title: IR-10dai Replicate 2 [P]

city: Ghent

first: Bruno

last: Verstraeten

state: Oost-Vlaanderen

address: Technologiepark 71

institute: Ghent University

description: Infected roots

data_processing:
Bases were called using the Real Time Analysis 2 software of the NextSeq500.
The quality of raw reads from each sample was controlled by FastQC (Andrews, 2010), then using Trimmomatic (Bolger, Lohse, & Usadel, 2014), the reads were checked with a 5-base sliding window and bases with lower phred score than 20 and reads shorter than 40 nt were removed.
The trimmed reads from each sample were aligned to the Beta vulgaris subsp. vulgaris reference genome (Refbeet1.1, (Dohm et al., 2014)) using STAR software (Dobin et al., 2013).
The number of trimmed reads mapped to each annotated gene per condition was counted using the summarizeOverlaps functions of the GenomicAlignments R package .
The reads were normalized and differentially expressed genes (DEGs) were obtained using the R-package ‘DESeq2’ (Love, Huber, & Anders, 2014). In this software the counts were normalized using the median of ratios method. Adjusted P-values for each estimate of False Discovery Rates (FDRs) were added using the Benjamin-Hochberg algorithm. Transcripts were considered to be significant DEGs when the adjusted P-value was < 0.05.
Genome_build: RefBeet1.1
Supplementary_files_format_and_content: A count table containing counts for all genes for all samples

organism: Beta vulgaris

source_name: Resistant_25 days_Infected roots

growth_protocol: Seeds of a susceptible sugar beet line (7112*SB36, SBSI of Iran) and a resistant cultivar containing the Hs1pro-1 gene (Nemakil, Syngenta) were used in this study. Sugar beet seeds germinated on sterile soil for 3 days at 28 °C. The seedlings were transferred to SAP substrate (Sand Absorbent Polymer; (Reversat, Boyer, Sannier, & Pando-Bahuon, 1999)) and were kept in a growth chamber at 25 ± 2 °C, with 16 h light/8 h darkness.

extract_protocol:
Whole root tissue of infected and control plants was collected at 4 and 10 dai, were then washed in water, frozen in liquid nitrogen and stored at -80 °C until RNA extraction. For each time point two independent biological replicates were collected and each replicate consisted of a pool of six individual plants. Total RNA was extracted from whole roots of infected and uninfected plants at 4 and 10 dai with the RNeasy Plant Mini Kit (Qiagen) according to the manufacturer’s protocol, with an additional sonication step for 30 seconds after addition of buffer RLT, and were then stored at -80 °C. The quantity and quality of each RNA sample was controlled by a NanoDrop 2000c (Thermo Fisher Scientific, Belgium).
For each sample, 2 µg of total RNA was used for library preparation using QuantSeq 3’ mRNA-Seq Library Prep Kit (Lexogen) as following: An oligodT primer containing an Illumina-compatible-linker sequence at its 5´ end was hybridized to the RNA and reverse transcription was performed. After first strand synthesis the RNA was removed and second strand synthesis was initiated by random priming and a DNA polymerase. The random primer also contained an Illumina-compatible-linker sequence at its 5´ end. The second strand synthesis was followed by a magnetic bead-based purification step. Amplification of libraries was performed by addition of external barcodes during the PCR amplification step. In the final step the double-stranded libraries were purified by magnetic beads to remove all reaction components. The libraries were checked using an Agilent Bioanalyzer 2100 (Agilent). After cluster generation, 16 libraries were sequenced on a NextSeq 500 Illumina sequencing platform to produce single-end 76 bp reads. The samples were multiplexed to minimize lane effects. Library construction and sequencing were carried out by NXTGNT, the UGent sequencing reference center (Faculty of Pharmaceutical Sciences, Ghent University, Belgium).

treatment_protocol: A pure population of H. schachtii that originated from one cyst was multiplied on a susceptible sugar beet cultivar (Jolgeh, SBSI of Iran) in sterile potting soil under the same conditions. The J2s were harvested from cysts, soaked in 3 mM ZnCl2 to stimulate hatching. Fifteen-day-old roots of the sugar beet plants were inoculated with 300 fresh J2s of H. schachtii. Control plants were mock-inoculated with water.

tag: plant age

value: 25 days

tag: cultivar phenotype

value: Resistant

value: infected with Heterodera schachtii

tag: replicate number

value: roots


title: UnR-10dai Replicate 2 [N]

status: Public on Aug 08 2020

type: SRA

country: Belgium

description: Uninfected roots

library_source: transcriptomic

molecule: total RNA

source_name: Resistant_25 days_Uninfected roots

tag: treatment

value: none (uninfected)


title: F105H NET-seq Rep 2

city: Birmingham

first: David

last: Schneider

state: AL

address: Kaul Human Genetics Bldg, Rm 442, 720 20th St South

country: USA

institute: University of Alabama at Birmingham

data_processing:
Library strategy: NET-seq
Basecalls were converted to fastq files using Illumina’s bcl2fastq software (ver. 2.18.0.012)
The first two nucleotides were clipped from the 5’ end of each read and the 3’ adapter sequence CTGTAGGCACCAT was removed from each read via trim galore (ver. 0.4.1)
The sequences were aligned to the Saccharomyces cerevisiae genome assembly 3 via BWA MEM (ver 0.6) with a minimum alignment score of 20. All other parameters are default
Output files were converted to .bed files using samtools (ver 1.3.1)
Reads mapping to the rDNA gene were extracted, and depth at each position was counted
Genome_build: SacCer 3
Supplementary_files_format_and_content: Bed files include raw read depth coverage at each position along the rDNA.

organism: Saccharomyces cerevisiae

growth_protocol: Three liters per sample were grown to A600 = 0.3 in YEPD with nutation and collected via filtration

extract_protocol:
Cells were lysed via 10 1 minute grinds at maximum amplitude in a mikro dismembrator II. The grinding cup was chilled in liquid nitrogen for 1 minute between each grind.
Pol I elongation complexes were isolated from the cell lysate via immunoprecipitation with anti-HA magnetic beads (tagged gene = RPA135 C-terminal (3x-HA 7x- his) . Associated RNA transcripts were then isolated using an acidic phenol-chloroform extraction. The RNA transcripts were then 3’ appended with a DNA linker and reverse transcribed to produce complementary DNAs (cDNAs). The cDNAs were circularized and amplified by PCR to produce high-throughput sequencing libraries.

tag: genotype

value: rpa190-f1205h RNA polymerase I


title: IR-10dai Replicate 1 [O]

description: Infected roots


title: WT NET-seq Rep 3

source_name: WT

value: WT


title: F105H NET-seq Rep 1


title: WT NET-seq Rep 2


title: IS-10dai Replicate 2 [L]

description: Infected roots

source_name: Susceptible_25 days_Infected roots

value: Susceptible


title: XRN2 KD rep 1 RNA-seq

city: Montreal

first: Stéphane

last: Richard

state: Quebec

address: 3755 Côte-Ste-Catherine Road

department: Departments of Oncology and Medicine

country: Canada

institute: Lady Davis Institute - JGH, McGill University

description: Supplemental Data 2

data_processing:
Sequenced reads were first trimmed with fastq-mcf to remove the adapter sequence and then mapped to the human genome (hg19) through Bowtie2 for DRIP-seq and through STAR v2.7.1a for RNA-seq.
DRIP-seq: Peaks were called for each replicate through MACS in broad mode and then merged into a single peak list through the DiffBind R package.
RNA-seq: Gene expression was quantified for each replicate through Homer v4.11.1.
Differential R-loop sites (DRIP-seq) and differentially expressed genes (RNA-seq) were identified by DESeq2 v1.26.0.
Bedtools v2.28.0 was used to annotate the nearest and neighboring genes of the DRIP-seq peaks and to extract the intergenic RNA-seq reads for antisense transcription detection.
Genome_build: hg19
Supplementary_files_format_and_content: xlsx: DRIP-seq peak read counts and differential binding analysis, RNA-seq gene expression read counts and differential expression analysis.

organism: Homo sapiens

source_name: Osteosarcoma

extract_protocol: Total cellular RNA was isolated from siRNA-transfected U2OS cells using GenElute Mammalian total RNA Miniprep Kit (Sigma-Aldrich) according to the manufacturer’s instructions.

tag: cell line

value: XRN2 KD


title: PRMT5 KD rep 3 RNA-seq

description: Supplemental Data 2

value: PRMT5 KD


title: UnS- 10dai Replicate 2 [J]

description: Uninfected roots

source_name: Susceptible_25 days_Uninfected roots


title: IS-10dai Replicate 1 [K]

description: Infected roots


title: XRN2 KD rep 3 RNA-seq

description: Supplemental Data 2


title: XRN2 KD rep 2 RNA-seq

description: Supplemental Data 2


title: UnS- 10dai Replicate 1 [I]

description: Uninfected roots


title: IR-4dai Replicate 2 [H]

description: Infected roots

source_name: Resistant_19 days_Infected roots

value: 19 days


title: IR-4dai Replicate 1 [G]

description: Infected roots


title: Mut_295

city: San Francisco

first: Samuel

last: Pleasure

state: CA

address: 675 Nelson Rising Lane

institute: UCSF

data_processing:
Samples were sequenced, demultiplexed, aligned, and analyzed by the UCSF Functional Genomics Core Facility
sequenced reads were mapped to Ensembl Mouse GRCm38.78 using STAR_2.4.2a
Genome_build: Ensembl Mouse GRCm38.78
Supplementary_files_format_and_content: tab delimited text file containing samples and their raw (unnormalized ) read counts per gene.
Supplementary_files_format_and_content: tab delimited text file containing gene level differential expression tests

organism: Mus musculus

source_name: dorsal forebrain

extract_protocol:
Total RNA was extracted using Qiagen RNEasy Mini Kit (Qiagen) and prepared for RNAseq.
Illumina Truseq Stranded mRNA Library Prep Kit

tag: strain

value: mixed background with C57BL/6J

tag: age

value: Emx1-cre/+;Sufu-fl/fl (Sufu-cKO)

tag: Sex

value: male


title: PRMT5 KD rep 2 RNA-seq

description: Supplemental Data 2


title: UnR-4dai Replicate 2 [F]

description: Uninfected roots

source_name: Resistant_19 days_Uninfected roots


title: PRMT5 KD rep 1 RNA-seq

description: Supplemental Data 2


title: DDX5 KD rep 2 RNA-seq

description: Supplemental Data 2

value: DDX5 KD


title: DDX5 KD rep 3 RNA-seq

description: Supplemental Data 2


title: IS-4dai Replicate 1 [C]

description: Infected roots

source_name: Susceptible_19 days_Infected roots


title: UnR-4dai Replicate 1 [E]

description: Uninfected roots


title: DDX5 KD rep 1 RNA-seq

description: Supplemental Data 2


title: UnS- 4dai Replicate 2 [B]

description: Uninfected roots

source_name: Susceptible_19 days_Uninfected roots


title: UnR-10dai Replicate 1 [M]

description: Uninfected roots


title: Control rep 3 RNA-seq

description: Supplemental Data 2

value: Control


title: WT NET-seq Rep 1


title: Control rep 2 RNA-seq

description: Supplemental Data 2


title: Control rep 1 RNA-seq

description: Supplemental Data 2


title: MA008

city: Seattle

first: Nicole

middle: A P

last: Lieberman

state: WA

address: 850 Republican St S130

department: Laboratory Medicine

institute: University of Washington

data_processing:
Paired end/2x100 bp reads, illumina novaseq 6000
Illumina Casava1.7 software used for basecalling.
Reads were pseudoaligned by Kallisto v0.44 to human reference transcriptome GRCh38.
Genome_build: GRCh38
Supplementary_files_format_and_content: raw gene counts for each sample generated by kallisto

molecule: polyA RNA

source_name: brain organoids generated from iPSC

growth_protocol: Cerebral organoids were generated as previously described (Lancaster et. al. 2013) from two iPSC lines (FA0000010 and FA0000011, for brevity called FA10 and FA11). Briefly, cells were dissociated into single cell suspensions with Accutase (Stem Cell Technologies, cat#7920) and seeded at 4,500 cells/ well in 96-well low attachment U bottom plates, using EB media (DMEM/F12 (Thermo Fisher Scientific, cat. #11330-032) supplemented with 20% Knockout Serum Replacement (Thermo Fisher Scientific, cat. #10828-028), 3% ES quality batch-tested fetal bovine serum (Thermo Fisher Scientific, cat#10439024), 1% Glutamax (Thermo Fisher Scientific, cat. #35050-038), 1x Non-Essential Amino Acids, 0.1mM 2-mercaptoethanol, 4ng/ml bFGF (R&D Systems, cat.#233FB01M), and 50uM Y-27632. Fresh medium was replaced every other day until day 6,when  EB medium was replaced with Neural Induction (NI) medium (DMEM/F12, 1x N2 Supplement, 1X Glutamax, 1X NEAA and 1μg/ml Heparin (Sigma-Aldrich, cat#H3149) ) and the organoids were  transferred to 60 mm or 100 mm low-attachment plates. The organoids were allowed to form neuroepithelium tissue till day 11-14, with media changed every other day. Between day 11-24, the organoids were coated with Matrigel droplets and allowed to gel by keeping them at 37°C for 30 min. Matrigel-coated organoids were transferred to differentiation media (1:1 DMEM/F12:Neurobasal, 0.5% N2 Supplement, 2% B27 Supplement without Vitamin A (Life Technologies, cat#12587010), 0.25% Insulin solution (Life Technologies, cat#12585014), 50uM 2-mercaptoethanol, 1% Glutamax, 0.5% NEAA, 1% Penicillin-Streptomycin), for 4 days, then transferred to differentiation media containing B27 Supplement with Vitamin A (Thermo Fisher Scientific, cat#17504-044). Brain organoids were cultured for an additional 60 days with medium changed every 7 days, and used for experiments at 90 days or 270 days.

extract_protocol:
Direct-zoltm RNA MicroPrep (Zymo)
TruSeq stranded/poly-A enrichment (Illumina)

treatment_protocol: 90-day old organoids were infected with viruses, and RNA extracted after 10 days

tag: agent

value: WT.TdTomato measles virus

tag: experiment site

value: A

tag: organoid


title: miR-520b-1#

city: Beijing

first: Yang

last: Bin

address: 167 Beilishi Road

country: China

institute: Chinese Academy of  Medical Sciences and Peking Union Medical College

data_processing:
Basecalls performed using CASAVA version 1.8
aligned to the GRCh38 genome assembly using Hisat2（version:2.0.4）
Uniquely mapped reads with mapping quality >=20 were retrieved for downstream analysis.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: hg38
Supplementary_files_format_and_content: Excel files include RPKM values for each Sample ...

source_name: endothelial cells

extract_protocol:
Total RNA was harvested using Qiagen Kit.1 ug of total RNA was used for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: cell type

value: primary human umbilical vein endothelial cells


title: NC-3#


title: miR-520b-2#


title: FB009

value: WT.eGFP measles virus

value: B


title: miR-520b-3#


title: NC-2#


title: FB001


title: MA007


title: CON RNA-seq

city: Meizhou

last: Wang

address: Meisong Road

institute: Jiaying University

data_processing:
Basecalls performed using CASAVA version 1.8
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence using FASTP
mRNA-seq reads were aligned to the GRCh38 genome assembly using Hisat2 version 2.1.0 with default parameters
Gene abundance measurements were generationed and normalized  using stringtie version 1.3.4b Linux_x86_64 ; The normalized standard were Fragments Per Kilobase of exon per Megabase of library size (FPKM)
Differential expression were analysed  using edgeR function package in R program version 3.4.3
Genome_build: hg38
Supplementary_files_format_and_content: tab-delimited text files include normalized FPKM values and raw fragment counts for each Sample

source_name: Tu212 cells

growth_protocol: All cells were incubated at 37°C in 5% CO2

extract_protocol:
Total RNA were isolated by TRIzol (No.:15596026, Thermo Fisher Scientific, USA) and extracted by chloroform. RNA degradation and contamination were monitored on 1% agarose gels and purity was checked by the NanoPhotometer spectrophotometer (IMPLEN, CA USA). RNA concentration was measured using Qubit RNA Assay Kit in Qubit 2.0 Flurometer(Life Technologies, CA, USA) and integrity was assessed using the RNA Nano 6000 Assay Kit of the Agilent Bioanalyzeer 2100 system (Agilent Technologies, CA, USA ).
Sequencing libraries were generated using NEBNest UltraTM RNA Library Prep Kit for IlluminaR (NEB, USA) following manufacturer's recommendations and index codes were added to attribute sequences to each sample.

treatment_protocol: All cells were treated with pEZ-vector or pEZ-PRMT5 for 48 h

value: Epithelial-derived  cells

value: control


title: NC-1#


title: IS-4dai Replicate 2 [D]

description: Infected roots

value: infected with Heterodera schachtii

value: roots


title: PRMT5 RNA-seq

value: PRMT5 overexpression


title: FB004

value: T461I mutant measles virus


title: XRN2 KD rep 1 DRIP-seq

description: Supplemental Data 1

library_source: genomic

data_processing:
Library strategy: DRIP-Seq
Sequenced reads were first trimmed with fastq-mcf to remove the adapter sequence and then mapped to the human genome (hg19) through Bowtie2 for DRIP-seq and through STAR v2.7.1a for RNA-seq.
DRIP-seq: Peaks were called for each replicate through MACS in broad mode and then merged into a single peak list through the DiffBind R package.
RNA-seq: Gene expression was quantified for each replicate through Homer v4.11.1.
Differential R-loop sites (DRIP-seq) and differentially expressed genes (RNA-seq) were identified by DESeq2 v1.26.0.
Bedtools v2.28.0 was used to annotate the nearest and neighboring genes of the DRIP-seq peaks and to extract the intergenic RNA-seq reads for antisense transcription detection.
Genome_build: hg19
Supplementary_files_format_and_content: xlsx: DRIP-seq peak read counts and differential binding analysis, RNA-seq gene expression read counts and differential expression analysis.

molecule: genomic DNA

extract_protocol: Nucleic acids were extracted from U2OS cells by SDS/proteinase K treatment at 37°C overnight followed by phenol–chloroform extraction using MaXtractTM High Density (100 × 15 ml from QIAGEN) and ethanol precipitation at room temperature.

tag: chip antibody

value: Monoclonal S9.6

value: XRN2 KD


title: FB012


title: RAP+KU treatment group

city: Chongqing

first: Kexuan

last: Deng

address: 174 Shazheng ST, Shapingba

institute: Chongqing University

description:
R_K
Potato
miRNA

data_processing:
Reference genome and gene model annotation files were downloaded from genome website directly. Index of the reference genome was built using Bowtie v2.2.3 and paired-end clean reads were aligned to the reference genome using TopHat v2.0.12. We selected TopHat as the mapping tool for that TopHat can generate a database of splice junctions based on the gene model annotation file and thus a better mapping result than other non-splice mapping tools.
HTSeq v0.6.1 was used to count the reads numbers mapped to each gene. Then FPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene. FPKM, expected number of Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced, considers the effect of sequencing depth and gene length for the reads count at the same time, and is currently the most commonly used method for estimating gene expression levels.
Prior to differential gene expression analysis, for each sequenced library, the read counts were adjusted by edgeR program package through one scaling normalized factor. Differential expression analysis of two conditions was performed using the DEGSeq R package (1.20.0). The P values were adjusted using the Benjamini & Hochberg method. Corrected P-value of 0.005 and log 2 (Fold change) of 1 were set as the threshold for significantly differential expression.
Genome_build: The used potato genome is that of the doubled monoploid S. tuberosum Group Phureja DM1-3 (DM; AEWC00000000).
Supplementary_files_format_and_content: Readcount_TPM.txt: Matrix table with raw gene counts for every gene and every sample.

organism: Solanum tuberosum

source_name: potato seedling

growth_protocol: Potato seedlings were planted in MS medium (20g/L sucrose) and grown in growth chambers with 16 h light/8 h dark photoperiod at 22℃.

extract_protocol:
RNA was harvested using Trizol reagent.
Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample.

treatment_protocol: 4-week-old potato seedlings growing on the MS media were treated with TOR inhibitors (rapamycin (10μM) plus KU (10μM)) and DMSO for 48h.

tag: tissue

value: Seedling

value: Rapamycin plus KU63794


title: XRN2 KD rep 2 DRIP-seq

description: Supplemental Data 1


title: wsmxO516

city: Qingdao

first: Sijia

last: Kong

address: 238 Songling Road

institute: Ocean University of China

description: incubated for 24 hrs

data_processing:
Base-calling performed using CASAVA version 1.8.2.
Clean data (clean reads) were obtained by removing reads containing adapter, reads containing poly-N and low quality reads from raw data. Both building index of reference genome DSM 20549 and aligning clean reads to reference genome were used Bowtie2-2.2.3.
HTSeq v0.6.1 was used to count the reads numbers mapped to each gene. Then, FPKM (expected number of Fragments Per Kilobase of transcript sequence per Millions of base pairs sequenced) of each gene was calculated based on the length of the gene and reads count mapped to this gene using a protocol from Trapnell et al.
Genome_build: DSM 20549 (GCA_003664125.1)
Supplementary_files_format_and_content: *FPKM.txt: Tab-delimited text files include FPKM values for each Sample.

organism: Planococcus kocurii

source_name: Planococcus kocurii O516

extract_protocol:
The bacterial suspension was centrifuged at 4000 rpm and 4°C to remove the supernatant, and RNA was harvested using Trizol reagent according to instructions.
NEBNext® UltraTM Directional RNA Library Prep Kit for Illumina® (NEB, USA) was used with 3 ug of total RNA for the construction of sequencing libraries following the manufacturer’s recommendations. Briefly, a specialized kit was used to remove rRNA from total RNA to purify mRNA. Then, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA) to select cDNA fragments of preferentially 150~200 bp in length. PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, library quality was assessed on the Agilent Bioanalyzer 2100 system.

value: With sulfamethoxazole


title: UnS- 4dai Replicate 1 [A]

description: Uninfected roots

value: 19 days

value: Susceptible

value: none (uninfected)


title: MA004

value: A


title: PRMT5 KD rep 2 DRIP-seq

description: Supplemental Data 1

value: PRMT5 KD


title: DMSO control group

description:
DMS
Potato
miRNA

value: DMSO


title: nsmxO516

description: incubated for 24 hrs

value: Without sulfamethoxazole


title: SCMC H3K4me3

city: Bethesda

first: Javed

last: Khan

state: MD

address: 37 Convent Dr.

department: Genetics Branch

institute: NCI, NIH

data_processing:
ChIP enriched DNA reads were mapped to reference genome using BWA version 0.7.10
The ChIP-seq peaks were called by MACS2 version 2.1.0.  Duplicates were allowed
Enrichment of known and de novo motifs were found using HOMER version 4.8.2
Genome_build: Hg19
Supplementary_files_format_and_content: peak files are tab delimited text files (BED coordinates) for each sample.

source_name: SCMC

growth_protocol: Cells were maintained in DMEM with 10% FBS and Penn/Strep at 37 °C incubation until harvesting for ChIP.

extract_protocol:
harvesting protocol: ChIP samples harvested after fixation, sonicated, processed and ChIP-enriched via standard protocols (iDeal ChIP-seq kit for Transcription Factors, Diagenode).
DNA was extracted from ChIP-seq samples prepared following the iDeal ChIP-seq kit for Transcription Factors (Diagenode) protocol.
Standard Illumina barcodes were introduced during library preparation with the Illumina TruSeq ChIP Sample Prep Kit 48 Samples - Set A Cat# IP-202-1012

value: SCMC

tag: enrichment target

tag: condition

value: Histone modification

tag: target function

value: no treatment


title: MA003


title: DDX5 KD input DRIP-seq

description: Supplemental Data 1

value: DDX5 KD input


title: DDX5 KD rep 2 DRIP-seq

description: Supplemental Data 1

value: DDX5 KD


title: MA002


title: XRN2 KD input DRIP-seq

description: Supplemental Data 1

value: XRN2 KD input


title: DDX5 KD rep 1 DRIP-seq

description: Supplemental Data 1

value: Monoclonal S9.6


title: SCMC MTA2

value: Anti-MTA2/PID antibody, abcam, ab8106

value: NuRD Complex


title: CM024

data_processing:
Paired end/2x100 bp reads, illumina novaseq 6000
Illumina Casava1.7 software used for basecalling.
Reads were pseudoaligned by Kallisto v0.44 o mouse reference transcriptome mmGRCm38.
Genome_build: mmGRCm38
Supplementary_files_format_and_content: raw gene counts for each sample generated by kallisto

source_name: organotypic brain culture

growth_protocol: Cerebella were isolated from the brains of 7-day-old mice and cut with a McIlwain tissue chopper (WPI-Europe) to obtain350-µm-thick progressive slices. The brain slices were then dissociated in cold Hibernate®-A/5 g/L D-Glucose/1x Kynurenic acid buffer and laid out on Millipore cell culture insert membranes (Millicell cell culture insert, 30 mm, hydrophilic polytetraﬂuoroethylene, Millipore). Slices were cultured in GlutaMAX minimal essential medium supplemented with 25% horse serum, 5 g/L glucose, 1% HEPES (all Thermo Fisher Scientific), and 0.1 mg/L human recombinant insulin (R&D Systems) at 37°C in 5% CO2 in a humidiﬁed atmosphere. The medium was changed daily after the slicing procedure.

treatment_protocol: Slices from 5 mice were infected on the day of slicing with MeV IC323-EGFP-F L454W virus (5.103pfu/slice from IFNAR1KO and 100pfu/slice from SLAM/CD150tg x IFNAR1KO mice). Cultures were then treated daily from day 1 to day 4 with either serial dilutions of HRC4 peptide in Neurobasal medium or vehicle (untreated; “NT”), by adding 2µl of 10000 nM, 1000 nM or 100nM of HRC4 on top of each slice in each well. After several minutes, the drops containing the peptides were completely absorbed and reached the lower compartment of the Millipore well that contains the feeding medium. The final concentration in the medium of culture (1ml) was 100 nM, 10 nM or 1nM. At each time point, slices were collected, RNA extracted and RT-qPCR performed

value: B


title: SCMC RBBP4

value: RBBP4, Epigentek (catA-2703 lotA511004 pAb)


title: MA001


title: CM023


title: CM017

value: F-L454W measles virus


title: SCMC CHD4

value: CHD4 Invitrogen PA527472


title: Control rep 2 Rnase-H DRIP-seq

description: Supplemental Data 1

value: Control Rnase-H


title: FB018


title: PRMT5 KD input DRIP-seq

description: Supplemental Data 1

value: PRMT5 KD input


title: SCMC Input control

value: input

value: control


title: Control input DRIP-seq

description: Supplemental Data 1

value: Control input


title: RH5 H3K4me3

value: H3K4me3, Abcam (ab8580)

value: Histone modification


title: PRMT5 KD rep 1 DRIP-seq

description: Supplemental Data 1

value: PRMT5 KD


title: CM007

value: A


title: FB003

value: N462K mutant measles virus


title: Control rep 2 DRIP-seq

description: Supplemental Data 1

value: Monoclonal S9.6

value: Control


title: RH5 MTA2

value: Anti-MTA2/PID antibody, Abcam (ab8106)

value: NuRD Complex

value: no treatment


title: CM006

value: F-L454W measles virus


title: Control rep 1 DRIP-seq

description: Supplemental Data 1


title: FB011

value: B


title: MA010

value: L454W.E455G mutant measles virus


title: CM005


title: RH5 CHD4

value: CHD4, Invitrogen (PA527472)


title: MA009


title: RH5 RBBP4

value: RBBP4, Epigentek (catA-2703 lotA511004 pAb)


title: RH5 Input control

value: input

value: control


title: CM004

value: WT measles virus


title: Control rep 1 Rnase-H DRIP-seq

description: Supplemental Data 1

value: Control Rnase-H


title: RH4 MTA2


title: CM001


title: CM003


title: FB017


title: MA005

value: L454W mutant measles virus


title: CM008

value: A


title: MA006


title: TuMV_3

status: Public on Aug 07 2020

city: Nanjing

first: Shanwu

last: Lyu

address: Weigang 1

department: College of Horticulture

institute: Nanjing Agriculture University

data_processing:
Basecalls performed using CASAVA version 1.8
The Brassicaceae Genome Data v 1.5 (ftp://brassicadb.org/Brassica_rapa/Bra_Chromosome_V1.5/) was used as the reference genome.
After removing data corresponding to impurities using the Trimmomatic program (v 0.33) (Kim et al., 2018), clean reads from the raw data were mapped to the reference genome using a short read alignment software SOAPaligner/SOAP2 (Li et al., 2009) with two mismatches for mapping the reads to the reference genome.
The result mapped to the genome was recorded for further analysis. Expression levels of each unigene were calculated using the reads kilobases per million reads (RPKM) method (Mortazavi et al., 2008).
We used the Pearson correlation coefficients (PCC) to assess repeatability within replications. The longest transcript was used for the calculation of gene expression and coverage if there were several transcripts in a unigene. We used a false discovery rate (FRD) of ≤ 0.001 and the absolute value of log2Ratio ≥ 1 to screen for DEGs.
Genome_build: Brassicaceae Genome Data v 1.5
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

organism: Brassica rapa subsp. pekinensis

source_name: Chinese cabbage

growth_protocol: Seeds (chiif-401-42) were pre-germinated for 2 days at 25°C and were transferred into an incubator set at 20-25°C with a 16-h light/8-h dark photoperiod and a constant 85% relative humidity.

extract_protocol:
Total RNA was extracted from the TuMV-infected and control leaves using TRIzol reagent (Takara Bio Inc., Otsu, Japan) following the manufacturer’s protocol.
RNA quantity and quality assessment were performed using the Agilent 2100 Bioanalyzer (Agilent, Santa Clara, CA) and gel electrophoresis. After treatment with DNase I and enrichment using the oligo (dT) magnetic beads (for eukaryotes), the mRNA was fragmented into short fragments (about 200 bp). Subsequently, double-stranded cDNA was synthesized, and the sequence adaptors were ligated to the fragments.

treatment_protocol: The five-leaf stage seedlings were rubbed with the TuMV inoculum by grounding the TuMV-infected Chinese cabbage leaves with phosphate buffer (pH 7.4). The plants inoculated only with buffer served as control. At 21 days post-inoculation (dpi), >2.0 g each of the infected and the control leaves were collected.

value: TuMV inoculum

value: leaf


title: Ck_3

value: phosphate buffer


title: hSAECs RSV 1

city: Madison

first: Allan

last: Brasier

state: WI

address: 715 Highland

department: ICTR

institute: university of Wissconsin

data_processing:
quality assesssment of fastq files was performed using fastQC
Adapter removal was performed using TrimGalore
alignment was performed using Rsubread to GRCh38.p13 NCBI genome assembly
Peak calling was using genrich to create narrowpeak files
Genome_build: GRCh38.p13
Supplementary_files_format_and_content: peak

source_name: human small airway epithelial cells

growth_protocol: hSAECs were grown in SAGM small airway epithelial cell growth medium (Lonza, Walkersville, MD) in a humidified atmosphere of 5% CO2.

extract_protocol:
Total RNA was isolated using RNeasy with on-column DNase I digestion (Qiagen) according to the manufacturers recommendation
eluted DNA was pre-amplified for 5 cycles in PCR mix (30 µL NEB Next High-Fidelity 2x master mix (NEB, Cat. No.: M054 1S), 10 µl primer mix (Illumina, NEXtera DNA UD indexes, Cat. No.: 20026121), 20 µL purified DNA) for 5 cycles.
NovaSeq 6000 with 50 million reads per sample

treatment_protocol: RSV Long strain was used to infect cells at an Multiplicity of infection of 1.0

tag: sample type

value: telomerase immortalized

tag: infection

value: infected with Respiratory syncytial virus


title: hSAECs RSV2


title: TuMV_2

tag: treatment


title: hSAECs CON 1

value: uninfected


title: TuMV_1


title: hSAECs CON 2


title: Ck_1


title: sac3bc_mRNA_seq rep3

city: Shanghai

first: Chenjiang

last: You

state: Shanghai

address: 2005 Songhu Rd

institute: Fudan University

data_processing:
The base-calling was performed by Illumina's pre-installed software.
Data analysis was performed with the pRNASeqTools pipeline (https://github.com/grubbybio/RNASeqTools).
The raw reads were aligned to the Araport11 genome using STAR. Transcript levels were measured in reads per million total read counts.
Differentially expressed genes were identified using DEseq2 with fold change of 2 and p < 0.01 as the parameters.
Genome_build: Araport11
Supplementary_files_format_and_content: tab-delimited text files including raw read count for each gene

organism: Arabidopsis thaliana

source_name: 14 days old seedlings

growth_protocol: 16 h light+ 8 h dark at 22 ˚C

extract_protocol:
Total RNAs were isolated by Trizol RNA isolation method. Polyadenylated RNA was isolated from total RNA using the Magnetic mRNA Isolation Kit (New England Biolabs).
RNA-seq libraries were prepared using NEBNext mRNA Library Prep Reagent Set for Illumina (New England Biolabs) and sequenced on an Illumina Hiseq 2500 platform to generate high-quality single-end reads of 150bp in length.

treatment_protocol: No treatment

tag: genotype

value: 14 days old seedlings

tag: grwoth medium

value: 1/2 MS with 1%  Sucrose

tag: growth condition

value: 16 h light+ 8 h dark at 22 ˚C


title: sac3bc_mRNA_seq rep2


title: thp1-5_mRNA_seq rep3


title: thp1-5_mRNA_seq rep2


title: thp1-5_mRNA_seq rep1


title: Ck_2


title: Col_mRNA_seq rep2

value: Col


title: Col_mRNA_seq rep1


title: sac3bc_mRNA_seq rep1


title: thp1-5_small_RNA_seq rep1

description: small RNA

data_processing:
The base-calling was performed by Illumina's pre-installed software.
The resulting raw reads were analyzed by a homemade pipeline pRNASeqTools v0.4 (https://github.com/grubbybio/RNASeqTools).
The raw reads were trimmed to remove the 3’ adaptor sequences and filtered based on size (18-42 nt) and quality using cutadapt v1.9.1.
Trimmed reads then were aligned to the Arabidopsis genome (Araport11) using ShortStack v3.4 with parameters ‘-bowtie_m 1000 -ranmax 50 -mmap u -mismatches 0’.
Normalization was performed by calculating the RPMR (reads per million of 45S rRNA reads) value for each miRNA, and comparison was conducted by DESeq2.
Genome_build: Araport11
Supplementary_files_format_and_content: bigwig containing CPM-normalized 18-42 small RNA read counts along the whole genome

source_name: Inflorescence

extract_protocol:
Total RNA was isolated by Trizol RNA isolation method. small RNAs of 18-40 nucleotides (nt) were isolated from total RNA resolved on 15% urea-PAGE gels. Small RNA libraries were constructed following instructions from the NEBNext Multiplex Small RNA Library Prep Set for Illumina (E7300). The libraries were sequenced on Illumina Hiseq 2500.
Small RNA libraries were constructed following instructions from the NEBNext Multiplex Small RNA Library Prep Set for Illumina (E7300). The libraries were sequenced on Illumina Hiseq 2500.

value: Inflorescence

tag: growth medium

value: Soil


title: thp1-5_amS_small_RNA_seq rep2

description: small RNA


title: Col_small_RNA_seq rep1

description: small RNA


title: Col_small_RNA_seq rep2

description: small RNA


title: thp1-5_small_RNA_seq rep2

description: small RNA


title: miR-122 expressing cells (miRNA-seq)

city: Ithaca

first: Andrew

last: Grimson

state: NY

address: 445 Biotech Bldg

department: Molecular Biology and Genetics

institute: Cornell University

description:
miRNA profiling of miR-122-expressing cells.
processed data file: miRNA_read_counts.tsv

data_processing:
Illumina pipeline software v1.8 was used for base calling.
The mapper.pl (-d -e -h -i -j -k AGATCGGAAGAGCACACGTCT -l 18 -m -s reads.fa -v) was used to trim adapters and to collapse non-redundant reads to fasta format.
Reads matching to mature human miRNAs (miRBase and synthetic miRNAs (prefix: hsa-synv14)) were counted and normalized using quantify.pl (-t hsa -d -W).
Genome_build: hg19
Supplementary_files_format_and_content: miRNA_read_counts.tsv: Tab-delimited miRDeep2 output file containing raw and normalized read counts for human miRNAs downloaded from miRBase and the induced synthetic miRNAs (prefix: hsa-synv14).

source_name: miR-122 expressing cells

growth_protocol: Flp-In™ T-REx™ 293 (HEK293) cells were cultured in DMEM containing 10% FBS, 1% Pen/Strap and 0.1% Zeocin.

extract_protocol:
Total RNA from HEK293 cells was isolated with Trizol, with an extra chloroform extraction to remove residual phenol and addition of glyco-blue as a carrier to promote RNA precipitation.
Libraries were prepared from 1000ng total RNA using the NEBNext Multiplex Small RNA Library Prep Set for Illumina (New England Biolabs) by the RNA Sequencing Core at Cornell University.

treatment_protocol: The cells were transduced with lentiviral vectors with or without miRNA expression cassette. After 6 days of selection in 1 mg/mL Geneticin, the stably integrated cells, both control and experimental, were treated with 1 ug/mL Doxycycline to induce the expression of miRNAs for seven days.

tag: cell line

value: Flp-In T-REx 293 Cell

tag: induced mirna


title: thp1-5_amS_small_RNA_seq rep3

description: small RNA


title: amS_small_RNA_seq rep2

description: small RNA

value: amS


title: thp1-5_amS_small_RNA_seq rep1

description: small RNA


title: nup1-cs_small_RNA_seq rep1

description: small RNA


title: amS_small_RNA_seq rep1

description: small RNA


title: miR-1 expressing cells (miRNA-seq)

description:
miRNA profiling of miR-1-expressing cells.
processed data file: miRNA_read_counts.tsv

source_name: miR-1 expressing cells


title: Empty vector control (miRNA-seq)

description:
miRNA profiling of control cells.
processed data file: miRNA_read_counts.tsv

source_name: Empty vector control


title: miR-122 expressing cells rep 2 [AGO-eCLIP-seq]

description: AGO-eCLIP analysis of miR-122-expressing cells.

data_processing:
Library strategy: eCLIP-seq
Illumina pipeline software v1.8 was used for base calling.
Following removal of UMI sequences from the read sequence, the reads were trimmed to remove adapters using cutadapt (-a TGGAATTCTCGGGTGCCAAGG -m 18).
All reads mapping to ribosomal RNAs using Bowtie2-based alignment were removed and the remaining reads were aligned to the human genome (hg19) using TopHat (-g 1 -p 2 --library-type fr-secondstrand). PCR duplicates, as defined by reads containing identical UMI barcodes and mapping locations, were removed using an in-house Perl script. These BAM files were uploaded to GEO.
Peaks were called for each sample with CLIPper (available on GEO). The peak sets from replicates were used to obtain reproducible peaks (IDR < 0.1) using Irreproducibility discovery rate (IDR) analysis (available on GEO).
For each miRNA, the reproducible peaks from the cells expressing that miRNA and the control cells were pooled and overlapping peaks were merged using BEDTools to produce a set of consolidated eCLIP peaks for each miRNA. The eCLIP reads mapping to these consolidated peak sets were counted using featureCounts (-F SAF -s 1 -Q 50).
Genome_build: hg19
Supplementary_files_format_and_content: clipper.bed files: Set of CLIPper peaks in bed file format.
Supplementary_files_format_and_content: IDR files: Set of reproducible peaks in bed file format.
Supplementary_files_format_and_content: read.counts.tsv: Raw counts of reads mapping to the consolidated peak sets in a tab-separated file format.

extract_protocol:
UV-crosslinked cells were lysed and AGO2 was immunoprecipitated, and an RNA adaptor (P-TGGAATTCTCGGGTGCCAAGG/3InvdT) was ligated onto the 3′ end of co-immunoprecipitated RNA. AGO2-RNA complexes were run on SDS-PAGE and transferred to nitrocellulose membrane for size selection. Following the proteinase K treatment to digest AGO2 protein, the RNA was extracted using acid phenol:chloroform.
The extracted RNA was reverse transcribed, and an ssDNA adaptor with a 10-nucleotide unique molecular identifier (P-NNNNNNNNNNGATCGTCGGACTGTAGAACTCTGAAC/3InvdT) was ligated to the 3′ end of the cDNA. Illumina-compatible sequences and sample barcodes were added via PCR.

treatment_protocol: The cells were transduced with lentiviral vectors with or without miRNA expression cassette. After 6 days of selection in 1 mg/mL Geneticin, the stably integrated cells, both control and experimental, were treated with 1 ug/mL Doxycycline to induce the expression of miRNAs for seven (rep 1) or eight (rep 2) days and UV crosslinked.


title: miR-1 expressing cells rep 2 [AGO-eCLIP-seq]

description: AGO-eCLIP analysis of miR-1-expressing cells.


title: miR-122 expressing cells rep 1 [AGO-eCLIP-seq]


title: nup1-cs_small_RNA_seq rep2

description: small RNA


title: miR-373 expressing cells rep 3 (batch 2) [RNA-seq]

description:
RNA-seq analysis of miR-373-expressing cells.
PolyA RNA
processed data file: batch2.exon.read.count.tsv

data_processing:
Illumina pipeline software v1.8 was used for base calling.
Raw reads were trimmed to remove adapter sequences using fastx_clipper (-l 15 -Q33 -a GATCGGAAGAGCACACGTCTGAACTCCAGTC) and were aligned to the human genome (hg19) using TopHat v2.1.1 (--library-type fr-firststrand).
The featureCounts software was used to count reads mapping to exons (-F SAF -s 2 -Q 50) and introns (-F SAF -s 2 -Q 50 --fracOverlap 1).
The differential expression analysis of RNA-seq and the Combined Analysis of RNA-seq and PRO-seq (CARP) were performed using edgeR v3.24.3 and q-values were computed using qvalue v2.14.1 R package. The RNA-seq and PRO-seq data that were used for the CARP analyses were from the current Series and from the GSE140365 Series, respectively.
Genome_build: hg19
Supplementary_files_format_and_content: exon.read.count files: Tab-separated count files, containing counts of RNA-seq reads mapping to exons.
Supplementary_files_format_and_content: edgeR.rnaSeq files: Tab-separated edgeR output files for RNA-seq data, one for each miRNA used in this study.
Supplementary_files_format_and_content: edgeR.CARP files: Tab-separated edgeR output files for CARP analyses, one for each miRNA used in this study.

source_name: miR-373 expressing cells

extract_protocol:
Total RNA from HEK293 cells was isolated with Trizol, with an extra chloroform extraction to remove residual phenol and addition of glyco-blue as a carrier to promote RNA precipitation.
Directional RNA-seq libraries were prepared from 1000ng total RNA per sample using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (New England Biolabs), with initial polyA+ isolation, by the RNA Sequencing Core and the Transcriptional Regulation and Expression Facility at Cornell University.

tag: batch


title: Empty vector control rep 1 [AGO-eCLIP-seq]

description: AGO-eCLIP analysis of control cells.


title: adjacent non-neoplastic tissue sample NC

type: RNA

city: Gaozhou

first: Zhi-Hai

state: Guangdong

address: 89 Xiguan road

department: Urology

institute: Gaozhou People's Hospital

description: Gene expression in prostate cancer tissues surgucally removed tissue samples

hyb_protocol: DNA in hybridization solution was denatured at 95℃ for 3 min prior to loading onto a microarray. Arrays were hybridized was preformed in a Agilent Hybridization Oven overnight at a rotation speed of 20 rpm at 42℃ and washed with two consecutive solutions (0.2% SDS, 2× SSC at 42℃ for 5 min, and 0.2× SSC for 5 min at room temperature).

scan_protocol: Scanned on an Agilent G2565CA scanner.

data_processing: Feature Extraction v10.7 (Agilent Technologies, CA) software was used to extract all features of the data obtained from the scanned images and genespring software was used to analyze the raw data, which are normalized by percentile normalization.

source_name: prostate cancer sample

label_protocol: cDNA labeled with a fluorescent dye (Cy3-dCTP) was produced by Eberwine’s linear RNA amplification method and subsequent enzymatic reaction. This procedure has been previously described, and the procedure has been improved by using CapitalBio cRNA Amplification and Labeling Kit (CapitalBio) for producing higher yields of labeled cDNA.

extract_protocol: RNA was prepared using the TRIzol reagent following the manufacturer's recommendations.  RNA was quantified using a NanoDrop-1000 spectrophotometer.


title: Empty vector control rep 2 [AGO-eCLIP-seq]

description: AGO-eCLIP analysis of control cells.


title: amS_small_RNA_seq rep3

description: small RNA


title: miR-373 expressing cells rep 2 (batch 2) [RNA-seq]


title: prostate cancer sample CA

value: prostate cancer sample CA


title: HCT116_H3.3K9M

type: genomic

city: Grand Rapids

first: Scott

last: Rothbart

state: MI

address: 333 Bostwick Ave NE

department: Center for Epigenetics

institute: Van Andel Research Institute

hyb_protocol: bisulphite converted DNA was amplified, fragmented and hybridised to the Illumina Infinium MethylationEPIC Beadchip using standard Illumina protocol

scan_protocol: Arrays were imaged using iScan using standard recommended Illumina scanner setting

data_processing: openSesame from SeSAMe R package

label: Cy3, Cy5

label_protocol: Standard Illumina Protocol

growth_protocol: 37ºC, 5% CO2, McCoy's 5A (HCT116) or DMEM (HeLa) media with 10% FBS and no antibiotics

extract_protocol: Qiagen Dneasy kit

tag: compound


title: miR-372 expressing cells rep 3 (batch 2) [RNA-seq]

description:
RNA-seq analysis of miR-372-expressing cells.
PolyA RNA
processed data file: batch2.exon.read.count.tsv

source_name: miR-372 expressing cells


title: miR-302a expressing cells rep 2 (batch 2) [RNA-seq]

description:
RNA-seq analysis of miR-302a-expressing cells.
PolyA RNA
processed data file: batch2.exon.read.count.tsv

source_name: miR-302a expressing cells


title: miR-372 expressing cells rep 2 (batch 2) [RNA-seq]


title: miR-372 expressing cells rep 1 (batch 2) [RNA-seq]


title: miR-302a expressing cells rep 3 (batch 2) [RNA-seq]


title: miR-302a expressing cells rep 1 (batch 2) [RNA-seq]


title: miR-155 expressing cells rep 1 (batch 2) [RNA-seq]

description:
RNA-seq analysis of miR-155-expressing cells.
PolyA RNA
processed data file: batch2.exon.read.count.tsv

source_name: miR-155 expressing cells


title: miR-155 expressing cells rep 2 (batch 2) [RNA-seq]


title: miR-155 expressing cells rep 3 (batch 2) [RNA-seq]


title: HCT116_H3.3WT


title: miR-1 expressing cells rep 1 [AGO-eCLIP-seq]


title: Empty vector control rep 3 (batch 2) [RNA-seq]

description:
RNA-seq analysis of control cells.
PolyA RNA
processed data file: batch2.exon.read.count.tsv


title: miR-133a expressing cells rep 3 (batch 2) [RNA-seq]

description:
RNA-seq analysis of miR-133a-expressing cells.
PolyA RNA
processed data file: batch2.exon.read.count.tsv

source_name: miR-133a expressing cells


title: miR-133a expressing cells rep 2 (batch 2) [RNA-seq]


title: miR-133a expressing cells rep 1 (batch 2) [RNA-seq]


title: Empty vector control rep 2 (batch 2) [RNA-seq]


title: Empty vector control rep 1 (batch 2) [RNA-seq]


title: miR-122 expressing cells rep 2 (batch 1) [RNA-seq]

description:
RNA-seq analysis of miR-122-expressing cells.
PolyA RNA
processed data file: batch1.exon.read.count.tsv


title: miR-122 expressing cells rep 1 (batch 1) [RNA-seq]


title: Empty vector control rep 1 (batch 1) [RNA-seq]

description:
RNA-seq analysis of control cells.
PolyA RNA
processed data file: batch1.exon.read.count.tsv


title: miR-1 expressing cells rep 1 (batch 1) [RNA-seq]

description:
RNA-seq analysis of miR-1-expressing cells.
PolyA RNA
processed data file: batch1.exon.read.count.tsv


title: miR-1 expressing cells rep 2 (batch 1) [RNA-seq]


title: miR-373 expressing cells rep 1 (batch 2) [RNA-seq]


title: K562_Imatinib_rep2 [3T-fill]

city: Solna

first: Vicent

last: Pelechano

address: Nobels väg 16

department: MTC

country: Sweden

institute: ScilifeLab - Karolinska Institutet

description:
B-case
PolyA RNA

data_processing:
Library strategy: 3'T-fill
We employed bcf2fastq (v2.20.0) for converting raw image to sequence information (FASTQ files) and meanwhile demultiplex, allowing two mismatches in index 1 and one mismatch in index 2.
For TIF-Seq2 data, we collapsed all 5’-end or 3’-end sequence reads in each sample according to the indexes. Cutadapt (v1.16) was utilized to trim TIF-seq2 sequencing primer (-AGGTGACCGGCAGGTGT) and Illumina TruSeq adapter (-AGATCGGAAG). After extracting 8 bp of unique molecular identifiers (UMIs) with UMItools (v0.5.4) from the 5’ ends and removing extra A stretches in the 3’ ends caused by poly(A) slippery during PCR amplification, we kept reads over 20 bp for alignment. We used STAR (v2.5.3a) for aligning paired-end reads separately to the human reference genome hg38, supplying Gencode V27 transcript annotation splicing junction as reference annotation. Alignment setting was adjusted as below, --alignIntronMax 200000 --alignEndsType Extend5pOfRead1 --alignSJoverhangMin 10. We then linked paired-end reads and kept the uniquely mapped pairs that are in the same chromosome. Furthermore, a customised script adapted from UMItools was employed to remove PCR duplicates from the leftover reads, allowing 1 bp mismatch in the UMIs and 1 bp shifting in the transcription start sites.
For 3’T-fill sequencing data, we trimmed Illumina TruSeq adapter (-AGATCGGAAG) and extra A stretches in the 3’ ends with cutadapt v1.16. Reads over 20 bp are aligned to hg38 by using STAR v2.5.3a in paired-end mode, supplying Gencode V27 transcript annotation splicing junction as reference annotation with adjusted setting, --alignSJDBoverhangMin 1 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --alignEndsType Extend5pOfReads12.
A customised python script was employed to extract both boundaries of TIF-Seq2 read pairs and 3’ boundaries of 3’T-fill sequencing data, collapse the boundary tags and calculate the coverage (saved in *boundary.txt and bedGraph files).
For PacBio long-read sequencing data, highly accurate consensus sequences were produced by Pacbio ccs package from sequencing facility. We trimmed the PCR primers from both ends of the reads with cutadapt (v1.16) and extract UMIs with UMItools (v1.0.0), keeping the reads with at least 200 bp in length. Then the reads were aligned to human reference genome using minimap2 (v2.16) with the following setting (-ax splice -uf -C5 -O6,24 -B4). We used UMItools for removing the PCR duplicates with adjusted setting as --method cluster --spliced-is-unique. We further employed BEDTools (v2.27.1) bamToBed function to convert the alignment reads into BED format and a customised script to convert BED format to GFF format.
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: *_boundary.txt: Tab-delimited text files of coverage at transcription boundaries for TIF-Seq2.
Supplementary_files_format_and_content: *.3end.bedGraph: bedGraph files of read coverage at 3' ends for 3T-fill sequencing.
Supplementary_files_format_and_content: K562_target_Pacbio.gff: GFF file of validated cDNA sequence annotation for PacBio long-read sequencing data.

source_name: chronic myeloid leukemia cells

growth_protocol: Cells were cultured in RPMI 1640 medium supplemented with 10% FBS, 2 mM L-glutamine, 1% pen/strep (Life Technologies) at 37 °C with 5% CO2 in a humidified atmosphere.

extract_protocol:
RNA was extracted with Trizol and Direct-zol RNA kit from Zymo Research.
3’T-fill: We performed 3’T-fill as previously described (PMID: 23295673). In brief, 10µg DNA-free total RNA (16µL) was fragmented by adding 4µl fragmentation buffer(5X)(200mM Tris-acetate, pH 8.1, 500mM Potassium Acetate, 150mM Magnesium acetate) and incubated at 80˚C for 5min. Fragmented RNA was purified with 1.5X Ampure XP beads and used as the template for reverse transcription(RT). Sample were mixed with biotinylated dT primer (P5_dT16VN: 5’- [Btn]AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCTTTTTTTTTTTTTTTTVN-3’) (where V refers to A, C or G) and denatured at 65˚C for 5min. Denatured RNA was then mixed with 4µl 5X first strand buffer, 2µl DTT(0.1M) and 4µl freshly prepared actinomycin D(0.1µg/ul). After incubation at 42˚C for 2min, 0.5µl SuperScriptII (Invitrogen) was added, incubated at 42˚C for 50 min and finally 72˚C for 15min. cDNA was purified with 1.5X Ampure XP beads according to manufacturer instruction. For second strand synthesis, cDNA was mixed with of 0.5 µl RNase H (5U/µl)(NEB) and 2 µl DNA Polymerase I (10U/µl) (Thermo Scientific) and incubated at 16˚C for 2.5 hours. cDNA was purified with 0.9X Ampure XP beads according to manufacturer instruction. 20 µl biotin labelled cDNA was mixed with 20µl Dynabeads M280 streptavidin beads (Invitrogen) at room temperature for 15 min. Following end repair and dA tailing, each sample(8 µl) was mixed with 12.5 µl 2X Quick Ligation buffer (NEB), 2.5µL T4 DNA ligase (2000U/µL, NEB) 2µl of anneal duplexed adaptors at 2.5 µM(5’- GTGACTGGAGTTCAGACGTGTGCTCTTCCGATC*T-3’, 5’-Phos-GATCGGAAGAGCACACGTCTGAACTCCAGTCAC[AmC7]-3’) and incubated for at 20˚C for 20min. Adaptor ligated libraries were washed and amplified by PCR by adding 25 µl 2X Fusion High Fidelity Master Mix, 0.5 µl PE1 primer (10µM, 5’-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATC*T-3’)and 0.5µl PE2_MPX primers (10µM, 5’-CAAGCAGAAGACGGCATACGAGATXXXXXXGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC*T-3’; where X refer to the specific barcode). Samples were subjected to the following program: 98˚C for 30 seconds, 18 cycles of 98˚C for 10s, 65˚C for 10s, 72˚C for 10s and finally 72˚C for 5min. Library was purified with 1.8X Ampure XP beads according to manufacturer instruction. 3´T-fill libraries were sequenced simultaneously with TIF-Seq2 libraries (using same condition and sequencing oligonucleotides).

treatment_protocol: K562 cells (3 X 10e5 cells/mL) were exposed to 1µM Imatinib for 24 hours and corresponding DMSO treated control cells were used for TIFseq library preparation, 3’T fill library preparation and PacBio target sequencing.

tag: cell type

value: Chronic myeloid leukemia cells

value: Imatinib


title: miR-122 expressing cells rep 2 [RNA-seq]

description:
8907_10109_63523_HMTGTBGX3_RNA_miR122_2_GCCAAT
RNA-seq analysis of miR-122-expressing cells
codingRegion.read.count.tsv:RNA_miR122_rep2

data_processing:
library strategy: Ribo-seq
The raw reads were trimmed to remove adapter sequences using cutadapt (-a AGATCGGAAGAGCACACGTC -m 18).
Reads originating from rRNA were removed using bowtie2 with default parameters for RNA-seq datasets and using "-L 20" and other default parameters for Ribo-seq datasets.
Remaining reads were mapped to the hg19 genome and gencode v19 annotated transcript using following parameters: --no-novel-juncs --transcriptome-index indexFile -p 3 --library-type fr-firststrand.
Reads mapping to coding region excluding the ends (initial 45nt and ending 15nt) were counted using featureCounts (-F SAF -s 1 -Q 50 -T 10 --fracOverlap 1 ) software of Subread (v1.5.1) package.
Genome_build: hg19
Supplementary_files_format_and_content: Tab-separated file, containing counts of reads originating from coding region for all genes across all samples

extract_protocol:
Cycloheximide-treated cells were lysed and digested with nuclease to generate ribosome footprints (RBFs). Ribosomes and ribosome footprints were isolated by size exclusion with a MicroSpin S-400 column. The cycloheximide-treated cells from the same pool were also used to extract total RNA for RNA-seq libraries.
Ribosome profiling libraries and RNA-seq libraries (used for normalizing ribosome footprint (RBF) data) were generated using the TruSeq Ribo Profile (Mammalian) Kit (illumina). The rRNA was depleted from the ribosome footprints and the total RNA using Ribo-Zero rRNA Removal Kit (illumina). The ribosome footprints were size selected using Urea-PAGE before library preparation. The total RNA was fragmented with heat. The size selected RBFs and fragmented total RNA were ligated to a 3′ adaptor, and reverse transcribed. cDNA was purified by Urea-PAGE and circularized. The cDNA library was amplified and barcoded with Illumina-compatible primers.

treatment_protocol: The cells were transduced with lentiviral vectors with or without miRNA expression cassette. After 6 days of selection in 1 mg/mL Geneticin, the stably integrated cells, both control and experimental, were treated with 1 ug/mL Doxycycline to induce the expression of miRNAs for seven days. To stall ribosomes, cells were treated with 100 ug/ml cycloheximide for two minutes at 37°C.


title: K562_Imatinib_rep1 [3T-fill]

description:
A-case
PolyA RNA


title: K562_Imatinib_rep1 [TIF-seq2]

description:
A-case
PolyA RNA

data_processing:
Library strategy: TIF-Seq2
We employed bcf2fastq (v2.20.0) for converting raw image to sequence information (FASTQ files) and meanwhile demultiplex, allowing two mismatches in index 1 and one mismatch in index 2.
For TIF-Seq2 data, we collapsed all 5’-end or 3’-end sequence reads in each sample according to the indexes. Cutadapt (v1.16) was utilized to trim TIF-seq2 sequencing primer (-AGGTGACCGGCAGGTGT) and Illumina TruSeq adapter (-AGATCGGAAG). After extracting 8 bp of unique molecular identifiers (UMIs) with UMItools (v0.5.4) from the 5’ ends and removing extra A stretches in the 3’ ends caused by poly(A) slippery during PCR amplification, we kept reads over 20 bp for alignment. We used STAR (v2.5.3a) for aligning paired-end reads separately to the human reference genome hg38, supplying Gencode V27 transcript annotation splicing junction as reference annotation. Alignment setting was adjusted as below, --alignIntronMax 200000 --alignEndsType Extend5pOfRead1 --alignSJoverhangMin 10. We then linked paired-end reads and kept the uniquely mapped pairs that are in the same chromosome. Furthermore, a customised script adapted from UMItools was employed to remove PCR duplicates from the leftover reads, allowing 1 bp mismatch in the UMIs and 1 bp shifting in the transcription start sites.
For 3’T-fill sequencing data, we trimmed Illumina TruSeq adapter (-AGATCGGAAG) and extra A stretches in the 3’ ends with cutadapt v1.16. Reads over 20 bp are aligned to hg38 by using STAR v2.5.3a in paired-end mode, supplying Gencode V27 transcript annotation splicing junction as reference annotation with adjusted setting, --alignSJDBoverhangMin 1 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --alignEndsType Extend5pOfReads12.
A customised python script was employed to extract both boundaries of TIF-Seq2 read pairs and 3’ boundaries of 3’T-fill sequencing data, collapse the boundary tags and calculate the coverage (saved in *boundary.txt and bedGraph files).
For PacBio long-read sequencing data, highly accurate consensus sequences were produced by Pacbio ccs package from sequencing facility. We trimmed the PCR primers from both ends of the reads with cutadapt (v1.16) and extract UMIs with UMItools (v1.0.0), keeping the reads with at least 200 bp in length. Then the reads were aligned to human reference genome using minimap2 (v2.16) with the following setting (-ax splice -uf -C5 -O6,24 -B4). We used UMItools for removing the PCR duplicates with adjusted setting as --method cluster --spliced-is-unique. We further employed BEDTools (v2.27.1) bamToBed function to convert the alignment reads into BED format and a customised script to convert BED format to GFF format.
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: *_boundary.txt: Tab-delimited text files of coverage at transcription boundaries for TIF-Seq2.
Supplementary_files_format_and_content: *.3end.bedGraph: bedGraph files of read coverage at 3' ends for 3T-fill sequencing.
Supplementary_files_format_and_content: K562_target_Pacbio.gff: GFF file of validated cDNA sequence annotation for PacBio long-read sequencing data.

extract_protocol:
RNA was extracted with Trizol and Direct-zol RNA kit from Zymo Research.
TIF-Seq2: 2.5 µg total DNase treated RNA was dephosphorylated using Calf Intestinal Alkaline Phosphatase (CIP; NEB). Sample was phenol-chloroform purified and precipitated with ethanol. mRNA was decapped by incubating with Cap-Clip (CellScript) at 37˚C for 60min. After phenol-chloroform purification and ethanol precipitation, RNA was ligated overnight at 16˚C at 5’ with DNA/RNA oligonucleotide adaptor (TIF2-RNA: TCAGACGTGTGCTCTTCCGATCTrNrNrWrNrNrWrNrN) using T4 RNA ligase (NEB) in the presence of 10% DMSO, RiboLock RNase Inhibitor and 1mM ATP. Ligated RNA was purified with 1.8X Ampure XP beads according to the manufacturer instruction and then used as the template for reverse transcription. Ligated mRNAs were reverse transcribed using barcoded oligo-dT primers (TIF2-RT: TAGTTCAGTCTTCAGTACCTCGTGCGGCCGCXXXXXXACACTCTTTCCCTACACGACGCTCTTCCGATCTTTTTTTTTTTTTTTTVN; where X refer to a specific barcode). For the revers transcription reaction, mRNA was mixed with the corresponding TIF2-RT oligo and dNTPs, denatured at 65˚C for 5min and then put on ice. The sample was mixed with first strand buffer, Trehalose and RiboLock RNase inhibitor and incubated first at 42˚C for 2 min. Finally 2 µl of SuperScript III (Thermo Fisher Scientific) was added to each reaction and incubated at 42˚C for 50 min, at 50˚C for 30 min, at 55˚C for 30 min and inactivated at 70˚C for 15 min. RNA template was removed by incubating with RNase H and RNase cocktail (Ambion) at 37˚C for 30min. First-strand cDNA was purified with 2X Ampure XP beads. To avoid saturating the reaction, only half of the obtained cDNA was used for the following PCR, and the rest was stored as a backup. Used cDNA template was further split into two PCR reactions with Terra PCR Direct Polymerase (Takara) with the following program 98˚C for 2min, then 16 cycles of 98˚C for 20s, 60˚C for 30s, 68˚C for 5min (+10s/cycle) and finally 72˚C for 5min. The PCR above used as primers TIF2-Rv: TAGTTCAGTCTTCAGTACCTCGT and TIF2-Fw: TATAGCGGCCGCXXXXXXGTGAC[BtndT]GGAGTTCAGACGTGTGCTCTTCCGATC (where X refers to different barcodes). PCR products were purified with 1X Ampure XP beads and quantified with qubit dsDNA HS assay. PCR products from different samples were pooled together with equal mass. Pooled PCR products were subjected to NOT1-HFNEB) digestion at 37˚C for one hour. After inactivation of NOT1 at 65˚C for 20 min, samples were purified with 1X Ampure XP beads. To favour intramolecular ligation, PCR products were diluted to a final concentration less than 1ng/µl and ligated with a high concentration of T4 DNA ligase (NEB) at 16˚C for at least 16 hours. Unligated linear PRC products were removed by treatment with plasmid-safe. After inactivation at 70˚C for 30min, the circularized cDNA was purified with phenol-chloroform and ethanol precipitation. Circularized cDNA was fragmented by sonication (Covaris ME220; Covaris, Inc) (Duration 240s, peak power 30, duty factor 10, Avg Power(W) 6). The fragments were then purified with 1X Ampure XP beads. Biotin labelled fragments which contained the 5’ and 3’ connecting region was enriched using Dynabeads M280 streptavidin (Invitrogen) and incubating at room temperature for 30 min. Captured fragments were subjected to end repair and dA tailing. To add the required Illumina grafting sequences, each sample (20.5 µl beads) was incubated at room temperature for one hour with a mixture of 10 µl 5X quick ligation buffer (NEB), 19.5 µl nuclease-free water (Ambion), 3 µl T4 DNA ligase and 1 µl 1µM duplexed adaptors. Duplex adaptors where generated by annealing TIF2-forkFw (5’-AATGATACGGCGACCACCGAGATCTACACACACCTGCCGGTCACC*T-3) and TIF2-forkRv (5’-phos-GGTGACCGGCAGGTGTATCTCGTATGCCGTCTTCTGCTTG-3’) at 1µM. After cleaning the beads, the beads were resuspended in 20µL H2O and used for PCR amplification using 25µl Phusion High-fidelity MasterMix (2X) (NEB), 4 µl nuclease-free water and 1 µl PCRgraftP5 primer (10µM, 5’-AATGATACGGCGACCACCGAGATCTACAC-3') and 1 µl PCRgraftP7 primer (10µM, 5’- CAAGCAGAAGACGGCATACGAGAT-3’). Samples were subjected to PCR amplification with the following program: 98˚C for 30s, then 18 cycles of 98˚C for 20s, 65˚C for 30s, 72˚C for 30s and finally 72˚C for 5min. DNA library then went through two step beads-based size selection (first step 0.35X and second step 0.45X) with expected size distribution in the range of 300bp-1000bp. Purified DNA library was adjusted to 4nM, then denatured, diluted according to Illumina instruction and sequenced on Nextseq 500 platform. We used four custom sequencing oligos as follows: SeqR1+15T (5’-ACACTCTTTCCCTACACGACGCTCTTCCGATCTTTTTTTTTTTTTTTT-3’), SeqINDX1 (5’-GATCGGAAGAGCACACGTCTGAACTCCAGTCAC-3’), SeqINDX2 (5’-GATCGGAAGAGCGTCGTGTAGGGAAAGAGTGT-3’) and SeqR2 (5’-GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT-3’). Two mixtures of sequencing oligos were prepared as follows: mixture 1 contained SeqR1+15T(0.3µM) and SeqR2(0.3µM), mixture 2 composed of SeqINDX1 primer (0.3µM) and SeqINDX2 primer (0.3µM). Mixture 1 was loaded into both positions (#7 and #8 on reagent cartridge in a NextSeq 500 instrument) for custom read1 primer and for custom read2 primer. Mixture2 was loaded into the position for custom index primer (#9 on reagent cartridge). Sequencing was carried out in an Illumina NextSeq 500 instrument with stand-alone configuration and custom sequencing oligos. Paired end sequencing read lengths were set as read1 76bp, read2 76bp, index1 6bp and index2 6bp.


title: K562_DMSO_rep2 [3T-fill]

description:
B-ctrl
PolyA RNA

value: DMSO


title: K562_Imatinib_rep2 [TIF-seq2]

description:
B-case
PolyA RNA


title: K562_DMSO_rep1 [3T-fill]

description:
A-ctrl
PolyA RNA


title: Empty vector control rep 2 [RNA-seq]

description:
8907_10109_63519_HMTGTBGX3_RNA_empty_2_CGATGT
RNA-seq analysis of control cells
codingRegion.read.count.tsv:RNA_empty_rep2


title: Empty vector control rep 1 [RNA-seq]

description:
8907_10109_63518_HMTGTBGX3_RNA_empty_1_ATCACG
RNA-seq analysis of control cells
codingRegion.read.count.tsv:RNA_empty_rep1


title: K562_DMSO_rep1 [TIF-seq2]

description:
A-ctrl
PolyA RNA


title: miR-122 expressing cells rep 1 [RNA-seq]

description:
8907_10109_63522_HMTGTBGX3_RNA_miR122_1_ACAGTG
RNA-seq analysis of miR-122-expressing cells
codingRegion.read.count.tsv:RNA_miR122_rep1


title: miR-1 expressing cells rep 1 [Ribo-seq]

description:
8907_10109_63526_HMTGTBGX3_RBF_miR1_1_GATCAG
Ribo-seq analysis of miR-1-expressing cells
codingRegion.read.count.tsv:RBF_miR1_rep1

data_processing:
library strategy: Ribo-seq
library strategy: Ribo-seq
The raw reads were trimmed to remove adapter sequences using cutadapt (-a AGATCGGAAGAGCACACGTC -m 18).
Reads originating from rRNA were removed using bowtie2 with default parameters for RNA-seq datasets and using "-L 20" and other default parameters for Ribo-seq datasets.
Remaining reads were mapped to the hg19 genome and gencode v19 annotated transcript using following parameters: --no-novel-juncs --transcriptome-index indexFile -p 3 --library-type fr-firststrand.
Reads mapping to coding region excluding the ends (initial 45nt and ending 15nt) were counted using featureCounts (-F SAF -s 1 -Q 50 -T 10 --fracOverlap 1 ) software of Subread (v1.5.1) package.
Genome_build: hg19
Supplementary_files_format_and_content: Tab-separated file, containing counts of reads originating from coding region for all genes across all samples


title: miR-122 expressing cells rep 2 [Ribo-seq]

description:
8907_10109_63529_HMTGTBGX3_RBF_miR122_2_CTTGTA
Ribo-seq analysis of miR-122-expressing cells
codingRegion.read.count.tsv:RBF_miR122_rep2


title: miR-122 expressing cells rep 1 [Ribo-seq]

description:
8907_10109_63528_HMTGTBGX3_RBF_miR122_1_GGCTAC
Ribo-seq analysis of miR-122-expressing cells
codingRegion.read.count.tsv:RBF_miR122_rep1


title: miR-1 expressing cells rep 2 [Ribo-seq]

description:
8907_10109_63527_HMTGTBGX3_RBF_miR1_2_TAGCTT
Ribo-seq analysis of miR-1-expressing cells
codingRegion.read.count.tsv:RBF_miR1_rep2


title: miR-373 expressing cells rep 2 (batch 2) [PRO-seq]

description:
miR-373_rep2_batch2
processed data file: batch2.genebody.read.count

data_processing:
library strategy: PRO-seq
Illumina pipeline software v1.8 was used for base calling. The batch2 datasets were demultiplexed using in-line sample identifier (see extraction protocol), and UMI sequences were stripped from the reads and put them in read names, to be used later for removing PCR duplicates.
Data from both batch1 and batch2 were trimmed to remove adapter sequences using fastx_clipper (-a TGGAATTCTCGGGTGCCAAGG -l 0 -Q33) and cutadapt v1.12 (-a TGGAATTCTCGGGTGCCAAGG -m 0 -f fastq), respectively. The sequences shorter than 15bp were removed and remaining reads were reverse complemented.
The reads originating from PhiX or rRNAs were removed using bowtie (v1.1.2) alignments to respective reference sequences. The high quality reads were aligned to hg19 genome using bowtie-1.1.2 (-p7 -v2 -m1 -q), and the resultant BAM files were used for further analysis.
For batch2, the BAM files were further processed to remove PCR duplicates, defined as multiple reads with same UMI sequence and mapping location. These BAM files were provided to GEO for batch2 samples.
Reads mapping to the genebody were counted using featureCount software (-F SAF -s 1 -Q 50 -T 30) of Subread package (v1.5.1).
Genome_build: hg19
Supplementary_files_format_and_content: BigWig files, one file each strand of the genome
Supplementary_files_format_and_content: Tab-separated files, containing genebody read counts for all genes across all samples

extract_protocol:
The cells were permeabilized to prepare nuclei. The nascent RNA generated using biotin-run-on reaction of nuclei was used for library preparation.
The biotin-labelled RNA was extracted using Trizol, hydrolyzed using NaOH and enriched using streptavidin beads. 3' adapters were ligated and 5' ends were modified, followed by 5' adapter ligation. For batch-2, the 3' adapters containing sample identifiers (allowing to pool libraries after 3' adapter ligation) were used, and both 3' and 5' adapters contained unique molecular index (UMI) at the ligation junction. Purified and adapter ligated biotin-labelled RNAs were reverse transcribed and PCR amplified, followed by size selection and sequencing.

tag: molecule subtype

value: Nascent RNA


title: Empty vector control rep 2 [Ribo-seq]

description:
8907_10109_63525_HMTGTBGX3_RBF_empty_2_ACTTGA
Ribo-seq analysis of control cells
codingRegion.read.count.tsv:RBF_empty_rep2


title: miR-373 expressing cells rep 1 (batch 2) [PRO-seq]

description:
miR-373_rep1_batch2
processed data file: batch2.genebody.read.count


title: Empty vector control rep 1 [Ribo-seq]

description:
8907_10109_63524_HMTGTBGX3_RBF_empty_1_CAGATC
Ribo-seq analysis of control cells
codingRegion.read.count.tsv:RBF_empty_rep1


title: miR-373 expressing cells rep 3 (batch 2) [PRO-seq]

description:
miR-373_rep3_batch2
processed data file: batch2.genebody.read.count


title: miR-372 expressing cells rep 2 (batch 2) [PRO-seq]

description:
miR-372_rep2_batch2
processed data file: batch2.genebody.read.count


title: miR-372 expressing cells rep 1 (batch 2) [PRO-seq]

description:
miR-372_rep1_batch2
processed data file: batch2.genebody.read.count


title: Empty vector control rep 2 (batch 1) [RNA-seq]

description:
RNA-seq analysis of control cells.
PolyA RNA
processed data file: batch1.exon.read.count.tsv


title: miR-302a expressing cells rep 3 (batch 2) [PRO-seq]

description:
miR-302a_rep3_batch2
processed data file: batch2.genebody.read.count


title: miR-1 expressing cells rep 1 [RNA-seq]

description:
8907_10109_63520_HMTGTBGX3_RNA_miR1_1_TTAGGC
RNA-seq analysis of miR-1-expressing cells
codingRegion.read.count.tsv:RNA_miR1_rep1


title: miR-1 expressing cells rep 2 [RNA-seq]

description:
8907_10109_63521_HMTGTBGX3_RNA_miR1_2_TGACCA
RNA-seq analysis of miR-1-expressing cells
codingRegion.read.count.tsv:RNA_miR1_rep2


title: miR-302a expressing cells rep 2 (batch 2) [PRO-seq]

description:
miR-302a_rep2_batch2
processed data file: batch2.genebody.read.count


title: miR-302a expressing cells rep 1 (batch 2) [PRO-seq]

description:
miR-302a_rep1_batch2
processed data file: batch2.genebody.read.count


title: miR-155 expressing cells rep 1 (batch 2) [PRO-seq]

description:
miR-155_rep1_batch2
processed data file: batch2.genebody.read.count


title: miR-155 expressing cells rep 2 (batch 2) [PRO-seq]

description:
miR-155_rep2_batch2
processed data file: batch2.genebody.read.count


title: miR-155 expressing cells rep 3 (batch 2) [PRO-seq]

description:
miR-155_rep3_batch2
processed data file: batch2.genebody.read.count


title: miR-133a expressing cells rep 2 (batch 2) [PRO-seq]

description:
miR-133a_rep2_batch2
processed data file: batch2.genebody.read.count


title: miR-133a expressing cells rep 3 (batch 2) [PRO-seq]

description:
miR-133a_rep3_batch2
processed data file: batch2.genebody.read.count


title: Empty vector control rep 3 (batch 2) [PRO-seq]

description:
Empty_rep3_batch2
processed data file: batch2.genebody.read.count


title: miR-133a expressing cells rep 1 (batch 2) [PRO-seq]

description:
miR-133a_rep1_batch2
processed data file: batch2.genebody.read.count


title: Empty vector control rep 2 (batch 2) [PRO-seq]

description:
Empty_rep2_batch2
processed data file: batch2.genebody.read.count


title: miR-122 expressing cells rep 2 (batch 1) [PRO-seq]

description:
miR-122_rep2_batch1
processed data file: batch1.genebody.read.count


title: Empty vector control rep 1 (batch 2) [PRO-seq]

description:
Empty_rep1_batch2
processed data file: batch2.genebody.read.count


title: miR-122 expressing cells rep 1 (batch 1) [PRO-seq]

description:
miR-122_rep1_batch1
processed data file: batch1.genebody.read.count


title: miR-1 expressing cells rep 2 (batch 1) [PRO-seq]

description:
miR-1_rep2_batch1
processed data file: batch1.genebody.read.count


title: miR-1 expressing cells rep 1 (batch 1) [PRO-seq]

description:
miR-1_rep1_batch1
processed data file: batch1.genebody.read.count


title: Empty vector control rep 1 (batch 1) [PRO-seq]

description:
Empty_rep1_batch1
processed data file: batch1.genebody.read.count


title: miR-372 expressing cells rep 3 (batch 2) [PRO-seq]

description:
miR-372_rep3_batch2
processed data file: batch2.genebody.read.count


title: K562_DMSO_rep2 [TIF-seq2]

description:
B-ctrl
PolyA RNA


title: H3T3ph

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:
Illumina 50bp Single End FastQ data of Immunoprecipitated and Input DNA for Drosophila melanogaster S2 H3T3ph were aligned against the dm3 genome using Bowtie 0.12.5 (Lagmead et al, 2009), using options –v2 –m1.
Downstream analyses of aligned reads were performed using R 2.15 and Bioconductor (Gentleman et al, 2004). The htSeqTools  v.1.4.0 package (Planet et al, 2012) was used to detect and clean aligned data from overamplification artifacts using the filterDuplReads function with a False Discovery Rate of 0.01. Strand shift bias was corrected using the alignPeaks function with default settings.
Enriched H3T3ph sites were detected with the enrichedRegions function using the options minReads=10 and selecting as enriched regions those with a Benjamini-Hochberg pvalue < 0.05, and were annotated to overlapping and closest Drosophila melanogaster dm3 genomic features (Ensembl April 2014) using the AnnotatePeakInBatch function from the ChIPpeakAnno package (Zhu et al, 2010) with the options output='both', select='all', maxgap=0, PeakLocForDistance='middle', FeatureLocForDistance='TSS'.
The rtracklayer package 1.18.2 (Lawrence et al, 2009) was used to generate tracks for normalized IP/Input H3T3ph signal across the genome.
Additional quality control of raw and aligned data were performed using FastQC version 0.10 (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
Genome_build: dm3
Supplementary_files_format_and_content: Processed BED file contains list of enriched regions in H3T3ph vs Input, with columns indicating chromosome, start, end, width, reads for IP sample, reads for Input sample, BH-pvalue, RPKM for IP sample and RPKM for Input sample.

organism: Drosophila melanogaster

source_name: S2 cells

growth_protocol: S2 cells were grown on Schneider’s insect Medium (Sigma) supplemented with 100 μg/ml streptomycin (Gibco), 100 units/ml penicilin (Gibco) and 10% FBS (Gibco)

extract_protocol:
Lysates were clarified from sonicated nuclei and histone-DNA complexes were isolated with aH3T3ph antibodies. Finally DNA was purified by standard phenol extraction.
ChIP-seq, library construction, cluster generation and Sequencing analysis were performed using the Illumina HiSeq2000, sequencing type SR50, following manufacture’s protocols (www.illumina.com). In brief, libraries were prepared using Illumina’s ChIP-Seq Sample Prep Kit from 8.5 ng of DNA. Samples were size selected to ̴300-350 bp using DNA magnetic beads. Adaptor-modified DNA fragments were subjected to limited PCR amplification (18 cycles). Using Illumina’s Cluster Generation Kit, libraries were subject to cluster generation for subsequent sequencing.

tag: fraction

value: IP

tag: antibody

value: rabbit aH3T3ph Millipore 04-746


title: input

value: input


title: Empty vector control rep 2 (batch 1) [PRO-seq]

description:
Empty_rep2_batch1
processed data file: batch1.genebody.read.count


title: cov_17_antibody

city: Stanford

first: Madeleine

last: Scott

address: 729 Lagunita Dr

institute: Stanford

description:
RNA (ADT tag)
cov17_matrix.mtx.gz

data_processing:
Cellranger 3.1
Seurat 3.1.4
Genome_build: Hg38
Supplementary_files_format_and_content: gene expression matrices (mtx). The files barcodes.tsv.gz and features.tsv.gz are also included and for use with all *mtx files.

source_name: antibody tag

extract_protocol:
PBMC isolation directly after blood sample collection
10x Single Cell 3' v3

value: PBMC

tag: sample name

tag: age

tag: Sex

value: M

value: Healthy


title: cov_17_RNA

description:
RNA
cov17_matrix.mtx.gz

source_name: single cell mRNA


title: cov_18_antibody

description:
RNA (ADT tag)
cov18_matrix.mtx.gz

value: F


title: cov_18_RNA

description:
RNA
cov18_matrix.mtx.gz


title: P16 Fgfr1/Fgfr2 double mutant, rep2

city: Safed

first: David

last: Enshell-Seijffers

address: 8 Henrietta Szold

department: The Azrieli Faculty of Medicine

country: Israel

institute: Bar Ilan University

description:
Ribosomal depletion was performed
P16mutC

data_processing:
trimmed and quality filtered using Trimmomatic
mapped to the mouse genome (NCBI38/mm10) using Tophat (version 2)
Mapped read for each annotated ENSEMBL gene (GRCm38.p4) were counted using HTSeq-count tool
Read count normalization and differential gene expression analysis was performed using DESeq2
Genome_build: mm10

source_name: primary dermal papilla sorted cells

extract_protocol:
Total RNA was purified from FACS-sorted cells utilizing the RNeasy Plus Micro kit (QIAGEN)
Ribosomal depletion was performed on 100 ng of total RNA and libraries for Illumina sequencing were performed using the Nebnext Ultra Directional RNA kit (NEB, #E7420L)

value: Fgfr1/Fgfr2 double mutant

tag: hair cycle stage

value: Anagen

tag: time point

value: primary dermal papilla sorted cells


title: cov_12_antibody

description:
RNA (ADT tag)
cov12_matrix.mtx.gz

value: Moderate


title: P16 Fgfr1/Fgfr2 double mutant, rep3

description:
Ribosomal depletion was performed
P16mutD

tag: genotype


title: P16 Fgfr1/Fgfr2 double mutant, rep1

description:
Ribosomal depletion was performed
P16mutB


title: P16 wild type control, rep2

description:
Ribosomal depletion was performed
P16WTC

value: wild type control


title: cov_12_RNA

description:
RNA
cov12_matrix.mtx.gz


title: P16 wild type control, rep3

description:
Ribosomal depletion was performed
P16WTD


title: GM12878 nanoNOMe

city: Baltimore

first: Isac

last: Lee

address: 3400 N. Charles St.

institute: Johns Hopkins University

data_processing:
Library strategy: Oxford Nanopore Sequencing (NOMe-seq)
Raw current signals were converted to DNA sequences using Guppy version 3.0.3 (ONT), using the “high-accuracy” basecalling model.
DNA sequences were aligned to hg38 human reference genome without alternative contigs using NGMLR version 0.2.8 with default settings for aligning Oxford nanopore reads (-x ont)
CpG and GpC methylation were called using nanopolish version 0.11.1.
Nanopolish outputs were converted to methylation bed files (one line per read) as specified by https://github.com/isaclee/nanopore-methylation-utilities using mtsv2bedGraph.py
Methylation bed files were subsequently converted to bismark-style methylation counts (one line per locus)  using parseMethylbed.py frequency in https://github.com/isaclee/nanopore-methylation-utilities
Single-read anallysis was performed on select genomic contexts (CTCF binding sites and gene promoter regions), and the resulting data was deposited into Zenodo (https://zenodo.org/record/3969567)
Genome_build: hg38
Supplementary_files_format_and_content: Methylation Counts files contain the counts of methylation states for each observed genomic locus in the same format as bismark cytosine reports : <chromosome> <position> <strand> <count methylated> <count unmethylated> <C-context> <trinucleotide context> (https://rawgit.com/FelixKrueger/Bismark/master/Docs/Bismark_User_Guide.html)

source_name: nanoNOMe on GM12878 Cell line

growth_protocol: Cell lines were grown according to manufacturer's specifications

extract_protocol:
Samples were treated with proteinase K (NEB) at 55 °C for > 2 hours, and DNA was extracted via phenol:chloroform extraction and ethanol precipitation. After proteinase K treatment, and in all following steps, samples were handled with care using large orifice pipette tips to avoid excessive fragmentation of DNA.
Purified gDNA was prepared for nanopore sequencing following the protocol in the genomic sequencing by ligation kit LSK-SQK108 (ONT). Samples were first sheared to ~10 kb using G-tubes (Covaris): by centrifuging 2-3 μg of unfragmented gDNA at 5,000x g for 1 minute, then inverting the tube and centrifuging again. We sheared the DNA to 10 kb because it produces long fragments of DNA while maximizing the yield of nanopore sequencing. Shearing to larger sizes or unsheared DNA may be used to maximize the length of sequenced reads, with the caveat that sequencing yield will drop. In two samples (GM12878 samples 8 and 9), we targeted 20kb fragments, with an additional step of removing short fragments using the Short Read Eliminator module by Circulomics, following the manufacturer’s specifications. The sheared samples were end-repaired and dA-tailed using NEBnext Ultra II end-repair module (NEB), followed by clean-up using 1x v/v AMPure XP beads (Beckman Coulter). Sequencing adaptors, comprised of leader adaptor DNA and motor proteins, were ligated to the end-prepared DNA fragments using Blunt/TA Ligase Master Mix (NEB), followed by clean-up using 0.4x v/v AMPure XP beads and sequencing kit reagents. >400 ng of adaptor ligated samples per flow cell were loaded onto FLO-MIN106 or PRO-002 flowcells and run on MinION Mk1b, GridION, or PromethION sequencers for up to 72 hours.

treatment_protocol: NOMe-seq was performed on the cells with adjustments for nanopore sequencing. Cells were collected by trypsinization, then nuclei were extracted by incubating in resuspension buffer (100 mM Tris-Cl, pH 7.4, 100 mM NaCl, 30 mM MgCl2) with 0.25 % NP-40 for 5 minutes on ice. Intact nuclei were collected by centrifugation for 5 minutes at 500xg at 4 °C. Nuclei were subjected to a methylation labeling reaction using a solution of 1x M. CviPI Reaction Buffer (NEB), 300 mM sucrose, 96 μM S-adenosylmethionine (SAM; New England Biolabs, NEB), and 200 U M. CviPI (NEB) in 500 μL volume per 500,000 nuclei. The reaction mixture was incubated at 37 °C with shaking on a thermomixer at 1,000 rpm for 15 minutes. SAM was replenished at 96 μM at 7.5 minutes into the reaction. The reaction was stopped by the addition of an equal volume of stop solution (20 mM Tris-Cl, pH 7.9, 600 mM NaCl, 1% SDS, 10 mM disodium EDTA).

value: EBV-transformed lymphoblastoid cell line


title: cov_11_antibody

description:
RNA (ADT tag)
cov11_matrix.mtx.gz

value: Severe


title: cov_11_RNA

description:
RNA
cov11_matrix.mtx.gz


title: cov_10_antibody

description:
RNA (ADT tag)
cov10_matrix.mtx.gz


title: cov_10_RNA

description:
RNA
cov10_matrix.mtx.gz


title: P14 wild type control, rep3

description:
Ribosomal depletion was performed
P14WTE


title: cov_09_RNA

description:
RNA
cov09_matrix.mtx.gz


title: P16 wild type control, rep1

description:
Ribosomal depletion was performed
P16WTB


title: cov_09_antibody

description:
RNA (ADT tag)
cov09_matrix.mtx.gz


title: P14 Fgfr1/Fgfr2 double mutant, rep1

description:
Ribosomal depletion was performed
P14mutA


title: P14 Fgfr1/Fgfr2 double mutant, rep3

description:
Ribosomal depletion was performed
P14mutE


title: cov_08_antibody

description:
RNA (ADT tag)
cov08_matrix.mtx.gz


title: P14 wild type control, rep2

description:
Ribosomal depletion was performed
P14WTC


title: cov_07_antibody

description:
RNA (ADT tag)
cov07_matrix.mtx.gz


title: P14 wild type control, rep1

description:
Ribosomal depletion was performed
P14WTA


title: P7 KO2

first: Haiyang

last: Hu

address: 320 Yueyang Road

institute: PICB

description: expressed RNA in UXT-CKO retina of 7 days old mouse

data_processing:
Basecalling based on Illumina Casava1.7 software
After RNA sequencing reads quality checking, RNA-seq data were mapped to the mouse genome (mm10) using Hisat2 using default parameters。
The gene expression abundance was estimated by counting number of reads mapped to the cognate gene exons using HTSeq
Genome_build: mm10 (mouse)
Supplementary_files_format_and_content: tab-delimited text files include gene ID and expression level in reads count for each sample

source_name: Retina

extract_protocol:
C57BL/6 mice were purchased from the Model Animal Research Center of Nanjing University. The mice were maintained under specific pathogen-free (SPF) conditions at the Center for New Drug Safety Evaluation and Research, China Pharmaceutical University. The Uxt conditional knocking-out mice were generated by mating between Chx10-Cre and Uxtflox/flox mice. Chx10-Cre mice were purchased from Shanghai Biomodel Organism. Uxt flox mice were generated with CRISPR/Cas9-mediated homologous recombination of a 2289bp double stranded DNA donor containing two loxP sites flanking exon 3. Total RNA were extracted with Trizol reagent (Invitrogen). Total RNA was extracted from control and Uxt CKO retinas using the Trizol reagent (Invitrogen) and prepared into cDNA library according to the standard Illumina RNA-seq instruction.
A cDNA library was generated from polyA+ RNA according to the Illumina RNAseq protocol. The cDNA library was sequenced on an Illumina Hiseq2500 by a 2X150 bp run.

tag: strain

tag: tissue

value: Retina

value: UXT-CKO


title: P7 KO1


title: cov_07_RNA

description:
RNA
cov07_matrix.mtx.gz


title: cov_04_RNA

description:
RNA
cov04_matrix.mtx.gz


title: cov_04_antibody

description:
RNA (ADT tag)
cov04_matrix.mtx.gz


title: P7 CTL1

description: expressed RNA in retina of 7 days old mouse

value: control


title: cov_03_antibody

description:
RNA (ADT tag)
cov03_matrix.mtx.gz


title: P14 Fgfr1/Fgfr2 double mutant, rep2

description:
Ribosomal depletion was performed
P14mutC


title: cov_02_antibody

description:
RNA (ADT tag)
cov02_matrix.mtx.gz


title: P30 KO1

description: expressed RNA in UXT-CKO retina of 30 days old mouse


title: Col_mRNA_seq rep3

data_processing:
The base-calling was performed by Illumina's pre-installed software.
Data analysis was performed with the pRNASeqTools pipeline (https://github.com/grubbybio/RNASeqTools).
The raw reads were aligned to the Araport11 genome using STAR. Transcript levels were measured in reads per million total read counts.
Differentially expressed genes were identified using DEseq2 with fold change of 2 and p < 0.01 as the parameters.
Genome_build: Araport11
Supplementary_files_format_and_content: tab-delimited text files including raw read count for each gene

source_name: 14 days old seedlings

value: Col

value: 14 days old seedlings

tag: grwoth medium

value: 1/2 MS with 1%  Sucrose

tag: growth condition

value: 16 h light+ 8 h dark at 22 ˚C


title: P30 CTL1

description: expressed RNA in retina of 30 days old mouse


title: P30 KO2


title: P7 CTL2


title: cov_02_RNA

description:
RNA
cov02_matrix.mtx.gz

value: PBMC


title: P30 CTL2


title: cov_03_RNA

description:
RNA
cov03_matrix.mtx.gz


title: cov_01_antibody

description:
RNA (ADT tag)
cov01_matrix.mtx.gz


title: cov_01_RNA

description:
RNA
cov01_matrix.mtx.gz


title: P1 CTL1

description: expressed RNA in retina of 1 day old mouse


title: P1 KO1

description: expressed RNA in UXT-CKO retina of 1 day old mouse


title: cov_08_RNA

description:
RNA
cov08_matrix.mtx.gz

value: Healthy


title: P1 CTL2


title: P14 UXT KO2

description: expressed RNA in UXT-CKO retina of 14 days old mouse


title: H_10

city: Murcia

first: Ana

middle: María

last: Hurtado López

state: Murcia

address: Libertad, n

department: Hematology and Medical Oncology Department

institute: Hospital Morales Meseguer, Instituto Murciano de Investigación Biosanitaria (IMIB)

data_processing:
Artifacts (low quality reads, adaptor sequence,  contaminant DNA, or PCR duplicates) were removed
Trimmed reads were mapped to reference genome with TopHat, splice-aware aligner.
Transcript was assembled by Cufflinks with aligned reads that contain paired-end information.
After the read mapping, Cufflinks was used for transcript assembly. Expression profile was calculated  for each sample and transcript/gene as FPKM (Fragment per Kilobase of transcript per Million mapped  reads).for each sample and transcript/gene as FPKM (Fragment per Kilobase of transcript per Million mapped reads).
Genome_build: hg19
Supplementary_files_format_and_content: cvs format. FPKM for each sample

source_name: bone marrow

extract_protocol:
Bone marrow samples were colected from CMML patients and healthy donors. RNA was then isolated from buffy coat using Qiagen Rneasy Mini kits as per manufacturer's recommendations.   RNA libraries were prepared for sequencing using standard Illumina protocols
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: disease

value: Healthy donor

tag: gender

value: male


title: P14 UXT KO1


title: P14 CTL2

description: expressed RNA in retina of 14 days old mouse


title: H_20


title: H_17


title: H_19


title: P14 CTL1


title: EB14_5aza_d6

city: Boston

first: Bradley

middle: E

last: Bernstein

state: MA

address: 185 Cambridge Street

department: Pathology

institute: Massachusetts General Hospital

description:
EB_day14_5aza_d6
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

data_processing:
Sequencing RNA libraries, passed through general quality filter such as cluster density, total yield, and per-cycle base quality, were then split by library barcodes using bcl2fastq.
R2 reads were demultiplexed into separate files based on the cell barcode from R1 and mapped to a mouse reference genome (mm9) by Bowtie2. After eliminating reads sharing the same UMI by htseq-count, an accurate molecule count for each gene was generated and converted to the number of UMIs into transcript counts through binomial statistics (CEL-seq2).
Nanopore sequencing reads were converted to fastq files by Guppy (v2.3.5) and then demultiplexed to the each cell based on its matched cell barcode sequence with the highest mapping score by minimap2 with default parameters.
To detect tandem-loxP, different loxP flanking sequences were mapped to the demultiplexed nanopore sequence (minimap2) then assembled into a full tandem-loxP sequence based on their position in the nanopore sequence. Due to the sequencing error, we selected the top 1 combination with additional filter: 1) over 30 reads, 2) outlier from others, 3) full length with upstream and downstream sequence.
To detect the UCI, we first mapped the upstream and downstream of the UCI sequence to nanopore reads by minimap2 then extracted the sequence between M13F and WPRE_M13R_HSV. Due to the sequencing error, we selected the top 1 sequence with additional filter: 1) over 30 reads, 2) outlier from others, 3) represented by the sequence from UCI plasmid.
Genome_build: mm9 (MGSCv37)
Supplementary_files_format_and_content: Tab-delimited text files include UMI count values for each Sample.

source_name: mESC (Mouse embryoid body at day 0)

growth_protocol:
Mouse v6.5 ESCs were cultured with feeder cells (mouse embryonic fibroblasts (MEF)) in knockout DMEM media supplemented with 15% FBS, and 0.01% Leukemia inhibitory factor (LIF). The media was replenished every day and cells were split every two days.
For differentiation, we detached and dissociated ESCs to single cells. MEFs were further removed by incubating single cell suspensions on a 100mm tissue culture dish for 40 minutes at 37°C. We seeded 1000 cells per well micro-well plate (STEMCELL Technologies) in EB media (mESC media without LIF). For inducible barcoding experiments, we seeded FACS sorted RFP+/GFP-/BFPmid cells. After 4 days of EB differentiation in the micro-well plate, we collected EBs and transferred them to 100mm petri dishes and pursued spontaneous differentiation for another 10 days. EB media was changed every day (micro-well plate) or every other day (petri dish) during EB differentiation.

extract_protocol:
mESC and EBs were dissociated by trypsin to single cells and sorted into 384 well plates. After freeze and thaw for twice, we performed a reverse transcription and second strand synthesis, then pooled samples and ran IVT. For DNA extraction (30%), before pooling, we added unique primers (DNA adapter) involving identical cell barcode with RNA-adapter, targeting dual-barcode region of genomic DNA to each well, and then amplified the region by PCR. For RNA extraction (70%), after 2nd reverse transcription, we amplified RNA fragments by Illumina adapter.
RNA libraries were prepared for sequencing using standard Illumina protocols.

treatment_protocol: To perturb DNA methylation, we treated EBs with 5-azacytidine or DMSO (100 nM) and replenished them with every media change throughout all days of differentiation.

value: Embryoid body

tag: media

value: Serum-LIF

tag: differentiation day

value: Day 14

tag: treatment


title: Ctrl H3K9me3 rep1

city: Lund

first: Christopher

last: Douse

address: Sölvegatan 17

department: Medicine

institute: Lund University

data_processing:
Library strategy: CUT&RUN
Read alignment with bowtie2 (v2.3.4.3) --local --very-sensitive-local --no-mixed --no-discordant --phred33 -I 10 -X 700
For CUT&RUN, conversion to bedgraph format and normalization was done with bedtools genomecov (v2.27.1; -bg -scale), where the scale factor was 1e4 divided by the number of reads mapping to the yeast spike-in genome (R64-1-1).
See also GSE155693
Genome_build: hg38
Supplementary_files_format_and_content: bigwig files with normalized coverage plots
Supplementary_files_format_and_content: bed file listing 393 TASOR-regulated H3K9me3 loci

source_name: HeLa cells

growth_protocol: HeLa and HEK293T cells (ECACC) were grown in RPMI plus 10% fetal calf serum (FCS) and penicillin/streptomycin (100 U/mL). Cell lines were routinely tested for mycoplasma contamination using the MycoAlert detection kit (Lonza). Exogenous expression of TASOR variants was done with lentiviral vectors pHRSIN-pSFFV-V5-PPHLN-WPRE-pPGK-Hygro. Lentivirus was generated through the triple transfection of HEK 293T cells with the lentiviral transfer vector plus the two packaging plasmids pCMVΔR8.91 and pMD.G using TransIT-293 transfection reagent (Mirus) as recommended by the manufacturer. Viral supernatant was typically harvested 48 h post-transfection, cell debris removed using a 0.45 µm filter, and PPHLN KO HeLa cells transduced by spin infection at 1,000 g for 90 min.

extract_protocol:
DNA was cleaved by antibody-directed MNase (CUT&RUN) treatment of digitonin-permeabilised cells and purified from the cell supernatant
KAPA Hyperprep kit was used as per instructions. Final cleanup was done with 1.1x KAPA SPRI beads

tag: cell line

value: HeLa

value: Clone bearing HUSH-repressed GFP transgene reporter

value: rabbit anti-H3K9me3 abcam ab8898


title: H_16


title: H_13

value: female


title: EB14_5aza_d4

description:
EB_day14_5aza_d4
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

tag: cell type


title: LM_140

value: Chronic Myelomonocytic Leukemia


title: EB12_5aza

description:
EB_day12_5aza
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Day 12


title: P1 KO2

value: Retina

value: UXT-CKO


title: EB08_5aza

description:
EB_day8_5aza
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Day 8


title: LM_137


title: EB10_5aza

description:
EB_day10_5aza
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Day 10


title: EB06_5aza

description:
EB_day6_5aza
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Day 6


title: H_25


title: EB04_5aza

description:
EB_day4_5aza
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Day 4


title: EB02_5aza

description:
EB_day2_5aza
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Day 2


title: LM_135

value: male


title: LM_133


title: LM_102


title: EB12_dmso

description:
EB_day12_dmso
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Embryoid body

value: DMSO


title: LM_97


title: LM_131


title: EB14_dmso

description:
EB_day14_dmso
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt

value: Serum-LIF

value: Day 14


title: EB08_dmso

description:
EB_day8_dmso
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt


title: LM_143


title: LM_87


title: EB06_dmso

description:
EB_day6_dmso
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt


title: EB10_dmso

description:
EB_day10_dmso
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt


title: LM_86


title: LM_95


title: EB02_dmso

description:
EB_day2_dmso
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt


title: EB04_dmso

description:
EB_day4_dmso
scRNA-Seq
96 cells
processed data file: EB_dmso_5aza_rel_RNA.txt


title: LM_42


title: LM_82


title: LM_39


title: LM_52