title: H3Position_HumanPlate2

description: H3Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunodeficient host_Replicate 4


title: H3Position_HumanPlate1

status: Public on Oct 29 2025

type: SRA

city: Zurich

first: Francesc

last: Castro-Giner

address: Otto-Stern-Weg 7

country: Switzerland

institute: ETH Zurich

description: H3Position_HumanPlate1 GFP immunogen_LN_Replicate2

library_source: genomic

data_processing:
Mapping using bowtie2 (v2.3.5.1; parameters: -L 11 -N 1 -I S,1,0.75 --norc), allowing for one mismatch.
Normalized abundance of individual sgRNAs were obtained using MAGeCK (v0.5.9.4).
Assembly: lentiviral libraries encompassing 2215 sgRNAs against mouse immune interactome genes  or 2180 sgRNAs against the homologue human genes
Supplementary files format and content: counts.txt.gz are tab-delimited files containing raw counts for each library
Library strategy: CRISPR-seq

molecule: genomic DNA

organism: Homo sapiens

source_name: dCas9 (i.e. no knockout) - SgRNAs as barcodes for studying loss of clonality

extract_protocol:
All tissue samples were processed through a common DNA extraction protocol, as described (Scheidmann et al., 2022). Briefly, tumor and metastasis-carrying organs were lysed in a FastPrep tissue homogenizer (MP) using dedicated matrix beads-containing disposable tubes (MP) in presence of a DNA extraction buffer (50 mM Tris-Cl pH 8.0, 50 mM EDTA, 1% SDS). The lysate was then digested for at least 16 hours at 65 °C in presence of Proteinase K (Qiagen) and then for 30 minutes at 37 °C in presence of RNAase A (Qiagen). Separation of genomic DNA was carried out by first adding pre-chilled 7.5 M ammonium acetate, followed by isopropanol addition to the resulting supernatant, leading the DNA to precipitate. The pellet was then washed with 70% ethanol and the genomic DNA was resuspended in Tris-EDTA buffer (Ambion) and frozen at -20 °C. Finally, DNA extraction from cultivated cells was carried out using the Dneasy kit (Qiagen). For CRISPR genomics applications, DNA samples were quantified using NanoDrop ND-1000 UV-Vis Spectrophotometer (Thermo Fisher).
Samples were processed to reach a theoretical coverage of 500–1,000× per target. For mouse samples, 10-15 µg of genomic DNA were amplified in a one-step PCR with barcoded primers targeting the pLentiGuide vector backbone (P5 and P7). The primer sequences in the table below highlight an 8 bp barcode position in blue and vector binding site in bold, mapping between the sgRNA scaffold and the U6 promoter. All pPrimers were ordered as desalted Oligos oligos at IDT (Integrated DNA Technologies)(at Integrated DNA Technologies or at Microsynth) Microsynth and resuspended in nuclease-free water. For human samples, 10 µg of gDNA were pre-amplified in a nested PCR with the external primers targeting the huTHY1insert and the upstream portion of the U6 promoter (primer sequences in table below). In the second PCR step, barcoded primers targeting the pLentiguide vector backbone (P5 and P7) were used as internal primers (primer sequences in table above). For mouse samples, PCR conditions were as follows: 100 µl as reaction volume, KAPA HiFi Hot Start Ready Mix (2X; Roche)0.02 U/µl of Q5 polymerase (New England Biosciences), and 25-31 cycles of amplification. For human samples, only 23-27 cycles of amplification were applied in the first step using KAPA HiFi Hot Start Ready Mix (2X; Roche). Since some samples did not yield any clearly detectable product after the first PCR, all products were purified with AMPure XP Beads (Beckman Coulter) or Sera-Mag Select Beads (2X, Cytiva Europa GmbH) and used as input for a second PCR (23-27 cycles) with the same primers that were used for the first PCR. When loading a second PCR, pre-amplified fragments were re-quantified using NanoDrop. To reach the theoretical coverage after sequencing, 200 ng of pre-amplified product were re-amplified using nested barcoded primers (P5 and P7). PCR conditions for the second PCR were as follows: 200 ng of pre-amplified fragments per 25 µl reaction, 0.02 U/µl of Q5 polymerase (New England Biosciences), and 27 cycles of amplification. The amplified product containing the spacer sequence with a size of 202 bp was purified by SPRI bead cleanup and a right-sided size selection to remove the genomic DNA input. Using a dual barcoding system, the final NGS amplicons were multiplexed and sequenced on an Illumina platform with 21 dark cycles (DC) in Read 1 to skip the invariable region of the primer binding side (U6 promoter).

tag: tissue

tag: cell line

tag: cell type

value: tumor

tag: genotype

value: xenograft

tag: treatment

tag: crispr lib

value: Immuno Signalome (2180) sgRNA library

tag: sgrna library

tag: effect type

value: sgRNAs used as barcodes (no KO)

tag: vector

value: lentiGuide-THY

tag: full library_complexity


title: Py2T_KO_rep3 CD8-knockout Py2T Mammary fat pad tumor

library_source: transcriptomic

data_processing:
Trim Galore! (v0.6.6; parameters: –q 20 –length 20)
Mapping using STAR (v 2.5.9a) to mouse (GRCm39)  with splice junctions from the human GENCODE annotation (release M29)
The gene-level expression counts were computed with featureCounts from the subread package (v.2.0.3; parameters: -t exon -g gene_id --minOverlap 10 -Q 10) using the human gene annotations from GENCODE (release 40)
Assembly: GRCm39 and GENCODE release M29 and CAS9 sequence
Supplementary files format and content: counts.txt.gz are tab-delimited files containing raw counts for each library

molecule: total RNA

organism: Mus musculus

source_name: tumor

extract_protocol:
Individual tumor fragments were and homogenized using M-tubes (Miltenyi) and GentleMACS devices (Miltenyi) using the program RNA_001 and total RNA was isolated using the RNeasy kit (Qiagen).
The TruSeq Stranded mRNA (Illumina) was used for library preparation as described. Briefly, total RNA samples (500 ng) were poly-A enriched and then reverse-transcribed into double-stranded cDNA. The cDNA samples were fragmented, end-repaired and adenylated before ligation of TruSeq adapters containing unique dual indices (UDI) for multiplexing. Fragments containing TruSeq adapters on both ends were selectively enriched by PCR. The quality and quantity of the enriched libraries were validated using the Fragment Analyzer (Agilent).

tag: number of_cells

value: bulk

tag: origin

value: allograft

tag: strain

value: FVB

tag: transduction


title: H2Position_HumanPlate1

description: H2Position_HumanPlate1 No immunogen_LN_Replicate4


title: H1Position_HumanPlate2

description: H1Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunocompetent host_Replicate 3

source_name: CRISPR Knockout (by Cas9 transfection) of the 2180 sgRNA library (immune signalome library) through the StealTHY


title: mRNA_SCI_12h_SH_B

city: Vestec

first: Ruslan

last: Klassen

address: Průmyslová 595

department: Institute of Biotechnology

country: Czech Republic

institute: Czech Academy of Science

description: Library name: mRNA_SCI_12h_SH_B

data_processing:
Raw reads were trimmed for adapters and low-quality bases using TrimmomaticSE v0.36. Reads mapping to mtDNA and rRNA were removed with SortMeRNA v2.1. Remaining reads were aligned to the rat genome (Rnor_6.0) using STAR v2.5.2b with default settings. Gene-level counts were obtained with htseq-count in "union" mode using Ensembl annotation Rnor_6.0.93.
Assembly: Rnor 6.0
Supplementary files format and content: mRNA_count_table.txt - Matrix of raw counts with mature miRNA names in rows and samples in columns.

molecule: polyA RNA

organism: Rattus norvegicus

source_name: spinal cord tissue

growth_protocol: Adult male Wistar rats (10 weeks old, 300 ± 30 g) were obtained from the breeding facility of the Physiological Institute of the Academy of Sciences of the Czech Republic, Prague. Animals were housed under standard conditions (12-hour light/dark cycle) with ad libitum access to food and water. All procedures were approved by the Ethics Committee of the Institute of Experimental Medicine CAS and conformed to the Directive 2010/63/EU for animal experiments.

extract_protocol:
Animals were euthanized under deep anesthesia (ketamine 100 mg/kg, xylazine 20 mg/kg). Spinal cords were rapidly dissected, and a 2-cm-long segment centered on the injury epicenter was isolated. Tissue was immediately placed into RNAlater (Thermo Fisher Scientific) and stored at -80°C until RNA extraction. Total RNA was isolated using the AllPrep DNA/RNA/Protein Mini Kit (Qiagen) according to the manufacturer's protocol. RNA quantity and purity were assessed with NanoDrop 2000 (Thermo Fisher) and integrity evaluated using the Fragment Analyzer (Agilent); only samples with RQN >7 were used.
Libraries were prepared from 500 ng total RNA using the QuantSeq 3’ mRNA-Seq Library Prep Kit FWD (Lexogen), including 15 PCR amplification cycles. ERCC spike-ins (0.6 µl of 0.01× dilution) were added to each sample. Libraries were quantified using Qubit 2 fluorometer and Fragment Analyzer and sequenced on the Illumina NextSeq 500 platform using 85 bp single-end reads.

treatment_protocol: Spinal cord injury (SCI) was induced by balloon compression at the T8–T9 spinal level. Under anesthesia (3% isoflurane, buprenorphine 0.1 mg/kg), a 2F Fogarty catheter was inserted epidurally, inflated with 15 μl saline for 5 minutes, and then removed. Sham-operated animals underwent catheter insertion without balloon inflation. Post-surgery, animals received antibiotic (ampicillin 60 mg/kg daily for 5 days) and analgesic (buprenorphine 0.05–0.1 mg/kg as needed). Tissue for RNA extraction was collected at 3 hpi, 12 hpi, 24 hpi, 3 dpi, and 7 dpi, along with corresponding sham groups and intact controls.

value: spinal cord tissue

value: wild type

value: Sham control surgery


title: H10Position_HumanPlate1

description: H10Position_HumanPlate1 dCas9-GFP control_LN_Replicate4

value: NSG


title: Py2T_KO_rep2 CD8-knockout Py2T Mammary fat pad tumor


title: H1Position_HumanPlate1

description: H1Position_HumanPlate1 No immunogen_LN_Replicate2


title: mRNA_SCI_12h_SH_A

description: Library name: mRNA_SCI_12h_SH_A


title: G8Position_HumanPlate1

description: G8Position_HumanPlate1 dCas9-GFP immunogen_Liver_Replicate4


title: G9Position_HumanPlate1

description: G9Position_HumanPlate1 dCas9-GFP control_Liver_Replicate2


title: Py2T_CD8_rep3 CD8-transduced Py2T Mammary fat pad tumor


title: G6Position_HumanPlate1

description: G6Position_HumanPlate1 dCas9-THY1 immunogen_Liver_Replicate4


title: mRNA_SCI_12h_C

description: Library name: mRNA_SCI_12h_C

value: Laminotomy


title: Py2T_CD8_rep1 CD8-transduced Py2T Mammary fat pad tumor


title: mRNA_SCI_12h_D

description: Library name: mRNA_SCI_12h_D


title: G3Position_HumanPlate2

description: G3Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunodeficient host_Replicate 4


title: mRNA_SCI_12h_B

description: Library name: mRNA_SCI_12h_B


title: mRNA_SCI_12h_A

description: Library name: mRNA_SCI_12h_A


title: G5Position_HumanPlate1

description: G5Position_HumanPlate1 dCas9-THY1 immunogen_Liver_Replicate2


title: MVTCas9BROAD_FVB3 Rejected Cas9-BROAD MVT1 Mammary fat pad tumor

value: GFP-transduced


title: G1Position_HumanPlate2

description: G1Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunocompetent host_Replicate 3


title: G4Position_HumanPlate1

description: G4Position_HumanPlate1 GFP immunogen_Liver_Replicate4


title: MVTCas9BROAD_FVB2 Rejected Cas9-BROAD MVT1 Mammary fat pad tumor


title: G2Position_HumanPlate1

description: G2Position_HumanPlate1 No immunogen_Liver_Replicate4


title: mRNA_SCI_12h_SH_C

description: Library name: mRNA_SCI_12h_SH_C


title: G1Position_HumanPlate1

description: G1Position_HumanPlate1 No immunogen_Liver_Replicate2


title: G10Position_HumanPlate1

description: G10Position_HumanPlate1 dCas9-GFP control_Liver_Replicate4


title: G3Position_HumanPlate1

description: G3Position_HumanPlate1 GFP immunogen_Liver_Replicate2


title: Py2T_KO_rep1 CD8-knockout Py2T Mammary fat pad tumor


title: mRNA_SCI_3h_SH_D

description: Library name: mRNA_SCI_3h_SH_D


title: mRNA_SCI_3h_SH_C

description: Library name: mRNA_SCI_3h_SH_C


title: F8Position_HumanPlate1

description: F8Position_HumanPlate1 dCas9-GFP immunogen_Lungs_Replicate4


title: F9Position_HumanPlate1

description: F9Position_HumanPlate1 dCas9-GFP control_Lungs_Replicate2


title: MVT1NSG3 Rejection control MVT1 Mammary fat pad tumor


title: F7Position_HumanPlate1

description: F7Position_HumanPlate1 dCas9-GFP immunogen_Lungs_Replicate2


title: mRNA_SCI_3h_SH_B

description: Library name: mRNA_SCI_3h_SH_B


title: F6Position_HumanPlate1

description: F6Position_HumanPlate1 dCas9-THY1-immunogen_Lungs_Replicate4


title: G7Position_HumanPlate1

description: G7Position_HumanPlate1 dCas9-GFP immunogen_Liver_Replicate2


title: MVTCas9BROAD_FVB1 Rejected Cas9-BROAD MVT1 Mammary fat pad tumor


title: MVT1NSG2 Rejection control MVT1 Mammary fat pad tumor


title: Py2T_CD8_rep2 CD8-transduced Py2T Mammary fat pad tumor


title: MVT1NSG1 Rejection control MVT1 Mammary fat pad tumor


title: F5Position_HumanPlate1

description: F5Position_HumanPlate1 dCas9-THY1-immunogen_Lungs_Replicate2


title: mRNA_SCI_3h_D

description: Library name: mRNA_SCI_3h_D


title: F2Position_HumanPlate2

description: F2Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunocompetent host_Replicate 6


title: F3Position_HumanPlate2

description: F3Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunodeficient host_Replicate 3


title: mRNA_SCI_3h_C

description: Library name: mRNA_SCI_3h_C


title: MVT1_UT_rep2 Untransduced MVT1 Mammary fat pad tumor


title: mRNA_SCI_3h_SH_A

description: Library name: mRNA_SCI_3h_SH_A


title: MVT1_UT_rep3 Untransduced MVT1 Mammary fat pad tumor


title: F3Position_HumanPlate1

description: F3Position_HumanPlate1 GFP immunogen_Lungs_Replicate2


title: F2Position_HumanPlate1

description: F2Position_HumanPlate1 No immunogen_Lungs_Replicate4


title: MVT1_UT_rep1 Untransduced MVT1 Mammary fat pad tumor


title: F1Position_HumanPlate2

description: F1Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunocompetent host_Replicate 2


title: mRNA_SCI_3h_B

description: Library name: mRNA_SCI_3h_B


title: F10Position_HumanPlate1

description: F10Position_HumanPlate1 dCas9-GFP control_Lungs_Replicate4


title: mRNA_SCI_3h_A

description: Library name: mRNA_SCI_3h_A


title: F1Position_HumanPlate1

description: F1Position_HumanPlate1 No immunogen_Lungs_Replicate2


title: E9Position_HumanPlate1

description: E9Position_HumanPlate1 dCas9-GFP control_Tumor_Replicate2


title: MVT1_KO_rep3 CD8-knockout MVT1 Mammary fat pad tumor


title: miRNA_SCI_Ctrl_D

description: Library name: miRNA_SCI_Ctrl_D

library_source: other

data_processing:
Adapters, low-quality reads, and reads <16 bp or >28 bp were removed with cutadapt v1.18. Reads mapping to rRNA and UniVec were filtered out using SortMeRNA v2.1 with Bowtie allowing one mismatch. Reads were mapped to Rnor_6.0 genome using STAR in "end-to-end" mode with a 5% mismatch allowance. miRNA-level quantification was performed using featureCounts from Rsubread v2.0.1, counting only uniquely mapping reads.
Assembly: miRBase v22
Supplementary files format and content: miRNA_count_table.txt - Matrix of raw counts with mature miRNA names in rows and samples in columns.

molecule: other

extract_protocol:
Animals were euthanized under deep anesthesia (ketamine 100 mg/kg, xylazine 20 mg/kg). Spinal cords were rapidly dissected, and a 2-cm-long segment centered on the injury epicenter was isolated. Tissue was immediately placed into RNAlater (Thermo Fisher Scientific) and stored at -80°C until RNA extraction. Total RNA was isolated using the AllPrep DNA/RNA/Protein Mini Kit (Qiagen) according to the manufacturer's protocol. RNA quantity and purity were assessed with NanoDrop 2000 (Thermo Fisher) and integrity evaluated using the Fragment Analyzer (Agilent); only samples with RQN >7 were used.
Libraries were prepared from 300 ng total RNA using the RealSeq-AC miRNA Library Kit for Illumina (Somagenics), with 14 PCR cycles and 0.6× adapter concentration. Libraries were quantified and sequenced similarly to RNA-seq samples.

value: Wistar

tag: Sex

value: male

value: without treatment

tag: sample group


title: E8Position_HumanPlate1

description: E8Position_HumanPlate1 dCas9-GFP immunogen_Tumor_Replicate4

value: tumor


title: MVT1_KO_rep2 CD8-knockout MVT1 Mammary fat pad tumor


title: E6Position_HumanPlate1

description: E6Position_HumanPlate1 dCas9-THY1 immunogen_Tumor_Replicate4

value: xenograft


title: E5Position_HumanPlate1

description: E5Position_HumanPlate1 dCas9-THY1 immunogen_Tumor_Replicate2

value: Immuno Signalome (2180) sgRNA library


title: E7Position_HumanPlate1

description: E7Position_HumanPlate1 dCas9-GFP immunogen_Tumor_Replicate2

value: sgRNAs used as barcodes (no KO)


title: MVT1_KO_rep1 CD8-knockout MVT1 Mammary fat pad tumor


title: E4Position_HumanPlate1

description: E4Position_HumanPlate1 GFP immunogen_Tumor_Replicate4

value: lentiGuide-THY


title: F4Position_HumanPlate1

description: F4Position_HumanPlate1 GFP immunogen_Lungs_Replicate4


title: miRNA_SCI_Ctrl_B

description: Library name: miRNA_SCI_Ctrl_B


title: E3Position_HumanPlate2

description: E3Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunodeficient host_Replicate 3


title: E3Position_HumanPlate1

description: E3Position_HumanPlate1 GFP immunogen_Tumor_Replicate2


title: E2Position_HumanPlate2

description: E2Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunocompetent host_Replicate 6


title: miRNA_SCI_7D_SH_D

description: Library name: miRNA_SCI_7D_SH_D


title: miRNA_SCI_Ctrl_A

description: Library name: miRNA_SCI_Ctrl_A


title: MVT1_CD8_rep1 CD8-transduced MVT1 Mammary fat pad tumor

value: bulk


title: E2Position_HumanPlate1

description: E2Position_HumanPlate1 No immunogen_Tumor_Replicate4


title: E1Position_HumanPlate2

description: E1Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunocompetent host_Replicate 2


title: MVT1_CD8_rep2 CD8-transduced MVT1 Mammary fat pad tumor

value: allograft


title: miRNA_SCI_7D_SH_C

description: Library name: miRNA_SCI_7D_SH_C


title: D9Position_HumanPlate1

description: D9Position_HumanPlate1 dCas9-GFP control_LN_Replicate1


title: E1Position_HumanPlate1

description: E1Position_HumanPlate1 No immunogen_Tumor_Replicate2


title: miRNA_SCI_Ctrl_C

description: Library name: miRNA_SCI_Ctrl_C


title: E10Position_HumanPlate1

description: E10Position_HumanPlate1 dCas9-GFP control_Tumor_Replicate4


title: miRNA_SCI_7D_SH_B

description: Library name: miRNA_SCI_7D_SH_B


title: miRNA_SCI_7D_SH_A

description: Library name: miRNA_SCI_7D_SH_A


title: D7Position_HumanPlate1

description: D7Position_HumanPlate1 dCas9-GFP immunogen_LN_Replicate1


title: D8Position_HumanPlate1

description: D8Position_HumanPlate1 dCas9-GFP immunogen_LN_Replicate3


title: 4T1NSG1 Rejection control 4T1 Mammary fat pad tumor


title: 4T1NSG3 Rejection control 4T1 Mammary fat pad tumor


title: D5Position_HumanPlate1

description: D5Position_HumanPlate1 dCas9-THY1 immunogen_LN_Replicate1


title: miRNA_SCI_7D_C

description: Library name: miRNA_SCI_7D_C


title: 4T1NSG2 Rejection control 4T1 Mammary fat pad tumor


title: D6Position_HumanPlate1

description: D6Position_HumanPlate1 dCas9-THY1 immunogen_LN_Replicate3


title: D4Position_HumanPlate1

description: D4Position_HumanPlate1 GFP immunogen_LN_Replicate3


title: 4T1Cas9BROAD_BLB3 Rejected Cas9-BROAD 4T1 Mammary fat pad tumor

value: BALBc


title: D4Position_HumanPlate2

description: D4Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunodeficient host_Replicate 6


title: miRNA_SCI_7D_B

description: Library name: miRNA_SCI_7D_B

value: spinal cord tissue


title: MVT1_CD8_rep3 CD8-transduced MVT1 Mammary fat pad tumor

value: FVB


title: D3Position_HumanPlate2

description: D3Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunodeficient host_Replicate 2


title: miRNA_SCI_7D_A

description: Library name: miRNA_SCI_7D_A

value: wild type


title: 4T1_UT_rep3 Untransduced 4T1 Mammary fat pad tumor


title: miRNA_SCI_3D_SH_B

description: Library name: miRNA_SCI_3D_SH_B

value: Sham control surgery


title: D10Position_HumanPlate1

description: D10Position_HumanPlate1 dCas9-GFP control_LN_Replicate3

value: NSG


title: 4T1_UT_rep2 Untransduced 4T1 Mammary fat pad tumor


title: miRNA_SCI_3D_SH_A

description: Library name: miRNA_SCI_3D_SH_A


title: C8Position_HumanPlate1

description: C8Position_HumanPlate1 dCas9-GFP immunogen_Liver_Replicate3


title: C6Position_HumanPlate1

description: C6Position_HumanPlate1 dCas9-THY1 immunogen_Liver_Replicate3


title: C7Position_HumanPlate1

description: C7Position_HumanPlate1 dCas9-GFP immunogen_Liver_Replicate1


title: C9Position_HumanPlate1

description: C9Position_HumanPlate1 dCas9-GFP control_Liver_Replicate1


title: D1Position_HumanPlate1

description: D1Position_HumanPlate1 No immunogen_LN_Replicate1


title: miRNA_SCI_3D_D

description: Library name: miRNA_SCI_3D_D

value: Laminotomy


title: C4Position_HumanPlate2

description: C4Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunodeficient host_Replicate 6


title: C5Position_HumanPlate1

description: C5Position_HumanPlate1 dCas9-THY1 immunogen_Liver_Replicate1


title: 4T1_KO_rep3 CD8-knockout 4T1 Mammary fat pad tumor


title: C3Position_HumanPlate2

description: C3Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunodeficient host_Replicate 2


title: 4T1Cas9BROAD_BLB2 Rejected Cas9-BROAD 4T1 Mammary fat pad tumor

value: GFP-transduced


title: 4T1_KO_rep2 CD8-knockout 4T1 Mammary fat pad tumor


title: D3Position_HumanPlate1

description: D3Position_HumanPlate1 GFP immunogen_LN_Replicate1


title: C3Position_HumanPlate1

description: C3Position_HumanPlate1 GFP immunogen_Liver_Replicate1


title: miRNA_SCI_3D_C

description: Library name: miRNA_SCI_3D_C

value: Wistar

value: male


title: 4T1_KO_rep1 CD8-knockout 4T1 Mammary fat pad tumor

value: tumor


title: D2Position_HumanPlate1

description: D2Position_HumanPlate1 No immunogen_LN_Replicate3

value: xenograft


title: miRNA_SCI_3D_SH_D

description: Library name: miRNA_SCI_3D_SH_D


title: 4T1Cas9BROAD_BLB1 Rejected Cas9-BROAD 4T1 Mammary fat pad tumor


title: D2Position_HumanPlate2

description: D2Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunocompetent host_Replicate 5

value: Immuno Signalome (2180) sgRNA library


title: D1Position_HumanPlate2

description: D1Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunocompetent host_Replicate 1

value: sgRNAs used as barcodes (no KO)


title: miRNA_SCI_3D_B

description: Library name: miRNA_SCI_3D_B_


title: miRNA_SCI_3D_SH_C

description: Library name: miRNA_SCI_3D_SH_C


title: C2Position_HumanPlate2

description: C2Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunocompetent host_Replicate 5

value: lentiGuide-THY


title: 4T1_UT_rep1 Untransduced 4T1 Mammary fat pad tumor

value: bulk


title: 4T1_CD8_rep3 CD8-transduced 4T1 Mammary fat pad tumor

value: allograft


title: miRNA_SCI_3D_A

description: Library name: miRNA_SCI_3D_A


title: C4Position_HumanPlate1

description: C4Position_HumanPlate1 GFP immunogen_Liver_Replicate3


title: C2Position_HumanPlate1

description: C2Position_HumanPlate1 No immunogen_Liver_Replicate3


title: C1Position_HumanPlate1

description: C1Position_HumanPlate1 No immunogen_Liver_Replicate1


title: C1Position_HumanPlate2

description: C1Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunocompetent host_Replicate 1


title: 4T1_CD8_rep1 CD8-transduced 4T1 Mammary fat pad tumor

value: BALBc


title: miRNA_SCI_24h_SH_D

description: Library name: miRNA_SCI_24h_SH_D

value: spinal cord tissue


title: miRNA_SCI_24h_SH_C

description: Library name: miRNA_SCI_24h_SH_C


title: C10Position_HumanPlate1

description: C10Position_HumanPlate1 dCas9-GFP control_Liver_Replicate3


title: B8Position_HumanPlate1

description: B8Position_HumanPlate1 dCas9-GFP immunogen_Lungs_Replicate3


title: B9Position_HumanPlate1

description: B9Position_HumanPlate1 dCas9-GFP control_Lungs_Replicate1


title: B7Position_HumanPlate1

description: B7Position_HumanPlate1 dCas9-GFP immunogen_Lungs_Replicate1


title: activated B cells Day5_ Cpd10_B

city: New York

first: Prokopios Alexandros

last: Polyzos

address: 413 E 69th St

country: USA

institute: Weill Cornell Medicine

data_processing:
Alignment with STAR aligner (version 2.7.10a)
fliter and assign reads to genes mm10 – (GRCm38.95) with featureCounts (v2.0.6)
Differential analysis was performed with DESeq2 in R4.2.2
Assembly: mm10
Supplementary files format and content: tab-delimited text file with raw counts for all genes

source_name: spleen

growth_protocol: primary naive B cells were maintained in Embryonic Stem cell media (KO-DMEM) in humidified atmosphere with 5% CO2 at 37C and activated for 5 days with 80ng/ml of mIL4 and 5 μg/ml of anti-CD40.

extract_protocol:
RNA was harvested using Rneasy mini plus kit (Qiagen). 1ug of total RNA was used for the construction of sequencing libraries.
RNA libraries for RNA-seq were prepared using NEB Ultra II Directional RNA Library Prep (plus Poly A isolation module) Kit following manufacturer's protocols.

treatment_protocol: DMSO or 1uM of the foxo1 inhibitor (Cpd10) was added to the culture from Day 3 to Day 5 upon replenishment of the media

value: spleen

value: naive B cells

value: Foxo1 inhibition


title: activated B cells Day5_ Cpd10_C


title: B4Position_HumanPlate2

description: B4Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunodeficient host_Replicate 5

value: NSG


title: B5Position_HumanPlate1

description: B5Position_HumanPlate1 dCas9-THY1-immunogen_Lungs_Replicate1


title: miRNA_SCI_24h_SH_B

description: Library name: miRNA_SCI_24h_SH_B

value: wild type

value: Sham control surgery


title: activated B cells Day5_ Cpd10_A


title: miRNA_SCI_24h_SH_A

description: Library name: miRNA_SCI_24h_SH_A

value: Wistar


title: B4Position_HumanPlate1

description: B4Position_HumanPlate1 GFP immunogen_Lungs_Replicate3


title: B3Position_HumanPlate2

description: B3Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunodeficient host_Replicate 1


title: miRNA_SCI_24h_D

description: Library name: miRNA_SCI_24h_D

value: male

value: Laminotomy


title: activated B cells Day5_ DMSO_C

value: control


title: miRNA_SCI_24h_C

description: Library name: miRNA_SCI_24h_C


title: B3Position_HumanPlate1

description: B3Position_HumanPlate1 GFP immunogen_Lungs_Replicate1

value: tumor

value: xenograft

value: Immuno Signalome (2180) sgRNA library


title: B2Position_HumanPlate2

description: B2Position_HumanPlate2 StealTHY CRISPR KO _whole lungs in immunocompetent host_Replicate 4

value: sgRNAs used as barcodes (no KO)

value: lentiGuide-THY


title: activated B cells Day5_ DMSO_B


title: 4T1_CD8_rep2 CD8-transduced 4T1 Mammary fat pad tumor

value: bulk

value: allograft

value: BALBc


title: B2Position_HumanPlate1

description: B2Position_HumanPlate1 No immunogen_Lungs_Replicate3


title: B1Position_HumanPlate2

description: B1Position_HumanPlate2 MDA231 Thy1 pre-Cas9 transfection, preamplified by us

source_name: Thy1 pre-Cas9 transfection

value: In vitro

value: CRISPR KO


title: miRNA_SCI_24h_B

description: Library name: miRNA_SCI_24h_B_

value: spinal cord tissue


title: activated B cells Day5_ DMSO_A

value: spleen

value: naive B cells


title: B1Position_HumanPlate1

description: B1Position_HumanPlate1 No immunogen_Lungs_Replicate1


title: B10Position_HumanPlate1

description: B10Position_HumanPlate1 dCas9-GFP control_Lungs_Replicate3

value: NSG


title: Germinal Center B cells - MycGFP negative GEX

data_processing:
Raw fastq files were processed using cellranger-arc (v2.0.2) with default parameters. Reads were aligned to the mm10 mouse reference. ArchR (Granja et al., 2021) (v1.0.1) was used to perform unbiased clustering of the Myc-GFPneg and Myc-GFPpos sorted cells. Quality control was performed based on the number of unique fragments, fragment size distribution and fraction of reads in peaks (FRIP). Doublets were excluded using the demuxlet function from ArchR (v1.0.1).
Assembly: mm10
Supplementary files format and content: Fragment files: tab-separated values (.tsv)
Supplementary files format and content: Filtered feature matrices: Hierarchical Data Format 5 (HDF5) files (.h5)

source_name: Spleen

extract_protocol:
Splenic B cells from mice immunized with SRBC for 9 days were pre-enriched for B220pos cells by negative magnetic enrichment and stained for GC markers as described above before flow sorting. GC Myc-GFPpos and Myc-GFPneg fractions were sorted from 6 individual 3-month-old male mice and pooled before processing for Multiomic sequencing using the Chromium Next GEM Single Cell Multiome ATAC + Gene Expression kit (10X Genomics #N-1000283) according to the provider specifications. Briefly, two independent nuclei suspensions (one per Myc-GFP fraction) were prepared following the Low Cell Input protocol with confirmed Countess II Live/Dead ratio of <2% with a 3min lysis.
Transposition was performed for 1h at 37°C, after which samples were loaded onto the 10x Genomics Chromium platform to generate barcoded single-cell GEMs, targeting about 10000 nuclei recovered (about 8000 nuclei/μl) per sample. GEM incubation (37 °C for 45 min, 25 °C for 30 min; held at 4 °C) was performed in a C1000 Touch Thermal Cycler with 96-Deep Well Reaction Module (Bio- Rad) for the 10X barcode attachment as well as reverse transcription, template switch and extension. After the RT reaction, GEMs were broken up and the single strand cDNAs were cleaned up with DynaBeads MyOne Silane Beads (ThermoFisher; 37002D). Pre-Amplification (72 °C for 5 min; 98 °C for 3 min, 98 °C for 20 s, 63 °C for 30 s, 72 °C for 1 min for 7 cycles followed by 72 °C for 1 min; held at 4 °C) was performed before proceeding to the ATAC (A) and Gene expression (B) library constructions. (A) A unique sample index for each library was introduced to the pre-amplified tagmented DNA through 7 cycles of PCR amplification using the indexes provided in the kit (98 °C for 45 s; 98 °C for 20 s, 67 °C for 30 s, and 72 °C for 20 s x 7 cycles; 72 °C for 1 min; held at 4 °C). Indexed libraries were subjected to a second double-sided size selection, and libraries were then quantified using Qubit (ThermoFisher, RRID:SCR_020553). The quality was assessed on an Agilent Bioanalyzer 2100 (RRID:SCR_019389), obtaining an average library size of 441 (GC Myc-GFPneg) and 405 (Myc-GFPpos) bp. (B) The GC Myc-GFPneg and Myc-GFPpos cDNAs were amplified for 6 cycles (98 °C for 3 min; 98 °C for 15 s, 63 °C for 20 s, 72 °C for 1 min, 72 °C for 1 min; held at 4 °C). Quality of the cDNAs was assessed using Agilent Bioanalyzer 2100, obtaining an average product size of 1472 and 1503 bp, respectively. These cDNAs were enzymatically fragmented, end repaired, A-tailed, subjected to a double-sided size selection with SPRIselect beads (Beckman Coulter) and ligated to adaptors provided in the kit. A unique sample index for each library was introduced through 15 cycles of PCR amplification using the indexes provided in the kit (98 °C for 45 s; 98 °C for 20 s, 54 °C for 30 s, and 72 °C for 20 s x 14 cycles; 72 °C for 1 min; held at 4 °C). Indexed libraries were subjected to a second double-sided size selection, and libraries were then quantified using Qubit (ThermoFisher, RRID:SCR_020553). The quality was assessed on an Agilent Bioanalyzer 2100 (RRID:SCR_019389), obtaining an average library size of 441 (GC Myc-GFPneg) and 455 bp (GC Myc-GFPpos). Libraries were diluted to 2 nM and clustered on an Illumina NovaSeq 2000 (RRID:SCR_016387) on a pair-end read flow cell and sequenced for: (A) 50 cycles on R1 (10x barcode and the UMIs), followed by 8 cycles of i7 Index and 24 cycles of i5 Index (sample Index) and 49 bases on R2, with a coverage around 200M reads per sample. (B)28 cycles on R1 (10x barcode and the UMIs), followed by 10 cycles of i7 Index and 10 cycles of i5 Index (sample Index) and 90 bases on R2, with a coverage around 200M reads per sample.  Primary processing of sequencing images was done using Illumina’s Real Time Analysis software (RTA).
Other: 10X Multiome kit: https://www.10xgenomics.com/products/single-cell-multiome-atac-plus-gene-expression

value: Spleen

value: Germinal Center B cells


title: B6Position_HumanPlate1

description: B6Position_HumanPlate1 dCas9-THY1-immunogen_Lungs_Replicate3


title: miRNA_SCI_12h_SH_D

description: Library name: miRNA_SCI_12h_SH_D

value: Wistar

value: male

value: wild type

value: Sham control surgery


title: A8Position_HumanPlate1

description: A8Position_HumanPlate1 dCas9-GFP immunogen_Tumor_Replicate3

value: tumor

value: xenograft

value: Immuno Signalome (2180) sgRNA library

value: sgRNAs used as barcodes (no KO)


title: A7Position_HumanPlate1

description: A7Position_HumanPlate1 dCas9-GFP immunogen_Tumor_Replicate1

value: lentiGuide-THY


title: miRNA_SCI_12h_SH_C

description: Library name: miRNA_SCI_12h_SH_C


title: miRNA_SCI_12h_SH_B

description: Library name: miRNA_SCI_12h_SH_B


title: Germinal Center B cells - MycGFP negative ATAC

description: Filter feature matrices contain cell (columns) by feature (rows) counts.


title: A5Position_HumanPlate1

description: A5Position_HumanPlate1 dCas9-THY1 immunogen_Tumor_Replicate1


title: A4Position_HumanPlate2

description: A4Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunodeficient host_Replicate 5


title: A4Position_HumanPlate1

description: A4Position_HumanPlate1 GFP immunogen_Tumor_Replicate3


title: A3Position_HumanPlate2

description: A3Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunodeficient host_Replicate 1


title: N6 [miRNA]

city: Hefei

first: Yaqin

last: Zhang

state: China

country: China

institute: The First Affiliated Hospital of Anhui Medical University

description: miRNA

data_processing:
lncRNAs and mRNAs library construction methods: The sequencing data was filtered with SOAPnuke (v1.5.2) by (1) Removing reads containing sequencing adapter; (2) Removing reads whose low-quality base ratio (base quality less than or equal to 5) is more than 20%; (3) Removing reads whose unknown base ('N' base) ratio is more than 5%, afterwards clean reads were obtained and stored in FASTQ format. The clean reads were mapped to the reference genome using HISAT2 (v2.0.4). After that, Ericscript (v0.5.5) and rMATS (V3.2.5) were used to detect fusion genes and differential splicing genes (DSGs), respectively. Bowtie2 (v2.2.5) was applied to align the clean reads to the gene set, a database built by BGI (Beijing Genomic Institute in ShenZhen), in which known and novel, coding and noncoding transcripts were included, then expression level of gene was calculated by RSEM (v1.2.12). The heatmap was drawn by pheatmap (v1.0.8) according to the gene expression indifferent samples. Essentially, differential expression analysis was performed using theDESeq2 (v1.4.5) with Q value ≤ 0.05. To take insight to the change of phenotype, GO (http://www.geneontology.org/) and KEGG (https://www.kegg.jp/) enrichment analysis of annotated different expression gene was performed by Phyper (https://en.wikipedia.org/wiki/Hypergeometric_distribution) based on Hypergeometric test. The significant levels of terms and pathways were corrected by Q value with a rigorous threshold (Q value ≤ 0.05) by Bonferroni.
miRNAs library construction methods: The raw sequencing data are called raw tags. The raw tags were processed using the following steps: remove low quality tags; remove tags with 5 primer contaminants; remove tags without 3 primer; remove tags without insertion; remove tags with polyA; remove tags shorter than 18 nt. After filtering, the clean tags were mapped to the reference genome and other sRNA database including miRbase, siRNA, piRNA and snoRNA with Bowtie2. Particularly, cmsearch was performed for Rfam mapping. The software miRDeep2 was used to predict novel miRNA by exploring the secondary structure, and Piano was used to predict piRNAs. RNA hybrid, miRanda and TargetScan were used to predict target genes of miRNAs. The small RNA expression level is calculated by counting absolute numbers of molecules using unique molecular identifiers. Differential expression analysis was performed using the DEGseq, Qvalue ≤ 0.001 and the absolute value of Log2 Ratio ≥ 1 as the default threshold to judge the significance of expression difference. To annotate gene functions, all target genes were aligned against the Kyoto Encyclopedia of Genes (KEGG) and Gene Ontology (GO) database. GO enrichment analysis and KEGG enrichment analysis of target genes were performed using phyper, a function of R. The P-value was corrected using the Bonferroni method, and a corrected P-value ≤ 0.05 was taken as a threshold. GO terms or KEGG terms fulfilling this condition were defined as significantly enriched terms.
Assembly: Human: NCBI_GCF_000001405.38_GRCh38.p12
Supplementary files format and content: gene_expression-1.txt: Tab-delimited text file includes FPKM values, TPM values, and read counts for the lncRNA/mRNA samples.
Supplementary files format and content: gene_expression-2.txt: Tab-delimited text file includes FPKM values and read counts for the miRNA samples.

source_name: peripheral blood

extract_protocol:
In the first step, ultra-centrifugation was conducted to isolate exosomes from the plasma samples.
In the second step, total RNA was extracted from the exosomes. The specific steps of the second step are as follows: Total RNA was extracted from the tissues using Trizol (Invitrogen, Carlsbad, CA, USA) according to the manual instructions. For tissue samples, grind about 60mg with liquid nitrogen into powder and transfer the powder samples into the 2ml tube contains 1.5ml Trizol reagent. The mix was centrifuge at 12000×g for 5min at 4°C. The supernatant was transferred to a new 2.0ml tube which was added 0.3 ml of Chloroform/isoamyl alcohol (24:1) per 1.5ml of Trizol reagent. After the mix was centrifuged at 12000×g for 10min at 4°C, the aqueous phase was transferred to a new 1.5mL tube which was add equal volume of supernatant of isopropyl alcohol. The mix was centrifuged at 12000×g for 20min at 4°C and then removed the supernatant. After washed with 1 ml 75% ethanol, the RNA pellet was air-dried in the biosafety cabinet and then dissolved by add 25μL~100μL of DEPC-treated water. Subsequently, total RNA was qualified and quantified using a Nano Drop and Agilent 2100 bioanalyzer (Thermo Fisher Scientific, MA, USA).
Library sequencing was performed with the BGISEQ-500 platform.

tag: disease

value: Normal control

value: peripheral blood

value: serum exosome


title: miRNA_SCI_12h_SH_A

description: Library name: miRNA_SCI_12h_SH_A

tag: tissue

value: spinal cord tissue


title: A2Position_HumanPlate2

description: A2Position_HumanPlate2 StealTHY CRISPR KO _whole tumor in immunocompetent host_Replicate 4

tag: cell line


title: A2Position_HumanPlate1

description: A2Position_HumanPlate1 No immunogen_Tumor_Replicate3

tag: cell type


title: A1Position_HumanPlate1

description: A1Position_HumanPlate1 No immunogen_Tumor_Replicate1

tag: genotype


title: miRNA_SCI_12h_C

description: Library name: miRNA_SCI_12h_C

tag: treatment

value: Laminotomy


title: A10Position_HumanPlate1

description: A10Position_HumanPlate1 dCas9-GFP control_Tumor_Replicate3

value: NSG

tag: crispr lib


title: N4 [miRNA]

description: miRNA


title: miRNA_SCI_12h_B

description: Library name: miRNA_SCI_12h_B


title: N5 [miRNA]

description: miRNA


title: miRNA_SCI_12h_A

description: Library name: miRNA_SCI_12h_A


title: Germinal Center B cells - MycGFP positive GEX

value: Spleen


title: HCT116, auxin 4hr, biological replicate 1, input

city: La Jolla

first: Alon

last: Goren

state: CA

address: 9375 Gilman Dr

department: Department of Medicine

institute: University of California San Diego

description: Library name: HCT116_IAA_4hr_1_input

data_processing:
Samples were sequenced on Illumina NextSeq 550 or Nextseq 2000, samples were demultiplexed with Illumina bcl2fastq.
Adapters were removed with Trimmomatic, reads with Phred score <26 over a 4bp window were truncated.
Reads were aligned to a concatenated genome containing hg38 + sacCer3 + dm6, alignment files were separated by species. Duplicate reads were removed with samtools markdup.
HOMER tag directories were made; by default read counts for each species were filtered for primary alignments with MAPQ > 10, reads were extended to model approximate ChIP-seq fragment size using the option -fragLength 150. Samples were converted to bed files for normalization. Details of spike-in normalization are described in Supplement, normalized data are in bedGraph format.
Read normalized bigwig files were created with HOMER makeBigWig.pl from tag directories.
Assembly: hg38, dm6, sacCer3
Supplementary files format and content: Read-normalized BigWig files for hg38 genome

growth_protocol: HeLa-S3 cells were grown in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% fetal bovine serum (FBS), 1x penicillin-streptomycin, L-glutamine, and 110mg/L sodium pyruvate. Cells were incubated at 37°C and 5% CO2. HCT116-EF1a-OsTIR1-Rpb1 cells were cultured in McCoy’s 5A media (Thermo Scientific 16600082) with 10% FBS and 1x antibiotic-antimycotic. Cells were incubated at 37°C and 5% CO2.

extract_protocol:
HCT116 or HeLa-S3 cells were double crosslinked on plates with 2 mM DSG for 30 minutes (Fisher Scientific NC1736108) and 1% formaldehyde (Pierce ThermoFisher PI28906) for 10 minutes. Crosslinking was quenched with 1/20th volume of 2.625 M glycine (EMD Millipore 357002) and 0.5% BSA (NEB B9000S). Cells were washed by resuspending 3 times with 0.5% BSA/PBS + Protease inhibitor (PIC, Roche 11697498001), spinning down at 1200g for 5 minutes at 4°C and removing the supernatant. Washed pellets were snap frozen in dry ice then stored in -80°C. Cells were lysed with LB3 lysis buffer + PIC, then chromatin was sheared on the PIXUL sonicator (Active Motif 53130) for 108 minutes. S2 cells were crosslinked in 1% formaldehyde for 10 minutes at room temperature, then quenched with glycine and 10% BSA to a final concentration of 0.5%, lysed in LB3 buffer, and sonicated on the PIXUL for 60 minutes. S. cerevisiae cultures in log phase were crosslinked with 0.86% formaldehyde for 30 minutes on a room temperature rotating shaker. The cell wall was digested with Zymolase at 37°C for 60 minutes, cells were washed by pelleting 3x at 5000g and washing in PBS + Protease inhibitor, then resuspended in NP-S buffer + PI. Sheared S. cerevisiae chromatin was sonicated on the PIXUL for 108 minutes. All PIXUL sonication was done with settings Pulse (N) = 50, PRF (kHz) = 1.00, Burst Rate (Hz) = 20, only Process Time was varied. Sheared chromatin from each species were clarified by centrifugation at 15000g, for 10 minutes at 4°C, then combined and inputs were taken before immunoprecipitation for each condition, to assess the relative ratios of chromatin. To HeLa-S3 cells, approximately 100k S2 and 500k S. cerevisiae cells were added per immunoprecipitation reaction. To HCT116 cells, approximately 500k S2 and 100k S. cerevisiae cells were added to each immunoprecipitation reaction. To each IP, 10uL of Protein A beads (Thermo Fisher Scientific 10002D), 1% Triton X-100 and 1x PIC were added. 1 million HCT116 cells or HeLa-S3 cells were used per immunoprecipitation reaction.
To each IP, 10uL of Protein A beads (Thermo Fisher Scientific 10002D), 1% Triton X-100 and 1x PIC were added. 1 million HCT116 cells or HeLa-S3 cells were used per immunoprecipitation reaction. Below are the volumes per IP of each antibody used: CST H3K27ac (D5E4): 1uL CST H3K4me3 (C42D8): 1uL CST H3K4me1 (D1A9): 1uL CST H3K36me3 (D5A7): 1uL CST Rpb1 NTD (D8L4Y): 4uL CST H3K9ac (C5B11): 4uL Samples were incubated overnight on a rotator at 4°C. IP Washes were performed: 3x with 180uL of WBI, 3x with 180uL of WBIII, and 2x with 185uL TET, then eluted in 25uL of cold TT Buffer. Input chromatin containing target and spike-in species’ chromatin from each biological condition were diluted 1:62.5x in TT buffer to a final volume of 25uL. Libraries were prepared with the NEB Ultra II Library Prep Kit (NEB E7645L). NEXTflex Dual-Barcode ChIP-seq Adapters (NOVA-514151) were added at 0.25uM or 0.5uM or 0.625uM. Libraries were then de-crosslinked at 55°C for 1hr, then 65°C overnight and cleaned with Speedbeads, 8.6% PEG and 0.8M NaCl and eluted in TT buffer. Next libraries were PCR amplified with 1uM Solexa 1GA and 1GB primers and NEB Ultra II Q5 Mastermix (PCR protocol: 98C for 30’’, then 12 or 13 cycles of 98C for 10’’, 60C for 15’’, 72C for 30’’, then hold at 72C for 2’). A final cleanup was done with Speedbeads, 8.5% PEG and 1M NaCl, washed twice with 80% EtOH, then the final libraries were eluted in 20uL TT. Library size was estimated by running 6uL of eluted library on a 1.8% agarose gel and concentration was quantified with Qubit HS dsDNA kit (Invitrogen Q32854). Libraries were pooled and sequenced on Illumina NextSeq 550 or NextSeq 2000.

treatment_protocol: HeLa-S3 cells were synchronized in G1/S by adding 2 mM thymidine for 16 hr, washed with PBS, released for 3 hr in fresh medium, and pro-metaphase arrested with 10uM S-trityl-L-cysteine (STC) for 12 hr then harvested. HeLa-S3 cells and HCT116 cells were treated with 500nM Triptolide for 1 or 4 hours. HCT116 cells were treated with 500uM auxin for 1 or 4 hours.

tag: spike-in species

value: D. melanogaster, S. cerevisiae

value: human colon cancer

tag: chip antibody

value: Auxin 4hr


title: miRNA_SCI_24h_A

description: Library name: miRNA_SCI_24h_A_

value: Wistar

value: male

value: wild type


title: N2 [miRNA]

description: miRNA


title: HCT116, auxin 1hr, biological replicate 1, input

description: Library name: HCT116_IAA_1hr_1_input

value: Auxin 1hr


title: HCT116, DMSO treated, 0hr, biological replicate 1, input

description: Library name: HCT116_DMSO_0hr_1_input

value: DMSO


title: A6Position_HumanPlate1

description: A6Position_HumanPlate1 dCas9-THY1 immunogen_Tumor_Replicate3

value: tumor

value: xenograft

value: Immuno Signalome (2180) sgRNA library

tag: sgrna library

tag: effect type

value: sgRNAs used as barcodes (no KO)

tag: vector


title: miRNA_SCI_12h_D

description: Library name: miRNA_SCI_12h_D


title: N1 [miRNA]

description: miRNA

value: Normal control

value: peripheral blood

value: serum exosome


title: HCT116, triptolide 4hr, biological replicate 1, Rpb1

description: Library name: HCT116_TRP_4hr_1_Rpb1_1

value: CST Rpb1 NTD (D8L4Y)

value: Triptolide 4hr


title: Germinal Center B cells - MycGFP positive ATAC

value: Spleen

value: Germinal Center B cells


title: miRNA_SCI_3h_SH_C

description: Library name: miRNA_SCI_3h_SH_C

value: spinal cord tissue

value: Sham control surgery


title: HCT116, triptolide 1hr, biological replicate 1, Rpb1

description: Library name: HCT116_TRP_1hr_1_Rpb1_1

value: D. melanogaster, S. cerevisiae

value: human colon cancer

value: Triptolide 1hr


title: A9Position_HumanPlate1

description: A9Position_HumanPlate1 dCas9-GFP control_Tumor_Replicate1

value: NSG

value: CRISPR KO

value: lentiGuide-THY

tag: full library_complexity


title: miRNA_SCI_3h_SH_D

description: Library name: miRNA_SCI_3h_SH_D

tag: strain

value: Wistar

value: male

value: wild type


title: HCT116, auxin 4hr, biological replicate 1, Rpb1

description: Library name: HCT116_IAA_4hr_1_Rpb1_1

value: Auxin 4hr


title: M6 [miRNA]

description: miRNA

value: Primary biliary cholangitis

value: peripheral blood

value: serum exosome


title: miRNA_SCI_3h_SH_B

description: Library name: miRNA_SCI_3h_SH_B


title: HCT116, auxin 1hr, biological replicate 1, Rpb1

description: Library name: HCT116_IAA_1hr_1_Rpb1_1

value: CST Rpb1 NTD (D8L4Y)

value: Auxin 1hr


title: HCT116, triptolide 4hr, biological replicate 1, input

description: Library name: HCT116_TRP_4hr_1_input

value: Triptolide 4hr


title: HCT116, triptolide 1hr, biological replicate 1, input

description: Library name: HCT116_TRP_1hr_1_input


title: N3 [miRNA]

description: miRNA

value: Normal control


title: HCT116, triptolide 4hr, biological replicate 1, H3K36me3

description: Library name: HCT116_TRP_4hr_1_H3K36me3_1

value: D. melanogaster, S. cerevisiae

value: CST H3K36me3 (D5A7)


title: M5 [miRNA]

description: miRNA


title: miRNA_SCI_3h_SH_A

description: Library name: miRNA_SCI_3h_SH_A

value: spinal cord tissue

value: Sham control surgery


title: M4 [miRNA]

description: miRNA


title: HCT116, triptolide 1hr, biological replicate 1, H3K36me3

description: Library name: HCT116_TRP_1hr_1_H3K36me3_1

value: human colon cancer

value: Triptolide 1hr


title: HCT116, auxin 4hr, biological replicate 1, H3K36me3

description: Library name: HCT116_IAA_4hr_1_H3K36me3_1


title: miRNA_SCI_3h_D

description: Library name: miRNA_SCI_3h_D

value: Wistar

value: Laminotomy


title: HCT116, auxin 1hr, biological replicate 1, H3K36me3

description: Library name: HCT116_IAA_1hr_1_H3K36me3_1


title: miRNA_SCI_3h_C

description: Library name: miRNA_SCI_3h_C

value: male

value: wild type


title: M3 [miRNA]

description: miRNA

value: Primary biliary cholangitis


title: HCT116, triptolide 1hr, biological replicate 1, H3K4me1

description: Library name: HCT116_TRP_1hr_1_H3K4me1_1

value: CST H3K4me1 (D1A9)


title: HCT116, DMSO treated, 0hr, biological replicate 1, H3K36me3

description: Library name: HCT116_DMSO_0hr_1_H3K36me3_1

value: DMSO


title: M2 [miRNA]

description: miRNA

value: peripheral blood

value: serum exosome


title: HCT116, triptolide 4hr, biological replicate 1, H3K4me1

description: Library name: HCT116_TRP_4hr_1_H3K4me1_1

value: Triptolide 4hr


title: miRNA_SCI_3h_B

description: Library name: miRNA_SCI_3h_B


title: HCT116, auxin 1hr, biological replicate 1, H3K4me1

description: Library name: HCT116_IAA_1hr_1_H3K4me1_1

value: Auxin 1hr


title: HCT116, DMSO treated, 0hr, biological replicate 1, Rpb1

description: Library name: HCT116_DMSO_0hr_1_Rpb1_1

value: D. melanogaster, S. cerevisiae

value: CST Rpb1 NTD (D8L4Y)


title: HCT116, auxin 4hr, biological replicate 1, H3K4me1

description: Library name: HCT116_IAA_4hr_1_H3K4me1_1

value: Auxin 4hr


title: M1 [miRNA]

description: miRNA


title: HCT116, triptolide 1hr, biological replicate 1, H3K4me3

description: Library name: HCT116_TRP_1hr_1_H3K4me3_1

value: CST H3K4me3 (C42D8)


title: N6 [lncRNA_mRNA]

description: lncRNA and mRNA

value: Normal control


title: HCT116, triptolide 4hr, biological replicate 1, H3K4me3

description: Library name: HCT116_TRP_4hr_1_H3K4me3_1

value: human colon cancer


title: HCT116, DMSO treated, 0hr, biological replicate 1, H3K4me1

description: Library name: HCT116_DMSO_0hr_1_H3K4me1_1


title: N5 [lncRNA_mRNA]

description: lncRNA and mRNA


title: HCT116, auxin 4hr, biological replicate 1, H3K4me3

description: Library name: HCT116_IAA_4hr_1_H3K4me3_1


title: miRNA_SCI_3h_A

description: Library name: miRNA_SCI_3h_A

value: spinal cord tissue

value: Wistar


title: HCT116, auxin 1hr, biological replicate 1, H3K4me3

description: Library name: HCT116_IAA_1hr_1_H3K4me3_1


title: HCT116, DMSO treated, 0hr, biological replicate 1, H3K4me3

description: Library name: HCT116_DMSO_0hr_1_H3K4me3_1


title: C42_MOCK_rep3

city: Rochester

first: LIGUO

last: WANG

state: MN

address: 200 1st St SW

department: Division of Computational Biology

institute: Mayo Clinic

data_processing:
Image data were processed using the Illumina Standard Pipeline.
Base-calling and data filtering processed by the Mayo Clinic Genome Analysis Core (GAC) using the pipeline Cassava 1.7.
Raw sequences were mapped to the human reference genome (GRCh38) using STAR (2.5.2b) with the following parameters:  --limitBAMsortRAM 31532137230 --outSAMtype BAM   Unsorted      --outSAMstrandField intronMotif   --outSAMunmapped Within      --outSAMheaderCommentFile /tmp/7795325.1.ngs-pri/tmp.jiuaN3DG28   --alignIntronMax 200000   --alignMatesGapMax 200000   --alignSJDBoverhangMin 10   --chimSegmentMin 12   --chimJunctionOverhangMin 12 --twopass1readsN 18446744073709551615   --twopassMode Basic.
RseQC ("FPKM_count.py" command) was used to calculate raw fragment count, FPM (fragments per million) and FPKM (fragments per kilobase of exon per million).
DESeq2 was used to identify differentially expressed genes (adjusted pvalue < 0.05).
Assembly: GRCh38/hg38.
Supplementary files format and content: The processed file contains the following columns: chrom, gene_start, gene_end, gene_symbol, gene_size, gene_strand, Frag_count, FPM, and FPKM.

extract_protocol:
C4-2 cells treated with irradiation (6 Gy), enzalutamide (10 μM, 48 h) or combination.Then we used RNA seq data from different treatments for gene expression analysis
RNA libraries for RNA-seq were prepared using SMARTER mRNA-Seq Library Prep Kit following manufacturer's protocols. The SMART-seq Stranded Kit for total RNA generates strand-specific RNA-seq libraries using Takara Bio’s SMART technology which uses reverse transcription with random priming rather than oligo(dT) priming, thus capturing the full transcriptome instead of poly-A fractions

value: MOCK


title: N4 [lncRNA_mRNA]

description: lncRNA and mRNA


title: RNA_WT_Rep2

city: Bethesda

first: Shiv

last: Grewal

state: MD

address: 9000 Rockville Pike

department: National Cancer Institute

institute: National Institutes of Health

description:
Library name: Sample7
RNA_WT_Rep2

data_processing:
Single ended short reads from RNAseq experiments were quality trimmed using fastp (Chen et al. 2018) and aligned to the reference genome using the STAR aligner (Dobin, A. et al., 2013). Wig files, normalized to counts per million mapped reads, were generated by STAR.
Assembly: S. pombe V2 reference with the sequence of the full 40 kb mat region provided on a separate contig (MAT).
Supplementary files format and content: Wig format for stranded RNAseq.

organism: Schizosaccharomyces pombe

source_name: Yeast Whole Cells

growth_protocol: Sample were grown in YEA medium at 30°C.

extract_protocol:
Total RNA was extracted from cells using the MasterPure Yeast RNA Purification Kit (Biosearch Technologies) according to the manufacturer's instructions.
For RNA Seq-Sequencing, libraries were prepared using ScriptSeq v2 RNA-seq library preparation kit (Illumina) or Ultra II directional RNA library prep kit for Illumina (NEB) according to the manufacturer’s protocol and analyzed using an Agilent 2100 BioAnalyzer.

value: mat1Msmt0 ade6-210 ura4-D18 mat2P(Bg-Bs)::ura4+

value: Yeast Whole Cells

tag: growth

value: Growth in YEA medium at 30°C


title: HCT116, triptolide 4hr, biological replicate 1, H3K27ac

description: Library name: HCT116_TRP_4hr_1_H3K27ac_1_

value: CST H3K27ac (D5E4)


title: N3 [lncRNA_mRNA]

description: lncRNA and mRNA

value: peripheral blood


title: HCT116, triptolide 1hr, biological replicate 1, H3K27ac

description: Library name: HCT116_TRP_1hr_1_H3K27ac_1

value: Triptolide 1hr


title: RNA_upf1∆_Rep2

description:
Library name: Sample8
RNA_upf1∆_Rep2

value: mat1Msmt0 leu1-32 ade6-210 ura4-D18 mat2P(Bg-Bs)::ura4+ upf1D::KanMX6


title: C42_MOCK_rep2


title: HCT116, auxin 4hr, biological replicate 1, H3K27ac

description: Library name: HCT116_IAA_4hr_1_H3K27ac_1


title: C42_MOCK_rep1


title: N2 [lncRNA_mRNA]

description: lncRNA and mRNA

value: serum exosome


title: HCT116, auxin 1hr, biological replicate 1, H3K27ac

description: Library name: HCT116_IAA_1hr_1_H3K27ac_1

value: D. melanogaster, S. cerevisiae

value: Auxin 1hr


title: HCT116, DMSO treated, 0hr, biological replicate 1, H3K27ac

description: Library name: HCT116_DMSO_0hr_1_H3K27ac_1

value: DMSO


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 2, input

description: Library name: HeLa-S3_TRP_4hr_2_input_1

value: human cerival cancer

value: Triptolide 4hr


title: N1 [lncRNA_mRNA]

description: lncRNA and mRNA

value: Normal control


title: C42_IR_rep3

value: IR


title: M6 [lncRNA_mRNA]

description: lncRNA and mRNA

value: Primary biliary cholangitis


title: C42_IR_rep2


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 2, input

description: Library name: HeLa-S3_TRP_1hr_2_input_1


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 1, input

description: Library name: HeLa-S3_TRP_1hr_1_input_1


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 2, input

description: Library name: HeLa-S3_TRP_0hr_2_input_1

value: Triptolide 0hr


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 1, input

description: Library name: HeLa-S3_TRP_4hr_1_input_1


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 1, input

description: Library name: HeLa-S3_TRP_0hr_1_input_1


title: M5 [lncRNA_mRNA]

description: lncRNA and mRNA


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 2, Rpb1

description: Library name: HeLa-S3_TRP_4hr_2_Rpb1_1

value: CST Rpb1 NTD (D8L4Y)


title: C42_EZ_rep3

value: EZ


title: C42_IR_rep1


title: M4 [lncRNA_mRNA]

description: lncRNA and mRNA


title: M3 [lncRNA_mRNA]

description: lncRNA and mRNA


title: C42_EZ_rep2


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 1, Rpb1

description: Library name: HeLa-S3_TRP_4hr_1_Rpb1_1


title: M2 [lncRNA_mRNA]

description: lncRNA and mRNA


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 1, Rpb1

description: Library name: HeLa-S3_TRP_1hr_1_Rpb1_1


title: C42_EZ_rep1


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 2, Rpb1

description: Library name: HeLa-S3_TRP_0hr_2_Rpb1_1


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 2, Rpb1

description: Library name: HeLa-S3_TRP_1hr_2_Rpb1_1


title: M1 [lncRNA_mRNA]

description: lncRNA and mRNA


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 1, Rpb1

description: Library name: HeLa-S3_TRP_0hr_1_Rpb1_1


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 1, H3K4me1

description: Library name: HeLa-S3_TRP_4hr_1_H3K4me1_1

value: CST H3K4me1 (D1A9)


title: C42_EZ_IR_rep3


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_13-4-2

city: xiamen

first: Xiaogang

last: Chen

address: xiangan Campus，Xiamen University

institute: xiamen university

data_processing:
Gene expression was quantified as FPKM using EdgeR (v4.2.1), and differential expression analysis was conducted with DESeq2 (v1.44.0) (FC > 1.5, FDR < 0.05). Normalized counts were obtained via DESeq2’s VST method. GSEA was performed using ClusterProfiler (v4.12.1), analyzing log2FC values against GO, KEGG, Hallmark, and Reactome gene sets. Visualization utilized ComplexHeatmap (v2.20.0) and ggplot2 (v3.5.1) in R (v4.4.1).
Assembly: mm39
Supplementary files format and content: excel flies include RPKM values for each Sample

source_name: placenta

extract_protocol:
Total RNA from placental tissue was extracted using TRIzol reagent(Invitrogen, 15596018)
RNA-seq was performed on three biological replicates each from Adar1f/f and Adar1PKO placentas using the MGI-2000 platform (BGI, China). Paired-end reads were quality-filtered using Trim Galore (v0.6.10) and aligned to the mouse genome (mm39) with STAR (v2.7.9a)

value: placenta

tag: developmental stage


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 2, H3K4me1

description: Library name: HeLa-S3_TRP_1hr_2_H3K4me1_1


title: C42_EZ_IR_rep2


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_13-4-1


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 1, H3K4me1

description: Library name: HeLa-S3_TRP_1hr_1_H3K4me1_1


title: C42_EZ_IR_rep1


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 2, H3K4me1

description: Library name: HeLa-S3_TRP_0hr_2_H3K4me1_1


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_13-16-10


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 1, H3K4me1

description: Library name: HeLa-S3_TRP_0hr_1_H3K4me1_1


title: Group II Intron, 1 Nucleotide Deletion, unmodified

city: Chapel Hill

first: David

last: Qiu

state: NC

address: 104 Airport Dr

department: Chemistry

institute: UNC Chapel Hill

description: Library name: GroupII_1NtDel_Control

data_processing:
ShapeMapper 2.1.5
DanceMapper
Supplementary files format and content: Files with format _profile.txt are from ShapeMapper v2.1.5. Each row in these tables describes per-nucleotide mutation rates and normalized reactivity profiles
Supplementary files format and content: Files with format_DanceMap.txt are reactivity files from DanceMapper. Each row in these tables describes the modelled per-nucleotide reactivity for each population in an ensemble.

organism: Oceanobacillus iheyensis

source_name: bacteria

extract_protocol:
RNA was generated by in vitro transcription with T7 polymerase
MaP-RT was performed using targeted primers. Two rounds of PCR were performed to add Illumina adapter sequences and barcodes.

value: bacteria

value: 1 Nucleotide Deletion

value: Control


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 2, H3K4me3

description: Library name: HeLa-S3_TRP_4hr_2_H3K4me3_1

value: CST H3K4me3 (C42D8)


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 1, H3K4me3

description: Library name: HeLa-S3_TRP_4hr_1_H3K4me3_1

value: D. melanogaster, S. cerevisiae


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_13-16-1


title: Group II Intron, 1 Nucleotide Deletion, TMO modified

description: Library name: GroupII_1NtDel_TMO

value: TMO


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 2, H3K4me3

description: Library name: HeLa-S3_TRP_1hr_2_H3K4me3_1

value: Triptolide 1hr


title: Group II Intron, 3 Nucleotide Deletion, unmodified

description: Library name: GroupII_3NtDel_Control

value: 3 Nucleotide Deletion


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 1, H3K4me3

description: Library name: HeLa-S3_TRP_1hr_1_H3K4me3_1

value: human cerival cancer


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 1, H3K4me3

description: Library name: HeLa-S3_TRP_0hr_1_H3K4me3_1


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_13-1-5


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 2, H3K4me1

description: Library name: HeLa-S3_TRP_4hr_2_H3K4me1_1

value: Triptolide 4hr


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 2, H3K4me3

description: Library name: HeLa-S3_TRP_0hr_2_H3K4me3_1


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_13-1-3


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 2, H3K27ac

description: Library name: HeLa-S3_TRP_4hr_2_H3K27ac_1

value: CST H3K27ac (D5E4)


title: HeLa-S3, Triptolide treated, 4hr, biological replicate 1, H3K27ac

description: Library name: HeLa-S3_TRP_4hr_1_H3K27ac_1


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 2, H3K27ac

description: Library name: HeLa-S3_TRP_1hr_2_H3K27ac_1


title: Group II Intron, U Substitution, unmodified

description: Library name: GroupII_USub_Control

value: U Substitution


title: HeLa-S3, Triptolide treated, 1hr, biological replicate 1, H3K27ac

description: Library name: HeLa-S3_TRP_1hr_1_H3K27ac_1


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_10-6-6


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 2, H3K27ac

description: Library name: HeLa-S3_TRP_0hr_2_H3K27ac_1

value: Triptolide 0hr


title: RNA-Seq of mus musculus:Adar1 ko placentaE14.5_10-6-4


title: Group II Intron, 3 Nucleotide Deletion, TMO modified

description: Library name: GroupII_3NtDel_TMO


title: Group II Intron, U Substitution, TMO modified

description: Library name: GroupII_USub_TMO


title: RNA-Seq of mus musculus:Adar1 ko placentaE13.5_13-6-4


title: HeLa-S3, Triptolide treated, 0hr, biological replicate 1, H3K27ac

description: Library name: HeLa-S3_TRP_0hr_1_H3K27ac_1


title: RNA-Seq of mus musculus:Adar1 ko placentaE13.5_13-4-6


title: Group II Intron, native, TMO modified

description: Library name: GroupII_Native_TMO

value: Native


title: Group II Intron, native, unmodified

description: Library name: GroupII_Native_Control


title: RNA-Seq of mus musculus:Adar1 ko placentaE13.5_13-6-3


title: Group II Intron, native, DMS modified

description: Library name: GroupII_Native_DMS

value: DMS


title: control, replicate biosample B3

type: RNA

city: Shenyang

first: Qingbin

last: Hou

address: wenhua road 83

institute: General Hospital of Northern Theater Command

description:
replicate biosample 3
Agilent Mouse miRNA Microarray Kit,Release 21.0,8x60K(DesignID:070155)

hyb_protocol: 1.5 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) for 17 hours at 65Â°C in a rotating Agilent hybridization oven.  After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37Â°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.

scan_protocol: After washing, the arrays were scanned with the Agilent Scanner G2505C (Agilent Technologies).

data_processing: Feature Extraction software (version 10.7.1.1, Agilent Technologies) was used to process the original image to extract the original data. Quantile standardization is then performed on the raw data. The normalized data were filtered, with at least one set of 100% Detected probes left for follow-up analysis in each set of samples for comparison. The P value and fold change value of t-test were used to screen differential genes. The screening criteria were up-regulated or down-regulated fold change value > = 2.0 and p value < = 0.05. At the same time, scatter plot was used to screen the difference results of each group, volcano maps (for groups with biological duplications) and clustering maps. Next, target gene prediction was performed on the differential mirnas, and the predicted target genes were subjected to GO and KEGG enrichment analysis to determine the biological functions or pathways that the differential mirnas might affect

source_name: control

label_protocol: Cyanine-3 (Cy3) labeled cRNA was prepared from 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA).  Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.

growth_protocol: The abdominal aorta was harvested 28 days later

extract_protocol: RNA was prepared using the Versagene  RNA Purification kit (Gentra Systems, Minneapolis MN) following the manufacturer's recommendations.  The protocol includes differential lysis of red and white blood cells, and an on-column DNase digestion.  Globin message was further reduced using GLOBINclear (Ambion Inc., Austin, TX) to specifically remove both a- and b- globin. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).

treatment_protocol: 8-week old Male ApoE-/- mice were infused with saline (0.9% sodium chloride) or Ang II (1,000 ng/kg/min, APEBIO) via an osmotic pump (Alzet model 1004; AlzaCorp) for 28 days. Normal saline (NS) was used as a vehicle control. Micropumps were embedded under the neck of the mice.

value: abdominal aorta

tag: Sex

value: male

tag: group

value: control


title: control, replicate biosample B2

description:
replicate biosample 2
Agilent Mouse miRNA Microarray Kit,Release 21.0,8x60K(DesignID:070155)

tag: tissue


title: RNA-Seq of mus musculus:Adar1 ko placentaE13.5_13-4-5

value: placenta


title: control, replicate biosample B1

description:
replicate biosample 1
Agilent Mouse miRNA Microarray Kit,Release 21.0,8x60K(DesignID:070155)


title: abdominal aortic aneurysm mice, replicate biosample A3

source_name: abdominal aortic aneurysm mice

value: abdominal aortic aneurysm mice


title: RNA-Seq of mus musculus:Adar1 ko placentaE13.5_13-14-6


title: Follicle_8_post_hCG_0hr

city: Piscataway

first: Shuo

last: Xiao

state: NJ

address: 170 Frelinghuysen Rd

department: Department of Pharmacology and Toxicology

institute: Rutgers University

data_processing:
Following sequencing, BCL files were converted to merged, demultiplexed FASTQs. FASTQs were checked for size to ensure a comparable sequencing depth across all samples.
FASTQs mapped to the GRCm38 genome using HISAT2
After alignment, count data were batch corrected using ComBat-seq
TPMs were calculated based on batched-corrected counts
Count matrices were input to Partek Flow software and differential expression was performed using DESeq2 function embeded in Partek
Genome_build: GRCm38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every sample after batch correction
Supplementary_files_format_and_content: Matrix table with TPMs calculated from raw counts
Supplementary_files_format_and_content: Differientially-expressed gene lists generated by Partek DESeq2 (post hCG 0 hour as the baseline)

source_name: Ovarian follicle (in vitro cultured)

growth_protocol: Follicles within alginate beads were cultured using 96-well plates, with each well containing one follicle and 100 μl growth media. The growth media contains 50% αMEM Glutamax (Sigma-Aldrich), 50% F-12 Glutamax (Sigma-Aldrich), 3 mg/ml bovine serum albumin (BSA; Sigma-Aldrich), 10 mIU/ml recombinant follicle stimulating hormone (rFSH; from A. F. Parlow, National Hormone and Peptide Program, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, MD), 1 mg/ml bovine fetuin (Sigma-Aldrich), and 5 μg/ml ITS (Sigma-Aldrich). Follicles were cultured for 8 days at 37 °C in 5% CO2. Half of the growth media was replaced every other day.

extract_protocol:
Purify RNA from snap-frozen samples using RNA-SPRI beads (doi: 10.1002/0471142727.mb0422s107)
Smart-Seq2 cDNA synthesis was performed on isolated follicle RNA with Maxima H Minus reverse transcriptase (ThermoFisher). Whole transcriptome amplified (WTA) product was purified with AMPure XP beads (Beckman Coulter), quantified with Quanit-iT PicoGreen dsDNA Assay (ThermoFisher), normalized and used for preparing paired-end sequencing libraries with Nextera XT (Illumina, #FC-131) according to the manufacturer’s instructions.

treatment_protocol: On day 8 of eVIFG, grown antral follicles were removed from alginate beads by incubating alginate beads in L15 media containing 1% FBS and 10 IU/ml alginate lyase from Flavobacterium multivorum (Sigma-Aldrich) at 37 °C for 20 minutes. Follicles were then stimulated for ovulation by incubating with maturation media, which contains 50% αMEM Glutamax, 50% F-12 Glutamax, 10%FBS, 1.5 IU/ml hCG (Sigma-Aldrich], 10 ng/ml epidermal growth factor (EGF; BD Biosciences), and 10 mIU/ml rFSH. At 0, 1, 4, and 8 hours post-hCG treatment,  follicles were washed in 1X phosphate buffered saline (PBS; Invitrogen) for three times and were snap-frozen.

value: Ovarian follicle (in vitro cultured)

tag: batch


title: Follicle_6_post_hCG_0hr


title: Follicle_9_post_hCG_0hr


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Pmu dcas9-FLAG+Pmu sgRNA + 0.1mM IPTG

city: Ann Arbor

first: P. Lydia

last: Freddolino

state: MI

address: 1150 W. Medical Center Dr.

department: Biological Chemistry

institute: University of Michigan

data_processing:
*library strategy: Primer-amplified DNA sequencing
Read processing: Sequencing reads were assessed for quality with fastQC (version 0.11.8) and multiQC (version 0.92) to ensure the general quality of the datasets. For the newly acquired spacer sequences, the Read 1 sequences of the paired-end sequencing reads were trimmed to extract new spacer instances using Cutadapt (version 2.6) by identifying the flanking Repeat 26 sequences (attgtagcactgcgaaatgagaaagggagctacaac).
Deduplication: For samples with UMIs, the Read 2 sequences of the paired-end reads were processed with Cutadapt to extract the 8-nt UMI sequences by trimming known construct sequences. The newly spacer-UMI sequence combinations were paired with fastq-pair (version 0.4) and processed with UMI-tools (version 1.0.1) for deduplication after alignment. For samples with no UMIs during library construction, no UMI processing was performed (see the next step for an alternative approach to minimize PCR artifacts).
Spacer source identification: The preprocessed spacer sequences were aligned by bowtie2 (version 2.4.1) with the "very-sensitive" preset to the combined alignment reference containing the N.meningitidis 8013 genome sequence (RefSeq accession number NC_017501.1, assembly: GCF_000026965.1) and the MDA phage genome sequence. For samples with UMIs, aligned spacers were deduplicated using UMI-tools with the directional-adjacency method as noted above. For samples with no UMIs, to handle technical artifacts such as PCR jackpot sites that can over-amplify a subset of spacer sequences, a maximum cap of 5% of the total detected spacer counts on the corresponding Neisseria or phage genome was applied to each unique spacer sequence, i.e., for each unique spacer sequence, if its abundance among total spacers exceeded 5%, its abundance was set to 5% of the total detected spacer count of the given source genome.
Genomic context extraction: Any reads that mapped ambiguously were removed from further analysis using samtools (version 1.9). For each genomic location aligned with the 5' end of any new spacer, the genomic context was extracted as the reference genome sequences flanking the alignment location, including the sequence upstream 10 bp from the alignment sites.
Spacer counting: The numbers of spacers at each unique spacer-aligned location were counted using bedtools2 (version 2.26.0) and custom codes were used to format the genomic context sequences for sequence logo inputs.
Assembly: N.meningitidis 8013 genome sequence (RefSeq accession number NC_017501.1, assembly: GCF_000026965.1) MDA phage genome: J. Meyer et al., Characterization of MDAPhi, a temperate filamentous bacteriophage of Neisseria meningitidis. Microbiology (Reading) 162, 268-282 (2016).
Supplementary files format and content: For *.MDA.fa files: format: FASTA files, with each header formatted as ">MDA_kan:<start on genome>:<end on genome>(<strand>)[<spacer count>](rc)"; content: unique spacer sequences and their genomic contexts, with their source on MDA phage genome and counts listed in the headers
Supplementary files format and content: For *.all.fa files: format: FASTA files, with each header formatted as "><genome>:<start on genome>:<end on genome>(<strand>)[<spacer count>](rc)"; content: unique spacer sequences and their genomic contexts, with their source on either MDA phage genome or Nme genome and counts listed in the headers

organism: Neisseria meningitidis

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Pmu dcas9-FLAG+pTR Pmu sgRNA

growth_protocol: N. meningitidis (aka meningococcal) strains MC58, 8013, and 8013 mutant derivatives used in this study are listed in Supplemental Table 3. These strains were grown on GC Medium Base (GCB) (BD, 228950) plates with appropriate antibiotics and Kellogg’s supplements I and II (22.2 mM glucose [Fisher Scientific, D16-1], 0.68 mM glutamine [Sigma, G3126], 0.45 mM co-carboxylase [aka thiamine pyrophosphate, Sigma, C8754], 1.23 mM Fe(NO3)3 [Sigma, F8508]). Antibiotic concentrations used were 2.5 μg/mL for erythromycin (Erm), 100 μg/mL for kanamycin (Kan), 100 μg/mL for streptomycin (strp), 2.5 μg/mL for chloramphenicol (cm), and 100 μg/mL for spectinomycin (spc). All solid cultures were incubated at 37 ̊C in a 5% CO2 humidified atmosphere.  JM109 (Promega, L2005) and Mach1 (Invitrogen, C862003) chemically competent E. coli cells were used for plasmid cloning. E. coli BL21 (DE3) (Novagen, 69450-3) cells were used for protein purification. E. coli cells were grown at 37 ̊C in LB broth Lennox or on LB agar plates supplemented, when needed, with 50 μg/mL kanamycin, 50 μg/mL carbenicillin, 20 μg/mL for chloramphenicol or 100 μg/mL spectinomycin.

extract_protocol:
To detect spacer acquisition, 10 ng genomic DNA was used as template to PCR amplify CRISPR’s downstream end using Q5 DNA Polymerase with with primers described in Supplemental Table 1. The following PCR conditions were used: 98 ̊C for 30 s; 98 ̊C for 10 s, 60 ̊C for 10 s, 72 ̊C for 20 s (25 cycles); 72 ̊C for 2 min. PCR products were resolved on 2% agarose gel, stained by SYBR Safe (Invitrogen, S33102), and visualized using a Bio-Rad Chemidoc MP imager (12003154). The adapted “+1” “+2” bands are 66- and 132- nts larger than the un-adapted “0” product derived from parental CRISPR array, due to addition of repeat-spacer units (66-nt each). Adaptation efficiency was measured as the percentage of total CRISPR amplicons with new spacers. Quantification of PCR band intensity was done using Image Lab 6.0 software (Bio-Rad).
UMI library construction: Ligated products from above were used as template for 1st step Q5 PCR to amplify the leader-end of CRISPR array with primers OYZ3624+593 containing partial adaptor. Cycling conditions are 98 ̊C for 30 s; 98 ̊C for 10 s, 60 ̊C for 10 s, 72 ̊C for 25 s (25 cycles); 72 ̊C for 2 min. Following 2% agarose gel electrophoresis, the “+1” adapted band was gel extracted with QIAquick Gel Extraction Kit (Qiagen, 28706). Eluate is used as template for 2nd step Q5 PCR amplification using I5 and I7 index primers to add full Illumina adapter and indexes. 2nd PCR conditions are: 98 ̊C for 30 s; 98 ̊C for 10 s, 65 ̊C for 10 s, 72 ̊C for 25 s (15 cycles); 72 ̊C for 2 min.   Non-UMI library construction: For mutant strains carrying the Δcrispr::R26, Δcrispr::crispr(sp1), Δcrispr::crispr(sp1-2), Δcrispr::crispr(sp1-3), Δcrispr::crispr(sp1-5), Δcrispr::crispr(sp1-7), Δcrispr::crispr(sp1-8), Δcrispr::crispr(sp1-17), Δcrispr::crispr(sp1-25), Δcrispr::crispr(sp1-49) alleles, a non-UMI method was used due to the lack of unique restriction site (e.g., FatI) close enough to the newly acquired spacer. 10 ng of total genomic DNA from an adaptation experiment was used as template for 1st step PCR with primers are described in Supplemental Table 1. The following steps for “+1” adapted band gel extraction and 2nd step Q5 PCR were as described in the earlier paragraph.  All final libraries from 2nd step PCR were pooled together, cleaned up using QIAquick PCR Purification Kit, and then purified as one sample by agarose gel electrophoresis and gel extraction. This final pooled library was sequenced on an Illumina sequencing platform, NovaSeq (2x150 bp) at the Univ. of Michigan Advanced Genomics Core.

treatment_protocol: A phage transduction assay was initially performed. Rather than culturing cells on GCB plates with kanamycin, the cells were back-diluted to a concentration of 0.1 OD, and kanamycin was added at a concentration of 40 µg/mL to enrich for transductants. Adaptation assay was performed by incubating diluted culture with or without appropriate inducers, at 37 ̊C in a 5% CO2 humidified atmosphere. Inducer concentrations used were 0.1mM for IPTG (Fisher scientific, BP1755-10). After 24 hrs of induction, genomic DNA was isolated using QIAamp DNA Mini Kit (Qiagen, 51306) per manufacturer’s instructions with the following minor modification. After cell lysis at 56 ̊C for 40 min, RNase A (1mg/mL, Qiagen 19101) treatment was performed at RT for 2 min.

value: strain 8013 (NCBI:txid604162)

tag: cell type

value: bacteria

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Pmu dcas9-FLAG+pTR Pmu sgRNA

tag: treatment -_phage

value: MDAΦ 3+21 hr

tag: treatment -_iptg

value: 0.1 mM IPTG

tag: umi availability

value: Non-UMI

tag: molecule subtype

value: primer-amplified DNA


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Ppn dcas9-FLAG+Ppn sgRNA + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Ppn dcas9-FLAG+pTR Ppn sgRNA

tag: genotype

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Ppn dcas9-FLAG+pTR Ppn sgRNA


title: Follicle_5_post_hCG_0hr

tag: treatment


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Asu dcas9-FLAG+Asu sgRNA + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Asu dcas9-FLAG+pTR Asu sgRNA

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Asu dcas9-FLAG+pTR Asu sgRNA


title: abdominal aortic aneurysm mice, replicate biosample A2


title: Follicle_13_post_hCG_4hr


title: RNA-Seq of mus musculus:E100075418_L01_9177_1

extract_protocol:
Total RNA from placental tissue was extracted using TRIzol reagent(Invitrogen, 15596018)
RNA-seq was performed on three biological replicates each from Adar1f/f Ifnar-/- and Adar1f/f Ifnar-/-Elfcre/wt placentas using the MGI-2000 platform (BGI, China). Paired-end reads were quality-filtered using Trim Galore (v0.6.10) and aligned to the mouse genome (mm39) with STAR (v2.7.9a)

value: Adar1f/f Ifnar-/-


title: Follicle_12_post_hCG_4hr


title: Follicle_11_post_hCG_4hr


title: RNA-Seq of mus musculus:E18.5_6-1_KO_1

value: Adar1f/f Ifnar-/-Elfcre/wt


title: abdominal aortic aneurysm mice, replicate biosample A1

value: abdominal aorta


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Hpa dcas9-FLAG+Hpa sgRNA + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Hpa dcas9-FLAG+pTR Hpa sgRNA

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Hpa dcas9-FLAG+pTR Hpa sgRNA


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nme3 dcas9-FLAG+sgRNA + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme3 dcas9-FLAG+pTR sgRNA

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme3 dcas9-FLAG+pTR sgRNA


title: Follicle_12_post_hCG_1hr


title: RNA-Seq of mus musculus:E100075418_L01_9179_1

value: placenta


title: Follicle_7_post_hCG_0hr


title: RNA-Seq of mus musculus:Adar1 ko placenta E13.5_13-14-2


title: Follicle_3_post_hCG_0hr


title: RNA-Seq of mus musculus:E100075418_L01_9176_1


title: Follicle_2_post_hCG_0hr


title: Follicle_1_post_hCG_0hr


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Pmu dcas9 (D13A+H580A)-FLAG + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Pmu dcas9 (D13A+H580A)-FLAG

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Pmu dcas9 (D13A+H580A)-FLAG


title: Follicle_4_post_hCG_0hr

value: Ovarian follicle (in vitro cultured)


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nme dcas9-FLAG+sgRNA + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme dcas9-FLAG+pTR sgRNA

value: strain 8013 (NCBI:txid604162)

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme dcas9-FLAG+pTR sgRNA


title: Follicle_9_post_hCG_4hrs


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Ppn dcas9 (D13A+H582A) -FLAG + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Ppn dcas9 (D13A+H582A) -FLAG

value: bacteria

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Ppn dcas9 (D13A+H582A) -FLAG


title: Follicle_8_post_hCG_4hrs


title: Follicle_10_post_hCG_4hrs


title: RNA-Seq of mus musculus:E100075418_L01_9178_1


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nme2 dcas9-FLAG+sgRNA + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme2 dcas9-FLAG+pTR sgRNA

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme2 dcas9-FLAG+pTR sgRNA

value: MDAΦ 3+21 hr

value: 0.1 mM IPTG


title: RNA-Seq of mus musculus:E100075418_L01_9180_1


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Asu dcas9 (D10A+H579A)-FLAG + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Asu dcas9 (D10A+H579A)-FLAG

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Asu dcas9 (D10A+H579A)-FLAG

value: Non-UMI

value: primer-amplified DNA


title: Follicle_7_post_hCG_4hrs


title: Follicle_13_post_hCG_1hr


title: Follicle_5_post_hCG_4hrs


title: Follicle_4_post_hCG_4hrs


title: RNA-Seq of mus musculus:E100075418_L01_9170_1

value: Adar1f/f Ifnar-/-


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Hpa dcas9 (D13A+H581A)-FLAG + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Hpa dcas9 (D13A+H581A)-FLAG

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Hpa dcas9 (D13A+H581A)-FLAG


title: RNA-Seq of mus musculus:E100075418_L01_9175_1


title: Follicle_6_post_hCG_4hrs


title: Follicle_11_post_hCG_1hr


title: Follicle_3_post_hCG_4hrs


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nme3 dcas9 (D16A+H588A)-FLAG + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme3 dcas9 (D16A+H588A)-FLAG

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme3 dcas9 (D16A+H588A)-FLAG


title: Follicle_2_post_hCG_4hrs


title: RNA-Seq of mus musculus:E100075418_L01_9169_1

source_name: placenta


title: Follicle_1_post_hCG_4hrs


title: Follicle_10_post_hCG_1hr


title: Follicle_9_post_hCG_1hr


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nme2 dcas9 (D16A+H588A)-FLAG + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme2 dcas9 (D16A+H588A)-FLAG

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme2 dcas9 (D16A+H588A)-FLAG


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nme dcas9 (D16A+H588A)-FLAG + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme dcas9 (D16A+H588A)-FLAG

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nme dcas9 (D16A+H588A)-FLAG


title: RNA-Seq of mus musculus:E100075418_L01_9168_1

value: placenta


title: Follicle_7_post_hCG_1hr

value: Ovarian follicle (in vitro cultured)


title: Follicle_8_post_hCG_1hr


title: RNA-Seq of mus musculus:E100075418_L01_9173_1

value: Adar1f/f Ifnar-/-Elfcre/wt


title: Follicle_5_post_hCG_1hr


title: RNA-Seq of mus musculus:E100075418_L01_9174_1


title: Follicle_6_post_hCG_1hr


title: RNA-Seq of mus musculus:E100075418_L01_9172_1


title: FigS7CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, empty + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::empty

value: strain 8013 (NCBI:txid604162)

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::empty


title: Follicle_3_post_hCG_1hr

source_name: Ovarian follicle (in vitro cultured)


title: FigS6D: endo dcas9, ΔCRISPR(R17-S1), Cas1/2 + 0.1mM IPTG

source_name: endo dcas9, ΔCRISPR(R17-S1), iga-trpB::pLacNmecas1-cas2

value: bacteria

value: endo dcas9, {delta}CRISPR(R17-S1), iga-trpB::pLacNmecas1-cas2

value: UMI-1 (FatI)


title: RNA-Seq of mus musculus:E100075418_L01_9171_1


title: FigS6D: endo dcas9, ΔCRISPR(R24-S2), Cas1/2 + 0.1mM IPTG

source_name: endo dcas9, ΔCRISPR(R24-S2), iga-trpB::pLacNmecas1-cas2

value: endo dcas9, {delta}CRISPR(R24-S2), iga-trpB::pLacNmecas1-cas2

value: MDAΦ 3+21 hr

value: 0.1 mM IPTG


title: Follicle_1_post_hCG_1hr


title: Follicle_4_post_hCG_1hr


title: Input_W

description:
Library name: Sample96
Input_for_samples_89:90:91:92

data_processing:
Single ended fastq format sequences derived from ChIP-seq data were quality-trimmed using fastp (Chen et al. 2018).
Trimmed fastqs were aligned to the S. pombe reference (Wood et al. 2002) using the BWA aligner (Li H. and Durbin R. 2009) and fold enrichment bedgraphs were generated using Macs2 (Zhang Y et al, 2008).
Assembly: S. pombe V2 reference with the sequence of the full 40 kb mat region provided on a separate contig (MAT). Alignments of reads from strains lacking the REII element were to the standard genome as described but with the sequence of the REII element removed.
Supplementary files format and content: For ChIPseq, bedgraphs of Fold Enrichment of the IP over Input (FE) or raw pileups for the Inputs as generated by Macs2 (Zhang Y et al, 2008).

growth_protocol: Sample were grown in YEA medium at 30°C. For induction of gene under nmt1 promoter, cells were grown in PMG medium at 30°C.

extract_protocol:
For the H3K9me3 chromatin immunoprecipitation (ChIP) assay, 5x108 cells were crosslinked with 3% paraformaldehyde (PFA) for 30 minutes at room temperature, followed by quenching with 125 mM glycine. For Swi6 or Raf2 or H3K14ub ChIPs,1x109 cells were incubated for 2 hours at 18°C, fixed in 3% PFA, and washed in ice-cold PBS. Cells were treated with 10mM Dimethyl adipimidate (DMA) at room temperature for 45 minutes. Fixed cells were resuspended in ChIP lysis buffer (50 mM HEPES/KOH pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate) supplemented with 1 mM PMSF and protease inhibitors (Roche cOmplete, Mini Protease Inhibitor Cocktail) and lysed by bead-beating ( Biospec Mini-Beadbeater-16). The lysates were sonicated with a Diagenode Bioruptor for 14-cycles on medium power setting (30 s ON, 30 s OFF) to shear DNA into 0.4–0.6 kb fragments. Cellular debris were removed by centrifugation and 2% supernatants was kept aside for whole cell extract input control. The remaining lysates were pre-cleared with agarose beads for 1 hour at 4°C. After bead removal, lysates were incubated with the appropriate antibody (2–10 μg) overnight at 4°C with slow rotation. Antibodies used includes anti-H3K9me3 (Abcam), anti-c-Myc (Santa Cruz), and home-made affinity-purified anti-Swi6. Pre-washed protein A and protein G agarose beads were added to the lysates and incubated for 4 hours at 4°C. The beads were sequentially washed, twice with Wash Buffer I (50 mM HEPES/KOH pH 7.5, 140 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate), twice with Wash Buffer II (50 mM HEPES/KOH pH 7.5, 500 mM NaCl, 1 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate), twice with Wash Buffer III (10 mM Tris/HCl pH 8.0, 250 mM LiCl, 0.5% IGEPAL, 0.5% sodium deoxycholate, 1 mM EDTA), and once with TE buffer (50 mM Tris/HCl pH 8.0, 10 mM EDTA). The chromatin was eluted from the beads twice with 50 μl of elution buffer (50 mM Tris/HCl pH 8.0, 10 mM EDTA, 1% SDS) at 65°C for 30 minutes each, with shaking (~1,200 rpm). Eluted chromatin and input control samples were reverse crosslinked overnight at 65°C. Samples were treated sequentially with RNase A (10 μg, 2 hours, 37°C) and Proteinase K (20 μg, 2 hours, 37°C), and subsequently purified using the Qiagen PCR purification kit.
Sequencing libraries were prepared using the NEBNext Ultra II DNA Library Prep Kit for Illumina according to the manufacturer’s protocol and analyzed using an Agilent 4200 TapeStation system (Agilent).

value: mat1Msmt0 ade6-210 ura4-D18 mat2P(Bg-Bs)::ura4+

tag: chip antibody

value: Growth in YEA medium at 30°C

tag: fixation

value: 3% PFA


title: Follicle_11_post_hCG_8hrs


title: FigS6D: endo dcas9, ΔCRISPR(R8-S3), Cas1/2 + 0.1mM IPTG

source_name: endo dcas9, ΔCRISPR(R8-S3), iga-trpB::pLacNmecas1-cas2

value: endo dcas9, {delta}CRISPR(R8-S3), iga-trpB::pLacNmecas1-cas2

value: primer-amplified DNA


title: FigS6D: endo dcas9, Cas1/2 + 0.1mM IPTG

source_name: endo dcas9, iga-trpB::pLacNmecas1-cas2

value: endo dcas9, iga-trpB::pLacNmecas1-cas2


title: Input_V

description:
Library name: Sample95
Input_for_samples_79:80:81:82:83:84:85:86:87:88

value: Growth in YEA medium at 30°C + 2hr at 18°C before fixation

value: 3% PFA + 10mM DMA


title: Follicle_10_post_hCG_8hrs

value: Ovarian follicle (in vitro cultured)


title: Follicle_9_post_hCG_8hrs


title: Follicle_8_post_hCG_8hrs


title: Input_U

description:
Library name: Sample94
Input_for_samples_74:75:76:77:78


title: FigS5CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nmecas9+CRISPR Δ40nt leader+tracr + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+CRISPR Δ40nt leader+pTR tracr

value: strain 8013 (NCBI:txid604162)

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+CRISPR {delta}40nt leader+pTR tracr

value: Non-UMI


title: Follicle_7_post_hCG_8hrs


title: Input_T

description:
Library name: Sample93
Input_for_samples_70:71:72:73


title: Follicle_5_post_hCG_8hrs


title: Follicle_6_post_hCG_8hrs


title: FigS5CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nmecas9+sgRNA + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+pTR sgRNA

value: bacteria

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+pTR sgRNA


title: Follicle_4_post_hCG_8hrs


title: H3K9me3_sap49A175V_FE_Exp21_Rep1

description:
Library name: Sample92
H3K9me3_sap49A175V_FE_Exp21_Rep1

value: mat1Msmt0 ade6-210 ura4-DS/E mat2P(Bg-Bs)::ura4+ sap49-A175V

value: Abcam anti-H3K9me3 (cat# ab8898)


title: Follicle_2_post_hCG_1hr


description:
Library name: Sample91
H3K9me3_raf1R576H_sap49A175V_FE_Exp21_Rep1

value: mat1Msmt0 leu1-32 ade6-210 ura4 mat2P(Bg-Bs)::ura4+ raf1R576H::KanMX6 sap49-A175V


title: FigS5CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nmecas9+CRISPR Δ40nt Leader + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+CRISPR Δ40nt Leader

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+CRISPR {delta}40nt Leader

value: MDAΦ 3+21 hr

value: 0.1 mM IPTG


title: FigS5CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nmecas9+tracr + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+pTR tracr

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9+pTR tracr


title: Follicle_3_post_hCG_8hrs


title: Follicle_1_post_hCG_8hrs


title: H3K9me3_raf1R576H_FE_Exp21_Rep5

description:
Library name: Sample90
H3K9me3_raf1R576H_FE_Exp21_Rep5

value: mat1Msmt0 leu1-32 ade6-210 ura4 mat2P(Bg-Bs)::ura4+ raf1R576H::KanMX6

value: Growth in YEA medium at 30°C

value: 3% PFA


title: Follicle_2_post_hCG_8hrs


title: H3K9me3_WT_FE_Exp21_Rep8

description:
Library name: Sample89
H3K9me3_WT_FE_Exp21_Rep8

value: mat1Msmt0 ade6-210 ura4-D18 mat2P(Bg-Bs)::ura4+


title: SMAD2+/P135*(RD11)_D30, RNAseq

city: Boston

first: Jon

last: Seidman

state: Massachusetts

address: 77 Avenue Louis Pasteur

department: Genetics

institute: Harvard Medical School

data_processing:
Libraries were sequenced (Illumina HiSeq 2000 or Illumina HiSeq 2500, 50-base paired-end reads).
All data was combined into a single fastq file
Reads were aligned to the hg38 reference genome using STAR.
Mitochondrial and duplicate reads were discarded using Samtools and Picard’s MarkDuplicates, respectively
The aligned reads were quantitated by Feature counts and counts were normalized.
Assembly: hg38
Supplementary files format and content: bam files and raw counts document for each sample

growth_protocol: Induced pluripotent stem cells (iPSCs) were routinely generated by re-plating 6 non-adherent cell line cultures (typically 5 x 106 cells) on a matrigel coated plate in MTESR medium (Stemcell Technologies)

extract_protocol:
RNA was purified from iPSCs using Trizol (Life Technologies) methods. Purified poly-A RNA that had gone through two rounds of oligo-dT selection was converted into cDNA for construction of libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: 4 undifferentiated IPS samples were differentiated into cardiomyocytes using published cardiac differentiation protocols  (Mummery et al. 2012, Lian et al. 2012)

tag: cell line

value: Induced pluripotent stem cells

value: heterozygous

value: untreated


title: Wildtype_D30_R2, RNAseq

value: wildtype


title: Wildtype_D30_R1, RNAseq


title: FigS5CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, Nmecas9 + 0.1mM IPTG

source_name: ΔtracrΔcas9, ΔCRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9

tag: strain

value: strain 8013 (NCBI:txid604162)

value: bacteria

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::pTet Nmecas9

value: Non-UMI

value: primer-amplified DNA


title: FigS5CD: ΔtracrΔcas9, ΔCRISPR::R26, Cas1/2, empty + 0.1mM IPTG

value: {delta}tracr{delta}cas9, {delta}CRISPR::R26, NICS::pLacNmecas1-cas2, iga-trpB::empty


title: SMAD2+/W274C*(RG12)_R1, RNAseq


title: FigS4FG: ΔCRISPR::R26, Cas1/2 + 0.1mM IPTG

value: MDAΦ 3+21 hr

value: UMI-2 (BsmAI)


title: H3K9me3_ubc4-1_raf1-oe_FE_Exp20_Rep1

description:
Library name: Sample88
H3K9me3_ubc4-1_raf1-oe_FE_Exp20_Rep1

value: mat1Msmt0 leu1-32 ade6 ura4 mat2P(Bg-Bs)::ura4+ ubc4-1 lys1::Padh1-GFP-raf1-HphMX6

value: Abcam anti-H3K9me3 (cat# ab8898)

tag: growth


title: SMAD2+/W274C*(RB1)_R3, RNAseq


title: SMAD2+/R114C*(RF10)_R3, RNAseq


title: H3K9me3_WT_FE_Exp20_Rep7

description:
Library name: Sample86
H3K9me3_WT_FE_Exp20_Rep7

value: Growth in YEA medium at 30°C


title: FigS3EF: endo dcas9, Δtracr, Cas1/2, tracr + 0.1mM IPTG

source_name: endo dcas9, Δtracr, iga-trpB::pLacNmecas1-cas2, NICS::tracr

value: endo dcas9, {delta}tracr, iga-trpB::pLacNmecas1-cas2, NICS::tracr

value: 0.1 mM IPTG

value: UMI-1 (FatI)


title: SMAD2+/R114C*(RF10)_R1, RNAseq

value: Induced pluripotent stem cells

value: heterozygous


title: SMAD2+/R114C*(RA10)_R3, RNAseq

value: untreated


title: SMAD2+/R114C*(RA10)_R2, RNAseq


title: SMAD2+/P135*(RD11)_R3, RNAseq


title: FigS3EF: endo dcas9, Δtracr, Cas1/2 + 0.1mM IPTG

source_name: endo dcas9, Δtracr, iga-trpB::pLacNmecas1-cas2

value: strain 8013 (NCBI:txid604162)

value: endo dcas9, {delta}tracr, iga-trpB::pLacNmecas1-cas2


title: SMAD2+/P135*(RB9)_D30, RNAseq


title: H3K14ub_ubc4-1_FE_Exp19_Rep1

description:
Library name: Sample84
H3K14ub_ubc4-1_FE_Exp19_Rep1

value: mat1Msmt0 leu1-32 ade6 ura4 mat2P(Bg-Bs)::ura4+ ubc4-1

value: anti-H3K14ub (provided by Dr. Jiemin Wong)

value: Growth in YEA medium at 30°C + 2hr at 18°C before fixation

value: 3% PFA + 10mM DMA


title: SMAD2+/P135*(RD11)_R2, RNAseq


title: FigS3EF: endo dcas9, Δtracr, Cas1/2

value: bacteria

value: MDAΦ 3+21 hr

value: primer-amplified DNA


title: SMAD2+/W274C*(RG12)_R3, RNAseq


title: SMAD2P135*/P135*_R1, RNAseq

value: homozygous


title: Wildtype_R4_2M, RNAseq

value: Wildtype


title: SMAD2P135*/P135*_R3, RNAseq