title: ChIP-Seq analysis of TRB1 associated sequences_(Wt(-)_anti-TRB1)

city: Brno

first: Jan

last: Hapala

address: Kamenice 5

country: Czech Republic

institute: Masaryk University - CEITEC

description: DNA (genomic, chloroplast, mitochondrial)

data_processing:
Illumina Casava1.8.2 software used for basecalling.
ChIP-seq reads were aligned to the tair9 genome assembly using Bowtie2 version 2.2.2 with with --very-sensitive option; output: BAM files.
Data were filtered using the following specifications: mapping quality >= 25 (samtools view -q 25).
Technical replicates were merged (samtools merge): reads01+reads02 → rep1control, reads03+reads04 → rep1sample, reads05+reads06 → rep2sample, reads07 → rep2control.
Mitochondria- and chloroplast- mapped reads were separated from the genomic reads.
BAM files were sorted (samtools sort) and indexes calculated.
BAM files were converted into BED files (bedtools bamtobed).
Normalization of ChIP-seq data performed with NCIS.
Peak calling 1: with MACS, version 1.4.3 20131216 with parameters -g: 90000000, band width --bw 250,  keeping all duplicate reads --keep-dup all, ChIP/control scaling ratio --ratio 1.215273 and 1.742786 for the first and second replicate, respectively.
Peak calling 2: with PePr, version 1.0.2, parameters: --peaktype=sharp.
Peak coordinates extracted from MACS peak XLS files where FDR <= 5%: tail -n +19 peak_file.xls | awk '{if ($9 < 5.0){print}}' | cut -f 1,2,3 | awk '{print $1,$2,$3,"MACS_peak_"NR}' > $4/${REP1_NAME}_below5FDR.bed.
PePr peaks filtered: if estimated fragment length < 30bp (columns 5, 6), peak was removed; the rest retained: awk '{if ($5 >= 30 && $6 >= 30) print}' pepr_peaks.bed
MACS peak files intersected: bedtools intersect -a rep1_macs_peaks.bed -b rep2_macs_peaks.bed > macs_peaks.bed
MACS intersection intersected with PePr peaks: bedtools intersect -a pepr.bed -b macs.bed > TRB1_genomic_peaks.bed
TAIR10 gene-region-separated datasets in FASTA format converted into BED files.
Intersection of TRB1_genomic_peaks and TAIR10 datasets was calculated (both directions).
Genome_build: tair9
Supplementary_files_format_and_content: pepr_peaks.bed; peaks identified by PePr comparing both replicates
Supplementary_files_format_and_content: rep1_macs_peaks.bed; peaks identified by MACS in replicate 1
Supplementary_files_format_and_content: rep2_macs_peaks.bed; peaks identified by MACS in replicate 2
Supplementary_files_format_and_content: TRB1_genomic_peaks.bed; final set of peaks (intersection of the three above)
Supplementary_files_format_and_content: *_peaks_intersect.bed; bedtools intersect -u -a TRB1_GENOMIC_PEAKS.bed -b TAIR10_DATASET.bed
Supplementary_files_format_and_content: *_intersect.bed; bedtools intersect -u -a TAIR10_DATASET.bed -b TRB1_GENOMIC_PEAKS.bed
Supplementary_files_format_and_content: *_telobox_sorted.bed; extracted telobox motifs, sorted
Supplementary_files_format_and_content: *_telobox_peaks_intersect.bed; bedtools intersect -u -a TELOBOXES.bed -b TRB1_GENOMIC_PEAKS.bed

organism: Arabidopsis thaliana

source_name: seedlings

growth_protocol: Seedlings were grown under constant light, long day (16h/21°C, 8h/19°C), 50-60% relative humidity, for 2 weeks.

extract_protocol:
The ChIP assay was performed as described Bowler et al. (2004) with modifications. Chromatin extracts were prepared from seedlings treated by 1% formaldehyde. The chromatin from isolated nuclei was sheared to an average length of 250-500 bp by sonication (Bioruptor, Diagenode) and spun. Matrix GFP-TRAP_A (Chromtec) was blocked against non-specific interaction with 200 mM ethanolamine, 1% BSA and TR10-24-G and TR10-24-C (i.e. DNA sequences, that are not recognised by TRB proteins (Schrumpfova et al., 2004). The pretreated matrix was incubated with chromatin diluted with ChIP dilution buffer (16.7 mM Tris-HCl, pH=8,0, 1.2 mM EDTA, 167 mM NaCl, 0.1% TritonX-100, PMSF and protease inhibitors) for 4°C / 4h and consequently washed with low salt; high salt; LiCl; TE buffers (in contrast to Bowler et al. (2004) levels of detergents (TritonX-100, NP-40 and sodium deoxycholate) were lowered to 0.1%). The cross-linking was reversed by 0.2 M NaCl O/N and followed by proteinase K (Serva) treatment, phenol/chlorophorm extraction and RNase A (Serva) treatment according to Bowler et al. (2004).
Fifty microliters of immunoprecipitated DNA (ranging from 0.2-6 ng DNA (measured by Qubit dsDNA HS Assay Kit (Invitrogen)) was used for library preparation using NEBNext ChIP Seq Liabrary Prep Master Mix (NEB), with Agencourt XP beads (Beckman) in the ratios described in the protocol only with one exception: the Adapter was diluted 1:17 in water. DNA fragments were selected by 2% e-gel (Invitrogen) in the 270-300bp size range. Subsequently, a PCR reaction (18 cycles) with indexed Primers 1-11 from NEBNext Multiplex Oligo set (NEB) was performed. Libraries were sequenced (50 bp single-end reads) on an Illumina HiSeq 2500 using TruSeq SBS Kit v3-HS (Illumina) sequencing reagents.  The quality of the final libraries was checked on the Bioanalyzer (Agilent) and the quantity was checked with Qubit dsDNA HS Assay Kit (Invitrogen). The libraries were pooled equimolar and diluted to 10 pM (denaturation in NaOH) in Hyb Buffer 1 (HT1) from TruSeq SR Cluster Kit v3- cBot HS (Illumina).

value: Wt

value: seedlings

tag: age

value: Wt/Wt

tag: chip-antibody


title: DGRP822 M_E1_3_L1

status: Public on Dec 21 2015

type: SRA

city: Bethesda

first: Brian

last: Oliver

state: MD

address: 50 South Drive

department: LBG

country: USA

institute: NIDDK, NIH

description:
EP2F111
Drosophila Genetic Reference Panel 822, Male, Environment 1, Fly 3, Library 1

library_source: transcriptomic

data_processing:
We performed single-end 76 bp sequencing on the HiSeq2000 Sequencing System (Illumina, San Diego, CA) according to the manufacturer.  De-multiplexed reads (Chastity > 0.6) FASTQ output was from Illumina CASAVA v.1.8.2. Reads were mapped to FlyBase release 5 version 57 (FlyBase file: dmel-all-chromosome-r5.57.fasta) and release 6 version 01 (FlyBase file: dmel-all-chromosome-r6.01.fasta) of the Drosophila melanogaster genome and appended sequence corrected ERCC sequences (ERCC_reference.fa, See Jiang et al., Genome Research, 2011, PMID 21816910) and annotations using TopHat2 (v2.0.10) with parameters  "-g 1 --library-type fr-firststrand" (Kim et al., Genome Biol, 2013, PMID 23618408).  We aligned to Drosophila melanogaster chromosome scaffolds only (release 5 chrU and chrUextra were excluded).  We used a Perl script to convert the dmel-all-r5.57.gff to a GTF file (dme5_57_ERCC.gtf) compatible with TopHat2 and HTSeq (v0.5.4p1) (Anders et al., Bioinformatics, 2015, PMID 25260700).  We extracted features, including pre-miRNAs, pseudogenes, mRNAs, ncRNAs, rRNAs, snoRNAs, snRNAs, tRNAs, genes, CDSs and exons, and changed the 304 features for “miRNA” to “exon”.   We changed the following features: 9 mRNA transcripts without strand assignment were assigned to the parent gene strand; 11 Exons and 9 CDSs without strand assignment were assigned to the parent transcript strand; and 10 CDSs, with ends longer than the parent, were truncated to fit the parent ends.  TopHat2 called Bowtie 2 (v2.1.0) to make indexes. We used SAMtools (v0.1.19) (Li et al., Bioinformatics, 2009, PMID 19505943) to index chromosomes, to sort and merge alignments, and for read statistics.  Mapped reads at the gene-level were counted by HTSeq with parameter "--stranded=reverse -i gene_id -t exon".
Supplementary_files_format_and_content: There are five supplemental files.  The tab-delimited 5_57_HTSeq_raw_read_counts.txt and 6_01_HTSeq_raw_read_counts.txt files contain the gene-level output, for each library, for genome releases 5.57 and 6.01 respectively. Column 1 and 2 are FlyBase FBgn#s and gene symbols, or ERCC  identifiers as applicable.  The remaining columns are library names (as GEO GSM#s).  The GEO_run_summary.xls file contains detailed information on each sequencing run and a readme sheet with field definitions.  The dme5_57_ERCC.gtf file contains the annotation for the alignment to release 5.57 plus the ERCC spike-ins.  ERCC_reference.fa is a FASTA file of the sequence corrected ERCCs.

molecule: polyA RNA

organism: Drosophila melanogaster

source_name: Whole body

growth_protocol: We grew and aged Drosophila Genetic Reference Panel (DGRP) inbred lines derived from the Raleigh, NC, USA wild population (Mackay et al., Nature, 2012, PMID 22318601; Huang et al., Genome Res, 2014, PMID 24714809).  We randomly chose the following 16 DGRP lines after excluding 5 lines from the population that were slow-growing: DGRP-93, DGRP-229, DGRP-320, DGRP-352, DGRP-370, DGRP-563, DGRP-630, DGRP-703, DGRP-761, DGRP-787, DGRP-790, DGRP-804, DGRP-812, DGRP-822, DGRP-850, and DGRP-900. We collected 8 virgin male and 8 virgin female flies from the 16 DGRP genotypes in three replicated environments to produce RNA sequence profiles.  We controlled the environmental conditions in the following ways. We seeded the fly cultures with 5 male and 5 female parents.  We reared the progeny in a single incubator on standard Drosophila food (http://flystocks.bio.indiana.edu/Fly_Work/media-recipes/bloomfood.htm) at 25°C, 60% humidity, and a 12:12-hour light:dark  cycle.  We collected and maintained male and female virgins at 20 flies to a same-sex vial for four days prior to RNA extraction to control for social exposure.

extract_protocol:
Single flies frozen on dry ice were placed into each well of Axygen 96 Deep Well Plates (Corning, Corning, NY) pre-loaded with 200µL of 1 mm glass beads (Biospec Products, Bartlesville, OK).  Fly placement in the wells of each plate was randomized within a replicated environment. We isolated total RNA using the RNeasy 96 Plate Kit according to the manufacturer using either vacuum or centrifugation technology (Qiagen, Valencia, CA), with the following modifications. We added 200µl RLT buffer to each well, sealed the plates with Axygen Sealing Mats (Corning, Corning, NY), and homogenized flies at room temperature for 30sec using a fixed-speed Mini Bead Beater (Biospec Products, Bartlesville, OK).  We added 200µl of 70% ethanol to the crude homogenate, mixed by pipetting several times, and transferred the supernatant to RNeasy plates.  We measured total RNA yield using the Quant-iTTM RiboGreen(R) RNA Assay Kit (Invitrogen, Carlsbad, CA) in black FLUORTRAC 600 384-well PS plates (Greiner Bio-One, Inc., Longwood, FL) using a Gemini EM Fluorescence Microplate Reader (Molecular Devices, Sunnyvale, CA).  We used Dynabeads Oligo (dT) 25 (Life Technologies, Carlsbad, CA) to purify polyA RNA according to manufacturer instructions, except that we used 50-200ng of total RNA depending on yield, adjusted the volume to 50µl with distilled H2O, added 10µl of the washed Dynabeads in a 50µl binding buffer slurry, heated the samples at 65°C for 5min, and immediately chilled the samples on ice.  To prepare RNA-Seq stranded libraries we modified an existing protocol (Wang et al., PLoS One, 2011, PMID 22039485).  All steps were done in 96-well plate format. We incubated all reactions in a Tetrad PTC-225 Thermal Cycler (MJ Research, Waltham, MA). To clean up enzymatic reactions and size select at any of several steps below, we used 0.1% w/v carboxyl-modified Sera-Mag Magnetic Speed-beads (MagNA beads, Thermo Fisher Scientific, Waltham, MA) in XP buffer [20% PEG 8000, 2.5 M NaCl, (Sigma Aldrich, St. Louis, MO)] following the protocol in (Rohland et al., Genome Res, 2012, PMID 22267522), except that we incubated samples with beads for 10 min and used 80% ethanol for washing.  The volume of MagNA beads in XP buffer and corresponding concentration of PEG in the solution was 1.6× XP buffer for no size-selection (except where noted) and 1× XP buffer for selecting fragments > 200bp. Unless noted, we left the beads in the sample after each step.  We used 96-well PCR plates (USA Scientific, Ocala, Fl) paired with Alpaqua 96R Ring Magnet Plates (Alpaqua, Beverly, MA) for magnetic bead separation. We measured RNA quantity with Quant-iTTM RiboGreen(R) and DNA with Quant-iTTM PicoGreen(R) (Invitrogen, Carlsbad, CA) in 384-well plates, PS, FLUOROTRAC 600, black (Greiner Bio-One Inc, Longwood, Fl) using a Gemini EM Fluorescence Microplate Reader (Molecular Devices, Sunnyvale, CA) according to manufacturer instructions.  We used Agilent Bioanalyzer RNA chips and High Sensitivity DNA chips on the 2100 Bioanalyzer system (Agilent, Santa Clara, CA) according to manufacturer instructions and visually inspected electrophoregrams for RNA degradation, library size (DNA), and adapter dimer contamination (DNA).
We fragmented polyA RNA bound to Dynabeads at 94°C for 8min in 16µl of 1.25× first strand MMuLV RT buffer (New England Biolabs, Beverly, MA), with 100ng random primers (Invitrogen, Carlsbad, CA), and 10pg ERCC spike-in controls (Jiang et al., Genome Res, 2011, PMID 21816910) from pools 78A and 78B (Zook et al., PLoS ONE, 2012, PMID 22859977) obtained from Marc Salit (National Institute of Standards and Technology, Gaithersburg MD).  We chilled the samples immediately on ice for 2min and eluted from the beads.  We transferred 15µl of the eluate to a fresh 96-well PCR plate. We added 5µl of the first strand synthesis mixture [0.3mM dNTPs, 5mM DTT, and 10U M-MuLV RT (New England Biolabs, Beverly, MA), and 0.5U SuperRase-In (Life Technologies, Carlsbad, CA)] to the fragmented RNA in the fragmentation buffer and performed a reverse transcription reaction. We bound RNA/cDNA hybrid with 32µl MagNA bead XP buffer slurry, washed, and eluted samples in 16µl of distilled H2O.  We then performed a second strand synthesis with dUTP by adding 5µl of 1× NEB buffer2 (New England Biolabs, Beverly, MA), with 1mM each of dATP, dCTP, dGTP and 2mM dUTP (Thermo Fisher Scientific, Waltham, MA), 10U DNA PolI, 2.5U RNAseH, and 2.5mM DTT (New England Biolabs, Beverly, MA) and incubated at 16°C for 5 hours.  We rebound, washed, and eluted as above. We repaired ends by adding 4µl of NEBNext End Repair Module (New England Biolabs, Beverly, MA) to the eluate following manufacturer instructions.  We then rebound samples to MagNA beads, washed, and eluted as above. Next we performed dA-tailing by adding 4µl of 1× Blue Buffer, with 1U Klenow 3’-5’ exo- (Enzymatics, Beverly, MA) and 1mM dATP, to the eluate and incubated at 37°C for 30min.  We then rebound samples to MagNA beads and washed as above, eluted with 21µl distilled H2O, and transferred 10µl of cDNA to each of two fresh plates. One plate was used in the following steps and one plate was frozen as a back-up.  We ligated RNA Adapter Indexes (Illumina, San Diego, CA) to dsDNA by adding 1µl of an adapter to each 10µl sample and 13µl of 1× Rapid Ligation Buffer with 30U T4 DNA Ligase (Enzymatics, Beverly, MA), and incubated at 25°C for 10min. We stopped reactions with a final concentration of 0.01M EDTA. We added 25µl of MagNA beads in XP buffer (final PEG = 13.6%), bound cDNA to the beads, washed and eluted in 30µl of distilled H2O as above.  We size-selected libraries by adding 1× XP buffer (30µl), washed, eluted samples with 24µl distilled H2O, and transferred 23µl of cDNA to a fresh plate. 11.5µl of the eluate was transferred to another plate and used in the next step; the remaining samples were frozen as a back-up.  We mixed dsDNA product with 2.5U (0.5µl) of Uracil DNA Glycosylase (New England Biolabs, Beverly, MA) and 3µl of PCR Primer Cocktail (Illumina, San Diego, CA) and incubated at 37°C for 30min to digest the second strand DNA.  We added 15µl 2× KAPA HiFi HotStart ReadyMix (Kapa Biosystems, Woburn, MA) directly to the UDG digested DNA mixture and performed a PCR amplification with the following programmed cycle: 95°C for 2min, followed by 12 cycles of 98°C for 20sec, 65°C for 30sec, and 72°C for 30sec; then 72°C for 5min. We purified the product by adding 30µl MagNA beads in 1× XP buffer, eluted in 30µl distilled H2O and transferred 29µl of cDNA library to a fresh plate as described above. To assay plate-level failure, we took 11 samples from each 96-well plate and examined electropherograms for a strong signal in the 300-350 bp library target size and weak signal in the 100-150bp range (primer dimer products).  Each 96-well plate was composed of 4 sets of samples with 24 unique indexes. We pooled equal amounts (5-15 ng) of each library into pools for multiplex sequencing.  All multiplexed libraries were again quantified and checked for quality as above.  Excess library pools were frozen for re-sequencing if required.

tag: strain

tag: developmental stage

value: Adult

tag: Sex

value: Male

tag: tissue

value: Whole body


title: DGRP229 F_E1_5_L1

description:
EP2F101
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 5, Library 1

value: Female


title: DGRP787 F_E1_6_L1

description:
EP2F091
Drosophila Genetic Reference Panel 787, Female, Environment 1, Fly 6, Library 1


title: ChIP-Seq analysis of TRB1 associated sequences_(TRB1(+)_anti-TRB1)

library_source: genomic

molecule: genomic DNA

tag: genotype

value: anti-TRB1 (5.2)


title: ChIP-Seq analysis of TRB1 associated sequences_(TRB1-GFP(+)_GFP-Trap)

value: GFP-Trap A matrix


title: ChIP-Seq analysis of TRB1 associated sequences_(Wt(-)_GFP-Trap)


title: DGRP822 F_E1_2_L1

description:
EP2F081
Drosophila Genetic Reference Panel 822, Female, Environment 1, Fly 2, Library 1


title: DGRP812 M_E1_1_L1

description:
EP2F071
Drosophila Genetic Reference Panel 812, Male, Environment 1, Fly 1, Library 1


title: DGRP787 M_E1_2_L1

description:
EP2F051
Drosophila Genetic Reference Panel 787, Male, Environment 1, Fly 2, Library 1


title: DGRP850 F_E1_4_L1

description:
EP2F061
Drosophila Genetic Reference Panel 850, Female, Environment 1, Fly 4, Library 1


title: DGRP229 F_E1_3_L3

description:
EP2F043
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 3, Library 3


title: Th17_ChIRP_RMRP_WT_Input

city: New York

first: Wendy

last: Huang

address: 540 First Ave Skirball 2-14

institute: NYU School of Medicine

description:
Affinity purified genomic DNA
no probe
Th17_ChIRP_RMRP_WT_Input.bw,Th17_ChIRP_RMRP_WT.bw

data_processing:
Library strategy: ChIRP-seq
After demultiplexing, reads were stripped adaptors and mapped to the mouse (mm9)  genome using Bowtie2. Even and Odd datasets were merged as described previously (PMID:21963238) and visualized as bigWig files. Peaks from the merged datasets were called using MACS2.
Genome_build: mm9
Supplementary_files_format_and_content: bigWig files of merged (even and odd probe sets) RMRP ChIRP-seq read density
Supplementary_files_format_and_content: MACS2 Peaks.bed

organism: Mus musculus

source_name: In vitro Th17

extract_protocol:
Lysates were produced from soncated cell material and RMRP RNA enriched with antisense oligonucleotide enrichment.
Libraries were prepared according to the standard ChIRP-seq protocol (PMID: 21963238)

treatment_protocol: All cells were fixed with 1% gluteraldehyde for 10 minutes at room temperature.

tag: cell type

value: T cell

value: WT


title: Th17_ChIRP_RMRP_WT_Even

description:
Affinity purified genomic DNA
probes 2, 4
Th17_ChIRP_RMRP_WT_Input.bw,Th17_ChIRP_RMRP_WT.bw


title: Th17_ChIRP_RMRP_WT_Odd

description:
Affinity purified genomic DNA
probes 1, 3, 5
Th17_ChIRP_RMRP_WT_Input.bw,Th17_ChIRP_RMRP_WT.bw


title: DGRP703 F_E1_8_L3

description:
EP2F013
Drosophila Genetic Reference Panel 703, Female, Environment 1, Fly 8, Library 3


title: MBD3_rep2

city: RTP

first: Takashi

last: Shimbo

address: 111 TW Alexander Dr.

institute: NIH/NIEHS

data_processing:
Reads were filtered based on a mean base quality score >20. Reads were mapped to hg19 genome using Bowtie 0.12.8
Duplicate reads were removed using MarkDuplicates.jar from picard-tools-1.107 package.
Non-duplicate, uniquely mapped paired-end reads were converted to a single fragment. BigWig files were generated to visualize the genomic coverage.
Genome_build: hg19
Supplementary_files_format_and_content: bigWig

organism: Homo sapiens

growth_protocol: Cells were grown on 15 cm plates in DMEM medium with 10% FBS

extract_protocol: Tagmentation and PCR amplification were performed using Nextera XT kit (Illumina).

value: Breast

tag: chip antibody

tag: cell line


title: DGRP352 M_E1_1_L1

description:
EP2E121
Drosophila Genetic Reference Panel 352, Male, Environment 1, Fly 1, Library 1


title: DGRP787 F_E1_1_L1

description:
EP2E111
Drosophila Genetic Reference Panel 787, Female, Environment 1, Fly 1, Library 1


title: Th17_ChIRP_RMRP_WT_DDX5ko_Input

description:
Affinity purified genomic DNA
no probe
Th17_ChIRP_RMRP_WT_DDX5ko_Input.bw,Th17_ChIRP_RMRP_WT_DDX5ko.bw


title: DGRP370 M_E1_5_L1

description:
EP2E101
Drosophila Genetic Reference Panel 370, Male, Environment 1, Fly 5, Library 1


title: Th17_ChIRP_RMRP_WT_DDX5ko_Even

description:
Affinity purified genomic DNA
probes 2, 4
Th17_ChIRP_RMRP_WT_DDX5ko_Input.bw,Th17_ChIRP_RMRP_WT_DDX5ko.bw


title: DGRP352 M_E1_8_L1

description:
EP3A081
Drosophila Genetic Reference Panel 352, Male, Environment 1, Fly 8, Library 1


title: Th17_ChIRP_RMRP_RM262_Odd

description:
Affinity purified genomic DNA
probes 1, 3, 5
Th17_ChIRP_RMRP_RM262_Input.bw,Th17_ChIRP_RMRP_RM262.bw


title: Th17_ChIRP_RMRP_RM262_Input

description:
Affinity purified genomic DNA
no probe
Th17_ChIRP_RMRP_RM262_Input.bw,Th17_ChIRP_RMRP_RM262.bw


title: DGRP812 F_E1_6_L1

description:
EP2E091
Drosophila Genetic Reference Panel 812, Female, Environment 1, Fly 6, Library 1


title: DGRP563 M_E1_1_L1

description:
EP2E071
Drosophila Genetic Reference Panel 563, Male, Environment 1, Fly 1, Library 1


title: DGRP804 M_E1_4_L3

description:
EP2E063
Drosophila Genetic Reference Panel 804, Male, Environment 1, Fly 4, Library 3


title: Th17_ChIRP_RMRP_RM262_Even

description:
Affinity purified genomic DNA
probes 2, 4
Th17_ChIRP_RMRP_RM262_Input.bw,Th17_ChIRP_RMRP_RM262.bw


title: DGRP812 M_E1_5_L1

description:
EP2E031
Drosophila Genetic Reference Panel 812, Male, Environment 1, Fly 5, Library 1


title: DGRP370 M_E1_7_L1

description:
EP2E041
Drosophila Genetic Reference Panel 370, Male, Environment 1, Fly 7, Library 1


title: DGRP370 M_E1_2_L1

description:
EP2E021
Drosophila Genetic Reference Panel 370, Male, Environment 1, Fly 2, Library 1


title: DGRP563 F_E1_7_L1

description:
EP2E011
Drosophila Genetic Reference Panel 563, Female, Environment 1, Fly 7, Library 1


title: DGRP900 M_E1_6_L1

description:
EP2D121
Drosophila Genetic Reference Panel 900, Male, Environment 1, Fly 6, Library 1


title: DGRP320 M_E1_1_L1

description:
EP2E051
Drosophila Genetic Reference Panel 320, Male, Environment 1, Fly 1, Library 1


title: DGRP703 M_E1_2_L1

description:
EP2D111
Drosophila Genetic Reference Panel 703, Male, Environment 1, Fly 2, Library 1


title: DGRP900 M_E1_2_L1

description:
EP2D101
Drosophila Genetic Reference Panel 900, Male, Environment 1, Fly 2, Library 1


title: Th17_ChIRP_RMRP_WT_DDX5ko_Odd

description:
Affinity purified genomic DNA
probes 1, 3, 5
Th17_ChIRP_RMRP_WT_DDX5ko_Input.bw,Th17_ChIRP_RMRP_WT_DDX5ko.bw


title: DGRP229 F_E1_6_L1

description:
EP2D091
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 6, Library 1


title: DGRP787 F_E1_4_L1

description:
EP2D081
Drosophila Genetic Reference Panel 787, Female, Environment 1, Fly 4, Library 1


title: G38_DDX5Tko_Th17_ChIPseqInput

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 mm10 whole genome using Bowtie
Peaks from the merged datasets were called using MACS.
Genome_build: mm10
Supplementary_files_format_and_content: BED file from MACS peak called.

extract_protocol:
Total  and IP enriched sonicated chromatin from cultured Th17 cells were harvested using Qiagen PCR purification kit.
ChIP-seq libraries were prepared for sequencing using standard Illumina protocols. 1% PhiX spiked in during the sequencing process and that allows us to calculate QC metrics for each lane and determine if poor performance is library or sequencer related.

value: DDX5fl/fl, CD4Cre+

tag: chip antibody vendor

tag: chip antibody cat. #

tag: chip antibody lot#


title: DGRP630 M_E1_3_L1

description:
EP2D051
Drosophila Genetic Reference Panel 630, Male, Environment 1, Fly 3, Library 1


title: DGRP790 M_E1_2_L1

description:
EP2D061
Drosophila Genetic Reference Panel 790, Male, Environment 1, Fly 2, Library 1


title: DGRP850 F_E1_8_L1

description:
EP2E081
Drosophila Genetic Reference Panel 850, Female, Environment 1, Fly 8, Library 1


title: G34_DDX5Tko_Th17_ChIPseqDDX5

value: goat anti-DDX5

value: Abcam


title: DGRP761 M_E1_6_L1

description:
EP2D011
Drosophila Genetic Reference Panel 761, Male, Environment 1, Fly 6, Library 1


title: DGRP804 M_E1_1_L1

description:
EP2D021
Drosophila Genetic Reference Panel 804, Male, Environment 1, Fly 1, Library 1


title: DGRP93 F_E1_5_L1

description:
EP2D041
Drosophila Genetic Reference Panel 93, Female, Environment 1, Fly 5, Library 1


title: G39_WT_Th17_ChIPseqInput

value: DDX5+/+, CD4Cre+


title: DGRP93 F_E1_4_L1

description:
EP2C121
Drosophila Genetic Reference Panel 93, Female, Environment 1, Fly 4, Library 1


title: DGRP563 M_E1_5_L1

description:
EP2D031
Drosophila Genetic Reference Panel 563, Male, Environment 1, Fly 5, Library 1


title: DGRP787 M_E1_3_L1

description:
EP2C101
Drosophila Genetic Reference Panel 787, Male, Environment 1, Fly 3, Library 1


title: G36_WT_Th17_ChIPseqDDX5


title: RRBseq_Smad1_5_KO_3

city: Boston

first: Ruben

last: Dries

address: 360 Longwood Ave

department: Computational Biology

institute: DFCI

data_processing:
Illumina Casava Pipeline 1.8.2 was used for BaseCalling
We cleaned sequencing reads with Trim Galore v0.3.7 using the parameters --phred64 --fastqc --stringency 3 --rrbs --non_directional --paired
We aligned to the mouse mm10 genome with bismark v0.14.1 using the parameters --bowtie2 --phred64-quals -X 1000 --non_directional
We extracted methylation information from the aligned reads with bismark command bismark_methylation_extractor using parameters --paired_end --no_overlap --ignore 3 --ignore_r2 3 --comprehensive --merge_non_CpG
We extracted cytosine coverage information from the aligened reads with bismark command coverage2cytosine using default parameters
Further processing and analysis of the data was performed in R v3.1.3 using the methylKit v0.9.4 package and in-house custom scripts
Genome_build: mm10
Supplementary_files_format_and_content: BedGraph file containing methylation percentage scores for all CpGs with at least 5x coverage

extract_protocol:
gDNA extraction was performed using the Wizard Genomic DNA purification kit (Promega)
gDNA concentration was measured with Quant-it Picogreen dsDNA assay and 1 μg input was used in the MSP1 digest. Following overnight incubation at 37°C, digestion reactions were terminated by adding 0.5 M EDTA and purified on a GeneJET PCR purification column. Libraries were prepped using the NEBNext Ultra DNA library preparation kit for Illumina and methylated adapters (Index primers Set 1). Subsequently, adapter ligated fragments were BS converted using the EZ DNA Methylation Gold kit (Zymo Research). 14 cycles of PCR were performed and the products were purified using AMPure XP beads. Quality of the final libraries was checked on a High sensitivity DNA chip (Agilent) and concentration was measured with qPCR. Sequencing was done on an Illumina HiSeq2500 PE 2x50bp

value: mixed CD-1, 129/Ola, C57Bl/6

tag: culture conditions

value: 2i + Lif on gelatin


title: DGRP320 M_E1_2_L1

description:
EP2C111
Drosophila Genetic Reference Panel 320, Male, Environment 1, Fly 2, Library 1


title: DGRP229 F_E1_2_L1

description:
EP2C091
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 2, Library 1


title: RRBseq_Smad1_5_KO_4


title: RRBseq_Smad1_5_KO_1


title: DGRP630 M_E1_5_L1

description:
EP2C081
Drosophila Genetic Reference Panel 630, Male, Environment 1, Fly 5, Library 1


title: RRBseq_Smad1_5_FL_2

value: Smad1/5 fl/fl


title: RRBseq_Smad1_5_FL_1


title: DGRP822 M_E1_6_L1

description:
EP2C071
Drosophila Genetic Reference Panel 822, Male, Environment 1, Fly 6, Library 1


title: RRBseq_BRE_pos_3

description:
FACS sorted based on SSEA1 and GFP
BRE_pos_CpG_methylation.bdg

value: BRE-GFP

value: mixed CBA, C57Bl/6

value: FCS + Lif on feeders


title: DGRP703 F_E1_4_L1

description:
EP2C061
Drosophila Genetic Reference Panel 703, Female, Environment 1, Fly 4, Library 1

value: Adult

value: Whole body


title: RRBseq_BRE_neg_3

description:
FACS sorted based on SSEA1 and GFP
BRE_neg_CpG_methylation.bdg


title: DGRP900 F_E1_3_L1

description:
EP2C051
Drosophila Genetic Reference Panel 900, Female, Environment 1, Fly 3, Library 1

value: Female


title: DGRP787 M_E1_4_L1

description:
EP2C041
Drosophila Genetic Reference Panel 787, Male, Environment 1, Fly 4, Library 1

value: Male


title: RRBseq_BRE_pos_2


title: DGRP630 F_E1_2_L1

description:
EP2C031
Drosophila Genetic Reference Panel 630, Female, Environment 1, Fly 2, Library 1


title: RRBseq_BRE_pos_1


title: DGRP352 F_E1_2_L1

description:
EP2C021
Drosophila Genetic Reference Panel 352, Female, Environment 1, Fly 2, Library 1


title: DGRP822 M_E1_7_L1

description:
EP2C011
Drosophila Genetic Reference Panel 822, Male, Environment 1, Fly 7, Library 1


title: RRBseq_BRE_neg_2


title: DGRP812 M_E1_4_L3

description:
EP2B123
Drosophila Genetic Reference Panel 812, Male, Environment 1, Fly 4, Library 3


title: RRBseq_BRE_neg_1


title: RNAseq_Smad1_5_KO_3

data_processing:
Illumina Casava Pipeline 1.8.2 was used for BaseCalling
We checked the quality of our reads with FASTQC v0.11.3_devel
We aligned to the mouse mm10 genome with STAR v2.4.1d using the parameters --sjdbOverhang 150
counts: featureCounts v1.4.6 was used to count the number of genes for Ensembl GRCm38.p4 annotated genes with parameters -t exon and -g gene_name.
differential expression: normalization of gene expression levels and differential gene expression between conditions was calculated with DESeq2 v1.6.3.
Genome_build: mm10
Supplementary_files_format_and_content: Matrix for all samples with raw counts as tab-delimited text file. Matrix for all samples with Transcript Per Million (TPM) counts as tab-delimited text file.

molecule: total RNA

extract_protocol:
RNA was extracted using RNAeasy minikit (Qiagen).
Libraries were prepared using the Illumina's TruSeq Stranded Total RNA kit protocol. In short, the first step involves the removal of ribosomal RNA (rRNA) using biotinylated, target-specific oligos combined with Ribo- Zero rRNA removal beads. The Ribo-Zero Human/Mouse/Rat kit depletes samples ofcytoplasmic rRNA and the Ribo-Zero Gold kit depletes samples of both cytoplasmic andmitochondrial rRNA. Ribo-Zero Globin kit depletes globin-encoding mRNA in addition tothe rRNA species targeted with Ribo-Zero Gold. Ribo-Zero Plant kit targets cytoplasmic andchloroplast rRNA. Following purification, the RNA is fragmented into small pieces usingdivalent cations under elevated temperature. The cleaved RNA fragments are copied into first strand cDNA using reverse transcriptase and random primers, followed by sec- ondstrand cDNA synthesis using DNA Polymerase I and RNase H. These cDNA fragmentsthen have the addition of a single 'A' base and subsequent ligation of the adapter. The products are purified and enriched with PCR to create the final cDNA library.


title: RNAseq_Smad1_5_KO_4


title: RNAseq_Smad1_5_KO_1


title: RNAseq_Smad1_5_KO_2


title: RRBseq_Smad1_5_KO_2


title: DGRP630 F_E1_5_L1

description:
EP2B081
Drosophila Genetic Reference Panel 630, Female, Environment 1, Fly 5, Library 1


title: DGRP93 M_E1_2_L1

description:
EP2B091
Drosophila Genetic Reference Panel 93, Male, Environment 1, Fly 2, Library 1


title: DGRP229 F_E1_4_L1

description:
EP2B111
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 4, Library 1


title: DGRP93 M_E1_6_L1

description:
EP2B071
Drosophila Genetic Reference Panel 93, Male, Environment 1, Fly 6, Library 1


title: RNAseq_Smad1_5_FL_2


title: DGRP703 M_E1_7_L1

description:
EP2B061
Drosophila Genetic Reference Panel 703, Male, Environment 1, Fly 7, Library 1


title: DGRP804 M_E1_5_L1

description:
EP2B101
Drosophila Genetic Reference Panel 804, Male, Environment 1, Fly 5, Library 1


title: DGRP761 M_E1_7_L1

description:
EP2B041
Drosophila Genetic Reference Panel 761, Male, Environment 1, Fly 7, Library 1


title: RNAseq_BRE_pos_3

description: FACS sorted based on SSEA1 and GFP


title: RNAseq_Smad1_5_FL_1


title: RNAseq_BRE_neg_3

description: FACS sorted based on SSEA1 and GFP


title: DGRP93 M_E1_1_L1

description:
EP2B031
Drosophila Genetic Reference Panel 93, Male, Environment 1, Fly 1, Library 1


title: RNAseq_BRE_neg_2

description: FACS sorted based on SSEA1 and GFP


title: DGRP900 F_E1_4_L1

description:
EP2B021
Drosophila Genetic Reference Panel 900, Female, Environment 1, Fly 4, Library 1


title: RNAseq_BRE_pos_2

description: FACS sorted based on SSEA1 and GFP


title: DGRP850 M_E1_2_L1

description:
EP2B011
Drosophila Genetic Reference Panel 850, Male, Environment 1, Fly 2, Library 1


title: RNAseq_BRE_neg_1

description: FACS sorted based on SSEA1 and GFP


title: G246_RORgko_Th17_SL85764

description: processed data file: G340_CuffDiff_WTvsRORgko.tabular.txt

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 Tophat2
Transcripts FPKM were calculated by Cufflink using Reference Annotation: mm9.gtf from UCSC
Differential gene expression were called by CuffDiff using Reference Annotation: mm9.gtf from UCSC
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

source_name: RORgKO_In vitro Th17

extract_protocol:
Total RNAs from cultured Th17 cells were harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: strain background

tag: genotype/variation

value: RORgKO

value: T cell

value: in vitro polarized T helper 17 cells


title: DGRP320 M_E1_4_L1

description:
EP2B051
Drosophila Genetic Reference Panel 320, Male, Environment 1, Fly 4, Library 1


title: DGRP900 F_E1_5_L1

description:
EP2A111
Drosophila Genetic Reference Panel 900, Female, Environment 1, Fly 5, Library 1


title: DGRP787 F_E1_5_L1

description:
EP2A081
Drosophila Genetic Reference Panel 787, Female, Environment 1, Fly 5, Library 1


title: G141_RmrpG262T_Th17_SL85755

source_name: RmrpG262T_In vitro Th17


title: DGRP563 F_E1_4_L1

description:
EP2A101
Drosophila Genetic Reference Panel 563, Female, Environment 1, Fly 4, Library 1


title: G105_DDX5Tko_Th17_SL85753

description: processed data file: G263_CuffDiff_WTvsDDX5Tko.tabular.txt

source_name: DDX5Tko_In vitro Th17


title: DGRP804 M_E1_6_L1

description:
EP2A091
Drosophila Genetic Reference Panel 804, Male, Environment 1, Fly 6, Library 1


title: G83_DDX5Tko_Th17_SL85751


title: G100_WT_Th17_SL85752

source_name: WT_In vitro Th17

value: WT


title: DGRP352 M_E1_4_L3

description:
EP2A063
Drosophila Genetic Reference Panel 352, Male, Environment 1, Fly 4, Library 3


title: DGRP320 F_E1_7_L1

description:
EP2A051
Drosophila Genetic Reference Panel 320, Female, Environment 1, Fly 7, Library 1


title: DiabEC2

city: Kuopio

first: Mohan

last: Babu

address: Neulaniementie 2

department: Biotechnology and Molecular Medicine

country: Finland

institute: A.I.Virtanen Institute, University of Eastern Finland

data_processing:
bismark 0.10.0 (relying on bowtie 2-2.0.5) for read alignment to mm10 genome (derived from UCSC)
methylKit 0.9.2 R package for differential methylation calling (R version 3.0.1). Prefiltering using function filterByCoverage with parameters lo.count=10 and  hi.perc=99.0, differential methylation calculated using function calculateDiffMeth.
Genome_build: mm10
Supplementary_files_format_and_content: Tab delimited text files.  <comparison>_Diff-all-promoter.txt files contain differential methylation results for proximal promoter regions (-1kb upstream and +500 bp downstream of the transcription start site (TSS)). Columns: row number, chromosome, start coordinate, end coordinate, strand, pvalue, qvalue, methylation difference.

source_name: Gastrocnemius muscle

extract_protocol:
Genomic DNA was isolated from FACS sorted macrophages and endothelial cells using AllPrep DNA/RNA/Protein Mini Kit (Qiagen Finland, Helsinki, Finland) according to manufacturer's instructions.
Reduced representation bisulfite sequencing (RRBS) library was generated as described previously (Boyle P, Clement K, Gu H, Smith ZD, Ziller M, Fostel JL, Holmes L, Meldrim J, Kelley F, Gnirke A, Meissner A: Gel-free multiplexed reduced representation bisulfite sequencing for large-scale DNA methylation profiling. Genome Biol 2012, 13(10):R92).

treatment_protocol: Unilateral hindlimb ischemia was induced by ligating femoral artery proximal to the bifurcation of superficial and deep femoral artery in control, hyperlipidemic and type 2 diabetic mice. Ishcemic muscle macrophages and endothelial cells were FACS sorted at 1 week time point after femoral artery ligation

value: C57BL/6J background

value: Enodthelial cells

tag: condition

value: type 2 diabetic


title: DiabEC3


title: DiabEC1


title: DGRP229 F_E1_8_L2

description:
EP2A032
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 8, Library 2


title: HypEC2

value: Hyperlipidemic


title: DGRP370 F_E1_5_L1

description:
EP2A011
Drosophila Genetic Reference Panel 370, Female, Environment 1, Fly 5, Library 1


title: DGRP320 F_E1_3_L3

description:
EP1H123
Drosophila Genetic Reference Panel 320, Female, Environment 1, Fly 3, Library 3


title: HypEC1


title: DGRP352 F_E1_6_L1

description:
EP1H101
Drosophila Genetic Reference Panel 352, Female, Environment 1, Fly 6, Library 1


title: ControlEC3

value: normal


title: DGRP229 F_E1_1_L1

description:
EP1H091
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 1, Library 1


title: G72_WT_Th17_SL85750

tag: age


title: DiabMac3

value: macrophages


title: DGRP703 M_E1_3_L1

description:
EP1H071
Drosophila Genetic Reference Panel 703, Male, Environment 1, Fly 3, Library 1


title: DGRP352 F_E1_1_L1

description:
EP1H081
Drosophila Genetic Reference Panel 352, Female, Environment 1, Fly 1, Library 1


title: DGRP563 M_E1_3_L1

description:
EP2A041
Drosophila Genetic Reference Panel 563, Male, Environment 1, Fly 3, Library 1


title: DGRP370 F_E1_8_L1

description:
EP1H061
Drosophila Genetic Reference Panel 370, Female, Environment 1, Fly 8, Library 1


title: DiabMac1


title: ControlEC1


title: DGRP790 F_E1_8_L1

description:
EP1H041
Drosophila Genetic Reference Panel 790, Female, Environment 1, Fly 8, Library 1


title: DGRP320 M_E1_3_L1

description:
EP1H051
Drosophila Genetic Reference Panel 320, Male, Environment 1, Fly 3, Library 1


title: DGRP850 M_E1_3_L1

description:
EP2A021
Drosophila Genetic Reference Panel 850, Male, Environment 1, Fly 3, Library 1


title: DGRP761 M_E1_3_L1

description:
EP1H031
Drosophila Genetic Reference Panel 761, Male, Environment 1, Fly 3, Library 1


title: HypEC3


title: DGRP352 M_E1_3_L1

description:
EP1H111
Drosophila Genetic Reference Panel 352, Male, Environment 1, Fly 3, Library 1


title: HypMac2


title: ControlMac3


title: DGRP850 F_E1_1_L1

description:
EP1H011
Drosophila Genetic Reference Panel 850, Female, Environment 1, Fly 1, Library 1


title: DGRP822 F_E1_4_L1

description:
EP1G121
Drosophila Genetic Reference Panel 822, Female, Environment 1, Fly 4, Library 1


title: DGRP630 F_E1_7_L1

description:
EP1H021
Drosophila Genetic Reference Panel 630, Female, Environment 1, Fly 7, Library 1


title: HypMac1


title: ControlMac2


title: ControlEC2


title: ControlMac1


title: DGRP93 M_E1_4_L1

description:
EP1G111
Drosophila Genetic Reference Panel 93, Male, Environment 1, Fly 4, Library 1


title: DiabMac2


title: DGRP630 M_E1_4_L1

description:
EP1G091
Drosophila Genetic Reference Panel 630, Male, Environment 1, Fly 4, Library 1


title: DGRP850 M_E1_6_L1

description:
EP1G101
Drosophila Genetic Reference Panel 850, Male, Environment 1, Fly 6, Library 1


title: DGRP850 M_E1_4_L1

description:
EP1G081
Drosophila Genetic Reference Panel 850, Male, Environment 1, Fly 4, Library 1


title: HypMac3


title: DGRP630 M_E1_2_L3

description:
EP1G073
Drosophila Genetic Reference Panel 630, Male, Environment 1, Fly 2, Library 3


title: DGRP352 F_E1_5_L1

description:
EP1G051
Drosophila Genetic Reference Panel 352, Female, Environment 1, Fly 5, Library 1


title: G241_WT_Th17_SL85763

description: processed data file: G340_CuffDiff_WTvsRORgko.tabular.txt


title: Ischemic muscle DNA from Hyperlipidemic mice

data_processing:
Color space data were preprocessed and aligned to the reference genome using Life Technologies' LifeScope software
Relative methylation scores were calculated with MEDIPS package in R/Bioconductor
Genome_build: mm9
Supplementary_files_format_and_content: wig files were generated using MEDIPS package in R/Bioconductor. Scores represent the relative methylation values.

extract_protocol:
Tissue samples were snap frozen in liquid nitrogen at day 7 after ischemia induction and later DNA was isolated using DNeasy Tissue kit (Qiagen).
The quality and quantity of DNA was analyzed with nanodrop. 5 µg of genomic DNA was sheared into small fragments with a mean size of 150 bp by using a Covaris™ S2 sonicator System (www.covaris.com). Quality of the fragmentation was analyzed with Bioanalyzer (www.agilent.com). Fragmented DNA samples with sufficient yield and an appropriate size distribution were used for preparation of DNA fragment libraries.  Methylated DNA was captured according to the manufacturer’s protocol [The Methyl Miner™ methylated DNA Enrichment Kit (Invitrogen)]. Methylated DNA was isolated from fragmented whole genomic DNA (5 ng –25 μg) via binding to the methyl-CpG binding domain of human MBD2 protein, which is coupled to paramagnetic Dynabeads RM-280 Streptavidin via a biotin linker. The amount of input DNA was 3 ug. The captured DNA fragments were eluted with 2000 mM NaCl. End Polishing Enzyme 1 and End Polishing Enzyme 2 were used to convert DNA that has damaged or incompatible 5'-protruding and/or 3'-protruding ends to 5'-phosphorylated, blunt-ended DNA. End Polishing Enzyme 1 and ATP were also included for phosphorylation of the 5'-ends of the blunt-ended DNA to allow for subsequent adaptor ligation. End repaired samples were purified with the PureLink™ PCR Purification columns (Invitrogen). The double-stranded P1 and P2 adaptors were hybridized and ligated to the ends of fragmented DNA. These adaptors contained defined sequences required for SOLiD sequencing. For multiplexing, the SOLiD™ Fragment Library Barcoding Kit Module 1–16 was used. The samples were nick translated and subsequently amplified using Library PCR Primers 1 and 2 and Platinum® PCR Amplification Mix. After amplification, PCR samples were purified with the Agencourt AM XP beads. The amplified and purified DNA was run on a 1 % agarose gel. The correct sized ligation products (150 to 300 bp) were cut from the gel according to the 50 bp ladder (Invitrogen). The size-selected product was elute extracted (QIAquick gel extraction kit, Qiagen) from the gel. The quality and size of the samples were then analyzed with Bioanalyzer.  Libraries were quantitated with Qubit fluorometer (Invitrogen) and quantitative PCR to warrant the accuracy. SOLiD Library TaqMan Quantitation Kit was used for determining the molar concentration of amplified template in a SOLiD library. The uniformity of fragment size of libraries was confirmed with Bioanalyzer. For SOLiD fragment libraries the target size of the fragments was 150-300 bp. Templated bead preparation was performed by emulsion PCR (ePCR), which involves monoclonal amplification of individual species of template DNA from a complex library pool. During ePCR, multiple copies of a single DNA sequence were coated onto a single 1-μm magnetic bead (SOLiD™ P1 DNA Beads) within a water-in-oil emulsion droplet. SOLiD™ EZ Bead™ instrumentation was used for templated bead preparation. Before ePCR, equal molar amounts (250 pM) of each of the bar coded libraries were mixed together and one ePCR reaction was performed with the library mixture (template 1 pM). After ePCR and enrichment of the template beads, 800 mlj beads were deposited onto one single chamber sequencing glass slide. Subsequently, samples were sequenced on the SOLiD4 next generation sequencing instrument (Applied Biosystems) on one flow cell with SOLiD4 chemistry for 50 bp reads.

treatment_protocol: Mice were anesthetized by subcutaneous injection of mixture containing Rompun® (10 mg/kg) (Xylazine, Orion pharma, Espoo, Finland) and Ketamine (100 mg/kg) (Ketalar, Pfizer Oy, Espoo, Finland) before surgery. Unilateral hindlimb ischemia was induced by ligating femoral artery proximal to the bifurcation of superficial and deep femoral artery as described previously.

value: LDLR-/-ApoB100/100 , with C57BL/6J background

value: Gastrocnemius muscle


title: Ischemic muscle DNA from Normal mice


title: DGRP804 F_E1_1_L1

description:
EP1G041
Drosophila Genetic Reference Panel 804, Female, Environment 1, Fly 1, Library 1


title: Run297_L6_H01_single cell

city: Stockholm

first: Sten

last: Linnarsson

address: Scheeles väg 1

department: Medical Biochemistry and Biophysics

country: Sweden

institute: Karolinska Institutet

data_processing:
Read quality filtering and barcode extraction: Every read that was considered valid by the Illumina HiSeq control software was processed and filtered as follows: a) any 3' bases with a quality score of 'B' were removed; b) the well-identifying barcode was extracted from the 5' end of the read; c) if the read ended in a poly(A)-sequence leaving less than 25 transcript-derived bases, the read was discarded; d) the 8-base UMI was extracted from the 5' end and if any of the UMI bases had a Phred score less than 17 the read was discarded; e) a maximum of nine template-switching generated Gs were removed from the 5' end of the transcript-derived sequence; f) if the remaining sequence consisted of less than six non-A bases, or a dinucleotide repeat with less than six other bases at either end, the read was discarded.
Alignment:the reads were aligned to the genome using the Bowtie aligner, allowing for up to three mismatches and up to 24 alternative mappings for each read. The genome included an artifical chromosome, containing a concatamer of the ERCC control sequences. Any reads with no alignments were re-aligned against another artifical chromosome, containing all possible splice junctions arising from the exons defined by the known transcript variants. Reads mapping within these splice junctions were translated back to the corresponding actual genomic positions. The UCSC transcript models22 were used for the expression level calculation. If a locus had several transcript variants, the exons of these were merged to a combined model that represented all expression from the locus. To account for incompete cap site knowledge, the 5' ends of all models were extended by 100 bases, but not beyond the 3' end of any upstream nearby exon of another gene of the same orientation.
Annotation and quantitation: The annotation step was performed barcode-by-barcode. For every unique mapping (genomic position and strand) the number of reads in each UMI was counted. Any multiread that had one or more repeat mappings that was outside exons, was assigned randomly as one of these repeats and contributed to the summarized read count of that repeat class. Else, if it had one or more mappings to exons, it was assigned at the exon where it was closest to the transcript model 5' end, even if the sequence was repeat-like. If it had no exon mapping, it was assigned randomly at one of the mappings. After assigning reads, the number of molecules at each mapping position was determined by the number of distinct UMIs observed. To account for UMIs that stem from PCR-induced mutations or sequencing errors, any UMI that had fewer reads than 1/100 of the average of the non-zero UMIs was excluded. The raw UMI count was corrected for the UMI collision probability (important only at high UMI counts) as described10. The expression level of each transcript model was taken as the total number of molecules at all its possible mapping positions.
Genome_build: UCSC mm10
Supplementary_files_format_and_content: excel file containing counts of detected cDNA molecules for each transcript model in each cell.

source_name: cortical neuron

growth_protocol: We used a dual reporter mouse line expressing discosoma red fluorescent protein (DsRed) under regulatory elements of the cholecystokinin promoter and green fluorescent protein (GFP) knocked into  the glutamate decarboxylase 67 gene .  Animals were group-housed under 12:12 h/h  light/dark cycle with ad libitum access to water and food. Animals of both sexes were used  for neurophysiology experiments during postnatal days 17-23.

extract_protocol:
At the end of each patch-clamp protocol, the micropipette was clamped to a holding potential. The entire soma of each recorded neuron was aspirated into the micropipette slowly by applying negative pressure, and ejected into a drop of lysis buffer.
20 microL lysis buffer (0.15% Triton X-100, 1 U/microL TaKaRa RNase inhibitor, 4 microM reverse transcription primer C1-P1-T31, 5% C1 Loading Reagent and 1:50,000 Life Technologies ERCC  Spike-In Mix 1), reverse transcription mix (1x SuperScript II First-Strand Buffer supplemented with 3 mM MgCl2, 1.5 mM dNTP, 4 mM DTT, 3.3% C1 Loading Reagent, 1.8 microM template-switching oligo C1-P1-RNA-TSO, 1.5 U/microL TaKaRa RNase inhibitor and 18 U/microL Life Technologies Superscript II reverse transcriptase) and PCR mix (1.1x Clontech Advantage2 PCR buffer, 440 microM dNTP, 530 nM PCR primer C1-P1-PCR-2, 5% C1 Loading Reagent and 2x Advantage2 Polymerase Mix) were added to the designated wells according to the manufacturer's instructions, but using the indicated mixes in place of the corresponding commmercial reagents. The plate was then returned to the Fluidigm C1 and the mRNA Seq: RT + Amp (1772x/1773x) script was executed, which took about 8.5 hours and included lysis, reverse transcription and 21 cycles of PCR. When the run finished, the amplified cDNA was harvested in a total of 13 microL C1 Harvesting Reagent and quantified on an Agilent BioAnalyzer. The typical yield was 1 ng/microL. Amplified cDNA was simultaneously fragmented and barcoded by tagmentation, i.e. using Tn5 DNA transposase to transfer adapters to the target DNA. 96 different 10x transposome stocks (6.25 microM barcoded adapter C1-TN5-x, 40% glycerol, 6.25 microM Tn5 transposase, where x denotes a well-specific barcode) were prepared, each with a different barcode sequence.  6 microL harvested cDNA was mixed with 5 microL tagmentation buffer (50 mM TAPS-NaOH pH 8.5, 25 mM MgCl2 and 50% DMF), 11.5 microL nuclease-free water and 2.5 microL 10x transposome stock. The mix was incubated for 5 minutes at 55 degeees C then cooled on ice. 100 microL Dynabeads MyOne Streptavidin C1 beads were washed in 2xBWT (10 mM Tris HCl pH 7.5, 1 mM EDTA, 2 M NaCl, 0.02% Tween-20) then resuspended in 2 mL 2xBWT. 20 microL beads was added to each well and incubated at room temperature for 5 minutes. All fractions were pooled, the beads were immobilized and the supernatant removed (thus removing all internal fragments, and retaining only the 5'- and 3'-most fragments). The beads were then resuspended in 100 microL TNT (20 mM Tris pH 7.5, 50 mM NaCl, 0.02% Tween), washed in 100 microL Qiagen Qiaquick PB, then twice in 100 microL TNT. The beads were then resuspended in 100 microL restriction mix (1x NEB NEBuffer 4, 0.4 U/microL PvuI-HF enzyme), designed to cleave 3' fragments carrying the PvuI recognition site. The mix was incubated for one hour at 37 degrees C, then washed three times in TNT. Finally, the single-stranded library was eluted by resuspending the beads in 100 microL 100 mM NaOH, incubating for 5 minutes, removing the beads, then adding 100 microL 100 mM HCl and 50 microL Neutralization buffer (200 mM Tris pH 7.5, 0.05% Tween-20). At this point, the typical yield was 1-5 nM single-stranded library. Aliquots were rom these were run on  with 187ds_).

treatment_protocol: Coronal slices of primary somatosensory cortex were prepared in ice-cold artificial cerebrospinal fluid. Neurons were visualized by differential interference contrast microscopy and co-existence of DsRed and GFP in interneurons was confirmed by epifluorescence microsopy, and whole-cell patch clamp recordings were made.

value: a dual reporter line expressing DsRed under regulatory elements of the cholecystokinin promoter and GFP knocked into the glutamate decarboxylase 67 gene

value: postnatal days 17-23

tag: cell id

value: cortical neuron


title: Run297_L6_H03_single cell


title: Run297_L6_H05_single cell


title: Run297_L6_G05_single cell


title: DGRP804 F_E1_7_L1

description:
EP1G031
Drosophila Genetic Reference Panel 804, Female, Environment 1, Fly 7, Library 1

tag: strain

value: Adult

value: Female

value: Whole body


title: DGRP787 F_E1_8_L1

description:
EP1F121
Drosophila Genetic Reference Panel 787, Female, Environment 1, Fly 8, Library 1

tag: developmental stage


title: Run297_L6_G01_single cell


title: DGRP822 M_E1_8_L1

description:
EP1G021
Drosophila Genetic Reference Panel 822, Male, Environment 1, Fly 8, Library 1

tag: Sex

value: Male


title: DGRP563 F_E1_1_L1

description:
EP1G011
Drosophila Genetic Reference Panel 563, Female, Environment 1, Fly 1, Library 1

tag: tissue


title: Run297_L6_F05_single cell


title: DGRP93 F_E1_7_L1

description:
EP1F111
Drosophila Genetic Reference Panel 93, Female, Environment 1, Fly 7, Library 1


title: Run297_L6_F03_single cell


title: DGRP563 M_E1_4_L1

description:
EP1F101
Drosophila Genetic Reference Panel 563, Male, Environment 1, Fly 4, Library 1


title: DGRP790 F_E1_1_L1

description:
EP1F081
Drosophila Genetic Reference Panel 790, Female, Environment 1, Fly 1, Library 1


title: Ischemic muscle DNA from type 2 diabetic mice

value: IGF-II/LDLR-/-ApoB100/100 , with C57BL/6J background


title: DGRP787 M_E1_6_L1

description:
EP1F091
Drosophila Genetic Reference Panel 787, Male, Environment 1, Fly 6, Library 1


title: Run297_L6_F01_single cell


title: Run297_L6_E04_single cell


title: DGRP352 F_E1_3_L1

description:
EP1F061
Drosophila Genetic Reference Panel 352, Female, Environment 1, Fly 3, Library 1


title: DGRP804 F_E1_6_L1

description:
EP1F071
Drosophila Genetic Reference Panel 804, Female, Environment 1, Fly 6, Library 1


title: Run297_L6_E01_single cell


title: DGRP850 F_E1_2_L1

description:
EP1F051
Drosophila Genetic Reference Panel 850, Female, Environment 1, Fly 2, Library 1


title: Run297_L6_D05_single cell


title: DGRP370 M_E1_4_L1

description:
EP1F041
Drosophila Genetic Reference Panel 370, Male, Environment 1, Fly 4, Library 1


title: Run297_L6_D01_single cell


title: DGRP93 F_E1_3_L3

description:
EP1F033
Drosophila Genetic Reference Panel 93, Female, Environment 1, Fly 3, Library 3


title: DGRP630 F_E1_3_L1

description:
EP1E121
Drosophila Genetic Reference Panel 630, Female, Environment 1, Fly 3, Library 1


title: Run297_L6_B06_single cell


title: Run297_L6_B03_single cell


title: Run297_L6_B04_single cell


title: DGRP822 F_E1_5_L1

description:
EP1E111
Drosophila Genetic Reference Panel 822, Female, Environment 1, Fly 5, Library 1


title: Run297_L6_B01_single cell


title: DGRP822 M_E1_4_L1

description:
EP1E101
Drosophila Genetic Reference Panel 822, Male, Environment 1, Fly 4, Library 1


title: Run297_L6_E02_single cell


title: DGRP93 M_E1_5_L1

description:
EP1E091
Drosophila Genetic Reference Panel 93, Male, Environment 1, Fly 5, Library 1


title: Run297_L6_A05_single cell


title: DGRP703 M_E1_8_L3

description:
EP1E083
Drosophila Genetic Reference Panel 703, Male, Environment 1, Fly 8, Library 3


title: Run297_L6_A04_single cell


title: Run297_L6_A03_single cell


title: DGRP804 M_E1_3_L1

description:
EP1E051
Drosophila Genetic Reference Panel 804, Male, Environment 1, Fly 3, Library 1


title: Run297_L6_A01_single cell


title: Run291_L7_G11_single cell


title: Run291_L7_G10_single cell


title: Run297_L6_D04_single cell


title: DGRP229 F_E1_7_L1

description:
EP1F011
Drosophila Genetic Reference Panel 229, Female, Environment 1, Fly 7, Library 1


title: DGRP563 F_E1_8_L3

description:
EP1E033
Drosophila Genetic Reference Panel 563, Female, Environment 1, Fly 8, Library 3


title: DGRP790 M_E1_5_L3

description:
EP1E043
Drosophila Genetic Reference Panel 790, Male, Environment 1, Fly 5, Library 3


title: DGRP320 F_E1_4_L3

description:
EP1E023
Drosophila Genetic Reference Panel 320, Female, Environment 1, Fly 4, Library 3


title: DGRP850 M_E1_8_L1

description:
EP2A071
Drosophila Genetic Reference Panel 850, Male, Environment 1, Fly 8, Library 1


title: DGRP822 M_E1_5_L1

description:
EP1F021
Drosophila Genetic Reference Panel 822, Male, Environment 1, Fly 5, Library 1


title: DGRP761 F_E1_8_L1

description:
EP1E071
Drosophila Genetic Reference Panel 761, Female, Environment 1, Fly 8, Library 1


title: Run297_L6_C06_single cell


title: DGRP320 F_E1_5_L1

description:
EP1D111
Drosophila Genetic Reference Panel 320, Female, Environment 1, Fly 5, Library 1


title: DGRP370 F_E1_3_L3

description:
EP1E063
Drosophila Genetic Reference Panel 370, Female, Environment 1, Fly 3, Library 3


title: Run291_L7_G04_single cell


title: DGRP900 F_E1_6_L1

description:
EP1D121
Drosophila Genetic Reference Panel 900, Female, Environment 1, Fly 6, Library 1


title: Run291_L7_G02_single cell


title: DGRP370 F_E1_2_L1

description:
EP1D101
Drosophila Genetic Reference Panel 370, Female, Environment 1, Fly 2, Library 1


title: Run291_L7_G03_single cell


title: Run291_L7_F12_single cell


title: DGRP812 F_E1_1_L1

description:
EP1D081
Drosophila Genetic Reference Panel 812, Female, Environment 1, Fly 1, Library 1


title: DGRP563 F_E1_5_L1

description:
EP1D091
Drosophila Genetic Reference Panel 563, Female, Environment 1, Fly 5, Library 1


title: Run291_L7_F09_single cell


title: Run291_L7_F11_single cell


title: DGRP352 M_E1_6_L1

description:
EP1D061
Drosophila Genetic Reference Panel 352, Male, Environment 1, Fly 6, Library 1


title: Run291_L7_F03_single cell


title: DGRP822 F_E1_3_L1

description:
EP1D051
Drosophila Genetic Reference Panel 822, Female, Environment 1, Fly 3, Library 1


title: Run291_L7_G09_single cell


title: DGRP761 M_E1_5_L3

description:
EP1E013
Drosophila Genetic Reference Panel 761, Male, Environment 1, Fly 5, Library 3


title: Run291_L7_F02_single cell


title: Run291_L7_F01_single cell


title: Run291_L7_E11_single cell


title: DGRP761 M_E1_4_L1

description:
EP1D041
Drosophila Genetic Reference Panel 761, Male, Environment 1, Fly 4, Library 1


title: DGRP761 M_E1_8_L1

description:
EP1D021
Drosophila Genetic Reference Panel 761, Male, Environment 1, Fly 8, Library 1


title: Run291_L7_E10_single cell


title: Run291_L7_E09_single cell


title: DGRP804 M_E1_2_L1

description:
EP1D011
Drosophila Genetic Reference Panel 804, Male, Environment 1, Fly 2, Library 1


title: Run291_L7_E04_single cell


title: Run291_L7_E03_single cell


title: DGRP320 F_E1_8_L3

description:
EP1C123
Drosophila Genetic Reference Panel 320, Female, Environment 1, Fly 8, Library 3


title: Run291_L7_D11_single cell


title: Run291_L7_D12_single cell


title: DGRP320 F_E1_1_L1

description:
EP1C091
Drosophila Genetic Reference Panel 320, Female, Environment 1, Fly 1, Library 1


title: DGRP812 F_E1_5_L1

description:
EP1C111
Drosophila Genetic Reference Panel 812, Female, Environment 1, Fly 5, Library 1


title: DGRP229 M_E1_2_L1

description:
EP1C081
Drosophila Genetic Reference Panel 229, Male, Environment 1, Fly 2, Library 1


title: Run291_L7_D09_single cell


title: DGRP850 F_E1_7_L1

description:
EP1C071
Drosophila Genetic Reference Panel 850, Female, Environment 1, Fly 7, Library 1


title: Run291_L7_D06_single cell


title: DGRP370 F_E1_6_L1

description:
EP1C061
Drosophila Genetic Reference Panel 370, Female, Environment 1, Fly 6, Library 1


title: Run291_L7_D04_single cell


title: DGRP790 M_E1_7_L1

description:
EP1C051
Drosophila Genetic Reference Panel 790, Male, Environment 1, Fly 7, Library 1


title: DGRP790 M_E1_3_L1

description:
EP1D031
Drosophila Genetic Reference Panel 790, Male, Environment 1, Fly 3, Library 1


title: DGRP93 F_E1_1_L1

description:
EP1C041
Drosophila Genetic Reference Panel 93, Female, Environment 1, Fly 1, Library 1


title: Run291_L7_D01_single cell


title: DGRP703 M_E1_4_L3

description:
EP1C033
Drosophila Genetic Reference Panel 703, Male, Environment 1, Fly 4, Library 3


title: DGRP787 F_E1_3_L1

description:
EP1C021
Drosophila Genetic Reference Panel 787, Female, Environment 1, Fly 3, Library 1


title: DGRP352 M_E1_7_L1

description:
EP1C011
Drosophila Genetic Reference Panel 352, Male, Environment 1, Fly 7, Library 1


title: Run291_L7_D03_single cell


title: DGRP761 F_E1_4_L2

description:
EP1B122
Drosophila Genetic Reference Panel 761, Female, Environment 1, Fly 4, Library 2


title: Run291_L7_C09_single cell


title: Run291_L7_C11_single cell


title: DGRP563 F_E1_2_L1

description:
EP1B111
Drosophila Genetic Reference Panel 563, Female, Environment 1, Fly 2, Library 1


title: Run291_L7_C10_single cell


title: DGRP787 M_E1_1_L1

description:
EP1B101
Drosophila Genetic Reference Panel 787, Male, Environment 1, Fly 1, Library 1


title: DGRP787 M_E1_7_L1

description:
EP1B091
Drosophila Genetic Reference Panel 787, Male, Environment 1, Fly 7, Library 1


title: Run291_L7_C04_single cell


title: DGRP812 M_E1_2_L1

description:
EP1B081
Drosophila Genetic Reference Panel 812, Male, Environment 1, Fly 2, Library 1


title: RNAseq_BRE_pos_1

description: FACS sorted based on SSEA1 and GFP

tag: genotype

value: BRE-GFP

value: mixed CBA, C57Bl/6

value: FCS + Lif on feeders


title: Run291_L7_D10_single cell

tag: cell type


title: DGRP703 F_E1_7_L1

description:
EP1B071
Drosophila Genetic Reference Panel 703, Female, Environment 1, Fly 7, Library 1


title: Run291_L7_C02_single cell


title: Run297_L6_C02_single cell


title: Run291_L7_C01_single cell


title: DGRP630 F_E1_8_L1

description:
EP1B061
Drosophila Genetic Reference Panel 630, Female, Environment 1, Fly 8, Library 1


title: DGRP761 F_E1_5_L1

description:
EP1B051
Drosophila Genetic Reference Panel 761, Female, Environment 1, Fly 5, Library 1


title: Run291_L7_B12_single cell


title: Run291_L7_B10_single cell


title: DGRP900 F_E1_1_L1

description:
EP1B031
Drosophila Genetic Reference Panel 900, Female, Environment 1, Fly 1, Library 1


title: DGRP352 F_E1_7_L1

description:
EP1B041
Drosophila Genetic Reference Panel 352, Female, Environment 1, Fly 7, Library 1


title: Run291_L7_B09_single cell


title: DGRP563 M_E1_6_L1

description:
EP1B021
Drosophila Genetic Reference Panel 563, Male, Environment 1, Fly 6, Library 1


title: Run291_L7_B06_single cell


title: KO

city: Princeton

first: Sneha

last: Rath

state: NJ

address: Lewis Thomas Laboratory

department: Molecular Biology

institute: Princeton University

description: processed data file: Dataset 7.txt

data_processing:
TopHat 2 set to map stranded reads with default parameters
HTseq-count run in union mode
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimined text files with (1) header row; (2) RefSeq ID and accepted file names; and (3) read counts from HT-seq.

growth_protocol: All cell lines were grown using ATCC or provider-recommended conditions in MEM + 10% FBS (HeLa), RPMI + 10% FBS (T47D and WT and RNase L KO MEFs) or IMDM + 10% FBS (WT and RNase L KO HAP1). All media were purchased from Gibco, Life Technologies.

extract_protocol:
Total RNA was extracted (RNeasy kit, Qiagen). RNA integrity was verified by an RNA 6000 Nano Chip, using BioAnalyzer and 2100 Expert software (Agilent Technologies).
The mRNA was enriched by oligo-dT pulldown from total RNA, followed by fragmentation, adapter ligation, PCR amplification, and finally sequencing on Illumina HiSeq 2000 platform. For sequencing introns, the oligo-dT pulldown step was replaced with Ribo-Zero rRNA removal (Illumina).

treatment_protocol: For RNase L overexpression, an 80%-confluent 6-well plate of cells was transfected with 2 µg of WT or the H672N mutant plasmid using Lipofectamine 2000 reagent. Cells were harvested at 24 hours and 25% of the final volume was used for western blot, and the remainder for RNA extraction. For each RNase L knockdown, a combination of two DsiRNAs - 100 nM f/c HSC.RNAI.N021133.12.1, which targets the CDS/Exon 1 and 100 nM f/c HSC.RNAI.N021133.12.1, which targets the 3'-UTR/Exon 6 of RNase L - was used. Commercial scrambled DsiRNAs (200 nM f/c) were used as negative control. All DsiRNAs were purchased from IDT and transfected for 36 hours using Lipofectamine 2000 reagent. PBS or 1 µg/ml poly I:C was transfected subsequently for 6h. For rapid activation of RNase L in T47D, the cells were permeabilized with 0.05 mg/ml digitonin (Sigma) with or without 5 µM 5'-p-A2',5'A2',5'A2',5'A for 1, 3, or 9 minutes. Cells were harvested in 500 ml of RLT buffer (Qiagen) and stored at -80 °C until purification according to Qiagen protocol. RNase L cleavage assay in HeLa extracts. Seven 10 cm dishes of HeLa cells were harvested by trypsinization at ~80-90% confluence.  Cells were washed with 2x10 ml cold PBS and the cell pellet was frozen on dry ice and stored at -80 °C overnight. The cell pellet (~100 ul) was resuspended in 200 ul phosphate buffer (15 mM sodium phosphate pH 7, 137.5 mM NaCl, 6.125 mM MgCl2, 1.25 mM b-mercaptoethanol, and 0.125% Triton X-100) and incubated for 10 min at 4 °C with gentle rotation. The crude lysate was centrifuged for 10 min at 10,000g, 4 °C and the supernatant taken as S10 extract (A260=0.258). RNA cleavage assays were initiated by adding 16 ul of S10 to a tube containing 1 ul of 20 mM ATP, 2 ul of 1 uM RNase L or plain buffer (20 mM HEPES pH 7.5, 300 mM NaCl, 1 mM MgCl2, 10% glycerol, 2 mM DTT), and 1 ul of 200 uM 5'-p-A2',5'A2',5'A2',5'A.  Reactions were incubated at 20 °C and 4 ul aliquots were placed at time intervals into 350 ul of RLT buffer (Qiagen). Samples were stored at -80 °C and purified using RNeasy kit (Qiagen).

tag: tissue source

value: epithelium

value: RNase L -/-

tag: sample description

value: RNase L-KO HAP1 cells, Ribo-Zero sequencing

tag: experiment group

value: EXPERIMENT 7


title: Run291_L7_C06_single cell


title: Run291_L7_B04_single cell


title: Run291_L7_C03_single cell


title: Run291_L7_B02_single cell


title: Run291_L7_B03_single cell


title: DGRP850 M_E1_5_L3

description:
EP1A113
Drosophila Genetic Reference Panel 850, Male, Environment 1, Fly 5, Library 3


title: Run291_L7_B01_single cell


title: Hap1WT-2naive

description: processed data file: Dataset 6.txt

tag: cell line

value: wild type

value: HAP1 cells, poly-A sequencing

value: EXPERIMENT 6


title: Hap1WT-1naive

description: processed data file: Dataset 6.txt


title: DGRP93 M_E1_7_L1

description:
EP1A091
Drosophila Genetic Reference Panel 93, Male, Environment 1, Fly 7, Library 1


title: Run291_L7_A11_single cell

value: a dual reporter line expressing DsRed under regulatory elements of the cholecystokinin promoter and GFP knocked into the glutamate decarboxylase 67 gene

value: postnatal days 17-23


title: DGRP352 M_E1_5_L1

description:
EP1A101
Drosophila Genetic Reference Panel 352, Male, Environment 1, Fly 5, Library 1

value: Adult


title: Run291_L7_A12_single cell

value: cortical neuron


title: 2-5A 9min

description: processed data file: Dataset 5.txt

value: T47D cells semi-permeabilized with digitonin +2-5A for 9 minutes

value: EXPERIMENT 5


title: 2-5A 3min

description: processed data file: Dataset 5.txt

value: T47D cells semi-permeabilized with digitonin +2-5A for 3 minutes


title: DGRP229 M_E1_5_L1

description:
EP1A071
Drosophila Genetic Reference Panel 229, Male, Environment 1, Fly 5, Library 1

value: Male

value: Whole body


title: DGRP850 F_E1_5_L1

description:
EP1A081
Drosophila Genetic Reference Panel 850, Female, Environment 1, Fly 5, Library 1

value: Female


title: DGRP370 F_E1_4_L1

description:
EP1A061
Drosophila Genetic Reference Panel 370, Female, Environment 1, Fly 4, Library 1


title: WT

description: processed data file: Dataset 7.txt

value: epithelium

value: HAP1 cells, Ribo-Zero sequencing


title: Run291_L7_A10_single cell


title: 2-5A 1min

description: processed data file: Dataset 5.txt

value: T47D cells semi-permeabilized with digitonin +2-5A for 1 minute


title: Hap1KO-2naive

description: processed data file: Dataset 6.txt

value: RNase L -/-

value: RNase L-KO HAP1 cells, poly-A sequencing


title: Control 9m

description: processed data file: Dataset 5.txt

source_name: T47D_Control 9m

value: wild type

value: T47D cells semi-permeabilized with digitonin for 9 minutes


title: Run291_L7_A09_single cell

value: a dual reporter line expressing DsRed under regulatory elements of the cholecystokinin promoter and GFP knocked into the glutamate decarboxylase 67 gene


title: Hap1KO-1naive

description: processed data file: Dataset 6.txt

value: EXPERIMENT 6


title: Run291_L7_A06_single cell

value: postnatal days 17-23


title: DGRP804 F_E1_2_L1

description:
EP1A031
Drosophila Genetic Reference Panel 804, Female, Environment 1, Fly 2, Library 1

value: Adult


title: Control 1m

description: processed data file: Dataset 5.txt

source_name: T47D_Control 1m

value: T47D cells semi-permeabilized with digitonin for 1 minute


title: Control 3m

description: processed data file: Dataset 5.txt

source_name: T47D_Control 3m

value: T47D cells semi-permeabilized with digitonin for 3 minutes

value: EXPERIMENT 5


title: Run291_L7_A04_single cell

value: cortical neuron


title: DGRP804 F_E1_8_L1

description:
EP1A011
Drosophila Genetic Reference Panel 804, Female, Environment 1, Fly 8, Library 1

value: Whole body


title: Scr-IC2

description: processed data file: Dataset 4.txt

value: HeLa

value: 36h scrambled RNA control

value: 36h scrambled RNA control followed by poly-IC treatment for 6h

value: EXPERIMENT 4


title: Scr-IC1

description: processed data file: Dataset 4.txt

value: epithelium


title: Run283_L4_F03_single cell


title: RNAi-IC2

description: processed data file: Dataset 4.txt

value: 36h RNAi for RNase L

value: 36h RNAi for RNase L followed by poly-IC treatment for 6h


title: Run283_L4_E05_single cell


title: Scr1

description: processed data file: Dataset 4.txt


title: Run291_L7_A02_single cell


title: RNAi-IC1

description: processed data file: Dataset 4.txt


title: RNAi-2

description: processed data file: Dataset 4.txt


title: DGRP804 M_E1_7_L1

description:
EP1A041
Drosophila Genetic Reference Panel 804, Male, Environment 1, Fly 7, Library 1

value: Male


title: Run283_L4_D03_single cell

value: a dual reporter line expressing DsRed under regulatory elements of the cholecystokinin promoter and GFP knocked into the glutamate decarboxylase 67 gene


title: RNAi-1

description: processed data file: Dataset 4.txt


title: DGRP804 F_E1_5_L1

description:
EP1A021
Drosophila Genetic Reference Panel 804, Female, Environment 1, Fly 5, Library 1

value: Female


title: Run283_L4_E03_single cell

value: postnatal days 17-23


title: Scr2

description: processed data file: Dataset 4.txt


title: H672_overexpression_(rep 2)

description: processed data file: Dataset 3.txt

value: 24h Overexpression of WT human RNase L

value: 24h Overexpression of H672N RNase-inactive human RNase L

value: EXPERIMENT 3


title: Run283_L4_D02_single cell


title: WT_overexpression_(rep 2)

description: processed data file: Dataset 3.txt


title: H672N_overexpression_(rep 1)

description: processed data file: Dataset 3.txt


title: Run291_L7_A03_single cell


title: Run283_L4_C03_single cell


title: Run283_L4_C01_single cell


title: WT_overexpression_(rep 1)

description: processed data file: Dataset 3.txt


title: Run291_L7_A01_single cell


title: G55_WT_Th17_RIPseqInput

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 Tophat2
Transcripts FPKM were calculated by Cufflink using Reference Annotation: mm9.gtf from UCSC
Transcripts FPKM were calculated by Cufflink using Reference Annotation: NONCODEv4_mouse_lncRNA.gtf from NONCODE
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample ...

extract_protocol:
Total RNAs and IP enriched RNAs from cultured Th17 cells were harvested using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: WT

value: T cell

value: in vitro polarized T helper 17 cells

tag: molecule subtype

value: total RNA


title: Run283_L4_G04_single cell

value: cortical neuron


title: Run283_L4_A03_single cell


title: G59_WT_Th17_RIPseqGFPL10

tag: rip antibody

value: rabbit anti-GFP

tag: rip antibody vendor

value: Life Technology

tag: rip antibody cat. #

tag: rip antibody lot#

value: total RNA post GFPL10 IP


title: Recomb-+RNL+2-5A_30sec

description: processed data file: Dataset 2.txt

value: HeLa

value: epithelium

tag: genotype/variation

value: wild type

value: HeLa S10 extract treated with recombinant human RNase L + 2-5A for 5 seconds

value: EXPERIMENT 2


title: Recomb-+RNL+2-5A_5sec

description: processed data file: Dataset 2.txt


title: Recomb-Control

description: processed data file: Dataset 2.txt

value: Control HeLa RNA from S10 extracts


title: G19_WT_Th17_RIPseqDDX5

tag: strain background

tag: age

value: goat anti-DDX5

value: Abcam

value: total RNA post DDX5 IP


title: 2-5A_2h

description: processed data file: Dataset 1.txt

value: HeLa S10 extract treated with 2-5A for 1 hr

value: EXPERIMENT 1


title: Control_1h

description: processed data file: Dataset 1.txt

value: Control HeLa RNA from S10 extracts exposed to room temperature for 1 hr


title: DGRP563 M_E1_7_L1

description:
EP1B011
Drosophila Genetic Reference Panel 563, Male, Environment 1, Fly 7, Library 1

tag: strain

tag: developmental stage

value: Adult

value: Male

value: Whole body


title: Untreated_0m

description: processed data file: Dataset 1.txt


title: G23_WT_Th17_RIPseqRORgt

value: WT

tag: tissue

value: T cell

value: in vitro polarized T helper 17 cells

value: rat anti-RORgt

value: eBioscience

value: total RNA post RORgt IP


title: ASO_3

city: Ann Arbor

first: Richard

middle: C

last: McEachin

state: United States

address: 2800 Plymouth Road

department: DCM&B

institute: University of Michigan

data_processing:
Base Calling: Illumina HiSeq
Quality Control: FastQC
Alignment: Tophat2.0.3,  --b2-very-sensitive
Quality Control: FastQC
Abundance Measurement: CuffLinks/CuffDiff2.0.2, UCSC mm10.gtf transcriptome, with -u, -N, --compatible-hits-norm, and -b (relative to the UCSC mm10.fa)
Format Output: Format cufflinks output using a locally derived Perl script
Genome_build: mm10
Supplementary_files_format_and_content: FPKM

source_name: muscle

extract_protocol:
TruSeq SBS Kit v3, TruSeq PE Cluster Kit v3 - cBot - HS, TruSeq Multiplex Sequencing Primer Box
Trizol

tag: genotype

tag: treatment

value: ASO - antisense oligonucleotide targeting of the androgen receptor

value: muscle


title: KRKR_1

value: KRKR - knockin mutation into exon 1 of the AR as amino acids 385 and 518


title: KRKR_2


title: nfUV3b

first: Mats

last: Ljungman

state: Michigan

address: NCRC, B520 Room 1346 2800 Plymouth Rd.

description: molecule: bru-labeled RNA

data_processing:
Library strategy: Bru-Seq
Illumina Casava v1.8.2 software used for basecalling.
Reads were mapped to the Human ribosomal DNA complete repeating unit (U13369.1) using Bowtie v.0.12.8 and parameters -n 3 -k 1 -m 1
Reads that did not map to the ribosomal DNA were mapped to the hg19 genome using TopHat v1.4.1 and parameters –min-isoform-fraction 0, --max-multihits 1, --no-closure-search, --no-coverage-search, --bowtie-n, --initial-read-mismatches 3
A coverage determination was then made for every base in the genome using Bedtools v.2.16.2 such that a base covered by one read was recorded as having a coverage of 1/read_length
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads 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: hg19
Supplementary_files_format_and_content: The columns of the abundance measurement files indicate: (1) chromosome; (2) start; (3) end; (4) name; (5) score; (6) strand; (7) gene length; (8) feature type – exon, intron, etc; (9) number of introns and exons; (10) feautre length; (11) hits count; (12) density – count/length; (13) RPKM - count/(length/1E3)/(mapped read count/1E6)

source_name: Foreskin fibroblasts (HF1)

growth_protocol: HF1 cells were grown as monolayers in MEM supplied with 10% fetal bovine serum and antibiotics (Invitrogen).  K562 cells were grown in suspension in IMDM with 10% FBS. All cells were maintained at 37 C in a humidified 5% CO2 atmosphere.

extract_protocol:
Total RNA was isolated by using TRIzol reagent (Invitrogen), and Bru-labeled RNA was isolated from the total RNA by incubation with anti-BrdU antibodies (BD Biosciences) conjugated to magnetic beads (Dynabeads, Goat anti-Mouse IgG; Invitrogen) under gentle agitation at room temperature for 1 h. For more detail, see Paulsen et. al 2013
Bru-labeled RNA was mixed with first strand buffer and random primers and fragmented by heating at 85 ̊C for 10 minutes. The first strand cDNA was then  synthesized, in the presence of Actinomycin D to result in strand specific reads (only applicable to certain samples as indicated in sample table above). After purifying the first strand cDNA using AMPure RNAclean beads (Beckman Coulter), the second strand cDNA was synthesized. The resulting cDNA was purified with AMPure XP beads, after which the Illumina TruSeq RNA Sample Prep Kit was used to repair the cDNA ends, adenylate and ligate adaptors to the cDNA. The samples were then run on a 3% agarose gel and size-selected by excising gel slices in the 300bp region. These gel slices were purified using the QIAEX II Gel Extraction Kit (Qiagen) and then the Illumina TruSeq Kit PCR reagents were used to enrich the DNA fragments. After a final purification using AMPure XP beads, the quality and concentration of the DNA libraries were determined using an Agilent Bioanalyzer.

treatment_protocol: Cells were washed in PBS and irradiated in 100 ul PBS on 100 mm plates with a UVC lamp producing 1 J/m2/s.  Cells were then incubated in conditioned media containing 2 mM bromouridine (BrU) at 37C for a 30 min.

tag: uv exposure

tag: time between uv exposure and bru labeling

value: 0 hours

tag: bru labeling time

value: 0.5 hours

tag: time between bru labeling and rna extraction

tag: time between uv exposure and rna extraction

tag: labeling agent

value: Bromouridine (Bru)


title: KRKR_3


title: ASO_1


title: tnfpreUV2

description: molecule: bru-labeled RNA


title: nfUV3a

description: molecule: bru-labeled RNA


title: MUT_3

value: SUMO MUTANT - SUMOylation of the AR blocked


title: ASO_2


title: nfUV4

description: molecule: bru-labeled RNA


title: MUT_2


title: HF1 nf0h4

description: molecule: bru-labeled RNA

value: No exposure


title: K562UV1001

description: molecule: bru-labeled RNA


title: WT_1


title: WT_2


title: MUT_1


title: K562UV251

description: molecule: bru-labeled RNA


title: K5626h1

description: molecule: bru-labeled RNA

value: 6 hours


title: K5620h1

description: molecule: bru-labeled RNA


title: WT_3


title: PTL_3

status: Public on Dec 20 2015

city: Zhenjiang

first: youwen

last: tan

address: Danjianmen 300

country: China

institute: the third hospital of zhenjiang affiliated Jiangsu University

data_processing:
The raw sequences were processed using the Illumina pipeline program. After masking of adaptor sequences and removal of contaminated reads, the clean reads were filtered for miRNA prediction with the software package ACGT101-miR-v4.2 (LC Sciences, Houston, Texas, USA) and subsequently analyzed according to (http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3248653/). Secondary structure prediction of individual miRNAs was performed by Mfold software (Version 2.38; http://mfold.rna.albany.edu/?q=mfold/RNA-Folding-Form) using the default folding conditions.
Genome_build: GRCh37.p13

source_name: serum

extract_protocol: The supernatant serum was stored at −80°C until analysis. Total RNA was isolated using LCS TRK1001 miRNeasy kit (LC Sciences, Hangzhou, China). The libraries were constructed from total RNA using the Illumina® Truseq Small RNA Sample Preparation Kit (Illumina, San Diego, CA, USA) according to the manufacturer’s protocol. The quantity and purity of total RNAs were monitored using a NanoDrop® ND-1000 spectrophotometer. The integrity of total RNAs was analyzed using an Agilent 2100 Bioanalyzer system and RNA 6000 Nano LabChip Kit.

value: venousblood

tag: patient

value: cirrhosis patients


title: PTL_5

value: HCC patients


title: PTL_2

value: healthy controls


title: LacI tile 7 after negative selection

first: Alexander

last: Garruss

state: MA

address: 77 Avenue Louis Pasteur, Rm 238

department: Genetics

institute: Harvard Medical School

data_processing:
Library strategy: DNA-seq
Basecalls performed using CASAVA version 1.8.2
Paired sequencing reads were collapsed and filtered for sequencing errors using FLASH v. 1.2.11 with a maximum overlap of 300
Collapsed reads were then translated in three frames, aligned to wild-type protein LacI with BLAT v.35 using default settings, and the best translated alignment by percentage match was retained. Protein sequences were then trimmed according to the amplicon/tile number and cloning flanking sequence.
Sequences with mis-matches in the fixed flanking sequences or different than the expected length (containing insertions or deletions) were discarded. Only sequences harboring a single amino-acid change found in either the pre- or post-selection were retained for further analysis. Protein sequences were then counted for pre- and post-selection, respectively, for each amplicon. A psuedo-count of one was added. Counts between pre- and post-selection were then quantile normalized using the normalize.quantiles function from preprocessCore in R v.3.1.1. Log2 fold-changes were computed from the ratio of pre-selection divided by post-selection quantile normalized counts.
genome build: available on series record in file lacI_reference_DNA.txt
Supplementary_files_format_and_content: Depletion values for amino-acid and position pairs; tabular format; log2(pre/post negative selection)

molecule: other

organism: Escherichia coli

source_name: plasmid

growth_protocol: For library transformations, the screening strain was made electrocompetent by harvesting early log-phase cells (10 mL per transformation at OD600 nm = 0.15–0.25), removing salt through two washes with ice cold 10% glycerol, and resuspended in 50 μL cold 10% glycerol. Ten ng of library plasmid were electroporated into the competent cells, which were recovered for 1 hr in 1 mL SOC medium. To estimate the number of transformants, 1 μL recovered cells were plated on selective LB spectinomycin medium, and the remainder of the recovered cells were added to 10 mL of LB spectinomycin medium and selected overnight. Screening strain cells expressing LacI variants that do not bind to operator DNA constitutively express tolC and gfp; these were eliminated through negative selection by overnight selection with colicin E1 protein. We added five μL of saturated library transformation culture to 150 μL LB spectinomycin medium supplemented with tenfold serial dilutions of purified colicin E1 protein (2.73 mg/mL), in the range from 1:100 to 1:1,000,000; a control population of the same library was grown overnight without colicin E1. Enrichment of DNA-bound lacI variants was verified by flow cytometry the next day by measuring the fraction of GFP+ cells in the colicin E1 incubated and control populations. A 1:100,000 dilution of colicin E1 (20.7 ng/mL) was generally found to be optimal.

extract_protocol:
Plasmids were prepped from each library using a Spin Miniprep Kit (Qiagen #27104). A nested amplicon PCR strategy was used to append barcodes and sequencing adapters. In a first round of PCR, amplicon primers were used to amplify 300 bp regions of the lacI gene and add standard Illumina MiSeq sequencing primer binding sites. The first six bases to be read in sequencing read 1 were degenerate to increase complexity and aid in base calling. In a second round of PCR, barcoding primers were used to append barcodes and flow cell adapter sequences.
Amplicons were purified, quantified using Kapa Biosystems Library Quanitification kit (part number KK4824), and normalized. Libraries were sequenced on the Illumina MiSeq using MiSeq Reagent Kit v3, 600 cycle (part number MS-102-3003) according to the manufacturer’s protocol.

treatment_protocol: The lacI gene variant libraries were constructed by cloning oligonucleotides encoding the desired mutations into plasmid pSC101_lacI_specR amplified by PCR with primers (Supplementary Table 6; Taylor, et al. Nature Methods. 2015) to appropriately linearize, add BsaI recognition sequence (5’-GGTCTCN), and remove the wild type lacI coding sequence segment to be replaced.Constructing the single residue replacement library involved replacing lacI codons from 3–359 with one missense codon encoding each of the remaining 19 natural amino acids (6,802 sequences). Oligos encoding these mutations were synthesized on a B3 Synthesizer (CustomArray, Inc., Bothell, WA), and were organized into the following tiles spanning the lacI gene: (#1)3–38, (#2)39–74, (#3)75–110, (#4)111–146, (#5)147–182, (#6)183–218, (#7)219–254, (#8)255–290, (#9)291–326, (#10)327–359.

value: K12 MG1655 EcNR2

tag: sequenced molecule

value: plasmid DNA


title: LacI tile 6 after negative selection


title: LacI tile 8 after negative selection


title: LacI tile 9 after negative selection


title: LacI tile 10 after negative selection


title: LacI tile 3 after negative selection


title: LacI tile 5 after negative selection


title: LacI tile 9 no selection


title: LacI tile 1 after negative selection


title: LacI tile 8 no selection


title: LacI tile 7 no selection


title: LacI tile 6 no selection


title: LacI tile 5 no selection


title: LacI tile 3 no selection


title: LacI tile 4 after negative selection


title: LacI tile 1 no selection


title: LacI tile 2 no selection


title: LacI tile 4 no selection


title: LacI tile 10 no selection


title: Zscan10_KO_rep2

city: Kobe

first: Hitoshi

last: Niwa

state: Hyogo

address: 2-2-3 Minatojima-minamimachi,Chuo-ku

department: Center for Developmental Biology

country: Japan

institute: RIKEN

description:
Gene expression in subcloned Zscan10-null ES cells
rRNA depleted RNA

data_processing:
Sequenced reads were aligned to the mouse genome (mm10) using TopHat after trimming of adaptor sequences with trim_galore and quality check with FastQC.
Gene expression level measured as FPKM was determined with Cuffdiff.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text file includes FPKM values for each Sample

source_name: Embryonic stem cells, Zscan10 KO, replicate2

extract_protocol:
Total RNAs were prepared by QuickGene RNA cultured cell HC kit (KURABO) following the manufacturer's instruction and treated with Ribo-Zero Gold Sample Prep Kit (Illumina) for deplete ribosomal RNA.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: cell type

value: ESC


title: Zscan10_KO_rep1

source_name: Embryonic stem cells, Zscan10 KO, replicate1

tag: genotype/variation


title: LacI tile 2 after negative selection


title: Ogg1_Mutyh_DKO_HC

city: Trondheim

first: Katja

last: Scheffler

address: Edvard Griegs Gate 8

department: Department of Neuromedicine and Movement Sciences

country: Norway

institute: Norwegian University of Science and Technology

data_processing:
Libray preparison:mRNA is enriched by using the oligo(dT) magnetic beads, following to be fragmented into short fragments (about 200 bp). Then the first strand of cDNA is synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End reparation and 3'-end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The fragments are enriched by PCR amplification.
Sequencing: The library products are ready for sequencing via Illumina HiSeqTM 2000.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence. The clean reads were mapped to mouse reference sequences (Li R., Yu C., Li Y., et al . 2009, Bioinformatics).
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include RPKM values for each sample

source_name: hippocampus

extract_protocol:
Total RNA was isolated from mouse hippocampus using RNeasy kit (Qiagen) according to the manufacturer’s instructions. RNA quality and concentration were measured using a RNA Pico chip on an Agilent Bioanalyzer.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Ogg-/- Mutyh-/-

value: brain

value: 6 month


title: Mutyh_KO_HC

value: Mutyh-/-


title: ANME-1 Mcr-producing M. acetivorans grown on methanol, Replicate 3

city: State College

first: Thomas

middle: K.

last: Wood

state: PA

address: Room 161 Fenske Laboratory, University Park

department: Chemical Engineering & Biochemistry and Molecular Biology

institute: Pennsylvania State University

description:
total RNA (with rRNA partially removed)
TMML3

data_processing:
Reads were mapped to the reference genome of M. acetivorans (Genbank accession NC_003552.1) using STAR.
The mapped reads were assembled using Cufflink v2.2.1 to identify potential novel transcripts. Assembled, unannotated novel transcripts for all the strains were combined with the list of known genes.
Differential expression of genes and potential novel transcripts were determined using Cuffdiff at a significance cutoff at q < 0.07 with a false discovery rate of 0.05.
Expression changes are determined by the ratio of FPKM of culture replicates grown on methane to FPKM of culture replicates grown on methanol.
Genome_build: NC_003552.1
Supplementary_files_format_and_content: Read count of each replicate and the average FPKM value (adjusted to sequencing depth) for each mapped locus.

organism: Methanosarcina acetivorans

growth_protocol: All starter cultures of M. acetivorans with pES1-MATmcr3 (200 mL) were grown on methanol for 5 days, and harvested by centrifugation. Cell pellets were washed three times with HS medium, and resuspended using 5 mL HS medium, 2 µg/mL puromycin, and 0.1 mM FeCl3. For growth on methane, methane was filled into the headspace of the cultures. For growth on methanol, 150 mM methanol was added. All cultures were incubated at 37C for 5 days.

extract_protocol:
Rapid centrifugation in the presence of 50 µL RNAlater solution (Ambion, Austin, TX) per mL of culture, followed by flash-freezing of cell pellets. Total RNA was isolated using RNeasy Mini kit (Qiagen, Valencia, CA), and were further digested with terminator 5’-phosphate-dependent exonuclease (Epicentre, Madison, WI) to partially remove ribosomal RNA. Digested RNA were cleaned up using AgenCourt RNAClean XP beads (AgenCourt Bioscience, Beverly, MA).
RNA libraries were prepared for sequencing using standard Illumina protocols (TruSeq Stranded mRNA Library kit).

value: containing pES1-MATmcr3; ANME-1 Mcr-producing

tag: components of growth medium

value: HS medium, 2 µg/mL puromycin, and methanol

tag: gas in headspace


title: ANME-1 Mcr-producing M. acetivorans grown on methanol, Replicate 2

description:
total RNA (with rRNA partially removed)
TMML2


title: Ogg1_KO_HC

tag: strain

tag: age