title: Rat alveolar macrophages, 24hr, CNC3, replicate 3

city: Tsukuba

first: Katsuhide

last: Fujita

address: 16-1 Onogawa

department: Research Institute of Science for Safety and Sustainability

institute: National Institute of Advanced Industrial Science and Technology

description:
Gene expression after 24hr in CNC3-exposed rat alveolar macrophages
CF060_03

hyb_protocol: Each labeled cRNA probe was used separately for hybridization to a 4×44 K Whole Human Genome Microarray (G2519F-014850; Agilent Technologies) at 65 °C for 17 h.

scan_protocol: Hybridized microarray slides were washed according to the manufacturer’s instructions and scanned with a DNA Microarray Scanner (Agilent) at 5 μm resolution.

data_processing:
The scanned images were analyzed numerically using Agilent Feature Extraction Software, version 12.0.3.1.
log2 average normalized signal intensity

label_protocol: Cyanine-3-labeled cRNA was prepared from RNA using a Low Input Quick Amp Labeling kit (Agilent) according to the manufacturer’s instructions, followed by RNeasy column purification (Qiagen). Dye incorporation and cRNA yield were checked with the NanoDrop 2000 spectrophotometer.

growth_protocol: Rat alveolar macrophage cells (NR8383) obtained the American Type Culture Collection were cultured at 37 °C in 75-cm2 cell culture flasks in F-12K with 10 % (v/v) heat-inactivated fetal bovine serum (FBS), 100 units/mL penicillin, and 100 μg/mL streptomycin in a humidified atmosphere of 95% air and 5% CO2.

extract_protocol: Total RNA was extracted using QIAzol Lysis Reagent (Qiagen, Japan) following the manufacturer’s instructions. RNA was quantified using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific Inc., US), and sample qualities were monitored with an Agilent 2100 Bioanalyzer (Agilent Technologies, US).

treatment_protocol: The F-12K + FBS medium was removed by centrifugation, and cell cultures were then treated with working solutions of three types of CNCs (CNC1, CNC2, or CNC3) . These cultures were subsequently seeded into individual wells of a 6-well plate and incubated for 24 hours at 37 ºC in a humidified incubator with 5% CO2. At this point, the cell concentrations were approximately 3.0 x 10^5 cells/mL.

tag: cell

value: Rat alveolar macrophages  (NR8383)

tag: time


title: 22Rv1 HIF1A set 2 5UTR DMS Vehicle biol rep 1

status: Public on Feb 16 2025

type: SRA

city: San Francisco

first: Duygu

last: Kuzuoglu Ozturk

state: California

address: 1450 3rd Street, Helen Diller Cancer Building

country: USA

institute: UCSF

description: Library name: Sample21

library_source: other

data_processing:
Reads were trimmed using trim galore (default parameters), and then aligned to the reference 5’UTR sequence using bowtie, using the parameters -local --no-unal --no-discordant --no-mixed -X 1000 -L 12.
Bam files were then input into the perl-based rnaframework toolkit (v2.8.9).Mutations were counted using rf-count -m and normalized using rf-norm (parameters -sm 4 -nm 2 -rb AC -nf float,float) with experiment-wide normalization factors obtained from rf-normfactor (parameters -sm 4 -nm 2 -rb AC).
Normalized replicate xml files were combined using rf-combine.
Structural prediction was done using rf-fold with parameters -g -ct --folding-method 2 -dp. Predicted secondary structures were compared using rf-compare.
Assembly: hg38
Supplementary files format and content: xlsx files with normalized per-condition and delta bp reactivities for endogenous AR, HIF1a, and control transcripts (DNAJA1, APEX1, RPS5) corresponding to supplemental tables from paper
Library strategy: DMS MaPSeq

molecule: other

organism: Homo sapiens

growth_protocol: 22Rv1 cells were cultured in RPMI 1640 medium supplemented with 10% Fetal Bovine serum and Penicillin/Streptomycin (GIBCO).

extract_protocol:
Cells were resuspended in 2.8ml RPMI media, mixed with 800ul warm 1M bicine (pH 8.57 at room temperature) buffer followed by 400ul 16% DMS diluted in EtOH to have a final 1.6% DMS concentration in the fume hood. Cells were then incubated in a 37 ˚C water bath for 5 min, and DMS was quenched by adding 2 ml cold beta-mercaptoethanol (BME), followed by centrifugation at 300 g for 5min at 4C. Cells were washed once with 7ml 30% BME in PBS and lysed in 1ml Trizol (Invitrogen). RNA was isolated following the manufacturer’s protocol. Any trace of DNA was removed by Turbo DNase I followed by phenol:chloroform extraction, and RNA was used as a template for cDNA synthesis using Induro reverse transcriptase (NEB). cDNA was cleaned up using SPRIselect beads and is used as a template to amplify the full-length 5’UTRs of corresponding transcripts using Prime Star Max DNA polymerase (Takara).
The libraries were prepared by the NEBNext® Ultra™ II DNA Library Prep (NEB) and sequenced on Novaseq X.

treatment_protocol: 22Rv1 cells were seeded on a 6 cm plate and incubated overnight. The following day, the cells were treated with zotatifin (10nM) or enzalutamide (1mM) for 2 hours and harvested immediately by trypsinization.

tag: cell line

tag: cell type

value: Prostate cancer cell line

tag: treatment


title: M8-inferior

city: Bethesda

first: Matthew

middle: J

last: Brooks

state: MD

address: 6 Center Dr Bldg 6, Rm 303

department: NEI

institute: NIH

description: 10X Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, barcoded processing, gene counting, and aggregation were made using the Cell Ranger software v7.0.1.
Data matrixes were imported into Seurat v4.3.0 (Satija, et al. 2015) in the R v4.2.1 statistical environment. Initial matrix filtering of >5 cells per gene, >200 features per cell, and cells with mitochondria reads <20% was performed.
Sample count matrixes of the same age were merged and ambient RNA was removed using decontX function in the celda v1.14.2 package. DecontX adjusted counts matrixes were split into individual samples prior to doublet detection and removal using DoubletFinder v2.0.3 with a target doublet rate of 5%.
Sample decontX adjusted count matrixes were merged into a single Seurat object and normalized and variance stabilized using SCTransform v2 (Choudhary and Satija, 2022). PCA (50 dimensions), FindNeighbors (30 dimensions), RunUMAP (30 dimensions), and FindClusters were performed using Seurat.
Cell types were assigned using scType (https://github.com/IanevskiAleksandr/sc-type) with sctype 'eye' database "Astrocytes", "Immune cells", "Retinal pigment epithelial cells", and custom cell type gene lists (https://github.com/NEI-NNRL/2023_Mouse_Retina_Aging/blob/main/src/Retina_Cell_type_MB_v2.xlsx). High resolution clustering (108 clusters) was performed to assess and remove clusters exhibiting expression of canonical cell type markers from more than one cell type.
Assembly: GRCm38.p6/ENSEMBL v98
Supplementary files format and content: HDF5 file of filtered CellRanger output

molecule: total RNA

organism: Mus musculus

source_name: Retina

extract_protocol:
Mouse retinas were dissected and dissociated as previously described (Fadl, et al. Mol Vis 2020). A 1mm punch of superior or inferior retina dissociate at each age was used as input for 10X Genomics Chromium controller using v2 (Chromium Next GEM Single Cell 3ʹ GEM, Library and Gel Bead Kit v2) chemistry. Approximately 10,000 live cells per sample were loaded into the Chromium chip following the Cell Suspension Volume Calculator Table to capture transcripts from approximately 6,000 cells.
Libraries were performed according to the manufacturer’s instructions (Single Cell 3’ v2 protocol, 10x Genomics)

tag: tissue

value: Retina

tag: age

value: 8 months

tag: region

value: inferior

tag: genotype

tag: Sex

value: male


title: 22Rv1 HIF1A set 2 5UTR DMS Enza biol rep 1

description: Library name: Sample23

value: Enzalutamide


title: 5xFAD, Microglia, circRNA, rep4

city: Shanghai

first: Li

last: Yang

address: 131 Dong-An Road

department: Institutes of Biological Sciences

country: China

institute: Fudan University

library_source: transcriptomic

data_processing:
Pair-end RNA-seq reads were firstly trimmed by Trimmomatics (v0.32, parameters: PE -threads 5 -phred33 ILLUMINACLIP:TruSeq3PE-2.fa:2:30:10 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:36) to remove adaptors and low quality bases.
The reads were mapped to the mm10 rDNA sequences for rRNAs (5S, 5.8S,18S, 28S) and pre-rRNAs (45S) to remove reads from ribosomal RNA by Bowtie2 (version: 2.3.5, parameters: -p 10 --score-min L,-16,0 --rfg 0,7 --rdg 0,7 --mp 7,7) and the rest paired reads were then mapped to the mouse mm10 genome reference with HISAT2 (version 2.1.0, parameters: --no-softclip --score-min L,-16,0 --mp 7,7 --rfg 0,7 --rdg 0,7 --max-seeds 20 --dta -k 1)
Fragment counts of genes were obtained using featureCounts (v1.5.1, parameters: -s 2 -p --fraction -O -T 16 -t exon -g gene_id) based on gencode vM25 gene annotation and normalized to FPKM (fragments per kilobase of gene per million fragments mapped)
Assembly: mm10
Supplementary files format and content: tab-delimited text file include normalized gene expression values (FPKM) for samples

source_name: Microglia

extract_protocol:
Total RNA from tissues was extracted using TRIzol reagent (Invitrogen), 1 ug of total RNA was used for the construction of  sequencing library.
Total RNAs were processed with RiboMinus kit (Human/ Mouse Module, Invitrogen) to deplete most ribosomal RNAs (ribo- RNA). The ribo- RNA libraries for sequencing were prepared with illumina TruSeq® Stranded Total RNA Library Prep Gold (#20020598) kit. All libraries were sequenced with NovaSeq 6000 system at Sequanta Technologies. FastQC was applied for raw read qualities assessment.

value: Microglia

value: AAV PHP.eB-circPOLR2A(9,13)


title: WT, H3K56ac, ChIP, 2

city: beijing

first: Yulin

state: Beijing

address: Institute of Microbiology

institute: University of Chinese Academy of Sciences

library_source: genomic

data_processing:
Basecalls were performed using Peaks were called using MACS2 (v.2.2.8, macs2 callpeak) with parameter -t -c -g 4.3e8 -q 0.05 --broad --max-gap 500 -n. Peaks annotation was performed by the package makeTxDbFromGFF (makeTxDbFromGFF function - RDocumentation) of R.
Assembly: Neurospora crassa OR74A
Supplementary files format and content: bigWig,

molecule: genomic DNA

organism: Neurospora crassa

extract_protocol:
The purity of the sample was determined by NanoPhotometer ® (IMPLEN, CA, USA)；The concentration and integrity of RNA samples were detected by Agilent 2100 RNA nano 6000 assay kit (Agilent Technologies, CA, USA).
Sequencing libraries were generated using TIANSeq DirectFast Library Kit (illumina) (NG101)  following the manufacturer’s recommendations and index codes were added to attribute sequences to each sample.

tag: strain

value: WT

value: 2% glucose with 1mg/mL HU in DD14


title: M3-superior

description: 10X Genomics

value: 3 months

value: superior


title: 22Rv1 DNAJA1 5UTR DMS Zota biol rep 2

description: Library name: Sample20

value: Zotatifin


title: Rat alveolar macrophages, 24hr, CNC3, replicate 2

type: RNA

country: Japan

organism: Rattus norvegicus


title: Rat alveolar macrophages, 24hr, CNC3, replicate 4


title: WT, H3, ChIP, 1


title: 22Rv1 HIF1A set 2 5UTR DMS Vehicle biol rep 2

description: Library name: Sample22


title: Rat alveolar macrophages, 24hr, CNC3, replicate 1


title: Cat_#1_ConA

city: Ube

first: Yoichi

last: Mizukami

institute: Yamaguchi　University

data_processing:
FASTQ files produced by Illumina NovaSeq 6000 were imported to CLC Genomics Workbench software (ver.22.0.1).
The 3' end of the sequenced read was trimmed by one nucleotide. Reads with more than 2 ambiguous nucleotides and reads with low quality scores were removed using CLC Genomics Workbench software (ver.22.0.1).
The reads were trimmed and mapped to the feline reference genome (felCat9) by using the CLC Genomics Workbench ver. 22.0.1.
Transcripts Per Million (TPM) were calculated using CLC Genomics Workbench software (ver.22.0.1).
Assembly: felCat9
Supplementary files format and content: tab-delimited text files include TPM values for each sample.

organism: Felis catus

source_name: peripheral blood lymphocytes

extract_protocol:
Total RNA was isolated from the melanoma cells using the RNeasy Plus Universal Kit  (QIAGEN). Polyadenylated mRNA was purified using oligo dT beads included in the NEBNext Poly (A) mRNA Magnet Isolation Module (New England Biolabs, NEB). Purified mRNA was used for the construction of sequencing libraries using NEBNext Ultra II RNA library Prep kit (NEB) and NEBNext Multiplex Oligos for Illumina (NEB).
Extracted mRNA was incubated with NEBNext Random Primers in NEBNext First Strand Synthesis Reaction Buffer at 94°C for 15 min, and reverse transcription was performed with NEBNext Strand Synthesis Enzyme Mix. After reverse transcription, index sequences included in NEBNext Multiplex Oligos for Illumina were inserted to cDNA fragments with PCR amplification. Following purification with AMPureXP beads (Beckman Coulter), the quality and the concentration of the libraries was evaluated using a Tapestation 4200 D1000 (Agilent). The confirmed libraries were mixed to equal molecular amounts against the clustering and sequencing on an Illumina NovaSeq 6000 with 50 bp paired-end cycle sequencing kit (Illumina).

value: peripheral blood lymphocytes

value: ConA


title: Rat alveolar macrophages, 24hr, CNC2, replicate 4

description:
Gene expression after 24hr in CNC2-exposed rat alveolar macrophages
CF060_02


title: WT, H3K56ac, ChIP, 1


title: WT, H3, ChIP, 2


title: Rat alveolar macrophages, 24hr, CNC2, replicate 3


title: 5xFAD, Microglia, circRNA, rep3

value: AAV PHP.eB-circPOLR2A(9,12)


title: chk1chk2, H3T11ph, ChIP, 2

value: chk1chk2 knockout


title: chk1chk2, H3T11ph, ChIP, 1


title: 5xFAD, Microglia, circRNA, rep1

value: AAV PHP.eB-circPOLR2A(9,10)


title: 22Rv1 DNAJA1 5UTR DMS Vehicle biol rep 2

description: Library name: Sample18

value: Prostate cancer cell line


title: chk1chk2, H3K56ac, ChIP, 2


title: 5xFAD, Microglia, EGFP, rep2

value: AAV PHP.eB-EGFP


title: Rat alveolar macrophages, 24hr, CNC2, replicate 2

value: Rat alveolar macrophages  (NR8383)


title: 22Rv1 AR-Nterm 5UTR DMS Zota biol rep 2

description: Library name: Sample16


title: chk1chk2, H3K56ac, ChIP, 1


title: 5xFAD, Microglia, EGFP, rep1


title: Rat alveolar macrophages, 24hr, CNC1, replicate 4

description:
Gene expression after 24hr in CNC1-exposed rat alveolar macrophages
CF060_01


title: WT, Microglia, EGFP, rep3


title: chk1chk2, H3, ChIP, 1


title: WT, Microglia, EGFP, rep2


title: Rat alveolar macrophages, 24hr, CNC1, replicate 3


title: 22Rv1 AR-Nterm 5UTR DMS Enza biol rep 2

description: Library name: Sample14


title: 22Rv1 AR-Nterm 5UTR DMS Enza biol rep 1

description: Library name: Sample13


title: 22Rv1 DNAJA1 5UTR DMS Zota biol rep 1

description: Library name: Sample19


title: 22Rv1 Zota RPF biol rep 3

description: Library name: Sample12

data_processing:
Ribo-seq reads were first trimmed using cutadapt (v2.3) to remove the linker sequence AGATCGGAAGAGCAC.
The fastx_barcode_splitter script from the Fastx toolkit (v0.0.13) was then used to split the samples based on their barcodes.
The UMIs were removed from the beginning and end of each read (two and five nucleotides, respectively) and appended to the name of each read using umi_tools extract (v0.3.3).
Bowtie2 (v2.3.5) was then used to remove reads that map to ribosomal RNAs and tRNAs, and the remainder of the reads were then aligned to mRNAs (we used the isoform with the longest CDS for each gene as the representative).
After alignment, umi_tools dedup (v0.3.3) was used to deduplicate reads.
For the associated total RNA-Seq, reads were aligned to the longest CDS using Bowtie2.
Ribo-Seq and RNA-Seq reads (bam files) were then input into Ribolog. For the Ribo-Seq data, reads between 23-32 nt in length were selected.
The P-site offsites and codon counts were calculated for each sample using psite_info_rW  and psite_to_codon_count, respectively. Codon counts were corrected for stalling using CELP_bias.
TE ratios for each transcript were calculated as the ratio between (median of ratios-normalized) Ribo-Seq/total RNA-Seq transcript counts.TE changes between conditions were modeled using logistic regression, specifically utilizing the function logit_seq.
Assembly: hg38
Supplementary files format and content: csv file of the raw RNA-seq counts for each sample
Supplementary files format and content: csv file of the observed Ribo-Seq codon counts summarized to transcript level for each sample
Supplementary files format and content: csv file of translational efficiency (TE) results from Ribolog's logit_seq function
Supplementary files format and content: csv file of DESeq2 output from RNA-seq zotatifin vs vehicle comparison

growth_protocol: 22Rv1 cells were injected subcutaneously into the left or right hind flank of male immunodeficient mice (NSG) at 6-8 weeks.

extract_protocol:
6 hour post-treatment tumors were harvested and snap frozen in liquid nitrogen and crushed using a mortar and pestle, ensuring the samples were always frozen.
Crushed tissues were lysed in 20mM Tris pH 7.5, 150 mM NaCl, 5mM MgCl2, 1mM DTT, 100 μg/ml CHX, 1% Triton and 25U/ml Turbo DNase I on ice for 30min by vortexing several times. Lysates were spun down at 10,000g for 10 minutes, and the RNA concentration of clear lysates was measured by Qubit RNA HS Assay kit (Invitrogen). 30 μg and 1-2ug of RNA containing lysates were used to process Riboseq and RNAseq libraries, respectively. 30 μg RNA containing lysate was digested with RNAse T1 (120U) and RNase A (0.2ug) at room temperature for 30 min on a rotator. RNase digestion was inhibited by adding 10ul SuperaseIN RNase Inhibitor and immediately putting the samples on ice. Next, the samples were fractionated by sucrose gradient as described above into 14 samples, and only two fractions that show precise monosome distribution were used for RNA isolation. RNAs were isolated by mixing with TRizol-LS and precipitated by isopropanol and glycogen. Next, purified RNA samples were run on a 15% TBE-urea gel, stained by SyberGold, and the bands above 24nt and below 34 nt were extracted from the gel. Dephosphorylation and linker ligation of the footprint fragments followed by reverse transcription, circularization of cDNA, and the library construction were prepared as described previously (PMID: 28579404). For the RNAseq libraries, RNA was isolated by TRizol from the tumor lysates. Only 10ng of total RNA was used following the Takara Smarter Stranded Total RNA-Seq kit protocol

treatment_protocol: Mice were randomized to receive different treatments once the tumors reached 70-100 mm3 in volume. Animals were treated with either zotatifin (dissolved in 5% dextrose, at 0.3mg/kg) or vehicle (5% dextrose) by intraperitoneal injection.


title: 5xFAD, Microglia, circRNA, rep2

value: AAV PHP.eB-circPOLR2A(9,11)


title: WT, Microglia, EGFP, rep1


title: Rat alveolar macrophages, 24hr, CNC1, replicate 2


title: 22Rv1 Zota RPF biol rep 1

description: Library name: Sample10


title: M24_retina

data_processing:
The demultiplexing, barcoded processing, gene counting, and aggregation were made using the Cell Ranger software v7.0.1. Data matrixes were imported into Seurat v4.3.0 (Satija, et al. 2015) in the R v4.2.1 statistical environment. Initial matrix filtering of >5 cells per gene, >500 features per cell, and cells with mitochondria reads <20% was performed. Sample count matrixes of the same age were merged and ambient RNA was removed using decontX function in the celda v1.14.2 package. DecontX adjusted counts matrixes were split into individual samples prior to doublet detection and removal using DoubletFinder v2.0.3 with a target doublet rate of 7.5%. Sample decontX adjusted count matrixes were merged into a single Seurat object and normalized and variance stabilized using SCTransform v2 (Choudhary and Satija, 2022). PCA (50 dimensions), FindNeighbors (30 dimensions), RunUMAP (30 dimensions), and FindClusters were performed using Seurat. Cell types were assigned using scType (https://github.com/IanevskiAleksandr/sc-type) with sctype 'eye' database "Astrocytes", "Immune cells", "Retinal pigment epithelial cells", and custom cell type gene lists (https://github.com/NEI-NNRL/2023_Mouse_Retina_Aging/blob/main/src/Retina_Cell_type_MB_v2.xlsx). High resolution clustering (112 clusters) was performed to assess and remove clusters exhibiting expression of canonical cell type markers from more than one cell type.
Assembly: GRCm38.p6/ENSEMBL v98
Supplementary files format and content: HDF5 file of filtered CellRanger output

extract_protocol:
Mouse retinas were dissected and dissociated as previously described (Fadl, et al. Mol Vis 2020). One sample from each timepoint was used to deplete CD73 positive cells (rod photoreceptors) using Anti-CD73 conjugated magnetic beads. Whole retina and CD73 negative dissociate was used as input for 10X Genomics Chromium controller using v3.1 (Chromium Next GEM Single Cell 3' GEM, Library and Gel Bead Kit v3.1) chemistry. Approximately 17,000 live cells per sample were loaded into the Chromium chip following the Cell Suspension Volume Calculator Table to capture transcripts from approximately 10,000 cells.
Libraries were performed according to the manufacturer’s instructions (Single Cell 3’ v3.1 protocol, 10x Genomics).

value: Retina

value: 24 months

value: whole retina

value: male


title: 22Rv1 DNAJA1 5UTR DMS Vehicle biol rep 1

description: Library name: Sample17


title: Rat alveolar macrophages, 24hr, CNC1, replicate 1


title: 22Rv1 AR-Nterm 5UTR DMS Vehicle biol rep 2

description: Library name: Sample12


title: 22Rv1 Zota RPF biol rep 2

description: Library name: Sample11


title: M24_CD73neg

value: CD73 depleted


title: 22Rv1 AR-Nterm 5UTR DMS Vehicle biol rep 1

description: Library name: Sample11


title: Rat alveolar macrophages, 24hr, Control for CNC1, CNC2 and CNC3, replicate 4

description:
Gene expression after 24hr in rat alveolar macrophages as vehicle cotrol
CF060_C1

value: Vehicle control for CNC1, CNC2, and CNC3


title: Rat alveolar macrophages, 24hr, Control for CNC1, CNC2 and CNC3, replicate 2


title: 22Rv1 AR-Cterm 5UTR DMS Zota biol rep 2

description: Library name: Sample10


title: 22Rv1 AR-Cterm 5UTR DMS Enza biol rep 1

description: Library name: Sample7

value: Enzalutamide


title: M12_CD73neg

value: 12 months


title: 22Rv1 Zota RNA biol rep 3

description: Library name: Sample9


title: Rat alveolar macrophages, 24hr, Control for CNC1, CNC2 and CNC3, replicate 3


title: Rat alveolar macrophages, 24hr, CNC2, replicate 1


title: 22Rv1 AR-Cterm 5UTR DMS Vehicle biol rep 2

description: Library name: Sample6


title: 22Rv1 AR-Nterm 5UTR DMS Zota biol rep 1

description: Library name: Sample15

value: Zotatifin


title: Ink4a-ast,JAG1,rep3

city: Seoul

first: Nayoung

last: Hong

address: Anam-ro 145

country: South Korea

institute: Korea University

data_processing:
QuantSeq 3' mRNA-Seq reads were aligned using Bowtie2.
Differentially expressed gene were determined based on counts from unique and multiple alignments using coverage in Bedtools.
The RC (Read Count) data were processed based on Quantile normalization method using EdgeR within R using Bioconductor.
Assembly: GRCm38/mm10
Supplementary files format and content: Excel spreadsheet format which includes contents; 1) Ensembl gene ID, 2) Normalized read count, 3) Read count.

source_name: Brain

growth_protocol: Cells were cultured in DMEM containing 10% fetal bovine serum, 1% penicillin and streptomycin, 1% L-glutamine, and 50 μg/ml gentamicin sulfate.

extract_protocol:
Total RNA was isolated from cells using Trizol reagent according to the manufacturer's instructions.
The construction of library was performed using QuantSeq 3' mRNA-Seg Library Prep Kit (Lexogen, Inc., Austria) according to the manufacturer's instructions.

value: Brain

value: Ink4a -/- murine astrocyte

value: astrocyte

value: JAG1 overexpression


title: 22Rv1 AR-Cterm 5UTR DMS Zota biol rep 1

description: Library name: Sample9

value: Prostate cancer cell line


title: 22Rv1 Vehicle RPF biol rep 2

description: Library name: Sample5


title: chk1chk2, H3, ChIP, 2

value: chk1chk2 knockout

value: 2% glucose with 1mg/mL HU in DD14


title: M18_CD73neg

value: 18 months


title: 22Rv1 Zota RNA biol rep 2

description: Library name: Sample8


title: M18_retina

value: Retina


title: M3_retina

value: 3 months


title: Rat alveolar macrophages, 24hr, Control for CNC1, CNC2 and CNC3, replicate 1

value: Rat alveolar macrophages  (NR8383)


title: 22Rv1 AR-Cterm 5UTR DMS Enza biol rep 2

description: Library name: Sample8


title: 22Rv1 Vehicle RPF biol rep 3

description: Library name: Sample6


title: 22Rv1 Vehicle RNA biol rep 3

description: Library name: Sample3


title: Ink4a-ast,JAG1,rep2


title: M3_CD73neg

value: male


title: Ink4a-ast,Con,rep3

value: WT


title: M12_retina

value: whole retina


title: 22Rv1 Vehicle RNA biol rep 2

description: Library name: Sample2


title: Ink4a-ast,Con,rep2


title: 22Rv1 AR-Cterm 5UTR DMS Vehicle biol rep 1

description: Library name: Sample5


title: 22Rv1 Vehicle RPF biol rep 1

description: Library name: Sample4


title: 22Rv1 HIF1A set 1 5UTR DMS Vehicle biol rep 1

description: Library name: Sample1


title: Ink4a-ast,Con,rep1


title: S1_Control

first: Shumei

last: Ma

address: No. 1111, Zhongshan North 1st Road

institute: Shanghai Institute of Pharmaceutical Industry Co., Ltd.

data_processing:
The Cell Ranger software pipeline (version 3.1.0) provided by 10×Genomics was used to demultiplex cellular barcodes, map reads to the genome and transcriptome using the STAR aligner, and down-sample reads as required to generate normalized aggregate data across samples, producing a matrix of gene counts versus cells
Genome_build: mm10
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every cell

molecule: polyA RNA

source_name: healthy mouse lung tissue

extract_protocol:
Single cells were suspended in calcium- and magnesium-free PBS containing 0.04% weight/volume BSA. Then Cells were added to each channel. After generation of GEMs, reverse transcription reactions were engaged barcoded full-length cDNA followed by the disruption of emulsions using the recovery agent and cDNA clean up with DynaBeads Myone Silane Beads (Thermo Fisher Scientific). Subsequently, the amplified cDNA was fragmented, end-repaired, A-tailed,  and index adaptor ligated and library amplification.
RNA libraries  were prepared with Chromium Single cell 3’ Reagent v3 Kits according to the manufacturer’s protocol.

value: 3 month

value: healthy mouse lung

value: Control


title: induction, day 10, time 48, litter 2

status: Public on Feb 15 2025

city: Seattle

first: Edward

last: Kelly

state: Washington

address: 1959 NE Pacific St

institute: University of Washington

description: Library name: C482_1

data_processing:
data aligned using STAR aligner
Assembly: mRatBN7.2
Supplementary files format and content: counts per million reads normalized from STAR aligner gene counts

source_name: testis

growth_protocol: testis cells maintained in microculture environemnt with 4.5 mg/mL matrigel and seeded at cell denisty of 5 million cells/mL. Media is serum free and 50% media is replaced every 24 hours.

extract_protocol:
RNA isolated via intital wash with corning cell recovery solution to remove excess matrigel, followed by lysis and disruption with Qiagen RLT buffer. Lyzed samples stored at -80 until analysis.
Lyzed samples sento to Novogene for isolation and sequencing using totalRNA  isolation and messenger RNA purification using poly-T oligo-attached magnetic beads. mRNA was then fragmented and the first strand cDNA was synthesized using random hexamer primers, followed by second strand cDNA synthesis using dTTP for nondirectional library, and sequenced by a NovaSEq 6000 system

treatment_protocol: hormone treatment added directly to fresh media

value: testis

value: primary cells

value: FSH and LH


title: 22Rv1 Zota RNA biol rep 1

description: Library name: Sample7


title: 22Rv1 HIF1A set 1 5UTR DMS Zota biol rep 2

description: Library name: Sample4

value: Zotatifin


title: Ink4a-ast,JAG1,rep1

value: Brain

value: Ink4a -/- murine astrocyte

value: astrocyte

value: JAG1 overexpression


title: 22Rv1 HIF1A set 1 5UTR DMS Zota biol rep 1

description: Library name: Sample3

value: Prostate cancer cell line


title: induction, day 10, time 12, litter 2

description: Library name: C122_1


title: S2_Model

value: Model


title: 22Rv1 HIF1A set 1 5UTR DMS Vehicle biol rep 2

description: Library name: Sample2


title: induction, day 10, time 2, litter 1

description: Library name: C21_1


title: induction, day 10, time 0, litter 3

description: Library name: C03_1

value: control


title: 22Rv1 Vehicle RNA biol rep 1

description: Library name: Sample1


title: induction, day 10, time 0, litter 2

description: Library name: C02_1


title: baseline, in vitro day 7, tissue, litter 1, replicate 2

description: Library name: D2_A4_Day7


title: induction, day 10, time 2, litter 2

description: Library name: C22_1


title: induction, day 10, time 48, litter 1

description: Library name: C481_1


title: induction, day 10, time 12, litter 3

description: Library name: C123_1


title: induction, day 10, time 12, litter 1

description: Library name: C121_1


title: induction, day 10, time 48, litter 3

description: Library name: C483_1


title: baseline, in vitro day 7, tissue, litter 1, replicate 3

description: Library name: D3_A4_Day7


title: S3_PD00


title: baseline, in vitro day 5, tissue, litter 1, replicate 2

description: Library name: D2_A3_Day5


title: baseline, in vitro day 5, tissue, litter 1, replicate 1

description: Library name: D1_A3_Day5


title: baseline, in vitro day 7, tissue, litter 1, replicate 1

description: Library name: D1_A4_Day7


title: baseline, in vitro day 3, tissue, litter 1, replicate 2

description: Library name: D3_A2_Day3


title: baseline, in vitro day 3, tissue, litter 1, replicate 3

description: Library name: D4_A2_Day3


title: baseline, tissue, litter 1, replicate 3

description: Library name: C_827_S3


title: baseline, tissue, litter 1, replicate 2

description: Library name: B_827_S2


title: induction, day 10, time 2, litter 3

description: Library name: C23_1


title: baseline, in vitro day 3, tissue, litter 1, replicate 1

description: Library name: D1_A2_Day3


title: baseline, tissue, litter 1, replicate 1

description: Library name: A_827_S1


title: R&Z_WT_CD_50d_2

city: Dresden

first: Meritxell

last: Huch

address: Pfotenhauerstrasse 108

country: Germany

institute: Max Planck Institute of Molecular Cell Biology and Genetics

description:
Library name: i707.i517
i707.i517

data_processing:
Basecalling by Illumina's standard pipeline
QC step, sickle reads with quality > Q20
QC step, trim_galore removing Illumina adaptors
Mapping step: STAR 2.5.0a extra paramaters outSAMmultNmax 1 --quantMode GeneCounts --outSAMunmapped Within
Quanitification step: featureCounts 1.5.0-p3 using NCBI mm10 gtf (mm10.ncbiRefSeq.gtf.gz)
Assembly: mm10
Supplementary files format and content: FeatureCounts output: Gene Name, Chromosome, Start,End,Strand,Length,Counts

source_name: adult liver

extract_protocol:
RNA was extracted from liver tissue using Quiagen Rneasy mini kit
SMARTSEQ2 protocol

value: adult liver

value: Wild-type

tag: chronic damage

value: No

tag: time point

value: 3 months

tag: batch


title: R&Z_WT_CD_170d_3

description:
Library name: i712.i517
i712.i517

value: 7 months


title: R&Z_WT_7M_5

description:
Library name: i705-i503
i705-i503

value: yes


title: R&Z_KO_7M_6

description:
Library name: i706.i517
i706.i517

value: Rnf43 and Znrf3 mutant


title: R&Z_KO_3M_3

description:
Library name: i704-i502
i704-i502


title: induction, day 10, time 0, litter 1

description: Library name: C01_1

value: testis

value: primary cells

value: WT


title: R&Z_KO_3M_2

description:
Library name: i701-i503
i701-i503


title: ATAC_E11.5Pdx1H_2

city: Beijing

first: Cheng-ran

last: Xu

address: NO.5 YIHEYUAN ROAD HAIDIAN DISTRICT, BEIJING, P.R.CHINA

department: School of Basic Medical Sciences

institute: Peking University

description: Library name: ATAC_E11.5Pdx1H_2

data_processing:
For bulk-cell ATAC-seq data, Overrepresented sequences were identified using FastQC (v0.11.3), and then removed along with sequencing adaptors, primers, and other overrepresented sequences using Trimmomatic (v0.38). The remaining sequencing reads were aligned to the Mus musculus genome (mm10) using bowtie2 (v2.2.5) with default parameters. PCR duplicate reads were subsequently removed using samtools (v1.3.1), and only reads with a mapping quality score ≥30 were retained for downstream analysis. Prior to peak calling, reads generated from biological replicates were combined. ATAC-seq peaks were then identified using MACS2 (v2.2.6), retaining peaks with a false discovery rate (FDR) less than 0.01. Genome regions overlapping with the ENCODE blacklist were excluded from further analysis. Tag intensities were calculated as TP10M values and stored in bedGraph files using Homer (v4.7.2).
Assembly: mm10
Supplementary files format and content: Bedgraph

extract_protocol:
Bulk-cell ATAC-seq was conducted following the published protocol  with modifications in cell sorting and nuclei preparation. Specifically, 2,000 cells per ATAC-seq were directly sorted into 50 ul lysis buffer in a 200 ul tube for nuclei preparation. Libraries were constructed using the TruePrep DNA Library Prep Kit V2 for Illumina (Vazyme, TD502). PCR amplification was performed for 12 cycles under the following conditions: 72°C for 3 minutes, 98°C for 30 seconds, then 12 cycles of 98°C for 15 seconds, 60°C for 30 seconds, and 72°C for 3 minutes, followed by a final extension at 72°C for 5 minutes.
The final DNA was purified using 1× VAHTS DNA Clean Beads and sequenced as 150 bp paired-end reads on the Illumina NovaSeq 6000 platform.


title: ATAC_E11.5Pdx1H_1

description: Library name: ATAC_E11.5Pdx1H_1


title: baseline, in vitro day 5, tissue, litter 1, replicate 3

description: Library name: D3_A3_Day5


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_28

description:
sm3_rc.cell_3.tsv
sm3_rc.gene_3.tsv
sm3_rc_3.mtx

data_processing:
For smart-seq3 snRNA-seq, Trimmomatic (v0.39) was utilized to remove primer and adaptor sequences from the reads. Subsequently, the trimmed forward reads (Read 1) were aligned to the mm10 mouse genome using HISAT2 (v2.1.0). The resulting aligned reads were annotated to gene features using featureCounts (v2.0.0), considering both the 5' and internal fragments. Cells were filtered out if they exhibited fewer than 4,000 detected genes. Gene expression levels were quantified using TPM.
Assembly: mm10
Supplementary files format and content: Sparce matrix

source_name: fetal pancreas

extract_protocol:
For Smart-seq3 scRNA-seq, single cells were directly sorted into 384-well plates containing lysis buffer. cDNA synthesis and amplification were carried out according to the Smart-seq3 protocol.
For Smart-seq3 scRNA-seq, libraries were constructed using the TruePrep DNA Library Prep Kit V2 for lllumina (Vazyme, TD501), and sequencing was conducted on the MGIseq-2000 platform, generating 100 bp paired-end reads.

value: fetal pancreas

tag: dataset

value: Smartseq3 scRNA-seq


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_30


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_29


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_25


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_24


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_27


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_26


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_23


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_22


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_21


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_20


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_19


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_17


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_16


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_15


title: R&Z_KO_CD_50d_3

description:
Library name: i711-i503
i711-i503


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_18


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_12


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_13


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_11


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_10


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_6


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_9


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_8


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_5


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_3


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_4


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_1


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_2


title: Pdx1CreNeurod1OE_E15.5_3_92


title: Pdx1CreNeurod1OE_E15.5_3_94


title: Pdx1CreNeurod1OE_E15.5_3_91


title: Pdx1CreNeurod1OE_E15.5_3_95


title: Pdx1CreNeurod1OE_E15.5_3_90


title: Pdx1CreNeurod1OE_E15.5_3_88


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_14


title: Pdx1CreNeurod1OE_E15.5_3_84


title: Pdx1CreNeurod1OE_E15.5_3_89


title: Pdx1CreNeurod1OE_E15.5_3_86


title: Pdx1CreNeurod1OE_E15.5_3_93


title: Pdx1CreNeurod1OE_E15.5_3_81


title: Pdx1CreNeurod1OE_E15.5_3_80


title: Pdx1CreNeurod1OE_E15.5_3_85


title: Pdx1CreNeurod1OE_E15.5_3_79


title: Pdx1CreNeurod1OE_E15.5_3_83


title: Pdx1CreNeurod1OE_E15.5_3_77


title: Pdx1CreNeurod1OE_E15.5_3_75


title: Pdx1CreNeurod1OE_E15.5_3_74


title: Pdx1CreNgn3KONeurod1OE_E15.5_3_7


title: Pdx1CreNeurod1OE_E15.5_3_78


title: Pdx1CreNeurod1OE_E15.5_3_73


title: Pdx1CreNeurod1OE_E15.5_3_71


title: Pdx1CreNeurod1OE_E15.5_3_72


title: Pdx1CreNeurod1OE_E15.5_3_68


title: Pdx1CreNeurod1OE_E15.5_3_70


title: Pdx1CreNeurod1OE_E15.5_3_67


title: Pdx1CreNeurod1OE_E15.5_3_87


title: Pdx1CreNeurod1OE_E15.5_3_66


title: Pdx1CreNeurod1OE_E15.5_3_65


title: Pdx1CreNeurod1OE_E15.5_3_62


title: Pdx1CreNeurod1OE_E15.5_3_64


title: Pdx1CreNeurod1OE_E15.5_3_69


title: Pdx1CreNeurod1OE_E15.5_3_61


title: Pdx1CreNeurod1OE_E15.5_3_55


title: Pdx1CreNeurod1OE_E15.5_3_82


title: Pdx1CreNeurod1OE_E15.5_3_56


title: Pdx1CreNeurod1OE_E15.5_3_63


title: Pdx1CreNeurod1OE_E15.5_3_58


title: Pdx1CreNeurod1OE_E15.5_3_53


title: Pdx1CreNeurod1OE_E15.5_3_54


title: Pdx1CreNeurod1OE_E15.5_3_60


title: Pdx1CreNeurod1OE_E15.5_3_57


title: Pdx1CreNeurod1OE_E15.5_3_50


title: Pdx1CreNeurod1OE_E15.5_3_76


title: Pdx1CreNeurod1OE_E15.5_3_49


title: Pdx1CreNeurod1OE_E15.5_3_51


title: Pdx1CreNeurod1OE_E15.5_3_48


title: Pdx1CreNeurod1OE_E15.5_3_47


title: Pdx1CreNeurod1OE_E15.5_3_44


title: Pdx1CreNeurod1OE_E15.5_3_45


title: Pdx1CreNeurod1OE_E15.5_3_52


title: Pdx1CreNeurod1OE_E15.5_3_43


title: Pdx1CreNeurod1OE_E15.5_3_42


title: Pdx1CreNeurod1OE_E15.5_3_41


title: Pdx1CreNeurod1OE_E15.5_3_40


title: Pdx1CreNeurod1OE_E15.5_3_38


title: Pdx1CreNeurod1OE_E15.5_3_59


title: Pdx1CreNeurod1OE_E15.5_3_37


title: Pdx1CreNeurod1OE_E15.5_3_36


title: Pdx1CreNeurod1OE_E15.5_3_33


title: Pdx1CreNeurod1OE_E15.5_3_31


title: Pdx1CreNeurod1OE_E15.5_3_30


title: Pdx1CreNeurod1OE_E15.5_3_32


title: Pdx1CreNeurod1OE_E15.5_3_29


title: Pdx1CreNeurod1OE_E15.5_3_28


title: Pdx1CreNeurod1OE_E15.5_3_26


title: Pdx1CreNeurod1OE_E15.5_3_25


title: Pdx1CreNeurod1OE_E15.5_3_24


title: Pdx1CreNeurod1OE_E15.5_3_46


title: Pdx1CreNeurod1OE_E15.5_3_23


title: Pdx1CreNeurod1OE_E15.5_3_35


title: Pdx1CreNeurod1OE_E15.5_3_21


title: Pdx1CreNeurod1OE_E15.5_3_20


title: Pdx1CreNeurod1OE_E15.5_3_19


title: Pdx1CreNeurod1OE_E15.5_3_17


title: Pdx1CreNeurod1OE_E15.5_3_16


title: Pdx1CreNeurod1OE_E15.5_3_39


title: Pdx1CreNeurod1OE_E15.5_3_14


title: Pdx1CreNeurod1OE_E15.5_3_15


title: Pdx1CreNeurod1OE_E15.5_3_13


title: Pdx1CreNeurod1OE_E15.5_3_12


title: Pdx1CreNeurod1OE_E15.5_3_11


title: Pdx1CreNeurod1OE_E15.5_3_34


title: Pdx1CreNeurod1OE_E15.5_3_10


title: Pdx1CreNeurod1OE_E15.5_3_9


title: Pdx1CreNeurod1OE_E15.5_3_8


title: Pdx1CreNeurod1OE_E15.5_3_5


title: Pdx1CreNeurod1OE_E15.5_3_22


title: Pdx1CreNeurod1OE_E15.5_3_4


title: Pdx1CreNeurod1OE_E15.5_3_1


title: Pdx1CreNeurod1OE_E15.5_3_7


title: Pdx1CreNeurod1OE_E15.5_3_2


title: Pdx1CreNeurod1OE_E15.5_3_6


title: Pdx1CreNeurod1OE_E15.5_3_27


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_202


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_203


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_205


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_200


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_199


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_191


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_204


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_194


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_198


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_192


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_193


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_190


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_196


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_195


title: Pdx1CreNeurod1OE_E15.5_3_18


title: Pdx1CreNeurod1OE_E15.5_3_3


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_189


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_197


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_187


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_188


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_186


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_201


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_183


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_184


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_181


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_182


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_180


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_179


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_178


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_185


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_176


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_177


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_169


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_170


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_168


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_167


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_166


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_160


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_163


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_162


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_165


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_172


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_171


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_158


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_173


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_161


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_159


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_156


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_155


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_154


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_175


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_153


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_174


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_151


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_150


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_149


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_148


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_147


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_143


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_142


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_145


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_144


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_141


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_140


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_146


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_139


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_157


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_138


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_136


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_135


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_133


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_132


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_131


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_134


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_130


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_152


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_127


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_129


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_123


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_125


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_164


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_128


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_124


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_122


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_121


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_120


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_137


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_117


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_118


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_114


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_113


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_110


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_119


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_109


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_115


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_112


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_108


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_106


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_111


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_107


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_104


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_105


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_103


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_101


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_98


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_97


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_95


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_96


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_99


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_100


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_116


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_102


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_93


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_126


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_91


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_92


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_89


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_94


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_84


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_80


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_81


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_87


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_85


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_79


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_78


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_82


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_75


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_77


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_76


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_74


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_70


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_73


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_69


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_72


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_67


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_66


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_90


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_68


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_64


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_86


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_88


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_62


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_83


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_60


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_61


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_63


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_58


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_59


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_57


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_56


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_52


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_54


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_53


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_71


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_50


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_51


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_48


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_47


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_65


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_46


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_45


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_44


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_42


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_43


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_41


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_39


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_38


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_40


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_37


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_55


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_36


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_35


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_34


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_32


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_31


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_30


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_27


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_28


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_26


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_49


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_25


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_24


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_22


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_23


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_21


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_20


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_19


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_18


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_15


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_16


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_14


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_13


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_12


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_33


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_11


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_9


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_10


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_6


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_8


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_7


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_29


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_2


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_3


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_1


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_5


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_4


title: Pdx1CreNeurod1OE_E15.5_2_115


title: Pdx1CreNeurod1OE_E15.5_2_114


title: Pdx1CreNeurod1OE_E15.5_2_116


title: Pdx1CreNeurod1OE_E15.5_2_112


title: Pdx1CreNeurod1OE_E15.5_2_111


title: Pdx1CreNeurod1OE_E15.5_2_113


title: Pdx1CreNgn3KONeurod1OE_E15.5_2_17


title: Pdx1CreNeurod1OE_E15.5_2_109


title: Pdx1CreNeurod1OE_E15.5_2_117


title: Pdx1CreNeurod1OE_E15.5_2_110


title: Pdx1CreNeurod1OE_E15.5_2_107


title: Pdx1CreNeurod1OE_E15.5_2_108


title: Pdx1CreNeurod1OE_E15.5_2_106


title: Pdx1CreNeurod1OE_E15.5_2_105


title: Pdx1CreNeurod1OE_E15.5_2_104


title: Pdx1CreNeurod1OE_E15.5_2_103


title: Pdx1CreNeurod1OE_E15.5_2_102


title: Pdx1CreNeurod1OE_E15.5_2_100


title: Pdx1CreNeurod1OE_E15.5_2_101


title: Pdx1CreNeurod1OE_E15.5_2_98


title: Pdx1CreNeurod1OE_E15.5_2_99


title: Pdx1CreNeurod1OE_E15.5_2_97


title: Pdx1CreNeurod1OE_E15.5_2_95


title: Pdx1CreNeurod1OE_E15.5_2_94


title: Pdx1CreNeurod1OE_E15.5_2_92


title: Pdx1CreNeurod1OE_E15.5_2_91


title: Pdx1CreNeurod1OE_E15.5_2_96


title: Pdx1CreNeurod1OE_E15.5_2_90


title: Pdx1CreNeurod1OE_E15.5_2_89


title: Pdx1CreNeurod1OE_E15.5_2_88


title: Pdx1CreNeurod1OE_E15.5_2_84


title: Pdx1CreNeurod1OE_E15.5_2_85


title: Pdx1CreNeurod1OE_E15.5_2_86


title: Pdx1CreNeurod1OE_E15.5_2_82


title: Pdx1CreNeurod1OE_E15.5_2_83


title: Pdx1CreNeurod1OE_E15.5_2_81


title: Pdx1CreNeurod1OE_E15.5_2_78


title: Pdx1CreNeurod1OE_E15.5_2_80


title: Pdx1CreNeurod1OE_E15.5_2_79


title: Pdx1CreNeurod1OE_E15.5_2_77


title: Pdx1CreNeurod1OE_E15.5_2_74


title: Pdx1CreNeurod1OE_E15.5_2_76


title: Pdx1CreNeurod1OE_E15.5_2_75


title: Pdx1CreNeurod1OE_E15.5_2_73


title: Pdx1CreNeurod1OE_E15.5_2_72


title: Pdx1CreNeurod1OE_E15.5_2_71


title: Pdx1CreNeurod1OE_E15.5_2_67


title: Pdx1CreNeurod1OE_E15.5_2_68


title: Pdx1CreNeurod1OE_E15.5_2_66


title: Pdx1CreNeurod1OE_E15.5_2_93


title: Pdx1CreNeurod1OE_E15.5_2_70


title: Pdx1CreNeurod1OE_E15.5_2_65


title: Pdx1CreNeurod1OE_E15.5_2_64


title: Pdx1CreNeurod1OE_E15.5_2_60


title: Pdx1CreNeurod1OE_E15.5_2_63


title: Pdx1CreNeurod1OE_E15.5_2_62


title: Pdx1CreNeurod1OE_E15.5_2_87


title: Pdx1CreNeurod1OE_E15.5_2_59


title: Pdx1CreNeurod1OE_E15.5_2_61


title: Pdx1CreNeurod1OE_E15.5_2_56


title: Pdx1CreNeurod1OE_E15.5_2_53


title: Pdx1CreNeurod1OE_E15.5_2_57


title: Pdx1CreNeurod1OE_E15.5_2_55


title: Pdx1CreNeurod1OE_E15.5_2_54


title: Pdx1CreNeurod1OE_E15.5_2_52


title: Pdx1CreNeurod1OE_E15.5_2_48


title: Pdx1CreNeurod1OE_E15.5_2_50


title: Pdx1CreNeurod1OE_E15.5_2_69


title: Pdx1CreNeurod1OE_E15.5_2_49


title: Pdx1CreNeurod1OE_E15.5_2_46


title: Pdx1CreNeurod1OE_E15.5_2_47


title: Pdx1CreNeurod1OE_E15.5_2_38


title: Pdx1CreNeurod1OE_E15.5_2_45


title: Pdx1CreNeurod1OE_E15.5_2_42


title: Pdx1CreNeurod1OE_E15.5_2_43


title: Pdx1CreNeurod1OE_E15.5_2_44


title: Pdx1CreNeurod1OE_E15.5_2_37


title: Pdx1CreNeurod1OE_E15.5_2_35


title: Pdx1CreNeurod1OE_E15.5_2_32


title: Pdx1CreNeurod1OE_E15.5_2_58


title: Pdx1CreNeurod1OE_E15.5_2_39


title: Pdx1CreNeurod1OE_E15.5_2_31


title: Pdx1CreNeurod1OE_E15.5_2_30


title: Pdx1CreNeurod1OE_E15.5_2_41


title: Pdx1CreNeurod1OE_E15.5_2_34


title: Pdx1CreNeurod1OE_E15.5_2_51


title: Pdx1CreNeurod1OE_E15.5_2_29


title: Pdx1CreNeurod1OE_E15.5_2_33


title: Pdx1CreNeurod1OE_E15.5_2_27


title: Pdx1CreNeurod1OE_E15.5_2_25


title: Pdx1CreNeurod1OE_E15.5_2_26


title: Pdx1CreNeurod1OE_E15.5_2_24


title: Pdx1CreNeurod1OE_E15.5_2_21


title: Pdx1CreNeurod1OE_E15.5_2_23


title: Pdx1CreNeurod1OE_E15.5_2_20


title: Pdx1CreNeurod1OE_E15.5_2_22


title: Pdx1CreNeurod1OE_E15.5_2_19


title: Pdx1CreNeurod1OE_E15.5_2_18


title: Pdx1CreNeurod1OE_E15.5_2_40


title: Pdx1CreNeurod1OE_E15.5_2_36


title: Pdx1CreNeurod1OE_E15.5_2_17


title: Pdx1CreNeurod1OE_E15.5_2_16


title: Pdx1CreNeurod1OE_E15.5_2_15


title: Pdx1CreNeurod1OE_E15.5_2_14


title: Pdx1CreNeurod1OE_E15.5_2_13


title: Pdx1CreNeurod1OE_E15.5_2_11


title: Pdx1CreNeurod1OE_E15.5_2_12


title: Pdx1CreNeurod1OE_E15.5_2_10


title: Pdx1CreNeurod1OE_E15.5_2_9


title: Pdx1CreNeurod1OE_E15.5_2_8


title: Pdx1CreNeurod1OE_E15.5_2_28


title: Pdx1CreNeurod1OE_E15.5_2_7


title: Pdx1CreNeurod1OE_E15.5_2_2


title: Pdx1CreNeurod1OE_E15.5_2_1


title: Pdx1CreNeurod1OE_E15.5_2_3


title: Pdx1CreNeurod1OE_E15.5_2_5


title: WT_E15.5_744

description:
Smartseq3 scRNA-seq
sm3_rc.cell.tsv
sm3_rc.gene.tsv
sm3_rc.mtx

data_processing:
For CUT&RUN data, overrepresented sequences were identified using FastQC (v0.11.3), and then removed along with sequencing adaptors, primers, and other overrepresented sequences using Trimmomatic (v0.38). The remaining sequencing reads were aligned to the Mus musculus genome (mm10) using bowtie2 (v2.2.5) with default parameters. PCR duplicate reads were subsequently removed using samtools (v1.3.1), and only reads with a mapping quality score ≥30 were retained for downstream analysis.  Peak calling was performed on individual samples using MACS2 (v2.2.6), retaining peaks with a P value less than 10-3. Reproducibility between two biological replicates was assessed using idr (v2.0.4.2). Different irreproducible discovery rate (IDR) thresholds were applied depending on the CUT&RUN targets. For CUT&RUN targeting NGN3 in Ngn3low cells, peaks with an IDR larger than 250 were retained for downstream analysis. For CUT&RUN targeting NGN3 in Ngn3high cells and NEUROD1, the threshold was set to 400. Genome regions overlapping with the ENCODE blacklist were filtered out from the final identified peaks. Tag intensities were calculated as Tags per ten million (TP10M) values and stored in bedGraph files using Homer (v4.7.2).
For bulk-cell ATAC-seq data, Overrepresented sequences were identified using FastQC (v0.11.3), and then removed along with sequencing adaptors, primers, and other overrepresented sequences using Trimmomatic (v0.38). The remaining sequencing reads were aligned to the Mus musculus genome (mm10) using bowtie2 (v2.2.5) with default parameters. PCR duplicate reads were subsequently removed using samtools (v1.3.1), and only reads with a mapping quality score ≥30 were retained for downstream analysis.  Prior to peak calling, reads generated from biological replicates were combined. ATAC-seq peaks were then identified using MACS2 (v2.2.6), retaining peaks with a false discovery rate (FDR) less than 0.01. Genome regions overlapping with the ENCODE blacklist were excluded from further analysis. Tag intensities were calculated as TP10M values and stored in bedGraph files using Homer (v4.7.2).
For snATAC-seq, preprocessing was conducted using cellranger-atac (v1.2.0) with default parameters against the refdata-cellranger-atac-mm10-1.2.0 genome. Cells with detected UMI counts greater than 1,000 and reads in promoter regions (RIPR) between 15% and 45% were retained for downstream analysis. The mm10 genome was segmented into discrete 5 kb intervals, serving as bins for each cell. The tag intensities for each bin were calculated based on the frequency of sequence tags and imported into the R package Signac (v1.3.0) for downstream analysis.
For smart-seq2 snRNA-seq, sequencing reads were aligned to the mm10 mouse genome assembly using TopHat (v2.1.0). Gene-level quantification of mapped reads was performed with HTSeq (v0.6.6) using the parameters “-s no -a 30”. Cells exhibiting fewer than 0.2 million mapped reads or fewer than 4,000 detected genes were excluded from further analysis. Gene expression levels were quantified using TPM (transcripts per million).
For smart-seq3 snRNA-seq, Trimmomatic (v0.39) was utilized to remove primer and adaptor sequences from the reads. Subsequently, the trimmed forward reads (Read 1) were aligned to the mm10 mouse genome using HISAT2 (v2.1.0). The resulting aligned reads were annotated to gene features using featureCounts (v2.0.0), considering both the 5' and internal fragments. Cells were filtered out if they exhibited fewer than 4,000 detected genes. Gene expression levels were quantified using TPM.
For 10x Genomics scRNA-seq, preprocessing was conducted using Cell Ranger (v3.0.1) with default parameters aligned to the refdata-cellranger-mm10-1.2.0 reference genome. The “filtered_feature_bc_matrix” output from Cell Ranger was imported into the Seurat package (version 4.1.1) for downstream analysis in R. WT and CKO or OE cells within the same sample were initially distinguished based on TP10K (transcripts per 10 thousand) values of Uty, Eif2s3y, and Ddx3y for male cells, and the TP10K value of Xist for female cells. These values were log-transformed, with cells exhibiting male-related values greater than 0.2 and female-related values equal to 0 classified as male, while cells with female-related values greater than 1 and male-related values equal to 0 classified as female. The downstream analysis, including dimensional reduction, cell clustering, and differential expression analysis, was conducted using the standard Seurat pipeline.
Assembly: mm10
Supplementary files format and content: Sparce matrix, Bedgraph

extract_protocol:
For Smart-seq2 scRNA-seq, single cells were manually picked from FACS-purified bulk cells using a mouth pipette. cDNA synthesis and amplification were performed following the protocol by Picelli et al. For Smart-seq3 scRNA-seq, single cells were directly sorted into 384-well plates containing lysis buffer. cDNA synthesis and amplification were carried out according to the Smart-seq3 protocol .For 10x Genomics scRNA-seq, the procedure was performed according to the manufacturer’s instructions using the Single Cell 3′ Reagent Kit v3.1. Sorted EpCAM+ pancreatic cells from wild-type (WT), conditional knockout (CKO), or overexpression (OE) embryos of different genders were pooled and processed as one sample. For NGN3-CKO samples, E14.5 EpCAM+ pancreatic cells were pooled per replicate: replicate 1 consisted of two WT male litters and one CKO female litter, and replicate 2 consisted of three WT male litters and three CKO female litters. For NGN3-OE samples, E14.5 EpCAM+ pancreatic cells were pooled from four WT female litters and four OE male litters, and E17.5 EpCAM+ pancreatic cells were pooled from two WT female litters and three OE male litters. For NeuroD1-CKO samples, E15.5 EpCAM+ pancreatic cells were pooled per replicate: replicate 1 consisted of one WT male litter and one CKO female litter, and replicate 2 consisted of two WT female litters and one CKO male litter. The gender of the embryos was determined by genotyping for Y-chromosome-associated gene Sry.
For Smart-seq2 scRNA-seq, libraries were prepared with 2 ng of cDNA using the TruePrep DNA Library Prep Kit (Vazyme, TD502), and sequenced on the Illumina HiSeq 4000 system, generating 150 bp paired-end reads. For Smart-seq3 scRNA-seq, libraries were constructed using the TruePrep DNA Library Prep Kit V2 for lllumina (Vazyme, TD501), and sequencing was conducted on the MGIseq-2000 platform, generating 100 bp paired-end reads. For 10x Genomics scRNA-seq, libraries were sequenced as 150-bp paired-end reads on an Illumina HiSeq 4000 system.
CUT&RUN was conducted following the published protocol with modifications in DNA purification and library construction steps. We used 30,000 cells per CUR&RUN assay with primary antibodies against FLAG (1:25, CST, 2368S), p300 (1;100, CST, 54062S), and NeuroD1 (1:100, Millpore, ABE991). DNA was purified using a spin column (VISTECH, DC2005) and processed for library construction with KAPA Hyper Prep Kit (KAPA biosystems, KK8502). Briefly, after end repair and adaptor ligation, the reaction was purified using VAHTS DNA Clean Beads (Vazyme, N411-03-AA) and subjected to PCR amplification with 2× KAPA HiFi HotStart ReadyMix (KAPA Biosystems, KM2602) and Illumina universal primers. The PCR program was: 98°C for 45 seconds, followed by 19 cycles of 98°C for 15 seconds, 60°C for 10 seconds, and 72°C for 1 minute, with a final extension at 72°C for 5 minutes. The final DNA was purified using 1× VAHTS DNA Clean Beads.
CUT&RUN libraries were subjected to 150 bp paired-end sequencing or 51 bp single-end sequencing on the Illumina NovaSeq 6000 or Illumina Hiseq 2500 platform.
Bulk-cell ATAC-seq was conducted following the published protocol  with modifications in cell sorting and nuclei preparation. Specifically, 2,000 cells per ATAC-seq were directly sorted into 50 ul lysis buffer in a 200 ul tube for nuclei preparation. Libraries were constructed using the TruePrep DNA Library Prep Kit V2 for Illumina (Vazyme, TD502). PCR amplification was performed for 12 cycles under the following conditions: 72°C for 3 minutes, 98°C for 30 seconds, then 12 cycles of 98°C for 15 seconds, 60°C for 30 seconds, and 72°C for 3 minutes, followed by a final extension at 72°C for 5 minutes.
The final DNA was purified using 1× VAHTS DNA Clean Beads and sequenced as 150 bp paired-end reads on the Illumina NovaSeq 6000 platform.
For snATAC-seq, the procedure was performed according to the manufacturer’s instructions using the Chromium Next GEM Single Cell ATAC Library & Gel Bead Kit v1.1 (10x Genomics). For the Ngn3-CKO samples, E14.5 EpCAM+ pancreatic cells of WT and CKO embryos with different genders (two WT male litters and two CKO female litters) were pooled and loaded as one sample. To ensure experimental success and an adequate number of nuclei, extra pancreatic cells from WT embryos were included, with a ratio of to EpCAM+ cells of 1:4. For the NeuroD1-CKO samples, E15.5 EpCAM+ pancreatic cells of WT and CKO embryos with different genders (two WT female litters and three CKO male litters) were pooled and loaded as one sample.
Libraries were sequenced as 50-bp paired-end reads on an NovaSeq 6000 system.
snATAC-seq

value: WTEpcam+


title: WT_E15.5_740


title: WT_E15.5_743


title: WT_E15.5_742


title: WT_E15.5_747


title: WT_E15.5_748


title: WT_E15.5_739


title: WT_E15.5_746


title: WT_E15.5_738


title: WT_E15.5_737


title: WT_E15.5_735


title: WT_E15.5_736


title: WT_E15.5_733


title: WT_E15.5_734


title: Pdx1CreNeurod1OE_E15.5_2_6


title: WT_E15.5_731


title: WT_E15.5_730


title: WT_E15.5_732


title: WT_E15.5_728


title: Pdx1CreNeurod1OE_E15.5_2_4


title: WT_E15.5_749


title: WT_E15.5_729


title: WT_E15.5_726


title: WT_E15.5_725


title: WT_E15.5_723


title: WT_E15.5_727


title: WT_E15.5_724


title: WT_E15.5_722


title: WT_E15.5_741


title: WT_E15.5_721


title: WT_E15.5_716


title: WT_E15.5_715


title: WT_E15.5_719


title: WT_E15.5_717


title: WT_E15.5_713


title: WT_E15.5_720


title: WT_E15.5_710


title: WT_E15.5_711


title: WT_E15.5_709


title: WT_E15.5_712


title: WT_E15.5_708


title: WT_E15.5_707


title: WT_E15.5_705


title: WT_E15.5_706


title: WT_E15.5_704


title: WT_E15.5_703


title: WT_E15.5_702


title: WT_E15.5_701


title: WT_E15.5_700


title: WT_E15.5_699


title: WT_E15.5_696


title: WT_E15.5_697


title: WT_E15.5_693


title: WT_E15.5_694


title: WT_E15.5_695


title: WT_E15.5_714


title: WT_E15.5_692


title: WT_E15.5_690


title: WT_E15.5_688


title: WT_E15.5_718


title: WT_E15.5_687


title: WT_E15.5_685


title: WT_E15.5_689


title: WT_E15.5_684


title: WT_E15.5_691


title: WT_E15.5_683


title: WT_E15.5_681


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title: WT_E15.5_5


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title: Pdx1CreNgn3KONeurod1OE_E12.5_254


title: Pdx1CreNgn3KONeurod1OE_E12.5_249


title: Pdx1CreNgn3KONeurod1OE_E12.5_248


title: Pdx1CreNgn3KONeurod1OE_E12.5_244


title: Pdx1CreNgn3KONeurod1OE_E12.5_243


title: Pdx1CreNgn3KONeurod1OE_E12.5_241


title: Pdx1CreNgn3KONeurod1OE_E12.5_245


title: Pdx1CreNgn3KONeurod1OE_E12.5_242


title: Pdx1CreNgn3KONeurod1OE_E12.5_239


title: Pdx1CreNgn3KONeurod1OE_E12.5_262


title: Pdx1CreNgn3KONeurod1OE_E12.5_240


title: Pdx1CreNgn3KONeurod1OE_E12.5_236


title: Pdx1CreNgn3KONeurod1OE_E12.5_238


title: Pdx1CreNgn3KONeurod1OE_E12.5_258


title: Pdx1CreNgn3KONeurod1OE_E12.5_235


title: Pdx1CreNgn3KONeurod1OE_E12.5_229


title: Pdx1CreNgn3KONeurod1OE_E12.5_231


title: Pdx1CreNgn3KONeurod1OE_E12.5_233


title: Pdx1CreNgn3KONeurod1OE_E12.5_232


title: Pdx1CreNgn3KONeurod1OE_E12.5_250


title: Pdx1CreNgn3KONeurod1OE_E12.5_230


title: Pdx1CreNgn3KONeurod1OE_E12.5_228


title: Pdx1CreNgn3KONeurod1OE_E12.5_234


title: Pdx1CreNgn3KONeurod1OE_E12.5_226


title: Pdx1CreNgn3KONeurod1OE_E12.5_225


title: Pdx1CreNgn3KONeurod1OE_E12.5_223


title: Pdx1CreNgn3KONeurod1OE_E12.5_224


title: Pdx1CreNgn3KONeurod1OE_E12.5_222


title: Pdx1CreNgn3KONeurod1OE_E12.5_221


title: Pdx1CreNgn3KONeurod1OE_E12.5_227


title: Pdx1CreNgn3KONeurod1OE_E12.5_247


title: Pdx1CreNgn3KONeurod1OE_E12.5_219


title: Pdx1CreNgn3KONeurod1OE_E12.5_237


title: Pdx1CreNgn3KONeurod1OE_E12.5_217


title: Pdx1CreNgn3KONeurod1OE_E12.5_215


title: Pdx1CreNgn3KONeurod1OE_E12.5_218


title: Pdx1CreNgn3KONeurod1OE_E12.5_246


title: Pdx1CreNgn3KONeurod1OE_E12.5_213


title: Pdx1CreNgn3KONeurod1OE_E12.5_212


title: Pdx1CreNgn3KONeurod1OE_E12.5_214


title: Pdx1CreNgn3KONeurod1OE_E12.5_211


title: Pdx1CreNgn3KONeurod1OE_E12.5_209


title: Pdx1CreNgn3KONeurod1OE_E12.5_210


title: Pdx1CreNgn3KONeurod1OE_E12.5_207


title: Pdx1CreNgn3KONeurod1OE_E12.5_208


title: Pdx1CreNgn3KONeurod1OE_E12.5_204


title: Pdx1CreNgn3KONeurod1OE_E12.5_206


title: Pdx1CreNgn3KONeurod1OE_E12.5_202


title: Pdx1CreNgn3KONeurod1OE_E12.5_201


title: Pdx1CreNgn3KONeurod1OE_E12.5_199


title: Pdx1CreNgn3KONeurod1OE_E12.5_198


title: Pdx1CreNgn3KONeurod1OE_E12.5_197


title: Pdx1CreNgn3KONeurod1OE_E12.5_220


title: Pdx1CreNgn3KONeurod1OE_E12.5_196


title: Pdx1CreNgn3KONeurod1OE_E12.5_194


title: Pdx1CreNgn3KONeurod1OE_E12.5_193


title: Pdx1CreNgn3KONeurod1OE_E12.5_200


title: Pdx1CreNgn3KONeurod1OE_E12.5_195


title: Pdx1CreNgn3KONeurod1OE_E12.5_192


title: Pdx1CreNgn3KONeurod1OE_E12.5_216


title: Pdx1CreNgn3KONeurod1OE_E12.5_191


title: Pdx1CreNgn3KONeurod1OE_E12.5_189


title: Pdx1CreNgn3KONeurod1OE_E12.5_187


title: Pdx1CreNgn3KONeurod1OE_E12.5_186


title: Pdx1CreNgn3KONeurod1OE_E12.5_190


title: Pdx1CreNgn3KONeurod1OE_E12.5_185


title: Pdx1CreNgn3KONeurod1OE_E12.5_184


title: Pdx1CreNgn3KONeurod1OE_E12.5_183


title: Pdx1CreNgn3KONeurod1OE_E12.5_181


title: Pdx1CreNgn3KONeurod1OE_E12.5_180


title: Pdx1CreNgn3KONeurod1OE_E12.5_182


title: Pdx1CreNgn3KONeurod1OE_E12.5_175


title: Pdx1CreNgn3KONeurod1OE_E12.5_179


title: Pdx1CreNgn3KONeurod1OE_E12.5_178


title: Pdx1CreNgn3KONeurod1OE_E12.5_177


title: Pdx1CreNgn3KONeurod1OE_E12.5_173


title: Pdx1CreNgn3KONeurod1OE_E12.5_171


title: Pdx1CreNgn3KONeurod1OE_E12.5_172


title: Pdx1CreNgn3KONeurod1OE_E12.5_170


title: Pdx1CreNgn3KONeurod1OE_E12.5_169


title: Pdx1CreNgn3KONeurod1OE_E12.5_166


title: Pdx1CreNgn3KONeurod1OE_E12.5_174


title: Pdx1CreNgn3KONeurod1OE_E12.5_165


title: Pdx1CreNgn3KONeurod1OE_E12.5_176


title: Pdx1CreNgn3KONeurod1OE_E12.5_164


title: Pdx1CreNgn3KONeurod1OE_E12.5_188


title: Pdx1CreNgn3KONeurod1OE_E12.5_156


title: Pdx1CreNgn3KONeurod1OE_E12.5_162


title: Pdx1CreNgn3KONeurod1OE_E12.5_160


title: Pdx1CreNgn3KONeurod1OE_E12.5_155


title: Pdx1CreNgn3KONeurod1OE_E12.5_158


title: Pdx1CreNgn3KONeurod1OE_E12.5_154


title: Pdx1CreNgn3KONeurod1OE_E12.5_161


title: Pdx1CreNgn3KONeurod1OE_E12.5_163


title: Pdx1CreNgn3KONeurod1OE_E12.5_159


title: Pdx1CreNgn3KONeurod1OE_E12.5_151


title: Pdx1CreNgn3KONeurod1OE_E12.5_168


title: Pdx1CreNgn3KONeurod1OE_E12.5_167


title: Pdx1CreNgn3KONeurod1OE_E12.5_142


title: Pdx1CreNgn3KONeurod1OE_E12.5_157


title: Pdx1CreNgn3KONeurod1OE_E12.5_148


title: Pdx1CreNgn3KONeurod1OE_E12.5_147


title: Pdx1CreNgn3KONeurod1OE_E12.5_146


title: Pdx1CreNgn3KONeurod1OE_E12.5_141


title: Pdx1CreNgn3KONeurod1OE_E12.5_145


title: Pdx1CreNgn3KONeurod1OE_E12.5_150


title: Pdx1CreNgn3KONeurod1OE_E12.5_203


title: Pdx1CreNgn3KONeurod1OE_E12.5_140


title: Pdx1CreNgn3KONeurod1OE_E12.5_137


title: Pdx1CreNgn3KONeurod1OE_E12.5_138


title: Pdx1CreNgn3KONeurod1OE_E12.5_136


title: Pdx1CreNgn3KONeurod1OE_E12.5_152


title: Pdx1CreNgn3KONeurod1OE_E12.5_153


title: Pdx1CreNgn3KONeurod1OE_E12.5_130


title: Pdx1CreNgn3KONeurod1OE_E12.5_131


title: Pdx1CreNgn3KONeurod1OE_E12.5_132


title: Pdx1CreNgn3KONeurod1OE_E12.5_129


title: Pdx1CreNgn3KONeurod1OE_E12.5_149


title: Pdx1CreNgn3KONeurod1OE_E12.5_135


title: Pdx1CreNgn3KONeurod1OE_E12.5_134


title: Pdx1CreNgn3KONeurod1OE_E12.5_144


title: Pdx1CreNgn3KONeurod1OE_E12.5_128


title: Pdx1CreNgn3KONeurod1OE_E12.5_139


title: Pdx1CreNgn3KONeurod1OE_E12.5_127


title: Pdx1CreNgn3KONeurod1OE_E12.5_126


title: Pdx1CreNgn3KONeurod1OE_E12.5_125


title: Pdx1CreNgn3KONeurod1OE_E12.5_124


title: Pdx1CreNgn3KONeurod1OE_E12.5_123


title: Pdx1CreNgn3KONeurod1OE_E12.5_122


title: Pdx1CreNgn3KONeurod1OE_E12.5_120


title: Pdx1CreNgn3KONeurod1OE_E12.5_121


title: Pdx1CreNgn3KONeurod1OE_E12.5_133


title: Pdx1CreNgn3KONeurod1OE_E12.5_143


title: Pdx1CreNgn3KONeurod1OE_E12.5_116


title: Pdx1CreNgn3KONeurod1OE_E12.5_117


title: Pdx1CreNgn3KONeurod1OE_E12.5_118


title: Pdx1CreNgn3KONeurod1OE_E12.5_113


title: Pdx1CreNgn3KONeurod1OE_E12.5_114


title: Pdx1CreNgn3KONeurod1OE_E12.5_111


title: Pdx1CreNgn3KONeurod1OE_E12.5_110


title: Pdx1CreNgn3KONeurod1OE_E12.5_109


title: Pdx1CreNgn3KONeurod1OE_E12.5_107


title: Pdx1CreNgn3KONeurod1OE_E12.5_106


title: Pdx1CreNgn3KONeurod1OE_E12.5_205


title: Pdx1CreNgn3KONeurod1OE_E12.5_103


title: Pdx1CreNgn3KONeurod1OE_E12.5_105


title: Pdx1CreNgn3KONeurod1OE_E12.5_112


title: Pdx1CreNgn3KONeurod1OE_E12.5_101


title: Pdx1CreNgn3KONeurod1OE_E12.5_100