title: RNA_seq_8hpi_rep2

data_processing:
Basecalling with Illumina Casava 1.8 software
Base correction, filtering, 3' adapter trimming with Fastp
Alignment to rRNA and other non-coding RNA with STAR v2.7.0a
Alignment to genome with STAR v2.7.0a
Read quantitation using custom scripts
Assembly: Amoeba genome assembly: https://zenodo.org/records/6800059; Amoeba mitochondrial genome: NC_001637.1; Virus genome: NC_014649.1
Supplementary files format and content: csv file contains 9 columns. The first column is transcript name. The 2nd to 9th columns are read counts in each sample.

extract_protocol:
The cell pellets were resuspended with 600 µl lysis buffer (20mM Tris pH 7.5, 150mM NaCl, 5mM MgCl2, 1mM dithiothreitol, 1% Triton X-100, 100 µg/ml of cycloheximide, and 100 µg/ml chloramphenicol), treated with 5 μl of TURBO DNase (2 U/μl, ThermoFisher Scientific) on ice for 10 min, and then centrifuged at 20,000 × g, at 4°C for 10 min.
Total RNA was purified using TRIzol-LS and Direct-zol RNA Microprep kit (Zymo Research). RNA-Seq libraries were prepared with TruSeq Stranded Total RNA Library Prep Gold (Illumina).


title: HEK293T cell, EMX1, G5T, loop2-13-18-7-M, PE2, rep3

status: Public on Nov 17 2025

type: SRA

city: Hangzhou

first: Lijia

last: Ma

state: Zhejiang

address: Shilongshan Road No.18, Cloud Town, Xihu District

department: School of Life Sciences

country: China

institute: Westlake university

description:
5856
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1

library_source: genomic

molecule: genomic DNA

organism: Homo sapiens

growth_protocol: The culture conditions for HEK293T cells typically involve maintaining the cells in a high-glucose Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin antibiotic solution. The medium is further enriched with 2 mM L-glutamine for optimal cell growth and health. Cells should be cultured at 37°C in a humidified atmosphere with 5% CO2. Regular media replacement is essential, usually every 2-3 days, to prevent the accumulation of metabolic waste and to supply fresh nutrients. When the cell confluence reaches about 80-90%, subculturing is recommended to avoid overgrowth, commonly achieved by detaching the cells using trypsin and splitting them into fresh culture flasks at a suitable dilution ratio, typically around 1:10.

extract_protocol:
Genomic DNA was extracted by the addition of 20 µl of freshly prepared lysis buffer (10 mM Tris-HCl, pH 7.5; 0.05% SDS (ThermoFisher Scientific); 25 µg/ml proteinase K (Tiangen)) directly into each tube which 30,000 cells were sorted from each sample. The genomic DNA mixture was incubated at 37 °C for 1.5 h, followed by an 80 °C enzyme inactivation step for 30 min.
Use site-specific primers with second-generation sequencing adapters to amplify target sites via PCR for library construction.

treatment_protocol: HEK293T cells were seeded on 12-well poly-d-lysine coated plates (Corning). Between 16 and 24 h after seeding, cells were transfected at approximately 70% confluency with lipofectamine 3000 (ThermoFisher Scientific) according to the manufacturer’s protocols and 3249 ng All in one(PE2) plasmid, and 1083 ng sgRNA plasmid (for PE2 and PE3). Unless otherwise stated, cells were cultured for 2 days following transfection, after which the medium was removed, the cells were washed with 1 ml 1×PBS (ThermoFisher Scientific), then digested with 0.25% Trypsin-EDTA (1X) (Gibco) for 1 min, 1 ml DMEM complete medium was added to terminate the reaction. Centrifugation at 300 g for 5 min, discard supernatant and resuspended with 150 ul 1×PBS (ThermoFisher Scientific). After resuspended, all positive cells (single or double fluorescence positive) were sorted and collected into a 1.5 ml low adsorption centrifuge tube (ThermoFisher Scientific) by MULTI-APPLICATION CELL SORTER MA900 (Sony). Centrifugation at 11,200 g for 1 min, discard supernatant and store at -20°C for a long time.

tag: cell line

tag: treatment

value: base editing by prime editing


title: HEK293T cell, HEK4, G2T, loop2-22-33-1-M, PE2, rep1

description:
5854
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-13-20-7-M, PE2, rep3

description:
5857
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-18-1-M, PE2, rep1

description:
5851
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: RNA_seq_8hpi_rep1

city: Wako

first: Shintaro

last: Iwasaki

state: Saitama

address: 2-1, Hirosawa

department: Center for Pioneering Research

country: Japan

institute: RIKEN

library_source: transcriptomic

molecule: total RNA

organism: Acanthamoeba castellanii

source_name: amoeba cell

growth_protocol: A. castellanii, strain Neff [American Type Culture Collection (ATCC), 30010] was cultured with 10 ml peptone-yeast-glucose (PYG) medium in a 10cm dish at 28°C.

treatment_protocol: Cells were infected with APMV at MOI of 10 and harvested at 2, 4 and 8 hours after infection. For mock infection, PYG medium was used. Samples were treated with 100μg/mL CHX for 1 minute and scraped down and pelleted at 300 × g at 4°C for 3 min, washed with 1 ml of PBS containing 100 µg of CHX and 100 µg of chloramphenicol, and then pelleted again 300 × g at 4°C for 2 min.

tag: cell type

value: amoeba cell

tag: strain

value: Acanthamoeba castellanii strain Neff

value: Infected with Acanthamoeba polyphage mimivirus at MOI of 10

tag: time


title: RNA_seq_4hpi_rep2


title: RNA_seq_4hpi_rep1


title: HEK293T cell, HEK4, G2T, loop2-22-16-1-M, PE2, rep1

description:
5850
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-18-1-M, PE2, rep1

description:
5845
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: RNA_seq_mock_rep2

value: Mock infection

value: mock


title: HEK293T cell, HEK4, G2T, loop2-10-27-1-M, PE2, rep1

description:
5847
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-16-1-M, PE2, rep1

description:
5844
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: RNA_seq_2hpi_rep1


title: HEK293T cell, HEK4, G2T, loop2-10-12-1-M, PE2, rep1

description:
5842
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-14-1-M, PE2, rep1

description:
5843
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: RNA_seq_mock_rep1


title: HEK293T cell, EMX1, G5T, loop2-13-22-7-M, PE2, rep1

description:
5858
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scRNA-seq, adult, 5'

city: Barcelona

first: Arnau

last: Sebe-Pedros

address: Dr. Aiguader 88

department: Systems Biology

country: Spain

institute: Centre for Genomic Regulation

description: Library name: 26_Adult_5GEX

library_source: transcriptomic single cell

data_processing:
Demultiplexing was done using Illumina's bcl2fastq.
scATAC: processing, mapping and counting was done using scATAC-pro workflow (scATAC-pro_v1.4.1 doi.org/10.1186/s13059-020-02008-0)
Multiome: processing, mapping and counting was done using 10x CellRanger (cellranger-arc-2.0.1)
scRNA-seq: processing, mapping and counting was done using 10x CellRanger (cellranger-7.0.0)
Assembly: jaNemVect1.1
Supplementary files format and content: bed file: scATAC-seq peaks, scRNA-seq peaks
Supplementary files format and content: rds files: peaks and gene score matrices for metacells and cell types
Supplementary files format and content: bigwig files: scATAC-seq pseudobulk coverage

molecule: polyA RNA

organism: Nematostella vectensis

source_name: whole-adult

growth_protocol: Adult polyps were maintained at 18°C in 1/3 filtered seawater ( Nematostella medium, NM), and spawned by a temperature and light shock. Fertilized egg packages were treated with a 3% L-cysteine in NM solution to remove the egg jelly. Embryos were raised at 21°C until mid-gastrula stage (26 hours post fertilization, hpf) and collected based on their morphology.

extract_protocol:
scATACseq Gastrula samples: Nuclei from 300 pooled gastrula were isolated in 1x OmniATAC lysis buffer supplemented with Pitstop2 (10mM Tris-HCl pH 7.5, 10mM NaCl, 3mM MgCl2, 1%BSA, 0.1% NP-40, 0.1% Tween-20, 0.01% digitonin, 70uM Pitstop2), 3min while gently dounce homogenizing in cold conditions. Nuclei were then wash it in ATACwash buffer (10mM Tris-HCl pH 7.5, 10mM NaCl, 3mM MgCl2, 2%BSA, 0.1% Tween-20), twice in cold PBS-1%BSA, and finally gently resuspended in 1x Diluted buffer from 10x genomics (sample 1_Gastrula_Fresh). When nuclei from gastrula samples were fixed, nuclei were isolated in 1x OmniATAC lysis buffer without digitonin and mildly fixed in 0.1% methanol-free formaldehyde (PFA) prior to purification by FACS (FACS Influx, 100um nozzle, 12psi). Then, sorted nuclei were  permeabilized in 0.1x OmniATAC lysis buffer with Pitstop2 for 2min on ice while gently pipetting (sample 2_Gastrula_Fix). After washing, nuclei were processed as described above for fresh nuclei.
scATACseq Adult samples: 2-4 adult polyps were gently dounce homogenized in ice-cold TST lysis buffer (10mM Tris-HCl pH 7.5, 146mM NaCl, 1mM CaCl2, 21mM MgCl2, 0.03% Tween-20, 1x cOmplete protease inhibitor) until a homogenous nuclei suspension was achieved for a maximum of 12min. The resulting nuclei suspension was wash it in ST-2%BSA buffer (without tween-20) and mildly fixed in 0.1% PFA prior to FACSorting (FACS Influx, 100um nozzle, 12psi). FACS-purified nuclei were then permeabilized in 0.1x OmniATAC lysis buffer with Pitstop2 for 2min on ice while gently pipetting, wash it and resuspended in 1x diluted buffer from 10x genomics (samples 3-15_Adult_Fix; samples 17-19_Adult_Fix). When nuclei from adult samples were processed fresh (without PFA fixation), NP-40 was added to TST lysis buffer (0.01% NP-40 final concentration) after dounce homogenization and further dissociated 5min by pipetting. In this case, nuclei were purified from debris using an OptiPrep® continuous density gradient. Fresh purified nuclei were permeabilized in ice-cold 1x OmniATAC lysis buffer with Pitstop2 for 4min while gently pipetting, wash it and resuspended in 1x diluted buffer from 10x genomics (sample 16_Adult_Fresh).
scATACseq Elav+ neurons: One month old NvElav1::mOrange positive polyps were dissociated at 25°C in calcium- and magnesium-free NM (CMF/NM) containing 5mM EDTA and 0.25% α-chymotrypsin . Single cell suspensions were then stained with Hoechst 33342 (1µg/ml) and TO-PRO-3 (50nM) to remove debris and non-viable cells by FACS (FACS Aria II, 100um nozzle). Nuclei from sorted mOrange-positive cells were isolated, mildly fixed in 0.1%PFA and permeabilized (samples 20-21 Elav_fix), or directly permeabilized in OmniATAC lysis buffer with Pitstop2 (samples 22-24 Elav_fresh). Finally, permeabilized nuclei were washed in ATAC wash buffer, resuspended in 1x Diluted buffer and encapsulated using 10x Chromium platform.
scMultiome (ATAC+GEX): 2 adult wild-type polyps were dissociated and stained for FACSorting as described above for NvElav1::mOrange samples. Nuclei from single viable cells were isolated and permeabilized for 3min in ice-cold 0.1x OmniATAC lysis buffer supplemented with Pitstop (70mM), RNAse inhibitor (1U/ul) and DTT (1mM). Nuclei were then washed in ATAC wash buffer, resuspended in 1x Diluted Nuclei Buffer with RNAse inhibitor (1U/ul) and DTT (1mM), and encapsulated using 10x Chromium platform.
scRNAseq 5' GEX: A single cell suspension from 2-3 month old polyps were obtained after ACMEsorbitol (0.4M) fixation and dissociation as previously described (Garcia-Castro, H., et al., 2021; Najle,S.R and Grau-Bove,X., et al, 2023).
scATAC-seq libraries from whole-gastrula stage were prepared using the Chromium scATAC v2 (Next GEM) kit from 10x Genomics, following the manufacturer's instructions
scATAC-seq libraries from whole-adult and elav+ cells were prepared using the Chromium scATAC v1.1 (Next GEM) kit from 10x Genomics, following the manufacturer's instructions
scMultiome, libraries (ATAC+GEX) from whole-adult cells were prepared using the Chromium single cell multiome kit (Next GEM) from 10x Genomics, following the manufacturer's instructions. 8 PCR cycles were performed in Step 5.1 for ATAC library construction. 9 PCR cycles of cDNA amplification were performed in Step 6.1 for GEX library construction
scRNAseq 5' GEX library from whole-adult cells was prepared using the Chromium Next GEM 5’ GEX scRNAseq v2 kit from 10x Genomics, following the manufacturer's instructions and with 14 PCR cycles of cDNA amplification

tag: tissue

value: whole-adult


title: Multiome, adult, fresh, GEX

description: Library name: 25_Adult_Fresh_MultiomeGEX


title: Multiome, adult, fresh, ATAC

description: Library name: 25_Adult_Fresh_MultiomeATAC

library_source: genomic single cell


title: HEK293T cell, HEK4, G2T, loop2-22-33-3-M, PE2, rep1

description:
5841
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-12-3-M, PE2, rep1

description:
5835
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, Elav, fresh, 24

description: Library name: 24_Elav_Fresh

source_name: elav+ neurons

value: elav+ neurons


title: HEK293T cell, HEK4, G2T, loop2-22-21-1-M, PE2, rep1

description:
5852
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-18-3-M, PE2, rep1

description:
5838
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, Elav, fix, 20

description: Library name: 20_Elav_Fix


title: scATAC-seq, Elav, fresh, 23

description: Library name: 23_Elav_Fresh


title: HEK293T cell, HEK4, G2T, loop2-22-27-3-M, PE2, rep1

description:
5840
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, Elav, fresh, 22

description: Library name: 22_Elav_Fresh


title: HEK293T cell, HEK4, G2T, loop2-10-33-3-M, PE2, rep1

description:
5834
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: RNA_seq_2hpi_rep2


title: HEK293T cell, HEK4, G2T, loop2-22-14-3-M, PE2, rep1

description:
5836
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, Elav, fix, 21

description: Library name: 21_Elav_Fix


title: scATAC-seq, adult, fix, 19

description: Library name: 19_Adult_Fix


title: scATAC-seq, adult, fix, 15

description: Library name: 15_Adult_Fix


title: scATAC-seq, adult, fix, 18

description: Library name: 18_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-10-21-3-M, PE2, rep1

description:
5832
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, adult, fix, 17

description: Library name: 17_Adult_Fix


title: scATAC-seq, adult, fix, 14

description: Library name: 14_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-22-21-3-M, PE2, rep1

description:
5839
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-18-3-M, PE2, rep1

description:
5831
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, adult, fresh, 16

description: Library name: 16_Adult_Fresh


title: HEK293T cell, HEK4, G2T, loop2-10-16-3-M, PE2, rep1

description:
5830
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, adult, fix, 12

description: Library name: 12_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-22-27-5-M, PE2, rep1

description:
5826
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-33-5-M, PE2, rep1

description:
5827
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-33-5-M, PE2, rep1

description:
5821
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-18-5-M, PE2, rep1

description:
5824
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, adult, fix, 6

description: Library name: 6_Adult_Fix


title: scATAC-seq, adult, fix, 13

description: Library name: 13_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-10-27-3-M, PE2, rep1

description:
5833
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-14-3-M, PE2, rep1

description:
5829
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-14-5-M, PE2, rep1

description:
5816
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, adult, fix, 11

description: Library name: 11_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-22-21-5-M, PE2, rep1

description:
5825
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-12-3-M, PE2, rep1

description:
5828
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, gastrula, fresh

description: Library name: 1_Gastrula_Fresh

source_name: whole-gastrula

value: whole-gastrula


title: scATAC-seq, gastrula, fixed

description: Library name: 2_Gastrula_Fix


title: scATAC-seq, adult, fix, 10

description: Library name: 10_Adult_Fix


title: scATAC-seq, adult, fix, 3

description: Library name: 3_Adult_Fix


title: scATAC-seq, adult, fix, 9

description: Library name: 9_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-10-27-5-M, PE2, rep1

description:
5820
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-16-5-M, PE2, rep1

description:
5823
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-16-5-M, PE2, rep1

description:
5817
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, adult, fix, 8

description: Library name: 8_Adult_Fix


title: scATAC-seq, adult, fix, 7

description: Library name: 7_Adult_Fix


title: scATAC-seq, adult, fix, 5

description: Library name: 5_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-22-12-7-M, PE2, rep1

description:
5808
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-21-5-M, PE2, rep1

description:
5819
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-21-7-M, PE2, rep1

description:
5805
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: scATAC-seq, adult, fix, 4

description: Library name: 4_Adult_Fix


title: HEK293T cell, HEK4, G2T, loop2-10-12-5-M, PE2, rep1

description:
5815
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-33-7-M, PE2, rep1

description:
5814
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P22 Peanut Knockout 4

city: Saint Louis

first: Jade

middle: Olivia Enright

last: Hostetler

state: MO

address: 660 S Eucllid Ave

department: Ophthalmology

country: USA

institute: Washington University in St. Louis

description:
Library name: Sample 14
Column name in P22RNAseq.csv : KO_4

data_processing:
Reads trimmed for adaptor sequence/low-quality sequence using Trim Galore.
Trimmed reads were then aligned to the mm10 reference genome using STAR.
Gene-level quantification of reads was performed using HTSeq.
Differential gene expression analysis was performed on counts using EdgeR.
Assembly: mm10
Supplementary files format and content: csv files include TMM-normalized cpm values for each sample.

organism: Mus musculus

source_name: Retina

extract_protocol:
Retinas were dissected and flash frozen in Trizol reagent.RNA was harvested using chloroform and extraction performed using Zymo RNA Clean and Concentrator .
P0 RNAseq library preparation was performed with 500ng to 1ug of total RNA. Ribosomal RNA was removed by an RNase-H method using RiboErase kits (Kapa Biosystems). mRNA was then fragmented in reverse transcriptase buffer and heating to 94 degrees for 8 minutes. mRNA was reverse transcribed to yield cDNA using SuperScript III RT enzyme (Life Technologies, per manufacturer's instructions) and random hexamers. A second strand reaction was performed to yield ds-cDNA. cDNA was blunt ended, had an A base added to the 3' ends, and then had Illumina sequencing adapters ligated to the ends.  P22 RNAseq library contruction was performed following manufacturer protocol of Takara SMARTer Stranded Total RNA High Input using Illumina TruSeq stranded primers.

value: Retina

value: retinal cells

tag: genotype

value: Peanut -/-

tag: age


title: HEK293T cell, HEK4, G2T, loop2-22-27-7-M, PE2, rep1

description:
5813
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P22 Peanut Knockout 2

description:
Library name: Sample 12
Column name in P22RNAseq.csv : KO_2


title: HEK293T cell, HEK4, G2T, loop2-10-14-7-M, PE2, rep1

description:
5802
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-21-7-M, PE2, rep1

description:
5812
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P22 Peanut Heterozygote 2

description:
Library name: Sample 10
Column name in P22RNAseq.csv : Het_2

value: Peanut +/-


title: HEK293T cell, HEK4, G2T, loop2-22-18-7-M, PE2, rep1

description:
5811
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-16-7-M, PE2, rep1

description:
5810
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-22-14-7-M, PE2, rep1

description:
5809
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, DNMT1, C2A, epegRNA+loop2-23-31-7-M, PE2, rep1

description:
5765
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-33-7-M, PE2, rep1

description:
5807
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-27-7-M, PE2, rep1

description:
5806
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-18-7-M, PE2, rep1

description:
5804
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P0 Peanut Knockout 2

description:
Library name: Sample 6
Column name in P0RNAseq.csv : KO_2


title: P22 Peanut Knockout 3

description:
Library name: Sample 13
Column name in P22RNAseq.csv : KO_3


title: HEK293T cell, HEK4, G2T, loop2-10-16-7-M, PE2, rep1

description:
5803
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, FANCF, A7C, epegRNA+loop2-29-32-7-M, PE2, rep2

description:
5753
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P0 Peanut Heterozygote 4

description:
Library name: Sample 4
Column name in P0RNAseq.csv : Het_4


title: P22 Peanut Knockout 1

description:
Library name: Sample 11
Column name in P22RNAseq.csv : KO_1


title: P0 Peanut Heterozygote 2

description:
Library name: Sample 2
Column name in P0RNAseq.csv : Het_2


title: HEK293T cell, RUNX1, C1A, epegRNA+loop2-27-33-7-M, PE2, rep2

description:
5770
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, FANCF, A7C, loop2-29-32-7-M, PE2, rep2

description:
5752
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK4, G2T, loop2-10-12-7-M, PE2, rep1

description:
5801
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, epegRNA+loop2-34-31-7-M, PE2, rep2

description:
5748
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P22 Peanut Heterozygote 1

description:
Library name: Sample 9
Column name in P22RNAseq.csv : Het_1


title: P0 Peanut Knockout 4

description:
Library name: Sample 8
Column name in P0RNAseq.csv : KO_4


title: HEK293T cell, RUNX1, C1A, loop2-27-33-7-M, PE2, rep2

description:
5769
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P0 Peanut Knockout 3

description:
Library name: Sample 7
Column name in P0RNAseq.csv : KO_3


title: HEK293T cell, DNMT1, C2A, loop2-23-31-7-M, PE2, rep1

description:
5764
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P0 Peanut Knockout 1

description:
Library name: Sample 5
Column name in P0RNAseq.csv : KO_1


title: Replicate 2, GFP reporter C50 cell, GFP positive, dDAVP

city: Bethesda

first: Lihe

last: Chen

state: MD

address: 10 center drive MSC 1603, Building 10, Room 6N228

institute: NHLBI

description:
Library name: Sample22
C50_dDAVP GFP+2

data_processing:
RNA-seq raw sequencing reads were aligned by STAR to the mouse Ensembl genome (release 106). Transcript per million (TPM) and expected counts were generated by RSEM. Computational analyses were performed on the National Institutes of Health Biowulf High-Performance Computing platform
Assembly: Ensemble release 106
Supplementary files format and content: LC7188_GFP_TPM.txt

source_name: mpkCCD

growth_protocol: Mouse cortical collecting duct cells (mpkCCDc11 secondary clone 38, mpkCCDC11-38) were grown on transwells (Cat. No. 3460 and 3450, Corning) in complete DMEM/Ham’s F12 medium (DMEM/F12) containing 5 μg/mL insulin, 50 nM dexamethasone, 1 nM triiodothyronine, 10 ng/mL epidermal growth factor, 60 nM sodium selenite, 5 μg/mL transferrin, and 2% fetal bovine serum for 4 days. Then, the medium was changed to a serum-free/growth factor-free simple medium [with 50 nM dexamethasone, 60 nM sodium selenite, 5 μg/mL transferrin] to ensure complete polarization and cell differentiation. Cells were treated with vehicle or 0.1 nM or 1 nM dDAVP on the basolateral side for 24 or 48 hours in simple medium. To test Forskolin effect on the cells, 10 μM Forskolin were added on the both sides for 48 hours in simple medium.

extract_protocol:
RNA was extracted using Direct-zol RNA MicroPrep kit
Trizol-extracted RNA was proceeded to cDNA generation using the SMART-Seq HT Kit (Takara Bio). cDNA was generated using 1 ng of total RNA with 11 PCR cycles. 1 ng of the resulting cDNA was “tagmented” and barcoded by using the Nextera XT DNA Sample Preparation Kit (Illumina) to generate sequencing libraries for high throughput sequencing. Libraries were purified by AmPure XP magnetic beads (Beckman Coulter), and quantified using a Qubit 2.0 Fluorometer (Thermo Fisher Scientific). An equal amount of index libraries were pooled and sequenced (paired-end 50 bp) on an Illumina platform.

value: mpkCCD

value: dDAVP


title: HEK293T cell, RNF2, A4C, epegRNA+loop2-26-33-7-M, PE2, rep2

description:
5757
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, HEK3, G2C, epegRNA+loop2-22-31-7-M, PE2, rep1

description:
5761
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 1, GFP reporter C50 cell, GFP negative, dDAVP

description:
Library name: Sample19
C50_dDAVP GFP-1


title: HEK293T cell, VEGFA, A3C, tetraloop-34-43-7-M, PE2, rep1

description:
5746
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, RUNX1, C1A, loop2-27-33-7-M, PE2, rep1

description:
5717
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P0 Peanut Heterozygote 3

description:
Library name: Sample 3
Column name in P0RNAseq.csv : Het_3


title: Replicate 1, GFP reporter C43 cell, GFP positive, dDAVP

description:
Library name: Sample17
C43_dDAVP GFP+1


title: Replicate 1, GFP reporter C50 cell, GFP positive, dDAVP

description:
Library name: Sample21
C50_dDAVP GFP+1


title: HEK293T cell, RUNX1, C1A, epegRNA+loop2-27-33-7-M, PE2, rep1

description:
5718
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, tetraloop-34-31-7-M, PE2, rep1

description:
5744
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, GFP reporter C43 cell, GFP positive, dDAVP

description:
Library name: Sample18
C43_dDAVP GFP+2


title: HEK293T cell, HEK3, G2C, loop2-22-31-7-M, PE2, rep2

description:
5709
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, GFP reporter C50 cell, GFP negative, dDAVP

description:
Library name: Sample20
C50_dDAVP GFP-2


title: HEK293T cell, VEGFA, A3C, tetraloop-34-25-7-M, PE2, rep1

description:
5743
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: P0 Peanut Heterozygote 1

description:
Library name: Sample 1
Column name in P0RNAseq.csv : Het_1


title: HEK293T cell, RNF2, A4C, loop2-26-33-7-M, PE2, rep2

description:
5705
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, tetraloop-34-43-7-M, PE2, rep3

description:
5695
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 1, GFP reporter C43 cell, GFP negative, dDAVP

description:
Library name: Sample15
C43_dDAVP GFP-1


title: HEK293T cell, VEGFA, A3C, epegRNA+loop2-34-31-7-M, PE2, rep1

description:
5697
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, FANCF, A7C, epegRNA+loop2-29-32-7-M, PE2, rep1

description:
5702
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, GFP reporter C11 cell, GFP positive, dDAVP

description:
Library name: Sample14
C11_dDAVP GFP+2


title: HEK293T cell, VEGFA, A3C, epegRNA(3xPBS-RTT), PE2, rep2

description:
5698
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, GFP reporter C1 cell, GFP negative, dDAVP

description:
Library name: Sample8
C1_dDAVP GFP-2


title: Replicate 1, GFP reporter C11 cell, GFP positive, dDAVP

description:
Library name: Sample13
C11_dDAVP GFP+1


title: HEK293T cell, VEGFA, A3C, tetraloop-34-37-7-M, PE2, rep1

description:
5745
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, GFP reporter C11 cell, GFP negative, dDAVP

description:
Library name: Sample12
C11_dDAVP GFP-2


title: HEK293T cell, VEGFA, A3C, tetraloop-34-31-7-M, PE2, rep3

description:
5693
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 1, GFP reporter C11 cell, GFP negative, dDAVP

description:
Library name: Sample11
C11_dDAVP GFP-1


title: Replicate 1, GFP reporter C1 cell, GFP negative, dDAVP

description:
Library name: Sample7
C1_dDAVP GFP-1


title: HEK293T cell, VEGFA, A3C, loop2-34-31-1-M, PE2, rep1

description:
5690
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, tetraloop-34-25-7-M, PE2, rep3

description:
5692
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, WT-mpkCCD cell, dDAVP

description:
Library name: Sample5
WT_mpkCCD dDAVP_2


title: Replicate 1, WT-mpkCCD cell, dDAVP

description:
Library name: Sample4
WT_mpkCCD dDAVP_1


title: Replicate 3, WT-mpkCCD cell, no dDAVP

description:
Library name: Sample3
WT_mpkCCD Control_3

value: no treatment


title: HEK293T cell, VEGFA, A3C, loop2-34-16-1-M, PE2, rep1

description:
5685
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-34-20-1-M, PE2, rep1

description:
5687
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-22-1-M, PE2, rep1

description:
5681
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, tetraloop-34-37-7-M, PE2, rep3

description:
5694
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-34-37-1-M, PE2, rep1

description:
5691
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, GFP reporter C43 cell, GFP negative, dDAVP

description:
Library name: Sample16
C43_dDAVP GFP-2


title: HEK293T cell, VEGFA, A3C, loop2-34-22-1-M, PE2, rep1

description:
5688
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 3, WT-mpkCCD cell, dDAVP

description:
Library name: Sample6
WT_mpkCCD dDAVP_3


title: Replicate 2, WT-mpkCCD cell, no dDAVP

description:
Library name: Sample2
WT_mpkCCD Control_2


title: HEK293T cell, VEGFA, A3C, loop2-22-25-1-M, PE2, rep1

description:
5682
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Replicate 2, GFP reporter C1 cell, GFP positive, dDAVP

description:
Library name: Sample10
C1_dDAVP GFP+2


title: Replicate 1, WT-mpkCCD cell, no dDAVP

description:
Library name: Sample1
WT_mpkCCD Control_1


title: eWAT_metr_ce_5

city: Ithaca

first: Philip

last: Ruppert

state: NY

address: 244 Garden Avenue

department: Division of Nutritional Sciences

institute: Cornell University

description:
Library name: C017_eWAT_metr_ce
Mouse ID: JWK-1312

data_processing:
Sequencing reads were aligned to the mouse reference genome (Gencode vM25) (12) using STAR aligner (v2.7.9a) (13) with the following parameters: --outSAMunmapped Within, --outFilterType BySJout, --outSAMattributes NH HI AS NM MD, --outFilterMultimapNmax 10, --outFilterMismatchNoverReadLmax 0.04, --alignIntronMin 20, --alignIntronMax 1000000, --alignMatesGapMax 1000000, --alignSJoverhangMin 8, and --alignSJDBoverhangMin 1. Gene-level quantification was performed using featureCounts (v2.0.3) (14). In parallel, transcript-level quantification was performed using Salmon (v1.9.0) (15) and Gencode vM25, with parameters --seqBias, --useVBOpt, and --numBootstraps 30. Transcript abundance estimates were imported and summarized to the gene level using the tximport package (16). Quality control was summarized with MultiQC v1.19. All RNA-seq samples passed the QC.
Differential gene expression analysis was performed using DESeq2 (v1.46.0) (17) with shrinkage of fold-changes using apeglm (18). Genes with fewer than 10 total counts across all samples were excluded prior to normalization. The following interaction model was applied: ~ diet + temperature + diet:temperature, allowing detection of diet effects, temperature effects, and their statistical interaction. Specifically, the model tested: (i) the main effect of diet (MetR vs. Ctrl), (ii) the main effect of temperature (CE vs. RT), and (iii) the diet × temperature interaction (whether the effect of MetR differs between RT and CE). Genes with an adjusted p-value < 0.05 and an absolute log2FC ≥ 0.585 were considered significantly differentially expressed. Gene set enrichment analysis (GSEA) was performed using the gseKEGG() function from the clusterProfiler package (v4.14.6) (19) with parameters: organism = "mmu", pvalueCutoff = 1, pAdjustMethod = "BH", minGSSize = 0, seed = TRUE, and eps = 0. Quality control metrics and preprocessing approaches were done as previously described (20).
Assembly: mm10
Supplementary files format and content: Liver_Matrix_Raw_counts.txt, iBAT_iWAT_Matrix_Raw_counts.txt and gWAT_Matrix_Raw_counts.txt

source_name: eWAT

extract_protocol:
Total RNAs from tissues were isolated using the TRI reagent (Sigma) followed by clean-up with RPE buffer (Qiagen, Germany). The quality of RNA was validated by the Agilent RNA 6000 Nano-Kit in Agilent 2100 Bioanalyzer according to the manufacturer's protocol (Agilent Technologies, Waldbronn, Germany). mRNA sequencing was performed in-house.
After quality control 500 ng RNA in a final volume of 25 μL DEPC-treated water was prepared and sample preparation was performed as described in the NEBNext Poly(A) mRNA Magnetic Isolation Module kit and NEBNext Ultra II RNA Library Prep Kit (cat no: #E7770, New England Biolabs, Ipswich, MA, USA). The amplified libraries were validated by Agilent 2100 Bioanalyzer using a DNA 1000 kit (Agilent Technologies, Inc., Santa Clara, CA, USA) and quantified by qPCR using the KaPa Library Kits (KaPa Biosystems, Wilmington, MA, USA). Hereafter, 2x50bp paired-end sequencing was performed on Illumina Novaseq 6000.

treatment_protocol: 2×2 design: Control vs. MetR for 7 days at room temperature (RT: 22 °C) +/- cold exposure (CE: 4 °C) for 24 h.

value: eWAT

value: WT


title: Replicate 1, GFP reporter C1 cell, GFP positive, dDAVP

description:
Library name: Sample9
C1_dDAVP GFP+1


title: eWAT_metr_ce_3

description:
Library name: C015_eWAT_metr_ce
Mouse ID: JWK-1330


title: HEK293T cell, VEGFA, A3C, loop2-34-16-3-M, PE2, rep1

description:
5671
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1

value: base editing by prime editing


title: HEK293T cell, VEGFA, A3C, loop2-34-22-3-M, PE2, rep1

description:
5674
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_ce_1

description:
Library name: C013_eWAT_metr_ce
Mouse ID: JWK-1305


title: HEK293T cell, VEGFA, A3C, loop2-22-20-3-M, PE2, rep1

description:
5666
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_ce_7

description:
Library name: C019_eWAT_metr_ce
Mouse ID: JWK-1303


title: HEK293T cell, VEGFA, A3C, loop2-22-20-1-M, PE2, rep1

description:
5680
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-34-37-5-M, PE2, rep1

description:
5663
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_ce_6

description:
Library name: C018_eWAT_metr_ce
Mouse ID: JWK-1313


title: eWAT_ctrl_ce_3

description:
Library name: C008_eWAT_ctrl_ce
Mouse ID: JWK-1316


title: HEK293T cell, VEGFA, A3C, loop2-22-18-1-M, PE2, rep1

description:
5679
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-16-1-M, PE2, rep1

description:
5678
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_ce_4

description:
Library name: C016_eWAT_metr_ce
Mouse ID: JWK-1311


title: HEK293T cell, VEGFA, A3C, loop2-34-25-1-M, PE2, rep1

description:
5689
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-25-3-M, PE2, rep1

description:
5668
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_rt_7

description:
Library name: C005_eWAT_metr_rt
Mouse ID: JWK-1288


title: eWAT_metr_rt_5

description:
Library name: C003_eWAT_metr_rt
Mouse ID: JWK-1286


title: HEK293T cell, VEGFA, A3C, loop2-22-22-3-M, PE2, rep1

description:
5667
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_ce_7

description:
Library name: C012_eWAT_ctrl_ce
Mouse ID: JWK-1329


title: HEK293T cell, VEGFA, A3C, loop2-22-18-3-M, PE2, rep1

description:
5665
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_ce_6

description:
Library name: C011_eWAT_ctrl_ce
Mouse ID: JWK-1328


title: HEK293T cell, VEGFA, A3C, loop2-34-22-5-M, PE2, rep1

description:
5661
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_ce_1

description:
Library name: C006_eWAT_ctrl_ce
Mouse ID: JWK-1314


title: eWAT_ctrl_ce_4

description:
Library name: C009_eWAT_ctrl_ce
Mouse ID: JWK-1317


title: HEK293T cell, VEGFA, A3C, loop2-34-25-5-M, PE2, rep1

description:
5662
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_ce_2

description:
Library name: C007_eWAT_ctrl_ce
Mouse ID: JWK-1315


title: HEK293T cell, VEGFA, A3C, loop2-22-20-5-M, PE2, rep1

description:
5654
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-34-18-5-M, PE2, rep1

description:
5659
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_ce_2

description:
Library name: C014_eWAT_metr_ce
Mouse ID: JWK-1285


title: eWAT_metr_rt_1

description:
Library name: B999_eWAT_metr_rt
Mouse ID: JWK-1266


title: HEK293T cell, VEGFA, A3C, loop2-34-16-5-M, PE2, rep1

description:
5658
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_rt_7

description:
Library name: B998_eWAT_ctrl_rt
Mouse ID: JWK-1310


title: HEK293T cell, VEGFA, A3C, loop2-34-37-7-M, PE2, rep1

description:
5651
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-37-5-M, PE2, rep1

description:
5657
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_rt_6

description:
Library name: C004_eWAT_metr_rt
Mouse ID: JWK-1287


title: HEK293T cell, VEGFA, A3C, loop2-22-31-5-M, PE2, rep1

description:
5656
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-34-31-7-M, PE2, rep1

description:
5650
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_metr_rt_4

description:
Library name: C002_eWAT_metr_rt
Mouse ID: JWK-1326


title: eWAT_ctrl_ce_5

description:
Library name: C010_eWAT_ctrl_ce
Mouse ID: JWK-1327


title: eWAT_metr_rt_3

description:
Library name: C001_eWAT_metr_rt
Mouse ID: JWK-1302


title: eWAT_metr_rt_2

description:
Library name: B1000_eWAT_metr_rt
Mouse ID: JWK-1278


title: HEK293T cell, VEGFA, A3C, loop2-34-25-7-M, PE2, rep1

description:
5649
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-18-5-M, PE2, rep1

description:
5653
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-34-22-7-M, PE2, rep1

description:
5648
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-22-5-M, PE2, rep1

description:
5655
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_rt_3

description:
Library name: B994_eWAT_ctrl_rt
Mouse ID: JWK-1300


title: HEK293T cell, VEGFA, A3C, loop2-34-20-5-M, PE2, rep1

description:
5660
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_metr_ce_4

description:
Library name: B295_iWAT_metr_ce
Mouse ID: JWK-1312

source_name: iWAT

value: iWAT


title: HEK293T cell, VEGFA, A3C, loop2-22-16-5-M, PE2, rep1

description:
5652
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-18-7-M, PE2, rep1

description:
5640
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_metr_ce_3

description:
Library name: B294_iWAT_metr_ce
Mouse ID: JWK-1311


title: eWAT_ctrl_rt_5

description:
Library name: B996_eWAT_ctrl_rt
Mouse ID: JWK-1277


title: eWAT_ctrl_rt_6

description:
Library name: B997_eWAT_ctrl_rt
Mouse ID: JWK-1306


title: iWAT_metr_ce_1

description:
Library name: B291_iWAT_metr_ce
Mouse ID: JWK-1305


title: HEK293T cell, RNF2, A4C, loop2-26-33-7-M, PE2, rep1

description:
5628
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_ctrl_ce_4

description:
Library name: B287_iWAT_ctrl_ce
Mouse ID: JWK-1317


title: eWAT_ctrl_rt_4

description:
Library name: B995_eWAT_ctrl_rt
Mouse ID: JWK-1301


title: HEK293T cell, VEGFA, A3C, loop2-34-18-7-M, PE2, rep1

description:
5647
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_rt_2

description:
Library name: B993_eWAT_ctrl_rt
Mouse ID: JWK-1299


title: HEK293T cell, EMX1, G5T, loop2-25-27-1-M, PE2, rep3

description:
5613
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_ctrl_ce_1

description:
Library name: B284_iWAT_ctrl_ce
Mouse ID: JWK-1314


title: HEK293T cell, EMX1, G5T, loop2-13-39-1-M, PE2, rep3

description:
5608
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-13-27-1-M, PE2, rep3

description:
5606
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-34-16-7-M, PE2, rep1

description:
5646
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: eWAT_ctrl_rt_1

description:
Library name: B992_eWAT_ctrl_rt
Mouse ID: JWK-1298


title: HEK293T cell, VEGFA, A3C, loop2-22-31-7-M, PE2, rep1

description:
5644
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, epegRNA(3xPBS-RTT), PE2, rep1

description:
5621
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_metr_ce_6

description:
Library name: B297_iWAT_metr_ce
Mouse ID: JWK-1303


title: iWAT_metr_rt_6

description:
Library name: B282_iWAT_metr_rt
Mouse ID: JWK-1287


title: HEK293T cell, EMX1, G5T, loop2-13-24-1-M, PE2, rep3

description:
5605
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-25-7-M, PE2, rep1

description:
5643
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_metr_rt_4

description:
Library name: B280_iWAT_metr_rt
Mouse ID: JWK-1326


title: iWAT_metr_ce_5

description:
Library name: B296_iWAT_metr_ce
Mouse ID: JWK-1313


title: HEK293T cell, EMX1, G5T, loop2-25-39-3-M, PE2, rep1

description:
5601
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_metr_rt_3

description:
Library name: B279_iWAT_metr_rt
Mouse ID: JWK-1302


title: HEK293T cell, VEGFA, A3C, loop2-22-22-7-M, PE2, rep1

description:
5642
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, loop2-22-20-7-M, PE2, rep1

description:
5641
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_metr_ce_2

description:
Library name: B292_iWAT_metr_ce
Mouse ID: JWK-1285


title: HEK293T cell, HEK3, G2C, loop2-22-31-7-M, PE2, rep1

description:
5632
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-25-20-3-M, PE2, rep2

description:
5597
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-13-39-3-M, PE2, rep3

description:
5595
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_ctrl_rt_7

description:
Library name: B276_iWAT_ctrl_rt
Mouse ID: JWK-1310


title: HEK293T cell, RNF2, A4C, epegRNA+loop2-26-33-7-M, PE2, rep1

description:
5629
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-13-22-1-M, PE2, rep3

description:
5604
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_ctrl_ce_7

description:
Library name: B290_iWAT_ctrl_ce
Mouse ID: JWK-1329


title: iWAT_ctrl_rt_5

description:
Library name: B274_iWAT_ctrl_rt
Mouse ID: JWK-1277


title: iWAT_ctrl_ce_6

description:
Library name: B289_iWAT_ctrl_ce
Mouse ID: JWK-1328


title: iWAT_ctrl_ce_5

description:
Library name: B288_iWAT_ctrl_ce
Mouse ID: JWK-1327


title: iWAT_ctrl_rt_4

description:
Library name: B273_iWAT_ctrl_rt
Mouse ID: JWK-1301


title: HEK293T cell, EMX1, G5T, loop2-25-39-1-M, PE2, rep1

description:
5615
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-25-33-5-M, PE2, rep3

description:
5588
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_ctrl_ce_3

description:
Library name: B286_iWAT_ctrl_ce
Mouse ID: JWK-1316


title: iWAT_ctrl_ce_2

description:
Library name: B285_iWAT_ctrl_ce
Mouse ID: JWK-1315


title: HEK293T cell, EMX1, G5T, loop2-25-24-5-M, PE2, rep3

description:
5586
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-25-18-1-M, PE2, rep4

description:
5609
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_metr_rt_7

description:
Library name: B283_iWAT_metr_rt
Mouse ID: JWK-1288


title: HEK293T cell, FANCF, A7C, loop2-29-32-7-M, PE2, rep1

description:
5624
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_metr_ce_5

description:
Library name: B267_iBAT_metr_ce
Mouse ID: JWK-1312

source_name: iBAT

value: iBAT


title: HEK293T cell, EMX1, G5T, loop2-13-20-5-M, PE2, rep3

description:
5580
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_metr_ce_3

description:
Library name: B265_iBAT_metr_ce
Mouse ID: JWK-1330


title: iWAT_metr_rt_2

description:
Library name: B278_iWAT_metr_rt
Mouse ID: JWK-1278


title: HEK293T cell, EMX1, G5T, loop2-25-33-3-M, PE2, rep3

description:
5600
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-25-27-7-M, PE2, rep3

description:
5576
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_metr_ce_1

description:
Library name: B263_iBAT_metr_ce
Mouse ID: JWK-1305


title: iWAT_metr_rt_1

description:
Library name: B277_iWAT_metr_rt
Mouse ID: JWK-1266


title: HEK293T cell, EMX1, G5T, loop2-25-24-3-M, PE2, rep3

description:
5598
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_ctrl_ce_7

description:
Library name: B262_iBAT_ctrl_ce
Mouse ID: JWK-1329


title: iWAT_metr_rt_5

description:
Library name: B281_iWAT_metr_rt
Mouse ID: JWK-1286


title: iWAT_ctrl_rt_6

description:
Library name: B275_iWAT_ctrl_rt
Mouse ID: JWK-1306


title: HEK293T cell, EMX1, G5T, loop2-13-33-3-M, PE2, rep3

description:
5594
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, DNMT1, C2A, epegRNA, PE2, rep3

description:
5525
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-13-24-3-M, PE2, rep3

description:
5592
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iWAT_ctrl_rt_3

description:
Library name: B272_iWAT_ctrl_rt
Mouse ID: JWK-1300


title: HEK293T cell, EMX1, G5T, loop2-13-27-3-M, PE2, rep3

description:
5593
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-25-39-5-M, PE2, rep1

description:
5589
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, DNMT1, C2A, epegRNA, PE2, rep1

description:
5523
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_ctrl_ce_2

description:
Library name: B257_iBAT_ctrl_ce
Mouse ID: JWK-1315


title: iWAT_ctrl_rt_2

description:
Library name: B271_iWAT_ctrl_rt
Mouse ID: JWK-1299


title: iWAT_ctrl_rt_1

description:
Library name: B270_iWAT_ctrl_rt
Mouse ID: JWK-1298


title: iBAT_metr_ce_7

description:
Library name: B269_iBAT_metr_ce
Mouse ID: JWK-1303


title: iBAT_ctrl_ce_1

description:
Library name: B256_iBAT_ctrl_ce
Mouse ID: JWK-1314


title: HEK293T cell, EMX1, G5T, loop2-25-20-5-M, PE2, rep2

description:
5584
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_ctrl_ce_3

description:
Library name: B258_iBAT_ctrl_ce
Mouse ID: JWK-1316


title: HEK293T cell, EMX1, G5T, epegRNA(3xPBS-RTT), PE2, rep2

description:
5521
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: ΔUhpTU + pBAD-Ctr-Sal R2

city: Shanghai

first: Chuan

last: Wang

address: 131 # Dong An Road

institute: Fudan University

description:
Library name: deltaUhpTU + pBAD-Ctr-Sal R2
The uhpTU-deleted Salmonella enterica strain carrying the empty pBAD plasmid was cultured in M9CA medium supplemented with 0.1% (w/v) glucose-6-phosphate (G6P) at 37°C until reaching an OD₆₀₀ of approximately 0.5. L-arabinose was then added to a final concentration of 0.2% (w/v) to induce expression. Cultures were incubated for an additional 10 minutes before total RNA was extracted. This sample represents biological replicate 2 of the control group.

data_processing:
RNA-seq was performed by BGI Group, Shenzhen, Guangdong, China. Briefly, a strand-specific cDNA library was constructed for each sample to keep the direction of RNA transcription. The obtained cDNA libraries were sequenced on an Illumina HiSeq 4000 machine.
SOAP (v2.21, -m 0 -x 1000 -s 28 -l 32 -v 5 -r 1 -p 3).The rest reads with adaptor，or with unknown bases more than 5%  ,or with more than 20% low-quality bases were cleared by a software developed by BGI group. After, clear reads were mapped to reference genome with Bowtie2： v2.2.5 -q --phred64 --sensitive --dpad 0 --gbar 99999999 --mp 1,1 --np 1 --score-min L,0,-0.1 -I 1 -X 1000 --), and expression value was caculated  (http://deweylab.biostat.wisc.edu/RSEM）
Assembly: Salmonella Typhimurium LT2 Genome  (NC_003197) Chromosome (https://www.ncbi.nlm.nih.gov/nuccore/NC_003197.2) and plasmid (https://www.ncbi.nlm.nih.gov/nuccore/NC_003277.2) , E.coli K-12 MG1655 genome (NC_000913.3) chromosome, (https://www.ncbi.nlm.nih.gov/nuccore/556503834)
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

organism: Salmonella enterica subsp. enterica

source_name: Typhimurium str. SL1344

growth_protocol: M9CA broth plus 0.1% glucose-6-phosphate (G6P) or LB miller medium

extract_protocol:
Bacterial total RNA was isolated using the hot phenol method. Briefly, Cells were resuspended in lysozyme, 10% SDS and 3M sodium acetate (pH5.2). The cleared lysate was mixed with saturated phenol (pH4.5) and incubated at 64°C for 6 min with shaking. After mixing with chloroform and centrifugation in a Phase Lock Gel tube (#WM5-2302820, TIANGEN Biotech), the aqueous phase was collected and mixed the with 30:1 ethanol: sodium acetate (pH 6.5) and precipitated at -80°C overnight. RNA pellets were washed with 80% ethanol and dissolved in ultra-pure water (#10977, Thermo Fisher Scientific), and the RNA concentration was determined using NanoDrop 2000.
Ribosomal RNAs was removed by Biotin-labeled rRNA probe. RNA was fragmented with BGI fragmentation buffer and was reverse transcribed with random hexamer. Double chain cDNA was then blunted and phosphorylated for the 5’ end, and overhang A base in the 3’ end and ligated with T-overhang adapter. After PCR application, samples were sequenced in BGISeq-500 platform

treatment_protocol: UhpU expression was induced with 0.2% (w/v) L-arabinose for 10 minutes

value: Typhimurium str. SL1344

value: {delta}UhpTU + pBAD-Ctr-Sal


title: wtOSC_ONT_directRNAseq

city: Vienna

first: Dominik

last: Handler

address: Dr. Bohr-Gasse 3

country: Austria

institute: IMBA-Institute of Molecular Biotechnology of Austrian Academy of Sciences

description:
Library name: wtOSC_ONT_directRNAseq
Nanopore direct-RNA sequencing

data_processing:
Direct RNA multi-read FAST5 files were basecalled on GPUs with ONT Guppy v5.0.7+2332e8d65 using the appropriate direct RNA configuration for the flowcell/kit (Oxford Nanopore Technologies). Basecalling emitted all reads (--disable_qscore_filtering) while preserving raw signal and disabling basecaller trimming where applicable (--fast5_out --trim_strategy none). Post-basecalling, per-batch FASTQ files were concatenated and processed to annotate read bodies upstream of the poly(A) tail and to estimate poly(A) lengths. Basecalled FAST5 files were converted to gzip-compressed FAST5 (compress_fast5, ONT) and poly(A) tails were identified with tailfindr in R, yielding per-read tail metrics (tail_end_base_index, tail_length)(Krause, Niazi et al. 2019). For each read, sequence and quality strings were truncated at tail_end_base_index to retain the transcribed body upstream of the poly(A) tract, and metadata were appended to read headers (LENGTH=, avgQUAL=, polyAlength=) with seqkit(Shen, Sipos et al. 2024) and mawk. Reads without a detectable tail were flagged (polyAlength=notDetected) and listed for QC. For 3′ UTR annotation, only direct RNA reads with a detected poly(A) tail were used.
Assembly: OSC_r1.01
Supplementary files format and content: All mapping and unique mapping bigwig files for sense and antisense

organism: Drosophila melanogaster

source_name: OSC

growth_protocol: Ovarian somatic cells (OSCs) were maintained at 27 °C in a humidified incubator with 2.5% CO₂, as previously described (Niki, Yamaguchi et al. 2006, Saito, Inagaki et al. 2009) Cells were cultured in M3 basal medium supplemented with 10% fetal bovine serum (FBS), insulin, glutathione, and fly extract.

extract_protocol:
Total RNA was isolated from 90% confluent T75 flasks of OSC cells. Cells were washed three times with PBS, lysed with 10 ml of Trizol, and incubated on a shaker for 30–60 minutes. The lysate was transferred to a 15 ml Falcon tube containing phase lock gel, mixed with 2 ml of chloroform by careful inversion, and centrifuged at 4800 g for 10 minutes at 20°C using a swing-out rotor. The aqueous phase was transferred to a fresh Falcon tube, 7 ml of isopropanol was added, and the mixture was incubated at room temperature for 15 minutes. RNA was then pelleted by centrifugation at 20,000 g for 1 hour at 4°C in a fixed-angle rotor. The supernatant was removed, and the pellet was washed with 75% ethanol. After complete removal of ethanol, the RNA pellet was resuspended in 500 µL of nuclease-free water without pipetting for mixing, and carefully transferred to an Eppendorf tube. RNA concentration was measured using Qubit.
mRNA was selected using oligo(dT) Dynabeads. A 200 µL aliquot of Dynabead slurry was washed twice with 2x binding buffer (10 mM Tris HCl pH 7.5, 1.0 M LiCl, and 2 mM EDTA) and resuspended in 100 µL of 2x binding buffer. Total RNA was heat-denatured at 65°C for 5 minutes and snap-cooled on ice. Washed beads were added to the heat-denatured RNA, and the mixture was incubated with overhead rotation for 15 minutes. After incubation, the supernatant was removed, and the beads were washed three times with 500 µL of wash buffer (10 mM Tris HCl pH 7.5, 0.15 M LiCl, and 1 mM EDTA), resuspending the beads each time. mRNA was eluted into 12 µL of nuclease-free water by incubation @ 80°C for 1 minute. mRNA concentration was measured using Qubit. Nanopore direct RNA sequencing libraries were prepared from 10 µL of eluted mRNA following the RNA002 protocol (Oxford Nanopore Technologies). Half of the prepared library was used on a fresh flow cell for sequencing.

value: OSC

value: ovarian somatic cells

value: wildtype


title: iBAT_metr_rt_7

description:
Library name: B255_iBAT_metr_rt
Mouse ID: JWK-1288


title: iBAT_metr_ce_6

description:
Library name: B268_iBAT_metr_ce
Mouse ID: JWK-1313


title: GSC23 cells, BRD2_BD1/2_1

city: San Diego

first: raghavendra

last: vadla

state: CA

address: sanford consortium for regenerative medicine, 2880 Torrey Pines Scenic Drive

institute: UCSD

description:
Library name: Sample 49
raw_counts_1.txt
GSC23_BRD2KO+Y113F-Y386F_1

data_processing:
Fastq file quality was ensured with FastQC, and reads were aligned to the hg38 index using STAR. Alignment quality was confirmed by assessing the final STAR logs. Index and bigwig files were generated from the BAM files with samtools and bamCoverage, respectively. Each BAM file was viewed in IGV to assess the quality and features of mapped reads. The Subread package was used to generate count files
Assembly: GRCh38 (GENCODE v39)
Supplementary files format and content: tab-delimited text includes raw counts for each sample

source_name: Brain

extract_protocol:
RNA was extracted using Rneasy mini kit (Qiagen)
Novogene sequencing

value: Brain

value: Glioma stem cells


title: HEK293T cell, EMX1, G5T, loop2-13-39-5-M, PE2, rep3

description:
5583
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, epegRNA(3xPBS-RTT), PE2, rep1

description:
5520
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_metr_ce_4

description:
Library name: B266_iBAT_metr_ce
Mouse ID: JWK-1311


title: HEK293T cell, EMX1, G5T, loop2-25-39-7-M, PE2, rep1

description:
5578
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: GSC23 cells, BRD2_BD1/2_2

description:
Library name: Sample 50
raw_counts_1.txt
GSC23_BRD2KO+Y113F-Y386F_2


title: HEK293T cell, EMX1, G5T, loop2-25-33-7-M, PE2, rep3

description:
5577
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_metr_rt_4

description:
Library name: B252_iBAT_metr_rt
Mouse ID: JWK-1326


title: GSC23 cells, BRD2_WT_3

description:
Library name: Sample 42
raw_counts_1.txt
GSC23_BRD2KO+WT_3


title: iBAT_metr_ce_2

description:
Library name: B264_iBAT_metr_ce
Mouse ID: JWK-1285


title: HEK293T cell, RNF2, A4C, epegRNA, PE2, rep3

description:
5498
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-13-27-7-M, PE2, rep3

description:
5570
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-25-18-7-M, PE2, rep2

description:
5572
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: GSC23 cells, BRD2_BD2_2

description:
Library name: Sample 47
raw_counts_1.txt
GSC23_BRD2KO+Y386F_2


title: HEK293T cell, EMX1, G5T, loop2-13-22-7-M, PE2, rep3

description:
5569
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_ctrl_ce_6

description:
Library name: B261_iBAT_ctrl_ce
Mouse ID: JWK-1328


title: iBAT_ctrl_ce_5

description:
Library name: B260_iBAT_ctrl_ce
Mouse ID: JWK-1327


title: HEK293T cell, DNMT1, C2A, epegRNA, PE2, rep2

description:
5524
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: ΔUhpTU + pBAD-UhpU-Sal R1

description:
Library name: deltaUhpTU + pBAD-UhpU-Sal R1
The uhpTU-deleted Salmonella enterica strain carrying the pBAD-UhpU plasmid was cultured in M9CA medium supplemented with 0.1% (w/v) glucose-6-phosphate (G6P) at 37°C until reaching an OD₆₀₀ of approximately 0.5. L-arabinose was then added to a final concentration of 0.2% (w/v) to induce UhpU expression. After 10 minutes of induction, total RNA was extracted. This sample represents biological replicate 1 of the UhpU pulse-expression group.

value: {delta}UhpTU + pBAD-UhpU-Sal


title: ΔUhpTU + pBAD-UhpU-Sal R2

description:
Library name: deltaUhpTU + pBAD-UhpU-Sal R2
The uhpTU-deleted Salmonella enterica strain carrying the pBAD-UhpU plasmid was cultured in M9CA medium supplemented with 0.1% (w/v) glucose-6-phosphate (G6P) at 37°C until reaching an OD₆₀₀ of approximately 0.5. L-arabinose was then added to a final concentration of 0.2% (w/v) to induce UhpU expression. After 10 minutes of induction, total RNA was extracted. This sample represents biological replicate 2 of the UhpU pulse-expression group.


title: ΔUhpTU + pBAD-Ctr-Sal R1

description:
Library name: deltaUhpTU + pBAD-Ctr-Sal R1
The uhpTU-deleted Salmonella enterica strain carrying the empty pBAD plasmid was cultured in M9CA medium supplemented with 0.1% (w/v) glucose-6-phosphate (G6P) at 37°C until reaching an OD₆₀₀ of approximately 0.5. L-arabinose was then added to a final concentration of 0.2% (w/v) to induce expression. Cultures were incubated for an additional 10 minutes before total RNA was extracted. This sample represents biological replicate 1 of the control group.


title: HEK293T cell, EMX1, G5T, epegRNA(3xPBS-RTT), PE2, rep3

description:
5522
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: GSC23 cells, BRD2_BD1/2_3

description:
Library name: Sample 51
raw_counts_1.txt
GSC23_BRD2KO+Y113F-Y386F_3


title: iBAT_ctrl_ce_4

description:
Library name: B259_iBAT_ctrl_ce
Mouse ID: JWK-1317


title: GSC23 cells, GSK620_2

description:
Library name: Sample 38
raw_counts_2.txt
GSC23_GSK620_2


title: GSC23 cells, BRD2_BD2_3

description:
Library name: Sample 48
raw_counts_1.txt
GSC23_BRD2KO+Y386F_3


title: GSC23 cells, GSK620_1

description:
Library name: Sample 37
raw_counts_2.txt
GSC23_GSK620_1


title: GSC23 cells, BRD2_BD2_1

description:
Library name: Sample 46
raw_counts_1.txt
GSC23_BRD2KO+Y386F_1


title: wtOSC_HiC

description: Library name: wtOSC_HiC

data_processing:
Illumina reads were processed by removing 3′ adapters with cutadapt (Martin 2011) and filtering for minimal length (>18 nt) and sequence complexity (bbduk, entropy = 0.35, entropy window = 18, k = 4)(Bushnell 2014).
Hi-C read-pairs were mapped to the OSC_r1.01 genome assembly using BWA-MEM (Li 2013) with the -P option for Hi-C paired-end data. Alignments were processed using samtools (Danecek, Bonfield et al. 2021) and parsed with pairtools (v.1.1.3)(Abdennur, Fudenberg et al. 2023). Duplicate fragments were filtered with pairtools dedup to generate sets of valid pairs, duplicates, unmapped reads, and summary statistics. For chromosome-scale contact map construction, valid Hi-C pairs were converted into a balanced matrix format using Cooler (v.0.10.4)(Abdennur and Mirny 2020). Bin intervals were generated at 1 kb resolution with cooler makebins. Matrices were built with cooler cload pairs and balanced using cooler balance with a maximum median absolute deviation (MAD) threshold of 5. To enable multi-resolution analysis of genome architecture, balanced matrices were zoomified with cooler zoomify at 1 kb, 2 kb, 5 kb, 10 kb, 25 kb, 50 kb, and 100 kb resolutions. The resulting .mcool files were used for downstream visualization and quantitative analysis of Hi-C contact patterns. Hi-C contact matrix was visualized using HiGlass(Kerpedjiev, Abdennur et al. 2018).
Assembly: OSC_r1.01
Supplementary files format and content: Multiresolution Hi-C contact matrices in .mcool (HDF5) format containing balanced and raw contact counts at the following genomic resolutions: 1 kb, 2 kb, 5 kb, 10 kb, 25 kb, 50 kb, and 100 kb.

extract_protocol:
Two OSC aliquots (5 × 10⁶ cells each; 1 × 10⁷ total) were processed in parallel. Cells were resuspended in 1 mL ice-cold lysis buffer, incubated for 30 min at 4 °C with rotation, and pelleted (2,500 g, 5 min, 4 °C). Pelleted Nuclei were resuspended in 1 mL lysis buffer, pelleted again, and finally resuspended in 400 µL 1.2× NEB buffer 3.1, minimizing additional wash steps to reduce clumping.
Nuclei were digested overnight at 37 °C with rotation (800 rpm) using 30 U DpnII (NEB) in a final volume of 80 µL. After digestion, pellets were resuspended in 52 µL fill-in mix containing 2.2 µL biotin-14-dATP (1 mM, Jena Bioscience), 1.1 µL dNTP mix (2 mM dCTP, dGTP, dTTP each), 11 µL Klenow fragment, 5.72 µL NEB buffer 2, and nuclease-free water to volume, and incubated for 1.5 h at 37 °C with rotation. Filled-in nuclei were resuspended in a 250 µL ligation mix (27.5 µL Thermo T4 DNA ligase buffer, 2.2 µL 10% Triton X-100, 2.25 µL BSA [10 mg/mL], 232.5 µL H₂O, and 11 µL T4 DNA ligase). Samples were incubated at RT for 5 h with rotation. Aliquots (10 µL) were collected at digestion and ligation steps as process controls. Post-ligation, reactions were diluted to 500 µL with PBS, supplemented with NaCl (final concentration TBD) and 10 µL proteinase K (20 mg/mL), and incubated overnight at 65 °C. DNA was purified by two rounds of phenol–chloroform extraction, followed by a chloroform wash, and ethanol precipitation in the presence of glycogen. Pellets were resuspended in 150 µL H₂O (controls in 10 µL), treated with RNase A (2 µL; 10 U/µL, 30 min, 37 °C), and quantified using Qubit. 130 microliters of DNA were transferred to Covaris microTUBEs and sheared on a Covaris E220 (<10 °C, intensifier inserted, 105 W peak power, 5% duty factor, 200 cycles/burst, 55 s). DNA was rinsed from Covaris tubes with 80 µL EB and subjected to two-step Ampure XP bead size selection (0.44× and 0.18×). DNA was eluted in 100 µL H₂O. Biotinylated DNA was captured with Dynabeads MyOne Streptavidin C1, pre-washed in Tween wash buffer (5 mM Tris-HCl pH 7.5, 0.5 mM EDTA, 1 M NaCl, 0.05% Tween-20). Beads were resuspended in 100 µL 2× binding buffer (10 mM Tris-HCl pH 7.5, 1 mM EDTA, 2 M NaCl), mixed with DNA, and incubated for 1.5 h at RT with rotation. Beads were washed twice in 500 µL Tween wash buffer and twice in H₂O, and resuspended in 50 µL H₂O. Bead-bound DNA was subjected to end repair with NEBNext Ultra II End Prep (7 µL End Prep buffer, 3 µL enzyme mix; 30 min at 20 °C then 30 min at 65 °C), adaptor ligation (2.5 µL NEB Illumina adaptor, 1 µL ligation enhancer, 30 µL ligation master mix; 15 min at 20 °C), and USER enzyme treatment (3 µL; 15 min, 37 °C). Beads were washed four times with 500 µL Tween wash buffer. DNA was eluted in 20 µL pre-warmed elution buffer (10 mM EDTA, 95% formamide, 65 °C), ethanol-precipitated with glycogen and NaCl, and resuspended in 15 µL H₂O. PCR amplification was performed with KAPA HiFi polymerase (25 µL master mix, 2 µL EvaGreen, 2.5 µL 10 µM barcoded primer, 1 µL 25 µM universal forward primer, 4.5 µL water). Cycling proceeded for 5 cycles, stopping when real-time fluorescence monitoring reached 3,600 RFU. PCR products were diluted, purified using Zymo DNA Clean & Concentrator-5, and eluted in 15 µL H₂O. Final libraries were quantified with Qubit, quality-checked on a Fragment Analyzer, and sequenced (PE75, illumina NextSeq 550).

treatment_protocol: Hi-C libraries were generated as previously described(Wang, Sun et al. 2018) with minor modifications. OSCs were harvested, washed in PBS, and resuspended in M3 medium containing 1% formaldehyde at a density of 5 × 10⁶ cells/mL. After incubation for 10 min at room temperature (RT) with gentle rotation, 2 M Tris-HCl (pH 8.0) was added to a final concentration of 0.125 M to quench crosslinking. Samples were incubated for 5 min at RT, followed by 20 min on ice. Cells were pelleted, washed twice with PBS, and stored in aliquots of 5 × 10⁶ cells at –80 °C.


title: HEK293T cell, EMX1, G5T, epegRNA(2xPBS-RTT), PE2, rep3

description:
5519
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_metr_rt_5

description:
Library name: B253_iBAT_metr_rt
Mouse ID: JWK-1286


title: iBAT_metr_rt_6

description:
Library name: B254_iBAT_metr_rt
Mouse ID: JWK-1287


title: GSC23cells, BRD2_BD1_3

description:
Library name: Sample 45
raw_counts_1.txt
GSC23_BRD2KO+Y113F_3


title: HEK293T cell, EMX1, G5T, epegRNA(2xPBS-RTT), PE2, rep2

description:
5518
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: GSC23 cells, BRD2_BD1_2

description:
Library name: Sample 44
raw_counts_1.txt
GSC23_BRD2KO+Y113F_2


title: PROseq-wtOSC_permeabilized_biotin-U

description: Library name: PROseq-wtOSC_permeabilized_biotin-U

data_processing:
Paired end Illumina reads were processed by removing the first mate, clipping 5′ adapters with cutadapt  from the 2nd mate read(Martin 2011), filtering for minimal length (>18 nt) and sequence complexity (bbduk, entropy = 0.35, entropy window = 18, k = 4)(Bushnell 2014). Reads were first aligned to Drosophila rRNA precursor sequences and the mitochondrial genome using bowtie. Remaining unmapped reads were mapped to the OSC_r1.01 genome using STAR(Dobin, Davis et al. 2013) in two pass mode with splice junction support (--alignEndsType Local, --outFilterType BySJout, --alignSJoverhangMin 15, --alignSJDBoverhangMin 1, --outFilterMismatchNmax 1, --outFilterMultimapNmax 1000, --winAnchorMultimapNmax 2000). Alignments were separated into uniquely and all mapping fractions. For visualization, HOMER(Heinz, Benner et al. 2010) was used to generate tag directories (makeTagDirectory … -format bed -keepAll -single -fragLength given) and UCSC compatible tracks (makeUCSCfile … -fsize 1e20 -strand +/- -fragLength given -noadj -normLength 0) were generated per strand. Counts were normalized to 10M uniquely aligned reads  and final BigWig tracks were created with bedGraphToBigWig (Kent Utilities)(Perez, Barber et al. 2025), producing strand specific (sense and antisense) coverage profiles for genome browser display.
Assembly: OSC_r1.01.fa.gz
Supplementary files format and content: All mapping and unique mapping bigwig files for sense and antisense
Library strategy: PRO-Seq

molecule: nuclear RNA

extract_protocol:
PROseq libraries were prepared, modified from Mahat et al.(Mahat, Kwak et al. 2016) from both isolated nuclei and permeabilized cells. Cells were harvested, washed with ice-cold PBS, and counted. Nuclei were isolated by resuspension in Douncing buffer (10 mM Tris-HCl pH 7.4, 300 mM Sucrose, 3 mM CaCl₂, 2 mM MgCl₂, 0.1% Triton X-100, supplemented with 0.5 mM DTT, protease, and RNase inhibitors), dounced 25 times with a tight pestle, and pelleted. Permeabilized cells were prepared by resuspension in permeabilization buffer (10 mM Tris-HCl pH 7.4, 300 mM Sucrose, 10 mM KCl, 5 mM MgCl₂, 1 mM EGTA, 0.05% Tween, 0.1% NP40 substitute, supplemented with 0.5 mM DTT, protease, and RNase inhibitors), incubated on ice for 5 minutes, and pelleted. Both nuclei and permeabilized cells were resuspended in storage buffer (10 mM Tris-HCl pH 8.0, 25% glycerol, 5 mM MgCl₂, 0.1 mM EDTA, supplemented with 5 mM DTT), aliquoted, and snap-frozen in liquid nitrogen. Nuclear run-on reactions were performed using 2xNRO buffer supplemented with biotin-11-CTP and biotin-11-UTP (for 2-biotin mixes) or single biotinylated nucleotides (for 1-biotin mixes), ATP, GTP, and RNaseOUT. Samples were incubated at 30°C for 3 minutes, followed by RNA extraction using Trizol-LS and chloroform. RNA was precipitated with Glycoblue and 100% ethanol, incubated for 15 minutes at room temperature, and pelleted by centrifugation (16000 x G, 1 hour, 4°C). Pellets were washed with 75% ethanol and stored at -80°C.
Isolated RNA was fragmented by heat denaturation (65°C for 40 seconds) and alkaline hydrolysis (1N NaOH for 10 minutes on ice), then purified using P30 columns. Biotinylated RNA was enriched using MyOne C1 streptavidin beads, washed with high-salt, PROseq binding, and low-salt buffers, and eluted with Trizol and chloroform extraction. RNA was precipitated with Glycoblue and 100% ethanol. After decapping and hydroxyl repair using TAP and PNK enzymes followed by phenol purification, libraries were generated under sRNA seq ligation conditions. 3′ adaptor ligation was performed with T4 RNA ligase 2, truncated KQ, using adaptors containing four random nucleotides at the 5′ end(Jayaprakash, Jabado et al. 2011). After a second streptavidin purification, 5′ adaptor ligation was performed with T4 RNA ligase, using adaptors containing four random nucleotides at the 3′ end, followed by a third streptavidin purification. Reverse transcription was carried out with SuperScript II using an adaptor specific primer. Libraries were PCR amplified with KAPA HiFi polymerase and barcoded Illumina primers in reactions monitored by EvaGreen real time fluorescence. Reactions were stopped after reaching defined RFU thresholds (12–17 cycles). Products were separated on 2.5% agarose gels, gel purified (Zymo), and multiplexed for sequencing (PE50, Illumina HiSeq2500).


title: GSC23 cells, BRD2_BD1_1

description:
Library name: Sample 43
raw_counts_1.txt
GSC23_BRD2KO+Y113F_1


title: HEK293T cell, EMX1, G5T, epegRNA(2xPBS-RTT), PE2, rep1

description:
5517
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_metr_rt_3

description:
Library name: B251_iBAT_metr_rt
Mouse ID: JWK-1302


title: GSC23 cells, BRD2_WT_1

description:
Library name: Sample 40
raw_counts_1.txt
GSC23_BRD2KO+WT_1


title: iBAT_metr_rt_2

description:
Library name: B250_iBAT_metr_rt
Mouse ID: JWK-1278


title: HEK293T cell, FANCF, A7C, epegRNA, PE2, rep1

description:
5490
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: PROseq-wtOSC_permeabilized_biotin-CU

description: Library name: PROseq-wtOSC_permeabilized_biotin-CU


title: GSC23 cells, GSK620_3

description:
Library name: Sample 39
raw_counts_2.txt
GSC23_GSK620_3


title: PROseq-wtOSC_permeabilized_biotin-C

description: Library name: PROseq-wtOSC_permeabilized_biotin-C


title: iBAT_ctrl_rt_2

description:
Library name: B243_iBAT_ctrl_rt
Mouse ID: JWK-1299


title: GSC23 cells, DMSO_1

description:
Library name: Sample 34
raw_counts_2.txt
GSC23_DMSO_1

value: DMSO


title: HEK293T cell, RNF2, A4C, epegRNA, PE2, rep2

description:
5497
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, EMX1, G5T, loop2-25-22-7, PE2, rep3

description:
5444
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: SMA R6-Mo N3 replicate 4

city: Rozzano

first: Simona

last: Lodato

address: via Manzoni 56

country: Italy

institute: IRCCS Humanitas Research Hospital

description:
Library name: SMA_SCRAMBLE_N3_4
BULK-SAM-168

data_processing:
Sequence reads were aligned to GRCh38 and counted using STAR (v 2.7.10a, --chimSegmentMin 2, --alignIntronMax 1000000, --alignSJDBoverhangMin 6, --outSAMmapqUnique 254, --outFilterMismatchNmax 3).
Assembly: GRCh38
Supplementary files format and content: .txt table of raw counts

growth_protocol: Spinal Cord Organoids were generated by harvesting iPSCs using Accutase and approximately 9,000 iPSCs were plated in each well of an ultra-low attachment 96-well plate in pluripotent stem cell media containing low recombinant human FGF-basic (4 ng/ml) and Y-27632 (50 μM ROCK inhibitor). On day 2, half of the medium was changed to fresh medium containing a final concentration of 4 ng/ml bFGF, 50 μM Y-27632, 3 μM CHIR 99021 (GSK3 inhibitor), 2 μM SB-431542 (Activin/BMP/TGF-β pathway inhibitor), and 0.2 μM LDN-193189 (inhibitor of the bone morphogenetic protein [BMP] pathway). On day 4, the medium was replaced, adding the same small molecules (excluding bFGF and Y-27632). On day 6, the EBs were transferred into ultra-low attachment 24-well plates and cultured in NIM plus 100 nM retinoic acid. On day 12 EBs embedded in Cultrex and cultured with SCO differentiation media (SCODM, composed of 1:1 DMEM/F12 and Neurobasal with 0.5% N2 supplement, 1% B27, 2-mercaptoethanol, 0.025% insulin, 1% Glutamax, and 0.5% MEM-NEAA). 1 μM Smoothened Agonist was added to the media. On day 14, SCOs were transferred to spinning bioreactors and cultured with SCODM supplemented with RA and SAG. On day 28, concentrations of RA and SAG were reduced to 50% and neurotrophic factors BDNF, GDNF, and IGF-1 were added at 10 ng/ml each.

extract_protocol:
Total RNA was extracted using the ReliaPrep RNA Cell Miniprep System (Promega, Z6010).
Libraries were prepared starting from 50 ng of total RNA derived from 9 control, 6 SMA, and 16 morpholino-treated SMA (8 r6-MO, 8 r6-MO-scrambled) organoids, using the SMARTer Stranded RNA-Seq Kit, TaKaRa) and following the manufacturer’s instructions. Samples underwent a high coverage paired-end 150-bp strand-specific sequencing using a NextSeq 2000 platform (Illumina).

treatment_protocol: Anti-sense oligonucleotide morpholino (MO)10-34 sequence conjugated with r6 peptide (r6-MO). Details of specific MO used: (i) r6-MO targeting the SMN2 ISS-N1 region downstream of exon 7; (ii) negative control r6-MO with scrambled sequence. SMA SCOs were treated with growing concentrations of MO or r6-MO. The solution containing 2.4 nmoles MO or r6-MO was added directly to each well and incubated for 48h. Afterward, the culture medium was changed and fresh aliquots of 2.4 nmoles MO or r6-MO were added for an additional 48 h. The medium was then replaced with fresh medium for 72 h. Lastly, the organoids were treated again with 4.8 nmoles of MO or r6-MO for 48 h.

value: SMA


title: iBAT_ctrl_rt_1

description:
Library name: B242_iBAT_ctrl_rt
Mouse ID: JWK-1298


title: wtOSC_rRNA-depleted_RNAseq

description: Library name: wtOSC_rRNA-depleted_RNAseq

data_processing:
Illumina reads were processed by removing 3′ adapters with(Martin 2011), trimming the first five nucleotides, and filtering for minimal length (>18 nt) and sequence complexity (bbduk, entropy = 0.35, entropy window = 18, k = 4)(Bushnell 2014). Reads were first aligned to Drosophila rRNA precursor sequences and the mitochondrial genome using bowtie(Langmead, Trapnell et al. 2009). Remaining unmapped reads were mapped to the OSC_r1.01 genome using STAR in two pass mode with splice junction support (--alignEndsType Local, --outFilterType BySJout, --alignSJoverhangMin 15, --alignSJDBoverhangMin 1, --outFilterMismatchNmax 1, --outFilterMultimapNmax 1000, --winAnchorMultimapNmax 2000)(Dobin, Davis et al. 2013). Alignments were separated into uniquely and all mapping fractions. For visualization, HOMER(Heinz, Benner et al. 2010) was used to generate tag directories (makeTagDirectory … -format bed -keepAll -single -fragLength given) and UCSC compatible tracks (makeUCSCfile … -fsize 1e20 -strand +/- -fragLength given -noadj -normLength 0) were generated per strand. Counts were normalized to 10M uniquely aligned reads and final BigWig tracks were created with bedGraphToBigWig (Kent Utilities)(Perez, Barber et al. 2025), producing strand specific (sense and antisense) coverage profiles for genome browser display.
Assembly: OSC_r1.01
Supplementary files format and content: All mapping and unique mapping bigwig files for sense and antisense

extract_protocol:
Total RNA was isolated using Trizol (Invitrogen) according to the manufacturer’s instructions.
Ribosomal RNA was depleted using an RNAseH based protocol(Morlan, Qu et al. 2012, Adiconis, Borges-Rivera et al. 2013). In brief, standard 50 mer DNA oligos complementary to rRNA precursor and mitochondrial rRNA sequences were annealed to 1 µg total RNA in Hybridase buffer with EDTA. Samples were denatured (95 °C, 3 min), cooled gradually to 45 °C, and incubated with Thermostable Hybridase RNAseH (1 h, 45 °C) to specifically degrade rRNA. Following digestion, samples were treated with Turbo DNAse (30 min, 37 °C) and purified using RNA Clean & Concentrator 5 columns (Zymo). Elution was carried out in NEBNext Ultra II First Strand Synthesis buffer for downstream library preparation. RNA seq libraries were generated using the NEBNext Ultra II Directional RNA Library Prep Kit (New England Biolabs) according to the manufacturer’s instructions, with the exception of the final amplification. Libraries were PCR amplified with KAPA HiFi polymerase and barcoded Illumina primers, with amplification monitored in real time using EvaGreen fluorescence. Reactions were stopped once predetermined RFU thresholds were reached to avoid over amplification. Libraries were then purified, quantified by Qubit, assessed for fragment size, and multiplexed for Illumina sequencing.


title: iBAT_metr_rt_1

description:
Library name: B249_iBAT_metr_rt
Mouse ID: JWK-1266


title: PROseq-wtOSC_nuclei_biotin-C

description: Library name: PROseq-wtOSC_nuclei_biotin-C


title: HEK293T cell, RNF2, A4C, epegRNA, PE2, rep1

description:
5496
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_ctrl_rt_6

description:
Library name: B248_iBAT_ctrl_rt
Mouse ID: JWK-1310


title: HEK293T cell, EMX1, G5T, loop2-25-22-7-M, PE2, rep5

description:
5413
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: GSC23 cells, BRD2_WT_2

description:
Library name: Sample 41
raw_counts_1.txt
GSC23_BRD2KO+WT_2


title: Rat retina – Nano-LD (biol rep 7)

city: Messina

first: Luigi

last: Donato

state: Italy

address: via Consolare Valeria, 1

department: BIOMORF

institute: University of Messina

description:
Library name: mRNA-RAT-nano-LD-7_S65
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).

data_processing:
Raw reads were trimmed for adapter sequences and low-quality bases using Trimmomatic (v0.39).
High-quality reads were aligned to the Rattus norvegicus reference genome (rn6, Ensembl release XX) using STAR (v2.7.9a) with default parameters.
Aligned reads were assigned to genes using featureCounts (Subread v2.0.1).
Count matrices were normalized and analyzed for differential expression using DESeq2 (v1.30.1) in R (v4.0).
Pathway and gene ontology enrichment analyses were performed using clusterProfiler (v3.18).
Assembly: Rattus norvegicus (rn6)
Supplementary files format and content: count_matrix.csv: rows = genes, columns = samples; raw and normalized counts.

organism: Rattus norvegicus

growth_protocol: Adult rats were housed under standard conditions (12 h light/dark cycle, temperature 22±1°C, food and water ad libitum) until treatment.

extract_protocol:
Retinas were rapidly dissected under RNase-free conditions. Total RNA was extracted using the Qiagen RNeasy Mini Kit, following manufacturer’s instructions, including on-column DNase I treatment. RNA quality was assessed by Agilent Bioanalyzer; samples with RIN ≥ 7 were used.
RNA-seq libraries were prepared using the Watchmaker RNA Library Prep Kit (Twist Bioscience). Poly(A)-enriched mRNA was fragmented, reverse-transcribed, end-repaired, ligated to adapters, and PCR amplified according to manufacturer’s protocol. Libraries were indexed and pooled for sequencing.

treatment_protocol: Light-induced retinal degeneration was induced by high-intensity light exposure (15,000 lux, 24 h). Rats were treated with intravitreal injection of cerium oxide nanoparticles (nanoceria, 1 mM) or vehicle. Control animals received no treatment.

value: Retina

value: Nanoceria LD


title: SMA scramble N3 replicate 4

description:
Library name: SMA_R6-MO_N3_4
BULK-SAM-164

value: Scramble


title: HEK293T cell, FANCF, A7C, epegRNA, PE2, rep3

description:
5492
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: iBAT_ctrl_rt_5

description:
Library name: B247_iBAT_ctrl_rt
Mouse ID: JWK-1306

value: WT


title: GSC23 cells, DMSO_3

description:
Library name: Sample 36
raw_counts_2.txt
GSC23_DMSO_3


title: SMA scramble N3 replicate 3

description:
Library name: SMA_R6-MO_N3_3
BULK-SAM-163


title: Rat retina – Control (biol rep 7)

description:
Library name: mRNA-RAT-Ctrl-7_S60
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).

value: Control


title: HEK293T cell, FANCF, A7C, epegRNA, PE2, rep2

description:
5491
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1

value: base editing by prime editing


title: Rat retina – Vehicle + Light damage (biol rep 6)

description:
Library name: mRNA-RAT-veh-LD-6_S57
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).

value: Vehicle + Light damage


title: iBAT_ctrl_rt_4

description:
Library name: B245_iBAT_ctrl_rt
Mouse ID: JWK-1301

value: iBAT


title: SMA scramble N3 replicate 1

description:
Library name: SMA_R6-MO_N3_1
BULK-SAM-161


title: Liver_metr_ce_2

description:
Library name: B086_Liver_metr_ce
Mouse ID: JWK-1285

source_name: Liver

value: Liver


title: HEK293T cell, VEGFA, A3C, tetraloop-34-22-7-M, PE2, rep3

description:
5315
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: SMA NT N3 replicate 3

description:
Library name: SMA_NT_N3_3
BULK-SAM-160

value: NT


title: GSC23 cells, DMSO_2

description:
Library name: Sample 35
raw_counts_2.txt
GSC23_DMSO_2


title: iBAT_ctrl_rt_3

description:
Library name: B244_iBAT_ctrl_rt
Mouse ID: JWK-1300


title: HEK293T cell, EMX1, G5T, loop2-25-22-7, PE2, rep2

description:
5443
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: PROseq-wtOSC_nuclei_biotin-U

description: Library name: PROseq-wtOSC_nuclei_biotin-U

value: OSC

value: ovarian somatic cells


title: HEK293T cell, VEGFA, A3C, tetraloop-34-22-7-M, PE2, rep2

description:
5314
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: SMA R6-Mo N3 replicate 3

description:
Library name: SMA_SCRAMBLE_N3_3
BULK-SAM-167


title: HEK293T cell, EMX1, G5T, loop2-25-22-7-M, PE2, rep6

description:
5414
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Liver_metr_ce_7

description:
Library name: B091_Liver_metr_ce
Mouse ID: JWK-1303


title: Liver_metr_ce_6

description:
Library name: B090_Liver_metr_ce
Mouse ID: JWK-1313


title: Rat retina – Vehicle + Light damage (biol rep 4)

description:
Library name: mRNA-RAT-veh-LD-4_S45
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).


title: HEK293T cell, EMX1, G5T, loop2-25-22-7-M, PE2, rep4

description:
5412
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: PROseq-wtOSC_nuclei_biotin-CU

description: Library name: PROseq-wtOSC_nuclei_biotin-CU

value: wildtype


title: Rat retina – Vehicle-LD (biol rep 7)

description:
Library name: mRNA-RAT-veh-LD-7_S63
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).

value: Vehicle LD


title: Liver_ctrl_ce_7

description:
Library name: B084_Liver_ctrl_ce
Mouse ID: JWK-1329


title: SMA R6-Mo N3 replicate 2

description:
Library name: SMA_SCRAMBLE_N3_2
BULK-SAM-166


title: SMA NT N3 replicate 2

description:
Library name: SMA_NT_N3_2
BULK-SAM-159


title: Liver_metr_ce_5

description:
Library name: B089_Liver_metr_ce
Mouse ID: JWK-1312


title: HEK293T cell, VEGFA, A3C, loop2-34-20-7-M, PE2, rep3

description:
5402
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Liver_ctrl_ce_6

description:
Library name: B083_Liver_ctrl_ce
Mouse ID: JWK-1328


title: SMA NT N3 replicate 1

description:
Library name: SMA_NT_N3_1
BULK-SAM-158


title: Rat retina – Nano (biol rep 7)

description:
Library name: mRNA-RAT-nano-7_S64
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).

value: Nanoceria


title: Liver_metr_ce_3

description:
Library name: B087_Liver_metr_ce
Mouse ID: JWK-1330


title: Liver_metr_ce_4

description:
Library name: B088_Liver_metr_ce
Mouse ID: JWK-1311


title: HEK293T cell, VEGFA, A3C, loop2-34-20-7-M, PE2, rep2

description:
5401
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: SMA R6-Mo N3 replicate 1

description:
Library name: SMA_SCRAMBLE_N3_1
BULK-SAM-165


title: SMA scramble N3 replicate 2

description:
Library name: SMA_R6-MO_N3_2
BULK-SAM-162


title: Liver_metr_ce_1

description:
Library name: B085_Liver_metr_ce
Mouse ID: JWK-1305


title: Rat retina – Vehicle + Light damage (biol rep 5)

description:
Library name: mRNA-RAT-veh-LD-5_S51
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).


title: HEK293T cell, VEGFA, A3C, tetraloop-34-22-7-M, PE2, rep1

description:
5313
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Rat retina – Vehicle + Light damage (biol rep 3)

description:
Library name: mRNA-RAT-veh-LD-3_S39
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).


title: HEK293T cell, VEGFA, A3C, loop2-34-20-7-M, PE2, rep1

description:
5400
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, epegRNA+loop2-34-31-7, PE2, rep3

description:
5312
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: SMA R6-Mo CS83 replicate 4

description:
Library name: SMA_SCRAMBLE_CS83_4
BULK-SAM-157


title: Liver_ctrl_ce_5

description:
Library name: B082_Liver_ctrl_ce
Mouse ID: JWK-1327


title: HEK293T cell, VEGFA, A3C, epegRNA+loop2-34-31-7, PE2, rep2

description:
5311
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Rat retina – Vehicle + Light damage (biol rep 1)

description:
Library name: mRNA-RAT-veh-LD-1_S4
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).


title: SMA R6-Mo CS83 replicate 2

description:
Library name: SMA_SCRAMBLE_CS83_2
BULK-SAM-155


title: HEK293T cell, VEGFA, A3C, loop2-34-31-7, PE2, rep2

description:
5308
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: HEK293T cell, VEGFA, A3C, epegRNA+loop2-34-31-7, PE2, rep1

description:
5310
The G5T mutation was introduced into the EMX1 gene in HEK293T cells via PE2 with the loop2-25-22-7 , rep1


title: Liver_ctrl_ce_2

description:
Library name: B079_Liver_ctrl_ce
Mouse ID: JWK-1315


title: Rat retina – Nanoceria + Light damage (biol rep 6)

description:
Library name: mRNA-RAT-nano-LD-6_S59
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).

value: Nanoceria + Light damage


title: SMA R6-Mo CS83 replicate 1

description:
Library name: SMA_SCRAMBLE_CS83_1
BULK-SAM-154

value: SMA


title: Liver_ctrl_ce_3

description:
Library name: B080_Liver_ctrl_ce
Mouse ID: JWK-1316


title: Rat retina – Vehicle + Light damage (biol rep 2)

description:
Library name: mRNA-RAT-veh-LD-2_S33
Rat retina RNA-Seq; polyA selection; paired-end 150 bp; four lanes (L001–L004).

value: Retina


title: SMA scramble CS83 replicate 4

description:
Library name: SMA_R6-MO_CS83_4
BULK-SAM-153


title: Liver_ctrl_ce_1

description:
Library name: B078_Liver_ctrl_ce
Mouse ID: JWK-1314