title: small RNAs from wild type worms input control for the IP experiment

status: Public on Oct 02 2009

first: Julie

middle: M.

last: Claycomb

address: 373 Plantation Street Biotech2 Suite 219

department: Program in Molecular Medicine

institute: University of Massachusetts Medical School

description:
small RNAs, 18 to 26 nt
18 to 26 nt small RNAs from wild type animals

data_processing: Small RNA sequences bigger than 17 nt were mapped to the Caenorhabditis elegans genome (WS192, www.wormbase.org). Sequences that matched tRNA or rRNA sequences were then removed. Only full-length complete matches were allowed. siRNA-targeted loci were defined as a region containing at least 25 reads per million matching reads. Relative abundance at each locus was determined by calculating (22G-RNAs in mutant(or)IP)/(22G-RNAs in wt+22G-RNAs in mutant(or)IP ).

organism: Caenorhabditis elegans

source_name: wild type adult animals

growth_protocol: animals were grown at 20 C for approximately 60h. Csr-1(tm892), ego-1(om97) and DA1316 animals were counter selected on ivermectin

extract_protocol:
[CSR-1 IP samples] RNA was eluted from CSR-1 immunopercipitated complex by extraction with TRI Reagent (MRC Reagents, Inc). RNA was extracted as described in the manufacturer’s protocol. Small RNAs from the immunopercipitated complex, and wild type input, were resolved in a 15% polyacrylamide 7M Urea Gel, along with 20 picomoles of RNA standard (18- and 26-nt) in separate lanes. Ethidium Bromide staining was used to visualize the RNA standards. A gel fragment was excised from the sample lanes in the migration range between the two standards. RNA was eluted from the gel fragment in (0.3M NaCl-TE pH7.5) solution overnight and ethanol-precipitated using 20mg of glycogen as the carrier. Gel purified RNA was incubated with 0.05 Unit/µl Tobacco Acid Pyrophosphatase (Epicenter Biotechnologies) in 10µl reaction buffer containing 1 Unit/µl SUPERase Inhibitor (Ambion) for 1h at 37°C. After phenol extraction, the RNA was ethanol precipitated. The gel purified RNA and 1µM of each standard were incubated with 20µM of 3’-end linker, 1 Unit of SuperRNaseIN, 10% DMSO and 3 Units T4 RNA ligase (Takara) in 10µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2, 6mg/mL BSA, 10mM DTT). The 3’ ligated products were gel purified and the RNAs were incubated in the presence of 30µM of 5’ adapter oligonucleotide, 1 Unit SuperRNaseIN (Ambion) and 1.5 Units of T4 RNA ligase in ligation buffer (50mM Tris-HCI (pH7.5), 10mM MgCI2, 10mM DTT, 1mM ATP and 10% Dimethyl sulfoxide). The ligated products were gel purified as described above and reverse transcribed in a standard 50µl reaction (SuperScript III, Invitrogen). The cDNA was amplified by PCR and purified in a 10% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing platform (Illumina, Inc.)
[csr-1(tm892), ego-1(om97) and DA1316 samples] RNA was resolved in a 15% polyacrylamide 7M Urea Gel, along with 20 picomoles of RNA standard (18- and 26-nt) in separate lanes. Ethidium Bromide staining was used to visualize the RNA standards. A gel fragment was excised from the sample lanes in the migration range between the two standards. RNA was eluted from the gel fragment in (0.3M NaCl-TE pH7.5) solution overnight and ethanol-precipitated using 20mg of glycogen as the carrier.  Gel purified RNA was treated with 1 Unit/µl of Alkaline Phosphatase, Calf Intestine (NEB) in 100mM NaCl, 50mM Tris-HCl, 10mM MgCl2, 1mM Dithiothreitol, pH 7.9 at 25°C and 1 Unit/µl SuperRNaseIn (Ambion) for 1 hour at 37 °C. After phenol extraction, the gel purified RNA and 1µM of each standard were incubated with 20µM of 3’-end linker, 1 Unit of SuperRNaseIN, 10% DMSO and 3 Units T4 RNA ligase (Takara) in 10µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2, 6mg/mL BSA, 10mM DTT).  The 3’ ligated products were gel purified and treated with 1 Unit/µl Polynucleotide Kinase in 1x Polynucleotide Kinase buffer (70mM Tris-HCl, 10mM MgCl2, 5mM Dithiothreitol, pH 7.6 at 25°C), 2mM ATP, 1 Unit/µl SuperRNAseIN. After phenol extraction, RNAs were incubated in the presence of 30µM of 5’ adapter oligonucleotide, 1 Unit SuperRNaseIN (Ambion) and 1.5 Units of T4 RNA ligase in ligation buffer (50mM Tris-HCI pH7.5), 10mM MgCI2, 10mM DTT, 1mM ATP) and 10% Dimethyl sulfoxide). The ligated products were gel purified as described above and reverse transcribed in a standard 50µl reaction (SuperScript III, Invitrogen). The cDNA was amplified by PCR and purified in a 10% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing platform (Illumina, Inc.)

value: Wild type, N2


title: small RNAs from ego-1 mutant animals

type: SRA

city: Worcester

state: MA

country: USA

description:
small RNAs, 18 to 26 nt
18 to 26 nt small RNAs from ego-1 mutant animals

library_source: transcriptomic

molecule: total RNA

source_name: ego-1(om97) adult animals

tag: strain


title: Cdx2_ChIPSeq

status: Public on Oct 01 2009

city: Baltimore

first: Minoru

last: Ko

state: MD

address: 251 Bayview Blvd, Suite 100, 10C

department: National Institute on Aging

institute: NIH

description: Chromatin IP against CDX2-Flag fusion protein

library_source: genomic

data_processing:
Sequencing of Flag-ChIP DNA fragments produced 17.59 million 36-nucleotide tags, and 32.5% of them (5.72 million) were successfully mapped to the latest mouse genome sequence (mm9, NCBI/NIH) using ELAND program. The remaining tags either had >2 mismatches or matched to more than 5 different genome locations. Among tags with multiple matches we selected only those with the minimum number of mismatches. To identify significant peaks of tags we first estimated the median distance between closely located tags in positive and negative orientation, which appeared to be ca. 150 bp. Thus, we subtracted 150 bp from coordinates of tags with negative orientation. The expected location of CDX2 binding site was estimated as tag start + 91. Redundant tags with the same starting positions were not counted to avoid amplification-related bias. Peaks of tags were identified as regions where the density of tags (minimum 6 tags) was higher than the 2.5-percentile of the frequency distribution. The frequency distribution of the log-transformed distance between i-th tag and i+5 tag was estimated for each chromosome, and the upper portion of the distribution starting from the maximum frequency was used to estimate the standard deviation (from maximum). Then the maximum minus 2 standard deviations was used as a threshold; if log-distance was below that threshold it was considered a part of a peak. Within each peak we identified the densest region of tags (within 350-bp window) which was viewed as the expected position of TF binding to DNA. Tags >500 bp away from this position were not counted. The number of tags in peaks was then compared with the number of tags in the control sample for the same region corrected by the total coverage of tags. If the adjusted number of control tags was >50% of the number of tags in the peak or if it was >20 (10 fold higher than in average), then the peak was discarded. These conditions of peak filtering were optimized by searching CDX2 motifs in all peaks and then estimating the proportion of peaks with motifs among groups of peaks classified by the number of tags in the peak and the number of control tags. Total we found 59098 peaks with at least 6 tags, of which 15,855 had at least 10 tags.
CDX2 binding motifs were found using the CisFinder on http://lgsun.grc.nia.nih.gov/CisFinder/. We extracted 200-bp long sequences from mouse genome centered at 15,855 TFBS locations identified with ChIP-Seq and searched for DNA motifs that were over-represented in these sequences compared to contol (regions from 350 to 500 bp from TFBS). We selected motifs that were over-represented by at least 2-fold and clustered them using 0.9 correlation threshold.
Identified CDX2 binding sites were annotated using CisView  http://lgsun.grc.nia.nih.gov/cisview/. For each binding site we considered genes with nearest transcription start sites (TSS). Preference in annotation was given to genes with good symbols. If the distance to TSS of a gene exceeded 5-fold the distance to the nearest gene of the same quality, then the gene was not included into the annotation. We did not consider genes >200 Kb away from binding sites. To quantify the association between genes and CDX2 binding sites we estimated the binding score for each gene combining information on the number of tags and distance from the TSS. In the case of multiple binding sites, their scores were combined.

The 'GSM364858_Cdx2_eland_multi.txt' file is the output of the ELAND alignment program run in multiple-match mode.

molecule: genomic DNA

organism: Mus musculus

source_name: Embyonic stem cells

growth_protocol: ES cells grown without feeders on gelatin coated dish in DMEM containing 15% FBS, Heat Inactivated (GIBCO); LIF (ESGRO) 1000 U/ml; Doxycyclin 0.2 ug/ml; uvate (GIBCO); 0.1 mM Non Essential Amino Acids (GIBCO), 2 mM glutamate (GIBCO), 0.1 mM beta-mercapto ethanol (GIBCO), and 1X penicillin/streptomycin (GIBCO). Cells were incubated at 37C; 5% CO2. Medium was changed daily.

extract_protocol: Chromatin from 100000000 cells was incubated with anti-FLAG M2 affinity gel at 4C over night. Eluted product were incubated at 65?C for 8 h. Purified ChIP DNA was used for ChIP-Seq preparation according to manufacturer's manuals.

treatment_protocol: ES cells grown without feeders on gelatin coated dish in DMEM containing 15% FBS, Heat Inactivated (GIBCO); LIF (ESGRO) 1000 U/ml; 1mM sodium pyruvate (GIBCO); 0.1 mM Non Essential Amino Acids (GIBCO), 2 mM glutamate (GIBCO), 0.1 mM beta-mercapto ethanol (GIBCO), and 1X penicillin/streptomycin (GIBCO). Cells were incubated at 37C; 5% CO2. Medium was changed daily.

tag: cell type

value: Cdx2-inducible ES cells, ES[MC1R(20):tetCdx2(6)]

tag: passages

tag: antibody

value: Flag M2


title: small RNAs that co-immunopercepitate with the argonaute CSR-1

description:
small RNAs, 18 to 26 nt
18 to 26 nt small RNAs from CSR-1 immunopercipitate

source_name: CSR-1 immunopercipitate

value: CSR-1 immunopercipitate from wild type, N2


title: Strand-specific, shotgun sequencing of mRNA from the IMR90 cell line; mRNA-seq_imr90_r1

status: Public on Oct 13 2009

city: La Jolla

first: UCSD AND

last: SALK

state: CA

address: Health Sciences Drive

institute: University of California, San Diego

description:
sample_term_id: EFO_0001196
assay_term_id: OBI_0001271
nucleic_acid_term_id: SO_0000871
Library name: mRNA-seq_imr90_r1
EDACC Genboree Sample Page:
http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FUCSD%2FSAMPLE%2FEDACC.1121
EDACC Genboree Experiment Page:
http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FUCSD%2FEXPERIMENT%2FEDACC.1440

****************
For data usage terms and conditions, please refer to:
http://www.drugabuse.gov/funding/funding-opportunities/nih-common-fund/epigenomics-data-access-policies
****************

data_processing:
**********************************************************************

ANALYSIS FILE NAME: GSM438363_UCSD.IMR90.mRNA-Seq.mRNA-seq_imr90_r1.bed
ANALYSIS CENTER: EDACC
ANALYSIS ALIAS: mRNA-seq_imr90_r1.hg19.level.1.release.4
ANALYSIS TITLE: Mapping of IMR90 Cell Line mRNA-Seq Data
ANALYSIS DESCRIPTION: SOLiD reads produced by mRNA-Seq on the IMR90 Cell Line, Library IMR90 mRNA   were mapped to the human genome using Pash.
ANALYSIS TYPE: REFERENCE_ALIGNMENT
EDACC Genboree Analysis Page: http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FEDACC%2FANALYSIS%2FEDACC.9489
DATA_ANALYSIS_LEVEL: 1
EXPERIMENT_TYPE: mRNA-Seq
GENOME_ASSEMBLY: NCBI Build GRCh37/UCSC Build hg19
SOFTWARE: Pash
SOFTWARE_VERSION: 3.0
MAXIMUM_ALIGNMENT_LENGTH: Read length
MISMATCHES_ALLOWED: 10% of read length
ALIGNMENTS_ALLOWED: 1
TREATMENT_OF_MULTIPLE_ALIGNMENTS: If a read maps to more than 1 position it is removed from consideration.
TREATMENT_OF_IDENTICAL_ALIGNMENTS_OF_MULTIPLE_READS: None
ALIGNMENT_POSTPROCESSING: None
RELEASE_NUMBER: Human Epigenome Atlas 4

QUALITY SCORES:
NUMBER_OF_MAPPED_READS: 27,821,581
NUMBER_OF_MRNA-SEQ_EXPERIMENTS_SCORED_IN_THIS_RELEASE: 8
PERCENT_READS_MAPPING_TO_UCSC_GENES: 93.6
PERCENT_READS_MAPPING_TO_UCSC_GENES_PERCENTILE: 100
MAXIMUM_REPLICATE_CORRELATION: NA

**********************************************************************

ANALYSIS FILE NAME: GSM438363_UCSD.IMR90.mRNA-Seq.mRNA-seq_imr90_r1.wig
ANALYSIS CENTER: EDACC
ANALYSIS ALIAS: mRNA-seq_imr90_r1.hg19.level.2.release.4
ANALYSIS TITLE: Raw Signal Density Graphs of IMR90 Cell Line mRNA-Seq Data
ANALYSIS DESCRIPTION: SOLiD mRNA-Seq read mappings from the IMR90 Cell Line, Library IMR90 mRNA   were processed into density graphs of raw signal representing the aligned read density.
ANALYSIS TYPE: ABUNDANCE_MEASUREMENT
EDACC Genboree Analysis Page: http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FEDACC%2FANALYSIS%2FEDACC.9497
DATA_ANALYSIS_LEVEL: 2
EXPERIMENT_TYPE: mRNA-Seq
GENOME_ASSEMBLY: NCBI Build GRCh37/UCSC Build hg19
SOFTWARE: In house programs and scripts
SOFTWARE_VERSION: NA
READ_EXTENSION: 0bp
GENOMIC_WINDOW: 20bp
TREATMENT_OF_REGIONS_PRONE_TO_MULTIPLE_ALIGNMENTS: None
RELEASE_NUMBER: Human Epigenome Atlas 4
BROWSER_TRACK_NAME: IMR90 mRNA r1
BROWSER_TRACK_DESCRIPTION: UCSD IMR90 Cell Line mRNA-Seq Library mRNA-seq_imr90_r1 EA Release 4

QUALITY SCORES:
NUMBER_OF_MAPPED_READS: 27,821,581
NUMBER_OF_MRNA-SEQ_EXPERIMENTS_SCORED: 8
PERCENT_READS_MAPPING_TO_UCSC_GENES: 93.6
PERCENT_READS_MAPPING_TO_UCSC_GENES_PERCENTILE: 100
MAXIMUM_REPLICATE_CORRELATION: NA

**********************************************************************

organism: Homo sapiens

extract_protocol: Library construction protocol: Total RNA was isolated from IMR90 cells and mRNA isolated by RiboMinus depletion of rRNA. mRNA was fragmented, dephosphorylated, 5' phosphorylated, then sequentially ligated to the adenylated 3' RNA adapter then to the 5' RNA adapter. Following reverse transcription of the adapter ligated RNA, the library was enriched by 15 cycles of PCR.

tag: disease

value: ATCC

value: Cell Line

tag: line

tag: lineage

tag: batch

value: Fetal lung fibroblast

tag: passage

tag: medium

value: Minimum Essential Medium, Eagle with Earle's Balanced Salt Solution supplemented with 10% Fetal Bovine Serum (FBS), 1% Pen/Strep (penicillin, streptomycin), 1% GlutaMax (stable form of glutamine)

tag: Sex

value: Female

value: mRNA-Seq

value: mirVana miRNA isolation kit (Applied Biosystems), performed as per manufacturer's instructions to isolate total RNA, followed by treatment with DNaseI (Qiagen) for 30 min at room temperature

value: 2 x RiboMinus (Life Technologies) rRNA depletion (5S, 5.8S, 12S, 18S, 28S)

value: 15 min at 70 ˚C with RNA Fragmentation kit (Ambion/Applied Biosystems) as per manufacturer's instructions

value: 5' GUUCAGAGUUCUACAGUCCGACGAUC

value: 5' UCGUAUGCCGUCUUCUGCUUGidT, 5' adenylated (Illumina)

value: 5' CAAGCAGAAGACGGCATACGA

value: Fragmented RNA was treated with 5 U Antarctic phosphatase (New England Biolabs) for 40 min at 37˚C in the presence of 40 U RNaseOut followed by phosphatase heat inactivation at 65˚C for 5 min. The RNA was purified using 66 µl SPRI beads (Agencourt) and eluted in 11 µl 10 mM Tris buffer pH 8.0.

value: Phosphorylation was performed by addition of 10 U PNK (New England Biolabs), 1 mM ATP, and 20 U RNaseOut  (Life Technologies) and incubation at 37˚C for 1 h.

value: 1 µl of 1:10 diluted adenylated 3’ RNA adapter oligonucleotide was added to the phosphorylated RNA and incubated at 70˚C for 2 min followed by placement on ice. The 3’ RNA adapter ligation reaction was performed by addition of 2 µl 10x T4 RNA ligase 2 truncated ligation buffer, 1.6 µl 100 mM MgCl2, 20 U RNaseOut and 300 U T4 RNA ligase 2 truncated (New England Biolabs) and incubation at 22˚C for 1 h.

value: Ligation of the 5’ RNA adapter was performed by addition to the 3’ adapter ligated reaction of 1 µl 1:1 diluted, heat denatured (70˚C 2 min) 5’ RNA adapter oligonucleotide, 1 µl 10 mM ATP, and 10 U T4 RNA ligase (Promega), and incubation at 20˚C for 1 h. RNA was purified using 66 µl SPRI beads and eluted in 10 µl 10 mM Tris buffer pH 8.0.

value: To the RNA ligation products, 2 µl 1:5 diluted RT primer was added and heat denatured (70˚C 2 min), followed by incubation on ice. Added to the denatured RNA/primer solution was 4 µl 5x first strand buffer, 1 µl 12.5 mM dNTPs, 2 µl 100 mM DTT, and 40 U RNaseOut, followed by incubation at 48˚C for 1 min. To this, 200 U Superscript II reverse transcriptase (Life Technologies) was added, followed by incubation at 44˚C for 1 h.

value: The entire reverse transcription reaction was used in the PCR enrichment of the library.

value: Phusion hot-start high fidelity DNA polymerase (New England Biolabs). 1x Phusion polymerase buffer and 4 U Phusion hot-start high fidelity DNA polymerase was used in a 100 µl reaction.

value: 98˚C 30 sec; 98˚C 10 sec, 60˚C 30 sec, 72˚C 15 sec; 72˚C 10 min

value: PCR products were purified in two steps, first by purification using 180 µl SPRI beads and elution in 30 µl 10 mM Tris buffer pH 8.0, followed by purification with 39 µl SPRI beads and elution in 10 µl 10 mM Tris buffer pH 8.0.


title: ekl-1_CIPPNK_seq_prb_s2

first: Craig

last: Mello

address: 368 Plantatoin Street, Suite AS5-2047

description: CIPPNK

data_processing: Small RNA sequences longer than 17 nt were mapped to the Caenorhabditis elegans genome (WS192, www.wormbase.org). Sequences that matched tRNA or rRNA sequences were then removed. Only full-length perfect matches were allowed. siRNA-targeted loci were defined as a region containing at least 25 reads per million matching reads. Relative abundance at each locus was determined by calculating (22G-RNAs in mutant(or)IP)/(22G-RNAs in wt+22G-RNAs in mutant(or)IP ).

source_name: adult worms

growth_protocol: animals were grown at 20 C for approximately 60h.

extract_protocol: CIP-PNK protocol: RNA was resolved in a 15% polyacrylamide 7M Urea Gel, along with 20 picomoles of RNA standard (18- and 26-nt) in separate lanes. Ethidium Bromide staining was used to visualize the RNA standards. A gel fragment was excised from the sample lanes in the migration range between the two standards. RNA was eluted from the gel fragment in (0.3M NaCl-TE pH7.5) solution overnight and ethanol-precipitated using 20mg of glycogen as the carrier.  Gel purified RNA was treated with 1 Unit/µl of Alkaline Phosphatase, Calf Intestine (NEB) in 100mM NaCl, 50mM Tris-HCl, 10mM MgCl2, 1mM Dithiothreitol, pH 7.9 at 25°C and 1 Unit/µl SuperRNaseIn (Ambion) for 1 hour at 37 °C. After phenol extraction, the gel purified RNA and 1µM of each standard were incubated with 20µM of 3’-end linker, 1 Unit of SuperRNaseIN, 10% DMSO and 3 Units T4 RNA ligase (Takara) in 10µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2, 6mg/mL BSA, 10mM DTT).  The 3’ ligated products were gel purified and treated with 1 Unit/µl Polynucleotide Kinase in 1x Polynucleotide Kinase buffer (70mM Tris-HCl, 10mM MgCl2, 5mM Dithiothreitol, pH 7.6 at 25°C), 2mM ATP, 1 Unit/µl SuperRNAseIN. After phenol extraction, RNAs were incubated in the presence of 30µM of 5’ adapter oligonucleotide, 1 Unit SuperRNaseIN (Ambion) and 1.5 Units of T4 RNA ligase in ligation buffer (50mM Tris-HCI pH7.5), 10mM MgCI2, 10mM DTT, 1mM ATP) and 10% Dimethyl sulfoxide). The ligated products were gel purified as described above and reverse transcribed in a standard 50µl reaction (SuperScript III, Invitrogen). The cDNA was amplified by PCR and purified in a 10% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing platform (Illumina, Inc.)

tag: sample type

value: adult worms


title: mut7_CIPPNK_seq_prb_s8

description: CIPPNK

source_name: gravid adult worms

value: gravid adult worms


title: deb26_CIPPNK_seq_prb_s5

description: CIPPNK

value: rrf-1(neC1) ego-1(om97)


title: smg5_CIPPNK_seq_prb_s5

description: CIPPNK


title: rde4_CIPPNK_seq_prb_s8

description: CIPPNK


title: rde-3_CIPPNK_seq_prb_s7

description: CIPPNK


title: mago12_CIPPNK_seq_prb_s3

description: CIPPNK


title: RPO21  120 min

status: Public on Oct 15 2009

city: Jerusalem

first: Assaf

last: Weiner

address: Givat Ram

department: Computer Science

country: Israel

institute: The Hebrew University of Jerusalem

description: MNase-seq at12 0 minutes after shifting the RPO21 yeast strain from 25 C to 37 C

data_processing: Mapped Reads: Unique reads mapped to the yeast genome with minimal errors (as described in Yassour et al. PNAS, 2008)

organism: Saccharomyces cerevisiae

growth_protocol: For rpb1-1 temperature shifts, cells were grown to an OD of 0.6 in YPD at 25 C, then culture aliquots were immediately shifted to 37 C by addition of an equal volume of YPD at 49 C.

extract_protocol: After recovery of digested DNA, mononucleosomes were gel-purified and subjected to Solexa sequencing as described in Shivwaswamy et al

treatment_protocol: Yeast culture, fixation, and MNase titrations were carried out as previously described (Yuan et al. 2005)


title: WAGO-1_input_CIPPNK_seq_prb_s7

description: CIPPNK

source_name: Young adult worms

value: Young adult worms


title: WAGO-1_IP_CIPPNK_seq_prb_s6

description: CIPPNK

source_name: WAGO-1 IP from Young adult worms

value: WAGO-1 IP from Young adult worms


title: oocyte_TAP_seq_prb_s2

description: TAP

source_name: oocytes

value: oocytes

value: rom fem-1(hc17)


title: FLX sequencing of Piwi-associated small RNAs extracted from Drosophila ovarian somatic cells (OSC)

city: Koto-ku

first: Toutai

last: MITUYAMA

state: Tokyo

department: Artificial Intelligence Research Team

country: Japan

institute: National Institute of Advanced Industrial Science and Technology (AIST)

description: After immunoprecipitation, total RNAs were isolated from the immunoprecipitates with phenol:chloroform and precipitated with ethanol. RNAs were dephosphorylated with 16 CIP (NEB) and labeled with 32P-gamma-ATP with T4 polynucleotide kinase (TaKaRa) for visualization.

data_processing: The adapter sequences attached to 5' and 3' end of every sequence produced from Roshe/454 FLX system were removed. We used NCBI bl2seq program (-p BLASTN -g F -W 10 -e 10 -D 1) to detect fragments of the adapter sequences in the raw sequence and extracted 21,211 sequences. Sequences containing ambiguous Ns were discarded. Among the extracted sequences, 21,089 sequences were mapped onto Drosophila melanogaster genome (dm3). 21,089 sequences were boiled down to 10,890 unique sequence set.

organism: Drosophila melanogaster

source_name: ovarian somatic cell (OSC)

growth_protocol: not applicable

extract_protocol: Piwi was immunopurified from OSC using a specific antibody. Total RNAs were isolated from the immunoprecipitates with phenol:chloroform and precipitated with ethanol. Cloning of small RNAs associated with Piwi in OSC was carried out.

treatment_protocol: not applicable

value: ovarian somatic cell (OSC)

tag: method

value: immunoprecipitation

tag: target

value: Piwi protein


title: heterozygous affected female, biological rep3

status: Public on Sep 30 2009

type: RNA

city: Mannheim

first: Li

last: Li

address: Theodor-Kutzer-Ufer

country: Germany

institute: University Heidelberg

description: Gene expression data from kidney of 36d old heterozygous affected female PKD/Mhm rats

hyb_protocol: Processed according to the standard Affymetrix protocol.

scan_protocol: GeneChips were scanned using the Affymetrix GeneArray Scanner3000.

data_processing: The data were analyzed with JMP Genomics,  version 3.1 (Santa Clara, CA, USA). The data have been log2 transformed and quantile normalized for each microarray.

label: biotin

organism: Rattus norvegicus

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol.

growth_protocol: Rats were kept under standard laboratory conditions (12 h light cycle, 55% humidity, 20 ± 2ºC) in an animal care facility in Mannheim, Germany. They had free access to tap water and to standard 19% protein rat pellet feed (ssniff R/M-H, Soest, Lage, Germany).

extract_protocol: Trizol extraction of total RNA was performed according to the manufacturer's instructions.

treatment_protocol: Animals under anesthesia with isofluran inhalation were killed by cervical translocation. The left kidneys were snap frozen in liquid nitrogen for microarray analysis.

tag: Genotype

value: heterozygous affected

tag: gender

value: female


title: heterozygous affected male, biological rep3

description: Gene expression data from kidney of 36d old heterozygous affected male PKD/Mhm rats

source_name: heterozygous affected male PKD/Mhm rats

value: male


title: heterozygous affected female, biological rep2

source_name: heterozygous affected female PKD/Mhm rats


title: heterozygous affected female, biological rep1


title: wild type male, biological rep2

description: Gene expression data from kidney of 36d old wild type male PKD/Mhm rats

source_name: wild type male PKD/Mhm rats

value: wild type


title: wild type male, biological rep1


title: heterozygous affected male, biological rep2


title: Whole genome shotgun bisulfite sequencing of the H1 cell line; methylC-seq_h1_r2b

description:
Library name: methylC-seq_h1_r2b
EDACC Genboree Sample Page:
http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FUCSD%2FSAMPLE%2FEDACC.1106
EDACC Genboree Experiment Page:
http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FUCSD%2FEXPERIMENT%2FEDACC.989

****************
For data usage terms and conditions, please refer to:
http://www.drugabuse.gov/funding/funding-opportunities/nih-common-fund/epigenomics-data-access-policies
****************

data_processing:
**********************************************************************

ANALYSIS FILE NAME: GSM429322_UCSD.H1.Bisulfite-Seq.methylC-seq_h1_r2b.wig
ANALYSIS CENTER: EDACC
ANALYSIS ALIAS: methylC-seq_h1_r2b.hg19.level.2
ANALYSIS TITLE: Methylation Proportion Graphs of H1 Cell Line Bisulfite-Seq Data
ANALYSIS DESCRIPTION: Illumina Bisulfite-Seq read mappings from the H1 Cell Line, Library methylC-seq_h1_r2b were processed into graphs of methylation proportions. Methylation proportions were calculated as (methylated calls / (methylated calls + unmethylated calls)) for all CpGs covered by at least 4 reads. Reads from the + and - strands were combined for methylation proportion calculations.
ANALYSIS TYPE: ABUNDANCE_MEASUREMENT
EDACC Genboree Analysis Page: http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FEDACC%2FANALYSIS%2FEDACC.5120
DATA_ANALYSIS_LEVEL: 2
EXPERIMENT_TYPE: Bisulfite-Seq
GENOME_ASSEMBLY: NCBI Build GRCh37/UCSC Build hg19
SOFTWARE: In house programs and scripts
SOFTWARE_VERSION: NA
READ_EXTENSION: 0bp
TREATMENT_OF_IDENTICAL_ALIGNMENTS_OF_MULTIPLE_READS: If multiple reads map to the same start position on the + strand or stop position on the - strand, only a single read is retained.
GENOMIC_WINDOW: 2bp containing CpGs
TREATMENT_OF_REGIONS_PRONE_TO_MULTIPLE_ALIGNMENTS: None
RELEASE_NUMBER: Human Epigenome Atlas 2
BROWSER_TRACK_NAME: H1 BS 2b
BROWSER_TRACK_DESCRIPTION: UCSD H1 Cell Line Bisulfite-Seq Library methylC-seq_h1_r2b EA Release 2

QUALITY SCORES:
NUMBER_OF_Bisulfite-Seq_EXPERIMENTS_SCORED: 17
BISULFITE_CONVERSION_PERCENTAGE_BASED_ON_MAPPINGS: 97.28
BISULFITE_CONVERSION_PERCENTAGE_BASED_ON_MAPPINGS_PERCENTILE: 18

**********************************************************************

ANALYSIS FILE NAME: GSM429322_UCSD.H1.Bisulfite-Seq.combined.wig
ANALYSIS CENTER: EDACC
ANALYSIS ALIAS: methylC-seq_h1_r1a-methylC-seq_h1_r1b-methylC-seq_h1_r2a-methylC-seq_h1_r2b-methylC-seq_h1_r2c.hg19.level.2
ANALYSIS TITLE: Methylation Proportion Graphs of H1 Cell Line Bisulfite-Seq Data
ANALYSIS DESCRIPTION: Illumina Bisulfite-Seq read mappings from the H1 Cell Line combined libraries were processed into graphs of methylation proportions. Methylation proportions were calculated as (methylated calls / (methylated calls + unmethylated calls)) for all CpGs covered by at least 4 reads. Reads from the + and - strands were combined for methylation proportion calculations.
ANALYSIS TYPE: ABUNDANCE_MEASUREMENT
EDACC Genboree Analysis Page: http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FEDACC%2FANALYSIS%2FEDACC.5127
DATA_ANALYSIS_LEVEL: 2
EXPERIMENT_TYPE: Bisulfite-Seq
GENOME_ASSEMBLY: NCBI Build GRCh37/UCSC Build hg19
SOFTWARE: In house programs and scripts
SOFTWARE_VERSION: NA
READ_EXTENSION: 0bp
TREATMENT_OF_IDENTICAL_ALIGNMENTS_OF_MULTIPLE_READS: If multiple reads map to the same start position on the + strand or stop position on the - strand, only a single read is retained.
GENOMIC_WINDOW: 2bp containing CpGs
TREATMENT_OF_REGIONS_PRONE_TO_MULTIPLE_ALIGNMENTS: None
RELEASE_NUMBER: Human Epigenome Atlas 2
BROWSER_TRACK_NAME: H1 BS Combined
BROWSER_TRACK_DESCRIPTION: UCSD H1 Cell Line Bisulfite-Seq Combined Libraries EA Release 2
BISULFITE_CONVERSION_PERCENTAGE_BASED_ON_MAPPINGS: 97.1
SPECIAL NOTE: generated from combined data in GSM432685, GSM432686, GSM429321, GSM429322, GSM429323 (thus .wig files on these records are identical)

**********************************************************************

extract_protocol: Library construction protocol: Genomic DNA was purified from H1 human embryonic stem cells, sheared by sonication to 50-500 bp, ligated to methylated single-end Illumina adaptors, fragments of 140-210 bp isolated after agarose gel electrophoresis, bisulfite converted, amplified by PCR and sequenced using the Illumina Genome Analyzer II according to manufacturer's instructions

tag: molecule

value: genomic DNA

value: Thomson Laboratory

value: Cell Line

value: Replicate 1

value: embryonic stem cell

value: TeSR

value: Male

value: DNA Methylation

value: Qiagen DNeasy mini kit, performed as per manufacturer's instructions'

value: Diagenode Bioruptor

value: 20 cycles of:30 seconds on high power, 2 minutes off

value: 5 µg

value: A:5' P-GATCGGAAGAGCTCGTATGCCGTCTTCTGCTTG, B:5' ACACTCTTTCCCTACACGACGCTCTTCCGATCT

value: Standard Illumina genomic DNA library preparation ligation protocol (15 minutes at room temperature), except the adapters contained methylcytosine and were supplied by Illumina

value: Separation by electrophoresis on a 2% agarose gel followed by excision of 140-210 bp fragments

value: Standard Human Genetic Signatures MethylEasy Xceed bisulfite conversion kit protocol

value: 99.6% of cytosines converted based on shotgun sequencing of unmethylated lambda phage control spiked into original genomic DNA sample

value: One third of the 140-210 nt adapter-ligated, bisulfite converted DNA was used in each 50 µl PCR reaction

value: Stratagene Pfu Turbo Cx

value: 95˚C 2 min; 98˚C 30 sec, 4 cycles of 98˚C 15 sec, 60˚C 30 sec, 72˚C 4 min; 72˚C 10 min

value: 5' CAAGCAGAAGACGGCATACGAGCTCTTCCGATCT

value: 25 µM

value: Separation by electrophoresis on a 2% agarose gel followed by excision of 180-300 bp fragments.


title: Whole genome shotgun bisulfite sequencing of the H1 cell line; methylC-seq_h1_r2a

description:
Library name: methylC-seq_h1_r2a
EDACC Genboree Sample Page:
http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FUCSD%2FSAMPLE%2FEDACC.1106
EDACC Genboree Experiment Page:
http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FUCSD%2FEXPERIMENT%2FEDACC.1001

****************
For data usage terms and conditions, please refer to:
http://www.drugabuse.gov/funding/funding-opportunities/nih-common-fund/epigenomics-data-access-policies
****************

data_processing:
**********************************************************************

ANALYSIS FILE NAME: GSM429321_UCSD.H1.Bisulfite-Seq.methylC-seq_h1_r2a.wig
ANALYSIS CENTER: EDACC
ANALYSIS ALIAS: methylC-seq_h1_r2a.hg19.level.2
ANALYSIS TITLE: Methylation Proportion Graphs of H1 Cell Line Bisulfite-Seq Data
ANALYSIS DESCRIPTION: Illumina Bisulfite-Seq read mappings from the H1 Cell Line, Library methylC-seq_h1_r2a were processed into graphs of methylation proportions. Methylation proportions were calculated as (methylated calls / (methylated calls + unmethylated calls)) for all CpGs covered by at least 4 reads. Reads from the + and - strands were combined for methylation proportion calculations.
ANALYSIS TYPE: ABUNDANCE_MEASUREMENT
EDACC Genboree Analysis Page: http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FEDACC%2FANALYSIS%2FEDACC.5119
DATA_ANALYSIS_LEVEL: 2
EXPERIMENT_TYPE: Bisulfite-Seq
GENOME_ASSEMBLY: NCBI Build GRCh37/UCSC Build hg19
SOFTWARE: In house programs and scripts
SOFTWARE_VERSION: NA
READ_EXTENSION: 0bp
TREATMENT_OF_IDENTICAL_ALIGNMENTS_OF_MULTIPLE_READS: If multiple reads map to the same start position on the + strand or stop position on the - strand, only a single read is retained.
GENOMIC_WINDOW: 2bp containing CpGs
TREATMENT_OF_REGIONS_PRONE_TO_MULTIPLE_ALIGNMENTS: None
RELEASE_NUMBER: Human Epigenome Atlas 2
BROWSER_TRACK_NAME: H1 BS 2a
BROWSER_TRACK_DESCRIPTION: UCSD H1 Cell Line Bisulfite-Seq Library methylC-seq_h1_r2a EA Release 2

QUALITY SCORES:
NUMBER_OF_Bisulfite-Seq_EXPERIMENTS_SCORED: 17
BISULFITE_CONVERSION_PERCENTAGE_BASED_ON_MAPPINGS: 97.35
BISULFITE_CONVERSION_PERCENTAGE_BASED_ON_MAPPINGS_PERCENTILE: 24

**********************************************************************

ANALYSIS FILE NAME: GSM429321_UCSD.H1.Bisulfite-Seq.combined.wig
ANALYSIS CENTER: EDACC
ANALYSIS ALIAS: methylC-seq_h1_r1a-methylC-seq_h1_r1b-methylC-seq_h1_r2a-methylC-seq_h1_r2b-methylC-seq_h1_r2c.hg19.level.2
ANALYSIS TITLE: Methylation Proportion Graphs of H1 Cell Line Bisulfite-Seq Data
ANALYSIS DESCRIPTION: Illumina Bisulfite-Seq read mappings from the H1 Cell Line combined libraries were processed into graphs of methylation proportions. Methylation proportions were calculated as (methylated calls / (methylated calls + unmethylated calls)) for all CpGs covered by at least 4 reads. Reads from the + and - strands were combined for methylation proportion calculations.
ANALYSIS TYPE: ABUNDANCE_MEASUREMENT
EDACC Genboree Analysis Page: http://genboree.org/java-bin/project.jsp?projectName=XML%20Submissions%2FEDACC%2FANALYSIS%2FEDACC.5127
DATA_ANALYSIS_LEVEL: 2
EXPERIMENT_TYPE: Bisulfite-Seq
GENOME_ASSEMBLY: NCBI Build GRCh37/UCSC Build hg19
SOFTWARE: In house programs and scripts
SOFTWARE_VERSION: NA
READ_EXTENSION: 0bp
TREATMENT_OF_IDENTICAL_ALIGNMENTS_OF_MULTIPLE_READS: If multiple reads map to the same start position on the + strand or stop position on the - strand, only a single read is retained.
GENOMIC_WINDOW: 2bp containing CpGs
TREATMENT_OF_REGIONS_PRONE_TO_MULTIPLE_ALIGNMENTS: None
RELEASE_NUMBER: Human Epigenome Atlas 2
BROWSER_TRACK_NAME: H1 BS Combined
BROWSER_TRACK_DESCRIPTION: UCSD H1 Cell Line Bisulfite-Seq Combined Libraries EA Release 2
BISULFITE_CONVERSION_PERCENTAGE_BASED_ON_MAPPINGS: 97.1
SPECIAL NOTE: generated from combined data in GSM432685, GSM432686, GSM429321, GSM429322, GSM429323 (thus .wig files on these records are identical)

**********************************************************************

value: genomic DNA

value: Thomson Laboratory

value: Cell Line

value: Replicate 1

value: embryonic stem cell

value: TeSR

value: Male


title: wild type female, biological rep3

description: Gene expression data from kidney of 36d old wild type female PKD/Mhm rats

source_name: wild type female PKD/Mhm rats

value: wild type

value: female


title: wild type male, biological rep3

value: male


title: hESC-KLF4-1a

first: Bing

last: Ren

address: 9500 Gilman Drive

data_processing:
Base calling performed with Illumina Genome Analyzer Pipeline Software version 0.3
eland_multi
- Generated using ELAND
- ELAND parameters:
1.  Mapping length:  25 bases
2.  Maximum number of matches (0 mismatches, 1 mismatch, 2 mismatches): 250,250,250

bed (hg18)
- Generated using a custom perl script
- Input was the eland_multi file.
- Only reads mapping uniquely were retained.
- Read length is 36
- The score field contains the number of copies of the read

source_name: embryonic stem cells

tag: cell line

tag: chip antibody


title: heterozygous affected male, biological rep1

value: heterozygous affected


title: S2_DRSC_CG7878_RNAi-1

status: Public on Oct 09 2009

city: Farmington

first: Brenton

middle: R

last: Graveley

state: CT

address: 400 Farmington Avenue

department: Genetics and Genome Sciences

institute: University of Connecticut Health Center

description:
dsRNA treatment: CG7878
read length: 75
read length: 75
read length: 76
library_source: RNA
library_selection: RT and size selection

data_processing: Reads were aligned to the reference genome and a splice junction database to identify uniquely mapping reads and the data converted to SAM files and .bedgraph files which indicate the number of reads that map to each base in the genome.

growth_protocol: Schneider's medium + 10% heat-inactivated FCS

extract_protocol:
Oligo-dT selected, double stranded cDNA, Single Read Sequencing
TRIzol Dnase Rneasy

treatment_protocol: Cells were treated with dsRNA to the indicated gene


title: wild type female, biological rep2


title: wild type female, biological rep1


title: L2_combined_RNAseq_1, 2

status: Public on Sep 19 2009

city: Chicago

first: Kevin

middle: P.

last: White

state: IL

address: 900 E. 57th STR. 10th FL.

department: Institute for Genomics and Systems Biology

institute: University of Chicago

description: combined_RNAseq_1 and combined_RNAseq_2

data_processing: Reads were aligned to the flybase BDGPv5 reference genome using TopHat (http://tophat.cbcb.umd.edu/).  The bed file was generated by thresholding the bedgraph file (tag count>=1; minimum run length=50; maximum gap=50).

growth_protocol: y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. Embryos and larvae are collected using a filter mesh and a brush. They are rinsed with EWB and crushed in Trizol reagent. Pupae are collected on the side of a vial with a brush. Adult animals are sorted by sex on a CO2 pad and, rinsed before crushed.

extract_protocol:
Matching total RNA was collected from each time-point of this study in TRIzol reagent and isolated according to the manufacturer instructions. DNAse I treated RNA is then purified and concentrated using the RNeasy MinElute Cleanup Kit. PolyA RNAs are then purified using the micropolyA purist Kit from Ambion and converted into single stranded DNA after Reverse Transcription using random hexamers. The second strand synthesis is then carried out by adding to the reaction the RNAseH (Invitrogen #18021014) and DNA Polymerase II (NEB #M0209S) enzymes. At this stage, the double stranded DNA is then cleaned up on Qiagen Quiaquick columns and the ends are repaired by using the T4DNA Polymerase, Klenow Fragment and T4 PNK enzymes.
After another round of purififcation, an A residue is added by the use of the Klenow [3'>5' exo-] enzyme and again purified on Quiaquick columns. Adapters from Illumina for LM-PCR are then ligated to the end of the DNA molecules. The product of the reaction is then run on an Agarose gel (2% NuSieve) and a band corresponding to 100 bp is then extracted and purified. 20 cycles of PCR reaction using the phusion polymerase (Finnzyme F-530S) enzyme and the Illumina oligos are then performed and the product is again purified on gel. Solexa sequencing is then performed on Genome Analyzer with standard Illumina 36 cycles reaction kit.

treatment_protocol: No Treatment

tag: development stage


title: RNAPolII_ChIPSeq

status: Public on Sep 15 2009

city: Nijmegen

first: Gert Jan

last: Veenstra

address: Geert Grooteplein 28

department: Molecular Developmental Biology

country: Netherlands

institute: Radboud University

description:
Chromatin IP against RNA PolII
Processed data file: RNAPII.bed
Raw data file: 30K5HAAXX_1_sequence.fastq
Raw data file: 30K6UAAXX_2_sequence.fastq
Raw file type: fastq

data_processing: 32 bp reads were aligned to the JGI Xenopus tropicalis genome assembly (version 4.1) and filtered for uniqueness.

organism: Xenopus tropicalis

source_name: whole embryo

growth_protocol: Xenopus tropicalis embryos were obtained from a natural mating procedure after human chorion gonadotropin injection, dejellied in 2% cysteine and collected at the indicated stage

extract_protocol: Chromatin (300-600 embryos) was fixed in 1% formaldehyde, sheared using a Branson sonifier, immunoprecipitated using 1-5 µg of antibody, washed, decrosslinked, DNA was purified and sequenced using a Illumina Genome Analyzer according to the manufacturer.

tag: developmental stage

value: 11-12 embryo

tag: antibody

value: RNAPII (Diagenode, AC-055-100)


title: RNASeq

description:
PolyA selected, reverse transcribed total RNA
Processed_data_file: RNASeq.bed
Processed_data_file: RNASeq_expression_level.txt
Raw data file: 20FDNAAXX_1_sequence.fastq
Raw data file: 20FDNAAXX_4_sequence.fastq
Raw data file: 20FGLAAXX_2_sequence.fastq
Raw data file: 20FDNAAXX_2_sequence.fastq
Raw data file: 20FE3AAXX_8_sequence.fastq
Raw data file: 20FGLAAXX_3_sequence.fastq
Raw data file: 20FDNAAXX_3_sequence.fastq
Raw data file: 20FGLAAXX_1_sequence.fastq
Raw data file: 20FGLAAXX_4_sequence.fastq
Raw file type: fastq

data_processing: RNA-seq reads were mapped to JGI FilteredModels genes to determine a normalized expression level.

molecule: polyA RNA

growth_protocol: Xenopus tropicalis embryos were obtained from a natural mating procedure after human chorion gonadotropin injection, dejellied in 2% cysteine and collected at the indicated stage.

extract_protocol: Total RNA was isolated using Trizol and the Qiagen RNeasy Kit. Subsequently, polyadenylated RNA was selected with the Oligotex mRNA kit (Qiagen) and the mRNA selection was repeated to improve the removal of rRNA. cDNA was prepared with random hexamer primers using Superscript III (Invitrogen) and second strand cDNA was made with DNA polymerase I, DNA ligase and T4 DNA polymerase.

value: 10-13 embryo


title: cells, in vitro, rep1

status: Public on Sep 23 2009

city: Cambridge

first: Corbin

middle: Elizabeth

last: Meacham

address: 40 Ames Street

department: Koch Institute for Integrative Cancer Research

institute: MIT

description:
shRNAs were PCR amplified from genomic DNA using a common primer set. Amplified shRNAs were sequenced to assess hairpin representation
PCR amplification protocol: shRNAs were PCR amplified from genomic DNA using the following primers, which include adaptors for 454 sequencing: 5'GCCTCCCTCGCGCCATCAGCAGAAGGCTCGAGAAGGTATATTGCTGTTGACAGTGAGCG 3' 5'GCCTTGCCAGCCCGCTCAGCTAAAGTAGCCCCTTGAATTCCGAGGCAGTAGGCA 3'. The PCR product of approximately 180 basepairs was isolated and gel extracted. This PCR product was sent to 454 Lifesciences for sequencing.

data_processing: Reads were aligned to our shRNA library using blast and the best blast hit was considered a match. Reads that were truncated at the start of the unique shRNA sequence were excluded from this analysis. To normalize for the total number of reads in each sample, the number of reads for each unique shRNA was divided by the total number of reads for that sample. This value represents the fraction of the total number of reads. To calculate the fold change for each shRNA in the three in vitro and three in vivo samples, relative to acutely infected cells, the fraction value for a given shRNA in an experimental sample was divided by the fraction value for that shRNA in the input sample (acutely infected cells).

source_name: lymphoma cells

growth_protocol: normal growth conditions in vitro or in vivo

extract_protocol: Cells were digested in lysis buffer (10mM Tris pH 8.0, 100 mM NaCl, 10 mM EDTA pH 8.0, .5% SDS) and proteinase K (1 mg/mL), and DNA was extracted with isopropanol.

treatment_protocol: GFP-sorted, infected cells were maintained in culture for two weeks

tag: cell type

value: Eu-myc Arf-/- lymphoma cells

tag: protocol

value: transduced with shRNA library, maintained in culture

tag: host mouse strain


title: cells, tumor, rep1

treatment_protocol: GFP-sorted, infected cells were tail-vein injected into syngeneic recipient mice. After approximately two weeks, at the time of disease presentation, tumors were harvested

value: transduced with shRNA library, tail-vein injected into three syngeneic recipient mice, tumors harvested


title: cells, in vitro, rep2


title: cells, acutely infected, rep1

description:
Reference sample.
shRNAs were PCR amplified from genomic DNA using a common primer set. Amplified shRNAs were sequenced to assess hairpin representation
PCR amplification protocol: shRNAs were PCR amplified from genomic DNA using the following primers, which include adaptors for 454 sequencing: 5'GCCTCCCTCGCGCCATCAGCAGAAGGCTCGAGAAGGTATATTGCTGTTGACAGTGAGCG 3' 5'GCCTTGCCAGCCCGCTCAGCTAAAGTAGCCCCTTGAATTCCGAGGCAGTAGGCA 3'. The PCR product of approximately 180 basepairs was isolated and gel extracted. This PCR product was sent to 454 Lifesciences for sequencing.

treatment_protocol: Cells were infected with a GFP-tagged, pooled shRNA library and were GFP sorted 48 hours after infection. Immediately after sorting, cells were collected to serve as a reference sample.

value: transduced with shRNA library, collected immediately to serve as a reference sample


title: cells, tumor, rep3


title: cells, tumor, rep2


title: Input_control

city: University Park

first: Ross

last: Hardison

state: PA

address: 303 Wartik Lab

institute: Pennsylvania State University

description: Induced G1E ER4 cells

data_processing:
Alignment: Sequence reads were obtained and mapped to the mouse (Feb. 2006 mm8) genomes using the Illumina Genome Analyzer Pipeline. All reads mapping with two or fewer mismatches were retained.
Peaks: Peak detection was performed with the Model-based Analysis of ChIP-Seq (MACS) algorithm (http://liulab.dfci.harvard.edu/MACS/)

source_name: Chromatin input as control

growth_protocol: G1E-ER4 cells were cultured as described previously. Weiss MJ, Yu C, Orkin SH. Erythroid-cell-specific properties of transcription factor GATA-1 revealed by phenotypic rescue of a gene-targeted cell line. Mol Cell Biol. 1997;17: 1642-1651.

extract_protocol: Lysates were clarified from sonicated nuclei and GATA1-DNA complexes were isolated with antibody.  Libraries were prepared according to Illumina's instructions accompanying the ChIP Sequencing of DNA Kit (Part# 11257047). 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.

treatment_protocol: G1E-ER4 cells growing in log phase were induced for 24 hours with 20nM estradiol.

value: G1E ER4


title: H3K27me3_ChIPSeq

description:
Chromatin IP against H3K27me3
Processed_data_file: H3K27me3.bed
Processed_data_file: H3K27me3_enriched.bed
Raw data file: 202KJAAXX_1_sequence.fastq
Raw data file: 204BGAAXX_6_sequence.fastq
Raw data file: 204BGAAXX_8_sequence.fastq
Raw data file: 204BGAAXX_5_sequence.fastq
Raw data file: 204BGAAXX_7_sequence.fastq
Raw file type: fastq

data_processing: 32 bp reads were aligned to the JGI Xenopus tropicalis genome assembly (version 4.1) and filtered for uniqueness. Enriched regions for H3K4me3 and H3K27me3 were determined by a sliding window approach of 500bp using a threshold of threshold (determined by a FDR approach) of 9 and 12 reads for H3K4me3 and H3K27me3, respectively.


title: plasmid library, rep1

description:
shRNAs were PCR amplified from plasmid DNA using a common primer set. Amplified shRNAs were sequenced to assess hairpin representation
PCR amplification protocol: shRNAs were PCR amplified from plasmid DNA using the following primers, which include adaptors for 454 sequencing: 5'GCCTCCCTCGCGCCATCAGCAGAAGGCTCGAGAAGGTATATTGCTGTTGACAGTGAGCG 3' 5'GCCTTGCCAGCCCGCTCAGCTAAAGTAGCCCCTTGAATTCCGAGGCAGTAGGCA 3'. The PCR product of approximately 180 basepairs was isolated and gel extracted. This PCR product was sent to 454 Lifesciences for sequencing.

organism: synthetic construct

source_name: pooled retroviral shRNA library

tag: library

value: retroviral plasmid library (MSCV-based retroviral vector) of 2250 shRNAs


title: cells, in vitro, rep3


title: GATA1_ChIPSeq

description: Induced G1E ER4 cells

source_name: Chromatin IP against GATA1


title: H3K4me3_ChIPSeq

description:
Chromatin IP against H3K4me3
Processed_data_file: H3K4me3.bed
Processed_data_file: H3K4me3_enriched.bed
Raw data file: 20419AAXX_7_sequence.fastq
Raw data file: 204BGAAXX_2_sequence.fastq
Raw data file: 204BGAAXX_4_sequence.fastq
Raw data file: 204BGAAXX_1_sequence.fastq
Raw data file: 204BGAAXX_3_sequence.fastq
Raw file type: fastq


title: Pupae_combined_RNAseq_1,2

status: Public on Sep 12 2009

description: combined_RNAseq_1 and combined_RNAseq_2

source_name: Pupae

value: Pupae


title: Control-KD mRNA-Seq

status: Public on Sep 13 2009

first: Xinshu

last: Xiao

address: 77 Mass Ave. MIT, 68-211

department: Biology

data_processing: short reads mapping to human genome hg18 using Eland

source_name: 293T cells

growth_protocol: 293T cells were cultured with D-MEM medium supplemented with 10% fetal bovine serum.

extract_protocol: mRNA is purified from total RNA with oligo-dT magnetic beads. cDNA is synthesized with random primer priming. ds-cDNA is sequenced using Illumina Genome Analyzers as recommended by the manufacturer.

treatment_protocol: Day 0: plate cells in 10cm dishes. Day 1: transfect 20nM siRNA using Dharmafect 1 (Dharmacon) as transfection reagent. Day 2: transfect 50nM siRNA using Dharmafect 1 (Dharmacon) as transfection reagent. Day 4: harvest cells. Transfections were conducted using protocols suggested by the manufacturer of the transfection reagent.

tag: cell l ine


title: hnRNP H-KD mRNA-Seq


title: L3_combined_RNAseq_1,2

description: combined_RNAseq_1 and combined_RNAseq_2


title: E8-12_INPUT_CBP

data_processing: Reads were aligned to the flybase BDGPv5 reference genome, and the fragment count at any given position (10-bp resolution) was estimated as the number of uniquely aligned reads oriented towards it and within 300Â bp. Enriched intervals were identified by comparison of the mean fragment count in 1-kb windows against a sample-specific expected distribution estimated by randomization (H3K4me3, H3K27me3), or using a supervised Hidden Markov Model (H3K36me3, H3K9me3, H4K20me3).

growth_protocol: 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder.2. For a 8-12 hours egg laying, new plates are added after a 2hours pre-egglaying at 8 pm and flies are allowed to lay eggs for 4 hours.3. Embryos are collected at 7.45 am using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 8 am in 1.8% formaldehyde at room temperature for 15 min.

extract_protocol: Drosophila samples are crushed in presence of 1.8% of formaldehyde for 15 minutes. After washing, the material is sonicated for 15 minutes to solubilize the chromatin. After centrifugation, the supernatant containing soluble chromatin is collected for IPs. Specific antibodies are added to the chromatin extract while one aliquot will be treated for DNA extraction corresponding to the reference samples. ChIP are carried overnight at room temperature. Protein A Sepharose beads are used to pull down the chromatin bound antibodies. After washes and finally elution, the DNA corresponding to the IPed material is extracted and purified. The DNA is directly used for Solexa library preparation.

tag: reference

value: INPUT


title: E8-12_CBP_ChIPSeq_1

description: corresponding input record: GSM451802

value: CBP

tag: antibody provider

value: Dr. Mattias Mannervik in Stockholm University


title: E4-8_combined_RNAseq_1,2

description: combined_RNAseq_1 and combined_RNAseq_2


title: E16-20_combined_RNAseq_1,2

description: combined_RNAseq_1 and combined_RNAseq_2


title: AdultFemale_combined_RNAseq_1, 2

description: combined_RNAseq_1 and combined_RNAseq_2

source_name: AdultFemale

value: AdultFemale


title: E0-4_combined_RNAseq_1,2

description: combined_RNAseq_1 and combined_RNAseq_2


title: E12-16_combined_RNAseq_1, 2

description: combined_RNAseq_1 and combined_RNAseq_2

data_processing: Reads were aligned to the flybase BDGPv5 reference genome using TopHat (http://tophat.cbcb.umd.edu/).  The bed file was generated by thresholding the wiggle file (tag count>=1; minimum run length=50; maximum gap=50).


title: E20-24_combined_RNAseq_1, 2

description: combined_RNAseq_1 and combined_RNAseq_2


title: E8-12_combined_RNAseq_1,2

description: combined_RNAseq_1 and combined_RNAseq_2


title: L1_combined_RNAseq_1,2

description: combined_RNAseq_1 and combined_RNAseq_2


title: AdultMale_combined_RNAseq_1, 2

description: combined_RNAseq_1 and combined_RNAseq_2

source_name: AdultMale

value: AdultMale


title: L2_INPUT_PolII

growth_protocol: 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a collection of second instar larvae (L2), new plates are added after a 2hours pre-egglaying at 9 am and flies are allowed to lay eggs for 24 hours. Plates are then kept at 25C for another 24 hours.3. L2 larvae are collected at 8.45 am using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 9 am in 1.8% formaldehyde at room temperature for 15 min.


title: IgG_ChIA-PET (454)

status: Public on Sep 11 2009

city: Singapore

first: Yijun

last: Ruan

state: Singapore

address: 60 Biopolis Street, #02-01 Genome, Singapore 138672

department: Research Computing Group

country: Singapore

institute: Genome Institute of Singapore

description: A control ChIA-PET library against IgG (mock ChIP) in MCF-7 human breast cancer cells in estrogen-treated condition.

data_processing: Mapping analysis was performed by Compressed Suffix Arrays and results were further processed using the ChIA-PET pipeline software.

source_name: MCF-7 cells

growth_protocol: MCF-7 cells were grown to at least 80% confluence in DMEM/F12 (Invitrogen/Gibco) supplemented with 5% FBS (Invitrogen/Gibco), penicillin (Invitrogen), streptomycin (Invitrogen), and gentamycin (Invitrogen).

extract_protocol: We used 1% formaldehyde to crosslink the cells, and sonication to break the chromatin fibers.  Antibodies were used to enrich chromatin fragments by ChIP. After this either ChIA-PET or ChIP-Seq was performed.

treatment_protocol: In preparation for the 17 beta-estradiol (“estrogen”, E2, Sigma) treatment, cells were grown in hormone-free media: they were washed with PBS and incubated in phenol red-free medium (Invitrogen/Gibco) supplemented with 5% charcoal-dextran stripped FBS (Hyclone), penicillin, streptomycin, gentamycin, and L-glutamine (Invitrogen) for a minimum of 72 hours. Hormone-depleted cells were treated with estrogen to a final concentration of 100 nM for 45 min before the ChIP procedure.

value: ChIA-PET


title: ERα_ChIP-PET (454)

description: A control ChIP-PET library against ERα in MCF-7 human breast cancer cells in estrogen-treated condition.

value: ChIP-PET


title: S2_DRSC_CG1987_RNAi-1

description:
dsRNA treatment: CG1987
read length: 75
read length: 75
library_source: RNA
library_selection: RT and size selection


title: L2_PolII_ChIPSeq_1

description: corresponding input record: GSM451800

value: PolII

tag: antibody manufacturer

value: Covance

tag: antibody catalog number

tag: antibody lot number


title: H3K4me3_ChIP-Seq_SCS805 (Illumina)

description: H3K4me3 ChIP-Seq in MCF-7 human breast cancer cells in estrogen-treated conditions. From SCS773 and SCS805 merged together.

data_processing: Mapping analysis was performed by ELAND and results were further processed using the ChIA-PET pipeline software.

value: ChIP-Seq

tag: antibody target


title: H3K4me3_ChIP-Seq_SCS773 (Illumina)


title: RNAPII_ChIP-Seq_IHM035 (Illumina)

description: RNAPII ChIP-Seq in MCF-7 human breast cancer cells in estrogen-treated conditions. From IHM034, IHM035, and IHM036 merged together

value: RNA Polymerase II (RNAPII)


title: RNAPII_ChIP-Seq_IHM034 (Illumina)


title: ERα_ChIA-PET_Replicate2 (Illumina)

description: 1 biological replicate of a ChIA-PET library against ERα in MCF-7 human breast cancer cells in estrogen-treated condition.


title: ERα_ChIA-PET_Replicate1 (Illumina)


title: S2-DRSC small RNAs BioRep-2 TechRep-3

status: Public on Sep 10 2009

city: Woodbury

first: Chris

last: Zaleski

state: NY

address: 500 Sunnyside Blvd

institute: Cold Spring Harbor Labs

description:
20-200 ntds
RNA Seq
RNAs 20-200 n'tds
Stranded, single-ended, 5' bias

data_processing: Illumina fastq processed into “sam” format

tag: biorep

tag: techrep


title: S2-DRSC small RNAs BioRep-2 TechRep-2

tag: cell line


title: S2-DRSC small RNAs BioRep-2 TechRep-1


title: root_control_1

city: Canberra

first: Andrew

last: Spriggs

state: ACT

address: GPO Box 1600

department: Bioinformatics Group

country: Australia

institute: CSIRO Plant Industry

description:
High-throughput sequencing of Arabidopsis small RNA
processed data file (complete): uniqueSeqs-DM1.txt

data_processing: Using perl scripts, raw reads from fastq files were trimmed of any 3' adapter sequence present. Identical trimmed sequences were collated into a single row, with occurrences (number of reads) tallied. The Solexa read quality string with the highest overall quality out of a group of reads with identical trimmed sequences was retained to represent that group.  Sequences reads less than 18 basepairs long after 3' adapter trimming were rejected.

organism: Arabidopsis thaliana

source_name: whole root tissue

growth_protocol: Seeds were surface sterilised in 0.5% (w/v) sodium hypochlorite. Seeds were sown on 100 mm petri plates containing a modified Murashige-Skoog (MS) medium (Murashige and Skoog 1962) containing 3% sucrose (w/v) and 0.8% (w/v) agar and were subjected to at least 24 h of cold treatment in the dark (4°C) to break dormancy. Each plate contained approximately 40 seeds and grown at 21°C ± 1°C under cool-white fluorescent lights (120 μM m–2 s–1) with a 16 h photoperiod

extract_protocol: Frozen root tissue was ground to a fine white powder using a mortar and pestle. Total RNA extraction was performed using TRI Reagent (Molecular Research Center Inc., Cincinnati, Ohio, USA) following manufacturer's instructions.

treatment_protocol: Low-oxygen assays were performed as outlined in Christianson et al. 2009. Fifteen 2.5 week old plants were transferred to 100 mm petri plates containing 15 mls of MS liquid media supplemented with 3% sucrose one day prior to treatment. Immediately prior to treatment plants were transferred to liquid media which had been sparged with 0.1% O2 and then placed in 3.5L anaerobic chambers (Oxoid, Adelaide, SA, Australia) and purged with 0.1% O2, 99.9% N2 at a flow rate of ~5L/min for 15 min. The plants were left in 0.1% O2, in the dark with gentle shaking for a designated period. Control plants were subjected to the same treatment except that they were not transferred to 0.1% O2 sparged media, and the chambers were not sparged with 0.1% O2. Immediately after the treatment, plants were removed from the chamber and a scalpel used to isolate root tissue, which was then placed in liquid nitrogen and stored at ‑80ºC.

tag: ecotype

tag: stress

value: control

tag: age

value: 2.5 weeks old


title: S2-DRSC small RNAs BioRep-1 TechRep-4


title: S2-DRSC small RNAs BioRep-1 TechRep-3


title: S2-DRSC small RNAs BioRep-1 TechRep-2


title: Small RNAs from Petunia cosuppressed flower

status: Public on Sep 28 2009

city: Udine

first: Emanuele

last: De Paoli

state: UDINE

address: via delle Scienze 206

department: Agricultural, Food, Environmental and Animal Sciences

country: Italy

institute: University of Udine

description: Total RNA was submitted to Illumina (Hayward, CA, http://www.illumina.com) for small RNA library construction using approaches described in (Lu et al., 2007) with minor modifications. Low molecular weight RNA was gel-sized in the ~18-33 nt range. The small RNA library was sequenced with the Sequencing-by-synthesis (SBS) technology by Illumina.

data_processing: Small RNA reads were analyzed after removal of 3' adapter (5' -TCGTATGCCGTCTTCTGCTTG) and filtering of 5' adapter (5'-GTTCAGAGTTCTACAGTCCGACGATC) or 3' adapter artefacts.

organism: Petunia x hybrida

source_name: Cosuppressed flower

growth_protocol: Young leaves of petunia line V26 were co-cultivated with disarmed Agrobacterium tumefaciens LBA4404 harboring the complete coding region of CHS-A gene under the control of a 2x35S promoter and the nopaline synthase (NOS) terminator (pGSA2644). Transformants were selected and regenerated as described in Jorgensen et al (R.A. Jorgensen, P.D. Cluster, J. English , Q Que, C.A. Napoli, Plant Mol Biol 31, 957-73 (1996)) except that rooting of adventitious shoot was performed with 100 µg/ml kanamycin sulfate. After rooting, plantlets were transplanted to soil and grown in a greenhouse as described in Napoli and Ruehle (C.A. Napoli and J. Ruehle, J. Hered 87, 371–377 (1996)).

extract_protocol: Total RNA was extracted from pre-anthesis petals of silenced petunia plants (Petunia hybrida V26) with the Plant RNA Purification Reagent (Invitrogen). Quality of total RNA was evaluated by agarose gel electrophoresis, micro-electrophoresis (Agilent 2100 Bioanalyzer, Ambion) and O.D. measurements (Nanodrop ND-1000).

tag: strain

value: V26 transformed with one copy of the complete coding region of Chalcone synthase A (CHS-A) gene under the control of a 2x35S promoter and the nopaline synthase (NOS) terminator (pGSA2644); Pre-anthesis petal tissue


title: S2-DRSC small RNAs BioRep-1 TechRep-1


title: smallRNA-K.polysporus-WT

city: San Francisco

first: David

middle: Eric

last: Weinberg

address: 600 16th Street, GH S572F

institute: University of California, San Francisco

description: Small RNA sequencing, 18-30 nt size selection.

data_processing: After removing the adaptor sequences, reads representing the small RNAs were collapsed to a non-redundant set, and 14–30-nt sequences were mapped to the appropriate genome, allowing no mismatches and recovering all hits.  The processed files are collapsed to unique sequences (1 line per sequence); the second (numeric) value in each column is the number of times each unique sequence was sequenced.

organism: Vanderwaltozyma polyspora

source_name: log-phase liquid culture, WT

growth_protocol: Liquid cultures were grown at 25ºC (S. castellii and K. polysporus) or 30ºC (S. bayanus, S. cerevisiae and C. albicans) to log-phase.

extract_protocol: Total RNA was isolated using hot phenol from log-phase YPD cultures.  Small-RNA cDNA libraries were prepared as described in Grimson et al. (2008).

value: KpolWT


title: smallRNA-C.albicans-WT

organism: Candida albicans


title: Kc167_PolyA+ BioRep-1 TechRep-6

description:
>200 ntds
Poly-A+ RNA Seq
RNAs >200 n'tds
Stranded, single-ended, 5' bias


title: Kc167_PolyA+ BioRep-1 TechRep-4


title: Kc167_PolyA+ BioRep-1 TechRep-8


title: Kc167_PolyA+ BioRep-1 TechRep-7


title: root_low oxygen_1

description:
High-throughput sequencing of Arabidopsis small RNA
processed data file (complete): uniqueSeqs-DM2.txt

value: low oxygen


title: Kc167_PolyA+ BioRep-1 TechRep-3


title: smallRNA-S.bayanus-WT

organism: Saccharomyces bayanus


title: RNAPII_ChIP-Seq_IHM036 (Illumina)


title: Kc167_PolyA+ BioRep-1 TechRep-2


title: smallRNA-S.castellii-ago1KO

organism: Naumovozyma castellii

source_name: log-phase liquid culture, ∆ago1


title: smallRNA-S.castellii-dcr1KO

source_name: log-phase liquid culture, ∆dcr1


title: S2-DRSC small RNAs BioRep-2 TechRep-4


title: smallRNA-S.castellii-Input

source_name: log-phase liquid culture, input


title: ERα_ChIA-PET_Replicate1 (454)


title: Kc167_PolyA+ BioRep-1 TechRep-1


title: smallRNA-S.castellii-WT


title: Kc167 small RNAs BioRep-2 TechRep-2


title: cde1IP

status: Public on Oct 07 2009

city: Groningen

first: Eugene

last: Berezikov

address: Antonius Deusinglaan 1

department: European Research Institute for the Biology of Ageing

institute: University Medical Center Groningen

description: Small RNA sequence data

data_processing: 3' adapter sequences were trimmed and inserts longer than 18 nt were mapped to the C. elegans genome (ce4).

extract_protocol:
Imunoprecipitation of protein complexes was preformed by preclearing worm extracts with 20 ml of Protein A/G PLUS-Agarose (Santa Cruz Biotechnologies) agarose beads per 5 mg of total protein for 1 h at 4°C. The cleared extract was then incubated with 15 to 25 mg of antibody for 5mg of total protein for 1h at 4°C. 50 ml of Protein A/G PLUSAgarose beads were added, and the mixture incubated for 1 hour. The agarose beads were then washed three times with ice-cold lysis buffer. RNA was eluted from the Protein A/G PLUS-Agarose beads by extraction with TRI Reagent (MRC Reagents, Inc). 500 ml of TRI Reagent were added to 100 ml of beads and RNA was extracted as described in the manufacturer's protocol. Typical IPs for small RNA cloning were performed on 80 to 100 mg of total protein per sample to be cloned.
RNA was resolved in a 15% polyacrylamide 7M Urea Gel, along with 2 µmol of RNA standards (18- and 26-nt) in separate lanes. Ethidium Bromide Staining was used to visualize the RNA standards. A gel fragment was excised in the sample lanes between the two standards. RNA was eluted from the gel fragment in a [0.3M NaCl-TE (pH7.5)] solution overnight and ethanol-precipitated using 10 mg of glycogen as the carrier. Gel purified RNA was treated with 1 Unit/µl of Alkaline Phosphatase, Calf Intestine (NEB) in 100 mM NaCl, 50 mM Tris-HCl, 10 mM MgCl2, 1 mM Dithiothreitol, pH 7.9 @ 25°C and 1 Unit/µl SuperRNAseIn (Ambion) for 1 hour at 37 °C. After phenol extraction, the gel purified RNA and 1 µM of each standard were incubated with 20 µM of 3'-end linker, 1 Unit of SuperRNaseIN, 10% DMSO and 3 Units T4 RNA ligase (Takara) in 10 µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2, 6mg/mL BSA, 10mM DTT). The 3' ligated products were gel purified and treated with 1 Unit/µl Polynucleotide Kinase in 1x Polynucleotide Kinase buffer (70 mM Tris-HCl, 10 mM MgCl2, 5 mM Dithiothreitol, pH 7.6 @ 25°C), 2 mM ATP, 1 Unit/µl SuperRNAseIN. After phenol extraction, RNAs were incubated in the presence of 30 µM of 5' adapter oligonucleotide, 1 Unit SuperRNaseIN (Ambion) and 1.5 Unit of T4 RNA ligase in ligation buffer (50 mM Tris-HCI (pH7.5), 10 mM MgCI2, 10 mM DTT, 1 mM ATP) and 10% Dimethyl sulfoxide. The ligated products were gel purified as described above and reverse transcribed in a standard 50 µl reaction (SuperScript III, Invitrogen) and subjected to RNaseH treatment. The cDNA was amplified by PCR and purified in a 10% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing platform (Illumina, Inc.).

tag: Total small RNA from a CSR-1 IP from cde-1 mutant animals

tag: tissue

value: whole animal

tag: developmental stage

value: adult

tag: genome/variation

value: cde-1 mutant

tag: ip antibody


title: Kc167 small RNAs BioRep-2 TechRep-1


title: mRNA-dcr1KO-2

description: Small RNA sequencing by mRNA fragmentation, 5' and 3' adaptor ligation, and RT-PCR

data_processing: The first 25 nt of each 36-nt read were isolated and collapsed into a non-redundant set of 25-nt sequences with occurrence counts.  Sequences were mapped to the reference genome, allowing no mismatches and recovering all hits.  The processed files are collapsed to unique sequences (1 line per sequence); the second (numeric) value in each column is the number of times each unique sequence was sequenced.

growth_protocol: Two biological replicates of yeast strains DPB005 (WT), DPB007 (∆ago1), and DPB009 (∆dcr1) were grown at 25°C in YPD to OD600 0.6–0.8.

extract_protocol: Total RNA isolated using hot phenol was treated with DNaseI.  Poly-(A)+ mRNA was purified from total RNA using magnetic oligo-dT beads, and then fragmented by alkaline hydrolysis.  RNA fragments (25–45 nt) were gel-purified and 3´-dephosphorylated with T4 PNK in MES-NaOH buffer.  After phenol extraction and precipitation, RNA was ligated to pre-adenylated adaptor DNA.  Gel-purified ligation products were 5´-phosphorylated with T4 PNK.  After phenol extraction and precipitation, RNA was ligated to a 5´-adaptor RNA, gel-purified, converted to cDNA, amplified, and sequenced.

tag: rnai deletion


title: mRNA-dcr1KO-1


title: Kc167 small RNAs BioRep-1 TechRep-1


title: mRNA-ago1KO-2


title: mRNA-ago1KO-1


title: N2YNG

description: Small RNA sequence data

extract_protocol:
Total RNA was size selected on acrylamide gels (15-30 bases). The RNAs were treated with Tobacco Acid Pyrophosphatase and subsequently poly(A)-tailed using poly(A) polymerase followed by ligation of a RNA adaptor to the 5'-phosphate of the small RNAs. First-strand cDNA synthesis was performed using an oligo(dT)-linker primer and M-MLV-RNase H- reverse transcriptase. The cDNA was PCR-amplificated for 16 cycles according to the instructions of Illumina/Solexa. Equal amounts of bar-coded cDNA were mixed. The 125-140 bp fraction of the cDNA pool was obtained by fractionation of the cDNA on a preparative 6% polyacrylamide (PAA) gel. The eluted cDNA was finally purified using the Macherey & Nagel NucleoSpin Extract II kit and was sent for sequencing on an Illumina/Solexa platform. The following adaptor sequences with NNNN representing the barcodes flank the cDNA inserts 5'-end: 5'-
GCCTCCCTCGCGCCATCAGCTNNNNGACCTTGGCTGTCACTCA-3'; 3'-end:
5'-
GCCTTGCCAGCCCGCTCAGACGAGACATCGCCCCGCTTTTTTTTTTTTTTTT
TTTTTTTTT-3'.

tag: Total RNA from young adult wild-type adults

value: young adult

value: wild type


title: smallRNA-S.castellii-Ago1IP

source_name: log-phase liquid culture, Ago1IP


title: CDE1OLD

description: Small RNA sequence data

source_name: CDE1 aged adults

tag: Total RNA from old cde-1 mutant adults

value: aged adult


title: Cl.8 small RNAs BioRep-2 TechRep-4


title: Cl.8 small RNAs BioRep-2 TechRep-3


title: Cl.8 small RNAs BioRep-2 TechRep-2


title: Kc167 small RNAs BioRep-1 TechRep-2


title: Kc167_PolyA+ BioRep-1 TechRep-5


title: cde1input

description: Small RNA sequence data

extract_protocol:
Imunoprecipitation of protein complexes was preformed by preclearing worm extracts with 20 ml of Protein A/G PLUS-Agarose (Santa Cruz Biotechnologies) agarose beads per 5 mg of total protein for 1 h at 4°C. The cleared extract was then incubated with 15 to 25 mg of antibody for 5mg of total protein for 1h at 4°C (no antibody added for "input" samples). 50 ml of Protein A/G PLUSAgarose beads were added, and the mixture incubated for 1 hour. The agarose beads were then washed three times with ice-cold lysis buffer. RNA was eluted from the Protein A/G PLUS-Agarose beads by extraction with TRI Reagent (MRC Reagents, Inc). 500 ml of TRI Reagent were added to 100 ml of beads and RNA was extracted as described in the manufacturer's protocol. Typical IPs for small RNA cloning were performed on 80 to 100 mg of total protein per sample to be cloned.
RNA was resolved in a 15% polyacrylamide 7M Urea Gel, along with 2 µmol of RNA standards (18- and 26-nt) in separate lanes. Ethidium Bromide Staining was used to visualize the RNA standards. A gel fragment was excised in the sample lanes between the two standards. RNA was eluted from the gel fragment in a [0.3M NaCl-TE (pH7.5)] solution overnight and ethanol-precipitated using 10 mg of glycogen as the carrier. Gel purified RNA was treated with 1 Unit/µl of Alkaline Phosphatase, Calf Intestine (NEB) in 100 mM NaCl, 50 mM Tris-HCl, 10 mM MgCl2, 1 mM Dithiothreitol, pH 7.9 @ 25°C and 1 Unit/µl SuperRNAseIn (Ambion) for 1 hour at 37 °C. After phenol extraction, the gel purified RNA and 1 µM of each standard were incubated with 20 µM of 3'-end linker, 1 Unit of SuperRNaseIN, 10% DMSO and 3 Units T4 RNA ligase (Takara) in 10 µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2, 6mg/mL BSA, 10mM DTT). The 3' ligated products were gel purified and treated with 1 Unit/µl Polynucleotide Kinase in 1x Polynucleotide Kinase buffer (70 mM Tris-HCl, 10 mM MgCl2, 5 mM Dithiothreitol, pH 7.6 @ 25°C), 2 mM ATP, 1 Unit/µl SuperRNAseIN. After phenol extraction, RNAs were incubated in the presence of 30 µM of 5' adapter oligonucleotide, 1 Unit SuperRNaseIN (Ambion) and 1.5 Unit of T4 RNA ligase in ligation buffer (50 mM Tris-HCI (pH7.5), 10 mM MgCI2, 10 mM DTT, 1 mM ATP) and 10% Dimethyl sulfoxide. The ligated products were gel purified as described above and reverse transcribed in a standard 50 µl reaction (SuperScript III, Invitrogen) and subjected to RNaseH treatment. The cDNA was amplified by PCR and purified in a 10% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing platform (Illumina, Inc.).

tag: Total small RNA from IP input material of cde-1 mutant animals


title: mRNA-WT-2

value: WT


title: Cl.8 small RNAs BioRep-2 TechRep-1


title: mRNA-WT-1


title: CDE1YNG

description: Small RNA sequence data

source_name: CDE1 young adults

tag: Total RNA from young cde-1 mutant adults


title: Cl.8 small RNAs BioRep-1 TechRep-2

tag: biorep


title: Cl.8 small RNAs BioRep-1 TechRep-1

tag: techrep


title: smallRNA-S.cerevisiae-WT


title: The Physcomitrella Degradome

first: Michael

middle: J

last: Axtell

address: 280 Mueller Lab

description: Wild-type ten day old Physcomitrella patens (Gransden) protonemata were cultured, harvested, and total RNA extracted as previously described (Cho et al., 2008).  ~200 ng of poly-A enriched RNA was directly ligated to a chimeric DNA/RNA adapter containing a 3’ EcoP15I site using T4 RNA Ligase 1 (New England Biolabs).  After fractionation through S-400 spin columns (GE Healthcare) to deplete short RNAs, samples were reverse transcribed with Superscript III (Invitrogen) primed with a T20 tailed adapter.  After treatment with RNAse H (New England Biolabs), the primary cDNA was amplified with primers specific for the 5’ and 3’ adapters and digested with EcoP15I (New England Biolabs). The entire sample was then ligated to a dsDNA adapter using T4 DNA Ligase (New England Biolabs) followed by PAGE purification of the ~97nt population of 5’-end adapter ligated fragments.  After amplification, the sample was analyzed on the SOLiD™ 2 sequencing system (Applied Biosystems).  Because the SOLiD™ 2 system is incapable of analyzing samples where every nucleotide is identical (the first six nucleotides of the degradome samples are the EcoP15I recognition site), our sequencing runs mixed in an  irrelevant sample with SOLiD™ adapters from an unrelated experiment at a ratio of nine parts degradome to one part “filler”.  A detailed protocol including oligo sequences is available (Addo-Quaye et al., 2009 [in prep]).

data_processing: Raw colorspace reads were first parsed to separate the degradome reads from the “filler” by identifying the presence of the colors corresponding to the EcoP15I site.  Reads with the site were trimmed to remove this site.  Although EcoP15I cleavage is expected to chiefly generate 27nt fragments, we have consistently observed a distribution of EcoP15I-derived cleavage sizes ranging from ~25 to ~29nts (data not shown).  By computationally trimming the 3’ ends to leave 24nt reads we increased the probability that the resulting data is largely free of contamination by adapter sequences, and also removed data from the 3’ of the read which is typically of lower quality.  The SOLiD™ system’s two-base encoding scheme allows single color errors to cause ‘frameshifts’ if the read is translated to DNA before mapping to a reference.  Because we only wished to analyze those trimmed reads which could be exactly matched to either the Physcomitrella nuclear genome or nuclear transcriptome we directly translated into DNA space before mapping.  Values are the number of reads for each sequence -- all reported sequences were exactly matched to >= 1 genome location, >=1 location in the sense transcriptome, or both.

organism: Physcomitrium patens

source_name: Wild-type protonemata, 10-day old

growth_protocol: protonemal tissue was grown on cellophane-overlaid BCD media {Ashton, 1977} supplemented with 50mM ammonium tartarate under 16hr days, 8hr nights at 22ºC.

extract_protocol: Total RNA was extracted with Tri-Reagent, followed by polyA+ enrichment using oligo-dT agarose beads (Qiagen).

tag: genotype

value: Wild-type

value: protonemata

value: 10-day old


title: N2OLD

description: Small RNA sequence data

tag: Total RNA from aged wild-type adults


title: Cl.8 small RNAs BioRep-1 TechRep-3

tag: cell line


title: BG3 small RNAs BioRep-2 TechRep-2

tag: cell type


title: BG3 small RNAs BioRep-2 TechRep-1


title: csr1input

description: Small RNA sequence data

data_processing: 3' adapter sequences were trimmed and inserts longer than 18 nt were mapped to the C. elegans genome (Ce4).

extract_protocol:
Imunoprecipitation of protein complexes was preformed by preclearing worm extracts with 20 ml of Protein A/G PLUS-Agarose (Santa Cruz Biotechnologies) agarose beads per 5 mg of total protein for 1 h at 4°C. The cleared extract was then incubated with 15 to 25 mg of antibody for 5mg of total protein for 1h at 4°C (no antibody added for the "input" samples). 50 ml of Protein A/G PLUSAgarose beads were added, and the mixture incubated for 1 hour. The agarose beads were then washed three times with ice-cold lysis buffer. RNA was eluted from the Protein A/G PLUS-Agarose beads by extraction with TRI Reagent (MRC Reagents, Inc). 500 ml of TRI Reagent were added to 100 ml of beads and RNA was extracted as described in the manufacturer's protocol. Typical IPs for small RNA cloning were performed on 80 to 100 mg of total protein per sample to be cloned.
RNA was resolved in a 15% polyacrylamide 7M Urea Gel, along with 2 µmol of RNA standards (18- and 26-nt) in separate lanes. Ethidium Bromide Staining was used to visualize the RNA standards. A gel fragment was excised in the sample lanes between the two standards. RNA was eluted from the gel fragment in a [0.3M NaCl-TE (pH7.5)] solution overnight and ethanol-precipitated using 10 mg of glycogen as the carrier. Gel purified RNA was treated with 1 Unit/µl of Alkaline Phosphatase, Calf Intestine (NEB) in 100 mM NaCl, 50 mM Tris-HCl, 10 mM MgCl2, 1 mM Dithiothreitol, pH 7.9 @ 25°C and 1 Unit/µl SuperRNAseIn (Ambion) for 1 hour at 37 °C. After phenol extraction, the gel purified RNA and 1 µM of each standard were incubated with 20 µM of 3'-end linker, 1 Unit of SuperRNaseIN, 10% DMSO and 3 Units T4 RNA ligase (Takara) in 10 µl ligation buffer (50mM Tris-Cl pH7.5, 10mM MgCl2, 6mg/mL BSA, 10mM DTT). The 3' ligated products were gel purified and treated with 1 Unit/µl Polynucleotide Kinase in 1x Polynucleotide Kinase buffer (70 mM Tris-HCl, 10 mM MgCl2, 5 mM Dithiothreitol, pH 7.6 @ 25°C), 2 mM ATP, 1 Unit/µl SuperRNAseIN. After phenol extraction, RNAs were incubated in the presence of 30 µM of 5' adapter oligonucleotide, 1 Unit SuperRNaseIN (Ambion) and 1.5 Unit of T4 RNA ligase in ligation buffer (50 mM Tris-HCI (pH7.5), 10 mM MgCI2, 10 mM DTT, 1 mM ATP) and 10% Dimethyl sulfoxide. The ligated products were gel purified as described above and reverse transcribed in a standard 50 µl reaction (SuperScript III, Invitrogen) and subjected to RNaseH treatment. The cDNA was amplified by PCR and purified in a 10% acrylamide gel. PCR products generated for all the samples were sequenced on a Solexa sequencing platform (Illumina, Inc.).

tag: Total small RNA from IP input material of wild-type animals


title: BG3 small RNAs BioRep-1 TechRep-1


title: Total Nucleosome zim2 seq

status: Public on Aug 31 2009

city: Piscataway

first: Sam

middle: Guoping

last: Gu

state: NJ

address: Nelson Labs A125, 604 Allison Road

department: Molecular Biology and Biochemistry

institute: Rutgers University

description:
nucleosome core DNA
total nucleosome core DNA in the IP input
alignment in UCSC genome browser bed file format
library strategy: Micrococcal nuclease digestion and chromatin immunoprecipitation were used to obtain distinct populations of single nucleosome cores
library selection: Mnase and ChIP

data_processing:
alignment:The first 25 nt of each raw sequencing read was used to map the nucleosome end to the C. elegans genome (WS190) using Eland (Illumina) (Bentley et al., 2008) To avoid ambiguity, only perfect and unique matches were used.  For each unambiguous read, a putative nucleosome core sequence was defined by extending the sequence read to 147 bp.
processed: For each unambiguous read, a putative nucleosome core sequence was defined by extending the sequence read to 147 bp.

source_name: synchronized young adult population of mutant strain zim-2(tm574)

extract_protocol: Nucleosome core preparation   Synchronized populations of C. elegans (wild type strain N2 or mutant strain zim-2 (tm574)) were fed on E.coli OP50 (Brenner, 1974). Young adult worms were flash frozen and ground to fine powder in liquid nitrogen.  Worm grinds were directly resuspended in buffer A (15 mM Hepes-Na, pH 7.5, 60 mM KCl, 15 mM NaCl, 0.15 mM beta-mecaptoethonal, 0.15 mM spermine, 0.15 mM spermidine, 0.34 M sucrose), 0.5 mM PMSF, 1/100 dilution of protease set III (Cal Biochem), 1 mM DTT, and 2 mM CaCl2.  Micrococcal nuclease (MNase) (Roche), resuspended as 2 mg/ml in 10 mM Bis-Tris-Propane-HCl, 10 mM MgCl2, and 1 mM DTT, was added to the C. elegans extract.  Titrations of enzyme concentration at a series of time points were performed to find a condition where the majority of chromatin was digested to mono- and di-nucleosomes. Digestions (at 37°C) were stopped by adding EGTA to a final concentration of 20 mM.  Nucleosomes were solubilized by slowly adding drops of 2 M NaCl to a final concentration of 450 mM.  The extract from N2 animals was further fractionated in a 5-30% sucrose gradient (Beckman SW-41 rotor, 28,500 rpm for 18 hours).  Mononucleosome fractions were determined by DNA extraction and gel analysis, dialyzed in PBS buffer (137 mM NaCl, 10 mM Phosphate, 2.7 mM KCl, and pH 7.4) containing 1 mM PMSF, and stored at -80°C.  The ability of zim-2(tm574) mutant animals to enrich H3K9me3 on the pairing center arm of chromosome V was confirmed using a simplified protocol omitting the gradient fractionation.    Nucleosome immunoprecipitation   For each immunoprecipitation reaction, 2.5 μg antibody to either histone H3 (di- and trimethylated lysine 4, Abcam 6000) or histone H3 (trimethylated lysine 9, Abcam 8898) was captured to 10 μl Dynabead® conjugated with protein A (Invitrogen) that had been washed and resuspended in 95 μl PBS buffer with 0.01% Tween-20.  Nucleosome core fraction (N2) or crude MNase-treated extract (zim-2) was mixed with the conjugated antibodies with constant rotation at 4 °C for 6 hours.  The immunoprecipitant was washed thrice with 1 ml IP washing buffer (100 mM Hepes-Na, pH 7.5, 450 mM NaCl, 1% NP-40) at 4 °C.  The washing mix was rotated for three minutes for each wash. Immunoprecipitated nucleosome core DNA was isolated by incubating the beads with 400 μl lysis buffer (100 mM Tris-Cl, pH 7.5, 0.1 M NaCl, 1% SDS, 50 mM EDTA, 0.2 mg/ml protease K) at 65 °C for 1 hour, followed by organic extraction and DNA precipitation.     DNA library preparation  Nucleosome core DNA was treated with 0.5 U/μl T4 polynucleotide kinase (NEB) in 1x T4 DNA ligase buffer (NEB, containing 1 mM ATP) at 37 °C for 1 hour to phosphorylate 5’ ends and dephosphorylate 3’ ends.  Ends were then blunted in a reaction containing 0.75 U/μl of T4 DNA polymerase (NEB), dNTP (2mM each, Roche), and 1x NEB buffer 1 at 12 °C for 15 min.  Nucleosome core DNA was then ligated to previously annealed DNA oligos AF-SG-133 (5’ Pi-AGATCGGAAGAGCTCGTATGCCGTCTTCTGCTTG-OH 3’) and AF-SG-134 (5’ OH-CCCTACACGACGCTCTTCCGATCT-OH 3’) (NEB Quick Ligation Kit).  Nucleosome core DNA captured by linker oligos <search for "oligoes">  was purified using 6 % polyacrylamide (acrylamide:bis-acrylamide = 19:1) gels containing 8 M urea, followed by PCR amplification with primers AF-SG-135 (5’ OH-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT-OH 3’) and AF-SG-137 (5’ OH-CAAGCAGAAGACGGCATACGAGCT-OH 3’).  A series of PCR reactions with increasing cycle numbers was then carried out; we carefully choose cycle numbers for which product levels have not saturated (i.e. product levels still able to increase substantially with additional cycles); this ensures that the majority of amplified segments are still annealed to a true complement and avoids reannealing-distortion in the resulting sequence libraries (Parameswaran et al., 2007). .  After separating PCR products on 2 % agarose gels, DNA bands of the expected size (~210-240 bp) were extracted (QIAchange® Gel Extraction Kit (Invitrogen); omitting the 50°C heating step), followed by massive parallel DNA sequencing (Illumina GAII).

tag: strain

tag: chip antibody

value: IP input


title: H3K9me3 zim2 chipSeq

description:
nucleosome core DNA
anit-H3K9me3 precipitated nucleosome core DNA
alignment in UCSC genome browser bed file format
library strategy: Micrococcal nuclease digestion and chromatin immunoprecipitation were used to obtain distinct populations of single nucleosome cores
library selection: Mnase and ChIP

tag: developmental stage


title: H3K9me3 N2 chipSeq

source_name: synchronized young adult population of wild type strain N2


title: H3K4me2/3 N2 chipSeq

description:
nucleosome core DNA
anit-H3K4me2/3 precipitated nucleosome core DNA
alignment in UCSC genome browser bed file format
library strategy: Micrococcal nuclease digestion and chromatin immunoprecipitation were used to obtain distinct populations of single nucleosome cores
library selection: Mnase and ChIP


title: csr1IP

description: Small RNA sequence data

tag: Total small RNA from a CSR-1 IP from wild-type animals

tag: tissue

tag: genome/variation


title: H3K27Ac_s6_ChIPSeq_1

status: Public on Sep 03 2009

description: corresponding input record: GSM400669

tag: antibody

tag: antibody manufacturer

value: Abcam

tag: antibody catalog number

tag: antibody lot number

tag: development stage


title: HeLa-siWDR5-H3K4me3

status: Public on Aug 20 2009

city: Charlottesville

first: Chongzhi

last: Zang

state: VA

institute: University of Virginia

description:
HeLa siWDR5 H3K4me3
ChIP-Seq, genomic DNA

data_processing: Sequence reads were obtained and mapped to the human (March 2006, hg18) genomes using the Illumina Genome Analyzer Pipeline. More details as described in Wang et al., 2009

source_name: HeLa cell

growth_protocol: Growth protocol as described in Wang et al., 2009

extract_protocol: Extract protocol as described in Wang et al., 2009

treatment_protocol: Treatment protocol as described in Wang et al., 2009

value: HeLa cell

value: anti-H3K4me3 Ab

tag: sirna

tag: treatment


title: HeLa-siWDR5-H4K16ac-HDACi-0h

description:
HeLa siWDR5 H4K16ac w/o TSA+Bu
ChIP-Seq, genomic DNA

value: anti-H4K16ac Ab


title: HeLa-siControl-H4K16ac-HDACi-1h

description:
HeLa siControl H4K16ac w/ TSA+Bu 1h
ChIP-Seq, genomic DNA

value: siControl

value: Trichostatin + sodium butyrate for 1 hours


title: HeLa-siWDR5-H3K9ac-HDACi-1h

description:
HeLa siWDR5 H3K9ac w/ TSA+Bu 1h
ChIP-Seq, genomic DNA

value: anti-H3K9ac Ab


title: HeLa-siControl-H4K16ac-HDACi-0h

description:
HeLa siControl H4K16ac w/o TSA+Bu
ChIP-Seq, genomic DNA


title: HeLa-siControl-H3K9ac-HDACi-1h

description:
HeLa siControl H3K9ac w/ TSA+Bu 1h
ChIP-Seq, genomic DNA


title: HeLa-siWDR5-H3K9ac-HDACi-0h

description:
HeLa siWDR5 H3K9ac w/o TSA+Bu
ChIP-Seq, genomic DNA


title: HeLa-siControl-H3K9ac-HDACi-0h

description:
HeLa siControl H3K9ac w/o TSA+Bu
ChIP-Seq, genomic DNA


title: HeLa-siWDR5-H4K16ac-HDACi-1h

description:
HeLa siWDR5 H4K16ac w/ TSA+Bu 1h
ChIP-Seq, genomic DNA


title: HeLa-siControl-H3K4me3

description:
HeLa siControl H3K4me3
ChIP-Seq, genomic DNA


title: CD4-PolII-HDACi-2h

description:
CD4 RNA Pol II w/ TSA+Bu 2h
ChIP-Seq, genomic DNA

source_name: CD4+ T cell

value: CD4+ T cell

value: anti-RNA Pol II Ab

value: Trichostatin + sodium butyrate for 2 hours


title: CD4-PolII-HDACi-12h

description:
CD4 RNA Pol II w/ TSA+Bu 12h
ChIP-Seq, genomic DNA

value: Trichostatin + sodium butyrate for 12 hours


title: CD4-H4K16ac-HDACi-0h

description:
CD4 H4K16ac w/o TSA+Bu
ChIP-Seq, genomic DNA


title: CD4-H3K9ac-HDACi-0h

description:
CD4 H3K9ac w/o TSA+Bu
ChIP-Seq, genomic DNA


title: Act-CD4-HDAC6

description:
Activated CD4 HDAC6
ChIP-Seq, genomic DNA

value: anti-HDAC6 Ab


title: CD4-H3K9ac-HDACi-8h

description:
CD4 H3K9ac w/ TSA+Bu 8h
ChIP-Seq, genomic DNA

value: Trichostatin + sodium butyrate for 8 hours


title: CD4-PolII-HDACi-0h

description:
CD4 RNA Pol II w/o TSA+Bu
ChIP-Seq, genomic DNA


title: CD4-H4K16ac-HDACi-8h

description:
CD4 H4K16ac w/ TSA+Bu 8h
ChIP-Seq, genomic DNA


title: CD4-H3K9ac-HDACi-2h

description:
CD4 H3K9ac w/ TSA+Bu 2h
ChIP-Seq, genomic DNA


title: CD4-H4K16ac-HDACi-2h

description:
CD4 H4K16ac w/ TSA+Bu 2h
ChIP-Seq, genomic DNA


title: CD4-HDAC3

description:
CD4 HDAC3
ChIP-Seq, genomic DNA

value: anti-HDAC3 Ab


title: CD4-HDAC2

description:
CD4 HDAC2
ChIP-Seq, genomic DNA

value: anti-HDAC2 Ab


title: CD4-HDAC6

description:
CD4 HDAC6
ChIP-Seq, genomic DNA


title: CD4-Tip60

description:
CD4 Tip60
ChIP-Seq, genomic DNA

value: anti-Tip60 Ab


title: CD4-Rabbit-IgG

description:
CD4 Rabbit IgG
ChIP-Seq, genomic DNA

value: Rabbit IgG


title: Small RNAs from petunia WT flower

description: Low molecular weight RNA was gel-sized in the ~18-33 nt range and used to construct a small RNA library according to Lu et al. (2007). The library was  sequenced on the Illumina Genome Analyzer II at the Delaware Biotechnology Institute according to manufacturer’s instructions.

source_name: WT flower

growth_protocol: Plants were grown in a greenhouse as described in Napoli and Ruehle (J. Hered 87, 371–377, 1996).

extract_protocol: Total RNA was extracted from pre-anthesis petals of both WT (Petunia hybrida V26) and silenced petunia plants with the Plant RNA Purification Reagent (Invitrogen). Quality of total RNA was evaluated by agarose gel electrophoresis, micro-electrophoresis (Agilent 2100 Bioanalyzer, Ambion) and O.D. measurements (Nanodrop ND-1000).

value: V26;  Pre-anthesis petal tissue

tag: replicate

value: WT-biological replicate


title: Act-CD4-Tip60

description:
Activated CD4 Tip60
ChIP-Seq, genomic DNA

value: activated CD4+ T cell

value: anti Tip60 Ab


title: CD4-MOF

description:
CD4 MOF
ChIP-Seq, genomic DNA

value: anti-MOF Ab


title: CD4-p300

description:
CD4 p300
ChIP-Seq, genomic DNA

value: anti-p300 Ab


title: Total Nucleosome N2 seq

description:
nucleosome core DNA
total nucleosome core DNA in the IP input
alignment in UCSC genome browser bed file format
library strategy: Micrococcal nuclease digestion and chromatin immunoprecipitation were used to obtain distinct populations of single nucleosome cores
library selection: Mnase and ChIP

value: young adult

value: IP input


title: CD4-Goat-IgG

description:
CD4 Goat IgG
ChIP-Seq, genomic DNA

value: CD4+ T cell

value: Goat IgG


title: CD4-HDAC1

description:
CD4 HDAC1
ChIP-Seq, genomic DNA

value: anti-HDAC1 Ab


title: Illumina sequencing of small RNAs from mixed C. elegans embryos

status: Public on Sep 06 2009

city: Berlin

first: Jonas

last: Maaskola

address: Robert-Roessle Str. 10

department: Systems Biology of Gene Regulatory Elements

institute: Max-Delbrueck Centrum Berlin-Buch

description: Mixed embryos.

data_processing: Removal of 3' adapter sequences by searching for best matches of adapter prefixes up to edit distance 4.

source_name: Mixed C. elegans embryos

growth_protocol: Strains were maintained using standard methods (Brenner, 1974) on OP50 seeded NGM plates or in liquid culture.

extract_protocol: Library preparation as well as cluster generation and deep sequencing was performed according to the 5’ ligation dependent (5’monophosphate dependent) manufacturer’s protocol (Digital Gene Expression for small RNA, Illumina). Rougly 60,000 embryos (~10µg total RNA) were used for small RNA library preparation. Small RNAs were size selected between 18 and 40 nt according to the ssDNA marker in the small RNA sequencing kit (Illumina).

treatment_protocol: Mixed embryos obtained by standard bleaching procedure (Brenner, 1974).

value: Mixed embryos

tag: meoh-fixed

tag: sorted

tag: resorted


title: Section14_tip_Mo17

status: Public on Aug 15 2009

city: Ithaca

first: Lalit

last: Ponnala

address: 616 Rhodes Hall

institute: Cornell University

description:
mRNA-seq from tip
processed data file: Sample2.copynumber.txt
processed data file: Sample2.alignment.txt
The copynumber file shows the number of times each read is detected in our sample. The alignment file shows the alignment for each read in the copynumber file. Please see the README.txt file (linked as supplementary file on Series record) for a description of each file's contents.

data_processing: The data in the alignment table was generated by aligning the first 32bp of each raw read to the maize genome (release 4.53a, available at:    http://www.maizesequence.org/index.html). The software tool "Eland" provided by Illumina was used to perform the alignments, with settings -10,5,2, i.e. allowing up to 10 perfect alignments, 5 1-mismatch alignments, and 2 2-mismatch alignments for each read.

organism: Zea mays

source_name: Maize leaf

growth_protocol: Plants were grown for 9 days under a 80:20 mix of metal halide: capsylite halogen lamps at a fluence of 500 umol/m2/sec, 12L:12D, 31ºL, 22ºD, 50% rel. humidiy.

extract_protocol: Total RNA from 1 cm segments of leaf tip and base tissues from the third leaf of 9 day old (DAP) Mo17 seedlings was extracted using Trizol reagent (Invitrogen, TRIZOL® Reagent, Cat. No.  15596-018). Then, 100 ng mRNA was isolated from each total RNA samples using Dynabeads mRNA purification kit (Invitrogen Cat. No.  610-02). cDNA synthesis was primed with random hexamers and libraries constructed according to manufacturer's recommendations (Illumina mRNA-seq Sample Prep Kit part# 1004898 Rev.A).

tag: inbred line

value: leaf tip


title: E20-24_CBP_ChIPSeq_1

status: Public on Aug 13 2009

description: corresponding input record: GSM400673

growth_protocol: 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a 20-24 hours egg laying, new plates are added after a 2 hours pre-egglaying at 9 am and flies are allowed to lay eggs for 4 hours.3. Embryos are collected at 8.45 am using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 9 am in 1.8% formaldehyde at room temperature for 15 min.

tag: development point

value: CBP

tag: antibody provider

value: Dr. Mattias Mannervik in Stockholm University


title: E20-24_H3K4Me1_ChIPSeq_1

description: corresponding input record: GSM400673

value: Abcam


title: E12-16_PolII_ChIPSeq_1

description: corresponding input record: GSM408987

value: PolII

value: Covance


title: CD4-PCAF

description:
CD4 PCAF
ChIP-Seq, genomic DNA

tag: cell type

value: anti-PCAF Ab


title: DLD2

status: Public on Aug 25 2009

first: Christine

last: Mayr

address: 9 Cambridge Center

institute: Whitehead Institute

data_processing: Reads containing linker matches were mapped (after removing linkers) to predicted pre-miRNAs (MirBase version 12.0)

source_name: cell line

growth_protocol: RNA was harvested from exponentially growing cells

extract_protocol: Total RNA extracted using standard Trizol extraction. Total RNA fractionated on 15% polyacrylamide gel and 3' and 5' adaptors sequentially ligated requiring a 3' hydroxyl group and 5' phosphate

tag: cell line


title: CD4-CBP

description:
CD4 CBP
ChIP-Seq, genomic DNA

value: anti-CBP Ab


title: E0-4_CBP_ChIPSeq_1

first: Kevin

last: White

description: corresponding input record: GSM400657

data_processing: Reads were aligned to the flybase BDGPv5 reference genome, and the fragment count at any given position (10-bp resolution) was estimated as the number of uniquely aligned reads oriented towards it and within 300Â bp. Enriched intervals were identified by comparison of the mean fragment count in 1-kb windows against a sample-specific expected distribution estimated by randomization (H3K4me3, H3K27me3), or using a supervised Hidden Markov Model (H3K36me3, H3K9me3, H4K20me3).

growth_protocol: 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a 0-4 hours egg laying, new plates are added after a 2hours pre-egglaying at 10 am and flies are allowed to lay eggs for 4 hours. 3. Embryos are collected at 1.45 pm using a filter mesh and a brush. They are rinsed with EWB and cross-linked at 2 pm in 1.8% formaldehyde at room temperature for 15 min.

extract_protocol: Drosophila samples are crushed in presence of 1.8% of formaldehyde for 15 minutes. After washing, the material is sonicated for 15 minutes to solubilize the chromatin. After centrifugation, the supernatant containing soluble chromatin is collected for IPs. Specific antibodies are added to the chromatin extract while one aliquot will be treated for DNA extraction corresponding to the reference samples. ChIP are carried overnight at room temperature. Protein A Sepharose beads are used to pull down the chromatin bound antibodies. After washes and finally elution, the DNA corresponding to the IPed material is extracted and purified. The DNA is directly used for Solexa library preparation.


title: E20-24_PolII_ChIPSeq_1

description: corresponding input record: GSM400673


title: AdultFemale_H3K4Me1_ChIPSeq_1

description: corresponding input record: GSM400671

source_name: AdultFemale

growth_protocol: 1. y; bw cn sp flies are cultivated in cages with apple juice agar plates covered with yeast powder. 2. For a collection of Drosophila Adult Female 2-3 days old, new plates are added after a 2 hours pre-egglaying at 9 am and flies are allowed to lay eggs for 24 hours. Embryos covered plates are then cut and placed back into regular fly medium bottles. 3. Adult flies are collected 2 to 3 days after the first adult hatched. They are sorted for sex under a dissecting microscope after being put to sleep with CO2. Females and males are placed into different eppendorff tubes on ice and then rinsed with EWB before cross-linking in 1.8% formaldehyde at room temperature for 15 min.

value: AdultFemale