title: cdk19_kd_5fuRep2

city: Boulder

first: Dylan

last: Taatjes

state: Colorado

address: 3415 Colorado Ave.

department: Molecular and Cell Biology

institute: University of Colorado, Boulder

data_processing:
Fastq quality control via Fastqc ; Command = fastqc <.fastq>
Fastq adapter trimming via Trimmomatic; Command = java -jar trimmomatic-0.32.jar SE -threads 32 -phred33 -trimlog ILLUMINACLIP:TruSeq3-SE:2:30:10 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:50
Alignment to reference transcriptome via Tophat2 version 2.0.6; Command = tophat2 -p 32 --b2-very-sensitive --microexon-search --no-convert-bam --library-type fr-firststrand -G hg19.refGene.gtf
Gene counts quantified via Htseq count version 0.6.1; Command = htseq-count -t exon -i gene_id -r name -f bam -m intersection-nonempty --stranded=reverse
Differential gene expression and exploration via standard Deseq2 pipeline
Genome_build: hg19
Supplementary_files_format_and_content: Gene counts file. Tab delimited. Rows = gene Counts from htseq. Columns = samples

source_name: SJSA - osteosarcoma

growth_protocol: SJSA cells were grown in 15cm dishes to approximately 80% confluency in RPMI supplemented with 10% FBS at 37 degrees C with 5% CO2.

extract_protocol:
Cells were washed in cold PBS, then scraped in TRIzol RNA purification reagent, which was added directly to each plate according to the manufacterers' instructions. Subsequently, RNA was further purified with a Qiagen Rneasy kit using an on-column DNAse I digestion.
500ng per sample total RNA was used (measured with a NanoDrop 1000 and Agilent 2100 Bioanalyzer) to construct libraries following the Illumina TruSeq Stranded RNA Sample Preparation Guide

treatment_protocol: 5-FU in 100% DMSO (or vehicle; 100% DMSO only)  was added at a final concentration of 375uM directly to the media for each replicate. Two biological replicates were performed for both 5-FU and DMSO treatment.

value: SJSA - osteosarcoma


title: cdk19_kd_5fuRep1

status: Public on May 09 2017

type: SRA

country: USA

library_source: transcriptomic

molecule: polyA RNA

organism: Homo sapiens

tag: tissue


title: siRNA_cid11Δ

city: Munich

first: Mario

last: Halic

department: Biochemistry - Gene Center

country: Germany

institute: Ludwig-Maximilians-Universität

data_processing:
base calling with RTA1.18.64
Sequenced reads were trimmed for the adaptor sequence, then mapped to the S.pombe genome with Novoalign (http://www.novocraft.com) randomly assigned.
Reads mapping with two or fewer mismatches were retained.
Enrichment was calculated with our house Perl scripts
Normalization to reads per million for siRNA and ChIP seq, normalization to reads mapping to protein coding genes for RNAseq
Genome_build: Schizosaccharomyces_pombe.ASM294v1.18
Supplementary_files_format_and_content: can be viewed with IGV browser: http://www.broad.mit.edu/igv

molecule: total RNA

organism: Schizosaccharomyces pombe

growth_protocol: cells were grown in liquid, rich YES medium or EMM -Leucine

extract_protocol:
Endogenous 3xFLAG-tagged Ago1-bound siRNA were purified by protein affinity purification and subsequent size selection on an 18% acrylamide gel. 2 pmol of a preadenylated 3' adaptor oligonucleotide (miRNA Cloning Linker-1 from IDT, 5'-App CTG TAG GCA CCA TCA AT/ddC/-3') were ligated in a 10 µl reaction with 5 U T4 RNA ligase (TaKaRa), ligation buffer without ATP and 5 U RNasin (Promega) at 20°C for 2 hours. The 3' ligated products were purified on an 18% acrylamide urea gel with subsequent phenol-chloroform purification and ethanol purification. The 5' adaptor ligation was performed in a 10 µl reaction with 2 pmol 5' adaptor oligonucleotide (5'-GUU CAG AGU UCU ACA GUC CGA CGA UC-3'), 5 U RNasin (Promega), 0.06 µg BSA, 5 U T4 RNA ligase (Thermo Scientific) and 1x ligation buffer with ATP (Thermo Scientific) for 2 h at 20°C. The ligated products were gel purified and reverse transcribed with 10 pmol primer (RT primer: 5'- GTG ACT GGA GTT CAG ACG TGT GCT CTT CCG ATC GAT TGA TGG TGC CTA CAG-3') and the SuperScript III First Strand Synthesis System (Thermo Scientific). The cDNA was PCR-amplified with Q5 High-Fidelity 2x Master Mix (NEB) for 14-20 cycles using the Illumina P5 5' primer (5' -AAT GAT ACG GCG ACC ACC GAG ATC TAC ACT CTT TCC CTA CAC GAC G -3') and the Illumina P7 3' primer with inserted barcode (5'-CAA GCA GAA GAC GGC ATA CGA GAT XXXXXX GTG ACT GGA GTT CAG ACG TG -3').
RNA libraries were prepared with NEBnext Ultra Directional RNA Library Prep Kit for Illumina (NEB); DNA library construction with NEBNext Ultra II DNA Library Prep Kit for Illumina kit (NEB). siRNA library: ligation of 3’ adapter (5'-App CTG TAG GCA CCA TCA AT/ddC/-3') and 5’ adapter (5'-GUU CAG AGU UCU ACA GUC CGA CGA UC-3'), reverse transcription, PCR of cDNA with Illumina P5 5' primer and barcoded llumina P7 3' primer.

tag: genotype/variation

tag: antibody for ip

value: anti-FLAG (Sigma-Aldrich, catalog#A2220, lot# SLBQ4019V)

tag: strain number

tag: target molecule

value: siRNA


title: 45_Rep1_3357_2dpi_ear3_2011.fastq

city: Carbondale

first: Bryan

middle: Manyasi

last: Musungu

state: IL

address: 1125 Lincoln Dr.

department: Plant Biology

institute: Southern Illinois University Carbondale

description: Sample 17

data_processing:
Illumina Casava1.7 software used for basecalling.
Default CLC mapping settings, trimming and fastqc quality control of the reads, for both Z. mays and A. flavus, were done with CLC workbench 4.9 (Qiagen, Hilden, Germany).
RPKM and count values were then generated from CLC.
Genome_build: (AGPv3, INSDC Assembly GCA_000005005.5, Apr 2013) and A. flavus (JCVI-afl1-v2.0, INSDC Assembly GCA_000006275.1, Jan 2009).
Supplementary_files_format_and_content: RPKM/Count values for each Sample.

organism:
Zea mays
Aspergillus flavus

source_name: Zea mays kernel_48 hpi

growth_protocol: Maize B73 was grown in Clayton, NC during the years 2011 and 2013.  Seeds were planted on April 16.  Ears were hand pollinated from July 5-8 and covered with a paper bag.

extract_protocol:
Eight kernels per ear were ground and used for RNA isolation. Approximately one hundred milligrams of ground tissue was homogenized in a Virtis homogenizer (Virtis, Gardiner, NY) in Saturated Phenol, pH 6.6 for 2 min. Samples were then dissolved in Tris EDTA buffer, pH 8.0 (ACROS Organics, New Jersey, US), purified with 5:1 Acid Phenol: Chloroform, pH 4.5, and precipitated with ice-cold 100% ethanol overnight. Total RNA was purified again with RNeasy Mini Kit (Qiagen, Hilden, Germany) according to the manufacturer’s instructions. The quality and concentration of RNA was analyzed using an RNA Pico chip on an Agilent Bioanalyzer (Agilent, Santa Clara, CA).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: A  time course study was performed by pin bar inoculating one ear (per time point) of maize B73 with A. flavus NR3357 and harvesting at 0, 6, 12, 18, 24, 30, 36, 42, 48 and 72 hours post inoculation.

tag: maize development stage

value: Milk

tag: maize cultivar

tag: aspergillus strain

tag: time point

value: 48 hrs post inoculation (hpi)

value: kernel


title: 44_Rep1_3357_2dpi_ear2_2011.fastq

description: Sample 16


title: siRNA_cid16Δcid12Δ


title: siRNA_cid1Δ


title: siRNA_cid13Δ


title: ∆yenR rep3

city: Salt Lake City

first: Shin

last: Enomoto

state: Utah

address: 201 South Biology, 257 S 1400 E

department: Biology

institute: University of Utah

description: Grown in 10µg/ml OHHL

data_processing:
50-base single reads were filtered using NGSQCToolkit.
Mapped to reference genome using bowtie2 employing the --very-sensitive preset.
Read counts were generated with HTSeq using htseq-count with options -s no -a 0 -m intersection-nonempty -t gene.
Counts from the replicates were combined and normalized using a custom Perl script.
Genome_build: Sodalis praecaptivus strain:HS1 (Accession: PRJNA199998)
Supplementary_files_format_and_content: tab-delimited text file.

organism: Sodalis praecaptivus

source_name: Liquid culture

growth_protocol: Bacteria were grown in LB broth at 30°C with aeration to mid-log phase (OD 600 ~0.4), in the presence or absence of 10µg/ml of N-(3-oxohexanoyl) homoserine lactone(OHHL: Sigma Aldrich, K3255)

extract_protocol:
RNA was prepared by triton X-100 boiling and chloroform extraction method, followed by purification on Purelink RNA minikit (Ambion)
Illumina TruSeq RNA sample prep v2 with oligo dT selection

tag: genotype

value: delta-ypeR

tag: strain


title: cdk19_kd_dmsoRep2


title: 43_Rep1_3357_2dpi_ear1_2011.fastq

description: Sample 15


title: cdk19_kd_dmsoRep1


title: 56_Rep1_3357_3dpi_ear2_2011.fastq

description: Sample 20

source_name: Zea mays kernel_72 hpi

value: 72 hrs post inoculation (hpi)


title: timepoint42 RNA-seq.fastq

description: Sample 14

source_name: Zea mays kernel_42 hpi

value: 42 hrs post inoculation (hpi)


title: ctrl_5fu1Rep2


title: siRNA_cid12Δ


title: siRNA_cid16Δ_b


title: ∆yenR rep2

description: Grown in 10µg/ml OHHL


title: ctrl_5fu1Rep1


title: timepoint36 RNA-seq.fastq

description: Sample 13

source_name: Zea mays kernel_36 hpi

value: 36 hrs post inoculation (hpi)


title: timepoint30 RNA-seq.fastq

description: Sample 12

source_name: Zea mays kernel_30 hpi

value: 30 hrs post inoculation (hpi)


title: ∆yenR rep1

description: Grown in 10µg/ml OHHL


title: ∆ypeR rep3

description: Grown in 10µg/ml OHHL


title: timepoint24 RNA-seq.fastq

description: Sample 11

source_name: Zea mays kernel_24 hpi

value: 24 hrs post inoculation (hpi)


title: siRNA_cid14Δ_b

source_name: cid14Δ _b


title: ∆ypeR rep2

description: Grown in 10µg/ml OHHL


title: 33_Rep1_3357_1dpi_ear3_2011.fastq

description: Sample 10


title: siRNA_cid14Δ


title: 32_Rep1_3357_1dpi_ear2_2011.fastq

description: Sample 9


title: siRNA_wildtype_b

source_name: wild type_b

value: wild type


title: 31_Rep1_3357_1dpi_ear1_2011.fastq

description: Sample 8


title: siRNA_wildtype

source_name: wild type


title: Passage 2 replicate 3

city: New York

first: Peter

last: Palese

state: New York

address: 1 Gustave L. Levy Place, Box 1124

department: Department of Microbiology

institute: Icahn School of Medicine at Mount Sinai

data_processing:
Illumina CasavaX.X software used for basecalling
A previously published script was used to filter and trim reads containing the transposon sequence marker (TGCGGCCGCA; contained in all insertion sites). Reads not containing the insertion sequence were placed in separate fastq file.
Reads with and without insertions were separately mapped to the ZIKV genome (Genbank accession number KX830960) using Bowtie v2. The reads from the input library (Sample 1) were mapped to the rescue plasmid (MR766_ Intron_3127.txt), which contains an intron after nucleotide position 3127 in  ZIKV genome for technical reasons.
Statistics and visualization was performed with samtools and bedtools.
The total number of reads without inserts at each nucleotide position in the genome was counted to determine sequencing coverage, while the reads with inserts at each position in the genome were
The number of reads with insertions at each site was normalized to the sequencing coverage.
Then the total number of reads with insertions was normalized between the different samples.
The reads at each site in the passaged virus were divided by the total number of reads with insertions in the input.
Genome_build: KX830960
Supplementary_files_format_and_content: The normalized transposon insertion frequency at each position of the ZIKV genome is provided. For the Sample 1: Input, the values are the normalized abundance of reads with inserts at each genomic position. For the rest of the samples, the fold over input value is provided at each genomic position.

organism: Zika virus

source_name: African green monkey kidney cells (Vero)

extract_protocol:
RNA from the transfected and infected cells (Passage 1 and 2) were extracted from the cells with TRIzol Plus RNA purification kit (Thermo Fisher Scientific). The RNA was subjected to RT-PCR with SuperScript III RT-PCR Platinum Taq High Fidelity kit (Thermo Fisher Scientific) and ZIKV specific primers.
The cDNA and the plasmid input library were prepared with the NEB Next Ultra II kit and multiplex oligos (New England Biolabs) using the manufactures protocols.


title: Passage 2 replicate 2


title: Passage 1 replicate 3


title: H3K9me2_ChIP_cid14Δ + pREP1-Rdp1_2

library_source: genomic

molecule: genomic DNA

source_name: cid14Δ + pREP1-Rdp1_2

extract_protocol:
Immunoprecipitation of crosslinked (1% formaldehyde, 15 min) cells, decrosslinking, protein degradation and RNA degradation.
RNA libraries were prepared with NEBnext Ultra Directional RNA Library Prep Kit for Illumina (NEB); DNA library construction with NEBNext Ultra II DNA Library Prep Kit for Illumina kit (NEB). siRNA library: ligation of 3’ adapter (5'-App CTG TAG GCA CCA TCA AT/ddC/-3') and 5’ adapter (5'-GUU CAG AGU UCU ACA GUC CGA CGA UC-3'), reverse transcription, PCR of cDNA with Illumina P5 5' primer and barcoded llumina P7 3' primer.

value: cid14{delta} + pREP1-Rdp1_2

value: anti-H3K9me2 (Abcam, catalog# ab1220, lot# 1380686)

value: H3K9me2 bound DNA


title: 21_Rep1_3357_12hpi_ear3_2011.fastq

description: Sample 6

source_name: Zea mays kernel_12 hpi

value: 12 hrs post inoculation (hpi)


title: WT rep1

description: Grown in 10µg/ml OHHL

value: WT


title: timepoint18 RNA-Seq.fastq

description: Sample 7

source_name: Zea mays kernel_18 hpi

value: 18 hrs post inoculation (hpi)


title: WT rep2

description: Grown in 10µg/ml OHHL


title: 20_Rep1_3357_12hpi_ear2_2011.fastq

description: Sample 5


title: Passage 2 replicate 1


title: H3K9me2_ChIP_wildtype + pREP1

source_name: wild type + pREP1

value: wild type + pREP1


title: H3K9me2_ChIP_cid14Δ + pREP1

source_name: Cid14Δ + pREP1

value: Cid14{delta} + pREP1


title: ctrl_dmsoRep2

source_name: SJSA - osteosarcoma

value: SJSA - osteosarcoma


title: Passage 1 replicate 2


title: ctrl_dmsoRep1


title: Passage 1 replicate 1


title: Transfection replicate 3


title: 19_Rep1_3357_12hpi_ear1_2011.fastq

description: Sample 4

value: Milk

value: kernel


title: RNA_dcr1Δ

extract_protocol:
Total RNA was isolated applying the hot phenol method. The extracted nucleic acids were treated with DNAse I for 2 h at 37°C. DNAse was removed by a second phenol-chloroform-isoamylalcohol extraction and ethanol precipitation. 1 µg total RNA was degraded with Terminator nuclease (Epicentre) in buffer A at 30°C for 2 h.
RNA libraries were prepared with NEBnext Ultra Directional RNA Library Prep Kit for Illumina (NEB); DNA library construction with NEBNext Ultra II DNA Library Prep Kit for Illumina kit (NEB). siRNA library: ligation of 3’ adapter (5'-App CTG TAG GCA CCA TCA AT/ddC/-3') and 5’ adapter (5'-GUU CAG AGU UCU ACA GUC CGA CGA UC-3'), reverse transcription, PCR of cDNA with Illumina P5 5' primer and barcoded llumina P7 3' primer.

value: total RNA


title: ∆ypeR rep1

description: Grown in 10µg/ml OHHL

value: delta-ypeR


title: ∆ypeI rep1

value: delta-ypeI


title: timepoint12 RNA-seq.fastq

description: Sample 3


title: RNA_cid16Δ


title: Transfection replicate 2

data_processing:
Illumina MiSeq Control Software 2.6.2.1 software was used for sequencing.
A previously published script was used to filter and trim reads containing the transposon sequence marker (TGCGGCCGCA; contained in all insertion sites). Reads not containing the insertion sequence were placed in separate fastq file.
Reads with and without insertions were separately mapped to the ZIKV genome (Genbank accession number KX830960) using Bowtie v2. The reads from the input library (Sample 1) were mapped to the rescue plasmid (MR766_ Intron_3127.txt), which contains an intron after nucleotide position 3127 in  ZIKV genome for technical reasons.
Statistics and visualization was performed with samtools and bedtools.
The total number of reads without inserts at each nucleotide position in the genome was counted to determine sequencing coverage, while the reads with inserts at each position in the genome were
The number of reads with insertions at each site was normalized to the sequencing coverage.
Then the total number of reads with insertions was normalized between the different samples.
The reads at each site in the passaged virus were divided by the total number of reads with insertions in the input.
Genome_build: KX830960
Supplementary_files_format_and_content: The normalized transposon insertion frequency at each position of the ZIKV genome is provided. For the Sample 1: Input, the values are the normalized abundance of reads with inserts at each genomic position. For the rest of the samples, the fold over input value is provided at each genomic position.

extract_protocol:
RNA from the transfected and infected cells (Passage 1 and 2) was extracted from the cells with TRIzol Plus RNA purification kit (Thermo Fisher Scientific). The RNA was subjected to RT-PCR with SuperScript III RT-PCR Platinum Taq High Fidelity kit (Thermo Fisher Scientific) and ZIKV specific primers.
The cDNA and the plasmid input library were prepared with the NEB Next Ultra II kit and multiplex oligos (New England Biolabs) using the manufactures protocols.


title: WT rep3

description: Grown in 10µg/ml OHHL


title: ∆ypeI rep3


title: Transfection replicate 1

description:
DNA plasmid
KX830960


title: ∆ypeI+OHHL rep3

description: Grown in 10µg/ml OHHL


title: timepoint6 RNA-Seq.fastq

description: Sample 2

source_name: Zea mays kernel_6 hpi

value: 6 hrs post inoculation (hpi)


title: RNA_cid14Δ


title: ∆ypeI+OHHL rep2

description: Grown in 10µg/ml OHHL

source_name: Liquid culture


title: Plasmid input library

description:
DNA plasmid
KX830960

data_processing:
Illumina MiSeq Control Software 2.6.2.1 software used for sequencing.
A previously published script was used to filter and trim reads containing the transposon sequence marker (TGCGGCCGCA; contained in all insertion sites). Reads not containing the insertion sequence were placed in separate fastq file.
Reads with and without insertions were separately mapped to the ZIKV genome (Genbank accession number KX830960) using Bowtie v2. The reads from the input library (Sample 1) were mapped to the rescue plasmid (MR766_ Intron_3127.txt), which contains an intron after nucleotide position 3127 in  ZIKV genome for technical reasons.
Statistics and visualization was performed with samtools and bedtools.
The total number of reads without inserts at each nucleotide position in the genome was counted to determine sequencing coverage, while the reads with inserts at each position in the genome were
The number of reads with insertions at each site was normalized to the sequencing coverage.
Then the total number of reads with insertions was normalized between the different samples.
The reads at each site in the passaged virus were divided by the total number of reads with insertions in the input.
Genome_build: KX830960
Supplementary_files_format_and_content: The normalized transposon insertion frequency at each position of the ZIKV genome is provided. For the Sample 1: Input, the values are the normalized abundance of reads with inserts at each genomic position. For the rest of the samples, the fold over input value is provided at each genomic position.

molecule: other

source_name: E. Coli


title: RNA_wildtype


title: ∆ypeI+OHHL rep1

description: Grown in 10µg/ml OHHL


title: ∆ypeI rep2


title: unassigned reads from mock_2-8dpi

status: Public on May 08 2017

city: Norwich

first: Ghanasyam

last: Rallapalli

address: Norwich Research Park

country: United Kingdom

institute: The Sainsbury Laboratory

description: Mock 2dpi to 8dpi library samples were sequenced in one lane. nobarcode-sequences.mock_2-8DPI.fq is the file containing sequences unassigned to any barcode from Mock 2 to 8 dpi samples.

data_processing:
Library strategy: Tag-Seq
Illumina Casava1.7 software used for basecalling.
Illumina sequence library is quality filtered using FASTX Toolkit 0.0.13 with parameters -q20 and -p50 and reads containing “N” are discarded. Read qualities are converted from Illumina fastq to sanger fastq format
EXPRSS libraries are separated using perfect match to the barcode (at 5' end). Each sub-library is quality filtered (-q20 and -p50) and artefact filtered using FASTX-toolkit.
Quality-filtered libraries were aligned to the combined genome of Arabidopsis TAIR10 and Albugo strains. Therefore mock samples were analysed in pairwise manner with each Albugo species infection data, independently. Quality-filtered libraries of mock and Nc14 infected samples were aligned to the combined genomes of Arabidopsis TAIR10 and AlNc14; similarly mock and Nc2 infected samples were aligned to combined genomes of TAIR10 and AcNc2 using Bowtie version 0.12.8. Unaligned reads from previous step were mapped to the combined cDNA reference sequences of respective Arabidopsis and Albugo combinations using Novoalign v2.08.03 (http://www.novocraft.com/). Uniquely aligned read counts were selected for differential expression analysis. For gene expression analysis each Albugo (Nc14 or Nc2) infection time point data was compared against respective mock time point data resulted from pairwise analysis. Differential expression analysis was performed using the R statistical language version 2.11.1 with the Bioconductor package, edgeR version 1.6.15 with the exact negative binomial test using tagwise dispersions. Genes with false discovery rate (FDR) <0.01 were selected as differentially expressed.
Genome_build: Arabidopsis TAIR10, Albugo candida strain Nc2 version 01 and Albugo laibachii strain Nc14 version 1 (see supplementary files linked to Series record)
Supplementary_files_format_and_content: tab-delimited text files containing each genes tag counts for individual samples

organism: Arabidopsis thaliana

growth_protocol: Arabidopsis plants were grown at 22°C and 60% humidity under a 10-h photoperiod and a 14-h dark period in environmentally controlled growth cabinets.

extract_protocol:
Total RNA was extracted using the TRI reagent (Sigma) and 1-Bromo-3-chloropropane (Sigma), as per manufacturer’s guidelines. RNA was precipitated with half volume of isopropanol and half volume of high salt precipitation buffer (0.8 M sodium citrate and 1.2 M sodium chloride). RNA samples were treated with DNaseI (Roche) according to the manufacturer’s recommendation and phenol/chloroform extracted and ethanol precipitated.
[EXPRSS] Typically 3 µg of total RNA was used to generate first strand cDNA using a oligo(dT) primer comprising P7 sequence of Ilumina flowcell. Double strand cDNA is synthesized as described previously [Okayama & Berg, Molecular and cellular biology 1982, 2:161-170]. Purified cDNA is subjected to Covaris shearing to a target size of 200bp (parameters: Intensity – 5, Duty cycle – 20%, Cycles/Burst – 200, Duration – 60seconds). End repairing and A-tailing of sheared cDNA is carried out as described by Illumina. Y-shaped adapters are ligated to A-tailed DNA and subjected to size selection on 1X TAE agarose gel. The gel-extracted library is PCR enriched and quantified using qPCR with previously sequenced similar size range Illumina library

treatment_protocol: Four or five week old plants were inoculated with Albugo strains using 7.5 x 104 zoosporangia/ml. Mock plants were treated the same way but sprayed with cold water and leaves were harvested at 0, 2, 4, 6 and 8 days post-inoculation (dpi).

tag: line

value: multi-parent recombinant inbred line MAGIC-107

value: aerial shoots

tag: age

value: 4-5 weeks

tag: inoculation

tag: time

value: 2dpi to 8dpi


title: unassigned reads from albugo_8dpi

description: 8dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in one lane. nobarcode-sequences.albugo_8DPI.fq is the file containing sequences unassigned to any barcode from 8 dpi albugo samples.

value: AcNc02 and AlNc14


title: timepoint0 RNA-seq.fastq

description: Sample 1

source_name: Zea mays kernel_0 hpi

value: 0 hrs post inoculation (hpi)


title: unassigned reads from albugo_6dpi_lib1

description: 6dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_6dpi_lib1.fq is the file containing sequences unassigned to any barcode from 6 dpi albugo samples from lane1.


title: unassigned reads from albugo_6dpi_lib2

description: 6dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_6dpi_lib2.fq is the file containing sequences unassigned to any barcode from 6 dpi albugo samples from lane2.


title: unassigned reads from albugo_4dpi_lib2

description: 4dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_4dpi_lib2.fq is the file containing sequences unassigned to any barcode from 4 dpi albugo samples from lane2.


title: unassigned reads from albugo_4dpi_lib3

description: 4dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_4dpi_lib3.fq is the file containing sequences unassigned to any barcode from 4 dpi albugo samples from lane3.


title: unassigned reads from albugo_2dpi_lib1

description: 2dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_2dpi_lib1.fq is the file containing sequences unassigned to any barcode from 2 dpi albugo samples from lane1.


title: unassigned reads from albugo_2dpi_lib3

description: 2dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_2dpi_lib3.fq is the file containing sequences unassigned to any barcode from 2 dpi albugo samples from lane3.


title: unassigned reads from albugo_2dpi_lib2

description: 2dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_2dpi_lib2.fq is the file containing sequences unassigned to any barcode from 2 dpi albugo samples from lane2.


title: unassigned reads from albugo_4dpi_lib1

description: 4dpi Albguo infected library samples (AcNc02 and AlNc14) were sequenced in 3 independant lanes. nobarcode-sequences.albugo_4dpi_lib1.fq is the file containing sequences unassigned to any barcode from 4 dpi albugo samples from lane1.


title: unassigned reads from 0dpi

description: 0dpi library samples (AcNc02, AlNc14 and Mock) were sequenced in one lane. nobarcode-sequences_0DPI.fq is the file containing sequences unassigned to any barcode from 0 dpi samples.

value: AcNc02, AlNc14 and Mock


title: Mock_8dpi_biorep2

description:
Barcode sequence: ATCCT
Barcode position: 5' end of the read

source_name: aerial shoots


title: Mock_8dpi_biorep3

description:
Barcode sequence: CAGAGT
Barcode position: 5' end of the read


title: Mock_8dpi_biorep1

description:
Barcode sequence: GCAT
Barcode position: 5' end of the read


title: Mock_6dpi_biorep4

description:
Barcode sequence: AACAGAGT
Barcode position: 5' end of the read


title: Mock_6dpi_biorep3

description:
Barcode sequence: CGAGAT
Barcode position: 5' end of the read


title: Mock_6dpi_biorep2

description:
Barcode sequence: GTACT
Barcode position: 5' end of the read


title: siRNA_cid16Δ

value: anti-FLAG (Sigma-Aldrich, catalog#A2220, lot# SLBQ4019V)

value: siRNA


title: Mock_8dpi_biorep4

description:
Barcode sequence: TGTGCGAT
Barcode position: 5' end of the read


title: Mock_6dpi_biorep1

description:
Barcode sequence: TCGT
Barcode position: 5' end of the read


title: Mock_4dpi_biorep1

description:
Barcode sequence: CGGT
Barcode position: 5' end of the read


title: Mock_4dpi_biorep2

description:
Barcode sequence: ACTGT
Barcode position: 5' end of the read


title: Mock_4dpi_biorep3

description:
Barcode sequence: GACACT
Barcode position: 5' end of the read


title: Mock_2dpi_biorep4

description:
Barcode sequence: CATACGT
Barcode position: 5' end of the read


title: Mock_2dpi_biorep3

description:
Barcode sequence: TTGGAT
Barcode position: 5' end of the read


title: Mock_2dpi_biorep2

description:
Barcode sequence: GCTCT
Barcode position: 5' end of the read


title: Mock_2dpi_biorep1

description:
Barcode sequence: AGCT
Barcode position: 5' end of the read


title: Mock_0dpi_biorep3


title: Mock_0dpi_biorep1


title: Mock_0dpi_biorep2


title: AlNc14_8dpi_biorep2


title: AlNc14_8dpi_biorep3


title: AlNc14_8dpi_biorep4


title: AlNc14_8dpi_biorep1


title: Mock_4dpi_biorep4

description:
Barcode sequence: TTGCGCT
Barcode position: 5' end of the read


title: AlNc14_6dpi_biorep4_lib2


title: pO2-UN-2-1

city: Newcastle upon Tyne

first: Rajesh Reddy

last: Bommareddy

address: Ellision building

department: Health Sciences

institute: Northumbria University

description: pO2-UN-2 and pO2-UN-3_FPKM.txt

data_processing:
Raw sequence reads were checked for their quality using fastQC and reads were groomed to meet the Sanger-scaled quality values with ASCII offset 33. The groomed paired-end reads were then mapped to the reference genome of Y.lipolytica CLIB122 GCA_000002525.1 genome assembly  using Tophat2 . The gene and transcript expression levels were estimated and normalized based on FPKM (fragments per kilobase of exon per million fragments mapped) which were estimated using Cufflinks.
Genome_build: Y. lipolytica CLIB122 GCA_000002525.1 genome assembly
Supplementary_files_format_and_content: tab delimited text file is included with FPKM values for each sample

organism: Yarrowia lipolytica

source_name: cell mass

extract_protocol:
Cell mass was withdrawn from the fermentor and flash frozen on liquid nitrogen. RNA was extracted using the RiboPureTM RNA purification kit for yeast (Life technologies, Ambion). 1 ug of RNA was used by the Illumina Truseq kit to prepare sequencing libraries.
RNA libraries were prepared using standard Illumina protocols

value: ACA DC 50109

tag: group

value: uncontrolled


title: pO2-UN-2-2

description: pO2-UN-2 and pO2-UN-3_FPKM.txt


title: pO2-UN-3-2

description: pO2-UN-2 and pO2-UN-3_FPKM.txt


title: AlNc14_6dpi_biorep3_lib2


title: pO2-2-2

description: pO2-1 and pO2-2_FPKM.txt

value: controlled


title: pO2-2-1

description: pO2-1 and pO2-2_FPKM.txt


title: AlNc14_6dpi_biorep4_lib1


title: AlNc14_6dpi_biorep3_lib1


title: AlNc14_6dpi_biorep2_lib2


title: pO2-1-2

description: pO2-1 and pO2-2_FPKM.txt


title: pO2-1-1

description: pO2-1 and pO2-2_FPKM.txt


title: Mock_0dpi_biorep4


title: AlNc14_6dpi_biorep2_lib1


title: AlNc14_6dpi_biorep1_lib1


title: AlNc14_4dpi_biorep4_lib3


title: pO2-UN-3-1

description: pO2-UN-2 and pO2-UN-3_FPKM.txt


title: CS-Seq_A549_OE_06hpi_DIS3-1

first: Zuleyma

last: Peralta

state: NY

address: 1470 Madison Ave Rm 8-302

department: Genetics and Genomic Sciences

data_processing:
library strategy: Capsnatch sequencing
For Capsnatch-Seq: Due to the design of our 5’ RNA adapter, each capsnatch-Seq read starts with 12 adapter-derived nucleotides, consisting of 3 random nt, followed by a 6-nt unique barcode sequence, and another 3 random nts. Demultiplexing and removal of these 5’ adapter sequences was done using custom scripts, allowing up to two barcode base mismatches and no insertions/deletions. Remaining sequences were additionally trimmed at the 3’-end after reaching a base with a phred quality score lower than 10, or after encountering 15 bases with a phred score lower than 28. Finally, 3' Illumina adapter sequences were removed using cutadapt with a minimal overlap of 6bp and allowing for an adapter error rate of 15%. Reads less than 50nt in length after quality and adapter trimming were removed from further analysis. To identify uniformly host- and virus-derived 5’ RNA sequences, full-length adapter-trimmed reads were mapped to the human (hg38) and viral (IAV) reference genomes using TopHat 2.0.12 and Bowtie 1.0.1 with the corresponding gene annotations (GRCh38/Gencode-v23 for the human genome), requiring a minimal anchor size of 10 nucleotides and using default settings for all other arguments. To identify hybrid viral mRNA sequences, the first 20 nucleotides of each read were first masked prior to mapping to viral reference genomes using the same strategy. Hybrid reads were then identified as those masked reads mapping to the positive strand of a viral genome segment, with a start coordinate less than 20bp from the segment start.
For Directional RNA-Seq: Read sequences were trimmed at the 3’-end after reaching a base with a phred quality score lower than 10, or after encountering 15 bases with a phred score lower than 28, using custom scripts. Next, 3' Illumina adapter sequences were removed using cutadapt  with a minimal overlap of 6 bp, allowing for an adapter error rate of 15%. Reads less than 50 nt (for paired-end 100 nt reads) in length after quality and adapter trimming were removed from further analysis. Full-length adapter-trimmed reads were mapped to the human (hg38) and viral (IAV) reference genomes using STAR v2.5 with the corresponding gene annotations (Gencode GRCh37/V23 for the human genome), using detection of chimeric alignments with a minimum mapped length and a minimum chimeric overhang junction of 15 nt, and default settings for all other arguments. Total mapped fragment (i.e. paired-end read) counts per gene were determined using featureCounts with default settings
Genome_build: Human: GRCh38/Gencode-v23 annotations
Genome_build: IAV: A/Puerto Rico/8/1934 H1N1
Genome_build: Murine: mm10(GRCm38)/ENSEMBL v75 annotations.
Supplementary_files_format_and_content: Tab-separated text files summarizing for each gencode transcript the number of host-mapped or viral segment reads. One column shows raw counts obtained from featureCounts, second column containes FKPM normalized counts.

growth_protocol:
A549 cells (adenocarcinomic human alveolar basal epithelial cells) were originally obtained from the American Type Culture Collection (ATCC). Cells were maintained in culture at 37°C with 5% CO2 in Dulbecco’s minimal essential medium (DMEM, Gibco, Life Technologies) supplemented with 2 mM glutamine (Life Technologies), 10% fetal bovine serum (FBS; Hyclone), penicillin (100 U/ml; Life Technologies), and streptomycin (100 μg/ml; Gibco, Life Technologies)
We utilized the conditional system developed by (Pefanis et al., 2014). In short, this mouse model contains a conditional inversion (COIN) allele of Exos¬c3, allowing conditional ablation of RNA exosome function using tissue-specific or inducible Cre recombinase. Cre-mediated ablation of Exosc3 leads to concomitant green fluorescent protein (GFP) reporter induction from the Exosc3 locus. As such, there are two mouse types: COIN/+ (heterozygous for the conditional ablation), and COIN/COIN (homozygous for the conditional ablation). Spleens were obtained from each mouse type, and single cell suspensions were prepared by pressing the tissues through a 70 μm cell strainer, followed by homogenization using a 20g syringe. B cells were purified by negative selection using a MACS column (Miltenyi Biotec) according to the manufacturer's instructions, and then resuspended in DMEM at 500,000 cells/mL in 6-well plates. Cells were growth-stimulated with 20 ng/mL IL-4 (404-ML-010, R&D Systems) and 5 ug/mL purified anti-CD40 (Clone HM40-3, BD Biosciences).
A549 cells (adenocarcinomic human alveolar basal epithelial cells) were originally obtained from the American Type Culture Collection (ATCC). Cells were maintained in culture at 37°C with 5% CO2 in Dulbecco’s minimal essential medium (DMEM, Gibco, Life Technologies) supplemented with 2 mM glutamine (Life Technologies), 10% fetal bovine serum (FBS; Hyclone), penicillin (100 U/ml; Life Technologies), and streptomycin (100 μg/ml; Gibco, Life Technologies)

extract_protocol:
Capsnatch sequencing : 1 µg of DNase treated RNA was treated with 5U of Cap-Clip Acid Pyrophosphotase (CellScript) for 1.5hrs at 37°C to remove 5’-cap structures. Immediately after, 5 µl of freshly prepared Sodium Periodate was added to a final concentration of 500mM to block free 3’-OH ends of RNA. After incubating for 1.5hrs at 4°C the reaction was stopped by adding 1/10 volume of 1M L-lysine and incubating at room temperature for an additional 10 minutes. Next, RNA was purified with 1.8X volume of AMPure XP beads (Beckman Coulter), and 10 µM barcoded RNA adapters (5'- GUU CAG AGU UCU ACA GUC CGA CGA UCN NNX XXX XXN NN -3’, Bioo Scientific; N=randomized bases; X=barcode adapter) were ligated to the 5’ ends of RNAs in an overnight (~16-18hrs) reaction at 16°C containing 5U of T4 RNA ligase, 10 µM ATP, and 1 µl of RNAse Inhibitor (Promega, N2618). Adapter-ligated RNA was purified with 1.8X volume of AMPure XP beads, and up to six 1 µg samples with distinct 5’ adapter barcodes were pooled together. Ribosomal RNAs were removed from pooled samples using the Ribo-Zero Gold rRNA Removal Kit (Human/Mouse/Rat) (Illumina) according to the manufacturer’s protocol, using 4 µls of Reaction Buffer and 10 µls of rRNA Removal Solution. rRNA depleted samples were purified with 1.6X volume AMPure XP beads and reverse-transcribed using by hybridizing 10 µM of a custom 3’ primer (5'-AGA CGT GTG CTC TTC CGA TCT N*N*N*N*N*N*-3',
Bioo Scientific, N*=randomized bases) for 2 minutes at 65°C. Then, 25mM of dNTP Mix (Thermo Scientific), 1 µl of RNasin® Plus RNase Inhibitor (Promega), 5X First Strand Buffer, 200mM of DTT, and SuperScript® III Reverse Transcriptase (Invitrogen) were added to the reaction and incubated in a thermocycler for 25°C for 10 minutes, 42°C for 50 minutes and 70°C for 15 minutes. Next, cDNAs were purified with 1.8X volume AMPure XP beads, followed by PCR amplification using Kapa HiFi HotStart Ready Mix, 25µM SR forward primer (5'-AAT GAT ACG GCG ACC ACC GAG ATC TAC ACG TTC AGA GTT CTA CAG TCC G-s-A-3', Bioo Scientific) and a reverse indexed primer (5’-CAA GCA GAA GAC GGC ATA CGA GAT XXX XXX GTG ACT GGA GTT CAG ACG TGT GCT CTT CCG ATC*T-3’, IDT) for 20 cycles. PCR products were purified using 1.8X AMPure XP beads fragments of 200-400bp were size-selected using BluePippin 2% M1 gels (Sage Scientific). Following library validation on the Agilent Bioanalyzer samples were sequenced on the Illumina HiSeq 2500 platform in a 100 bp single-end read run format.
A549 cells were washed 3X in cold PBS, and total RNA was extracted using Trizol reagent.
B cells were washed 3X in cold PBS, and total RNA was extracted using Trizol reagent.

treatment_protocol:
Transfection of plasmid DNA was achieved using Lipofectamine 3000 (ThermoFisher Scientific). A549 cells were used 36 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.
Transfection was performed using the Lipofectamine RNAiMAX transfection reagents according to the manufacturer’s instructions (Invitrogen). Cells were transfected with siRNA pools (all from Dharmacon) targeting the genes encoding human EXOSC10 or DIS3, or with a control non-targeting pool, at a final concentration of 50 nM. Cells were used 48 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 4 hours post infection.
B cells were treated with 100 nM tamoxifen for 3 days in order to induce the conversion event. B cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.
Transfection of plasmid DNA was achieved using Lipofectamine 3000 (ThermoFisher Scientific). A549 cells were used 36 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.

tag: cell line

tag: cell type

tag: viral strain

value: A/Puerto Rico/8/1934 H1N1

tag: treatment

value: DIS3 overexpression

tag: timepoint

value: 06 hours post-infection

tag: genome build for data mapping

value: hg38(GRCh38) with GENCODE v23 annotations


title: AlNc14_4dpi_biorep4_lib2

tag: tissue


title: CS-Seq_A549_OE_06hpi_DIS3-2

value: adenocarcinomic human alveolar basal epithelial cells


title: CS-Seq_A549_OE_00hpi_EV-2

value: Empty Vector

value: 00 hours post-infection


title: AlNc14_4dpi_biorep3_lib2

value: multi-parent recombinant inbred line MAGIC-107

value: aerial shoots

value: 4-5 weeks


title: AlNc14_4dpi_biorep4_lib1


title: CS-Seq_A549_OE_06hpi_EV-2


title: CS-Seq_A549_OE_00hpi_EV-1


title: AlNc14_4dpi_biorep2_lib3


title: AlNc14_4dpi_biorep3_lib3


title: AlNc14_4dpi_biorep3_lib1


title: AlNc14_4dpi_biorep2_lib1


title: CS-Seq_A549_OE_00hpi_DIS3-2


title: AlNc14_4dpi_biorep1_lib3


title: CS-Seq_A549_siRNA_04hpi_siEXOSC10-2

treatment_protocol:
Transfection was performed using the Lipofectamine RNAiMAX transfection reagents according to the manufacturer’s instructions (Invitrogen). Cells were transfected with siRNA pools (all from Dharmacon) targeting the genes encoding human EXOSC10 or DIS3, or with a control non-targeting pool, at a final concentration of 50 nM. Cells were used 48 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 4 hours post infection.
Transfection was performed using the Lipofectamine RNAiMAX transfection reagents according to the manufacturer’s instructions (Invitrogen). Cells were transfected with siRNA pools (all from Dharmacon) targeting the genes encoding human EXOSC10 or DIS3, or with a control non-targeting pool, at a final concentration of 50 nM. Cells were used 48 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 4 hours post infection.
B cells were treated with 100 nM tamoxifen for 3 days in order to induce the conversion event. B cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.
Transfection of plasmid DNA was achieved using Lipofectamine 3000 (ThermoFisher Scientific). A549 cells were used 36 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.

value: 04 hours post-infection


title: CS-Seq_A549_OE_00hpi_DIS3-1


title: AlNc14_4dpi_biorep1_lib2


title: CS-Seq_A549_siRNA_04hpi_siEXOSC10-1


title: CS-Seq_A549_siRNA_04hpi_siDIS3-2


title: CS-Seq_A549_siRNA_04hpi_siCTRL-2

value: siCTRL


title: CS-Seq_A549_siRNA_04hpi_siDIS3-1


title: CS-Seq_A549_OE_06hpi_EV-1


title: AlNc14_6dpi_biorep1_lib2


title: CS-Seq_A549_siRNA_00hpi_siEXOSC10-2


title: CS-Seq_A549_siRNA_04hpi_siCTRL-1


title: AlNc14_2dpi_biorep4_lib3


title: AlNc14_2dpi_biorep4_lib2


title: AlNc14_2dpi_biorep4_lib1


title: CS-Seq_A549_siRNA_00hpi_siEXOSC10-1


title: AlNc14_4dpi_biorep2_lib2


title: CS-Seq_A549_siRNA_00hpi_siDIS3-2


title: AlNc14_2dpi_biorep3_lib2


title: CS-Seq_A549_siRNA_00hpi_siDIS3-1


title: AlNc14_2dpi_biorep3_lib3


title: CS-Seq_A549_siRNA_00hpi_siCTRL-1


title: CS-Seq_A549_siRNA_00hpi_siCTRL-2


title: AlNc14_2dpi_biorep2_lib3


title: AlNc14_2dpi_biorep2_lib2


title: AlNc14_2dpi_biorep2_lib1


title: AlNc14_2dpi_biorep3_lib1


title: AlNc14_2dpi_biorep1_lib3


title: AlNc14_0dpi_biorep3


title: AlNc14_0dpi_biorep2


title: AcNc02_8dpi_biorep2


title: AlNc14_4dpi_biorep1_lib1


title: AcNc02_8dpi_biorep4


title: AlNc14_2dpi_biorep1_lib1


title: AlNc14_2dpi_biorep1_lib2


title: AcNc02_8dpi_biorep1


title: AlNc14_0dpi_biorep4


title: AcNc02_8dpi_biorep3


title: AlNc14_0dpi_biorep1


title: AcNc02_6dpi_biorep2_lib1


title: AcNc02_6dpi_biorep4_lib1


title: AcNc02_6dpi_biorep1_lib2


title: AcNc02_6dpi_biorep2_lib2


title: AcNc02_6dpi_biorep4_lib2


title: AcNc02_6dpi_biorep3_lib1


title: AcNc02_4dpi_biorep3_lib3


title: AcNc02_4dpi_biorep4_lib1


title: AcNc02_4dpi_biorep3_lib2


title: AcNc02_4dpi_biorep3_lib1


title: AcNc02_6dpi_biorep1_lib1


title: AcNc02_4dpi_biorep2_lib2


title: AcNc02_4dpi_biorep1_lib3


title: AcNc02_4dpi_biorep2_lib1


title: AcNc02_4dpi_biorep1_lib2


title: AcNc02_6dpi_biorep3_lib2


title: AcNc02_4dpi_biorep1_lib1


title: AcNc02_2dpi_biorep4_lib3


title: AcNc02_2dpi_biorep4_lib2


title: AcNc02_2dpi_biorep4_lib1


title: AcNc02_2dpi_biorep3_lib1


title: AcNc02_2dpi_biorep3_lib3


title: AcNc02_2dpi_biorep3_lib2


title: AcNc02_4dpi_biorep4_lib2


title: AcNc02_4dpi_biorep4_lib3


title: AcNc02_0dpi_biorep4


title: AcNc02_2dpi_biorep1_lib1


title: AcNc02_2dpi_biorep2_lib1


title: AcNc02_2dpi_biorep1_lib2


title: AcNc02_4dpi_biorep2_lib3


title: AcNc02_2dpi_biorep2_lib3


title: AcNc02_0dpi_biorep2


title: AcNc02_2dpi_biorep1_lib3


title: TruSeq_MurB_ConKO_Inf_C_NoTam_2

data_processing:
library strategy: Directional RNA sequencing
For Capsnatch-Seq: Due to the design of our 5’ RNA adapter, each capsnatch-Seq read starts with 12 adapter-derived nucleotides, consisting of 3 random nt, followed by a 6-nt unique barcode sequence, and another 3 random nts. Demultiplexing and removal of these 5’ adapter sequences was done using custom scripts, allowing up to two barcode base mismatches and no insertions/deletions. Remaining sequences were additionally trimmed at the 3’-end after reaching a base with a phred quality score lower than 10, or after encountering 15 bases with a phred score lower than 28. Finally, 3' Illumina adapter sequences were removed using cutadapt with a minimal overlap of 6bp and allowing for an adapter error rate of 15%. Reads less than 50nt in length after quality and adapter trimming were removed from further analysis. To identify uniformly host- and virus-derived 5’ RNA sequences, full-length adapter-trimmed reads were mapped to the human (hg38) and viral (IAV) reference genomes using TopHat 2.0.12 and Bowtie 1.0.1 with the corresponding gene annotations (GRCh38/Gencode-v23 for the human genome), requiring a minimal anchor size of 10 nucleotides and using default settings for all other arguments. To identify hybrid viral mRNA sequences, the first 20 nucleotides of each read were first masked prior to mapping to viral reference genomes using the same strategy. Hybrid reads were then identified as those masked reads mapping to the positive strand of a viral genome segment, with a start coordinate less than 20bp from the segment start.
For Directional RNA-Seq: Read sequences were trimmed at the 3’-end after reaching a base with a phred quality score lower than 10, or after encountering 15 bases with a phred score lower than 28, using custom scripts. Next, 3' Illumina adapter sequences were removed using cutadapt  with a minimal overlap of 6 bp, allowing for an adapter error rate of 15%. Reads less than 50 nt (for paired-end 100 nt reads) in length after quality and adapter trimming were removed from further analysis. Full-length adapter-trimmed reads were mapped to the human (hg38) and viral (IAV) reference genomes using STAR v2.5 with the corresponding gene annotations (Gencode GRCh37/V23 for the human genome), using detection of chimeric alignments with a minimum mapped length and a minimum chimeric overhang junction of 15 nt, and default settings for all other arguments. Total mapped fragment (i.e. paired-end read) counts per gene were determined using featureCounts with default settings
Genome_build: Human: GRCh38/Gencode-v23 annotations
Genome_build: IAV: A/Puerto Rico/8/1934 H1N1
Genome_build: Murine: mm10(GRCm38)/ENSEMBL v75 annotations.
Supplementary_files_format_and_content: Tab-separated text files summarizing for each gencode transcript the number of host-mapped or viral segment reads. One column shows raw counts obtained from featureCounts, second column containes FKPM normalized counts.

organism: Mus musculus

extract_protocol:
Directional RNA sequencing : 1 µg  of DNase-treated RNA was depleted of rRNAs using the Ribo-Zero Gold rRNA Removal Kit (Human/Mouse/Rat) (Illumina) according to the manufacturer’s instructions, and purified post-depletion with 1.6X volume AMPureXP beads. Barcoded directional RNA-Seq libraries were then prepared using the NEBNext Ultra Directional RNA Library Prep kit for Illumina (New England Biolabs), per kit instructions. PCR products were purified with 1.8X volume AMPure XP beads and fragments of 300-500bp were size-selected using BluePippin 2% M1 gels (Sage Scientific). Afterwards, libraries were sequenced on the Illumina HiSeq 2500 platform in a 100 bp single-end read run format.
A549 cells were washed 3X in cold PBS, and total RNA was extracted using Trizol reagent.
B cells were washed 3X in cold PBS, and total RNA was extracted using Trizol reagent.

treatment_protocol:
B cells were treated with 100 nM tamoxifen for 3 days in order to induce the conversion event. B cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.
Transfection was performed using the Lipofectamine RNAiMAX transfection reagents according to the manufacturer’s instructions (Invitrogen). Cells were transfected with siRNA pools (all from Dharmacon) targeting the genes encoding human EXOSC10 or DIS3, or with a control non-targeting pool, at a final concentration of 50 nM. Cells were used 48 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 4 hours post infection.
B cells were treated with 100 nM tamoxifen for 3 days in order to induce the conversion event. B cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.
Transfection of plasmid DNA was achieved using Lipofectamine 3000 (ThermoFisher Scientific). A549 cells were used 36 hours after transfection. A549 cells were infected with Influenza A/Puerto Rico/8/1934 H1N1Viral at a multiplicity of infection (MOI) of 3 and cells were analyzed at 6 hours post infection.

value: murine primary B cell isolated from spleens

value: mm10(GRCm38) with ENSEMBL v75 annotations


title: TruSeq_MurB_ConKO_Inf_C_Tam_1

value: Tamoxifen


title: TruSeq_MurB_ConKO_Inf_CC_Tam_2

value: C/C


title: TruSeq_MurB_ConKO_Inf_C_NoTam_1


title: TruSeq_MurB_ConKO_Inf_C_Tam_2


title: AcNc02_0dpi_biorep3


title: TruSeq_MurB_ConKO_Inf_CC_Tam_1


title: TruSeq_MurB_ConKO_Inf_CC_NoTam_1


title: TruSeq_MurB_ConKO_Inf_CC_NoTam_2


title: AcNc02_0dpi_biorep1


title: TS685_II_B

city: Solna

first: Vicent

last: Pelechano

address: Nobels väg 16

department: MTC

country: Sweden

institute: ScilifeLab - Karolinska Institutet

data_processing:
Base-calling was done during processing of sequencing data with the Illumina pipeline casava v1.8.2.
Reads were mapped to S. cerevisiae genome (SGD R64-1-1) using the GSNAP version 2012-01-11.
Genome_build: R64-1-1 (sacCer3)
Supplementary_files_format_and_content: bedGraph

organism: Saccharomyces cerevisiae

growth_protocol: S. cerevisiae strains were grown to mid-log phase (OD~1) in YPD (1% yeast extract, 2% peptone, 2% glucose) at 30ºC.

extract_protocol:
Total RNA was isolated by a standard hot phenol method and treated with RNase-free DNaseI using Turbo DNA-free kit (Ambion).
Genome-wide quantification of gene expression and poly(A) site usage was performed using 3'T-fill approach (Wilkening et al. 2013; PMID: 23295673) and the QuantSeq 3'mRNA-Seq v016.24 and v016.96 (Lexogen) for batches A and B, respectively, according to manufacturer's instructions. Polyadenylated in vitro transcripts (ATCC 87482, 87483 and 87484) were added as external controls to the RNA samples (as described in Wilkening et al. 2013; PMID: 23295673).


title: TS801_II_B


title: TS685_I_B


title: TS801_I_B


title: TS801_III_B


title: AcNc02_2dpi_biorep2_lib2


title: TS1248_III_B

value: YJR141W-PH::KanR; his3^1leu2^0 ura3^0 met15^0 yjr141w-ph BY4741


title: PTC_13_RNAseq

city: Lund

first: Kajsa

last: Paulsson

address: Sölvegatan 19

department: Department of Clinical Genetics

institute: Lund University

data_processing:
Illumina Casava1.7 software used for basecalling.
Data processing of RNA using the  TCGA UNC V2 RNAseq protocol (https://cghub.ucsc.edu/docs/tcga/UNC_mRNAseq_summary.pdf)  have been previously described for The Cancer Genome Atlas (TCGA)
Expected read counts in the '.genes.results' files generated by TCGA UNC V2 RNAseq workflow were used as the input for DESeq2, limma-voom  and edgeR, respectively.
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include Expected read counts for each Sample

extract_protocol:
RNA was extracted from the cell lines using the RNeasy Mini Kit (Qiagen, Hilden, Germany)
Paired-end RNA libraries were constructed using the TruSeq RNA Sample Preparation Kit v2 (Illumina)

value: papillary thyroid cancer cell line

value: thyroid

tag: passages


title: ∆spxA1/∆spxA2 H2O2 2

city: Gainesville

first: Jessica

middle: K

last: Kajfasz

state: FL

address: 1395 Center Drive

department: Oral Biology

institute: University of Florida

description: S. mutans ∆spxA1/∆spxA2 mutant 0.5mM H2O2 treatment replicate 2

data_processing:
DEMULTIPLEXING - bcltofastq-1.8.4
CLEANING - Trimmomatic-0.32 "SLIDINGWINDOW:4:20 TRAILING:13 LEADING:13 ILLUMINACLIP:adapters.fasta:2:30:10 MINLEN:25"
MAPPING - SHRiMP-2.2.3 ""--qv-offset 33 --all-contigs"
Normalization & Diff. EXP - Cufflinks-2.0.2 "--FDR 0.05 –u -b GENOME"
Differential expression analysis is performed using Cufflinks version 2.0.2; specifically, cuffdiff2 and usage of the general transfer format (GTF) annotation file
Genome_build: NC_004350.2
Supplementary_files_format_and_content: The processed .txt data files contain comparisons of the pooled data (two replicates for each sample type) between two samples, represented as the log2 fold change of expression of each coding region (SMU).

organism: Streptococcus mutans UA159

source_name: Streptococcus mutans ∆spxA1/∆spxA2 mutant

growth_protocol: Cells were grown in brain-heart infusion (BHI) medium to OD600 = 0.4

extract_protocol:
Total RNA was isolated from homogenized S. mutans cell lysates by hot acid-phenol:chloroform extractions.  RNA was precipitated with ice-cold isopropanol and 3 M sodium acetate (pH 5) at 4°C before RNA pellets were dissolved in nuclease-free H2O and treated with DNase I.  For each total RNA extract, 10 μg RNA aliquots were subjected to a second DNase I treatment using the DNA-free kit (Ambion).
The TruSeq RNA Sample Preparation Kit V2 (Illumina, San Diego, CA) was used for next generation sequencing library construction following the instructions from the manufacturer.
Single end reads of 100nt were generated for each sample and aligned to the organism specific reference genome.

value: spxA1/spxA2 mutant

value: Cells were grown in BHI medium to OD600 of 0.4, exposed to 0.5mM H2O2 for 5 minutes, then harvested by centrifugation, and stored at -80°C until use

tag: media

value: BHI; brain heart infusion with 0.5mM H2O2


title: PTC_5_RNAseq


title: TS1248_I_B


title: PTC_4_RNAseq


title: BY4741_III_B

value: Mat a; his3^1leu2^0 ura3^0 met15^0


title: ∆spxA1/∆spxA2 H2O2 1

description: S. mutans ∆spxA1/∆spxA2 mutant 0.5mM H2O2 treatment replicate 1


title: ∆spxA2 H2O2 2

description: S. mutans ∆spxA2 mutant 0.5mM H2O2 treatment replicate 2

source_name: Streptococcus mutans ∆spxA2 mutant

value: spxA2 mutant


title: BY4741_II_B


title: ∆spxA2 H2O2 1

description: S. mutans ∆spxA2 mutant 0.5mM H2O2 treatment replicate 1


title: ATC_14_RNAseq

value: Anaplastic thyroid cancer cell line


title: BY4741_I_B


title: ATC_10_RNAseq


title: ATC_12_RNAseq


title: ∆spxA1 H2O2 2

description: S. mutans ∆spxA1 mutant 0.5mM H2O2 treatment replicate 2

source_name: Streptococcus mutans ∆spxA1 mutant

value: spxA1 mutant


title: ∆spxA1 H2O2 1

description: S. mutans ∆spxA1 mutant 0.5mM H2O2 treatment replicate 1


title: UA159 H2O2 2

description: S. mutans UA159 parent strain 0.5mM H2O2 treatment replicate 2

source_name: Streptococcus mutans UA159 wild type

value: wild type


title: TS801_III

data_processing:
Base-calling was done during processing of sequencing data with the Illumina pipeline casava v1.8.2.
Reads were mapped to S. cerevisiae genome (SGD R64-1-1) using the GSNAP version 2012-01-11.
Genome_build: R64-1-1 (sacCer3)


title: TS1248_II


title: UA159 H2O2 1

description: S. mutans UA159 parent strain 0.5mM H2O2 treatment replicate 1


title: UA159 T0 2

description: S. mutans UA159 parent strain untreated replicate 2

value: Cells were grown in BHI medium to OD600 of 0.4, then harvested by centrifugation, and stored at -80°C until use

value: BHI; brain heart infusion


title: ATC_2_RNAseq


title: ATC_1_RNAseq


title: UA159 T0 1

description: S. mutans UA159 parent strain untreated replicate 1


title: TS685_III


title: BY4741_III


title: ATC_15_RNAseq


title: CEC_like#3_RNAseq

status: Public on May 07 2017

city: los angeles

first: Qin

last: An

state: CALIFORNIA

address: Room 6554 Gonda Building, UCLA

department: Department of Human Genetics

institute: ucla

data_processing:
All obtained reads from each sample were mapped against hg19 genome with Bowtie/Tophat 2.0.6.
The mapped reads were subsequently quantified by HTSeq-0.6.1
Expression level of each gene was quantified and normalized to RPKM (Reads Per Kilobase of exon per Megabase of library size) using R. Genes with an maximum RPKM <0 were filtered out.
Genome_build: hg19
Supplementary_files_format_and_content: [.txt] tab-delimited text files include RPKM values

source_name: ES cell-derived CEC-like cells

growth_protocol: Passage human ES cells with Collagenase IV and Dispase, Resuspend the Embryonic Body with KSR medium plus 10μm SB431542 and 500ng/ml Noggin for 7 days in low attachment plates. Then change into N2 medium plus 10μm SB431542 and 500ng/ml Noggin for another 7 days. Two weeks later, change the medium with half of the fresh CEC medium and half of bovine CEC conditional medium for several days to see original area of the coming CEC-like cells( usually it takes about 7-14 days).

extract_protocol:
RNA were extracted using the RNeasy Micro Kit (Qiagen, USA). RNA concentration was detected by nanodrop spectrophotometer.
Library construction was followed by Illumina TruSeqTM RNA Sample Preparation Guide protocol. 100 ng of total RNA was measured and input for library construction, poly-T oligo-attached magnetic beads was used to purify the poly-A containing mRNA molecules. RNA fragments were then reverse-transcribed into first strand cDNA and second strand cDNA. After end repaired and A-tail, cDNAs were ligated with illumine true-seq barcode adapters for PCR amplification. After PCR enrichment, the final libraries were tested by Qubit Fluorometer (Invitrogen, USA), bioanalyzer (Angilent Technology) and KAPA library quantification kit(KAPA biosystems). We submitted 10 nmol of each sample to sequencing according to the manufacturer’s instruction with Illumina Hi-seq 2000.

value: ES cell-derived CEC-like cells

tag: differentiation time

value: 1 months


title: CEC_like#2_RNAseq

value: 3 months


title: CEC_like#1_RNAseq

value: 70 days


title: Lmx1b ChIP-seq (BMO8 validation)

status: Public on May 06 2017

city: Loma Linda

first: Kerby

middle: C

last: Oberg

state: CA

address: 24785 Stewart St

department: Pathology

institute: Loma Linda University

description: Lmx1b bound chromatin

data_processing:
Reads were aligned to the mouse genome (mm10) using the default settings for BWA algorithm.
Alignments were extended in silico at their 3’-ends to a length of 200 bp and assigned to 32-nt bins along the genome.
Lmx1b peak locations were determined using the MACS algorithm (v1.4.2) with a cutoff of p-value = 1e-5.
Genome_build: mm10

source_name: limb buds

growth_protocol: harvested on embryonic day 12 at noon (e12.5)

extract_protocol:
After disruption with a Dounce homogenizer lysates were sonicated and the DNA sheared to an average length of 300-500 bp.  Chromatin (30 ug) was precleared with protein A agarose beads. Incubated with 20 ul antibody against Lmx1b (BMO8)53 kindly provided Dr. Witzgall.. Crosslink was reversed Proteinase K digestion and incubation overnight at 65 °C. ChIP DNA was purified by phenol-chloroform extraction and ethanol precipitation.
After end-polishing, dA-addition, adaptor ligation and final PCR amplification step, the resulting DNA libraries were quantified and sequenced on Illumina’s NextSeq 500.

treatment_protocol: Rapid freezing (liquid nitrogen), frozen samples pooled and then fixed (1% formaldehyde) during thawing (15 min).

tag: Stage

value: limb buds

tag: antibody

value: Lmx1b polyclonal (BM08), Suleiman et al., 2007


title: ATC_8_RNAseq

value: thyroid


title: WT_0h_3

city: Boston

first: Kai

middle: W

last: Wucherpfennig

state: MA

address: 1 Jimmy Fund Way

department: Cancer Immunology and Virology

institute: Dana Farber Cancer Institute

description:
Processed data file: STAR_Gene_Counts.csv
Processed data file: Cuff_Gene_Counts.csv

data_processing:
Illumina Bcl2fastq v2.18.
Sequencing reads were aligned to mm9 reference genome using STAR (v2.5).
Raw gene counts were generated from STAR output and normalized RPKM counts were generated using cufflinks v2.2.1.
Genome_build: mm9 (MGSCv37)
Supplementary_files_format_and_content: STAR_Gene_Counts.csv: Comma-separated text file includes raw gene counts for each sample.
Supplementary_files_format_and_content: Cuff_Gene_Counts.csv: Comma-separated text file includes normalized RPKM counts for each sample.

source_name: Spleen and lymph nodes

growth_protocol: CD8 T cells were isolated form spleens and lymph nodes by negative selection (EasySep CD8+ negative selection kit, Stem Cell) and cultured in RMPI-1640 supplemented with 10% fetal calf serum, 50 units/ml Penicillin-Streptomycin, 10 mM HEPES, 2 mM L-glutamine, and 55 microM beta-mercaptoethanol. Cells were activated using CD3/CD28 Dynabeads (Thermo Fisher) for 2 days, then cultured in supplemented RPMI-1640 with 10ng/mL IL-2 (Biolegend) for a further 5 days.

extract_protocol:
RNA was isolated from cells at each time point (0, 4, 18h) using Direct-zol RNA miniprep kit (Zymo) as per the manufacturer's instructions.
RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: After 7 days activation, cells were stimulated with 20ng/mL IFN-beta (Biolegend) for 0, 4, or 18h.

tag: strain/background

tag: genotype/variation

value: wild-type

value: CD8 T cell

tag: activation

value: anti-CD3/CD28 beads for 2d, IL-2 for 5d

value: IFN-beta

tag: treatment duration


title: aCEC67_RNAseq

source_name: cultured adult CECs

value: cultured adult CECs


title: WT_4h_3


title: KO_0h_3

value: miR-31 knockout


title: ATC_7_RNAseq

value: Anaplastic thyroid cancer cell line


title: TS801_I

tag: genotype

tag: strain


title: sRNA_No_Stress

city: Gif Sur Yvette

first: Jeremie

last: Bazin

address: Bâtiment 630, rue de Noetzlin Plateau du Moulon

country: France

institute: CNRS

description:
small RNA
shortStack.txt

data_processing:
Library strategy: Ribo-Seq and sRNA seq
Base calling and demultiplexing performed with Illumina CASAVA version 1.8.
R/Bioconductor (version 2.15.1) and FastX_toolkit (http://hannonlab.cshl.edu) were used in short read preprocessing steps (Quality filtering, adaptor trimming).
For polyA RNA-seq and Ribo-seq and dsRNA-seq, the processed short reads were mapped to the Arabodopsis genome (Version TAIR10) using TopHat v1.3.1 (Trapnell et al., 2009) with command "tophat -p 8 -g 1 --segment-length 15 -i 30 -I 3000 -G".
Read count data were calculated for each specific mRNA feature.
For sRNA-seq, trimmed and quality filtered reads were mapped with the short read aligner Bowtie 2.2.9 using the following arguments (-k 50, non-deterministic). ShortStack v3 (Axtell et al 2013) was used to detect small RNA cluster and to calculate their phasing score
Genome_build: Genome_build: TAIR10

source_name: 14 days-old seedling roots

growth_protocol: Transgenic Arabidopsis thaliana (Col-0) expressing 35S:His6FLAG-RPL18B line 12-2-4 (background Columbia-0) was used in this study.  Seeds were surface sterilized and sown on a nylon mesh (100 µm, Nitex-Sefar) in square 12 cm square petri plates containing a solid modified Murashige and Skoog (MS/10) medium (0.15 mM MgSO4, 2.1 mM NH4NO3, 1.9 mM KNO3, 0.5 mM NaH2PO4, 0.34 mM CaCl2, 0.5 µM KI, 10 µM FeCl2, 10 µm H3BO3, 10 µM MnSO4, 3 µM ZnSO4, 0.1 µM CuSO4, 0.1 µM CoCl2, 1 µM Na2MoO4, 5.9 µM thiamine, 4.9 µM pyridoxine, 8.1 µM nicotinic acid, 55 µM inositol, and 3.4 mM 2-(N-morpholino)ethanesulfonic acid (MES) [pH 5.7], 0.8% (w/v) agar). In the Pi-deficient medium, NaCl (0.5 mM) replaced the equivalent amount of sodium provided by NaH2PO4. Growth was in a chamber at 23 °C under a 16 h light (80 μE m-2 sec-1) and 8 h dark cycle for 7 d.

extract_protocol:
Small RNA-seq libraries were prepared from total RNA extracted using TRIzol  from 100 mg of root tissue. RNA was resuspended in 10 µl of 10 mM Tris (pH 7.0) and separated on a 7 M urea-denaturing  polyacrylamide gel electrophoresis (PAGE) in a 15% polyacrylamide gel. Marker oligos were used to demarcate fragments of 18-28 nt, which were collected. Gel slices were processed in 250 µl gel extraction buffer [20 mM Tris (pH 7.0), 250 mM NaOAc, 1 mM; EDTA, 0.25% (w/v); SDS] by freezing at -80°C for at least 30 min and then incubation for 15 min at 23-25 °C with rocking. The eluate was recovered by centrifugation for 10 min at 13,000 x g and the RFs recovered by ethanol precipitation.
sRNA libraries were prepared using NEXTflex™ Small RNA Sequencing Kit (BIOO Scientific).

treatment_protocol:
Transgenic Arabidopsis thaliana (Col-0) expressing 35S:His6FLAG-RPL18B line 12-2-4 (background Columbia-0) was used in this study.  Seeds were surface sterilized and sown on a nylon mesh (100 µm, Nitex-Sefar) in square 12 cm square petri plates containing a solid modified Murashige and Skoog (MS/10) medium (0.15 mM MgSO4, 2.1 mM NH4NO3, 1.9 mM KNO3, 0.5 mM NaH2PO4, 0.34 mM CaCl2, 0.5 µM KI, 10 µM FeCl2, 10 µm H3BO3, 10 µM MnSO4, 3 µM ZnSO4, 0.1 µM CuSO4, 0.1 µM CoCl2, 1 µM Na2MoO4, 5.9 µM thiamine, 4.9 µM pyridoxine, 8.1 µM nicotinic acid, 55 µM inositol, and 3.4 mM 2-(N-morpholino)ethanesulfonic acid (MES) [pH 5.7], 0.8% (w/v) agar). For  Pi-replete treatment, seedlings were transfered of the same medium on a new plate for 7 days . Growth was in a chamber at 23 °C under a 16 h light (80 μE m-2 sec-1) and 8 h dark cycle
For nutrient treatments, seedlings on the mesh were transferred to new plates with Pi replete (+Pi) MS/10 or Pi-deficient MS/10 (-Pi) medium and grown for 7 d more before the complete root system. All harvested tissue was directly immersed in liquid nitrogen and stored at -80 ˚C for subsequent use.

tag: line

tag: molecule

value: small RNA

tag: sample type

value: control


title: Lmx1b ChIP-seq

description: Lmx1b bound chromatin

tag: developmental stage


title: TS685_II


title: Total_NoStress

description:
PolyA RNA
count_ncRNA.txt
count_uORF.txt
countDF_CDS_Bygene.txt
countDF_five_prime_UTR_rpkm.txt
countDF_three_prime_UTR_rpkm

extract_protocol:
Pulverized frozen root tissue (2.5 mL) was thawed in 5 mL of PEB and a S16 fraction was obtained as detailed by Mustroph et al. 2009 PNAS.  RNA was extracted from 500µl of the S16 fraction using Trizol
Poly(A)+ RNA-seq libraries were prepared from 15 µg of DNAse I (Thermo Scientific) treated RNA extracted from the S16 fraction as described by Wang et al., PlosOne 2011

value: PolyA RNA


title: Input DNA

description: WT and mutant pooled chromatin


title: Wild type mouse 3

city: Worcester

first: Rui

last: Li

state: Massachusetts

address: 364 Plantation Street, LRB

department: MCCB

institute: University of Massachusetts Medical School

data_processing:
CLC Genomic Workbench 7.5
Aligned reads to the genome with 0.95 similarity and 0.9 coverage threshold
Quantified gene expression with "uniq exon read count"
Used DESeq2 for differential expression analysis (p value adjusted by FDR method)
Genome_build: Mus_musculus.GRCm38.dna.fa/Mus_musculus.GRCm38.78.gtf
Supplementary_files_format_and_content: xlsx format; gene expression values (uniq exon count) and DESeq2 analysis result (KO vs WT)

source_name: Brain

growth_protocol: Generated from breeding Kdm6a-LoxP females with Thy1-Cre males

extract_protocol:
Qiagen Rneasy Mini
Illumina TruSeq RNA library

tag: Sex

value: Male

tag: age

value: Adult

value: wild type


title: RS_P_Stress

description:
Ribosome Protected Fragment (RPF)
count_ncRNA.txt
count_uORF.txt
countDF_CDS_Bygene.txt
countDF_five_prime_UTR_rpkm.txt
countDF_three_prime_UTR_rpkm

extract_protocol:
Ribosome-Protected Footprint Isolation and Ribo-seq Library Construction. Pulverized frozen root tissue (2.5 mL) was thawed in 5 mL of PEB and a S16 fraction was obtained as briefly described above and detailed by Mustroph et al. 2009 PNAS. Polysomes were isolated by TRAP and digested with RNAse. EZview Red anti-FLAG M2 magnetic beads (250 μL) were prewashed twice with RNAse Digestion Buffer [RNID; 20 mM Tris·HCl (pH 8.0), 140 mM KCl, 35 mM MgCl2, 50 μg/mL cycloheximide, 50 μg/mL chloramphenicol]. These were added to the S16 fraction and binding of FLAG epitope-tagged ribosomes associated with RNA was achieved by by incubation with gentle rocking for 2 h at 4 °C. The supernatant was removed with a magnet and the beads were washed four times by resuspension in 5 mL RNID followed by capture. Ribosome footprint fragments (RFs) were obtained by resuspension of the beads in 1 mL of RNID buffer additionally containing 500 units of RNase I (Ambion; ca. 15 U/µg RNA) and incubation for 90 min at 23-25 °C with rocking. RFs were extracted from ribosomes with TRIzol. For fragment size selection, the RNA was resuspended in 10 µl of 10 mM Tris (pH 7.0) and separated on a 7 M urea-denaturing  polyacrylamide gel electrophoresis (PAGE) in a 15% polyacrylamide gel. Marker oligos were used to demarcate fragments of 26-32 nt, which were collected. Gel slices were processed in 250 µl gel extraction buffer [20 mM Tris (pH 7.0), 250 mM NaOAc, 1 mM; EDTA, 0.25% (w/v); SDS] by freezing at -80°C for at least 30 min and then incubation for 15 min at 23-25 °C with rocking. The eluate was recovered by centrifugation for 10 min at 13,000 x g and the RFs recovered by ethanol precipitation.
RFs were dephosphorylated using T4 polynucleotide kinase, resuspended in 25 µl 10 mM Tris (pH 8.0) and denatured for 2 min at 75°C, equilibrated to 37°C and brought to a volume of 50 µl in 1 T4 polynucleotide kinase reaction buffer with 25 U T4 polynucleotide kinase (NEB M0201S) and 12.5 U SuperaseIn. This was incubated 1 h at 37°C and heat-inactivated 10 min at 70°C. Dephosphorylated RFs were ethanol precipitation and linkers were ligated for 2.5 hr at 37°C in a 20 µl reaction consisting of the RFs, 12.5% w/v PEG 8000, 10% DMSO, 1x T4 Rnl2(tr) reaction buffer, 20 U SuperaseIn, 500 ng preadenylylated miRNA cloning linker (IDT, miRNA cloning linker #1), and 200 U T4 Rnl2(tr) (NEB, M0242L). Ligation products were separated by a denaturing 15% PAGE, the ligated-RFs were excised, recovered and resuspended in 10.0 µl of 10 mM Tris (pH 8). Reverse transcription was then performed in a 20 µL reaction by addition of 4 µL of 5× first-strand buffer, 1.0 μL of SUPERase•In, 1.0 μL of 100 mM DTT, and 1.0 μL of SuperScript III (Invitrogen) and incubation for 30 min at 48 °C. The reaction was heat- inactivated for 15 min at 98 °C, separated on a 10% (wt/vol) 7 M urea-denaturing PAGE in 0.5× TBE and stained with SYBR Gold. The 130 nt fragment region was excised and recovered. The cDNA was resuspended in 15 μL of RNase-free water and subjected to ssDNA circularization. The 20 μL reaction containing 1× CircLigase buffer, 1 μL of 1 mM ATP, 1 μL of 50 mM MnCl2 and 1 μL (100 U) CircLigase(Epicentre) was incubated at 60 °C for 1 h and heat-inactivated at 80 °C for 10 min. Contaminating rRNA fragments were partially removed by subtractive hybridization with biotinylated oligos corresponding to rRNA fragments abundant in RF libraries (101). Briefly, 5 μL of the circularization reaction was combined with 1 μL of 20× SSC, 1 μL of 200 μM subtraction pool and 3 μL of water; denatured at 100 °C for 90 s; and annealed to 37 °C. The biotinylated oligo/rRNA hybrids were recovered with Dynabeads MyOne Streptavidin C1 (Invitrogen) using 0.25 mg beads per sample according to the manufacturer’s instructions. The RFs of the subtracted libraries were precipitated and resuspended in 20 μL of water.  PCR amplification of libraries was performed in an 50 μL reaction of 1× Phusion HF buffer (New England Biolabs), 1 μL of 10 mM (each) dNTPs, 0.5 μL of 50 mM (each) library primers and indexing primers, 0.5 μL of Phusion HF polymerase (New England Biolabs), and 5 μL of rRNA-subtracted circularized template. Amplification was as follows: 30 s at 98°C and 12 cycles of 10s at 98°C, 10s at 60°C, and 5s at 72 °C. PCR products were separated on a 8% (wt/vol) PAGE using 1× Tris-Borate-EDTA. The amplified RF library (∼120-125 bp) was excised and recovered. Poly(A)+ RNA-seq libraries were prepared from 15 µg of DNAse I (Thermo Scientific) treated RNA extracted from the S16 fraction as described by Wang et al., PlosOne 2011

treatment_protocol:
Transgenic Arabidopsis thaliana (Col-0) expressing 35S:His6FLAG-RPL18B line 12-2-4 (background Columbia-0) was used in this study.  Seeds were surface sterilized and sown on a nylon mesh (100 µm, Nitex-Sefar) in square 12 cm square petri plates containing a solid modified Murashige and Skoog (MS/10) medium (0.15 mM MgSO4, 2.1 mM NH4NO3, 1.9 mM KNO3, 0.5 mM NaH2PO4, 0.34 mM CaCl2, 0.5 µM KI, 10 µM FeCl2, 10 µm H3BO3, 10 µM MnSO4, 3 µM ZnSO4, 0.1 µM CuSO4, 0.1 µM CoCl2, 1 µM Na2MoO4, 5.9 µM thiamine, 4.9 µM pyridoxine, 8.1 µM nicotinic acid, 55 µM inositol, and 3.4 mM 2-(N-morpholino)ethanesulfonic acid (MES) [pH 5.7], 0.8% (w/v) agar). For  Pi-deficiency treatment, seedlings were transfered of the same medium where NaCl (0.5 mM) replaced the equivalent amount of sodium provided by NaH2PO4 for 7 days . Growth was in a chamber at 23 °C under a 16 h light (80 μE m-2 sec-1) and 8 h dark cycle
For nutrient treatments, seedlings on the mesh were transferred to new plates with Pi replete (+Pi) MS/10 or Pi-deficient MS/10 (-Pi) medium and grown for 7 d more before the complete root system. All harvested tissue was directly immersed in liquid nitrogen and stored at -80 ˚C for subsequent use.

value: ribosome

value: low P


title: Wild type mouse 2


title: Kdm6a deficient mouse 1

value: Kdm6a deficient


title: Kdm6a deficient mouse 2


title: CEC_like#4_RNAseq

tag: cell type

value: ES cell-derived CEC-like cells

value: 2 months


title: dsRNA_P_Stress

description:
dsRNA
count_ncRNA.txt
count_dsRNA_Bygenes.txt

extract_protocol:
Total RNA extracted using TRIzol  from 100 mg of root tissue
dsRNA sequencing (dsRNA-seq) libraries were prepared as previously described  (Zheng et al., 2010 PLos Biol)  . Briefly, total RNA was subjected to two rounds of rRNA depletion (Ribominus, Invitrogen (Carlsbad, CA)), and then treated with a single-strand specific ribonuclease (RNase One, Promega (Madison, WI)) per manufacturer’s instructions. The RNA samples were then used as the substrate for sequencing library construction using the Small RNA Sample Prep v1.5 kit (Illumina, San Diego, CA) as per manufacturer’s instructions.

value: double stranded RNA


title: aCEC66_RNAseq


title: dsRNA_No_Stress


title: YKWdgd

city: Nanning

first: Wang

last: Yikui

address: north road

country: China

institute: Institute Of Vegetable Research, Guangxi Academy Of Agricultural Sciences

description:
long purple eggplant ‘B3-3’, having two morphologically different types of flowers (termed distylous) on the same plant, which is cultivated mainly in South China
Parallel analysis of RNA ends (PARE)

data_processing:
Raw sequencing reads were obtained using Illumina’s Sequencing Control Studio software version 2.8 (SCS v2.8) following real-time sequencing image analysis and base-calling by Illumina’s Real-Time Analysis version 1.8.70 (RTA v1.8.70).
A proprietary pipeline script, ACGT101-miR v4.2 (LC Sciences, Houston, TX, USA), was used for the analysis of sequencing data (Li et al. 2010; Wei et al. 2011). Unique small RNA sequences were first mapped to known miRNAs in the miRNA and pre-miRNA database (ftp://mirbase.org/pub/mirbase/CURRENT/; Version 21.0). The remaining unique small RNA sequences were mapped to genome of S. melongena in Eggplant Genome DataBase (http://eggplant.kazusa.or.jp/). Potentially novel miRNAs were identified by folding the flanking genome sequence of the unique small RNAs using MIREAP (http://sourceforge.net/projects/mireap/), followed by the prediction of secondary structures by Mfold program (Zuker 2003).
The publicly available CleaveLand 3.0 software package and the ACGT301-DGEv1.0 program (LC Sciences, Houston, TX, USA) were used to detect the potentially sliced targets of the known and novel miRNAs identified by high-throughput sequencing and degradome analysis (Addo-Quaye et al. 2008; Addo-Quaye et al. 2009). The alignments of the identified miRNA sequences to the reference sequence (gene sequences on Solanum melongena L.genome sequence SME_r2.5.1 downloaded from http://compbio.dfci.harvard.edu/tgi/cgi-bin/tgi/gimain.pl?gudb=eggplant)were then scored as follow: mismatched pairs or single nucleotide bulges were each scored as 1 and GU pairs were scored as 0.5. The mismatched and GU pair scores within the core segment (at positions 2-13) were doubled. All targets were allocated into five categories according to the abundance of the resulting mRNA tag relative to the overall profile of the degradome reads matching the target (Addo-Quaye et al. 2008; Addo-Quaye et al. 2009). The optimized score thresholds were set to 4 for all five category to select the resulting output (Feng et al. 2014).
Genome_build: genome of S. melongena in Eggplant Genome DataBase (http://eggplant.kazusa.or.jp/).
Supplementary_files_format_and_content: [miRNA_results.txt] miRNA sequence and expression data.
Supplementary_files_format_and_content: [Table S4.txt] miRNA target prediction results,target position and degradome fragments abundance on cleavage position.

organism: Solanum virginianum

source_name: pooled samples of  flower development 1-3 days after budding and  flower development 3-6 days after budding

growth_protocol: The long purple eggplant ‘B3-3’, having two morphologically different types of flowers (termed distylous) on the same plant, which is cultivated mainly in South China, was used in the present study. In one morph, termed thrum or brevistylous flower (S-morph), the stamens are long and the pistils are short, whereas in the other morph, termed pin or longistylous flower (L-morph), the stamens are short and the pistils are long. The B3-3 line was maintained in the Guangxi Academy of Agricultural Sciences and cultivated in a field using conventional methods. The L-morph and S-morph pistils were collected, respectively, starting from the budding stage to blooming.

extract_protocol:
Total RNA was isolated from each sample using an RNA Extraction Kit (Norgen Biotek Corporation, Thorold, Canada) according to the manufacturer’s instructions.
Small RNA libraries were generated from the mixture of samples using the Illumina TruSeq Small RNA Preparation kit (Illumina, San Diego, USA) according to the manufacturer’s guidelines.
A mixed eggplant pistil degradome library from six samples (S- and L-morph pistils in 1, 2, and 5 phases, respectively) was constructed following the methods described previously by German et al. (2008).

treatment_protocol: The tissues were immediately frozen in liquid N2 after excision and stored at -80°C until the RNA was extracted. We divided the period of the flower development into five phases, 1-3, 3-6, 7-9, 10-12 and 13-15 days after budding, respectively. The pistil samples of S-morph and L-morph flower were prepared; each sample consisted of a mixture of pistils harvested in phase 1 and 2,.

tag: tissue

value: pistil

tag: tissue morph

value: mixture of S- and L-morph pistils


title: Wild type mouse 1


title: Total_P_stress


title: C57BL/6-207_MOCK_PFU0_04d_2

status: Public on May 05 2017

city: La Jolla

first: Max

last: Chang

address: 9500 Gilman Drive

institute: University of California, San Diego

data_processing:
Raw sequencing reads were aligned to the mouse mm10 genome with STAR
Read counts per gene were calculated using FeatureCounts
DESeq2 was used to generate FPKM values
Genome_build: mm10
Supplementary_files_format_and_content: comma-separated values containing FPKM quantification for all samples

source_name: Mouse Lung Tissue

extract_protocol:
Mouse lungs were homogenized in liquid nitrogen. 1 ml Trizol was added per 50-100 mg of tissue and the homogenate was incubated at room temperature for 5 minutes. 200 μl chloroform were added per ml of Trizol, shaken vigorously for 15-30 seconds and incubated at room temperature for 15 minutes. Samples were centrifuged for 15 minutes at 4˚C and 12,000 rcf. The aqueous phase was carefully collected in a separate tube. 500 μl isopropanol was added per 1 ml Trizol, inverted to mix and incubated at room temperature for 5-10 minutes. Samples were transferred to Qiagen RNeasy Spin Columns and RNA was isolated according to the manufacturer’s instructions.
Libraries were constructed using the Illumina TruSeq Stranded Total RNA Library Prep Kit.

treatment_protocol: Mice were anesthetized with an intraperitoneal injection of a mixture of ketamine and xylazine (100 mg/kg and 10 mg/kg, respectively), and then inoculated with 1 × 104 PFU of infectious virus diluted in 50 μl phosphate-buffered saline (PBS) intranasally (i.n.). At 12h, 1d, 2d, 3d and 4d post infection, 3 mice per experimental group were euthanized. Mice inoculated with PBS and euthanized after 4d served as mock controls.

value: Lung

tag: infection

value: uninfected (mock control)

tag: time point

tag: biological replicate


title: C57BL/6-208_MOCK_PFU0_04d_3


title: C57BL/6-202_H5N1_PFU1E4_04d_3

value: A/Vietnam/1203/04 (H5N1) HALo


title: C57BL/6-195_H3N2_PFU1E4_04d_2


title: C57BL/6-201_H5N1_PFU1E4_04d_2


title: C57BL/6-206_MOCK_PFU0_04d_1


title: WYKmiR

description: long purple eggplant ‘B3-3’, having two morphologically different types of flowers (termed distylous) on the same plant, which is cultivated mainly in South China

value: flower development 1-3 days after budding and  flower development 3-6 days after budding


title: C57BL/6-200_H5N1_PFU1E4_04d_1


title: C57BL/6-190_H1N1_PFU1E4_04d_3

value: A/California/04/09 (H1N1)


title: C57BL/6-188_H1N1_PFU1E4_04d_1


title: C57BL/6-194_H3N2_PFU1E4_04d_1


title: C57BL/6-184_H5N1_PFU1E4_03d_3


title: C57BL/6-189_H1N1_PFU1E4_04d_2


title: Kdm6a deficient mouse 3

tag: genotype/variation


title: RS_NoStress

value: control


title: C57BL/6-183_H5N1_PFU1E4_03d_2


title: C57BL/6-178_H3N2_PFU1E4_03d_3


title: C57BL/6-182_H5N1_PFU1E4_03d_1


title: C57BL/6-177_H3N2_PFU1E4_03d_2


title: C57BL/6-176_H3N2_PFU1E4_03d_1


title: C57BL/6-172_H1N1_PFU1E4_03d_3


title: C57BL/6-196_H3N2_PFU1E4_04d_3


title: C57BL/6-160_H3N2_PFU1E4_02d_3


title: C57BL/6-171_H1N1_PFU1E4_03d_2


title: C57BL/6-165_H5N1_PFU1E4_02d_2


title: C57BL/6-170_H1N1_PFU1E4_03d_1


title: C57BL/6-159_H3N2_PFU1E4_02d_2


title: C57BL/6-158_H3N2_PFU1E4_02d_1


title: C57BL/6-166_H5N1_PFU1E4_02d_3


title: C57BL/6-153_H1N1_PFU1E4_02d_2


title: C57BL/6-164_H5N1_PFU1E4_02d_1


title: C57BL/6-154_H1N1_PFU1E4_02d_3


title: C57BL/6-148_H5N1_PFU1E4_01d_3


title: C57BL/6-152_H1N1_PFU1E4_02d_1


title: C57BL/6-146_H5N1_PFU1E4_01d_1


title: C57BL/6-147_H5N1_PFU1E4_01d_2


title: C57BL/6-142_H3N2_PFU1E4_01d_3


title: C57BL/6-141_H3N2_PFU1E4_01d_2


title: C57BL/6-140_H3N2_PFU1E4_01d_1


title: C57BL/6-136_H1N1_PFU1E4_01d_3


title: C57BL/6-134_H1N1_PFU1E4_01d_1


title: C57BL/6-135_H1N1_PFU1E4_01d_2


title: C57BL/6-124_H3N2_PFU1E4_12h_3


title: C57BL/6-129_H5N1_PFU1E4_12h_2


title: C57BL/6-128_H5N1_PFU1E4_12h_1


title: C57BL/6-118_H1N1_PFU1E4_12h_3


title: C57BL/6-123_H3N2_PFU1E4_12h_2


title: C57BL/6-122_H3N2_PFU1E4_12h_1


title: C57BL/6-116_H1N1_PFU1E4_12h_1


title: Neutrophils_WT_rep3

city: Vienna

first: Vitaly

last: Sedlyarov

address: Lazarettgasse 14, AKH BT 25.3

country: Austria

institute: Research Center for Molecular Medicine

data_processing:
Raw RNA-Seq reads were pre-processed (including demultiplexing, barcode, adaptor and quality trimming, quality control using FastQC (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/). Remaining reads were mapped to the GRCm38/mm10 mouse genome assembly using GSNAP (Wu and Watanabe 2005) split mapping resulting in more than 95 % of uniquely mapped reads in each sample. Sample integrity was judged by 5’-3’ coverage plots (SeQC). Transcripts were quantified using the Mix2 RNA-Seq data analysis software (Lexogen). Differential expression analysis was performed using DESeq2 based on read counts in gene models (Love, 2014).
Genome_build: mm10
Supplementary_files_format_and_content: normalized counts

source_name: Peritoneal neutrophils

growth_protocol: Neutrophils were isolated from peritoneum of 8 - 10 week old mice after recruitment by casein injection (16 h prior isolation)

extract_protocol:
Total RNA was isolated using 1 ml TRIzol reagent (Life Technologies) according to the manufacturer’s protocol. Contaminating DNA was removed by digestion with 20 U recombinant DNase I (Roche).
RNA-Seq library was prepared using NEBNext Ultra Directional RNA Library Prep Kit (NEB).

value: neutrophil

tag: tag

value: WT


title: Neutrophils_TTPKO_rep3

value: TTP-/-


title: Neutrophils_TTPKO_rep2


title: Control Rep 2

city: Birmingham

first: Sara

middle: Ann

last: Gibson

state: AL

address: MCLM 313 1918 University Blvd

department: Cell, Developmental and Integrative Biology

institute: University of Alabama at Birmingham

description: Control vs. Treatment.xlsx

data_processing:
Genewiz took the sequenced reads and trimmed to remove possible adapter sequences and nucleotides with poor quality bases (error rate < 0.05) at the ends. After trimming, sequence reads shorter than 30 nucleotides were discarded.
Genewiz took the remaining sequence reads and aligned to the Mus musculsus genome with CLC Genomics Server program. Gene hit counts were measured and RPKM values were calculated.
Genewiz then performed quantile normalization and log2-transformation on the RPKM values. A student t test was performed and p-values, fold-change, and False Discovery Rate (FDR) were calculated
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text file includes RPKM values for each Sample and fold change and p-/q-values.

source_name: Primary CD4+ T cells

growth_protocol: Naïve CD4+ T cells were isolated from murine spleens using the Naïve CD4+ T cell isolation kit from StemCell Technologies and grown for 72 h in RPMI containing 10% FBS for 72 h in Th17 conditions (anti-CD3, anti-CD28, IL-6, IL-23 and TGFb1)

extract_protocol:
Total RNA was extracted using Trizol, quality checked by Nanodrop and 5 mg of each sample sent to Genewiz.
Genewiz took the RNA libraries and prepared them for sequencing using standard Illumina protocols

treatment_protocol: Cells were treated with DMSO (Control) or 2 mM CX-4945 (Treatment).

value: enriched splenic CD4+ T cells

tag: conditions

value: anti-CD3, anti-CD28, IL-6, IL-23, TGFb1

tag: mouse strain

tag: treatment

value: DMSO


title: Treatment Rep 1

description: Control vs. Treatment.xlsx


title: Control Rep 1

description: Control vs. Treatment.xlsx


title: Treatment Rep 3

description: Control vs. Treatment.xlsx


title: Neutrophils_TTPKO_rep1

tag: genotype


title: Treatment Rep 2

description: Control vs. Treatment.xlsx


title: Neutrophils_WT_rep2


title: Control Rep 3

description: Control vs. Treatment.xlsx


title: C57BL/6-130_H5N1_PFU1E4_12h_3

tag: strain


title: Neutrophils_WT_rep1


title: H4_160111_run0456_Mm2h4cR1_male_t2h_DMSO0.0.40pct_rep1

city: Bethesda

first: Brian

last: Oliver

state: MD

address: 50 South Drive

department: LBG

institute: NIDDK, NIH

data_processing:
Basecalling was performed using CASAVA v1.8.2 (Illumina, San Diego, CA). BCL files were converted to FASTQ and demultiplexed using bcl2fastq v1.8 (Illumina, San Diego, CA).
The RNA-seq reads of mixed species were first mapped to a merged genome and annotation including D. grimshawi (dgri_caf1 assembly; Drosophila_grimshawi.GCA_000005155.1.29.dna_sm.genome.fa) D. melanogaster (FlyBase release 6 version 04 ; FlyBase file: dmel-all-chromosome-r6.04.fasta), and A. stephensi (Anopheles-stephensi-Indian_SCAFFOLDS_AsteI2.fa), appended with ERCC sequences (Zook et al., 2012. PMID: 22859977) using STAR (v2.4.2a) (Doblin et al., Bioinformatics, 2012, PMID 23104886). The unmapped reads and reads mapping to D. grimshawi or A. stephensi are not reported in this GEO submission or affiliated SRA entry.
Segregated D. melanogaster reads were mapped to the major chromosome arms (chr2L, chr2R, chr3L, chr3R, chr4, chrX, chrY, and chrM) of Drosophila genome assembly release 6 plus ISO1 MT and ERCC sequences using TopHat v2.1.0 (Trapnell et al., 2009. PMID: 19289445) with default options except for the following: Libraries were stranded so mapping was done with --library-type fr-firststrand, and we provided a GTF with known set of gene models (FlyBase r6.09 and ERCC) using -G option.
The ERCC reference was based on the DNA Sequence Library SRM2374 from NIST, the FASTA sequence was corrected according to differences found in (Jiang et al. 2011, PMID: 21816910). We also used the reverse complement of ERCC-00009, ERCC-00014, ERCC-00057, ERCC-00059, ERCC-00099, ERCC-00108, ERCC-00116 for mapping to correct the strand orientation and ERCC-00085 was re-annotated as Subpool C.
Unique alignments were created by keeping only reads that aligned to a single location (NH:i:1) using grep (v2.20). HTseq v0.6.1p1 (Anders et al., 2014. PMID: 25260700) was used to create gene level read counts with default options except (-s reverse) was used to get strand specific read counts.
Genome_build: Drosophila melanogaster Release 6 plus ISO1 MT (GeneBank assembly accession: GCA_000001215.4)
Supplementary_files_format_and_content: Tab delimited file containing gene level read counts from Htseq.

organism: Drosophila melanogaster

source_name: whole body

growth_protocol: The Drosophila melanogaster w1118 fly stock (FBst0005905) was obtained from the Bloomington Drosophila Stock center (Cook et al., 2010, PMID: 20160480). Flies were grown on Cornmeal, sucrose and yeast media (Agar 10 g/L, Cornmeal 27 g/L, Sugar 43 g/L, Yeast 15 g/L, p-Hydroxy-benzoic acid methyl ester, 2.8 g/ 10.8 ml of Ethanol) obtained from (LabExpress, Ann Arbor, MI) at 25oC at low density (10 female and 5 male parents per vial, transferred to fresh vials daily). After 10 - 11 days post egg laying, adult progeny started to eclose, and flies were removed from the vials and sorted by sex with a fine brush (Robert Simmons, Cranbury, New Jersey. Sapphine S85 or S61), vials were checked every 4 h for newly eclosed flies. We collected 450 virgin w1118 flies per sex, and placed them in batches of 30 per vial with CDF (400kcal/L) for 3 days to age (Lee and Micchelli, 2013, PMID: 23844001).

extract_protocol:
Total RNA isolation followed the RNeasy for spin technology protocol (Qiagen, Valencia, CA). Since these samples were multiplexed with RNA from distantly related species (D. grimshawi and Anopheles stephensi), RNA of the two Drosophila species were mixed with a single Anopheles embryo for repeated bead maceration and a second round of RNA extraction, following the RNeasy protocol as above.
Libraries were prepared using the Truseq PolyA+ stranded mRNA kit (Illumina, San Diego, CA. Cat. No. RS-122-2101 or RS-122-2102), following the manufacturer’s protocol. We used a half reaction size with 100ng total RNA as input for all samples. External RNA Control Consortium (ERCC) spike-ins transcribed from a certified plasmid collection (SRM2374) were added to the Fragment-Prime-Finish Mix during the mRNA fragmentation step.  We used the previously described RNAs from Pool 78A for DMSO treated flies and 78B for control flies (20pg /sample)  (Zook et al., 2012. PMID: 22859977).  Libraries were multiplexed using 24 differently barcoded adapters from TruSeq (Illumina, San Diego, CA). Libraries were quantified with Quant-iT PicoGreen (Life Technologies, Carlsbad, CA), and pooled for multiplexed sequencing. We sequenced in an Illumina HiSeq2500 (Illumina, San Diego, CA.)

treatment_protocol: Three day post-eclosion flies were anesthetized with CO2 and manually transferred to individual wells in the Whole Animal Feeding Flat (WAFFL, US Application No. 62/080,181, PCT Application No. PCT/US2015/060575) with a fine brush. The top half of the plate (rows A-D) contained females (n= 48) and the bottom half of the plate (rows E-H) contained males (n= 48). One WAFFL was used per time point and each food plate contained all tested concentrations of DMSO. The WAFFLs were sealed using AXYMAT AM-2ML-RD silicone mats (Axygen,Tewksbury MA). Flies were allowed to recover from the anesthetic for at least 3 h before being fed with 8 ul of CDF liquid (400kcal/L) for 24 h provided on a round bottomed No.3799 96-well plate (Costar, Corning, NY), then flies were starved for 6 h by replacing the food with a 96 well plate containing 8 ul of PBS pH 7.4 (Gibco, Life technologies, Grand Island, NY). After starvation, the food plate was change to CDF liquid with 0, 0.1, 0.2, or 0.4% (v/v) DMSO and Sulforhodamine B 0.5 mg/ml (Sigma, St Louis, MO. Cat. No. D2650 and S1402). Flies were allowed to feed for 0, 2, 4 or 8 h. We visually examined flies for a bolus of dyed food in the fly abdomen. A saturated K2SO4 solution (Sigma Aldrich, St Louis, MO) was used to maintain humidity between 95 - 97 % RH in an enclosed chamber which contained the WAFFLs. Relative humidity and temperature of 25oC was measured with a traceable hygrometer (VWR, Randor, PA).                                                                                At each specific end point, the WAFFL was removed from the food plate, blotted on a paper towel to remove leftover food from the bottom and placed at -20oC for 20 min. Once flies were immobilized, the WAFFL was opened and attached to the transfer adapter and receiver plate and then flies were transferred to the receiver plate by centrifugation. The receiver plate then was attached to 96 deep well plate (Axygen, Tewksbury MA. Cat. No. P-DW-11-C-S) then transferred by centrifugation. RLT buffer 600 ul (Qiagen, Valencia, CA) and 100 ul of 1.0 mm dia glass beads (BioSpec Products, Bartlesville, OK) were added to each well, and wells were sealed with two layers of plastic (Applied Biosystems, Warrington, UK). Flies were macerated on MiniBead beater for 1 min x 2 resting on ice (Biospec Products, Bartlesville, OK) and stored at -80oC until RNA extraction. From the treated macerated flies we randomly selected 3 replicates per concentration (n=4), time point (n= 4) and sex (n=2) for a total of 96 samples to be used for preparing libraries.

value: Male

tag: time (h)

tag: dmso % v/v

value: 3 day old adult

tag: fly line


title: C57BL/6-117_H1N1_PFU1E4_12h_2


title: H2_160111_run0456_Mm0h4cR2_male_t0h_DMSO0.0.40pct_rep2

tag: Sex

tag: developmental stage


title: H12_160111_run0456_Mm8h4cR3_male_t8h_DMSO0.0.40pct_rep3


title: H9_160111_run0456_Mm4h4cR3_male_t4h_DMSO0.0.40pct_rep3


title: H7_160111_run0456_Mm4h4cR1_male_t4h_DMSO0.0.40pct_rep1


title: H8_160111_run0456_Mm4h4cR2_male_t4h_DMSO0.0.40pct_rep2


title: H6_160111_run0456_Mm2h4cR3_male_t2h_DMSO0.0.40pct_rep3


title: H10_160111_run0456_Mm8h4cR1_male_t8h_DMSO0.0.40pct_rep1


title: H1_160111_run0456_Mm0h4cR1_male_t0h_DMSO0.0.40pct_rep1


title: H11_160111_run0456_Mm8h4cR2_male_t8h_DMSO0.0.40pct_rep2


title: G6_160111_run0456_Mm2h2cR3_male_t2h_DMSO0.0.20pct_rep3


title: G7_160111_run0456_Mm4h2cR1_male_t4h_DMSO0.0.20pct_rep1


title: G5_160111_run0456_Mm2h2cR2_male_t2h_DMSO0.0.20pct_rep2


title: G4_160111_run0456_Mm2h2cR1_male_t2h_DMSO0.0.20pct_rep1


title: H3_160111_run0456_Mm0h4cR3_male_t0h_DMSO0.0.40pct_rep3


title: G3_160111_run0456_Mm0h2cR3_male_t0h_DMSO0.0.20pct_rep3


title: G9_160111_run0456_Mm4h2cR3_male_t4h_DMSO0.0.20pct_rep3