title: Pol2_KO_CT18_ChIPSeq

description: Chromatin IP against Pol2

source_name: Mll3∆/∆ MEF

value: Mll3∆/∆ MEF

value: Abcam

tag: antibody cat#


title: Mll3_KO_CT6_ChIPSeq

status: Public on Jan 01 2013

type: SRA

city: Philadelphia

first: Utham Kashyap

last: Valekunja

state: PA

address: 3400 Civic Center Boulevard,

department: Department of Systems Pharmacology & Therapeutics, Perelman School of Medicine,

country: USA

institute: University of Pennsylvania

description: Chromatin IP against Mll3

library_source: genomic

data_processing: Raw paired-end 50bp reads (in FASTQ format) were aligned to the UCSC mm9 genome build with Bowtie (http://bowtie-bio.sourceforge.net/index.shtml) using pre-built indexes downloaded from the host website. Parameters were optimised to utilise the software’s multi-threading capabilities (we typically used a 128-core cluster for alignments). Alignment files were then converted from the standard Bowtie output format to ALN files (for use with Cisgenome) using a custom Perl script. Alignments were then analysed using a 2-sample comparison (ChIPed sample vs. Input control for each time-point) with Cisgenome v1, using a cut-off false discovery rate of 10% and a sliding window size of 100. Output files were converted into various formats for browsing datasets and to display data. The Cisgenome and UCSC browsers were used for data visualisation, using custom tracks

molecule: genomic DNA

organism: Mus musculus

extract_protocol: Chromatin was sheared using a Diagenode Bioruptor at 4oC, using conditions optimised to produce 150-300bp fragments with the use of 15 ml Falcon tubes holding 2ml of lysate each (30 cycles of 30 secs on HIGH, 30 secs OFF). Sheared chromatin was transferred to 1.5 ml microcentrifuge tubes and spun at 14,000 rpm for 15 min at 4°C to pellet debris. Cleared supernatants were transferred to clean microcentrifuge tubes and 100 µl from each sample lysate was saved for use as an input control for sequencing and to check fragment size distribution. standard Illumina indexing primers were used, as per the manufacturer’s instructions. All libraries were multiplexed, and 4 indexed pooled samples (corresponding to four time-points, CT0, 6, 12, 18) sequenced per lane.

tag: strain

tag: tissue

tag: chip antibody

tag: antibody vendor

tag: time point


title: H3K4Me3_KO_CT18_ChIPSeq

description: Chromatin IP against H3K4Me3


title: Pol2_WT_CT18_ChIPSeq

description: Chromatin IP against Pol2

source_name: Wild Type MEF

value: Wild Type MEF


title: Pol2_WT_CT6_ChIPSeq

description: Chromatin IP against Pol2


title: Mll3_WT_CT18_ChIPSeq

description: Chromatin IP against Mll3


title: Mll3_WT_CT6_ChIPSeq

description: Chromatin IP against Mll3


title: H3K4Me3_WT_CT18_ChIPSeq

description: Chromatin IP against H3K4Me3


title: H3K4Me3_KO_CT6_ChIPSeq

description: Chromatin IP against H3K4Me3


title: Mll3_KO_CT18_ChIPSeq

description: Chromatin IP against Mll3


title: Mll3_CT18_ChIPSeq

description: Chromatin IP against Mll3

source_name: Liver tissue

value: Liver


title: H3K4Me3_WT_CT6_ChIPSeq

description: Chromatin IP against H3K4Me3


title: 511_RNA-polII_CHIP-seq

city: ILLKIRCH Cedex

first: Stephanie

last: Le Gras

address: 1, rue Laurent Fries

country: France

institute: IGBMC

data_processing:
Illumina Casava1.8 software used for basecalling.
Sequence reads were mapped to reference genome mm9/NCBI37 using Bowtie v0.12.7 with the following parameters -m 1 --strata --best.
Genome_build: mm9
Supplementary_files_format_and_content: Wig files were generated using with an in-house script (Variable step, span=25, reads were elongated to 200b)

source_name: liver

extract_protocol: ChIP-seq libraries were prepared using ChIP-Seq Sample Prep Kit (#IP-102-1001, Illumina) following the manufacturer's protocol with some modifications. Briefly, 10 ng of ChIP enriched DNA or control DNA were end repaired using T4 DNA polymerase, Klenow DNA polymerase and T4 PNK. A single ‘A’ nucleotide was added to the 3’ ends of the blunt DNA fragments with a Klenow fragment (3' to 5'exo minus). The ends of the DNA fragments were ligated to double stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR (30 sec at 98°C; [10 sec at 98°C, 30 sec at 65°C, 30 sec at 72°C] x 14 cycles; 5 min at 72°C) and then purified using Agencourt AMPure XP beads (#A63881, Beckman). DNA library size selection was performed by excising the 250-350 bp fragments from a 2% agarose gel followed by purification using a QIAquick Gel Extraction Kit (#28906, Qiagen).

treatment_protocol: Total liver from the indicated mouse genetic backgrounds was homogenized by douncing in buffer with 25mM Hepes pH=7,8, 1,5mM MgCL2, 10mM KCl, and 0,1% NP40. Nuclei were purified by centrifugation (4000 rpm, 5 min) and resuspended in fixation buffer (PBS1X, PIC PMSF and DTT, 1% PFA) and incubated 15 min at RT. Cold glycine was added to stop the reaction and nuclei were subsequently washed twice in PBS1X followed by centrifugation (4000 rpm, 5min). Nuclei were incubated 10min at 4°C in lysis buffer (EDTA 10mM, Tris HCl pH=8,0 50mM, SDS 1% +PIC +PMSF) and sonicated to average fragment size of 200-1000 nt. ChIP experiments were performed as previously described (Choukrallah et al., 2011).

value: Wild-type 129/SV mouse

tag: age

value: 5 Week old

value: Pol II :Santa-Cruz (Sc9001)


title: set1Δ/Δ Set3-9myc yeast ChIP Rep3

city: Vienna

first: Denes

last: Hnisz

address: Dr Bohr-gasse

department: Medical Biochemistry

country: Austria

institute: Max F Perutz Laboratories

description: Sample 19

data_processing:
Basecalls performed using CASAVA version 1.4
Multiplexing adaptor sequences were removed.
Reads were mapped onto the chromosomal Assembly 21 of the C. albicans genome with Bowtie, allowing only for uniquely mapping reads.
The RPKM values of all ORFs (using their default chromosomal coordinates) of Assembly 21 were calculated with Cufflinks (Trapnell C, Nat Biotechnology, 2010).
Genome_build: Assembly 21 of the C. albicans genome
Supplementary_files_format_and_content: txt file containing RPKM values for all ORFs in all samples

organism: Candida albicans

source_name: set3Δ set1Δ/Δ_Set3-9myc yeast ChIP

extract_protocol: Cultures were crosslinked by the addition of formaldehyde at a final concentration of 1% for 15 minutes at room temperature. Crosslinking was quenched by the addition of 125mM glycine for 5 minutes. Cells were washed twice with ice-cold TBS and pellets were frozen in liquid N2.  Pellets corresponding to 80 OD600 cells were resuspended in 480 μl ice-cold ChIP lysis buffer (50 mM HEPES/KOH pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% Na-deoxycholate, protease inhibitors). After addition of around 500μl glass beads (425–600 mm, Sigma), cells were broken at 6 m/s for 60 seconds 8 times on a FastPrep instrument (MP Biomedicals). Samples were cooled on ice for five minutes between cycles. The bottom of the tubes was punctured with a 27 3/4G needle and lysates were collected by centrifugation at 1500g for 1 min at 4C into a fresh tube. The lysates were diluted to 2.4 ml with ChIP lysis buffer. 300 μl aliquots were sonicated 15 times with 30s ON/30s OFF cycles at high setting on a Bioruptor (Diagenode). After sonication the aliquots were combined and centrifuged once at 14000g for 5 min at 4C. Supernatants were collected and used as input chromatin lysates. Set3-9myc and Hos2-9myc ChIPs were performed using EZ-View anti-myc coupled agarose beads (Sigma). After overnight incubation at 4C washing and DNA purification was performed exactly as described in (Hernday AD, Noble SM, Mitrovich QM, Johnson AD (2010) Methods Enzymol 470: 737-758.)

tag: genotype/variation

value: set3-delta set1-delta/delta

tag: morphological phase

value: yeast phase

value: anti-myc (EZ-View anti-c-myc Affininty Gel)

tag: chip antibody vendor

value: Sigma

tag: chip antibody cat. #


title: Hos2-9myc yeast ChIP Rep1

description: Sample 11

source_name: hos2Δ_Hos2-9myc yeast ChIP


title: Mll3_CT6_ChIPSeq

description: Chromatin IP against Mll3


title: Pol2_KO_CT6_ChIPSeq

description: Chromatin IP against Pol2


title: Hos2-9myc yeast ChIP Rep2

description: Sample 12


title: Hos2-9myc yeast input

description: Sample 10

source_name: hos2Δ_Hos2-9myc yeast input


title: Set3-9myc hypha ChIP Rep1

description: Sample 8

source_name: set3Δ_Set3-9myc hypha ChIP

value: hypha phase


title: Set3-9myc hypha ChIP Rep2

description: Sample 9


title: Mll3_CT0_ChIPSeq

description: Chromatin IP against Mll3


title: Set3-9myc hypha input

description: Sample 7

source_name: set3Δ_Set3-9myc hypha input


title: Set3-9myc yeast input

description: Sample 3

source_name: set3Δ_Set3-9myc yeast input


title: Set3-9myc yeast ChIP Rep1

description: Sample 4

source_name: set3Δ_Set3-9myc yeast ChIP


title: Set3-9myc yeast ChIP Rep2

description: Sample 5


title: Hos2-9myc hypha ChIP Rep1

description: Sample 14

source_name: hos2Δ_Hos2-9myc hypha ChIP


title: Hos2-9myc hypha ChIP Rep2

description: Sample 15


title: set1Δ/Δ Set3-9myc yeast ChIP Rep1

description: Sample 17


title: set1Δ/Δ Set3-9myc yeast input

description: Sample 16

source_name: set3Δ set1Δ/Δ_Set3-9myc yeast input


title: Set3-9myc yeast ChIP Rep3

description: Sample 6


title: untagged input

description: Sample 1

source_name: wild type_untagged input

value: wild type


title: Hos2-9myc hypha input

description: Sample 13

source_name: hos2Δ_Hos2-9myc hypha input


title: untagged ChIP

description: Sample 2

source_name: wild type_untagged ChIP


title: set1Δ/Δ Set3-9myc yeast ChIP Rep2

description: Sample 18


title: Mll3_CT12_ChIPSeq

description: Chromatin IP against Mll3


title: S-sample

city: Tai'an

first: Dong

last: fei

state: Shandong

address: No.61 Daizong street

country: China

institute: Shandong Agricultural University

library_source: transcriptomic

data_processing:
Illumina Casava1.7 software used for base-calling.
Sequenced reads were trimmed for adaptor sequence, and masked for low quality reads, reads without 3’ primer, reads with 5’ primer contaminants, reads without the inserted tag, reads with poly(A), reads shorter than 18 nt or low-quality sequence, then mapped to Ovis aries genome using SOAPv1.11 software
Genome_build: UCSC, 2010, ISGC Ovis aries 1.0/oviAri1

molecule: total RNA

organism: Capra hircus

source_name: peak lactation

extract_protocol: Mammary gland tissues were obtained by surgery and immediately frozen in liquid nitrogen. RNA was harvested using Trizol reagent. Illumina/Solexa RNA Sample Prep Kit (Cat#FC-122-1001) was used with 2 ug of total RNA for the construction of sequencing libraries.

value: dairy goat

value: mammary gland

tag: developmental stage

value: peak lactation


title: 72_non.H3K4me3

city: Bethesda

first: Apratim

last: Mitra

state: MD

address: 6 Center Dr Building 6B Room 2B206

department: Division of Developmental Biology

institute: NICHD

data_processing:
Alignment: Sequence reads were aligned to the May 2006 version of the chicken genome (galGal3) using Maq version 0.7.1 (Li et al. 2008). A valid alignment could have a maximum of two mismatches and if a read aligned equally well to multiple places in the genome, one was chosen at random. Redundant reads were removed before further analysis to avoid amplification bias. Read counts were summarized using non-overlapping windows of 200 bp for visualization.
Peak calling: Summarized read counts were subjected to peak calling with SICER (Zang et al. 2009). The source code was modified to include support for the chicken genome. Fragment length was specified to be about 190 bp as estimated from our ChIP-Seq experiments. A window size of 200 bp (default) and gap size of 400 bp was used for the analysis. The E-value for estimating significant peaks was set to 100.

organism: Gallus gallus

source_name: thymus tissue

growth_protocol: Two specific-pathogen-free inbred lines of White Leghorn either resistant (L63) or susceptible (L72) to MD were hatched, reared and maintained in Avian Disease and Oncology Laboratory (ADOL, Michigan, USDA).

extract_protocol: ChIP was carried out using thymus samples from MDV infected and controls birds as follows: About 30mg thymus samples from three individuals were cut into small pieces (1 mm3) and digested with MNase to obtain mononucleosomes. PNK and Klenow enzymes (NBE, Ipswich, MA, USA) were used to repair the ChIP DNA ends pulled down by the specific antibody.   A 3′ A was then added using Taq polymerase for adaptor ligation. A pair of Solexa adaptors (Illumina, USA) was ligated to the repaired ends. Seventeen cycles of PCR was performed on ChIP DNA using the adaptor primers and fragments with a length of about 190 bp (mononucleosome + adaptors) were isolated from agarose gel. Subsequently, cluster generation and sequencing analysis using the purified DNA was performed on the Solexa 1G Genome Analyzer (Illumina, USA) following manufacturer protocols.

treatment_protocol: The chickens were injected intra-abdominally with a partially attenuated very strong strain of MDV (648A passage 40, vv+) at 5 days after hatch with a viral dosage of 500 plaque-forming units (PFU). Chickens were terminated at 10dpi to collect thymus tissues. All procedures followed the standard animal ethics and use guidelines of ADOL.

value: line 72

value: thymus

tag: infection status

value: control

tag: ChIP


title: 72_inf.H3K4me3

value: infected


title: 63_non.H3K27me3

value: line 63


title: 63_non.H3K4me3


title: 72_non.H3K27me3


title: 63_inf.H3K4me3


title: C17_2b_24h_T3

status: Public on Dec 31 2012

city: Oullins

first: Fabrice

last: Chatonnet

address: 165 Chemin du Grand Revoyet

institute: INSERM

description: PolyA RNA

data_processing:
Basecalls were done with applied solid.sage.v106.
Reads were aligned on a library of 28 nt fragments of the RefSeq release from 07/20/2011 digested by NaeIII with applied solid.sage.v106. Alignment was done assuming one mismatch maximum, dividing the number of reads matching with several transcripts equally between transcript and grouping all different transcripts for a same gene under a unique gene symbol. This file is linked to the Series record as a supplementary file.
Reads number were normalized between samples by the total number of reads to obtain read frequencies (reads per million).
Genome_build: mm9
Supplementary_files_format_and_content: DGE_processed_data.txt is a matrix presenting for each gene and each sample the normalized (reads per million) number of reads for every gene, assuming one mismatch maximum, dividing the number of reads matching with several transcripts equally between transcript and grouping all different transcripts for a same gene under a unique gene symbol. This file is linked to the Series record as a supplementary file.

source_name: C17.2b neural progenitor cells, T3, 24h

growth_protocol: C17.2 cells were passed every four days and grown in DMEM medium containing 12% FBS and 2 mM Glutamine and Penicillin / Streptomycin. Cells were grown for 24h in serum-free medium before treatment.

extract_protocol: Cells were lysed and total RNA was extracted using a commercial kit. PolyA RNA were then reverse-transcribed into cDNA.

treatment_protocol: Cells were treated with 10-7M T3 or left untreated for 6, 12 or 24h.

tag: cell line

tag: cell type

value: C17.2 neural progenitor cells stably expressing tagged thyroid hormone receptor beta

tag: treatment

value: 10^-7 M T3 thyroid hormone

tag: treatment duration


title: C17_2b_6h_T3

description: PolyA RNA

source_name: C17.2b neural progenitor cells, T3, 6h


title: C17_2b_6h_un

description: PolyA RNA

source_name: C17.2b neural progenitor cells, untreated, 6h


title: C17_2b_12h_T3

description: PolyA RNA

source_name: C17.2b neural progenitor cells, T3, 12h


title: 63_inf.H3K27me3

tag: strain


title: H-sample

source_name: late lactation

tag: tissue

value: late lactation


title: C17_2b_24h_un

description: PolyA RNA

source_name: C17.2b neural progenitor cells, untreated, 24h


title: C17_2a_12h_un

description: PolyA RNA

source_name: C17.2a neural progenitor cells, untreated, 12h

value: C17.2 neural progenitor cells stably expressing tagged thyroid hormone receptor alpha


title: C17_2a_24h_T3

description: PolyA RNA

source_name: C17.2a neural progenitor cells, T3, 24h


title: C17_2a_24h_un

description: PolyA RNA

source_name: C17.2a neural progenitor cells, untreated, 24h


title: C17_2a_6h_un

description: PolyA RNA

source_name: C17.2a neural progenitor cells, untreated, 6h


title: C17_2b_12h_un

description: PolyA RNA

source_name: C17.2b neural progenitor cells, untreated, 12h


title: mtr3-1

city: Cleveland

first: Daniel

middle: Louis

last: Kiss

state: OH

address: 2109 Adelbert Road

department: Molceular Biology and Microbiology

institute: Case Western Reserve University

description: RNA-seq of total RNA, selected for poly-A+.

data_processing:
All individual samples were groomed using the FASTQ groomer in the NGS: QC and Manipulation tab, and aligned to the sacCer2 reference genome using the Map with Bowtie for Illumina function of the NGS: Mapping tab of the Galaxy website http://main.g2.bx.psu.edu/.
The aligned files were then uploaded into Avadis NGS software (STRAND Scientific Intelligence Inc.), reads were normalized, then numerical values were exported and analyzed using Microsoft Excel and Access suites.  Genes included in mitosis-related GO terms were examined in detail, and their levels compared to WT were calculated.  These values are included in the Excel file 'Table_S3_family_YEAST.xls', which is linked to the Series record as a supplementary file.

organism: Saccharomyces cerevisiae

source_name: 2 hour temperature-shifted culture, mtr3-1

extract_protocol: RNA libraries were generated using an Illumina poly-A select mRNA kit, and a unique 3-base barcode was included into each library.  All libraries were simultaneously sequenced in a single lane of an Illumina GAIIx at the Microarray and Genomic Analysis Core Facility of the Huntsman Cancer Institute, University of Utah.  36 cycles of sequencing were performed, and each read resulted in 1.1 billion filter-passed bases.  The resulting data were split by barcodes using SplitBarcode (Christopher Maxwell, U. of Utah).

treatment_protocol: Yeast strains were grown in 50 mL of YPD in a 250 mL flask at 30Â°C and 250 rpm to a final OD600 of 0.4, and were temperature shifted for 2 hours at 37Â°C.

tag: genotype

value: mtr3 mutation

value: temperature shift (OD600 of 0.4 @ 30 degrees, then 2 hours @ 37 degrees)


title: dis3^mtr

source_name: 2 hour temperature-shifted culture, dis3^mtr

value: dis3 mutation


title: mtr4-1

source_name: 2 hour temperature-shifted culture, mtr4-1

value: mtr4 mutation


title: 72_inf.H3K27me3


title: WT

source_name: 2 hour temperature-shifted culture, WT

value: WT


title: FLAG_HA_FH_Fbxl10

city: Copenhagen

first: Xudong

last: Wu

address: Ole Maaløes Vej 5

country: Denmark

institute: Biotech Research and Innovation Centre

data_processing:
Basecalls performed using CASAVA version 1.8.2 – except:  version 1.8.0 for FLAG_HA_IgG, version 1.7.0 for FLAG_HA_FH_Fbxl10
ChIP-seq reads were aligned to the mm9 genome assembly using Bowtie version 0.12.7 with parameters: -p 20 -S -m 1 mm9 <FILE>. For “FLAG_HA_FH_Fbxl10” the quality-score was specified by adding the “--solexa1.3-quals” flag
SAM-files were converted to BAM using the “samtools view” command from “Samtools” version 0.1.18, and
BAM-files were converted to BED-files using the “bamToBed” command from “bedtools” version 2.15.0
Genome_build: mm9
Supplementary_files_format_and_content: BED files (BED6)

source_name: embryonic stem cells

growth_protocol: Mouse ES cells were cultured feeder-free on 0.1% gelatin-coated plates in 2i medium (Dulbecco’s modified Eagle’s medium [DMEM; Hi-Glucose]/Neuralbasel medium 1:1, non-essential amino acids, L-glutamine, b-mercaptoethanol, penicillin/streptomycin, sodium pyruvate, N2, B27 and leukemia inhibitory factor (LIF), GSK3β and MEK1 inhibitors. All cell cultures were maintained at 37 °C with 5% CO2.

extract_protocol: Chromatin immunoprecipitation assays (ChIP) were performed and analysed as previously described {Pasini et al. (2010). Nature. 2010 Mar 11;464(7286):306-10. ES cell DNA was sonicated to an average size between 200-400 bp. The sequencing libraries were constructed using the NEBNext™ DNA Sample Prep Master Mix, NEB combined with Illumina adaptors. The DNA was purified using a Qiaquick PCR purification kit (Qiagen) and amplified by 18 cycles of PCR. The amplified DNA from ChIP-seq experiments was analyzed by Solexa/Illumina high-throughput sequencing. The tags were mapped to the mouse genome (assembly mm9) with the Bowtie alignment tool and the Solexa Analysis Pipeline.

treatment_protocol: Cells were either untransfected or transfected with a scrambled or Fbxl10 specific shRNA

tag: chip antibody

value: FLAG (Sigma, A2220) and HA (Sigma, A2095)

value: embryonic stem cells

tag: transfection/genotype


title: HSF_perox

city: Morahalom

first: Balazs

last: Horvath

address: Vallalkozok u. 7.

department: NGSP

country: Hungary

institute: SeqOmics Biotechnology Ltd.

data_processing:
Basecalls performed using Abi 5500xl ICS (Instrument Controller version 2011.09)
Data were filtered using the following specifications: minimum number of nucleotides in reads = 50
RNA reads were aligned to the NC_003070/NC_003070/NC_003070/NC_003070/NC_003076/NC_003076/Y08501_MT genome assembly using CLC Genomics Workbench version 4.7.2 with the configurations described in the readme.txt file on the Series record.
Calculate expression levels and discover novel exons using CLC Genomics Workbench version 4.7.2
Annotation using CLC Genomics Workbench version 4.7.2 (Data from www.geneontology.org)
Genome_build: NC_003070/NC_003070/NC_003070/NC_003070/NC_003076/NC_003076/Y08501_MT
Supplementary_files_format_and_content: Final processed data file, with min. 30 total gene reads
Supplementary_files_format_and_content: Comparing all samples (RPKM) temporary
Supplementary_files_format_and_content: Comparing all samples (RPKM) added function fields temporary
Supplementary_files_format_and_content: Comparing all samples (RPKM) added  annotation from www.geneontology.org
Supplementary_files_format_and_content: Primary output from CLC Genomics Workbench version 4.7.2

organism: Arabidopsis thaliana

growth_protocol: Plants were grown in sterile conditions using half-stength MS culture medium (0.5 MS) containing 0.5% sucrose,

extract_protocol: Extracting total RNA made with RNeasy (Qiagen), plus AMBION DNAse treatment. Libraries were processed according to the SOLID transcriptome sequencing protocol (Publication Part Number 4452437). Briefly, total RNA was DNaseI treated and rRNA-depleted using RiboMinus Plant Kit. After the fragmentation of RNA, the 50-200nt fraction was cleaned and was the subject of adaptor hybridization and reverse transcription. The cDNA library was cleaned with Qiagen MinElute PCR purification Kit and size-selected on a 6% TBE-Urea denaturing polyacrylamide gel.  The 150-250nt cDNA fraction was amplified to generate sufficient template for SOLID sequencing. To determine the accurate concentration of each library, a SOLID Library TaqMan Quantitation Kit was used according to the manufacturer's instructions. After, each library was clonally amplified on SOLiD P1 DNA Beads by emulsion PCR (ePCR). ePCR beads were enriched for template-positive beads by hybridization with capture beads. Template-enriched beads were extended at the 3' end in the presence of terminal transferase and 3' bead linker. Approximately 60 million beads per sample with clonally amplified DNA were deposited onto one lane of the 5500 SOLID sequencing flowchip . The flowchip was then loaded onto a SOLID 5500xl Instrument and the 50-base sequences were obtained according to manufacturer's protocol.

treatment_protocol: Plants were treated by 5µM estradiol with or without 1mM H2O2 for 6h in liquid 0.5 MS medium

tag: cultivar

value: overexpressing the HSFA4A transcription factor

value: 1mM H2O2 for 6h


title: Col_wt

value: wild type

value: untreated


title: Col_wt_perox


title: HSF

source_name: HSF


title: C17_2a_12h_T3

description: PolyA RNA

source_name: C17.2a neural progenitor cells, T3, 12h


title: C17_2a_6h_T3

description: PolyA RNA

source_name: C17.2a neural progenitor cells, T3, 6h


title: case25 nasal polyp

city: Guang Zhou

first: Peng

last: Tian

state: Guang dong

address: Yan Jiang Xi Road 107#

department: Department of Biochemistry, Department of Otorhinolaryngology-Head and Neck Surgery

institute: Sun Yat-sen 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. All of the reads were aligned to the human genome (hg19; downloaded at UCSC genome bioinformatics (Rhead et al.)) with Bowtie (version 0.12.5; parameters: -q -p 5 -k 2 --best -v 2), allowing for 2 mismatches.
The uniquely mapped reads were used for internal priming filtering by examining the genomic sequence 1 to 20 bases downstream of poly(A) cleavage sites. The uniquely mapped reads were considered to be internal priming candidates and then removed if they contained more than 12 “A”s or one of the following patterns: 5'-AAAAAAAA-3' and 5'-GAAAA+GAAA+G-3' (in which “+” means “or more”) in the 20 nt region immediately downstream from poly(A) cleavage sites.
All of the reads of the samples were iteratively clustered as described previously, and then the poly(A) cleavage sites that are located next to each other within 24 nt were clustered. Next, cleavage clusters with two or more reads were assigned as poly(A) sites.
We tested tandem 3'UTR switching events among different samples through adopting a method to test linear trend alternative to independence for two-way tables with ordered classifications [40]. For a co-expressed (in both polyp tissue and mucosa tissue ) UCSC gene with two or more tandem polyA sites, we performed the test using the following steps: 1) calculate the tandem UTR length for each tandem polyA site; 2) list the number of reads for each tandem polyA site for each sample in a table: take the tandem polyA sites as columns (from the site with the shortest UTR to that with the longest) and take the two samples as rows (mucosa tissue sample and polyp tissue); 3) if the total number of reads in the table is less than 30, neglect this gene for the test; 4) let the lengths of the tandem UTRs denote the scores for the columns; let 1 denote the row score for the mucosa tissue sample and let 2 denote the row score for the nasal polyp tissue; 5) calculate the Pearson correlation   using the number of reads in the table as the values and using the scores for the rows and columns as coordinates; 6) calculate a statistic: M2= (n-1)r2; for large samples, this statistic is approximately chi-squared with df = 1, and a P-value can be obtained. The Benjamini-Hochberg FDR was estimated using the R software. Moreover, tandem 3'UTR length switching with significant P-values paired to a false discovery rate cutoff of 1% were considered to be significantly different between the two samples. A positive value of r indicates a longer tandem 3'UTR in nasal polyp tissue and vice versa.
Software used to generate the .pla files: A series of in-house programs have been applied to generate these files. These programs use the raw Illumina fastaq file as input, and do the filtering, mapping, clustering, and finally generate these files (see Fu et al. Genome Res, 2011 for details).
Genome_build: hg19
Supplementary_files_format_and_content: Each processed data.pla file has number of reads, ucscID, cleavage site, number of cleavage sites, number of reads per gene, position of stop codon, position of clevage site, polyA signal,etc.  Further, the seventh column represent read number of a particular poly(A) site, which can be regarded as expression level of that poly(A) site. Since the geneID (the 5th column) is also included for each polyA site in this file, the gene expression levels of each gene can be studied by using this file. Thus, .pla files can be regarded as either polyA site or gene expression profiling.

organism: Homo sapiens

source_name: nose

extract_protocol: The sequencing libraries were constructed using a protocol from Yonggui et al., Genome Research, 2011. Briefly, total RNA was randomly fragmented by heating. Using template-switching technology and an improved reverse transcription (RT) reaction mixture, high-quality 3'-anchored first-strand cDNA was generated with Super Script II reverse transcriptase (Invitrogen Life Technologies, Karlsruhe, Germany). Concurrently, a 5' template-switching adaptor tagged with Illumina adaptors was added . Next, ds-cDNA was synthesized by PCR amplification with known sequencing primers and Platinum® Taq DNA Polymerase High Fidelity (Invitrogen, Carlsbad, CA, USA). Fragments of 300–500 bp were selected from the PCR products by performing PAGE separation, excision, and gel extraction with a QIAquick Gel Extraction Kit (Qiagen, Valencia, CA). The final pooled fragments were sequenced from the 3' end with an Illumina Solexa GA IIx (Illumina, San Diego, CA).

tag: patient

tag: patient condition

value: chronic rhinosinusitis with nasal polyps

value: nasal polyp

tag: gender

value: male

tag: age

value: 46 years


title: case13 nasal polyp


title: case25 nasal mucosa

value: nasal uncinate process mucosa


title: Patient 1

city: North Adelaide

first: Jozef

last: Gecz

state: South Australia

address: 72 King William Rd

department: Genetics Medicine

country: Australia

institute: SA Pathology

data_processing:
The first and last 14 bases of all reads were trimmed to improve alignment and error rate. Reads were aligned to the human genome HG19, splice junctions database, mitochondrion DNA and rRNA. Splice junctions were built from exon annotation which comprises of 45 bases on each side of the exon-exon junction (Supplied with processed data). If the exon is less than 45 bp, the whole exon is included. Reads were mapped using ELAND (Illumina). Mapped reads with more than 2 mismatches were excluded from downstream analysis.
Genome Build:
UPF2_P1_Aligned.txt: HG19

molecule: polyA RNA

source_name: Lymphoblastoid cell line

growth_protocol: The Epstein-Barr virus (EBV) immortalized B- cell lines (B-LCL) used in this study were established from peripheral blood lymphocytes of patients and controls. Once established the LCL cell lines were cultured in RPMI 1640 (Sigma) supplemented with 10% FCS, 2mM L-Glutamine, 0.017mg/ml benzylpenicillin and grown at 37 0 C with 5% CO2

extract_protocol: Total RNA was extracted from lymphoblastoid cell lines using Trizol (Inivtrogen) and RNeasy mini-kit (QIAGNE). PolyA+ RNA was purified from total RNA using oligo-dT magnetic beads. RT-PCR was performed to make cDNA with random primers. All outlined steps are part of the mRNA-seq 8 sample preparation kit (cat# RS-100-0801, Illumina) and were performed by Geneworks (Australia). ds-cDNA was sequenced using Illumina Genome Analyzer II as recommended my manufacturer, and was also performed by Geneworks (Australia).

value: B-LCL

tag: affected status

value: UPF2 Patient


title: case13 nasal mucosa


title: ChAPSeq_TRb

description: Affinity-purified bound fraction via IgG beads precipitation.

data_processing:
Basecalls were performed using CASAVA version 1.7 (ChAPSeq_TRa, ChAPSeq_TRb) or version 1.8 (ChIPSeq_RXR).
ChAP-seq or ChIP-seq reads were aligned to the mm9 genome assembly using Bowtie v0.12.7 with the following parameters:  -m 1 --strata –best:
Peaks were called using the MACS algorithm on the Galaxy website with the following setting: effective genome size (2.70e+09), tag size (36), bandwith (300), model fold (5), pvalue cut-off (10^-4), ranges for calculating regional lambda (peak_region,1000,5000,10000).
Genome_build: mm9
Supplementary_files_format_and_content: Peak.bed files are the peaks called by MACS (length of peak, summit position and score = -10*log(p-value) are indicated).
Supplementary_files_format_and_content: wig files were generated during the peak calling with a 10 bp window, except for RXR for which wig was generated during alignment with a 25 bp window.

source_name: C17.2b neural progenitor cells, TRb ChAP

growth_protocol: C17.2 cells were passed every four days and grown in DMEM medium containing 12% FBS and 2 mM Glutamine and Penicillin/Streptomycin. Cells were grown for 24h in serum-free medium before fixation.

extract_protocol: Lysates were clarified from sonicated cells and TRa-DNA or TRb-DNA complexes were isolated with IgG magnetic beads. RXR-DNA complexes were isolated with antibody. ChIP-seq libraries were prepared using the ChIP-Seq Sample Prep Kit (#IP-102-1001, Illumina) following the manufacturer's protocol with some modifications. Briefly, 10 ng of ChIP-enriched DNA were end repaired using T4 DNA polymerase, Klenow DNA polymerase and T4 PNK. A single 'A' nucleotide was added to the 3' ends of the blunt DNA fragments with a Klenow fragment (3' to 5'exo minus). The ends of the DNA fragments were ligated to double-stranded adapters using T4 DNA Ligase. The ligated products were enriched by PCR and purified using Agencourt AMPure XP beads (#A63881, Beckman). DNA library size selection was performed by excising the 250-350 bp fragments from a 2% agarose gel followed by purification using a QIAquick Gel Extraction Kit (#28906, Qiagen). Prior to analyses, DNA libraries were checked for quality and quantified using a 2100 Bioanalyzer (Agilent). The libraries were loaded in the flowcell at 8pM concentration and clusters were generated using the Cbot and sequenced.

treatment_protocol: Untreated cells.

value: C17.2 neural progenitor cells stably expressing tagged thyroid hormone receptor beta

tag: chap tag

value: Protein G


title: Znf335a_ChIPSeq

city: Boston

first: Christopher

middle: A

last: Walsh

state: MA

address: 3 Blackfan Circle

department: Genetics

institute: Children's Hospital Boston

data_processing:
Base calls performed using ELAND
ChIP-seq reads were aligned to the mm9 genome assembly using Bowtie version 0.12.7 with default parameters
peaks were called using MACS with default parameters
Genome_build: mm9
Supplementary_files_format_and_content: wig files were generated using the SPP package (Karchenko et al, doi:10.1038/nbt.1508)

source_name: lateral telencephalon

extract_protocol: Lysates were clarified from sonicated nuclei and Znf335-DNA complexes were isolated with antibody.  Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 18 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer following the manufacturer's protocols.

tag: developmental stage

tag: antibody

tag: antibody vendor

value: Bethyl

tag: antibody catalog#


title: Patient 2

data_processing:
The first and last 14 bases of all reads were trimmed to improve alignment and error rate. Reads were aligned to the human genome HG19, splice junctions database, mitochondrion DNA and rRNA. Splice junctions were built from exon annotation which comprises of 45 bases on each side of the exon-exon junction (Supplied with processed data). If the exon is less than 45 bp, the whole exon is included. Reads were mapped using ELAND (Illumina). Mapped reads with more than 2 mismatches were excluded from downstream analysis.
Genome Build:
UPF2_P2_Aligned.txt: HG19

tag: cell line


title: ChAPSeq_TRa

source_name: C17.2a neural progenitor cells, TRa ChAP

tag: cell type

value: C17.2 neural progenitor cells stably expressing tagged thyroid hormone receptor alpha

tag: treatment


title: ChIPSeq_RXR

description: Affinity-purified bound fraction via Protein A beads precipitation.

source_name: C17.2a neural progenitor cells, RXR ChIP

value: pan-RXR

value: Santa Cruz

tag: antibody catalog number


title: RNAPII-SER2P

city: Barcelona

first: Conchi

last: EI

address: Baldiri Reixac 15

country: Spain

institute: IBMB

data_processing:
Basecalls performed with CASAVA
ChIP-seq reads were mapped to mm9 whole genome using bowtie v0.12.7 with default parameters and –best --chunkmbs 512 --sam
SAM to BAM conversion was done with samtools 0.1.18
Peaks were called with SICER 1.1
Genome_build: mm9
Supplementary_files_format_and_content: BED files were generated with SICER 1.1

source_name: neural stem cells

growth_protocol: Mouse NSCs, provided by Dr. K. Helin, were dissected out from cerebral cortex of mouse embryos (E12.5) and cultured in a poly-D-lysine (5mg/ml, 2h 37°C) and laminine (5 mg/ml 37°C, 4 h 37ºC) precoated dishes growing with a media composed by equal parts of DMEM F12 (without Phenol Red, Gibco) and Neural Basal Media (Gibco) containing Penicillin/Streptomycin and Glutamax (1%), N2 and B27 supplements (Gibco), non essential aminoacids (0.1mM), sodium pyruvat (1mM), Hepes (5 mM), Heparin (2 mg/l), bovine serum albumin (25 mg/l) and βmercaptoethanol (0.01 mM). We add fresh recombinant human EGF (R&D systems) and FGF (Invitrogen) to 20 ng/ml and 10 ng/ml final concentrations respectively. Under our growing conditions NSCs preserve the ability to self-renew and to generate a wide range of differentiated neural cell types upon the activation of appropriate signals (Calloni et al, 2009; Gossrau et al, 2007; Sasaki et al, 2006).

extract_protocol: Before sequencing, ChIP DNA was prepared by simultaneously blunting, repairing and phosphorylating ends using a mixture of T4 DNA polymerase, DNA polymerase I and T4 kinase according to manufacturer’s instruction (Illumina). The DNA was adenylated at 3´end and recovered by Qiaquick PCR purification kit (Qiagen) according to the manufacturer’s recommendations. Adaptors were added by ligation and the ligated fragments were amplified by PCR, resolved in a gel and purified by Qiagen columns. The purified DNA was quantified by picogreen (Invitrogen) and diluted to a working concentration of 10 nM. Samples were loaded into individual lanes of flow cell.

treatment_protocol: TGFb (Millipore) was used at a final concentration of 5 ng/ml for 3h

tag: tissue

value: cerebral cortex

tag: development stage

tag: genotype/variation

value: polyclonal rabbit  RNAPII Phospho S2 ChIP grade

tag: vendor/catalog

value: Abcam, ab5095


title: INPUT

value: wild type


title: 2-DAP outgrown buds

city: Campinas

first: Renato

last: Vicentini

state: SP

address: Cidade Universitaria "Zeferino Vaz"

department: Centro de Biologia Molecular e Engenharia Genetica - CBMEG

country: Brazil

institute: Universidade Estadual de Campinas - UNICAMP

description: small RNA populations in the sugarcane vegetative axillary buds with 2-DAP

data_processing: Raw sequences were retrieved in a FASTQ formatted file and the adapter sequences were removed using Perl Scripts. After trimming of adapter sequences, the inserts were sorted into separate files according to their lengths.

organism: Saccharum officinarum

source_name: 2-DAP outgrown buds

extract_protocol: Ten micrograms of total RNA, prepared using TRizol reagent (Invitrogen) according to the manufacturer’s instructions, were used to generate sRNA library following Illumina’s modified protocol. The sRNA fraction of 19–28 nt was purified by size fractionation on a 15% TBE–Urea polyacrylamide gel. A 5`-adenylated single-stranded adapter was first ligated to the 3’-end of the RNA using T4 RNA ligase without ATP, followed by a second single-stranded adapter ligation at the 5’-end of the RNA using T4 RNA ligase in the presence of ATP. The resulting products were fractioned on a 10% TBE–Urea polyacrylamide gel and then used for cDNA synthesis and PCR amplification.

value: 2-DAP outgrown buds

tag: cultivar


title: 0-DAP inactive buds

description: small RNA populations in the sugarcane vegetative axillary buds with 0-DAP

source_name: 0-DAP inactive buds

value: 0-DAP inactive buds


title: MHV68 infected 1

city: St. Louis

first: Jing

last: Xia

state: MO

address: Campus Box 1045 One Brookings Drive

department: Department of Computer Science and Engineering

institute: Washington University in St. Louis

description: small RNA-Seq

data_processing:
The raw sequence reads were first processed to remove reads with no 3' sequencing adaptor, of low quality, or shorter than 18 nt. The sequence adaptor trimming was done by an in-house program that recursively searches for the longest subsequence of the adaptor appearing within a sequence read. If a raw sequence read did not have a subsequence of the adaptor that was longer than 4 nt, it was considered to have no adaptor.
Genome_build: NCBI37/mm9
Supplementary_files_format_and_content: txt format. Content: 1st column is SEQUENCE;  2nd column is COUNT

source_name: NIH 3T12 cells infectd with MHV68

growth_protocol: MHV68 WUMS (ATCC VR1465) was passaged and titered on NIH 3T12 cells (19). NIH 3T12 cells and bone marrow-derived macrophages were infected at a multiplicity of infection (MOI) of 10 PFU for1hat 37°C and 5% CO2, washed three times with medium, and replated in media. Subsequent RNA extractions on mock and infected cells were done 18 h later.

extract_protocol: Total RNA libraries for sequencing from mock- or MHV68-infected cells were prepared using the small RNA sample prep kit (Illumina) by following the Small RNA v.1.5 Sample Preparation Guide. Twenty micrograms of total RNA was used as input, and small RNA was size selected b  running total RNA on a 15% denaturing polyacrylamide gel and cutting out the RNA corresponding to 18 to 30 nt.

treatment_protocol: Total RNA was prepared using the mirVana miRNA isolation kit (Ambion) according to the manufacturer’s instructions for isolation of total RNA. RNA was treated with DNase I (Ambion) and run on a 15% denaturing acrylamide gel (Invitrogen) followed by staining with ethidium bro- mide to check for RNA integrity.

tag: strain

value: mouse bone marrow-derived macrophages

value: NIH 3T12


title: Mock treated 1

description: small RNA-Seq

source_name: NIH 3T12 cells  with mock treated


title: MHV68 infected 3

description: small RNA-Seq


title: Mock treated 2

description: small RNA-Seq


title: Ring1B_Scr_exp2

tag: chip antibody

value: Ring1B (custom-made)

value: embryonic stem cells

tag: transfection/genotype

value: untransfected


title: input DNA


title: JMJD3

value: affinity purified antibody against mouse JMJD3

value: Dr. K. Helin


title: MHV68 infected 2

description: small RNA-Seq


title: Ring1B_Suz12-/-


title: FLAG_HA_IgG

value: IgG (Thermo scientific, 23212, lot LI150449)


title: Ring1B_Ring1b-/-


title: IgG_shFbxl10


title: Znf335b_ChIPSeq

value: Bethyl


title: IgG_Scr

value: shScr


title: H3K27me3

growth_protocol: Mouse NSCs, provided by Dr. K. Helin, were dissected out from cerebral cortex of mouse embryos (E12.5) and cultured in a poly-D-lysine (5mg/ml, 2h 37°C) and laminine (5 mg/ml 37°C, 4 h 37ºC) precoated dishes growing with a media composed by equal parts of DMEM F12 (without Phenol Red, Gibco) and Neural Basal Media (Gibco) containing Penicillin/Streptomycin and Glutamax (1%), N2 and B27 supplements (Gibco), non essential aminoacids (0.1mM), sodium pyruvat (1mM), Hepes (5 mM), Heparin (2 mg/l), bovine serum albumin (25 mg/l) and βmercaptoethanol (0.01 mM). We add fresh recombinant human EGF (R&D systems) and FGF (Invitrogen) to 20 ng/ml and 10 ng/ml final concentrations respectively. Under our growing conditions NSCs preserve the ability to self-renew and to generate a wide range of differentiated neural cell types upon the activation of appropriate signals.

value: polyclonal rabbit trimethyl H3K27

value: Millipore, 07449


title: Ring1B_shFbxl10


title: Ring1B_Scr_exp1


title: Fbxl10_shFbxl10

value: Fbxl10 (custom-made)


title: Fbxl10_Scr


title: RNA-seq of mES total RNA after non-targeting shRNA replicate 2

status: Public on Dec 30 2012

city: Kansas CIty

first: Alexander

middle: (Garrett)

last: Garruss

state: Missouri

address: 1000 East 50th Street

institute: Stowers Institute for Medical Research

data_processing:
Base-calling was done with Casava v1.8 using default settings for quality filtering.
Alignment of fastq files was done with Bowtie aligner v0.12.7 allowing uniquely mapping reads only and allowing up to two mismatches for ChIP-seq samples.  RNA-seq samples were aligned using TopHat v1.4.1 and the Ensembl 63 (ES) and Ensembl 65 (EB5) GTF file allowing unique matches only.
Peak-calling was done with MACS v1.4.1 and filtered at FDR < 5 (0.05)
Genome_build: UCSC mm9
Supplementary_files_format_and_content: Aligned files are BED converted from BAM from bowtie alignments.  Peak files are MACS tab-delimited text files, filtered by FDR < 5 (0.05).

source_name: murine embryonic stem cells

growth_protocol: Mouse embryonic stem cells (KH2 and V6.5) were cultured on irradiated mouse embryonic fibroblast (MEF) feeder layers in 0.1% gelatin-coated tissue culture flask.  Cells were grown in DMEM (D6546, Sigma) supplemented with 15% ES-certified fetal bovine serum, 2 mM L-glutamine, 0.1 mM nonessential amino acids, 0.1 mM β-mercaptoethanol, and ecombinant LIF.  For ChIP and RNA analysis, cells were grown for one passage off feeders on tissue culture plates for 30 minutes.

extract_protocol: ES cells (V6.5) were infected with lentivirus carrying either Non-targeting shRNA or Ell3 shRNA in the presence of 8 ug/ml of polybrene.  24 hours later, ES cells were selected with 2 ug/ml of puromycin for additional 48 hours and then were grown one passage off feeders for 30 min before harvesting.  Total RNA was isolated with the RNeasy (74106 Qiagen) kit, treated with DNase I (M0303L NEB), and re-purified with RNeasy. For total RNA-seq analyses, 2.5 µg of total RNA was depleted of ribosomal RNA with the Ribo-Zero kit from Epicentre (#RZH1046).  Ribosomal RNA-depleted samples were used for library preparation with the TruSeq RNA Sample Prep Kit (FC-122-1001 Illumina) for next-generation sequencing.

treatment_protocol: The Non-targeting shRNA construct (SHC002) was purchased from Sigma.  Lentiviral particle preparation and infection were performed as previously described (Lin et al. 2011).  Briefly, around 70% confluent 293T cells in a 150 mm tissue culture plate were co-transfected with 8μg of the shRNA construct or a Non-targeting control shRNA, 6 μg of PsPAX2 packaging plasmids and 2 μg of pMD2.G envelope plasmids using FuGENE 6 (Roche 04709691001) or X-tremeGENE 9 (Roche 06365787001).  The media was replaced with fresh DMEM supplemented with 10% FBS after 16 hours of transfection.  The lentiviral supernatants were collected 48 and 72 hours after the transfection, filtered through 0.45 μm filters and concentrated at 18K rpm for 2 hours.  The V6.5 ES cells were infected with concentrated lentiviral particles with polybrene at the concentration of 8 μg/ml. At 24 hours after infection, the ES cells were subjected to selection with 2 ug/ml of puromycin for an additional 48 hours.

value: non-targeting shRNA

value: C57BL/6 (female) x 129/S (male)

value: mES


title: RNA-seq of mES total RNA after Ell3 shRNA replicate 1

treatment_protocol: Mouse Ell3 (RMM4534-NM_145973) shRNA constructs were purchased from Open Biosystems.  Lentiviral particle preparation and infection were performed as previously described (Lin et al. 2011).  Briefly, around 70% confluent 293T cells in a 150 mm tissue culture plate were co-transfected with 8μg of the shRNA construct or a Non-targeting control shRNA, 6 μg of PsPAX2 packaging plasmids and 2 μg of pMD2.G envelope plasmids using FuGENE 6 (Roche 04709691001) or X-tremeGENE 9 (Roche 06365787001).  The media was replaced with fresh DMEM supplemented with 10% FBS after 16 hours of transfection.  The lentiviral supernatants were collected 48 and 72 hours after the transfection, filtered through 0.45 μm filters and concentrated at 18K rpm for 2 hours.  The V6.5 ES cells were infected with concentrated lentiviral particles with polybrene at the concentration of 8 μg/ml. At 24 hours after infection, the ES cells were subjected to selection with 2 ug/ml of puromycin for an additional 48 hours.

value: Ell3 shRNA


title: ChIP-seq of WCE in mES after Ell3 shRNA

extract_protocol: For ChIP-Seq, 5×107 cells were used per immunoprecipitation according to previously described protocols (Lee et al., 2006). ChIP-Seq libraries were prepared with Illumina's ChIP-Seq sample prep kit (0801-0303).

treatment_protocol: Mouse Ell3 (RMM4534-NM_145973) shRNA constructs were purchased from Open Biosystems.  Lentiviral particle preparation and infection were performed as previously described (Lin et al. 2011).  Briefly, around 70% confluent 293T cells in a 150 mm tissue culture plate were co-transfected with 8μg of the shRNA construct or a Non-targeting control shRNA, 6 μg of PsPAX2 packaging plasmids and 2 μg of pMD2.G envelope plasmids using FuGENE 6 (Roche 04709691001) or X-tremeGENE 9 (Roche 06365787001).  The media was replaced with fresh DMEM supplemented with 10% FBS after 16 hours of transfection.  The lentiviral supernatants were collected 48 and 72 hours after the transfection, filtered through 0.45 μm filters and concentrated at 18K rpm for 2 hours.  The V6.5 ES cells were infected with concentrated lentiviral particles with polybrene at the concentration of 8 μg/ml. At 24 hours after infection, the ES cells were subjected to selection with 2 ug/ml of puromycin for an additional 48 hours.  For Pol II occupancy analysis, the 3-day control and Ell3-knockdown ES cells were grown one passage off feeders before harvesting for paraformaldehyde cross-linking.


title: RNA-seq of embryoid bodies at day 5 total RNA after non-targeting shRNA replicate 2

source_name: murine embryoid bodies day 5

growth_protocol: Embryoid bodies (EBs) were formed by culturing 150,000/ml ES cells in ES medium without LIF (ES differentiation medium) on non-adhesive bacteriological petri dishes for indicated days(Kurosawa, 2007).  Media was changed daily after two days. For neural differentiation, day-5 EBs were plated on laminin-coated 6-well tissue culture plate in ES differentiation medium with 1 mM all-trans retinoic acid for an additional 14 days.  Media was changed every other day

value: murine EB at day 5


title: ChIP-seq of WCE in mES after non-targeting shRNA

treatment_protocol: The Non-targeting shRNA construct (SHC002) was purchased from Sigma.  Lentiviral particle preparation and infection were performed as previously described (Lin et al. 2011).  Briefly, around 70% confluent 293T cells in a 150 mm tissue culture plate were co-transfected with 8μg of the shRNA construct or a Non-targeting control shRNA, 6 μg of PsPAX2 packaging plasmids and 2 μg of pMD2.G envelope plasmids using FuGENE 6 (Roche 04709691001) or X-tremeGENE 9 (Roche 06365787001).  The media was replaced with fresh DMEM supplemented with 10% FBS after 16 hours of transfection.  The lentiviral supernatants were collected 48 and 72 hours after the transfection, filtered through 0.45 μm filters and concentrated at 18K rpm for 2 hours.  The V6.5 ES cells were infected with concentrated lentiviral particles with polybrene at the concentration of 8 μg/ml. At 24 hours after infection, the ES cells were subjected to selection with 2 ug/ml of puromycin for an additional 48 hours.  For Pol II occupancy analysis, the 3-day control and Ell3-knockdown ES cells were grown one passage off feeders before harvesting for paraformaldehyde cross-linking.


title: RNA-seq of mES total RNA after non-targeting shRNA replicate 1


title: SESN3 induced 4C

city: Utrecht

first: Elzo

last: de Wit

address: Uppsalalaan 8

country: Netherlands

institute: Hubrecht Institute

description: barcode: GAGCTAAATATTTCCTGATC

data_processing:
Library strategy: 4C-Seq
The initial step in the 4C-seq analysis is the alignment of the sequencing reads to a reduced genome of sequences that flank DpnII sites (fragment ends), using custom perl scripts. Due to their ambiguous nature in reporting contacts, repetitive fragment ends were excluded from subsequent analysis. The reduced genome was based on hg19.
Genome_build: hg19
Supplementary_files_format_and_content: Processed data files are in wiggle track format. Entries in the wiggle track represent genomic regions that overlap with DpnII fragment ends. Only for DpnII fragments ends for which a sequence capture is found a value is reported.This value represents read count. For a correct analysis a comparison must be made to a full set of DpnII restriction sites (not provided).

source_name: SESN3 induced

growth_protocol: DL23 cells were grown on RPMI-1640 medium with 10% FCS and standard supplements. 4-hydroxy-tamoxifen (4OHT) from Sigma, was dissolved in ethanol and added to cells at a final concentration of 1uM.

extract_protocol: 4C procedure, as published before (Splinter et al., 2001, Genes Dev). Cells are cross linked using 1% formaldehyde for 10min at room temperature, nuclei are isolated, after which chromatin is digested with DpnII and subsequently ligated under diluted conditions. After reversal of the cross links the DNA is purified and treated with the second restriction enzyme treatment (Csp). After a second re-ligation step the sample is purified and a 4C PCR is applied.

treatment_protocol: cells are cross linked using 1% formaldehyde for 10minutes at room temperature

value: DLD1 colon carcinoma


title: CLU uninduced 4C

description: barcode: AATCCACCTCCCAGAAGATC

source_name: CLU uninduced


title: ChIP-seq of Pol2 in mES after Ell3 shRNA

value: PolII

tag: chip antibody manufacturer

value: Santa Cruz

tag: chip antibody catalog #


title: SCARA3 uninduced 4C

description: barcode: ACAAGAATATTCCACTGATC

source_name: SCARA3 uninduced


title: KLF6 induced 4C

description: barcode: CCGTTCAGAATGACTTGATC

source_name: KLF6 induced


title: RNA-seq of mES total RNA after Ell3 shRNA replicate 2


title: CLU induced 4C

description: barcode: AATCCACCTCCCAGAAGATC

source_name: CLU induced


title: SESN3 uninduced 4C

description: barcode: GAGCTAAATATTTCCTGATC

source_name: SESN3 uninduced


title: RNA-seq of embryoid bodies at day 5 total RNA after non-targeting shRNA replicate 1


title: IRS2 uninduced 4C

description: barcode: TTCTTCGTTGCGCAGGAT

source_name: IRS2 uninduced


title: ChIP-seq of Pol2 in mES after non-targeting shRNA


title: PMAIP induced 4C

description: barcode: ACAACTCTCTAACGTTGATC

source_name: PMAIP induced


title: KLF5 uninduced 4C

description: barcode: CTGACAAATATTGCTTGATC

source_name: KLF5 uninduced


title: ChIP-seq of WCE in mES


title: KLF5 induced 4C

description: barcode: CTGACAAATATTGCTTGATC

source_name: KLF5 induced


title: KLF6 uninduced 4C

description: barcode: CCGTTCAGAATGACTTGATC

source_name: KLF6 uninduced


title: ChIP-seq of Ell3 in mES

value: A fragment of human ELL3 (aa 1-248) was expressed as His-tag fusion protein in pET-16b, purified on NTA-agarose according to Qiagen’s protocol and sent to Pocono Rabbit Farm and Labs for immunization into rabbits.


title: SCARA3 induced 4C

description: barcode: ACAAGAATATTCCACTGATC

source_name: SCARA3 induced


title: PMAIP uninduced 4C

description: barcode: ACAACTCTCTAACGTTGATC

source_name: PMAIP uninduced


title: plusRA_GRO-seq

status: Public on Dec 29 2012

city: La Jolla

first: MICHAEL

middle: G

last: ROSENFELD

state: CA

address: 9500 Gilman Drive

institute: HHMI/UCSD

data_processing:
The image analysis and base calling were performed by using Illumina's Genome Analysis pipeline.
The sequencing reads were aligned to hg18 using BFAST, and we kept only 1 aligned read/ genome position for downstream analyses.
An equal number of reads (4.4 million) was randomly extracted from each sample, and the sequencing reads were counted over the entire gene body on the sense strand with respect to the gene orientation using BedTools.
Genome_build: hg18
Supplementary_files_format_and_content: BED aligned files and BEDGRAPH files

source_name: NTera2/D1 cells

growth_protocol: Ntera2/D1 cells (American Type Culture Collection) were kept in minimum essential medium α (Invitrogen) plus 10% FBS.

extract_protocol: After harvesting the cells, RNA polymerases were allowed to run-on for 5 minutes at 30C (~100bp) in the presence of sarkosyl and BrUTP. The RNA was then base hydrolyzed and purified. BrUTP-labeled run-on RNA was immunopurified with anti-BrdUTP-coated agarose beads. Run-on RNA was then polyadenylated to generate primer sites for first strand cDNA synthesis. Complementary poly-T primers contain two adaptor sequences separated by an Ape1 restriction site, which after circularization and Ape1 digestion, will terminate both ends, thus permitting cDNA generation, PCR-amplification, and library generation for deep sequencing.

treatment_protocol: For induction, the cells were treated with 1 μM atRA (Sigma) for 2 days.

value: RA


title: ∆sigH mutant-Control-mRNA

city: Madison

first: Pallab

last: Ghosh

address: 1656 Linden Dr

institute: UW-Madison

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence.  Raw RNA-seq data files in FASTQ format were assembled against the standard M. paratuberculosis K10 using the CLC Genomics Workbench 4.8 (CLC bio, Aarhus, Denmark).  Gene expression for each of the different sample conditions was calculated using “reads per kilobase million” (RPKM) expression values (Mortazavi et al., 2008). The following formula was used to determine the RPKM values:

RPKM = Number of Reads/ (Kilobase length of gene × Millions mapped reads)

This RPKM metric enables comparisons between datasets with a varying number of total reads.  All reads mapping to annotated non-coding RNA (ncRNA) were removed from the datasets before determining RPKM values. Datasets were quantile normalized and Kal’s Z-test (Kal et al., 1999) was used for the comparative gene expression analysis.  Genes were considered significantly, differentially expressed if they showed ± 2.5 fold of change with FDR-P values < 0.05.
Genome_build: SRR060191 (http://www.ncbi.nlm.nih.gov/sra/?term=SRR060191). J. Bacteriol. December 2010 vol. 192 no. 23 6319-6320.
Supplementary_files_format_and_content: Excel files include RPKM values for each Sample

organism: Mycobacterium avium subsp. paratuberculosis K-10

source_name: ∆sigH mutant-Control-mRNA

growth_protocol: M. paratuberculosis strains were grown in Middlebrook 7H9 broth supplemented with 0.5% glycerol, 2 µg/ml mycobactin J and 10% ADC.

extract_protocol:
Total RNA was extracted using TRIzol regent (Invitrogen)
Libraries were prepared according to Illumina's instructions accompanying the Illumina TruSeq RNA kit.

treatment_protocol: M. paratuberculosis cultures from mid-log phase were exposed to 10 mM diamide for 3 h. Control cultures were not treated with diamide.

value: delta-sigH mutant


title: ∆sigH mutant Diamide treated-mRNA

source_name: ∆sigH mutant Diamide treated-mRNA


title: minusRA_GRO-seq


title: WT K10-Control-mRNA

source_name: WT K10-Control-mRNA

value: wild type


title: IRS2 induced 4C

description: barcode: TTCTTCGTTGCGCAGGAT

source_name: IRS2 induced


title: fruit tissue from Prunus mume

status: Public on Dec 27 2012

city: Shenzhen

first: Zhijun

last: Zheng

address: #11 Building 2/F│Beishan Industrial Zone│Yantian District

institute: BGI

description: fruit tissue from Prunus mume

data_processing:
filter reads with many Ns or with adapter or with small  size using in-house script
filter reads which were generated by PCR duplication with in-house software
map reads to the Prunus mume genome using Tophat (v 1.3.3) with parameters set to be "--max-intron-length 50000  -m 1 -r 20 --mate-std-dev 20  --closure-search  --coverage-search --microexon-search"
convert *bam files generated by Tophat to wig format
Supplementary_files_format_and_content: wig

organism: Prunus mume

source_name: young fruit

growth_protocol: Samples were kept at natural environment

extract_protocol:
RNA was purified using TRIzol from the five fresh tissues included bud, leaf, root, stem and young fruit. Standard 200bp library protocol provide by Illumina
200bp index library

tag: species

value: Prunus mume

tag: type

value: landrace

value: fruit


title: GFP RIP-seq non-strand-specific

status: Public on Dec 28 2012

city: Cambridge

first: Yue

last: Li

state: Massachusetts

address: 32 Vassar Street, 32-D528

institute: Massachusetts Institute of Technology

description: library strategy: RIP-seq control

data_processing:
Basecalls performed using CASAVA version 1.4
RIP-seq reads are preprocessed by a custom script to trim off low quality bases from the 3' end of each read, filter out adaptors, rRNA, mtRNA reads.
The preprocessed reads were then aligned to hg19 genome assembly using TopHat 1.4.1 with default setting, allowing no more than 20 multihits. We also provided TopHat with a list of exon annotations from Ensembl (v65) for the parameter option for -GTF to enable alignment to the transcriptome first and then to the genome.
For the strand-specific sequencing, the library type --libType was set to "fr-secondstrand'' consistent with the RIP-seq experiments where the second strand of the cDNA (i.e., the opposite strand of the CCNT1-bound RNA) was sequenced.
To account for PCR artefacts, only reads aligned to the distinct genome coordinates are retained. This post-alignment filtering is achieved using samtools rmdup from Samtools and MarkDuplicates from Picard
Genome_build: hg19
Supplementary_files_format_and_content: BED converted from the BAM files

extract_protocol:
CCNT1-RNA complexes were isolated using a triple tag system that supports lentiviral stable expression and mammalian affinity purification
Tagged CCNT1 was generated using a triple tag system that supports lentiviral stable expression and mammalian affinity purification (MAPLE) (Mak et al., 2010).  The HEK293 cells stably expressing tagged CCNT1 was purified by M2 agarose beads, followed by RNA extraction by Trizol.  The library synthesis was carried out according to the RIP-seq protocol described in Zhao et al. (2010) except that one of the two experiments was done with non-strand-specific sequencing.

tag: library type

value: Non-strand-specific


title: stem tissue from Prunus mume

description: stem tissue from Prunus mume

source_name: stem

value: stem


title: CCNT1 RIP-seq strand-specific

description: library strategy: RIP-seq

tag: cell line

value: Strand-specific


title: root tissue from Prunus mume

description: root tissue from Prunus mume

source_name: root

value: root


title: Scr-Input

status: Public on Dec 26 2012

city: Sheffield

first: Ian

last: Sudbery

address: Firth Court, Western Bank

department: Molecular Biology and Biotechnology

country: United Kingdom

institute: University of Sheffield

description: Input DNA from mESCs stably expressing control shRNAs.

data_processing:
Reads were mapped to mm9 using Bowtie 0.12.7 allowing 2 mismatches.
Aligned reads were down-sampled so that all samples within an experimental condition contained the same number of reads.
Peaks were called and wig files generated using MAC14 comparing against input.
Genome_build: mm9
Supplementary_files_format_and_content: MAC14 wig files over the whole genome.

source_name: Embryonic stem cells, control shRNA, input

growth_protocol: Regular ES media supplemented with 10ng/mL LIF. For V6.5, cells were grown for at least 2 passages under feeder-free conditions.

extract_protocol:
Double-crosslinked chromatin preparation. Chromatin immunoprecipitation was performed as previously described (Schmidt et al., 2009 (PMID 19275939)), with minor modifications. Cells were fixed for 1 hour in 2mM EGS, followed by 15 minutes in 1% formaldehyde, which was quenched by the addition of glycine to a final concentration of 125 uM. Sonication was performed using a BioRuptor sonicator (Diagenode) to produce fragments of approximately 500bp-1kb. Immunoprecipitation was performed overnight at 4oC with approximately 3 µg of antibody and chromatin corresponding to 5 x 10^6 cells. Antibody-bound proteins were isolated on protein A agarose beads (Repligen) or protein A magnetic Dynabeads (Invitrogen), washed extensively, eluted, and cross-links reversed according to the Upstate protocol. Samples were then sequentially treated with RNase and proteinase K before being purified on a PureLink PCR micro column (Invitrogen). Anti-KDM2A and anti-KDM2B antibodies were generated in-house. The RING1B mouse monoclonal antibody has been described previously (Atsuta et al., 2001 (PMID 11289226)).
Sequencing libraries were generated as described in Blackledge et al., 2010 (PMID 20417597) and sequenced on the Illumina HiSeq2000 platform with 51bp reads.

treatment_protocol: KDM2B knockdown: Recombinant lentiviral particles containing scrambled or KDM2B shRNA were produced. Lentiviral infection of E14 cells was performed overnight in the presence of 4ug/mL polybrene. The following day, cells were diluted into fresh growth media and allowed to settle onto gelatine-coated dishes. Puromycin selection (1.5ug/mL) was started 48 hours following transduction, and stable lines were isolated and expanded.

tag: cell type

value: embryonic stem cells (ESCs)

tag: shRNA

value: shScrambled

tag: shRNA sequence

value: CCTAAGGTTAAGTCGCCCTCG


title: Scr-Ring1b

description: Chiq-seq of Ring1b in mESCs stably expressing control shRNA.

source_name: Embryonic stem cells, control shRNA, Ring1b ChIP


title: KD-Kdm2b

description: ChIP-seq of Kdm2b in mESCs stably expressing shRNA targeting Kdm2b.

source_name: Embryonic stem cells, Kdm2b shRNA, Kdm2b ChIP

value: CGCTGTGGAAATATCTGTCAT


title: KD-Ring1b

description: ChIP-seq of Ring1b in mESCs stably expressing shRNA targeting Kdm2b.

source_name: Embryonic stem cells, Kdm2b shRNA, Ring1b ChIP


title: bud tissue from Prunus mume

description: bud tissue from Prunus mume

source_name: bud

tag: tissue

value: bud


title: Scr-Kdm2b

description: ChIP-seq of Kdm2b in mESCs stably expressing control shRNA.

source_name: Embryonic stem cells, control shRNA, Kdm2b ChIP


title: WT-Kdm2b

description: ChIP-seq of Kdm2b in V6.5 cells.

source_name: Embryonic stem cells, WT, Kdm2b ChIP


title: miRNAs_in_HCT_WT

status: Public on Dec 25 2012

city: Palo Alto

first: shuo

last: gu

address: 269 Campus Drive

department: Pediatrics

institute: Stanford University

data_processing:
Step 1. Fastq file was converted to Fasta file usying FASTX-toolkit
Step 2. Different samples were sorted by the barcode. Of note, most sequencing results contain additional samples non-related to this study under different barcodes.
Step 3. Custom Perl scripts were used to identify and remove the 4nt barcode from the 5’ end and linker sequences from the 3’end
Step 4.RNA sequences were then aligned to human/mouse miRNA library sequences (miRBase) or shRNA sequences by bowtie with command line " bowtie  -v 3 --best --norc ref input output"
Genome_build: miRbase release 17

extract_protocol: Total RNAs were extracted from cells directly or Agos-IP samples by Trizol, and then ligated to Linker-1 (IDT) in the absence of ATP with T4 RNA ligase 1 (New England Biolabs). Ligated RNA was size selected (38-46nt DNA/RNA hybrid) on 12% PAGE. The 5’ ends RNA were ligated to barcoded adapters. Dual-ligated RNA was reverse transcribed; PCR amplified and size selected on 4% NuSieve (Lonza) agarose. All sequencing was performed on the Illumina/GAII platform.

treatment_protocol: Transfection assays were done using TransIT-LT1 (MirusBio) (for MEF cells) or Lipofectamine 2000 (Invitrogen) (for 293 cells) following the manufacturer’s protocol. Up to 5ug plasmids expressing various shRNAs with or without Flag_Ago2 expressiong plasmids(500ng)  were transfected into HEK293 or MEFcells in 10cm dishes. For details about the shRNA sequences used, please see shRNA sequences.txt supplementary file on the Series record.

value: HCT cell line

tag: passages

tag: ip antibody


title: miRNAs_in_HCT_mutant

value: HCT Dicer mutant cell line


title: GFP RIP-seq strand-specific

description: library strategy: RIP-seq

extract_protocol:
CCNT1-RNA complexes were isolated using a triple tag system that supports lentiviral stable expression and mammalian affinity purification
Tagged CCNT1 was generated using a triple tag system that supports lentiviral stable expression and mammalian affinity purification (MAPLE) (Mak et al, 2010.  The HEK293 cells stably expressing tagged CCNT1 was purified by M2 agarose beads, followed by RNA extraction by Trizol.  The library synthesis was carried out according to the RIP-seq protocol described in Zhao et al. (2010) except that one of the two experiments was done with non-strand-specific sequencing.

tag: sequencing type

value: control


title: WT-Input

description: Input DNA from V6.5 cells.

source_name: Embryonic stem cells, WT, input


title: T47D-overexpressing ELF5-ELF5 DNA-24hours-rep1

city: Darlinghurst

first: Mark

last: Cowley

state: NSW

address: 384 Victoria St.

department: Genome Informatics & Clinical Genomics

institute: Garvan Institute of Medical Research

description: T47D cell lines, overexpressing ELF5 with V5 tag, ELF5-bound enriched DNA, replicate 1, 24 hours post dox treatment

data_processing:
Sequences generated by the Illumina genome analyzer were aligned against NCBI Build 36.3 of the human genome using MAQ with default parameters
Uniquely aligning reads were converted to BED format using a custom script.
Peaks were called using MACS (version 1.4.0alpha2; Zhang et al, 2008) using only uniqely aligned reads

source_name: T47D cell line

growth_protocol: Human breast carcinoma T47D-EcoR (Musgrove et al., 2001) cell lines were grown in RPMI 1640 medium (Invitrogen) supplemented with 10 % Tet System Approved fetal bovine serum (FBS) (Clontech). Cells infected with pHUSH-ProEX-based produced by the Platinum E cell line (Morita et al., 2000). Transient transfection with plasmid constructs used FuGENE reagent (Roche) or Lipofectamine LTX (Invitrogen) according to manufacturer's instructions. Cells were maintained in the presence of puromycin (Sigma) at a concentration of 2 ug/mL.

extract_protocol: ChIP-Seq was performed as previously described (Schmidt et al., 2009). Cells were cross-linked in 1% formaldehyde, sonicated and the chromatin was purified. ChIP was performed using a 50%-50% mixture of ELF5-V5 tag specific antibody and the Santa Cruz N20 anti Elf5 antibody (The ELF5-V5 antibody was from Invitrogen, Cat no: R960-25, Lot no:673900; ELF 5 Santa Cruz, Cat no: sc-9645, Lot no: G192) and enriched DNA was amplified. 36bp fragments were sequenced.

treatment_protocol: Drug treatments were performed with Dox (Clontech) at 0.1 ug/ml, R5020 (Du Pont) at 10 nM or 17beta-estradiol (Sigma) at 10 mM. Dox-containing medium was changed every 24 hours to ensure optimum Dox activity. Adherent cells on tissue culture plates were incubated in fresh, warm, serum-free medium supplemented with 1% formaldehyde, at 37 C for 10 minutes. Cells were washed 2 times with cold PBS and scraped into 600 ul PBS with protease inhibitors (P8340, Sigma). Fixed cells were spun for 2 minutes at 6,000xg, washed as before and snap frozen in liquid nitrogen.

tag: treatment

value: doxycycline

tag: timepoint

tag: enrichment

value: enriched DNA


title: T47D-overexpressing ELF5-ELF5 DNA-48hours-rep1

description: T47D cell lines, overexpressing ELF5 with V5 tag, ELF5-bound enriched DNA, replicate 1, 48 hours post dox treatment


title: sh_HCV variants

source_name: sh_HCV variants

value: HEK293 cell lines


title: KD-Input

description: Input DNA from mESCs stably expressing shRNA targeting Kdm2b.

source_name: Embryonic stem cells, Kdm2b shRNA, input

tag: chip antibody


title: sh_miR_30 variants

source_name: sh_miR_30 variants


title: rule_validation_human


title: synthetic sh_miR_30_Ago2_IP

source_name: synthetic sh_miR_30_Ago2_IP

value: Anti-FLAG (Sigma #A2220)


title: leaf tissue from Prunus mume

description: leaf tissue from Prunus mume

source_name: leaf

value: Prunus mume

value: landrace

value: leaf


title: WT K10-Diamide treated-mRNA

source_name: WT K10-Diamide treated-mRNA

tag: genotype/variation

value: wild type

tag: strain


title: CCNT1 RIP-seq non-strand-specific

description: library strategy: RIP-seq

value: Non-strand-specific


title: Endogenous small RNAs from Larix leptolepis seedling

city: Beijing

first: Liwang

last: Qi

address: Xiang shan Road

department: The Research Institute of Forestry

institute: The Chinese Academy of Forestry

data_processing: The raw data were obtained using Illumina’s Pipeline v1.5 software following sequencing image analysis by Pipeline Firecrest Module and base-calling by pipeline Bustard Module.The extracted sequencing reads were stored in file seedling. Rawdata_sequence.txt (s_1_sequence.txt). and were then used in the standard data analysis. After the raw sequence reads were extracted from image data, a series of digital filters, namely ACGT101-miR program (Version 3.5, LC Sciences) were employed to remove various un-mappable sequencing reads. Specifically, 533ADT, 53ADT, and junk sequences (>=80% A, C, G or T; >=3 N; only A, C, or only G, T) were initially filtered out, sequences possessed a length of ﹤15-nt were cut out, followed by “hit” mRNA, RFam including rRNA, tRNA, snRNA, and snoRNA, and Repbase. The resulting sequences were referred to as “Mappable sequences” and were saved in the file called seedling.FHG_unique.txt, which is used for miRNA mapping. The small RNA sequences were blasted against Larix ESTs(TSA:JR139456--JR196814).

organism: Larix kaempferi

source_name: 1.5 year-old seedling

growth_protocol: Plants were grown in greenhouse after transplant from nursery lands

extract_protocol: Total RNAs of needle, stem, and root tissues were extracted respectively using Trizol reagent (Invitrogen, CA, USA) according to the manufacturer’s protocol. Subsequently, the three RNA samples were pooled to form a mixed-sample with equal amounts (10μg) of each tissue type. Approximately 20 μg total RNA was ligated with proprietary 5’ and 3’ adaptors developed by Illumina Inc and size-fractionated on a 15% TBE urea polyacrylamide gel and the 90 base-pair fraction was excised and re-covered. The RNA was converted to single-stranded cDNA using Superscript II reverse transcriptase (Invitrogen, CA, USA) and subsequently amplified in 15 cycles using Illumina’s small RNA primer set and Phusion polymerase (New England Lab, USA). The PCR products were size-fractionated and recovered for sequencing with Illumina's Genome Analyzer IIX (Illumina, San Diego, CA, USA)following manufacturer’s instructions.

value: Mixed tissue (root,stem,and needle)

tag: age

value: 1.5 year-old


title: WT-Kdm2a

description: ChIP-seq of Kdm2a in V6.5 cells.

source_name: Embryonic stem cells, WT, Kdm2a ChIP

value: embryonic stem cells (ESCs)


title: synthetic sh_miR_30

source_name: synthetic sh_miR_30


title: sh_HCV variants_repeat

source_name: sh_HCV variants_repeat

value: MEF cell lines


title: sh_LSW


title: sh_miR_30


title: sh_miR_30_Ago2_IP


title: rule_validation_mouse


title: sh_hAAT_25


title: sh_hAAT_25_repeat


title: sh_LSW_repeat


title: synthetic sh_miR_30_repeat

source_name: synthetic sh_miR_30_repeat


title: sh_Bantam/sh_Bantam_P_repeat


title: sh_miR_30_repeat


title: Secondary hair folicle from Inner Mongolia cashmere goat (individual C)

status: Public on Dec 23 2012

data_processing:
filter reads with many Ns or with adapter or with small size using in-house script
filter reads which were generated by PCR duplication with in-house software
map reads to the  Capra hircus genome using Tophat (v 1.2.0) with parameters set to be "--max-intron-length 150000 -m 1 -r 0 --mate-std-dev 20  --coverage-search --microexon-search"
convert *bam files generated by Tophat to wig format
Cnnnnnnnn are the contigs of the Capra hircus genome
Supplementary_files_format_and_content: wig

source_name: Secondary hair folicle From individual C

growth_protocol: goats were kept at natural environment

extract_protocol: total RNA extraction protocol, standard 200bp library protocol provide by Illumina.

tag: landrace

value: Inner Mongolia cashmere goat

value: Secondary hair folicle


title: Primaryhair folicle from Inner Mongolia cashmere goat (individual C)

source_name: Primary hair folicle From individual C

value: Primary hair folicle


title: Secondary hair folicle from Inner Mongolia cashmere goat (individual A)

source_name: Secondary hair folicle From individual A


title: Primary hair folicle from Inner Mongolia cashmere goat (individual A)

source_name: Primary hair folicle From individual A


title: synthetic sh_miR_30_Ago2_IP_repeat

source_name: synthetic sh_miR_30_Ago2_IP_repeat


title: Ovarian tissue from female Yunnan black goat

source_name: Ovarian

value: 3 years old

tag: gender

value: female

value: Yunnan black goat

value: Ovarian


title: Spleen tissue from female Yunnan black goat

source_name: Spleen

value: Spleen


title: Muscle tissue from female Yunnan black goat

source_name: Muscle

value: Muscle


title: Liver tissue from female Yunnan black goat

source_name: Liver

value: Liver


title: Lung tissue from female Yunnan black goat

source_name: Lung

value: Lung


title: Secondary hair folicle from Inner Mongolia cashmere goat (individual B)

source_name: Secondary hair folicle From individual B


title: Heart tissue from female Yunnan black goat

source_name: Heart

value: Heart


title: Lymph tissue from female Yunnan black goat

source_name: Lymph

value: Lymph


title: sh_Bantam/sh_Bantam_P

value: HEK293 cell lines


title: Brain tissue from female Yunnan black goat

source_name: Brain

value: Brain


title: DB110 HP

status: Public on Dec 22 2012

city: Padova

first: Stefano

last: Campanaro

address: Via Ugo Bassi 58/b

department: Biology

country: Italy

institute: University of Padua

description: DB110 strain is a derivative of the wild type strain SS9

data_processing:
SOLiD reads were aligned using PASS software; only those uniquely aligned on genome were considered
Using self-written perl script number of reads aliged on each gene was determined for each sample and an input file for DEGseq was generated
Differentially expressed genes were determined using DEGseq software 2.11.1
Genome_build: Photobacterium_profundum_SS9_uid62923
Supplementary_files_format_and_content: tab-delimited text files include number of reads for each Sample normalized respect to DB110_HP sample and considering the total number of reads aligned on the genome

organism: Photobacterium profundum

source_name: Bacterial culture

growth_protocol: Bacterial cultures were growth at 0.1 MPa and 28 MPa in Difco Marine 2216 Broth

extract_protocol: For RNA-seq analysis bacterial cultures were grown as explained above at two different conditions (0.1 MPa; 16°C and 28 MPa; 16°C); for each growth condition total RNA was extracted from three independent cultures using trizol (Gibco) and RNeasy columns (Qiagen). Genomic DNA was removed using DNAsi (Ambion). RNA quality was determined using the 2100 Bioanalyzer (Agilent). Equal RNA quantities obtained from the three cultures grown in the same condition were pooled to reduce biological differences between replicates and to obtain a more robust sample and representative of each condition examined.  Starting from 10 μgs of total RNA, 16S and 23S rRNAs were removed using the MICROBExpressTM kit (Ambion), after rRNA depletion, RNA was fragmented incubating the sample at 82°C for 6 mins in a fragmentation buffer (40 mM Tris Acetate pH 8.1, 100mM KOAc, 30 mM MgOAc-4H20). Fragments obtained were checked by gel electrophoresis, had an average length of 200 bases and range in quantity from 1 to 2 μgs (estimated using NanoDrop 1000 Spectrophotometer - Thermo Scientific).  All the RNA obtained from each sample was retrotranscribed using One-Cycle cDNA Kit (Invitrogen), second strand synthesis was performed using the same kit. Protocol was modified because first strand synthesis was performed using random hexamers. DNA obtained was purified using AMPureR kit (Agencourt) and quantified using NanoDrop 3300 Fluorospectrometer (Thermo Scientific).

treatment_protocol: Samples DB110_HP and TW30_HP were grown in  ~2 cm of stainless steel pressure vessels at 28 MPa (280 Atm), samples DB110_LP and TW30_LP were grown at 0.1 MPa (atmospheric pressure)

value: hydrostatic pressure Mpa 28

tag: genotype

value: wild type


title: TW30 LP

description: TW30 is a toxR mutant strain

value: hydrostatic pressure Mpa 0.1

value: toxR


title: Bladder tissue from female Yunnan black goat

source_name: Bladder

value: Bladder


title: Frog kidney

city: Toronto

first: Nuno

middle: L

last: Barbosa-Morais

state: ON

address: 160 College Street, Room 908

department: The Donnelly Centre

country: Canada

institute: University of Toronto

description: Sample 18

data_processing:
Full genomic sequences for the analyzed species were downloaded from the UCSC Genome Browser database. Full transcriptomic sequences for all species were downloaded from Ensembl. For each gene, a canonical transcript was selected for gene expression analysis based on the hierarchy derived from the BioMart associated transcript names. For the cases in which such information was not available, the longest protein-coding transcript was selected as the gene representative.
For each species, we assembled all canonical transcript (as defined above) sequences. In order to calculate the effective number of unique mappable positions in each transcript (i.e. the effective length) we performed the following steps. For each read length k, we extracted the L-k+1 (L being the transcript length) k-mer sequences from each canonical transcript and then aligned the full set of k-mers against the respective genome using Bowtie, allowing for a maximum of two mismatches. k-mers with no or one unique genomic alignment were then likewise aligned back to the canonical transcriptome. For each transcript, the number of such k-mers having a unique transcriptomic alignment was determined. This corresponds to the transcript’s effective number of unique mappable positions for k-mer mRNA-Seq reads. For each sample, the corresponding mRNA-Seq data were aligned against the respective genome using Bowtie, allowing for a maximum of two mismatches. Reads with one unique genomic alignment were then aligned against the canonical transcriptome and, for each transcript, the number of reads with one unique transcriptomic alignment were counted. Gene expression levels were determined as reads per thousand mappable positions of target transcript sequence per million of reads, where the reads uniquely align to the analyzed transcriptome. This procedure for estimating gene expression levels is a corrected version of the widely used RPKM (reads per kilobase of target transcript sequence per million of total reads) metric, and is referred to as “cRPKM” [Labbe RM et al. Stem Cells (2012)].
Genome_build: mm9, galGal3, anoCar2, xenTro2, tetNig2
Supplementary_files_format_and_content: Tab-delimited text files include cRPKM values (as defined above), as well as the mappability and the total read count for each canonical transcript, for each Sample. To generate bigWig files we aligned the mRNA-Seq data against the respective genomes using TopHat; genomeCoverageBed (part of the BEDTools suite) was run on the resulting BAM files for the quantification of genomic coverage, outputting bedGraph files; bedGraphToBigWig (dowloaded from UCSC) was then used to generate bigWig files from the bedGraph files.

organism: Xenopus tropicalis

extract_protocol:
About 20 μg of each tissue was homogenized in 700 μl QIAzol Lysis Reagent (Qiagen) using ceramic beads (Precellys). RNA was extracted according to manufacturer’s recommendations. Briefly, 140 μl of chloroform was added to the homogenate. After phase separation, 450 μl of isopropanol was added to the upper, aqueous phase. The yield and quality of the total RNA were monitored by spectrophotometry at 260, 280, and 230 nm using a Bioanalyzer Eukaryote Total RNA Nano Series II chip (Agilent).
10 μg extracted total RNA was DNase-treated (Turbo DNase, Ambion) and polyadenylated RNA was enriched twice from total RNA using polyATtract mRNA isolation system IV (Promega). RNA was reversed transcribed and converted into double-stranded cDNA (SuperScript cDNA synthesis kit, Invitrogen), sheared by sonication followed by end-repairment, A-tailing, paired end adapter (Illumina) ligation and, prior to PCR amplification, cDNA was UNG-treated to maintain strand-specificity. cDNA was amplified by 15 cycles of PCR and size selected (200-300 bp).

value: mixed

value: adult

value: kidney

tag: molecule subtype

value: poly-A enriched RNA


title: Frog skeletal muscle

description: Sample 20

source_name: Frog_skeletal muscle_pooled

value: skeletal muscle


title: TW30 HP

description: TW30 is a toxR mutant strain


title: Frog heart

description: Sample 19

value: heart


title: Primary hair folicle from Inner Mongolia cashmere goat (individual B)

source_name: Primary hair folicle From individual B

value: Inner Mongolia cashmere goat

value: Primary hair folicle


title: Frog liver

description: Sample 17

value: liver


title: Chicken skeletal muscle

description: Sample 11

source_name: Chicken_skeletal muscle_pooled


title: Chicken heart

description: Sample 10


title: Chicken kidney

description: Sample 9


title: CapSeq, insert size 100-130 nt mutation, adult stage

city: Worcester

first: Craig

last: Mello

address: 368 Plantatoin Street, Suite AS5-2047

department: Program in Molecular Medicine

institute: University of Massachusetts Medical School

description: Caenorhabditis elegans long capped RNA

data_processing:
remove 5' barcode and its variants containing a single mutation, and then remove the 3' linker or the partial linker of at least 2nt at the very 3' end. And then for CapSeq, additional 8 nt was removed from the 3' end because of random priming.
Bowtie 0.12.7, blastn 2.2.25 and custom PERL (5.10.1) scripts were used to map and analyze the sequence. In general, all post-mapping analyses were performed using custom PERL scripts. Please contact Weifeng Gu at weifeng.gu@umassmed.edu for detailed PERL scripts or the related database.
Small RNA was normalized to 5 million of matched non-structural reads and mRNA was normalized to 10 million of sense protein coding reads
Gbrowse 1.70 is used to visualize the alignments.
Genome_build: WormBase release WS215, Repbase 15.10, and miRBase release 16 for C. elegans, and NCBIM37.67, miRBase 18 and non-coding RNA database fRNAdb 3.4 for mouse.
Supplementary_files_format_and_content: Fasta format for sequence with id containing the RNA read number (_xNo.)

organism: Caenorhabditis elegans

source_name: avr-14/avr-15/glc-1 triple mutation, adult stage

growth_protocol: Normal condition on OP50 plate at 20 oC

extract_protocol: A. CapSeq protocol:  To destroy 5.8S, 18S, and 26S rRNA, 0.5-2 µg of total RNA was treated with 0.1 U/µl Terminator exonuclease (Epicentre, TER51020) in 20 µl volume containing 1U/µl of SUPERase·InTM (Ambion, AM2696) and 1 X buffer A at 30 oC for 1-2 hr. The efficiency of the above reaction could be monitored by resolving the sample on a 5% denaturing PAGE gel visualized by Ethidium Bromide.  To de-phosphorylate tRNA and 5S rRNA, the above reaction was diluted into 100 µl with 10 µl of 10X DNase I buffer, 20 U more of SUPERase·InTM, 30 U of CIP (NEB, M0290S), 5 U of DNase I (Ambion, AM2222), and H2O, and incubated at 37 oC for 30 minutes. The RNA was extracted using phenol/chloroform with phase-lock column (5PRIM, 2302830), and precipitated with 0.3M NaAc (pH 5.2) and at least 1 volume of isopropanol plus 30 µg of glycoblue (Ambion, AM9515), a procedure called as RNA cleanup. To expose the 5' phosphate in the cap structure, the RNA was treated with 0.25 U/µl TAP (tobacco acid pyrophosphatase, Epicentre, T19050) in 10 µl reaction containing 1U/µl SUPERase·InTM and  1 X TAP buffer at 37 oC for 1 hour, and then cleaned up, but without additional glycoblue because glycoblue was already added in the previous step.  The RNA was ligated with 5 µM of barcoded 5’ linkers in 10 µl reaction containing 1 X buffer, 1 U/µl of SUPERase·InTM, 2 U/µl of T4 RNA ligase (Takara, 2050A), 10% DMSO, and 0.1 µg/µl BSA at 15 oC for 8 hours, and cleaned up without additional glycoblue.   To make cDNA, the precipitated RNA was annealed with 50 pmole of RT oligo plus 10 nmole of dNTP (each nucleotide) in a 13 µl reaction at 65 oC for 5 min, and then chilled on ice for at least 2 minutes. The sample was incubated with 5 U/µl Superscript III (Invitrogen, 18080), 10 mM DTT, 1 U/µl SUPERase·InTM, and 1 X buffer, all of which brought in 7 µl volume to the previous annealing reaction. The RT was incubated at 15, 25, and 37 oC each for 15 min, and then at 50 oC for 30 min to finish the RT reaction. The RT was heat-inactivated at 85 oC for 5 min. To destroy RNA, 1 µl each of RNase A (Ambion, AM2270) and RNase H (Ambion, AM2292) is added to the RT reaction at 37 oC for 20 min. To increase the cDNA quantity, the RT reaction was diluted into 100 µl PCR reaction containing 1 X PCR buffer, 0.01 U/µl ExTaq (Takara, RR001B), 0.05 µM of oligo CMO13729, 0.2 mM dNTP, and H2O, and a linear PCR was performed for 12 cycles with condition 94 oC 20 s, 52  oC 20 s, and 72 oC 30 s. The cDNA of desired size (here ~ 130 to 170 nt  for most samples or ~ 50-130 nt for ya0217), visualized using SYBRGold (Invitrogen, S-11494) with 5S and 5.8S RNA from the total RNA as size marker, was purified from a 15% denaturing PAGE gel. The cDNA was eluted using TE buffer (10 mM Tris pH 7.5, 1mM EDTA) containing 0.3 M NaCl for 6 hours to overnight with constant vortexing, filtered through 0.45 µm Spin-x column (Costar, 19442-758), and precipitated with isopropanol and glycoblue, as above. The whole procedure after PCR above was defined as PAGE gel purification.  To obtain the optimal PCR cycle number to produce the cDNA, a testing PCR was performed in 50 µl reaction containing 1 X buffer, 0.2 µM each of oligo CMO13279 and solexa3sh, 20% of the eluted linear PCR product, 0.25 mM dNTP and 0.025 U/µl ExTaq, for 15 cycles with condition 94 oC 20 s, 52 oC 20 s, and 72 oC 30 s. And then 5 µl each of oligo solexa3 and CMO13278, each at 10 µM stock concentration, was added. 3 µl of PCR product was sampled at 3, 6, 9 and 12 more cycles, and then resolved on a 8% native PAGE gel visualized using Ethidium Bromide with 10 bp DNA marker (Invitrogen 10821-015).  The optimal PCR cycle number was defined as the one at which the PCR reaction produces the cDNA of the desired size (here insert size 100 to 130 nt for most samples or 20-100 for ya0217) without obvious bulged products (diffusive band running much more slowly). This was the condition to make the final cDNA amplicons.  To check the quality of the library, TA cloning followed by colony PCR was performed to obtain individual cDNA species sequenced using the traditional method. Finally, single end 76 or 100 nt sequence was obtained using Illumina Genome Analyzer or Hi-Seq.    Oligo used in Capseq:  RT oligo: CAGAAGACGGCATACGANNNNNNNN (N, random nucleotide)  PCR oligo:  solexa3: CAAGCAGAAGACGGCATACGA  solexa3sh: GCAGAAGACGGCATACGA  CM13279: GTTCTACAGTCCGACGATC   CM13278: AATGATACGGCGACCACCGACAGGTTCAGAGTTCTACAGTCCGACGATC  5' linker: DNA/RNA hybrid oligo (RNA preceded with ‘r’, otherwise DNA)  TCTAC-rArGrUrCrCrGrArCrGrArUrC-barcode  Barcodes: A-rTrGrArC, B-rCrArGrT, C-rGrCrTrG, D-rArTrCrA    B. Small RNA cloning:  We used ligation dependent method, as described (Gu et al., Molecular cell 36, 231-244, 2009) to make small RNA libraries. PRG-1 IP was performed using the same antibody and method, as described (Batista et al., Mol Cell 31, 67-78, 2008; Gu et al., Molecular cell 36, 231-244,2009). Unlike CIP-PNK cloning or ligation independent cloning, TAP cloning, which clones much less degradation products, was used to clone RNA from PRG-1 IP, the oxidized sample for enriching for the 3’ modified RNA (Gu et al., 2009), and the corresponding control samples. To clone capped small RNA, 18-40 nt RNA was first purified from 40 µg of total RNA using a 15% denaturing PAGE gel, and then treated with 0.5 U/µl CIP in 100 µl reaction containing 0.5 U/µl SUPERase·InTM and 1X CIP buffer at 37 oC for 1 hour. The dephosporylated RNA was cleaned up, ligated with a 3' linker, gel-purified, and split into two parts: one quarter treated with PNK and the rest treated with TAP. The treatment and the following cloning protocol were described previously (Gu et al.,Molecular cell 36, 231-244, 2009). Except the previously published oxidized sample (the first step of beta-elimination) and its control, other samples were cloned using 5' 4nt-barcoded linker. Single-end 36nt sequence was obtained using illumina genome analyzer.

tag: 5' barcode

value: CAGT

tag: 3' linker

value: TCGTATGCC

value: young adult


title: Lizard heart

description: Sample 14

organism: Anolis carolinensis

tag: tissue


title: Chicken liver

description: Sample 8


title: Lizard kidney

description: Sample 13


title: Chicken brain

description: Sample 7

value: brain


title: CapSeq, insert size 20-100 nt YA

description: Caenorhabditis elegans long capped RNA

source_name: wild type, N2, Young adult stage

value: ATCA

value: wild type


title: Mouse heart

description: Sample 5


title: CIP-TAP, insert 18-40 nt adult

description: Caenorhabditis elegans capped small RNA

source_name: wild type, N2, adult

value: GCTG

value: CTGTAG


title: Kidney tissue from female Yunnan black goat

source_name: Kidney

value: 3 years old

value: female

value: Yunnan black goat

value: Kidney


title: CIP-PNK, insert size 18-40 nt adult

description: Caenorhabditis elegans normal small RNA

value: TGAC

value: adult


title: Mouse kidney

description: Sample 4

value: mixed

value: kidney

value: poly-A enriched RNA


title: Lizard skeletal muscle

description: Sample 15

source_name: Lizard_skeletal muscle_pooled

value: skeletal muscle


title: DB110 LP

value: hydrostatic pressure Mpa 0.1


title: Mouse liver

description: Sample 3

value: liver


title: PRG1 IP, insert size 18-40 nt adult

description: Caenorhabditis elegans PRG1 IP for 21U-RNA

value: CAGT


title: Frog brain

description: Sample 16

value: brain


title: Mouse brain rep1

description:
Sample 1
The 'Mouse_brain.bw' supplementary file merges data from both Mouse brain rep1 and Mouse brain rep2 samples.


title: Pufferfish brain

description: Sample 21

organism: Tetraodon nigroviridis


title: CapSeq, insert size 100-130 nt YA

description: Caenorhabditis elegans long capped RNA

value: ATCA

value: TCGTATGCC

value: wild type

value: young adult


title: CapSeq, insert size 100-130 nt 6m

description: mouse testis long capped RNA

source_name: wild type, 6 months

value: GCTG

value: 6 months


title: CapSeq, insert size 100-130 nt 4wk

description: mouse testis long capped RNA

source_name: wild type, 4 week

value: 4 week


title: CapSeq, insert size 100-130 nt L2

description: Caenorhabditis elegans long capped RNA

source_name: wild type, N2, L3 stage

value: L2 age


title: Mouse skeletal muscle

description: Sample 6

source_name: Mouse_skeletal muscle_pooled

value: adult