title: ESC_DUX_Clone2_DOX_16h_RPA_ChIP

status: Public on Jun 07 2021

city: Bethesda

first: Desiree

last: Tillo

state: MD

address: 41 Center Dr, Room D310

institute: NIH/NCI

data_processing:
Basecalling was performed using bcl2fastq version 2.20.0
Reads were adapter and quality trimmed usin cutadapt v1.18 with the following parameters: --nextseq-trim=2 --trim-n -n 5 -O 5 -q 10,10 -m 35.
Next, reads were aligned  using BWA mem v0.7.17 (Li & Durbin, 2009) to version mm10 of the mouse genome, and PCR duplicates were removed using the 'filterdup' command from macs2 v2.1.1 (Zhang et al., 2009), with the parameter --keep-dup=”auto”.  Fragment length was estimated using phantompeakcalltools v2.0 (Landt et al., 2012) on deduplicated and aligned reads with mapQ>5.
Normalized reads per kilobase per million (RPKM) signal tracks were generated using the ‘bamCoverage’ utility from deepTools  (Ramirez et al., 2016) with the following parameters --binSize 25 --smoothLength 75 --normalizeUsing RPKM -extend_reads [estimated fragment length from phantompeakqualtools].
Genome_build: mm10
Supplementary_files_format_and_content: bigwig files containing RPKM normalized read density.

source_name: mouse ESCDux

growth_protocol: ESCDux  were grown on a feeder layer of growth-arrested MEFs or on gelatin 0.1% in high-glucose DMEM (Invitrogen) supplemented with 15% FBS, 1:500 LIF (made in house), 0.1 mM nonessential amino acids, 1% glutamax, 1mM Sodium Pyruvate, 55 mM β-mercaptoethanol, and 1% penicillin/streptomycin (all from Life Technologies) at 37°C and 5% CO2. Cells were routinely passaged with Trypsin 0.05% (Gibco).

extract_protocol:
Twenty million of untreated or DOX-treated ESCDUX cells were fixed using 1% Formaldehyde (Sigma, F1635) at 37°C for 10 min. Fixation was then quenched with 125mM glycine (Sigma). Cell pellets were washed twice with cold PBS and samples were snap-frozen and stored in −80°C. Frozen pellets were resuspended in 1mL RIPA buffer (10 mM Tris-HCl pH 7.5, 1 mM ethylenediaminetetraacetic acid (EDTA), 0.1% sodium dodecyl sulphate, 0.1% sodium deoxycholate, 1% Triton X-100, and 1 Complete Mini EDTA-free proteinase inhibitor tablet (Roche)). Sonication was performed using Covaris S220 (duty cycle 20%, peak incident power 175, and cycle/burst 200 for 30 min at 4 °C). Chromatin was pre-clarified with 40 μL prewashed Dynabeads Protein A (ThermoFisher) for 30 min at 4 °C and then incubated with 40 μL Dynabeads Protein A bound to 10mg of anti-RPA32 antibody (Abcam ab10359) or 10mg of Guinea Pig anti-Rabbit IgG (ABIN101961, Antibodies-online) in 100uL PBS overnight at 4°C. Beads were then collected in a magnetic separator (DynaMag-2 Invitrogen), washed twice with cold RIPA buffer, twice with RIPA buffer containing 0.3M NaCl, twice with LiCl buffer (0.25 M LiCl, 0.5% Igepal-630, 0.5% sodium deoxycholate), once with TE (10 mM Tris pH 8.0, 1mM EDTA) + 0.2% Triton X-100, and once with TE. Crosslinking was reversed by incubating the chromatin bound beads at 65°C for 4 hours in the presence of 0.3% SDS and 1mg/mL of Proteinase K (Qiagen).
The entire ChIP DNA was used to prepare Illumina sequencing libraries. End-repair was performed in 75 μl of T4 ligase reaction buffer, 0.4 mM of dNTPs, 4 U of T4 DNA polymerase (NEB), 13.5 U of T4 Polynucleotide Kinase (NEB) and 1.5 U of Klenow fragment (NEB) at 24°C for 30 min in a ThermoMixer C at 400 rpm. End-repair reaction was cleaned using 2X Agencourt AMPure XP beads and eluted in 15 μl of EB that was used for A-tailing reaction in 30 μl of NEBNext dA-Tailing reaction buffer (NEB) with 7.5 U of Klenow fragment exo- (NEB) at 37°C for 30 min. The 30 μl of the A-tailing reaction were mixed with Quick Ligase buffer 2X (NEB), 3,000 U of Quick ligase and 5 nM of annealed adaptor (Illumina truncated adaptor) in a volume of 75 μl and incubated at 25C for 20 min. Adaptor was prepared by annealing the following HPLC oligos: 5’-phosphate/GATCGGAAGAGCACACGTCT-3’ and 5’-ACACTCTTTCCCTACACGACGCT CTTCCGATC∗T-3’ (∗phosphorothioate bond). Ligation was stopped by adding 50mM of EDTA and cleaned with 1.8X Agencourt AM- Pure XP beads and eluted in 15ul of EB that was used for PCR amplification in a 50 uL reaction with 1 μM primers TruSeq barcoded primer p5, AATGATACGGCGACCACCGAGATCTACACNNNNNNNNACACTCTTTCCCTACACGACGCTCTTCCGATC∗T, TruSeq barcoded primer p7, CAAGCAGAAGACGGCATACGAGANNNNNNNGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC∗T, (NNNNNNNN represents barcode and ∗ a phosphothiorate bond), and 2X Kapa HiFi HotStart Ready mix (Kapa Biosciences). The temperature settings during the PCR amplification were 45 s at 98°C followed by 15 cycles of 15 s at 98°C, 30 s at 63°C, 30 s at 72°C and a final 5 min extension at 72°C . PCR reactions were cleaned with Agencourt AMPure XP beads (Beckman Coulter), run on a 2% agarose gel and a smear of 200-500bp was cut and gel purified using QIAquick Gel Extraction Kit (QIAGEN). Library concentration was determined with KAPA Library Quantification Kit for Illumina Platforms (Kapa Biosystems). Sequencing was performed on the Illumina Nextseq500 (75bp single-end reads).

treatment_protocol: Doxycyline was added for 16h to the media when indicated

value: DUX-overexpressing ESC

value: Doxycycline 16h

value: anti-RPA32/RPA2 Antibody (Abcam, Cat# ab10359, RRID:AB_297095)


title: RET7

status: Public on Jun 08 2021

type: genomic

city: Vejle

first: Luisa

last: Matos do Canto

address: Beriderbakken 4

department: Biochemistry and Immunology

country: Denmark

institute: Vejle Hospital

description: naive biopsies

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

scan_protocol: Arrays were imaged using the Illumina HiScan system with the standard recommended Illumina scanner setting

data_processing: data were processed using R language. Briefly, β values were normalized and batch effects were removed. Probes with low quality (detection p-value >0.05), mapped in X/Y chromosomes and cross-reactive were filtered out, as previously described by Pidsley R et al. (2016 - DOI: 10.1186/s13059-016-1066-1)

label: Cy5 and Cy3

molecule: genomic DNA

organism: Homo sapiens

source_name: rectal adenocarcinoma

label_protocol: Standard Illumina Protocol

extract_protocol: genomic DNA was extracted and purified from pre-treatment biopsies of rectal tissue using a phenol-chloroform-isoamyl alcohol (25:24:1) solution-based protocol.

tag: response

value: pIR


title: RET43

description: naive biopsies

value: pCR


title: RET44

description: naive biopsies


title: adipose 1week 5%CR, rep1

type: RNA

city: bunkyo-ku

first: Kenji

last: Saito

department: Department of Applied Biological Chemistry

country: Japan

institute: The Univeristy of Tokyo

description:
a-1w-5(1)
Rats fed reduced amount of diet.

hyb_protocol: Following fragmentation, 15 ug of cRNA were hybridized for 16 hr at 45C on GeneChip. GeneChips were washed and stained in the Affymetrix Fluidics Station 450.

scan_protocol: GeneChips were scanned using the Gene Array Scanner.

data_processing: The data were analyzed with Microarray Suite version 5.0 (MAS 5.0) using Affymetrix default analysis settings and global scaling as normalization method.

label: biotin

molecule: total RNA

organism: Rattus norvegicus

source_name: rat adipose tissue

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 10 ug total RNA (Expression Analysis Technical Manual, Affymetrix).

growth_protocol: 5-week-old rats were purchased from Japan SLC (Tokyo, Japan).

extract_protocol: Total RNA was prepared using RNeasy Plant Mini Kit (Qiagen) according to the manufacturer's instructions.

tag: strain

value: Wistar

tag: tissue

value: Adipose tissue

tag: gender

value: Male

tag: dietary restriction

value: 5% reduction

tag: time

value: 1 week


title: ESC_DUX_Clone1_DOX_16h_RPA_ChIP

type: SRA

country: USA

library_source: genomic

organism: Mus musculus

tag: cell type

tag: treatment

tag: chip antibody


title: ESC_DUX_Clone1_RPA_ChIP


title: ESC_DUX_Clone2_RPA_ChIP


title: ESC_DUX_Clone2_IgG_ChIP

value: anti-IgG (Antibodies-Online, Cat# ABIN101961,RRID:AB_10775589)


title: adipose 1week 20%CR, rep1

description:
a-1w-20(1)
Rats fed reduced amount of diet.

value: 20% reduction


title: RET9

description: naive biopsies


title: adipose 1week 10%CR, rep1

description:
a-1w-10(1)
Rats fed reduced amount of diet.

value: 10% reduction


title: adipose 1week 30%CR, rep1

description:
a-1w-30(1)
Rats fed reduced amount of diet.

value: 30% reduction


title: ESC_DUX_Clone1_IgG_ChIP


title: ESC_DUX_Clone1_ENDseq

data_processing:
Library strategy: ENDseq
Basecalling was performed using bcl2fastq version 2.20.0
END-seq reads were aligned to the mouse reference genome mm10 using bowtie (v1.1.2) with parameters -n 3 -l 50 -k 1.
Functions “view” and “sort” of samtools (v 1.6) were used to convert and sort the aligned sam files to sorted bam files
Bam files were further converted to bigwig files using bedtools bamToBed command, followed by UCSC toolkit function bedGraphToBigWig.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig files containing RPKM normalized read density, separated by strand.

growth_protocol: Wild-type (R1 and E14) ESC and ESCDux  were grown on a feeder layer of growth-arrested MEFs or on gelatin 0.1% in high-glucose DMEM (Invitrogen) supplemented with 15% FBS, 1:500 LIF (made in house), 0.1 mM nonessential amino acids, 1% glutamax, 1mM Sodium Pyruvate, 55 mM β-mercaptoethanol, and 1% penicillin/streptomycin (all from Life Technologies) at 37°C and 5% CO2. Cells were routinely passaged with Trypsin 0.05% (Gibco).

extract_protocol:
END-seq was performed as described. Briefly, for untreated DOX-treated ESCDUX, a total of 30 million cells in single cell suspension were embedded in a single agarose plug. Lysis and digestion of embedded cells was performed using Proteinase K (50C, 1 hour then 37C for 7 hours). Agarose plugs were rinsed in TE buffer and treated with RNase A at 3C, 1 hour. Next, DNA ends were blunted. For these reactions, DNA was retained in the plugs to prevent shearing. The first blunting reaction was performed using ExoVII (NEB, M0379S) for 1hr, 37C. Plugs were washed twice in NEB Buffer 4 (1X), immediately followed by the second blunting reaction using ExoT (NEB, M0265S) for 1 hour, 24°C. After this final blunting, two washes were performed in NEBNext dA606 Tailing Reaction Buffer (NEB, B6059S), followed by A-tailing (Klenow 3ʹ- > 5ʹ exo-, NEB, M0212S). After A-tailing, we performed a ligation with the “END-seq hairpin adaptor 1,” listed in reagents section, using NEB Quick Ligation Kit (NEB, M2200S).
Agarose plugs were then melted and dissolved. DNA was sonicated using to a median shear length of 170bp using a Covaris S220 sonicator for 4 min at 10% duty cycle, peak incident power 175, 200 cycles per burst, 4°C. Following the sonication, DNA was precipitated with ethanol and dissolved in 70 μl TE buffer. 35 μL of Dynabeads were washed twice with 1 mL Binding and Wash Buffer (1xBWB) (10 mM Tris-HCl pH8.0, 1 mM EDTA, 1 M NaCl, 0.1% Tween20). After the wash, beads were recovered using a DynaMag-2 magnetic separator (12321D, Invitrogen) and supernatants were discarded. Washed beads were resuspended in 130 μL 2xBWB (10 mM Tris- HCl pH8.0, 2 mM EDTA, 2 M NaCl) combined with the 130 μL of sonicated DNA followed by an incubation at 24C for 30 min. Next, the supernatant was removed, and the biotinylated DNA bound to the beads was washed thrice with 1 mL 1xBWB, twice with 1 mL EB buffer, once with 1 mL T4 ligase reaction buffer (NEB) and then resuspended in 50 μL of end-repair reaction mix (0.4 mM of dNTPs, 2.7 U of T4 DNA polymerase (NEB), 9 U of T4 Polynucleotide Kinase (NEB) and 1 U of Klenow fragment (NEB)) and incubated at 24C for 30 min. Once again, the supernatant was removed using a magnetic separator and beads were then washed once with 1 mL 1xBWB, twice with 1 mL EB buffer, once with 1 mL NEBNext dA-Tailing reaction buffer (NEB) and then resuspended in 50 μL of with NEBNext dA-Tailing reaction buffer (NEB) and 20 U of Klenow fragment exo- (NEB). The A-tailing reaction was incubated at 37C for 30 min. The supernatant  was removed using a magnetic separator and washed once with 1 mL NEBuffer 2 and resuspended in 115 mL of Ligation reaction with Quick Ligase buffer (NEB), 6,000 U of Quick Ligase (NEB) and ligated to “END-seq hairpin adaptor 2” by incubating the reaction at 25C for 30 min. Reaction was stopped by adding 50 mM of EDTA, and beads washed 3X BWB, 3X EB, andeluted in 8 μL of EB. Hairpin adaptors were digested using USER enzyme (NEB, M5505S) at 37C for 30 minutes. PCR amplification was performed in 50 μL reaction with 10 mM primers 5ʹCAAGCAGAAGACGGCATACGAGATXXXXXXGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC∗T-3ʹ and 5ʹAATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATC∗T-3ʹ, and 2X Kapa HiFi HotStart Ready mix (Kapa Biosciences). ∗ represents a phosphothioratebond and NNNNNN a Truseq index sequence. PCR program: 98C, 45 s; 15 cycles [98C, 15 s; 63C, 30 s; 72C, 30 s]; 72C, 5 min. PCR reactions were cleaned with AMPure XP beads, and after running the reactions on a 2% agarose gel, 200-500 bp fragments were isolated. Libraries were purified using QIA-quick Gel Extraction Kit (QIAGEN). Library concentration was determined with KAPALibrary Quantification Kit for Illumina Platforms (Kapa Biosystems) and the sequencing wasperformed on Illumina NextSeq 500 or 550 (75bp single end reads).

value: ESC

tag: cell line

value: ESCDux


title: ESC_DUX_Clone1_DOX_16h_ENDseq


title: RET34

description: naive biopsies


title: R1_RFPpos_CTCF

data_processing:
Library strategy: CUT&RUN
Basecalling was performed using bcl2fastq version 2.20.0
Data were processed using a modified version of Cut&RunTools (Zhu et al., 2019).  Reads were adapter trimmed using fastp v.0.20.0 (Chen et al., 2018).  An additional trimming step was performed to remove up to 6bp adapter from each read.
Next, reads were aligned to a hybrid mm10 and E. coli (MG1655) genome using bowtie2 (Langmead et al., 2012) with the ‘dovetail’ and ‘sensitive’ settings enabled.  Aligned fragments were segregated into mouse and carry-over (E. coli) subsets.  macs2 (Zhang et al., 2008) was used to call peaks with q-value cutoff < 0.01.
Normalized reads per kilobase per million (RPKM) signal tracks were generated using the ‘bamCoverage’ utility from deepTools  (Ramirez et al., 2016) with parameters ‘center_reads’ and ‘extend_reads’ options enabled.  A bin-size of 25bp and smooth length of 75bp was used for Nanog, Otx2, and Sox2 data, and bin-size 75 and smooth-length 300bp for H3K27ac data.
Genome_build: mm10
Supplementary_files_format_and_content: bigwig files containing RPKM normalized read density.

source_name: mouse ESC_R1

extract_protocol:
The CUT&RUN protocol was slightly modified from (Skene et al., 2018; Meers et al., 2019). In brief, trypsinized or cell sorted ESC (between 150,000-500,000 cells depending on the experiment) were washed three times with Wash Buffer (20 mM HEPES-KOH pH 7.5, 150 mM NaCl, 0.5 mM spermidine, Roche complete Protease Inhibitor tablet EDTA free) and bound to activated Concanavalin A beads (Polysciences) for 10 minutes at room temperature. Cells were then permeabilized in Digitonin Buffer (0.05 % Digitonin and 0.1% BSA in Wash Buffer) and incubated with the antibody against CTCF (07-729, Millipore) at 4°C for 2 hours. For negative controls, Guinea Pig anti-Rabbit IgG (ABIN101961, Antibodies-online) was used. Cells were washed with Digitonin Buffer following antibody incubation, and further incubated with purified hybrid protein A-protein G-Micrococcal nuclease (pAG-MNase) at 4°C for 1 hour. Samples were washed in Digitonin Buffer, resuspended in 150 μl Digitonin Buffer and equilibrated to 0°C on ice water for 5 minutes. To initiate MNase cleavage, 3 μl 100 mM CaCl2 was added to cells and after 1 hour of digestion, reactions were stopped with the addition of 150 μl 2x Stop Buffer (340 mM NaCl, 20 mM EDTA, 4 mM EGTA, 0.02 % Digitonin, 50 μg/ml RNase A, 50 μg/ml Glycogen). Samples were incubated at 37°C for 10 minutes to release DNA fragments and centrifuged at 16,000 g for 5 minutes. Supernatants were collected and a mix of 1.5 μl 20% SDS / 2.25 μl 20 mg/ml Proteinase K was added to each sample and incubated at 65°C for 35 minutes. DNA was precipitated with ethanol and sodium acetate and pelleted by high-speed centrifugation at 4°C, washed, air-dried and resuspended in 10 μ 0.1x TE.
The entire precipitated DNA obtained from CUT&RUN was used to prepare Illumina compatible sequencing libraries. In brief, end-repair was performed in 50 μl of T4 ligase reaction buffer, 0.4 mM dNTPs, 3 U of T4 DNA polymerase (NEB), 9 U of T4 Polynucleotide Kinase (NEB) and 1 U of Klenow fragment (NEB) at 20°C for 30 minutes. End-repair reaction was cleaned using AMPure XP beads (Beckman Coulter) and eluted in 16.5 μl of Elution Buffer (10 mM Tris-HCl pH 8.5) followed by A-tailing reaction in 20 μl of dA-Tailing reaction buffer (NEB) with 2.5 U of Klenow fragment exo- (NEB) at 37°C for 30 minutes. The 20 μl of the A-tailing reaction were mixed with Quick Ligase buffer 2X (NEB), 3000 U of Quick Ligase (NEB) and 10 nM of annealed adaptor (Illumina truncated adaptor) in a volume of 50 μl and incubated at room temperature for 20 min. The adaptor was prepared by annealing the following HPLC-purified oligos: 5′-Phos/GATCGGAAGAGCACACGTCT-3′ and 5′-ACACTCTTTCCCTACACGACGCTCTTCCGATC∗T-3′ (∗phosphorothioate bond). Ligation was stopped by adding 50 mM of EDTA, cleaned with AMPure XP beads and eluted in 14 μl of Elution Buffer. All volume was used for PCR amplification in a 50 μl reaction with 1 μM primers TruSeq barcoded primer p7, 5′-CAAGCAGAAGACGGCATACGAGATXXXXXXXXGTGACTGGAGTTCAGACGTGTGCTCTTCCGATC∗T-3′ and TruSeq barcoded primer p5 5′-AATGATACGGCGACCACCGAGATCTACACXXXXXXXXACACTCTTTCCCTACACGACGCTCTTCCGATC*T-3′ (∗ represents a phosphothiorate bond and XXXXXXXX a barcode index sequence), and 2X Kapa HiFi HotStart Ready mix (Kapa Biosciences). The temperature settings during the PCR amplification were 45 s at 98°C followed by 15 cycles of 15 s at 98°C, 30 s at 63°C, 30 s at 72°C and a final 5 min extension at 72°C. PCR reactions were cleaned with AMPure XP beads (Beckman Coulter), run on a 2% agarose gel and a band of 300bp approximately was cut and gel purified using QIAquick Gel Extraction Kit (QIAGEN). Library concentration was determined with KAPA Library Quantification Kit for Illumina Platforms (Kapa Biosystems). Sequencing was performed on the Illumina NextSeq550 (75bp pair-end reads).

treatment_protocol: Doxycyline was added for 16h or 24h to the media when indicated

tag: genotype

value: wild type

tag: antibody

value: anti-CTCF Antibody (Millipore, Cat# 07-729, RRID:AB_441965)


title: R1_RFPneg_CTCF


title: RET41

description: naive biopsies


title: ESC_DUX_Clone 2_CTCF


title: E14_RFPneg_CTCF

source_name: mouse ESC_E14


title: E14_IgG


title: E14_RFPpos_CTCF


title: ESC_DUX_Clone 1_IgG


title: ESC_DUX_Clone 2_ DOX_24h_RFPpos_CTCF

value: Doxycycline 24h


title: ESC_DUX_Clone 1_ DOX_24h_RFPpos_CTCF


title: ESC_DUX_Clone 2_ DOX_16h_RFPpos_CTCF

value: Doxycycline 16h


title: ESC_DUX_Clone 2_IgG


title: ESC_DUX_Clone 2_ DOX_24h_RFPneg_CTCF


title: ESC_DUX_Clone2_ENDseq


title: ESC_DUX_Clone 1_ DOX_24h_RFPneg_CTCF


title: ESC_DUX_Clone 1_CTCF


title: ESC_DUX_Clone 1_ DOX_16h_RFPneg_CTCF


title: ESC_DUX_Clone 1_ DOX_16h_RFPpos_CTCF


title: RET33

description: naive biopsies


title: RET29

description: naive biopsies


title: RET28

description: naive biopsies


title: P111_G1.7_C3_NoRash_NoRx_R2

city: Baltimore

first: Theresa

middle: K

last: Hodges

address: 801 W. Baltimore Street

department: Institute for Genome Sciences

institute: University of Maryland Baltimore

description: Gene expression data from fibroblasts not treated

hyb_protocol: Following fragmentation, biotinylated cRNA were hybridized for 16 hr on GeneChip® Human Genome U133 Plus 2.0 array.

scan_protocol: The arrays were washed and stained in the Affymetrix fluidics station and scanned in a 3000 7G scanner.

data_processing: Probe level analysis was performed on the Affymetrix raw data (.cel files) with the Robust Multi-Array Average (RMA) method implemented in the Bioconductor package 'affy' to obtain a log2 expression value for each gene probe set.

source_name: Human, adult, dermal fibroblasts from skin biopsy sample

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

growth_protocol: Human, adult, fibroblasts collected from skin biopsy samples were seeded in cell culture flasks and grown under standard culture conditions.

extract_protocol: Total RNA was extracted on the fourth day of the experiment using a standard TRIzol®-based extraction method.

treatment_protocol: Cells were transferred to 35mm cell culture dishes, grown to confluence, and then treated once daily for three days with erlotinib 1μM DMSO, or no treatment.

tag: age

value: adult

value: dermal fibroblasts


title: RET42

description: naive biopsies

value: pCR


title: RET11

description: naive biopsies


title: ESC_DUX_Clone2_DOX_16h_ENDseq


title: P111_G1.5_C3_Rash_NoRx_R2


title: P111_G1.24_C3_Rash_Veh_R4

description: Gene expression data from fibroblasts treated with DMSO 1μM

value: DMSO


title: P111_G1.27_C3_Rash_E1uM_R1

description: Gene expression data from fibroblasts treated with erlotinib 1μM daily for three days

value: erlotinib


title: P111_G1.28_C3_NoRash_E1uM_R2


title: RET13

description: naive biopsies


title: P111_G1.36_C3_Rash_E1uM_R4


title: P111_G1.4_C3_NoRash_NoRx_R1


title: P111_G1.19_C3_Rash_Veh_R3


title: P111_G1.22_C3_NoRash_Veh_R4


title: P111_G1.21_C3_NoRash_Veh_R3


title: P111_G1.29_C3_Rash_E1uM_R2


title: P111_G1.18_C3_NoRash_Veh_R2


title: P111_G1.14_C3_Rash_Veh_R1


title: RET10

description: naive biopsies


title: P111_G1.2_C3_Rash_NoRx_R1


title: P111_G1.16_C3_Rash_Veh_R2


title: P111_G1.12_C3_Rash_NoRx_R4


title: P111_G1.34_C3_NoRash_E1uM_R3


title: P111_G1.13_C3_NoRash_Veh_R1


title: P111_G1.26_C3_NoRash_E1uM_R1


title: P111_G1.11_C3_NoRash_NoRx_R4


title: P111_G1.10_C3_NoRash_NoRx_R3


title: P104_G2.8_C3_Rash_Veh_R4


title: R1_IgG


title: adipose 1week control, rep1

description:
a-1w-C(1)
Rats fed control diet.

value: Wistar

value: Adipose tissue

value: Male


title: AhR-KO-5_MPP

city: Rochester

first: Anthony

last: Franchini

state: NY

address: 601 Elmwood Ave

institute: University of Rochester

library_source: transcriptomic

data_processing:
DATA FORMATTING (Illumina -> FastQ) & DEMULTIPLEXING - bcltofastq-1.8.4
CLEANING - Trimmomatic-0.36 "SLIDINGWINDOW:4:20 TRAILING:13 LEADING:13 ILLUMINACLIP:adapters.fasta:2:30:10 MINLEN:15"
MAPPING - STAR_2.5.2b "--twopassMode Basic --runMode alignReads --genomeDir ${GENOME} --readFilesIn ${SAMPLE} --outSAMtype BAM SortedByCoordinate --outSAMstrandField intronMotif --outFilterIntronMotifs RemoveNoncanonical"
READ COUNTING - subread-1.5.0p3, featurecounts, "-s 0 -t exon -g gene_name"
DIFFERENTIAL EXP. - DESeq2_1.12.4 Multi-Mapping -s 0 -M -t exon -g gene_name
Genome_build: GRCm38.p5 + Gencode-M12 Annotation
Supplementary_files_format_and_content: Raw read counts for each sample in tab-delimited text file

growth_protocol: Eight to twelve week old C57BL/6 or AHR KO mice were used in this study

extract_protocol:
Murine HSCs and MPPs were FACS sorted from mice under homeostatic conditions.
The total RNA concentration from sorted mouse DCs was determined with the NanopDrop 1000 spectrophotometer (NanoDrop, Wilmington, DE) and RNA quality assessed with an Agilent Bioanalyzer 2100 (Agilent, Santa Clara, CA). There were three independent DC samples for each treatment group. The TruSeq RNA Sample Preparation Kit V2 (Illumina, San Diego, CA) was used for next generation sequencing library construction per manufacturer’s protocols. Briefly, mRNA was purified from 100ng total RNA with oligo-dT magnetic beads and fragmented. First-strand cDNA synthesis was performed with random hexamer priming followed by second-strand cDNA synthesis. End repair and 3` adenylation was then performed on the double stranded cDNA. Illumina adaptors were ligated to both ends of the cDNA, purified by gel electrophoresis and amplified with PCR primers specific to the adaptor sequences to generate amplicons of approximately 200-500bp in size.

treatment_protocol: Mice were not treated and bone marrow cells were isolated under homeostatic conditions.

value: Eight to twelve weeks


title: P104_G2.6_C3_Rash_Veh_R2


title: P106_G2.56_C4_NoRash_E1uM_R4


title: P106_G2.54_C4_NoRash_E1uM_R2


title: P106_G2.55_C4_NoRash_E1uM_R3


title: AhR-KO-3_MPP


title: AhR-KO-4_MPP


title: ESC_DUX_Clone 2_ DOX_16h_RFPneg_CTCF


title: AhR-KO-2_MPP


title: P106_G2.51_C4_NoRash_Veh_R3


title: AhR-KO-1_MPP


title: P106_G2.53_C4_NoRash_E1uM_R1


title: WT-5_MPP


title: WT-4_MPP


title: P106_G2.48_C4_NoRash_NoRx_R4


title: WT-2_MPP


title: WT-3_MPP


title: P106_G2.46_C4_NoRash_NoRx_R2


title: P106_G2.52_C4_NoRash_Veh_R4


title: AhR-KO-3_LT-HSC


title: AhR-KO-7_LT-HSC


title: P107_G2.24_C2_NoRash_E1uM_R4


title: P106_G2.49_C4_NoRash_Veh_R1


title: P107_G2.22_C2_NoRash_E1uM_R2


title: P107_G2.21_C2_NoRash_E1uM_R1


title: P104_G2.5_C3_Rash_Veh_R1


title: P104_G2.4_C3_Rash_NoRx_R4


title: P107_G2.23_C2_NoRash_E1uM_R3


title: AhR-KO-1_LT-HSC


title: P104_G2.2_C3_Rash_E1uM_R2


title: P106_G2.47_C4_NoRash_NoRx_R3


title: P106_G2.50_C4_NoRash_Veh_R2


title: P107_G2.18_C2_NoRash_Veh_R2


title: P107_G2.15_C2_NoRash_NoRx_R3


title: P107_G2.17_C2_NoRash_Veh_R1


title: P104_G2.3_C3_Rash_NoRx_R3


title: WT-1_MPP


title: P107_G2.20_C2_NoRash_NoRx_R4


title: AhR-KO-4_LT-HSC


title: P107_G2.16_C2_NoRash_Veh_R4


title: P107_G2.14_C2_NoRash_NoRx_R2


title: WT-8_LT-HSC


title: WT-6_LT-HSC


title: AhR-KO-2_LT-HSC


title: ARPE-19_RNA-seq

city: Shanghai

first: Jianing

last: Wang

address: Dongchuan Road

country: China

institute: Shanghai JiaoTong University

data_processing:
RNA-seq reads were trimmed using Trim Galore, reads containing adapter were discarded, and HISAT2 software was used for mapping to hg38 genome. Then Stringtie (version 1.3.3) was used to assemble gene expression from aligned reads to quantify the number of reads mapped to each gene. The expression level of genes was assessed by Fragments per kilobase of exon per million reads mapped (FPKM).
Genome_build: hg38 (GRCh38)
Supplementary_files_format_and_content: ARPE-19_hg38_gene_count_matrix.txt: Tab-delimited text file includes FPKM values for each gene.

source_name: ARPE-19 cell line

growth_protocol: The cells were cultured with DMEM/F-12 medium (Biological Industries, Israel) containing 20% fetal bovine serum (Gibco BRL/Invitrogen, Carlsbad, CA, USA), 100μg/ml streptomycin, and 100IU/ml penicillin, and incubated with 37℃ and 5% CO2.

extract_protocol:
Total RNA was extracted from ARPE-19 cell samples by using TRIzol reagents (Invitrogen, Waltham, Massachusetts, USA) as recommended by the manufacturer.
One microgram of total RNA was used to construct the RNA-seq libraries by KAPA Stranded RNA-Seq Kit following the manufacturer’s protocol. The quality of libraries was assessed with Agilent 2100 Bioanalyzer (Agilent, United States).

value: Epithelial Cells

value: Normal culture

tag: cell cycle

value: Interphase


title: P104_G2.12_C3_Rash_E1uM_R4


title: WT-5_LT-HSC


title: WT-4_LT-HSC


title: P104_G2.11_C3_Rash_NoRx_R3


title: LNCaP RNASeq

city: Bern

first: Kellie

last: Cotter

state: Bern

address: Murtenstrasse 35

department: Dept for BioMedical Research

country: Switzerland

institute: University of Bern

data_processing:
miCLIP data was anlalyzed for crosslinking-induced mutation sites as described previously (Linder et al., Nat Methods 2015).
RNA-Seq reads were aligned to the human reference genome hg19 by STAR.
Count tables were generated with DESeq2
Genome_build: hg19
Supplementary_files_format_and_content: Bedgraph files of unique CIMS miCLIP tags. Score represents the number of reads covering that position.
Supplementary_files_format_and_content: Bed files of m6A sites detected in CIMS miCLIP
Supplementary_files_format_and_content: Matrix table containing normalized counts data from RNA-Seq

molecule: polyA RNA

source_name: LNCaP cell line

growth_protocol: LNCaP cells were maintained in RPMI medium, supplemented with 10% FBS, and 1% penicillin-streptomycin on poly-L-lysine coated plates. RWPE cells were maintained in Keratinocyte Serum Free Medium supplemented with bovine pituitary extract and human recombinant EGF), and 1% penicillin-streptomycin.

extract_protocol:
Total RNA from LNCaP and RWPE cells was isolated with TRIzol according to the manufacturer instructions. PolyA+ mRNA was isolated using the PolyATtract mRNA Isolation System from Promega.
Libraries were prepared using 1 ug of polyA+ RNA as input using the Illumina protocol.

value: LNCaP


title: UE, meta [miPC060]

city: Seoul

first: Sun

last: Shin

address: Banpodaero 222

department: Dept. of Microbiology

country: South Korea

institute: The Catholic University of Korea

description: Test

data_processing:
Matrix non-normalized' raw data were processed using the QuantStudio Real-Time PCR Software (Thermo Fisher Scientific) to determine a cycle threshold (Ct) value for each miRNA. Quantile-normalized Ct 'Matrix normalized' data and fold-change from the resultant ΔCt against U6 snRNA between the localized and metastatic cancer across all TLDA data using the HTqPCR R package.
Matrix normalized worksheet reports quantile-normalized signal. After getting delta Ct against U6, 'Fold Change' worksheet reports test/control ratios.

label: FAM

source_name: Exosome extracted from urine

label_protocol: Megaplex reverse transcription reactions and pre-amplification reactions were performed to increase the quantity of cDNA for miRNA expression analysis using the Megaplex PreAmp Primers Human Pool A and TaqMan PreAmp Master Mix (Thermo Fisher Scientific).

extract_protocol: Exosomes were isolated from 5 mL of urine using the ExoQuick-TC exosome precipitation solution (System biosciences), and Total RNA was isolated from the pellets using TRIzol.

tag: disease

value: metastatic PCa


title: RWPE RNASeq

source_name: RWPE cell line

value: RWPE


title: UE, meta [miPC059]

description: Test


title: UE, meta [miPC058]

description: Test


title: RWPE miCLIP

data_processing:
Library strategy: miCLIP
miCLIP data was anlalyzed for crosslinking-induced mutation sites as described previously (Linder et al., Nat Methods 2015).
RNA-Seq reads were aligned to the human reference genome hg19 by STAR.
Count tables were generated with DESeq2
Genome_build: hg19
Supplementary_files_format_and_content: Bedgraph files of unique CIMS miCLIP tags. Score represents the number of reads covering that position.
Supplementary_files_format_and_content: Bed files of m6A sites detected in CIMS miCLIP
Supplementary_files_format_and_content: Matrix table containing normalized counts data from RNA-Seq

extract_protocol:
Total RNA from LNCaP and RWPE cells was isolated with TRIzol according to the manufacturer instructions. PolyA+ mRNA was isolated using the PolyATtract mRNA Isolation System from Promega.
Libraries were prepared using 1 ug of polyA+ RNA as input using the Illumina protocol.
miCLIP: 20 ug of mRNA was used as input for miCLIP following the previously reported protocol (Linder et al., Nat Methods 2015)


title: LNCaP miCLIP


title: UE, meta [miPC056]

description: Test


title: UE, meta [miPC057]

description: Test


title: treat_In_3k27ac

city: Singapore

first: Gokce

last: OGUZ

address: 60 Biopolis St, GENOME, #02-01

department: Bioinformatics Consulting and Training Platform

country: Singapore

institute: Genome Institute of Singapore

data_processing:
Raw data analyzed by nf-core-ChIPseq version 1.1.0, a nextflow(version 19.01.0) based pipeline
The paired-end reads were mapped to hg38 using bwa-mem 0.7.17-r1188
BEDtools(v2.27.1), Bamtools(v2.5.1) and SAMtools(v1.9) were used to detect and remove the aligned reads that are marked as duplicates, mapping to the blacklisted regions, multiple locations, different chromosomes, containing more than 4 mismatches, insert size bigger than 2kb.
Normalized bigwigs are generated by scaling to 1 million mapped reads via BEDtools(v2.27.1) and UCSC.
Narrow peaks called by MACS2(version 2.1.2) using input sequences as a background control.
Consensus peaksets generated by BEDtools(v2.27.1) and annotated to gene features using HOMER(v4.11).
Genome_build: hg38
Supplementary_files_format_and_content: peak text files &bigWig files

source_name: Patient Derived Cancer Cells

growth_protocol: patient-derived cancer cells (PDC) were cultured in DMEM medium (Gibco) supplemented with 0.5%FBS, NEAA and insulin.

extract_protocol:
A million cells (per sample) were cross-linked using 1% formaldehyde for 10min and reactions were stopped by adding 0.125 M glycine for 5 min at room temperature. Cell pellets were washed with cold TBSE buffer (20 mM Tris-HCl pH7.5, 150 mM NaCl, 1 mM EDTA) for 3 times. Then cells were lysed in SDS lysis buffer (50 mM Tris-HCl pH8.0, 10 mM EDTA, 1% SDS) for 5 min on ice. Sonication conditions were optimized for tumor cells using Branson Digital Sonifier to achieve a shear length of 250–500 bp. Solubilized chromatin was diluted 10 times in dilution buffer (10 mM Tris-HCl pH7.4, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.01% SDS) and precleared with Dynabead (Life Technologies) for 4 hr at 4°C. Precleared chromatin was then incubated with appropriate antibodies overnight at 4°C. The complexes of chromatin and antibodies were captured with Dynabead at 4 °C for 2hr. Immune complexes were then washed thrice with dilution buffer and once with TE buffer (10 mM Tris-HCl pH7.5, 1 mM EDTA), sequentially. Elution and reverse-crosslinking were performed in elution buffer (50 mM Tris-HCl pH7.5, 10 mM EDTA, 1% SDS) at 68°C with vortex overnight followed by digestion with proteinase K (20 mg/ml) for 2hr at 42℃. Eluted DNA was purified using Phenol-CHCl3-Isoamylalchol and then precipitated with ethanol with existence of NaCl and yeast tRNA. DNA was re-suspended in water with RNase A and incubated in 37 °C for 10 min.
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Illumina HiSeq 2500 following the manufacturer's protocols.

treatment_protocol: PDC cells were  pre-cultured in DMEM medium (Gibco) supplemented with 0.5%FBS, NEAA and insulin overnight, then treated with 50ng/ml rIL6/8 for another 8hr.

value: treated with IL6/8

value: H3K27Ac (abcam, cat #:ab4729)

value: colorectal cancer


title: ctrl_In_3k27ac

value: untreated


title: ctrl_In_HM


title: treat_In_HM


title: UE, meta [miPC055]

description: Test


title: UE, meta [miPC054]

description: Test


title: treat_PS

value: P-STAT3 (Cell Signaling, cat #:9145)


title: UE, meta [miPC053]

description: Test


title: XR-Seq A2780

city: Jerusalem

first: Sheera

last: Adar

state: Israel

address: Ein Kerem

department: Department of Microbiology and Molecular Genetics

country: Israel

institute: Institute for Medical Research Israel Canada, Faculty of Medicine, Hebrew University of Jerusalem

description: Excised oligomer released in Nucleotide Excision Repair (XR-seq)

data_processing:
Library strategy: XR-Seq
Base calling using Casava v1.8.2. Demultiplexing performed allowing 1 mismtach in the index
Flanking adapter sequences were removed from the reads using trimmomatic version 0.36 with the command options: ILLUMINACLIP:adapter_sequence.txt:2:30:10 (Bolger et al. 2014).
Filter 50 nt length reads
Reads were aligned to the hg38 human genome using bowtie 1.0.0 (Langmead et al. 2009) with the command options: -q --nomaqround --phred33-quals -p 32 -m 4 -n 2 -e 70 -l 20 --chunkmbs 800 --best –S
reads mapped to chromosome Y or mitochondrial chromosome were filtered
PCR duplicates were removed using PicardCommandLine MarkDuplicates version 2.23.4 (http://broadinstitute.github.io/ picard/)
All bam files of replicates from condition were merged to create a pooled bam file
Per base read counts were calculated using bedtools genomecov and normalized per billion mapped reads per chromosome. Read counts were calculated in 25 nt windows using awk  and converted to bigwig using the bedGraphToBigWig command to create stranded signal maps in bigwig file format
Genome build: hg38
bigwig files of the data normalized to mapped reads per chromosome counts and separated by strands.

source_name: ovarian carcinoma

growth_protocol: A2780 and Acis cells (Sigma, 93112519 and 93112517) were grown in RPMI 1640 Medium supplemented with 10% FBS, 2 mM Glutamine, 1 mM Sodium Pyruvate, 100 units/ml Penicillin, 0.25 mg/ml Streptomycin. Every 3 passages Acis cells were treated with 1 µM cisplatin (Pharmachemie BV, Teva group, Haarlem, The Netherlands, 031 30 25429 05).

extract_protocol:
cells were harvested, collected by centrifugation and resuspended in lysis buffer and lysed using a Dounce homogenizer.
XR-seq was performed as previously described (Hu. et al Genes and Dev. 2016, Adar S. et al PNAS 2016). Briefly, primary excision products were pulled-down by TFIIH co-immunoprecipitation using anti-p62 antibody (Santa-Cruz Biotechnology, sc-293) and anti-p89 antibody (Santa-Cruz Biotechnology, sc-292). Adapters were ligated on both ends of the excision oligomer and immunoprecipitated using cisplatin-specific antibody (abcam, 103261). Cisplatin-induced damages were reversed in-vitro by incubating the samples with 200 mM NaCN at 65°C overnight. DNA was PCR amplified and purified by native polyacrylamide gel electrophoresis (PAGE) to create the library.

treatment_protocol: Cells were grown to ~80% confluence in 60 mm (for RNA- and ATAC-Seq) or 150 mm (for XR-Seq) dishes, culture medium was removed, cells were washed once in PBS. Then, cells were incubated with media containing 200µM cisplatin for 3h.

value: ovarian carcinoma

tag: cisplatin treatment

tag: treatment duration


title: treat_PB

value: P-BRD4 (homemade polyclonal antibody from iDNA)


title: XR-Seq A2780cis


title: treat_B

value: BRD4 (Cell Signaling, cat #:13440)


title: WT-1_LT-HSC


title: UE, meta [miPC052]

description: Test


title: UE, meta [miPC051]

description: Test


title: UE, meta [miPC049]

description: Test


title: treat_In_TF


title: UE, meta [miPC048]

description: Test


title: ctrl_PB


title: UE, meta [miPC047]

description: Test


title: ctrl_B


title: UE, meta [miPC046]

description: Test


title: UE, meta [miPC045]

description: Test


title: exRNA_fraction_4_10

city: Tampa

first: Jing

last: Jia

state: Florida

address: 12902 Magnolia Drive

department: Tumor biology

institute: Moffitt cancer center

data_processing:
FastQC (v0.11.0) was used for quality check
Cutadapt (v2.3.1) was used to trim adaptor sequences.
We collected fasta files for a variety of RNAs: The miRNA databases release 22.1 were downloaded from miRBase; piRNAs from piRNA Quest, tRNAs from gtRNAdb and circRNAs from cirBase. The lincRNAs, snoRNAs and snRNAs came from the GRCh38 ncRNA release 96. For protein coding cDNAs and other cDNAs detection, reads were mapped to the Gencode version 24. rRNAs mapping was based on both of the GRCh38 ncRNA and 45S, 5S and mt-rRNA sequences. DNASTAR lasergene (v15.3) was used for mapping
Data normalization was performed with DESeq2 package (v1.26.0)
Genome_build: hg38
Supplementary_files_format_and_content: excel files include normalized data for each sample

extract_protocol:
RNA was extracted using miRNeasy Micro Kit (Cat. 217084)
RNA libraries were prepared for sequencing using CATS RNA-seq Kit (Cat. C05010049)

tag: cell type source

value: plasma

tag: fraction

value: supernatant

value: Yes


title: exRNA_fraction_4_09


title: ctrl_In_TF

tag: tissue


title: exRNA_fraction_4_08


title: UE, meta [miPC044]

description: Test


title: UE, meta [miPC043]

description: Test


title: UE, meta [miPC042]

description: Test


title: exRNA_fraction_4_06


title: UE, meta [miPC041]

description: Test


title: exRNA_fraction_4_05


title: exRNA_fraction_4_07


title: UE, meta [miPC040]

description: Test


title: UE, meta [miPC039]

description: Test


title: exRNA_fraction_4_04


title: UE, meta [miPC038]

description: Test


title: exRNA_fraction_4_03

value: No


title: exRNA_fraction_4_02


title: UE, meta [miPC050]

description: Test


title: ctrl_PS


title: exRNA_fraction_4_01


title: UE, localized [miPC035]

description: Control

value: localized PCa


title: UE, localized [miPC036]

description: Control


title: UE, localized [miPC034]

description: Control


title: UE, localized [miPC031]

description: Control


title: exRNA_fraction_3_07

value: extracellular vesicles


title: exRNA_fraction_3_08


title: exRNA_fraction_3_10


title: UE, localized [miPC032]

description: Control


title: exRNA_fraction_3_09


title: UE, localized [miPC030]

description: Control


title: UE, localized [miPC029]

description: Control


title: exRNA_fraction_3_04


title: exRNA_fraction_3_03


title: exRNA_fraction_3_05


title: exRNA_fraction_3_02


title: exRNA_fraction_3_01


title: UE, localized [miPC027]

description: Control


title: UE, localized [miPC026]

description: Control


title: exRNA_fraction_2_10

value: fibrins


title: exRNA_fraction_2_08


title: exRNA_fraction_2_07


title: UE, localized [miPC024]

description: Control


title: UE, localized [miPC021]

description: Control


title: exRNA_fraction_2_06


title: UE, localized [miPC022]

description: Control


title: UE, localized [miPC020]

description: Control


title: UE, localized [miPC023]

description: Control


title: exRNA_fraction_2_09


title: exRNA_fraction_2_05


title: UE, localized [miPC033]

description: Control


title: exRNA_fraction_3_06


title: exRNA_fraction_2_04


title: UE, localized [miPC017]

description: Control


title: UE, localized [miPC019]

description: Control


title: exRNA_fraction_2_01


title: exRNA_fraction_2_03


title: UE, localized [miPC025]

description: Control


title: exRNA_fraction_2_02


title: ATAC-Seq A2780cis cisplatin

data_processing:
Adapter sequence was trimmed from each read using Cutadapt version 1.15 with the command options: -m 10
Reads were aligned to the genome using Bowtie2 version 2.4.1 with the command options: --very-sensitive  -k 10
reads mapped to chromosome Y or mitochondrial chromosome were filtered
PCR duplicates were removed using PicardCommandLine MarkDuplicates version 2.23.4 (http://broadinstitute.github.io/ picard/)
All bam files of replicates from condition were merged to create a pooled bam file
Peaks were called separately for each replicate and pooled bam files using macs2 callpeak command version 2.2.7.1
Peak files from replicates of the same condition were merged with the pooled bam peak file using bedtools intersect to create one set of peaks per condition.
Per base read counts were calculated using bedtools genomecov and normalized per billion mapped reads per chromosome. Read counts were converted to bigwig using the bedGraphToBigWig command to create signal maps in bigwig file format
Genome build: hg38
bigwig files of the data normalized to mapped reads per chromosome counts.
peak text files

extract_protocol:
cells were washed with PBS and incubated in the 37°C incubator with media containing 200 units/mL DNase (Worthington, LS002006) in 1.2 mM MgCl2 and 0.5 mM CaCl2 for 30 min
ATAC-seq was performed using an ATAC-seq kit (Active motif, 53150) according to manufacturer’s protocol. 100,000 cells were taken per reaction. ATAC-seq libraries were purified and size selected with HighPrep PCR beads (MagBio, AC-60005)


title: exRNA_fraction_1_09

value: platelets cell debris


title: RNA-Seq A2780cis cisplatin rep3

data_processing:
Reads were aligned to the genome using STAR version 2.7.3
PCR duplicates were removed using PicardCommandLine MarkDuplicates version 2.23.4 (http://broadinstitute.github.io/picard)
Read counts per gene were calculated using htseq-count v0.11.1 with the command option: --stranded=reverse
DESeq2 was applied using R version 4.0.0
All bam files of replicates from condition were merged to create a pooled bam file
Per base read counts were calculated using bedtools genomecov and normalized per billion mapped reads per chromosome. Read counts were calculated in 75 nt windows using awk  and converted to bigwig using the bedGraphToBigWig command to create stranded signal maps in bigwig file format
Genome build: hg38
bigwig files of the data normalized to mapped reads per chromosome counts and separated by strands.
Matrix table with raw gene counts for every gene and every sample

extract_protocol:
Total RNA was extracted using GENEzol™ TriRNA Pure with DNaseI Kit (Geneaid, GZXD).
Ribo-depleted RNA-Seq libraries were prepared using KAPA Stranded RNA-Seq Kit with RiboErase HMR (Kapa, KK8483) according to the manufacturer’s protocol, starting with 1 μg total RNA, and using Illumina TrueSeq adapters.


title: ATAC-Seq A2780 cisplatin


title: exRNA_fraction_1_07


title: ATAC-Seq A2780 control

value: control


title: RNA-Seq A2780cis cisplatin rep2


title: exRNA_fraction_1_08


title: exRNA_fraction_1_06


title: exRNA_fraction_1_05


title: RNA-Seq A2780cis cisplatin rep1


title: RNA-Seq A2780 cisplatin rep2


title: exRNA_fraction_1_04


title: exRNA_fraction_1_03


title: RNA-Seq A2780 cisplatin rep3


title: exRNA_fraction_1_02


title: UE, localized [miPC018]

description: Control


title: exRNA_fraction_1_01


title: RNA-Seq A2780cis control rep3


title: RNA-Seq A2780cis control rep2


title: exRNA_fraction_1_10


title: ATAC-Seq A2780cis control


title: RNA-Seq A2780cis control rep1


title: RNA-Seq A2780 control rep3


title: total RNAseq, PAS-T4G, replicate 3

city: Basel

first: Fabio

last: Mohn

address: Maulbeerstrasse 66

institute: Friedrich Miescher Institute for Biomedical Research

data_processing:
Raw data were demultiplexed and converted to fastq format using bcl2fastq2 (v1.17) Illumina. Fastq files were aligned using STAR (v2.7.3a) with the parameters --runMode alignReads --outFilterType BySJout --outFilterMultimapNmax 100 --outFilterMismatchNoverLmax 0.05 --outSAMmultNmax 1 --outMultimapperOrder Random --outSAMtype BAM SortedByCoordinate --outSAMattributes NH HI NM MD AS nM --outSAMunmapped Within. STAR sam output was converted to bam using samtools (v1.9). Bam files were used to calculate strand-specific coverage using bedtools genomeCoverageBed (v2.27.1), resulting bedgraph files were normalized to 1 million genome mapping reads (RPM), and converted to bigWig format using bedGraphToBigWig (v4).
smallRNA-seq:Raw data were demultiplexed and converted to fastq format using bcl2fastq2 (v1.17) Illumina. Qiagen 3' adaptors were trimmed using cutadapt (v1.18) with the parameters --discard-untrimmed -m 18. Fastq files were aligned using Bowtie (v1.2.2) with parameters -f -M 10000 -v 0 -S --best --strata. Bowtie sam output was converted to bam using samtools (v1.9). Bam files were converted to bed using bedtools bamtobed (v2.27.1), bed files were used to calculate strand-specific coverage using bedtools genomeCoverageBed (v2.27.1), resulting bedgraph files were normalized to 1 million genome mapping reads (RPM), and converted to bigWig format using bedGraphToBigWig (v4).
RNA-seq: Raw data were demultiplexed and converted to fastq format using bcl2fastq2 (v1.17) Illumina. Fastq files were aligned using STAR (v2.7.3a) with the parameters --runMode alignReads --outFilterType BySJout --outFilterMultimapNmax 100 --outFilterMismatchNoverLmax 0.05 --outSAMmultNmax 1 --outMultimapperOrder Random --outSAMtype BAM SortedByCoordinate --outSAMattributes NH HI NM MD AS nM --outSAMunmapped Within. STAR sam output was converted to bam using samtools (v1.9). Bam files were used to calculate strand-specific coverage using bedtools genomeCoverageBed (v2.27.1), resulting bedgraph files were normalized to 1 million genome mapping reads (RPM), and converted to bigWig format using bedGraphToBigWig (v4).
Genome_build: Spombe.ASM294v2.24
Supplementary_files_format_and_content: bigWig files containing normalized (to 1 mio genome mapping reads per library) genome mapping read counts for visualization in UCSC or IGV. Counts per million (CPM) table for the RNAseq samples as a text file.

organism: Schizosaccharomyces pombe

source_name: S. pombe cell culture

growth_protocol: S. pombe cell cultures were grown exponentially in YES media at 30 degrees celsius.

extract_protocol:
Total RNA was extracted with the MasterPure RNA Purification Kit from epicenter (MPY03100).
Total RNA libraries were prepared with TruSeq Stranded Total RNA kit (Illumina, 20020599) according to manufacturer’s instructions.

tag: strain

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+-TTAGTT


title: RNA-Seq A2780 control rep2


title: RNA-Seq A2780 control rep1


title: total RNAseq, PAS-T4G, replicate 1


title: total RNAseq, PAS-T4C, replicate 3

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+-TTACTT


title: total RNAseq, PAS-T4C, replicate 1


title: total RNAseq, PAS-T4G, replicate 2


title: Patient52_tumor [R52B] miRNA-Seq

first: Caroline G.

last: Lee

address: MD7, Level 2, 8 Medical Drive

department: Biochemistry

institute: National University of Singapore

data_processing:
Clean Reads: Raw sequences were generated as clean reads from Illumina Nextseq by real-time base calling and quality filtering. The clean reads were recorded in FASTQ format, containing the read information, sequences and quality encoding. The clean reads were located in a FASTQ format file.
Trimmed Reads: Subsequently, the 3’ adapter sequence was trimmed from the clean reads and the reads with lengths shorter than 15 nt were discarded. As the 5’-adaptor was also used as the sequencing primer site, the 5’-adaptor sequence is not present in the sequencing reads. The trimmed reads were recorded in FASTA format.
Aligned Reads: The trimmed reads (length>=15 nt) were aligned to the human pre-miRNA in miRBase 21, using NovoAlign software (NovoCraft, Selangor, Malaysia). For miRNA alignment, the maximum mismatch was 1. Reads with counts less than 2 were discarded when calculating the miRNA expression. The alignment results were saved in text files.
Normalization: The miRNA expression levels were measured and normalized as transcripts per million of total aligned miRNA reads (TPM).
miRNA Expression Profiles: The miRNA read counts are used to estimate the expression level of each miRNA. The sequence counts for all the miRNAs in the samples are compiled in an Excel file
Genome_build: miRBase 21 Release
Supplementary_files_format_and_content: premiRNA_alignment_raw.txt
Supplementary_files_format_and_content: premiRNA_alignment_normalization.txt

source_name: Liver

extract_protocol:
The frozen tissues were added with RLT buffer (Qiagen, Germany) with 1% β-mercaptoethanol and then homogenized by gentleMACS dissociator (Miltenyi Biotec). RNeasy mini kit (Qiagen, Germany) was used to extract the total RNA following the manufacturer’s protocol.
Total RNA of each sample was used to prepare the miRNA sequencing library, which included the following steps: 1) 3'-adapter ligation; 2) 5'-adapter ligation; 3) cDNA synthesis; 4) PCR amplification; 5) size selection of ~130-150 bp PCR amplified fragments (corresponding to ~15-35 nt small RNAs).

value: hepatocellular carcinoma

value: Tumor

tag: gender

value: F


title: Patient59_adjacent non-tumor [R59A] miRNA-Seq

value: Adjacent non-tumor

value: M


title: total RNAseq, PAS-T4C, replicate 2


title: RNA-Seq A2780 cisplatin rep1

value: ovarian carcinoma


title: Patient52_adjacent non-tumor [R52A] miRNA-Seq


title: total RNAseq, PAS-T4A, replicate 2

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+-TTAATT


title: total RNAseq, swd22D, replicate 2

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ swd22D::kanMX


title: total RNAseq, PAS-T4A, replicate 1


title: Patient50_tumor [R50B] miRNA-Seq


title: total RNAseq, swd22D, replicate 1


title: Patient49_tumor [R49B] miRNA-Seq


title: Patient50_adjacent non-tumor  [R50A] miRNA-Seq


title: total RNAseq, ppn1D, replicate 3

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ ppn1D::kanMX


title: Patient47_tumor [R47B] miRNA-Seq


title: Patient49_adjacent non-tumor [R49A] miRNA-Seq


title: total RNAseq, ppn1D, replicate 2


title: total RNAseq, swd22D, replicate 3


title: Patient46_tumor [R46B] miRNA-Seq


title: Patient47_adjacent non-tumor [R47A] miRNA-Seq


title: total RNAseq, ppn1D, replicate 1


title: total RNAseq, dis2D, replicate 3

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ dis2D::kanMX


title: Patient59_tumor [R59B] miRNA-Seq


title: total RNAseq, dis2D, replicate 2


title: total RNAseq, PAS-T4A, replicate 3


title: total RNAseq, dis2D, replicate 1


title: Patient45_tumor [R45B] miRNA-Seq


title: Patient45_adjacent non-tumor [R45A] miRNA-Seq


title: total RNAseq, ssu72D, replicate 3

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ ssu72D::kanMX


title: Patient44_tumor [R44B] miRNA-Seq


title: Patient44_adjacent non-tumor [R44A] miRNA-Seq


title: total RNAseq, ssu72D, replicate 2


title: Patient43_tumor [R43B] miRNA-Seq


title: total RNAseq, ssu72D, replicate 1


title: total RNAseq, pfs2-F115H, replicate 3

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ pfs2-F115H::kanMX


title: Patient43_adjacent non-tumor  [R43A] miRNA-Seq


title: Patient42_tumor [R42B] miRNA-Seq


title: Patient42_adjacent non-tumor [R42A] miRNA-Seq


title: total RNAseq, pfs2-F115H, replicate 1


title: Patient41_tumor [R41B] miRNA-Seq


title: total RNAseq, yth1-R59C, replicate 3


title: total RNAseq, pfs2-F115H, replicate 2


title: total RNAseq, yth1-R59C, replicate 2


title: Patient46_adjacent non-tumor  [R46A] miRNA-Seq


title: Patient40_adjacent non-tumor [R40A] miRNA-Seq


title: total RNAseq, yth1-R59C, replicate 1


title: total RNAseq, wt, replicate 3


title: total RNAseq, wt, replicate 2


title: Patient40_tumor [R40B] miRNA-Seq


title: Patient39_tumor [R39B] miRNA-Seq


title: total RNAseq, wt, replicate 1


title: Patient38_tumor [R38B] miRNA-Seq


title: Patient38_adjacent non-tumor [R38A] miRNA-Seq


title: Patient39_adjacent non-tumor [R39A] miRNA-Seq


title: Patient37_adjacent non-tumor [R37A] miRNA-Seq


title: Patient35_tumor [R35B] miRNA-Seq


title: Patient36_tumor [R36B] miRNA-Seq


title: Patient36_adjacent non-tumor [R36A] miRNA-Seq


title: Patient34_adjacent non-tumor [R34A] miRNA-Seq


title: Patient34_tumor [R34B] miRNA-Seq


title: Patient35_adjacent non-tumor [R35A] miRNA-Seq


title: Patient33_adjacent non-tumor [R33A] miRNA-Seq


title: Patient32_tumor [R32B] miRNA-Seq


title: Patient41_adjacent non-tumor [R41A] miRNA-Seq


title: Patient31_adjacent non-tumor [R31A] miRNA-Seq


title: Patient32_adjacent non-tumor [R32A] miRNA-Seq


title: Patient31_tumor [R31B] miRNA-Seq


title: Patient30_adjacent non-tumor [R30A] miRNA-Seq


title: Patient30_tumor [R30B] miRNA-Seq


title: Patient29_tumor [R29B] miRNA-Seq


title: WT3_24h

city: Oxford

first: Adam

last: Handel

address: John Radcliffe Hospital

department: Weatherall Institute of Molecular Medicine

country: United Kingdom

institute: University of Oxford

data_processing:
Sequencing data was trimmed using Trimmomatic and aligned to the mm10 genome using STAR (version 2.5.3a)
Reads were assigned to genes using HTSeq (intersection non-empty)
Sample KO1 was excluded from downstream analysis as it was an outlier on PCA
Genome_build: mm10
Supplementary_files_format_and_content: Gene counts

source_name: Naïve CD4 T cells

extract_protocol:
RNA was extracted using the RNeasy Plus Micro kit (Qiagen).
RNA was processed using the RNA-Seq Poly A method.

treatment_protocol: Two thousand naïve CD4 cells were isolated by FACS sorting directly into lysis buffer (RLT buffer (Qiagen)) for timepoint 0 hrs. At the same time, 106 cells/mL were activated in vitro using a combination of plate bound anti-CD3 (2 ug/mL) (BioLegend) and soluble anti-CD28 (2 ug/mL) (BioLegend) antibodies for 24 hrs before 2000 live cells were sorted directly into lysis buffer.

value: Anti-CD3 and anti-CD28 for 24h

value: Naive CD4 T cells


title: WT2_24h


title: WT1_24h


title: WT3_0h

value: Untreated


title: WT2_0h


title: Patient29_adjacent non-tumor [R29A] miRNA-Seq


title: Patient28_adjacent non-tumor  [R28A] miRNA-Seq


title: Patient28_tumor [R28B] miRNA-Seq


title: Patient27_tumor [R27B] miRNA-Seq


title: Patient37_tumor [R37B] miRNA-Seq


title: KO3_24h


title: WT1_0h


title: KO1_24h


title: Patient27_adjacent non-tumor [R27A] miRNA-Seq


title: Patient24_adjacent non-tumor [R24A] miRNA-Seq


title: KO2_24h


title: Patient26_adjacent non-tumor  [R26A] miRNA-Seq


title: KO2_0h


title: Patient26_tumor [R26B] miRNA-Seq


title: Patient33_tumor [R33B] miRNA-Seq


title: KO1_0h


title: Patient24_tumor [R24B] miRNA-Seq


title: Patient23_tumor [R23B] miRNA-Seq


title: Patient23_adjacent non-tumor [R23A] miRNA-Seq


title: Patient22_tumor [R22B] miRNA-Seq


title: pdx1.gRNA.rep3

city: Durham

first: Charles

last: Gersbach

address: 101 Science Drive

department: Biomedical Engineering

institute: Duke University

data_processing:
Sequences for TruSeq Illumina adapters were removed from the raw reads using Trimmomatic v0.32 (Bolger et al., 2014).
Reads were aligned against the latest mouse reference genome (GRCm38) using the aligner STAR v2.4.1a following the proposed 2-pass strategy, removing alignments containing non-canonical splice junctions (--outFilterIntronMotifs RemoveNoncanonical)
Aligned reads were assigned to genes in the GENCODE vM13 comprehensive gene annotation using the featureCounts command in the subread package with default settings (v1.4.6-p4)
Differential expression analysis was performed using DESeq2 (v1.22.0) running on R (v3.5.1). Briefly, raw counts were imported and filtered to remove genes with low or no expression, that is, keeping genes having two or more counts per million (CPMs) in two or more samples.
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited files with raw gene counts
Supplementary_files_format_and_content: bam files with aligned reads

source_name: liver

extract_protocol:
On day of harvest, mice were euthanized by CO2 inhalation. Livers were harvested and immediately snap frozen using liquid N2. Total RNA was extracted using Qiagen RNAuniversal Kit.
Library Preparation and sequencing was performed by GENEWIZ.

treatment_protocol: Approximately 8 week old animals were injected with PBS or AAV.Cbh.gRNA (either control or targeted)

tag: grna target

tag: total viral genomes/mouse)

value: AAV expressing gRNA pdx1


title: pdx1.gRNA.rep4

tag: treatment


title: Patient22_adjacent non-tumor [R22A] miRNA-Seq


title: pdx1.gRNA.rep2


title: Patient21_adjacent non-tumor  [R21A] miRNA-Seq


title: Patient20_tumor [R20B] miRNA-Seq


title: Patient21_tumor [R21B] miRNA-Seq


title: Patient20_adjacent non-tumor [R20A] miRNA-Seq


title: pdx1.gRNA.rep1


title: controlgRNA.rep3 (2)

value: non-targeting

value: AAV expressing gRNA control


title: controlgRNA.rep2 (2)


title: controlgRNA.rep4 (2)


title: Patient19_tumor [R19B] miRNA-Seq

value: hepatocellular carcinoma

value: Tumor


title: controlgRNA.rep1 (2)


title: KO3_0h

tag: cell type


title: Patient19_adjacent non-tumor [R19A] miRNA-Seq

value: Adjacent non-tumor

value: F


title: Patient18_tumor [R18B] miRNA-Seq

value: M


title: PBS.rep4 (2)

value: PBS


title: PBS.rep3 (2)


title: PBS.rep1 (2)


title: Patient17_adjacent non-tumor [R17A] miRNA-Seq


title: Patient16_tumor [R16B] miRNA-Seq


title: K27ac.Pdx1.rep8

data_processing:
Sequences for TruSeq Illumina adapters were removed from the raw reads using Trimmomatic v0.32 (Bolger et al., 2014).
Single-end reads were mapped using Bowtie (v1.0.0 PMID: 19261174) against Mouse (mm10) genome with flags -v 2 -X 2000 --best --strata -m 1.
Duplicates were removed using Picard MarkDuplicates v1.130
Low mappability or blacklisted regions identified by the ENCODE project (Consortium, 2012) were filtered out from the final BAM files
Using the sequenced input controls, binding regions were identified using the callpeak function in MACS2 v2.1.0.20151222 (Zhang et al., 2008).
Signal files were generated with deeptools bamCoverage (v3.0.1 PMID:24799436) ignoring duplicates, extending reads 200bp and converting the read counts to RPKMs.
For the differential binding analysis, first, a union peakset was computed merging individual peak calls using bedtools2 v2.25.0 (Quinlan and Hall, 2010). Then, reads in peaks were estimated using featureCounts and the difference in binding was assessed with DESeq2.
Genome_build: mm10
Supplementary_files_format_and_content: bigWig signal in RPMs

extract_protocol:
On day of harvest, mice were euthanized by CO2 inhalation. Livers were harvested and immediately snap frozen using liquid N2.  Livers were then treated for chromatin extraction as described in Savic et al, 2013. Briefly, following fixation, livers were rocked at 4˚C in 10 ml Farnham Lysis Buffer for 10 minutes followed by centrifugation. Pellets were resuspended in 4 mL RIPA buffer. 1 mL RIPA and 1 mL RIPA containing resuspended liver was placed into a 15 mL tube with 800 mg of Diagenode sonication beads (Diagenode, C01020031). This tube was sonicated for 30 cycles of 30s on and 30s off using a Bioruptor Pico.
ChIP-seq libraries were prepared using a Kapa Hyper Prep kit.


title: K27ac.Pdx1.rep9


title: K27ac.Pdx1.rep7


title: Patient15_tumor [R15B] miRNA-Seq


title: Patient15_adjacent non-tumor  [R15A] miRNA-Seq


title: K27ac.Pdx1.rep6


title: K27ac.controlgRNA.rep5


title: Patient14_tumor [R14B] miRNA-Seq


title: Patient14_adjacent non-tumor [R14A] miRNA-Seq


title: K27ac.controlgRNA.rep3


title: Patient18_adjacent non-tumor  [R18A] miRNA-Seq


title: Patient17_tumor [R17B] miRNA-Seq


title: Patient16_adjacent non-tumor [R16A] miRNA-Seq


title: Patient59_adjacent non-tumor

hyb_protocol: 1 μg of each labeled cRNA was fragmented by adding 5 μl 10 × Blocking Agent and 1 μl of 25 × Fragmentation Buffer, then heated the mixture at 60 °C for 30 min, finally 25 μl 2 × Hybridization buffer was added to dilute the labeled cRNA. 50 μl of hybridization solution was dispensed into the gasket slide and assembled to the circRNA expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven.

scan_protocol: The scanning was performed using Agilent DNA Microarray Scanner (part number G2505C) after washing and fixing the hybridized arrays.

data_processing: Agilent Feature Extraction software (version 11.0.1.1) was used to analyze acquired array images. Quantile normalization and subsequent data processing were performed using the R software limma package.

label_protocol: Total RNAs were digested with Rnase R (Epicentre, Inc.) to remove linear RNAs and enrich circular RNAs. Then, the enriched circular RNAs were amplified and transcribed into fluorescent cRNA utilizing a random priming method (Arraystar Super RNA Labeling Kit; Arraystar). The labeled cRNAs were purified by RNeasy Mini Kit (Qiagen). The concentration and specific activity of the labeled cRNAs (pmol Cy3/μg cRNA) were measured by NanoDrop ND-1000.

extract_protocol: The frozen tissues were added with RLT buffer (Qiagen, Germany) with 1% β-mercaptoethanol and then homogenized by gentleMACS dissociator (Miltenyi Biotec). RNeasy mini kit (Qiagen, Germany) was used to extract the total RNA following the manufacturer’s protocol.


title: PBS.rep2 (2)


title: Patient59_tumor


title: K27ac.controlgRNA.rep2


title: Patient13_tumor [R13B] miRNA-Seq


title: K27ac.controlgRNA.rep1


title: Patient13_adjacent non-tumor [R13A] miRNA-Seq


title: Patient52_tumor


title: Patient52_adjacent non-tumor


title: K27ac.PBS.rep4


title: Patient12_tumor [R12B] miRNA-Seq


title: Patient49_tumor


title: Patient50_tumor


title: Patient47_adjacent non-tumor


title: K27ac.PBS.rep12


title: Patient47_tumor


title: Patient46_tumor


title: Patient45_adjacent non-tumor


title: Patient12_adjacent non-tumor  [R12A] miRNA-Seq


title: Patient50_adjacent non-tumor


title: K27ac.PBS.rep11


title: Patient49_adjacent non-tumor


title: Patient11_tumor [R11B] miRNA-Seq


title: Patient45_tumor


title: Patient44_tumor


title: K27ac.PBS.rep10


title: Patient11_adjacent non-tumor [R11A] miRNA-Seq


title: Patient10_tumor [R10B] miRNA-Seq


title: Patient43_adjacent non-tumor


title: input.Pdx1.rep9

value: input


title: input.Pdx1.rep8


title: Patient44_adjacent non-tumor


title: Patient43_tumor


title: Patient46_adjacent non-tumor


title: Patient42_adjacent non-tumor


title: Patient42_tumor


title: Patient9_tumor [R09B] miRNA-Seq


title: input.Pdx1.rep6


title: Patient40_tumor


title: Patient41_tumor


title: Patient39_tumor


title: Patient41_adjacent non-tumor


title: Patient7_tumor [R07B] miRNA-Seq


title: input.controlgRNA.rep5


title: Patient40_adjacent non-tumor


title: input.controlgRNA.rep3


title: Patient10_adjacent non-tumor [R10A] miRNA-Seq


title: Patient7_adjacent non-tumor [R07A] miRNA-Seq


title: input.Pdx1.rep7


title: Patient6_tumor [R06B] miRNA-Seq


title: Patient37_adjacent non-tumor


title: Patient37_tumor


title: Patient36_adjacent non-tumor


title: AK12282 Combo Rep2

city: Houston

first: Haoqiang

last: Ying

state: Texas

address: 1515 Holcombe Blvd

institute: M. D. Anderson Cancer Center

description:
Sample_40-AK12282_Combo2
40-AK12282_Combo2_S18_L005

data_processing:
fastq files were preprocessed using fastp software to remove low quality reads
clean fastq files were mapped to mouse transcriptome (mm10) using salmon v0.14.1 to generate counts and TPM for each transcript
raw counts and TPM for each transcript were aggregated by gene symbols to generate a gene expression TPM table
Genome_build: mm10
Supplementary_files_format_and_content: tab delimited txt file of TPM gene expression table

source_name: AK12282 Combo

growth_protocol: 500000 iKras cell were seeded in the 10cm dish and cultured with RPMI1640 with 10% FBS.

extract_protocol:
Total RNA was extracted using the Qiagen RNeasy kit following the manufacturer's instruction.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: iKras cell the treatment with vehicle, 1 mM 2DG, 25 nM Trametinib, or combination for 24 h.

tag: cell line

value: PDAC

value: p48Cre; tetO_KrasG12D; Rosa_rtTAL/+; p53L/+


title: Patient35_adjacent non-tumor


title: input.controlgRNA.rep1


title: Patient9_adjacent non-tumor  [R09A] miRNA-Seq


title: Patient35_tumor


title: input.controlgRNA.rep2


title: Patient39_adjacent non-tumor


title: AK12282 Combo Rep1

description:
Sample_39-AK12282_Combo1
39-AK12282_Combo1_S16_L005

tag: genotype


title: Patient6_adjacent non-tumor [R06A] miRNA-Seq


title: input.PBS.rep4 (2)

tag: antibody


title: Patient38_adjacent non-tumor

tag: age


title: input.PBS.rep12 (2)


title: Patient5_tumor [R05B] miRNA-Seq


title: AK12282 2DG Rep2

description:
Sample_38-AK12282_2DG2
38-AK12282_2DG2_S14_L005


title: Patient5_adjacent non-tumor  [R05A] miRNA-Seq


title: input.PBS.rep11


title: Patient4_tumor [R04B] miRNA-Seq


title: AK12282 2DG Rep1

description:
Sample_37-AK12282_2DG1
37-AK12282_2DG1_S13_L005


title: AK12282 Tra Rep2

description:
Sample_36-AK12282_MEKi2
36-AK12282_MEKi2_S22_L005

source_name: AK12282 Tra


title: Patient34_tumor


title: input.PBS.rep10


title: Patient34_adjacent non-tumor


title: Patient4_adjacent non-tumor [R04A] miRNA-Seq


title: AK12282 Tra Rep1

description:
Sample_35-AK12282_MEKi1
35-AK12282_MEKi1_S21_L005


title: Patient33_adjacent non-tumor


title: Patient38_tumor


title: Cas9.Pdx1.rep9


title: Patient3_tumor [R03B] miRNA-Seq


title: Patient3_adjacent non-tumor [R03A] miRNA-Seq


title: Cas9.Pdx1.rep8


title: AK12282 Dox OFF Rep2

description:
Sample_34-AK12282_DOX_OFF2
34-AK12282_DOX_OFF2_S20_L005

source_name: AK12282 Dox OFF


title: Cas9.Pdx1.rep7


title: Patient2_tumor [R02B] miRNA-Seq


title: Patient32_tumor


title: AK12282 Dox OFF Rep1

description:
Sample_33-AK12282_DOX_OFF1
33-AK12282_DOX_OFF1_S19_L005


title: AK12282 Dox ON Rep2

description:
Sample_32-AK12282_DOX_ON2
32-AK12282_DOX_ON2_S17_L005

source_name: AK12282 Dox ON


title: Patient2_adjacent non-tumor  [R02A] miRNA-Seq


title: Patient33_tumor


title: Patient32_adjacent non-tumor


title: AK12282 Dox ON Rep1

description:
Sample_31-AK12282_DOX_ON1
31-AK12282_DOX_ON1_S15_L005


title: Patient31_tumor


title: Cas9.controlgRNA.rep5


title: Patient1_adjacent non-tumor [R01A] miRNA-Seq


title: AK192 Combo Rep1

description:
Sample_29-AK192_Combo1
R406-29-AK192Combo1_S54_L006

source_name: AK192 Combo


title: Patient30_adjacent non-tumor


title: Patient1_tumor [R01B] miRNA-Seq


title: Cas9.controlgRNA.rep3


title: Patient31_adjacent non-tumor


title: Patient36_tumor


title: Cas9.controlgRNA.rep2


title: Cas9.controlgRNA.rep1


title: H3K27ac_siChe-1

city: Rome

first: Matteo

last: Pallocca

address: Via Fermo Ognibene

country: Italy

institute: Italian National Cancer Institute "Regina Elena"

data_processing:
BaseSpace Illumina Cloud Environment was used for basecalling and demultiplexing.
Reads were mapped with Bowtie vv. 2.2.3
Peaks were called with MACS2
Genome_build: hg38
Supplementary_files_format_and_content: bigWig enrichment files generated with MACS2 bdgcmp command

extract_protocol: For Che-1 (Bruno et al., 2002), RNA pol II (Millipore, #05-623) H3K27AC (Millipore, #07-360), H3K9me3 (Abcam, #8898) ChIP-seq, 10ng of total ChIP DNA was used for library construction using the TruSeq ChIP Library Preparation Kit (Illumina,CA) and sequenced using NextSeq 500 system (Illumina, CA) to obtain 100bp single-end reads.

value: bone marrow

tag: genotype/variation

value: Che-1 silencing

tag: chip antibody

value: H3K27ac (Millipore, Cat# 07-360; RRID:AB_310550)


title: Cas9.Pdx1.rep6


title: AK192 Combo Rep2

description:
Sample_30-AK192_Combo2
R406-30-AK192Combo2_S53_L006


title: Cas9.PBS.rep12


title: Cas9.PBS.rep11


title: AK192 2DG Rep1

description:
Sample_27-AK192_2DG1
R406-27-AK1922DG1_S56_L006

source_name: AK192 2DG


title: Che-1

value: control

value: Che-1 (Bruno et al., 2002)


title: Cas9.PBS.rep4


title: H3K27ac_CN


title: AK192 Tra Rep1

description:
Sample_25-AK192_MEKi1
R406-25-AK192MEKi1_S58_L006

source_name: AK192 Tra


title: Patient29_tumor

tag: disease


title: AK192 Tra Rep2

description:
Sample_26-AK192_MEKi2
R406-26-AK192MEKi2_S57_L006


title: input


title: Cas9.PBS.rep10


title: H3K9me3

value: H3K9me3 (Abcam, Cat# ab8898; RRID:AB_306848)


title: AK192 Dox OFF Rep2

description:
Sample_24-AK192_DOXOFF2
R406-24-AK192DOXOFF2_S59_L006

source_name: AK192 Dox OFF


title: H3K4me3

value: H3K4me3 (Abcam, Cat# ab1012; RRID:AB_442796)


title: AK192 Dox OFF Rep1

description:
Sample_23-AK192_DOXOFF1
R406-23-AK192DOXOFF1_S60_L006


title: AK192 2DG Rep2

description:
Sample_28-AK192_2DG2
R406-28-AK1922DG2_S55_L006


title: Patient30_tumor


title: Patient27_adjacent non-tumor


title: Patient27_tumor


title: Patient29_adjacent non-tumor


title: Patient28_tumor


title: pcsk9.gRNA.rep3

extract_protocol:
On day of harvest, mice were euthanized by CO2 inhalation. Livers were harvested and immediately snap frozen using liquid N2. Total RNA was extracted using Qiagen RNAuniversal Kit.
Library Preparation and sequencing was performed by NOVAGENE

value: AAV expressing gRNA pcsk9


title: H3K4me1

value: H3K4me1 (Abcam, Cat# ab8895; RRID:AB_306847)


title: pcsk9.gRNA.rep4


title: Patient26_tumor

value: hepatocellular carcinoma

value: Tumor


title: Pol_II_siChe-1

value: PolII  (Millipore, #05-623)


title: AK192 Dox ON Rep1

description:
Sample_21-AK192_DOXON1
R406-21-AK192DOXON1_S62_L006

source_name: AK192 Dox ON


title: Patient28_adjacent non-tumor

value: Adjacent non-tumor

value: F


title: Pol_II


title: pcsk9.gRNA.rep2


title: Patient26_adjacent non-tumor

value: M


title: AK192 Dox ON-3day Rep3 [ON-D3-5]

source_name: AK192 Dox ON-3day

treatment_protocol: For the tumor samples, the mice were randomized into three groups (Kras-ON/Vehicle, Kras-OFF/Vehicle, Kras ON/Trametinib) on the 7th day after orthotopic injection. After 1 or 3 days of treatment, the tumors were collected for RNA extraction.

value: PDAC

value: 3 day


title: pcsk9.gRNA.rep1


title: controlgRNA.rep4

value: non-targeting

value: AAV expressing gRNA control


title: AK192 Dox ON-3day Rep2 [ON-D3-3]

value: p48Cre; tetO_KrasG12D; Rosa_rtTAL/+; p53L/+


title: AK192 Dox ON-3day Rep1 [ON-D3-2]


title: AK192 Dox ON Rep2

description:
Sample_22-AK192_DOXON2
R406-22-AK192DOXON2_S61_L006


title: ATAC_CN

data_processing:
BaseSpace Illumina Cloud Environment was used for basecalling and demultiplexing.
Reads were mapped with Bowtie vv. 2.2.3
Peaks were called with MACS2, with parameters --nomodel --shift -100 --extsize 200
Genome_build: hg38
Supplementary_files_format_and_content: bigWig enrichment files generated with MACS2 bdgcmp command

extract_protocol:
To profile open chromatin, we used the ATAC-seq protocol developed by (Buenrostro et al., 2013) with minor modifications. Malignants and control Bone Marrow aspirates, were isolated for PCs by magnetic cell separation using human CD138 positive selection kit (Miltenyi-Biotec- Germany) and Macs separator (Macs Miltenyi-biotec- Germany. To perform the analysis 50,000 cells were washed once with 1X PBS and centrifuged at 500g for 5 minutes at 4°C. For flash-frozen cell pellets were removed from -80°C and immediately re-suspended in ice-cold cell lysis buffer. Nuclear morphological evaluation was performed by Trypan blue coloration, before starting the tagmentation. Pellets were re-suspended in transposase 25 μl reaction buffer containing 2,5 μl of Tn5 transposase and 12.5 μl of TD buffer (Nextera DNA Sample preparation kit from Illumina) on ice. Samples were incubated at 37°C for 30 min. The samples were purified with MiniElute PCR Purification Kit (Qiagen).
To amplify transposed DNA fragments we used NEBNext High-Fidelyty 2x PCR Master Mix (New England Labs) and the Customized Nextera PCR Primers (see Table 3). Libraries were purified using Agencourt Ampure XP (Beckman) magnetic beads (1:1 ratio) to remove remaining adapters and double purified (1:0,5 and 1:1,8-05) for right side selection. Libraries were controlled using a High Sensitivity DNA Kit Bioanalyzer (Agilent Technologies).

value: bone marrow

value: control


title: ATAC_siChe-1

value: Che-1 silencing


title: Patient24_adjacent non-tumor


title: AK192 Dox ON-1day Rep2 [ON-D1-4]

source_name: AK192 Dox ON-1day

value: 1 day


title: AK192 Dox ON-1day Rep3 [ON-D1-5]


title: KMS27_siChe-1_2

data_processing:
BaseSpace Illumina Cloud Environment was used for basecalling and demultiplexing.
RNA-seq was analyzed via the web pipeline RAP, which employs the Tuxedo suite for mapping (Tophat) and transcripts reconstruction (Cufflinks, Cuffdiff)
For spikein-normalized data, HTSeq count were normalized using spikein raw counts as scaling factors.
Genome_build: hg19
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample.
[deseq2_spikein_normalized.tsv] all genes spikein-normalized differential expression
[spikein_count.csv] spikein raw count table
[all_cuffdiff_transcripts.tsv] transcript abundance

extract_protocol:
total RNA was extracted from KMS27 cells using Qiazol (Invitrogen), purified from genomic DNA contamination through a DNase I (Qiagen) digestion step and further enriched by Qiagen RNeasy columns for gene expression profiling (Qiagen). Quantity and integrity of the extracted RNA were assessed by NanoDrop Spectrophotometer (NanoDrop Technologies) and by Agilent 2100 Bioanalyzer (Agilent Technologies), respectively.
RNA libraries for sequencing were generated in triplicate according to the Illumina TruSeq Stranded Total RNA kit with an initial ribosomal depletion step using Ribo Zero Gold (Illumina, Inc., San Diego, CA, US). Starting material was 500ng of total RNA.


title: controlgRNA.rep1


title: Patient22_tumor


title: Patient23_adjacent non-tumor


title: Patient24_tumor


title: controlgRNA.rep2


title: Patient22_adjacent non-tumor


title: KMS27_siControl_3


title: KMS27_siChe-1_1


title: Patient21_tumor


title: Patient23_tumor


title: KMS27_siControl_2


title: AK192 Dox OFF-3day Rep2 [OFF-D3-4]

source_name: AK192 Dox OFF-3day


title: PBS.rep4

value: PBS


title: PBS.rep3


title: AK192 Dox ON-1day Rep1 [ON-D1-3]


title: controlgRNA.rep3


title: PBS.rep2


title: KMS27_siControl_1


title: Patient21_adjacent non-tumor


title: PBS.rep1


title: 52 NAC.High_3h [TRA00038981]

city: Boston

first: Hui

last: Pan

address: 1 Joslin Place

department: Bioinformatics and Biostatistics Core

institute: Joslin Diabetes Center

description: Mouse Nucleus accumbens RNA-seq by Illumina HiSeq 2500

data_processing:
Reads were aligned to the mouse genome using STAR.
Reads were counted with featureCounts.
Genome_build: GRCm38.p4
Supplementary_files_format_and_content: CSV file contains read counts.

source_name: High_Nucleus accumbens

extract_protocol:
Brain tissues were harvested, snap frozen in liquid nitrogen and kept at -80C. Total RNA was isolated from hypothalamus, hippocampus and nucleus accumbens.
Stranded cDNA libraries with unique index tags for each sample (48 multiplexed) were made using a directional RNA-seq kit (Wafergen).

tag: Sex

value: male

value: 3 months

tag: time point

value: After 3 h of clamp

tag: dose

value: High

value: Nucleus accumbens


title: AK192 Dox OFF-3day Rep1 [OFF-D3-3]


title: AK192 Dox OFF-1day Rep3 [OFF-D1-4]

source_name: AK192 Dox OFF-1day


title: AK192 Dox OFF-3day Rep3 [OFF-D3-5]


title: Patient20_tumor

source_name: Liver

value: hepatocellular carcinoma

tag: tissue

value: Tumor

value: F


title: Patient20_adjacent non-tumor

value: Adjacent non-tumor


title: 51 NAC.Low_3h [TRA00038980]

source_name: Low_Nucleus accumbens

tag: strain

value: Low


title: AK192 Dox OFF-1day Rep2 [OFF-D1-3]

value: PDAC


title: 50 NAC.Basal_3h [TRA00038979]

source_name: Basal_Nucleus accumbens

value: Basal


title: 49 HCA.High_3h [TRA00038978]

description: Mouse Hippocampus RNA-seq by Illumina HiSeq 2500

value: Hippocampus


title: AK192 Dox OFF-1day Rep1 [OFF-D1-1]

value: p48Cre; tetO_KrasG12D; Rosa_rtTAL/+; p53L/+


title: K9me3.Pcsk9.rep6

source_name: liver


title: AK192 Tra-3day Rep3 [MEKI-D3-5]

source_name: AK192 Tra-3day

value: 3 day


title: K9me3.Pcsk9.rep5


title: K9me3.Pcsk9.rep7


title: 48 HCA.Low_3h [TRA00038977]


title: AK192 Tra-3day Rep2 [MEKI-D3-4]


title: Patient19_tumor

tag: gender


title: K9me3.Pcsk9.rep3

tag: grna target


title: Patient18_tumor

value: M


title: Patient18_adjacent non-tumor


title: K9me3.controlgRNA.rep9

value: non-targeting

tag: total viral genomes/mouse)


title: 46 HTM.High_3h [TRA00038975]

description: Mouse Hypothalamus RNA-seq by Illumina HiSeq 2500

value: male

value: Hypothalamus


title: 47 HCA.Basal_3h [TRA00038976]

value: 3 months

value: After 3 h of clamp


title: K9me3.controlgRNA.rep8


title: Patient17_adjacent non-tumor


title: AK192 Tra-1day Rep3 [MEKI-D1-5]

source_name: AK192 Tra-1day

tag: treatment

value: 1 day


title: Patient17_tumor

value: hepatocellular carcinoma


title: Patient19_adjacent non-tumor


title: 45 HTM.Low_3h [TRA00038974]


title: K9me3.controlgRNA.rep11


title: AK192 Tra-1day Rep2 [MEKI-D1-4]


title: K9me3.controlgRNA.rep10


title: Patient16_adjacent non-tumor


title: Patient16_tumor

value: Tumor


title: K9me3.PBS.rep2


title: K9me3.PBS.rep1


title: AK192 Tra-1day Rep1 [MEKI-D1-3]


title: 43 NAC.High_3h [TRA00038960]

value: High

value: Nucleus accumbens


title: KMS27_siChe-1_3

value: bone marrow

value: Che-1 silencing


title: 41 NAC.Basal_3h [TRA00038958]

value: Basal


title: 44 HTM.Basal_3h [TRA00038973]


title: 42 NAC.Low_3h [TRA00038959]

value: Low


title: 40 HCA.High_3h [TRA00038942]

value: Hippocampus


title: 39 HCA.Low_3h [TRA00038941]


title: AK192 Tra-3day Rep1 [MEKI-D3-2]

value: PDAC

value: p48Cre; tetO_KrasG12D; Rosa_rtTAL/+; p53L/+

value: 3 day


title: Patient15_adjacent non-tumor

value: Adjacent non-tumor

value: M


title: K9me3.PBS.rep12


title: Patient15_tumor


title: input.Pcsk9.rep7


title: Patient13_tumor


title: 38 HCA.Basal_3h [TRA00038940]

value: male

value: 3 months


title: input.Pcsk9.rep6


title: 37 HTM.High_3h [TRA00038939]

value: After 3 h of clamp

value: Hypothalamus


title: input.Pcsk9.rep5


title: K9me3.PBS.rep4


title: input.Pcsk9.rep3


title: Patient12_adjacent non-tumor

value: hepatocellular carcinoma


title: input.controlgRNA.rep9

value: non-targeting


title: Patient14_tumor

value: Tumor


title: 36 HTM.Low_3h [TRA00038938]


title: 35 HTM.Basal_3h [TRA00038935]


title: 34 NAC.High_3h [TRA00038921]

value: High

value: Nucleus accumbens


title: Patient12_tumor


title: Patient13_adjacent non-tumor


title: input.controlgRNA.rep11


title: Patient11_tumor


title: 33 NAC.Low_3h [TRA00038920]


title: input.controlgRNA.rep10


title: input.PBS.rep4


title: 32 NAC.Basal_3h [TRA00038919]

value: Basal


title: 31 HCA.High_3h [TRA00038918]


title: Patient10_tumor


title: Patient11_adjacent non-tumor


title: input.PBS.rep2


title: 30 HCA.Low_3h [TRA00038917]

value: Low


title: input.controlgRNA.rep8


title: Patient9_adjacent non-tumor


title: input.PBS.rep1


title: sRNAseq, ssu72D

description: 3' adapter sequence: TGGAATTCTCGGGTGCCAAGG

data_processing:
Raw data were demultiplexed and converted to fastq format using bcl2fastq2 (v1.17) Illumina. Qiagen 3' adaptors were trimmed using cutadapt (v1.18) with the parameters --discard-untrimmed -m 18. Fastq files were aligned using Bowtie (v1.2.2) with parameters -f -M 10000 -v 0 -S --best --strata. Bowtie sam output was converted to bam using samtools (v1.9). Bam files were converted to bed using bedtools bamtobed (v2.27.1), bed files were used to calculate strand-specific coverage using bedtools genomeCoverageBed (v2.27.1), resulting bedgraph files were normalized to 1 million genome mapping reads (RPM), and converted to bigWig format using bedGraphToBigWig (v4).
smallRNA-seq:Raw data were demultiplexed and converted to fastq format using bcl2fastq2 (v1.17) Illumina. Qiagen 3' adaptors were trimmed using cutadapt (v1.18) with the parameters --discard-untrimmed -m 18. Fastq files were aligned using Bowtie (v1.2.2) with parameters -f -M 10000 -v 0 -S --best --strata. Bowtie sam output was converted to bam using samtools (v1.9). Bam files were converted to bed using bedtools bamtobed (v2.27.1), bed files were used to calculate strand-specific coverage using bedtools genomeCoverageBed (v2.27.1), resulting bedgraph files were normalized to 1 million genome mapping reads (RPM), and converted to bigWig format using bedGraphToBigWig (v4).
RNA-seq: Raw data were demultiplexed and converted to fastq format using bcl2fastq2 (v1.17) Illumina. Fastq files were aligned using STAR (v2.7.3a) with the parameters --runMode alignReads --outFilterType BySJout --outFilterMultimapNmax 100 --outFilterMismatchNoverLmax 0.05 --outSAMmultNmax 1 --outMultimapperOrder Random --outSAMtype BAM SortedByCoordinate --outSAMattributes NH HI NM MD AS nM --outSAMunmapped Within. STAR sam output was converted to bam using samtools (v1.9). Bam files were used to calculate strand-specific coverage using bedtools genomeCoverageBed (v2.27.1), resulting bedgraph files were normalized to 1 million genome mapping reads (RPM), and converted to bigWig format using bedGraphToBigWig (v4).
Genome_build: Spombe.ASM294v2.24
Supplementary_files_format_and_content: bigWig files containing normalized (to 1 mio genome mapping reads per library) genome mapping read counts for visualization in UCSC or IGV. Counts per million (CPM) table for the RNAseq samples as a text file.

source_name: S. pombe cell culture

extract_protocol:
Total RNA was extracted with the MasterPure RNA Purification Kit from epicenter (MPY03100).
Small RNA libraries were prepared with the QIAseq miRNA Library Kit (331505) according to manufacturer’s instructions.

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ nmt1+::ade6-hp+::natMX ssu72D::kanMX


title: 29 HCA.Basal_3h [TRA00038916]

value: Hippocampus


title: 28 HTM.High_3h [TRA00038915]


title: Patient14_adjacent non-tumor


title: Patient9_tumor


title: Patient7_tumor


title: flag.Pcsk9.rep7

value: anti-flag


title: 27 HTM.Low_3h [TRA00038914]


title: Patient7_adjacent non-tumor


title: flag.Pcsk9.rep6


title: 26 HTM.Basal_3h [TRA00038913]


title: sRNAseq, pfs2-F115H

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ nmt1+::ade6-hp+::natMX pfs2-F115H::kanMX


title: Patient10_adjacent non-tumor

value: Adjacent non-tumor


title: Patient6_tumor

value: M


title: flag.Pcsk9.rep5


title: sRNAseq, swd22D

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ nmt1+::ade6-hp+::natMX swd22D::kanMX


title: sRNAseq, yth1-R59C

value: h- leu1-32 ura4-D18 ade6-M210 trp1+::ade6+ nmt1+::ade6-hp+::natMX yth1-R59C


title: Patient6_adjacent non-tumor


title: flag.Pcsk9.rep3


title: sRNAseq, wt

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ nmt1+::ade6-hp+::natMX


title: Patient5_tumor


title: 24 NAC.Low_3h [TRA00038888]

value: male

value: 3 months

value: After 3 h of clamp


title: sRNAseq, dis2D

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ nmt1+::ade6-hp+::natMX dis2D::kanMX


title: input.PBS.rep12


title: sRNAseq, ppn1D

value: h- leu1-32 ura4-D18 ade6-704  trp1+::ade6+ nmt1+::ade6-hp+::natMX ppn1D::kanMX


title: flag.controlgRNA.rep9

value: non-targeting


title: alh-4_ko_mutant_rep3

city: Hong Kong

first: Ho Yi

last: Mak

address: Clear Water Bay

department: Division of Life Science

institute: The Hong Kong University of Science and Technology

data_processing:
Illumina bcl2fastq v2.19 with default parameters was used for basecalling to generate demultiplexed FASTQ files.
To improve mappability of reads, low quality bases were trimmed using Trimmomatic v0.36 (Bolger et al., 2014) with parameters ‘SLIDINGWINDOW:20:20 MINLEN:20’.
Trimmed reads were mapped to mm10 using HISAT2 v2.1.0 (Kim et al., 2015) with default parameters. Unmapped reads and read alignments with a mapping quality below 10 were removed using samtools v1.3 (Li et al., 2009).
After read mapping, StringTie v1.3.3 (Pertea et al., 2015) was used with the default parameters and the Illumina ce10 iGenome annotation GTF file to estimate the expression of annotated transcripts.
Ballgown v2.2.0 (Frazee et al., 2015) was used to calculate gene expression FPKM values and output the gene expression table.
Genome_build: ce10
Supplementary_files_format_and_content: Matrix table with FPKM value for every gene and every sample

organism: Caenorhabditis elegans

source_name: alh-4_ko_mutant_whole body

growth_protocol: C. elegans strains were synchronized, fed with E. coli OP50 and grown to L4 stage on standard Nematode Growth Medium (NGM) plates at 20℃ using standard protocol.

extract_protocol:
Worms were havested freshly and snap freezed with dry ice before being lysed for library preparation
RNA libraries were prepared for sequencing using the Smart-seq2 Protocol adapted for worm RNA-seq (DOI: 10.21769/bioprotoc.2729) with the following modifications: the number of worms for each replicate and the reagent for worm lysis were scaled up by 10 fold and the worms were broken with a mini-pestle in a 1.5ml disposable tube on ice instead of cutting.

treatment_protocol: No treatment

tag: developmental stage

value: Bristol N2 with alh-4 deletion (hj221)

value: Whole body


title: Patient5_adjacent non-tumor

value: hepatocellular carcinoma


title: 22 HCA.High_3h [TRA00038886]

value: High

value: Hippocampus


title: alh-4_ko_mutant_rep1


title: alh-4_ko_mutant_rep2


title: 23 NAC.Basal_3h [TRA00038887]

value: Basal

value: Nucleus accumbens


title: flag.controlgRNA.rep8

value: anti-flag


title: 20 HCA.Basal_3h [TRA00038884]


title: n2_wild_type_rep2

source_name: n2_wild_type_whole body

value: Bristol N2

value: wild type


title: flag.controlgRNA.rep11


title: Patient4_tumor

value: Tumor

value: F


title: n2_wild_type_rep3


title: 21 HCA.Low_3h [TRA00038885]

value: male

value: 3 months

value: Low


title: Patient4_adjacent non-tumor

value: Adjacent non-tumor


title: 19 HTM.High_3h [TRA00038883]

value: After 3 h of clamp

value: Hypothalamus


title: n2_wild_type_rep1

value: Whole body


title: flag.PBS.rep4


title: flag.PBS.rep2


title: Patient3_tumor

value: hepatocellular carcinoma

value: M


title: Patient3_adjacent non-tumor


title: Patient2_tumor


title: 17 NAC.High_3h [TRA00038869]

value: High


title: Patient2_adjacent non-tumor


title: 18 HTM.Low_3h [TRA00038882]


title: T3, biological replicate3

city: Beijing

first: Qing

last: Wei

address: NO.1 Beichen West Road, Chaoyang District

institute: Institute of Microbiology, Chinese Academy of Sciences

description: Gene expression data from TRQ-treated Staphylococcus aureus grown till exponential phase

data_processing:
The raw paired end reads were trimmed and quality controlled by Trimmomatic with default parameters.
Then clean reads were separately aligned to reference genome with orientation mode using HISAT.
To identify DEGs (differential expression genes) between two different samples, the expression level for each transcript was calculated using the fragments per kilobase of read per million mapped reads (RPKM) method. The method Bowtie2 and RSEM were used for differential expression analysis
Genome_build: Staphylococcus aureus COL

organism: Staphylococcus aureus

source_name: Exponential phase bacterial culture of Staphylococcus aureus treated by TRQ

growth_protocol: Bacterial cells were grown for 12 hr at 37C.

extract_protocol:
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
RNA-seq strand-specific libraries were prepared following TruSeq RNA sample preparation Kit from Illumina (San Diego, CA), using 5 μg of total RNA. Briefly, rRNA was removed by RiboZero rRNA removal kit (Epicenter), fragmented using fragmentation buffer. cDNA synthesis, end repair, A-base addition and ligation of the Illumina-indexed adaptors were performed according to Illumina’s protocol. Libraries were then size selected for cDNA target fragments of 200~300 bp on 2% Low Range Ultra Agarose followed by PCR amplified using Phusion DNA polymerase (NEB) for 15 PCR cycles.

treatment_protocol: Bacterial cells were collected and kept on ice to extract RNA.

value: TRQ-treated in lB medium

tag: phase

value: Exponential phase


title: flag.PBS.rep1

value: anti-flag


title: T2, biological replicate2


title: flag.PBS.rep12


title: Input GATA6-/-indFOXA2 Day 4

city: Charleston

first: Stephen

middle: A

last: Duncan

state: SC

address: 173 Ashley Avenue, BSB 6th Floor, Room 657A

department: Regenerative Medicine and Cell Biology

institute: Medical University South Carolina

data_processing:
FASTQC (v0.11.7) of raw sequencing file
Aligned reads to hg19 genome using Bowtie 2 (v.2.2.5)
Removed unaligned reads with Partek Flow software (v.9.0.20.0526)
Peaks identified by MACS2 (v2.1.1)
Genome_build: hg19
Supplementary_files_format_and_content: Bedgraph file generated by MACS2 (v2.1.1)

source_name: K3 Induced pluripotent stem cell

growth_protocol: Pluripotent stem cells were seeded on Geltrex-coated tissue culture plates. After 24 hours, the cells were subjected to hepatic differentiation. Briefly, iPSCs were supplemented with Activin A (100ng/ml), fibroblast growth factor 2 (FGF2 20ng/ml), and bone morphogenetic protein 4 (BMP4, 10ng/ml) for 2 days. Activin A (100ng/ml) alone was supplemented for a further 2 days to commit the culture to definitive endoderm.

extract_protocol:
Cells were cross-linked with 1% formaldehyde for 10 minutes at room temperature. The cross-linking reaction was quenched by adding 125mM glycine and incubating at room temperature for 5 minutes. Chromatin was isolated from cell pellets using MNase enzymatic digestion following the manufacturers protocol (Magnetic CHIP kit, ThermoFisher/Pierce). Briefly, cell nuclei were isolated using a membrane lysis buffer. MNase was then added to the isolated nuclei at optimized concentrations and incubated at 37°C for 15 minutes. The enzymatic reaction was stopped with EDTA and digested chromatin was then extracted by brief sonication. Chromatin isolated from 4 individual differentiations were pooled for each condition. Pooled chromatin was incubated with primary antibodies overnight at 4°C. The following day, protein A/G magnetic beads were added to the sample-antibody mixture and incubated at 4°C for 2 hours. Beads were then washed three times in a low salt wash buffer, and once in a high salt wash buffer, before reverse cross-linking in elution buffer. Eluted samples were subjected to proteinase K digestion and the DNA isolated using DNA-binding columns.
CHIP DNA samples were submitted to Beijing Genomics Institute (BGI), who performed library preparation and sequencing on the BGISEQ-500 platform. 50bp single end reads were generated and provided as FASTQ files.

treatment_protocol: Doxycycline was supplemented to induce FOXA2-3xFLAG cDNA expression in the denoted samples

tag: cell type

value: K3 Induced pluripotent stem cell

tag: genotype

value: GATA6Ex4{delta}1/{delta}2;ind FOXA2

tag: differentiation day

value: Day 4 - Definitive Endoderm

tag: doxycyline treatment

value: treated post day 1


title: T1, biological replicate1

tag: genotype/variation

value: wild type


title: 16 NAC.Low_3h [TRA00038868]

value: male

tag: age

value: 3 months

value: Nucleus accumbens


title: Input GATA6+/+ Day 4

value: untreated

tag: chip antibody


title: 15 NAC.Basal_3h [TRA00038867]

tag: Sex

tag: time point

value: After 3 h of clamp

value: Basal


title: 14 HCA.High_3h [TRA00038866]

tag: dose

tag: tissue

value: Hippocampus


title: Patient1_tumor

tag: disease

value: hepatocellular carcinoma

value: Tumor

tag: gender

value: F


title: 25 NAC.High_3h [TRA00038889]

tag: strain

value: High


title: C3, biological replicate3

description: Gene expression data from Staphylococcus aureus grown till exponential phase

source_name: Exponential phase bacterial culture of Staphylococcus aureus

tag: treatment

value: liquid LB medium

value: Exponential phase


title: Input GATA6-/- Day 2

value: K3 Induced pluripotent stem cell

value: GATA6Ex4{delta}1/{delta}2;ind GATA6

value: Day 2 - Mesendoderm


title: Input GATA6+/+ Day 2


title: Patient1_adjacent non-tumor

value: Adjacent non-tumor


title: C1, biological replicate1

value: wild type


title: VARI068 CRISPR RhoC KO replicate 4

city: Ann Arbor

first: Sofia

middle: D

last: Merajver

state: MI

address: 1500 E. Medical Ct. Dr., 7314 CC

department: Int Med-Hematology/Oncology

institute: University of Michigan

data_processing:
Read data in FASTQ format were quality assessed with FastQC/MultiQC (v.0.11.3) and contamination checked with fastq_screen (v.0.11.1)
Reads were adapter-trimmed using CutAdapt (v.1.8.1) and aligned to the GRCh37 hg19 human genome using Tophat/Bowtie2 (v.2.0.13/v.2.2.1, options –b2_very_sensitive and the default max intron length of 500000).
Raw read counts were extracted for each gene using HTSeq (v.0.6.0).
DESeq2 (v1.14.1), run within the R (v.3.3.3) Bioconductor package (Biobase v.2.34), was used to model differential expression in genes between modeled conditions. The main factors used in the model were cell line and CRISPR knockout status (cRhoC or WT). DESeq2 utilizes generalized linear models for each gene and infers a log2 fold change between conditions using maximum likelihood estimation and (by default) a Wald test for significance. Default parameters for DESeq2 were used, specifying a standardized normal prior on the non-intercept coefficients (betaPrior=TRUE).
Genes were annotated with NCBI Entrez GeneIDs and text descriptions.
Genome_build: hg19
Supplementary_files_format_and_content: raw counts, normalized counts, differential expression measurements
Supplementary_files_format_and_content: kraken2 output

source_name: cell line

growth_protocol: Cell lines were incubated at 37°C. Normal growth media for each cell line was replaced with DMEM for 24 hrs prior to RNA extraction.

extract_protocol:
Extraction performed with RNeasy Plus Mini Kit (Qiagen) following normal protocols.
TruSeq stranded mRNA kit (Illumina), using 520ng of total RNA as input and 13 cycles of PCR.

tag: cell line

value: CRISPR RhoC KO


title: VARI068 CRISPR RhoC KO replicate 3


title: 12 HCA.Basal_3h [TRA00038864]

value: male

value: 3 months


title: VARI068 CRISPR RhoC KO replicate 1


title: VARI068 wild type replicate 4

value: wild-type


title: 11 HTM.High_3h [TRA00038863]

value: After 3 h of clamp

value: Hypothalamus


title: GATA6 GATA6+/+ Day 4

value: Day 4 - Definitive Endoderm

value: untreated

value: GATA6 (Cell Signaling - 5851)


title: VARI068 wild type replicate 3