title: Exp-UVC_K2S,Cell-HaCaT,Treatment-Control

description: UV_HaCat_NormalizedCounts_DESeq2.csv - UVC_K2S


title: Exp-UVC_F2S,Cell-HDF,Treatment-Control

status: Public on May 01 2025

type: SRA

city: Bremen

first: Jyoti

last: Jyoti

state: Bremen

address: Campus Ring 6

department: School of Science

country: Germany

institute: Constructor University

description: UV_HDF_NormalizedCounts_DESeq2.csv - UVC_F2S

library_source: transcriptomic

data_processing:
CLC Genomics Workbench 20.0.4 (CLC bio, Qiagen)
The resulting reads were reduced to succesful reads (Q30 bases run quality limit 0.15) and trimmed to 75 bp.
Trimmed sequence reads were mapped to human_GRCh38.p7_NCBI_20160606 (mismatch cost: 2; insertion/deletion cost: 3; length fraction: 0.8; similarity fraction: 0.8)
Assembly: GRCh38.p7
Supplementary files format and content: comma separated .csv files include normalized gene expression values. One file contains samples belonging to one batch of experiments.

molecule: total RNA

organism: Homo sapiens

source_name: HDF

growth_protocol: Cells were cultured in DMEM, high glucose, supplemented with 10% FBS and 1% penicillin streptomycin in an incubator with 37°C, 5% CO2 and saturated humidity.

extract_protocol:
To keep the homogeneity of the transmitting power flux density or SAR as high as possible, the cells outside a defined area were excluded during harvesting. After exposure, the medium of the cell dishes was discarded and a template made with a 3D printer (at 27 GHz an ellipse with a radius of 24 mm/23 mm; at 40.5 GHz a capped circle with a radius of 24 mm and a distance of the capped area to the center of 19 mm) was applied to the outside of the bottom of the dish and the areas to be omitted were marked. The cells were removed according to the marking with a cell scraper and the cell monolayer was then rinsed twice with PBS.
Cells were harvested by washing the remaining cell monolayer once with PBS and transferring 350 ul lysis buffer to each of the dishes. The cell lysates were collected using a cell scraper and transferring them into reaction tubes. The lysates were frozen at -80C until further processing. DNA and RNA extraction was done using the DNA/RNA Allprep Mini Kit (Qiagen) or the RNeasy Mini Kit (Qiagen) according to the manufacturer’s protocol.
The transcriptome library was generated using TruSeq Stranded mRNA technology according to the manufacturer's protocol (Illumina).

treatment_protocol:
For each exposure experiment 120,000 HDF or 200,000 HaCaT cells were seeded onto a 60 mm cell culture dish. Cells were used in passages 6-7 (HDF) and 35 (HaCaT). One day (48h experiments) or three days (2h experiments) after seeding, the dishes were transferred to the exposure incubators and cells were exposed or sham-exposed for 2h or 48h, with a frequency of 27 GHz or 40.5 GHz, and a power flux density of 1 mW/cm2 or 10 mW/cm2. In all experiments, the temperature was measured continuously using a fiber optic sensor. The internal temperature of the incubator in which the exposure takes place was reduced in the 10 mW/cm2 experiments manually, so that an average temperature of 37C could be maintained in the petri dishes and a lid heater was used to prevent condensation in the exposed petri dish and maintain blinding conditions.
Temperature controls: Cells were exposed or sham exposed at 40.5 GHz and 10 mW/cm2 for 48h without compensating for the temperature increase. The starting temperature of the medium was 37.0C in these experiments and increased to an average of 38.3C in the exposed samples. No lid heaters were used, so that the heating of the medium in the exposed samples led to medium condensation.
UV controls: Three replicates each of HaCaT and HDF cells were exposed or sham-exposed at a wavelength of 254 nm and an illuminance of 1 mW/cm2. The medium was removed and collected, rinsed twice with PBS and the cells were thinly coated with 600 ul PBS per dish. Exposure was performed without the dish lid for 20 sec, for sham exposure the dishes were stored in the dark at room temperature. The PBS was then aspirated, the cells were layered with the previously collected medium, and incubated for 2h at 37C and 5%CO2.

tag: cell type

value: HDF

tag: treatment

value: Control

tag: date


title: Exp-V8F_1A,Cell-HDF,Power-10,Frequency-27,Time-48,Treatment-sham

description: P10_HDF_NormalizedCounts_DESeq2.csv - S90

value: sham

tag: trial

tag: incubator

value: A

tag: frequency [ghz]

tag: power.density [mw/cm2]

tag: exposure.time [h]

tag: aliquot

tag: passage

value: b


title: Exp-V7F_6A,Cell-HDF,Power-10,Frequency-27,Time-2,Treatment-sham

description: P10_HDF_NormalizedCounts_DESeq2.csv - S86

value: a


title: Exp-V5F_3B,Cell-HDF,Power-10,Frequency-40.5,Time-48,Treatment-sham

description: P10_HDF_NormalizedCounts_DESeq2.csv - S89

value: B

value: c


title: Exp-V5F_3A,Cell-HDF,Power-10,Frequency-40.5,Time-48,Treatment-exposed

description: P10_HDF_NormalizedCounts_DESeq2.csv - S88

value: exposed


title: Exp-V7F_6B,Cell-HDF,Power-10,Frequency-27,Time-2,Treatment-exposed

description: P10_HDF_NormalizedCounts_DESeq2.csv - S87


title: Exp-V6F_2B,Cell-HDF,Power-10,Frequency-40.5,Time-2,Treatment-sham

description: P10_HDF_NormalizedCounts_DESeq2.csv - S77


title: Exp-V7F_4A,Cell-HDF,Power-10,Frequency-27,Time-2,Treatment-sham

description: P10_HDF_NormalizedCounts_DESeq2.csv - S82


title: Exp-V7F_4B,Cell-HDF,Power-10,Frequency-27,Time-2,Treatment-exposed

description: P10_HDF_NormalizedCounts_DESeq2.csv - S83


title: Exp-V6F_2A,Cell-HDF,Power-10,Frequency-40.5,Time-2,Treatment-exposed

description: P10_HDF_NormalizedCounts_DESeq2.csv - S76


title: Exp-V6K_7B,Cell-HaCaT,Power-10,Frequency-40.5,Time-48,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S73

source_name: HaCaT

tag: cell line

value: HaCaT


title: Exp-V5K_4A,Cell-HaCaT,Power-10,Frequency-27,Time-48,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S70


title: Exp-V5K_3A,Cell-HaCaT,Power-10,Frequency-27,Time-48,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S68


title: Exp-V8K_5A-Neu,Cell-HaCaT,Power-10,Frequency-27,Time-2,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S110

value: ac


title: Exp-V8K_6A,Cell-HaCaT,Power-10,Frequency-27,Time-2,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S64


title: Exp-V8K_4B,Cell-HaCaT,Power-10,Frequency-27,Time-2,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S61


title: Exp-V6K_7A,Cell-HaCaT,Power-10,Frequency-40.5,Time-48,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S72


title: Exp-V8K_5B,Cell-HaCaT,Power-10,Frequency-27,Time-2,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S63


title: Exp-V5K_2A,Cell-HaCaT,Power-10,Frequency-27,Time-48,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S66


title: Exp-V8K_6B,Cell-HaCaT,Power-10,Frequency-27,Time-2,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S65


title: Exp-V8K_4A,Cell-HaCaT,Power-10,Frequency-27,Time-2,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S60


title: Exp-V5K_3B,Cell-HaCaT,Power-10,Frequency-27,Time-48,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S69


title: Exp-V5K_2B,Cell-HaCaT,Power-10,Frequency-27,Time-48,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S67


title: Exp-V7K_7B,Cell-HaCaT,Power-10,Frequency-40.5,Time-2,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S59


title: Exp-V7K_6B,Cell-HaCaT,Power-10,Frequency-40.5,Time-2,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S57


title: Exp-V7K_6A,Cell-HaCaT,Power-10,Frequency-40.5,Time-2,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S56


title: Exp-V7K_7A,Cell-HaCaT,Power-10,Frequency-40.5,Time-2,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S58


title: Exp-V7K_5B,Cell-HaCaT,Power-10,Frequency-40.5,Time-2,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S55


title: Exp-V7K_5A,Cell-HaCaT,Power-10,Frequency-40.5,Time-2,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S54


title: Exp-V6K_8A,Cell-HaCaT,Power-10,Frequency-40.5,Time-48,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S74


title: Exp-V6K_5A,Cell-HaCaT,Power-10,Frequency-40.5,Time-48,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S52


title: Exp-V5K_4B,Cell-HaCaT,Power-10,Frequency-27,Time-48,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S71


title: Exp-V6K_5B,Cell-HaCaT,Power-10,Frequency-40.5,Time-48,Treatment-sham

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S53


title: Exp-V4F_3A,Cell-HDF,Power-1,Frequency-27,Time-48,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S48

value: bc


title: Exp-V4F_3B,Cell-HDF,Power-1,Frequency-27,Time-48,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S49


title: Exp-V4F_2B,Cell-HDF,Power-1,Frequency-27,Time-48,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S47


title: Exp-V4F_2A,Cell-HDF,Power-1,Frequency-27,Time-48,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S46


title: Exp-V4F_1B,Cell-HDF,Power-1,Frequency-27,Time-48,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S45


title: Exp-V1F_3A,Cell-HDF,Power-1,Frequency-40.5,Time-48,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S40


title: Exp-V1F_4B,Cell-HDF,Power-1,Frequency-40.5,Time-48,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S43


title: Exp-V1F_3B,Cell-HDF,Power-1,Frequency-40.5,Time-48,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S41


title: Exp-V1F_2A,Cell-HDF,Power-1,Frequency-40.5,Time-48,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S38


title: Exp-V1F_2B,Cell-HDF,Power-1,Frequency-40.5,Time-48,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S39


title: Exp-V3F_3B,Cell-HDF,Power-1,Frequency-27,Time-2,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S37


title: Exp-V3F_3A,Cell-HDF,Power-1,Frequency-27,Time-2,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S36


title: Exp-V3F_1B,Cell-HDF,Power-1,Frequency-27,Time-2,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S33


title: Exp-V3F_1A,Cell-HDF,Power-1,Frequency-27,Time-2,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S32


title: Exp-V3F_2A,Cell-HDF,Power-1,Frequency-27,Time-2,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S34


title: Exp-V3F_2B,Cell-HDF,Power-1,Frequency-27,Time-2,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S35


title: Exp-V2F_3B,Cell-HDF,Power-1,Frequency-40.5,Time-2,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S31


title: Exp-V6K_8B,Cell-HaCaT,Power-10,Frequency-40.5,Time-48,Treatment-exposed

description: P10_HaCat_NormalizedCounts_EDASeq.csv - S75


title: Exp-V2F_2A,Cell-HDF,Power-1,Frequency-40.5,Time-2,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S28


title: Exp-V2H_1A,Cell-HDF,Power-1,Frequency-40.5,Time-2,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S26


title: Exp-V2H_1B,Cell-HDF,Power-1,Frequency-40.5,Time-2,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S27


title: Exp-V4K_4B,Cell-HaCaT,Power-1,Frequency-40.5,Time-48,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S24


title: Exp-V4K_4A,Cell-HaCaT,Power-1,Frequency-40.5,Time-48,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S23


title: Exp-V4K_3A,Cell-HaCaT,Power-1,Frequency-40.5,Time-48,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S21


title: Exp-V4K_3B,Cell-HaCaT,Power-1,Frequency-40.5,Time-48,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S22


title: Exp-V4F_1A,Cell-HDF,Power-1,Frequency-27,Time-48,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S44


title: Exp-V4K_1A,Cell-HaCaT,Power-1,Frequency-40.5,Time-48,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S19

value: ba


title: Exp-V3K_4A,Cell-HaCaT,Power-1,Frequency-40.5,Time-2,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S17


title: Exp-V3K_3B,Cell-HaCaT,Power-1,Frequency-40.5,Time-2,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S16


title: Exp-V3K_4B,Cell-HaCaT,Power-1,Frequency-40.5,Time-2,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S18


title: Exp-V4K_1B,Cell-HaCaT,Power-1,Frequency-40.5,Time-48,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S20


title: Exp-V1F_4A,Cell-HDF,Power-1,Frequency-40.5,Time-48,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S42


title: Exp-V3K_1B,Cell-HaCaT,Power-1,Frequency-40.5,Time-2,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S14


title: Exp-V2K_6B,Cell-HaCaT,Power-1,Frequency-27,Time-2,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S12


title: Exp-V3K_1A,Cell-HaCaT,Power-1,Frequency-40.5,Time-2,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S13


title: Exp-V2K_6A,Cell-HaCaT,Power-1,Frequency-27,Time-2,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S11


title: Exp-V2K_4B,Cell-HaCaT,Power-1,Frequency-27,Time-2,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S8


title: Exp-V2F_3A,Cell-HDF,Power-1,Frequency-40.5,Time-2,Treatment-sham

description: P1_HDF_NormalizedCounts_DESeq2.csv - S30


title: Exp-V2K_4A,Cell-HaCaT,Power-1,Frequency-27,Time-2,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S7


title: Exp-V2K_5B,Cell-HaCaT,Power-1,Frequency-27,Time-2,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S10


title: Exp-V1K_4B,Cell-HaCaT,Power-1,Frequency-27,Time-48,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S6


title: Exp-V2F_2B,Cell-HDF,Power-1,Frequency-40.5,Time-2,Treatment-exposed

description: P1_HDF_NormalizedCounts_DESeq2.csv - S29


title: Exp-V1K_4A,Cell-HaCaT,Power-1,Frequency-27,Time-48,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S5


title: Exp-V1K_3B,Cell-HaCaT,Power-1,Frequency-27,Time-48,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S4


title: Exp-V1K_3A,Cell-HaCaT,Power-1,Frequency-27,Time-48,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S3


title: Exp-V1K_2B,Cell-HaCaT,Power-1,Frequency-27,Time-48,Treatment-exposed

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S2


title: Exp-V1K_2A,Cell-HaCaT,Power-1,Frequency-27,Time-48,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S1


title: SHAPE_s4U_COX1_P2_1_DC_B

city: New Haven

first: Matthew

last: Simon

state: CT

address: 266 Whitney Ave

department: Molecular Biophysics and Biochemistry

country: USA

institute: Yale University

description:
COX1 cellular RNA
Shapemapper2 was used for reactivity profiling
Tot_MTCO1_SHAPE.xlsx

data_processing:
Reads were filtered for unique sequences using FastUniq
Reads were trimmed using cutadapt (--minimum-length=20)
reads were aligned to the GRCh38 genome using HISAT-3N aligned with default parameters.
Reads were processed using Picard tools, including FixMateInformation, SortSam and BuildBamIndex
Reads were filtered for those that aligned uniquely (flag: 83/163, 99/147), with MAPQ ≥ 2, and without insertions.
HTSeq-count was used to identify mapped reads in UCSC transcripts (union mode)
T-to-C mutations were identified using Rsamtools and a custom R script (available on Bitbucket). Only mutations at positions with a base quality score of greater than 40 that were at least three nt from the end of the read were counted. Reads were excluded where there were greater than five T-to-C mutations and these mutations did not account for at least one third of the observed mutations (NM tag).
SNP sites were filtered from our data in two ways: 1, bcftools was used to identify T-to-C SNP sites in control samples, and these sites were excluded from analysis; 2, sites exhibiting high T-to-C mutation rates in non-s4U treated controls were excluded from analysis.
To examine the distribution of reads with each minimum number of T-to-C mutations, bam files were filtered using Picard tools.
STAR aligner was used to make genome-coverage tracks (inputAlignmentsFromBam mode, outWigType bedGraph).
Tracks were converted to binary format using IGVtools (toTDF).
Assembly: BDGP6 or GRCh38
U reads were filtered based on read count (reads >= 25), prior to HDProbe analysis
Supplementary files format and content: Tracks are provided in tdf format to visualize enrichment levels for controls and T-to-C mutation counts for all Tether-seq samples.

source_name: HEK293T cells

growth_protocol: HEK-293T cells were cultured at 37°C in DMEM supplemented with 10% FBS and 1% Penicillin/Streptomycin. S2 cells were cultured at 27°C in Schneider's Drosophila Medium with 10% heat-inactivated FBS and 1% Penicillin/Streptomycin.

extract_protocol:
RNA was harvested under non-denaturing conditions using a hypotonic lysis buffer (HLB). The pelleted cells were resuspended in HLB (20 mM HEPES, pH 8, 2 mM MgCl2, 0.2 mM EGTA, 10% glycerol, fresh 1 mM DTT) and freeze-thawed 3x and then placed on ice. After adding NaCl to 400 mM, samples were iced for 5 min and then centrifuged for 15 min at 4°C.The supernatant was collected, treated with proteinase K and 1% SDS and incubated at 37°C for 45 min. RNA was extracted 2x with phenol:chloroform:isoamyl (24:24:1) and 2x with chloroform. The aqueous layer was treated with Turbo DNAse and then buffer exchanged into folding buffer.
RNA was reverse transcribed by first annealing to the RT primer (2 pmol) by heating at 65°C for 5 minutes and then cooling on ice. The RT samples (20 μL) were then prepared by adding 0.4 volumes of freshly prepared 2.5x MaP buffer (125 mM Tris-HCl, pH 8, 187.5 mM KCl, 15 mM MnCl2, 25 mM DTT, 1.25 mM dNTPs) and 1 μL of SuperScript II and incubating at 42°C for 3 hours. The enzyme was heat-inactivated by heating at 70°C for 15 minutes and the cDNA product was purified with 3 volumes of SPRI beads. The RT cDNA was amplified using Phusion polymerase and 10 pmols of PCR primers and purified with 3 volumes of SPRI beads. The PCR product was further amplified using 10 pmols of primers containing sequencing indexes and isolated using 1.5 volumes of SPRI beads.

treatment_protocol: SHAPE-MaP was performed largely following the protocol described previously (Smola et al. Nature Protocols 2018). Three samples were prepared to probe the secondary structure of the COX1 transcript: +1M7 (+), –1M7 (–) and denatured control (DC).

value: HEK293T cells

tag: s4u feed

value: Yes (100 μM, 2 hours)


title: SHAPE_s4U_COX1_P2_1_no_1M7_B


title: Exp-V3K_3A,Cell-HaCaT,Power-1,Frequency-40.5,Time-2,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S15


title: SHAPE_s4U_COX1_P2_1_DC_A


title: SHAPE_s4U_COX1_P2_1_1M7_A


title: SHAPE_s4U_COX1_P2_1_no_1M7_A


title: SHAPE_s4U_COX1_P4_1M7_A


title: SHAPE_s4U_COX1_P3_DC_B


title: SHAPE_s4U_COX1_P4_1M7_B


title: SHAPE_s4U_COX1_P4_no_1M7_A


title: SHAPE_s4U_COX1_P4_DC_A


title: SHAPE_s4U_COX1_P4_DC_B


title: Exp-V2K_5A,Cell-HaCaT,Power-1,Frequency-27,Time-2,Treatment-sham

description: P1_HaCat_NormalizedCounts_EDASeq.csv - S9


title: SHAPE_s4U_COX1_P3_no_1M7_B


title: SHAPE_s4U_COX1_P3_DC_A


title: SHAPE_s4U_COX1_P3_1M7_A


title: SHAPE_s4U_COX1_P2_DC_B


title: SHAPE_s4U_COX1_P2_DC_A


title: SHAPE_s4U_COX1_P2_1M7_B


title: SHAPE_s4U_COX1_P2_no_1M7_B


title: SHAPE_s4U_COX1_P2_1M7_A


title: SHAPE_s4U_COX1_P3_1M7_B


title: SHAPE_s4U_COX1_P2_no_1M7_A


title: SHAPE_s4U_COX1_P1_no_1M7_B


title: SHAPE_s4U_COX1_P1_1M7_B


title: SHAPE_s4U_COX1_P1_DC_A


title: SHAPE_s4U_COX1_P1_no_1M7_A


title: SHAPE_s4U_COX1_P1_1M7_A


title: SHAPE_s4U_COX1_P4_no_1M7_B


title: SHAPE_s4U_COX1_P2_1_1M7_B


title: SHAPE_COX1_P2_1_no_1M7_B

treatment_protocol: SHAPE-MaP was performed largely following the protocol described previously (Smola et al. Nature Protocols 2018). Three samples were prepared to probe the secondary structure of the COX1 transcript: +1M7 (+), –1M7 (–) and denatured control (DC). Two additional samples were prepared with either 1-NH2 or 2-NH2 (1 mM)  and were also probed using 1M7.

value: No


title: SHAPE_COX1_P2_1_2-NH2_B


title: SHAPE_COX1_P2_1_DC_A


title: SHAPE_COX1_P2_1_no_1M7_A


title: SHAPE_COX1_P2_1_1M7_B


title: SHAPE_COX1_P2_1_DC_B


title: SHAPE_COX1_P2_1_1-NH2_A


title: SHAPE_COX1_P2_1_1M7_A


title: SHAPE_COX1_P2_1_1-NH2_B


title: SHAPE_s4U_COX1_P3_no_1M7_A


title: Tether_HEK293T_Mol1_No2PT_B

description:
Enriched RNA
Reads filtered for number of T-to-C mutations (0-5+) and strandedness (min/pos) for each transcript and U position

extract_protocol:
RNA was harvested under non-denaturing conditions using a hypotonic lysis buffer (HLB). The pelleted cells were resuspended in HLB (20 mM HEPES, pH 8, 2 mM MgCl2, 0.2 mM EGTA, 10% glycerol, fresh 1 mM DTT) and freeze-thawed 3x and then placed on ice. After adding NaCl to 400 mM, samples were iced for 5 min and then centrifuged for 15 min at 4°C.The supernatant was collected, treated with proteinase K and 1% SDS and incubated at 37°C for 45 min. RNA was extracted 2x with phenol:chloroform:isoamyl (24:24:1) and 2x with chloroform. The aqueous layer was treated with Turbo DNAse and then buffer exchanged into folding buffer.
Tether-seq libraries were generated following standard protocols using the SMARTer Stranded Total RNA-Seq Kit-Pico Input Mammalian, V2 (Takara Bio USA, cat. 634413).

treatment_protocol: 8 μg of total RNA was subjected to tethering conditions: Samples (40 μL) were prepared with non-denatured s4U-labeled RNA (8 μg), 0-250 μM disulfide small molecule and 0-500 μM 2PT competitor in 20 mM HEPES, pH 8, 1 mM EDTA, and 5% DMF. After incubating at room temperature for 30 minutes, mCPBA (0-15 mM, 10% ethanol final) was added and reactions were incubated at 45-50°C for 45 minutes. Urea was added to 5 M and mCPBA was replenished to a final concentration of 15 mM and final volume of 90 μL. Samples were again incubated at 45-50°C for 45 minutes. DTT (30 mM final) was added and samples were heated at 55°C for 15 minutes to quench excess oxidant and reduce off tether. Samples were biotinylated and enriched as previously described (Duffy et al., 2015). RNA was then treated with oxidative-nucleophilic-aromatic-substitution chemistry using mCPBA as the oxidant as previously described (Schofield et al., Nature Methods 2018).


title: U-2OS Clone#12 WGS

city: Munich

first: Stephan

last: Hamperl

state: Bayern

address: Feodor-Lynen-Strasse 21

department: Institute of Epigenetics and Stem Cells

institute: Helmholtz-Zentrum Munich

library_source: genomic

data_processing:
The human genome (version GRCh38 primary assembly) was downloaded from the Ensembl database, and the sequence of the reporter construct was concatenated to the genome fasta file.
Paired-end sequencing reads were aligned to the combined genome by bwa (version 0.7.17) with the parameter -B 3 (Li and Durbin, 2009)
Mapped reads were filtered by samtools (version 1.16.1) with the parameter -q 12 (Danecek et al., 2021).
Structural variants were called on the filtered reads and the combined genome file using tiddit (version 3.3.2) with the parameters --sv -p 1 -r 1 --min_contig 4500 (Lindstrand et al., 2017).
Genomic positions detected by reads split between the reporter and another chromosome were extracted from the variant call file (vcf). To exclude genomic positions that mapped internally to the reporter sequence, only those positions were considered that are within 300 bp of the reporter ends.
Adjacent genomic positions with a maximum distance of 200 bp were merged using bedtools merge (version 2.31.0) with the parameter -d 200 (Quinlan and Hall, 2010) and the average of the start and end position of the merged region was used as insertion site.
Assembly: hg38
Supplementary files format and content: bed file with insertion sites

molecule: genomic DNA

source_name: U-2OS Clone#12

growth_protocol: U-2OS Tet-ON Clone#12 cells were grown in DMEM (GIBCO) containing 10% FBS, 2mM L-glutamine, and penicillin/streptomycin, in a 5% CO2 environment at 37°C

extract_protocol:
Genomic DNA was isolated by phenol-chloroform extraction. 20 µg/sample of DNA were sonicated on a Covaris E220 focused sonicator for 10 min.
Libraries were constructed using NEBNext Ultra II DNA Library Kit with Purification Beads (E7103S, E7103L) and NEBNext Multiplex Oligos for Illumina (E6440S) according to manufacturer's instructions
Whole Genome Sequencing

value: U-2OS Clone#12

value: U-2OS Tet-ON


title: hCiPSC-derived RPE cells, C40 cell line, rep3

city: Beijing

first: Ke

last: Zhang

address: Huayuan North Road

country: China

institute: Peking university

description:
10753-1_hCiPSCs_C40_3-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt

data_processing:
The sequencing data was filtered with SOAPnuke, afterwards clean reads were obtained and stored in FASTQ format.
Experssion level of gene was calculated by featureCount (v2.14.2).
The raw data was filtered by using Fastp.
The heatmap was drawn by pheatmap (v1.0.8) [19] according to the gene expression difference.
Differential expression analysis was performed using the DESeq2(v1.4.5).
Assembly: GRCh38
Supplementary files format and content: tab-delimited text file includes FLPM values for each samples

extract_protocol:
Total RNA was isolated using the Direct-zol RNA MiniPrep Kit from Zymo Research, and checked for integrity using an Agilent Bioanalyzer 2100.
mRNA is enriched by oligo (dT) -attached magnetic beads from total RNA, and strand specific transcriptome library is constructed.
The transcriptome library was sequenced by DNBSEQ high-throughput platform and Illumina Novaseq6000/BGISEQ-500.

value: hCiPSC-derived RPE cells

tag: genotype

value: no treatment


title: hCiPSC-derived RPE cells, C40 cell line, rep1

description:
10753-1_hCiPSCs_C40_1-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: Tether_HEK293T_Mol1_No2PT_A


title: SHAPE_s4U_COX1_P1_DC_B


title: hCiPSC-derived RPE cells, C17 cell line, rep2

description:
10753-1_hCiPSCs_C17_2-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C40 cell line, rep2

description:
10753-1_hCiPSCs_C40_2-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C17 cell line, rep3

description:
10753-1_hCiPSCs_C17_3-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C6 cell line, rep3

description:
10753-1_hCiPSCs_C6_3-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C6 cell line, rep2

description:
10753-1_hCiPSCs_C6_2-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C7 cell line, rep3

description:
10753-1_hCiPSCs_C7_3-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C6 cell line, rep1

description:
10753-1_hCiPSCs_C6_1-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C3 cell line, rep3

description:
10753-1_hCiPSCs_C3_3-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C3 cell line, rep2

description:
10753-1_hCiPSCs_C3_2-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C7 cell line, rep2

description:
10753-1_hCiPSCs_C7_2-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: hCiPSC-derived RPE cells, C3 cell line, rep1

description:
10753-1_hCiPSCs_C3_1-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: SHAPE_COX1_P2_1_2-NH2_A

description:
COX1 cellular RNA
Shapemapper2 was used for reactivity profiling
Tot_MTCO1_SHAPE.xlsx


title: U-2OS Clone#12 cells, H3 INPUT R1/2/3

data_processing:
Paired-end reads were trimmed using trim_galore (version 0.6.10) (Krueger et al., 2023) with the parameter --quality 28
Reads were mapped to the genome using bowtie2 (version 2.5.1) with the parameters --end-to-end --very-sensitive --no-unal --no-mixed --no-discordant --dovetail -I 10 -X 700
Aligned reads were filtered by samtools (version 1.17) with the parameter -q 12
Duplicated reads were removed using picard MarkDuplicates (version 3.0.0) with the parameter -REMOVE_DUPLICATES TRUE
Genome coverages were created using deeptools bamCoverage (version 3.5.2) (Ramírez et al., 2016) with parameters --blackListFileName hg38-blacklist.v2.bed --ignoreForNormalization MT --binSize 20 --smoothLength 60 –extendReads --maxFragmentLength 700 --normalizeUsing "CPM"
Assembly: hg38
Supplementary files format and content: bigWig bigWig with normalized ChIP-seq coverage

growth_protocol: U-2OS Clone#12 cells were grown in DMEM (GIBCO) containing 10% FBS, 2mM L-glutamine, and penicillin/streptomycin, in a 5% CO2 environment at 37°C

extract_protocol:
Cells were fixed with 1% formalehyde for 10 min, quenched and sonicated with a Covaris E220 focused ultra sonicator for 10 min
Libraries were constructed using NEBNext Ultra II DNA Library Kit with Purification Beads (E7103S, E7103L) and NEBNext Multiplex Oligos for Illumina (E6440S) according to manufacturer's instructions

treatment_protocol: U-2OS Clone#12 cells were synchronized by double thymidine block. Cells were released into S-phase by replacing thymidine containing medium by fresh culturing medium. Simultaneous to S-phase release  0 or 1 µg/mL Doxycycline was added. Cells were harvested 4h after release.

tag: chip antibody

value: Histone H3 (Abcam #1791)

value: DMSO


title: U-2OS Clone#12 cells, H3 ChIP DOX R3

value: DMSO+DOX


title: U-2OS Clone#12 cells, H3 INPUT DOX R1/2/3


title: U-2OS Clone#12 cells, H3 ChIP R3


title: U-2OS Clone#12 cells, H3 ChIP DOX R1


title: U-2OS Clone#12 cells, RNAPII Ser2P INPUT R1/2/3

value: RNAPII Ser2P (Abcam #ab5095)


title: U-2OS Clone#12 cells, RNAPII Ser2P INPUT DOX R1/2/3


title: U-2OS Clone#12 cells, H3 ChIP R1


title: U-2OS Clone#12 cells, H3 ChIP R2


title: U-2OS Clone#12 cells, H3 ChIP DOX R2


title: hCiPSC-derived RPE cells, C17 cell line, rep1

description:
10753-1_hCiPSCs_C17_1-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: U-2OS Clone#12 cells, RNAPII Ser2P ChIP R3


title: U-2OS Clone#12 cells, RNAPII Ser2P ChIP DOX R1


title: U-2OS Clone#12 cells, RNAPII Ser2P ChIP DOX R2


title: U-2OS Clone#12 cells, RNAPII Ser2P ChIP DOX R3


title: U-2OS Clone#12 cells, RNAPII Ser2P ChIP R1


title: mDA neurons, SLC30A10 patient 2, Mn treated, replicate 3

city: London

first: Dimitri

last: Budinger

address: 30 guilford street

country: United Kingdom

institute: university college london

data_processing:
Sequenced reads were trimmed for adaptor sequence, and filtered for low-quality sequence, then mapped to GRCh38 whole genome using HISAT2 (v.2.2.1).
Fragments counts for genes were extracted with featureCounts (v.1.6.4) excluding duplicates, multimapping reads and chimeric fragments.
Differential gene expression was analysed using edgeR (v.3.24.1), filtering low counts with 0.35 minimum CPM in at least 3 samples.
Differential gene expression was performed comparing disease status (patients vs. controls) in both basal and manganese-treated conditions. ZIP14 and ZnT10 patient lines were also compared together to controls for hypermanganesaemia phenotypes analyses.
Assembly: Genome_build: GRCh38
Supplementary files format and content: Tabular file including raw counts for each control and patient lines in basal and manganese treated conditions. File generated in Galaxy.org

source_name: Induced pluripotent stem cells

growth_protocol: iPSC-derived midbrain dopaminergic neurons were produced following the protocol in Ng et al., 2021, Science Translational Medicine

extract_protocol:
Total RNA was isolated from midbrain dopaminergic neurons using the RNeasy mini kit (Qiagen) and RNA libraries were prepared from 100 ng of total RNA using KAPA mRNA HyperPrep kit (Roche).
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: Midbrain dopaminergic neurons were collected in basal or manganese treated conditions (100uM for 48h)

tag: tissue

value: Induced pluripotent stem cells

value: Midbrain dopaminergic neurons

value: Day 65 of differentiation

value: SLC30A10 mutant 2

value: Manganese 100μM 48h


title: U-2OS Clone#12 cells, RNAPII Ser2P ChIP R2


title: mDA neurons, SLC30A10 patient 2, Mn treated, replicate 2


title: hCiPSC-derived RPE cells, C7 cell line, rep1

description:
10753-1_hCiPSCs_C7_1-ZK-6G_S1_L002
new_mean_gene_fpkm(1)(1).txt


title: mDA neurons, SLC30A10 patient 2, Mn treated, replicate 1


title: mDA neurons, SLC39A8 patient 2, Mn treated, replicate 3

value: SLC39A8 mutant 2


title: mDA neurons, SLC30A10 patient 1, Mn treated, replicate 1

value: SLC30A10 mutant 1


title: mDA neurons, SLC30A10 patient 1, Mn treated, replicate 2


title: mDA neurons, SLC30A10 patient 1, Mn treated, replicate 3


title: mDA neurons, SLC39A8 patient 2, Mn treated, replicate 1


title: mDA neurons, SLC39A8 patient 1, Mn treated, replicate 2

value: SLC39A8 mutant 1


title: mDA neurons, SLC39A8 patient 1, Mn treated, replicate 3


title: mDA neurons, SLC39A8 patient 2, Mn treated, replicate 2


title: mDA neurons, SLC39A8 patient 1, Mn treated, replicate 1


title: mDA neurons, SLC39A14 patient 2, Mn treated, replicate 2

value: SLC39A14 mutant 2


title: mDA neurons, SLC39A14 patient 2, Mn treated, replicate 1


title: mDA neurons, SLC39A14 patient 2, Mn treated, replicate 3


title: mDA neurons, SLC39A14 patient 1, Mn treated, replicate 3

value: SLC39A14 mutant 1


title: mDA neurons, SLC39A14 patient 1, Mn treated, replicate 1


title: mDA neurons, control 1, Mn treated, replicate 3

value: Healthy control


title: mDA neurons, SLC39A14 patient 1, Mn treated, replicate 2


title: mDA neurons, control 3, Mn treated, replicate 2


title: mDA neurons, control 2, Mn treated, replicate 3


title: mDA neurons, control 1, Mn treated, replicate 2


title: mDA neurons, control 1, Mn treated, replicate 1


title: mDA neurons, SLC30A10 patient 2, basal, replicate 1

value: No treatment


title: mDA neurons, SLC30A10 patient 1, basal, replicate 3


title: mDA neurons, SLC30A10 patient 2, basal, replicate 2


title: mDA neurons, SLC39A8 patient 2, basal, replicate 3


title: mDA neurons, SLC30A10 patient 1, basal, replicate 2


title: mDA neurons, SLC30A10 patient 1, basal, replicate 1


title: mDA neurons, SLC39A8 patient 1, basal, replicate 3


title: mDA neurons, SLC39A8 patient 2, basal, replicate 1


title: mDA neurons, SLC39A8 patient 2, basal, replicate 2


title: mDA neurons, SLC39A14 patient 2, basal, replicate 2


title: mDA neurons, SLC39A14 patient 2, basal, replicate 3


title: mDA neurons, SLC39A8 patient 1, basal, replicate 1


title: mDA neurons, SLC39A14 patient 2, basal, replicate 1


title: mDA neurons, SLC39A14 patient 1, basal, replicate 3


title: mDA neurons, control 2, Mn treated, replicate 1


title: mDA neurons, SLC39A14 patient 1, basal, replicate 1


title: mDA neurons, control 2, basal, replicate 3


title: mDA neurons, control 3, basal, replicate 2


title: mDA neurons, SLC39A8 patient 1, basal, replicate 2


title: Humanised mouse spleen MEKi experiment, HTO

city: Nantes

first: Christine

last: Chauveau

address: 30 Boulevard Jean Monnet

country: France

institute: CR2TI UMR 1064

description:
Hashtag-derived oligonucleotide (HTO)
HTO library: read1 file contains cell barcode and UMI; read2 file contains Hashtag Antibody Derived Tag reads

library_source: other

data_processing:
CellRanger v7.0.0. multi function to aggregate reads from RNA and HTO across sequencing lanes into a single set of  barcodes.tsv.gz, features.tsv.gz and matrix.mtx.gz as well as identify the V(d)J sequences from the TCR library.
Assembly: GrCH38
Supplementary files format and content: CellRanger output files: barcodes.tsv.gz, features.tsv.gz, matrix.mtx.gz, filtered_contig_annotations.csv

molecule: protein

source_name: spleen

extract_protocol:
CD45+ cells were sorted from splenocytes. Single cells were encapsulated in  Gel-Beads-in emulsion (GEMs) using the 10X Controller with a Chromium Single Cell 5’ GEM Library and Gel Bead kit (Garvan Weizmann Centre for Cellular Genomics).
The cell hashing protocol described by Stoeckius et al  (Stoeckius et al.,2017)  was used to generate a hashtag oligo (HTO) library, adapted with a 5’v2 immune profiling protocol with TCR. https://citeseq.files.wordpress.com/2019/02/cite-seq_and_hashing_protocol_190213.pdf
CITE-seq VdJ Immunoprofiling

treatment_protocol: Human skin grafted humanised mice were treated with MEK inhibitor (tra) or not (sham)

value: spleen

value: Human CD45+

value: mixed MEKi treated, non-treated

tag: library type

value: HTO

tag: hto

value: TotalSeq-C0251 anti-human Hashtag 1 , TotalSeq-C0252 anti-human Hashtag 2, TotalSeq™-C0253 anti-human Hashtag 3 , TotalSeq™-C0254 anti-human Hashtag 4


title: mDA neurons, SLC39A14 patient 1, basal, replicate 2

value: Induced pluripotent stem cells

tag: cell type


title: mDA neurons, control 2, basal, replicate 1

value: Midbrain dopaminergic neurons

tag: treatment


title: Humanised mouse spleen MEKi experiment, TCR

description:
V(d)J RNA sequence
TCR library: : read1 file contains cell barcode and UMI; read2 file contains V(d)J reads

molecule: other

value: TCR


title: WT_SBP009_Rep3_WGBS_DNAme

city: Montreal

first: Alain

last: Pacis

state: QC

address: 1010 Sherbrooke #1800

country: Canada

institute: Canadian Centre for Computational Genomics (C3G)

data_processing:
Adaptor sequences and low-quality score bases were first trimmed using Trimmomatic.
The resulting reads were mapped to the human reference genome (GRCh38) and lambda phage genome using Bismark, which uses a bisulfite-converted reference genome for read mapping.
Only reads that had a unique alignment were retained and PCR duplicates were marked using Picard tools.
Methylation levels for each CpG site were estimated by counting the number of sequenced C (‘methylated’ reads) divided by the total number of reported C and T (‘unmethylated’ reads) at the same position of the reference genome using Bismark’s methylation extractor tool.
We performed a strand-independent analysis of CpG methylation where counts from the two Cs in a CpG and its reverse complement (position on the plus strand and position i+1 on the minus strand) were combined and assigned to the position of the C in the plus strand.
To assess MethylC-seq bisulfite conversion rate, the frequency of unconverted cytosines (C basecalls) at lambda phage CpG reference positions was calculated from reads uniquely mapped to the lambda phage reference genome.
CpG sites with at least 5X coverage were included for all downstream analyses.
Assembly: GRCh38
Supplementary files format and content: Processed file contains methylation levels for CpG sites with a coverage >= 5. It includes 12 columns: "chromosome" "start" "end" "mC (+ strand)" "mC+C (+ strand)" "mC/(mC+C) % (+ strand)" "mC (- strand)" "mC+C (- strand)" "mC/(mC+C) % (- strand)" "mC (combined)" "mC+C (combined)" "mC/(mC+C) % (combined)"

source_name: iPSCs were derived from lymphoblasts

growth_protocol: iPSCs were maintained on a Matrigel (Corning, Cat. #08-774-552) coated plate with mTeSR™1 medium (STEMCELL Technologies, Cat. #85870) along with Y-27632 (ROCK inhibitor, STEMCELL Technologies, Cat. # 72307) after each passage. During the course of induction of iPSC to NPCs (20 days), STEMdiff™ SMADi Neural Induction Kit (STEMCELL Technologies, Cat. #08581) was used. The method for induction of iPSC to NPCs was based on the STEMCELL™ neural induction EB-based method and was performed following the manufacturer’s protocol. On Day 0, a single-cell suspension of iPSC was generated using Gentle Cell Dissociation Reagent (STEMCELL Technologies, Cat. # 100-0485), and 10,000 cells per microwell were seeded in an ultralow attachment 96-well plate to generate Embryoid bodies (EBs), cultured in STEMdiff™ Neural Induction Medium + SMADi (STEMdiff™ SMADi Neural Induction Kit, Cat. # 08582) + 10 μM Y-27632. From Day 1 to Day 4, a daily partial (3/4) medium change was performed using STEMdiff™ Neural Induction Medium + SMADi. On Day 5, EBs were harvested using a wild-board 1ml serological pipettes and 40μm strainer and transferred to a single well of a 6-well plate coated with Poly-L-ornithine hydrobromide (PLO) and laminin. A daily full medium change was performed from Day 6 to Day 11. After the neural induction efficiency was determined higher than 75%, neural rosettes were manually selected using STEMdiff™ Neural Rosette Selection Reagent (STEMCELL Technologies, Cat. # 05832) on Day 12 and replated onto a single well of PLO/Laminin coated 6-well plate. With continuous daily full medium change, selected rosette-containing clusters attached, and NPC outgrowths formed a monolayer between the clusters. The NPCs were ready for passage 1 when cultures were approximately 80 - 90% confluent (typically on Day 19). NPCs were maintained and expanded (7-10 days) using DMEM/F12 with Glutamax (ThermoFisher, Cat. #10565042), N2 1x (ThermoFisher, Cat. # 17502001), B27 1x (ThermoFisher, Cat. #17504001), FGF2 20 ng/ml (PEPROTECH, Cat. #100-18B), and EGF 20 ng/ml (Gibco, Cat. # AF-100-15-1MG). For the final differentiation of NPCs into neurons, NPCs were detached using Accutase (STEMCELL Technologies, Cat. #7922) and then differentiated onto PLO/laminin-coated plates with final differentiation media of BrainPhys™ Neuronal Medium (STEMCELL Technologies, Cat. #05790) supplemented with Glutamax 1x (ThermoFisher, Cat. #35050061), N2 1x (ThermoFisher, Cat. #17502001), B27 1x (ThermoFisher, Cat. #17504001), 200nM ascorbic acid 200nM (STEMCELL Technologies, Cat. #72132), cyclic AMP 500 μg/ml (Sigma-Aldrich, Cat #A9501-1G), brain-derived neurotrophic factor 20 ng/ml (BDNF, GIBCO, Cat. #PHC7074), Wnt3a 10 ng/ml (R&D Systems, Cat. #5036-WN-500), and laminin 1 μg/ml (Gibco, Cat. #23017015) for 14 days (medium change of 3 times per week). Subsequently, the cells were maintained in STEMdiff™ Forebrain Neuron Maturation Kit for 11 days (STEMCELL Technologies, Cat. #08605) --final differentiation media was replaced by STEMdiff™ Forebrain Neuron Maturation Kit in the same dishware.

extract_protocol:
DNA was extracted from bulk neuronal culture 25 days post-differentiation using DNeasy Blood & Tissue Kit (Cat. #69504).
Whole genome sequencing libraries were generated from 1000 ng of genomic DNA spiked with 0.1% (w/w) unmethylated λ DNA (Roche Diagnostics) and fragmented to 300–400 bp peak sizes using the Covaris focused-ultrasonicator E210. Fragment size was controlled on a Bioanalyzer High Sensitivity DNA Chip (Agilent) and NxSeq AmpFREE Low DNA Library Kit (Lucigen) was applied. End repair of the generated dsDNA with 3′ or 5′ overhangs, adenylation of 3′ ends, adaptor ligation, and clean-up steps were carried out as per Lucigen’s recommendations. The cleaned-up ligation product was then analyzed using LabChip GX/GX II (Caliper). Samples were then bisulfite converted using the EZ-DNA Methylation Gold Kit (Zymo Research) according to the manufacturer’s protocol. DNA was amplified by 9 cycles of PCR using the Kapa HiFi Uracil+ Kit (Roche) DNA polymerase (KAPA Biosystems) according to the manufacturer’s protocol. The amplified libraries were purified using Ampure XP Beads (Beckman Coulter), validated on LabChip GX/GX II (Caliper), and quantified by qPCR amplification with Illumina adapters. The values are read using Lightcycler LC480 (Roche).

treatment_protocol: For Het-AKAP11-KO clones only: To generate three isogenic knockout clones from the WT iPSC (SBP009), Ribonucleoprotein (RNP)-mediated CRISPR-Cas9 genome editing was performed using the Alt-R CRISPR-Cas9 system (IDT). Two synthetic crRNA guides were designed (TACTGGTATAAGGTACACCT PAM:TGG and CTGATGCAAGAATGTGTCCA PAM:AGG) to form a duplex with Alt-R® CRISPR-Cas9 tracrRNA, ATTO 550 (IDT, Cat. # 1075928) and coupled to the Alt-R© S.p. Cas9 Nuclease V3 (IDT). The Lipofectamine™ Stem Transfection (Invitrogen, Cat. #STEM00001) and PLUS™ Reagent (Invitrogen, Cat. #11514015) ratios were optimized with a lower volume than the manufacturer’s protocol. The transfected cells were incubated for 48 hours. Single ATTO550+ cells were then sorted into 96-well plates.

value: iPSCs were derived from lymphoblasts

value: neurons

value: WT


title: mDA neurons, SLC30A10 patient 2, basal, replicate 3

value: Day 65 of differentiation

value: SLC30A10 mutant 2


title: mDA neurons, control 1, basal, replicate 2


title: WT_SBP009_Rep1_WGBS_DNAme


title: WT_SBP009_Rep3

data_processing:
Adaptor sequences and low-quality score bases (Phred score < 30) were first trimmed using Trimmomatic.
The resulting reads were aligned to the GRCh38 human reference genome assembly, using STAR.
Read counts were obtained using HTSeq with parameters-m intersection-nonempty-stranded=reverse.
For all downstream analyses, we excluded lowly expressed genes with an average read count lower than 20 across all samples, resulting in a total of 23,590 genes.
Raw counts were normalized using edgeR’s TMM algorithm and were then transformed to log2-counts per million (logCPM) using the voom function implemented in the limma R package.
Assembly: GRCh38
Supplementary files format and content: RNA-seq raw counts matrix is a tab-delimited text file, with the genes in rows and the samples in columns.

extract_protocol:
RNA was extracted from bulk neuronal culture 25 days post-differentiation using RNeasy Mini Kit and its protocol (Cat. #74104).
Library preparation from total RNA was performed using NEB rRNA-depleted (HMR) stranded library preparation kit according to the manufacturer’s instructions


title: mDA neurons, control 1, basal, replicate 3


title: Het-AKAP11-KO-Clone 21_WGBS_DNAme


title: Het_AKAP11-KO-Clone 21


title: Humanised mouse spleen MEKi experiment, GEX

description:
polyA RNA
GEX (3' mRNA) library: read1 file contains cell barcode and UMI; read2 file contains transcript

molecule: polyA RNA

value: mRNA


title: Het-AKAP11-KO-Clone 16_WGBS_DNAme


title: Het_AKAP11-KO-Clone 16


title: WT_SBP009_Rep1


title: WT_SBP009_Rep2


title: Het_AKAP11-KO-Clone 16_Input

data_processing:
ChIP-seq reads were first trimmed for adapter sequences and low-quality score bases using Trimmomatic.
The resulting reads were mapped to the human reference genome (GRCh38) using BWA-MEM48 in paired-end mode at default parameters (-k 19 -w 100 -d 100).
Only reads that had a unique alignment (mapping quality > 20) were retained and PCR duplicates were marked using Picard tools.
Peaks were called using MACS2.
Bigwig files were generated using HOMER's makeUCSCfile then UCSC's bedGraphToBigWig tool.
Assembly: GRCh38
Supplementary files format and content: Processed files contains MACS2 output, which includes 10 columns: "chromosome" "start" "end" "name" "score" "strand" "signalValue" "pValue" "qValue" "peak"

extract_protocol:
About 10 million neuronal cells per sample were grown and directly crosslinked on the plate with 1% formaldehyde (Sigma) for 10 minutes at room temperature and the reaction was stopped using 125nM Glycine for 5 minutes. Fixed cell preparations were washed with ice-cold PBS, scraped off the plate, pelleted, washed twice again in ice-cold PBS, and the flash-frozen pellets were stored at −80°C.
Library preparation was carried out using Kapa Hyper Prep library preparation reagents (Kapa Hyper Prep kit, Roche 07962363001) following the manufacturer’s protocol.


title: Het_AKAP11-KO-Clone 20


title: Het_AKAP11-KO-Clone 20_Input


title: Het_AKAP11-KO-Clone 21_Input


title: WT_SBP009_Rep3_ChIP

value: rabbit polyclonal anti-H3K27ac Diagenode Cat. #C15410196, RRID:AB_2637079


title: Het-AKAP11-KO-Clone 20_WGBS_DNAme


title: mDA neurons, control 1, basal, replicate 1

value: Healthy control


title: WT_SBP009_Rep1_ChIP


title: Het_AKAP11-KO-Clone 21_ChIP


title: WT_SBP009_Rep2_ChIP


title: Het_AKAP11-KO-Clone 20_ChIP


title: WT_SBP009_Rep2_WGBS_DNAme


title: Het_AKAP11-KO-Clone 16_ChIP


title: EMseq_501mel shNIPBL_#2 6D rep2

city: New York

first: Elissa

middle: WP

last: Wong

state: NY

address: 1275 York Avenue

department: HOPP

institute: Memorial Sloan Kettering Cancer Center

data_processing:
EM-sequencing reads were processed using Bismark pipeline71. Raw reads with low-quality (less than 20) and adapter sequences were removed by Trim Galore (v0.6.4).
C(G) in the trimmed sequences was converted to T(A) and mapped to similarly converted reference human genome (hg19), lambda, and pUC19 separately using default Bowtie2 settings implemented in Bismark. Duplicated reads were discarded.
The remaining alignments were then used for cytosine methylation calling by Bismark methylation extractor. Batch effects were adjusted using limma (v3.46.0). Any CpGs with coverage less than 10 were removed prior downstream analyses.
Assembly: hg19
Supplementary files format and content: Coverage and methylation bigwig

source_name: skin

growth_protocol: All cells lines were cultured in media supplemented with 10% FBS, 2 mM L-glutamine, 100 U/ml penicillin and 100 µg/ml streptomycin. All cells were cultured at 37 ºC in 5% CO2.

extract_protocol:
High-molecular-weight genomic DNA was extracted using Puregene cell core kit (Qiagen).
EM-seq was performed with 100ng of purified genomic DNA using NEBNext® Enzymatic Methyl-seq Kit (NEB, E7120) and following manufacturer’s instructions.
After quantification and size distribution assessment using a Qubit Flex fluorometer (Thermo Fisher Scientific) and a TapeStation (Agilent), respectively, the purified libraries were sequenced on an Illumina NovaSeq 6000 to generate around 200 M PE100 reads per library.

treatment_protocol: Cells that were transduced with lentivirus carrying puromycin-resistant shRNA plasmids were treated with 2 µg/ml puromycin.

value: skin

value: melanoma


title: EMseq_501mel shNIPBL_#1 6D rep1


title: EMseq_501mel shLuc 6D rep2

value: shLuc


title: EMseq_501mel shLuc 6D rep1


title: ChIPseq_501mel shNIPBL_#2 6D MITF

data_processing:
ChIP-seq reads were aligned to hg19 using BWA (v0.7.17-r1188). Unmapped and low MAPQ reads (< 30) were filtered out using SAMtools (v1.9) and PCR duplicates were removed using Picard tools (v2.18.16). Depth normalized bigWig tracks were generated using deepTools (v3.1.1) bamCoverage with `bamCoverage -b ${input_bam} -o ${output_bigwig} --normalizeUsing RPGC --ignoreForNormalization chrX chrM -bl hg19-blacklist.v2.bed`.
Assembly: hg19
Supplementary files format and content: bigWig

extract_protocol:
20 x 106 cells were harvested and fixed in 1% formaldehyde for 10 min with gentle shaking. Glycine was added to final concentration of 125 mM and incubated for 5 min. Cells were washed once with PBS, resuspended in LB1 [50mM HEPES (pH 7.5), 140mM NaCl, 1mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100 and 1× complete EDTA-free protease inhibitor] and incubated with rotation at 4 °C for 10min. Samples were pelleted, resuspended in LB2 [10mM Tris-HCl (pH 8.0), 200mM NaCl, 1mM EDTA, 0.5mM EGTA and 1× complete protease inhibitor] and incubated rotating at 4 °C for 10min. Nuclei were pelleted, resuspended in LB3 with 0.5% SDS [0.5% SDS, 20mM Tris-HCl (pH 8.0), 150mM NaCl, 2mM EDTA, 1% Triton X-100 and 1× complete protease inhibitor] and sonicated using Covaris E220 focused ultrasonicator (peak power = 150 W, duty factor = 2%, cycle per burst = 200) for 12 min. Soluble chromatin extracts were cleared by centrifugation and 1% was kept as input DNA. Sheared chromatin was diluted to contain 0.16% SDS for antibody binding and incubated overnight at 4 °C with 2 ug anti-SMC1A (Bethyl Laboratories Inc, A300-055A), anti-MITF (Active Motif, 91201) or anti-CTCF (Cell Signlaing Technology, 3418S). Spike-in chromatin (Active motif, 53083) and antibody (Active motif, 61686) were included to normalize SMC1A ChIP-seq.  20 ul Pierce ChIP-grade magnetic beads were used for precipitation at 4 °C for 6 hr. Beads were washed 3x LiCl buffer [250 mM LiCl, 10 mM Tris-HCl (pH 8.0), 1% sodium deoxycholate, 1% NP-40 and 1 mM EDTA], 1x RIPA wash buffer [500 mM LiCl, 50 mM HEPES-KOH (pH 7.6), 0.7% sodium deoxycholate, 1% NP-40 and 1 mM EDTA] and 1x TE [10 mM Tris-HCl (pH 8.0) and 1 mM EDTA] + 50 mM NaCl. Chromatin was eluted in elution buffer [1% SDS, 50 mM Tris-HCl (pH 8.0) and 10 mM EDTA] and incubated at 65 °C for 30 min. Eluate was reverse cross-linked at 65 °C overnight, treated with RNaseA (Roche) at 37 °C for 1 hr and Proteinase K (Roche) at 55 °C for 2 hr before recovered using a miniElute PCR purification kit from Qiagen.
Library was prepared using KAPA HTP library kit (KAPA biosystems).
ChIP libraries were sequenced to obtain 30-40 M reads for each sample on NovaSeq 6000 with PE50.

value: shNIPBL_#2, 6D

value: MITF antibody


title: ChIPseq_501mel shLuc 6D MITF

value: shLuc, 6D


title: ChIPseq_501mel shNIPBL_#2 6D CTCF

value: CTCF antibody


title: WT_SBP009_Input


title: ChIPseq_501mel shNIPBL_#1 6D CTCF

value: shNIPBL_#1, 6D


title: ChIPseq_501mel shLuc 6D CTCF


title: CAGEseq_HCT116 shNIPBL_#2 5D rep1

data_processing:
CAGE-seq reads were analyzed based on the Nextflow CAGE-seq analysis pipeline.
Index sequences, EcoP15I recognition site and the first G following the EcoP15I site at the 5’-end from reads were removed using Cutadapt (v2.6).
rRNA sequences were filtered out using SortMeRNA (v4.2.0) and filtered reads were aligned to the hg19 genome using STAR (v2.7.1a).
At this step, two sets of BAM files were created for each sample: one with uniquely mapped reads (adding --outFilterMultimapNmax 1 to STAR command) and one with multi-mapped reads (adding --outFilterMultimapNmax 10 to STAR command). Unmapped reads from BAM files were removed.
BAM files were converted to BED files with the summed up 1 bp unclustered CAGE tags using SAMtools (v1.9) and bamtobed (v.2.26.0). We pooled CAGE tags across samples, filtered them by TPM threshold of 0.5 and then clustered them with paraclu to generate a BED file with the clustered CAGE transcription start sites (CTSSs). CAGE tags were intersected with the clusters identified by paraclu and a raw count table with columns for each sample and rows for each tag cluster was generated. DESeq2 (v1.30.1) was used to perform differential CAGE expression using the raw count table. CTSS clusters were annotated using R/Bioconductor packages ChIPseeker (v1.26.2) and AnnotationDbi (v1.52) using GENCODE (v19) gene annotations.
Assembly: hg19
Supplementary files format and content: Differential expression analysis by DESeq2

source_name: Bowel

extract_protocol:
Cells were lyzed directly on culture dish with TRIzol RNA isolation reagent (Thermo fisher scientific). Total RNA was extracted by thorough mixing with chloroform and isopropanol precipitation of upper aqueous phase. 5 µg total RNA was reverse transcripted using a primer that carried a 15-nt random region and EcoP15I recognition sequence and SuperScriptIII RNase H- reverse transcriptase (Thermo fisher scientific) in the presence of trehalose/sorbitol to increase efficiency of reverse transcription. Diol groups on RNA/cDNA duplex were oxidated and labelled with biotin hydrazide (Vector laboratories). RNase I (Promega) treatment was performed to cleave single-stranded RNA regions that were not hybridized with first-strand cDNA. tRNA-coated streptavidin beads (Thermo fisher scientific) were used to capture biotinylated RNA/cDNA duplex. Sample was washed three times with wash buffer A [4.5 M NaCl, 50 mM EDTA (pH 8.0) and 0.1 % Tween20], once with wash buffer B [10 mM Tris–HCl (pH 8.5), 1 mM EDTA (pH 8.0), 0.5 M NaOAc (pH 6.1) and 0.1 % Tween20] and once with wash buffer C [0.3 M NaCl, 1 mM EDTA (pH 8.0) and 0.1 % Tween20]. Single-stranded cDNAs were released from streptavidin beads by heating at 95 °C for 5 min. Released cDNA was treated with RNase H (Thermo fisher scientific) and RNase I (Promega) at 37 °C for 45 min and proceed for library construction.
Library was constructed with modified primers for compatible sequencing on Illumina NextSeq/NovaSeq sequencer. Ligation of bar-coded 5’ linker was performed. Second-strand cDNA was synthesized using Deep Vent (exo-) DNA polymerase (NEB) and biotinylated second-strand primer. cDNA was treated with shrimp alkaline phosphatase (Thermo fisher scientific) to remove phosphate group from 5’ linker. EcoP15I (NEB) was used to cleave 27 nt downstream of the recognition site at the 5’ linker. 3’ linker was ligated to the 27-nt-long tags, purified with tRNA-coated streptavidin beads and amplified with 2x Phusion high-fidelity PCR mix (NEB). CAGE libraries were treated with Exonuclease I (NEB) to degrade any single-stranded DNA before sequencing.
Libraries were sequenced on NextSeq/NovaSeq using SR50 to obtain around 40 M reads for each library.

treatment_protocol: Cells that were transduced with lentivirus carrying puromycin-resistant shRNA plasmids were treated with 0.5-2 µg/ml puromycin.

value: Bowel

value: Colorectal cancer


title: CAGEseq_HCT116 shLuc 5D rep2


title: ChIPseq_501mel shNIPBL_#1 6D SMC1A spike-in

value: SMC1A antibody


title: CAGEseq_HCT116 shLuc 5D rep1


title: ChIPseq_501mel shLuc 6D SMC1A spike-in


title: ChIPseq_501mel shNIPBL_#2 6D SMC1A spike-in


title: CAGEseq_H2228 shNIPBL_#2 5D rep2

source_name: Lung

value: Lung

value: Lung cancer


title: CAGEseq_A549 shLuc 5D rep2


title: CAGEseq_A549 shNIPBL_#2 5D rep2


title: CAGEseq_A549 shNIPBL_#2 5D rep1


title: CAGEseq_H2228 shLuc 5D rep1


title: CAGEseq_H2228 shNIPBL_#2 5D rep1


title: CAGEseq_A549 shLuc 5D rep1


title: EMseq_501mel shNIPBL_#2 6D rep1


title: EMseq_501mel shNIPBL_#1 6D rep2


title: AS9_rep2

city: Nashville

first: Lindsey

middle: "Joey"

last: Stolze

state: TN

address: 1211 Medical Center Dr

institute: Vanderbilt University Medical Center

description: 10X Genomics Multiome Kit

data_processing:
Postprocessing and analysis were performed on the Terra Platform (app.terra.bio) using Scanpy v1.9.3. Samples were inspected individually for mapping quality based on the number of mapped reads per nucleus, percentage of mapped mitochondrial reads, and shape of the unique molecular identifier (UMI) decay curve, of which all samples demonstrated adequate sequencing quality. To identify nuclei with high levels of spliced reads indicative of increased cytosolic fragment contamination, we used Scrinvex (v13, https://github.com/getzlab/scrinvex) to assign reads to exonic and intronic regions. We then calculated the ratio of exonic reads to the total number of mapped reads within each sample. Nuclei with an exon ratio greater than the 75th percentile + IQR range were removed from downstream analyses. Doublets were detected using Scrublet, a threshold of 0.25 was set based on the simulated distribution, and nuclei with a doublet score >0.25 were removed from the analysis. Finally, nuclei with reads mapped to fewer than 200 genes and nuclei with more than 5% of reads mapped to mitochondrial genes were removed. Counts were normalized to 10,000 unique molecules per nucleus and log-normalized. Highly variable genes were identified for clustering (minimum mean 0.0125, maximum mean 3, minimum dispersion 0.5, 4,283 genes). The effects of the total read count and the percentage of mitochondrial reads were then regressed out using default parameters (Scanpy preprocessing regress_out), and the data were scaled to a maximum value of 10. Principal components (PCs) were calculated for highly variable genes (scanpy.tl.pca(adata, svd_solver='arpack')) and corrected for batch effects using Harmonypy (version 0.0.9), with default parameters considering each sample as a unique batch. Batch-corrected PCs were used to calculate the nearest neighbors (scanpy.pp.neighbors(adata, n_neighbors=10, n_pcs=40)) and generate a UMAP representation of the data for two dimensional visualization purposes (scanpy.tl.umap(adata)). Graph-based clustering was performed using Leiden algorithm (scanpy.tl.leiden(adata)) at a basal resolution of 0.2.
Assembly: GRCh38
Supplementary files format and content: Seurat object in RDS format

molecule: nuclear RNA

source_name: human heart

extract_protocol:
Myocardial tissue was obtained with a scalpel from the basal septum of the left ventricle from patients (N=11) undergoing surgical aortic valve replacement for AS or unmatched donor hearts from 2011-2012 (N=9; hearts not used for transplantation) and immediately snap frozen in liquid nitrogen. The tissue was subsequently stored in a -80°C freezer until being used for analysis. Briefly, frozen ventricular tissue was mounted onto a CryoStat using Optimal Cutting Temperature (OCT) compound, followed by 100 µm sectioning. OCT was removed, and sections were placed in cold lysis buffer, followed by gentle dounce homogenization to release intact nuclei. After pelleting at 40g for 1 minute to remove large debris, the suspension underwent sequential filtering using 100 µm and 20 µm filters. The filtered suspension was centrifuged at 550g for 5 minutes, washed in a nuclear wash buffer, and recentrifuged at 550g for 5 minutes. The nuclear wash buffer was removed via aspiration and the nuclei were resuspended in cold nuclear resuspension buffer containing RNase inhibitors. Nuclei were stained with trypan blue and counted using a hemocytometer. Nuclei were loaded into a 10X Genomics microfluidic platform (Chromium Next GEM Single Cell Multiome ATAC + Gene Expression), targeting an estimated recovery of 8,000 nuclei per sample. Libraries were processed according to the manufacturer’s instructions.
The sequencing files were de-multiplexed using Cellranger-ARC mkfastq (v2.0.0). The RNA reads were then trimmed using Cutadapt (v2.8) with default parameters to remove homopolymers (A30, T30, G30, and C30) and the template switch oligo (CCCATGTACTCTGCGTTGATACCACTGCTT) and its complement (AAGCAGTGGTATCAACGCAGA GTACATGGG). To generate an RNA count matrix, Cellranger (v7.1.0) count was used with default settings to align the trimmed reads to the human genome (GRCh38) and CellBender (v2.0) was used with default settings to filter samples for ambient RNA byproducts and call valid cell barcodes.
Single-nuclear RNA and ATAC seq (10X Multiome kit)

treatment_protocol: Control vs Aortic Stenosis

value: human heart

value: aortic stenosis


title: ChIPseq_501mel shNIPBL_#1 6D MITF


title: CAGEseq_H2228 shLuc 5D rep2


title: AS7_rep2

description: 10X Genomics Multiome Kit


title: AS8_rep1

description: 10X Genomics Multiome Kit


title: AS6_rep2

description: 10X Genomics Multiome Kit


title: AS6_rep1

description: 10X Genomics Multiome Kit


title: AS7_rep1

description: 10X Genomics Multiome Kit


title: AS9_rep1

description: 10X Genomics Multiome Kit


title: AS5_rep2

description: 10X Genomics Multiome Kit


title: AS4_rep2

description: 10X Genomics Multiome Kit


title: CAGEseq_HCT116 shNIPBL_#2 5D rep2


title: AS5_rep1

description: 10X Genomics Multiome Kit


title: AS4_rep1

description: 10X Genomics Multiome Kit


title: AS3_rep1

description: 10X Genomics Multiome Kit


title: AS2_rep2

description: 10X Genomics Multiome Kit


title: AS3_rep2

description: 10X Genomics Multiome Kit


title: AS2_rep1

description: 10X Genomics Multiome Kit


title: AS1_rep1

description: 10X Genomics Multiome Kit


title: AS11_rep2

description: 10X Genomics Multiome Kit


title: AS11_rep1

description: 10X Genomics Multiome Kit


title: S16_014_rep3

description: 10X Genomics Multiome Kit

value: control


title: S16_014_rep4

description: 10X Genomics Multiome Kit


title: AS1_rep2

description: 10X Genomics Multiome Kit


title: S16_013_rep4

description: 10X Genomics Multiome Kit


title: S16_014_rep1

description: 10X Genomics Multiome Kit


title: AS8_rep2

description: 10X Genomics Multiome Kit


title: S16_013_rep3

description: 10X Genomics Multiome Kit


title: S16_013_rep2

description: 10X Genomics Multiome Kit


title: AS10_rep1

description: 10X Genomics Multiome Kit


title: AS10_rep2

description: 10X Genomics Multiome Kit


title: S14_012_rep3

description: 10X Genomics Multiome Kit


title: S14_012_rep4

description: 10X Genomics Multiome Kit


title: S14_012_rep2

description: 10X Genomics Multiome Kit


title: S12_010_rep3

description: 10X Genomics Multiome Kit


title: S14_012_rep1

description: 10X Genomics Multiome Kit


title: S12_010_rep2

description: 10X Genomics Multiome Kit


title: S12_010_rep4

description: 10X Genomics Multiome Kit


title: S12_008_rep3

description: 10X Genomics Multiome Kit


title: S12_008_rep1

description: 10X Genomics Multiome Kit


title: S12_008_rep2

description: 10X Genomics Multiome Kit


title: S12_008_rep4

description: 10X Genomics Multiome Kit


title: S12_007_rep4

description: 10X Genomics Multiome Kit


title: S12_007_rep2

description: 10X Genomics Multiome Kit


title: S12_007_rep1

description: 10X Genomics Multiome Kit


title: S12_007_rep3

description: 10X Genomics Multiome Kit


title: S12_003_rep3

description: 10X Genomics Multiome Kit


title: S16_014_rep2

description: 10X Genomics Multiome Kit


title: S12_003_rep1

description: 10X Genomics Multiome Kit


title: S12_003_rep2

description: 10X Genomics Multiome Kit


title: S16_013_rep1

description: 10X Genomics Multiome Kit


title: S11_001_rep4

description: 10X Genomics Multiome Kit


title: S11_002_rep2

description: 10X Genomics Multiome Kit


title: S11_002_rep3

description: 10X Genomics Multiome Kit


title: S11_001_rep3

description: 10X Genomics Multiome Kit


title: S11_002_rep1

description: 10X Genomics Multiome Kit


title: S11_001_rep1

description: 10X Genomics Multiome Kit


title: S11_001_rep2

description: 10X Genomics Multiome Kit


title: testis with normal spermatogenesis_3 (characterization of the hTP cells)

city: Bergen

first: Tomasz

last: Stokowy

address: Nygardsgaten 5

department: IT Division

country: Norway

institute: University of Bergen

description: hTestis 27

data_processing:
Alignment to reference genome GRCh38
Counting of reads per gene using FeatureCounts
Normalisation using DESeq2
Assembly: GRCh38
Supplementary files format and content: Counts_gencode_hTPc_Testis_v26_240324.txt, read counts per gene
Supplementary files format and content: Normalized_RNA_Testis_240324.txt, normalized gene expression

source_name: testis

extract_protocol:
The cell samples were lysed in 350 µl of RLT buffer with β-mercaptoethanol and RNA, DNA and protein were extracted according to the manufacturer’s protocol AllPrep DNA/RNA/Protein Mini Kit (Qiagen, Hilden, Germany). The RNA samples were additionally purified according to the manufacturer’s protocol of Turbo DNA-free Kit (Thermo Fisher Scientific, MT, USA).
Sequencing libraries of cell samples were prepared using the the TruSeq Stranded Total RNA with Ribo-Zero Human (Illumina) and sequenced on an Illumina NovaSeq6000  platform
Paired end 150bp sequencing

value: testis

tag: group

value: testis with normal spermatogenesis

value: no modification


title: testis with normal spermatogenesis_1 (characterization of the hTP cells)

description: hTestis 20


title: testis with normal spermatogenesis_2 (characterization of the hTP cells)

description: hTestis 26


title: human testicular primary cells_5 (characterization of the hTP cells)

description: hTPc 19105/4

source_name: human testicular primary cells

value: human testicular primary cells

value: native human testicular primary cells


title: human testicular primary cells_6 (characterization of the hTP cells)

description: hTPc 19105/5


title: human testicular primary cells_3 (characterization of the hTP cells)

description: hTPcIM 19105/6


title: S12_010_rep1

description: 10X Genomics Multiome Kit


title: human testicular primary cells_4 (characterization of the hTP cells)

description: hTPc 19105/3


title: human testicular primary cells_2 (characterization of the hTP cells)

description: hTPcIM 19105/4


title: human testicular primary cells_1  (characterization of the hTP cells)

description: hTPcIM 19105/2


title: S12_003_rep4

description: 10X Genomics Multiome Kit


title: hTPc wild type_3

data_processing:
Alignment to reference genome GRCh38
Counting of reads per gene using FeatureCounts
Normalisation using DESeq2
Assembly: GRCh38
Supplementary files format and content: Counts_genocode_hTPc_WT_GFP_TKTL_v26_240324.txt, read counts per gene
Supplementary files format and content: Normalized_RNA_hTPc_WT_GFP_TKTL_240324.txt, normalized gene expression

value: WT


title: hTPc wild type_2


title: hTPc negative control with GFP expression_3

value: negative control

value: GFP expression


title: hTPc wild type_1


title: hTPc negative control with GFP expression_2


title: hTPc negative control with GFP expression_1


title: hTPc with overexpression of TKTL1 gene_3

value: TKTL1 overexpression gene activation

value: overexpression of TKTL1 gene


title: S11_002_rep4

description: 10X Genomics Multiome Kit


title: hTPc with overexpression of TKTL1 gene_1


title: SHAPE_COX1_P4_DC_B

description:
COX1 cellular RNA
Shapemapper2 was used for reactivity profiling

value: HEK293T cells

value: No


title: SHAPE_COX1_P4_no_1M7_B


title: hTPc with overexpression of TKTL1 gene_2


title: SHAPE_COX1_P4_2-NH2_B


title: SHAPE_COX1_P4_1M7_B


title: SHAPE_COX1_P4_DC_A


title: SHAPE_COX1_P4_no_1M7_A


title: SHAPE_COX1_P4_1-NH2_B


title: SHAPE_COX1_P4_2-NH2_A


title: SHAPE_COX1_P4_1M7_A


title: SHAPE_COX1_P3_DC_B


title: SHAPE_COX1_P3_2-NH2_B


title: SHAPE_COX1_P3_no_1M7_B


title: SHAPE_COX1_P3_1-NH2_B


title: SHAPE_COX1_P3_1M7_B


title: SHAPE_COX1_P3_DC_A


title: SHAPE_COX1_P3_2-NH2_A


title: SHAPE_COX1_P3_no_1M7_A


title: SHAPE_COX1_P3_1-NH2_A


title: SHAPE_COX1_P1_no_1M7_B


title: SHAPE_COX1_P1_1-NH2_B


title: SHAPE_COX1_P3_1M7_A


title: SHAPE_COX1_P1_2-NH2_B


title: SHAPE_COX1_P1_1M7_B


title: PBMC, scCLEAN, sequencing replicates 1..3

city: New Orleans

first: Amitabh

middle: C

last: Pandey

state: LA

address: 1430 Tulane Ave

department: Medicine, Section of Cardiology

institute: Tulane University School of Medicine

description:
Library name: 1335_2_B01_untreated
10x Genomics

library_source: transcriptomic single cell

data_processing:
The demultiplexing, UMI counting, and aggregations were performed with CellRanger v6.0.0 for Illumina sequenced data and pigeon v1.3.0 was used for PacBio sequenced data.
Assembly: GRCh38; GENCODE v32
Supplementary files format and content: tarball for each dataset; h5 files for Illumina processed samples and tab separated and matrix files for PacBio processed samples

source_name: PBMC

extract_protocol:
single cell supsension was achieved and these cells were then ready for the library construct. Cell viablity was assessed using Countess system.
Chromium Next GEM Single Cell 3ʹ Kit v3.1

value: PBMC

value: immune

value: polyA RNA


title: nuclei from cells treated with Ad-Dam-3HA-eGFP, MNase rep 2

city: Bethesda

first: David

middle: J.

last: Clark

state: MD

address: 6 Center Drive

department: NICHD

institute: NIH

description: Library name: MCF7_MNase_P4

data_processing:
The pair-end reads aligned to the reference genome using bowtie2 (2.5.1).
We filtered the single-nucleosome fragments with read fragment length between 120 bp and 180 bp, and counted the dyads (center of the read fragments) at each genome location.
The dyad count is stored in the bigwig files.
Assembly: hg38
Supplementary files format and content: bigwig files containing the dyad count in each genome location

source_name: Breast; Mammary gland

extract_protocol:
Nuclei from MCF7 cells (treated with Ad-Dam-3HA-eGFP for 72h) were treated with MNase, digested DNA was purified using the Purelink genomic DNA kit (K182002) from Invitrogen and analysed on agarose gel. For accurate nucleosome mapping we chose digests with the only major band at ~150 bp.
Paired-end libraries were prepared using the New England Biolabs kits and protocols.
Ocampo et al., 2016, Nucleic Acids Res. 2016 Jun 2;44(10):4625-35.

value: Breast; Mammary gland

value: Epithelial cell


title: SHAPE_COX1_P1_DC_A

data_processing:
Reads were filtered for unique sequences using FastUniq
Reads were trimmed using cutadapt (--minimum-length=20)
reads were aligned to the GRCh38 genome using HISAT-3N aligned with default parameters.
Reads were processed using Picard tools, including FixMateInformation, SortSam and BuildBamIndex
Reads were filtered for those that aligned uniquely (flag: 83/163, 99/147), with MAPQ ≥ 2, and without insertions.
HTSeq-count was used to identify mapped reads in UCSC transcripts (union mode)
T-to-C mutations were identified using Rsamtools and a custom R script (available on Bitbucket). Only mutations at positions with a base quality score of greater than 40 that were at least three nt from the end of the read were counted. Reads were excluded where there were greater than five T-to-C mutations and these mutations did not account for at least one third of the observed mutations (NM tag).
SNP sites were filtered from our data in two ways: 1, bcftools was used to identify T-to-C SNP sites in control samples, and these sites were excluded from analysis; 2, sites exhibiting high T-to-C mutation rates in non-s4U treated controls were excluded from analysis.
To examine the distribution of reads with each minimum number of T-to-C mutations, bam files were filtered using Picard tools.
STAR aligner was used to make genome-coverage tracks (inputAlignmentsFromBam mode, outWigType bedGraph).
Tracks were converted to binary format using IGVtools (toTDF).
Assembly: BDGP6 or GRCh38
U reads were filtered based on read count (reads >= 25), prior to HDProbe analysis
Supplementary files format and content: Tracks are provided in tdf format to visualize enrichment levels for controls and T-to-C mutation counts for all Tether-seq samples.

extract_protocol:
RNA was harvested under non-denaturing conditions using a hypotonic lysis buffer (HLB). The pelleted cells were resuspended in HLB (20 mM HEPES, pH 8, 2 mM MgCl2, 0.2 mM EGTA, 10% glycerol, fresh 1 mM DTT) and freeze-thawed 3x and then placed on ice. After adding NaCl to 400 mM, samples were iced for 5 min and then centrifuged for 15 min at 4°C.The supernatant was collected, treated with proteinase K and 1% SDS and incubated at 37°C for 45 min. RNA was extracted 2x with phenol:chloroform:isoamyl (24:24:1) and 2x with chloroform. The aqueous layer was treated with Turbo DNAse and then buffer exchanged into folding buffer.
RNA was reverse transcribed by first annealing to the RT primer (2 pmol) by heating at 65°C for 5 minutes and then cooling on ice. The RT samples (20 μL) were then prepared by adding 0.4 volumes of freshly prepared 2.5x MaP buffer (125 mM Tris-HCl, pH 8, 187.5 mM KCl, 15 mM MnCl2, 25 mM DTT, 1.25 mM dNTPs) and 1 μL of SuperScript II and incubating at 42°C for 3 hours. The enzyme was heat-inactivated by heating at 70°C for 15 minutes and the cDNA product was purified with 3 volumes of SPRI beads. The RT cDNA was amplified using Phusion polymerase and 10 pmols of PCR primers and purified with 3 volumes of SPRI beads. The PCR product was further amplified using 10 pmols of primers containing sequencing indexes and isolated using 1.5 volumes of SPRI beads.


title: SHAPE_COX1_P1_no_1M7_A


title: nuclei from cells treated with Ad-Dam-3HA-eGFP, MNase rep 1

description: Library name: MCF7_MNase_P3


title: PBMC, 10xv3, sequencing replicates 1..3

description:
Library name: 1335_1_A01_treated
10x Genomics


title: wild type nuclei, 6.25 nM Dam, No spermidine, rep 2

description: Library name: MCF7_H45_damM_200U

data_processing:
The pair-end reads aligned to the reference genome using bowtie2 (2.5.1), the read ends and occupancy is calculated using bedtools (2.31.1).
The end counts and occupancy is further processed to calculate the methylated fraction of GATC sites at each genome location using the snakemake workflow snakemakeMethylFrac, and the methylated fractions are stored in the bigwig files.
The snakemake protocol for data processing is avaiable at https://github.com/zhuweix/snakemakeMethylFrac.
Assembly: hg38 and T2T chm13v1.1, as shown in the file name of the processed files.
Supplementary files format and content: bigwig files of the methylated fraction of GATC sites in each genome location.
Library strategy: qDA-seq (Prajapati et al. (2024) NSMB

extract_protocol:
Genomic DNA was purified from Human MCF7 nuclei by using Purelink genomic DNA kit (K182002) from invitrogen.
Genomic DNA was digested with excess DpnI and sonicated to ~300 bp fragments. Paired-end libraries were prepared using the New England Biolabs kits and protocols.


title: SHAPE_COX1_P1_1-NH2_A


title: SHAPE_COX1_P1_2-NH2_A


title: SHAPE_COX1_P4_1-NH2_A


title: SHAPE_COX1_P1_1M7_A


title: wild type nuclei, 3.12 nM Dam, No spermidine, rep 2

description: Library name: MCF7_H45_damM_100U


title: wild type nuclei, 1.56 nM Dam, No spermidine, rep 2

description: Library name: MCF7_H45_damM_50U


title: SHAPE_COX1_P2_DC_B


title: wild type nuclei, No Dam, No spermidine, rep 2

description: Library name: MCF7_H45_damM_0U


title: SHAPE_COX1_P2_no_1M7_B


title: SHAPE_COX1_P2_1-NH2_B


title: SHAPE_COX1_P1_DC_B


title: wild type nuclei, 6.25 nM Dam, No spermidine, rep 1

description: Library name: MCF7_H44_damM_200U


title: wild type nuclei, 3.12 nM Dam, No spermidine, rep 1

description: Library name: MCF7_H44_damM_100U


title: wild type nuclei, 1.56 nM Dam, No spermidine, rep 1

description: Library name: MCF7_H44_damM_50U


title: SHAPE_COX1_P2_1M7_B


title: wild type nuclei, 0.78 nM Dam, No spermidine, rep 1

description: Library name: MCF7_H44_damM_25U


title: SHAPE_COX1_P2_DC_A


title: SHAPE_COX1_P2_no_1M7_A


title: wild type nuclei, MNase rep 1

description: Library name: MCF7_MNase_P1

extract_protocol:
MCF7 nuclei were treated with MNase, digested DNA was purified using the Purelink genomic DNA kit (K182002) from Invitrogen and analysed on agarose gel. For accurate nucleosome mapping we chose digests with the only major band at ~150 bp.
Paired-end libraries were prepared using the New England Biolabs kits and protocols.
Ocampo et al., 2016, Nucleic Acids Res. 2016 Jun 2;44(10):4625-35.


title: SHAPE_COX1_P2_2-NH2_A


title: wild type nuclei, MNase rep 2

description: Library name: MCF7_MNase_P2


title: SHAPE_COX1_P2_1-NH2_A


title: MCF7, wild type live cells, 48 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF7_H31_48h

data_processing:
The pair-end reads aligned to the reference genome using bowtie2 (2.5.1), the read ends and occupancy is calculated using bedtools (2.31.1).
The end counts and occupancy is further processed to calculate the methylated fraction of GATC sites at each genome location using the snakemake workflow snakemakeMethylFrac, and the methylated fractions are stored in the bigwig files.
The snakemake protocol for data processing is avaiable at https://github.com/zhuweix/snakemakeMethylFrac.
Assembly: hg38 and T2T chm13v1.1, as shown in the file name of the processed files.
Supplementary files format and content: bigwig files of the methylated fraction of GATC sites in each genome location.

extract_protocol:
Genomic DNA was purified from Human MCF7 cells or nuclei by using Purelink genomic DNA kit (K182002) from invitrogen.
Genomic DNA was digested with excess DpnI and sonicated to ~300 bp fragments. Paired-end libraries were prepared using the New England Biolabs kits and protocols.
qDA-seq (Prajapati et al. (2024) NSMB


title: SHAPE_COX1_P2_1M7_A


title: MCF7, wild type live cells, 72 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF7_H31_72h


title: wild type nuclei, 0.78 nM Dam, No spermidine, rep 2

description: Library name: MCF7_H45_damM_25U


title: MCF7, wild type live cells, 12 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF7_H31_12h


title: MCF7, wild type live cells, 24 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF7_H31_24h


title: Tether_HEK293T_Mol2_2PT_100_B

description:
Enriched RNA
Reads filtered for number of T-to-C mutations (0-5+) and strandedness (min/pos) for each transcript and U position

extract_protocol:
RNA was harvested under non-denaturing conditions using a hypotonic lysis buffer (HLB). The pelleted cells were resuspended in HLB (20 mM HEPES, pH 8, 2 mM MgCl2, 0.2 mM EGTA, 10% glycerol, fresh 1 mM DTT) and freeze-thawed 3x and then placed on ice. After adding NaCl to 400 mM, samples were iced for 5 min and then centrifuged for 15 min at 4°C.The supernatant was collected, treated with proteinase K and 1% SDS and incubated at 37°C for 45 min. RNA was extracted 2x with phenol:chloroform:isoamyl (24:24:1) and 2x with chloroform. The aqueous layer was treated with Turbo DNAse and then buffer exchanged into folding buffer.
Tether-seq libraries were generated following standard protocols using the SMARTer Stranded Total RNA-Seq Kit-Pico Input Mammalian, V2 (Takara Bio USA, cat. 634413).

value: Yes (100 uM, 2 hours)


title: MCF7, wild type live cells, 72 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF7_H29_72h


title: Tether_HEK293T_Mol2_2PT_100_A


title: Tether_HEK293T_Mol2_2PT_10_C


title: Tether_HEK293T_Mol2_2PT_10_B


title: SHAPE_COX1_P2_2-NH2_B


title: MCF7, wild type live cells, 0 h, rep 2

description: Library name: MCF7_H31_0h


title: MCF7, wild type live cells, 24 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF7_H29_24h


title: Tether_HEK293T_MolEH_2PT_10_B


title: MCF7, wild type live cells, 12 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF7_H29_12h


title: Tether_HEK293T_MolEH_2PT_10_A


title: Tether_HEK293T_MolEH_2PT_10_C


title: wild type nuclei, No Dam, No spermidine, rep 1

description: Library name: MCF7_H44_damM_0U


title: wild type live cells, 48 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF10A_P5_48h

extract_protocol:
Genomic DNA was purified from Human MCF10A cells by using Purelink genomic DNA kit (K182002) from invitrogen.
Genomic DNA was digested with excess DpnI and sonicated to ~300 bp fragments. Paired-end libraries were prepared using the New England Biolabs kits and protocols.
qDA-seq (Prajapati et al. (2024) NSMB


title: MCF7, wild type live cells, 0 h, rep 1

description: Library name: MCF7_H29_0h


title: wild type live cells, 36 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF10A_P5_36h


title: Tether_HEK293T_Ox_4_C


title: wild type live cells, 24 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF10A_P5_24h


title: Tether_HEK293T_Mol2_2PT_100_C


title: wild type live cells, 12 h Dam adenoviral vector transduction , rep 2

description: Library name: MCF10A_P5_12h


title: Tether_HEK293T_Mol1_B


title: wild type live cells, 0 h, rep 2

description: Library name: MCF10A_P5_0h


title: Tether_HEK293T_Ox_3_A


title: wild type live cells, 48 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF10A_P4_48h


title: MCF7, wild type live cells, 48 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF7_H29_48h


title: wild type live cells, 12 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF10A_P4_12h


title: wild type live cells, 0 h, rep 1

description: Library name: MCF10A_P4_0h


title: wild type live cells, 36 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF10A_P4_36h


title: Tether_HEK293T_Mol3_C

treatment_protocol: 8 μg of total RNA was subjected to tethering conditions as described in the supporting information, followed by biotinylation and streptavidin enrichment as previously described (Duffy et al., 2015). RNA was then treated with oxidative-nucleophilic-aromatic-substitution chemistry using mCPBA as the oxidant as previously described (Schofield et al., Nature Methods 2018).


title: CAGEseq_SW620 shNIPBL_#2 5D rep2

value: Bowel

value: Colorectal cancer


title: Tether_HEK293T_Mol1_A


title: Tether_HEK293T_Ox_4_B


title: CAGEseq_CAMA1 shNIPBL_#2 5D rep2

source_name: Breast

value: Breast

value: Breast cancer


title: CAGEseq_CAMA1 shLuc 5D rep1

value: shLuc


title: Tether_HEK293T_no_s4U_ctl_Input_3

description:
Input RNA
Used for SNP calling in TimeLapse pipeline

treatment_protocol: 8 μg of total RNA was subjected to tethering conditions: Samples (40 μL) were prepared with non-denatured s4U-labeled RNA (8 μg), 0-250 μM disulfide small molecule and 0-500 μM 2PT competitor in 20 mM HEPES, pH 8, 1 mM EDTA, and 5% DMF. After incubating at room temperature for 30 minutes, mCPBA (0-15 mM, 10% ethanol final) was added and reactions were incubated at 45-50°C for 45 minutes. Urea was added to 5 M and mCPBA was replenished to a final concentration of 15 mM and final volume of 90 μL. Samples were again incubated at 45-50°C for 45 minutes. DTT (30 mM final) was added and samples were heated at 55°C for 15 minutes to quench excess oxidant and reduce off tether. Samples were biotinylated as previously described (Duffy et al., 2015). Input RNA was then isolated and then treated with oxidative-nucleophilic-aromatic-substitution chemistry using mCPBA as the oxidant as previously described (Schofield et al., Nature Methods 2018).


title: Tether_HEK293T_Mol3_B


title: Tether_HEK293T_MolEH_C


title: Tether_HEK293T_MolEH_B


title: Tether_HEK293T_Mol3_A


title: Tether_HEK293T_Ox_3_B


title: Tether_HEK293T_Ox_4_A


title: Tether_HEK293T_MolEH_A


title: Tether_HEK293T_Mol2_2PT_10_A


title: CAGEseq_CAMA1 shLuc 5D rep2


title: Tether_HEK293T_no_s4U_ctl_Input_4


title: wild type live cells, 24 h Dam adenoviral vector transduction , rep 1

description: Library name: MCF10A_P4_24h


title: Tether_HEK293T_Mol2_C


title: CAGEseq_SW620 shLuc 5D rep2


title: CAGEseq_CAMA1 shNIPBL_#2 5D rep1


title: CAGEseq_SW620 shLuc 5D rep1


title: Tether_HEK293T_Mol2_B


title: CAGEseq_C106  shNIPBL_#2 5D rep1


title: CAGEseq_C106  shNIPBL_#2 5D rep2


title: Tether_HEK293T_Mol4_C


title: Tether_HEK293T_Mol2_A


title: CAGEseq_C106 shLuc 5D rep1


title: CAGEseq_H358 shNIPBL_#2 5D rep2

value: Lung

value: Lung cancer


title: Tether_HEK293T_Mol4_A


title: CAGEseq_C106  shLuc 5D rep2


title: CAGEseq_H358 shLuc 5D rep2


title: Tether_HEK293T_Ox_2_C


title: Tether_HEK293T_Ox_2_B


title: CAGEseq_22RV1 shNIPBL_#2 5D rep2

source_name: Prostate

value: Prostate

value: Prostate cancer


title: CAGEseq_SW620 shNIPBL_#2 5D rep1


title: Tether_HEK293T_s4U_ctl_2

value: HEK293T cells


title: CAGEseq_22RV1 shNIPBL_#2 5D rep1


title: CAGEseq_H358 shLuc 5D rep1


title: Tether_HEK293T_no_s4U_ctl_2

value: No


title: Tether_HEK293T_PNA_B_2PT_A

extract_protocol:
RNA was harvested with Trizol reagent, followed by chloroform extraction and isopropanol precipitation. Genomic DNA was depleted using Turbo DNAse, followed by cleanup with RNAclean beads.
Tether-seq libraries were generated following standard protocols using the SMARTer Stranded Total RNA-Seq Kit-Pico Input Mammalian, V2 (Takara Bio USA, cat. 634413).

treatment_protocol: 7.5 μg of total RNA was subjected to tethering conditions: Samples (40 μL) were prepared with s4U-labeled RNA (7.5 μg), 0-250 μM disulfide PNA and 0-500 μM 2PT competitor in 20 mM HEPES, pH 8, 1 mM EDTA, and 5% DMF. Samples were heated to 80°C for 3 min, cooled to RT over 15 min, and then left to incubate at RT for 30 minutes. Following tethering, mCPBA (0-15 mM, 10% ethanol final) was added and reactions were incubated at 45-50°C for 45 minutes. Urea was added to 5 M and mCPBA was replenished to a final concentration of 15 mM and final volume of 90 μL. Samples were again incubated at 45-50°C for 45 minutes. DTT (30 mM final) was added and samples were heated at 55°C for 15 minutes to quench excess oxidant and reduce off tether. Samples were biotinylated and enriched as previously described (Duffy et al., 2015). RNA was then treated with oxidative-nucleophilic-aromatic-substitution chemistry using mCPBA as the oxidant as previously described (Schofield et al., Nature Methods 2018).


title: Tether_HEK293T_no_s4U_ctl_Input_2


title: Tether_HEK293T_PNA_B_C


title: Tether_HEK293T_PNA_B_B


title: Tether_HEK293T_PNA_B_2PT_B


title: Bovine,RNA-seq,8cell,DRBW control,rep2

city: Yangling

first: zhang

last: yong

state: Shanxi

address: No.3 Taicheng Road

institute: Northwest A&F University

data_processing:
We aligned all RNA-seq datasets to distinct genomes (bovine, bosTau9) utilizing Hisat2 v2.1.1. The quantification of gene expression was subsequently performed using StringTie 2.2.1.
Assembly: bosTau9
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

organism: Bos grunniens x Bos taurus

source_name: ovaries

extract_protocol:
Bovine ovaries, sourced from Holstein cows at a local abattoir, were preserved in pre-warmed saline solution. Cumulus oocyte complexes (COCs) were aspirated from antral follicles ranging from 2 to 10 mm, utilizing a 21-G needle connected to a vacuum pump. The COCs with uniform oocyte cytoplasm and at least three closed layers of cumulus cells were selected and washed in the collection medium: modified M199 medium (Sigma-Aldrich, M2520) supplement with 0.22 mM sodium pyruvate, 10 μg/mL gentamicin, and 1 mg/mL PVA. Then, the washed COCs were transferred to oocyte maturation medium: modified M199 medium (Sigma-Aldrich, M2154) was supplemented with 2 mM Ala-glutamine, 0.22 mM sodium pyruvate, 10 μg/mL gentamicin, 50 ng/mL EGF, 40 ng/mL FGF, 0.01 IU/mL pFSH, 0.01 IU/mL pLH, β-estradiol 1μg/mL, 3.10 mM L-carnitine, 2.77 mM myo-inositol, 0.1 mM cysteamine, and 6 mg/mL fatty acid-free bovine serum albumin (BSA). The maturation process occurred in a humidified atmosphere containing 5% CO2 at 38.8°C.  Bovine IVF embryos at various stages were collected at defined time points: 18-20 hpi (zygote), 30-32 hpi (2-cell), 42-44 hpi (4-cell), 72 hpi (8-cell), 96 hpi (16-cell), 132 hpi (morula). The zona pellucida of all oocytes and embryos underwent a brief incubation in 0.5% pronase (Sigma- Aldrich, P8811). Polar bodies of MII oocytes, zygotes, and early cleavage-stage embryos were eliminated by repeated pipetting using a fire-polished glass needle in DPBS supplemented with 0.01 mg/mL BSA (DPBS-0.001% BSA). The ICMs or TEs of bovine blastocysts were mechanically separated with a laser assistant. ICMs were placed in a 1:4 dilution of rabbit anti- bovine serum (Sigma-Aldrich, B-8270) for 40 minutes, washed twice in DPBS-0.001% BSA, and incubated for 40 minutes in a 1:4 dilution of guinea pig Complement Serum (Sigma-Aldrich, S1639). The lysed outer cells (mostly trophectodermal cells) were manually removed using a finely pulled pipette. Isolated TEs and ICMs were washed two times with DPBS-0.001% BSA.
For the generation of Smart-seq2 libraries, zona pellucida-free oocytes or embryos lysis in a buffer containing RNase inhibitor, following the Smart-seq2 protocol as described (Picelli,2013). After completion of purification and fragmentation, the library was quantificatied and sequenced.

value: ovaries

tag: developmental stage

value: control 8-cell embryo


title: CAGEseq_22RV1 shLuc 5D rep2


title: CAGEseq_22RV1 shLuc 5D rep1


title: Bovine,RNA-seq,8cell,DRBW control,rep1


title: Tether_HEK293T_PNA_B_A


title: Bovine,RNA-seq,8cell,DRBW,rep2

value: DRB wash treated 8-cell embryo


title: Tether_HEK293T_PNA_A_2PT_B


title: Tether_HEK293T_PNA_A_2PT_A


title: Tether_HEK293T_Ox_2_A


title: Tether_HEK293T_PNA_A_B


title: Tether_HEK293T_PNA_A_C


title: Bovine,RNA-seq,Zygote,rep4

data_processing:
We aligned all RNA-seq datasets to distinct genomes (bovine, bosTau9) utilizing Hisat2 v2.1.1. The quantification of gene expression was subsequently performed using StringTie 2.2.1.
Assembly: bosTau9
Supplementary files format and content: BigWig files of each stage

extract_protocol:
Bovine ovaries, sourced from Holstein cows at a local abattoir, were preserved in pre-warmed saline solution. Cumulus oocyte complexes (COCs) were aspirated from antral follicles ranging from 2 to 10 mm, utilizing a 21-G needle connected to a vacuum pump. The COCs with uniform oocyte cytoplasm and at least three closed layers of cumulus cells were selected and washed in the collection medium: modified M199 medium (Sigma-Aldrich, M2520) supplement with 0.22 mM sodium pyruvate, 10 μg/mL gentamicin, and 1 mg/mL PVA. Then, the washed COCs were transferred to oocyte maturation medium: modified M199 medium (Sigma-Aldrich, M2154) was supplemented with 2 mM Ala-glutamine, 0.22 mM sodium pyruvate, 10 μg/mL gentamicin, 50 ng/mL EGF, 40 ng/mL FGF, 0.01 IU/mL pFSH, 0.01 IU/mL pLH, β-estradiol 1μg/mL, 3.10 mM L-carnitine, 2.77 mM myo-inositol, 0.1 mM cysteamine, and 6 mg/mL fatty acid-free bovine serum albumin (BSA). The maturation process occurred in a humidified atmosphere containing 5% CO2 at 38.8°C.  Bovine IVF embryos at various stages were collected at defined time points: 18-20 hpi (zygote), 30-32 hpi (2-cell), 42-44 hpi (4-cell), 72 hpi (8-cell), 96 hpi (16-cell), 132 hpi (morula). The zona pellucida of all oocytes and embryos underwent a brief incubation in 0.5% pronase (Sigma- Aldrich, P8811). Polar bodies of MII oocytes, zygotes, and early cleavage-stage embryos were eliminated by repeated pipetting using a fire-polished glass needle in DPBS supplemented with 0.01 mg/mL BSA (DPBS-0.001% BSA). The ICMs or TEs of bovine blastocysts were mechanically separated with a laser assistant. ICMs were placed in a 1:4 dilution of rabbit anti- bovine serum (Sigma-Aldrich, B-8270) for 40 minutes, washed twice in DPBS-0.001% BSA, and incubated for 40 minutes in a 1:4 dilution of guinea pig Complement Serum (Sigma-Aldrich, S1639). The lysed outer cells (mostly trophectodermal cells) were manually removed using a finely pulled pipette. Isolated TEs and ICMs were washed two times with DPBS-0.001% BSA.
For the generation of Smart-seq2 libraries, zona pellucida-free oocytes or embryos lysis in a 708 buffer containing RNase inhibitor, following the Smart-seq2 protocol as described (Picelli,2013). After completion of purification and fragmentation, the library was quantificatied and sequenced.


title: Bovine,RNA-seq,Zygote,rep3


title: Tether_HEK293T_PNA_A_A


title: CAGEseq_H358 shNIPBL_#2 5D rep1


title: Bovine,RNA-seq,Zygote,rep1


title: Bovine,RNA-seq,Zygote,rep2


title: Bovine,RNA-seq,TBE_8cell,rep1

value: α-amanitin treated 8-cell embryo


title: Bovine,RNA-seq,TBE_8cell,rep2


title: Bovine,RNA-seq,8cell,DRBW,rep1


title: Bovine,RNA-seq,TBE_4cell,rep2

value: α-amanitin treated 4-cell embryo


title: Tether_HEK293T_Ox_1_C

value: Yes (100 uM, 2 hours)


title: Human T158M neuron, B2

city: Davis

first: Rachisan Gabriel

middle: Djiake

last: Tihagam

state: CA

address: 919 Drake Drive,, 113

department: BMG

institute: University of virginia

data_processing:
The Galaxy platform Galaxy version 23.0.rc1
Sequence reads were trimmed for adaptor sequence/low-quality sequence using Trimmomatic (Galaxy Version 23.0.rc1) with the parameters:  ILLUMINACLIP:/TruSeq3-PE.fa:2:30:10 SLIDINGWINDOW:4:15 MINLEN:35 LEADING:3 TRAILING:3
Trimmed sequence reads were mapped to GRCh38/hg38 whole genome using HISAT2 (Galaxy Tool version 23.0.rc1)
Gene expression counts were quantified from the mapped reads using featureCounts (Galaxy Tool version 23. 0.rc1)
Assembly: human
Supplementary files format and content: tab-delimited text files include count values for each Sample

source_name: Neuron

extract_protocol:
RNA was isolated with Rneasy Mini Kit at day 28 post-differentiation.
RNA libraries for RNA-seq were prepared by Novogene

value: Neuron

value: T158M neuron

value: pluripotent stem cell

value: WT

value: LTV-miRsp


title: Tether_HEK293T_s4U_ctl_1

source_name: HEK293T cells


title: Tether_HEK293T_no_s4U_ctl_Input_1

treatment_protocol: 7.5 μg of total RNA was subjected to tethering conditions: Samples (40 μL) were prepared with s4U-labeled RNA (7.5 μg), 0-250 μM disulfide PNA and 0-500 μM 2PT competitor in 20 mM HEPES, pH 8, 1 mM EDTA, and 5% DMF. Samples were heated to 80°C for 3 min, cooled to RT over 15 min, and then left to incubate at RT for 30 minutes. Following tethering, mCPBA (0-15 mM, 10% ethanol final) was added and reactions were incubated at 45-50°C for 45 minutes. Urea was added to 5 M and mCPBA was replenished to a final concentration of 15 mM and final volume of 90 μL. Samples were again incubated at 45-50°C for 45 minutes. DTT (30 mM final) was added and samples were heated at 55°C for 15 minutes to quench excess oxidant and reduce off tether. Samples were biotinylated as previously described (Duffy et al., 2015). Input RNA was then isolated and then treated with oxidative-nucleophilic-aromatic-substitution chemistry using mCPBA as the oxidant as previously described (Schofield et al., Nature Methods 2018).


title: Bovine,RNA-seq,TBE_4cell,rep1


title: Bovine,RNA-seq,TBE_2cell,rep2

value: α-amanitin treated 2-cell embryo


title: Tether_HEK293T_Ox_1_A


title: Tether_HEK293T_Ox_1_B


title: Tether_HEK293T_Mol4_B


title: Human T158M neuron, A2

value: LTV-empty


title: Tether_HEK293T_no_s4U_ctl_1


title: Bovine,RNA-seq,D7_TE,rep2

source_name: Trophectoderm

value: Trophectoderm


title: Bovine,RNA-seq,D7_TE,rep1


title: Bovine,RNA-seq,D7_ICM,rep1

source_name: Inner cell mass

value: Inner cell mass


title: Bovine,RNA-seq,Morula,rep2

source_name: Morula

value: Morula


title: A549,HSV-1,input-1

city: hangzhou

first: Hongyu

last: Lin

address: Zijingang Campus of Zhejiang University, 866 Yuhangtang Road, Xihu District

institute: Zhejiang University School of Medicine

data_processing:
The clean reads were aligned to human genome (GRCh38.p14) using BWA mem (v 0.7.12), and the IP enriched peaks were then analyzed by MACS2 (version 2.1.0).
Assembly: hg38
Supplementary files format and content: bigWig, narrowPeak (except for Input sample)

growth_protocol: DMEM/F-12, supplemented with 10% (v/v) FBS. Cells were cultured at 37C at 5% CO2.

extract_protocol:
Infected A549 cells were collected, fixed and undergone chromatin cross-linking according to the manufacturer’s instructions using the SimpleChIP® Enzymatic Chromatin IP Kit
The purified DNA was then used to construct a ChIP-seq library by Novogene (Beijing, China)

treatment_protocol: A549 cells were infected with HSV-1 for 8 hours

tag: antibody


title: A549,HSV-1,input-3


title: Bovine,RNA-seq,8cell,siKLF17,rep1

value: siKLF17 8-cell embryo


title: Bovine,RNA-seq,TBE_2cell,rep1


title: A549,HSV-1,input-2


title: Bovine,RNA-seq,8cell,siKLF17 control,rep1


title: A549,HSV-1,ChIP-3


title: Bovine,RNA-seq,Morula,rep1


title: Bovine,RNA-seq,8cell,siDuxA,rep2

value: siDUXA 8-cell embryo


title: Human T158M neuron, B1


title: Human T158M neuron, A1