title: CD45+/CX3CR1+ immune cells in NRAS* MBMs in Rela cKO mice (replicate 3)

city: San Juan de Alicante

first: Francisco Javier

last: Rodríguez Baena

state: Alicante

address: Avenida Ramón y Cajal S/N Campus de San Juan

institute: Instituto de Neurociencias CSIC-UMH

description:
Library name: Rela_cKO-3_immune_cells_in_Nras_MBMs
10x Genomics

data_processing:
Cell suspensions from several MBM from 3 different mice per sample (males and females), were subjected to surface antibody staining with anti-CD45-APC (Biolegend 103112) and DAPI for 30 minutes on ice and darkness.
Cells suspensions were subjected to FACS isolation using a gating strategy that consisted in selection of a DAPI negative population (live cells) and double discrimination of singlets and doublets.
CX3CR1-YFP+ and CD45-APC+ cells were sorted into individual tubes per group, 15,000 cells per tube.
Cell integrity and number was assessed by trypan blue and bright-field microscopy in every condition.
Live cells were encapsulated intro droplets and libraries were prepared using 10x Chromium Single Cell Reagents kit v3 single index or Chromium Single Cell Reagents kit v3.1 dual index according to the manufacturer’s protocol (10x Genomics, Inc.; PN-1000269).
Pooled libraries were loaded in a MiSeq Nano with required cycles (28+8+0+91), PF cluster minimal yield of 800M reads reaching a minimum of 88.08 Q30, or a Novaseq 6000 instrument (Illumina) with a S1 flowcell with standard run mode and number of cycles; 28+88+10+10 with a PF cluster minimal yield of 1,300M reads reaching a minimum of 94.14 Q30.
Six 10x-barcoded libraries were sequenced to about 500M/sample (Rela WT replicate 1 (Rela WT1): 505,708,488; Rela WT replicate 2 (Rela WT2): 536,491,838; Rela WT replicate 3 (Rela WT3): 518,092,208; Rela cKO replicate 1 (Rela cKO1): 557,385,410; Rela cKO replicate 2 (Rela cKO2): 481,331,789; Rela cKO replicate 3 (Rela cKO3): 465,771,156).
Raw FASTQ from gene expression libraries were aligned on the prebuilt 10x mouse GRCm38 (GENCODE v32/Ensembl 98) scRNA-seq reference genome (mm10-2020-A) and processed using CellRanger Count with default parameters.
We retrieved 6,030 cells in Rela WT1, 7,662 cells in Rela WT2, 2,718 cells in Rela WT3, 7,509 in Rela cKO1, 7,980 in Rela cKO2 and 6,628 in Rela cKO3.
Sequencing depth was between 60,317 and 190,615 mean read pairs per cell, with sequencing saturation between 50.20% and 85.7%.
Multiplet artifacts were removed using Scrublet v0.2.3 python package58 with default parameters.
Upon doublet removal, we retrieved 5,996 cells in Rela WT1, 7,611 cells in Rela WT2, 2,670 cells in Rela WT3, 7,419 in Rela cKO1, 7,834 in Rela cKO2 and 6,587 in Rela cKO3.
All subsequent single cell transcriptomics analyses were performed using R Statistical Software (v4.1.1) and the Seurat v4 (v4.0.3) package59.
For quality control, genes expressed in fewer than 3 cells were removed.
Cells with unique feature counts (detected genes) over 5,000 or less than 1,400 were excluded.
Finally, cells with more than 5% mitochondrial counts were filtered out.
The remaining cells, 29,745 high-quality cells (4,233 cells in Rela WT1, 6,152 cells in Rela WT2, 1,422 cells in Rela WT3, 5,986 in Rela cKO1, 6,468 in Rela cKO2 and 5,484 in Rela cKO3), were then used for downstream analyses.
For each sample, UMI count data were normalized using a regularized negative binomial regression with SCTransform (sctransform 0.4.1).
SCTransform was executed with the additional flag method="glmGamPoi" to fit a Gamma-Poisson Generalized Linear Model which improves the speed of the learning procedure.
Furthermore, we regressed out mitochondrial content and percentage of Xist and Tsix genes (X-linked genes) in a second non-regularized linear regression.
Next, to ensure that the sctransform residuals for the top variable genes across all samples are computed, we used the functions SelectIntegrationFeatures with nfeatures=3000 (getting the top 3,000 highly variable genes (HVGs) across all the samples) and PrepSCTIntegration (recomputing the residuals for missing values across the previous 3,000 HVGs using the stored model parameters).
Once we have the residuals of the common 3,000 HVGs across the samples, a Principal Component Analysis (PCA) on the SCT-normalized data of each sample.
Next, we used reciprocal PCA (RPCA) dimensional reduction to identify integration anchors with FindIntegrationAnchors ("SCT" as normalization method, 20 neighbors to pick anchors (k.anchor=20) and the rest of the parameters set by default) and then the six datasets were integrated using the pre-computed anchorset and SCT normalized data with IntegrateData function.
Principal Component Analysis was performed on top 3,000 HVGs integrated values to reduce dimensionality of the dataset.
Non-linear dimensional embedding was performed using Uniform Manifold Approximation and Projection (UMAP) on the 30 most significant dimensions of the PCA.
Shared Nearest-Neighbor (SNN) graph was constructed using FindNeighbors function on the 30 most significant principal components (PCs) and cell clusters were identified using FindClusters function.
The RNA assay data was used for the expression plots and related analysis.
Differentially expressed genes between clusters were obtained by Seurat-implemented Wilcoxon rank sum test within FindAllMarkers function with default parameters and clusters were identifed.
A cluster consisting of 129 non-immune cells, which accounted for 0.43% of the total high-quality cells, was excluded from the analysis.
The remaining clusters, comprising a total of 29,616 high-quality cells, were subsequently annotated into five major cell populations: Microglia, Non-Microglial Myeloid cells, NK cells, T cells, and B cells.
Myeloid populations (Microglia and Non-MG Myeloid) were subset and UMAP was recomputed using previous 12 most significant PCs on the remaining 26,685 high-quality cells (3,900 cells in Rela WT1, 5,767 cells in Rela WT2, 1,273 cells in Rela WT3, 5,198 in Rela cKO1, 5,596 in Rela cKO2 and 4,951 in Rela cKO3).
Myeloid cell clusters were identified using FindClusters function and differential expression analysis with FindAllMarkers with default parameters.
This analysis led to the identification of 5 cell populations (Non-MG Myeloid plus 4 microglia subpopulations).
Module enrichment score (AddModuleScore with default parameters) in the myeloid subset was calculated using the top 50 gene signature from each microglia subpopulation, selected from genes detected in over 25% of the population, average log2fc > 0.25, and sorted by ascending adjusted p-value.
Differentially expressed genes comparing Rela cKO vs Rela WT for each myeloid subpopulation were obtained by Seurat-implemented Wilcoxon rank sum tests (FindMarkers function with logfc.threshold=0.1 and min.pct=0.1).
Differences comparing Rela cKO vs Rela WT in top 50 gene signature scores and the expression of HM-like markers, DAM-like, PM and PIM markers in the myeloid cells from MBM were assessed by Mann-Whitney U test (wilcox.test from stats v4.1.1 package (Wilcoxon rank sum test)) with default parameters.
Lymphoid populations (NK cells, T cells, and B cells) were subset and UMAP was recomputed using previous 10 most significant PCs on the remaining 2,931 high-quality cells (332 cells in Rela WT1, 326 cells in Rela WT2, 99 cells in Rela WT3, 788 in Rela cKO1, 870 in Rela cKO2 and 516 in Rela cKO3).
Lymphoid cell clusters were identified using FindClusters function and differential expression analysis with FindAllMarkers with default parameters.
This analysis led to the identification of 5 cell populations (NK cells, NK T cells, CD8 T cells, CD4 T cells, and B cells).
Module enrichment score (AddModuleScore with default parameters) in the lymphoid subset was performed for the Tpex gene signature (Tox, Tcf7, Pdcd1 and Slamf6) and the MHC gene signature (H2-K1, H2-D1, H2-M3, H2-Q4, H2-Q6, H2-Q7, H2-T22, H2-Ab1, H2-Aa, H2-Eb1, H2-Eb2 and H2-Ea).
Inference and analysis of cell-cell communication between myeloid and lymphoid populations was performed with CellChat48, starting from a Seurat object containing the above 29,616 high-quality cells with metadata for the 5 minor lymphoid and 5 minor myeloid populations, with default parameters.
Assembly: GRCm38 (mm10)
Supplementary files format and content: Single cell Seurat object (major populations): RelA-cKO_Nras-MBM_CD45+_CX3CR1+_immune_major_populations.rds
Supplementary files format and content: Single cell Seurat object (myeloid populations): RelA-cKO_Nras-MBM_CD45+_CX3CR1+_immune_myeloid_populations.rds
Supplementary files format and content: Rela WT-1 - Cell barcode: Rela_WT-1_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_barcodes.tsv.gz
Supplementary files format and content: Rela WT-1 - Gene name: Rela_WT-1_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_features.tsv.gz
Supplementary files format and content: Rela WT-1 - Expression matrix: Rela_WT-1_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_matrix.mtx.gz
Supplementary files format and content: Rela WT-2 - Cell barcode: Rela_WT-2_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_barcodes.tsv.gz
Supplementary files format and content: Rela WT-2 - Gene name: Rela_WT-2_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_features.tsv.gz
Supplementary files format and content: Rela WT-2 - Expression matrix: Rela_WT-2_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_matrix.mtx.gz
Supplementary files format and content: Rela WT-3 - Cell barcode: Rela_WT-3_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_barcodes.tsv.gz
Supplementary files format and content: Rela WT-3 - Gene name: Rela_WT-3_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_features.tsv.gz
Supplementary files format and content: Rela WT-3 - Expression matrix: Rela_WT-3_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_matrix.mtx.gz
Supplementary files format and content: Rela cKO-1 - Cell barcode: Rela_cKO-1_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_barcodes.tsv.gz
Supplementary files format and content: Rela cKO-1 - Gene name: Rela_cKO-1_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_features.tsv.gz
Supplementary files format and content: Rela cKO-1 - Expression matrix: Rela_cKO-1_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_matrix.mtx.gz
Supplementary files format and content: Rela cKO-2 - Cell barcode: Rela_cKO-2_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_barcodes.tsv.gz
Supplementary files format and content: Rela cKO-2 - Gene name: Rela_cKO-2_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_features.tsv.gz
Supplementary files format and content: Rela cKO-2 - Expression matrix: Rela_cKO-2_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_matrix.mtx.gz
Supplementary files format and content: Rela cKO-3 - Cell barcode: Rela_cKO-3_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_barcodes.tsv.gz
Supplementary files format and content: Rela cKO-3 - Gene name: Rela_cKO-3_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_features.tsv.gz
Supplementary files format and content: Rela cKO-3 - Expression matrix: Rela_cKO-3_immune_cells_in_Nras_MBMs_filtered_feature_bc_matrix_matrix.mtx.gz

source_name: Brain

growth_protocol: NRASQ61k-1014 cells referred in the text as Nras* and were maintained in HAMS-F12 supplemented with 10% FBS (Sigma) and 1% penicillin/streptomycin (Sigma). Cells were injected via the intracarotid artery of mice, injecting 200k in 100 uL. This protocol was performed in cold to prevent microglia ex vivo activation. Brains were sliced in 1 mm thick cuts and placed under the stereomicroscope, MBM were dissected manually and placed in ice-cold HBSS containing 15 mM HEPES and 0,5% Glucose. Control tissue or MBM were mechanically disrupted with 5X slow strokes with a Dounce tissue grinder (Wheaton, 357542) on ice. Samples were centrifuged at 350g during 5 min at 4°C and later at 8.000 g during 30 sec. Samples were subjected to RBC lysis buffer (Biolegend, 420301) for 3 min at RT and then spined again as described.

extract_protocol:
After MBM sample preparation as described above, cell suspensions were directly subjected to surface antibody staining with anti-CD45-APC (Biolegend 103112) and DAPI for 30 minutes on ice and darkness. Cells suspensions were subjected to FACS isolation using a gating strategy that consisted in selection of a DAPI negative population (live cells) and double discrimination of singlets and doublets. CX3CR1-YFP+ and CD45-APC+ cells were sorted into individual tubes per group, 15.000 cells per tube and later encapsulated.
Live cells were encapsulated intro droplets and libraries were prepared using Chromium Single Cell Reagents kit v3.1 dual index according to the manufacturer’s protocol (10X genomics, PN-1000269).

value: Brain

value: CD45+/CX3CR1+ immune cells

value: Rela cKO

value: No treatment


title: Control-1, snATAC

status: Public on Feb 06 2025

type: SRA

city: Atlanta

first: Hsiao-Lin

middle: V.

last: Wang

state: GA

address: 615 Michael Street,, Whitehead Biomedical Research Building, Room 355,

department: School of Medicine, Department of Human Genetics

country: USA

institute: Emory University

description:
Library name: sample 27
10x Genomics Chromium Single Cell Multiome

library_source: genomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting and aggregation were made using the 10x Genomics Cell Ranger ARC (cellranger-arc-2.0.0)
Assembly: hg38
Supplementary files format and content: Tab-separated values files, matrix files, alignment files, and ATAC-seq fragment files

molecule: genomic DNA

organism: Homo sapiens

source_name: dorsolateral prefrontal cortex

extract_protocol:
Tissue sections were frozen and stored at -80°C. 20 mg frozen tissues were thawed in 1 mL cold homogenization buffer (260 mM sucrose, 30 mM KCl, 10 mM NaCl2, 20 mM Tricine-KOH pH 7.8, 1 mM DTT, 0.5 mM Spermidine, 0.2 mM Spermine, 0.3% NP40, cOmplete Protease inhibitor (Roche), and Ribolock) and homogenized in a pre-chilled Dounce. Cell lysates were passed through a 70 µm Flowmi cell strainer before separation using a discontinuous iodixanol gradient and centrifuge at 1480 g at 4°C for 20 min in a swinging bucket centrifuge with the brake off. The nuclei band located at the interface between 30% and 40% iodixanol was collected and washed in RSB-T wash buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% Tween-20)
Libraries were generated using the 10x Genomics Chromium Single Cell Multiome ATAC + Gene Expression kit following the manufacturer’s instructions, with the following modifications. Per sample, 16,100 nuclei were resuspended in 1x diluted nuclei buffer (10x Genomics) with 2% BSA (Sigma) with a capture target of 10,000 nuclei. ATAC libraries were sequenced to target of 25,000 read-pairs per nucleus and RNA libraries were sequenced to 20,000 read-pairs per nucleus on an Illumina NovaSeq 6000 instrument at the Florida State University Translational Science Laboratory. The matched RNA and ATAC libraries were processed using the 10x Genomics Cell Ranger ARC (cellranger-arc-2.0.0) pipeline with default parameters and aligned to the hg38 human genome assembly (refdata-cellranger-arc-GRCh38-2020-A-2.0.0).

tag: tissue

value: dorsolateral prefrontal cortex

tag: cell type

value: cortical cells

tag: genotype

value: WT

tag: batch


title: RPE_control_veh

city: Utrecht

first: Frank

last: Riemers

address: Uppsalalaan 8

country: Netherlands

institute: Utrecht University

description: SORT-seq

library_source: transcriptomic single cell

data_processing:
Paired end reads obtained by automated CEL-seq2 were aligned to the transcriptome using bwa with default parameters.
The mapped reads were counted using STAR. Reads mapping to multiple loci were discarded.
Read counts were processed and analyzed using the Seurat package
Assembly: hg38
Supplementary files format and content: Tab-separated counts per plate

molecule: total RNA

source_name: RPE

extract_protocol:
Cells were collected by trypsinization, washed with PBS after which cells were fixed with 0.5mM dithiobis(succinimidyl propionate)(DSP) in Sodium Phosphate buffered Saine (pH 8.4) for 45 minutes at room temperature at a concentration of 1 million cells per 2.5ml. Next, DSP was neutralized by incubating the cells with quench buffer (100mM Tris, pH 7.5, 150mM NaCl) for 10 minutes and cell clumps were removed usmg a 70µm cell strainer. Cells were icubated for 30 minutes with BP buffer (PFBB:PBS, 1:1, supplemented with 0.1% Triton•X-100 and 2 U/mL RNAsin plus) to permeabilize the cells, after which samples were incubated overnight with antibodies in BP buffer. Samples were filtered using a 40 um cell strainer and incubated with DAPI (0.2 ug/mL) prior to loading of samples on the flow cytometer.  Cells were sorter into the captuture plates on a BD Influx cell sorter.
Single Cell Discoveries, where single-cell RNA sequencing was performed according to the SORT-seq protocol. Cells were heat-lysed at 65° C followed by cDNA synthesis. After second-strand cDNA synthesis, all the barcoded material from one plate was pooled into one library and amplified using in vitro transcription (IVT). Following amplification, library preparation was done following the CEL-Seq2 protocol to prepare a cDNA library for sequencing using TruSeq small RNA primers (Illumina). The DNA library was paired-end sequenced on an Illumina Nextseq™ 500, high output, with a 1x75 bp Illumina kit

treatment_protocol: 4 conditions were employed in this experiment: RPE_veh_control received no doxycycline and no gemcitabine/prexasertib, RPE_dox_control received 0.2 ug/mL doxycycline for 24 hours and no gemictabine/perxasertib, RPE_veh_gp received no doxycycline and 4 nM gemcitabine/10 nM Prexasertib for 16 hours, and RPE_dox_gp received 0.2 ug/mL doxycycline for 24 hours followed by 4 nM gemcitabine/10 nM prexasertib for 16 hours. REP_veh_control and RPE_dox_control were sorted into one 384 well plate each, while RPE_veh_gp and RPE_dox_gp were sorted into 3 384-well plates each.

tag: cell line

value: RPE

value: TetOn HRAS G12V

tag: treatment


title: TDPhigh-7, snRNA

description:
Library name: sample 25
10x Genomics Chromium Single Cell Multiome

molecule: polyA RNA

value: C9orf72 HRE


title: Liver_PolyA_Slow_Medium_B1

city: Philadelphia

first: Thomas

middle: G

last: Brooks

address: 3400 Civic Center Blvd

department: ITMAT

institute: University of Pennsylvania

description: Library name: Liver_PolyA_Slow_Medium_B1

library_source: transcriptomic

data_processing:
Ribosomal content was removed from all samples computationally (see https://github.com/itmat/normalization). PCR duplicates were removed using umi-tools v1.1.1. Reads were quantified with Salmon v1.9.0 (options: "-l ISF --seqBias --gcBias --posBias --dumpEq") using the GRCm38 genome and Ensembl v102 annotation. Quantifications were aggregated using R v4.4.0 and tximeta v1.22.1.
Assembly: GRCm38, ensembl v102
Supplementary files format and content: Processed file contains gene-level read count estimates of all samples. The format is a tab-separated spreadsheet. The first column gives gene identifiers.

organism: Mus musculus

source_name: Liver

growth_protocol: A wild-type, twelve-week-old male C57BL/6J mouse was bred in-house at the animal facility of the University of Pennsylvania.

extract_protocol:
The mouse  was euthanized through carbon dioxide induced asphyxiation and liver and heart  samples were dissected and snap-frozen in liquid nitrogen. RNA was extracted from the tissue using TRIzol (ThermoFisher Scientific, catalog no. 15596018) and RNeasy Mini Kit (Qiagen, catalog no. 74104), according to manufacturers’ protocols.
Except where noted, we constructed all libraries according to the manufacturer’s protocols and used 13 cycles of PCR during the final library amplification step. To allow us to assess duplication rates with UMI sequences, we used the xGen UDI-UMI adapters (IDT, catalog no. 10005903) for all libraries, diluting the initial 15 μM stock of adapters to 3 μM in NGS adapter buffer (10 mM Tris pH 8.0, 0.1 mM EDTA, 100 mM NaCl).  We used the TruSeq Stranded mRNA library preparation kit (Illumina, catalog no. 20020594) and the TruSeq Stranded Total RNA Human/Mouse/Rat library preparation kit (Illumina, catalog no. 20020596) to prepare the PolyA and rRNA pull-down libraries, respectively. For the rRNA digestion libraries, we used the Illumina Stranded Total RNA Prep Ligation with Ribo-Zero Plus kit (Illumina, catalog no. 20040525). This kit uses a different strategy than the TruSeq kits for ligating indexed adapters to cDNA fragments that is not compatible with the xGen adapters. In order to use the same adapters, and their included UMIs, across all libraries, we modified the protocol for the Illumina Stranded Total RNA Prep Ligation as follows: 1) During the ‘Ligate Adapters’ step, replace the RNA Index Anchors with the diluted xGen adapters, and 2) during the ‘Amplify Library’ step, replace the Index Adapters with the PCR Primer Cocktail (PPC) from the TruSeq Stranded Total RNA kit. We performed all other steps according to the manufacturer’s protocol. All PolyA and ribo-depleted libraries used 200 ng of input RNA.  For the ‘No Select’ libraries, we used 20 ng of total RNA as input into the TruSeq Stranded Total RNA library preparation protocol, skipping the ‘Deplete rRNA’ step and starting with the ‘Fragment and Denature RNA’ step (see supplementary methods for detailed protocol). All remaining steps in the library preparation used reagents from the TruSeq Stranded Total RNA library preparation kit and followed that protocol. Since the majority of reads from the No Select library come from rRNA reads, we prepared and sequenced three No Select libraries from each sample/condition in order to get a sufficiently diverse sampling of non rRNA molecules.  For the experiments where we varied RNA fragmentation, we used the following times for the 94°C incubation during the fragment RNA step: 2 minutes (short), 8 minutes (medium), or 12 minutes (long). Similarly, for experiments where we varied PCR ramp rates, we used the following Thermocycler ramp rate settings during the library amplification step: 3°C/s (fast), 1.5°C/s (medium), or 0.5°C/s (slow).

value: Liver

tag: Sex

value: female

tag: ribosomal depletion

value: PolyA selection

tag: fragment length

value: Medium

tag: pcr ramp_rate

value: Slow


title: CD45+/CX3CR1+ immune cells in NRAS* MBMs in Rela cKO mice (replicate 2)

country: Spain

description:
Library name: Rela_cKO-2_immune_cells_in_Nras_MBMs
10x Genomics


title: TDPhigh-6, snRNA

description:
Library name: sample 24
10x Genomics Chromium Single Cell Multiome


title: DNMT3Amut_Metformin_replicate3

city: Cambridge

first: Muxin

last: Gu

email: muxingu@gmail.com, mg445@cam.ac.uk

address: Jeffrey Cheah Biomedical Centre, Puddicombe Way

department: Haematology

country: United Kingdom

institute: University of Cambridge

data_processing:
RNA-seq reads were mapped to the human genome assembly GRCm38 using STAR version 6.43, tolerating mismatch rate of 0.01 and allowing maximal intron lengths of 50kb. Read counts were calculated by STAR using Ensembl annotation of GRCm38 version 110 and normalized as fragments per kilobase of gene length per million uniquely mapped reads (FPKM). Differential expression analysis was done by DESeq2 using paired sample design and significant genes were identified using adjusted p-values of 0.05 as cut-off.
Assembly: GRCm38
Supplementary files format and content: STAR read counts

source_name: Bone marrow

growth_protocol: Metformin hydrochloride (Sigma, PHR1084) was dissolved in PBS and administrated at dose 125 mg/kg via intraperitoneal injection daily (5 days on, 2 days off) as previously reported. Treatment was performed for 7 weeks in total. For in vitro studies metformin was prepared in sterile water. 125 mg/kg/day dose when translated to human equivalents, corresponds to 10.4 mg/kg/day. Notably, this pharmacological dosage is lower than the levels commonly prescribed for the treatment of type II diabetes.

extract_protocol:
RNA was extracted with RNeasy kit (Qiagen, 74104) with with DNase I treatment (Qiagen, 79254) according to the manufacturer's instructions.
PolyA enrichment was performed with NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490S, New England Biolabs), and library preparation with NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (E7760S, New England Biolabs). NEBNext Multiplex Oligos (E6440S, New England Biolabs) were used as indexes. Starting input of RNA was 1ug. Libraries were sequenced on the Illumina NovaSeqX platform with 50-bp paired-end sequencing.

value: Bone marrow

value: HSC

value: Metformin


title: DNMT3Amut_Metformin_replicate2


title: DNMT3Amut_Metformin_replicate5


title: DNMT3Amut_Metformin_replicate4


title: fresh_veh1-12_gp13-24

description:
Mixed-sample-fresh
SORT-seq

extract_protocol:
Cells were collected by trypsinization, washed with PBS after which the RAID cells were fixed with 0.5mM dithiobis(succinimidyl propionate)(DSP) in Sodium Phosphate buffered Saine (pH 8.4) for 45 minutes at room temperature at a concentration of 1 million cells per 2.5ml. Next, DSP was neutralized by incubating the cells with quench buffer (100mM Tris, pH 7.5, 150mM NaCl) for 10 minutes and cell clumps were removed usmg a 70µm cell strainer. Cells were icubated for 30 minutes with BP buffer (PFBB:PBS, 1:1, supplemented with 0.1% Triton•X-100 and 2 U/mL RNAsin plus) to permeabilize the cells, after which samples were incubated overnight with antibodies in BP buffer. Samples were filtered using a 40 um cell strainer and incubated with DAPI (0.2 ug/mL) prior to loading of samples on the flow cytometer. Fresh cells were directly loaded on the flow cytometry. For both fresh and fixed cells, Geminin1-110-expressing cells were selected and sorted into the captuture plates on a BD Influx cell sorter. The reducing agent already present in the capture plates was sufficient to decrosslink the RAID fixed cells prior to single cell RNA sequencing.
Single Cell Discoveries, where single-cell RNA sequencing was performed according to the SORT-seq protocol. Cells were heat-lysed at 65° C followed by cDNA synthesis. After second-strand cDNA synthesis, all the barcoded material from one plate was pooled into one library and amplified using in vitro transcription (IVT). Following amplification, library preparation was done following the CEL-Seq2 protocol to prepare a cDNA library for sequencing using TruSeq small RNA primers (Illumina). The DNA library was paired-end sequenced on an Illumina Nextseq™ 500, high output, with a 1x75 bp Illumina kit. CELSeq 2, CB: 8bp, UMI: 6bp R1 starts with a 6bp UMI  followed by a 8bp cell barcode. R2 ends with a 6bp random primer.

treatment_protocol: 2 conditions were employed in this experiment: RPE_veh received no doxycycline and no gemcitabine/prexasertib, RPE_gp received no doxycycline and 4 nM gemcitabine/10 nM Prexasertib for 16 hours. For each condition, cells were either freshly sorted (fresh) or sorted after reversible fixing and intracellular staining (RAID).

value: fresh / veh


title: CD45+/CX3CR1+ immune cells in NRAS* MBMs in Rela cKO mice (replicate 1)

description:
Library name: Rela_cKO-1_immune_cells_in_Nras_MBMs
10x Genomics

value: Brain

value: CD45+/CX3CR1+ immune cells

value: Rela cKO


title: TDPhigh-5, snRNA

description:
Library name: sample 23
10x Genomics Chromium Single Cell Multiome

value: dorsolateral prefrontal cortex

value: cortical cells


title: DNMT3Amut_Vehicle_replicate5

value: Vehicle


title: CD45+/CX3CR1+ immune cells in NRAS* MBMs in Rela WT mice (replicate 2)

description:
Library name: Rela_WT-2_immune_cells_in_Nras_MBMs
10x Genomics

value: Rela WT

value: No treatment


title: DNMT3Amut_Vehicle_replicate4


title: CD45+/CX3CR1+ immune cells in NRAS* MBMs in Rela WT mice (replicate 3)

description:
Library name: Rela_WT-3_immune_cells_in_Nras_MBMs
10x Genomics


title: DNMT3Amut_Vehicle_replicate3


title: TDPhigh-4, snRNA

description:
Library name: sample 22
10x Genomics Chromium Single Cell Multiome

value: C9orf72 HRE


title: TDPhigh-3, snRNA

description:
Library name: sample 21
10x Genomics Chromium Single Cell Multiome


title: DNMT3Amut_Vehicle_replicate2


title: RPE_dox_veh

description: SORT-seq

value: RPE

value: TetOn HRAS G12V


title: DNMT3Amut_Vehicle_replicate1


title: B16,RI,4

city: Guangzhou

first: Yanpei

last: Zhang

state: Guangdong

address: No.1023,South Shatai Road, Baiyun District, Guangzhou, Guangdong, China

country: China

institute: Southern Medical University

description: Library name: 10547-1_RI_10S-ZYP-6G

data_processing:
Illumina Casava1.7 software used for basecalling.
HISAT2 software was used to conduct genome mapping analysis of preprocessed sequencing sequences. Usually we compare clean No RNA reads to the genome, which is the basis of subsequent analysis. HISAT2 uses global and local search methods, which can perform efficient mapping and effectively compare splICED reads in RNA Seq sequencing data. The default parameters are set, and the reference genome version is GRCm38.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Assembly: mm10 (GRCm38)
Supplementary files format and content: gene_count_matrix.csv: The CSV file includes the expression values for each sample.

source_name: tumor

extract_protocol:
Tumors were harvested, and total RNA was isolated and reverse transcribed into cDNA to construct an indexed Illumina library.
RNA libraries were prepared for sequencing using standard Illumina protocols.

value: tumor

value: melanoma cells

value: WT

value: radiation+immune checkpoint inhibitor


title: TDPhigh-8, snRNA

description:
Library name: sample 26
10x Genomics Chromium Single Cell Multiome


title: WT_Metformin_replicate4

value: Bone marrow

value: HSC

value: Metformin


title: TDPhigh-2, snRNA

description:
Library name: sample 20
10x Genomics Chromium Single Cell Multiome


title: WT_Metformin_replicate3


title: B16,RI,2

description: Library name: 10547-1_RI_8S-ZYP-6G


title: TDPhigh-1, snRNA

description:
Library name: sample 19
10x Genomics Chromium Single Cell Multiome


title: WT_Metformin_replicate5


title: WT_Metformin_replicate2


title: B16,ICI,3

description: Library name: 10547-1_ICI_5S-ZYP-6G

value: immune checkpoint inhibitor


title: TDPmed-5, snRNA

description:
Library name: sample 18
10x Genomics Chromium Single Cell Multiome


title: WT_Metformin_replicate1


title: TDPmed-4, snRNA

description:
Library name: sample 17
10x Genomics Chromium Single Cell Multiome


title: DNMT3Amut_Metformin_replicate1


title: WT_Vehicle_replicate3


title: B16,RI,1

description: Library name: 10547-1_RI_7S-ZYP-6G


title: WT_Vehicle_replicate5


title: WT_Vehicle_replicate1


title: B16,ICI,1

description: Library name: 10547-1_ICI_1S-ZYP-6G


title: CD45+/CX3CR1+ immune cells in NRAS* MBMs in Rela WT mice (replicate 1)

description:
Library name: Rela_WT-1_immune_cells_in_Nras_MBMs
10x Genomics

value: Brain

value: CD45+/CX3CR1+ immune cells


title: TA-MAC_rep4

data_processing:
Raw data (raw reads) of FASTQ format were mapped to a mouse reference transcriptome (Mus_musculus.GRCm38.cdna.all.fa) built with Kallisto v.0.46.1. Read quantification to reference transcriptome was performed with Kallisto. The following steps were performed using R and RStudio. Tximport was used to import abundance.tsv files to R environment. EdgeR was used for differential expression analysis to obtain DEG list objects and normalization. The MatrixStats package was used to determine the statistics on the data. Data was filtered by choosing transcripts with at least 10 reads and later, at least 1 CPM in at least 3 samples. Normalization factor TMM (trimmed mean of M-values) was applied. Limma and edgeR were used to obtain a final DEG list adjusted by BH (Benjamini-Hochberg) and sorted by p.value<0,05 and LFC > 1. The graphical constructs of the RNA-seq data were performed using gplots, plotly, gprofiler2, clusterprofiler and GSEABase.
Assembly: Mus_musculus.GRCm38.cdna.all.fa
Supplementary files format and content: Comma-separated files of abundance containing raw counts

source_name: Melanoma Brain Metastasis

growth_protocol: B16F10 cells (CRL-6475) were obtained from ATCC and were grown in DMEM (Sigma) with the same supplementation. Cells were kept at 37°C in a 5% CO2 atmosphere and the media was replaced every 2/3 days. Melanoma cells were passaged when 80% confluency was reached. Cells were discarded up to seven consecutive passages and replaced by fresh stocks. All cell lines were tested and confirmed negative for mycoplasma monthly at the host institution.

extract_protocol:
MBM samples preparation This protocol was performed in cold to prevent microglia ex vivo activation. Brains were sliced in 1 mm thick cuts and placed under the stereomicroscope, MBM were dissected manually and placed in ice-cold HBSS containing 15 mM HEPES and 0,5% Glucose. Control tissue or MBM were mechanically disrupted with 5X slow strokes with a Dounce tissue grinder (Wheaton, 357542) on ice. Samples were centrifuged at 350g during 5 min at 4°C and later at 8.000 g during 30 sec. Samples were subjected to RBC lysis buffer (Biolegend, 420301) for 3 min at RT and then spined again as described. Samples were subsequently subjected to flow cytometry, FACS or scRNAseq library preparation. FACS and flow cytometry: Prior to antibody staining, samples were blocked with Fc-block CD16/CD32 (Biolegend, 101320, 1:50) in FACS Buffer sterile filtered (1% FCS, 2mM EDTA, 25mM HEPES in PBS) for 10 min on ice and darkness to block nonspecific binding. For cell surface staining, cells were resuspended in 26 the appropriate antibody cocktail and incubated for 30 min on ice protected from light. Samples were centrifuged and washed with FACS buffer. Viability was assessed by staining with DAPI (1 μg/mL). Information and dilution for antibodies used for flow cytometry are listed in supplementary table 10. Immune cell profiling by flow cytometry was carried out by analyzing 50.000 live singlets in each sample with a FACS Aria III instrument and BD FACSDiva (BD Bioscience). After MBM sample preparation as described previously, samples were resuspended in FACS Buffer and subjected to surface antibody staining for TMEM119 (ab210405) and CD49D (Biolegend, 103627) and DAPI for 30 minutes on ice and darkness and subjected to FACS. Cells were collected in extraction buffer from the Arcturus PicoPure RNA Isolation Kit (Thermo Fisher). Then, following manufacturer’s instructions RNA was eluted in TE buffer and 1 μL subjected to Bioanalyzer High Sensitivity RNA chip (Agilent) for quality testing. Four samples containing isolated cells from MBM and control brains from 3 independent mice (males and females)
Stand-specific (reverse) libraries were prepared with cDNA converted using the SMARTer universal low input RNA synthesis kit. Sequencing was performed in single-end configuration with a read length of 150 bp.

treatment_protocol: Melanoma cells injection Intracarotid artery: Animals were anesthetized with isoflurane and laid on a heating mat under a stereomicroscope and viscous ophthalmic gel was applied in both eyes. Animals were injected with buprenorphine 1mg/mL 5 minutes prior the surgery. A ventral cut was performed in the left side of the neck and skin and fat was retracted to expose the trachea and pectoral muscle. The carotid artery was isolated with thin forceps, and detached from the bottom of the tissue and from the vagus nerve which was left untouched. Two sutures were placed under the artery with open knots. Then, the upper external branch of the artery was isolated and firmly closed with suture and 24 a piece of wet cotton placed under the common carotid artery. The bottom suture was closed, and cells were injected in the common carotid artery (2·10^5 Nras* and 1·10^5 B16F10 in 100 μL sterile PBS). Finally, animals were injected with 500-1000 μL of saline subcutaneously to prevent dehydration in the following hours post-surgery. Animals were monitored in the following hours and days for proper recovery.

value: Melanoma Brain Metastasis

value: FACS isolated myeloid cells

tag: group

value: T-MAC (Tumor associated macrophages)


title: TDPmed-2, snRNA

description:
Library name: sample 15
10x Genomics Chromium Single Cell Multiome

value: dorsolateral prefrontal cortex

value: cortical cells


title: WT_Vehicle_replicate4

value: Vehicle


title: TA-MAC_rep3


title: Fig6_mouse-liver_RtABE-V2-3e12-8w_2

city: Shanghai

first: Li

last: Yang

address: 131 Dong-An Road

department: Institutes of Biological Sciences

institute: Fudan University

data_processing:
RNA-seq reads were first processed by trimmomatic (version 0.39, parameters: ILLUMINACLIP: TruSeq3PE-2.fa:2:30:10:8:true LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:50) to remove low quality bases and adaptor sequences.
RNA editing was then analyzed and visualized using the RADAR pipeline
Assembly: hg38/mm10
Supplementary files format and content: tar.gz file includes high-confidence editing events called by RADAR from each sample
Library strategy: ribo– RNA-seq

source_name: mouse liver

extract_protocol:
Total RNAs were extracted from 293FT cells and mouse livers tissues using the TransZol Up Plus RNA Kit (ER501-01, TransGen).
The extracted RNAs were treated with the RiboErase HMR Kit (KAPA) to remove ribosomal RNA. Indexed RNA libraries were prepared using the RNA HyperPrep Kit (KAPA). The resulting RNA libraries were subjected to sequencing on the Illumina NovaSeq 6000 system (2×150) at Sequanta Technologies, Shanghai, China.

value: mouse liver


title: TA-MAC_rep2


title: TDPmed-1, snRNA

description:
Library name: sample 14
10x Genomics Chromium Single Cell Multiome

value: C9orf72 HRE


title: Fig6_mouse-liver_RtABE-V2-3e12-8w_1


title: Fig6_mouse-liver_RtABE-V2-3e12-4w_3


title: TA-MAC_rep1


title: TMG_rep4

value: TMG (tumor microglia)


title: TDPneg-5, snRNA

description:
Library name: sample 12
10x Genomics Chromium Single Cell Multiome


title: TMG_rep1


title: TMG_rep3


title: CMG_rep4

value: CMG (Control microglia)


title: B16,ICI,2

description: Library name: 10547-1_ICI_3S-ZYP-6G

value: tumor

value: melanoma cells

value: WT


title: CMG_rep3


title: WT_Vehicle_replicate2

value: Bone marrow

value: HSC


title: TMG_rep2


title: CMG_rep2


title: Fig6_mouse-liver_RtABE-V2-3e12-4w_1


title: CMG_rep1


title: TDPneg-6, snRNA

description:
Library name: sample 13
10x Genomics Chromium Single Cell Multiome


title: Fig6_mouse-liver_RtABE-V2-3e12-4w_2


title: TDPmed-3, snRNA

description:
Library name: sample 16
10x Genomics Chromium Single Cell Multiome


title: TDPneg-3, snRNA

description:
Library name: sample 10
10x Genomics Chromium Single Cell Multiome


title: TDPneg-4, snRNA

description:
Library name: sample 11
10x Genomics Chromium Single Cell Multiome


title: Mouse2_day30

library_source: other

data_processing:
CRISPR data were analyzed by Mageck 0.5.9.4 using the two mouse replicates as input samples and plasmid gRNA counts as control.
Assembly: GRCm38
Supplementary files format and content: CRISPR count, Mageck output

molecule: other

source_name: Blood

growth_protocol: HSPCs were plated into M3434 for seven days and then culturedin vitrofor another 7 days in X-VIVO 20 media (BE04-448Q, Lonza Bioscience) supplemented with 5% serum (Gibco, 10438-026), 10ng/ml IL3 (Peprotech, AF-213-13-1000), 10 ng/ml IL6 (Peprotech, AF-216-16-1000), 50 ng ml/ml SCF (Peprotech, AF-250-03-1000) and 1% penicillin–streptomycin–glutamine (Gibco, 12090216). Two days after transduction, BFP expression cells were flow-sorted and cultured for total of 30 days. On day6 post-transduction, 20% of cells were collected for DNA extraction, the remaining cells were replated in fresh X-vivo20 media with supplements. On day 10, 40% of cells were collected for DNA extraction, and the remaining cells replated. From day 15 to day 30, 50% of cells were collected for DNA analysis and 50% replated in fresh media.

extract_protocol:
Genomic DNA extraction and Illumina sequencing of gRNAs were described previously (Tzelepis et al., 2016). 19bp single-end sequency was performed with the custom sequencing primer 5′-TCTTCCGATCTCTTGTGGAAAGGACGAAACACCG-3′. Enrichment and depletion of guides and genes were analysed using the MAGeCK statistical package63by comparing read counts from each cell line with counts from the plasmid as the initial population.
For the screen we used our murine v2 lentiviral guideRNA library containing 90 230 gRNAs against 18 424 genes. 3*107cells were then transduced with a pre-determined volume of the mouse v2 genome-wide gRNA lentiviral supernatant that gave rise to 30% transduction efficiency measured by BFP expression.

value: Blood

value: HSPC


title: Fig6_mouse-liver_RtABE-V2-1e12-4w_3


title: TDPneg-2, snRNA

description:
Library name: sample 9
10x Genomics Chromium Single Cell Multiome


title: Mouse2_day25


title: TDPneg-1, snRNA

description:
Library name: sample 8
10x Genomics Chromium Single Cell Multiome


title: Fig6_mouse-liver_RtABE-V2-1e12-4w_2


title: Fig6_mouse-liver_RtABE-V2-1e12-4w_1


title: Mouse2_day15


title: Mouse2_day10


title: Control-7, snRNA

description:
Library name: sample 7
10x Genomics Chromium Single Cell Multiome


title: Fig6_mouse-liver_NT_rep3


title: Mouse2_day6


title: Mouse1_day25


title: Control-6, snRNA

description:
Library name: sample 6
10x Genomics Chromium Single Cell Multiome


title: Control-5, snRNA

description:
Library name: sample 5
10x Genomics Chromium Single Cell Multiome


title: Fig6_mouse-liver_NT_rep2


title: Fig6_mouse-liver_NT_rep1


title: Control-4, snRNA

description:
Library name: sample 4
10x Genomics Chromium Single Cell Multiome


title: Mouse1_day30


title: Mouse1_day6


title: Mouse1_day15


title: Control-3, snRNA

description:
Library name: sample 3
10x Genomics Chromium Single Cell Multiome


title: Fig6_mouse-liver_MCP-ADAR2DD-E488Q-1e12-4w_2


title: Mouse1_day10


title: Mouse1_day20


title: Control-2, snRNA

description:
Library name: sample 2
10x Genomics Chromium Single Cell Multiome


title: Fxr cko NEC pups_Intestinal epithelium_scRNAseq

city: Tianjin

first: yuchao

last: Jing

address: No.22, Meteorological Tai Road, Tianjin

institute: Tianjin Medical University

description:
Library name: sample4
10x Genomics

data_processing:
Cell Ranger pipeline was used to perform alignment against the mouse genome mm10 (using STAR), filtering, barcode counting and unique molecular identifiers (UMIs) counting.
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Intestinal epithelium

extract_protocol:
IECs were isolated from small intestines by washing twice with gut wash buffer composed of HBSS supplemented with 2 mM L-glutamine, 1 mM sodium pyruvate, 20 mM HEPES, 100 U/mL penicillin, 100 mg/mL streptomycin, and 2% fetal bovine serum (FBS). Tissues were then cut into 1- 2 cm segments and were incubated with gut wash buffer containing 2 mM dithiothreitol at 37℃ for 10 min. Subsequently, the tissues were vigorously shaken and incubated with fresh gut wash buffer containing 5 mM EDTA at 37℃ for 7 min. The fractions containing EDTA were collected and tissues were subsequently incubated with fresh EDTA for extra 7 min. The two EDTA-containing fractions were filtered through 100-μm filters and centrifuged at 300 g for 5 min. Cell pellets were then digested with Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 0.1 U/mL collagenase I (Invitrogen, USA) and 0.1 mg/mL DNase I (Solarbio) for 8 min in 37℃ water bath. The digestion process was neutralized with 2.5% FBS-DMEM, and the pellets were washed twice with gut wash buffer.
Library was performed according to the manufacter's instructions (single cell 3’v2 protocol. 10x Genomics)

value: Intestinal epithelium

value: mouse intestinal epithelial cells

value: Fxr cko

value: NEC model


title: Fig6_mouse-liver_MCP-ADAR2DD-E488Q-1e12-4w_3


title: NEC pups_Intestinal epithelium_scRNAseq

description:
Library name: sample2
10x Genomics

value: Wild type


title: Ctrl pups_Intestinal epithelium_scRNAseq

description:
Library name: sample1
10x Genomics

value: naïve


title: Control-1, snRNA

description:
Library name: sample 1
10x Genomics Chromium Single Cell Multiome

value: dorsolateral prefrontal cortex

value: cortical cells


title: Fig6_mouse-liver_MCP-ADAR2DD-E488Q-1e12-4w_1

value: mouse liver


title: Mouse2_day20


title: RNA_seq_SingleCell_Aged_3

city: Kyoto

first: Takuya

last: Yamamoto

address: 53 Kawahara-cho, Shogoin, Sakyo-ku

department: CiRA

country: Japan

institute: Kyoto University

data_processing:
The 10X Genomics Cell Ranger pipeline (version 6.0.1) was used to perform sample demultiplexing, alignment to the mm10 mouse reference genome, barcode/UMI processing, and gene counting for each cell.
A noise reduction method, RECODE (version 0.1.1.dev202111082351) (Imoto et al., bioRxiv, 2022, doi: https://doi.org/10.1101/2022.05.02.490246), was applied to the quality-checked UMI count matrices for 5,135 from young mice and 5,709 cells from aged mice separately with 1,000 < nFeature < 6,000, nCount < 50,000, and low mitochondrial gene expression (< 25%).
Assembly: mm10
Supplementary files format and content: *.h5: HDF5 files contain lists of gene names, cell barcodes and UMI counts for each sample.

source_name: intestinal epithelial cells

extract_protocol:
The proximal half of the small intestines were dissected, washed with cold PBS (Mg-/Ca-) and opened. Villi were removed by scraping with glass slides. The tissues were cut into 3-5mm fragments and washed 5-6 times in cold PBS (Mg-/Ca-) with gentle shaking by hand. The pieces were then incubated for 30 min at 4°C in PBS (Mg-/Ca-) containing 5mM EDTA. Crypts were then mechanically isolated by vigorous shaking, and filtered through a 70-um mesh to remove remaining villus and tissue fragments. The isolated crypts were dissociated into single cells by treating the cells with TrypLE Express (Gibco) containing DNase I (10U uL-1, Roche1) for 30 min at 37°C and gentle pipetting. The dissociated cells were resuspended in cold SMEM (Gibco) and incubated with the following antibodies for 30 min at 4°C: EPCAM-APC (G8.8) (Biolegend), CD24-Pacific Blue (M1/69) (Biolegend), CD45-PE/Cy7 (30-F11) (Biolegend), Ter119-PE/Cy7 (TER-119) (Biolegend) and CD31-PE/Cy7 (Mec13.3) (Biolegend), all at 1:500. Finally, cells were resuspended in SMEM supplemented with 7-AAD (BD Bioscience, dilution 1:100), 100 U mL-1 RNase inhibitor (Invitrogen) and 10 uM Y27632 (Tocris Bioscience). Cells were sorted using FACSAriaII or FACSAriaIIu (BD Biosciences). Intestinal epithelial cells were isolated as Epcamhigh CD31−Ter119−CD45−7-AAD−. Single-cell suspensions of all samples were resuspended in PBS-0.4% BSA and barcoded with a 10x Chromium Controller (10x Genomics).
RNA from the barcoded cells for each sample was subsequently reverse-transcribed and sequencing libraries were constructed with reagents from a Chromium Single Cell 3’ reagent kit v3.1 (10x Genomics). Sequencing was performed with Illumina NextSeq 500.

tag: strain

tag: age

value: 26 months

value: male

value: intestinal epithelial cells


title: Fxr fl/fl  NEC pups_Intestinal epithelium_scRNAseq

description:
Library name: sample3
10x Genomics

value: Fxr fl/fl


title: Fig5e_293FT_NT

value: human embryonal kidney cell

value: WT


title: RNA_seq_SingleCell_Young_2

value: 3 months


title: Murine lineage-negative (Lin-) cells from hematochimeric mice reconstituted by UBA1wt

city: Segrate

first: Roberta

last: Alfieri

state: MI

address: Via Fratelli Cervi 93

department: ITB

country: Italy

institute: CNR

data_processing:
Preliminary analysis using the CellRanger using default parameters. Filtering, Normalization, Scaling using the R package Seurat. Identification of clusters using the Louvain algorithm privded by the R package Seurat. Identification of differentially expressed genes using the Seurat function FindAllMarkers with default parameters. Genome_build: GRCh38 or GRCm38
Assembly: GRCh38, GRCm38.
Supplementary files format and content: matrix.mtx, barcodes.tsv, features.tsv

extract_protocol:
Cell suspensions were diluted in PBS with 0.04% BSA to a final concentration of 1 × 106 cells/mL. Vitality was assessed for each individual sample. Approximately 20,000 cells were loaded onto a single-cell chip for gel bead-in-emulsion (GEM) generation using the 10x Genomics Chromium Controller (10× Genomics).
3′mRNA-seq gene expression libraries were constructed using the Chromium Single Cell 3′ Library & Gel Bead Kit v2 (10x Genomics) according to the manufacturer’s guidelines. All reactions were performed in a C1000 Touch Thermal Cycler (Bio-Rad Laboratories). Twelve cycles were used for cDNA amplification, while the number of total cycles for the sample index PCR was calculated based on the cDNA concentration. Amplified cDNA and final libraries were evaluated using a Tape Station 4200 (Agilent Technologies) with a high sensitivity chip. scRNA-seq libraries were sequenced on an Illumina NovaSeq6000 with the standard sequencing protocol of R1 28; I1 10; I2 10; R2 90 nt read length.

value: Bone marrow

tag: individual

value: murine Lin-


title: RNA_seq_SingleCell_Aged_1

value: 22 months


title: Murine lineage-negative (Lin-) cells from hematochimeric mice reconstituted by UBA1mut


title: Fig5_293FT_split-ADAR2DD-E488Q


title: RNA_seq_SingleCell_Young_3


title: Fig5_293FT_RtABE-V2-STAT1-3


title: RNA_seq_Bulk_Paneth cells_Aged_Female_5_rep_2

data_processing:
The sequenced reads were trimmed to remove low-quality bases and adaptor sequences using cutadapt v3.0. The sequenced reads were mapped to the mm10 mouse reference genome using STAR version 2.7.9a with the GENCODE vM23 annotation gtf file. The read counts per gene were calculated using HTSeq-count v0.13.5. Uniquely mapped reads (MAPQ ≥ 20) were used. Uniquely mapped reads to mitochondrial genes were excluded for further analysis.
Each gene expression level was calculated as reads per million mapped reads (RPM) using DESeq2 version 1.30.1.
Assembly: mm10
Supplementary files format and content: *.tsv: Tab-separated text files include raw counts or CPM (= RPM) values.

source_name: Paneth cells

extract_protocol:
The proximal half of the small intestines were dissected, washed with cold PBS (Mg-/Ca-) and opened. Villi were removed by scraping with glass slides. The tissues were cut into 3-5mm fragments and washed 5-6 times in cold PBS (Mg-/Ca-) with gentle shaking by hand. The pieces were then incubated for 30 min at 4°C in PBS (Mg-/Ca-) containing 5mM EDTA. Crypts were then mechanically isolated by vigorous shaking, and filtered through a 70-um mesh to remove remaining villus and tissue fragments. The isolated crypts were dissociated into single cells by treating the cells with TrypLE Express (Gibco) containing DNase I (10U uL-1, Roche1) for 30 min at 37°C and gentle pipetting. The dissociated cells were resuspended in cold SMEM (Gibco) and incubated with the following antibodies for 30 min at 4°C: EPCAM-APC (G8.8) (Biolegend) and CD24-Pacific Blue (M1/69) (Biolegend), both at 1:500. Finally, cells were resuspended in SMEM supplemented with 7-AAD (BD Bioscience, dilution 1:100), 100 U mL-1　RNase inhibitor (Invitrogen) and 10 uM Y27632 (Tocris Bioscience). Cells were sorted using FACSAriaII or FACSAriaIIu (BD Biosciences). ISCs were isolated as Lgr5GFPhighEpcamhighCD24low7-AAD−, progenitors were isolated as Lgr5GFPmedium/lowEpcamhighCD24low7-AAD−, and Paneth cells were isolated as CD24highSidescatterhighLgr5GFP−Epcamhigh7-AAD−.
The cells were lysed and the total RNAs were purified using an RNeasy micro kit (Qiagen). 1 ng of total RNAs from each sample was used for synthesis and amplification of cDNA according to the single cell 3 prime RNA-seq (SC3-seq) method described previously (Nakamura et al., Nucleic Acids Res., 2015). RNA-seq libraries were sequenced on a Nextseq 500 (Illumina) with a NextSeq 500 High Output v2 Kit (Illumina).

value: 23 months

value: female

value: Paneth cells


title: RNA_seq_SingleCell_Young_1


title: LLC1-sgLkb1, RI, 3

data_processing:
Illumina Casava1.7 software used for basecalling.
HISAT2 software was used to conduct genome mapping analysis of preprocessed sequencing sequences. Usually we compare clean No RNA reads to the genome, which is the basis of subsequent analysis. HISAT2 uses global and local search methods, which can perform efficient mapping and effectively compare splICED reads in RNA Seq sequencing data. The default parameters are set, and the reference genome version is GRCm38.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Assembly: mm10 (GRCm38)
Supplementary files format and content: expression matrix.xlsx: The CSV file includes the expression values for each sample.

value: tumor

value: Lewis lung carcinoma cells

value: LKB1 knockdown

value: radiation+immune checkpoint inhibitor


title: RNA_seq_Bulk_Paneth cells_Aged_Female_5_rep_3


title: Stromal cells of Nras* MBMs in mice

description: 10x Genomics

data_processing:
10x-barcoded sequencing library was generated using the 10x Single Cell 3' v3 kit (10x Genomics, inc.) and sequenced to 535,522,726 reads.
Raw FASTQ from gene expression library was aligned on the mm10-2020-A mouse genome and processed using CellRanger Count (10X Genomics Cell Ranger v6.0.1) with default parameters and 7,477 cells were retrieved.
Sequencing depth was 71,623 mean read pairs per cell, with a sequencing saturation of 76.3%.
Multiplet artifacts were removed using Scrublet v0.2.3 python package (Wolock et al., 2019) (Python v3.7.10 (Van Rossum and Drake, 2009)) with default parameters and upon doublet removal, we retrieved 7,434 cells.
All subsequent single cell transcriptomics analyses were performed using R Statistical Software (v4.1.1) and the Seurat v4 (v4.0.3) package (Hao et al., 2021; R: The R Project for Statistical Computing).
For quality control, genes expressed in fewer than 3 cells were removed.
Cells with unique feature counts (detected genes) over 5,000 or less than 1,400 were excluded.
Finally, cells with more than 5% mitochondrial counts were filtered out.
The remaining cells, 3,393 high quality cells were then used for downstream analyses.
UMI count data was normalized using a regularized negative binomial regression with SCTransform (sctransform 0.4.1).
SCTransform was executed with the additional flag method="glmGamPoi" to fit a Gamma-Poisson Generalized Linear Model which improves the speed of the learning procedure.
Furthermore, we regressed out mitochondrial content and percentage of Xist and Tsix genes (X-linked genes) in a second non-regularized linear regression.
Principal Component Analysis (PCA) was performed on the SCT-normalized data.
Non-linear dimensional embedding was performed using Uniform Manifold Approximation and Projection (UMAP) on the 30 most significant dimensions of the PCA.
Shared Nearest-Neighbor (SNN) graph was constructed using FindNeighbors function on the 30 most significant principal components (PCs), and cell clusters were identified using FindClusters.
The SCT assay data was used for the expression plots and related analysis.
Differentially expressed genes between clusters were obtained by Seurat-implemented Wilcoxon rank sum test within FindAllMarkers function with only.pos=TRUE, min.pct=0.25 and logfc.threshold=0.25 parameters.
Assembly: GRCm38 (mm10)
Supplementary files format and content: Single cell Seurat object: Nras_MBMs_stroma.rds
Supplementary files format and content: Cell barcode: barcodes.tsv.gz
Supplementary files format and content: Gene name: features.tsv.gz
Supplementary files format and content: Expression matrix: matrix.mtx.gz

extract_protocol:
After MBM sample preparation as described above, cell suspensions were subjected to FACS isolation using a gating strategy that consisted in selection of a DAPI negative population (live cells). Cells were sorted into individual tubes per group, 15.000 cells per tube and later encapsulated
Live cells were encapsulated intro droplets and libraries were prepared using Chromium Single Cell Reagents kit v3 single index according to the manufacturer’s protocol (10X genomics, PN-1000269).

value: Brain

value: Stromal cells

value: No treatment


title: RNA_seq_SingleCell_Aged_2

value: 25 months

value: male

value: intestinal epithelial cells


title: Fig5_293FT_RtABE-V1-STAT1-3

value: human embryonal kidney cell

value: WT


title: Fig5_293FT_RtABE-V2-PPIA-5


title: RNA_seq_Bulk_Paneth cells_Aged_Female_4_rep_1


title: RNA_seq_Bulk_Paneth cells_Aged_Female_4_rep_2


title: LLC1-sgLkb1, RI, 2


title: RNA_seq_Bulk_Paneth cells_Aged_Female_3_rep_1


title: Fig5_293FT_RtABE-V1-PPIA-5


title: LLC1-sgLkb1, ICI, 3

value: immune checkpoint inhibitor


title: RNA_seq_Bulk_Paneth cells_Aged_Female_3_rep_3


title: RNA_seq_Bulk_Paneth cells_Aged_Female_3_rep_2


title: RNA_seq_Bulk_Paneth cells_Aged_Female_2_rep_1

value: 24 months


title: Fig5_293FT_dRanCas13b-ADAR2DD-E488Q


title: RNA_seq_Bulk_Paneth cells_Aged_Female_1_rep_3


title: Fig5_293FT_REPAIRx


title: RNA_seq_Bulk_Paneth cells_Aged_Female_1_rep_2


title: LLC1-sgLkb1, ICI, 2


title: RNA_seq_Bulk_Paneth cells_Aged_Female_4_rep_3

value: 23 months


title: LLC1-sgLkb1, RI, 1


title: RNA_seq_Bulk_Paneth cells_Aged_Female_5_rep_1

value: female


title: RNA_seq_Bulk_Paneth cells_Aged_Female_2_rep_3

value: Paneth cells


title: RNA_seq_Bulk_Paneth cells_Aged_Male_7_rep_3


title: Fig5_293FT_LEAPER-V2


title: Fig5_293FT_dCas13X-ADAR2DD-E488Q


title: RNA_seq_Bulk_Paneth cells_Aged_Male_6_rep_2

value: 26 months


title: LLC1, RI, 3

value: tumor

value: Lewis lung carcinoma cells


title: RNA_seq_Bulk_Paneth cells_Aged_Male_7_rep_2


title: LLC1, ICI, 3


title: LLC1, ICI, 2


title: Fig5_293FT_dRanCas13b-ADAR2DD-E488Q-ecDHFR


title: Fig5_293FT_cadRNA


title: LLC1, ICI, 1


title: yeast, Eluate Rnh, noIAA, rep1

city: Paris

first: Benoit

last: Palancade

address: 15 rue Hélène Brion

department: Institut Jacques Monod

country: France

institute: Université Paris Cité, CNRS

library_source: genomic

data_processing:
DRIP-seq reads were trimmed from 3' and 5' end until the final base had a quality score >30 using Cutadapt 3.4, discarding reads left with <15bp
DRIP-seq reads were aligned to the sacCer3 genome using Bowtie2 version 2.5.0, using the following parameters : D=20, R=3, N=0, L=20, I=S,1,0.5, X=500.
Aligned reads were filtered to retained only reads mapped in a proper pair with a quality score >30 using Samtools version 1.8, and duplicates were removed using Rmdup version 2.0.1.
Aligned reads were splitted based on their strandedness using BamCoverage version 3.5.4, with a bin size of 20bp, no scaling, and only including reads originating from fragments from the forward or reverse strand.
Assembly: sacCer3 (12/17/2014)
Supplementary files format and content: Bigwig files

organism: Saccharomyces cerevisiae

source_name: yeast cell

growth_protocol: Cells were grown at 30°C in Synthetic Complete Medium until OD600nm=0.8.

extract_protocol:
100 OD of cells were mixed a synthetic DNA:RNA hybrid spike-in, in a 1:1 ratio with cell count, and 5% of C. glabrata cells prior to lysis. Genomic DNA was phenol-extracted and isolated by ethanol precipitation. DNA:RNA hybrids were then immunoprecipitated using the S9.6 antibody.
Library were created using the xGen ssDNA & Low-Input DNA Library Prep Kit (Integrated DNA Technologies) according to the manufacturer instructions, with 16 cycles of amplification. Post-amplification libraries were size selected at 350-1000bp using SPRIselect beads (Beckman Coulter). Libraries were further quantified by Qubit and Tapestation (Agilent).
ssDNA-seq

treatment_protocol: Auxin (3-indole acetic acid, IAA, Sigma, 0.5 mM final) was added for 1h to the culture, prior to harvesting. For synchronization, G1 arrest was triggered at 30°C by three sequential additions of alpha-factor (2µg/ml, 4µg/ml then 2µg/ml, Biotem) spaced by 40min.

value: yeast cell

tag: background

value: DNA treated in vitro with RNase H

tag: fraction

value: Immunoprecipitated DNA obtained following in vitro RNaseH treatment


title: RNA_seq_Bulk_Paneth cells_Aged_Female_1_rep_1


title: RNA_seq_Bulk_Paneth cells_Aged_Male_7_rep_1


title: RNA_seq_Bulk_Paneth cells_Aged_Female_2_rep_2


title: LLC1, RI, 2

value: radiation+immune checkpoint inhibitor


title: RNA_seq_Bulk_Paneth cells_Aged_Male_6_rep_1


title: yeast, Eluate Rnh, IAA, rep1

value: cells treated with 0.5mM IAA, DNA treated in vitro with RNase H


title: RNA_seq_Bulk_Paneth cells_Aged_Male_5_rep_3

value: male


title: Fig4-5e_293FT_RtABE-K475Q-E488Q-RtABE-V1


title: LLC1, RI, 1


title: Fig4-5e_293FT_RtABE-K475I-S486A-RtABE-V2


title: yeast, Eluate Rnh, IAA G1, rep1

value: cells treated with 0.5mM IAA & 2-4 µg/mL alpha-Factor, DNA treated in vitro with RNase H


title: RNA_seq_Bulk_Paneth cells_Aged_Male_5_rep_2


title: LLC1-sgLkb1, RT, 5

value: LKB1 knockdown

value: radiation


title: Fig5_293FT_CLUSTER

value: human embryonal kidney cell

value: WT


title: yeast, Input, noIAA, rep1

value: noIAA

value: Input DNA


title: Fig4-5e_293FT_MCP-ADAR2DD-E488Q


title: RNA_seq_Bulk_Paneth cells_Aged_Male_6_rep_3

value: Paneth cells


title: RNA_seq_Bulk_Paneth cells_Aged_Male_5_rep_1

value: 24 months


title: LLC1-sgLkb1, RT, 4

value: tumor


title: Fig4_293FT_RtABE-WT


title: RNA_seq_Bulk_Paneth cells_Aged_Male_4_rep_3


title: LLC1-sgLkb1, RT, 1

value: Lewis lung carcinoma cells


title: yeast, Eluate, noIAA, rep2

value: yeast cell

value: Immunoprecipitated DNA


title: yeast, Input, IAA G1, rep1

value: cells treated with 0.5mM IAA & 2-4 µg/mL alpha-Factor


title: RNA_seq_Bulk_Paneth cells_Aged_Male_4_rep_1


title: yeast, Input, IAA, rep1

value: cells treated with 0.5mM IAA


title: Fig4_293FT_RtABE-V351G-E488Q


title: yeast, Eluate, noIAA, rep1


title: Fig4_293FT_RtABE-T375S-E488Q


title: RNA_seq_Bulk_Paneth cells_Aged_Male_4_rep_2


title: Fig4_293FT_RtABE-S486A


title: Fig4_293FT_RtABE-S486A-E488Q


title: yeast, Eluate, IAA, rep2


title: RNA_seq_Bulk_Paneth cells_Aged_Male_3_rep_2


title: RNA_seq_Bulk_Paneth cells_Aged_Male_3_rep_3


title: yeast, Eluate, IAA, rep1


title: Fig4_293FT_RtABE-R348A-E488Q


title: RNA_seq_Bulk_Paneth cells_Aged_Male_3_rep_1


title: yeast, Eluate, IAA G1, rep1


title: RNA_seq_Bulk_Paneth cells_Aged_Male_2_rep_2

value: 26 months


title: Fig4_293FT_RtABE-N473S-E488Q


title: yeast, noIAA, rep3

data_processing:
Reads were trimmed for adapter sequence from 3' end, and from both 3' and 5' end until the final base had a quality score >30 using Cutadapt 3.4, discarding reads left with <15bp
Trimmed reads were aligned to the sacCer3 genome using RNA STAR version 2.5.0, using the following parameters : sjdbOverhang=99, quantMode=Per gene read count, and ENCODE options for advanced options of the mapping stage.
Aligned reads were filtered to retain only uniquely mapped reads using bamTools version 2.5.2. Reads were then scaled to a scaling factor defined as 1 / millions of uniquely mapped reads for each sample using bedtools GenomeCoverage version 2.30.0
Assembly: sacCer3 (12/17/2014)
Supplementary files format and content: Bigwig files

extract_protocol:
Total RNA was extracted using the NucleoSpin RNA kit (Macherey-Nagel) following manufacturer's instructions.
Ribosomal RNA was removed from total RNA, followed by ethanol precipitation. After fragmentation, the first strand cDNA was synthesized using random hexamer primers. During the second strand cDNA synthesis, dUTPs were replaced with dTTPs in the reaction buffer. The directional library was obtained following end repair, A-tailing, adapter ligation, size selection, USER enzyme digestion, amplification, and purification.

treatment_protocol: Auxin (3-indole acetic acid, IAA, Sigma, 0.5 mM final) was added for 1h to the culture, prior to harvesting.


title: Fig4_293FT_RtABE-K475I-E488Q


title: RNA_seq_Bulk_Paneth cells_Aged_Male_2_rep_1

value: male


title: RNA_seq_Bulk_Paneth cells_Aged_Male_1_rep_3

value: 27 months


title: Fig4_293FT_RtABE-E488Q


title: yeast, noIAA, rep2


title: yeast, noIAA, rep1


title: GBC31T

type: RNA

city: New Delhi

first: Pramod

middle: Kumar

last: Garg

state: Delhi

address: Ansari Nagar

department: Gastroenterology

country: India

institute: AIIMS, New Delhi

description: Gene expression

hyb_protocol: The protocol of one-color micro-array-based gene expression (Agilent technologies; SurePrint G3 Human Gene Expression 8x60K v2 Microarray) was followed as per manufacturer’s recommendations

scan_protocol: Hybridized arrays were scanned with Agilent’s G2505B microarray scanner system

data_processing: hisat2 and stringtie

source_name: tissue sample

extract_protocol: RNA extraction was done using mirVana microRNA isolation kit (Ambion).

treatment_protocol: Tissue samples were collected during surgery

tag: disease state

value: adenocarcinoma

tag: gender

value: female


title: Fig4_293FT_RtABE-E488Q-K594I

tag: cell type

value: human embryonal kidney cell


title: GBC14T

description: Gene expression

label_protocol: The protocol of one-color micro-array-based gene expression (Agilent technologies; SurePrint G3 Human Gene Expression 8x60K v2 Microarray) was followed as per manufacturer’s recommendations


title: GBC18T

description: Gene expression


title: GBC22T

description: Gene expression


title: Fig4_293FT_NT

tag: genotype

value: WT


title: RNA_seq_Bulk_Paneth cells_Aged_Male_1_rep_2


title: RNA_seq_Bulk_Paneth cells_Aged_Male_1_rep_1


title: yeast, IAA, rep2

value: 0.5mM IAA, 1h


title: Fig3d-i_NT


title: GBC21T

description: Gene expression


title: RNA_seq_Bulk_Paneth cells_Young_Female_6_rep_3

value: 3 months


title: yeast, IAA, rep1


title: GBC18T-1

description: Gene expression


title: Fig3d-i_293FT_TEVc-NES-2A-NES-TEVn-agRNA


title: Fig3_293FT_without-TEVp-agRNA


title: Overexpression BRN3A3

city: Varanasi

first: Jagat

last: Roy

state: Uttar Pradesh

address: Institute of Science

department: Zoology

institute: Banaras Hindu University

description:
S-22
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

data_processing:
The resulting sequencing Fastq files were mapped to the Human genome (hg19) using Tophat with Bowtie.
The aligned SAM/BAM file for each was processed for guided transcript assembly using Cufflink 2.1.1 and gene counts were determined.
Mapped reads were assembled using Cufflinks. Transcripts from all samples were subjected to Cuffmerge to get final transcriptome assembly across samples.
In order to ascertain differential expression of gene transcripts between different samples, Cuffdiff 2.1.1 was used (Trapnell et al., 2012). A P-value ≤0.05 was taken to indicate significantly differentially expressing genes between different groups compared.
Assembly: hg19
Supplementary files format and content: Cuffdiff 2.1.1 result showing differential gene expression between different samples in excel sheet

source_name: SiHa

growth_protocol: HPV positive cervical cancer cell line, ‘SiHa’ cells were used in the present study, which was maintained in DMEM medium supplemented with 10% FBS with 1X antibiotic solution and incubated into a CO2 incubator with 5% CO2 level at 37 ºC. Cells were washed with 1X PBS and trypsinized and transferred every three days called passage and for seeding as per experimental requirement.

extract_protocol:
Transfected SiHa cells were transferred into a nuclease-free microcentrifuge tube (MCT), washed with 1x PBS, and the addition of 1.0mL TRIzol® (Sigma). Three biological replicates for each group (for both OE and KD) were maintained. RNA extraction was done by using instructions of the manufacturer and DNaseI (Roche) treatment was given followed by the purity and concentration check by NanoDrop® 2000 (Thermo Scientific, Inc) and also checked on the 1% agarose gel. Purified RNA was used for RNA sequencing.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Overexpression (OE) of BRN3A was done through expression vector pCDNA3.1. BRN3A -ORF cloned in pCDNA3.1 (Genetix Biotech Asia). This empty vector was used as a control. Overnight grown sub-confluent SiHa cells (1.0 X 106 cells/well) were seeded in a six-well plate followed by transient transfection of the vectors using FuGENE®6 (Promega) transfection reagents according to the manufacturer’s instructions. To Knockdown (KD) BRN3A expression, siRNA oligos (5’-GCCAUAAUCUGCAACUUCAdTdT-3’), were used (GeneX India Bioscience) along with universal siRNA control (Sigma) used as control. Transfection was done through Mission siRNA Transfection Reagent (Sigma) using Opti-MEM (Gibco®) transfection reagent as per manufacturer’s instructions with subtle modification.

value: SiHa

value: Overexpression of BRN3A3


title: RNA_seq_Bulk_Paneth cells_Young_Female_6_rep_1

value: Paneth cells


title: yeast, IAA, rep3


title: GBC32T

description: Gene expression


title: Overexpression BRN3A2

description:
S-21
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: Overexpression of BRN3A2


title: RNA_seq_Bulk_Paneth cells_Young_Female_5_rep_3

value: 2 months


title: Fig3_293FT_TEVn-NES-2A-NES-TEVc-agRNA


title: RNA_seq_Bulk_Paneth cells_Young_Female_5_rep_2


title: GBC17T

description: Gene expression


title: GBC12T

description: Gene expression


title: GBC13T

description: Gene expression


title: GBC19T

description: Gene expression


title: Overexpression con3

description:
S-19
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: Overexpression control3


title: RNA_seq_Bulk_Paneth cells_Young_Female_6_rep_2


title: GBC7T

description: Gene expression


title: Overexpression BRN3A1

description:
S-20
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: Overexpression of BRN3A1


title: GBC25T

description: Gene expression


title: Fig3_293FT_eagRNA-V5


title: RNA_seq_Bulk_Paneth cells_Young_Female_4_rep_3


title: Overexpression con1

description:
S-17
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: Overexpression control1


title: Fig3_293FT_eagRNA-V4


title: Fig3_293FT_TEVp-NES-agRNA


title: LLC1-sgLkb1, ICI, 1

tag: tissue

value: tumor

value: Lewis lung carcinoma cells

value: LKB1 knockdown

value: immune checkpoint inhibitor


title: RNA_seq_Bulk_Paneth cells_Young_Female_4_rep_2

value: female


title: GBC5T

description: Gene expression

value: adenocarcinoma

value: male


title: GBC8T

description: Gene expression

value: Control


title: siRNA BRN3A5

description:
S-16
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

tag: cell line

value: knock down of BRN3A3


title: GBC10T

description: Gene expression


title: Fig3_293FT_eagRNA-V3

value: human embryonal kidney cell

value: WT


title: RNA_seq_Bulk_Paneth cells_Young_Female_4_rep_1

value: Paneth cells


title: Fig3_293FT_eagRNA-V6


title: RNA_seq_Bulk_Paneth cells_Young_Female_5_rep_1


title: Overexpression con2

description:
S-18
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: SiHa

tag: treatment

value: Overexpression control2


title: RNA_seq_Bulk_Paneth cells_Young_Female_3_rep_2

value: 2 months

tag: Sex


title: siRNA BRN3A2

description:
S-15
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: knock down of BRN3A2


title: RNA_seq_Bulk_Paneth cells_Young_Female_3_rep_1


title: Fig2-5c_293FT_NT_rep2


title: Fig3_293FT_eagRNA-V1


title: siRNA con3

description:
S-13
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: siRNA control 3


title: RNA_seq_Bulk_Paneth cells_Young_Female_2_rep_3


title: siRNA con2

description:
S-12
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: siRNA control 2


title: RNA_seq_Bulk_Paneth cells_Young_Female_2_rep_2


title: Fig3_293FT_eagRNA-V2


title: GBC6T

description: Gene expression


title: GBC31N-1

description: Gene expression


title: GBC31N

description: Gene expression


title: GBC6N

description: Gene expression


title: Fig2-5c_293FT_NT_rep1


title: siRNA BRN3A1

description:
S-14
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: knock down of BRN3A1


title: Fig2_293FT_MCP-ADAR2DD-E488Q-PPIB_1


title: SC, 17DMD, differentiation, biol rep 4

city: München

first: Sophie

last: Franzmeier

address: Veterinärstraße 13

country: Germany

institute: LMU Munich

description:
Library name: Sample 16
Column name in Raw_counts_all samples.tsv: NR_17DMD_38

data_processing:
RNAseq reads were demultiplexed using the program bcl2fastq provided by Illumina. R version 4.1.2 was used for all further analysis. Quality control was done using the command line tools fastQC and multiQC.
Assembly: Umi-tool dedup was used for deduplication, and the reads were aligned to the Sus scrofa reference genome susScr3 using STAR aligner version 2.7.10b. The number of reads mapping to each gene was also obtained from STAR using the parameter quantMode GeneCounts.
Supplementary files format and content: Data in tab separated values format including raw counts for all samples

organism: Sus scrofa

source_name: Muscle tissue

growth_protocol: DMD and WT SC were cultivated in Ham's F10 medium (GIBCO Thermo Fisher) supplemented with 20% fetal bovine serum (FBS, GIBCO Thermo Fisher), 1% Penicillin-Streptomycin (PenStrep, GIBCO Thermo Fisher), and 10 µg recombinant human basic fibroblast growth factor (PEPro Tech) in an humidified atmosphere at 37°C with 5% CO2.

extract_protocol:
Total RNA was extracted from cells in PROL and DIFF conditions using the NEB Monarch RNA Miniprep Kit, the protocol was performed accordingly to the instructions provided by the manufacturer. 25 ng of total RNA was used.
polyA+ RNA s were selected during cDNA synthesis through annealing to a polydT oligo containing an unique molecular idenitifer (UMI) and a well-barcode. Second strand synthesis was done by RT-PCR using a template switch oligo with an integrated Illumina adapter sequence. After purification and exonuclease°I treatment, the library pool was amplified by PCR and purified again. To achieve optimal fragment lengths, NexteraXT DNA library prep Kit (Illumina) was used. With the Nextera transposon DNA was tagmented, a process that fragments and tags DNA with adapter sequences. For amplification of the library, a 3' enrichment PCR was performed during which i7 and i5 adapters were attached. After purification, the quantity of the library pool (Qubit Fluorometer 4.0, Thermo Fisher) and average fragment size (LabchipGX Touch 24, Perkin Elmer/Revvity) were measured. The resulting library was sequenced on a NextSeq550Dx using a 75-cycle high output kit (Read 1:16°cycle – UMI + barcode; Read 2: 76°cycles – Insert).

treatment_protocol: Myogenic differentiation was induced by cultivating the cells in Ham's F10 medium supplemented with 1% PenStrep and 5% horse serum (Biozol).

value: Muscle tissue

value: Myogenic satellite cell

value: DMD

value: Differentiation


title: RNA_seq_Bulk_Paneth cells_Young_Female_1_rep_2

value: female


title: siRNA con1

description:
S-11
Total RNA seq data from SiHa Cell line transfected for overexpression and knock down purpose

value: siRNA control 1


title: GSD3

description: Gene expression

value: Control


title: RNA_seq_Bulk_Paneth cells_Young_Female_2_rep_1


title: RNA_seq_Bulk_Paneth cells_Young_Female_1_rep_1


title: SC, 15DMD, differentation, biol rep 2

description:
Library name: Sample 14
Column name in Raw_counts_all samples.tsv: NR_15DMD_36


title: GBC23T

description: Gene expression

value: male


title: RNA_seq_Bulk_Paneth cells_Young_Female_1_rep_3


title: Fig2_293FT_MCP-ADAR2DD-E488Q-No_agRNA

value: human embryonal kidney cell

value: WT


title: SC, 16DMD, differentiation, bio rep 3

description:
Library name: Sample 15
Column name in Raw_counts_all samples.tsv: NR_16DMD_37


title: GSD20

description: Gene expression


title: Fig2_293FT_eGFP-ADAR2DD-E488Q


title: SC, 14DMD, differentiation, biol rep 1

description:
Library name: Sample 13
Column name in Raw_counts_all samples.tsv: NR_14DMD_35


title: GSD21

description: Gene expression


title: RNA_seq_Bulk_Paneth cells_Young_Male_6_rep_2

value: 3 months

value: Paneth cells


title: Fig2_293FT_NT_rep3


title: SC, 15DMD, proliferation, biol rep 2

description:
Library name: Sample 10
Column name in Raw_counts_all samples.tsv: NR_15DMD_32

value: Proliferation


title: Fig2_293FT_MCP-ADAR2DD-E488Q-CTNNB1_1


title: GBC9N

description: Gene expression


title: Fig2_293FT_ADAR2DD-E488Q_rep1


title: Fig2_293FT_ADAR2DD-E488Q_rep3


title: RNA_seq_Bulk_Paneth cells_Young_Male_6_rep_3


title: SC, 17DMD, proliferation, biol rep 4

description:
Library name: Sample 12
Column name in Raw_counts_all samples.tsv: NR_17DMD_34


title: Fig2_293FT_ADAR2DD-E488Q_rep2


title: SC, 35WT, differentiation, biol rep 4

description:
Library name: Sample 8
Column name in Raw_counts_all samples.tsv: NR_35WT_30


title: Fig1_293FT_NT


title: SC, 16DMD, proliferation, biol rep 3

description:
Library name: Sample 11
Column name in Raw_counts_all samples.tsv: NR_16DMD_33


title: GSD1

description: Gene expression


title: RNA_seq_Bulk_Paneth cells_Young_Male_6_rep_1


title: RNA_seq_Bulk_Paneth cells_Young_Male_5_rep_2

value: 2 months


title: RNA_seq_Bulk_Paneth cells_Young_Male_5_rep_3


title: GSD17

description: Gene expression


title: 0-2 DAP-C

city: Mohali

first: Rita

last: Sharma

state: Punjab

address: Sector 81

department: Food Biotechnology

institute: National Agri-Food Biotechnology Institute

description: Transcriptome

data_processing:
Low-quality reads with Phred score<30 were filtered
Adapter triming using NGS QC Toolkit (v.2.3)
Read alignment to the reference genome using  HISAT2
Estimation of gene raw counts using FeatureCounts (v.2.2.0)
Pairwise differential expression analysis using DESeq2
Assembly: Phytozome v5.1
Supplementary files format and content: TPM values for each stage in excel format

organism: Sorghum bicolor

source_name: Pistil

growth_protocol: Soghum bicolor M35-1 plants were raised in an open field

extract_protocol:
Total RNA was isolated using a modified trizol method
Libraries were prepared using TruSeq® Stranded Total RNA kit and NEBNext® UltraTM II Directional RNA library prep kit using manufacturer’s instructions

value: Seed

value: After pollination

tag: cultivar


title: RNA_seq_Bulk_Paneth cells_Young_Male_5_rep_1


title: SC, 33WT, differentiation, biol rep 2

description:
Library name: Sample 6
Column name in Raw_counts_all samples.tsv: NR_33WT_28


title: Fig1_293FT_MCP-ADAR2DD-E488Q-A3FCDA1


title: RNA_seq_Bulk_Paneth cells_Young_Male_4_rep_1


title: 0-2 DAP-A

description: Transcriptome


title: Fig1_293FT_MCP-ADAR2DD-E488Q


title: SC, 14DMD, proliferation, biol rep 1

description:
Library name: Sample 9
Column name in Raw_counts_all samples.tsv: NR_14DMD_31


title: SC, 31WT, differentiation, biol rep 1

description:
Library name: Sample 5
Column name in Raw_counts_all samples.tsv: NR_31WT_27


title: 0-2 DAP-B

description: Transcriptome


title: SC, 34WT, differentiation, biol rep 3

description:
Library name: Sample 7
Column name in Raw_counts_all samples.tsv: NR_34WT_29


title: 1-2 DBP-C

description: Transcriptome

value: Pistil

value: Before pollination


title: GSD2

description: Gene expression


title: RNA_seq_Bulk_Paneth cells_Young_Male_3_rep_3


title: GSD18

description: Gene expression


title: GSD4

description: Gene expression


title: Fig1_293FT_MCP-ADAR2DD-E488Q-A3DCDA1


title: Fig2_293FT_ADAR2DD-E488Q-MCP


title: 1-2 DBP-B

description: Transcriptome


title: SC, 35WT, proliferation, biol rep 4

description:
Library name: Sample 4
Column name in Raw_counts_all samples.tsv: NR_35WT_26

value: Muscle tissue

value: Myogenic satellite cell


title: Ex-Fig1_293FT_NT


title: RNA_seq_Bulk_Paneth cells_Young_Male_3_rep_2


title: Ex-Fig1_293FT_MCP-ADAR2DD-E488Q


title: RNA_seq_Bulk_Paneth cells_Young_Male_4_rep_2


title: GBC15T

description: Gene expression


title: 1-2 DBP-A

description: Transcriptome


title: RNA_seq_Bulk_Paneth cells_Young_Male_2_rep_3


title: SAMPLE 7,SAMPLE 9,SAMPLE 11,SAMPLE 13

first: Zhengshi

address: No.301 Middle Yanchang Road

institute: Shanghai Tenth People's Hospital

description:
SC273
12a_1Tag05,12a_2 Tag06,12a_3 Tag07,12a_4 Tag08

data_processing:
Raw sequencing reads of the cDNA library were processed through the BD Rhapsody Whole Transcriptome Assay Analysis Pipeline (v1.8), which included filtering by reads quality, annotating reads, annotating molecules, determining putative cells and generating single-cell expression matrix. The pipeline also determined sample origin of every single cells via Sample determination algorithm according to the sequencing reads of SampleTag library. Among all the output files, Matrix of UMI counts for each gene per cell were used for downstream analysis. Genome Reference Consortium Human Build 38 (GRCh38) was used as reference for BD pipeline.
Assembly: GRCh38
Supplementary files format and content: Tab-separated values files and matrix files

source_name: Thyroid

extract_protocol:
Thyroid tissuewere cut into about 1-2mm3 pieces and digest in SoloTM Tumor Dissociation Kit (Sinotech Genomics, JZ-SC-58201) at 37°C for 30-60min. Then we stopped enzymatic digestion with excess RPMI-1640 medium, and filtered cell with 40 μm Cell strainer. Single cell solution were kept on ice before loading to BD Rhapsody cartridge for single cell transcriptome capturation.
Cells were firstly stained with two fluorescent dye, Calcein AM and Draq7, for precisely determination of cell concentration and viability via BD Rhapsody™ Scanner (BD Biosciences) before single cell multiplexing labelling. Cell of each sample were sequentially labeled with BD Human Single-Cell Multiplexing Kit (Cat. No. 633781) which utilizing the cell-hashing technology [1] mainly to provide higher sample throughput and eliminate batch effect for single cell library preparation and sequencing. Briefly, cells from each sample were labeled by antibodies with different sample tags respectively and washed twice with Stain Buffer (FBS) (BD Biosciences, Cat. No. 554656) before pooling all samples together. Pooled samples were loaded in one BD Rhapsody micro-well cartridge based on Fan et al [2]. Cell capture beads were then loaded excessively to ensure that nearly every micro-well contains one bead, and the excess beads were washed away from the cartridge. After lysing cells with lysis buffer, Cell capture beads were retrieved and washed prior to performing reverse transcription. Microbeads-captured single cell transcriptome and SampleTag molecules were generated into cDNA library and SampleTag library separately containing cell labels and unique molecular identifiers (UMI) information

value: Thyroid

value: Surgery


title: Ganglioneuroblastoma 20

type: genomic

first: Ying

last: Li

address: Lane 2555, Xiu-Pu Road, Pudong New District, Shanghai 200120, China

institute: Shanghai Cinopath Medical Laboratory Co., Ltd.

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
206350350009_R07C01

hyb_protocol: RA1 resuspended DNA was loaded onto BeadChips arrays. After overnight incubation at 48℃, single-base extension and allele-specific staining was performed on a Teflow chamber rack system (Tecan, Maennedorf, Switzerland).

scan_protocol: After allele-specific staining BeadChip arrays were coated with XC4/ethanol , dried for 1 hour and scanned on a NextSeq 550 (Illumina).

data_processing:
Genomic DNA extracted fromneuroblastoma or ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
Genotype: AA,AB,BB,or NC (no call)

label: Biotin and 2,4-Dinitrophenol

source_name: Ganglioneuroblastoma

label_protocol: 200ng of genomic DNA was whole-genome amplified in an overnight reaction at 37℃ using amplification master mix (AM1) and primer/neutralization mix (MA2-MSM). After incubation the amplified DNA was fragmented with fragmentation mix (FAM), precipitated with isopropanol and precipitation mix (PA1) and resuspended in resuspension solution (RA1).

extract_protocol: Add Deparaffinization solution at 56℃ to get rid of paraffin on the surface of the sample, add buffer FTB, a combination of RNA-free enzyme water and proteinase K cleavage, incubate at 90℃ for 1h to decrosslink, and add proteinase K again to cleave the tissue by incubation at 65℃, and then transfer it to the adsorption column to filter out impurities.

value: Male

value: Ganglioneuroblastoma

value: carcinoma


title: RNA_seq_Bulk_Paneth cells_Young_Male_2_rep_2

value: male


title: SC, 31WT, proliferation, biol rep 1

description:
Library name: Sample 1
Column name in Raw_counts_all samples.tsv: NR_31WT_23

value: WT


title: Ganglioneuroblastoma 19

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205065750048_R04C01


title: pig abdominal adhesion tissue scRNA-seq Cntrl group replicate 3

city: Stanford

first: Michael

last: Januszyk

state: CA

address: 257 Campus Dr

institute: Stanford University

data_processing:
FASTQ files were aligned to the pig genome (sscrofa11.1) using Cell Ranger (10X Genomics; version 7.1.0).
Cell barcodes representative of quality cells were differentiated from apoptotic cell barcodes or background RNA based on a threshold of having at least 500 unique transcripts profiled and less than 50,000 total transcripts. Unique molecular identifiers (UMIs) from each cell barcode were retained for all downstream analysis, normalized with a scale factor of 10,000 UMIs per cell, and subsequently natural log transformed with a pseudocount of 1 using the R package Seurat (version 5.0.1) (Chen et al., 2013).
The first 30 principal components of the aggregated data were then used for uniform manifold approximation and projection (UMAP) analysis. Cell annotation was performed manually. Cell-type marker lists were generated using Seurat’s native FindMarkers function with a log fold change threshold of 0.25 using the ROC test to assign predictive power to each gene.
Assembly: Sus_scrofa.Sscrofa11
Supplementary files format and content: 10X genomics CellRanger output triad (barcodes/features/matrix) from the cannonical outs/filtered_feature_bc_matrix folder, all in their .gz formats

source_name: abdominal adhesion tissue

extract_protocol:
Pig abdominal adhesions specimens were harvested fresh and immediately minced on ice. The tissue was then digested for 40 min in a 37 °C water bath agitator in 1 mg/mL collagenase (collagenase type IV, ThermoFisher) digest buffer in Medium 199 (HyClone, GE Healthcare) consisting of 3-5% fetal bovine serum (Gibco FBS, ThermoFisher), DNase I (Worthington), Poloxamer 188 (Cat. P5556-100ML, Sigma), HEPES, and CaCl2. The digest was quenched with quench media (DMEM [Gibco DMEM, ThermoFisher] with 10% FBS), then centrifuged at 300 × g for 5 min at 4 °C, resuspended in quench media, and filtered through 100, 70, and 40 μm cell strainers (Falcon cell strainer, ThermoFisher). Cells were counted and re-suspended for further processing.
Single-cell RNA-seq (scRNA-seq) was then performed at the Stanford Genomics Facility (SGF) for droplet-based microfluidics using the 10x Chromium Single Cell platform (Single Cell 3’ v3, 10x Genomics, USA). Droplets of the cellular suspensions, reverse transcription master mix, and partitioning oil were mixed, loaded onto a single cell chip, and processed on the Chromium Controller. Reverse transcription was performed, and cDNA was amplified using a BioRad C1000 Touch thermocycler, with cDNA size selected using SpriSelect beads (Beckman Coulter, USA). An Agilent Bioanalyzer High Sensitivity DNA chip was used to analyze cDNA for qualitative control purposes; cDNA was then fragmented using the proprietary fragmentation enzyme blend for 5min at 32°C, followed by end repair and A-tailing at 65°C for 30 min. DNA was double-sided size selected using SpriSelect beats. Sequencing adaptors were ligated to the cDNA at 20°C for 15min. cDNA was amplified using a sample-specific index oligo as primer, followed by another round of double-sided size selection using SpriSelect beads.
Final libraries were analyzed on an Agilent Bioanalyzer High Sensitivity DNA chip for qualitative control purposes. Libraries were sequenced on a NextSeq 500 Illumina platform targeting 50,000 reads per cell.

treatment_protocol: Adhesion model pigs treated with T-5224-hydrogel following laparotomy vs control (saline irrigation, surgical standard of care)

value: abdominal adhesion tissue

value: all cells (unsorted)

value: yukutan minipig


title: SAMPLE 3, SAMPLE 15

description:
SC126
21c Tag02 ,21a Tag01

source_name: Thyroid,Lymph node

value: Thyroid,Lymph node


title: RNA_seq_Bulk_Paneth cells_Young_Male_2_rep_1

value: 2 months

value: Paneth cells


title: SAMPLE 8,SAMPLE 10,SAMPLE 12, SAMPLE 14

description:
SC261
12b_1Tag9,12b_2 Tag10,12b_3 Tag11,12b_4 Tag12


title: Ganglioneuroblastoma 18

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205329990028_R08C01


title: pig abdominal adhesion tissue scRNA-seq Cntrl group replicate 4


title: Neuroblastoma 25

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
204127480025_R08C01

source_name: Neuroblastoma

value: Neuroblastoma


title: SC, 34WT, proliferation, biol rep 3

description:
Library name: Sample 3
Column name in Raw_counts_all samples.tsv: NR_34WT_25

value: Proliferation


title: SAMPLE 1,SAMPLE 2,SAMPLE 4,SAMPLE 5, SAMPLE 6

description:
SC96
1a Tag01,1b Tag02,4a Tag03,4b Tag04,4c Tag12


title: pig abdominal adhesion tissue scRNA-seq Cntrl group replicate 2


title: Neuroblastoma 23

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
206350350126_R08C01


title: Neuroblastoma 24

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205495650030_R08C01


title: RNA_seq_Bulk_Paneth cells_Young_Male_1_rep_1


title: RNA_seq_Bulk_Paneth cells_Young_Male_1_rep_2


title: RNA_seq_Bulk_Paneth cells_Young_Male_3_rep_1


title: BM mononuclear cells from patient 8, enriched in the CD34+ HSPC fraction, scRNAseq

source_name: Bone marrow

value: Bone marrow

value: human CD34+/-


title: pig abdominal adhesion tissue scRNA-seq Cntrl group replicate 1


title: pig abdominal adhesion tissue scRNA-seq Rx group replicate 4


title: Neuroblastoma 22

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
202560410027_R03C01

value: Female


title: RNA_seq_Bulk_Progenitor cells_Aged_Female_5

source_name: Progenitor cells

value: 23 months

value: female

value: Progenitor cells


title: SC, 33WT, proliferation, biol rep 2

description:
Library name: Sample 2
Column name in Raw_counts_all samples.tsv: NR_33WT_24

value: Muscle tissue

value: Myogenic satellite cell


title: BM mononuclear cells from patient 5, enriched in the CD34+ HSPC fraction, scRNAseq


title: BM mononuclear cells from patient 3, enriched in the CD34+ HSPC fraction, scRNAseq


title: pig abdominal adhesion tissue scRNA-seq Rx group replicate 3


title: RNA_seq_Bulk_Progenitor cells_Aged_Female_4


title: BM mononuclear cells from patient 9, enriched in the CD34+ HSPC fraction, scRNAseq


title: Neuroblastoma 21

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205065750048_R03C01

value: Male

value: carcinoma


title: RNA_seq_Bulk_Paneth cells_Young_Male_1_rep_3

value: male


title: BM mononuclear cells from patient 2, enriched in the CD34+ HSPC fraction, scRNAseq


title: pig abdominal adhesion tissue scRNA-seq Rx group replicate 1

value: abdominal adhesion tissue

value: all cells (unsorted)

value: yukutan minipig


title: RNA_seq_Bulk_Progenitor cells_Aged_Female_3


title: Ganglioneuroblastoma 17

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
206443730158_R05C01

value: Ganglioneuroblastoma


title: RNA_seq_Bulk_Progenitor cells_Aged_Female_1


title: RNA_seq_Bulk_Progenitor cells_Aged_Female_2


title: BM mononuclear cells from patient 1, enriched in the CD34+ HSPC fraction, scRNAseq


title: Ganglioneuroblastoma 16

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
206443730125_R04C01


title: Zebrafish, RIP-seq, Rbm24a Knock-in IP

city: Tsingtao

first: li

middle: pan

last: feng

state: shandong

address: Ao Shan Wei Street

department: College of Life Sciences

institute: shandong university

data_processing:
Each library was amplified by a 15-cycle PCR, and 150 bp paired-end sequencing was subjected to Illumina Nova seq 6000 to obtain the raw data.
Assembly: GRCz11
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample
Supplementary files format and content: Bigwig file for each Sample

organism: Danio rerio

source_name: whole embryo

growth_protocol: Zebrafish embryos were maintained in 1/3x Ringer's at 28.5C

extract_protocol:
Total RNAs were extracted from WT and Mrbm24a embryos at 4-cell sphere 24-hpf stage, using TRIzol reagent (Invitrogen), and were precipitated by cold isopropanol (50%, v/v) in the presence of carrier glycogen (20 µg/each sample).
1 μg total RNA was used for following library preparation. The poly(A) mRNA isolation was performed  using Oligo(dT) beads. The mRNA and RIP IP RNA fragmentation was performed using divalent cations and high  temperature. Priming was performed using Random Primers. First strand cDNA and the second-strand  cDNA were synthesized. The purified double-stranded cDNA was then treated to repair both ends and  add a dA-tailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Size selection of  Adaptor-ligated DNA was then performed using DNA Clean Beads. Each sample was then amplified by  PCR using P5 and P7 primers and the PCR products were validated.  Then libraries with different indexs were multiplexed and loaded on an Illumina Nova seq 6000 instrument for sequencing using a 2x150 paired-end (PE) configuration according to  manufacturer’s instructions.

value: whole embryo

tag: developmental stage

value: 4-cell stage

value: embryonic cell

value: Rbm24a Knock-in

tag: antibody

value: GFP


title: Sorted human CD34+ HSPCs from the BM of UBA1mut hematochimeric mice, scRNAseq

value: Bone marrow

value: Hematochimeric mice_A

value: human CD34+


title: Neuroblastoma 20

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205329990033_R01C01

value: Female

value: Neuroblastoma


title: GBC5N

description: Gene expression

value: Control


title: Ganglioneuroblastoma 15

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
206350350009_R04C01

value: Male

value: carcinoma


title: RNA_seq_Bulk_Progenitor cells_Aged_Male_5

value: 24 months

value: male

value: Progenitor cells


title: Sorted human CD34+ HSPCs from the BM of UBA1wt hematochimeric mice, scRNAseq

value: Hematochimeric mice_C


title: Zebrafish, RIP-seq, Rbm24a Knock-in input

value: whole embryo

value: input


title: Zebrafish, RIP-seq, wild type IP

value: 4-cell stage

value: embryonic cell

value: wild type


title: RNA_seq_Bulk_Progenitor cells_Aged_Male_4

value: 23 months


title: Human BM cells from UBA1 wt mice, scRNAseq

value: Bone marrow

value: human CD34+/-


title: Ganglioneuroblastoma 14

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205495650067_R05C01

value: Ganglioneuroblastoma


title: Neuroblastoma 19

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205841280139_R06C01

value: Neuroblastoma


title: RNA_seq_Bulk_Progenitor cells_Aged_Male_3

value: 26 months


title: Human BM cells from UBA1 mut mice, scRNAseq


title: Zebrafish, RIP-seq, wild type input


title: Neuroblastoma 18

description:
Genomic DNA extracted from neuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
206863220159_R03C01

value: Male


title: RNA_seq_Bulk_Progenitor cells_Aged_Male_2


title: pig abdominal adhesion tissue scRNA-seq Rx group replicate 2

value: abdominal adhesion tissue

value: all cells (unsorted)

value: yukutan minipig


title: Ganglioneuroblastoma 13

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
206535470151_R05C01

value: carcinoma


title: RNA_seq_Bulk_Progenitor cells_Aged_Male_1

value: 27 months

value: male

value: Progenitor cells


title: Ganglioneuroblastoma 12

description:
Genomic DNA extracted from ganglioneuroblastoma was genotyped using CytoSNP-850K v1.2 Genotyping BeadChips (Illumina).
205841280011_R08C01

value: Female