title: FNC_3

data_processing:
CircRNAs were identified and quantified using pseudo-reference annotated reads from A-tailing libraries by CARP [35]. First, candidate circRNA identification was performed by applying four different algorithms with default parameters (CIRCexplorer2, CIRI-quant, find_circ and MapSplice) on A-tailing RNA-seq reads. Candidate circRNAs identified in each sample were then pooled and “CARP PseudoRef” was utilized to construct a pseudo-reference covering 149 bp of the flanking BSJ site for each candidate circRNA. RNA-seq reads were aligned to the pseudo-reference to detect candidate BSJ reads by “CARP Mapping.” “CARP Remap” was used to detect and filter out false reads that could also be mapped to the genome or transcriptome. The remaining reads were used for circRNA quantification by “CARP ReadsCount.” To determine if the circRNA was enriched in the A-tailing library, the ratio of the reads count in the A-tailing library to the control library was calculated. Expression of the last exon was calculated to represent linear RNA expression by CARP. The ratio of the last exon of the host gene, which does not form a circRNA, was also calculated for comparison by CARP. To remove circRNAs that are sensitive to A-tailing/RNase R treatment, a cutoff was set for A-tailing/Control ratio for the last exons > 95%. CircRNAs with A-tailing/Control ratios higher than the cutoff were considered Bona fide circRNAs while those with ratios lower than the cutoff were considered false positives and removed, guaranteeing an FDR < 0.05.
Assembly: mm9
Supplementary files format and content: genes.fpkm is gene expression for each total RNA-seq library.

source_name: CAD

extract_protocol:
Mouse brain cortex, hippocampus tissue samples, as well as CAD and N2a cell pellets were homogenized in 1mL TRIzol (Invitrogen, 15596018) and incubated at room temperature for 5 min. 200µL Chloroform was added, thoroughly mixed, incubated at room temperature for 15 min, and centrifuged at 13,000g for 20 min at 4ºC.  Total RNA was precipitated from 600µL supernatant in 60µL NaAc (3M, pH 5.2), 4µL glycogen (5 mg/mL), and 600µL 100% Isopropanol for at least 30 min at -80ºC. 500ng RNA was reverse transcribed (RT) using SuperScript III First Stand Synthesis System (Invitrogen, 18080051) with random hexamers for downstream qPCR analysis. All RT and qPCR experiments were performed in triplicate and analyzed using the relative quantification (ΔΔCt) method with Actin or Gapdh as the normalization control. All tested primers are provided in Additional File 1. To quantify circRNA levels, divergent primers were designed to span the circRNA back splice junction (BSJ) specifically amplifying the circRNAs and not the counterpart linear RNA.
A-tailing RNA was quantified by Qubit RNA High Sensitivity Assay (Thermo Fisher Scientific) and quality was determined by Bioanalyzer 2100 Eukaryote Total RNA Pico (Agilent Technologies). rRNA depleted RNA-seq libraries were generated according to previously described protocols [35]. Briefly, the Ribo-Zero rRNA Removal Kit was used to deplete ribosomal RNAs. Libraries were constructed using the NEBNext UltraTM RNA Library Prep Kit according to the manufacturer’s instructions. Library quality and concentrations were assessed by Tapestation High Sensitivity D1000 ScreenTapes (Agilent Technologies, 5067-5584) and qPCR, respectively. Libraries were equimolarly pooled and sequenced on the Hiseq platform with 150 PE/read length and a target of 80M total reads (40M in each direction) per sample (Admera Health LLC).

value: CAD

value: si NC


title: 5CG12

status: Public on Oct 22 2024

type: SRA

city: Atlanta

first: Bing

last: Yao

address: 615 Michael Street

department: Department of Human Genetics

country: USA

institute: Emory University

library_source: transcriptomic

data_processing:
CircRNAs were identified and quantified using A-tailing reads by CARP28. Candidate circRNA identification was performed by applying four different algorithms with default parameters (CIRCexplorer2, CIRI-quant, find_circ and MapSplice) on A-tailing RNA-seq reads. Candidate circRNAs identified in each sample were pooled and “CARP PseudoRef” was utilized to construct a pseudo-reference covering 149bp of the flanking BSJ site for each candidate circRNA. RNA-seq reads were aligned to the pseudo-reference to detect candidate BSJ reads by “CARP Mapping.” “CARP Remap” was used to detect and filter out false reads that could also be mapped to the genome or transcriptome. The remaining reads were used for circRNA quantification by “CARP ReadsCount.”
Normalized reads count for each circRNA obtained by “CARP ReadsCount” were used for DE circRNA analysis by Z-Test approached used in our published circMeta algorithm82. Significant DE circRNAs were defined by FDR < 0.05.
CircRNA full-length was determined by “CARP CircAS” using A-tailing RNA-seq data. Junction reads in the circRNA body were summarized and Z-test was used for circRNA isoform switch detection using P < 0.05 and |Inclusion level difference| > 20% as cutoff.
Sequencing reads from the control library were aligned to mm9 by TopHat 2.1.0 using default parameters. Aligned bam files were used for DE genes between 5xFAD and CTL at each time point using Cuffdiff. DE genes were defined by q-value < 0.05.
Assembly: mm9
Supplementary files format and content: CircRNA expression in A-tailing library are stored in CircRNA.5mCTL.5mFAD.7mCTL.7mFAD.txt
Supplementary files format and content: Gene expression in non-treated libearies are stored in Gene.5mCTL.5mFAD.7mCTL.7mFAD.txt

molecule: total RNA

organism: Mus musculus

source_name: Cortex

extract_protocol:
Mouse brain cortex samples or cell pellets were homogenized in 1mL TRIzol (Invitrogen) and incubated at room temperature for 5min. 200µL Chloroform was added in sample’s tube, thoroughly mixed, and incubated at room temperature for 15min. Next, all samples were centrifuged at 13,000g for 20min at 4ºC.  The total RNA was precipitated in 60µL NaAc (3M, pH 5.2), 4µL glycogen (5 mg/mL), and 600µL 100% Isopropanol for at least 30min at -80ºC with 600µL supernatant.
4µL E-PAP and 40U RNase inhibitor were added to 10µg total RNA in a 50µL reaction by following the poly(A) tailing kit (Thermo Fisher Scientific). The reactions were incubated for 1hour at 37ºC, purified and eluted with 25µL nuclease-free water. For the RNase R treatment, the RNA products were treated with 10U RNase R, 3µL 10X RNase R Buffer with specific LiCl (0.2M Tris–HCl pH 8.0, 1mM MgCl2, and 1M LiCl), and 80U RiboLock RNase Inhibitor (40 U/μL, Thermo Fisher Scientific). Finally, the RNA products were purified and eluted in 30µL nuclease-free water to proceed the rRNA-depleted RNA-seq library.
We used the Qubit RNA High Sensitivity Assay to quantify the A-tailing RNA y ) and the Bioanalyzer 2100 Eukaryote Total RNA Pico to value the quality of the A-tailing RNA. The Ribo-Zero rRNA Removal Kit was used to deplete ribosomal RNAs and the NEBNext UltraTM RNA Library Prep Kit was used to prepare libraries.

tag: tissue

value: Cortex

tag: genotype

tag: treatment

value: NO


title: WT5 WGBS

city: Valencia

first: Natalia

last: Tapia

state: València

address: Jaime Roig 11

country: Spain

institute: Spanish National Research Council

library_source: genomic

data_processing:
FastQC (fastqc_v0.11.8) was used to perform basic statistics on the quality of the raw reads.
Reads were pre-processed through Trimmomatic (Trimmomatic-0.36) software use the parameter (SLIDINGWINDOW: 4:15 ; LEADING:3, TRAILING:3 ; ILLUMINACLIP: adapter.fa: 2: 30: 10 ; MINLEN:36)
Bismark software (version 0.24.0; Krueger et al., 2011) was used to perform alignments of bisulfite-treated reads to the reference genome. The reference genome was firstly transformed into bisulfite-converted version (C-to-T and G-to-A converted) and then indexed using bowtie2 (Langmead et al., 2012).
Sequence reads that produced a unique best alignment from the two alignment processes (original top and bottom strand) were then compared to the normal genomic sequence and the methylation state of all cytosine positions in the read was inferred. The same reads that aligned to the same regions of genome were regarded as duplicated ones.
Assembly: grcm38
Supplementary files format and content: bigwig

molecule: genomic DNA

source_name: E18.5 isolated villi from intestinal epithelium

growth_protocol: Tissue, no cell culture.

extract_protocol:
Genomic DNA was extracted using QIAamp DNA micro kit (Qiagen) and bisulfite treated using EZ DNA Methylation Gold Kit (Zymo Research).
Genomic DNA spiked with lambda DNA were fragmented to 200-400 bp. The DNA fragments were bisulfite treated to generate single strand DNA. Methylation sequencing adapters were ligated, followed by double strand DNA synthesis. The library was ready after size selection and PCR amplification.

treatment_protocol: The villi of E18.5  intestinal epithelium was isolated as follows. Briefly, the small intestine was dissected from E18.5 embryos, opened longitudinally, and placed on ice-cold PBS containing 30mM EDTA and 1.5mM DTT for 20min on ice. Next, the intestine was transferred to a tube containing 30mM EDTA in PBS, incubated at 37ºC for 10min and, then, vigorously shaken for 30s to release the intestinal epithelium from the lamina propia. After removing the intact intestinal muscle, the villi suspension was used for DNA extraction and WGBS.

tag: strain

value: C57BL6/JRccHsd N>5

value: wild-type


title: CHA447-3B

city: Rochester

first: Surendra

last: Dasari

state: MN

address: 200 First Street SW

department: Quantitative Health Sciences Research

institute: Mayo Clinic

data_processing:
Data were processed using a previoulsy published method[https://pubmed.ncbi.nlm.nih.gov/24972667/]. In brief, reads were quality controlled using FASTQC. Reads were mapped against hg38 genome using STAR aligner. HT-Seq was utilized to analyzed the aligned reads and generate gene-wise counts using Ensemble version 78 transcriptome. Extracted read data was processed using edgeR software. Timmed means of M-values method was utilized to normalized reads that were provided in the file named 20231208-For-GEO-Raw-Counts.tsv".
Assembly: hg38
Supplementary files format and content: 20231208-For-GEO-Raw-Counts.tsv; tab-delimited; Raw gene-count matrix

organism: Homo sapiens

source_name: Skeletal muscle

extract_protocol:
mRNA was extracted using Trizol and stored in RNAlater.
mRNA library was constructed using Illumina TruSeq Stranded kit following manufacturer's recommendations.
Paired end library strategy was utilized

value: Skeletal muscle

tag: age

value: older

tag: Sex

value: male

tag: training time_point


title: 5CG8

value: WT


title: 5CG9


title: 5AT18

value: A-tailing Rnase R


title: FNC_2

tag: cell line


title: HOO343-2B


title: CUR267-1B

value: young

value: female

value: rest


title: GM19209_chr_rep1b

city: Sherbrooke

first: Karine

last: Choquet

state: Québec

address: 3201 rue Jean-Mignault

department: Biochemistry and Functional Genomics

country: Canada

institute: Université de Sherbrooke

library_source: other

data_processing:
Basecalling + poly(A) tail length estimation (Dorado v0.7.0)
Alignment (minimap2)
Determination of read splicing status (custom scripts available at https://github.com/churchmanlab/nano-COP)
Assembly: GRCh38
Supplementary files format and content: *_hg38_multi_introns_isoforms_counts.withSKP.RefSeq.txt: text files containing the read count for full-length splicing statuses
Supplementary files format and content: *_polyA_estimates.tsv: text files containing the poly(A) tail length estimate for each read
Supplementary files format and content: *_read_counts_per_gene_and_allele.RefSeq_all.txt: text file containing the read counts per allele
Library strategy: Direct RNA nanopore sequencing

molecule: other

source_name: Lymphoblastoid cell line

extract_protocol:
Cellular fractionation to separate cytoplasm and chromatin, Qiazol extraction to isolate cytoplasmic and chromatin-associated RNA
Poly(A) selection followed by direct RNA sequencing library preparation (SQK-RNA002)

value: Lymphoblastoid cell line


title: CHA447-2B


title: GM19144_chr_rep1b


title: WT3 WGBS

value: E18.5 isolated villi from intestinal epithelium


title: 5CG14


title: 5CG7


title: CHA447-1B


title: 5AT16


title: COR775-1B


title: GM19152_chr_rep1b


title: FKD_3

value: si circGIGYF2


title: 5AT17


title: CLO122-1B


title: CAM113-2B


title: 5AT15


title: GM18861_chr_rep1b


title: CAM113-1B


title: FKD_2

value: CAD


title: GM18853_chr_rep1b


title: BUR210-3B


title: FKD_1


title: CG_WT_3

data_processing:
CircRNAs were identified and quantified using pseudo-reference annotated reads from A-tailing libraries by CARP [35]. First, candidate circRNA identification was performed by applying four different algorithms with default parameters (CIRCexplorer2, CIRI-quant, find_circ and MapSplice) on A-tailing RNA-seq reads. Candidate circRNAs identified in each sample were then pooled and “CARP PseudoRef” was utilized to construct a pseudo-reference covering 149 bp of the flanking BSJ site for each candidate circRNA. RNA-seq reads were aligned to the pseudo-reference to detect candidate BSJ reads by “CARP Mapping.” “CARP Remap” was used to detect and filter out false reads that could also be mapped to the genome or transcriptome. The remaining reads were used for circRNA quantification by “CARP ReadsCount.” To determine if the circRNA was enriched in the A-tailing library, the ratio of the reads count in the A-tailing library to the control library was calculated. Expression of the last exon was calculated to represent linear RNA expression by CARP. The ratio of the last exon of the host gene, which does not form a circRNA, was also calculated for comparison by CARP. To remove circRNAs that are sensitive to A-tailing/RNase R treatment, a cutoff was set for A-tailing/Control ratio for the last exons > 95%. CircRNAs with A-tailing/Control ratios higher than the cutoff were considered Bona fide circRNAs while those with ratios lower than the cutoff were considered false positives and removed, guaranteeing an FDR < 0.05.
Assembly: mm9
Supplementary files format and content: genes.fpkm is gene expression for each total RNA-seq library. DEseq.xls is circRNA expression and DE analysis for A-tailing library.

source_name: Hippocampus

value: Hippocampus

value: 7 months


title: BUR210-2B


title: 5AT14

value: Cortex


title: 5AT13


title: GM18510_chr_rep1b


title: CG_WT_2


title: 5AT12


title: GM18501_chr_rep1b


title: 5AT11


title: FNC_1

value: si NC


title: CG_WT_1


title: BUR210-1B


title: BUC505-3B


title: GM19209_chr_rep1a


title: CG_FAD_3


title: 5AT10


title: BUC505-2B


title: GM19144_chr_rep1a


title: 5AT9


title: BUC505-1B


title: GM19152_chr_rep1a


title: GM18522_chr_rep1b


title: BRA782-3B


title: 5AT8


title: AT_WT_3


title: 5AT7


title: GM18861_chr_rep1a


title: BRA782-2B


title: AT_WT_2


title: GM18522_chr_rep1a


title: BRA782-1B


title: Differentiated Spermatogonia H3K36me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 2

city: Basel

first: Antoine

last: Peters

address: Maulbeerstrasse 66

country: Switzerland

institute: Friedrich Miescher Institute

data_processing:
Sequence reads were trimmed for adaptor sequence/low-quality sequence using TrimGalore (--quality 20 --illumina --stringency 3 )
Trimmed reads were then aligned to the mouse genome build mm10 using STAR (version 2.5.0a) with settings (--alignIntronMin 1 --alignIntronMax 1 --alignEndsType EndToEnd --alignMatesGapMax 1000 --outFilterMatchNminOverLread 0.85)
Aligned reads were deduplicated using SAMtools (version 1.10) with standard settings
Uniquely mapped reads were counted on genomic regions using the function qCount (mapqMin = 255L) from QuasR package (version 1.36.0).
Assembly: mm10
Supplementary files format and content: BigWig (.bw) processed files were generated from BAM files using the function qExportWig from QuasR package (version 1.36.0) with standard settings

source_name: Differentiated Spermatogonia

extract_protocol:
For spermatogonia, testicular cell suspension was prepared by incubation of seminiferous tubules in 200 U/ml Collagenase type I (Worthington Biochemical LS004196), 5 μg/ml DNAse I (Roche 10104159001), and 0.05%Trypsin (Gibco 25200056) in GBSS (Sigma G9779) as described in (60). Cells were stained with 1/200 anti-CD117/c-kit PE (eBioscience 12-1171-83), anti-CD324/E-Cadherin eFluor 660 (eBioscience 50-3249-82) and anti-CD49f/Integrin alpha 6 PE-Cyanine7 (eBioscience 25-0459-82) for 1 hour at 32oC with constant shaking and protected from light. After washing cells were incubated with 20 μg/ml with Hoechst 33342 (Thermo Fischer Scientific H3570) for 1 hour at 32oC with constant shaking and protected from light and then with 30nM DRAQ7 (Biostatus DR71000) for 5 minutes on bench. The stained cells were strained through a 40 μm nylon filter into 5 ml polypropylene tubes and were sorted on a BD FACSAria III cell sorter fitted with a 70 μm nozzle (Becton Dickinson) using a 375nm laser to excite Hoechst 33342, a 561nm laser to excite PE and PE-Cy7 and a 633nm laser to excite DRAQ7 and eFluor600. Cells were first gated for FSC and SSC to exclude debris. Then the live cells were selected based on the absence of DRAQ7 signal detected using 755LP, 780/60BP. Then we gated for cells positive for Hoechst 33342 emission which was detected using a 670LP (Hoechst-Red) and 450/20 BP (Hoechst-Blue). We gated for the Hoechst-Redlow and Hoechst-Bluemid population which is enriched for 2N spermatogonia. Fluorescence for PE was detected using a 582/15BP filter, for PE-Cy7 was detected using 735LP,780/60BP filter and for eFluor660 using a 660/20BP filter. Undifferentiated spermatogonia were sorted as CD324high, CD49fhigh, CD117low and differentiated spermatogonia were sorted as CD324low, CD49flow, CD117high. Briefly 5000-7500 spermatogonia cells were FACS-sorted directly in 20 μl nuclear isolation buffer (Sigma-Aldrich NUC101) in Eppendorf® LoBind microcentrifuge tubes. The nuclei were centrifuged at 500g for 5 minutes at 4°C in a swing bucket centrifuge and the volume of the sample was reduced to 10 μl. The sample was snap frozen in liquid nitrogen and stored at –80°C freezer until further processing. The sample was thawed on ice and 1 μl of TS buffer (5% Triton X-100, 5% Sodium deoxycholate) was added for 10 minutes to ensure permeabilization of nuclei. 40 μl of MNAse digestion buffer (3.33 gelUnits/μl Micrococcal Nuclease (NEB M0247S), 1.25X Micrococcal Nuclease buffer, 6.25% PEG 6000, 0.85mM DTT) was added to the sample for 10 minutes at 25 °C to fragment the chromatin. Reaction was stopped by addition of 5 μl 100 uM EDTA and the sample was placed on ice. The sample was diluted by addition of 145 μl ChIP buffer (20 mM Tris-HCl pH 8.0, 2 mM EDTA, 150 mM NaCl, 0.1% Triton X-100, 1X EDTA-free protease inhibitor cocktail). To ensure release of chromatin fragments, the sample was sonicated using the Diagenode Bioruptor Plus® for 3 cycles of 5 seconds ON and 30 seconds OFF at low output. Then chromatin was pre-cleared with 10 μl of 1:1 protein A:protein G Dynabeads (Thermo Fisher Scientific, 10001D and 10003D). The precleared chromatin sample was incubated with 0.5 μg of anti-H3K4me3 (Millipore 17-614) or 0.75 μg anti-H3K36me3 (Cell Signaling #4909) conjugated with 10 μl of 1:1 protein A: protein G Dynabeads overnight at 4 °C on a rotator. Chromatin-Antibody-Dynabeads complexes were washed once with 500 μl of ChIP buffer, 3 times with 1ml of Low Salt wash buffer (20 mM Tris-HCl (pH 8.0), 0.05% SDS, 1% Triton X-100, 2 mM EDTA and 150 mM NaCl) and 3 times with 1 ml High Salt buffer (20 mM Tris-HCl (pH 8.0), 0.05% SDS, 1% Triton X-100, 2 mM EDTA and 250 mM NaCl). Chromatin was eluted in 40 μl of Elution buffer (100 mM NaHCO3 and 1% SDS) for 90 minutes at 65 °C. DNA from eluted material was purified by 2X volumes of Ampure XP DNA purification beads (Beckman Coulter, A63881) and resuspended in 25 μl nuclease free H2O.
NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® (NEB E7645L)

tag: chip antibody

tag: cell type

value: Differentiated Spermatogonia

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg


title: GM18853_chr_rep1a


title: CG_FAD_2


title: AT_WT_1


title: GM18510_chr_rep1a


title: AT_FAD_3


title: Differentiated Spermatogonia H3K36me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 1


title: Differentiated Spermatogonia H3K36me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 2

value: Dnmt3a f/+ ; Dnmt3b f/+


title: BRA730-2B

value: Skeletal muscle

value: older

value: male


title: AT_FAD_2


title: GM18501_chr_rep1a


title: BRA730-1B


title: Differentiated Spermatogonia H3K36me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 1


title: BOH319-2B


title: AT_FAD_1


title: CG_FAD_1


title: CSL, scRNAseq

city: Zwijnaarde

first: Jordy

last: De Coninck

address: Technologiepark 71

department: Department of Biomedical Molecular Biology

country: Belgium

institute: University Gent

description: 10x Genomics

library_source: transcriptomic single cell

data_processing:
Demultiplexing was done with 10X CellRanger software (10x Genomics, Cellranger mkfastq)
Obtained reads were used in the CellRanger count pipeline (v 4.0.0) which performs alignment against mouse reference genome (GRCm38), filtering, counting and unique molecular identifier (UMI) counting.
Assembly: GRCm38.99 (with custom GFP)
Supplementary files format and content: .h5 file containing raw gene count matrices
Supplementary files format and content: .rds Seurat object

molecule: polyA RNA

source_name: Skin

extract_protocol:
The tumors developed in K14CreTg/+-Rosa26-Zeb2Tg/+-Trp53Fl/Fl mice were dissected, measured, finely chopped, and digested in 2 mg/ml collagenase A (Roche), 0.125 U/ml dispase II (Roche) and 1 µg/ml DNAse I (Roche) for 1 h at 37°C with gentle agitation. The digested tumor was passed through a 100-µm cell strainer, and the cells were washed in PBS containing 2% FCS. The cell suspension was stained DAPI for 1 h at 4°C. Tumor cells were sorted on a BD FACS ARIA II (BD bioscience) by forward and side scatter, doublet discrimination and DAPI exclusion. Tumor cells were selected by GFP positivity. The GFP+ tumor cells were cultured in DMEM supplemented with 10% FCS, penicillin/streptomycin, non-essential amino acids, and Na-pyruvate at 37°C under 5% CO2.
Library prep was performed by the VIB Single Cell Core (VIB, Ghent-Leuven, Belgium). Sequencing libraries were loaded at 2.1pM loading concentration on a HiSeq4000 with custom sequencing metrics (single-indexed sequencing run, 28/8/0/98 cycles for R1/i7/i5/R2) (Illumina).

value: Skin

value: Epithelial

value: Krt14Cre Tg/+ -Rosa26-Zeb2 Tg/+ /Trp53 Fl/Fl

tag: group

value: MIX Tumor


title: Undifferentiated Spermatogonia H3K36me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 2

source_name: Undifferentiated Spermatogonia

value: Undifferentiated Spermatogonia


title: BIL937-2B

value: young

value: female


title: BOH319-1B

value: rest


title: LA491, KT;Stag2 flox, 16 weeks

city: La Jolla

first: Jesse

middle: R

last: Dixon

state: CA

address: 10010 N. Torrey Pines Rd.

institute: Salk Institute for Biological Studies

description: Hi-C library

data_processing:
Hi-C  data were aligned to the mm10 reference genome using BWA-MEM. Reads were filtered (MAPQ >= 30) and manually paired. PCR duplicate reads were removed using Picard. Contact matrices were generated and normalized using the iterative correction method.
Assembly: mm10
Supplementary files format and content: mcool files
Supplementary files format and content: hic files

source_name: Lung

extract_protocol:
Tumors were dissociated using collagenase IV, dispase, and trypsin at 37 degrees C for 30 minutes as previously described (40). Cells were stained with DAPI and antibodies against CD45 (30-F11), CD31 (390), F4/80 (BM8), and Ter119 (all from BioLegend) to exclude hematopoietic and endothelial cells. FACSAria sorters (BD Biosciences) were used for cell sorting.
In situ Hi-C libraries were prepared from FACS-sorted neoplastic cells with low input ranging from 2 x 105 to 4 x 105 cells. In detail, cells were first pelleted at 300 G and resuspended in 1 mL Room Temp PBS containing 3% BSA, then fixed as previously described, (Rao et al. Cell 2014) only with a higher centrifuge speed at 2,500 G for pelleting the fixed cells. Fixed cell pellets were processed through the standard Arima Hi-C kit (Catalog # A510008), until the step of Covaris DNA sonication to an average size of 400 bp. Sheared DNA was purified and size-selected by a two-step AMPure XP bead (Beckman Coulter #A63882) cleanup at 0.6x and 1.0x, and then inputted into the 2S Plus DNA Library Kit from Integrated DNA Technologies (Catalog #10009878) for low-input library preparation, until the step before library PCR amplification. Libraries underwent a biotin pulldown using streptavidin to enrich chromatin interactions before an initial five-cycle amplification was performed and a qPCR quantification determining the number of additional cycles required for each library to reach a final concentration around 20 nM in 20 µL. Amplified libraries were then examined by Agilent 4200 TapeStation for confirmation of size distribution before submission for sequencing. Libraries were first sequenced using the Illumina MiniSeq PE50 for quality control (analysis described in the next section), and then deeply sequenced using the Illumina NovaSeq PE50 aiming at ~4  x 108  read pairs for each library that passed the MiniSeq QC.

value: Lung

value: DAPI-, lineage-, Tomato+

value: KRAS LSL-G12D;R26 LSL-Tomato; STAG2 flox/flox

value: Adeno-SPC-Cre


title: SL, scRNAseq

description: 10x Genomics

value: MSL Tumor


title: LA450, KT;Stag2 flox, 16 weeks

description: Hi-C library


title: Bleomycin 18

city: Worcester

first: Jillian

last: Richmond

state: MA

address: 364 Plantation Street

department: Dermatology

institute: University of Massachusetts Medical School

data_processing:
Paired-end RNA sequencing data with remaining adapter or quality scores less than 25 were trimmed and sequences with less than 35 base pairs were removed
Trimmed data were aligned to the mouse reference genome GRCm38 using HiSAT2 (Kim et al. 2015)
Aligned data underwent quality check via FastQC (Andrews et al. 2010)
Duplicates were marked using Picard ("Picard Toolkit." 2019) and features (i.e. genes, transcripts and exons) were counted using featureCounts (Liao et al. 2014).
Genome_build: mm10
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample
Supplementary_files_format_and_content: tab-delimited text files include logCPM values for each sample

source_name: skin

extract_protocol:
RNA was extracted from FFPE curls using Qiagen FFPE RNEasy kit per the manufacturer's protocol.
Libraries were prepared uising the Takara SMART-Seq Stranded Total RNA kit in accordance with manufacturer protocol specifications.

treatment_protocol: C57BL/6 mice received injections in their back skin (s.c./i.d.) of bleomycin or PBS vehicle daily for the number of days indicated in this time course. Skin was excised, fixed in 4% PFA overnight, and paraffin embedded.

value: bleo

tag: day


title: Undifferentiated Spermatogonia H3K36me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 2

value: Dnmt3a f/+ ; Dnmt3b f/+


title: BIL937-1B

value: Skeletal muscle


title: LA1104, KT, 16 weeks

description: Hi-C library

value: KRAS LSL-G12D;R26 LSL-Tomato


title: Undifferentiated Spermatogonia H3K36me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 1


title: CCL, scRNAseq

description: 10x Genomics

value: Skin

value: Tumor derived GFP+ cells

value: cSCC Cell line


title: Undifferentiated Spermatogonia H3K36me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 1

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg


title: Bleomycin 17

value: older


title: BOH319-3B

value: male


title: 5M_20um_protein

city: Philadelphia

first: Liran

last: Mao

state: PA

address: 422 Curie Boulevard

department: Perelman School of Medicine

institute: University of Pennsylvania

data_processing:
For ATAC and CUT&Tag data, linkers 1 and 2 are used for targeted filtering of read 2, aligning utilizing BWA followed by sorting and indexing via Samtools to facilitate efficient data handling and retrieval. This reformatting process assigned genome sequences to the first read and incorporated barcodes A and B into the second read. We aligned these fastq files against mouse (GRCm38) reference genomes. The conversion produced tsv-like fragments files, enriched with spatial and genomic information through the integration of barcode pairs, facilitating comprehensive downstream analysis.   For RNA sequencing data, we refined read 2 to extract barcode A, barcode B, and the Unique Molecular Identifier (UMI). Using the Spatial Transcriptomics (ST) pipeline version 1.7.2, this processed data was mapped against the appropriate mouse (GRCm38) genome references. This step produced a gene matrix that captures both gene expression and spatial positioning data, encoded through the combination of barcodes A and B, enabling detailed spatial transcriptomic analysis.
Assembly: GRCm38
Supplementary files format and content: TSV files for downstream analysis. For CUT&Tag and ATAC-seq, the tsv file contains tissue location info (barcode A x barcode B) and fragments info on the genome.For RNA-seq, Tab-delimited text files include counts for each pixels for each Sample.
Library strategy: spatial-CITE-seq

molecule: protein

source_name: Embryo

extract_protocol:
The workflow of simultaneous profiling of chromatin accessibility or a panel of cell surface proteins with two histone modifications and gene expression on the same tissue cryosections is shown schematically in Fig. 1a and Extended Data Fig. 1a-b. For joint analysis of chromatin accessibility, two histone modifications, and transcriptome (Extended Data Fig. 1a, Methods), a frozen tissue section was first fixed with formaldehyde and in situ Tn5 transposition was performed to insert a unique barcoded ligation linker to the accessible DNA loci. The same tissue section was then incubated with two primary antibodies targeting different histone modifications simultaneously, such as the combination of H3K27me3 with H3K27ac or H3K4me3. Afterwards, species specific nanobody-Tn5 fusion proteins loaded with unique barcoded ligation linkers were added to enable the demultiplexing of different histone modification loci. Next, the tissue section was incubated with a biotinylated DNA adaptor containing a universal ligation linker and poly-T sequence that binds to the messenger RNA (mRNA) to perform in situ reverse transcription (RT). Next, barcodes A (A1-A50) and barcodes B (B1-B50) were sequentially flowed over the tissue using microchannels and were conjugated to the universal ligation liker via templated ligation, which formed a two-dimensional grid of spatially barcoded tissue pixels (n =2,500), allowing all of modalities from the same pixel share the same spatial barcodes. Finally, barcoded complementary DNA (cDNA) and genomic DNA (gDNA) fragments were released by reverse crosslinking. cDNAs were separated from gDNA by streptavidin beads. Sequencing libraries for cDNAs and gDNA were separately constructed. For co-profiling of protein, gene expression, and histone modifications (Extended Data Fig. 1b, Methods), the fixed frozen tissue section was first incubated with two primary antibodies targeting different histone modifications, followed by nanobody-Tn5 in situ transposition. The same tissue section was then labeled with a panel of barcoded poly-A-tailed oligo-conjugated antibodies, which recognize surface antigens. Next, in situ reverse transcript was performed using the Biotinylated poly-T RT primer to capture complimentary DNA of both poly-A-tailed oligo-conjugated antibodies and mRNA. The subsequent steps for spatial barcoding and library construction were similar to above discussed methods. The protein library and RNA library can be separated by SPRI bead purification.
Frozen tissue slides were thawed for 1 min at 37 °C. Tissue was fixed with formaldehyde (0.2%, with 0.05 U μl–1 RNase Inhibitor) for 5 min and quenched with 1.25 M glycine for a further 5 min. After fixation, tissue was washed twice with 1 ml of 0.5× DPBS-RI and cleaned with ddH2O. The sequential order for spatial multiple profiling is as follows: 1. ATAC-seq; 2. Nanobody-based CUT&Tag; 3. In situ reverse transcription; 4. Ligation of barcode A; 5. Ligation of barcode B; 6. Reverse crosslink; 7. gDNA and cDNA separation; 8. Library construction; 9. Library QC and sequencing.

value: Embryo


title: PBS 5

value: PBS


title: 5M_20um_RNA

data_processing:
For ATAC and CUT&Tag data, linkers 1 and 2 are used for targeted filtering of read 2, aligning utilizing BWA followed by sorting and indexing via Samtools to facilitate efficient data handling and retrieval. This reformatting process assigned genome sequences to the first read and incorporated barcodes A and B into the second read. We aligned these fastq files against mouse (GRCm38) reference genomes. The conversion produced tsv-like fragments files, enriched with spatial and genomic information through the integration of barcode pairs, facilitating comprehensive downstream analysis.   For RNA sequencing data, we refined read 2 to extract barcode A, barcode B, and the Unique Molecular Identifier (UMI). Using the Spatial Transcriptomics (ST) pipeline version 1.7.2, this processed data was mapped against the appropriate mouse (GRCm38) genome references. This step produced a gene matrix that captures both gene expression and spatial positioning data, encoded through the combination of barcodes A and B, enabling detailed spatial transcriptomic analysis.
Assembly: GRCm38
Supplementary files format and content: TSV files for downstream analysis. For CUT&Tag and ATAC-seq, the tsv file contains tissue location info (barcode A x barcode B) and fragments info on the genome.For RNA-seq, Tab-delimited text files include counts for each pixels for each Sample.
Library strategy: spatial-RNA-seq


title: SL721, KT, 16 weeks

description: Hi-C library

value: Lung


title: BER395-1B

value: young

value: rest


title: BER395-2B


title: PBS 6


title: BER395-3B


title: Differentiated Spermatogonia H3K4me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 2

value: Differentiated Spermatogonia


title: Differentiated Spermatogonia H3K4me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 1


title: BEL997-3B


title: Differentiated Spermatogonia H3K4me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 2


title: 5M_20um_ATAC_H3K27ac_H3K27me3

data_processing:
For ATAC and CUT&Tag data, linkers 1 and 2 are used for targeted filtering of read 2, aligning utilizing BWA followed by sorting and indexing via Samtools to facilitate efficient data handling and retrieval. This reformatting process assigned genome sequences to the first read and incorporated barcodes A and B into the second read. We aligned these fastq files against mouse (GRCm38) reference genomes. The conversion produced tsv-like fragments files, enriched with spatial and genomic information through the integration of barcode pairs, facilitating comprehensive downstream analysis.   For RNA sequencing data, we refined read 2 to extract barcode A, barcode B, and the Unique Molecular Identifier (UMI). Using the Spatial Transcriptomics (ST) pipeline version 1.7.2, this processed data was mapped against the appropriate mouse (GRCm38) genome references. This step produced a gene matrix that captures both gene expression and spatial positioning data, encoded through the combination of barcodes A and B, enabling detailed spatial transcriptomic analysis.
Assembly: GRCm38
Supplementary files format and content: TSV files for downstream analysis. For CUT&Tag and ATAC-seq, the tsv file contains tissue location info (barcode A x barcode B) and fragments info on the genome.For RNA-seq, Tab-delimited text files include counts for each pixels for each Sample.
Library strategy: nanobody-based Spatial CUT&Tag


title: Differentiated Spermatogonia H3K4me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 1


title: Bleomycin 16


title: BEL997-2B


title: Undifferentiated Spermatogonia H3K4me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 2

value: Undifferentiated Spermatogonia


title: SL698, KTC sgPaxip1

city: Stanford

first: Emily

middle: Lorin

last: Ashkin

address: 279 Campus Drive

institute: Stanford University School of Medicine

data_processing:
Adapter-trimmed reads were aligned to the mm10 genome using Bowtie2 (2.1.0). Aligned reads were filtered for quality using SAMtools (v.1.9), duplicate fragments were removed using Picard (v.2.21.9-SNAPSHOT) and peaks were called using MACS2 (v.2.1.0.20150731) with aq-value cut-off of 0.01 and with a no-shift model. Peaks from replicates were merged, read counts were obtained using bedtools (v.2.17.0) and normalized using DESeq2 (v.1.26.0).
Assembly: mm10
Supplementary files format and content: tab-delimited text files includes raw counts for each sample
Supplementary files format and content: tab-delimited text files includes RLE-normalized counts for each sample

extract_protocol:
After cells were sorted and counted, 50-100,000 cells were resuspended in 1 mL of cold ATAC-seq resuspension buffer (RSB; 10mM Tris-HCl pH 7.4, 10 mM NaCl, and 3 mM MgCl2 in water) (68). Cells were centrifuged 500 x g for 5 min at 4 °C. After centrifugation, 900 μL of supernatant was aspirated. The remaining 100 μL of supernatant was carefully aspirated with a P20 pipette tip to avoid disturbing the cell pellet. Cell pellets were then resuspended in 50 μL of ATAC-seq RSB containing 0.1% NP40, 0.1% Tween-20, and 0.01% digitonin and mixed by pipetting up and down five times. This reaction was incubated on ice for 3 minutes, and after lysis, 1 mL of ATAC-seq RSB containing 0.1% Tween-20 was added, and tubes were inverted six times to mix. Nuclei were then centrifuged for 10 min at 500 x g at 4 °C. Supernatant was again removed using the two-pipette steps, as described before, and nuclei were resuspended in 50 μL of transposition mix (25 μL 2X TD buffer, 2.5 μL transposase (100 nM final), 16.5 μL PBS, 0.5 μL 10% Tween-20, and 5 μL water) by pipetting up and down six times. Transposition reactions were incubated at 37  °C for 30 minutes at 1000 rpm. Reactions were purified using Qiagen MinElute Reaction Cleanup Kit.
ATAC-seq library preparation was performed as described in Buenrostro, et al.

value: DAPI-, lineage-, Tomato+

value: Kras LSL-G12D; R26 LSL-Tomato, H11 LSL-Cas9


title: Undifferentiated Spermatogonia H3K4me3 Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 1


title: SL697, KTC sgPaxip1


title: BEL997-1B

value: Skeletal muscle


title: SL448, KTC sgInert

value: Lenti-Cre-sgInert


title: BEC999-2B


title: Undifferentiated Spermatogonia H3K4me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 1

value: Dnmt3a f/+ ; Dnmt3b f/+


title: PBS 4


title: Bleomycin 12

value: bleo


title: Bleomycin 13


title: SL113,KTC sgInert


title: Undifferentiated Spermatogonia H3K4me3 Dnmt3a f/+ ; Dnmt3b f/+  rep 2


title: Sperm Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg rep 2

data_processing:
Sequence reads were trimmed for adaptor sequence/low-quality sequence using TrimGalore with parameters (--rrbs --illumina --stringency 3)
Trimmed sequence reads were aligned to the mouse genome build mm10 using the qAlign function from the QuasR package (bisulfite="dir")
Methylated and unmethylated read counts for each CpG were extracted from the BAM files using the function qMeth from the QuasR package.
Assembly: mm10
Supplementary files format and content: BigWig files (*.bw) processed files were generated for methylation levels of each CpG (files ending with *MethylationLevel.bw) and total number of reads covering each CpG (*TotalNReads.bw).

source_name: Sperm

extract_protocol:
For sperm cell sorting, mouse cauda epididymides were dissected into a Petri dish and fat patches were removed with forceps and scissors. Each epididymis was punctured with a needle and carefully squeezed into 100 ul of PBS with the help of two forceps. Sperm was transferred into an Eppendorf tube and was allowed to liquefy at room temperature for 5 minutes. To break sperm tails, cells were briefly sonicated with a Brandson Tip digital sonicator with 10% amplitude and 3 cycles of 0.5 sec ON / 2 sec OFF. Cells were stained with 2 ul/ml Hoechst 33342 (H3570, ThermoFisher) for 1h at 25°C with constant shaking and protected from light. Sperm cells were filtered through a 40 μm Nylon filter into 5 ml polypropylene tubes. Sperm cells were sorted for 1C DNA content. Pellets of 1 million sperm cells were incubated in 200μl of DNA extraction buffer (20mM Tris-HCl pH 7.5, 300mM NaCl, 2% SDS, 10mM EDTA, 200 μg/ml Proteinase K) for 16 hours at 55oC. An equal volume of phenol:chloroform:isoamyl alcohol (25:24:1) solution was added to the samples. After a vigorous vortexing for 30 seconds and centrifugation at 16.000g for 10 minutes, the upper aqueous phase was transferred to new tubes. 0.05 μg/μl glycogen, 1/10 volume of 3M sodium acetate and 2.5 volumes of 100% ethanol were added, and the samples were incubated fot 2 hours at -80oC. DNA precipitated by centrifugation at 16000g for 30m minutes at 4oC. DNA pellet was washed once with 70% ethanol and resuspended in 20 μl. DNA was further purified using 2 volumes of Ampure XP beads (Beckman Coulter). For spermatogonia, testicular cell suspension was prepared by incubation of seminiferous tubules in 200 U/ml Collagenase type I (Worthington Biochemical LS004196), 5 μg/ml DNAse I (Roche 10104159001), and 0.05%Trypsin (Gibco 25200056) in GBSS (Sigma G9779) as described in (60). Cells were stained with 1/200 anti-CD117/c-kit PE (eBioscience 12-1171-83), anti-CD324/E-Cadherin eFluor 660 (eBioscience 50-3249-82) and anti-CD49f/Integrin alpha 6 PE-Cyanine7 (eBioscience 25-0459-82) for 1 hour at 32oC with constant shaking and protected from light. After washing cells were incubated with 20 μg/ml with Hoechst 33342 (Thermo Fischer Scientific H3570) for 1 hour at 32oC with constant shaking and protected from light and then with 30nM DRAQ7 (Biostatus DR71000) for 5 minutes on bench. The stained cells were strained through a 40 μm nylon filter into 5 ml polypropylene tubes and were sorted on a BD FACSAria III cell sorter fitted with a 70 μm nozzle (Becton Dickinson) using a 375nm laser to excite Hoechst 33342, a 561nm laser to excite PE and PE-Cy7 and a 633nm laser to excite DRAQ7 and eFluor600. Cells were first gated for FSC and SSC to exclude debris. Then the live cells were selected based on the absence of DRAQ7 signal detected using 755LP, 780/60BP. Then we gated for cells positive for Hoechst 33342 emission which was detected using a 670LP (Hoechst-Red) and 450/20 BP (Hoechst-Blue). We gated for the Hoechst-Redlow and Hoechst-Bluemid population which is enriched for 2N spermatogonia. Fluorescence for PE was detected using a 582/15BP filter, for PE-Cy7 was detected using 735LP,780/60BP filter and for eFluor660 using a 660/20BP filter. Undifferentiated spermatogonia were sorted as CD324high, CD49fhigh, CD117low and differentiated spermatogonia were sorted as CD324low, CD49flow, CD117high.DNA was extracted from 10000 undifferentiated or differentiated spermatogonia cell samples by QIAamp DNA Micro Kit (Qiagen # 56304) according to the manufacturer’s instructions.
DNA was digested with 0.9μl MspI Fast Digest (Thermo Scientific FD0544) in 1X Tango buffer in a volume of 18 μl for 6 hours at 37oC. DNA ends were repaired by adding 1 μl Klenow Fragment, exo– (Thermo Scientific EP0421) and 1µl of nucleotide end-repair mix (1mM dATP; 0.1mM dGTP; 0.1mM dCTP prepared in 2X Tango buffer) and incubated for 45min at 37ᵒC, followed by enzyme heat inactivation at 75ᵒC for 15min. Then 6.25nM of 5mC sequencing adapter (NEB E7535S) was ligated with 1 μl T4 DNA Ligase, HC (Thermo Scientific EL0013) and 0.5 μl ATP solution (Thermo Scientific R0441) for 16 hours at room temperature. The looped adaptor ligated DNA fragments were subjected to USER (NEB M5508) digestion at 37 ᵒC for 20 minutes and used directly for bisulfite treatment using the Imprint DNA Modification Kit (Sigma MOD50) according to the manufacturer’s 2-step modification procedure. The modified DNA was amplified with 13 cycles of PCR using Kapa U+ polymerase (Roche 07959052001) and indexed primers (NEB) and purified with 1.8X Ampure XP beads.

value: Sperm

value: Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg


title: BEC999-1B

value: older


title: BEC565-2B

value: female


title: BEC565-1B


title: Bleomycin 11


title: SL1046,KTC sgInert


title: Bleomycin 14


title: Sperm Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg rep 1


title: PBS 3


title: LA450, KT;Stag2 flox, 16 weeks

data_processing:
Paired-end RNA-seq reads were aligned to the mm10 mouse genome using STAR under standard input parameters. Two KT;Stag2flox samples were excluded from subsequent analysis based on two parameters: 1) >20% the average number of nucleotide counts in Exon 8 of STAG2, and 2) >40% mRNA expression of Stag2 from RNA-seq results (TPM). The differentially expressed genes between different tumor genotypes were called by DESeq2 using the HTSeq-derived counts as input (64). Unsupervised hierarchical clustering and heatmap visualization of DEGs were performed using the ‘pheatmap’ package in R (v.1.0.12, https://github.com/raivokolde/pheatmap).  The DESeq2-calculated fold changes were used to generate ranked gene lists for input into GSEA (v.4.0.3). GSEA was performed using normalized RNA-seq counts against gene signatures from the MSigDB database (65). Default parameters were used with the following exception: max size= 20,000. Plots were made using the Rtoolbox package in R (https://github.com/PeeperLab/Rtoolbox). clusterProfiler (v.4.3.1.900) was used to perform GO analyses, GO term enrichment, and functional enrichment analysis (66). GO terms were then categorized using EMBL’s European Bioinformatics Institute (EMBL-EBI) goslim_mouse annotations and statistics analysis by “all” relationships (“occurs_in”, “has_input”, etc.).
Assembly: mm10
Supplementary files format and content: tab-limited text file includes raw counts for each Sample

extract_protocol:
Tumors were dissociated using collagenase IV, dispase, and trypsin at 37 degrees C for 30 minutes as previously described (40). Cells were stained with DAPI and antibodies against CD45 (30-F11), CD31 (390), F4/80 (BM8), and Ter119 (all from BioLegend) to exclude hematopoietic and endothelial cells. FACSAria sorters (BD Biosciences) were used for cell sorting.
Total RNA was prepared from FACS-sorted neoplastic cells ranging from 5 x 104 to 4 x 105  cells. RNA quality was assessed using the RNA 6000 Pico Assay Kit on the Agilent 2100 Bioanalyzer (Agilent). RNA used for RNA-seq had a mean RIN of 6.6. RNA-seq libraries were generated by Novogene, Inc. Total RNA (from 2.5-10 ng/sample) was used for cDNA synthesis using the Ovation RNA-seq System (NuGEN Technologies, Inc.). Two micrograms of NuGEN-amplified double-stranded cDNA was sheared using a Covaris sonicator to an average length of 490 bp and subjected to library preparation using the Takara V4 Ultra low input mRNA non-directional library preparation according to the manufacturer's protocol and sequenced on a NovaSeq PE150 (~6G raw data per sample).

value: KRAS LSL-G12D;R26 LSL-Tomato; STAG2 flox/flox

value: Adeno-SPC-Cre


title: LA491, KT;Stag2 flox, 16 weeks

value: Lung


title: Bleomcyin 9


title: Sperm Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre tg rep 1

value: Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre


title: Bleomycin 15


title: Bleomycin 10


title: EA699, KT;Stag2 flox, 8 weeks


title: ATK159-2B


title: Sperm Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 2

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg


title: ATK159-1B

value: rest


title: Bleomycin 8


title: Sperm Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 1


title: SL821, KT, 16 weeks

value: KRAS LSL-G12D;R26 LSL-Tomato


title: SL696 KTC sgPaxip1

value: DAPI-, lineage-, Tomato+


title: ARM341-2B

value: male


title: Sperm Dnmt3a f/+ ; Dnmt3b f/+  rep 1


title: LA1104, KT, 16 weeks


title: Sperm Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre tg rep 2


title: ARM341-1B

value: Skeletal muscle


title: SL721, KT, 16 weeks


title: Bleomycin 6

value: younger


title: ARK583-2B

value: young


title: PBS 1

value: PBS


title: Sperm Dnmt3a f/+ ; Dnmt3b f/+  rep 2

value: Sperm

value: Dnmt3a f/+ ; Dnmt3b f/+


title: EA696, KT;Stag2 flox, 8 weeks


title: Differentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg rep 2

value: Differentiated Spermatogonia

value: Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg


title: Differentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg rep 1


title: Bleomycin 5

value: bleo

value: older


title: SL725, KT, 8 weeks

value: Lung

value: Adeno-SPC-Cre


title: Differentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre tg rep 2

value: Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre


title: AFZ519-2B


title: SL723, KT, 8 weeks


title: Bleomycin 7


title: ARM897-2B

value: female


title: SL113, KT, 8 weeks

value: KRAS LSL-G12D;R26 LSL-Tomato


title: Bleomycin 4


title: Bleomycin 3


title: Differentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre tg rep 1


title: ADL867-2B


title: PBS 2


title: 52

city: East Lansing

first: Tim

last: Zacharewski

state: Michigan

address: 1129 Farm Ln

department: Biochemistry and Molecular Biology, Integrative Toxicological Sciences

institute: Michigan State University

description: Adult male mouse dosed every 4 days for 28 days

data_processing:
Read quality was assessed using FastQC v0.11.7 and adapter trimming was performed using Trimmomatic v0.39.
Alignment was performed with STAR v2.7.3a. Genome indices were created using Mus_musculus.GRCm39.dna.chromosome.fa and Mus_musculus.GRCm39.104.gtf.
Gene counts were determined using STAR v2.7.3a.
Assembly: mm10 (build 39)
Supplementary files format and content: STAR-aligned counts were transformed with the variance stabilizing transformation method in DESeq2.
Supplementary files format and content: Tab-delimited text files represent the number of aligned PE reads to each gene.

source_name: Kidney

extract_protocol:
Frozen mouse-right kidney (~100 mg) was homogenized in 1 mL of TriZOL with chrome-steel beads using a Mixer Mill 300 (Life Sciences, Carlsbad, CA) for 4 minutes. Following, total RNA was extracted per the manufacturer’s instructions with an additional 5:1 phenol:chloroform extraction step (Sigma Aldrich, St. Louis, MO). RNA purity (260/280 ratio) and concentration (ng/µL) were assessed with a NanoDrop 1000 spectrophotometer. RNA quality was determined using an Agilent 2100 Bioanalyzer. All RNA samples had RNA integrity numbers (RINs) ≥7.
Library preparation was performed using a Ultra2 NEB RNA kit.

treatment_protocol: Mice were orally gavaged with TCDD in sesame oil vehicle every 4 days for 28 days.

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: percent

value: Innovive cage

tag: bedding

value: ALHPA-dri (Shepherd Specialty Papers)

tag: water

value: Aquavive water (Innovive)

tag: name

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

tag: formula

tag: vendor

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

tag: dose

value: microgram per kilogram

value: Carbon dioxide euthanasia

tag: library name

value: NEBNext Ultra RNA kit


title: ARM897-1B

tag: tissue

value: Skeletal muscle

value: older

value: female

value: rest


title: AFZ519-1B

value: young

value: male


title: 53

tag: strain

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940


title: Differentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 1

value: Differentiated Spermatogonia

tag: genotype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg


title: Bleomycin 2

tag: treatment

value: bleo

tag: age


title: ADL867-1B

value: Skeletal muscle

value: older

tag: Sex

tag: training time_point


title: Differentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/+  rep 2

tag: cell type

value: Dnmt3a f/+ ; Dnmt3b f/+


title: 49

value: Harlan Teklad

value: Yes

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 90_S16

data_processing:
Data were processed using a previoulsy published method[https://pubmed.ncbi.nlm.nih.gov/24972667/]. In brief, reads were quality controlled using FASTQC. Reads were mapped against hg38 genome using STAR aligner. HT-Seq was utilized to analyzed the aligned reads and generate gene-wise counts using Ensemble version 78 transcriptome. Extracted read data was processed using edgeR software. Timmed means of M-values method was utilized to normalized reads that were provided in the file named "Grover-PIIBS-Normed-Counts.tsv".
Assembly: hg38
Supplementary files format and content: Grover-PIIBS-Normed-Counts.tsv; tab-delimited

source_name: Colon Muscle

extract_protocol:
mRNA was extracted using Trizol and stored in RNAlater.
mRNA library was constructed using Illumina TruSeq Stranded kit following manufacturer's recommendations.

value: Colon Muscle

value: Muscle

value: PIIBS


title: Differentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/+  rep 1

value: Differentiated Spermatogonia


title: ARK583-1B

value: Skeletal muscle

value: young

value: female

value: rest


title: Bleomycin 1

value: bleo

value: older


title: Responder6

city: Shanghai

first: Peiyi

last: Xie

address: Fenglin Road No.180

country: China

institute: Zhongshan Hospital, Fudan University

data_processing:
Sequence reads were trimmed using Cutadapt and bases with average quality scores below 20 were filtered
Sequencing reads were mapped by using HISAT2 with BWT algorithm
Assembly: hg18
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

source_name: HCC

growth_protocol: Total RNA was extracted employing the RNeasy kit (Qiagen, CA, USA). RNA quality was evaluated by Tapestation (Agilent, USA). The library was prepared using the Illumina Truseq RNA Kit (Illumina, CA, USA). RNA-seq libraries were prepared employing Illumina TruSeq Stranded mRNA sample preparation kits according to the protocol of manufacturer.

extract_protocol:
RNA was extracted by RNAiso Plus (9109, Takara) following the manufacturer’s instructions, 1.5 ug of total RNA was used to construct sequencing libraries
RNA libraries for RNA-seq were prepared by using Agilent 2100 Bioanalyzer

value: HCC

value: human heptocellular carcinoma

tag: patient


title: Differentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 2

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg


title: Responder3


title: 47

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage


title: Undifferentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg rep 1

value: Undifferentiated Spermatogonia

value: Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg


title: Undifferentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/- ; Stra8-iCre tg rep 2


title: Responder5

value: HCC

value: human heptocellular carcinoma


title: 50

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 79_S33

value: Colon Muscle

value: Muscle

value: HV


title: 43

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Accustandard

value: Room temperature

value: sesame oil


title: 46

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: milliliter

value: oral gavage


title: 82_S32

value: Colon Muscle

value: Muscle

value: PIIBS


title: Responder4

value: HCC

value: human heptocellular carcinoma


title: Undifferentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre tg rep 2

value: Undifferentiated Spermatogonia

value: Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre


title: 44

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 78_S61

value: Colon Muscle

value: HV


title: Nonresponder4


title: Responder1


title: 76_S30

value: Muscle


title: 72_S62


title: 41

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage


title: Responder2

value: HCC

value: human heptocellular carcinoma


title: 40

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: microgram per kilogram


title: Nonresponder3


title: Undifferentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 2

value: Undifferentiated Spermatogonia

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg


title: 8_S67

value: Colon Muscle

value: Muscle

value: HV


title: Nonresponder2


title: Undifferentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/+  rep 2

value: Dnmt3a f/+ ; Dnmt3b f/+


title: 38

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: Undifferentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg rep 1

value: Undifferentiated Spermatogonia


title: Undifferentiated Spermatogonia Dnmt3a f/+ ; Dnmt3b f/+  rep 1


title: 39

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: microgram per kilogram


title: 66_S25

value: Colon Muscle

value: Muscle

value: HV


title: 68_S60

value: PIIBS


title: 37

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter


title: Undifferentiated Spermatogonia Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre tg rep 1

value: Undifferentiated Spermatogonia

value: Dnmt3a f/- ; Dnmt3b f/+ ; Stra8-iCre


title: 6_S27


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 20

data_processing:
Sequence reads were trimmed for adaptor sequence/low-quality sequence using TrimGalore with parameters (--quality 20 --nextera --stringency 3)
Trimmed reads were then aligned to the mouse genome build mm10 using STAR (version 2.5.0a) with settings (--outFilterMismatchNmax 6)
For allelic discrimination a SNP table of JF1 versus C57BL/6J mouse genome was acquired from (Takada, T., Ebata, T., Noguchi, H., Keane, T.M., Adams, D.J., Narita, T., Shin, I.T., Fujisawa, H., Toyoda, A., Abe, K. et al. (2013) The ancestor of extant Japanese fancy mice contributed to the mosaic genomes of classical inbred strains. Genome Res, 23, 1329-1338.). SNPs were injected into mm10 genome and alignment was done separately for C57BL/6J (R - reference paternal) and JF1 (A - alternative maternal) genomes. Allelic assignment for each read was done by comparing number of mismatches between alignments to each genome.
Aligned reads were deduplicated using SAMtools (version 1.10) with standard settings
Uniquely mapped reads were counted on genes (TxDb.Mmusculus.UCSC.mm10.ensGene) and tiles using the function qCount (mapqMin = 255L) from QuasR package (version 1.36.0).
Assembly: mm10
Supplementary files format and content: BigWig (.bw) processed files were generated from BAM files using the function qExportWig from QuasR package (version 1.36.0) with standard settings separately for maternal (files ending with XV_A.bw), paternal (XV_R.bw) and total alignments.

source_name: 4 cell stage embryo

extract_protocol:
To obtain early embryos, 7-10-week-old JF1 female mice were injected intraperitoneally with HyperOva (Cosmo Bio, KYD-010-EX-X5) 63 hours pre collection and human chorionic gonadotropin (hCG, Chorulon MSD) 15 hours pre collection. Oviducts were dissected and oocytes from 2 animals were collected in 30 μl drops of HTF medium (EmbryoMax® Human Tubal Fluid Millipore, MR-070-D). Sperm was collected from 2 cauda epididymides in 50 μl drops of HTF medium. 2-5 μl of sperm sample was transferred to the oocyte drops and incubated for 3 hours at 37 °C. Zygotes with visible pronuclei washed 3 times in HTF medium and transferred to KSOM medium (Millipore, MR-106-D) and cultured in low oxygen chamber filled with the mixture gas (5% O2/ 5% CO2). Early 4 cell stage embryos were collected 50-55 hours post fertilization.
Each 4-cell stage embryo was collected at 50-55 hours after IVF, was washed in PBS + 0,02% PVA,  transferred into a well of a 96-well plate containing SS2 Lysis buffer ( 0,09% Triton-X 100, 0,5U/μl SUPERAseIN (Life Technologies AM2696), 2.5mM oligodT primer (Microsynth), 2.5mM dNTP mix(Promega), ERCC RNA Spike-In Mix (1:3.2x107 Thermo Fischer scientific 4456740)), snap frozen on dry ice and then stored in -80°C until further use. The sample was incubated at 72°C for 3 minutes and immediately put back on ice. Then SS2 RT mix (10U/µl SuperScript II reverse transcriptase (Thermo Fischer 18064014), 0.25U/μl SUPERAseIN,  1X SuperScript II first-strand buffer , 5mM DT, 1M Betaine (Sigma B0300-1VL), 6mM MgCl2, 1mM TSO (Exiqon)) was added to the sample and incubated for 90 minutes at 42°C, then 10 cycles of 2 minutes at 50°C and 2 minutes at 42°C and finally for 15 minutes at 70°C. Sample was preamplified for 16 cycles using 1X KAPA HiFi HotStart ReadyMix (KAPA Biosystems KK2602) and 0.1µM ISPCR primers (Microsynth). DNA was purified using 1X volumes of Ampure XP DNA purification beads. 1ng of pre-amplified DNA was added to Tn5 tagmentation mix (1x TAPS-DMF buffer, homemade Tn5 (1:1,200)) in total volume 20μl and incubated at 55°C for 7minutes. Then the reaction was stopped by adding 5μl of 0.2% SDS and kept at 25°C for 7 min. Adapter-ligated fragment amplification was done using Nextera XT index kit (Illumina) in a total volume 50μl (1x Phusion HF Buffer, 2 µl of Phusion High Fidelity DNA Polymerase (Thermo Fischer, F530L), dNTP mix (0.3mM each) (Promega)) with 10 cycles of PCR. Library was purified by 1X volumes of Ampure XP DNA purification beads and resuspended in 25 μl nuclease free H2O.

value: 4 cell stage embryo

tag: maternal strain

tag: maternal genotype

value: wildtype

tag: paternal strain

tag: paternal genotype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

tag: developmental stage

value: early 4 cell stage embryos (50-55hpf)


title: 56_S40

value: Colon Muscle

value: Muscle


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 17


title: Nonresponder1

value: HCC

value: human heptocellular carcinoma


title: 64_S38

value: HV


title: 35

value: Charles Rivers Laboratories

value: Male

value: day

value: percent

value: Innovive cage

tag: bedding

value: ALHPA-dri (Shepherd Specialty Papers)

tag: water

value: Aquavive water (Innovive)

tag: name

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

tag: formula

tag: vendor

value: oral gavage

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 75_S51

tag: tissue

value: Colon Muscle

value: Muscle

tag: genotype


title: 60_S66

value: HV


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 19

value: 4 cell stage embryo

value: wildtype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

value: early 4 cell stage embryos (50-55hpf)


title: 55_S14


title: 34

tag: strain

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: percent

value: Innovive cage

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

tag: dose

tag: library name


title: LNCaP-sgC_ENZA_YY1

city: New York

first: Jorge

last: Moscat

state: NY

address: 1188 York Avenue

department: Pathology and Laboratory Medicine

institute: WEILL CORNELL MEDICINE

data_processing:
Quality control of raw sequencing reads was performed using FastQC
Low-quality reads were removed using Trimmomatic with a sliding window of 4bp and quality threshold of 15
Reads passing QC were aligned to GRCh38 using Bowtie2
BAM file creation, sorting, removal of PCR duplicates and indexing were performed using samtools
CUT&RUN peaks were called using MACS2 with IgG as the control for each set of data.
Assembly: Homo Sapiens - Human (GRCh38)
Supplementary files format and content: CUT&RUN signal (bigWig) called by MACS2 IgG as control
Library strategy: CUT&RUN

source_name: Prostate carcinoma

growth_protocol: LNCaP cells were cultured in RPMI1640 medium supplemented with 10% fetal bovine serum, 1x glutamine, 1x pen/strep

extract_protocol:
The cells were incubated overnight with conA beads and specific antibodies including K-metstat internal control at 4C. Chromatin were digested using pAG-MNase for 15 minutes at RT. The fragmented DNA was column purified.
CUT&RUN libraries were prepared using NEBNext Ultra II DNA Library Prep Kit for illumina. 5ng of CUT&RUN DNA were used to prepare the libraries using specific index primers and universal PCR primer and amplified for 14 cycles in a thermocycler. Post amplification, the libraries were size selected for 300-400bp using SPRIselect beads from NEB. The concentration and size of the libraries were measured using Qubit and Agilent High Sensitivity DNA kit respectively.

treatment_protocol: Enzalutamide 10 uM for 72 h.

value: Prostate carcinoma

tag: cell line

value: LNCaP

tag: cell type

value: Epithelial

value: sgControl

tag: treatment

value: ENZA 72 hours

tag: cut&run antibody

value: YY1 (Cell Signalling, ##46395)


title: LNCaP-sgPRKCI_ENZA_YY1

value: sgPRKCI


title: 29

value: Charles Rivers Laboratories

tag: Sex

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: oral gavage

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 26

value: Male

value: day

value: degree celsius

value: percent

value: Innovive cage

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 16

value: 4 cell stage embryo

tag: maternal strain

tag: maternal genotype

value: wildtype

tag: paternal strain

tag: paternal genotype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

value: early 4 cell stage embryos (50-55hpf)


title: 28

value: Charles Rivers Laboratories

tag: bedding

value: ALHPA-dri (Shepherd Specialty Papers)

tag: water

value: Aquavive water (Innovive)

tag: name

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: oral gavage


title: 27

value: Male

value: day

value: degree celsius

value: percent

tag: formula

tag: vendor

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 50_S36

tag: tissue

value: Colon Muscle

value: Muscle

tag: genotype

value: PIIBS


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 15

value: 4 cell stage embryo

value: wildtype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

tag: developmental stage

value: early 4 cell stage embryos (50-55hpf)


title: LNCaP-sgC_ENZA_IgG

value: Prostate carcinoma

tag: cell line

value: LNCaP

value: Epithelial

value: sgControl

value: ENZA 72 hours

value: IgG (Epicypher, #13-0042)


title: Caco-2, undifferentiated, rep2

city: San Diego

first: April

middle: Elizabeth

last: Williams

state: California

address: 10010 N Torrey Pines Rd

department: IGC

data_processing:
Raw paired-end FASTQ reads were trimmed using Trim Galore! v 0.6.6 with Cutadapt v 3.4 to remove library adapter sequences and low-quality bases (using command “-g 20 -length 20”). The trimmed reads were then aligned by STAR v2.79a to the hg38 reference genome.
Gene/smORF counts were quantified from the aligned reads using HOMER analyzeRepeats, with the smORFs appended to the annotated reference GTF file.
Assembly: GRCh38.p13
Supplementary files format and content: A counts file generated using HOMER, where the first 8 columns contain annotation information such as feature (gene/smORF) ID and coordinates, and the remaining columns contain the counts of each feature

growth_protocol: Cells were seeded into larger format plates according to the following densities: CCD18-Co cells 3 x 106 cells/23mL in 15cm plate, CaCo2 cells 4.2 x 106 cells/10mL in 10cm plate, HCEC-1CT cells 5 x 106 cells/8.7mL in 10cm plate, PBMC 39 x 106 cells/39mL in T75 flask.

extract_protocol:
Cells were washed twice after treatment with ice cold PBS supplemented with 100ug/mL cycloheximide (CHX; Fisher Scientific, AAJ66004X) and snap frozen in liquid nitrogen and stored at -80°C. After thawing briefly on ice, the cells were lysed with 300uL of ice-cold lysis buffer (20mM Tris-HCl, pH 7.4, 150mM NaCl, 5mM MgCl2, 1% Triton X-100, 1mM DTT, 25U Turbo DNase (Thermofisher Cat# AM2238), and 100ug/mL CHX – added fresh) for 10 minutes with periodic vortexing and pipetting through an 18G needle to ensure the cells are dispersed. Cell lysates were clarified by centrifugation for 15,000g for 10 minutes at 4°C, flash frozen in liquid nitrogen, and stored in -80°C for up to 1 week.
Ribosome footprinting was carried out by digesting 50 ug of RNA in 300 uL lysate with 0.375 U/ug RNase I (Lucigen, N6901K) for 45 min at room temperature. Digestion reactions were quenched with 200 U Superase-In RNase I inhibitor (Thermo Fisher, AM2694) on ice. Following digestion, monosomes were purified from small RNA fragments using Micro-Spin S-400 HR columns (GE Life Sciences), and ribosome protected RNA fragments (RPFs) were extracted by acid phenol chloroform and isopropanol precipitation. Sequencing libraries were prepared as in McGlincy and Ingolia with some modifications. First, the RiboZero Mammalian Kit (Illumina) was used to deplete rRNA after RPF extraction and just prior to RPF size selection by gel extraction. Second, the Zymo clean & concentrator step after adaptor ligation is omitted and the reaction was carried over straight into reverse transcription. For the reverse transcription step to form cDNA, Episcript RT (Lucigen, ERT12910K) was used. Following reverse transcription, excess primer was degraded using Exonuclease I (Lucigen, X40520K) and the RNA templates were degraded using Hybridase (Lucigen, H39500). For the cDNA circularization step, CircLigase I (Lucigen, CL4111K) was used. PCR amplification was then carried out using Phusion Hot Start II High-Fidelity Master Mix (Thermo Fisher, F565L) for 9–12 cycles.

treatment_protocol: Cells were then either allowed or not allowed to differentiate, and then treated or not treated with following stimulants: lipid micelles, micelles without lipid.

value: undifferentiated


title: LNCaP-sgPRKCI_ENZA_IgG

tag: cell type

value: sgPRKCI


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 18


title: 47_S65


title: Caco-2, undifferentiated, rep1

tag: treatment


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 13


title: 25

tag: strain

value: Charles Rivers Laboratories

value: Male

value: day

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

tag: dose

value: microgram per kilogram

value: Carbon dioxide euthanasia

tag: library name


title: Caco-2, undifferentiated, rep3

value: undifferentiated


title: 24

value: Charles Rivers Laboratories

tag: Sex

value: Male

value: day

value: NEBNext Ultra RNA kit


title: Caco-2, differentiated, micelles without lipid, rep3

value: differentiated, micelles without lipid


title: 38_S10

value: Colon Muscle

value: Muscle

value: PIIBS


title: Caco-2, differentiated, micelles without lipid, rep2


title: 23

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 12

value: 4 cell stage embryo

tag: maternal strain

tag: maternal genotype

value: wildtype

tag: paternal strain

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

value: early 4 cell stage embryos (50-55hpf)


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 14

tag: paternal genotype


title: 22

value: Charles Rivers Laboratories

value: Male

value: day

tag: bedding

tag: water

tag: name

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: Caco-2, differentiated, lipid micelles, rep3

value: differentiated, lipid micelles


title: 32_S35

tag: tissue

value: Colon Muscle

value: Muscle

value: PIIBS


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 11

value: 4 cell stage embryo

value: wildtype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg


title: 28_S5

tag: genotype


title: Caco-2, differentiated, lipid micelles, rep2


title: 21

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

tag: formula

tag: vendor


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 10

tag: developmental stage

value: early 4 cell stage embryos (50-55hpf)


title: 3_S63

value: Colon Muscle

tag: cell type


title: Caco-2, differentiated, lipid micelles, rep1

tag: cell line

tag: treatment

value: differentiated, lipid micelles


title: 20

tag: strain

value: day

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

tag: dose

value: microgram per kilogram

value: Carbon dioxide euthanasia

tag: library name

value: NEBNext Ultra RNA kit


title: 19

value: Charles Rivers Laboratories

tag: Sex

value: Male

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 8

value: 4 cell stage embryo

tag: maternal strain

tag: maternal genotype

value: wildtype

tag: paternal strain

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

value: early 4 cell stage embryos (50-55hpf)


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 7

tag: paternal genotype


title: 21_S72

value: Colon Muscle

value: Muscle

value: PIIBS


title: Caco-2, differentiated, control, rep3

value: differentiated, control


title: 39_S69

value: HV


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 6


title: Caco-2, differentiated, control, rep2


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 5


title: Caco-2, differentiated, micelles without lipid, rep1

value: differentiated, micelles without lipid


title: 20_S28


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 9


title: 16

value: day

tag: bedding

tag: water

value: Aquavive water (Innovive)

tag: name

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Accustandard

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 17

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

tag: formula


title: 2_S39

tag: tissue

value: Colon Muscle

value: Muscle

tag: genotype

value: PIIBS


title: Caco-2, differentiated, control, rep1

value: differentiated, control


title: 27_S50

tag: cell type


title: 15

value: day

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

tag: vendor

value: Accustandard

value: Room temperature


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 4

value: 4 cell stage embryo

value: wildtype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

tag: developmental stage

value: early 4 cell stage embryos (50-55hpf)


title: 16_S47

value: HV


title: PBMC pool2

data_processing:
Raw single-end FASTQ reads were trimmed using FASTX Toolkit 0.0.14 to remove library adapters and low-quality bases. The trimmed reads were then first aligned (using STAR 2.7.11a) to a list of tRNA/rRNA sequences to filter out tRNA/rRNA contaminants, and the remaining unmapped reads were aligned to the hg38 reference genome.
smORF counts were quantified from the aligned reads using HOMER analyzeRepeats, using a smORFs GTF file obtained from the pipeline described in Martinez et al (Martinez TF, Chu Q, Donaldson C, Tan D, Shokhirev MN, Saghatelian A: Accurate annotation of human protein-coding small open reading frames. Nat Chem Biol 2020, 16:458-468.).
Assembly: GRCh38.p13
Supplementary files format and content: A counts file generated using HOMER, where the first 8 columns contain annotation information such as feature (gene/smORF) ID and coordinates, and the remaining columns contain the counts of each feature

source_name: PBMC

extract_protocol:
Cells were washed twice after treatment with ice cold PBS supplemented with 100ug/mL cycloheximide (CHX; Fisher Scientific, AAJ66004X) and snap frozen in liquid nitrogen and stored at -80°C. After thawing briefly on ice, the cells were lysed with 300uL of ice-cold lysis buffer (20mM Tris-HCl, pH 7.4, 150mM NaCl, 5mM MgCl2, 1% Triton X-100, 1mM DTT, 25U Turbo DNase (Thermofisher Cat# AM2238), and 100ug/mL CHX – added fresh) for 10 minutes with periodic vortexing and pipetting through an 18G needle to ensure the cells are dispersed. Cell lysates were clarified by centrifugation for 15,000g for 10 minutes at 4°C, flash frozen in liquid nitrogen, and stored in -80°C for up to 1 week.
For each Abm HCEC-1CT, Caco2, CCD118co, and PBMC pool of unstimulated and stimulated cells, ribosome footprinting was carried out by digesting 50 ug of RNA in 300 uL lysate with 0.375 U/ug RNase I (Lucigen, N6901K) for 45 min at room temperature. Digestion reactions were quenched with 200 U Superase-In RNase I inhibitor (Thermo Fisher, AM2694) on ice. Following digestion, monosomes were purified from small RNA fragments using Micro-Spin S-400 HR columns (GE Life Sciences), and ribosome protected RNA fragments (RPFs) were extracted by acid phenol chloroform and isopropanol precipitation. Sequencing libraries were prepared as in McGlincy and Ingolia with some modifications. First, the RiboZero Mammalian Kit (Illumina) was used to deplete rRNA after RPF extraction and just prior to RPF size selection by gel extraction. Second, the Zymo clean & concentrator step after adaptor ligation is omitted and the reaction was carried over straight into reverse transcription. For the reverse transcription step to form cDNA, Episcript RT (Lucigen, ERT12910K) was used. Following reverse transcription, excess primer was degraded using Exonuclease I (Lucigen, X40520K) and the RNA templates were degraded using Hybridase (Lucigen, H39500). For the cDNA circularization step, CircLigase I (Lucigen, CL4111K) was used. PCR amplification was then carried out using Phusion Hot Start II High-Fidelity Master Mix (Thermo Fisher, F565L) for 9–12 cycles.

treatment_protocol: Cells were then treated with following stimulants (all 1uL/mL of media) for 24hr before being harvested: LPS (100ng/mL) (Sigma-Aldrich, Cat# L4391-1MG), IL-22 (10ng/mL) (RnD Bio, Cat# 782-IL), TNFa (10ng/mL) (RnD Bio, Cat#210-TA-005), IFN-g (10ng/mL) (RnD Bio, Cat#285-IF-100), IL-1b (10ng/mL) (RnD Bio, Cat#201-LB010), OSM (10ng/mL) (RnD Bio, Cat# 295-OM-010) and TGFb (10ng/mL) (RnD Bio, Cat#240-B-010).

tag: cell line

value: PBMC

tag: treatment


title: 14

tag: strain

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: sesame oil

value: milliliter

value: oral gavage

tag: dose

value: microgram per kilogram

value: Carbon dioxide euthanasia

tag: library name

value: NEBNext Ultra RNA kit


title: PBMC pool1


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 3

value: 4 cell stage embryo

tag: maternal strain

tag: maternal genotype

value: wildtype


title: 18

tag: Sex

value: day

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Accustandard

value: Room temperature


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 2

tag: paternal strain

tag: paternal genotype

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

value: early 4 cell stage embryos (50-55hpf)


title: Caco-2 pool2


title: 142_S8

value: Colon Muscle

value: Muscle

value: HV


title: Caco-2 pool1


title: 141_S54


title: 13

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

value: Innovive cage

tag: bedding

value: ALHPA-dri (Shepherd Specialty Papers)

tag: water

tag: name

value: sesame oil

value: milliliter

value: oral gavage

value: microgram per kilogram


title: 11

value: day

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

tag: formula

value: Accustandard

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 22

tag: tissue

value: 4 cell stage embryo

value: wildtype

value: Dnmt3a f/+ ; Dnmt3b f/+

value: early 4 cell stage embryos (50-55hpf)


title: 139_S43

value: Colon Muscle

tag: cell type

value: Muscle

tag: genotype

value: HV


title: Early 4 cell stage embryo Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg sperm derived rep 1

value: Dnmt3a f/- ; Dnmt3b f/- ; Stra8-iCre tg

tag: developmental stage


title: 18_S23


title: 9

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

tag: vendor

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

value: microgram per kilogram


title: 138_S15

value: PIIBS


title: 10

tag: strain

value: day

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Accustandard

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: HCEC-1CT pool2

tag: cell line

tag: treatment


title: CCD-18Co pool2


title: CCD-18Co pool1


title: 8

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

tag: dose

tag: library name


title: 136_S7

value: Colon Muscle

value: Muscle


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 19

value: 4 cell stage embryo

tag: maternal strain

tag: maternal genotype

value: wildtype

value: Dnmt3a f/+ ; Dnmt3b f/+

value: early 4 cell stage embryos (50-55hpf)


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 20

tag: paternal strain

tag: paternal genotype


title: 133_S34

value: PIIBS


title: 134_S58


title: HCEC-1CT pool1


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 18


title: Caco-2, undifferentiated, rep1

description: Undifferentiated Caco-2 cells

value: undifferentiated


title: 132_S2

value: HV


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 17


title: 7

tag: Sex

value: day

value: Innovive cage

tag: bedding

value: ALHPA-dri (Shepherd Specialty Papers)

tag: water

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Room temperature

value: sesame oil

value: milliliter

value: oral gavage

value: microgram per kilogram


title: Caco-2, differentiated, control, rep2

description: Differentiated Caco-2 cells without further treatment

value: differentiated, control


title: 2

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

tag: name

tag: formula

value: Accustandard

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 15

tag: tissue

value: 4 cell stage embryo

value: wildtype

value: Dnmt3a f/+ ; Dnmt3b f/+

value: early 4 cell stage embryos (50-55hpf)


title: 1

value: day

value: Innovive cage

value: ALHPA-dri (Shepherd Specialty Papers)

value: Aquavive water (Innovive)

value: Harlan Teklad 22/5 Rodent Diet 8940

value: Harlan Teklad

value: Yes

value: Room temperature


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 14

tag: developmental stage


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 21


title: SC34 - Hematoma/callus at day 5 post-musculosketal traumatic injury

city: Créteil

first: Céline

last: Colnot

address: 8 rue du Général Sarrail

country: France

institute: INSERM U955

description: 10x Genomics

data_processing:
Fastq files were processed with the CellRanger pipeline with the default’s parameters
Reads were align against the mm10 reference genome
Filtered features barcodes matrices were used to perform data analysis
Assembly: mm10
Supplementary files format and content: Cell Ranger output files

source_name: Periosteum

extract_protocol:
Injured tibias from 3 or 4 mice were collected and the surrounded tissues were removed. The activated periosteum was scraped and collected with the hematoma/fracture callus. Collected tissues were minced and placed 5 min in ice-cold Nuclei Buffer (NUC101, Merck) before mechanical nuclei extraction using a glass douncer. Nuclei suspension was filtered, centrifuged and resuspended in RNAse-free PBS (AM9624, ThermoFischer Scientific) with 2% Bovine Serum Albumin (A2153, Merck) and 0.2 U/µL RNAse inhibitor (3335399001, Roche). Sytox™ AADvanced™ (S10349, ThermoFischer Scientific) was added (1/200) to label nuclei and Sytox-AAD+ nuclei were sorted using Sony SH800.
The snRNA-seq libraries were generated using Chromium Single Cell Next GEM 3′ Library & Gel Bead Kit v.3.1 (10x Genomics) according to the manufacturer’s protocol. Briefly, 10 000 to 20 000 nuclei were loaded in the 10x Chromium Controller to generate single-nuclei gel- beads in emulsion. After reverse transcription, gel-beads in emulsion were disrupted. Barcoded complementary DNA was isolated and amplified by PCR. Following fragmentation, end repair and A-tailing, sample indexes were added during index PCR. The purified libraries were sequenced on a Novaseq (Illumina) with 28 cycles of read 1, 8 cycles of i7 index and 91 cycles of read 2. Sequencing data were processed using the Cell Ranger Count pipeline and reads were mapped on the HG38 2020-A reference human genome, with intronic and exonic sequences.
Library was prepared following manufacturer's recommendations.

value: Periosteum

tag: genotype


title: Caco-2, differentiated, micelles without lipid, rep2

description: Differentiated Caco-2 cells treated with micelles composed of taurocholate

value: differentiated, micelles without lipid


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 16


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 13


title: 127_S42

value: Colon Muscle

tag: cell type

value: Muscle

value: HV


title: 130_S52


title: Caco-2, differentiated, micelles without lipid, rep1

tag: cell line


title: Caco-2, undifferentiated, rep2

description: Undifferentiated Caco-2 cells

tag: treatment

value: undifferentiated


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 12

value: 4 cell stage embryo

value: wildtype


title: 3

tag: strain

value: Charles Rivers Laboratories

value: Male

value: degree celsius

value: percent

tag: vendor

value: Accustandard

value: sesame oil

value: milliliter

value: oral gavage

tag: dose

value: microgram per kilogram

value: Carbon dioxide euthanasia

value: NEBNext Ultra RNA kit


title: 120_S3

value: Colon Muscle

value: PIIBS


title: splenic H2-Aa deficient (cit) DC, scRNA-seq

city: Dallas

first: Hexin

last: Shi

state: Texas

address: 5323 Harry Hines Blvd

institute: UT southwestern medical center

description: DC were collected from 3 female, 4-month-old H2-Aa deficient (citation) mice

data_processing:
BCL files were demultiplexed using cellranger(v5.0.1)
Alignment, barcode counting, UMI counting were performed using cellranger(v5.0.1)
Normalization, PCA, clustering, and differential expression was performed using Seurat
mm10
count matrix in a sparse matrix format
Assembly: barcode id of the sample
Supplementary files format and content: gene ids of the sample

source_name: spleen

extract_protocol:
Splenic DC were enriched from splenocyte suspensions using EasySep Mouse CD11c Positive Selection Kit II (Stemcell) according to the manufacturer’s instructions. Then, about 1.5 × 10^4 live CD45.2+ CD11c+ cells were FACS sorted from WT or H2-Aa cit/cit DC enriched samples.
scRNA-seq libraries were generated using the Chromium Next GEM Single Cell 3’ kit (10X Genomics) according to the manufacturer’s protocol. Purified libraries were sequenced on one S4 lane of a NovaSeq sequencing instrument

value: spleen

value: primary dendritic cells

value: H2-Aa cit/cit

value: naive


title: splenic WT DC, scRNA-seq

description: DC were collected from 3 female, 4-month-old WT mice


title: 131_S18

value: Muscle


title: Caco-2, differentiated, lipid micelles, rep1

description: Differentiated Caco-2 cells treated with lipid micelles containing BSA, Oleic acid, Cholesterol

value: differentiated, lipid micelles


title: 12_S17

value: HV


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 11

value: 4 cell stage embryo

tag: maternal strain

tag: maternal genotype

value: wildtype

value: Dnmt3a f/+ ; Dnmt3b f/+

value: early 4 cell stage embryos (50-55hpf)


title: MRC8

description: TRAMP-Prkci-flox-PBcre prostate tissue piece#3

data_processing:
Raw data was downloaded in Ubuntu using BaseSpaceCLI.
We used cellranger mkfastqto demultiplex raw base call (BCL) files generated by Illumina sequencers into FASTQ files
We used cellranger countto take FASTQ files fromcellranger mkfastqand perform alignment to mm10, filtering, barcode counting, and UMI counting
We used cellranger aggrto aggregate all sample outputs ofcellranger count, normalizing  to the same sequencing depth and then recomputing the feature-barcode matrices and analysis on the combined data
We used Seurat in R to analyze the aggregated data
Assembly: mm10
Supplementary files format and content: Seurat object generated in R (TRAMP_TRAMPLKO_rds)

source_name: Prostate tumor

growth_protocol: In vivo endogenous mouse model

extract_protocol:
To prepare prostate tumor cells suspension, prostates were digested with digestion buffer (5 mg/ml Collagenase II  in Advanced DMEM-F12 with Y-27632) for 20 min at 37°C with agitation, filtered through 70- and 40-μm-mesh cell strainers, washed with Advanced DMEM-F12 plus Glutamax 1x, Hepes 10 mM, 10 uM Y-27632, and Pen/Strep 1x (ADF 4+). Cells were further digested eith TryplE during 20 min at 37°C, washed in ADF 4+, and resuspended with Advanced DMEM-F12 containing 10 mM HEPES, 1X Glutamax, 10 mM Y-27632, Pen/Strep 1x and 5% FBS. Dead cells were removed by EasySep Dead Cell Removal (Annexin V) kit (STEMCELL technologies).
scRNA-seq libraries were generated using the Chromium Single Cell 30 Reagent Kit v2 (10X Genomics). Cells were loaded onto the 10X Chromium Single Cell Platform (10X Genomics),  (Single Cell 3′ library and Gel Bead Kit v.2) as described in the manufacturer’s protocol (10x User Guide, Revision B). On average, approximately 20,000 cells were loaded. Generation of gel beads in emulsion (GEMs), barcoding, GEM-RT clean-up, complementary DNA amplification and library construction were all performed as per the manufacturer’s protocol. Individual sample quality was checked using a Bioanalyzer Tapestation (Agilent). Qubit was used for library quantification before pooling. The final library pool was sequenced on an Illumina NovaSeq6000 instrument using a S1 flowcell.

treatment_protocol: No treatment

value: Prostate tumor

value: Prostate tumor cells

value: TRAMP+;Prkci-flox/flox-Pbcre+


title: MRC7

description: TRAMP-Prkci-flox-PBcre prostate tissue piece#2


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 10

tag: paternal strain

tag: paternal genotype


title: SC28 - Hematoma/callus at day 1 post-fracture

description: 10x Genomics

value: Periosteum


title: LNCaP-sgPRKCI_ENZA_H3K27me3

data_processing:
Quality control of raw sequencing reads was performed using FastQC
Low-quality reads were removed using Trimmomatic with a sliding window of 4bp and quality threshold of 15
Reads passing QC were aligned to GRCh38 using Bowtie2
BAM file creation, sorting, removal of PCR duplicates and indexing were performed using samtools
CUT&RUN peaks were called using MACS2 with IgG as the control for each set of data.
Assembly: Homo Sapiens - Human (GRCh38)
Supplementary files format and content: CUT&RUN signal (bigWig) called by MACS2 IgG as control

value: Prostate carcinoma

value: LNCaP

value: Epithelial

value: sgPRKCI

value: Enzalutamide 72 hours

tag: cut&run antibody


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 9


title: 111_S45

value: Colon Muscle

value: PIIBS


title: 11_S4

value: Muscle


title: MRC6

description: TRAMP-Prkci-flox-PBcre prostate tissue piece#1


title: LNCaP-sgC_ENZA_H3K27me3

value: sgControl


title: Caco-2, differentiated, lipid micelles, rep2

value: differentiated, lipid micelles


title: LNCaP-sgPRKCI_ENZA_EZH2

value: EZH2 (Epicypher, #13-2026)


title: LNCaP-sgC_ENZA_EZH2


title: 1_S55


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 8

value: 4 cell stage embryo

value: wildtype

value: Dnmt3a f/+ ; Dnmt3b f/+


title: MRC5

description: TRAMP prostate tissue

value: Prostate tumor

value: Prostate tumor cells

value: TRAMP+


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 7

value: early 4 cell stage embryos (50-55hpf)


title: 10_S48

value: HV


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 6


title: TCD-24

description: Prostate organoids Pten-Prkci-DKO

data_processing:
Sequenced reads were trimmed to remove low-quality bases at ends and were then mapped to the reference genome (GRCm39) using STAR with function  --quantMode
Counts were filtered and normalized using DESeq2.
Assembly: Mus musculus - Mouse (GRCm39)
Supplementary files format and content: Normalized counts matrix with gene expression value for all samples.

source_name: Prostate

growth_protocol: Prostate organoids were grown in Advanced DMEM-F12 with Glutamax 1x, Hepes 10 uM, Pen/Strep 1x, B27 1x, N2 1x, A-8301 200 nM, hEGF 50 ng/ml, Y27632 10 uM, Rspondin 10%, Noggin 1x, DHT 1 nM.

extract_protocol:
RNA was harvested using Trizol reagent and purified using Quick-RNA MiniPrep kit (Zymo Research).
Library preparations and sequencing were conducted at Genewiz (AZENTA)

treatment_protocol: Cre-excision of floxed genes was induced by adenovirus cre (Ad-CMV-iCre, Vector Biosystems).

value: Prostate

value: organoids

value: Epithelial

value: Pten-Prkci-DKO


title: 074_TRNA_S68

value: Colon Muscle

value: PIIBS


title: TCD-25

description: Prostate organoids Pten-Prkci-DKO


title: TCD-23

description: Prostate organoids Pten-Prkci-DKO


title: LNCaP-sgC_ENZA_H3K4me3

value: Prostate carcinoma

value: LNCaP

value: sgControl

value: Enzalutamide 72 hours


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 5

value: 4 cell stage embryo

value: wildtype


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 4

value: Dnmt3a f/+ ; Dnmt3b f/+


title: 057_TRNA_S24

value: Muscle


title: 069_TRNA_S37


title: LNCaP-sgPRKCI_ENZA_IgG

value: sgPRKCI

value: IgG (Epicypher, #13-0042)


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 3

value: early 4 cell stage embryos (50-55hpf)


title: TCD-21

description: Prostate organoids Pten-KO

value: Prostate

value: Pten-KO


title: TCD-22

description: Prostate organoids Pten-KO

value: organoids

value: Epithelial


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 2


title: LNCaP-sgC_ENZA_IgG


title: LNCaP-sgPRKCI_ENZA_H3K4me3


title: 048_TRNA_S64

value: Colon Muscle


title: TCD-20

description: Prostate organoids Pten-KO


title: LNCaP-sgPRKCI_DMSO_H3K27me3

value: DMSO 72 hours


title: 043_TRNA_S31

value: PIIBS


title: Early 4 cell stage embryo Dnmt3a f/+ ; Dnmt3b f/+ sperm derived  rep 1


title: LNCaP-sgC_DMSO_H3K27me3

value: Prostate carcinoma

value: LNCaP