title: miwi2KO_P10_MACSsortedGermCells_BSseq

city: Pasadena

first: Sergei

middle: A

last: Manakov

address: 1200 E California blvd, MC 147-75

department: Biology and Biological Engeneering

institute: California Institute of Technology

data_processing:
Analysis of RNA-seq libraries. We used of polyA(+) non-stranded protocol to generate RNA-seq libraries. Trimming of Illumina adaptors with Reaper was the first step in library processing (Davis et al. 2013). Next, library reads were first aligned to sequences of rRNA transcripts (GenBank identifiers: 18S, NR_003278.3; 28S, NR_003279.1; 5S, D14832.1; and 5.8S, K01367.1) with Bowtie 0.12.7 with up to three mismatches (Langmead et al. 2009). Reads with valid alignments to rRNA were considered contamination and removed from father analysis. The remaining reads were mapped to mm10 reference genome either in the multimapping mode (up to 10,000 valid alignments were allowed). The alignments in the multimmaping mode were used to evaluate expression of TE families. Annotation of individual TE copies and families were taken from RepeatMasker (http://www.repeatmasker.org) annotation of mm10 genome as available via UCSC Genome Bioinformatics Site (Karolchik et al. 2014). Per-family read count was computed as previously described (Pezic et al. 2014). Briefly, reads with 5’ coordinates intersecting individual copies of TEs in a family were totaled up weighing each read alignment instance by the total number of alignments of that read.
Analysis of bisulfite sequencing libraries. We used the same computational procedure to analyze Miwi2 knock-out and heterozygote control libraries that we generated and previously published Mili knock-out and wild type control libraries (Molaro et al. 2014). First, Illumina adaptors were trimmed using Trim Galor! tool (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/). Trimmed reads were then aligned to mm10 genome with Bismark tool (Krueger and Andrews 2011). Default Bismark parameters and Bowtie2 (v 2.2.2) engine (Langmead and Salzberg 2012) were used to align both single-end and paired-end libraries. Following the alignment, mean CpG-methylation level of individual CpG sites and of regions of interests (e.g. TE copies or fragments of TEs) was determined using MethPipe computational tool suite (Song et al. 2013). Reads aligned to the same genomic position were collapsed and replicate samples were merged prior to evaluation of the mean CpG methylation of individual CpG sites and TE copies.
Genome_build: mm10
Supplementary_files_format_and_content: “rnaSeq_RMSK.tab.txt” files. These files provide an RPKM-normalized and a raw number of RNA-seq reads that overlap (by the 5'-nucleotide of the read) invidual repeats (e.g. transposons) that are annotated in RepeatMasker (RMSK) track in the UCSC genome browser. Description of fields: (1) Indexing field to speed chromosome range queries (2) Smith Waterman alignment score (3) Base mismatches in parts per thousand (4) Bases deleted in parts per thousand (5) Bases inserted in parts per thousand (6) Genomic sequence name (i.e. chromosome) (7) Start in genomic sequence (8) End in genomic sequence (9) -#bases after match in genomic sequence (10) Relative orientation + or – (11) Name of repeat (12) Class of repeat (13) Family of repeat (14) Start (if strand is +) or -#bases after match (if strand is -) in repeat sequence (15) End in repeat sequence (16) -#bases after match (if strand is +) or start (if strand is -) in repeat sequence (17) First digit of id field in RepeatMasker .out file. Best ignored. (18) mappablity adjusted RPKM (19) read count
Supplementary_files_format_and_content: “meth_RMSK.tab.txt” files. These files provide number of consensus CpG sites (i.e. CpG sites that are covered by at least one read in the most shallow BS-seq library in our study, which is DP107) in individual repats (e.g. transposons) that are annotated in RepeatMasker (RMSK) track in the UCSC genome browser. Description of fields: (1) Genomic sequence name (i.e. chromosome) (2) Start in genomic sequence (3) End in genomic sequence (4) Name of repeat (5) Class of repeat (6) Family of repeat (7) Smith Waterman alignment score (8) Base mismatches in parts per thousand (9) Bases deleted in parts per thousand (10) Bases inserted in parts per thousand (11) -#bases after match in genomic sequence (12)  Start (if strand is +) or -#bases after match (if strand is -) in repeat sequence (13) End in repeat sequence (14) -#bases after match (if strand is +) or start (if strand is -) in repeat sequence (15) # of consensus CpG sites (16) # of consensus CpG covered by reads (17) # of reads that support methylation of covered CpG sites  (18) # of reads in total that covered consensus CpG sites (19) average methylation level in the repeat (20) strand of the repeat

extract_protocol:
Total RNA extraction. Total RNA was isolated from total testis using Ribozol (Amresco, N580). For isolation of RNA from embryonic testes, embryos from several pregnant mothers were harvested, genotyped, and pooled according to genotype for RNA isolation. All P10 samples are either from individual animals, or from littermates. RNA was DNase-treated using Turbo DNA-free kit (Ambion, AM1907).
Small RNA extraction. Total RNA was isolated from embryonic testes using Ribozol. Thirty ug total RNA was loaded on 12% urea-polyacrylamide (PAA) gel. Fraction 19-30 nt was excised and snap-frozen in liquid nitrogen in 400 ul 0.4M NaCl. RNA was eluted from the gel over night at 16C, shaking at 1,000 rpm and precipitated in 3 volumes of absolute ethanol with added GlycoBlue.
Genomic DNA extraction (for BS-seq). Spermatocytes were isolated from P10 testis by MACS-sorting from testes as described in Pezic et al (2014), resuspended in 100 μl of TE, and lysed in 1 ml of genomic DNA lysis buffer (10 mM Tris-HCl pH 8.0, 100 mM EDTA pH 8.0, 0.5% SDS, supplemented with 250 U RNaseI (Ambion, AM2295)). Lysate was incubated at 37°C for 1 hour, transferred to a glass douncer and homogenized further by 15 strokes with tight pestle. Proteinase K (NEB) was added to final concentration of 100 μg/ml, and incubated at 50°C for 3 hours. After cooling to RT, equal volume of phenol pH 8.0 was added, mixed well, and incubated for 10 min at RT with agitation. Water phase was recovered after centrifugation, and phenol extraction repeated one more time, followed by phenol/chloroform and chloroform extraction. Water phase was then mixed with 2 volumes of absolute ethanol, with addition of 5M NaCl to final concentration of 300 mM. DNA was precipitated for 2 hours at -80°C, or over night at -20°C. Pelleted DNA was washed twice with 70% EtOH, dried, and resuspended in water. A260/280 and A260/230 ratios were measured to check purity.
RNA-seq (polyA+) libraries. For library cloning, 1 μg DNase-treated RNA was polyA+ selected using DynaBeads mRNA Purification Kit (Invitrogen, cat.no. 610.06). Total RNA libraries were cloned using either Illumina TruSeq RNA sample prep kit (version 2, 48 rxns, RS-122-2001) or NEBNext Ultra RNA library prep kit for Illumina (New England Biolabs, E7530).
Small RNA-seq libraries. Pre-adenylated 3’ linker (/5`Phos/TGGAATTCTCGGGTGCCAAGGAACTC/3`ddC/; 5’DNA adenylation kit, NEB) was ligated to RNA over night at 4C using Truncated RNA Ligase 2 (NEB). Ligation reactions were loaded onto 10% urea-PAGE, 45-56 nt fraction was excised and nucleic acids extracted as above. 5’ linker (rGrUrUrCrArGrArGrUrUrCrUrArCrArGrUrCrCrGrArCrGrArUrC) was ligated to the samples using RNA Ligase 1 (NEB) over night at 4C. Ligation reactions were loaded on 10% Urea-PAA gel, 72-83 nt fraction was excised and nucleic acids as above. Extracted samples were reverse-transcribed (primer sequence: GGAGTTCCTTGGCACCCGAGA) and library amplified by PCR using standard Illumina primers. Final libraries were excised from 1.5% agarose gel, and sequenced on Illumina platform.
BS-seq libraries. Genomic DNA (200 ng) was sonicated using Bioruptor (Diagenode) in 100ul 1X TE, and libraries cloned using NuGEN Ovation UltraLow MethylSeq Multiplex System 1-8 kit (NuGEN 0335-32) according to manufacturer’s instructions. Bisulfite conversion of DNA is performed after adapter ligation using this kit, and it was done using Active Motif MethylDetector Kit (#55001) as instructed by the manufacturer’s protocol.

tag: strain background

tag: genotype/variation

tag: tissue/cell type

value: MACS sorted (a-EpCAM G8.8 antibody, sc-53532) spermatocytes

value: AGATCGGAAGAGCAC

tag: molecule subtype

value: bisulphite converted genomic DNA


title: miliKO_P10_Testis_polyAplusRNA_replicate1

status: Public on Jul 11 2015

type: SRA

country: USA

library_source: transcriptomic

molecule: total RNA

organism: Mus musculus

tag: age

value: miliKO

value: total testis

value: GATCGGAAGAGCACA

value: polyA+ RNA fraction


title: miwi2HET_P10_MACSsortedGermCells_BSseq

library_source: genomic

molecule: genomic DNA


title: miwi2HET_E16.5_Testis_smallRNA

value: TGGAATTCTCGGGTG

value: small RNA fraction (19-30 nt range)


title: miwi2KO_E16.5_Testis_smallRNA


title: miliHET_P10_Testis_polyAplusRNA_replicate1

value: miliHET


title: SET2 cells persistent to BMS911543_replicate 1

city: New York

first: Jeffrey

last: Zhao

state: NY

address: 1275 York Avenue

department: Genomics Core

institute: Memorial Sloan Kettering Cancer Center

description:
s_SET2_BMS_per1
library run on two lanes to get enough reads

data_processing:
All sequenced library were mapped to the human genome (hg19) using rnaStar. Unmapped reads were mapped again to the same genome using BWA MEM (version 0.7.5.1)
HTSeq v0.5.3p3 (options -s y -m intersection-strict) is used for feature counting using the Homo_sapiens.GRCh37.69_ENSEMBL gene model also used at the mapping stage
DESeq (v1.10.1) is used for the normalization of count table.
Genome_build: hg19
Supplementary_files_format_and_content: text file with normalized count tables from each experiment

organism: Homo sapiens

source_name: SET2 cell line

growth_protocol: SET2 cells were cultured in RPMI1640 media + 20% FCS. For persistent cell lines, the respective type I JAK inhibitor was added to the culture media for continuous growth in presence of inhibitor. For acute exposures with inhibitor, DMSO was used as a control

extract_protocol:
SET2 cells were lysed in Trizol and processed according to instructions with reagent.
poly(A) RNA was isolated using Dynabeads® mRNA DIRECTTM Micro Kit (Life Technologies) from 1 μg of total RNA. mRNA was then fragmentated using RNaseIII and purified. The fragmented samples’ quality and yield were evaluated using Agilent BioAnalyzer. Subsequently, the fragmented material underwent whole transcriptome library preparation according to the Ion Total RNA-Seq Kit v2 protocol (Life Technologies), with 16 cycles of PCR.  Samples were barcoded, template-positive Ion PITM Ion SphereTM Particles (ISPs) were prepared using the ion one touch system II and Ion PITMTemplate OT2 200kit v2 Kit (Life Technologies). Enriched particles were sequenced on a Proton sequencing system using 200 bp version 2 chemistry.

tag: inhibitor exposure

value: chronic

tag: cell line


title: miliHET_P10_Testis_polyAplusRNA_replicate2


title: SET2 cells persistent to INC424_replicate 2


title: naïve SET2 cells, 48h INC424 replicate 1

value: acute


title: SET2 cells persistent to BMS911543_replicate 2


title: 76 mer control 75%

city: Athens

first: Robert

middle: J

last: Schmitz

state: GA

address: B416 Davison Life Sciences

department: Genetics

institute: University of Georgia

data_processing:
Data were mapped to the Caldicellulosiruptor kristjanssonii wild type reference genome
MethylC-seq analysis were performed as described in Schmtiz et al. Epigenome-wide inheritance of DNA methylation variants in a recombinant inbred population. Genome Research (2013).
Genome_build: Caldicellulosiruptor kristjanssonii wild type reference genome
Supplementary_files_format_and_content: tab delimited text file

organism: synthetic construct

source_name: pUC19 vector DNA

growth_protocol: Caldicellulosiruptor kristjanssonii wild-type strain was grown anaerobically in liquid low osmolarity complex growth (LOC) medium17 (final pH 7.0) with maltose (0.5% w/v; Sigma M5895) as the carbon source.

extract_protocol: Genomic DNA (gDNA) was purified from Arabidopsis thalina leaf tissue using a Qiagen DNease Plant Kit (Qiagen, Valencia, CA), according to the manufacturer’s instructions. MethylC-Seq libraries were constructed according to Urich MA, Nery JR, Lister R, Schmitz RJ, Ecker JR. (2014) MethylC-seq: Base resolution whole genome bisulfite sequencing library preparation. Nature Protocols (in press)

tag: tissue

value: pUC19 vector DNA


title: 76 mer control 50%


title: control_E16.5_Testis_polyAplusRNA_replicate2


title: UCH2_2

city: Cambridge

first: Charles

last: Lin

state: MA

address: 415 Main St.

institute: Broad Institute

description: paired-end ATAC-Seq

data_processing:
Library strategy: ATAC-Seq
reads were aligned to the hg19 genome building using bowtie2 with parameters -p 4 -k 1
mitochondrial reads were removed using the command "samtools view -h 'ALIGNED_BAM' | grep -vF chrM | samtools view -bS - >'%s' 2>/dev/null"
bam was resorted using samtools sort
bam reads were fixed using samtools fixmate
duplicates were removed using samtools with the command samtools rmdup
wiggles were made using MACS 1.4.2 pileups
Genome_build: hg19
Supplementary_files_format_and_content: .wig wiggles of read pileups across the whole genome

growth_protocol: For UCH2 cells: IMDM:RPMI 1640 mixed 4 parts IMDM:1 part RPMI; 10% FBS not heat inactivated, + Pen-strep. Cells are grown on 0.1% gelatin coated plates and are split 1:2 when nearing confluence, about every 5 days.

extract_protocol:
50,000 cells were pelleted, washed with PBS and re-pelleted.  Cell pellets were frozen on dry ice. Cells were subjected to ATAC transposase reaction for 60 minutes at 37deg
Genomic DNA was purified and amplified with Nextera custom ATAC primers. Side qPCR reactions were performed to minimize PCR amplification bias. Amplified libraries were run either on TapeStation or BioA to determine size range. Libraries were quantified by KAPA qPCR kit and sequenced


title: 76 mer control 100%


title: miwi2HET_P10_Testis_polyAplusRNA


title: control_E16.5_Testis_polyAplusRNA_replicate1


title: MUGCHOR

description: paired-end ATAC-Seq

source_name: MUGCHOR

growth_protocol: For MUGCHOR cells: IMDM:RPMI 1640 mixed 4 parts IMDM:1 part RPMI; 10% FBS not heat inactivated, 1X ITS, 1X glutamine (2mm), + Pen-strep. Cells are grown on regular plastic and are split 1:2, when nearing confluence, about every 4-5 days.

value: MUGCHOR


title: miwi2KO_P10_Testis_smallRNA


title: 76 mer control 25%


title: miliKO_P10_Testis_polyAplusRNA_replicate3


title: miliKO_P10_Testis_polyAplusRNA_replicate2


title: 76 mer control 0%


title: miwi2HET_P10_Testis_smallRNA


title: C. kristjanssonii Tet treated

organism: Caldicellulosiruptor acetigenus

source_name: genomic DNA

value: bacterial cells

tag: genotype

value: wild-type


title: miwi2KO_P10_Testis_polyAplusRNA


title: Fig2_SacCer_20% Scc1PK_IP

status: Public on Jul 10 2015

city: Sheffiled

first: Bin

last: Hu

address: West Bank

department: MBB

country: United Kingdom

institute: University of Sheffield

data_processing:
Library sequencing was carried out on the Ion Torrent Proton
The quality of the raw sequence data was examined using FastQC (Galaxy tool version 1.0.0) and trimming of reads was carried out using “trim sequences” (Galaxy tool version 1.0.0)
Any reads smaller than 50bp were removed using ‘Filter FASTQ’
The final reads were aligned to the Saccharomyces cerevisiae genome (sacCer3, SGD) or Candida glabrata genome (CBS138, genolevures) using Bowtie2 (Galaxy tool version 0.2) with the default (--sensitive) parameters
To generate an alignment in which all the sequences exclusively align to the Saccharomyces cerevisiae genome, the whole reads were first aligned to Candida glabrata genome and the unaligned reads were retrieved as a separate Fastq file. Subsequently, these unaligned reads were re-aligned to Saccharomyces cerevisiae genome and the resulting aligned BAM file therefore contained reads that were unique to Saccharomyces cerevisiae. A same strategy was used to generate alignments unique to Candida glabrata.
Genome_build: SacCer3 or Candida glabrata (ASM254v2)
Supplementary_files_format_and_content: bigwig files

organism: Nakaseomyces glabratus

source_name: Saccharomyces cerevisiae and Candida glabrata mixed cells

growth_protocol: Yeast cells were grown in YPD to reach a density of 0.3-0.6 OD600.

extract_protocol:
To crosslink cells, 45ml of yeast culture was mixed with 4.2ml of fixation solution (50mM Tris-HCl pH 8.0, 100mM NaCl, 0.5mM EGTA, 1mM EDTA, 30% Formaldehyde) and incubated at 18°C for 30 minutes. The crosslinking reaction was quenched by incubating with 2ml of 2.5M Glycine for 5 min. Fixed cells were harvested, washed with ice-cold PBS and re-suspended in 1ml of ChIP lysis buffer (50mM Hepes-KOH pH 8.0, 140mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% Sodium deoxycholate, 1mM PMSF, Roche protease inhibitor). For each ChIP, 10 OD600 units of Saccharomyces cerevisiae cells mixed with 5 OD600 units of Candida glabrata cells were pelleted and re-suspended in 0.3ml of ChIP lysis buffer. Cells were mixed with glass beads and disrupted by FastPrep®-24 (MP Biomedicals, USA). The entire lysis was collected and sonicated at 4°C for 40 minutes using a Bioruptor (Diagenode, Belgium). The cell debris were removed by centrifugation and supernatants containing sheared chromatin with a size range from 100-800bp were adjusted to a final volume of 1ml with ChIP lysis buffer. Extracts were pre-cleared for 1 hour at 4°C with 30µl of Protein G Dynabeads (Invitrogen).  80µl of supernatant was taken as whole cell extract (W) and stored at -80°C.  Five µg of anti-PK antibody (Bio-Rad) and 50 µl of Protein G Dynal beads (Invitrogen) was used for immunoprecipitation (6 hours-overnight, rotation at 4°C). The beads were subsequently washed for 5 minutes with the following buffers: 2Ã—ChIP lysis buffer; 3x ChIP high-salt lysis buffer (50mM Hepes-KOH pH 8.0, 500mM NaCl, 1mM EDTA, 1% Triton X-100, 0.1% Sodium deoxycholate, 1mM PMSF); 2Ã—ChIP wash buffer (10mM Tris-HCl pH 8.0, 0.25M LiCl, 0.5% NP-40, 0.5% sodium deoxycholate, 1mM EDTA, 1mM PMSF) and 1x TE buffer (10mM Tris-HCl pH 8.0, 1mM EDTA, 50mM NaCl). The immunoprecipitated chromatin was eluted by incubation of beads with 120µl of TES buffer (50mM Tris-HCl pH 8.0; 10mM EDTA; 1% SDS) at 65°C for 15 minutes. The supernatants were collected and termed the IP sample. The whole cell extract sample (W) was mixed with 40µl of TES3 buffer (50mM Tris-HCl pH 8.0; 10mM EDTA; 3% SDS). Both samples were de-crosslinked at 65°C overnight. RNA was degraded by incubating with 2-µl of RNAse A (10mg/ml, Roche) at 37°C for 1 hour and protein was subsequently removed by incubation with 10µl of Proteinase K (18mg/ml, Roche) at 65°C for 2 hours. DNA was purified using ChIP DNA Clean & Concentrator kit (Zymo Research, USA).
NEBNext Fast DNA Library Prep set for Ion Torrent, E6270

tag: sac_cer strain

value: K14061, Scc1PK tag

tag: gla_gra strain

value: K23308, Scc1 PK tag

tag: sac_cer and gla_gla mixed percentage

tag: condition for cell growth

value: exponentially grown

tag: antibody

value: ChIP -of Scc1_PK9 with anti-PK antibody (Biorad, MCA1360)


title: Fig2_SacCer_20% Scc1PK_WCE

value: none, input


title: Fig2_SacCer_40% Scc1PK_IP


title: Fig2_SacCer_0% Scc1PK_WCE


title: Fig2_SacCer_0% Scc1PK_IP


title: Fig1_SacCer_50% Scc1PK9_WCE


title: Fig1_CanGla_100% Scc1PK9_WCE

source_name: Candida glabrata  cells


title: Fig2_SacCer_40% Scc1PK_WCE


title: UCH2_1

description: paired-end ATAC-Seq


title: Fig1_CanGla_100% Scc1PK9_IP


title: H13_exp

city: Beijing

first: Haibo

last: Yang

address: No.1 Beichen West Road, Chaoyang District

country: China

institute: Institute of Microbiology, Chinese Academy of Sciences

description: Measurement of replication dynamics (marker frequency analysis; MFA) for Haloferax mediterranei H13

data_processing:
Illumina Casava1.8 software used for basecalling.
Clean reads recovered after filtering low quality reads and artificially redundant reads were mapped to the H. mediterranei genome with the Burrows-Wheeler Alignment tool (BWA)
The number of reads was calculated for every 1-kb window and plotted against the chromosome position
Genome_build: CP001868.2-CP001871
Supplementary_files_format_and_content: tab-delimited text files include number of reads for every 1kb window

organism: Haloferax mediterranei

source_name: Genomic DNA of Haloferax mediterranei H13 at exponential phase

growth_protocol: Cells  were grown at 37°C until OD600 reached 0.35 in AS-168 medium supplemented with uracil

extract_protocol:
Proteinase K, ethanol precipitation, RNaseA treatment, phenol-chloroform extraction as described previously [PMID: 24271389]
DNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: no treatment

value: delta-oriC4-cdc6H::trpA-oriP2-orc13 mutant of DFA50 delta-oriC1 delta-oriC2  delta-oriC3


title: ZmMADS47 RNAi mutant

city: Shanghai

first: zhenyi

last: qiao

address: 333 Nanchen Road, Life Science Building, Room 322

department: School of Life Science

institute: Shanghai University

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to B73 RefGen_v2 using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m 10 --best --strata
Fragments Per Kilobase of exon per Megabase of library size (FPKM) were calculated. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of fragments falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: B73 RefGen_v2
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

organism: Zea mays

source_name: kernel

extract_protocol:
RNA was extracted with Trizol reagent, followed by clean-up and DNase I treatment with QIAGEN RNeasy mini kit in accordance with the prescribed protocol provided with the kit.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: endosperm

value: 15 DAP

value: ZmMADS47 knockdown


title: wild type

value: wild type


title: Cas5t bound RNAs

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: RNA was extracted from Cas5t-immunopurified complexes using standard Trizol extraction. Total RNA fractionated on 10% polyacrylamide gel and size selected to enrich for RNAs of 20-70 nt, treated with thermosensitive alkaline phosphatase, and ligated to a 3' adaptor. Gel purified ligated products were then treated with T4 polynucleotide kinase and ligated to a 5' adaptor. The ligated RNAs were reverse transcribed, treated with RNaseH, and amplified by 16 cycles of PCR. Samples were sequenced on an Illumina Genome Analyzer IIx for 76 cycles.

data_processing:
Reads were trimmed to remove 3' linker sequence and aligned to the reference genome using bowtie-0.12.0-beta1 with parameters -m 1 -v 2 to identify uniquely mapping reads. Plus and minus strand wiggle tracks (bedGraphs) were made from the uniquely aligned reads.
Genome_build: pyrFur2
Supplementary_files_format_and_content: bedGraph - contains read density graphs for aligned reads to the plus and minus strand of the genome

organism: Pyrococcus furiosus

source_name: Cultured Organism

extract_protocol:
Protein RNA complexes were purified using antibodies specific to Cst2, Cas5t, Cas2, or Cas3-2. RNAs were then fractionated on 10% polyacrylamide gel and size selected to enrich for RNAs of 20-70 nt,
RNAs were treated with thermosensitive alkaline phosphatase, and ligated to a 3' adaptor. Gel purified ligated products were then treated with T4 polynucleotide kinase and ligated to a 5' adaptor. The ligated RNAs were reverse transcribed, treated with RNaseH, and amplified by 16 cycles of PCR.

tag: cell

value: Cultured organisms

tag: strain


title: Cas2 bound RNAs

description: RNA was extracted from Cas2-immunopurified complexes using standard Trizol extraction. Total RNA fractionated on 10% polyacrylamide gel and size selected to enrich for RNAs of 20-70 nt, treated with thermosensitive alkaline phosphatase, and ligated to a 3' adaptor. Gel purified ligated products were then treated with T4 polynucleotide kinase and ligated to a 5' adaptor. The ligated RNAs were reverse transcribed, treated with RNaseH, and amplified by 16 cycles of PCR. Samples were sequenced on an Illumina Genome Analyzer IIx for 76 cycles.


title: FigS7_scc2-4_IP

organism: Saccharomyces cerevisiae

value: K14710, scc2-4 ts allele, Scc1PK tag


title: Cas3-2 bound RNAs

description: RNA was extracted from Cas3-2-immunopurified complexes using standard Trizol extraction. Total RNA fractionated on 10% polyacrylamide gel and size selected to enrich for RNAs of 20-70 nt, treated with thermosensitive alkaline phosphatase, and ligated to a 3' adaptor. Gel purified ligated products were then treated with T4 polynucleotide kinase and ligated to a 5' adaptor. The ligated RNAs were reverse transcribed, treated with RNaseH, and amplified by 16 cycles of PCR. Samples were sequenced on an Illumina Genome Analyzer IIx for 76 cycles.


title: Cst2 bound RNAs

description: RNA was extracted from Cst2-immunopurified complexes using standard Trizol extraction. Total RNA fractionated on 10% polyacrylamide gel and size selected to enrich for RNAs of 20-70 nt, treated with thermosensitive alkaline phosphatase, and ligated to a 3' adaptor. Gel purified ligated products were then treated with T4 polynucleotide kinase and ligated to a 5' adaptor. The ligated RNAs were reverse transcribed, treated with RNaseH, and amplified by 16 cycles of PCR. Samples were sequenced on an Illumina Genome Analyzer IIx for 76 cycles.


title: FigS7_scc2-4_WCE


title: C. kristjanssonii untreated


title: FigS5_Scc1PK9_IP G1 releasing 15min

value: 60 minutes after releasing from alpha-factor arrest


title: FigS5_Scc1PK9_IP G1 releasing 30min

value: 45 minutes after releasing from alpha-factor arrest


title: FigS5_Scc1PK9_IP G1 releasing 60min

value: 15 minutes after releasing from alpha-factor arrest


title: FigS5_Scc1PK9_WCE G1 releasing 15min


title: FigS5_Scc1PK9_IP G1

value: alpha-factor arrest


title: Input WT Chromatin7

city: Martinsried

first: Tobias

last: Straub

address: Großhadener Str. 9

department: Biomedical Center, Bioinformatics

country: Germany

institute: LMU Munich

description: Sample 26

data_processing:
Alignment to dm3 without chromosome Uextra using bowtie version 1.1.1  (parameter -m 1)
Read extension to 200 bp and coverage vector calculation
Peaks were called using Homer findPeaks version 4.7 (-gsize 121400000 -style factor -size 200 -fragLength 200 -C 0)
bedGraph files were generated from coverage vectors using export.bedGraph (bioconductor package 'rtracklayer')
Genome_build: dm3
Supplementary_files_format_and_content: bedGraph and peaks.bed files

organism: Drosophila melanogaster

source_name: 0-12 hour old embryos

extract_protocol:
Un-staged embryos were dechorinated using 120 ml final, 1:5 diluted, sodium hypochloride for 3 min. The embryos were thoroughly washed and fixed in the fixing solution [30 ml n-Heptane solution and 10 ml fixing buffer containing 0.1 M NaCl, 0.05 M HEPES pH 8.0, 1 mM EDTA, 0.5 mM EGTA, 3.7% Formaldehyde for 15 min at 16-18°C on a rotating wheel. Fixation was then quenched by adding 125 mM, final, Glycine to the tube. The embryos were subsequently washed using PBS [+ 0.01% Triton-X100] for 10 min and stored at -80°C until further use. For nuclei isolation, embryos were slowly thawed and dounced using glass homogenizer A and B (20 strokes each) in ice-cold NX-I buffer [15 mM HEPES pH 7.6, 10 mM KCl, 2 mM MgCl2, 0.5 mM EGTA, 0.1 mM EDTA, 350 mM sucrose, 1 mM DTT, 0.2 mM PMSF, Protease inhibitors Leupeptin, Pepstatin and Aprotinin (10 µg/ml)]. The lysate was filtered through Miracloth and nuclei were spun down at 3500 rpm, 10 min, 4°C. The pellet (brown) was washed in NX-I buffer. Finally, the nuclei were resuspended in RIPA [1% Triton X-100, 0.1% Sodium deoxycholate, 140 mM NaCl, 10 mM Tris pH 8.0, 1 mM EDTA, 0.1% SDS, 1 mM PMSF] and washed 3 times. Nuclei were then counted and aliquoted to ~109 nuclei/ml. Fractions can be stored at -80°C until shearing. For shearing and ChIP, thawed nuclei were adjusted to ~2x108 in RIPA and sheared in Covaris S220 system (Covaris Inc. MA, USA) at power 110 Watts, duty factor 20%, and cycles per burst 200 for 25 min. Chromatin was pre-cleared using protein A+G beads (1:1) mix for 1h, 4°C. 200 µl of chromatin was taken for chromatin immunoprecipitation in 500 µl reaction. The ChIP antibodies are all custom antibodies produced in our lab.  4 µl of antibodies (Rb1-ACF1, Rb2-ACF1, Rb-RSF1) were incubated per reaction overnight at 4°C. RIPA equilibrated protein A+G (1:1) mix was then added to pull down the immune-complexes for 3h, 4°C. For Rat monoclonal antibody, the chromatin immune-precipitation step was performed using presorbed protein G beads, in an excess of antibody, for 3hr at 4°C. Beads were washed subsequently 5 times in 1 ml RIPA buffer. Residual RNA was digested by RNase (10 µg) at 37°C for 20 min. Subsequent protein digestion (25 µg Proteinase K) and reversal of cross-linking were performed together at 68°C for 2 hrs. DNA was purified using GenElute™ PCR Clean-Up Kit (Sigma, Cat.no NA1020).
ChIP DNA was quantified using Qubit® dsDNA HS Assay Kit (Life technologies, Cat.no.Q32851) and sequencing libraries were prepared using MicroPlex Library Preparation™ kit (Diagenode, Cat. No. C05010011) by taking 2 ng of starting material whenever possible. PCR amplification was monitored by quantifying amplified libraries (maximum 19 cycles).

tag: developmental stage

value: 0-12 hour old embryos

value: wt

value: Oregon-R

tag: chip antibody

tag: chromatin batch number


title: FigS5_Scc1PK9_IP G1 releasing 45min

value: K23308, Scc1 PK tag

value: 30 minutes after releasing from alpha-factor arrest


title: Input rsf1 Chromatin1

description: Sample 25


title: Fig1_SacCer_50% Scc1PK9_IP

value: K14061, Scc1PK tag

value: exponentially grown

value: ChIP -of Scc1_PK9 with anti-PK antibody (Biorad, MCA1360)


title: FigS5_Scc1PK9_WCE G1 releasing 45min

value: none, input


title: FigS5_Scc1PK9_WCE G1 releasing 60min


title: IgG_vehicle_ChIPseq

city: Innsbruck

first: Klocker

last: Helmut

address: Anichstrasse 35

department: Department of Urology

country: Austria

institute: Medical University of Innsbruck

description: chip antibody: Rabbit IgG

data_processing:
ChIP-seq reads were aligned to thehg18 genome assembly using bowtie version 0.12.7
Sam files were converted to bed files.
Reads overlapping tandem repeats or segmentally duplicated regions (>1000bp) were removed.
Macs (version 1.4) was run to identify chipseq peaks
Genome_build: hg18
Supplementary_files_format_and_content: peak calls.  Peaks were called by macs using AR_androgen_ChIPseq as chipseq, and AR_vehicle_ChIPseq as control.

source_name: Prostate cancer cell line

growth_protocol: DUCaP cells were rountinely maintained in T175 flask in RPMI 1640 medium supplemented with 10% FBS and 2 mM L-Glutamine

extract_protocol:
Cellular nuclei were pelleted after the cells were incubated with hypotonic solution and Dounce-homogenized to release the nuclei. Nuclear lysates were clarified from sonicated nuclei, followed by preclearing with protein A/G agrose beads. AR-DNA complexes were immunoprecipitated with AR antibody and isolated with protein A/G agrose beads.
Sequencing libraries were prepared according to Illumina's ChIP-Seq Sample Prep Kit. Briefly, ChIP-DNA was end-repaired, adaptor ligated and PCR amplified, followed by cluster generation and sequencing-by-synthesis using the Genome Analyzer II (Illumina).

treatment_protocol: Cells were cultured in 10% charcoal/dextran-treated fetal calf serum (Fischer) for 72 hr before stimulated by synthetic androgen R1881 for 1 hr

value: DUCaP

value: Dura mater metastasis of a cancer patient


title: Fig7_E1155Q_WCE


title: IgG_androgen_ChIPseq

description: chip antibody: Rabbit IgG


title: Input acf1 Chromatin2

description: Sample 21


title: Input WT Chromatin5

description: Sample 23


title: Input WT Chromatin6

description: Sample 24


title: AR_androgen_ChIPseq

description: chip antibody: AR (cell signaling, #3202 and Millipore UB 06-680)


title: Input WT Chromatin4

description: Sample 22


title: Input acf1 Chromatin1

description: Sample 20


title: Fig7_E1155Q_IP

value: ChIP -of smc3(E1155Q)_PK6 with anti-PK antibody (Biorad, MCA1360)


title: AR_vehicle_ChIPseq


title: FigS5_Scc1PK9_WCE G1 releasing 30min


title: WT_MEFs_Adriamycin_rep3

city: Freiburg im Breisgau

first: Thomas

last: Manke

address: Stuebeweg 51

department: Bioinformatics and Deep-Sequencing Unit

institute: Max-Planck-Institute for Immunobiology and Epigenetics

data_processing:
Reads mapped to the mouse genome (GRCm38) using the Tophat (v2.0.13)
counted for Ensembl transcripts (release 78) using featureCounts (v1.4.5-p1)
Differential gene expression levels were determined using DESeq2(v1.4.1)
Genome_build: GRCm38
Supplementary_files_format_and_content: tab-delimited text files include log2FoldChange values for all genes comparing Control vs MOF depletion (DESeq2 output)

source_name: MEFs

growth_protocol: Cells were grown at 37C, 5% CO2. MEFs were maintained in DMEM with 10% FCS and antibiotics and podocytes in RMPI + 10% FCS +antibiotics. MEFs plated at 10,000 cells/cm2.

extract_protocol:
RNA was extracted using the Rneasy kit (Qiagen)
RNA libraries were prepared for sequencing using standard Illumina protocols with polyA enrichment

treatment_protocol: Differentiated podocytes (0.25 ug/ml) and freshly passaged MEFs (0.1 ug/ml)  were treated with Adriamycin for 24 hours.

tag: mof

tag: drug

value: Adriamycin

tag: passage

tag: cell type

value: MEFs


title: Fig7_QQ_WCE


title: Input WT Chromatin2

description: Sample 18


title: Fig7_QQ_IP

value: ChIP -of smc3(K112Q, K113Q)_PK6 with anti-PK antibody (Biorad, MCA1360)


title: Input WT Chromatin1

description: Sample 17


title: Fig7_WT_WCE


title: MofKO_MEFs_Adriamycin_rep2

value: ko


title: MofKO_MEFs_Adriamycin_rep3


title: Fig6_scc2-45_WCE

value: K22390, scc2-45 ts allele, Scc1PK tag


title: RSF1-Rb WT rep2

description: Sample 16

value: 0-12 hour old embryos


title: WT_MEFs_Adriamycin_rep2

value: wt


title: Fig7_WT_IP

value: ChIP -of Smc3_PK6 with anti-PK antibody (Biorad, MCA1360)


title: FigS5_Scc1PK9_WCE G1

value: K23308, Scc1 PK tag

value: alpha-factor arrest


title: RSF1-Rb WT rep1

description: Sample 15

value: Oregon-R


title: Fig6_scc2-45_IP

value: exponentially grown


title: RSF1-Rb rsf1 rep1

description: Sample 14


title: WT_MEFs_Adriamycin_rep1


title: Fig6_WT_WCE


title: Fig6_WT_IP

value: ChIP -of Scc1_PK9 with anti-PK antibody (Biorad, MCA1360)


title: WT_MEFs_vehicle_rep3

value: vehicle (DMSO)


title: Fig6_Scc1_PK9_WCE

value: none, input


title: ACF1-Preimmune-Rb WT rep1

description: Sample 13


title: MofKO_MEFs_vehicle_rep3


title: Fig6_Scc1_PK9_IP


title: ACF1-Preimmune-Rb2 WT rep1

description: Sample 11


title: ACF1-Preimmune-Rb2 WT rep2

description: Sample 12


title: WT_MEFs_vehicle_rep2


title: Input WT Chromatin3

description: Sample 19


title: Fig4_G1 releasing 120min_Scc1PK9_WCE

value: K14061, Scc1PK tag

value: 120 minutes after releasing from alpha-factor arrest


title: WT_MEFs_vehicle_rep1


title: ACF1-Rb2 WT rep2

description: Sample 9


title: MofKO_MEFs_vehicle_rep1


title: ACF1-Rb2 WT rep3

description: Sample 10


title: Fig4_G1 releasing 120min_Scc1PK9_IP


title: ACF1-Rb1 WT rep1

description: Sample 7


title: scramble_MPC5podocytes_vehicle_rep3

source_name: podocytes

value: scramble

value: podocytes


title: ACF1-Rb2 WT rep1

description: Sample 8


title: Fig6_Scc1_WCE

value: K699, no PK tag


title: Fig6_Scc1_IP

value: ChIP -of Scc1 without PK tag with anti-PK antibody (Biorad, MCA1360)


title: ACF1-Rb2 acf1rep2

description: Sample 6


title: scramble_MPC5podocytes_vehicle_rep2


title: ACF1-Rb2 acf1rep1

description: Sample 5


title: Fig4_G1 releasing 105min_Scc1PK9_WCE

value: 105 minutes after releasing from alpha-factor arrest


title: Fig4_G1 releasing 105min_Scc1PK9_IP


title: Fig4_G1 releasing 90min_Scc1PK9_WCE

value: 90 minutes after releasing from alpha-factor arrest


title: ACF1-Rb1 acf1 rep1

description: Sample 4

value: 0-12 hour old embryos


title: MofKO_MEFs_vehicle_rep2

value: ko

value: MEFs


title: Fig4_G1 releasing 90min_Scc1PK9_IP

value: K23308, Scc1 PK tag


title: scramble_MPC5podocytes_Adriamycin_rep3

value: Adriamycin


title: ACF1-3F1 WT rep3

description: Sample 3

value: wt

value: Oregon-R


title: ACF1-3F1 WT rep2

description: Sample 2


title: MofKO_MEFs_Adriamycin_rep1


title: scramble_MPC5podocytes_Adriamycin_rep1


title: scramble_MPC5podocytes_Adriamycin_rep2


title: Mof-knockdown_MPC5podocytes_vehicle_rep3

value: knockdown

value: vehicle (DMSO)


title: Fig4_G1 releasing 75min_Scc1PK9_WCE

value: K14061, Scc1PK tag

value: 75 minutes after releasing from alpha-factor arrest

value: none, input


title: ACF1-3F1 WT rep1

description: Sample 1


title: Fig4_G1 releasing 60min_Scc1PK9_WCE

value: 60 minutes after releasing from alpha-factor arrest


title: MCF10A_10E2_MYC-ER_control_ChIP-Seq

city: Wuerzburg

first: Martin

last: Eilers

address: Am Hubland

department: Chair for Biochemistry and Molecular Biology

institute: University of Wuerzburg

data_processing:
Basecalling was performed with the RTA package within the Genome Analyzer Data Collection Software (SCS2.10).
Fastq files were generated by CASAVA, considering only high quality sequences (PF-cluster).
Overall sequencing quality was checked with the FastQC script.
Reads were aligned to the human genome hg19 with BOWTIE  v0.12.8 using default parameters.
Peaks were called with MACS-1.4.2 using default parameters and input as control. Input sample is accessioned as GSM1423726.
Genome_build: hg19
Supplementary_files_format_and_content: .wig files generated with MACS using a 10 bps resolution

source_name: MCF10A cells

growth_protocol: MCF10A cells were maintained in DMEM/F-12, supplemented with 5% horse serum, 100ug/ml EGF, 0.5ug/ml hydrocortisone, 10ug/ml insulin, 100ng/ml cholera toxin and 1% penicillin/streptomycin.

extract_protocol:
Cells were cross-linked with 1% formaldehyde. After a hypotonic lysis step, nuclei were resuspended in sonication buffer containing 0.1% SDS and 0.1% Na-deoxycholate and sonication was performed in a Branson sonifier. Anti-ER-a antibody (HC20, Santa Cruz) was coupled to Protein A-Dynabeads, 10E2 mouse anti-MIZ1 antibody was coupled to Protein G-Dynabeads (Invitrogen). Bound chromatin was eluted with 1% SDS and after decrosslinking DNA was purified with phenol-chloroform.
ChIP DNA was end-repaired, A-tailed and ligated to Illumina adaptors using NEBNext ChIP-Seq Sample Preparation Kit. After size selection on an agarose gel and purification with a Qiagen gel extraction kit, PCR-amplification was performed. Library quality was tested with an Experion DNA Chip (BioRad) and quantified using a picogreen assay.

treatment_protocol: Cells were lentivirally transduced with MYC-ER and MYC-ERVD and stable pools were treated with 100nM 4-OHT or solvent control  for 24h.

tag: cell line

value: mammary gland/breast

value: 10E2 (anti-MIZ1)


title: Fig4_G1 releasing 45min_Scc1PK9_WCE

value: 45 minutes after releasing from alpha-factor arrest


title: Mof-knockdown_MPC5podocytes_vehicle_rep2

value: podocytes


title: Mof-knockdown_MPC5podocytes_vehicle_rep1


title: scramble_MPC5podocytes_vehicle_rep1

value: scramble


title: Mof-knockdown_MPC5podocytes_Adriamycin_rep3


title: Fig4_G1 releasing 45min_Scc1PK9_IP

value: ChIP -of Scc1_PK9 with anti-PK antibody (Biorad, MCA1360)


title: Fig4_G1 releasing 75min_Scc1PK9_IP


title: Fig4_G1 releasing 30min_Scc1PK9_WCE

value: 30 minutes after releasing from alpha-factor arrest


title: MCF10A_10E2_MYC-ER_OHT_ChIP-Seq

tag: tissue


title: Mof-knockdown_MPC5podocytes_Adriamycin_rep1


title: Fig4_G1 releasing 15min_Scc1PK9_WCE

value: K23308, Scc1 PK tag

value: 15 minutes after releasing from alpha-factor arrest


title: Fig4_G1 releasing 15min_Scc1PK9_IP


title: Fig4_G1 releasing 60min_Scc1PK9_IP


title: MCF10A_10E2_MYC-ERVD_OHT_ChIP-Seq


title: MCF10A_10E2_MYC-ERVD_control_ChIP-Seq


title: MCF10A_HC20_MYC-ER_control_ChIP-Seq

tag: antibody manufacturer

value: Santa Cruz

tag: antibody catalog number

tag: antibody lot number


title: Mof-knockdown_MPC5podocytes_Adriamycin_rep2

value: Adriamycin


title: Fig4_G1 releasing 30min_Scc1PK9_IP

tag: sac_cer strain

tag: gla_gra strain

tag: sac_cer and gla_gla mixed percentage


title: Fig4_G1 Scc1PK9_IP

tag: condition for cell growth

value: alpha-factor arrest

tag: antibody


title: MCF10A_HC20_MYC-ERVD_control_ChIP-Seq


title: Fig2_SacCer_100% Scc1PK_WCE

value: exponentially grown


title: MCF10A_10E2_Vector_ChIP-Seq


title: RNA-Seq 786-O Vector Alone 3

city: Oxford

first: Rafik

last: Salama

address: Roosevelt Drive

institute: University of Oxford

description: 786-0 VHL defective Clear Cell Renal Cell Line

data_processing:
ChIP-seq: Illumina adaptor sequences were trimmed from the ChIP-seq reads using Trimgalore (0.3.3) and the reads were then aligned to Genome Reference Consortium GRCh37 (hg19) using BWA (0.7.5a-r405)
ChIP-seq: Low quality mapping was removed (MapQ < 15) using SAMtools (0.1.19) and reads mapping within the Duke Encode black list regions (http://hgwdev.cse.ucsc.edu/cgi-bin/hgFileUi?db=hg19&g=wgEncodeMapability) were excluded using BEDTools (2.17.0)
ChIP-seq: Duplicate reads were marked using Picard tools (1.106) (http://picard.sourceforge.net/) and BigWig files generated for visualization
ChIP-seq peaks were identified using T-PIC (Tree shape Peak Identification for ChIP-Seq) and MACS (Model-based analysis of ChIP-Seq)
Illumina adaptors were trimmed from the RNA-seq reads using Trim Galore 0.3.3 (http://www.bioinformatics.babraham.ac.uk/projects/trim_galore/).
Reads were then aligned to Genome Reference Consortium GRCh37 using Tophat 2.0.8b (http://ccb.jhu.edu/software/tophat/index.shtml) and bowtie 1.0.0 (http://bowtie-bio.sourceforge.net/index.shtml) and non-uniquely mapping fragments excluded using SAMtools (0.1.19)
The total read counts for each gene (as defined by UCSC genes definition) were extracted using HTSeq (0.5.4p3) with ‘intersection-strict’ mode. DESeq2 R package was used to identify significantly induced or repressed genes.
Genome_build: hg19

source_name: Renal Cancer Cell Line

growth_protocol: 786-O and HEK293 cells were grown in Dulbecco modified Eagle medium supplemented with 10% foetal calf serum, 2 mM L-glutamine, 100U penicillin and streptomycin 50 U/ml (v/v) (Sigma-Aldrich)

extract_protocol:
ChIP-seq: Cells were fixed with 1% formaldehyde and quenched with glycine (125mM). After washing, cells were lysed (1%SDS, 10mM EDTA, 50mM Tris-HCl pH 8.1) and sonicated (Diagenode Biorupter, Belgium). Chromatin was immunoprecipitated using rabbit polyclonal antisera to HIF-1? (PM14), HIF-2? (PM9), HIF-1? (NB-100-110, Novus Biologicals, UK) or pre-immune serum as control. Eluted chromatin was heated to reverse cross-linking, treated with proteinase-K and purified with phenol:chloroform:isoamyl alcohol and ethanol precipitation
RNA-seq: Total RNA was prepared using the mirVana miRNA Isolation Kit (Ambion; Life Technologies Ltd, Paisley, UK) and treated with DNaseI (TURBO DNA‐free™, Ambion). Directional PolyA+ RNA libraries were then prepared using the ScriptSeq™ v2 RNA‐Seq kit (Epicentre, Madison, WI, USA).
Standard Illumina protocol

treatment_protocol: HIF-1a and HIF-2a cDNA sequences were first cloned into pRRL.IRES.EGFP (kind gift from Kamil Kranc, Glasgow) to generate bicistronic vectors. These pRRL vectors were then co-transfected into HEK293T cells with plasmids pCMV-?R8.2 and pCMC-VSVG and the resultant viral particles were isolated by centrifugation and ultrafiltration. 786-0 cells (ATCC) were then transduced with either control (pRRL), HIF-1a (pRRL-HIF-1a) or HIF-2a (pRRL-HIF-2a) expressing virus.

tag: transfection

value: control (pRRL)


title: Fig4_G1 Scc1PK9_WCE

value: K14061, Scc1PK tag


title: RNA-Seq 786-O Vector Alone 2


title: Fig2_SacCer_100% Scc1PK_IP


title: MCF10A_HC20_MYC-ERVD_OHT_ChIP-Seq


title: RNA-Seq 786-O Vector Alone 1


title: Fig2_SacCer_80% Scc1PK_WCE

value: none, input


title: shMyt3-transduced

city: Vancouver

first: Brad

last: Hoffman

state: BC

country: Canada

institute: University of British Columbia

description: Sample 2

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm9 whole genome using bowtie2 v2.2.1
Transcript splice junctions were determined using Tophat v2.0.11
Transcripts were assembled and differences in expression were determined using Cufflinks v2.2.1
Genome_build: mm9
Supplementary_files_format_and_content: cuffdiff output.  Sample names used in processed data file are given in the "Description" field above. All sample comparisons are provided in this file of concatenated output. Gene IDs are UCSC Genome Browser ids.

growth_protocol: Transduced Isolated whole pancreatic islets were cultured on 804G-treated plates for 7 days in RPMI

extract_protocol:
RNA was harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit (Cat#FC-122-1001) was used with 800ng of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: After isolation islets were transduced with adenoviruses expressing a control shRNA, shMyt3 or a Myt3 over-expression construct overnight. Media was changed the next day to fresh RPMI

tag: strain

tag: age

value: 8-12 Weeks

value: pancreas

tag: tissue compartment

value: Islets


title: Fig2_SacCer_80% Scc1PK_IP


title: shScramble-transduced

description: Sample 1


title: Myt3-transduced

description: Sample 3


title: Fig2_SacCer_60% Scc1PK_WCE


title: Fig2_SacCer_60% Scc1PK_IP


title: RNA-Seq 786-O with HIF-2a over-expression 2

value: HIF-2a (pRRL-HIF-2a)


title: RNA-Seq 786-O with HIF-1a re-expression 3

value: HIF-1a (pRRL-HIF-1a)


title: Fig1_SacCer_100% Scc1PK_IP

source_name: Saccharomyces cerevisiae cells


title: RNA-Seq 786-O with HIF-2a over-expression 3


title: RNA-Seq 786-O with HIF-1a re-expression 2


title: Fig1_SacCer_100% Scc1PK_WCE


title: RNA-Seq 786-O with HIF-1a re-expression 1


title: ChIP-Seq of HIF-1b in 786-O with HIF-2a over expression

tag: chip antibody

value: Novus Biological, NB100-110


title: Input DNA in 786-O with HIF-2a over expression


title: Input DNA in 786-O with HIF-1a re-expression


title: Input DNA in 786-O Vector Alone


title: ChIP-Seq of HIF-2a in 786-O Vector Alone

value: HIF-2alpha Antibody (PM9)


title: ChIP-Seq of HIF-1b in 786-O with HIF-1a re-expression


title: ChIP-Seq of HIF-2a in 786-O with HIF-1a re-expression


title: ChIP-Seq of HIF-1a in 786-O with HIF-1a re-expression

value: HIF-1alpha Antibody (PM14)


title: ChIP-Seq of HIF-2a in 786-O with HIF-2a over expression


title: pro∆GRAS_Pac_2

status: Public on Jul 09 2015

city: Rehovot

first: Gilgi

last: Friedlander

address: Hertzel st

country: Israel

institute: Weizmann Institute of Science

data_processing:
Base call and quality scoring was done using RTA (1.18.64).
Reads were aligned to the tomato genome sequence SL2.50 (downloaded from the sol genomics network http://solgenomics.net/organism/Solanum_lycopersicum/genome) using Tophat (v2.0.10).
Counting reads on ITAG2.4 genes (downloaded from sol genomics network) was done with HTSeq-count (version 0.6.1p1)
Differential analysis was carried out using DESeq2 (1.6.3)
Genome_build: SL2.50
Supplementary_files_format_and_content: Tab delimited file including the raw counts, the normalized counts and ratios and p-values (and corrected p-values).

organism: Solanum lycopersicum

source_name: Seedlings young leaves

growth_protocol: Tomato plants were grown in a greenhouse under 24/20 ºC (day/night) at natural day length conditions

extract_protocol:
RNA was extracted using the GHC-Phenol chloroform method. cDNA was generated using the SuperScript® II Reverse Transcriptase (Invitrogene)
500ng of total RNA was processed using the TruSeq RNA Sample Preparation Kit v2 protocol (Illumina) (Part # 15026495). Libraries were evaluated by Qubit and TapeStation. Sequencing libraries were constructed with barcodes to allow multiplexing of 8 samples on one lane of flow cell v4 (run on Illumina HiSeq Rapid 2500).

treatment_protocol: M82 and pro∆GRAS seedlings were treated with Paclobutrazol (10mg/l) once a day for three days followed by a single GA3 application (100 µM).

tag: genotype

value: pro{delta}GRAS

tag: treatments

value: Paclobutrazol 10mg/l


title: pro∆GRAS_Pac+GA_2

value: Paclobutrazol 10mg/l + GA 100 µM


title: pro∆GRAS_Pac+GA_1


title: M82_Pac+GA_2


title: pro∆GRAS_Pac_1


title: M82_Pac_2


title: ChIP-Seq of HIF-1b in 786-O Vector Alone

tag: cell line


title: RNA-Seq 786-O with HIF-2a over-expression 1


title: forebrain_H3K27me3_ChIPseq

city: Bronx

first: Shira

last: Rockowitz

address: 1300 Morris Park Ave

institute: Albert Einstein College of Medicine

data_processing:
The ChIP-seq reads were aligned to the mouse genome (GRCm37/mm9).
Peak detection was performed with the MACS2 and SICER algorithms.
ChIP-seq peak calling: H3K27me3 peaks called using input_1 as control, H3K4me1/3 and H3K27ac peaks called using input_2 as control, Pax6 peaks called using Input_3 and input_4 (in forebrain) as control, Pol2 peaks called using input_3 as control.
Supplementary_files_format_and_content: ChIP-seq: Tab-delimited text file includes peaks for the samples.
Supplementary_files_format_and_content: SICER-output: chrom, start, end, ChIP_island_read_count, CONTROL_island_read_count, p_value, fold_change, FDR_threshold.
Genome_build: MGSCv37 (mm9)

source_name: e12.5 embryonic forebrain, H3K27me3 ChIP

extract_protocol:
ChIP-seq: The E12.5 forebrain and newborn (P1) lenses of CD-1 mice (Charles River Laboratories) were dissected, tissue was freshly crosslinked and homogenized. Sonicated lysates were clarified and histone-DNA complexes were isolated with antibody.
Libraries were prepared according to Illumina's instructions.

tag: strain/background

tag: developmental stage

value: embryo

tag: tissue

value: forebrain


title: M82_Pac_1


title: lens_H3K27me3_ChIPseq

source_name: newborn lens, H3K27me3 ChIP

value: newborn

value: lens


title: forebrain_input_1

data_processing:
The ChIP-seq reads were aligned to the mouse genome (GRCm37/mm9).
Peak detection was performed with the MACS2 and SICER algorithms.
ChIP-seq peak calling: H3K27me3 peaks called using input_1 as control, H3K4me1/3 and H3K27ac peaks called using input_2 as control, Pax6 peaks called using Input_3 and input_4 (in forebrain) as control, Pol2 peaks called using input_3 as control.
Supplementary_files_format_and_content: ChIP-seq: Tab-delimited text file includes peaks for the samples.
Genome_build: MGSCv37 (mm9)

source_name: e12.5 embryonic forebrain, input


title: PM23_T3

city: Paris

first: Laurence

last: Tiret

address: 91 Bd de l'Hôpital

country: France

institute: INSERM

description: sequenced twice in 2 different lanes

data_processing:
Filtering of bad quality reads using fastq_illumina_filter
Trimming  using sickle with default parameters (quality threshold=20, length threshold=20).
Quality of reads  checked using FastQC
Paired-end reads mapped to the human transcriptome using Bowtie2 v2.1.0 .
Quantification of transcript abundance  using eXpress
Genome_build: Reference human transcriptome  generated from the UCSC reference genome (hg19) and the Illumina iGenome annotation file of transcripts using RSEM
Supplementary_files_format_and_content: Data used for analysis are the eff_counts provided by eXpress. Data are provided at the gene level by summing isoform counts for each gene.
Supplementary_files_format_and_content: Rows of the matrix correspond to genes identified by Gene symbol and Ensembl identifier. Columns correspond to individuals at each time point (T0 or T3)

source_name: obese women_after bariatric surgery

extract_protocol:
Total RNA was extracted using the mirVana™ miRNA Isolation Kit (Ambion®). 500 ng of total RNA were used to prepare the library with TruSeq RNA sample Prep Kit V2 (Illumina). Briefly, mRNA was selected from total RNA by poly-T oligo-attached magnetic beads and fragmented. Reverse transcription was done with random hexamers by SuperScript II reverse transcriptase.
RNA libraries were prepared for sequencing using standard Illumina protocols.The generated libraries were submitted to quality control on High Sensitivity DNA chip from the Agilent Bioanalyzer 2100 (Agilent Technologies, Wokingham, UK) and quantified by fluorimetry using the Quant-iT™ dsDNA Assay Kit (Life Technology). A dilution at 10nM was performed for each library and pools of 12 libraries were constituted. Each pool included the libraries from 3 individuals for the 2 time points, each library being identifiable by a specific index. The resulting pools were loaded on a High Sensitivity DNA chip (Agilent) and quantified by integration. 9 pM of this solution was used for cluster generation using the TruSeq PE Cluster Kit v3 (Illumina) on a CBot instrument. A 75 bp paired-end run was performed on the Illumina HiSeq2000 platform with the TruSeq SBS kit V3 (Illumina). Sequencing was performed on two 8-laned flow cells. Each flow cell included 4 pools sequenced twice in 2 different lanes.

treatment_protocol: Venous blood samples were collected in the morning after a 12-hour fast. Monocytes were isolated from whole blood using Dynabeads® CD14 (Invitrogen™).

tag: subject id

tag: time point

value: T3; 3 months after bariatric surgery

tag: type of surgical procedure

value: adjustable gastric banding

tag: cell type

value: peripheral monocytes


title: PM22_T3

description: sequenced twice in 2 different lanes

value: Roux-en-Y gastric bypass


title: PM21_T3

description: sequenced twice in 2 different lanes


title: PM20_T3

description: sequenced twice in 2 different lanes


title: PM18_T3

description: sequenced twice in 2 different lanes


title: PM19_T3

description: sequenced twice in 2 different lanes


title: PM16_T3

description: sequenced twice in 2 different lanes


title: PM13_T3

description: sequenced twice in 2 different lanes


title: PM17_T3

description: sequenced twice in 2 different lanes


title: PM11_T3

description: sequenced twice in 2 different lanes


title: PM09_T3

description: sequenced twice in 2 different lanes


title: PM12_T3

description: sequenced twice in 2 different lanes


title: M82_Pac+GA_1


title: PM05_T3

description: sequenced twice in 2 different lanes


title: PM08_T3

description: sequenced twice in 2 different lanes


title: PM02_T3

description: sequenced twice in 2 different lanes


title: PM06_T3

description: sequenced twice in 2 different lanes


title: lens_input_1

source_name: newborn lens, input


title: PM01_T3

description: sequenced twice in 2 different lanes


title: PM23_T0

description: sequenced twice in 2 different lanes

source_name: obese women_before bariatric surgery

value: T0; before bariatric surgery


title: PM22_T0

description: sequenced twice in 2 different lanes


title: PM19_T0

description: sequenced twice in 2 different lanes


title: PM20_T0

description: sequenced twice in 2 different lanes


title: PM17_T0

description: sequenced twice in 2 different lanes


title: PM15_T0

description: sequenced twice in 2 different lanes


title: PM16_T0

description: sequenced twice in 2 different lanes


title: PM21_T0

description: sequenced twice in 2 different lanes


title: PM18_T0

description: sequenced twice in 2 different lanes


title: PM11_T0

description: sequenced twice in 2 different lanes


title: PM08_T0

description: sequenced twice in 2 different lanes


title: PM10_T0

description: sequenced twice in 2 different lanes


title: PM12_T0

description: sequenced twice in 2 different lanes


title: PM15_T3

description: sequenced twice in 2 different lanes


title: PM10_T3

description: sequenced twice in 2 different lanes


title: PM02_T0

description: sequenced twice in 2 different lanes


title: PM05_T0

description: sequenced twice in 2 different lanes


title: OSCs transfected with construct: Actin promoter – flam (D.simulans) - luciferase-lacz, PIWI IP

city: Gneveva

first: Ramesh

last: Pillai

address: 30, Quai Ernest-Ansermet

department: Department of Molecular Biology

country: Switzerland

institute: University of Geneva

description:
small RNA
Piwi IP

data_processing:
Reads were sorted into individual libraries based on the barcodes and the 3’ adapter sequences were clipped from the reads using cutadapt (DOI:http://dx.doi.org/10.14806/ej.17.1.200). Reads were then aligned to the specific reporter sequence using bowtie (Langmead et al., 2009) allowing no mismatches.
For the mouse libraries the reads were sorted into individual libraries based on the barcodes and 3’ adapter was trimmed. Reads were mapped to the mouse genome mm9. The software used for processing the data (genomic coordinates etc) from the raw data files are in-house tools from Sachidanandam lab. They are described in a publication: Olson, A.J., Brennecke, J., Aravin, A.A., Hannon, G.J. & Sachidanandam, R. Analysis of large-scale sequencing of small RNAs. Pac Symp Biocomput, 126-36 (2008).
genome build: mm9 for mouse libraries, the reads from insect libraries were mapped to specific transfected or knocked-in reporter sequence
processed data files format and content: Mouse proccessed files contain mapping of the reads to mouse genome (mm9). The processed files for insect libraries contain mapping of the reads to specific reporter sequence which was transfected into the cells or knocked-in into the fly genome .

source_name: OSCs

extract_protocol:
For preparation of small RNA libraries total RNA was isolated from the lysates or RNA was immunoprecipitated from the protein complexes with the indicated antibodies against PIWI protein and the RNAs were resolved by 15% or 20% urea-PAGE. Bands corresponding to piRNAs or the indicated size were excised from the gel and extracted with 400 µl of 0.3M NaCl solution at 25°C overnight. For sequencing mouse long RNAs, testicular RNAs were isolated by TRIZOL.
Small RNA sequencing libraries  were prepared using NEBNext® Multiplex Small RNA Library Prep Set for Illumina® (Cat No:E7300) according to manufacturer instructions.  For mouse total RNA libraries strand-specific RNA-seq libraries were prepared using TruSeq Stranded Total RNA Sample Preparation Kit with Ribo-Zero (mouse) (Ilumina). Libraries were sequenced on Illumina HiSeq platform (EMBL Heidelberg Gene core facility)

treatment_protocol: OSCs and BmN4 cells were transfected with specific reporters. Transgenic flies were created by knocking-in the piRNA reporter sequence into Chr3 via ΦC31-mediated integration (Genetic Services Inc., USA).  The pi-Chr17-ΔMyb mice was generated using Cas9-sgRNA mediated genome editing system, in collaboration with Taconic Artemis company, resulting in a deletion of chr17: 27461948 -27462175 region (mm9).

value: transfected OSCs


title: PM01_T0

description: sequenced twice in 2 different lanes


title: transgenic flies with piRNA reporter, Ago3 IP [RR211]

description:
small RNA
Ago3 IP

source_name: female fly abdomen

value: female fly abdomen


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +432-lacz, Piwi IP [RR223]

description:
small RNA
Piwi IP


title: PM13_T0

description: sequenced twice in 2 different lanes


title: transgenic flies with piRNA reporter, Ago3 IP [RR210]

description:
small RNA
Ago3 IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +636-lacz, Piwi IP [RR224]

description:
small RNA
Piwi IP


title: transgenic flies with piRNA reporter, Aub IP [RR212]

description:
small RNA
Aub IP


title: transgenic flies with piRNA reporter, Aub IP [RR185]

description:
small RNA
Aub IP


title: transgenic flies with piRNA reporter, Aub IP [RR184]

description:
small RNA
Aub IP


title: transgenic flies with piRNA reporter, Piwi IP [RR209]

description:
small RNA
Piwi IP


title: PM09_T0

description: sequenced twice in 2 different lanes


title: OSCs transfected with construct: Actin promoter - egfp - lacz, Piwi IP [RR166]

description:
small RNA
Piwi IP


title: transgenic flies with piRNA reporter, total small RNAs from ovaries [DH36]

description:
small RNA
total RNA

source_name: fly ovaries

value: fly ovaries


title: transgenic flies with piRNA reporter, Piwi IP [RR182]

description:
small RNA
Piwi IP


title: transgenic flies with piRNA reporter, total small RNAs from ovaries [DH37]

description:
small RNA
total RNA


title: OSCs transfected with construct: Actin promoter - egfp - lacz, Piwi IP [RR155]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: flam promoter - flam-1624 to +112 - luciferase, Piwi IP [RR192]

description:
small RNA
Piwi IP


title: PM06_T0

description: sequenced twice in 2 different lanes


title: OSCs transfected with construct: Actin promoter - flam+1 to +718-luciferase-lacz-tRNA, Piwi IP [RR195]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +519-lacz, Piwi IP [RR225]

description:
small RNA
Piwi IP


title: transgenic flies with piRNA reporter, Aub IP [RR213]

description:
small RNA
Aub IP


title: OSCs transfected with construct: Actin promoter -luciferase-lacz, Piwi IP [RR193]

description:
small RNA
Piwi IP


title: transgenic flies with piRNA reporter, Piwi IP [RR183]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+168 to +718-lacz, Piwi IP [RR226]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - flam+1 to +718-egfp-lacz, Piwi IP [RR156]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +718-lacz-malat, Piwi IP [RR198]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - flam+1 to +718-egfp-lacz, Piwi IP [RR167]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +718-lacz-tRNA, Piwi IP [RR197]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - flam+1 to +718-luciferase-lacz, Piwi IP [RR194]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - flam+1 to +718-egfp-lacz, Piwi IP [RR161]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp - lacz, Piwi IP [RR160]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: flam promoter - flam-1624 to +718 - luciferase, Piwi IP [RR154]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +718-lacz, Piwi IP [RR157]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +718-lacz, Piwi IP [RR168]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +718-lacz, Piwi IP [RR162]

description:
small RNA
Piwi IP


title: OSCs transfected with construct: Actin promoter - egfp-flam+1 to +718-lacz, Piwi IP [RR151]

description:
small RNA
Piwi IP


title: S65

city: Detroit

first: Stephen

last: Krawetz

state: MI

address: 275 E. Hancock

department: Center for Molecular Medicine and Genetics

institute: Wayne State University School of Medicine

description:
sperm RNA
Phenotype key
Group: IVF treatment/birth outcome
Group I: TIC/achived LB within one spermatogenic cycle
Group II-i: IUI/TIC - delayed past first 90 day cycle
Group II-ii: ART preceded by unsuccessful IUI or TIC
Group II-iii: ART
Group III-i: Site 2 (Harvard) samples
Group III-ii: Patients never achieving LB/female factor observed

data_processing:
Basecalling Illumina RTA 1.17.21.3
demultiplexing (Casava 1.8.2, Illumina)
Alignment to hg19+ERCC+18S/28S rRNA using Bowtie39 (v. 2.2.3.0) and splice junction mapper Tophat38 (v.2.0.12)
Total number of reads (primary) aligning within each sample to 278605 RNA elements - comprising all UCSC annotated elements plus 842 previously identified elements occuring uniquely in sperm - were summed using MultiBamCov (bedtools 2.17.0)
Coverage for each element was normalized by element length, and percentile rank within each sample for every element was calculated
Genome_build: hg19 + ERCC + 18S/28S rRNA
Supplementary_files_format_and_content: ClusterData_PercRank_GEOSubmission.xlsx contains for each sample, percentile rank (within sample) of element abundance for each of ~280K genomic elements.

source_name: sperm

extract_protocol:
Neat frozen (-80°C) semen samples from ejaculate obtained with permission of male partner from couples undergoing fertility treatment at two clinics - Serono(S) and Harvard(H) - were  thawed then immediately subjected to purification through a 50% gradient of Puresperm (NidaCon International AB) in order to obtain pure sperm
Sperm RNA was isolated and purified using a modified RNeasy (Qiagen) protocol and finally the RNA quality assessed for DNA contamination as described in paper
A total of 2 ng of unfractionated input RNA along with the recommended concentration of ERCC RNA spike-in controls (Life Technologies) was reverse transcribed and the resulting cDNA amplified using SeqPlex RNA Amplification (Sigma-Aldrich Co.) allowing for the use of samples with low quantity of spermatozoa
Amplified primer dimers were removed prior to library construction using Agencourt AMPure XP (Beckman Coulter) and libraries were constructed with 50 ng of amplified cDNA using NEBNext Ultra DNA Library Prep Kit for Illumina (New England Biolabs) kit.
6bp inline Illumina index primers allowed for multiplexing of samples (Index in file name)
Paired-end sequencing (2x51  + 6Index) was performed  using an Illumina HiSeq 2500 sequencer

tag: phenotype group (birth outcome)

value: Group III-ii

tag: source site

value: Serono - Toronto

tag: sequencing date

value: sperm


title: H5

value: Group III-i

value: Harvard


title: OSCs transfected with construct:  luciferase (pGl3 control), Piwi IP [RR131]

description:
small RNA
Piwi IP


title: S67


title: OSCs transfected with construct: flam promoter - flam-1624 to +718 - luciferase, Piwi IP [RR130]

description:
small RNA
Piwi IP


title: S66


title: OSCs transfected with construct:  luciferase (pGl3 control), Piwi IP [RR128]

description:
small RNA
Piwi IP


title: H3


title: H4


title: H2


title: S62

value: Group II-iii


title: H1


title: S63


title: S61


title: S60


title: S57


title: S59


title: S58


title: S53


title: S54


title: S55


title: S56


title: OSCs transfected with construct: flam promoter - flam-1624 to +718 - luciferase, Piwi IP [RR133]

description:
small RNA
Piwi IP


title: S64


title: S49

value: Group II-ii


title: S51


title: S52


title: S48


title: S50


title: S47


title: S46


title: S43


title: S45


title: S40


title: S41


title: S37


title: S38


title: S42


title: S35

value: Group II-i


title: S34


title: S33


title: S39


title: S36


title: S30


title: S29


title: S31


title: lens_epithilium_RNAseq

description: Processed data file: gene.rpkm.matrix.txt

data_processing:
The RNA-seq reads were aligned to the mouse genome (GRCm37/mm9) using TopHat (v2).
The Cuffdiff tool in the Cufflinks package (v2) was used to determine RPKM (Reads Per Kb exons per Million mapped reads).
Supplementary_files_format_and_content: RNA-seq: Tab-delimited text file includes RPKMs for each sample.
Supplementary_files_format_and_content: gene.rpkm.matrix.txt: gene_name, e12fb_RPKM, lensepi_RPKM, lensfiber_RPKM.
Genome_build: MGSCv37 (mm9)

source_name: newborn lens epithelium

extract_protocol:
RNA-seq: The E12.5 forebrain and newborn (P1) lenses of CD-1 mice (Charles River Laboratories) were dissected and homogenized, total RNA was extracted.
Libraries were prepared according to the following directionalRNASeq dUTP Method:
http://wasp.einstein.yu.edu/index.php/Protocol:directional_WholeTranscript_seq

value: newborn

value: lens epithelium


title: S27


title: lens_fiber_RNAseq

source_name: newborn lens fiber

value: lens fiber


title: forebrain_RNAseq

source_name: e12.5 embryonic forebrain

value: embryo

value: forebrain


title: S28


title: S44


title: forebrain_input_2


title: S24


title: lens_Pol2_ChIPseq

data_processing:
The ChIP-seq reads were aligned to the mouse genome (GRCm37/mm9).
Peak detection was performed with the MACS2 and SICER algorithms.
ChIP-seq peak calling: H3K27me3 peaks called using input_1 as control, H3K4me1/3 and H3K27ac peaks called using input_2 as control, Pax6 peaks called using Input_3 and input_4 (in forebrain) as control, Pol2 peaks called using input_3 as control.
Supplementary_files_format_and_content: ChIP-seq: Tab-delimited text file includes peaks for the samples.
Supplementary_files_format_and_content: narrowPeak: chrom, start, end, name, score, strand, fold-change, -log10pvalue, -log10qvalue, relative summit position to peak start.
Genome_build: MGSCv37 (mm9)

source_name: newborn lens, Pol2 ChIP

value: lens

value: RNA polymerase II (Abcam, ab817, lot# GR153063-402)


title: S26


title: lens_input_2


title: S22


title: lens_H3K27ac_ChIPseq

source_name: newborn lens, H3K27ac ChIP


title: forebrain_Pol2_ChIPseq

source_name: e12.5 embryonic forebrain, Pol2 ChIP


title: S20


title: forebrain_H3K27ac_ChIPseq

source_name: e12.5 embryonic forebrain, H3K27ac ChIP


title: S17


title: S21


title: S18


title: S32


title: forebrain_H3K4me1_ChIPseq

source_name: e12.5 embryonic forebrain, H3K4me1 ChIP


title: S16


title: S15


title: S14


title: S13


title: S25


title: forebrain_input_3


title: lens_Pax6_ChIPseq

source_name: newborn lens, Pax6 ChIP

value: Pax6 (Millipore, ab2237, lot# NG1731333)


title: forebrain_Pax6_ChIPseq_rep2

description: Processed data file: fb_Pax6_macs2_peaks.narrowPeak

source_name: e12.5 embryonic forebrain, Pax6 ChIP


title: lens_input_3


title: pi-Chr17-ΔMyb (-/-), total testicular RNA [RR190]

description:
long RNAs
total RNA
RR181_RR190_mappers.csv

source_name: mouse testis

value: testis


title: S23


title: pi-Chr17-ΔMyb (-/-), Miwi IP [RR176]

description:
small RNA
Miwi IP
RR171-RR176_mappers.csv


title: forebrain_Pax6_ChIPseq_rep1


title: forebrain_input_4

source_name: e12.5 embryonic forebrain, input


title: pi-Chr17-ΔMyb (+/-), total testicular RNA [RR181]


title: pi-Chr17-ΔMyb (-/-), Mili IP [RR173]

description:
small RNA
Mili IP
RR171-RR176_mappers.csv


title: S11


title: S12


title: pi-Chr17-ΔMyb (+/-), Mili IP [RR171]


title: GF4

city: Cork

first: Roman

middle: M

last: Stilling

address: Western Road

department: Anatomy and Neuroscience

country: Ireland

institute: University College Cork

data_processing:
Base calling was achieved using standard Illumina CASAVA pipeline
Fastq-format reads were quality filtered and trimmed using Trimmomatic (v0.32)
Aligment to the mouse reference genome (GRCm38.p3) was achieved using the STAR aligner (v2.3.1u )
Ensembl database release 78 gene models were also used for counting mapped reads per gene using HTSeq-Count (v0.6.0)
Differential gene expression was calculated for pairwise comparisons using the DESeq2 R-package (v1.6.2) with default parameters. Genes with an FDR-adjusted p-value ≤ 0.1 were considered differentially regulated.
Genome_build: GRCm38.p3
Supplementary_files_format_and_content: .txt - tab delimited files with read counts (HTSeq)
Supplementary_files_format_and_content: .xlsx - Excel file with expression values, fold change, p-values and gene symbol (DESeq2) - three excel sheets in one excel workbook

molecule: polyA RNA

source_name: hand dissected unilateral amydala from fresh brain tissue

extract_protocol:
Total RNA was extracted using the mirVanaTM miRNA isolation kit (Ambion/life technologies) and Dnase treated (Turbo DNA-free, Ambion/life technologies) according to the manufacturers recommendations. RNA concentrations and quality were determined using Nanodrop 1000 and Bioanalyzer. Equal amounts of RNA from two to three animals were then pooled to yield four samples per group
was performed externaly by Beckman Coulter Genomics (BCG)

tag: strain

value: Swiss Webster

value: Amygdala

value: 10 weeks

tag: colonization status

value: germ-free


title: S5

value: Group I

value: Serono - Toronto

value: sperm


title: GF1

tag: age


title: S4


title: S19

value: Group II-i


title: S10


title: exGF4

value: germ-free colonized from p21 (weaning)


title: lens_H3K4me3_ChIPseq

source_name: newborn lens, H3K4me3 ChIP

value: newborn

tag: chip antibody


title: lens_H3K4me1_ChIPseq

source_name: newborn lens, H3K4me1 ChIP


title: CON3

value: conventional


title: forebrain_H3K4me3_ChIPseq

source_name: e12.5 embryonic forebrain, H3K4me3 ChIP

value: embryo


title: S9

source_name: sperm


title: S8


title: exGF3


title: BmN4 transfected with Ago3-artif reporter - Siwi IP [DH18]

description:
small RNA
Siwi IP

organism: Bombyx mori

source_name: BmN4 cells

value: transfected BmN4 cells


title: BmN4 transfected with Ago3-artif reporter - Ago3 IP [DH17]

description:
small RNA
Ago3 IP


title: BmN4 transfected with Ago3-artif reporter - Ago3 IP [DH16]

description:
small RNA
Ago3 IP


title: BmN4 transfected with Ago3-endo reporter - Siwi IP [DH14]

description:
small RNA
Siwi IP


title: BmN4 transfected with Ago3-endo reporter - Ago3 IP [DH12]

description:
small RNA
Ago3 IP


title: Pc-C-BioTAP_IP_S2_replicate1

city: Boston

first: Peter

last: Park

address: 10 Shattuck St

department: Center for Biomedical Informatics

institute: Harvard Medical School

data_processing:
Sequencing reads were aligned to the fly genome fb5_22 using Bowtie v.0.12.7 with the unique option enabled (Langmead et al. 2009). For paired-end reads, 1000 bp of insertion was used.
Consistency of replicates was checked by genome-wide correlation analysis and by visual inspection.
The aligned reads were called using SPP R package (ver.1.10, Kharchenko et al., Nat.Biotech 2008). The binding characteristics and tag shift were estimated using SPP get.binding.characteristics() function. The reads showing abnormality were removed using SPP remove.local.tag.anomalies() function.
The smoothed fold enrichment profiles at log2 scale were calculated using the get.smoothed.enrichment.mle function (window size = 150 bp for the Gaussian kernel function and step size = 20 bp) in the SPP R package.
Genome_build: Flybase reference r5.22
Supplementary_files_format_and_content: wig and IP fold enrichment profile compared to input at log2 scale

source_name: S2 cells (male embryo-derived cell line)

extract_protocol:
We followed the BioTAP-XL protocol as described in (Alekseyenko et al. 2015). In brief, embryos of BioTAP transgenic fly lines (Pc-C-BioTAP, BioTAP-N-E(z) and BioTAP-N-Scm) were collected between 12 and 24 h after fertilization and were stored for up to 3 days at 4°C. Every 3 days, embryonic nuclei were crosslinked and extracts were prepared as described, snap-frozen with liquid nitrogen and stored at -80°C. These steps were repeated until extracts from ~50g embryos were pooled. Stable S2 cell lines expressing BioTAP transgenes (Pc-C-BioTAP and BioTAP-N-Scm) were incubated in four 2.8L Fernbach glass flasks at 26.5 °C and 90 rpm. In each flask, cells were grown in 500ml of HyClone CCM3 serum-free media to a density of ~1x107 cells/ml. Crosslinked nuclear extracts from S2 cell lines were prepared from ~2x1010 cells grown in four flasks. After sonication, tandem affinity purification was performed to isolate the BioTAP-tagged bait along with its protein interaction partners and associated genomic DNA.  Interacting proteins were identified by on-bead trypsinization of bound complexes followed by liquid chromatography mass-spectrometry (LC-MS/MS) of the resulting peptides.
Libraries for sequencing were prepared using standard Illumina protocols.

value: Embryo-derived cell-line

value: Late embryonic stage

tag: antibody

value: Pc-C-BioTAP

tag: genotype

value: wildtype S2 cells, co-transfected with pFly-Pc-C-BioTAP and pHygro (for selection)


title: Pc-C-BioTAP_input_S2_replicate1


title: Pc-C-BioTAP_input_emb_replicate2

source_name: Drosophila embryo

value: Bloomington stock strain 9736

value: Developmental Stage Embryo 12-24h

value: Embryo 12-24h


title: BioTAP-N-Scm_IP_S2_replicate1

value: BioTAP-N-Scm

value: wildtype S2 cells, co-transfected with pFly-BioTAP-N-Scm and pHygro (for selection)


title: BmN4 transfected with Ago3-endo reporter - Ago3 IP [DH13]

description:
small RNA
Ago3 IP


title: Pc-C-BioTAP_IP_emb_replicate1


title: BioTAP-N-E(z)_IP_emb_replicate1

value: BioTAP-N-E(z)


title: Pc-C-BioTAP_input_emb_replicate1


title: DP_V5anchor_rep2

status: Public on Jul 08 2015

city: Saint Louis

first: Eugene

last: Oltz

state: Missouri

address: 660 Euclid Ave, Campus Box 8118

department: Pathology & Immumonolgy

institute: Washington University in St Louis

description: chromosome conformation capture DNA

data_processing:
Sequence tags were alligned to the reference genome (mouse MM9) using Bowtie
Reads-per-million (RPM) values were calculated using the r3C-seq Bioconductor package
Genome_build: MM9

source_name: Rag deficient thymus

extract_protocol:
4C chromatin was prepared by primary digestion with HindIII (NEB) enzyme and secondary digestion with NlaIII enzyme (NEB), intramolecular ligation reactions were performed with T4 DNA Ligase (NEB). Inverse PCR was performed to generate 4C library. 100ng of 4C DNA was used for sequencing.
Libraries were prepared according to the Illumina protocol. After end-repair and indexed adapter ligation, fragments were size-selected using Agencourt AMPure XP beads (Beckman Coulter, Brea, CA, USA) prior to amplification. The size-selected and amplified fragment libraries were verified on a 2100 Bioanalyzer (Agilent) prior to being pooled (7-10 per lane) and sequenced

value: CD4/CD8 positive thymocytes

tag: treatment

value: generated by IP injections of anti-CD3ε for ten days


title: S6

value: Group I

value: Serono - Toronto

value: sperm


title: exGF2

value: Swiss Webster

value: Amygdala

value: 10 weeks

value: germ-free colonized from p21 (weaning)


title: GF2

value: germ-free


title: pi-Chr17-ΔMyb (-/-), Miwi IP [RR175]

value: testis


title: pi-Chr17-ΔMyb (+/-), Miwi IP [RR174]


title: pi-Chr17-ΔMyb (-/-), Mili IP [RR172]


title: exGF1


title: GF3


title: S3


title: S2


title: CON4

value: conventional


title: S1


title: S7


title: CON2


title: CON1


title: BmN4 transfected with Ago3-endo reporter - Siwi IP [DH15]

description:
small RNA
Siwi IP

value: transfected BmN4 cells


title: BioTAP-N-Scm_IP_S2_replicate2

value: Embryo-derived cell-line

value: Late embryonic stage


title: Pc-C-BioTAP_IP_emb_replicate2

value: Bloomington stock strain 9736

value: Developmental Stage Embryo 12-24h

value: Embryo 12-24h

value: Pc-C-BioTAP


title: BioTAP-N-Scm_input_S2_replicate1

value: wildtype S2 cells, co-transfected with pFly-BioTAP-N-Scm and pHygro (for selection)


title: BmN4 transfected with Ago3-artif reporter - Siwi IP [DH19]

description:
small RNA
Siwi IP


title: BioTAP-N-Scm_input_S2_replicate2


title: BioTAP-N-E(z)_IP_emb_replicate2

value: BioTAP-N-E(z)


title: BioTAP-N-E(z)_input_emb_replicate1


title: DP_Ebanchor_rep3

description: chromosome conformation capture DNA

value: CD4/CD8 positive thymocytes

value: generated by IP injections of anti-CD3ε for ten days


title: DP_Ebanchor_rep1

description: chromosome conformation capture DNA


title: BioTAP-N-E(z)_input_emb_replicate2


title: NMDA TND10 RNA Rep1

city: Cambridge

first: Sudhir

last: Thakurela

address: 7 Divinity Avenue

institute: HSCRB

description:
RNASeq
NormalizedReadCount_DMSONMDA_RNASeq.txt

data_processing:
For all the sequenced samples  reads were aligned to Mouse genome (mm9) using TopHat with default parameters. For ChIPSeq samples reads were mapped using bowtie and resultant bam files were used to call peaks using MACS. Peaks called were then subjected to enrichment calculation suing the bam files for each sample. For RNASeq data the aligned reads were then provided as an input to HTSeq_count utility from HTSeq package. The raw read count files obtained from HTSeq-count were then processed for differential expression using DESeq package. The raw read counts were normalized for library size differences using estimateSizeFactors function from DESeq package.
Genome_build: mm9
Supplementary_files_format_and_content: peaks for FAIRE and ChIP, and normalized counts for RNA

source_name: Day10 Neurons

extract_protocol: Neurons were washed three times with BSS (balanced salt solution; 25 mM Tris, 120 mM NaCl, 15 mM glucose, 5.4 mM KCl, 1.8 mM CaCl2, 0.8 mM MgCl2, pH 7). After treating the neurons with 200 µM NMDA (Sigma) or only DMSO for 6 hours, cells were washed three times with BSS again and processed for RT-qPCR, ChIP and FAIRE analysis.

value: Day10 Neurons

value: NMDA


title: BioTAP-N-Scm_IP_emb_replicate1

value: BioTAP-N-Scm


title: BioTAP-N-Scm_input_emb_replicate1


title: Control TND10 H3K27ac Rep2

description:
ChIPSeq
NormalizedEnrichment_DMSONMDA_H3K27Ac.txt

value: DMSO


title: DP_V5anchor_rep3

description: chromosome conformation capture DNA


title: NMDA TND10 H3K27ac Rep2


title: DP_V5anchor_rep1

description: chromosome conformation capture DNA


title: DN_Ebanchor_rep1

description: chromosome conformation capture DNA

value: double negative thymocytes


title: Control TND10 RNA Rep2


title: DN_Ebanchor_rep3

description: chromosome conformation capture DNA


title: Control TND10 H3K27ac Rep1


title: TND1 H3K27ac Rep2

description:
ChIPSeq
NormalizedEnrichment_PotentialPeaks_H3K27Ac.txt

source_name: Day1 Neurons

value: Day1 Neurons


title: Control TND10 RNA Rep1


title: NMDA TND10 RNA Rep2


title: NMDA TND10 H3K27ac Rep1


title: TND10 H3K27ac Rep1


title: DP_Ebanchor_rep2

description: chromosome conformation capture DNA


title: DN_V5anchor_rep1

description: chromosome conformation capture DNA


title: DN_Ebanchor_rep2

description: chromosome conformation capture DNA


title: DN_V5anchor_rep3

description: chromosome conformation capture DNA


title: CREB

first: David

middle: E

last: Sanin

address: Stübeweg 51

department: Immunometabolism

institute: Max-Planck-Institute für Immunbiologie und Epigenetik

description: Pooled from 3 samples

data_processing:
Basecalling and de-multiplexing of indexed reads was performed by CASAVA version 1.8.2 (Illumina) to produce fastq format file for each sample.
The raw fastq files were trimmed to remove Illumina adapter sequences and low quality bases using Cutadapt version 1.2.1 and Sickle version 1.200 with a minimum window quality score of 20. After trimming, reads shorter than 10 bp were removed.
Reads were mapped to the latest reference genome (downloaded from Ensembl (mm10 GRCm38) ftp://ftp.ensembl.org/pub/release-76/fasta/mus_musculus/dna/) with Bowtie2.
Mappings were filtered by mapping quality (<q 15) and also by removing mappings flagged as multiply mapped with Samtools. Duplicate reads were identified and subsequently removed with Picard (http://picard.sourceforge.net).
MACS v1.4 was used to detect peaks in the treatment samples, relative to the input sample, which would be indicative of protein-DNA binding sites. Peaks were subsequently annotated with nearby genomic features with HOMER.
Genome_build: mm10
Supplementary_files_format_and_content: Processed files were created using MACS

source_name: Bone marrow derived macrophages

growth_protocol: Bone marrow macrophages(BMMs) were produced by incubating 5x10^6 murine bone marrow cells for 7 days in the presence of CSF-1 in DMEM (Gibco, Paisley, UK) supplemented with FCS (Biosera, Uckfield, UK)

extract_protocol:
Aliquots of 5x106 cells were lysed (10 min 4 °C) in lysis buffer containing 50 mM Tris-HCl pH 8.1, 10 mM EDTA and 1 % SDS, supplemented with Protease Inhibitor Cocktail® (SIGMA-ALDRICH, Gillingham, UK) and Phosphatase Inhibitor Cocktail Set II® (Calbiochem, Watford, UK), then diluted (2:3) in a buffer containing 20 mM Tris-HCl, 150 mM NaCl pH 8.1, 1 % Triton X-100 and 1 mM EDTA. Lysates were sonicated for 45 min on ice using a Bioruptor plus® (Diagenode, Seraign, Belgium) 30s ON and 30s OFF at HIGH setting. Sonicated cellular lysate was centrifuged (15000 g 10 min 4°C), supernatants were pre-cleared with 30μl of Protein A Agarose/Salmon sperm DNA (50 % slurry) (Merk Millipore, Watford, UK) for 30 min at 4 °C with agitation. Agarose was removed by brief centrifugation (500 g 4 °C 1 min) and supernatants were incubated overnight with gentle agitation at 4 °C with anti-CREB (clone 48H2) (Cell Signaling Technology, Leiden, The Netherlands). Samples were then incubated with 30 μl of Protein A Agarose/Salmon sperm DNA (50 % slurry) for 1 hour at 4 °C with gentle agitation. The agarose pellet with bound DNA was recovered and washed for 5 min each with 1ml each of low salt buffer, then high Salt buffer, then LiCl buffer and finally twice with TE buffer (all Merk Millipore, Watford, UK). DNA was recovered by incubating agarose pellets for 2 hours at 68 ºC in an elution buffer containing 20 mM Tris–HCl, pH 7.5, 5 mM EDTA, 50 mM NaCl, 1% SDS and 50 μg/ml proteinase K (Promega, Madison, USA) with vigorous shaking. Agarose was removed by gentle centrifugation and DNA extracted from supernatants using phenol-chloroform.
DNA libraries, sequencing, peak calling, and enrichment analysis were carried out by the Centre for Genomic Research at the University of Liverpool, Liverpool, UK. Briefly, DNA for each sample was end repaired, A-tailed and ligated to an appropriate dilution of Illumina adapter. The ligated product was then amplified to generate the final library, which were pooled in equimolar ratios using Qubit and Bioanalyzer data, and quantified by qPCR, using an Illumina Library Quantification Kit (KAPA, KK4854) on a Roche LightCycler 480II system.
Template DNA was denatured according to the protocol described in the Illumina cBot User Guide and loaded at 8.5 pM concentration. Sequencing was carried out on 1 lane of an Illumina HiSeq 2500 with version 1 chemistry generating 2 × 100bp paired end reads

treatment_protocol: BMMs were treadted for 30 minutes with 50ug/ml of S. mansoni cercarial secretions

value: CREB (clone:48H2)


title: CREB 2

description: Pooled from 3 samples


title: DN_V5anchor_rep2

description: chromosome conformation capture DNA


title: INPUT

description: Pooled from 3 samples


title: INPUT 2

description: Pooled from 3 samples


title: E.coli_m6A_IP2

city: Hong Kong

first: Xin

last: Deng

address: To Yuen Building

department: Biomedical Sciences

country: Hong Kong

institute: City University of Hong Kong

data_processing:
Bacterial RIP-seq reads were mapped to the E. coli or P. aeruginosa genomes, using TopHat (version 2.0.0) with two mismatches allowed.
Only the uniquely mapped reads were kept for the subsequent analyses.
The enriched peaks were identified using MACS software (version 2.0.0)
Genome_build: E.coli (ASM584v2) and P. aeruginosa (ASM676v1)

organism: Escherichia coli K-12

source_name: liquid culture in LB

growth_protocol: grow until mid-log phase in LB medium

extract_protocol:
In a 0.5-mL IP reaction, 5 mg purified bacterial mRNA and 15 mL of 0.5 mg/mL anti-m6A antibody was incubated for 2 h at 4 °C in IPP buffer (150 mM NaCl, 0.1% NP-40, 10 mM Tris-HCl, pH 7.4, 1 U/mL RNasin). The reaction was exposed by UV 254 nm for 3 times, before RNase T1 (0.1 U/mL) digestion for 15 min at 22 °C. After quench the digestion reaction on ice for 5 min, 200 mL pre-blocked protein A bead slurry was added into reaction for 1 h at 4 °C. After washing for 3 times with IP wash buffer (50 mM HEPES-KOH, pH 7.5, 300 mM KCl, 0.05% NP-40, with proteinase inhibitor and RNasin), the beads were treated by a second round of RNase T1 digestion (15 U/mL) at 22 °C for 15 min. The beads were cooled down on ice for 5 min and then washed for 3 times with high salt wash buffer (50 mM HEPES-KOH, pH 7.5, 500 mM KCl, 0.05% NP-40, with proteinase inhibitor and RNasin). The beads were then treated with Antarctic phosphatase (0.5 U/mL) for 20 min at 37 °C. After dephosphorylation, beads were washed twice with phosphatase wash buffer (50 mM Tris-HCl, pH 7.5, 20 mM EGTA, 0.5% NP-40) and twice with PNK buffer without DTT (50 mM Tris-HCl, pH 7.5, 50 mM NaCl, 10 mM MgCl2). Polynucleotide kinase (1 U/mL) and 200 mM ATP was then added to the beads at 37 °C for 15 min. The RNA fragment was further purified by proteinase K digestion and TRIzol extraction. For IP samples, small RNA libraries were made by using NEBNext® Small RNA Library Prep Set for Illumina® (NEB). The input samples were followed above procedures without anti-m6A antibody pull-down and RNase T1 digestion. Libraries for input samples were made by using TruSeq RNA Sample Preparation Kits (Illumina). All samples were sequenced using the HiSeq 2000 system (Illumina).
NEXTflex™ ChIP-Seq Kit (Bioo Scientific)

treatment_protocol: mRNA purification by MICROexpress bacterial mRNA enrichment kit (Life technologies)


title: TND1 H3K27ac Rep1


title: TND10 H3K27ac Rep2


title: E.coli_m6A_IP1


title: NP H3K27ac Rep1

source_name: Neuronal Progenitors

value: Neuronal Progenitors


title: nuc2_run2

first: Graham

last: Johnson

state: Michigan

department: CMMG

institute: WSUSOM

description:
sperm nuclei 2
processed data file: nuc2_subcount.txt
processed data file: unadjusted RPKMs.txt

data_processing:
Illumina Casava v 1.8.2 software used for basecalling and demultiplexing
Sequencing reads were aligned to the mouse genome assembly Mm10 with Tophat2 (version 2.0.12) using the following parameters: tophat2 -r 30 --mate-std-dev 50 --no-coverage-search -G genes.gtf.
Novel transcript structures were assembled using Cufflinks (version 2.1.1). Assembled transcripts exceeding twice the average fragment length (135 bp) were combined with UCSC gene annotations (Mm10). Transcript abundance was calculated using uniquely aligned sequencing reads and the Samtools (version 0.1.19; 34) and Bedtools (version v2.19.1-2) suites.
Coverage of the most abundant isoform of each transcript was calculated in reads aligned per kilobases mapped (RPKM) using Bedtools.
Differential enrichment and subtractive differential enrichment analyzes were performed with the HT-Seq and DEseq2 software packages. The latter required randomly subsampling uniquely aligned reads to equivalent levels using Samtools prior to alignment with TopHat2. Subsequently nuclei naïve count values from HT-Seq were averaged, rounded to whole integers and subtracted from corresponding whole cell naïve values. Negative adjusted whole cell counts were assumed to be zero. Differential enrichment analysis was carried out using DESeq2 as before.
Genome_build: Mm10
Supplementary_files_format_and_content: unadjusted RPKM values, tab-delimited text files (Ht-seq counts)

source_name: sucrose fractionated mouse sperm nuclei RNA

extract_protocol:
Mature spermatozoa from transgenic line HP3.1 were isolated from cauda epididymides and vas deferens harvested from individual four month old transgenic mice on ice into 50 mM Tris-HCl, pH 7.4, buffer. The cells were washed twice following filtration through an 80 micron mesh, resuspended in 0.5 mL 50 mM Tris-HCl, 7.4, buffer and subjected to sonication with a TekMar TM-50 sonic disruptor (TekMar, Cincinnati, OH) at 70% maximum output for two minutes on ice to separate heads from tails. Separation was confirmed by light microscopy. The sperm solution containing 1 – 5 x 107 cells was diluted to a volume of 7.5 mL in 1 M sucrose buffered with 50 mM Tris-HCl, pH 7.4, buffer containing 5 mM MgCl2. A triple-step gradient was prepared by overlaying diluted samples onto cushions consisting of 2 M sucrose buffered with 50 mM Tris-HCl pH 7.4 containing 5 mM MgCl2 and 0.45 g/mL CsCl buffered with 25 mM Tris-HCl, pH 7.4, buffer containing 0.5% Triton X-100. Nuclei were recovered by ultracentrifugation at 75,600 x g for 45 minutes at 4° C and subsequently washed twice with 25 mM Tris-HCl, pH 7.4, buffer containing 0.5% Triton X-100. Following resuspension in 0.5 mL RLT buffer (Qiagen, Valencia, CA) supplemented with 1.5% β-mercaptoethanol (Amresco, Rockford, IL), 0.2 mm stainless steel beads were added to the samples and nuclei were lyzed with the Disruptor Genie (Scientific Industries, Inc., Bohemia, NY). After the addition of an equal volume of Qiazol (Qiagen) nucleic acids were recovered using the RNeasy system (Qiagen). Whole cell sperm RNAs were equivalently extracted following visual confirmation of the absence of contaminating cell-types omitting sonication and ultracentrifugation. Extracted total RNAs were DNased (Turbo Dnase, Ambion, Grand Island, NY) and subsequently subjected to RT-PCR with intron-spanning primers to Prm2 (Foward 5'-GCACCATGGTTCGCTACCG-3', Reverse 5'- GTGGCCTCACATGATGTTGCT-3'; 31).
Stock synthetic spike-in ERCC RNAs (Invitrogen, Carlsbad, CA) were diluted 1:10,000 and pooled with total sperm RNAs prior to reverse transcription and amplification using the Seq-plex system (Sigma, St. Louis, MO). Following universal priming site removal pre-amplified cDNA libraries were subjected to sequencing library construction (DNA Ultra-Low, NEB, Ipswich, MA) followed by 50 cycles of paired-end sequencing on the Illumina Hi-Seq 2500 platform.

value: C57BL/6 transgenic line HP3.1

value: sperm

tag: cell fraction

value: sucrose-fractionated sperm nuclei

value: adult


title: whole_run2

description:
whole sperm
processed data file: adj_w_subcount.txt
processed data file: w_subcount.txt
processed data file: unadjusted RPKMs.txt

source_name: whole mouse sperm RNA

value: whole sperm

tag: developmental stage


title: TND10 FAIRE Rep1

description:
FAIRESeq
NormalizedEnrichment_PotentialPeaks_FAIRE.txt


title: nuc1_run2

description:
sperm nuclei 1
processed data file: nuc1_subcount.txt
processed data file: unadjusted RPKMs.txt


title: E.coli_CK


title: TND1 FAIRE Rep2


title: TND10 FAIRE Rep2


title: nuc2_run1


title: nuc1_run1


title: NP H3K27ac Rep2


title: whole_run1


title: P. aerguinosa_m6A_IP2

organism: Pseudomonas aeruginosa PAO1


title: NP FAIRE Rep1


title: IPSDM M2 rep1

city: New York

first: Chenyi

last: Xue

institute: Columbia University

description: Sample 19

data_processing:
Base calling was done by RTA v1.13.48.0
Sequence reads were aligned to hg19 genome using STAR with default options.
Alignment was filtered using the following specifications: 1) the mapping quality score of each read is ≥ 30, 2) reads from the same pair were mapped to the same chromosome with expected orientations and mapping distance between the read pair was < 500,000 bp, 3) read was uniquely mapped
Fragments Per Kilobase of exon per Million reads mapped (FPKM) was estimated by Cufflinks 2.1.1. Differential expression was tested by Cuffdiff with options "-u --max-bundle-frags 50000000 --library-type fr-firststrand". Annotation from RefSeq genes and the stringent set of lincRNAs from Cabili et al. 2011 was provided.
Genome_build: hg19
Supplementary_files_format_and_content: DIFF files were obtained from Cuffdiff output.Tab-delimited text file include normalized FPKM values from HMDM, IPSDM and iPSC samples
HMDM_MAC_M1_gene_exp.diff contains differential expression result from HMDM and M1-HMDM. sample_1 = HMDM, sample_2 = M1-HMDM
HMDM_MAC_M2_gene_exp.diff contains differential expression result from HMDM and M2-HMDM. sample_1 = HMDM, sample_2 = M2-HMDM
HMDM_M1_M2_gene_exp.diff contains differential expression result from M1-HMDM and M2-HMDM. sample_1 = M1-HMDM, sample_2 = M2-HMDM
IPSDM_M1_M2_gene_exp.diff contains differential expression result from M1-IPSDM and M2-IPSDM. sample_1 = M1-IPSDM, sample_2 = M2-IPSDM
IPSDM_MAC_M1_gene_exp.diff contains differential expression result from IPSDM and M1-IPSDM. sample_1 = IPSDM, sample_2 = M1-IPSDM
IPSDM_MAC_M2_gene_exp.diff contains differential expression result from IPSDM and M2-IPSDM. sample_1 = IPSDM, sample_2 = M2-IPSDM
HMDM_IPSDM_gene_exp.diff contains differential expression result from HMDM and IPSDM. sample_1 = HMDM, sample_2 = IPSDM
iPS_IPSDM_gene_exp.diff contains differential expression result from iPSC and IPSDM. sample_1 = iPSC, sample_2 = IPSDM
HMDM_IPS_IPSDM.norm.expr.txt contains normalized FPKM from all samples

source_name: Macrophage

growth_protocol: iPSC lines were maintained in hESC media consisting of Dulbecco modified Eagle medium/F12 (50:50; Life Technologies, Grand Island, NY) supplemented with 20% knockout serum replacement, 100 μM nonessential amino acids, 50 U/ml penicillin, 50 μg/ml streptomycin (Life Technologies), 10−4 M β-mercaptoethanol (Sigma-Aldrich, St. Louis, MO), and 10 ng/ml human basic fibroblast growth factor (bFGF) on irradiated mouse embryonic feeder cells (MEF, GlobalStem, Rockville, MD). PBMC were cultured in 20% FBS/RPMI with 100 ng/ml human M-CSF (Peprotech, Rocky Hill, NJ) for 7 days to induce differentiation to macrophages. iPSC were differentiated to macrophages according to established protocols and maintained in 20% FBS/RPMI with 100 ng/ml human M-CSF.

extract_protocol:
RNA samples were extracted using All Prep DNA/RNA/miRNA Universal Kit (Qiagen, Valencia, CA). Extracted RNA samples underwent quality control (QC) assessment using the Agilent Bioanalyzer (Agilent, Santa Clara, CA)
With a minimum of 300 ng input RNA, we generated first-strand cDNA using random hexamer-primed reverse transcription, followed by second-strand cDNA synthesis using RNase H and DNA polymerase, and ligation of sequencing adapters using the TruSeq RNA Sample Preparation Kit (Illumina, San Diego, CA). Fragments of ~350 bp were selected by gel electrophoresis. To minimize lane and batch effects in RNA-Seq experiments, barcoded libraries were mixed and pooled in equimolar (2 nmol/L) amounts and diluted to 4 pmol/L for cluster formation on a single flow cell lane, followed by 100 bp paired-end sequencing on Illumina’s HiSeq 2000.

value: M2-polarized IPSDM


title: IPSDM M2 rep2

description: Sample 20


title: P. aeruginosa_m6A_IP1


title: TND1 FAIRE Rep1


title: NP FAIRE Rep2


title: P. aeruginosa_CK


title: IPSDM M2 rep3

description: Sample 21