCTA
processed data file: H2A.Z_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: Input DNA, EtOH, rep2, MCF7

description:
CTGATC
processed data file: Input_q10_nodup-rep1-2_MCF7_hg18_norm.bw

source_name: MCF7_Input DNA_EtOH

value: vehicle (EtOH) for 30 min


title: H2A.Z, E2, rep1, MCF7

description:
GTGCAC
processed data file: H2A.Z_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: H3K4me3, E2, rep1, MCF7

description:
TTAGTT
processed data file: H3K4me3_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw

value: Active Motif


title: H3K4me1, E2, rep2, MCF7

description:
GATGCT
processed data file: H3K4me1_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: C2_AICAR_1


title: Input DNA, EtOH, rep1, MCF7

description:
CTAACT
processed data file: Input_q10_nodup-rep1-2_MCF7_hg18_norm.bw


title: H3K27ac, EtOH, rep2, MCF7

description:
GTTCCG
processed data file: H3K27ac_EtOH_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: H3K4me3, EtOH, rep2, MCF7

description:
GCATGC
processed data file: H3K4me3_EtOH_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: H3K4me3, EtOH, rep1, MCF7

description:
TCCAAG
processed data file: H3K4me3_EtOH_q10_nodup_rep1-2__MCF7_hg18_norm.bw


title: H3K4me1, EtOH, rep2, MCF7

description:
GAAGGA
processed data file: H3K4me1_EtOH_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: H2A.Z, EtOH, rep2, MCF7

description:
GTACGT
processed data file: H2A.Z_EtOH_q10_nodup_rep1-2__MCF7_hg18_norm.bw


title: H2A.Z, EtOH, rep1, MCF7

description:
GGTACC
processed data file: H2A.Z_EtOH_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: HCSC_Hep3B-C_mRNA

city: Shanghai

first: Min

last: Ding

address: Changhai Road

institute: Eastern Hepatobiliary Surgery Hospital

data_processing:
High-quality reads were aligned to the human reference genome (NCBI Build 36.1) using NextGENe® software (Softgenetics, State College, PA, USA).
The matched reads were aligned to Human Refseq mRNA (NCBI).
Reads shorter than 20 bps and those with the quality score less than 14 were excluded.
The sequences aligned with individual transcript were counted digitally.
Genome_build: NCBI Build 36.1
Supplementary_files_format_and_content: tab-delimited text files include read counts for each Sample

source_name: hepatic cancer stem cells

extract_protocol: The sequencing library was prepared with the standard Illumina protocol.


title: HCSC_Huh7-C_mRNA


title: HCC_Hep3B_mRNA

source_name: hepatoma cell line


title: HCC_Huh7_mRNA


title: H3K4me1, E2, rep1, MCF7

description:
GCTTCA
processed data file: H3K4me1_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw


title: GABPA-FLAG ChIP-Seq in Modified mouse ES cells

city: Huntsville

first: Eric

last: Mendenhall

state: Alabama

address: 301 Sparkman Dr

department: Biological Sceinces

country: USA

institute: University of Alabama Huntsville

description: Rep 1

data_processing:
Imaging and basecalls from Illumina RTA
ChIP-Seq Fastq files were aligned to hg19 or mm9 using Bowtie. RNA-seq FASTQ files were aligned using Tophat v1.4.1
ChIP-Seq bedgraph files were generated usign Bedtools. RNA-seq transcripts assembled using Cufflinks v1.3.0 with Gencode 9
ChIP-seq Peaks were called with MACS usign mfold 15, tsize of 50, p-vaule of 1e-10. For RNA-seq RPKM and raw counts processed using Cufflinks
Genome_build: hg19, mm9
Supplementary_files_format_and_content: bedgraph files contains locations and range of scores. RPKM.txt files contains for normalized values for gene models

source_name: mouse V6.5 ES Cells

growth_protocol: HepG2 and Mcf7 (ATCC HB-8065) were grown according to the approved ENCODE cell culture protocols (http://hgwdev.cse.ucsc.edu/ENCODE/protocols/cell)

extract_protocol: Cross-linked chromatin was immunoprecipitated with an antibody. The Protein:DNA crosslinks were then reversed and the DNA fragments were recovered and sequenced. Please see protocol notes below and go to http://hudsonalpha.org/myers-lab/protocols for the most current version of the protocol. Biological replicates from each experiment were completed. For RNA-seq polyA RNA was isolated using oligo dT beads, and libraries prepared.

treatment_protocol: HepG2  cells were transfected at ~75% confluence using FUGENE reagent (Promega, E2311), selected using G418 (Invitrogen, 10131035) and expanded.

value: Embryonic Stem Cell

value: FLAG M2 Sigma F1804


title: RNASeq_WT_P18_rep2

city: Bethesda

first: Sumin

last: Oh

state: MD

address: 9000 Rockville Pkie

department: NIDDK

institute: NIH

data_processing:
Basecalls performed using the CASAVA software
RNA-seq reads were aligned to the mm9 genome assembly using the STAR program (Dobin et al., Bioinformatics, 2012)
processed data files format and content: RNA-Seq: bigWig files can be used to check RNA-seq peaks through the UCSC genome browser.
Genome_build: mm9

source_name: Mammary gland

growth_protocol: We mated Ezh2f/f mice (Ezhkova et al., 2011; Wang et al., 2010) with the control transgenic mouse line A (Wagner et al., 1997) in order to generate mice with mammary-specific deletion of Ezh2. Ezh1 knockout mice were obtained from Rockefeller University (Ezhkova et al., 2011). Stat5 f/f (Cui et al., 2004) and MMTV-creA mice were used to generate mice lacking STAT5 in mammary stem cells (S5KO). To generate the mice lacking both EZH2 and STAT5 (E2S5 DKO), we bred Ezh2f/fStat5 f/f females with Ezh2f/fStat5f/f;MMTV-cre males. Genotyping was confirmed by PCR using DNAs from ear clips. All animals were handled and housed in accordance with the guidelines of the National Institutes of Diabetes, Digestive and Kidney Diseses Animal Care and Use Committee.

extract_protocol:
RNA-seq: Poly(A) RNA were purified twice with Trizol and RNeasy® Plus Mini Kit (Qiagen). cDNA was synthesized using SuperScript II (Invitrogen) and TruSeq RNA Sample Preparation Kit (Illumina).
Libraries were prepared for sequencing using standard Illumina protocols.

value: Mixed C57BL/6 x FVB/N

tag: tissue

value: Mammary gland

tag: state

value: pregnancy day of 18

value: wild type


title: HCC_Hep3B_miRNA

data_processing:
After cleaning away some contaminant reads, removal of redundancy and reads smaller than 18nt, the clean reads were mapped to the human genome (NCBI Build 36.1) by the program SOAP.
Reads were mapped to Ribosomal RNA (rRNA), Small cytoplasmic RNA (scRNA), Small nuclear RNA (snRNA), Small nucleolar RNA (snoRNA) and Transfer RNA (tRNA) from GeneBank. Then the remaining reads were searched against the miRBase (release 17).
Genome_build: NCBI Build 36.1
Supplementary_files_format_and_content: tab-delimited text files include read counts for each Sample

source_name: Hep3B cell

value: hepatoma cell line


title: RNASeq_WT_P18_rep3


title: HCC_Huh7_miRNA

source_name: Huh7 cell


title: P1_CAP_1

value: fibroblast

value: complex I deficient (patient NDUFS2)


title: RNASeq_WT_P18_rep1


title: RNASeq_S5KO_P13_rep3

value: pregnancy day of 13

value: Stat5f/f; MMTV-cre


title: RNASeq_WT_P13_rep3


title: RNASeq_WT_P13_rep2


title: RNASeq_E2S5DKO_P13_rep3

value: Ezh2f/f,Stat5f/f; MMTV-cre


title: RNASeq_S5KO_P13_rep1


title: RNASeq_E2S5DKO_P13_rep2


title: H3K4me1, EtOH, rep1, MCF7

description:
GAGGAG
processed data file: H3K4me1_EtOH_q10_nodup_rep1-2_MCF7_hg18_norm.bw

value: human breast adenocarcinoma


title: RNASeq_S5KO_P13_rep2


title: H3K4me3, E2, rep2, MCF7

description:
GCCTTG
processed data file: H3K4me3_E2_q10_nodup_rep1-2_MCF7_hg18_norm.bw

value: 100 nM of 17-b-estradiol for 30min


title: RNASeq_E2KO_P13_rep3

value: Ezh2f/f; MMTV-cre


title: RNASeq_E2S5DKO_P13_rep1


title: RNASeq_E2KO_P13_rep2


title: RNASeq_E1KO_P13_rep1


title: RNASeq_E2KO_P13_rep1


title: RNASeq_E1KO_P13_rep2


title: ChIPSeq_WT_STAT5_rep1

data_processing:
Basecalls performed using the CASAVA software
ChIP-seq reads were aligned to the mm9 genome assembly using the Bowtie2 program (Langmead et al., Nature method, 2012)
processed data files format and content: ChIP-seq: BedGraph files can be used to check ChIP-seq peaks through the UCSC genome browser.
Genome_build: mm9

growth_protocol: We mated Ezh2f/f mice (Ezhkova et al., 2011; Wang et al., 2010) with the control transgenic mouse line A (Wagner et al., 1997) in order to generate mice with mammary-specific deletion of Ezh2. Ezh1 knockout mice were obtained from Rockefeller University (Ezhkova et al., 2011). Genotyping was confirmed by PCR using DNAs from ear clips. All animals were handled and housed in accordance with the guidelines of the National Institutes of Diabetes, Digestive and Kidney Diseses Animal Care and Use Committee.

extract_protocol:
Frozen-stored mammary tissues from p13 were broken into powder with mortar and pestle. Nuclei were extracted in Farnham lysis buffer (5mM PIPES PH8.0, 85mM KCl, 0.5% NP-40 and protease inhibitor). Antibodies against EZH1 (gift from Sartorelli, Mousavi et al. Mol. Cell 2012), EZH2 (Diagenode, cat# C15410039 and lot# 003), STAT5 (Santa Cruz Biotechnology, cat# sc-1081; L-20, lot# L2012, 10ug) H3K27me3 (Millipore, 07-449) and PolII (Abcam, cat# ab5408, lot# GR106949-6) were used for ChIP.
Libraries were prepared for sequencing using standard Illumina protocols.

value: STAT5A (Santa Cruz, sc-1081)


title: ChIPSeq_WT_STAT5_rep2


title: HCSC_Hep3B-C_miRNA

source_name: Hep3B stem-like cell

value: hepatic cancer stem cells


title: ChIPSeq_WT_PolII_rep2

value: Pol II (abcam, ab5408)


title: RNASeq_WT_P13_rep1


title: RNASeq_E1KO_P13_rep3


title: ChIPSeq_WT_PolII_rep1


title: ChIPSeq_WT_EZH2_rep2

value: EZH2 (Diagenode, C15410039)


title: ChIPSeq_WT_EZH1_rep2

value: Mixed C57BL/6 x FVB/N

value: EZH1 (a generous gift from Dr. Sartorelli)


title: ChIPSeq_WT_EZH2_rep1

value: Mammary gland


title: ChIPSeq_WT_H3K27me3_rep1

value: wild type

value: H3K4me3 (Millipore, CS200580)


title: ChIPSeq_H3K27me3_input


title: ChIPSeq_STAT5_input


title: ChIPSeq_WT_EZH1_rep1


title: ChIPSeq_E2KO_STAT5_rep1


title: ChIPSeq_E2KO_STAT5_rep2


title: aub_sh_ago3_sh_germ_GFP_Piwi_GFP_IPseq_ox_R2

city: Vienna

first: Kirsten-André

last: Senti

state: Wien

address: Dr. Bohrgasse 3

country: Austria

institute: IMBA

description: IP small RNA

data_processing:
genome_mapping_read_counts.txt: table indicating read counts for all genome mapping small RNA sequences in the indicated libraries
small RNA-seq and CAP-seq libraries: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0), PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15, fastq files were adapter 'AGATCGGAAGAGCACACGTCT' clipped using cutadapt, version 1.0, default settings, after adapter removal, the first and last 4 bases were trimmed (both linkers contain 4 random nucleotides at their respective termini), reads were mapped with bowtie v0.12.7 to D. mel. genome (dm3, release 5) and BigWig files and 1kb bin overlaps were generated with Samtools v 0.1.16 and BedTools v2.16.1.
RNA-seq libraries: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0), PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15, reads were trimmed to high-quality bases 5-45, N containing reads were discarded. Mapping to the genome was done allowing up to 3MM. For RPKM calculations, files containing 2nd reads were reverse complemented and combined with the 1st reads and then aligned to the genome with bowtie v0.12.7 to D. mel. genome (dm3, release 5). For visualization,  BigWig files  were generated with BedTools v2.16.1.
ChIP-seq: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0). PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15. Reads were trimmed to bases 5-45, N-containing reads  were discarded. Reads were mapped with bowtie v0.12.7 to D. mel. genome (dm3, release 5) allowing up to 3MM and BigWig files were generated with BedTools v2.16.1.
Genome_build: Drosophila melanogaster dm3, release 5

organism: Drosophila melanogaster

source_name: adult ovary

growth_protocol: Flies were kept at strandard conditions at 25º C.

extract_protocol:
small RNAs from beads were isolated by acid phenol extraction and Ethanol precipitation, further cloning of small RNAs was done as for total small RNA-seq
small RNAs were PAGE selected and 3' and 5' linkers was ligated. 1st strand cDNA synthesis was performed using SuperScript III (Invitrogen) and the library was amplified using the Kapa HiFi Library Amp real time Kit (Peqlab) with Illumina TruSeq compatible primers.
IP from ovary lysate/acid Phenol RNA extraction from beads/ sizeselected small RNA cloning

tag: developmental stage

value: 4-6 days old adult females

value: ovary

tag: germline knockdown and other transgenes

value: aubsh1 attP33  ago3 sh attP2 nanos-GFP-Piwi attP40

tag: chip or ip antibody

value: Memorial Sloan-Kettering Monoclonal AntibodyFacility; clone names: Htz-GFP-19F7 and Htz-GFP-19C8, 1:1 ratio, bioreactor supernatant purity


title: ChIPSeq_E2KO_PolII_rep1

tag: strain

value: pregnancy day of 13


title: ago3_sh_germ_GFP_Piwi_GFP_IPseq_nonox_R2

description:
IP small RNA
small RNAs were oxidized as in Vagin et al. , Sience, 2006

value: ago3 sh attP2 nanos-GFP-Piwi attP40


title: control_sh_germ_GFP_Piwi_GFP_IPseq_ox_R2

value: white sh attP2 (control) nanos-GFP-Piwi attP40


title: aub_sh_ago3_sh_germ_GFP_Piwi_GFP_IPseq_nonox_R2


title: ChIPSeq_E2KO_PolII_rep2

value: Ezh2f/f; MMTV-cre


title: ChIPSeq_E2KO_EZH2_rep2


title: ChIPSeq_E2KO_H3K27me3_rep1


title: HCSC_Huh7-C_miRNA

source_name: Huh7 stem_like cell

tag: cell type


title: ChIPSeq_E2KO_EZH2_rep1


title: control_sh_germ_GFP_Piwi_GFP_IPseq_nonox_R2


title: ago3_sh_germ_GFP_Piwi_GFP_IPseq_nonox_R1


title: ChIPSeq_E2KO_EZH1_rep2


title: aub_sh_ago3_sh_germ_GFP_Piwi_GFP_IPseq_nonox_R1


title: aub_sh_germ_GFP_Piwi_GFP_IPseq_nonox_R2

value: aub sh attP2 nanos-GFP-Piwi attP40


title: aub_sh_germ_GFP_Piwi_GFP_IPseq_nonox_R1


title: ChIPSeq_E2KO_EZH1_rep1


title: GFP-Piwi_BAC_w1118_GFP_IPseq

value: GFP-Piwi BAC transgene / w1118


title: ChIPSeq_EZH_input


title: ChIPSeq_PolII_input


title: ChIPSeq_E1KO_EZH1_rep1


title: aub_sh_Ago3_IPseq

value: aub sh1 attP2


title: ago3_sh_Piwi_IPseq

value: ago3 sh attP2

value: MAB Piwi (8C2-E4)


title: cd150plus_B9

city: Barcelona

first: Lars

last: Velten

address: C. Dr. Aiguader 88, P05

department: Bioinformatics and Genomics

country: Spain

institute: CRG

description: Sample 20

data_processing:
Demultiplexing and trimming of overhanging T
Trim off 8 bases of the reverse read (molecular barcode) and store with first read. Discard reverse read. Perl5
Trim off polyA from 3' end of forward read. Discard reads not ending in a poly A tail. Python2.7/HTSeq
Align reads to Mus Musculus genome GRCm38, to which ERCC spike in seqeunce has been appended. Gsnap
Filter out reads of non-unique alignment, low alignemt quality, alignment to A-rich genomic regions, A-rich reads. Python2.7/HTSeq
Count unique molecular barcodes aligning to different genomic features. Python2.7/HTSeq
Remove barcode counts stemming from similar molecular barcodes with a Hamming distance of less than 1. Perl5
Collapse putative polyadenylation sites stemming from genomic positions within 12bp distance. Perl5
For the summary files, 27 cells were removed because less than 1000 molecular barcodes were observed. R-3.0.2
Genome_build: Mus Musculus GRCm38 (ensembl version 38.73), with the sequences of the ERCC control RNA spike ins appended to it
Supplementary_files_format_and_content: .counts, tab-separated file of ENSEMBL gene IDs and read counts; readcounts.csv is a merged version of all *.counts.gz files.

extract_protocol:
Cells were sorted into 0.4µL lysis buffer (1%NP-40) containing a 1:1.000.000 dilution of ERCC spike-ins
QUARTZ-Seq (Sasagawa et al., Genome Biology 2013)

tag: strain background

value: Bone marrow

value: Hematopoietic stem and progenitor

tag: surface marker


title: aub_sh_Piwi_IPseq


title: cd150plus_B3

description: Sample 18


title: cd150plus_B4

description: Sample 19


title: piwi_sh_Piwi_IPseq

value: piwi sh2 attP2


title: cd150plus_B11

description: Sample 16


title: piwi_sh_Aub_IPseq

value: MAB Aub (8A3-E7)


title: cd150plus_A8

description: Sample 15


title: control_sh_Ago3_IPseq

value: 4-6 days old adult females

value: white sh attP2 (control)


title: control_sh_Aub_IPseq

value: ovary


title: piwi_sh_Ago3_IPseq


title: cd150plus_B12

description: Sample 17


title: cd150plus_A11

description: Sample 13


title: cd150plus_A3

description: Sample 14


title: cd150plus_A10

description: Sample 12


title: aub_sh_ago3_sh_totsRNAseq

description:
total small RNA
small RNAs were oxidized as in Vagin et al. , Sience, 2006

data_processing:
small RNA-seq and CAP-seq libraries: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0), PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15, fastq files were adapter 'AGATCGGAAGAGCACACGTCT' clipped using cutadapt, version 1.0, default settings, after adapter removal, the first and last 4 bases were trimmed (both linkers contain 4 random nucleotides at their respective termini), reads were mapped with bowtie v0.12.7 to D. mel. genome (dm3, release 5) and BigWig files and 1kb bin overlaps were generated with Samtools v 0.1.16 and BedTools v2.16.1.
RNA-seq libraries: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0), PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15, reads were trimmed to high-quality bases 5-45, N containing reads were discarded. Mapping to the genome was done allowing up to 3MM. For RPKM calculations, files containing 2nd reads were reverse complemented and combined with the 1st reads and then aligned to the genome with bowtie v0.12.7 to D. mel. genome (dm3, release 5). For visualization,  BigWig files  were generated with BedTools v2.16.1.
ChIP-seq: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0). PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15. Reads were trimmed to bases 5-45, N-containing reads  were discarded. Reads were mapped with bowtie v0.12.7 to D. mel. genome (dm3, release 5) allowing up to 3MM and BigWig files were generated with BedTools v2.16.1.
Genome_build: Drosophila melanogaster dm3, release 5

extract_protocol:
smallRNA cloning was performed according to Brennecke et al, Cell, 2007
small RNAs were PAGE selected and 3' and 5' linkers was ligated. 1st strand cDNA synthesis was performed using SuperScript III (Invitrogen) and the library was amplified using the Kapa HiFi Library Amp real time Kit (Peqlab) with Illumina TruSeq compatible primers.
sizeselected 18-29nt smallRNA cloning

value: aubsh1 attp40 ago3 sh attP2


title: cd150minus_D9

description: Sample 11


title: control_sh_Piwi_IPseq


title: ago3_sh_totsRNAseq

extract_protocol:
smallRNA cloning was performed according to Brennecke et al, Cell, 2007
total RNA was isolated with Trizol. small RNAs were PAGE selected and 3' and 5' linkers was ligated. 1st strand cDNA synthesis was performed using SuperScript III (Invitrogen) and the library was amplified using the Kapa HiFi Library Amp real time Kit (Peqlab) with Illumina TruSeq compatible primers.
sizeselected 18-29nt smallRNA cloning


title: cd150minus_D5

description: Sample 9


title: ago3_sh_Aub_IPseq


title: cd150minus_D8

description: Sample 10


title: cd150minus_D4

description: Sample 8


title: cd150minus_D3

description: Sample 7


title: piwi_sh_CAPseq

description: total RNA

data_processing:
Library strategy: CAP-Seq
small RNA-seq and CAP-seq libraries: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0), PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15, fastq files were adapter 'AGATCGGAAGAGCACACGTCT' clipped using cutadapt, version 1.0, default settings, after adapter removal, the first and last 4 bases were trimmed (both linkers contain 4 random nucleotides at their respective termini), reads were mapped with bowtie v0.12.7 to D. mel. genome (dm3, release 5) and BigWig files and 1kb bin overlaps were generated with Samtools v 0.1.16 and BedTools v2.16.1.
RNA-seq libraries: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0), PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15, reads were trimmed to high-quality bases 5-45, N containing reads were discarded. Mapping to the genome was done allowing up to 3MM. For RPKM calculations, files containing 2nd reads were reverse complemented and combined with the 1st reads and then aligned to the genome with bowtie v0.12.7 to D. mel. genome (dm3, release 5). For visualization,  BigWig files  were generated with BedTools v2.16.1.
ChIP-seq: basecalled with the instruments RTA (Run time analysis version 1.12.48.0): SCS (PN:RTA DS:Controlling software on instrument VN:1.13.48.0). PRINSEQ-lite version 0.20.2 was used to remove sequences that contained at least one ambiguous nucleotide (i.e. 'N') as well as low complexity sequences (mostly homopolymer and di/trinucleotide repeats) using the DUST algorithm with a threshold of 15. Reads were trimmed to bases 5-45, N-containing reads  were discarded. Reads were mapped with bowtie v0.12.7 to D. mel. genome (dm3, release 5) allowing up to 3MM and BigWig files were generated with BedTools v2.16.1.
Genome_build: Drosophila melanogaster dm3, release 5

extract_protocol:
Cap-seq was performed according to Nachaev et al, Science, 2010 and Gu et al, Cell, 2013
Purified Capped RNA fragments were ligated with 3' and 5' linkers. 1st strand cDNA synthesis was performed using SuperScript III (Invitrogen) and the library was amplified using the Kapa HiFi Library Amp real time Kit (Peqlab) with Illumina TruSeq compatible primers.


title: control_sh_RZ1_RNAseq

description: Ribozero (1x) depleted total RNA

extract_protocol:
RNA-seq was performed according to Zhang et al, Silence, 2012
Total RNA was extracted using Trizol and depleted for ribosomal RNA (Ribo-Zero Kit, Epicentre). For 2nd strand synthesis, dUTP was added to generate stranded libraries. UDG digested dsDNA was cloned using the NEBNext™ DNA Sample Prep Reagent Set 1 (NEB). The Kapa HiFi Library Amp Real-Time Kit (Peqlab) with Illumina TruSeq compatible primers was used to amplify the library.

value: control (GFP sh attP2)


title: aub_sh_pA_RNAseq

description: polyA bound mRNA from total RNA

extract_protocol:
RNA-seq was performed according to Zhang et al, Silence, 2012
Total RNA was extracted using Trizol and polA RNA was purified with oligo-dT beads. For 2nd strand synthesis, dUTP was added to generate stranded libraries using the NEBNext Ultra DNA Library Prep Kit for Illumina (NEB). The Kapa HiFi Library Amp Real-Time Kit (Peqlab) with Illumina TruSeq compatible primers was used to amplify the library.


title: cd150minus_D12

description: Sample 6


title: cd150minus_D11

description: Sample 5


title: aub_sh_ago3_sh_RNAseq

description: polyA bound mRNA from total RNA


title: ago3_sh_pA_RNAseq

description: polyA bound mRNA from total RNA


title: cd150minus_C9

description: Sample 4


title: piwi_sh_pA_RNAseq

description: polyA bound mRNA from total RNA


title: cd150minus_C12

description: Sample 2


title: cd150minus_C6

description: Sample 3


title: control_sh_pA_RNAseq

description: polyA bound mRNA from total RNA


title: aub_sh_ago3_sh_Pol2ChIP_R3

description: genomic DNA (CHIP)

extract_protocol:
ChIP-seq was performed according to Lee et al, Nat Protoc, 2006
ChIP-seq libraries were generated using the NEBNext™ DNA Sample Prep Reagent Set 1 for Illumina (NEB) and the Kapa HiFi Library Amp Real-Time Kit (Peqlab) for amplification.

value: rabbit anti-rbp3 (custom made)


title: cd150minus_C10

description: Sample 1


title: ago3_sh_Pol2ChIP_R1

description: genomic DNA (CHIP)


title: aub_sh_ago3_sh_Pol2ChIP_R2

description: genomic DNA (CHIP)


title: aub_sh_Pol2ChIP_R3

description: genomic DNA (CHIP)


title: piwi_sh_Pol2ChIP_R3

description: genomic DNA (CHIP)


title: aub_sh_Pol2ChIP_R1

description: genomic DNA (CHIP)


title: ago3_sh_Pol2ChIP_R3

description: genomic DNA (CHIP)


title: aub_sh_ago3_sh_Pol2ChIP_R1

description: genomic DNA (CHIP)


title: ago3_sh_Pol2ChIP_R2

description: genomic DNA (CHIP)


title: piwi_sh_Pol2ChIP_R2

description: genomic DNA (CHIP)


title: aub_sh_Pol2ChIP_R2

description: genomic DNA (CHIP)


title: piwi_sh_Pol2ChIP_R1

description: genomic DNA (CHIP)


title: control_sh_Pol2ChIP_R4

description: genomic DNA (CHIP)


title: control_sh_Pol2ChIP_R2

description: genomic DNA (CHIP)


title: control_sh_Pol2ChIP_R1

description: genomic DNA (CHIP)


title: control_sh_gDNAseq

description: genomic DNA

extract_protocol:
Genomic DNA was isolated from control sh ovaries, sheared to average 400bp, size selected to 200-600bp, cloned and sequenced (paired-end 100nts)
gDNA-seq library was generated using the NEBNext Ultra DNA Library Prep Kit for Illumina (NEB) and the Kapa HiFi Library Amp Real-Time Kit (Peqlab) for amplification.


title: RNA-Seq HepG2 RAD21

description: Rep 1

source_name: HepG2 cells

tag: cell line

value: Hepatocellular Carcinoma


title: RNA-Seq HepG2 ATF1

description: Rep 1


title: RNA-Seq HepG2 WT Bio Replicate 1

description: Rep 1


title: RNA-Seq HepG2 WT Bio Replicate 2

description: Rep 2


title: RAD21-FLAG ChIP-Seq Technical Rep2 in Modified MCF7 Cells

description: Rep 2

source_name: MCF7 cells

value: Breast adenocarcinoma

value: FLAG M2 Sigma F1804


title: RAD21-FLAG ChIP-Seq Technical Rep1 in Modified MCF7 Cells

description: Rep 1


title: RAD21 ChIP-Seq in Modified MCF7 Cells

description: Rep 1

value: RAD21, abcam, ab992


title: CREB1-FLAG Technical Rep1 in Modified HepG2 cells

description: Rep 1


title: CREB1-FLAG Biological Rep2 in Modified HepG2 cells

description: Rep 2


title: control_sh_Pol2ChIP_R3

description: genomic DNA (CHIP)


title: RAD21 ChIP-Seq Rep1 in MCF7 cells

description: Rep 1


title: ATF1-FLAG ChIP-Seq Technical Rep2 in Modified HepG2 cells

description: Rep 2


title: RAD21 ChIP-SeqRep2 in MCF7 Cells

description: Rep 2


title: CREB1 ChIP=Seq in Modified HepG2 Cells

description: Rep 1

value: CREB1 Santa Cruz Biotechnology, sc-240


title: RAD21-FLAG ChIP-Seq in Modified HepG2 cells Rep2

description: Rep 2


title: GABPA-FLAG ChIP-Seq Rep 1 in Modified HepG2 cells

description: Rep 1


title: GABPA-FLAG ChIP-Seq Rep 2 in Modified HepG2 cells

description: Rep 2


title: CREB1 ChIP-Seq Rep1 in HepG2 Cells

description: Rep 1


title: GABPA ChIP-Seq Rep 1 in HepG2 cells

description: Rep 1

value: GABPA, Santa Cruz Biotechnology, sc-28312


title: GABPA ChIP-Seq Rep 2 in HepG2 cells

description: Rep 2


title: RAD21-FLAG ChIP-Seq in Modified HepG2 cells Rep1

description: Rep 1


title: RAD21-FLAG ChIP-Seq Biological Rep2 in Modified MCF7 Cells

description: Rep 2


title: Rad21 ChIP-Seq Rep 1 in HepG2 cells

description: Rep 1


title: Rad21 ChIP-Seq Rep 2 in HepG2 cells

description: Rep 2


title: ATF1-FLAG ChIP-Seq Technical Rep1 in Modified HepG2 cells

description: Rep 1


title: CREB1-FLAG Technical Rep2 in Modified HepG2 cells

description: Rep 2


title: CREB1 ChIP-Seq Rep2 in HepG2 Cells

description: Rep 2


title: GABPA ChIP-Seq Rep 1 in Modified HepG2 cells

description: Rep 1


title: RAD21 ChIP-Seq in Modified HepG2 cells

description: Rep 1


title: PI 567690-Drought

city: Columbia

first: Trupti

last: Joshi

state: MO

address: 1201 Rollins St.

department: Computer Science

institute: University of Missouri

data_processing:
The initial base calling and quality filtering of the reads generated with the Illumina analysis pipeline (fastq format) were performed using a custom Perl script and the default parameters of the Illumina pipeline (http://www.illunina.com).
Additional filtering for bad-quality bases was performed using the FASTX-toolkit available in FastQC software package (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Good quality mRNA-seq reads were aligned to the Glycine max reference genome (Gmax1.1 version) and Phytozome v9.0 gene model release using Tophat (version 1.4.1)
Tophat was then run with default parameters and a reference GTF file using the –G option, and replicates of each condition/sample were mapped independently to improve alignment sensitivity and accuracy for further analysis.
The gene expression (FPKM) levels were estimated using Cufflinks software (version v2.1.1) while differential gene expression analysis was performed using Cuffdiff (version v2.1.1)

organism: Glycine max

source_name: Leaf tissue

growth_protocol: Genotypes were growm in a mixture of soil and sand (2:1) was used in 26.5 L pots (top and bottom diameter were 30 cm and 27 cm, respectively and 37 cm in height).

extract_protocol:
RNeasy Plant mini-kit (Qiagen, 214 CA, USA) with on-Column DNA digestion performed using RNase-Free DNase Set (Qiagen, CA, USA)
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: At the V5 stage the plants were imposed with water stress for 15 days.  The leaf tissues were collected for the transcriptome study from the stressed plants at R2 stage

tag: phenotype

value: Drought Tolerant

value: R2 stage-Leaf tissue

tag: treatment

value: Drought stressed for 15 days


title: Pana-Drought

value: Drought Susceptible


title: PI 567690-Control

value: Control condition


title: Pana-Control


title: wt1

city: Milan

first: Jose

middle: Manuel

last: Garcia-Manteiga

address: Via Olgettina 58

department: Center for Translational Genomics and Bioinformatics

country: Italy

institute: OSR - San Raffaele Scientific Institute

description: wild type PC12 cells

data_processing:
Alignment with TopHat (2.0.4)
HT-Seq 0.5.3
Differential Expression Analysis (DESeq 1.9.12)
Genome_build: rn4
Supplementary_files_format_and_content: txt table containing reads overlap onto gene names of GTF annotation file from Ensembl72 ftp by HT-Seq count

organism: Rattus norvegicus

source_name: wild type PC12 cells

extract_protocol:
The RNA, extracted with the RNeasy Mini Kit (Qiagen, Valencia, CA) from two, carefully washed clones of PC12, the wtPC12 and the hrPC12 previously referred to as PC12-27 [24,25,29-31], was analyzed with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA)
Libraries, prepared starting from 2 mg of RNA/sample with the Illumina TruSeq RNA Sample Prep kit v2 procedure, were quantified by the Qubit BR assay (Life Technologies, Illkirch, France) and the Agilent 2100 Bioanalyzer, and sequenced on the Illumina HiSeq 2000 platform

tag: rest levels

value: low


title: wtb

description: wild type PC12 cells


title: H3K27ac, EtOH, rep1, MCF7

description:
TAATTA
processed data file: H3K27ac_EtOH_q10_nodup_rep1-2_MCF7_hg18_norm.bw

value: human breast adenocarcinoma

tag: treated with

value: vehicle (EtOH) for 30 min

tag: chip antibody

value: Abcam


title: C43

status: Public on Aug 30 2015

city: Baltimore

first: Jaehoon

last: Shin

state: Maryland

address: 733 N Broadway

institute: Johns Hopkins University

description: single cell RNA

data_processing:
Trimommatic - adapter trimming
Custom R codes to further trim the cDNA amplification adapter sequence
RSEM (Cole Dewey et al.) to map and calculate the TPM
Genome_build: modified mm9 to include the Nestin-CFPnuc transgene
Supplementary_files_format_and_content: text file with TPM.

source_name: Dentate gyrus of hippocampus_single_cell

extract_protocol:
not provided
NUGEN Ultralow V2 protocol

value: brain

tag: tissue compartment

value: Dentate gyrus of hippocampus

tag: age


title: C49

description: single cell RNA

tag: tissue


title: N3

description: single cell RNA


title: N15

description: single cell RNA


title: N6

description: single cell RNA


title: C8

description: single cell RNA


title: C3

description: single cell RNA


title: C72

description: single cell RNA


title: N1

description: single cell RNA


title: N14

description: single cell RNA


title: C85

description: single cell RNA


title: C20

description: single cell RNA


title: C140

description: single cell RNA


title: N8

description: single cell RNA


title: N4

description: single cell RNA


title: C139

description: single cell RNA


title: C77

description: single cell RNA


title: C113

description: single cell RNA


title: 27b

description: high REST PC12 cells

source_name: high REST PC12 cells

value: high


title: C10

description: single cell RNA


title: C132

description: single cell RNA


title: C31

description: single cell RNA


title: C29

description: single cell RNA


title: 27a

description: high REST PC12 cells


title: C127

description: single cell RNA


title: C57

description: single cell RNA


title: C75

description: single cell RNA


title: C129

description: single cell RNA


title: C37

description: single cell RNA


title: C124

description: single cell RNA


title: C123

description: single cell RNA


title: C19

description: single cell RNA


title: C13

description: single cell RNA


title: C86

description: single cell RNA


title: C101

description: single cell RNA


title: C60

description: single cell RNA


title: C17

description: single cell RNA


title: C1

description: single cell RNA


title: C4

description: single cell RNA


title: C112

description: single cell RNA


title: C71

description: single cell RNA


title: C28

description: single cell RNA


title: N7

description: single cell RNA


title: C131

description: single cell RNA


title: C67

description: single cell RNA


title: C38

description: single cell RNA


title: C126

description: single cell RNA


title: C12

description: single cell RNA


title: C125

description: single cell RNA


title: N9

description: single cell RNA


title: C21

description: single cell RNA


title: C54

description: single cell RNA


title: N10

description: single cell RNA


title: N23

description: single cell RNA


title: C103

description: single cell RNA


title: C42

description: single cell RNA


title: C64

description: single cell RNA


title: C11

description: single cell RNA


title: C9

description: single cell RNA


title: C15

description: single cell RNA


title: C53

description: single cell RNA


title: C76

description: single cell RNA


title: C26

description: single cell RNA


title: C5

description: single cell RNA


title: C32

description: single cell RNA


title: CFDAC2

city: Guangzhou

first: Zhenhui

last: Li

address: Tianhe District Wushan road

institute: South China Agricultural University

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to gallus gallus whole genome using TopHat.
To estimate the expression of each gene, the total number of reads mapped to its unigenes were calculated and normalized with FPKM (fragments per kilobase of transcript per million mapped reads).
Genome_build: CanFam3.1

organism: Canis lupus familiaris

source_name: adrenal cortex

extract_protocol:
Total RNA was extracted from each spleen using Trizol (Invitrogen, CA, USA) according to manufacturer’s instructions.Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 10 ug of total RNA for the construction of sequencing libraries.
mRNA libraries were prepared for sequencing using standard Illumina protocols

tag: breed

value: Chinese Field Dog

value: adrenal cortex

tag: group

value: Treated with non-disposal

value: adult one


title: C92

description: single cell RNA


title: BAC1

value: Beagle

value: Treated with chronic stress exposure


title: C59

description: single cell RNA


title: C138

description: single cell RNA


title: BAC2


title: BP2

source_name: pituitary

value: pituitary


title: C35

description: single cell RNA


title: C108

description: single cell RNA


title: BP1


title: C100

description: single cell RNA


title: N12

description: single cell RNA


title: N20

description: single cell RNA


title: R3

description: single cell RNA


title: N18

description: single cell RNA


title: C16

description: single cell RNA


title: C44

description: single cell RNA


title: C98

description: single cell RNA


title: CFDAC1


title: C52

description: single cell RNA


title: CFDP2


title: C80

description: single cell RNA


title: C56

description: single cell RNA


title: C22

description: single cell RNA


title: C81

description: single cell RNA


title: C14

description: single cell RNA


title: CFDP1


title: C45

description: single cell RNA


title: N25

description: single cell RNA


title: N19

description: single cell RNA


title: N5

description: single cell RNA


title: N11

description: single cell RNA


title: C24

description: single cell RNA


title: C63

description: single cell RNA


title: C99

description: single cell RNA


title: C83

description: single cell RNA


title: N22

description: single cell RNA


title: N17

description: single cell RNA


title: N13

description: single cell RNA


title: C47

description: single cell RNA


title: C70

description: single cell RNA


title: N16

description: single cell RNA


title: C118

description: single cell RNA


title: C74

description: single cell RNA


title: C40

description: single cell RNA


title: C88

description: single cell RNA


title: C142

description: single cell RNA


title: C111

description: single cell RNA


title: C2

description: single cell RNA


title: C89

description: single cell RNA


title: C33

description: single cell RNA


title: C66

description: single cell RNA


title: N2

description: single cell RNA


title: C104

description: single cell RNA


title: C69

description: single cell RNA


title: C27_2

description: single cell RNA


title: C27_1

description: single cell RNA


title: C36

description: single cell RNA


title: C110

description: single cell RNA


title: C18

description: single cell RNA


title: C6

description: single cell RNA


title: C61

description: single cell RNA


title: C7

description: single cell RNA


title: C84

description: single cell RNA


title: C79

description: single cell RNA


title: C137

description: single cell RNA


title: C25

description: single cell RNA


title: C128

description: single cell RNA


title: C133

description: single cell RNA


title: C117

description: single cell RNA


title: R1

description: single cell RNA


title: R4

description: single cell RNA


title: C30

description: single cell RNA


title: C105

description: single cell RNA


title: N26

description: single cell RNA


title: C115

description: single cell RNA


title: C107

description: single cell RNA


title: C73

description: single cell RNA


title: C34

description: single cell RNA


title: C39

description: single cell RNA


title: C121

description: single cell RNA


title: C122

description: single cell RNA


title: C78

description: single cell RNA


title: N24

description: single cell RNA


title: C106

description: single cell RNA


title: C120

description: single cell RNA


title: C135

description: single cell RNA


title: C48

description: single cell RNA


title: C62

description: single cell RNA


title: C87

description: single cell RNA


title: R2

description: single cell RNA


title: CFDP1

data_processing:
Illumina Casava1.7 software used for basecalling.
The raw reads obtained by high-sequencing were performed with quality control to result in clean reads. In brief, the quality control was that of trimming low-quality reads (ambiguous N and length < 18 nt), 3′ adapters, 5′ adapters and poly (A) sequences. Thereby, the lengths of 18-32 bp reads, as the clean ones, were obtained and calculated for their length distributions.
Meanwhile, the identical reads were collapsed to obtain the unique reads for the statistics of the small RNA’s species and abundance. Moreover, the clean reads were mapped to Rfam database (ftp://sanger.ac.uk/pub/databases/Rfam/) and GenBank noncoding RNA database (http://blast.ncbi.nlm.nih.gov/ ) to annotate the miscellaneous RNAs, thus filtering out the rRNA, scRNA, snoRNA, snRNA, tRNA and other non-coding RNA. The remaining sRNAs obtained above, including known miRNAs and unknown miRNAs, were then mapped to the reference genome to analyze their distributions on genome and calculate their expressive quantity.
In addition, the remaining sRNAs were mapped to the Canis Canine miRNA data of miRBase 20.0 to identify the known miRNAs. Meanwhile, the novel miRNAs were predicted using miRDeep2 (http://www.mdcberlin.de/en/research/research_teams/systems_biology_of_gene_regulatory_elements/projects/miRDeep/index.html) from unannotated sRNAs.
Genome_build: CanFam3.1

extract_protocol:
Total RNA was extracted from each spleen using Trizol (Invitrogen, CA, USA) according to manufacturer’s instructions.Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 10 ug of total RNA for the construction of sequencing libraries.
Small RNA libraries were prepared for sequencing using standard Illumina protocols


title: N21

description: single cell RNA


title: CFDP2


title: BAC1


title: C65

description: single cell RNA


title: C90

description: single cell RNA


title: C23

description: single cell RNA


title: C50

description: single cell RNA


title: CFDAC1


title: CFDAC2


title: C114

description: single cell RNA


title: C96

description: single cell RNA


title: C119

description: single cell RNA


title: BP1


title: BAC2


title: BP2


title: C109

description: single cell RNA


title: C58

description: single cell RNA


title: dct2-1_minus4-rep3

city: Saint Louis

first: Sarit

last: Weissmann

address: 975 North Warson Rd

institute: Donald Danforth Plant Science Center

description: processed data file: minus4-diff.txt

data_processing:
GSNAP was used to align reads to the maize genome
CuffDiff was used to identify differences in gene expression between dct2-1 and WT plants
Genome_build: B73 AGPv2
Supplementary_files_format_and_content: tab-delimited text files containing FPKM and log2(dct2 FPKM/WT FPKM) - CuffDiff output. Sample_1 = WT; Sample_2 = dct2.

organism: Zea mays

source_name: Leaf

growth_protocol: Plants were grown in a growth chamber with 16h light per day, with supplemental light of 500 umol m-2 s-1, 28⁰C, and 40% humidity.

extract_protocol:
Samples were pooled from 15 plants per biological replicate. Leaf segments were frozen in N2.  RNA was extracted using TriPure reagent (Roche) according to the manufacturer protocol.
RNA-seq library were constructed according to the protocol described by Wang et al 2014 Nature Biotechnology

value: leaf

tag: leaf segment

value: 9 days

tag: genotype


title: dct2-1_minus1-rep3

description: processed data file: minus1-diff.txt

tag: strain


title: dct2-1_minus4-rep1

description: processed data file: minus4-diff.txt


title: C93

description: single cell RNA


title: dct2-1_plus10-rep1

description: processed data file: plus10-diff.txt


title: dct2-1_plus4-rep3

description: processed data file: plus4-diff.txt


title: dct2-1_plus10-rep3

description: processed data file: plus10-diff.txt


title: WT_plus10-rep2

description: processed data file: plus10-diff.txt

value: wildtype


title: WT_plus10-rep3

description: processed data file: plus10-diff.txt


title: C116

description: single cell RNA


title: dct2-1_plus4-rep1

description: processed data file: plus4-diff.txt


title: C46

description: single cell RNA


title: C55

description: single cell RNA


title: C94

description: single cell RNA


title: dct2-1_minus1-rep1

description: processed data file: minus1-diff.txt


title: dct2-1_plus4-rep2

description: processed data file: plus4-diff.txt


title: WT_plus4-rep1

description: processed data file: plus4-diff.txt


title: dct2-1_minus1-rep2

description: processed data file: minus1-diff.txt


title: dct2-1_minus4-rep2

description: processed data file: minus4-diff.txt


title: C41

description: single cell RNA


title: WT_plus10-rep1

description: processed data file: plus10-diff.txt


title: WT_minus4-rep3

description: processed data file: minus4-diff.txt


title: WT_plus4-rep3

description: processed data file: plus4-diff.txt


title: C97

description: single cell RNA


title: WT_plus4-rep2

description: processed data file: plus4-diff.txt


title: C141

description: single cell RNA


title: WT_minus1-rep3

description: processed data file: minus1-diff.txt


title: C82

description: single cell RNA


title: WT_minus1-rep2

description: processed data file: minus1-diff.txt


title: C136

description: single cell RNA


title: C134

description: single cell RNA


title: C91

description: single cell RNA


title: C68

description: single cell RNA


title: HK30

city: Dublin

first: Tim

last: Downing

state: Other

address: DCU

department: Biotechnology

country: Ireland

institute: DCU

description:
pre-menopausal healthy from USA
Small RNAs
mirna.2.each.counts.txt, mirna.2.mix.counts.txt, miscRNA.2.each.counts.txt, miscRNA.2.mix.counts.txt

data_processing:
[Read QC] The single-end 32 base RNA sequence reads generated on were processed using the Illumina Casava pipeline tool (v1.8.3). Quality assessment was based on data passing the Illumina Chastity filtering. Reads containing adaptors or PhiX control signal were removed. Quality assessment was based on the remaining reads using FASTQC v0.10.0 to ensure base quality did not decline significantly across the read length. Small RNA 3' adapter (Illumina TruSeq) sequences and low quality bases were removed from the reads using the CLC Genomics Workbench v5.1.5. Reads smaller than 18 bases and those longer than 25 bases after trimming were excluded.
[Read annotation] Unique sequence groups were determined for each sample and these RNA sets were annotated as mature RNAs (IsomiRs) allowing for up to a two base variation in length up- or down-stream and up to one mismatch. The expression level of each small RNA was annotated using either miRBase Homo sapiens, or miRBase Mus musculus, or Ensembl Homo sapiens GRCh37 ncRNA database releases 57 and 67.
[Read expression analysis] Differential RNA expression was evaluated in transcripts with a minimum of one count per million in at least three samples using three normalization methods: reads per kilobase per million mapped reads (RPKM), trimmed mean of M-values (TMM) and quantile normalization. Firstly, the fraction of reads mapping to small RNA or other small RNA transcripts for each sample was normalized within each sample as RPKM. Reads mapping to multiple RNAs were associated with those RNAs. Secondly, count data for each sample was normalized using quantile normalization. Thirdly, the TMM value between sample pairs as a normalizing factor reduced technical variation signals between small RNAs by compensating for differences in read levels between samples. These were computed using R v2.15.0 Bioconductor package EdgeR v3.2.4, which applied a negative binomial distribution model to identify differentially expressed miRNAs. Reads absent in one group but present in the other group required at least 10 reads in total to avoid sequencing artifacts. Multi-dimensional scaling (MDS) of the RPKM, quantile and TMM normalized levels within and between groups did not identify any samples as individual outliers.
Genome_build: Homo sapiens GRCh37
Genome_build: Mus musculus GRCm37
Supplementary_files_format_and_content: The files labelled "mirna" contain small RNAs that could be annotated using the human and mouse genomes; the "miscRNA" files contain the remainder. The "each" files contain each RNA isoforms for each RNA type; and the "mix" files contain RNA isoforms aggregated by annotation type (ie by miRNA here). Each file contains a header with the RNA/isoform and the sample names (HK...). Each subsequent row shows the RNA/isoform (for the "each" files this includes the actual RNA sequence), the length, the RNA type (if know, or else the species used for annotation), and then the observed RNA counts for each sample. The ENST label is included as well for the "miscRNA" files (ie ENST00000482497 etc).

source_name: Peripheral blood

extract_protocol:
[RNA Isolation] All whole blood samples for analysis in this study were collected in PAXgene™ blood tubes (Qiagen, Hilden, Germany). Total RNA, including small RNAs and miRNAs, were  isolated and purified from 2.5ml of whole blood using the PAXgene blood miRNA kit (Qiagen, Hilden, Germany) per the manufacturer’s instructions
[RNA Purification] Total purified RNA integrity was analyzed by capillary electrophoresis using a Bioanalyzer 2100 (Agilent Technologies, Santa Clara, California) and concentration and purity were measured by UV absorbance at 260nm using a NanoDrop 2000 (Thermo Scientific, Waltham, Massachusetts, USA). Isolated total RNA was stored at -80°C prior to use.
Small RNA library preparation and sequencing was carried out externally by BaseClear (Leiden, Holland) - small RNAs of lengths below 30 bases were enriched by polyacrylamide gel size selection purification. Illumina HiSeq adaptors were then ligated to the 5’ and 3’ ends of the small RNAs. These were amplified for 17 PCR cycles using primers for the adaptors. Post-amplification products were run on an agarose gel and the fragments of 90-110 bp, corresponding to the small RNA plus the adaptors, were isolated [39]. The purified retrieved PCR products were assessed for quality and quantity using the Agilent 2100 bioanalyzer (Agilent Technologies).
the library was sequenced on an Illumina HiSeq 2000 platform

tag: disease state

value: pre-menopausal healthy

tag: geographical location

value: USA

value: peripheral blood


title: HK31

description:
post-menopausal healthy from USA
Small RNAs
mirna.2.each.counts.txt, mirna.2.mix.counts.txt, miscRNA.2.each.counts.txt, miscRNA.2.mix.counts.txt

value: post-menopausal healthy


title: WT_minus1-rep1

description: processed data file: minus1-diff.txt


title: WT_minus4-rep2

description: processed data file: minus4-diff.txt


title: C102

description: single cell RNA

value: brain


title: C130

description: single cell RNA

value: Dentate gyrus of hippocampus


title: C95

description: single cell RNA


title: HK32


title: HK29


title: C51

description: single cell RNA


title: HK20

description:
Small RNAs
mirna.2.each.counts.txt, mirna.2.mix.counts.txt, miscRNA.2.each.counts.txt, miscRNA.2.mix.counts.txt

value: Breast cancer


title: HK21


title: HK28


title: dct2-1_plus10-rep2

description: processed data file: plus10-diff.txt


title: HK22


title: HK14


title: HK26


title: HK17


title: HK27


title: HK18


title: HK15


title: HK25


title: HK16


title: HK11


title: HK12


title: HK03

description:
pre-menopausal healthy from Ireland
Small RNAs
mirna.2.each.counts.txt, mirna.2.mix.counts.txt, miscRNA.2.each.counts.txt, miscRNA.2.mix.counts.txt

value: Ireland


title: HK24


title: WT_minus4-rep1

description: processed data file: minus4-diff.txt


title: HK01


title: HK19


title: HK13


title: HK6

value: Testing data


title: HK2


title: HK06


title: HK05


title: HK23


title: HK02


title: Delto-Smu-15-2

status: Public on Aug 28 2015

city: Quebec City

first: Adam

last: Foster

state: Quebec

address: 8678 rue de la Comtoise

department: CFS

institute: Natural Resources Canada

data_processing:
Illumina Casava1.8 software used for basecalling.
Quality checking: base call quality , consequtive homopolymer, Kmer frequency, base call frequency, overrepresented sequence and basic statisitics was performed in RobiNA - transcriptomics data preprocessor version 1.2.1
Adapter trimining, sliding window trimmer, read length filter to remove reads shorter than 20 bp for all samples. RobiNA - transcriptomics data preprocessor version 1.2.1
Read mapping was performed using Bowtie v1 to reference genome allowing 2 mismatches in a 28 bp seed legnth in RobiNA - transcriptomics data preprocessor version 1.2.1.
Gene expression analysis step - edgeR with BH corrected p-value cutoff 0.05, auto dispersion and min Log-fold change of 1 within RobiNA - transcriptomics data preprocessor version 1.2.1
Unassembled reads were pooled for samples of the same species and de novo assembled using CLC Genomics Workbench version 7 with default settings. Contigs less than 500 bp in legnth were discarded.
Read mapping was performed using Bowtie v1 to reference genome allowing 2 mismatches in a 28 bp seed legnth in RobiNA - transcriptomics data preprocessor version 1.2.1.
Gene expression analysis step - edgeR with BH corrected p-value cutoff 0.05, auto dispersion and min Log-fold change of 1 for samples mapped to lists of contigs
Genome_build: P. trichocarpa V3.0 210
Supplementary_files_format_and_content: Read count files were generated using Bowtie v1 in RobiNA 1.2.1 mapping to either the P. trichocarpa V3 genome of a list of de novo assembled contigs; Digital gene expression analysis files were prouced by edgeR in RobiNA 1.2.1; de novo asssembled contigs (fasta) for each poplar species were produced using CLC Genomics Workbench  version 7 de novo assembler with reads that did not map to the P. trichocarpa v3 genome.

organism: Populus deltoides

source_name: Fully expanded leaf

growth_protocol: Trees were propgated in the greenhouse in 3 L pots with supplemented light at 16 h daylengths for 5 months. The five month old trees were transferred to Conviron growth chambers set at 22°C and 16h light with a photosynthetic flux of 90-100 µmol m-2 s-1 and incubated for 4 weeks for a final age of 6 months.

extract_protocol:
RNA was also isolated from 100 mg of liquid nitrogen chilled homogenized leaf tissue using the RNeasy Plant Mini Kit (Qiagen) with in column RNase-Free DNase Set (Qiagen) .
RNA was extracted from each tree sampled and assayed for quality with a 2100 Bioanalyzer and Nanodrop 1000 spectophotometer. All RNA samples had a ratio of the absorbance at 260 and 280nm (A260/280) above 2.0 and were confirmed to be free of genomic DNA. Libraries for each RNA sample were prepared by Brian Boyle at ‘L'Institut de biologie intégrative et des systèmes’ (IBIS) at Université Laval, Quebec City, Quebec, Canada. Each cDNA library was barcoded and six samples were pooled together. TruSeq RNA Sample Preparation Kit v2 was used to prepare DNA for Illumina sequencing

treatment_protocol: One monosporal Sphaerulina isolate from each species was utilized. Isolates were prepared from spore suspensions stored in 25% glycerol at -80ºC.  To obtain conidia for foliar application, storage cultures were seeded to potato dextrose agar (PDA; Difco 213400) plates and grown at 22ºC in the dark for 8 days. Conidia were harvested by flooding plates (20 per isolate) with sterile distilled water, then filtered through 100-µl nylon mesh sterile Falcon® cell strainers (Fisher Scientific, Waltham, MA, USA), centrifuged at 4000 x g for 10 minutes, washed with 40 ml of sterile distilled water and resuspended in sterile distilled water to 1x105 per mL prior to application. Using an airbrush, each tree was sprayed with 10 mL of atomized spore suspension. Three trees from each species were sprayed with water to serve as controls.

tag: inoculation

value: Sphaerulina musiva conidia

tag: sample time

value: 15 days after inoculation


title: HK8


title: Delto-Smu-4-2

value: 4 days after inoculation


title: Delto-Smu-4-3


title: Delto-Smu-15-1


title: HK7


title: HK1


title: HK3


title: HK5


title: Delto-mock-3

value: mock-water

value: 0 days after inoculation


title: Delto-mock-1


title: HKpop

value: Pooled data for testing


title: Delto-Smu-15-3


title: Delto-Smu-1-2

value: 1 day after inoculation


title: Delto-Smu-4-1


title: Balsam-Spop-15-2

organism: Populus balsamifera

value: Sphaerulina populicola conidia


title: Balsam-Spop-4-1


title: Delto-Smu-1-3


title: Balsam-Spop-1-3


title: Balsam-Spop-1-2


title: Delto-Smu-1-1


title: Balsam-Spop-15-3


title: Balsam-Spop-15-1


title: HK10

value: pre-menopausal healthy


title: DLD1.exo.1

city: Nashville

first: Yongchao

last: Dou

state: Tennessee

address: 2525 west end

institute: vanderbilt medical center

description: miRNA

data_processing:
Illumina software used for basecalling.
reads were mapped and annotaed by ncpro-seq
DESeq was used to do the differential analysis
Genome_build: UCSC hg19
Supplementary_files_format_and_content: Tab-delimited plain text file linked to Series record: the first column is MicroRNA gene IDs and followings are reads counts

source_name: CRC cell line

growth_protocol: Cells cultured in DMEM containing 10% FBS until 80% confluency. Media replaced with serum-free DMEM and cultured for 24 hrs.

extract_protocol:
Exosomes isolation: conditioned media centrifuged for 10 min at 300 X g, filtered through a 0.22um polyethersulfone filter, concentrated ~300-fold with a 100,000 MW cutoff centrifugal concentrator, high-speed centrifugation at 150,000 x g for 2 hours. RNA isolation:Cells grown serum-free for 24hrs (see growth protocol) or the isolated exosomes resuspended in PBS were subjected to RNA isolation through Trizol (Life Technologies)-chloroform extraction, followed by second RNA purification using RNA easy kit (Qiagen).
Library prep was done  using NEB small RNA low input protocol using 6 ul sample input. Each sample was individually barcoded with NEB index primers for illumina. PCR amplification was done for 15 cycles and size selection was done using Ampure XP beads protocol as per manufactures instruction. .  Accurate quantification for sequencing applications was done using the qPCR-based KAPA Biosystems Library Quantification kit. Each library were diluted to a final concentration of 12.5nM and pooled equimolar prior to clustering.  Cluster generation will be carried out on a cBot v1.4.36.0 using Illumina's Truseq Single Read (SR) Cluster Kit v3.0. Sequencing was performed to generate approximately 15 million single end (SE) reads per sample on an Illumina HiSeq2500. Image analysis and base calling was performed using the standard Illumina Pipeline consisting of Real time Analysis (RTA) version v1.13 and demultiplexed using bcl2fastq converter with default settings.

treatment_protocol: The medium was collected and replaced with ionomycin-containing media for 1 hr, after which ionomycin-containing media was collected and pooled with the previously collected serum-free medium

tag: cell line

tag: kras status

value: both wild type and mutant (G13D) KRAS alleles

tag: miRNA source

value: exosome


title: Balsam-Spop-4-3


title: Delto-mock-2


title: DLD1.exo.3

description: miRNA


title: DLD1.exo.2

description: miRNA


title: DLD1.cell.2

description: miRNA

value: cell


title: DLD1.cell.3

description: miRNA


title: HK4

value: peripheral blood


title: Balsam-Spop-1-1


title: Balsam-mock-3


title: Balsam-mock-2


title: DLD1.cell.1

description: miRNA


title: Balsam-Spop-4-2


title: Balsam-mock-1


title: DKS8.exo.2

description: miRNA

value: a wild type KRAS allele


title: DKS8.cell.2

description: miRNA


title: DKS8.exo.3

description: miRNA


title: DKS8.exo.1

description: miRNA


title: DKS8.cell.1

description: miRNA


title: DKO1.exo.2

description: miRNA

value: a mutant KRAS allele


title: DKO1.exo.1

description: miRNA


title: DKO1.exo.3

description: miRNA


title: DKS8.cell.3

description: miRNA


title: DKO1.cell.2

description: miRNA


title: DKO1.cell.3

description: miRNA


title: DKO1.cell.1

description: miRNA


title: HNF4a8_IP_DOX_TCF4_rep2

city: Elk Grove

first: joseph

middle: M

last: dhahbi

state: ca

address: 9700 West Taron Drive

department: Medical Education

institute: California Northstate University College of Medicine

description: processed data file: HNF4a8_DOX_IP_TCF4_FE.bw

data_processing:
Bowtie v1.8 was used to pre-process and map sequence reads to the human reference genome (GRCh37/hg19).
MACS was used to call significant peaks by comparing signal tracks from ChIP against input. Default parameters were used in the MACS analysis.
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: bigwig files; these are fold-enrichment bedGraph files generated by MACS2 ‘bdgcmp’ function which deducts noise by comparing two signal tracks.

source_name: HNF4α8 Line

growth_protocol: The Tet-On inducible HCT116 cell lines (PL, α2, and α8) were maintained in McCoy's 5A media (Corning Cellgro; cat# 10-050-CV) supplemented with 10% Fetal Bovine Serum (BenchMark; cat# 100-106, lot# A07D00C) and 1% penicillin-streptomycin.

extract_protocol:
Formaldehyde cross-linking and chromatin immunoprecipiation was performed as described in Hwang-Verslues et al., 2008 (Mol Endo 22:78-90) and Li AG et al., 2007 (Mol. Cell 28:408-421) with some modification.
Illumina-compatible NEXTflex ChIPseq Kit (Bioo Scientific) was used to generate libraries from purified ChIP DNA for sequencing in the Illumina's HiSeq2000. Illumina HiSeq2000 was used to sequence the libraries (50bp, single-end)

treatment_protocol: α2 and α8 lines were treated with 0.0 or 0.3 μg/mL of DOX for 24 hours.

tag: cell type

value: colon cancer

tag: passages

tag: treatment

value: 0.3 μg/mL DOX

tag: length of treatment

value: 24 hours

tag: antibody

value: anti-TCF7L2 (TCF4) (Millipore; cat# 6H5-3)


title: HNF4a8_IP_TCF4_rep2

description: processed data file: HNF4a8_IP_TCF4_FE.bw

value: 0.0 μg/mL DOX


title: HNF4a8_IP_TCF4_rep1


title: HNF4a8_input

value: none (input)


title: HNF4a2_IP_DOX_TCF4_rep2

description: processed data file: HNF4a2_DOX_IP_TCF4_FE.bw

source_name: HNF4α2 Line


title: HNF4a2_IP_DOX_a445_rep2

description: processed data file: HNF4a2_DOX_IP_a445_FE.bw

value: affinity-purified anti-HNF4α (α445)


title: HNF4a8_IP_DOX_TCF4_rep1


title: HNF4a2_IP_TCF4_rep2

description: processed data file: HNF4a2_IP_TCF4_FE.bw


title: HNF4a8_IP_DOX_a445_rep2

description: processed data file: HNF4a8_DOX_IP_a445_FE.bw


title: HNF4a8_IP_DOX_a445_rep1


title: HNF4a2_IP_DOX_TCF4_rep1


title: HNF4a2_input


title: HNF4a2_IP_DOX_a445_rep1


title: α8_0.3_rep3_RNAseq

data_processing:
Paired-end sequencing reads aligned to human reference genome (GRCh37/hg19) with Tophat v1.2, using default parameters but allowing 10 alignments.
Cufflinks package used to assemble transcripts and measure relative abundance in FPKM.
Assembled transcripts were compared with RefSeq from UCSC genome browser.
Differential expression was examined using the Cuffcompare and Cuffdiff in the Cufflinks package.
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample.These are the differential expression files generated by cuffdiff.

extract_protocol:
Total RNA was extracted using miRNeasy kit (Qiagen). Sequencing libraries were constructed using the iIllumina TruSeq RNA Sample Prep Kit v2.
RNA libraries were prepared following the manufactor's standard protocol from Illumina.   Illumina HiSeq2000 was used to sequence the libraries (50bp, paired-end)

treatment_protocol: PL and α2 lines were treated with 0.0 or 0.3 μg/mL of DOX and α8 was treated with 0.0, 0.1, or 0.3 μg/mL DOX for 24 hours.

value: 0.3 μg/mL Doxycycline


title: α8_0.3_rep1_RNAseq


title: α8_0.3_rep2_RNAseq


title: HNF4a2_IP_DOX_a445_rep2.1


title: α8_0.1_rep3_RNAseq

value: 0.1 μg/mL Doxycycline


title: α2_0.3_rep2_RNAseq


title: α8_0.0_rep3_RNAseq

value: 0.0 μg/mL Doxycycline


title: α8_0.0_rep1_RNAseq


title: HNF4a2_IP_TCF4_rep1


title: α8_0.0_rep2_RNAseq


title: α8_0.1_rep1_RNAseq


title: PL_0.3_rep3_RNAseq

source_name: Parental Line


title: α2_0.0_rep1_RNAseq


title: α2_0.0_rep3_RNAseq


title: PL_0.3_rep2_RNAseq


title: α2_0.0_rep2_RNAseq


title: PL_0.0_rep3_RNAseq


title: α2_0.3_rep1_RNAseq


title: PL_0.3_rep1_RNAseq


title: PL_0.0_rep1_RNAseq


title: α8_0.1_rep2_RNAseq


title: α2_0.3_rep3_RNAseq


title: 110_1

city: New York

first: Yanwen

last: Jiang

state: NY

institute: Weill Cornell Medical College

data_processing:
FASTQ files were generated by using the Illumina CASAVA 1.8.2 pipeline.
Read counts were derived from HTSeq.scripts.count module in HTSeq-0.6.0 with default parameters.
Peak calling were performed by ChIPseeqer-2.1 with default parameters.
Genome_build: mm9
Supplementary_files_format_and_content: reads count and peaks files are in .txt format

growth_protocol: Isolated primary cells were harvested directly from mouse.  No growth culture.

extract_protocol:
Chromatin was sheared to 200-400bp, incubated with H3K4me1/2 antibody (abcam 32356) overnight at 4C on a rocker, antibody chromatin complex was captured by ProteinA agarose beads, washed with low salt, high salt, LiCl, and TE buffer, eluted and then de-crosslinked at 65C with 0.3M NaCl final for at least 6 hr.  Final chormatin was purified using Qiagen PCR purification columns.  RNA was purified using the RNAeasy Plus Kit (QIAGEN) that included a genomic DNA elimination step. DNA was purified using PCR purification columns (QIAGEN).
RNA-seq libraries were generated using Illumina’s TruSeq RNA sample Prep Kit v2. ChIP-seq libraries were prepared using 10ng of DNA and Illumina’s TruSeq ChIP sample prep. Libraries were validated using the Agilent Technologies 2100 Bioanalyzer and Quant-iT dsDNA HS Assay (Life Technologies).

treatment_protocol: B220+ cells were purified from mouse lymphoma tumors by immunomagnetic enrichment with CD45R(B220) microbeads (Miltenyi Biotech)

value: B cell

tag: transduction

tag: stain

value: tumor


title: 87_3

value: empty vector


title: 87_5


title: 110_2


title: 87_4


title: 103_5


title: AO103_5


title: AO103_2


title: AO87_2


title: AO103_1


title: AO87_1


title: AO78_5


title: jaz5/10 12h

city: Exeter

first: ZhengRong

last: Yang

address: Stocker Road

country: United Kingdom

institute: Exeter University

data_processing:
Base Calling carried out using CASAVA v1.8.2
Fastq-Mcf was then used to remove adapter sequences and trim reads of a phred quality score < 20 with the following parameters: -q 20 -l 35 -k 0  -p 15  --max-ns 0 -m 3. ERCC reads were removed using Bowtie v1.0.0 with the following parameters: -S -X 600
Reads were aligned to the reference genomes for Arabidopsis thaliana using TopHat v2.0.8b with the following parameters: -r 200 --mate-std-dev 375 --library-type fr-unstranded.
Reads were quantified at each predicted gene location using htseq-count v0.6.0 with the following parameters: -m union -s yes
These single replicate data were analysed by gFOLD (Feng et al., 2012) using a cutoff of log2 1.6
Supplementary_files_format_and_content: Tab delimited matrix containing raw counts, gene length and the normalised count for each gene in each sample.

organism: Arabidopsis thaliana

source_name: Infected Arabidopsis thaliana leaf tissue

growth_protocol: Arabidopsis Sowing And Plant Growth Conditions: Arabidopsis thaliana seed, Col-0 or jaz5/10, were sown onto ~7cm square pots of sieved compost mix (Levingtons F2 compost+sand (LEV206): vermiculite (medium grade) mixed in a 6:1 ratio) and stratified for 3 days at 4°C in darkness. Seeds were germinated in a controlled environment growth chamber with a 10 h photoperiod (lights on 8am, off 6pm) of 120 µmol m-2 s-  at 23oC (day), 20oC (night) and relative humidity of 65%. Plants were pricked into individual 7cm pots in a 24 cell and plants were grown for 4.5 weeks. Cells were continuously moved in the controlled environment chamber to reduce positional effects.  Pseudomonas growth: Pseudomonas syringae pv. tomato strain DC3000 carrying the empty broad host range vector pVSP61 (Innes et al., 1993) was grown on solidified Kings B media (King et al. 1954) containing rifampicin 50 μg ml-1 and kanamycin 25 μg ml-1. For inoculation, overnight cultures were grown with shaking (200 rpm) at 28oC. Cells were harvested (2000g X 8 min), washed and resuspended in 10 mM MgCl2. Cell density was adjusted to OD600 0.15 ( ~0.75 x 108 colony forming units (cfu) ml-1).

extract_protocol:
Plant RNA extraction: RNA was extracted according to de Torres et al. (2006). The frozen single leaf was ground to a fine powder in a liquid nitrogen pre-cooled mortar. 600µl of Z6 buffer (8M guanidinium hydrochloride, 20mM MES, 20mM EDTA pH7.0) was added and the powder quickly mixed. When thawed, the sample was passed to a 1.5ml microfuge tube and the mortar washed with 100µl of Z6 buffer. The combined sample was extracted with 1 vol. of phenol/chloroform then centrifuged at 4oC for 5 min at maximum speed. The aqueous phase was removed to a new microfuge tube and 1/20 vol. of acetic acid (1M) and 0.7 vol. 100% ethanol added nucleic acids collected after 30min on ice. The resultant RNA pellet was washed in 70% ethanol and resuspended in 100µl of sterile ddH2O. Samples were cleaned up using a Qiagen RNeasy Plant mini kit according to the manufacturers instructions and samples eluted in 30μl in RNAse free water.
2 µl of diluted ERCC RNA control spike-in mix 1 or control spike-in mix 2 (Life Technologies) was added to 1 µg of sample RNA and the volume adjusted to 50 µl.   Sequencing Libraries were prepared usingthe standard  Illumina TruSeq RNA library preparation protocol. on a Sciclone NGS platform. Samples were processed in 96 well hard shell PCR plates (Biorad HSP-9631). Washes were done in 150 µl bead washing solution instead of the 200 µl used in the manual process. Incubations at 65°C and 16°C for mRNA denaturation, and second-strand synthesis were performed on the Sciclone deck. However, the 80°C and 94°C incubations for mRNA elution and fragmentation, and first-strand cDNA synthesis reactions were performed on a thermocycler with heated lid to avoid excess evaporation. Purification and size selection was by Ampure XP bead clean up rather than gel based size selection. Library quality and quantitiy was determined by LabChipGX high sensitivity assay, after diluting the libraries 2µl in 20 µl.

treatment_protocol: Leaf infiltration: Plants were randomised and for each genotype and time-point 3 fully expanded leaves from four individual plants were either harvested (non-induced) or inoculated with DC3000. For infiltration, a small nick was made on either side of central vascular vein on the abaxial surface and bacterial challenge (OD600 of 0.15) was performed with a 1ml needleless syringe. All inoculations were completed by 11 am and inoculated plants were left under a light bench in the laboratory (22oC). Samples were taken at 6, 8, 12 and 16 hpi. Leaves were harvested by cutting at the petiole/leaf blade junction and were immediately snap-frozen in foil untill used for RNA preparation.

tag: ecotype

tag: genotype/variation

value: jaz5/10 mutant

tag: agent

value: Pseudomonas syringae pv. tomato DC3000


title: jaz5/10 16h


title: PL_0.0_rep2_RNAseq


title: jaz5/10 8h


title: AO110_3


title: AO110_2


title: B. juncea drought stressed (BDS) (UNPAIRED)

city: New Delhi

first: Gopal

state: Delhi

address: First Floor

department: Department of Botany

country: India

institute: Delhi University

data_processing:
Illumina Casava1.7 software used for basecalling.
RNA-Seq raw reads were processed by NGS-QC toolkit and low-quality as well as adapter-contaminated sequences were discarded. High quality (paired and unpaired) reads were assembled de-novo using SOAPde-novo assembler  independently at eight different K-mers (21, 27, 33, 39, 45, 51, 57, 63). The eight assemblies were subsequently clustered by using CD-HIT-EST [68]. The clustering parameters used were ≥80% query coverage and ≥80% identity. To further clean the data transcripts present in only one of the K-mer assemblies were removed. This was followed by removal of transcripts with less than 1 FPKM in all the three conditions (BC, BDS and BHS). Finally all the transcripts less than 200 bp were removed and the remaining transcripts were functionally annotated using FASTAnnotater tool (http://fastannotator.cgu.edu.tw/) with an e-value cut-off of 0.00001 by taking non-redundant protein database of EMBL (European Molecular Biology Laboratory) as a reference. Gene ontology analysis of transcripts was derived through Uniprot hit accessions and prediction of biochemical pathways was performed by KEGG identifiers (http://www.genome.jp/kegg/).
Supplementary_files_format_and_content: cufdiff_isoform_exp.diff: List of of deferentially expressed transcripts
Supplementary_files_format_and_content: matrix_isoforms.fpkm_tracking: Normalized FPKM values of each sample (Q1=BC, Q2=BDS, Q3=BHTS)

organism: Brassica juncea

source_name: 7-day old whole seedling

growth_protocol: Seeds were surface sterilized with 2% sodium hypochlorite solution for 10 minutes (min) on a shaker and then washed five times with double distilled water for three min each. Sterile seeds were hydroponically grown on a muslin cloth wrapped around a small container in a growth chamber at 24°C±1 with 16 hours (h) day/8 h night photoperiod.

extract_protocol:
Total RNA was isolated using GITC-based method from abiotic stress treated and untreated whole seedlings, independently for each time point. Extracted RNA was quantified using spectrophotometer and an aliquot of heat denatured RNA was electrophoresed on denaturing agarose gel to check its integrity. RNA extracted from two different time points were pooled in equimolar amounts and three RNA-Seq libraries- BC (control seedlings), BDS (drought stressed seedlings) and BHS (high temperature stressed) were prepared utilizing NEBNext RNA-Seq library preparation Master Mix Set for Illumina procured from NEB, USA.
Poly A+ RNA was isolated from 10 µg of total RNA using Sera-Mag beads (GE Healthcare, UK) and fragmented chemically at high temperature. Fragmented RNA was qualitatively and quantitatively checked on Bioanalyzer (Agilent, USA). 250 ng of fragmented RNA was used for first strand reverse transcription using random primers followed by second strand synthesis. The ends of double stranded cDNA were repaired and mono-adenylated. Paired end adapters were ligated using Rapid T4 DNA ligase and then size fractionated. Approximately, 350 bp size region was eluted and PCR amplified for 12 cycles. The quality and quantity of prepared libraries was evaluated utilizing Bioanalyzer (Agilent, USA) Ultra-deep parallel sequencing was performed using Illumina Genome Analyzer IIx at University of Delhi South Campus, Delhi, India according to manufacturer’s instructions.

treatment_protocol: Seedlings were grown for seven days and then subjected to various abiotic stresses. Drought stress was imposed for 3 h and 12 h by replacing water with high osmolality solution (300 mM mannitol). For imposing high temperature stress, seedlings were placed in a BOD incubator (Scientific systems, India) at 42°C for 30 min and 4 h. Entire seedlings (including the roots) were harvested after specified time intervals, snap frozen in liquid nitrogen and stored at -80°C.  Untreated seedlings were taken as control.

tag: strain

value: var varuna

tag: tissue

value: whole seedling

tag: age


title: ATF4 KO, HisOH

first: Stefan

last: Muljo

address: 10 CENTER DR RM 11N306

institute: NIH/NIAID

description: RNA samples from 3 independent KO clones were pooled.

data_processing:
Base-calling was done by Illumina RTA 1.9.35, at Phred+64 qualities.
RNA-Seq reads were aligned to the human genome (hg19) using Tophat version 2.0.11.
Read densities (in the bigWig tracks) were normalized to RPM with Bedtools version 2.17.0.
Genome_build: GRCh37 (hg19)
Supplementary_files_format_and_content: bigWig files were generated using Bedtools version 2.17.0. Peak height represents read density in reads per million.

source_name: HAP1, ATF4 KO, 2 mM histidinol for 24 hours

growth_protocol: HAP1 cells were re-plated the day before treatment at 5x10^5 cells/well in 6-well plate tissue culture-treated plates. Cells were cultured in IMDM supplemented with 10% fetal calf serum, GlutaMAX (Life Technologies), and Penicillin-Streptomycin (Life Technologies).

extract_protocol:
Total RNA was extracted from cells with RNAzol RT (SIGMA, R4533). For ATF4 KO samples, total RNA was pooled from 3 independent ATF4 KO cell lines.
RNA-Seq libraries were prepared using the NEBNext Ultra Directional RNA Library Prep Kit (New England BioLabs), including poly(A) selection using the NEBNext Poly(A) mRNA Magnetic Isolation module. Final libraries were analyzed with a Qubit fluorometer (Life Technologies) and Bioanalyzer 2100 (Agilent Technologies) to assess quantity and quality. Libraries were clustered at a density of 6.5 pM with a cluster station. Paired-end sequencing was performed using a GAIIx Genome Analyzer (Illumina).

treatment_protocol: For untreated cells, media was refreshed 24 hours prior to harvest. For cells treated with histidinol (HisOH), fresh media containing 2 mM histidinol (SIGMA, H6647) was added to cultures 24 hours prior to harvest.

value: ATF4 KO

value: 2 mM histidinol, 24 hours


title: jaz5/10 6h


title: jaz5/10 non induced

source_name: Arabidopsis thaliana leaf tissue


title: WT, HisOH

description: Not pooled.

source_name: HAP1, WT, 2 mM histidinol for 24 hours

value: WT


title: Col-0 12h

value: wild type


title: Col-0 8h


title: ATF4 KO, untreated

source_name: HAP1, ATF4 KO, untreated

value: untreated


title: Col-0 non induced


title: Col-0 6h


title: B. juncea drought stressed (BDS) (PAIRED-END)


title: WT, untreated

description: Not pooled.

source_name: HAP1, WT, untreated


title: B. juncea high temperature stressed (BHS) (UNPAIRED)