title: 3565_GSC__INHEG_sg4_rep2

description:
3565sg4-2R_L1_1
3565sg4VS3565NT_rna_fpkm1.txt

data_processing:
For ribometh-seq, sdapters were removed by cutadapt v2.7 with parameters (-a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A GATCGTCGGACTGTAGAACTCTGAAC). Trimmed reads were mapped to ribosome DNA sequence by bowtie v2.2.9 and then sorted by samtools v1.10. BAM files were converted to BED format by bamToBed (bedtools v2.26). Positions of 5' (beginnings) and 3' (ends) were extracted and converted to bedGraph using custom python script. The methylation level was further quantified by MethScore C2 calculation.
For RNA-seq, reads were aligned and counted by STAR with --quantMode GeneCounts option and differential expressed genes were identified by edgeR.
For ribosome profiling, adapter sequences were removed from raw sequencing data using cutadapt software. Meanwhile, reads with length between 25 and 35 bp were kept for downstream analysis. Then reads were aligned to rRNA and tRNA sequences so as to remove rRNA and tRNA reads using bowtie software, remaining reads were used to align to reference genome and transcriptome (Ensembl Version 91) using hisat2 and bowtie software separately. Read counts were calculated using featureCounts software. Raw counts were further normalized as RPKM values using fpkm function in edgeR package.
Assembly: NOP58_KO RNA-seq: hg19; INHEG-OE RNA-seq, Ribo-seq: hg38
Supplementary files format and content: for RNA-seq, the count table is the read counts for each sample calcualted from STAR with --quantMode GeneCounts mode. For ribometh-seq, methScore are provided for each nucleotide at each sample.

value: patient-derived glioma stem cell


title: Sample.11.Hector's Dolphin.205128010033_R04C01

status: Public on Sep 28 2023

type: genomic

city: Newport

first: Keith

last: Hernandez

state: OR

address: 2030 SE Marine Science Drive

department: Marine Mammal Institute

country: USA

institute: Oregon State University

hyb_protocol: Bisulphite converted DNA was amplified, fragmented and hybridized to the Beadchip HorvathMammalMethylChip40 using standard Illumina protocol

scan_protocol: Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner settings

data_processing: SeSaME normalization as described in W. Zhou et al. 2018. PMID: 30085201

molecule: genomic DNA

organism: Cephalorhynchus hectori hectori

source_name: Skin

label_protocol: Standard Illumina Protocol

growth_protocol: DNA extracted from tissues

extract_protocol: Genomic DNA was extracted from skin using a phenol-chloroform protocol modified for small skin samples as described in C. S. Baker et al. 1994. PMID: 7921358

treatment_protocol: Standard Illumina Protocol

tag: tissue

value: Skin

tag: basename

tag: species common name

value: Hector's Dolphin

tag: female

tag: canbeusedforagingstudies

value: Yes


title: pool CHP134 cells treated with CX-5461 rep1

type: SRA

city: Memphis

first: William

middle: C.

last: Wright

state: Tennessee

address: 262 Danny Thomas Pl

department: Computational Biology

institute: St. Jude Children's Research Hospital

library_source: other

data_processing:
CRISPR KO screens were analyzed using Mageck-Vispr/0.5.7
Supplementary files format and content: Mageck default output describing resulting RRA scores

molecule: other

organism: Homo sapiens

source_name: Peripheral Nervous System

extract_protocol: sgRNAs for the human CRISPR KO library were first designed using CRISPick . The top 30 sgRNAs for each gene then underwent an additional round of filtering using in-house off-target analysis to identify highly unique sgRNAs.  Up to 6 sgRNAs per gene were selected for the library along with non-targeting controls making up ~10% of the final library.

treatment_protocol: 72H treatments

value: Peripheral Nervous System

tag: cell line

tag: cell type

value: Neuroblastoma

tag: genotype

tag: treatment

tag: molecule type

value: gRNA


title: Sample.12.Hector's Dolphin.205138660001_R03C02


title: MOE, replicate 1, snRNAseq

city: New York

first: Ariel

last: Pourmorady

state: NY

address: 3227 Broadway

department: Neurobiology and Behavior

institute: Columbia University

description: 10x Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the Cell Ranger software v2.1.1 (https://support.10xgenomics.com/single-cell-gene-expression/software/pipelines/latest/what-is-cell-ranger)
Assembly: mm10. "genes.gtf" file was used for snRNA alignment
Supplementary files format and content: Tab-separated values files and matrix files

molecule: nuclear RNA

organism: Mus musculus

source_name: main olfactory epithelium (MOE)

extract_protocol:
Nuclei must be purified under RNA clean conditions. A cell suspension of mouse MOE was obtained from an adult mouse following the dissociation conditions described in Monahan et al. 2017. Cells pellet were immediately resuspended in 300-uL of cold RNAse free lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% IGEPAL CA-630), and nuclei were pelleted in a the Eppendorf 5810R Centrifuge at 1000-g, for 10-min at 4-˚C. Nuclei were resuspended in 500-uL of Homogenization Buffer (10x HB: 100-mM Trizma Base, 800 mM KCl, 100 mM EDTA, 10 mM spermidine trihydrochloride, 10 mM spermidine tetrahydrochloride in ddH2O, pH to 9-9.4 with NaOH, instructions for preparation of 1x HB can be found in Zhang et al. 2-12 50) with RNAse inhibitor (NEB MO314L), followed by 500-uL 82% Optiprep solution (4.1-mL Optiprep solution (Sigma Aldrich D1556-250ML), 25-uL 1M CaCl2, 15-uL 1M MgAc, 50-uL 1M Tris pH 8, 810-uL Water) then placed on ice. The 1-mL homogenate was carefully added onto 1-mL of 48% Optiprep solution (2.4-mL Optiprep solution, 800-uL 1M Sucrose, 25-uL 1M CaCl2, 15-uL 1M MgAc, 50-uL 1M Tris pH 8, 1710-uL Water), and was spun down in a pre-cooled swinging bucket centrifuge (Eppendorf 5810R) at 3200-g, for 20-min, at 4-˚C, acceleration 5/9, deceleration 0/9 (no break). The supernatant was aspirated and disposed, without dislodging the pellet. Air dry pellet and resuspend in 500-μL 1 x PBS diluted with 0.04% BSA with RNAse inhibitor. Cell concentration was measured for accurate loading into the 10x pipeline.
Joint scRNA- and scATAC-seq libraries were prepared in collaboration with the Columbia Genome Center using the 10x Genomics Single Cell Multiome ATAC + Gene Expression kit according to manufacturer’s instructions. Both GEX and ATAC libraries were sequenced to ~350M reads on an Illumina NextSeq550 100PE and Illumina NovaSeq 6000150PE, respectively.

value: main olfactory epithelium (MOE)

tag: age

value: adult


title: Cortex_FD_Untreated_5906

city: Boston

first: Susan

last: Slaugenhaupt

state: MA

address: 185 Cambridge St

department: Center for Genomic Medicine

institute: Massachusetts General Hospital

library_source: transcriptomic

data_processing:
RNASeq reads were mapped against a custom referenceby STAR (version 2.5.22b) allowing 5% mismatch
Sequence reads were trimmed for adaptor sequence/low-quality sequence using Trimmomatic (v0.36)
Trimmed sequence reads were mapped to GrCM38/mm10 using CLC genomic benchwork (parameters- mismath cost: 2 insertion cost: 3 deletion cost: 3 length fraction: 0.8 similarity fraction: 0.8)
Read count extraction and normalization were performed using HTSeq(v0.11.2) and DESeq2 (v1.34.0).
Assembly: human ELP1 gene (ENSG00000070061) Ensemble Human transcriptome reference GRCh37.75 to the Ensembl Mouse Transcriptome GRCm38.83
Supplementary files format and content: Supplementary Table S1 - DEG_stats.txt

molecule: total RNA

source_name: Cortex

growth_protocol: We developed a mouse model that faithfully recapitulates the the FD phenotype seen in patients using the TgFD9 hypomorphic mouse and human transgene.

extract_protocol:
RNA was harvested using QIAzol Reagent as per manufacturrers instructions. 100 ng of total RNA was used for the construction of sequencing libraries.
Tru-Seq Stranded® mRNA library Prep Kit (Illumina, 20020594) following manufacturer's protocols.

value: Cortex

value: Familial Dysautonomia (FD)


title: 10x_Zou_2_Spleen

city: Houston

first: Dawei

last: Zou

state: TX

address: 6670 Bertner Ave

department: Surgery

institute: Houston Methodist Research Institute

description: 10x Genomics

data_processing:
Raw Illumina sequencing reads were aligned to reference mouse genome mm10 (Ensembl 93) using Cell Ranger v7.0.0 with default parameters, generating one Fastq dataset for cells from heart and one for cells from spleen. Genes were quantified as UMI counts using Cell Ranger count and initially visualized using Loupe Browser v5.0.0.
Downstream analysis was performed using Seurat V4.0.0 on filtered feature counts generated by Cell Ranger count. Low-quality single cells containing >2.5% mitochondrial transcripts or >50% ribosomal transcripts were removed. Additionally, genes expressed in fewer than three single cells were removed. We identified potential single-cell doublets using DoubletFinder V2.0.3, with an expectation of a 7.5% doublet rate assuming Poisson statistics.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: from spleen

extract_protocol:
The adoptively transferred TEa cells were stained with fluorochrome-conjugated antibodies, and sorted from the spleens and heart allografts of the recipients by a cell sorter
Single cell Gene Expression Library was prepared according to Chromium Next GEM Single Cell 3' Reagent Kits v3 (10x Genomics). In Brief, single nuclei suspension, reverse transcription (RT) reagents, Gel Beads containing barcoded oligonucleotides, and oil were loaded on a Chromium controller (10x Genomics) to generate single cell GEMS (Gel Beads-In-Emulsions) where full length cDNA was synthesized and barcoded for each single cell. Subsequently the GEMS are broken and cDNA from each single cell are pooled. Following cleanup using Dynabeads MyOne Silane Beads, cDNA is amplified by PCR.  The amplified product is fragmented to optimal size before end-repair, A-tailing, and adaptor ligation. Final library was generated by amplification.

value: from spleen

value: CD4+ T cells


title: 10x_Zou_1_Heart

description: 10x Genomics

source_name: from heart allograft

value: from heart allograft


title: Sample.10.Hector's Dolphin.205128010011_R01C02


title: 3565_GSC__INHEG_sg4_rep1

city: Beijing

first: Lihui

last: Liu

address: 15 Datun Road, Chaoyang District

department: Institute of Biophysics

country: China

institute: Chinese Academy of Sciences

description:
3565sg4-1R_L1_1
3565sg4VS3565NT_rna_fpkm1.txt

source_name: glioma stem cells

extract_protocol:
Cell culture medium were removed and RNA was harvested using Trizol reagent. For RiboMeth-seq, total RNA was fragmented in 50 mM sodium carbonate/bicarbonate buffer (pH 9.2) at 95°C. Next, Fragmented RNA was 3′-end dephosphorylated and purified with the RNA Clean & Concentrator-5 kit (Zymo Research). RNA was then 5′-end phosphorylated with T4 polynucleotide kinase (NEB) and purified again. For uvRIP-seq, RNA were harvested with Trizol reagent after NOP58 UV-crosslinked immunoprecipitaion.
For RiboMeth-seq, sequencing libraries were prepared with the VAHTSTM Small RNA Library Prep Kit for Illumina® (Vazyme) following the manufacturer's instruction. For RNA-seq, RNA libraries were prepared for sequencing using VAHTSTM Total RNA-seq (H/M/R) Library Prep Kit for Illumina (Vazyme) with total RNA. Ribo-seq: Ribosome profiling was performed using Ribosome Profiling Kit.  RPFs  was extracted using RNA clean&ConcentratorTM-5 kit.     EpiTM RiboRNA Depletion Kit  was used for rRNA depletion.  Sequencing libraries were constructed using QIAseq miRNA Library kit.


title: DRG_Control_Untreated_5372

source_name: Dorsal Root Ganglion

value: Dorsal Root Ganglion

value: WT


title: DRG_Control_Untreated_5375


title: 3565_GSC__INHEG_sg2_rep2

description:
3565sg2-2R_L1_1
3565sg2VS3565NT_rna_fpkm1.txt


title: 3565_GSC__INHEG_sg2_rep1

description:
3565sg2-1R_L1_1
3565sg2VS3565NT_rna_fpkm1.txt


title: gg8tTAtetOP2, control HiC, r3

description: Control Hi-C libraries performed on an individual biological replicate of GFP+ OSNs sorted from adult mice

library_source: genomic

data_processing:
Reads were aligned to the mm10 genome using the distiller pipeline (https://github.com/mirnylab/distiller-nf), uniquely mapped reads (mapq > 30) were retained and duplicate reads discarded. Contacts were then binned into matrices using cooler 45. Analysis was performed on data pooled from two (HiChIP, Hi-C) or three biological replicates (Liquid Hi-C), after confirming that the results of analysis of individual replicates were similar.
Assembly: mm10
Supplementary files format and content: Genomic Contacts binned at 50-kb resolution
Library strategy: Hi-C/Liquid Hi-C/HiChIP

source_name: Olfactory Sensory Neurons

extract_protocol:
MOE were dissociated using the Worthington Papain Dissociation kit, and cells were fixed in 1% formaldehyde in 1x PBS either before or after sorting for GFP+ cells. For Liquid Hi-C, nuclei were digested with Dpn-II for various lengths of time, on a thermomixer shaking at 900 rpm at 37˚C, placed on ice for 10-minutes, then fixed in 1% formaldehyde in 1x PBS. After cross-linking, nuclei were processed for HiC exactly as described in "LHX2- and LDB1-mediated trans interactions regulate olfactory receptor choice" (Horta et al. 2019). Briefly nuclei were lysed, permeabilized, quenched, and then digested over night in DpnII . The next day we performed biotin-fill in with biotinylated dUTP, ligation with T4, and shearing. Cross-links were reversed overnight and DNA was purified with a 2x AMPure bead purification. Library prep was performed on MyOne Streptavidin beads to purify biotinylated contacts.
The HiChIP protocol was kindly given by the Chang Lab and integrated into our Hi-C protocol to perform H3K27ac HiChIP on olfactory sensory neurons. MOE from 5-7 gg8-tTA>tetO-P2 mice were dissociated to obtain 4M GFP+ cells per replicate, for a total of 2 replicates. Cells were processed according to the in situ Hi-C protocol with the following exceptions, nuclei were digested for only 2 hours instead of overnight; complete nuclei digestion was verified by running reverse cross-linked digested nuclei on a DNA Agarose gel. After ligation, nuclei were pelleted at 2500-g, for 5-min, at 4-˚C and stored overnight at -20 ˚C. The next day, nuclei were resuspended in 130-uL of HiChIP nuclear lysis buffer (50 mM Tris pH 7.5, 10 mM EDTA, 1% SDS, 1X Protease Inhibitor in water), and sheared on a Covaris S220 with the following parameters: duty cycle – 2%, PIP – 140, cycles/burst – 200, time – 4 minutes. After shearing, samples were precleared and immunoprecipitation was performed with H3K27ac antibody (Abcam GR323193701),
Libraries were prepared using the Ovation® Ultralow V2 DNA-Seq Library Preparation Kit for HiC and Liquid HiC. For HiChIP, libraries were prepared using on-bead based tagmentation with Tn5 as described in "HiChIP: Efficient and sensitive analysis of protein-directed genome architecture" (Mumbach et al. 2016).

value: gg8-tTA >tetO-P2

value: Olfactory Sensory Neurons


title: Cortex_FD_Untreated_5905


title: Sample.14.Hector's Dolphin.205128010010_R06C02

value: Skin


title: 3565_GSC_EGFP_rep2

description:
3565NT-2R_L1_1
3565sg2VS3565NT_rna_fpkm1.txt 3565sg4VS3565NT_rna_fpkm1.txt

value: patient-derived glioma stem cell


title: Cortex_FD_Untreated_5901


title: Prostate WT

city: St Louis

first: Tiandao

last: Li

state: MO

address: 4444 Forest Park Ave

institute: Washington University

data_processing:
Raw fastq files were aligned against human genome hg38 assembly using Burrows-Wheeler Alignment tool and then processed using methylQA to generate bed and bigwig files.
The MACS2 peak caller was used to compare the ChIP-Seq signal to a corresponding input control to identify narrow regions of enrichment (peaks) using default parameters.
ChIP-Seq signals and peak locations were further visualized using UCSC Genome Browser.
The top 500-1000 ChIP-Seq peaks were assigned to the nearest genes using the annotatePeaks function from HOMER2, motif binding sites were determined using HOMer2, and GO and pathway enrichment analyses were performed by EnrichR.
Assembly: hg38, mm10
Supplementary files format and content: fastq, narrowPeak

source_name: Mouse prostate cells

growth_protocol: LNCaP cells were grown in DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics in humidified atmosphere with 5% CO2 at 37 degree C

extract_protocol:
The LNCaP and prostate ysates were immunoprecipitation with p54-H3 antibodies (2 ug) for 3 hours.
Ten nanograms of immunoprecipitated DNA was fragmented to 300 base pairs using a Covaris M220 Focused-ultrasonicator (Covaris, Inc., Woburn, MA) and then used to generate sequencing libraries using the Kapa Hyper Prep Kit (Roche Sequencing Solutions Inc., Pleasanton, CA). The size and quality of the library was evaluated using the Agilent BioAnalyzer (Agilent Technologies, Inc., Santa Clara, CA), and the library was quantitated with the Kapa Library Quantification Kit.  Each enriched DNA library was then sequenced on an Illumina NextSeq 500 sequencer to generate 40-50 million 75-base paired-end reads (Illumina, Inc., San Diego, CA).  The raw sequence data were aligned using BowTie 2 [1], and the binding sites were identified using the MACS peak-finding software

treatment_protocol: 5x10^7 LNCaP cells were treated with ACK1 (R-9b) and SHP2 (SHP099) inhibitors overnight (0.5 uM) and lysates were prepared by sonication in RLB bufffer.

value: Mouse prostate cells

tag: chip antibody


title: Prostate KO


title: Prostate Shp2i treated


title: 3565_GSC_EGFP_rep1

description:
3565NT-1R_L1_1
3565sg2VS3565NT_rna_fpkm1.txt 3565sg4VS3565NT_rna_fpkm1.txt


title: Cortex_FD_Untreated_5881


title: MGG4_GSC__NOP58_sg2_rep2

description:
MGG4NOP58-2-2_L4_M030
NOP58RNAseq.count.txt


title: Cortex_FD_Untreated_5877


title: Prostate 9b treated


title: Sample.7.Hector's Dolphin.205128010033_R02C01

value: Hector's Dolphin

value: Yes


title: Sample.4.Maui Dolphin.205128010011_R06C01

organism: Cephalorhynchus hectori maui

value: Māui Dolphin

value: Maui Dolphin


title: Sample.9.Hector's Dolphin.205128010011_R02C02


title: Sample.8.Hector's Dolphin.205128010033_R03C01


title: Prostate Input


title: Sample.5.Hector's Dolphin.205166520009_R05C01


title: MGG4_GSC__NOP58_sg2_rep1

description:
MGG4NOP58-2-1_L4_M028
NOP58RNAseq.count.txt


title: Sample.6.Hector's Dolphin.205138660001_R02C02


title: Cortex_FD_Untreated_5822


title: lilp, NK, biol replicate, 4

city: Rome

first: Mattia

last: Laffranchi

address: Viale Regina Elena 291

department: Molecular Medicine

country: Italy

institute: La Sapienza University of Rome

data_processing:
Illumina run was demultiplexed and converted to FastQ using bcl2fastq v2.20.0.422
Read 1 was mapped to mm10 using tophat 2.1.1
RPKM values were generated using Partek GS 7.0
ANOVA comparisons were calculated using Partek GS 7.0
Assembly: mm10
Supplementary files format and content: 210705_211108_sciume_rna-seq_results.gene.rpkm.GEO.xlsx (RPKM table)
Supplementary files format and content: 210705_211108_sciume_rna-seq_results.gene.7-samples.ANOVA.GEO.xlsx (ANOVA results)

source_name: large intestine

extract_protocol:
RNA was isolated by using single cell purification kit (Norgen, #51800) using RNase-Free DNase (Norgen #25710).
Sequencing libraries generated using SMARTer® Stranded Total RNA-Seq Kit v2 (Takara Bio). Libraries were sequenced on an Illumina Novaseq 6000 as paired end, 50 base reads (by DNA Link lnc).

tag: strain

value: large intestine

value: NK cells

value: DSS

tag: time

value: Day 7


title: MGG4_GSC__NOP58_sg1_rep2

description:
MGG4NOP58-1-2_L4_M011
NOP58RNAseq.count.txt


title: MGG4_GSC__NOP58_sg1_rep1

description:
MGG4NOP58-1-1_L3_M006
NOP58RNAseq.count.txt


title: Sample.3.Hector's Dolphin.205138660001_R05C02


title: Hordeum vulgare embryonic tissue total RNA-seq r1

city: Pullman

first: Sascha

last: Duttke

state: Washington

address: BLS 202, 100 Dairy Road

department: School of Molecular Biosciences

institute: Washington State University

description:
total RNA-seq (Ribo0), steady state transcriptome profiling in H.vulgare cells;
H.vulgare-10.tss.txt

data_processing:
csRNA-seq (small capped RNAs, ~20-60 nt) and sRNA-seq reads (“input”) were trimmed of their adapter sequences using HOMER (“homerTools trim -3 AGATCGGAAGAGCACACGTCT -mis 2 -minMatchLength 4 -min 20" none -f {csRNA_fastq_path}/*fastq.gz”)(Heinz et al. 2010). Reads were aligned to the appropriate genome using Hisat2 version 2.2.1 (Kim et. al. 2019) with parameters “hisat2 -p 30 --rna-strandness RF --dta -x {hisat2_genome_index} -U {path_rimmed_csRNA or sRNA} -S {output_sam} 2> {mapping_stats}”. Hisat2 indices were then created for each genome used ("hisat2-build -p 40 genome.dna.toplevel.fa {Hisat2_indexfolder}") with exception for barely which required the addition of "--large-index" due to genome size. Tag directory folders were then created using HOMER makeTagDirectories ("makeTagDirectories {outputName} -omitSN -checkGC -fragLength 150 -single")(Heinz et. al. 2010). Only reads with a single, unique alignment (MAPQ >=10) were considered in the downstream analysis. This consolidates the aligned read information into a read/chromosome format along with other important alignment information. Peaks were called and annotated using HOMER ("findcsRNATSS.pl {csRNA} -o {output} -i {sRNA} -genome hg38 -size 150 -ntagThreshold 10") (Duttke et al. 2019) and annotatePeaks.pl ("annotatePeaks.pl {tss/tsr.txt} hg38 -strand + -fragLength 1 -rlog -d {listOfTagDirs} > {output}") (Heinz et. al. 2010) respectively (additional details available in the manuscript).
Transcription start sites (TSSs) and Transcription start regions (TSRs) were determined in this study. TSRs  were called using findcsRNATSS.pl (detailed above) while TSSs were called using getTSSfromReads.pl ("getTSSfromReads.pl -d {csRNA_tagdir} -dinput {sRNA_tagdir} -min 7 > {output_file}") (Heinz et. al. 2010). To ensure high quality TSSs, at least 7 per 10^7 aligned reads were required and TSS were required to be within called TSR regions.
Paired end total ribosomal RNA-depeleted RNA-seq libraries were trimmed using skewer (“time -p skewer -m mp {read1} {read2} -t 40 -o {trimmed_fastq_output}”) (Jiang et al. 2014) and aligned using Hisat2 (Kim et al. 2019) to ensure all data were processed as similar as possible (“hisat2 -p 30 --rna-strandness RF --dta -x {hisat2_index} -1 {trimmed_RNAseq_R1} -2 {trimmed_RNAseq_R2} -S {output_sam} 2> {mapping_file}”). In this manuscript, total RNA-seq was exclusively used to determine RNA-stability as described in the csRNA-seq analysis.
5’GRO-seq peaks were called using HOMER’s “findPeaks {5GRO_tagdirectory} -i {GRO_tagdirectory} -style tss -F 3 -P 1 -L 2 -LP 1 -size 150 -minDist 200 -ntagThreshold 10 > 5GRO_TSRs.txt”. Detailed explanation of each parameter can be found at http://homer.ucsd.edu/homer/ngs/tss/index.html.
Assembly: TAIR10, Papaya1.0, Chlamydomonas_reinhardtii_v5.5, Drosophila melanogaster: dm6, ASM32608v1, Phypa V3, Selaginella moellendroffii: v1.0, Zea mays (rest): Zm-B73-REFERENCE-NAM-5.0, Zea mays : AGPv4.38 (STARR-seq)
Supplementary files format and content: bed files: Locations indicate TSS positions. TSS.txt files: text files containing tab delimited  information about found transcription start site culsters (TSRs). Motif file containing new line delimited motifs. Motif files: files containing motifs used with a ‘>’ indicating motif information followed by rows with nucleotide probability.

organism: Hordeum vulgare

growth_protocol: A.thaliana Col-0 mature leaves were collected from plants grown as described (M. Wang et al. 2023), while A.thaliana Col-0 suspension cells were grown in 250-mL baffled flasks containing 50 mL of growth medium (3.2 g/L Gamborg’s B-5 medium, 3 mM MES, 3% [v/v] Suc, 1.1 mg L−1 2,4-dichlorophenoxyacetic acid). The cultures were maintained at 23°C under continuous light on a rotary shaker (160 rpm) and kindly provided as a frozen pellet by Dr. Ashley M. Brooks. Barley (Hordeum vulgare) RNA was isolated by Dr. Pete Hedley from embryonic tissue (including mesocotyl and seminal roots; EMB) isolated from grain tissues 4 days past germination (Mascher et al. 2017). Physcomitrium (Physcomitrella) patens (Gransden) was grown on plates with BCDA medium in a growth cabinet at 21°C under 16h light. Selaginella moellendorffii was purchased from Plant Delights Nursery and grown at the window under normal daylight for 1 week prior to isolating RNA from stems and leaves. C. reinhardtii, which was kindly provided by Dr. Will Ansari and Dr. Stephen Mayfield (UC San Diego), was grown to late logarithmic phase in TAP (Tris–acetate–phosphate) medium at 23°C under constant illumination of 5000 lux on a rotary shaker. Adult 2nd and 3rd leaves from Z. mays L. cultivar B73 was kindly provided by Dr. Lauri Smith (UC San Diego). Plants were grown in 4-inch pots in a greenhouse (temp: 23°C-29°C) without supplemental lighting or humidification (humidity in the 15 hours following inoculation ranged between 70 and 90%) year round in La Jolla, CA. RNA from Z. mays L. cultivar B73 7d old shoot, root and leaves were grown in the Schmitz laboratory (University of Georgia) as described in (Ricci et al. 2019).

extract_protocol: csRNA-seq was performed as described in (S. H. Duttke et al. 2019). Small RNAs of ~20-60 nt were size selected from 0.4-3 µg of total RNA by denaturing gel electrophoresis. A 10% input sample was taken aside and the remainder enriched for 5’-capped RNAs. Monophosphorylated RNAs were selectively degraded by 1 hour incubation with Terminator 5´-Phosphate-Dependent Exonuclease (Lucigen). Subsequently, RNAs were 5’dephosporylated through 90 minutes incubation in total with thermostable QuickCIP (NEB) in which the samples were briefly heated to 75°C and quickly chilled on ice at the 60 minutes mark. Input (sRNA) and csRNA-seq libraries were prepared as described in (Hetzel et al. 2016) using RppH (NEB) and the NEBNext Small RNA Library Prep kit, amplified for 11-14 cycles.  Total RNA-Seq Library Preparation  Strand-specific, paired-end libraries were prepared from total RNA by ribosomal depletion using the Ribo-Zero Gold plant rRNA removal kit (Illumina, San Diego, CA). Samples were processed following the manufacturer’s instructions.


title: RNA_pH4.4_3

city: Freiburg

first: Rick

last: Gelhausen

state: Baden Württemberg

address: Georges-Köhler-Allee 106

department: Department of Computer Science

country: Germany

institute: Albert-Ludwigs-University Freiburg

data_processing:
Basecalling
Adapter sequence "AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC" was trimmed using cutadapt 2.1 (HRIBO 1.6.0)
Mapping with segemehl 0.3.4 (HRIBO 1.6.0)
rRNA filtering with samtools 1.9 (HRIBO 1.6.0)
Coverage file creation with HRIBO 1.6.0
wig files were generated with full read or single nucleotide mapping
wig files were normalized using the Counts Per Million (CPM) method and converted to bigwig format
Assembly: ASM584v2
Supplementary files format and content: bigwig format files containing CPM normalized read coverage for each sample (global mapping)

organism: Escherichia coli str. K-12 substr. MG1655

source_name: bacterial cell

growth_protocol: Cells were inoculated to a starting OD600 of 0.05 from overnight cultures and aerobically grown to exponential phase in 200 ml of unbuffered LB media (pH 7.6) in 1 liter glas flasks. After reaching an OD600 of 0.5, cultures were either grown for 30 further min at pH 7.6 under non-stress conditions, 30 min at pH 5.8, or first 15 min at pH 5.8 and then shifted again for 15 min to pH 4.4

extract_protocol:
RNA was harvested using the miRNeasy Mini Kit (QIAGEN) in combination with the RNase-Free DNase Set (QIAGEN) following manufacturer's protocols. Ribosomal RNA depletion was performed using the NEBNext rRNA Depletion Kit for bacteria (NEB) following manufacturer's protocols. Integrity of RNA samples was evaluated by using the RNA 6000 Nano Kit (Agilent).
cDNA libraries were prepared using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (NEB) following manufacturer's protocols. cDNA library quality was assessed by using a High Sensitivity DNA Kit (Agilent).

treatment_protocol: Once OD600 = 0.5 was reached, cells were grown further at pH 7.6, or 5M hydrochloric acid was directly added to growing cultutes to adjust pH values to either pH 5.8, or pH 4.4.

value: bacterial cell

tag: rna fraction

value: mRNA


title: Cortex_FD_Untreated_5753

value: Cortex

value: Familial Dysautonomia (FD)


title: Cortex_FD_Untreated_5441


title: Sample.1.Hector's Dolphin.205166520007_R05C02


title: MGG4_GSC_NT_rep2

description:
MGG4NT-2_L3_M012
NOP58RNAseq.count.txt


title: Sample.2.Hector's Dolphin.205128010011_R04C02


title: RNA_pH5.8_3

value: WT


title: MGG4_GSC_NT_rep1

description:
MGG4NT-1_L4_M005
NOP58RNAseq.count.txt


title: 3565_GSC__NOP58_sg2_rep2

description:
B3565NOP582-2_L2_M023
NOP58RNAseq.count.txt


title: Cortex_FD_Untreated_5378


title: lilp, NK, biol replicate, 3


title: RNA_pH7.6_3


title: lilp, NK, biol replicate, 1


title: Cortex_FD_Untreated_5376


title: scRNA_Mouse_NCRposILC_WT_and_STA4_KO

description:
BD Rhapsody™ Single-Cell Analysis System
mouse large intestine was minced and lamina propria cells were isolated after tissue digestion by using DNAseI and Liberase TL.

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the SevenBridges platform with the pipeline BD WTA Multiplex Rhapsody Analysis Pipeline Version 1.9.1 (https://www.sevenbridges.com/)
Supplementary files format and content: mm10
Supplementary files format and content: Comma-separated values files

extract_protocol:
RNA was extracted according to the manufacter’s instructions (BD Rhapsody™ Single-Cell Analysis System ).
Library was performed according to the manufacter’s instructions (BD Rhapsody™ Single-Cell Analysis System ).

value: Primary

value: Innate lymphpocytes

value: 6-12 weeks old


title: Cortex_FD_Untreated_5374


title: Splenic, NK, biol replicate, 3

source_name: spleen

value: spleen

value: no treatment

value: Day 0


title: Cortex_FD_Untreated_5373


title: RNA_pH5.8_2


title: RNA_pH4.4_2


title: Splenic, NK, biol replicate, 1


title: Cortex_FD_Untreated_5780


title: Splenic, NK, biol replicate, 2


title: lilp, NK, biol replicate, 2


title: 3565_GSC__NOP58_sg1_rep1

description:
B3565NOP58-1_L2_M006
NOP58RNAseq.count.txt

value: patient-derived glioma stem cell


title: 3565_GSC__NOP58_sg1_rep2

description:
B3565NOP58-2_L2_M010
NOP58RNAseq.count.txt


title: Cortex_Control_Untreated_5907


title: RNA_pH7.6_2


title: RNA_amPA-M_rep5

city: Oxford

first: Pawel

last: Swietach

address: Sherrington Building, Parks Road

department: Department of Physiology, Anatomy & Genetics

country: United Kingdom

institute: University of Oxford

data_processing:
RNA-seq reads were aligned to the mm10 genome using STAR
Duplicate reads were removed using samtools rmdup
Read counts were determined using featureCounts (Subread package)
Differential gene expression analysis was conducted using DESeq2
Assembly: mm10

molecule: polyA RNA

source_name: polyA RNA-seq in Pcca-/- male 8-week-old mouse heart

growth_protocol: Mice were bred and housed in individually ventilated cages and all animals had ad libitum access to water and food (Teklad #2918 global 18% protein rodent diet). Mice used were either wild-types (Pcca+/+ A138T; amWT) for control experiments, or homozygous (Pcca-/- A138T; amPA). Only heterozygous or wild-type dams were used for breeding and genotyping by PCR was performed.

extract_protocol:
Ventricular tissue from 8-week mice was snap-frozen and cryoground. Bulk RNA was extracted using the Zymo Quick-RNA Miniprep Plus Kit (R1057), following the manufacturer’s protocol with some modifications in the tissue digestion stage. All centrifugation steps were performed at 16,000 RCF at 4°C. 20 mg of cryoground tissue was mechanically homogenised on ice with disposable probes (Qiagen TissueRuptor II) in 350 µL of DNA/RNA Shield. 45 µL of Proteinase K mix was added to 300 µL of sample, incubated for 30 mins at 55°C, and centrifuged for 2 mins after incubation. The supernatant was transferred to new microcentrifuge tubes and 300 µL RNA Lysis Buffer was added at 1:1 to the supernatant. Thereafter, the RNA purification procedure followed the manufacturer’s protocol, with some differences: half volume ethanol (300 µL) was added, in order to isolate RNAs ≥ 200 nt for poly(A) enrichment; DNase I treatment was performed in-column; RNA was eluted in non-DEPC-treated nuclease-free water.
PolyA RNA was purified using the NEBNext Poly(A) mRNA magnetic isolation module (E7490) and DNA libraries were prepared using the NEBNext Ultra II Directional RNA library preparation kit for Illumina (E7765).

treatment_protocol: To harvest tissues, 8-week-old mice were killed humanely by i.p. injection of an appropriate volume adjusted for body weight of sodium pentobarbitone (stock 200 mg/ml) pre-mixed at a 2:1 ratio with sodium heparin (stock 5,000 I.U./mL).

value: heart

value: FVB Pcca-/- A138T mutation


title: RNA_pH4.4_1


title: Cortex_Control_Untreated_5902


title: RNA_amPA-M_rep4


title: 3565_GSC_NT_rep1

description:
B3565NT-1_L4_M005
NOP58RNAseq.count.txt


title: RNA_pH7.6_1


title: RNA_pH5.8_1


title: Cortex_Control_Untreated_5880


title: MGG4_GSC_RiboMeth

description:
MGG4-GSC
DGC.GSC.MethScoreC2.txt

data_processing:
For ribometh-seq, sdapters were removed by cutadapt v2.7 with parameters (-a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A GATCGTCGGACTGTAGAACTCTGAAC). Trimmed reads were mapped to ribosome DNA sequence by bowtie v2.2.9 and then sorted by samtools v1.10. BAM files were converted to BED format by bamToBed (bedtools v2.26). Positions of 5' (beginnings) and 3' (ends) were extracted and converted to bedGraph using custom python script. The methylation level was further quantified by MethScore C2 calculation.
For RNA-seq, reads were aligned and counted by STAR with --quantMode GeneCounts option and differential expressed genes were identified by edgeR.
For ribosome profiling, adapter sequences were removed from raw sequencing data using cutadapt software. Meanwhile, reads with length between 25 and 35 bp were kept for downstream analysis. Then reads were aligned to rRNA and tRNA sequences so as to remove rRNA and tRNA reads using bowtie software, remaining reads were used to align to reference genome and transcriptome (Ensembl Version 91) using hisat2 and bowtie software separately. Read counts were calculated using featureCounts software. Raw counts were further normalized as RPKM values using fpkm function in edgeR package.
Assembly: NOP58_KO RNA-seq: hg19; INHEG-OE RNA-seq, Ribo-seq: hg38
Supplementary files format and content: for RNA-seq, the count table is the read counts for each sample calcualted from STAR with --quantMode GeneCounts mode. For ribometh-seq, methScore are provided for each nucleotide at each sample.
Library strategy: RiboMeth-seq


title: Cortex_Control_Untreated_5900


title: RNA_amPA-M_rep3


title: RNA_amPA-M_rep2


title: Cortex_Control_Untreated_5875


title: Cortex_Control_Untreated_5821


title: G2_miRNA-seq

city: Huaibei

first: Mengmeng

state: Anhui

address: Dongshan Street

department: College of Life Science

institute: Huaibei Normal University

data_processing:
The remaining high-quality reads were assembled de novo using Trinity software.
The TPM algorithm in Salmon software was used to measure the relative abundance of transcripts in each group of samples.
Differentially expressed genes (DEGs) in the samples were screened by DESeq, q-value <0.05 and log2(FoldChange) >1 were used as screening criteria to determine the significance of gene expression.
Assembly: no genome build/assembly, Trinity-assembled transcript sequences provided in a FASTA file
Supplementary files format and content: TPM or RPM

organism: Pinellia ternata

source_name: leaf, heat-treated, 24h

growth_protocol: The tubers were planted in pots and placed in a constant temperature and light incubator with a maintained temperature of 25 ℃, a light intensity of 4000 lx, and a photoperiod of 16 hours of light followed by 8 hours of darkness.

extract_protocol:
Trizol reagent (Sangon Biotech, Shanghai, China) was used for the extraction of total RNA from tissue samples. Six samples (C and G, each with three biological replicates) were harvested for small RNA library construction and sequencing.
Equal amounts of RNA from each sample were sequenced on an Illumina HiSeq2500 sequencer (Illumina, San Diego, CA, USA) at the Sangon Biotech company (Shanghai China), to construct the strand-specific RNA-seq. 1.0 μg of total RNA was used to prepare a small RNA library according to the instruction of TruSeq Small RNA Sample Prep Kits.

treatment_protocol: The selected seedlings were subjected to a temperature of 40 ℃ for either 0 or 24 hours while keeping all other conditions the same as the control group.

value: leaf

value: Pinellia ternata leaf cells

value: heat-treated

value: Day 1


title: RNA_amWT-M_rep4

source_name: polyA RNA-seq in wild-type male 8-week-old mouse heart

value: wild-type


title: RNA_amPA-M_rep1


title: MGG4_DGC_RiboMeth

description:
MGG4-DGC
DGC.GSC.MethScoreC2.txt

source_name: differentiated glioma cells

value: patient-derived differentiated glioma cell


title: G3_miRNA-seq


title: 3565_GSC_NT_rep2

description:
B3565NT-2_L2_M011
NOP58RNAseq.count.txt


title: G1_miRNA-seq


title: RNA_amWT-M_rep5


title: LNCaP SHP2i

source_name: LNCaP

value: LNCaP

value: Human prostate cancer


title: C3_miRNA-seq

source_name: normal


title: Cortex_Control_Untreated_5776

value: Cortex

value: WT


title: 3565_GSC_RiboMeth

description:
3565-GSC
DGC.GSC.MethScoreC2.txt


title: C2_miRNA-seq


title: 3565_DGC_RiboMeth

description:
3565-DGC
DGC.GSC.MethScoreC2.txt


title: Cortex_Control_Untreated_5779


title: RNA_amWT-M_rep2


title: RNA_amWT-M_rep3


title: G3_RNA-seq


title: LNCaP Ctrl


title: Cortex_Control_Untreated_5377


title: RNA_amWT-M_rep1


title: MGG4_GSC__FBL_sg4_rep2

description:
MGG4-sgFBL4-2_L2_A006
MGG4.rawcount.txt

data_processing:
For ribometh-seq, sdapters were removed by cutadapt v2.7 with parameters (-a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A GATCGTCGGACTGTAGAACTCTGAAC). Trimmed reads were mapped to ribosome DNA sequence by bowtie v2.2.9 and then sorted by samtools v1.10. BAM files were converted to BED format by bamToBed (bedtools v2.26). Positions of 5' (beginnings) and 3' (ends) were extracted and converted to bedGraph using custom python script. The methylation level was further quantified by MethScore A2 calculation.
For RNA-seq, reads were aligned and counted by STAR with --quantMode GeneCounts option and differential expressed genes were identified by edgeR.
For uvRIP-seq, read were aligned by STAR and counted by featureCounts, then enriched transcripts by NOP58 antibody relative to input or IgG control were identified by DESeq2.
Genome_build: hg19
Supplementary_files_format_and_content: For RNA-seq, the count table is the read counts for each sample calcualted from STAR with --quantMode GeneCounts mode. For ribometh-seq, methScore are provided for each nucleotide at each sample. For uvRIP-seq, the count tables are the read counts for each sample  calcualted from featureCounts.

extract_protocol:
Cell culture medium were removed and RNA was harvested using Trizol reagent. For RiboMeth-seq, total RNA was fragmented in 50 mM sodium carbonate/bicarbonate buffer (pH 9.2) at 95°C. Next, Fragmented RNA was 3′-end dephosphorylated and purified with the RNA Clean & Concentrator-5 kit (Zymo Research). RNA was then 5′-end phosphorylated with T4 polynucleotide kinase (NEB) and purified again. For uvRIP-seq, RNA were harvested with Trizol reagent after NOP58 UV-crosslinked immunoprecipitaion.
For RiboMeth-seq, sequencing libraries were prepared with the VAHTSTM Small RNA Library Prep Kit for Illumina® (Vazyme) following the manufacturer's instruction. For RNA-seq and uvRIP-seq, RNA libraries were prepared for sequencing using VAHTSTM Total RNA-seq (H/M/R) Library Prep Kit for Illumina (Vazyme) with total RNA.


title: RNA_amPA-F_rep5

source_name: polyA RNA-seq in Pcca-/- female 8-week-old mouse heart


title: G2_RNA-seq


title: MGG4_GSC__FBL_sg4_rep1

description:
MGG4-sgFBL4-1_L1_A005
MGG4.rawcount.txt


title: LNCaP 9b


title: C3_RNA-seq

source_name: leaf, normal


title: G1_RNA-seq


title: RNA_amPA-F_rep3


title: RNA_amPA-F_rep4


title: Flp-In HEK293 RBMX OEKD

city: Newcastle upon Tyne

first: Graham

last: Smith

address: Framlington Place

department: Bioinformatics Support Unit

institute: Newcastle University

data_processing:
Samples were aligned to human genome GRCh38 using hisat2 v2.2.1
merged bam files were produced with samtools v1.14 and examined in IGV
Assembly: GRCh38

source_name: kidney of a human embryo

growth_protocol: HEK293 cells were grown in DMEM, high glucose, pyruvate supplemented with 10% FBS, and maintained at 37°C with 5% CO2.

extract_protocol: RNA was extracted from cells using RNeasy Plus Mini Kit (Qiagen #74134) following manufacturer’s instructions

treatment_protocol: Cells were seeded onto 6-well plates forward transfected with Lipofectamine™ RNAiMAX transfection reagent (Invitrogen, # 13778150) according to manufacturer′s instructions using 30 pmol of either control siRNA or RBMX siRNA for 72h at 37°C before harvesting. Half the samples were treated with 1µg/ml Tetracycline to induce expression of RBMXL2-FLAG.

value: kidney of a human embryo

value: Flp-In HEK293

value: epithelial

value: RBMX siRNA + RBMXL2 overexpression


title: MGG4_GSC__FBL_sg1_rep2

description:
MGG4-sgFBL1-2_L2_A004
MGG4.rawcount.txt

value: patient-derived glioma stem cell


title: Cortex_Control_Untreated_5752

source_name: Cortex


title: RNA_amPA-F_rep2

value: heart


title: C2_RNA-seq

value: Day 0


title: MGG4_GSC__FBL_sg1_rep1

description:
MGG4-sgFBL1-1_L2_A003
MGG4.rawcount.txt

source_name: glioma stem cells


title: RNA_amPA-F_rep1

value: FVB Pcca-/- A138T mutation


title: Flp-In HEK293 RBMX OEC1

value: control siRNA + RBMXL2 overexpression


title: C1_RNA-seq

value: leaf

value: Pinellia ternata leaf cells

value: heat-treated


title: Cortex_Control_Untreated_5375


title: MGG4_GSC_ctrl_rep2

description:
MGG4-sgNT-2_L3_A002
MGG4.rawcount.txt


title: hR3

description:
RBMX_TPM.txt
RBMX_isoformTPM.txt

data_processing:
Quality assessment with FastQC Version 0.11.2
(samples 7-8, batch 1) Trimming - trimmomatic (v0.32). ILLUMINACLIP:TruSeq3-PE.fa:2:30:10 LEADING:12 TRAILING:1
(samples 7-8, batch 1) Reads were aligned to genome with STAR version 2.5.2b
(samples 7-8, batch 1) Gene counts were generated with HTSEQ-count. -f bam -r name -s reverse -a 4 -i gene_id -m union, using annotations from RefSeq
(samples 7-8, batch 1) Splicing changes were assessed with MAJIQ
(samples 1-6, batch 2) Reads were aligned to genome with STAR version 2.4.2a
(samples 1-6, batch 2) Reads were aligned to transcriptome with Salmon version 0.9.1 using annotations version 90 (GRCh38.90) from Ensembl
(samples 1-6, batch 2) Gene and transcript level abundances were made from the Salmon quant.sf files using tximport version 1.12.3 in R
(samples 1-6, batch 2) Splicing changes were assessed with SUPPA2
Genome_build: GRCh38 (primary assembly)
Supplementary_files_format_and_content: tab delimited text containing transcript level TPM abundances. Counts are not normalized
Supplementary_files_format_and_content: tab delimited text containing gene-level counts. Counts are not normalized

source_name: Breast cancer cell line

growth_protocol: MDA-MB-231 cells were maintained in DMEM (no phenol red) plus 10% fetal bovine serum and 1% penicillin Streptomycin.

extract_protocol:
RNA was extracted from cells using RNeasy Plus Mini Kit (Qiagen) following manufacturer’s instructions and re-suspended in nuclease-free water. RNA samples were DNase treated using DNA-free kit (Invitrogen).
Illumina's stranded mRNA kit
RNA-Seq 75 bp Paired-end reads

treatment_protocol: MDA-MB 231 cells were treated with an siRNA specific for RBMX, or a control siRNA, for 72 hours.

value: MDA-MB-231 cells

value: depleted for RBMX by siRNA

tag: batch


title: MGG4_GSC_ctrl_rep1

description:
MGG4-sgNT-1_L3_A001
MGG4.rawcount.txt


title: C1_miRNA-seq


title: Carica papaya young leaves sRNA-seq r1

description:
sRNA-seq; small RNA mapping in C.papaya cells; Experiment SD1069
C.papaya-10.tss.txt

organism: Carica papaya

source_name: young leaves

value: young leaves


title: RNA_amWT-F_rep5

source_name: polyA RNA-seq in wild-type female 8-week-old mouse heart

value: wild-type


title: 3565_GSC__FBL_sg4_rep1

description:
B3565sgFBL4-1_L3_A005
B3565.rawcount.txt


title: Flp-In HEK293 RBMX C1

value: control siRNA


title: 3565_GSC__FBL_sg4_rep2

description:
B3565sgFBL4-2_L2_A006
B3565.rawcount.txt


title: RNA_amWT-F_rep4


title: Carica papaya young leaves csRNA-seq r2

description:
csRNA-seq; nascent TSS mapping in C.papaya cells; Experiment SD22
C.papaya-10.tss.txt

data_processing:
csRNA-seq (small capped RNAs, ~20-60 nt) and sRNA-seq reads (“input”) were trimmed of their adapter sequences using HOMER (“homerTools trim -3 AGATCGGAAGAGCACACGTCT -mis 2 -minMatchLength 4 -min 20" none -f {csRNA_fastq_path}/*fastq.gz”)(Heinz et al. 2010). Reads were aligned to the appropriate genome using Hisat2 version 2.2.1 (Kim et. al. 2019) with parameters “hisat2 -p 30 --rna-strandness RF --dta -x {hisat2_genome_index} -U {path_rimmed_csRNA or sRNA} -S {output_sam} 2> {mapping_stats}”. Hisat2 indices were then created for each genome used ("hisat2-build -p 40 genome.dna.toplevel.fa {Hisat2_indexfolder}") with exception for barely which required the addition of "--large-index" due to genome size. Tag directory folders were then created using HOMER makeTagDirectories ("makeTagDirectories {outputName} -omitSN -checkGC -fragLength 150 -single")(Heinz et. al. 2010). Only reads with a single, unique alignment (MAPQ >=10) were considered in the downstream analysis. This consolidates the aligned read information into a read/chromosome format along with other important alignment information. Peaks were called and annotated using HOMER ("findcsRNATSS.pl {csRNA} -o {output} -i {sRNA} -genome hg38 -size 150 -ntagThreshold 10") (Duttke et al. 2019) and annotatePeaks.pl ("annotatePeaks.pl {tss/tsr.txt} hg38 -strand + -fragLength 1 -rlog -d {listOfTagDirs} > {output}") (Heinz et. al. 2010) respectively (additional details available in the manuscript).
Transcription start sites (TSSs) and Transcription start regions (TSRs) were determined in this study. TSRs  were called using findcsRNATSS.pl (detailed above) while TSSs were called using getTSSfromReads.pl ("getTSSfromReads.pl -d {csRNA_tagdir} -dinput {sRNA_tagdir} -min 7 > {output_file}") (Heinz et. al. 2010). To ensure high quality TSSs, at least 7 per 10^7 aligned reads were required and TSS were required to be within called TSR regions.
Paired end total ribosomal RNA-depeleted RNA-seq libraries were trimmed using skewer (“time -p skewer -m mp {read1} {read2} -t 40 -o {trimmed_fastq_output}”) (Jiang et al. 2014) and aligned using Hisat2 (Kim et al. 2019) to ensure all data were processed as similar as possible (“hisat2 -p 30 --rna-strandness RF --dta -x {hisat2_index} -1 {trimmed_RNAseq_R1} -2 {trimmed_RNAseq_R2} -S {output_sam} 2> {mapping_file}”). In this manuscript, total RNA-seq was exclusively used to determine RNA-stability as described in the csRNA-seq analysis.
5’GRO-seq peaks were called using HOMER’s “findPeaks {5GRO_tagdirectory} -i {GRO_tagdirectory} -style tss -F 3 -P 1 -L 2 -LP 1 -size 150 -minDist 200 -ntagThreshold 10 > 5GRO_TSRs.txt”. Detailed explanation of each parameter can be found at http://homer.ucsd.edu/homer/ngs/tss/index.html.
Assembly: TAIR10, Papaya1.0, Chlamydomonas_reinhardtii_v5.5, Drosophila melanogaster: dm6, ASM32608v1, Phypa V3, Selaginella moellendroffii: v1.0, Zea mays (rest): Zm-B73-REFERENCE-NAM-5.0, Zea mays : AGPv4.38 (STARR-seq)
Supplementary files format and content: bed files: Locations indicate TSS positions. TSS.txt files: text files containing tab delimited  information about found transcription start site culsters (TSRs). Motif file containing new line delimited motifs. Motif files: files containing motifs used with a ‘>’ indicating motif information followed by rows with nucleotide probability.
Library strategy: csRNA-Seq


title: Cortex_Control_Untreated_5372

value: Cortex

value: WT


title: hR2

description:
RBMX_TPM.txt
RBMX_isoformTPM.txt


title: Flp-In HEK293 RBMX KD

value: kidney of a human embryo

value: Flp-In HEK293

value: epithelial

value: RBMX siRNA


title: Carica papaya young leaves csRNA-seq r1

description:
csRNA-seq; nascent TSS mapping in C.papaya cells; Experiment SD1008
C.papaya-10.tss.txt


title: RNA_amWT-F_rep2

value: heart


title: RNA_amWT-F_rep3


title: 3565_GSC__FBL_sg1_rep2

description:
B3565sgFBL1-2_L4_A004
B3565.rawcount.txt

value: patient-derived glioma stem cell


title: msil2/4 stem sample Rep2

city: Perpignan

first: Thierry

last: Lagrange

address: 58 Avenue Paul Alduy

country: France

institute: CNRS

data_processing:
Filtering out of reads corresponding to chloroplastic, mitochondrial and rRNA genes was performed with Bowtie 2 v2.3.4.
Remaining reads were mapped against the TAIR 10 genome using gtf annotation file and default parameters of TopHat2 v2.0.7.
Assembly and transcript quantification were performed with Cufflinks v2.2.1.
Low expressed transcripts - less than 1 RPKM (reads per kilobase per million mapped reads) in one of the libraries were filtered out.
The differential analysis was conducted by Cuffdiff software, belonging to the Trapnell suite.
Assembly: Arabidopsis Thaliana TAIR 10 genome
Supplementary files format and content: Xls files including RPKM read count values for each Sample

organism: Arabidopsis thaliana

source_name: inflorescence stem

growth_protocol: Arabidopsis Wild-type and msil2/4 double mutant plants were grown in long day light condition in growth chambers.

extract_protocol:
Total RNAs were purified using Rneasy plant mini plus kit (Qiagen).
The mRNAs were purified using the NEBNext® Poly(A) mRNA Magnetic Isolation Module
The RNA-seq libraries were prepared using the NEBNext® Ultra™ II Directional RNA Library Prep Kit.  Single-end reads, 1x75bp and 30x106 reads per library.

treatment_protocol: No particular treatment

value: inflorescence stem

value: MSIL2/4 knockdown


title: RNA_amWT-F_rep1


title: hR1

description:
RBMX_TPM.txt
RBMX_isoformTPM.txt


title: msil2/4 stem sample Rep1


title: 3565_GSC__FBL_sg1_rep1

description:
B3565sgFBL1-1_L2_A003
B3565.rawcount.txt


title: 3565_GSC_ctrl_rep2

description:
B3565sgNT-2_L4_A002
B3565.rawcount.txt


title: 3565_GSC_ctrl_rep1

description:
B3565sgNT-1_L2_A001
B3565.rawcount.txt


title: hC3

description:
RBMX_TPM.txt
RBMX_isoformTPM.txt


title: Wild-type stem sample Rep2


title: Wild-type stem sample Rep1


title: hC1

description:
RBMX_TPM.txt
RBMX_isoformTPM.txt


title: Hordeum vulgare embryonic tissue csRNA-seq r1

description:
csRNA-seq; nascent TSS mapping in H.vulgare cells; Experiment SD887
H.vulgare-10.tss.txt

source_name: embryonic tissue (including mesocotyl and seminal roots; EMB)


title: TXN58: [ileum, biological replicate 9]

status: Public on Sep 27 2023

city: Corvallis

first: Adrian

middle: F

last: Gombart

address: 307 Linus Pauling Science Center

department: Biochemistry & Biophysics

description: single-end, two raw files were generated from two separate Illumina runs

data_processing:
Two reads from separate Illumina runs were concatenated into one.
Concatenated reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, using bbduk with parameters k=13 ktrim=r useshortkmers=t mink=5 qtrim=r trimq=20 minlength=20
Sequences then mapped to GRCm38 using STAR version 37.95
BAM files were then converted to raw counts and genes were annotated using Rsamtools (v.2.6.0), GenomicFeatures (1.42.1), GenomicAlignments (v.1.26.0), Mus.musculus (v.1.3.1) in R.
Genes whose names contain "ribosomal protein", "RIKEN", "predicted gene", "pseudogene", "cDNA sequence", "nuclear encoded rRNA", "microRNA",  "DNA segment", "uncharacterized", "pre-RNA", "20S rRNA",  "non-protein coding" were removed.
Assembly: GRCm38
Supplementary files format and content: Matrix table with raw gene counts for every gene and every sample

source_name: ileum

extract_protocol:
Ileum was removed, flash frozen in liquid N2 and stored at -80C until day of RNA isolation; total RNA was harvested using Direct-zol RNA Miniprep Plus kit (Zymo Research).
RNA purity and integrity were evaluated using a Bioanalyzer RNA 6000 Nano chip (Agilent Technologies). Lexogen QuantSeq 3'mRNA-Seq Library Prep Kit-FWD for Illumina sequencing was used with 325 ng of total RNA for the construction of sequencing libraries.
Library preparation was started by oligo(dT) priming, with primers already containing the Illumina-compatible linker sequence for Read 2. After firststrand synthesis, the RNA was removed before random primers that contained the corresponding Illumina-compatible linker sequence for Read 1 initiated the second-strand synthesis.
Second strand synthesis was followed by a magnetic bead-based purification step. The libraries were PCR amplified introducing sequences required for cluster generation and i7 and i5 dual indices (Lexogen i7 6 nt Index Set and Lexogen i5 6 nt Unique Dual Indexing Add-on Kit) for 16-20 PCR cycles with the optimal number predetermined by qPCR with the PCR Add-on Kit for Illumina (Lexogen GmbH). After a second magnetic bead-based purification, libraries were quantified using the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific) and sized using an Agilent High Sensitive D5000 Screen Tape (Agilent Technologies) to determine molarity. Equal molar amounts of the libraries were multiplexed and then sequenced on an Illumina Hiseq3000 platform (Illumina, San Diego, CA, USA) at the Center for Genome Research and Biocomputing, Oregon State University using single-end sequencing with 100-bp reads.

value: ileum

tag: Sex

value: male

value: 26 weeks

value: HFD+TXN


title: TXN57: [ileum, biological replicate 8]

tag: tissue


title: TXN56: [ileum, biological replicate 7]


title: TXN55: [ileum, biological replicate 6]


title: TXN52: [ileum, biological replicate 3]


title: hC2

description:
RBMX_TPM.txt
RBMX_isoformTPM.txt

value: MDA-MB-231 cells


title: HXN45: [ileum, biological replicate 9]

value: HFD+HXN


title: TXN49: [ileum, biological replicate 1]


title: HXN46: [ileum, biological replicate 10]


title: HXN44: [ileum, biological replicate 8]


title: Hordeum vulgare embryonic tissue sRNA-seq r1

description:
sRNA-seq; small RNA mapping in H.vulgare cells; Experiment SD945
H.vulgare-10.tss.txt

value: embryonic tissue (including mesocotyl and seminal roots; EMB)


title: HXN42: [ileum, biological replicate 6]


title: HXN43: [ileum, biological replicate 7]


title: HXN37: [ileum, biological replicate 1]


title: HXN39: [ileum, biological replicate 3]


title: HXN38: [ileum, biological replicate 2]


title: HXN41: [ileum, biological replicate 5]


title: TXN53: [ileum, biological replicate 4]


title: HFD20: [ileum, biological replicate 8]

value: HFD


title: HFD17: [ileum, biological replicate 5]


title: HFD19: [ileum, biological replicate 7]


title: TXN51: [ileum, biological replicate 2]


title: TXN54: [ileum, biological replicate 5]


title: HFD15: [ileum, biological replicate 3]


title: HFD21: [ileum, biological replicate 9]


title: HXN40: [ileum, biological replicate 4]


title: HFD16: [ileum, biological replicate 4]


title: HFD18: [ileum, biological replicate 6]


title: HFD13: [ileum, biological replicate 1]


title: LFD10: [ileum, biological replicate 10]

value: LFD


title: LFD09: [ileum, biological replicate 9]


title: HFD14: [ileum, biological replicate 2]


title: LFD08: [ileum, biological replicate 8]


title: LFD06: [ileum, biological replicate 6]


title: LFD05: [ileum, biological replicate 5]


title: HFD22: [ileum, biological replicate 10]


title: LFD04: [ileum, biological replicate 4]


title: LFD07: [ileum, biological replicate 7]


title: LFD02: [ileum, biological replicate 2]


title: LFD01: [ileum, biological replicate 1]


title: neutrophil PBS rep1

city: Manhasset

first: Max

last: Brenner

address: 350 Community Dr

institute: Feinstein Institutes for Medical Research

description: 10x Genomics

data_processing:
The Cell Ranger count pipeline (v6.0.0, 10X Genomics) was used to align FASTQs to the mouse reference genome (gex-mm10-2020-A, 10X Genomics) and produce digital gene-cell counts matrices
Assembly: gex-mm10-2020-A
Supplementary files format and content: h5 files processed from cellranger pipeline

source_name: bone marrow

extract_protocol:
single-cell RNA sequencing (scRNA-seq) using the 10X Genomics Chromium platform
Library preparation was conducted according to the recommended protocol for the NextGEM Single-Cell 3’ Library Kit v3.1 (#1000121; 10X Genomics).

treatment_protocol: PBS or 1 µg/mL recombinant mouse CIRP (eCIRP) for 2 h

value: bone marrow

value: primary cells

value: neutrophil

value: PBS


title: TXN58: [epididymal white adipose, biological replicate 9]

source_name: epididymal white adipose

extract_protocol:
eWAT was removed, flash frozen in liquid N2 and stored at -80C until day of RNA isolation; total RNA was harvested using Direct-zol RNA Miniprep Plus kit (Zymo Research).
RNA purity and integrity were evaluated using a Bioanalyzer RNA 6000 Nano chip (Agilent Technologies). Lexogen QuantSeq 3'mRNA-Seq Library Prep Kit-FWD for Illumina sequencing was used with 325 ng of total RNA for the construction of sequencing libraries.
Library preparation was started by oligo(dT) priming, with primers already containing the Illumina-compatible linker sequence for Read 2. After firststrand synthesis, the RNA was removed before random primers that contained the corresponding Illumina-compatible linker sequence for Read 1 initiated the second-strand synthesis.
Second strand synthesis was followed by a magnetic bead-based purification step. The libraries were PCR amplified introducing sequences required for cluster generation and i7 and i5 dual indices (Lexogen i7 6 nt Index Set and Lexogen i5 6 nt Unique Dual Indexing Add-on Kit) for 16-20 PCR cycles with the optimal number predetermined by qPCR with the PCR Add-on Kit for Illumina (Lexogen GmbH). After a second magnetic bead-based purification, libraries were quantified using the Qubit dsDNA HS Assay Kit (Thermo Fisher Scientific) and sized using an Agilent High Sensitive D5000 Screen Tape (Agilent Technologies) to determine molarity. Equal molar amounts of the libraries were multiplexed and then sequenced on an Illumina Hiseq3000 platform (Illumina, San Diego, CA, USA) at the Center for Genome Research and Biocomputing, Oregon State University using single-end sequencing with 100-bp reads.

value: epididymal white adipose


title: neutrophil rmCIRP rep1

description: 10x Genomics

value: rmCIRP


title: TXN49: [epididymal white adipose, biological replicate 1]


title: TXN53: [epididymal white adipose, biological replicate 4]


title: TXN56: [epididymal white adipose, biological replicate 7]


title: TXN54: [epididymal white adipose, biological replicate 5]


title: 3565_GSC__NOP58_sg2_rep1

description:
B3565NOP582-1_L2_M012
NOP58RNAseq.count.txt

data_processing:
For ribometh-seq, sdapters were removed by cutadapt v2.7 with parameters (-a AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC -A GATCGTCGGACTGTAGAACTCTGAAC). Trimmed reads were mapped to ribosome DNA sequence by bowtie v2.2.9 and then sorted by samtools v1.10. BAM files were converted to BED format by bamToBed (bedtools v2.26). Positions of 5' (beginnings) and 3' (ends) were extracted and converted to bedGraph using custom python script. The methylation level was further quantified by MethScore C2 calculation.
For RNA-seq, reads were aligned and counted by STAR with --quantMode GeneCounts option and differential expressed genes were identified by edgeR.
For ribosome profiling, adapter sequences were removed from raw sequencing data using cutadapt software. Meanwhile, reads with length between 25 and 35 bp were kept for downstream analysis. Then reads were aligned to rRNA and tRNA sequences so as to remove rRNA and tRNA reads using bowtie software, remaining reads were used to align to reference genome and transcriptome (Ensembl Version 91) using hisat2 and bowtie software separately. Read counts were calculated using featureCounts software. Raw counts were further normalized as RPKM values using fpkm function in edgeR package.
Assembly: NOP58_KO RNA-seq: hg19; INHEG-OE RNA-seq, Ribo-seq: hg38
Supplementary files format and content: for RNA-seq, the count table is the read counts for each sample calcualted from STAR with --quantMode GeneCounts mode. For ribometh-seq, methScore are provided for each nucleotide at each sample.

value: patient-derived glioma stem cell


title: TXN51: [epididymal white adipose, biological replicate 2]


title: HXN44: [epididymal white adipose, biological replicate 8]


title: TXN57: [epididymal white adipose, biological replicate 8]


title: TXN55: [epididymal white adipose, biological replicate 6]


title: HXN43: [epididymal white adipose, biological replicate 7]


title: LFD03: [ileum, biological replicate 3]


title: TXN52: [epididymal white adipose, biological replicate 3]


title: HXN42: [epididymal white adipose, biological replicate 6]


title: HXN40: [epididymal white adipose, biological replicate 4]


title: HXN45: [epididymal white adipose, biological replicate 9]


title: HXN46: [epididymal white adipose, biological replicate 10]


title: HXN39: [epididymal white adipose, biological replicate 3]


title: 3 days resected spleen, biological rep 3

type: RNA

city: Moscow

first: Polina

last: Vishnyakova

address: Oparina 4

country: Russia

institute: National Medical Research Center for Obstetrics, Gynecology and Perinatology

description: Gene expression data from spleen 3 days after resection

hyb_protocol: Following fragmentation, 10 ug of cRNA were hybridized for 16 hr at 45C on GeneChip™ Human Gene 2.0 ST Array. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: GeneChips were scanned using the Affimetrix GeneChip Scanner 3000 7g

data_processing: The data were analyzed with Transcriptome Analysis Console (TAC) Software  version 4.0.2 using Affymetrix default analysis settings and RMA algorithm

label: biotin

source_name: spleen RNA

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

growth_protocol: Spleen samples were collected, frozen, lysed in QIAZOL reagent for the following RNA extraction

extract_protocol: Ordinary extraction of total RNA was performed according to the manufacturer's instructions (Qiagen).

treatment_protocol: spleen samples were cellected on different time periods after 70% liver resection/without resection (intact)

value: spleen

value: spleen 3 days after resection


title: 3 days resected spleen, biological rep 2


title: HXN38: [epididymal white adipose, biological replicate 2]

value: male


title: 7 days resected spleen, biological rep 1

description: Gene expression data from spleen 7 days after resection

value: spleen 7 days after resection


title: intact spleen, biological rep 4

description: Gene expression data from intact spleen

value: intact spleen


title: 3 days resected spleen, biological rep 1


title: HXN41: [epididymal white adipose, biological replicate 5]

value: 26 weeks


title: 7 days resected spleen, biological rep 3


title: HFD21: [epididymal white adipose, biological replicate 9]


title: HXN37: [epididymal white adipose, biological replicate 1]

value: HFD+HXN


title: intact spleen, biological rep 5

description: Gene expression data from intact spleen


title: 7 days resected spleen, biological rep 2


title: HFD20: [epididymal white adipose, biological replicate 8]


title: HFD18: [epididymal white adipose, biological replicate 6]


title: HFD19: [epididymal white adipose, biological replicate 7]


title: intact spleen, biological rep 6

description: Gene expression data from intact spleen


title: T7 E. coli tRNAs control treatment (minus DMS) 30mers / 1:2 / 10 replicate 3

city: University Park

first: McCauley

middle: O'Brien

last: Meyer

email: mcobmeyer35@gmail.com, mxm1357@psu.edu

state: PA

address: 241 Chemistry Building

department: Biochemistry and Molecular Biology

institute: Penn State University

description:
Library 60
T7_30mers_12_10_merged_out.tar.gz

data_processing:
The adapters on the 3'-end of the reads were removed by cutadapt version 2.6 with Python version 3.6.13
ShapeMapper2 version 2.1.5 was used to align the reads to the sequences of the tRNA genes retrieved from the genomic tRNA database.
ShapeMapper2 version 2.1.5 was used to calculate the normalized DMS reactivity with the min-mutation-separation set to 1 for structure prediction and 0 for natural mutational profiling. The DMS reactivity was divided by the average DMS reacvtivity of the top 10% of the reactive nucleotides.
Then the correlation of the DMS reactivity between replicate experiments was evaulated by a custom R script available at https://github.com/MacMeyer1/Scripts-to-make-figures-for-tRNAs-in-peptide-rich-droplets
If the DMS reactivity between replicates was well correlated, the raw fastq files containing the reads were combined and ShapeMapper2 version 2.1.5 was used to again align the reads and calculate the normalized DMS reactivity.
Supplementary files format and content: Each processed file is a tar.gz directory containing the output of ShapeMapper2 version 2.1.5.
Library strategy: DMS-Map-Seq

organism: synthetic construct

source_name: T7 transcripts of E. coli tRNAs

extract_protocol:
Trizol extraction
To facilitate ligation onto the 3’-end of RNAs, the ligation adapter must be 5’-end adenylated. The adapter adenylation reaction was prepared as follows: water, 10 μM Mth RNA ligase (M2611A), 200 μM ATP, 20 μM of the ligation adapter (called 3' adapter in supplemental material excel sheet), and 1X 5’ DNA Adenylation reaction buffer (B2610S). The reaction was incubated at 65 °C for 1 h, after which, the enzyme was heat inactivated at 85 C for 5 min. We performed the RT primer ligation reaction as follows: 25% PEG 8000, 1X T4 RNA Ligase Reaction Buffer, 3 μM adenylated DNA oligonucleotide from the previous step, 4 μL of RNA recovered from a DMS experiment, and 200 U of T4 RNA Ligase 2 truncated (New England Biolabs). The reaction was incubated at 16 °C overnight, and the next day, the ligated product was fractionated alongside an unligated starting material lane on a 10% denaturing (8.3 M urea 1X TBE) polyacrylamide gel. The gel was stained with SYBR-GOLD and bands were visualized with UV light. The ligated band was excised, cut into ~1 mm x 1mm squares, and placed in a 1.7 mL tube with 450 μL of TEN-250 buffer and crush and soaked at 4 °C overnight. The ligated product was recovered by ethanol precipitation. The ligated products were reverse transcribed by marathon reverse transcriptase in the presence of Mn2+ ions to induce mutations at DMS modification sites and encourage readthrough.9,10; The ligated product and reverse transcription primer were renatured together at 90 °C for 1 min and then cooled to 4 °C for 10 min to hybridize them. The marathon reverse transcription reaction contained the following: 50 mM Tris-HCl (pH 8.3), 200 mM KCl, 5 mM dithiothreitol, 20% glycerol, 2 mM MnCl2, 250 μM of each dNTP, less than 500ng of ligated product, 250 nM of reverse transcription primer (called RT primer in supplemental material excel sheet) and 10-20 U of Marathon Reverse Transcriptase (Kerafast). The reaction was incubated at 42 °C for 1 h, then ~200 mM NaOH was added, and the reaction was heated to 95 °C for 5 min to denature the reverse transcriptase and degrade the RNA template. Then an equal volume of 2X formamide loading dye (0.025% w/v bromophenol blue, 90% formamide, 10 mM EDTA) was added and the sample was fractionated on a 10% denaturing (8.3 M urea 1X TBE) polyacrylamide gel. The gel was stained with SYBR-GOLD, bands were visualized under UV-light, the RT product band was excised, cut into ~1 mm x 1mm squares, placed in a 1.7 mL tube with TEN-250 buffer and crush and soaked at 4 °C overnight. The reverse transcription product was recovered by ethanol precipitation.  The single-stranded cDNA from the reverse transcription was circularized using CircLigase II. The reaction was prepared as follows: 1X Circligase reaction buffer (Lucigen), 2.5 mM MnCl2, 1 M betaine, single-stranded cDNA from the reverse transcription, and 3.3 U of CircLigase II (Lucigen). The reaction was incubated at 60 °C for 2 h. The circularized product was used in indexing PCR without further purification. The PCR was prepared as follows, 1X Phusion HF reaction buffer, 0.5 μM indexing primer, 0.5 μM reverse primer (called PCR reverse in supplemental material excel sheet), 200 μM of each dNTP, circularized DNA, and 0.5 U of Phusion HF DNA Polymerase. The thermocycler program was based on that from Yamagami et al.2 and was performed as follows: initial denaturation at 98 °C for 30 sec, denature at 98 °C for 10 sec, anneal at 60 °C for 10 sec, elongate at 72 °C for 5 sec, repeat steps 2-4 for 17 cycles before a final extension at 72 °C for 5 min. PCR products were fractionated on an 8% non-denaturing TBE polyacrylamide gel ran at 180 V for 90-120 min. The gel was stained with SYBR-GOLD, bands were visualized with UV light, the PCR product band was excised, cut into ~1 mm x 1mm squares, and placed in a 1.7 mL with TEN-250 buffer and crush and soaked at 4 °C overnight. The indexed library was recovered by Nucleospin Gel and PCR Clean-Up kit (Macherey-Nagel). Afterwards, the concentrations of each library were measured via a micro-volume UV-vis spectrophotometer (Denovix). Before the libraries were sequenced, they were evaluated for quality and appropriate length by the TapeStation system (Agilent). To determine the concentration of each library, qPCR was performed. Libraries were sequenced by Illumina NextSeq (150 nt single-end sequencing and High-output flow cells) with the standard Illumina primer.

value: control


title: T7 E. coli tRNAs control treatment (minus DMS) 30mers / 1:2 / 10 replicate 2

description:
Library 59
T7_30mers_12_10_merged_out.tar.gz


title: HFD17: [epididymal white adipose, biological replicate 5]

institute: Oregon State University

value: HFD


title: 1 day resected spleen, biological rep 3

description: Gene expression data from spleen 1 days after resection

value: spleen 1 days after resection


title: 1 day resected spleen, biological rep 2


title: T7 E. coli tRNAs control treatment (minus DMS) 30mers / 1:2 / 10 replicate 1

description:
Library 58
T7_30mers_12_10_merged_out.tar.gz


title: T7 E. coli tRNAs control treatment (minus DMS) 10mers / 1:2 / 10 replicate 3

description:
Library 57
T7_10mers_12_10_merged_out.tar.gz


title: intact spleen, biological rep 3

description: Gene expression data from intact spleen


title: HFD16: [epididymal white adipose, biological replicate 4]


title: HFD22: [epididymal white adipose, biological replicate 10]


title: 1 day resected spleen, biological rep 1


title: intact spleen, biological rep 2

description: Gene expression data from intact spleen


title: intact spleen, biological rep 1

description: Gene expression data from intact spleen


title: HFD14: [epididymal white adipose, biological replicate 2]


title: HFD13: [epididymal white adipose, biological replicate 1]


title: T7 E. coli tRNAs control treatment (minus DMS) 10mers / 1: 1 / 0.5 replicate 1

description:
Library 52
T7_10mers_11_05_merged_out.tar.gz


title: T7 E. coli tRNAs control treatment (minus DMS) 10mers / 1: 1 / 0.5 replicate 3

description:
Library 54
T7_10mers_11_05_merged_out.tar.gz


title: LFD10: [epididymal white adipose, biological replicate 10]

value: LFD


title: T7 E. coli tRNAs control treatment (minus DMS) 10mers / 1:2 / 10 replicate 1

description:
Library 55
T7_10mers_12_10_merged_out.tar.gz


title: HFD15: [epididymal white adipose, biological replicate 3]


title: T7 E. coli tRNAs control treatment (minus DMS) 10mers / 1: 1 / 0.5 replicate 2

description:
Library 53
T7_10mers_11_05_merged_out.tar.gz


title: LFD09: [epididymal white adipose, biological replicate 9]


title: LFD07: [epididymal white adipose, biological replicate 7]


title: LFD08: [epididymal white adipose, biological replicate 8]


title: T7 E. coli tRNAs DMS treated 30mers / 1:2 / 10 replicate 1

description:
Library 49
T7_30mers_12_10_merged_out.tar.gz

value: Dimethyl sulfate


title: T7 E. coli tRNAs DMS treated 30mers / 1:2 / 10 replicate 2

description:
Library 50
T7_30mers_12_10_merged_out.tar.gz


title: T7 E. coli tRNAs DMS treated 30mers / 1:2 / 10 replicate 3

description:
Library 51
T7_30mers_12_10_merged_out.tar.gz


title: T7 E. coli tRNAs DMS treated 10mers / 1:2 / 10 replicate 1

description:
Library 46
T7_10mers_12_10_merged_out.tar.gz


title: T7 E. coli tRNAs DMS treated 10mers / 1:2 / 10 replicate 3

description:
Library 48
T7_10mers_12_10_merged_out.tar.gz


title: T7 E. coli tRNAs DMS treated 10mers / 1:2 / 10 replicate 2

description:
Library 47
T7_10mers_12_10_merged_out.tar.gz


title: LFD02: [epididymal white adipose, biological replicate 2]

description: single-end, two raw files were generated from two separate Illumina runs


title: LFD03: [epididymal white adipose, biological replicate 3]


title: LFD06: [epididymal white adipose, biological replicate 6]


title: LFD05: [epididymal white adipose, biological replicate 5]


title: T7 E. coli tRNAs control treatment (minus DMS) 10mers / 1:2 / 10 replicate 2

description:
Library 56
T7_10mers_12_10_merged_out.tar.gz


title: T7 E. coli tRNAs DMS treated 10mers / 1: 1 / 0.5 replicate 3

description:
Library 45
T7_10mers_11_05_merged_out.tar.gz


title: LFD04: [epididymal white adipose, biological replicate 4]


title: T7 E. coli tRNAs DMS treated 10mers / 1: 1 / 0.5 replicate 2

description:
Library 44
T7_10mers_11_05_merged_out.tar.gz


title: Lexis_KO3

city: Los Angeles

first: Zhengyi

last: Zhang

address: Westwood

institute: UCLA

description: RNA isolated from iWAT of Lexis KO mice

data_processing:
RNA-seq reads were aligned with HTSAT2 to the mouse genome, version mm9 (Pertea et al., 2016). Differential gene expression analysis was performed using R package DEseq2 (Love et al., 2014).
Assembly: mm9
Supplementary files format and content: Text file: Raw counts.
Supplementary files format and content: Excel file: Expression data

source_name: iWAT

extract_protocol:
RNA samples were isolated from iWAT of 12 weeks western diet fed Lexis-AdWT or Lexis-AdKO mice. Samples were homogenized using a tissue homogenizer in Trizol. RNA was purified using the Qiagen RNeasy Mini  Kit following the manufacturer’s instructions.
KAPA Stranded RNA-Seq Kit

value: iWAT

value: Lexis Knockout

value: Western diet


title: LFD01: [epididymal white adipose, biological replicate 1]

value: epididymal white adipose


title: Yphe control treatment (minus DMS) 30mers / 1:2 / 10 replicate 3

description:
Library 36
Yphe_30mers_12_10_merged_out.tar.gz

source_name: T7 transcript of an S. cerevisiae tRNA


title: T7 E. coli tRNAs DMS treated 10mers / 1: 1 / 0.5 replicate 1

description:
Library 43
T7_10mers_11_05_merged_out.tar.gz


title: Lexis_KO2

description: RNA isolated from iWAT of Lexis KO mice


title: Yphe control treatment (minus DMS) 30mers / 1:2 / 10 replicate 2

description:
Library 35
Yphe_30mers_12_10_merged_out.tar.gz


title: Lexis_KO1

description: RNA isolated from iWAT of Lexis KO mice


title: Yphe control treatment (minus DMS) 30mers / 1:2 / 10 replicate 1

description:
Library 34
Yphe_30mers_12_10_merged_out.tar.gz


title: Lexis_WT1

description: RNA isolated from iWAT of Lexis WT mice

value: Lexis Wild-type


title: Lexis_WT3

description: RNA isolated from iWAT of Lexis WT mice


title: Yphe control treatment (minus DMS) 10mers / 1:2 / 10 replicate 3

description:
Library 30
Yphe_10mers_12_10_merged_out.tar.gz


title: Yphe control treatment (minus DMS) 10mers / 1:2 / 10 replicate 2

description:
Library 29
Yphe_10mers_12_10_merged_out.tar.gz


title: Yphe DMS treated 30mers / 1:2 / 10 replicate 3

description:
Library 33
Yphe_30mers_12_10_merged_out.tar.gz


title: HEK293T cells, mutant ASPA C152W 1

city: Copenhagen

first: Leonor

last: Rib

address: Ole Maaløes Vej 5

country: Denmark

institute: University of Copenhagen

description:
DNBSEQ Eukaryotic Strand-specific mRNA library, 100bp reads
CW.A

data_processing:
The first 12 bases and reads shorter than 35 nt were removed with "Trimmomatic" v0.39 [3] using the settings "HEADCROP:12 LEADING:3 SLIDINGWINDOW:4:15 MINLEN:35". Reads were mapped using "STAR" 2.7.3a [4] against the human genome (hg38). Up to two mismatches were allowed during the mapping, and the minimum number of overlap bases to trigger mates merging and realignment was set to five. Otherwise, default settings were used. Duplicate reads were also removed with the bamRemoveDuplicatesType "UniqueIdentical" option in "STAR". The "featureCounts" function of the "Rsubread" R package v2.2.6 [5] was used to quantify reads in exons. The Gencode v38 comprehensive gene annotation including all genomic regions was used to assign reads to genes.
genome build:hg38
Processed data files format and content
tab-delimited text file including the gene raw counts of all samples

growth_protocol: HEK293T LP cells stably transfected to produce either wild-type ASPA or C152W ASPA were maintained in DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics in a humidified atmosphere with 5% CO2 at 37 C.

extract_protocol:
RNA was harvested using GeneJet RNA purification kit (Thermo). >2 ug of total RNA was shipped to BGI for construction of sequencing libraries.
Library construction and sequencing was performed by BGI (https://www.bgi.com/).

treatment_protocol: ASPA expression was induced with 2 ug/ml doxycyclin for 5 days and the mCherry positive cells were isolated by cell sorting.

value: Immortalized human embryonic kidney cells

value: Mutant ASPA C152W


title: Yphe DMS treated 30mers / 1:2 / 10 replicate 2

description:
Library 32
Yphe_30mers_12_10_merged_out.tar.gz


title: Lexis_WT2

description: RNA isolated from iWAT of Lexis WT mice


title: Yphe DMS treated 30mers / 1:2 / 10 replicate 1

description:
Library 31
Yphe_30mers_12_10_merged_out.tar.gz


title: Yphe control treatment (minus DMS) 10mers / 1:2 / 10 replicate 1

description:
Library 28
Yphe_10mers_12_10_merged_out.tar.gz


title: HEK293T cells, mutant ASPA C152W 2

description:
DNBSEQ Eukaryotic Strand-specific mRNA library, 100bp reads
CW.B


title: HEK293T cells, control ASPA wild-type 2

description:
DNBSEQ Eukaryotic Strand-specific mRNA library, 100bp reads
WT.B

value: WT ASPA


title: HEK293T cells, control ASPA wild-type 3

description:
DNBSEQ Eukaryotic Strand-specific mRNA library, 100bp reads
WT.C


title: Yphe control treatment (minus DMS) 10mers / 1:1 / 10 replicate 3

description:
Library 27
Yphe_10mers_11_10_merged_out.tar.gz


title: Yphe control treatment (minus DMS) 10mers / 1:1 / 10 replicate 2

description:
Library 26
Yphe_10mers_11_10_merged_out.tar.gz


title: Yphe control treatment (minus DMS) 10mers / 1:1 / 10 replicate 1

description:
Library 25
Yphe_10mers_11_10_merged_out.tar.gz


title: Yphe control treatment (minus DMS) 10mers / 1: 1 / 0.5 replicate 3

description:
Library 12
Yphe_10mers_11_05_merged_out.tar.gz


title: Yphe control treatment (minus DMS) 10mers / 1: 1 / 0.5 replicate 2

description:
Library 11
Yphe_10mers_11_05_merged_out.tar.gz


title: Yphe control treatment (minus DMS) 10mers / 1: 1 / 0.5 replicate 1

description:
Library 10
Yphe_10mers_11_05_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1:2 / 10 replicate 3

description:
Library 9
Yphe_10mers_12_10_merged_out.tar.gz


title: HEK293T cells, mutant ASPA C152W 3

description:
DNBSEQ Eukaryotic Strand-specific mRNA library, 100bp reads
CW.C


title: Yphe DMS treated 10mers / 1:1 / 10 replicate 3

description:
Library 6
Yphe_10mers_11_10_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1: 1 / 0.5 replicate 1

description:
Library 1
Yphe_10mers_11_05_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1: 1 / 0.5 replicate 3

description:
Library 3
Yphe_10mers_11_05_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1:1 / 10 replicate 1

description:
Library 4
Yphe_10mers_11_10_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1:2 / 10 replicate 1

description:
Library 7
Yphe_10mers_12_10_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1:1 / 10 replicate 2

description:
Library 5
Yphe_10mers_11_10_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1:2 / 10 replicate 2

description:
Library 8
Yphe_10mers_12_10_merged_out.tar.gz


title: Yphe DMS treated 10mers / 1: 1 / 0.5 replicate 2

description:
Library 2
Yphe_10mers_11_05_merged_out.tar.gz


title: HEK293T cells, control ASPA wild-type 1

description:
DNBSEQ Eukaryotic Strand-specific mRNA library, 100bp reads
WT.A


title: K-562 WT EV Replicate 2

city: Montreal

first: Eric

last: Lecuyer

state: Quebec

address: 110 Pine Ave W

country: Canada

institute: Montreal Clinical Research Institute

description:
merged_counts.tsv
DEG_results_FeatureCounts_biotypes_cellsvsEV.tsv
isoforms.predictProductivity.bed
isoforms_deseq2_cells_v_EV_results.annot.tsv

data_processing:
Base calling on fast5 files using guppy base caller software with a quality threshold of 7
Reads that pass the base-celling quality  threshold were demultiplexed to individual barcodes using "Fastq barcoding" pipeline of Oxford Nanopore Technologies Bioinformatics Platform.
Filter out reads shoirter than 20 nucleotides from fastq files using NanoFilt -l 20
Align reads to the genome (hg38) with minimap2 -ax splice
Read assigment using Rsubread featureCounts (isLongRead=True, isGTFAnnotationFile=TRUE)
Differential expression analysis of known genes (DESeq2 on counts obtained by featureCounts)
Analysis of novel genes and transcripts using FLAIR pipeline.
Assembly: hg38
Supplementary files format and content: tab-delimited file of raw counts obtained by featureCounts (merged_counts.tsv)
Supplementary files format and content: tab-delimited file of differential gene expression analysis with gene information (DEG_results_FeatureCounts_biotypes_cellsvsEV.tsv)
Supplementary files format and content: bed file with isoforms obtained by FLAIR (isoforms.predictProductivity.bed)
Supplementary files format and content: tab-delimited file with the result of differential isoform expression analysis using FLAIR (isoforms_deseq2_cells_v_EV_results.annot.tsv)

source_name: Bone; Marrow

growth_protocol: Roswell Park Memorial Institute (RPMI) 1640 with L-glutamate supplemented with 10% foetal bovine serum, and 100 U mL-1 penicillin and 100 µg mL-1 streptomycin.

extract_protocol:
Total RNA (TRIzol)
Nanopore PCR-cDNA Barcoding

treatment_protocol: Untreated

value: Bone; Marrow

value: Human chronic myeloid leukemia

value: WT

value: Untreated


title: K-562 WT EV Replicate 3


title: 52-S1 3

city: Bath

first: Matthew

middle: J

last: Shepherd

state: Somerset

address: University of Bath, Claverton Down

department: Department of Life Sciences

institute: University of Bath

description:
WTCHG_816601_70155087
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15, PFLU1583Δ48-74
52S1

data_processing:
Reference-based transcript assembly was performed using the Rockhopper RNA sequencing analysis software (https://cs.wellesley.edu/~btjaden/Rockhopper/). PTT file generated from NCBI GenBank file for P. fluorescens SBW25 complete genome (NC_012660.1) using gb2ptt (https://github.com/sgivan/gb2ptt), .gff and .fna for SBW25 were downloaded from NCBI GenBank from the same accession.
Differential gene expression analysis performed using DESeq2 with assembled transcript counts output from Rockhopper, to generate Log2Fold changes for ever coding sequence.
Assembly: NC_012660.1
Supplementary files format and content: Reference based transcript assembly rockhopper output as tab-delimited .txt file (Rockhopper_transcripts.txt). File details raw counts for every transcript, normalised counts, RPKM, and an estimate of expression calculated by normalising RPKM by the upper quartile of gene expression.
Supplementary files format and content: DESeq2 outputs, using raw transcript counts to estimate differential gene expression, comparing expression for each evolved strain of interest against the ancestral AR2 background.

organism: Pseudomonas fluorescens

source_name: bacterial cells

growth_protocol: Cultures were incubated in LB broth at 27°C and 180 rpm shaking, until P. fluorescens reached mid-log phase growth (OD595 ~1.5).

extract_protocol:
Whole-cell RNA was extracted from 20 OD units of P. fluorescens cultures in mid-log phase growth (OD595 ~1.5). Upon reaching the desired OD, growth and RNA expression were halted by addition of a ½ culture volume of ice-cold killing buffer (20 mM NaN3, 20 mM Tris-HCl, 5 mM MgCl2). Cells were pelleted, and the killing buffer removed. A lysis buffer of β-mercaptoethanol in buffer RLT from the Qiagen RNeasy extraction kit was used to resuspend pellets, which were then lysed by bead-milling at 4500 rpm for 45 s with lysing matrix B. Lysates were spun through columns from the Qiagen RNeasy extraction kit, and the extraction completed following the RNeasy kit protocol. A DNase I treatment step was included between washes with buffer RW1, by adding DNase I directly to the column from the RNase-free DNase kit (Qiagen) following kit protocol. Samples were eluted in nuclease-free water, and subsequently treated with TURBO DNase from the Turbo DNA-free kit (Invitrogen) following kit protocols.
RNA sequencing was provided by Oxford Genomics (Oxford UK), and followed their methods for library preparation and sequencing. Samples were ribodepleted, and the mRNA fraction converted to cDNA with dUTP incorporated during second strand synthesis. The cDNA was end-repaired, A-tailed and adapter-ligated. Prior to amplification, samples underwent uridine digestion. The prepared libraries were size selected, multiplexed and QC’ed before paired end sequencing over one unit of a flow cell in a NovaSeq 6000.

value: bacterial cells

value: <delta>fleQ, IS-<omega>Km-hah:PFLU2552, <delta>ntrC, PFLU1131-del15, PFLU1583<delta>48-74

tag: condition

value: Shaking LB broth, logarithmic growth phase


title: 52-S1 1

description:
WTCHG_816601_70135085
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15, PFLU1583Δ48-74
52S1


title: 52-S1 2

description:
WTCHG_816601_70145086
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15, PFLU1583Δ48-74
52S1


title: K-562 WT Cell Replicate 3


title: 2501-S1 3

description:
WTCHG_816601_70125084
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15, PFLU1131/2 promoter
2501S1

value: <delta>fleQ, IS-<omega>Km-hah:PFLU2552, <delta>ntrC, PFLU1131-del15, PFLU1131/2 promoter


title: K-562 WT Cell Replicate 2


title: 2501-S1 2

description:
WTCHG_816601_70115083
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15, PFLU1131/2 promoter
2501S1


title: K-562 WT EV Replicate 1


title: MDAMB468_2

city: Bethesda

first: Stefan

last: Ambs

state: MD

address: Building 37, Room 3050B

department: CCR

institute: NCI

data_processing:
ATAC-seq samples were pooled and sequenced on NextSeq using OMNI ATAC-seq protocol adapted from M.R. Corces, et al., 2017 and paired-end sequencing. All the samples have percent of Q30 bases above 95%. Samples were trimmed for adapters using Cutadapt before the alignment. The trimmed reads were aligned with Human - hg38 reference using Bowtie2 alignment.
Assembly: hg38
Supplementary files format and content: excel files include Z-scores for motifs for each Sample

growth_protocol: For normoxic (21% O2) condition, cells were cultured at 37 °C in a 5% CO2 humidified environment as an adherent monolayer. Hypoxia experiments were performed in a hypoxia chamber (BioSpherix) at 1% O2 at 37 °C in a 5% CO2 humidified environment.

extract_protocol:
Cell revival, cell lysis, transposition, and DNA extraction were done using a published method by Corces et al., 2017 (PMID: 28846090) with some modifications for cryopreserved cell processing (Caravaca et al., 2022) and sequencing was performed on an Illumina NextSeq 2000 machine at the Sequencing Facility, Leidos Biomedical Research, Inc.
Library complexity was measured by uniquely aligned reads using Picard’s MarkDuplicate utility. All the samples had library complexity with percent non-duplicated reads ranging from 38 to 69%. The peaks were called using Genrich and total number of peaks ranged between 54,835 and 83,046. The number of reads falling in peaks (FRiP score), ranges between 20.1 to 34.3%.

value: breast cancer cell line

value: TNBC

tag: ethnicity

value: AA

tag: er status

value: Neg

tag: cell lines


title: HCC70_2


title: 2501-G 3

description:
WTCHG_816601_70095081
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15
2501G

value: <delta>fleQ, IS-<omega>Km-hah:PFLU2552, <delta>ntrC, PFLU1131-del15


title: 2501-S1 1

description:
WTCHG_816601_70105082
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15, PFLU1131/2 promoter
2501S1


title: BT549_2

value: EA


title: MDAMB231_2


title: MDAMB157_2


title: MDAMB436_2


title: 2501-G 2

description:
WTCHG_816601_70085080
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15
2501G


title: HCC1806_2


title: MDAMB468_1


title: dntrC 3

description:
WTCHG_816601_70065078
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC
dntrC


title: HS578t_2


title: K-562 WT Cell Replicate 1

tag: cell line


title: MDAMB231_1


title: BT549_1


title: HCC70_1


title: EXP-JZ47

city: Shenyang

first: Chen

middle: yu

last: lan

address: Nan Jing Bei Street

institute: Department of Clinical Epidemiology and Evidence-based Medicine, the First Affiliated Hospital, China Medical University

hyb_protocol: bisulphite converted DNA was amplified, fragmented and hybridised to Illumina HumanMethylationEPIC BeadChip (Illumina, Inc., CA, USA) (850K array) using standard Illumina protocol

scan_protocol: Arrays were imaged using BeadArray Reader using standard recommended Illumina scanner setting

data_processing: Data was processed with the the Chip Analysis Methylation Pipeline (ChAMP) package in R. Probes were removed if they did not pass the detection cut-off (p < 0.01), were associated with known SNPs, were associated with the X and Y chromosomes, or were non-CpG probes. Following normalization, a single value decomposition (SVD) analysis was used to identify batch effects which were removed using ComBat with the SVA package using gene-wise linear models. DMPs with a P < 0.05 and a |Δβ| > 0.1 were selected for subsequent analysis. LEF-related differentially methylated genes (DMGs) were then identified, including genes associated with both hypo- and hypermethylation.

label: Cy3, Cy5

source_name: Whole Blood

extract_protocol: genomic DNA was extracted and purified from RA samples using QIAamp DNA Blood Mini Kit (QIAGEN)  according to standard instructions

value: Whole Blood

tag: gender

value: female

tag: age at diagnosis

tag: group

value: case


title: EXP-JZ31

value: male


title: dntrC 1

description:
WTCHG_816601_70045076
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC
dntrC

tag: cell type

tag: genotype


title: dntrC 2

description:
WTCHG_816601_70055077
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC
dntrC


title: EXP-JZ45

tag: age


title: MDAMB157_1


title: EXP-D66


title: EXP-D28


title: 2501-G 1

description:
WTCHG_816601_70075079
ΔfleQ, IS-ΩKm-hah: PFLU2552, ΔntrC, PFLU1131-del15
2501G

tag: strain


title: AR2 3

description:
WTCHG_816601_70035075
ΔfleQ, IS-ΩKm-hah: PFLU2552
AR2

value: <delta>fleQ, IS-<omega>Km-hah:PFLU2552


title: HCC2157_2


title: EXP-T32


title: EXP-T228


title: EXP-T26


title: HCC2157_1


title: BT549_Hypo_2

value: hypoxia

value: CC


title: EXP-E30


title: EXP-D33


title: MDAMB436_1


title: EXP-JZ6


title: EXP-T127


title: HCC70_Hypo_2


title: EXP-D35


title: EXP-D34


title: lov_KO_HMM_NaCl_blue, rep 3

city: Ames

first: Gwyn

middle: A

last: Beattie

state: IOWA

address: 2213 Pammel Dr

department: Plant Pathology, Entomology, & Microbiology

institute: Iowa State University

description: total RNA

data_processing:
The raw reads were filtered by BGI Genomics using SOAPnuke (v1.5.2) to remove reads containing sequencing adapters and low-quality reads (reads containing >20% of bases having quality scores ≤15 or an ambiguous sequence content of >1%). FASTQ files of the resulting “clean” reads were used for further quality assessment using FastQC v0.11.4 and analysis.
The clean reads from each sample were mapped to the B728a reference genome GCF_000012245.1 (2005 version) using Bowtie2 (v2.2.6) and TopHat2 (v2.1.0).
The number of reads that mapped to each gene in the genome were counted using HTSeq-count (v0.6.0) with parameters -i locus_tag -t gene -s reverse -r pos -m intersection-nonempty -a 10 -f bam. Genes with average read counts of <1 across all samples were excluded from subsequent analyses.
Counts were normalized for library size using upper-quartile normatilzation.
Normalized gene counts for all 66 samples were used to assess differential expression between specified strain x treatment combinations using the R package QuasiSeq (negative binomial model, strain x treatment interaction and replicate used as fixed effects, and the QLSpline method for FDR estimates), with q-value ≤ 0.00105 used to define statistical significance.
Assembly: Pseudomonas syringae B278a, GCF_000012245.1 assembly was downloaded from NCBI in October 2016; the original RefSeq annotation (submitted in 2005 by the Joint Genome Institute) was replaced replaced by a PGAP annotation in 2023 without changing the accession number, but the original annotation used here is still available under NCBI accession GCA_000012245.1
Supplementary files format and content: raw_counts.csv is is a matrix table with raw gene counts for every gene and every sample
Supplementary files format and content: norm_counts.csv is a matrix table with natural log-transformed normalized gene counts for all genes with average raw read counts of ≥1 across all samples
Supplementary files format and content: B728aLight_QuasiSeq_results.xlsx is an Excel file with the results of the QuasiSeq differential analysis results (Ln[fold change], Log2[fold change], fold change, q-value) for all genes along with transcript metadata including gene name, product description, and functional classification.

organism: Pseudomonas syringae pv. syringae B728a

source_name: bacteria

growth_protocol: Two independent cultures of each strain were grown on each of six separate days. For each culture, an initial culture was grown from a single colony in KB medium containing Rif (50 μg/mL) to a density of approx. 10^9 cells/mL, at which point 10 μL was sub-cultured into 5 mL KB medium and grown again to a density of approx. 10^9 cells/mL. A 0.2-mL aliquot was sub-cultured into 20 mL KB in a 125-mL flask and grown for 4–6 h with shaking in the dark at 20–22°C until an optical density at 600 nm (OD600) of approx. 0.5.

extract_protocol:
Immediately after treatment, 0.5 mL of each culture sample was added to 1 mL of RNAprotect (Qiagen) and vortexed for 5 sec. Following a 5-min incubation at room temperature, the cells from the two independent cultures for a single strain x treatment combination were combined and centrifuged at 5,000 x g for 10 min. The supernatant was decanted, and the cell pellet was stored at -70°C. For each strain, this process was repeated on six separate days. The cell pellets for a given strain x treatment combination collected on two separate days were thawed on ice and combined, and this pool was considered one biological replicate. In this manner, each strain x treatment combination was represented by three biological replicates, each of which contained cells derived from four independently exposed cultures. Total RNA was extracted using a RNeasy Mini Kit (Qiagen) with on-column Dnase I treatment, with one biological replicate for each strain x treatment combination extracted at the same time.
Library construction and sequencing were performed by BGI Genomics (China). For each sample, total RNA was depleted for ribosomal RNA using an Epicentre Ribo-Zero Magnetic Kit (Bacteria), fragmented, and then used for RNA-seq library construction using a TruSeq RNA Sample Prep Kit v2 (Illumina). Libraries were assessed for quality using a 2100 Bioanalyzer, quantified using real-time quantitative PCR, and sequenced on an Illumina HiSeq 4000 with 100-bp paired-end reads.

treatment_protocol: Two-milliliter culture aliquots were transferred into individual wells of replicate 6-well cell culture plates and plates were immediately exposed to blue light (20 μmol/m^2/s), red light (30 μmol/m^2/s), far-red light (188 μmol/m^2/s), or white light (34 μmol/m^2/s), or maintained in the dark for 15 min at 20–22°C without shaking. As a follow-up to prior microarray studies, the Δlov mutant was subject to two additional treatments in which cells cultivated in defined HMM medium were exposed for 15 min to blue light in the presence or absence of 0.23 M NaCl.

value: bacteria

tag: treatment

value: HMM + 0.23M NaCl + blue


title: HCC1806_Hypo_2


title: HS578t_1


title: HCC1806_1


title: EXP-E31


title: EXP-T24


title: EXP-D51


title: lov_KO_HMM_NaCl_blue, rep 2

description: total RNA


title: AR2 2

description:
WTCHG_816601_70025074
ΔfleQ, IS-ΩKm-hah: PFLU2552
AR2

value: bacterial cells

value: Shaking LB broth, logarithmic growth phase


title: MDAMB157_Hypo_2

value: breast cancer cell line


title: lov_KO_HMM_NaCl_blue, rep 1

description: total RNA


title: HS578T_Hypo_2


title: BT549_Norm_2

value: normoxia


title: AR2 1

description:
WTCHG_816601_70015073
ΔfleQ, IS-ΩKm-hah: PFLU2552
AR2


title: MDAMB436_Hypo_2


title: MDAMB231_Hypo_2


title: lov_KO_HMM_blue, rep 3

description: total RNA

value: HMM + blue


title: EXP-E54


title: lov_KO_HMM_blue, rep 2

description: total RNA


title: HCC70_Norm_2

value: AA

value: Neg


title: MDAMB468_Hypo_2


title: HCC1806_Norm_2


title: EXP-JZ41


title: EXP-JZ3


title: MDAMB468_Norm_2


title: HS578T_Norm_2


title: MDAMB436_Norm_2


title: CON-JZ24

value: control


title: BT549_Hypo_1


title: MDAMB231_Norm_2


title: CON-T16

value: Whole Blood


title: lov_KO_HMM_blue, rep 1

description: total RNA


title: HS578T_Hypo_1


title: MDAMB157_Norm_2


title: MDAMB436_Hypo_1


title: rhlA_KO_far-red, rep 2

description: total RNA

value: far-red


title: MDAMB231_Hypo_1


title: rhlA_KO_far-red, rep 3

description: total RNA


title: HCC70_Hypo_1


title: rhlA_KO_far-red, rep 1

description: total RNA


title: BT549_Norm_1


title: bsi_KO_far-red, rep 3

description: total RNA