title: C4-2B_sh4B

city: Hsinchu city

first: Meng-Chen

last: Wu

address: 101, Section 2 Kuang Fu Road

department: Institute of Molecular and Cellular Biology

institute: National Tsing Hua University

description: Gene expression profiles by Affymetrix microarray

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

scan_protocol: GeneChip Scanner 3000

data_processing: Data were processing using Transcriptome Analysis Console (TAC) Software. Robust Multi-array Average (RMA) method was used to background correction and normalization.

label: Biotin

label_protocol: Biotin

growth_protocol: C4-2B were cultured in RPMI 1640 medium contained with 10% fetal bovine serum at 37°C in 5% CO2.

extract_protocol: Total RNAs were extracted with TRIzol reagent following manufacturer's instructions.

treatment_protocol: Cell lines were established in C4-2B cells using LKO and sh4B lentivirus infection

tag: cell line

value: C4-2B (ATCC number:CRL-3315)

value: Caucasian prostate cancer

value: KDM4B knockdown


title: Phosphate & Iron deprivation time 9h Rep3

status: Public on Mar 24 2023

type: RNA

city: Montpellier

first: Sandrine

last: Ruffel

address: 2 Place Viala

department: Montpellier University

country: France

institute: INRA

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours on a phosphate and iron deprived medium

hyb_protocol: Following fragmentation, 5.5ug of cDNA were hybridized for 16 hr at 48C on GeneChip 1.1ST Array Strip. GeneChips were washed and stained in the Affymetrix GeneAtlas Fluidics Station.

scan_protocol: Array strips were scanned using the Affymetrix GeneAtlas Imaging Station.

data_processing: The data quality analysis was done with the Affymetrix Expression Console software. The normalization of the raw data was done with the Affymetrix Transcriptome Analysis Console software using the GCRMA method.

label: biotin

molecule: total RNA

organism: Arabidopsis thaliana

source_name: Arabidopsis thaliana 10 Days old

label_protocol: Biotinylated cDNA were prepared according to the standard Affymetrix protocol from 100ng total RNA (GeneChip® WT PLUS Reagent Kit Manual Target Preparation for GeneChip® Whole Transcript (WT) Expression Arrays).

growth_protocol: Arabidopsis Columbia-0 plants were grown on a growth medium containing 1mM Pi

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

treatment_protocol: One part of the plants was transferred to a nutrient medium not containing Pi (0Pi)  for 3, 6 and 9 h. One part of the plants was transferred to a nutrient medium not containing Fe (0Fe)  for 3, 6 and 9 h and one part was tranferred to a nutrient medium not containing Pi (0Pi) and Fe (0Fe)  for 3, 6 and 9 h.

tag: tissue

value: Roots

tag: ecotype

tag: age

value: 10 days


title: Phosphate deprivation time 9h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours in a phosphate deprived  medium


title: Iron deprivation time 6h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours on an iron deprived medium


title: Full nutrient time 9h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours on full nutrient medium


title: NIH3T3 cells, shCTRL_SCR1-D2_D1

type: SRA

city: Munich

first: Vanessa

last: Arfelli

state: Bavaria

address: Max-Lebsche-Platz 30

department: Medical Clinic and Polyclinic III

country: Germany

institute: Munich University Hospital, Ludwig-Maximilians University (LMU)

library_source: transcriptomic

data_processing:
Raw Fastq data were processed with the pipeline zUMIs, version 0.2.0 (Parekh et al., 2018)
cDNA reads were mapped to the mouse genome mm10 using the software STAR (Dobin et al., 2013).
Gene annotations were obtained from Ensembl (GRCm38.75)
Assembly: Mouse: mm10
Supplementary files format and content: Sample (columns) and gene (rows) -wise UMI expression table

organism: Mus musculus

growth_protocol: The cells used in this study were cultivated at 37 ºC and 5% CO2 in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% of fetal bovine serum (FBS) and 1% of the antibiotics penicillin-streptomycin

extract_protocol:
Libraries were constructed using a bulk RNA-seq method named Prime-seq, which uses direct cell lysis, bead cleanup, cDNA synthesis and pre-aplification with KAPA HiFi Polymerase (see Janjic et al, 2021)
The cells were counted by Vi-Cell XR (Beckman Coulter, Brea, CA, USA) and 1x104 cells were harvested in microtubes containing 100 µl of DPBS 1x. After 5 minutes of centrifugation at 1500 rpm, cells were resuspended in 100 µl of Buffer RLT Plus, RNeasy Plus Lysis Buffer (Qiagen, Hilden, Germany) supplemented with 1% b-mercapoethanol.
Prime-seq (Janjic et al, 2021 -  https://doi.org/10.1101/2021.09.27.459575)

treatment_protocol: NIH3T3 cells were transduced with retroviral particles carrying either shRNA-UHMK1 sequences (shUHMK1#1, shUHMK1#2, shUHMK1#3 and scrambled control) or the rat Uhmk1 coding sequence, in frame with a Flag peptide in the 5’, N-terminal of the protein (and the respective empty vector control, MIY).

tag: replicate number

tag: genotype

value: Control - scramble

tag: transduction

value: scrambled shRNA-UHMK1 sequence (shCTRL) which has target gene


title: Phosphate & Iron deprivation time 6h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours on a phosphate and iron deprived medium


title: bsub-str168_cam_r3_s_r_v2

city: Solna

first: Vicent

last: Pelechano

address: Nobels väg 16

department: MTC

country: Sweden

institute: ScilifeLab - Karolinska Institutet

data_processing:
Adapter and quality trimming with the bbduk program of the BBTools suite, with options {qtrim=r, ktrim=r, hdist=3, hdist2=2, K=20, mink=14, trimq=16, minlen=30, maq=16}, using BBTools default adapter set and polyG or polyA sequences for short reads.
Deduplication of reads with both identical unique molecular identifier (UMI) and insert sequence using the dedupe program of the BBTools suite with its default parameters.
UMI sequences found in the first eight bases of each read were extracted using UMI-tools (version 1) with default options (using --bc-pattern NNNNNNNN).
Alignment was performed with the bbmap program of the BBTools suite, with the {32bit=t -da -eoom k=11 strictmaxindel=10 intronlen=0 t=16 trd=t minid=0.94 nzo=t} parameters. Alignment files were sorted and indexed with SAMtools.
Deduplication based on UMIs was then performed with UMI-tools (version 1) with options { --soft-clip-threshold 1 --edit-distance 2 --method unique}.
In selected samples, a dual undex strategy was used (sequencing runs v17). In brief, in addition of the Illumina indexes with dedicated reads used in HT-5Pseq, an additional index sequence is added before the UMI sequence. This is introduced during the ssRNA ligation space using anoligo with the following structure Illumina compatible-CCCCCC-NNNNNNNN. Please see dual_index_demultiplexing.txt for details on demultiplexing strategy for those samples.
Bedgraphs for read 5' positions for forward (fwd) and reverse (rev) strands were generated with BEDTools genomecov program with options {-bg -strand [+/-] -5}.
Genome_build: The genome used for alignment is indicated in the processed file names with the G-xxxx patterns. Those correspond to the following NCBI accessions and builds: G-bamy (Bacillus amyloliquefaciens: GCF_000196735.1), G-bsub (Bacillus subtilis: GCF_000009045.1), G-cvib (Caulobacter vibrioides/crescentus: GCF_000006905.1), G-ecol (Escherichia coli: GCF_000005845.2), G-spcc (Synechocystis sp. PCC 6803: GCF_000009725.1), G-lpla (Lactobacillus plantarum: GCF_000203855.3), G-lreu (Lactobacillus reuteriGCF_000016825.1), G-zymo (mix of the following genomes [Bacillus subtilis: GCF_000009045.1, Enterococcus faecalis: GCF_000007785.1, Escherichia coli: GCF_000005845.2, Lactobacillus fermentum: GCF_000010145.1, Listeria monocytogenes: GCF_000196035.1, Pseudomonas aeruginosa: GCF_000006765.1, Salmonella enterica: GCF_000006945.2, Staphylococcus aureus: GCF_000013425.1]), G-46genera (genomes from 46 genera listed in the Supplementary file).
Genome_build: Reference indiced for individual bacterial species and custom mixtures were built with the bbmap program of the BBTools suite, with default options (and k=10). See details in the supplementary file "46genera.xls". In Brief: Akkermansia muciniphila ATCC BAA-835 (GCF_000020225.1); Alistipes finegoldii DSM 17242 (GCF_000265365.1); Alistipes obesi (GCF_000311925.1); Alistipes putredinis DSM 17216 (GCF_000154465.1); Alistipes shahii WAL 8301 (GCF_000210575.1); Anaerobutyricum hallii DSM 3353 (GCF_000173975.1); Atopobium vaginae DSM 15829 (GCF_000159235.2); Bacillus amyloliquefaciens DSM 7 = ATCC 23350 (GCF_000196735.1); Bacillus subtilis subsp. subtilis str. 168 (GCF_000009045.1); Bacillus subtilis subsp. subtilis str. 168 (GCF_000009045.1); Bacteroides caccae (GCF_002222615.2); Bacteroides cellulosilyticus (GCF_001318345.1); Bacteroides coprocola DSM 17136 (GCF_000154845.1); Bacteroides coprophilus DSM 18228 = JCM 13818 (GCF_000157915.1); Bacteroides faecichinchillae JCM 17102 (GCF_000614145.1); Bacteroides fluxus YIT 12057 (GCF_000195635.1); Bacteroides fragilis YCH46 (GCF_000009925.1); Bacteroides ovatus (GCF_001314995.1); Bacteroides plebeius DSM 17135 (GCF_000187895.1); Bacteroides salanitronis DSM 18170 (GCF_000190575.1); Bacteroides salyersiae WAL 10018 = DSM 18765 = JCM 12988 (GCF_000381365.1); Bacteroides stercoris ATCC 43183 (GCF_000154525.1); Bacteroides thetaiotaomicron VPI-5482 (GCF_000011065.1); Bacteroides uniformis ATCC 8492 (GCF_000154205.1); Bilophila wadsworthia ATCC 49260 (GCF_000701705.1); Caulobacter vibrioides CB15 (GCF_000006905.1); Clostridium phoceensis (GCF_001244495.1); Collinsella aerofaciens ATCC 25986 (GCF_000169035.1); Coprobacter fastidiosus NSB1 (GCF_000473955.1); Coprococcus comes ATCC 27758 (GCF_000155875.1); Culturomica massiliensis (GCF_900091655.1); Desulfovibrio fairfieldensis (GCF_001553605.1); Dorea formicigenerans ATCC 27755 (GCF_000169235.1); Dorea longicatena DSM 13814 (GCF_000154065.1); Enterococcus faecalis V583 (GCF_000007785.1); [Clostridium] innocuum 2959 (GCF_000371425.1); [Clostridium] saccharogumia DSM 17460 (GCF_000686665.1); Escherichia coli str. K-12 substr. MG1655 (GCF_000005845.2); Escherichia coli str. K-12 substr. MG1655 (GCF_000005845.2); Eubacterium ventriosum ATCC 27560 (GCF_000153885.1); Faecalibacterium prausnitzii A2-165 (GCF_000162015.1); Flavobacterium johnsoniae UW101 (GCF_000016645.1); Flavobacterium saccharophilum (GCF_900142735.1); Fusicatenibacter saccharivorans (GCF_001405555.1); Gardnerella vaginalis 409-05 (GCF_000025205.1); Synechocystis sp. PCC 6803 (cyanobacteria) (GCF_000009725.1); Gemmiger formicilis (GCF_900167555.1); Intestinimonas butyriciproducens (GCF_001454945.1); Lactobacillus plantarum WCFS1 (GCF_000203855.3); Lactobacillus reuteri DSM 20016 (GCF_000016825.1); Lactobacillus crispatus ST1 (GCF_000091765.1); Lactobacillus iners DSM 13335 (GCF_000160875.1); Lactobacillus fermentum IFO 3956 (GCF_000010145.1); Lactococcus lactis subsp. lactis Il140 (GCF_000006865.1); Lacunisphaera limnophila (GCF_001746835.1); Listeria monocytogenes EGD-e (GCF_000196035.1); Mediterranea massiliensis (GCF_900128475.1); Megasphaera genomosp. type_1 str. 28L (GCF_000177555.1); Microbacterium barkeri 2011-R4 (GCF_000299315.2); Mucilaginibacter gossypiicola (GCF_900110105.1); Odoribacter splanchnicus DSM 20712 (GCF_000190535.1); Parabacteroides distasonis ATCC 8503 (GCF_000012845.1); Parabacteroides goldsteinii DSM 19448 = WAL 12034 (GCF_000969835.1); Prevotella amnii DSM 23384 = JCM 14753 (GCF_000378745.1); Prevotella bivia DSM 20514 (GCF_000262545.1); Prevotella buccalis ATCC 35310 (GCF_000177075.1); Prevotella timonensis 4401737 = DSM 22865 = JCM 15640 (GCF_000455445.1); Pseudomonas fluorescens F113 (GCF_000237065.1); Pseudomonas protegens CHA0 (GCF_000397205.1); Pseudomonas aeruginosa PAO1 (GCF_000006765.1); Rhodanobacter fulvus Jip2 (GCF_000264315.1); Romboutsia timonensis (GCF_900106845.1); Roseburia hominis A2-183 (GCF_000225345.1); Roseburia inulinivorans DSM 16841 (GCF_000174195.1); Ruthenibacterium lactatiformans (GCF_000949455.1); Salmonella enterica subsp. enterica serovar Typhimurium str. LT2 (GCF_000006945.2); Sneathia sanguinegens (GCF_001517935.1); Staphylococcus aureus subsp. aureus NCTC 8325 (GCF_000013425.1); Streptococcus salivarius (GCF_000785515.1); Streptococcus thermophilus JIM 8232 (GCF_000253395.1); [Eubacterium] rectale ATCC 33656 (GCF_000020605.1); [Eubacterium] siraeum DSM 15702 (GCF_000382085.1) and  Ureaplasma parvum serovar 3 str. ATCC 27815 (GCF_000019345.1).
Supplementary_files_format_and_content: Processed files are bedgraphs containing read 5' counts on reverse (_rev) and forward (_fwd) strands of genomes indicated with _G-xxxx pattern. The genomes corresponding to each pattern are described in the genome build entry above.

organism: Bacillus subtilis

source_name: Liquid culture of Bacillus subtilis 168 trpC2.

growth_protocol: Bacteria were grown according to their requirements. Overnight Cultures were diluted to OD600 0.05-0.07, growth was continued and cells were harvested in log-phase by centrifugation. Resulting pellet was shock-frozen in Dry ice/Ethanol bath and stored for RNA analysis

extract_protocol:
Microbial cell pellets were resuspended in equal volume of LET (25 mM Tris pH 8.0, 100 mM LiCl, 20 mM EDTA) and water saturated Phenol pH 6.6 (Thermo Fisher),  lysed with acid washed glass beads (Sigma-Aldrich) and vortexing for three minutes in MultiMixer. Lysates were supplemented  with equal volumes of phenol/chloroform isoamyl alcohol pH 4.5 (25:24:1) and nuclease free water, l two-minute vortexing followed by centrifugation. Resulting aqueous phase was purified in two steps using phenol-chloroform isoamyl alcohol (25:24:1) followed by chloroform. After centrifugation, the clean aqueous phase was precipitated with sodium acetate-ethanol.
Standard and random fragmented 5PSeq libraries were generated as described in PMID: 26046441 with minor modifications using 150-9000ng total RNA as an input. To prepare random fragmented samples (negative controls), ribosomal RNA was depleted from DNA-free RNA and subsequent fragmented by incubating five minutes at 80 ̊C in fragmentation buffer (40mM Tris Acetate pH 8.1, 100mM KOAc and 30mM MgOAc). Reaction was purified using 2 volumes of RNACleanXP beads (Beckman Coulter) as recommended by the manufacturer. Free 5’OH sites were re-phosphorylated using 5 Units of T4 Polynucleotide kinase (PNK, NEB) and incubated at 37 ̊C for 60 minutes as recommended by the manufacturer. Re-phosphorylated fragmented RNA was purified using Phenol:Chloroform: Isoamyl Alcohol (24:25:1), followed by sodium acetate-ethanol precipitation. From this step forward, procedures for random fragmented and standard 5PSeq library preparation merge. RNA was Ligated to either rP5_RND or rP5_RNA oligo (specified in characteristics protocol).    containing unique molecular identifiers). Ribosomal RNA was depleted using Ribozero rRNA removal kit (Illumina) suitable for Bacteria and Yeast. rRNA depleted sample was purified using 1.8V of Ampure beads (Abcam), fragmented using heat (80˚C) for 5 min in 5x Fragmentation Buffer (200mM Tris Acetate pH 8.1,500mM KOAc,150mM MgOAc) and reverse transcribed using random hexamers to prime. Resulting cDNA was bound to streptavidin beads (M-280), subjected to enzymatic reactions of DNA end repair, fill-in of adenine to 5’ protruding ends of DNA fragments using Klenow Fragment (NEB). Common adaptor (P7-MPX) was ligated and 5PSeq Libraries were amplified by PCR (15- 17 cycles). Purified samples were quantified using Qubit (Thermo Fisher), library size was estimated from bioanalyzer traces. 5PSeq Libraries were pooled by mixing equal amounts of each sample, following enrichment of fragments of the length between 300-500nt.   HT-5PSeq Libraries were generated as recently described (bioRxiv 2020.06.22.165134; doi: https://doi.org/10.1101/2020.06.22.165134). In brief, DNA-free RNA was ligated with RNA oligos containing unique molecular identifiers. Ligated RNA was reverse transcribed priming with oligos containing a random hexamer and an Illumina compatible region. RNA was eliminated by addition of NaOH. Ribosomal RNA was depleted by adding in-house rRNA DNA oligo depletion mixes (table 2) to the cDNA and performing a duplex-specific nuclease (DSN, Evrogen) treatment. rRNA depleted cDNA was PCR amplified (15-17 cycles). Depletion of ribosomal RNA with Ribozero Illumina (for bacteria and yeast) was done after the single-stranded RNA ligation step. Ribosomal depleted RNA was purified and reverse transcribed using the same oligos as stated above, and then amplified by PCR. Libraries were quantified by fluorescence (Qubit, Thermo Fisher), size estimated using an Agilent Bioanalyzer and sequence using a NextSeq500 Illumina sequencer.
5PSeq that captures  transcripts with free 5' phospahte (without cap)

treatment_protocol: As for Chloramphenicol (CAM) treatment, Bacterial cells were incubated with Chloramphenicol (100µg/ml final) for 5 minutes at 37˚C with aeration and harvested at 4˚C on ice supplemented with Chloramphenicol (~100µg/ml final).   Mupirocin (MUP) was added to bacterial culture at a final concentration of 65µg/ml and incubated for 10 minutes at 37˚C with aeration.   Cultures for stationary growth phase samples were inoculated with single colony and harvested at indicated times post-inoculation.   Salt stress samples were generated as described in PMID: 28290514 with minor modifications Salt stress was initiated by mixing equal volumes of bacterial growth culture and 2M NaCl (~5.8% NaCl). Bacterial cells were exposed to salt stress for 10 minutes at room temperature and harvested by centrifugation.   Cultures for stationary growth phase samples were inoculated with single colony and harvested at indicated times post-inoculation.   Heat shock experiments were setup the following. Lactobacillus plantarum (ATCC 8014) growth culture was harvested by centrifugation and heat shock was induced by resuspending cell pellet in prewarmed broth media following 15-minute incubation at 60˚C.   To generate low nutrient samples, actively growing Lactobacillus plantarum (ATCC 8014) culture was harvested by centrifugation. Low nutrient stress was induced for 15 minutes beginning with excessive washing of the cell pellet with Low nutrient media (0.5x LB broth), followed by centrifugation. Washed cell pellet was resuspended in 0.5x LB broth, incubated at 37˚C for the remaining time.   Polyribosome fractions were generated as described in PMID: 27146487 with minor modifications. For short, exponentially grown bacterial cells were harvested by centrifugation (5 minutes) at 4˚C on ice supplemented with 100µg/ml Chloramphenicol. Lysis was done in 1xTN buffer (50mM TRIS/HCl pH 7.4,150mM NaCl, 1mM DTT, 100μg/ml CAM and Complete EDTA Free Protease inhibitor tablet) using glass beads and of vortexing (2minutes) followed by resting of sample on ice (5 minutes).  Vortexing/resting cycle was repeated two more times. Lysate was cleared by centrifugation at 1500g for 5 minutes at 4˚C. Resulting supernatant was loaded on to a 15-50% sucrose gradient with a 80% cushion and sedimentation was accomplished by ultracentrifugation for 90 minutes at 36,000rpm. Fractions were collected and A254 value was monitored. Subsequently, RNA was extracted from sucrose fractions by adding equal volumes of phenol/chloroform isoamyl alcohol pH 4.5 (25:24:1) and nuclease free water followed by 2-minute vortexing and centrifugation. Aqueous phase was further cleaned up by the addition of Chloroform, vortexing and centrifugation. Resulting aqueous phase was sodium acetate-ethanol precipitated.

tag: treatment

value: 100microg/ml Chloramphenicol incubated for 5 minutes at 37C

tag: culture conditions

value: bsub-str168_cam_rep3 was grown in 2xYT broth at 37C. Chloramphenicol treatment initiated at exponential growth OD600 0.6

tag: library protocol

value: Standard 5PSeq, RNA rP5_RND oligo for RNA ligation

tag: ribosomal rna delpetion strategy

value: Ribo-Zero rRNA Removal Kit (Bacteria) Illumina


title: c_3_01180AAC_TGACCA_batch2

city: Barcelona

first: Oscar

last: Reina Garcia

state: Barcelona

address: C/Baldiri Reixac 10

department: Biostatistics and Bioinformatics

country: Spain

institute: IRB Barcelona

data_processing:
FastQ files were aligned against the mm10 reference genome using STAR 2.5.2b using default options.
Gene counts wer generated with the featureCounts function from the RSubread package version 1.32.4 with options: annot.inbuilt='mm10',allowMultiOverlap=TRUE,countMultiMappingReads=FALSE,minMQS=1,ignoreDup=FALSE
Genome_build: mm10
Supplementary_files_format_and_content: Excel XLSX

molecule: polyA RNA

source_name: epidermal cells

growth_protocol: Mice were aged in our animal facilities until treatment was performed at 73 weeks of age

extract_protocol:
Trizol and Rneasy Mini kit
Libraries were prepared using the TruSeq stranded mRNA Library Prep (96 samples ref. 20020595 or 48 samples ref. 20020594) according to the manufacturer's protocol,  to convert total RNA into a library of template molecules of known strand origin and suitable for subsequent cluster generation and DNA sequencing.  Briefly, 1000-500 ng of total RNA were used for poly(A)-mRNA selection using poly-T oligo attached magnetic beads using two rounds of purification. During the second  elution of the poly-A RNA, the RNA was fragmented under elevated temperature and primed with random hexamers for cDNA synthesis.  Then, the cleaved RNA fragments were copied into first strand cDNA using reverse transcriptase (SuperScript II, ref. 18064-014, Invitrogen) and random primers. The  addition of Actinomycin D to the First Stand Synthesis Act D mix (FSA) prevents spurious DNA-dependent synthesis, while allowing RNA-dependent synthesis, improving  strand specificity.  After that, second strand cDNA was synthesized, removing the RNA template and synthesizing a replacement strand, incorporating dUTP in place of dTTP to generate ds  cDNA using DNA Polymerase I and RNase H.  These cDNA fragments, then had the addition of a single 'A' base to the 3’ends of the blunt fragments to prevent them from ligating to one another during the adapter  ligation. A corresponding single T nucleotide on the 3’ end of the adapter provides a complementary overhang for ligating the adapter to the fragments ensuring a low rate  of chimera (concatenated template) formation. Subsequent ligation of the multiple indexing adapter to the ends of the ds cDNA.  Finally, PCR selectively enriched those DNA fragments that had adapter molecules on both ends and to amplify the amount of DNA in the library. The PCR was performed with a PCR Primer Cocktail that anneals to the ends of the adapters.  Final libraries were analyzed using Bioanalyzer DNA 1000 or Fragment Analyzer Standard Sensitivity (ref: 5067-1504 or ref: DNF-473, Agilent) to estimate the quantity and   validate the size distribution, and were then quantified by qPCR using the KAPA Library Quantification Kit KK4835 (REF. 07960204001, Roche) prior to the amplification with Illumina’s cBot.

treatment_protocol: Mice were injected intraperitoneally with blocking antibodies for IL-17A/F 3 times a week for 12 weeks

tag: cell type

value: epidermal cells

tag: strain

value: 73 weeks

value: Control IgG

tag: sequencing batch


title: IL_1_01182AAC_GATCAG_batch2

value: IL-17A/F blocking antibodies


title: bsub-str168_frag_r1_s_r_v2

value: Untreated

value: bsub-str168_frag_rep1 was grown in 2xYT broth at 37C and harvested at exponential growth OD600 0.416

value: random fragmented ( Standard 5PSeq), RNA rP5_RND oligo for RNA ligation


title: C4-2B_LKO

country: Taiwan

organism: Homo sapiens

value: C4-2B (ATCC number:CRL-3315)

value: Caucasian prostate cancer

value: control


title: Full nutrient time 6h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours on full nutrient medium

value: Roots

value: 10 days


title: Phosphate deprivation time 6h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours in a phosphate deprived  medium


title: Phosphate & Iron deprivation time 3h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours on a phosphate and iron deprived medium


title: bsub-str168_cam_r2_s_r_v2

value: bsub-str168_cam_rep2 was grown in 2xYT broth at 37C. Chloramphenicol treatment initiated at exponential growth OD600 0.621


title: Iron deprivation time 3h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours on an iron deprived medium


title: c_3_01180AAC_TGACCA_batch1


title: NIH3T3 cells, sh#3_1836-D1_H1

value: UHMK1 knockdown

value: shUHMK1#3 (sh#3) sequence


title: bsub-str168_cam_r1_s_r_v2

value: 100microg/ml Chloramphenicol incubated for 5 minutes at 37C

value: bsub-str168_cam_rep1 was grown in 2xYT broth at 37C. Chloramphenicol treatment initiated at exponential growth OD600 0.416

value: Standard 5PSeq,RNA rP5_RND oligo for RNA ligation


title: c_4_01181AAC_CAGATC_batch1

value: epidermal cells

value: 73 weeks


title: NIH3T3 cells, sh#2_1137-D1_D2

value: shUHMK1#2 (sh#2) sequence


title: c_2_01179AAC_TTAGGC_batch2

value: Control IgG


title: bsub-str168_ctr_r3_s_r_v2

value: Untreated

value: bsub-str168_ctr_rep3 was grown in 2xYT broth at 37C and harvested at exponential growth OD600 0.6

value: Ribo-Zero rRNA Removal Kit (Bacteria) Illumina


title: Phosphate deprivation time 3h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours in a phosphate deprived  medium


title: NIH3T3 cells, sh#1_907-D1_H2

value: shUHMK1#1 (sh#1) sequence


title: c_2_01179AAC_TTAGGC_batch1


title: bsub-str168_ctr_r2_s_r_v2

value: bsub-str168_ctr_rep2 was grown in 2xYT broth at 37C and harvested at exponential growth OD600 0.621

value: Standard 5PSeq, RNA rP5_RND oligo for RNA ligation


title: Full nutrient time 3h Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours on full nutrient medium

value: Roots

value: 10 days


title: NIH3T3 cells, shCTRL_SCR2-D1_F1

value: Control - scramble

value: scrambled shRNA-UHMK1 sequence (shCTRL) which has target gene


title: Control full nutrient time 0 Rep3

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium


title: c_1_01178AAC_CGATGT_batch1


title: bsub_hs03_v17 [mixed sample]

description: Culture: The cultures were grown in LB Broth at 37˚C and harvested in log growth phase (UT); In case of mutation strains those were supplemented with 100µg/ml Spectinomycin (rnjA) or 5µg/ml Kanamycin (rnjB, rny) for selection; In case of HS cultures 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN)  rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

data_processing:
library strategy:  5PSeq that captures  transcripts with free 5' phospahte (without cap)
Adapter and quality trimming with the bbduk program of the BBTools suite, with options {qtrim=r, ktrim=r, hdist=3, hdist2=2, K=20, mink=14, trimq=16, minlen=30, maq=16}, using BBTools default adapter set and polyG or polyA sequences for short reads.
Deduplication of reads with both identical unique molecular identifier (UMI) and insert sequence using the dedupe program of the BBTools suite with its default parameters.
UMI sequences found in the first eight bases of each read were extracted using UMI-tools (version 1) with default options (using --bc-pattern NNNNNNNN).
Dual index fastq samples were processed as described in supplementary file dual_index_demultiplexing.txt
Alignment was performed with the bbmap program of the BBTools suite, with the {32bit=t -da -eoom k=11 strictmaxindel=10 intronlen=0 t=16 trd=t minid=0.94 nzo=t} parameters. Alignment files were sorted and indexed with SAMtools.
Deduplication based on UMIs was then performed with UMI-tools (version 1) with options { --soft-clip-threshold 1 --edit-distance 2 --method unique}.
In selected samples, a dual undex strategy was used (sequencing runs v17). In brief, in addition of the Illumina indexes with dedicated reads used in HT-5Pseq, an additional index sequence is added before the UMI sequence. This is introduced during the ssRNA ligation space using anoligo with the following structure Illumina compatible-CCCCCC-NNNNNNNN. Please see dual_index_demultiplexing.txt for details on demultiplexing strategy for those samples.
Bedgraphs for read 5' positions for forward (fwd) and reverse (rev) strands were generated with BEDTools genomecov program with options {-bg -strand [+/-] -5}.
Assembly: The genome used for alignment is indicated in the processed file names with the G-xxxx patterns.  Those correspond to the NCBI accessions and builds indicated in the Supplementary file.
Supplementary files format and content: Reference indiced for individual bacterial species and custom mixtures were built with the bbmap program of the BBTools suite, with default options (and k=10).
Supplementary files format and content: Processed files are bedgraphs containing read 5' counts on reverse (_rev) and forward (_fwd) strands of genomes indicated with _G-xxxx pattern. The genomes corresponding to each pattern are described in the genome build entry above.

source_name: A mixed sample of 8 cultures:  bsub-wt_ctr_r3_s_o_v17, bsub-wt_hs_r3_s_o_v17, bsub-rnja_ctr_r3_s_o_v17, bsub-rnja_hs_r3_s_o_v17, bsub-rnjb_ctr_r3_s_o_v17, bsub-rnjb_hs_r3_s_o_v17, bsub-rny_ctr_r3_s_o_v17, bsub-rny_hs_r3_s_o_v17

growth_protocol: Bacteria were grown according to their requirements. Overnight Cultures were diluted to OD600 0.01-0.07, growth was continued and cells were harvested in log-phase by centrifugation. Resulting pellet was shock-frozen in Dry ice/Ethanol bath and stored for RNA analysis

extract_protocol:
Microbial cell pellets were resuspended in equal volume of LET (25 mM Tris pH 8.0, 100 mM LiCl, 20 mM EDTA) and water saturated Phenol pH 6.6 (Thermo Fisher),  lysed with acid washed glass beads (Sigma-Aldrich) and vortexing for three minutes in MultiMixer. Lysates were supplemented  with equal volumes of phenol/chloroform isoamyl alcohol pH 4.5 (25:24:1) and nuclease free water, l two-minute vortexing followed by centrifugation. Resulting aqueous phase was purified in two steps using phenol-chloroform isoamyl alcohol (25:24:1) followed by chloroform. After centrifugation, the clean aqueous phase was precipitated with sodium acetate-ethanol.
Standard and random fragmented 5PSeq libraries were generated as described in PMID: 26046441 with minor modifications using 150-9000ng total RNA as an input. To prepare random fragmented samples (negative controls), ribosomal RNA was depleted from DNA-free RNA and subsequent fragmented by incubating five minutes at 80 ̊C in fragmentation buffer (40mM Tris Acetate pH 8.1, 100mM KOAc and 30mM MgOAc). Reaction was purified using 2 volumes of RNACleanXP beads (Beckman Coulter) as recommended by the manufacturer. Free 5’OH sites were re-phosphorylated using 5 Units of T4 Polynucleotide kinase (PNK, NEB) and incubated at 37 ̊C for 60 minutes as recommended by the manufacturer. Re-phosphorylated fragmented RNA was purified using Phenol:Chloroform: Isoamyl Alcohol (24:25:1), followed by sodium acetate-ethanol precipitation. From this step forward, procedures for random fragmented and standard 5PSeq library preparation merge. RNA was Ligated to either rP5_RND or rP5_RNA oligo (specified in characteristics protocol).    containing unique molecular identifiers). Ribosomal RNA was depleted using Ribozero rRNA removal kit (Illumina) suitable for Bacteria and Yeast. rRNA depleted sample was purified using 1.8V of Ampure beads (Abcam), fragmented using heat (80˚C) for 5 min in 5x Fragmentation Buffer (200mM Tris Acetate pH 8.1,500mM KOAc,150mM MgOAc) and reverse transcribed using random hexamers to prime. Resulting cDNA was bound to streptavidin beads (M-280), subjected to enzymatic reactions of DNA end repair, fill-in of adenine to 5’ protruding ends of DNA fragments using Klenow Fragment (NEB). Common adaptor (P7-MPX) was ligated and 5PSeq Libraries were amplified by PCR (15- 17 cycles). Purified samples were quantified using Qubit (Thermo Fisher), library size was estimated from bioanalyzer traces. 5PSeq Libraries were pooled by mixing equal amounts of each sample, following enrichment of fragments of the length between 300-500nt.   HT-5PSeq Libraries were generated as recently described (bioRxiv 2020.06.22.165134; doi: https://doi.org/10.1101/2020.06.22.165134). In brief, DNA-free RNA was ligated with RNA oligos containing unique molecular identifiers. Ligated RNA was reverse transcribed priming with oligos containing a random hexamer and an Illumina compatible region. RNA was eliminated by addition of NaOH. Ribosomal RNA was depleted by adding in-house rRNA DNA oligo depletion mixes (table 2) to the cDNA and performing a duplex-specific nuclease (DSN, Evrogen) treatment. rRNA depleted cDNA was PCR amplified (15-17 cycles). Depletion of ribosomal RNA with Ribozero Illumina (for bacteria and yeast) was done after the single-stranded RNA ligation step. Ribosomal depleted RNA was purified and reverse transcribed using the same oligos as stated above, and then amplified by PCR. Libraries were quantified by fluorescence (Qubit, Thermo Fisher), size estimated using an Agilent Bioanalyzer and sequence using a NextSeq500 Illumina sequencer.

value: wild type; rnjA delta, rnjB delta, rny delta

value: Untreated (UT) or Heat shock at 65˚C for 10 minutes (HS)

tag: molecule subtype

value: total RNA, 5' monophosphorylated


title: bsub_hs02_v17 [mixed sample]

source_name: A mixed sample of 8 cultures:  bsub-wt_ctr_r2_s_o_v17, bsub-wt_hs_r2_s_o_v17, bsub-rnja_ctr_r2_s_o_v17, bsub-rnja_hs_r2_s_o_v17, bsub-rnjb_ctr_r2_s_o_v17, bsub-rnjb_hs_r2_s_o_v17, bsub-rny_ctr_r2_s_o_v17, bsub-rny_hs_r2_s_o_v17


title: NIH3T3 cells, shCTRL_SCR1-D1_B1


title: Y12_3

data_processing:
FastQ files were aligned against the mm9 reference genome using STAR 2.5.2b using default options.
mm10 liftover BAM files were generated with CrossMaps
Gene counts were generated with the featureCounts function from the RSubread package version 1.32.4 with options: annot.inbuilt='mm10',allowMultiOverlap=TRUE,countMultiMappingReads=FALSE,minMQS=1,ignoreDup=FALSE
Genome_build: mm9
Supplementary_files_format_and_content: Excel XLSX

extract_protocol:
Trizol and Rneasy Mini kit
Libraries were prepared using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Human/Mouse/Rat Kit (REF. RS-122-2201/2202, Illumina) according to the manufacturer's protocol. Briefly, from 150 - 300 ng of total RNA were used for ribosomal RNA depletion. Then, ribosomal depleted RNA was fragmented for 4.5 min at 94  C. The remaining steps of the library preparation were followed according to the manufacturer's instructions.Final libraries were analyzed on an Agilent Technologies 2100 Bioanalyzer system using the Agilent DNA 1000 chip to estimate the quantity and validate the size distribution, and were then quantified by qPCR using the KAPA Library Quantification Kit KK4835 (REF. 07960204001, Roche) prior to the amplification with Illumina's cBot. Finally, libraries were sequenced on the Illumina HiSeq 2500 sequencing system using paired-end 50 base pair long reads.

treatment_protocol: 210 ug of control IgG1 (clone MOPC-2, BioXCell) were injected intraperitoneally in 100ul 3 times per week for 12 weeks.

value: epidermal cells

value: Young (17-25 weeks)


title: Y12_4


title: bsub_hs01_v17 [mixed sample]

source_name: A mixed sample of 8 cultures:  bsub-wt_ctr_r1_s_o_v17, bsub-wt_hs_r1_s_o_v17, bsub-rnja_ctr_r1_s_o_v17, bsub-rnja_hs_r1_s_o_v17, bsub-rnjb_ctr_r1_s_o_v17, bsub-rnjb_hs_r1_s_o_v17, bsub-rny_ctr_r1_s_o_v17, bsub-rny_hs_r1_s_o_v17


title: EpiSC 0.75

city: Leiden

first: Christian

last: Schröter

state: Zuid-Holland

address: Einsteinweg 55

department: Cell Systems and Drug Safety

country: Netherlands

institute: Leiden University, Leiden Academic Centre for Drug Research

description: 10x Genomics

library_source: transcriptomic single cell

data_processing:
Demultiplexing, alignment to the mouse genome mm10 (GENCODE vM23/Ensembl 98, from 10x Genomics) and read quantification was performed with CellRanger (10x Genomics, v4.0.0).
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: EpiSC

extract_protocol: Single cell transcriptomes were generated with the 10x Genomics Chromium Next GEM Single Cell 3ʹ Reagent Kit v3.1 according to the manufacturer's instructions. Briefly, cells were differentiated for three days, detached with Accutase and counted. The cell suspension was diluted to a concentration of 900 cells/μl, to recover around 2000 cells per sample. Cell suspensions were loaded on a Chromium Controller (10x Genomics) to partition cells with gel beads in emulsion.  Reverse transcription, cDNA recovery and amplification, and sequencing library construction were performed according to manufacturer’s instructions (10x Genomics ChromiumNextGEMSingleCell_v3.1_Rev_D). We chose 12 PCR cycles for cDNA amplification, and 10 PCR cycles for index PCR.

treatment_protocol: Mesoderm differentiation of EpiSCs was performed by culture in N2B27 supplemented with 1 µM CHIR99201, 8 ng/ml BMP, and indicated concentrations of FGF2 or AZD4547 for three days.

value: EpiSC

value: 0.75 ng/ml FGF2


title: EpiSC 12

description: 10x Genomics

value: 12 ng/ml FGF2


title: EpiSC azd

description: 10x Genomics


title: Phosphate & Iron deprivation time 9h Rep2


title: Full nutrient time 9h Rep2


title: Y12_1

value: Control IgG


title: bsub-rny_hs_r3_s_o_v17

description: Culture: bsub-rny_hs_r3_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

source_name: Liquid culture of ribonuclease Y deletion strain (rny) Bacillus subtilis 168 trpC2, supplemented with 5µg/ml Kanamycin for selection

value: rny delta

value: Heat shock at 65˚C for 10 minutes


title: Phosphate deprivation time 9h Rep2


title: NIH3T3 cells, shCTRL_SCR3-D1_B4

extract_protocol:
Libraries were constructed using a bulk RNA-seq method called Prime-seq, which uses direct cell lysis, bead cleanup, cDNA synthesis and pre-aplification with KAPA HiFi Polymerase (see Janjic et al, 2021)
The cells were counted by Vi-Cell XR (Beckman Coulter, Brea, CA, USA) and 1x104 cells were harvested in microtubes containing 100 µl of DPBS 1x. After 5 minutes of centrifugation at 1500 rpm, cells were resuspended in 100 µl of Buffer RLT Plus, RNeasy Plus Lysis Buffer (Qiagen, Hilden, Germany) supplemented with 1% b-mercapoethanol.
Prime-seq (Janjic et al, 2021 -  https://doi.org/10.1101/2021.09.27.459575)


title: bsub-rny_ctr_r3_s_o_v17

description: Culture: bsub-rny_ctr_r3_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C and harvested in log growth phase  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: Untreated


title: Phosphate & Iron deprivation time 6h Rep2


title: c_1_01178AAC_CGATGT_batch2

value: 73 weeks


title: Iron deprivation time 9h Rep2

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours on an iron deprived medium

value: Roots


title: Iron deprivation time 6h Rep2

value: 10 days


title: O12_4

value: Old (73 weeks)


title: EpiSC 3

description: 10x Genomics

value: 3 ng/ml FGF2


title: bsub-str168_ctr_r1_s_r_v2

value: bsub-str168_ctr_rep1 was grown in 2xYT broth at 37C and harvested at exponential growth OD600 0.416

value: Standard 5PSeq, RNA rP5_RND oligo for RNA ligation

value: Ribo-Zero rRNA Removal Kit (Bacteria) Illumina


title: EpiSC 0

description: 10x Genomics

value: no FGF2


title: Phosphate deprivation time 6h Rep2


title: O12_3

value: epidermal cells


title: Y12_2

value: Young (17-25 weeks)


title: Full nutrient time 6h Rep2


title: O12_2


title: VacV-Ii-GP61 infected skin (2)

city: Portland

first: Jeffrey

last: Nolz

state: OR

address: 3181 SW Sam Jackson Park Road

country: USA

institute: Oregon Health & Science University

data_processing:
Sequence file QC was done using FastQC (Babraham Institute)
Reads were trimmed using Trimmomatic
Trimmed sequencers were aligned to GRCm38/mm10 mouse reference genome with STAR aligner
The resulting bam file was then indexed using SAMTools
Assembly: mm10
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

source_name: whole ear skin

extract_protocol:
Whole ear skin was homogenized in 1 ml of Trizol and RNA was extracted according to the manufacturers protocol. RNA was further purified using the RNeasy Mini Kit (Qiagen). RNA integrity was determined by running samples on the Bioanalyzer (Agilent).
Libraries were prepared using the TruSeq Stranded mRNA kit according to the manufacturers protocol (Illumina)
Libraries were quantified for sequencing using an NGS Library Quantification Kit (Roche/Kapa Biosystems) on a StepOnePlus Real Time PCR Workstation (Thermo/ABI). Sequencing was done on a NovaSeq 6000 (Illumina).

treatment_protocol: LCMV-immune C57Bl/6 mice were infected on the right ear skin with VacV and on the left ear skin with VacV-Ii-GP61

value: whole ear skin

value: WT - LCMV Immune

value: VacV-Ii-GP61 infected skin

tag: time

value: Day 3


title: VacV-Ii-GP61 infected skin (3)


title: O12_1

value: Control IgG


title: VacV-Ii-GP61 infected skin (1)


title: bsub-rnja_ctr_r3_s_o_v17

description: Culture: bsub-rnja_ctr_r3_s_o_v17 was cultured in LB Broth supplemented with 100µg/ml Spectinomycin for selection at 37˚C and harvested in log growth phase; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

source_name: Liquid culture of ribonuclease J1 deletion strain (rnjA) Bacillus subtilis 168 trpC2, supplemented with 100µg/ml Spectinomycin for selection

value: rnjA delta

value: Untreated

value: total RNA, 5' monophosphorylated


title: VacV-infected skin (3)

value: VacV-infected skin


title: bsub-rnjb_hs_r3_s_o_v17

description: Culture: bsub-rnjb_hs_r3_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

source_name: Liquid culture of ribonuclease J2 deletion strain (rnjB) Bacillus subtilis 168 trpC2, supplemented with 5µg/ml Kanamycin for selection

value: rnjB delta

value: Heat shock at 65˚C for 10 minutes


title: VacV-infected skin (2)


title: bsub-wt_hs_r3_s_o_v17

description: Culture: bsub-wt_hs_r3_s_o_v17 was cultured in LB Broth at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers.  rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

source_name: Liquid culture of Bacillus subtilis 168 trpC2

value: wild type


title: ChIP_P40_S31: MalePoly9_H3K4me3

city: Freiburg

first: Ori

last: Staszewski

address: Breisacher Str. 64

department: Institute of Neuropathology

institute: University Medical Center Freiburg

library_source: genomic

data_processing:
CASAVA1.8.2 software used for basecalling
fastq files were quality controlled with FASTQC v 0.11
Alignment to GRCm36 was performed with STAR_2.5.2b
ChIPseq Peak calling was performed with PePr
bigWig files were generated using deeptools 3.1.1 bamCoverage
Genome_build: GRCm36
Supplementary_files_format_and_content: bigWig files with per sample ChIPseq signal

molecule: genomic DNA

source_name: Microglia

extract_protocol:
Mice were anesthetized and transcardially perfused with ice cold PBS. Brain hemispheres were homogenized and subjected to 37% / 70% percoll density gradient separation. Microglia cells were collected from the pellet of 37% percoll and washed extensively with FACS buffer (0.5% FBS in PBS) followed by staining with anti-CD11b (clone M1/70, eBioscience) and anti-CD45 (clone 30-F11, eBioscience) antibodies. Isolation of immune cells and flow cytometry.
ChIP-Seq libraries were constructed using the KAPA Library Preparation Kit with Real-time PCR Library Amplification as per manufacturers instructions.

value: Whole brain

tag: Sex

value: Male

value: Poly IC


title: ChIP_P40_S32: MalePoly9_H3K9ac


title: Iron deprivation time 3h Rep2

value: Roots

value: 10 days


title: Phosphate deprivation time 3h Rep2


title: bsub-wt_ctr_r3_s_o_v17

description: Culture: bsub-wt_ctr_r3_s_o_v17 was cultured in LB Broth at 37˚C and harvested in log growth phase; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: bsub-rnja_hs_r3_s_o_v17

description: Culture: bsub-rnja_hs_r3_s_o_v17 was cultured in LB Broth supplemented with 100µg/ml Spectinomycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: bsub-rny_hs_r2_s_o_v17

description: Culture: bsub-rny_hs_r2_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: rny delta


title: bsub-rnjb_ctr_r3_s_o_v17

description: Culture: bsub-rnjb_ctr_r3_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C and harvested in log growth phase  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: Phosphate & Iron deprivation time 3h Rep2


title: ChIP_P40_S30: MalePoly8_Input


title: Control full nutrient time 0 Rep2


title: ChIP_P40_S29: MalePoly8_H3K9ac


title: bsub-rny_ctr_r2_s_o_v17

description: Culture: bsub-rny_ctr_r2_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C and harvested in log growth phase  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: PAC-seq siRRP40 mESC Replicate 3

city: Boston

first: Claudia

middle: A

last: Mimoso

state: MA

address: 45 Shattuck St. LHRRB

institute: Harvard Medical School

description:
211012_mm10_mESC_PACseq_RRP40KD_N4_DepthNorm_F.bedGraph
211012_mm10_mESC_PACseq_RRP40KD_N4_DepthNorm_R.bedGraph

data_processing:
PAC-seq libraries were processed as described in Routh et al., 2017, with the following modifications: First, trimmed and filtered reads were mapped to the Drosophila genome (dm6) using HISAT2. Reads not mapping to the spike genome were aligned to the mouse genome (mm10) using HISAT2.
For final read processing, a minimum of 5 non-primer/non-template As was required for each unique poly(A) tail and a read count filter per nucleotide position was not applied.
Assembly: mm10
Supplementary files format and content: bedGraph
Library strategy: PAC-seq

source_name: F121-9 wt mESCs

growth_protocol: mESCs were cultured on gelatin in KnockOut DMEM (ThermoFisher, 10829018), supplemented with 15% KO serum replacement (ThermoFisher, 10828028), 1X penicillin-streptomycin (MP, TMS-005-C), 1X non-essential amino acids (MP, TMS-001-C), 1% b-ME (MP, ES-007-E), 1X GlutaMAX (ThermoFisher, 35050061), 1000 U/ml LIF (Cell Guidance Systems, GFM200), 1 µM MEK inhibitor (Stemgent, PD0325901), and 3 µM GSK3 inhibitor (Stemgent, CHIR99021) mESCs were fed daily and passaged every two days. mESCs were cultured at 37°C with 5% CO2.

extract_protocol:
For SCR and U1 AMO conditions, libraries were constructed from 250 ng of enriched 4sU-labelled RNA (as described under TT-seq extraction protocol). For untreated mESC, siNT and siRRP40 conditions, 1 μg of total RNA (as described under RNA-seq extraction protocol) was used for library construction.
PAC-seq libraries were constructed using the PolyA-ClickSeq Library Kit (ClickSeq Technologies)

treatment_protocol: See individual samples.

tag: cell line

value: F121-9 wt mESCs

value: Mouse embryonic stem cells

value: WT

value: siRNA pool against RRP40


title: ChIP_P40_S33: MalePoly9_Input

tag: tissue


title: ChIP_P40_S28: MalePoly8_H3K4me3


title: bsub-rnjb_hs_r2_s_o_v17

description: Culture: bsub-rnjb_hs_r2_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: bsub-rnjb_ctr_r2_s_o_v17

description: Culture: bsub-rnjb_ctr_r2_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C and harvested in log growth phase  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN)  which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: VacV-infected skin (1)

value: whole ear skin

value: WT - LCMV Immune

value: Day 3


title: bsub-rnja_hs_r2_s_o_v17

description: Culture: bsub-rnja_hs_r2_s_o_v17 was cultured in LB Broth supplemented with 100µg/ml Spectinomycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: rnjA delta

value: total RNA, 5' monophosphorylated


title: ChIP_P40_S27: MalePoly6_Input


title: PAC-seq siRRP40 mESC Replicate 1


title: PAC-seq siRRP40 mESC Replicate 2


title: ChIP_P40_S26: MalePoly6_H3K9ac


title: bsub-rnja_ctr_r2_s_o_v17

description: Culture: bsub-rnja_ctr_r2_s_o_v17 was cultured in LB Broth supplemented with 100µg/ml Spectinomycin for selection at 37˚C and harvested in log growth phase; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: Untreated


title: Full nutrient time 9h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours on full nutrient medium

tag: ecotype


title: PAC-seq siRRP40 mESC Replicate 4


title: bsub-wt_hs_r2_s_o_v17

description: Culture: bsub-wt_hs_r2_s_o_v17 was cultured in LB Broth at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: wild type

value: Heat shock at 65˚C for 10 minutes


title: PAC-seq siNT mESC Replicate 3

description:
211012_mm10_mESC_PACseq_NT_N4_DepthNorm_F.bedGraph
211012_mm10_mESC_PACseq_NT_N4_DepthNorm_R.bedGraph

value: Non-targeting siRNA control


title: PAC-seq siNT mESC Replicate 4


title: ChIP_P40_S25: MalePoly6_H3K4me3

value: Whole brain

tag: age

value: Male

value: Poly IC


title: bsub-wt_ctr_r2_s_o_v17

description: Culture: bsub-wt_ctr_r2_s_o_v17 was cultured in LB Broth at 37˚C and harvested in log growth phase; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: ChIP_P40_S24: MaleCtr7_Input

value: Control


title: PAC-seq siNT mESC Replicate 2

value: F121-9 wt mESCs


title: Iron deprivation time 9h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours on an iron deprived medium

value: Roots

value: 10 days


title: Phosphate deprivation time 9h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours in a phosphate deprived  medium


title: ChIP_P40_S23: MaleCtr7_H3K9ac


title: Phosphate & Iron deprivation time 6h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours on a phosphate and iron deprived medium


title: Full nutrient time 6h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours on full nutrient medium


title: Full nutrient time 3h Rep2

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours on full nutrient medium


title: Iron deprivation time 6h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours on an iron deprived medium


title: Phosphate & Iron deprivation time 9h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 9hours on a phosphate and iron deprived medium


title: Phosphate deprivation time 6h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 6hours in a phosphate deprived  medium


title: bsub-rny_hs_r1_s_o_v17

description: Culture: bsub-rny_hs_r1_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers.  rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: rny delta


title: ChIP_P40_S22: MaleCtr7_H3K4me3


title: PAC-seq  WT mESC Replicate 3

description:
211012_mm10_mESC_PACseq_WT_N3_DepthNorm_F.bedGraph
211012_mm10_mESC_PACseq_WT_N3_DepthNorm_R.bedGraph

value: Mouse embryonic stem cells

value: WT


title: PAC-seq siNT mESC Replicate 1


title: ChIP_P40_S21: MaleCtr6_Input


title: bsub-rny_ctr_r1_s_o_v17

description: Culture: bsub-rny_ctr_r1_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C and harvested in log growth phase  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN)  which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: PAC-seq  WT mESC Replicate 2


title: ChIP_P40_S20: MaleCtr6_H3K9ac


title: bsub-rnjb_hs_r1_s_o_v17

description: Culture: bsub-rnjb_hs_r1_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: rnjB delta


title: Phosphate & Iron deprivation time 3h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours on a phosphate and iron deprived medium


title: U1 AMO PRO-seq Replicate 3

description:
201029_mm10_mESC_PROseq_N3_1.34NormEach_U1_AMO_F.bedGraph
201029_mm10_mESC_PROseq_N3_1.34NormEach_U1_AMO_R.bedGraph

data_processing:
mESC libraries were mapped as follows: using a custom script (trim_and_filter_PE.pl), FASTQ read pairs were trimmed to 41 bp per mate and read pairs with a minimum average base quality score of 20 were retained. Read pairs were further trimmed using cutadapt 1.14 to remove adapter sequences and low-quality 3’ bases (--match-read-wildcards -m 20 -q 10).
R1 reads, corresponding to RNA 3’ ends, were then aligned to the spiked in Drosophila genome index (dm6) using Bowtie 1.2.2 (-v 2 -p 6 --best --un), with those reads not mapping to the spike genome serving as input to the primary genome alignment step (mm10, using Bowtie 1.2.2 options -v 2 --best).
For HEK293T libraries, the following mapping pipeline was used: Dual, 6nt Unique Molecular Identifiers (UMIs) were extracted from read pairs using UMI-tools. Read pairs were trimmed using cutadapt 1.14 to remove adapter sequences (-O 1 --match-read-wildcards -m 26). The UMI length was trimmed off the end of both reads to prevent read-through into the mate’s UMI, which will happen for shorter fragments. An additional nucleotide was removed from the end of read 1 (R1), using seqtk trimfq (https://github.com/lh3/seqtk), to preserve a single mate orientation during alignment.
The paired end reads were then mapped to a combined genome index, including both the spike (dm6) and primary (hg38) genomes, using bowtie2. Properly paired reads were retained. These read pairs were then separated based on the genome (i.e., spike-in vs primary) to which they mapped, and both these spike and primary reads were independently deduplicated using UMI-tools.
For all libraries, reads mapping to the reference genome were then sorted, via samtools 1.3.1 (-n), and subsequently converted to bedGraph format using a custom script (bowtie2stdBedGraph.pl) that counts each read once at the exact 3’ end of the nascent RNA. Because R1 in PRO-seq reveals the position of the RNA 3’ end, the “+” and “-“ strands were swapped to generate bedGraphs representing 3’ end positions at single nucleotide resolution.
BedGraphs were normalized using the normalize_bedGraph custom script. For libraries generated from control mESCs and HEK293Ts, no further normalization was performed. For libraries generated under SCR or U1 AMO conditions, we observed a consistent increase in reads mapping to the fly genome (dm6) after U1 inhibition (Average ratio of U1 AMO / SCR AMO = 1.34), indicating a global decrease in RNAPII transcription in U1 AMO cells compared to SCR AMO control. Accordingly, the average ratio of reads mapping to the spike genome between U1 and SCR AMO conditions was used to generate spike normalization factors
Combined bedGraphs were generated by summing counts per nucleotide across replicates for each condition (bedgraphs2stdBedgraph).
Assembly: mm10 (mESC); hg38 (HEK293T)
Supplementary files format and content: bedGraph
Library strategy: PRO-seq

molecule: nuclear RNA

source_name: Mouse embryonic stem cells

growth_protocol: mESCs were cultured on gelatin in KnockOut DMEM (ThermoFisher, 10829018), supplemented with 15% KO serum replacement (ThermoFisher, 10828028), 1X penicillin-streptomycin (MP, TMS-005-C), 1X non-essential amino acids (MP, TMS-001-C), 1% b-ME (MP, ES-007-E), 1X GlutaMAX (ThermoFisher, 35050061), 1000 U/ml LIF (Cell Guidance Systems, GFM200), 1 µM MEK inhibitor (Stemgent, PD0325901), and 3 µM GSK3 inhibitor (Stemgent, CHIR99021) mESCs were fed daily and passaged every two days. HEK293Ts were cultured in DMEM (VWR, 45000-312) with 10% FBS (Thermo, 1600044). mESCs and HEK293Ts were cultured at 37°C with 5% CO2.

extract_protocol:
Cells were permeabilized as in Reimer et al. 2021; run-ons were performed with biotinylated NTPs and then purified.
Purified biotinylated RNAs were ligated to sequencing adapters and amplified by PCR.

treatment_protocol: See individual samples

value: F121-9 mESCs with a fluorescent splicing reporter system as described in Vlaming et al., 2022

value: 20uM U1 AMO, 4 hours.


title: bsub-rnjb_ctr_r1_s_o_v17

description: Culture: bsub-rnjb_ctr_r1_s_o_v17 was cultured in LB Broth supplemented with 5µg/ml Kanamycin for selection at 37˚C and harvested in log growth phase  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN)  which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: Untreated

value: total RNA, 5' monophosphorylated


title: U1 AMO PRO-seq Replicate 2


title: ChIP_P40_S18: MaleCtr5_Input


title: U1 AMO PRO-seq Replicate 1


title: bsub-rnja_hs_r1_s_o_v17

description: Culture: bsub-rnja_hs_r1_s_o_v17 was cultured in LB Broth supplemented with 100µg/ml Spectinomycin for selection at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation;  ; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers.  rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: rnjA delta

value: Heat shock at 65˚C for 10 minutes


title: ChIP_P40_S17: MaleCtr5_H3K9ac

value: Whole brain


title: ChIP_P40_S16: MaleCtr5_H3K4me3

value: Male


title: bsub-rnja_ctr_r1_s_o_v17

description: Culture: bsub-rnja_ctr_r1_s_o_v17  was cultured in LB Broth supplemented with 100µg/ml Spectinomycin for selection at 37˚C and harvested in log growth phase; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers. rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: bsub-wt_hs_r1_s_o_v17

description: Culture: bsub-wt_hs_r1_s_o_v17 was cultured in LB Broth at 37˚C to log growth phase, 2 ml of culture was centrifuged and pellet was resuspended in 65˚C pr-warmed LB, incubated for 10 minutes at 65˚C on Heatblock and harvested by centrifugation; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers.  rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670

value: wild type


title: Phosphate deprivation time 3h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours in a phosphate deprived  medium


title: bsub-wt_ctr_r1_s_o_v17

description: Culture: bsub-wt_ctr_r1_s_o_v17 was cultured in LB Broth at 37˚C and harvested in log growth phase; Library: HT-5PSeq using indexed rP5_RNA (rP5_MPX_RNA) Adapters (CrArCrGrArCrGrCrUrCrUrUrCrCrGrArUrCrU rXrXrXrXrXrX rNrNrNrNrNrNrNrN) carrying a sample barcode (rX) and unique molecular identifiers (rN) which were removed from the reads into the read headers.  rRNA removal:  in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: Iron deprivation time 3h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours on an iron deprived medium


title: Full nutrient time 3h Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium + 3hours on full nutrient medium


title: Control full nutrient time 0 Rep1

description: Gene expression data from 10 days Arabidopsis roots on full nutrient medium


title: rfg1 mutant-1

city: Beijing

first: Guobo

last: Guan

address: NO.1 Beichen West  Road, Chaoyang District, Beijing

country: China

institute: Institute of Microbiology, Chinese Academy of Sciences

data_processing:
Illumina OLB_1.9.4 were used for basecalling
The raw reads were filtered by removing the adapter and low quality reads. Clean reads were mapped to the genome of C. albicans SC5314 using SOAP aligner/soap2 software (version 2.21)
Relative gene expression levels were calculated using the FPKM (fragments per kb per million reads) method.
Assembly: Candida albicans SC5314 genome (ASM18296v1)
Supplementary files format and content: text files containing FPKM values for each Sample

organism: Candida albicans

growth_protocol: Cells of the WT and rfg1/rfg1 strains were cultured on YPD-K medium at 30˚C for two days and then plated onto YPD-K media, pH4 and cultured at 30˚C for two days.

extract_protocol:
Cells were lysed by Mini-beadbeater and total RNA was harvested using GeneJET RNA Purification Kit (Thermo Scientific #K0731, #K0732)
RNA libraries were prepared for sequencing using standard Illumina protocols

value: RFG1 knockout

value: YPD-K, pH 4, 30C


title: WT-2


title: rfg1 mutant-2


title: SCR AMO PRO-seq Replicate 3

description:
201029_mm10_mESC_PROseq_N3_SCR_AMO_F.bedGraph
201029_mm10_mESC_PROseq_N3_SCR_AMO_R.bedGraph

value: F121-9 wt mESCs

value: 20uM SCR AMO, 4 hours.


title: SCR AMO PRO-seq Replicate 2


title: WT-1


title: ChIP_P40_S15: FemalePoly8_Input

value: Female

value: Poly IC


title: bsub_mup_r2_s_o_v15

data_processing:
Adapter and quality trimming with the bbduk program of the BBTools suite, with options {qtrim=r, ktrim=r, hdist=3, hdist2=2, K=20, mink=14, trimq=16, minlen=30, maq=16}, using BBTools default adapter set and polyG or polyA sequences for short reads.
Deduplication of reads with both identical unique molecular identifier (UMI) and insert sequence using the dedupe program of the BBTools suite with its default parameters.
UMI sequences found in the first eight bases of each read were extracted using UMI-tools (version 1) with default options (using --bc-pattern NNNNNNNN).
Alignment was performed with the bbmap program of the BBTools suite, with the {32bit=t -da -eoom k=11 strictmaxindel=10 intronlen=0 t=16 trd=t minid=0.94 nzo=t} parameters. Alignment files were sorted and indexed with SAMtools.
Deduplication based on UMIs was then performed with UMI-tools (version 1) with options { --soft-clip-threshold 1 --edit-distance 2 --method unique}.
In selected samples, a dual undex strategy was used (sequencing runs v17). In brief, in addition of the Illumina indexes with dedicated reads used in HT-5Pseq, an additional index sequence is added before the UMI sequence. This is introduced during the ssRNA ligation space using anoligo with the following structure Illumina compatible-CCCCCC-NNNNNNNN. Please see dual_index_demultiplexing.txt for details on demultiplexing strategy for those samples.
Bedgraphs for read 5' positions for forward (fwd) and reverse (rev) strands were generated with BEDTools genomecov program with options {-bg -strand [+/-] -5}.
Assembly: The genome used for alignment is indicated in the processed file names with the G-xxxx patterns.  Those correspond to the NCBI accessions and builds indicated in the Supplementary file.
Assembly: Reference indiced for individual bacterial species and custom mixtures were built with the bbmap program of the BBTools suite, with default options (and k=10).
Supplementary files format and content: Processed files are bedgraphs containing read 5' counts on reverse (_rev) and forward (_fwd) strands of genomes indicated with _G-xxxx pattern. The genomes corresponding to each pattern are described in the genome build entry above.

extract_protocol:
Microbial cell pellets were resuspended in equal volume of LET (25 mM Tris pH 8.0, 100 mM LiCl, 20 mM EDTA) and water saturated Phenol pH 6.6 (Thermo Fisher),  lysed with acid washed glass beads (Sigma-Aldrich) and vortexing for three minutes in MultiMixer. Lysates were supplemented  with equal volumes of phenol/chloroform isoamyl alcohol pH 4.5 (25:24:1) and nuclease free water, l two-minute vortexing followed by centrifugation. Resulting aqueous phase was purified in two steps using phenol-chloroform isoamyl alcohol (25:24:1) followed by chloroform. After centrifugation, the clean aqueous phase was precipitated with sodium acetate-ethanol.
Standard and random fragmented 5PSeq libraries were generated as described in PMID: 26046441 with minor modifications using 150-9000ng total RNA as an input. To prepare random fragmented samples (negative controls), ribosomal RNA was depleted from DNA-free RNA and subsequent fragmented by incubating five minutes at 80 ̊C in fragmentation buffer (40mM Tris Acetate pH 8.1, 100mM KOAc and 30mM MgOAc). Reaction was purified using 2 volumes of RNACleanXP beads (Beckman Coulter) as recommended by the manufacturer. Free 5’OH sites were re-phosphorylated using 5 Units of T4 Polynucleotide kinase (PNK, NEB) and incubated at 37 ̊C for 60 minutes as recommended by the manufacturer. Re-phosphorylated fragmented RNA was purified using Phenol:Chloroform: Isoamyl Alcohol (24:25:1), followed by sodium acetate-ethanol precipitation. From this step forward, procedures for random fragmented and standard 5PSeq library preparation merge. RNA was Ligated to either rP5_RND or rP5_RNA oligo (specified in characteristics protocol).    containing unique molecular identifiers). Ribosomal RNA was depleted using Ribozero rRNA removal kit (Illumina) suitable for Bacteria and Yeast. rRNA depleted sample was purified using 1.8V of Ampure beads (Abcam), fragmented using heat (80˚C) for 5 min in 5x Fragmentation Buffer (200mM Tris Acetate pH 8.1,500mM KOAc,150mM MgOAc) and reverse transcribed using random hexamers to prime. Resulting cDNA was bound to streptavidin beads (M-280), subjected to enzymatic reactions of DNA end repair, fill-in of adenine to 5’ protruding ends of DNA fragments using Klenow Fragment (NEB). Common adaptor (P7-MPX) was ligated and 5PSeq Libraries were amplified by PCR (15- 17 cycles). Purified samples were quantified using Qubit (Thermo Fisher), library size was estimated from bioanalyzer traces. 5PSeq Libraries were pooled by mixing equal amounts of each sample, following enrichment of fragments of the length between 300-500nt.   HT-5PSeq Libraries were generated as recently described (bioRxiv 2020.06.22.165134; doi: https://doi.org/10.1101/2020.06.22.165134). In brief, DNA-free RNA was ligated with RNA oligos containing unique molecular identifiers. Ligated RNA was reverse transcribed priming with oligos containing a random hexamer and an Illumina compatible region. RNA was eliminated by addition of NaOH. Ribosomal RNA was depleted by adding in-house rRNA DNA oligo depletion mixes (table 2) to the cDNA and performing a duplex-specific nuclease (DSN, Evrogen) treatment. rRNA depleted cDNA was PCR amplified (15-17 cycles). Depletion of ribosomal RNA with Ribozero Illumina (for bacteria and yeast) was done after the single-stranded RNA ligation step. Ribosomal depleted RNA was purified and reverse transcribed using the same oligos as stated above, and then amplified by PCR. Libraries were quantified by fluorescence (Qubit, Thermo Fisher), size estimated using an Agilent Bioanalyzer and sequence using a NextSeq500 Illumina sequencer.
other: 5PSeq that captures  transcripts with free 5' phospahte (without cap)

value: 65ug/ml Mupirocin incubated for 10 minutes at 37˚C

value: bsub_mup_r2 was cultured in LB Broth at 37˚C and Mupirocin was added in mid-log growth phase  for 10 minutes

tag: ribosomal rna depletion strategy

value: in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: PAC-seq  WT mESC Replicate 1

value: Mouse embryonic stem cells

tag: genotype

value: WT


title: ChIP_P40_S19: MaleCtr6_H3K4me3

value: Control


title: ChIP_P40_S14: FemalePoly8_H3K9ac


title: ChIP_P40_S13: FemalePoly8_H3K4me3


title: afin_mup_r2_s_o_v15

organism: Alistipes finegoldii

source_name: Liquid culture of Alistipes finegoldii (DSM 17242)

value: afin_mup_r2  was anaerobically cultured in GAM Broth (HyServe) at 37˚C at 37˚C and split 24 hours prior treatments and Mupirocin was added to culture for 10 minutes.


title: bsub_mup_r1_s_o_v15

value: bsub_mup_r1  was cultured in LB Broth at 37˚C and Mupirocin was added in mid-log growth phase  for 10 minutes


title: zymo_ctr_r1_m_r_v2-v4

organism:
Escherichia coli
Staphylococcus aureus
Enterococcus faecalis
Bacillus subtilis
Limosilactobacillus fermentum
Listeria monocytogenes
Saccharomyces cerevisiae
Cryptococcus neoformans
Salmonella enterica

source_name: Cell sudspension containing mixture of ten inactivated microorganisms (ZymoBIOMICS Microbial Community Standard, cat-no D6300, lot-no ZRC 190633) .

value: Standard 5PSeq, HT-5PSeq

value: Ribo-Zero rRNA Removal Kit (Bacteria) Illumina.  In-house Bacterial rRNA Depletion MIX for Cultures , oVP690


title: bsub_ctr_r2_s_o_v15

value: Untreated

value: bsub_ctr_r2 was cultured in LB Broth at 37˚C and harvested in  mid-log growth phase


title: ChIP_P40_S12: FemalePoly7_Input

value: Whole brain


title: U1 AMO TT-seq Replicate 2

description:
U1_AMO_N3_TTseq_sorted_dedup_coverage_mitorvm_plus_normD2.10_sorted.bedGraph
U1_AMO_N3_TTseq_sorted_dedup_coverage_mitorvm_minus_normD2.10_sorted.bedGraph

data_processing:
Using a custom script (trim_and_filter_PE.pl), FASTQ read pairs were trimmed to 120 bp per mate and read pairs with a minimum average base quality score of 20 retained. Read pairs were further trimmed using cutadapt 1.14 to remove adapter sequences and low-quality 3’ bases (--match-read-wildcards -m 20 -q 10).
Reads were first mapped to the dm6 version of the Drosophila genome using STAR 2.7.31. Reads not mapping to the spike genome were then used for alignment to mouse (mm10) or human (hg38) using parameters --quantMode TranscriptomeSAM GeneCounts --outMultimapperOrder Random --outSAMattrIHstart 0 --outFilterType BySJout --outFilterMismatchNmax 4 --alignSJoverhangMin 8 --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonicalUnannotated --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outFilterScoreMinOverLread 0 --outFilterMatchNminOverLread 0. Duplicates were also removed using STAR.  Stranded coverage bedGraph files were generated from deduplicated BAM files using STAR.
BedGraphs were normalized using the normalize_bedGraph custom script. For libraries generated from untreated mESCs and HEK293Ts, no further normalization was performed. For libraries generated under SCR or U1 AMO conditions: We observed a consistent increase in reads mapping to the fly genome (dm6) after U1 inhibition (Average ratio of U1 AMO / SCR AMO = 2.1), indicating a global decrease in RNA synthesis in U1 AMO cells compared to the SCR AMO control. Accordingly, the average ratio of reads mapping to the spike genome between U1 and SCR AMO conditions was used to generate spike normalization factors
BedGraph files were converted to the bigWig format, and merged bedGraphs for each experimental condition were generated using bigWigMerge (UCSC tools).
Assembly: mm10 (mESC); hg38 (HEK293T)
Supplementary files format and content: bedGraph
Library strategy: TT-seq

extract_protocol:
Cells were collected in Trizol, RNA was extracted, and genomic DNA contamination removed using DNAse. RNA was then fragmented. 4sU-containing RNA was biotinylated, pulled down, and washed. RNA was then eluted and purified.
4sU-labeled RNA was used as input to the Illumina TruSeq stranded total RNA kit using the manufacturer's protocol.

treatment_protocol: See individual samples. Cells were labeled with 4sU for the final 20 minutes.

value: F121-9 wt mESCs

value: F121-9 mESCs with a fluorescent splicing reporter system as described in Vlaming et al., 2022

value: 20uM U1 AMO, 4 hours.


title: bsub_ctr_r1_s_o_v15

value: bsub_ctr_r1 was cultured in LB Broth at 37˚C and harvested in  mid-log growth phase


title: bsub_cam_r2_s_o_v15

value: 100ug/ml Chloramphenicol for 10 minutes at 37˚C

value: bsub_cam_r2 was cultured in LB Broth at 37˚C and Chloramphenicol was added in mid-log growth phase  for 10 minutes


title: ChIP_P40_S10: FemalePoly7_H3K4me3

value: Female

value: Poly IC


title: lreu_mup_r2_s_r_v1

organism: Limosilactobacillus reuteri

source_name: Liquid culture of Lactobacillus reuteri (DSM17938.

value: 65microg/ml Mupirocin incubated for 10 minutes at 37C

value: lreu_mup_rep2 was grown in MRS broth at 37C. Mupirocin was added at exponential growth OD600 0.56

value: Standard 5PSeq

value: Ribo-Zero rRNA Removal Kit (Bacteria) Illumina


title: lreu_mup_r3_s_r_v1

value: lreu_mup_rep3 was grown in MRS broth at 37C. Mupirocin was added at exponential growth OD600 0.37


title: SCR AMO TT-seq Replicate 3

description:
SCR_AMO_N3_TTseq_sorted_dedup_coverage_mitorvm_plus_sorted.bedGraph
SCR_AMO_N3_TTseq_sorted_dedup_coverage_mitorvm_minus_sorted.bedGraph

value: Mouse embryonic stem cells

value: 20uM SCR AMO, 4 hours.


title: ChIP_P40_S11: FemalePoly7_H3K9ac


title: lreu_frag_r2_s_r_v1

value: Untreated

value: lreu_frag_rep2 was grown in MRS broth at 37C and harvested at exponential growth OD600 0.56


title: bsub_cam_r1_s_o_v15

source_name: Liquid culture of Bacillus subtilis 168 trpC2.

value: bsub_cam_r1 was cultured in LB Broth at 37˚C and Chloramphenicol was added in mid-log growth phase for 10 minutes

value: in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: lreu_mup_r1_s_r_v1

value: lreu_mup_rep1 was grown in MRS broth at 37C. Mupirocin was added at exponential growth OD600 0.34


title: U1 AMO TT-seq Replicate 1

value: F121-9 wt mESCs

value: F121-9 mESCs with a fluorescent splicing reporter system as described in Vlaming et al., 2022

value: 20uM U1 AMO, 4 hours.


title: ChIP_P40_S8: FemalePoly3_H3K9ac

value: Whole brain

value: Female


title: SCR AMO PRO-seq Replicate 1


title: fecul-C_mup-t30_r1_fc_r_v13

organism: human feces metagenome

value: 650ug/ml Mupirocin for 30 minutes at 37˚C

value: fecul-C_mup-t30_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Mupirocin  (650µg/ml) for 30 minutes.

value: using PAN-Prokaryote Ribopool ( sitoolsbiotech REF dp-K024-000003)


title: lreu_frag_r1_s_r_v1

value: lleu_frg_rep1 was grown in MRS broth at 37C and harvested at exponential growth phase OD600 0.34

value: Standard 5PSeq

value: Ribo-Zero rRNA Removal Kit (Bacteria) Illumina


title: ChIP_P40_S9: FemalePoly3_Input

value: Poly IC


title: fecul-C_mup-t05_r1_fc_r_v13

value: 650ug/ml Mupirocin for 5 minutes at 37˚C

value: fecul-C_mup-t05_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Mupirocin  (650µg/ml) for 5 minutes.


title: ChIP_P40_S7: FemalePoly3_H3K4me3


title: SCR AMO TT-seq Replicate 2

value: Mouse embryonic stem cells

value: 20uM SCR AMO, 4 hours.


title: SCR AMO TT-seq Replicate 1


title: lreu_ctr_r2_s_r_v1

value: Untreated

value: lleu_ctr_rep2 was grown in MRS broth at 37C and harvested at exponential growth OD600 0.56


title: ChIP_P40_S4: FemaleCtr7_Input

value: Control


title: RNAPII ChIP-seq mESC Replicate 2

description:
F121-9 wt mESCs
mm10_mESC_RNAPII_ChIPseq_CAM_N3_dedup_25ntbin.bedGraph

data_processing:
Using a custom script (trim_and_filter_PE.pl), FASTQ read pairs were trimmed to 50bp per mate, and read pairs with a minimum average base quality score of 20 were retained.
Reads were then mapped to the mouse genome (mm10) using bowtie version 1.2.2 (-v2 -k1 --allow-contain -X1000 -p 5 –best).
The custom script extract_fragments.pl was used to retain reads fragments corresponding to insert sizes of 50-500bp, remove duplicate reads and generate bedGraphs (25nt bins) reporting the read fragment center.
The custom script bedgraphs2stdBedGraph was used to merge replicate bedGraphs (n=3).
Assembly: mm10
Supplementary files format and content: bedGraph

extract_protocol:
Cells were crosslinked with formaldehyde for 2.5 minutes and quenched with glycine. After washing, cell pellets were resuspended in sonication buffer and sonicated using the Qsonica Q800R3 system. Chromatin was pre-cleared with Protein A agarose and incubated with the indicated antibody overnight. Protein A agarose beads were added and allowed to bind antibody-epitope complexes. IPs were washed extensively and eluted twice. DNA was purified using phenol/chloroform.
ChIP-seq libraries were generated using the NEBNext Ultra II DNA Kit (New England Biolabs), following manufacturer’s instructions.

value: WT

tag: antibody

value: RNAPII


title: U1 AMO TT-seq Replicate 3

value: F121-9 wt mESCs

value: F121-9 mESCs with a fluorescent splicing reporter system as described in Vlaming et al., 2022

tag: treatment

value: 20uM U1 AMO, 4 hours.


title: RNAPII ChIP-seq mESC Replicate 3


title: fecul-C_ery-t30_r1_fc_r_v13

value: 5ug/ml Erythromycin for 30 minutes at 37˚C

tag: culture conditions

value: fecul-C_ery-t30_r1  culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Erythromycin  (5µg/ml) for 30 minutes.

value: using PAN-Prokaryote Ribopool ( sitoolsbiotech REF dp-K024-000003)


title: fecul-C_ery-t05_r1_fc_r_v13

value: 5ug/ml Erythromycin for 5 minutes at 37˚C

value: fecul-C_ery-t05_r1  culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Erythromycin  (5µg/ml) for 5 minutes.

tag: library protocol


title: lreu_ctr_r1_s_r_v1

value: lleu_ctr_rep1 was grown in MRS broth at 37C and harvested at exponential growth OD600 0.34

value: Standard 5PSeq

tag: ribosomal rna delpetion strategy

value: Ribo-Zero rRNA Removal Kit (Bacteria) Illumina


title: ChIP_P40_S3: FemaleCtr7_H3K9ac

value: Whole brain

value: Female


title: lreu_cam_r3_s_r_v1

value: 100microg/ml Chloramphenicol incubated for 5 minutes at 37C

value: lleu_cam_rep3 was grown in MRS broth at 37C. Chloramphenicol treatment initiated at exponential growth OD600 0.37.


title: RNAPII ChIP-seq mESC Replicate 1

tag: cell type

value: Mouse embryonic stem cells

value: WT


title: lreu_cam_r2_s_r_v1

value: lleu_cam_rep2 was grown in MRS broth at 37C. Chloramphenicol treatment was initiated at exponential growth phase OD600 0.56.


title: RNA-seq siRRP40 mESC Replicate 4

description:
211012_mm10_mESC_RNAseq_DepthNorm_sorted_RRP40KD_N4_F.bedGraph
211012_mm10_mESC_RNAseq_DepthNorm_sorted_RRP40KD_N4_R.bedGraph

data_processing:
Using a custom script (trim_and_filter_PE.pl), FASTQ read pairs were trimmed to 120bp per mate, and read pairs with a minimum average base quality score of 20 retained. Read pairs were further trimmed using cutadapt 1.14 to remove adapter sequences and low-quality 3’ bases (--match-read-wildcards -m 20 -q 10).
Reads were first mapped to the dm6 version of the Drosophila genome using STAR 2.7.31. Reads not mapping to the spike genome were then used for alignment to mm10/hg38 using parameters --quantMode TranscriptomeSAM GeneCounts --outMultimapperOrder Random --outSAMattrIHstart 0 --outFilterType BySJout --outFilterMismatchNmax 4 --alignSJoverhangMin 8 --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonicalUnannotated --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --outFilterScoreMinOverLread 0 --outFilterMatchNminOverLread 0. Duplicates were also removed using STAR.
Stranded coverage bedGraph files were generated from deduplicated BAM files using STAR.  For siRNA control and RRP40 KD conditions, bedGraphs were depth normalized using normalize_bedGraph
BedGraph files were converted to the bigWig format, and merged bedGraphs for each experimental condition were generated using bigWigMerge (UCSC tools).
Assembly: mm10 (mESC); hg38 (HEK293T)
Supplementary files format and content: bedGraph

extract_protocol:
RNA was extracted in Trizol and purified.
Libraries were generated with the llumina TruSeq Stranded Total RNA kit using standard protocols.

value: F121-9 wt mESCs

value: siRNA pool against RRP40


title: ChIP_P40_S2: FemaleCtr7_H3K4me3

tag: strain

value: Control


title: ChIP_P40_S1: FemaleCtr2_Input


title: lreu_cam_r1_s_r_v1

value: lleu_cam_rep1 was grown in MRS broth at 37C. Chloramphenicol treatment was initiated at exponential growth  OD600 0.34.


title: ChIP_P40_S35: FemaleCtr2_H3K9ac

value: Male

value: Poly IC


title: ChIP_P40_S34: FemaleCtr2_H3K4me3


title: S.solfataricus.P2.ChIP-seq.CreN7.replicate2

city: London

first: Finn

last: Werner

address: Gower Street

department: Institute of Structural and Molecular Biology

country: United Kingdom

institute: University College London

description:
18452X20
NovaSeq SP Reagent Kit v1.5_50x50 bp Sequencing

data_processing:
Paired-end reads were mapped using Bowtie v1.1.2 [30] with parameters -v 2 -m 1 --fr over the first 50 nt allowing only for uniquely mapped  read pairs to be included
We sampled from the alignments to obtain a defined fragment size distribution (normal distribution, mean 120 bp, sd 18 bp) using a custom R script (https://github.com/fblombach/ChIP-seq/blob/master/read_sampling.R)
Coverage normalized to input ("normRatio") was calculated using deeptools bamCompare with SES scaling (10,000 bins, 200 bp bin width), chromatin input samples for these IPs were previously deposited at NCBI GEO (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE141286)
Assembly: NC_002754.1
Supplementary files format and content: bigwig files with occupancy normalised against the chromatin input samples, scaled by the Signal Extraction Scaling method

organism: Saccharolobus solfataricus P2

source_name: microbial cells

growth_protocol: S. solfataricus P2 was grown in Brock medium supplemented with 0.2% glucose, 0.1% tryptone, and 20 µg/ml uracil at 75 °C in Erlenmeyer shake flasks at 150 rpm

extract_protocol:
Qiagen QIAquick PCR purification
NEBNext Ultra II DNA library prep kit for Illumina

tag: chip antibody

value: microbial cells


title: RNA-seq siRRP40 mESC Replicate 2


title: S.solfataricus.P2.ChIP-exo.CreN7.replicate2

description:
NovaSeq SP Reagent Kit v1.5_50x50 bp Sequencing
19862X4

data_processing:
Paired-end reads were mapped using Bowtie v1.1.2 with parameters -v 2 -m 1 --fr allowing only for uniquely mapped  read pairs to be included
Bam files with alignments were filtered for first read in proper pairs (samtools view -f 66 -b)
Read 5'-end coverage was calculated using bedtools genomecov with bedgraph output
Bedgraph files were converted to bigwig format using ucsc-bedgaphToBigwig
Assembly: NC_002754.1
Supplementary files format and content: Bigwig files with raw 5'-end coverage


title: ChIP_P40_S6: FemaleCtr10_Input


title: S.solfataricus.P2.ChIP-seq.CreN7.replicate1

description:
18452X19
NovaSeq SP Reagent Kit v1.5_50x50 bp Sequencing


title: fecul-C_cam-t30_r1_fc_r_v13

value: 80ug/ml Chloramphenicol for 30 minutes at 37˚C

value: fecul-C_cam-t30_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Chloramphenicol (80µg/ml) for 30 minutes.

value: using PAN-Prokaryote Ribopool ( sitoolsbiotech REF dp-K024-000003)


title: fecul-C_cam-t05_r1_fc_r_v13

value: 80ug/ml Chloramphenicol for 5 minutes at 37˚C

value: fecul-C_cam-t05_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Chloramphenicol (80µg/ml) for 5 minutes.


title: RNA-seq siNT mESC Replicate 4

description:
211012_mm10_mESC_RNAseq_DepthNorm_sorted_NT_N4_F.bedGraph
211012_mm10_mESC_RNAseq_DepthNorm_sorted_NT_N4_R.bedGraph

value: Non-targeting siRNA control


title: ChIP_P40_S5: FemaleCtr10_H3K9ac

value: Whole brain

value: Female


title: RNA-seq siRRP40 mESC Replicate 1

value: Mouse embryonic stem cells

value: WT


title: fecul-B_mup-t30_r1_fc_r_v13

value: 650ug/ml Mupirocin for 30 minutes at 37˚C

value: fecul-B_mup-t30_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Mupirocin  (650µg/ml) for 30 minutes.


title: fecul-C_ctr_r1_fc_r_v13

value: Untreated

value: fecul-C_ctr_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior harvesting.


title: RNA-seq siNT mESC Replicate 3

value: F121-9 wt mESCs


title: fecul-B_mup-t05_r1_fc_r_v13

value: 650ug/ml Mupirocin for 5 minutes at 37˚C

value: fecul-B_mup-t05_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Mupirocin  (650µg/ml) for 5 minutes.


title: fecul-C_dox-t30_r1_fc_r_v13

value: 5ug/ml Doxycyclin for 30 minutes at 37˚C

value: fecul-C_dox-t30_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Doxycyclin  (5µg/ml) for 30 minutes.


title: S.solfataricus.P2.ChIP-exo.Cbp1.replicate2

description:
NovaSeq SP Reagent Kit v1.5_50x50 bp Sequencing
19862X2

value: microbial cells


title: fecul-C_dox-t05_r1_fc_r_v13

value: 5ug/ml Doxycyclin for 5 minutes at 37˚C

value: fecul-C_dox-t05_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Doxycyclin  (5µg/ml) for 5 minutes.

value: using PAN-Prokaryote Ribopool ( sitoolsbiotech REF dp-K024-000003)


title: CD103 int RNA-seq

city: Birmingham

first: Aaron

last: Silva Sanchez

state: AL

address: 1825 University Boulevard

department: Medicine

institute: The University of Alabama at Birmingham

description: Sorted cells

data_processing:
All RNA-seq analyses were performed using the Penn State Galaxy publicly available server using the tuxedo suite analysis tools. Fastq files were aligned to the mm10 reference genome using TopHat (Galaxy Tool Version 0.9; Tool Version TopHat v2.0.14 for all studies except for the glutamine studies, Galaxy Tool Version 2.1.0 and Tool Version TopHat v2.1.0) were used with default parameters.
Cufflinks, Cuffmerge, Cuffquant, and Cuffnorm were performed to define FPKM values for genes. Cufflinks (Galaxy Tool Version 2.2.2.0; Tool Version Cufflinks v2.2.1.0) was performed using default parameters for all settings except bias and multi-read correction was performed.
Cuffmerge (Galaxy Tool Version 2.2.1.0) was performed using default parameters. Cuffquant (Galaxy Tool Version 2.2.1.0; Tool Version Cuffquant v2.2.1) was performed using default parameters for all settings except bias and multi-read correction was performed.
Cuffdiff (Galaxy Tool Version 2.2.1.3; Tool Version Cuffdiff v2.2.1) was performed using default parameters (using Cuffquant output files). Cuffnorm (Galaxy Tool Version 2.2.1.1; Tool Version Cuffnorm v2.2.1) was performed using default parameters with a Cuffdiff format output.
Genome_build: mm10
Supplementary_files_format_and_content: Microsoft Excel Open XML Format Spreadsheet files showing FPKM values for each individual sample

source_name: Lin- MHCII+ CD11c+ CD103Int CCR7+  DCs

extract_protocol:
Collected lungs from 2-3 litters of naïve WT C57BL/6 mice, diggested with collagenase/dnase for 30 minutes, then enriched with CD11c magnetic beads, and stained for sorting for 15 minutes at 4 degrees.
RNA libraries were prepared for sequencing using standard Illumina protocols

value: Post-natal day 7

value: Lungs


title: S.solfataricus.P2.ChIP-exo.Cbp1.replicate1

description:
NovaSeq SP Reagent Kit v1.5_50x50 bp Sequencing
19862X1


title: CD103 hi RNA-seq

description: Sorted cells

source_name: Lin- MHCII+ CD11c+ CD103hi  DCs


title: RNA-seq siNT mESC Replicate 1

value: Mouse embryonic stem cells

value: Non-targeting siRNA control


title: RNA-seq siRRP40 mESC Replicate 3

value: WT

value: siRNA pool against RRP40


title: CD11b RNA-seq

description: Sorted cells

source_name: Lin- MHCII+ CD11c+ CD11b+ DCs


title: afin_mup_r1_s_o_v15

value: 65ug/ml Mupirocin incubated for 10 minutes at 37˚C

value: afin_mup_r1 was anaerobically cultured in GAM Broth (HyServe) at 37˚C and split 24 hours prior treatments and Mupirocinwas added to culture for 10 minutes.

value: in-house Bacterial rRNA Depletion MIX for Cultures , oVP670


title: fecul-B_ery-t05_r1_fc_r_v13

value: 5ug/ml Erythromycin for 5min

value: fecul-B_ery-t05_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Erythromycin  (5µg/ml) for 5 minutes.


title: afin_ctr_r2_s_o_v15

value: Untreated

value: afin_ctr_r2 was anaerobically cultured in GAM Broth (HyServe) at 37˚C and split 24 hours prior harvesting.


title: afin_ctr_r1_s_o_v15

value: afin_ctr_r1 was anaerobically cultured in GAM Broth (HyServe) at 37˚C and split 24 hours prior harvesting.


title: fecul-B_dox-t30_r1_fc_r_v13

value: 5ug/ml Doxycyclin for 30 minutes at 37˚C

value: fecul-B_dox-t30_r1 culture was grown anaerobically at 37˚C and diluted with fresh MCDA media at a 1:10 ratio  24 hours prior treatment. Doxycyclin  (5µg/ml) for 30 minutes.


title: 03_SKM1-ABBV-075_BRD4

city: North Chicago

first: Josh

last: Plotnik

state: IL

address: 1 North Waukegan Rd, AP10/210A

institute: AbbVie Inc.

data_processing:
Illumina Casava1.8.2 software used for basecalling
Reads were aligned to the human genome (hg19) using the BWA algorithm
Duplicate reads were removed and only uniquely mapped reads (mapping quality >= 25) were used for further analysis.
Alignments were extended in silico at their 3’-ends to a length of 200 bp, which is the average genomic fragment length in the size-selected library, and assigned to 32-nt bins along the genome.
Genome_build: hg19
Supplementary_files_format_and_content: BigWig files were generated from BedGraph to BigWig. Scores represent peak density.

growth_protocol: SKM-1 cells were grown in 10%FBS RPMI 1640

extract_protocol:
Lysates were clarified from sonicated, fixed cell pellets and protein-DNA complexes were isolated with antibody.
Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition, and adaptor ligation.

treatment_protocol: Cells were treated with indicated inhibitors 6 hrs or DMSO as vehicle control

value: SKM-1 adult acute myeloid leukemia cell line

tag: agent

value: 100 nM ABBV-075 6 hrs


title: 04_SKM1-ABBV-744_BRD4

value: 200 nM ABBV-744 6 hrs