title: Htz1 Swc3∆ rep1

city: Ithaca

first: Justin

middle: Sungsoo

last: Cha

state: NY

address: 526 Campus Rd

department: Computational Biology

institute: Cornell University

data_processing:
*library strategy: ChIP-exo
ChIP-exo reads were aligned using bwa-mem with the following settings: -v (1)  -T (30) -h (5)
Remove duplicate reads and generate a deduplicated BAM file using picard MarkDuplicates and samtools
Generate forward and reverse BIGWIG files from deduplicated BAM file using deeptools bamCoverage (forward: --binSize 1 --Offset 1 --samFlagInclude 64 --samFlagExclude 16, reverse: --binSize 1 --Offset 1 --samFlagInclude 80 --samFlagExclude 0)
Assembly: sacCer3
Supplementary files format and content: Bigwig for forward strand -- coverage of 5' end of Read 1 across the genome
Supplementary files format and content: Bigwig for reverse strand -- coverage of 5' end of Read 1 across the genome

organism: Saccharomyces cerevisiae

growth_protocol: S. cerevisiae strains containing FLAG-tagged H2A.Z or TAP-tagged Swr1 in WT or mutants were grown in YPD media to an O.D600 of 0.6-0.8 at 25˚C

extract_protocol:
70 mL cell pellets were resuspended in 1 mL FA lysis buffer (50 mM HEPES-KOH, pH7.5, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 0.1% sodium deoxycholate and complete protease inhibitor (Roche)) along with 0.5 mL of 0.5 mm acid washed zirconia beads. Lysis was performed by bead beating for 3 cycles of 3 min on and 5 min off in a Mini-Beadbeater-96 machine (Biospec). Whole cell lysates were centrifuged at 14,000 rpm for 3 min at 4 ˚C. Cytoplasmic supernatants were discarded and nuclear pellets were washed in 600 uL FA lysis buffer. Samples were centirufged as above. Supernatant was discarded. Chromatin was resuspended in 600 uL FA lysis buffer with 300 uL 0.1 mm zirconia beads. Nuclear samples were sonicated in a Bioruptor (Diagenode) for 8 cycles of 15 sec on and 30 sec off. Samples were centrifuged at 14,000 rpm for 10 min at 4 *C to collect sonicated chromatin fraction which was predominantly enriched with DNA fragments in a range of 200-500 bp in size (verified on agarose gels). Following antibodies were used according to the manufacturer's instructions to prepare ChIP-exo libraries: anti-FLAG (Sigma, M8823), IgG (Sigma, i5006). Chromatin obtained from 70 mL culture was used for a single ChIP experiment. Appropriate antibodies (3-5 ug per ChIP) were bound to protein A/G magnetic Sepharose resin (Citiva) at 4 ˚C for 6-8 hr. Unbound antibody was removed, then chromatin samples were immunoprecipitated overnight (12-16 hr) at 4 ˚C. For TAP-tagged samples, 50 uL IgG-Dynabeads or IgG-Magnetic Sepharose slurry was used per sample. Subsequent steps were performed as described previously (Rossi et al., 2018).
ChIP-exo libraries were gel purified and were sequenced in a paired-end mode with NextSeq 500 (Read_1 40 cycles, Read_2 36 cycles).

tag: strain background

tag: genetic modification

tag: epitope tag

tag: antibody

tag: growth media

value: YPDad


title: Swr1 Swc3∆

status: Public on Aug 26 2024

type: SRA

country: USA

library_source: genomic

molecule: genomic DNA

source_name: grown in YPD media to an O.D600 of 0.6-0.8

tag: cell type

value: MATa can1-100 his3-11,15 leu2-3, ura3-1, RAD5+, pdr5∆::Hyg

value: YPD


title: Control 55D mg4

city: Guelph

first: Roger

last: Moorehead

address: 50 Stone Rd E

department: Biomedical Sciences

country: Canada

institute: University of Guelph

library_source: transcriptomic

data_processing:
adapters removed, reads containing N>10% removed, Qscore<=5 removed by Novogene
data was provided by Novogene as .fq.gz files
data was analyzed using Genialis software and the standard RNA-seq pipeling
Assembly: GRCm39_ens109
Supplementary files format and content: xlsx file containing featurecounts data

molecule: total RNA

organism: Mus musculus

source_name: mammary gland

growth_protocol: mice were weaned at 21 days of age and placed on solid food

extract_protocol:
Total RNA was extracted using the miRVana miRNA extraction kit from Thermo Fisher Scientific
libraries were created by Novogene using Illumina kits

treatment_protocol: beginning at 21 days of age, mice were provide rodent chow with 2g of doxycycline/kg chow

tag: tissue

value: mammary gland

tag: cell line

value: tissue

tag: genotype

value: WT

tag: treatment

value: 2g doxycycline/kg chow for 34days


title: THPE.1_0.5uM

city: Ottawa

first: Matthew

last: Meier

state: Ontario

address: 251 Sir Frederick Banting Driveway

department: Health Canada

institute: Government of Canada

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_37

library_source: other

data_processing:
Reads were demutliplexed from BCL files and processed into FASTQ files on Illumina Basespace using DRAGEN v1.3.0
Fastq files were aligned to the Human Whole Transcriptome v2.1 probes references sequences (provided by BioSpyder) using STAR aligner 2.7.8b
The qCount function from QuasR was used to extract feature counts specified in a GTF file (provided by BioSpyder) from the aligned reads.
Assembly: Biospyder human whole transcriptome gene set
Supplementary files format and content: Tab-delimited table containing read counts per TempO-Seq probe

molecule: other

organism: Homo sapiens

growth_protocol: MCF-7 cells were routinely cultured in Dulbecco's modified Eagle's medium (EMEM; 30-2003, ATCC, Manassas, VA) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco, Thermo Fisher Scientific), at 37°C in a humidified atmosphere with 5% CO2. Cells were maintained in routine culture for 7 days post-thaw prior to treatment, as this was determined to be the length of time required for cells to express the human PGR marker.

extract_protocol:
Following the exposure, cells were washed with phosphate-buffered saline (PBS) and lysed for Templated Oligo-Sequencing (TempO-Seq®) using 2xTempO-Seq lysis buffer diluted with PBS. Cell lysates were used directly for TempO-Seq library building.
Gene expression changes were measured using the TempO-Seq Human Whole Transcriptome Panel v2.1 Assay, Index E, F, G (BioSpyder Technologies Inc, Carlsbad, California) following manufacturer’s instructions. Briefly, 1X TempO-Seq Enhanced Lysis Buffer was added to exposed MCF-7 cells to achieve cell lysis. Cell lysates were frozen and stored at -80°C for less than six months prior to library building. At the onset of library building, thawed cell lysates and technical controls (1) Human Universal Reference RNA—uhrRNA Agilent Cat No. 740000, Santa Clara, California, and 2) Human Brain Total RNA brRNA—ThermoFisher AM7962, Waltham, Massachusetts) were added to assay plates. Freshly prepared annealing mix, containing the whole human genome Detector Oligo (DO) Pool to be annealed to the sample RNA, was added to the assay plates. The mixture was incubated at 70°C for 10 minutes, then ramped to 45°C over 50 minutes, then held at 45°C for 16 hours, and finally cooled to 25°C to hybridize the DO pool. Assay plates were frozen overnight at -20°C. Freshly prepared Nuclease Mix was added to each well of the thawed assay plates and then the assay plates were incubated for 90 minutes at 37°C to ensure removal of unbound DOs. Freshly prepared Ligation Mix was added to each well of the assay plate and then plates were incubated for 1 hour at 37°C, then immediately at 80°C for 15 minutes to ligate adjacent DOs together. For DO amplification, PCR Pre-Mix and PCR primers were mixed with the ligated assay products and then amplified on a thermocycler using the following protocol: 1) 37°C 10 minutes; 2) 95°C 1 minute; 3) 25 cycles of 95°C 10 sec, 65°C 30 sec, 68°C 30 sec; 4) 68°C 2 minutes; 5) hold at 25°C. After amplification, assay plates were stored at -20°C prior to purification. When all three assay plates in the study completed amplification, the plates were thawed and amplicons were purified using the Macherey-Nagel NucleoSpin Gel and PCR Cleanup Kit. Libraries were pooled in equimolar amounts prior to sequencing.
Human TempO-Seq Whole Transcriptome - Index Sets E, F, and G were used to index libraries

treatment_protocol: At the start of the experiment, cells were seeded into 96-well plates at a density of 3.0 × 104 cells per well in EMEM supplemented with 10% FBS to promote cell attachment to the plate. On Day 0, the medium was removed and replaced with 100 µL/well of fresh phenol red-free DMEM (DMEM; 31053-028, Gibco, Thermo Fisher Scientific) containing the treatment and supplemented with 10% charcoal-dextran stripped FBS to eliminate estrogenic components of the medium. Cells were exposed in triplicates for 48 hours to: (1) BPA and 11 alternative chemicals sharing structural similarity (Table 1;Figure 1) at six different concentrations (0.1, 0.5, 1, 5, 10, 25 µM); (2) the positive control 17β-estradiol (E2; 0.001, 0.01, 0.1, 1, 10 nM); (3) the non-ER-acting controls dexamethasone (Dex; 0.0001, 0.001, 0.01, 0.1, 1 µM) and Bis(4-chlorophenyl) sulfone (BCPS; 0.1, 0.5, 1, 5, 10, 25 µM); (4) the vehicle control (final (v/v) 1% DMSO, or 1% MeOH). The top concentration of 25 µM was chosen as higher concentrations of BPA alternatives (i.e., 50-100 µM) have been shown to cause cell stress, reduced cell count, or precipitation.

tag: batch

tag: index

value: E

tag: group

tag: chemical

tag: dose


title: MTB-TAN 55D mg3

value: Neu transgenic


title: MTB-TAN 55D mg5


title: Htz1 Swc3∆ rep2

value: grown in YPD media to an O.D600 of 0.6-0.8


title: Control 55D mg5


title: Swr1 Swc3 177-255∆


title: Swr1 wild type


title: Htz1 Swc3 177-255∆ rep2


title: Control 55D mg2


title: THPE.1_1uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_36


title: Control 55D mg3


title: MTB-TAN 55D mg1


title: THPE.1_10uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_34


title: THPE.1_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_33


title: Htz1 Swc3 177-255∆ rep1


title: exp_TSCF-3

city: Qingdao

first: Yong

last: Zhao

state: Shandong

address: 700 Changcheng Road

department: College of Life Sciences

country: China

institute: Qingdao Agricultural University

data_processing:
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm10 whole genome using bowtie v0.12.2 with parameters -q -p 4 -e 100 -y -a -m
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using a protocol from Chepelev et al., Nucleic Acids Research, 2009. In short, exons from all isoforms of a gene were merged to create one meta-transcript. The number of reads falling in the exons of this meta-transcript were counted and normalized by the size of the meta-transcript and by the size of the library.
Genome_build: mm10
Supplementary_files_format_and_content: include gene expression

source_name: testis

extract_protocol:
Testis were removed, quikly frozen in luquid nitrogen, and RNA was harvested using Trizol reagent. PurelinkTM RNA Mini Kit (Cat#12183018A) was used to abtain 1 ug of total RNA for the construction of sequencing libraries.
RNA libraries were prepared for sequencing using standard Illumina protocols

tag: strain

value: ICR

value: testis

tag: age

value: 10 weeks

value: STZ+control-FMT


title: exp_TSCF-2


title: Control 55D mg1


title: 2,4BPA.4_1uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_32

value: 2,4'-Bisphenol A


title: exp_TSCF-1


title: Htz1 wild type rep2

tag: strain background


title: Htz1 wild type rep1

tag: genetic modification


title: BPBP.4_1uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_31

value: Bisphenol BP


title: exp_TSAF-2


title: exp_TSAF-3


title: RA.1_0.5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_30

value: Rosalic Acid


title: exp_TSAF-1


title: exp_TSA-3


title: exp_TSA-2


title: 2,4BPA.1_1uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_28


title: exp_TS-3

value: STZ


title: SKOV3 CPT1Ash, biol rep 3

city: xian

first: yaqin

last: zhu

address: xi changan street, 620

institute: Shaanxi normal university

data_processing:
Cutadapt (V1.9.1)
Firstly, reference genome sequences and gene model annotation files of relative species were downloaded  from genome website, such as UCSC, NCBI, ENSEMBL. Secondly, Hisat2 (v2.0.1) was used to index  reference genome sequence. Finally, clean data were aligned to reference genome via software Hisat2 (v2.0.1).
In the beginning transcripts in fasta format are converted from known gff annotation file and indexed  properly. Then, with the file as a reference gene file, HTSeq (v0.6.1) estimated gene and isoform  expression levels from the pair-end clean data.
Differential expression analysis used the DESeq2 Bioconductor package, a model based on the negative  binomial distribution. the estimates of dispersion and logarithmic fold changes incorporate data-driven prior distributions, Padj of genes were setted <0.05 to detect differential expressed ones.
DEXSeq (V 1.21.1)
Assembly: mm8
Supplementary files format and content: tab-delimited text files include raw counts for each Sample
Supplementary files format and content: tab-delimited text files include RPKM values for each Sample

source_name: ovary

extract_protocol:
Total RNA of each sample was extracted using TRIzol Reagent (Invitrogen)/RNeasy Mini Kit(Qiagen)/other kits.  1 μg total RNA with RIN value above 6.5 was used for following library preparation.
Next generation sequencing library preparations were constructed according to the manufacturer’s protocol.The poly(A) mRNA isolation was performed using Poly(A) mRNA Magnetic Isolation Module or rRNA removal Kit. The mRNA fragmentation and priming was performed using First Strand Synthesis Reaction Buffer and Random Primers. First strand cDNA was synthesized using ProtoScript II Reverse Transcriptase and the second-strand cDNA was synthesized using Second Strand Synthesis Enzyme Mix. The purified double_x0002_stranded cDNA by beads was then treated with End Prep Enzyme Mix to repair both ends and add a dA_x0002_tailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Size selection of Adaptor_x0002_ligated DNA was then performed using beads, and fragments of ~420 bp (with the approximate insert size of 300 bp) were recovered. Each sample was then amplified by PCR for 13 cycles using P5 and P7 primers, with both primers carrying sequences which can anneal with flow cell to perform bridge PCR and P7 primer carrying a six-base index allowing for multiplexing. The PCR products were cleaned up using beads, validated using an Qsep100 (Bioptic，Taiwan， China), and quantified by Qubit3.0 Fluorometer (Invitrogen, Carlsbad, CA, USA).

value: ovary

value: ovarian cancer cell

value: normal culture


title: SKOV3, biol rep 3


title: 2,4BPA.1_0.5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_29


title: SKOV3 CPT1Ash, biol rep 2


title: SKOV3 CPT1Ash, biol rep 1


title: exp_TSA-1


title: exp_TS-1


title: 2,4BPA.1_5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_27


title: 2,4BPA.1_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_25


title: exp_TS-2


title: SKOV3, biol rep 2


title: exp_Tcon-2

value: control


title: exp_Tcon-1


title: SKOV3, biol rep 1

value: WT


title: A_soma, rep3

city: Hangzhou

first: Chengyong

last: Shen

address: 268 Kaiquan Rd.

department: Institute of Translational medicine

institute: Zhejiang University

data_processing:
The sequencing data was filtered with SOAPnuke (v1.5.2).
Removing reads whose low-quality base ratio (base quality less than or equal to 5) is more than 20% and  base ('N' base) ratio is more than 5%.
Bowtie2 (v2.2.5) was applied to align the clean reads to the reference coding gene set.
The expression level of gene was calculated by RSEM (v1.2.12)
Assembly: mm10
Supplementary files format and content: tab-delimited text files include FPKM and raw count values for each Sample
Library strategy: RiboTag-seq

source_name: Spinal cord

extract_protocol:
RNA was extracted using RNeasy micro kit.
RNA libraries for RNA-seq were prepared by BGI standard protocol.

value: Spinal cord

value: ChAT-Cre;Ribo-tag

tag: fraction

value: HA-tag purified RNA


title: exp_TA-3

value: AOS 10mg/kg


title: 2,4BPA.1_10uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_26

value: E


title: TMBPS.4_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_104

value: F

value: Tetra Methyl Bisphenol S


title: exp_TA-2

value: ICR

value: testis

value: 10 weeks


title: exp_Tcon-3


title: 2,4BPA.4_5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_24

value: 2,4'-Bisphenol A


title: A_soma, rep1


title: exp_TA-1


title: A_soma, rep2


title: iCLIP-seq of ZBTB48, replicate 1

city: Toronto

first: Ulrich

last: Braunschweig

address: 160 College Street

department: Donnelly Centre

institute: University of Toronto

data_processing:
Raw reads that consisted of 3 random positions, a 4-nt multiplexing barcode, and another 2 random positions, followed by the cDNA sequence, were initially trimmed to 49 nt from the 3'-end, and duplicates were discarded. Reads were de-multiplexed, and the random positions, barcodes, and any 3'-bases matching Illumina adaptors were removed. Remaining reads longer than 25 nt were mapped to the mouse genome/transcriptome (Ensembl annotation of NCBIm37) using TopHat with default settings. To prevent false assignment of reads from repetitive regions, any reads with a mapping quality < 3 were removed from further analysis. Note that this record contains demultiplexed but un-trimmed reads.
CLIP Tool Kit (CTK) was used to identify RNA-binding sites at single-nucleotide resolution.We  called CITS peaks on individual iCLIP replicates as well as SMI samples. CITS with FDR ≤ 0.01 were considered as significant. The identified iCLIP CITS were further normalized using SMI CITS by utilizing underlying reads from each dataset and performing a differential analysis (iCLIP/SMI). Remaining peaks from individual replicates were merged for downstream analysis.   miCLIP-seq data were processed similarly to the iCLIP-seq data (except for the SMI step), and putative anti-m6A mediated CITS were identified (FDR ≤ 0.01).
To generate meta-gene plots, the entire length of each gene (divided into 3 parts: 5’UTR, CDS, 3’UTR) plus 1kb upstream and downstream was allocated into 100 bins on the x-axis, the number of bins in each part is proportional to the median length of that part in all protein-coding genes. The Y-axis denotes coverage enrichment ratio over input/bin/gene, the shaded bands around each line represent 1xSE. All calculations and visualizations were performed with the R package 'GenomicPlot' (URL: https://github.com/shuye2009/GenomicPlot), the smoothing was done using R function "smooth.spline" in the stats package with the parameter df=20.
Assembly: hg19
Supplementary files format and content: BED files containing merged crosslink-induced termination site (CITS) locations from multiple replicates.
Supplementary files format and content: PDF files contains a meta-gene plot showing the mean normalized FTO iCLIP read density in different parts of the gene upon either ZBTB48 knockdown or control non-targeting knockdown. Figure was generated directly from aligned reads.

growth_protocol: Cell cultures were maintained in Dulbecco's modified Eagle's medium (DMEM) (Wisent Bioproducts catalogue number 319-005-CL) supplemented with 10% FBS (Wisent Bioproducts catalogue number 080-705), sodium pyruvate, non-essential amino acids, and penicillin/streptomycin.

extract_protocol:
Lysates were treated with Turbo DNase (Thermo Fisher Scientiﬁc) and RNase I (Thermo Fisher Scientiﬁc) diluted 1/250 in PBS for 5 minutes at 37°C to digest genomic DNA and partially digest RNA to short fragments of an optimal size range. RNA-protein complexes were puriﬁed using 100 mL of protein G Dynabeads (Thermo Fisher Scientiﬁc) and 10ug antibody, with 2% (20ul) of the total lysate retained for use as a size-matched input (SMI) control. SMI samples were processed as previously described (Van Nostrand et al., 2016). Following stringent high salt washes, the immunoprecipitated RNA was 3′ end dephosphorylated and ligated to a pre-adenylated adaptor at the 3′ end, as described previously. After extensive additional washes and buffer exchange, the immunoprecipitated RNA was 5′ end-labeled using ATP [γ-32P]. The puriﬁed protein-RNA complexes and SMI control samples were separated with SDS-PAGE and transferred to nitrocellulose membrane. Protein was digested using proteinase K, the RNA was puriﬁed using phenol/chloroform. miCLIP procedure was performed as decribed. Total RNA was extracted from HEK293 cells using TRIzol as per manufacturer's protocol. RNA was treated with DNase I to eliminate any contaminating genomic DNA. We used 12 μg of total RNA for miCLIP-seq. 2 μL of RNA Fragmentation Reagents was added to the RNA suspended in 20 μL of sterile water. The mixture was incubated in a heat block at 75°C for exactly 12 minutes. 2.2 μL of stop solution was added to stop the reaction. The fragmented RNA was incubated with 10 μg of m6A antibody for 2 hours at 4°C. RNA-antibody complexes were UV crosslinked twice with 0.15 J/cm2 at 254 nm in a Stratalinker 1800.  Protein G Dynabeads (100 μL) were added, and the samples were rotated for 2 hours at 4°C. Remaining steps, including dephosphorylation, adapter ligation, SDS-PAGE, and RNA-extraction are similar to the iCLIP procecure.
Purified RNA was reverse transcribed into cDNA. The cDNA was size selected by resolving samples on a 6% TBE-Urea gel, circularized to add the adaptor to the 5′ end, digested at the internal BamHI site, and then PCR ampliﬁed using AccuPrime SuperMix I (Thermo Fisher Scientiﬁc). The ﬁnal PCR libraries were puriﬁed by agarose gel electrophoresis using gel extraction columns (QIAGEN). The eluted DNA from the low, middle, and high fractions was pooled together and submitted for sequencing.

treatment_protocol: HEK293 cells were crosslinked (150mJ/cm2) at 254 nm with a Stratalinker 1800.

value: human embryonic kidney cells (HEK293)

tag: protocol

value: iCLIP-seq

tag: clip antibody

value: anti-ZBTB48 (GeneTex GTX118671)

tag: barcoded reverse_transcription_primer


title: iCLIP-seq of ZBTB48, replicate 2

tag: cell type


title: P10_soma, rep2

tag: tissue


title: BPBP.4_5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_23

tag: cell line

value: Bisphenol BP


title: P10_soma, rep3

tag: genotype

tag: treatment


title: RA.1_1uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_22

tag: batch

tag: index

value: Rosalic Acid


title: P10_soma, rep1


title: Universal.1

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_21

tag: group

tag: chemical

value: No cells; Universal human positive control


title: 16-04_Superficial,2N,Batch_2

city: Orangeburg

first: Stephen

middle: D.

last: Ginsberg

state: New York

address: 140 Old Orangeburg Road

department: Centre for Dementia Research

institute: Research Foundation for Mental Hygiene

data_processing:
The reads with adapter contamination/ low quality were trimmed using Trimmomatic 0.39.
Trimmed sequence reads were aligned to the reference  genome (GRCm39-mm10) using STAR Aligner. Gene counts were extracted using RSEM.
Downstream Analysis was performed using R with the gene counts obtained from the RSEM. Genes with over 0.1 counts per million were retained followed by TMM normalization.
Assembly: mm10
Supplementary files format and content: tab delimited text files with gene counts of all samples in a region per one file
Supplementary files format and content: tab delimited text files with gene counts of one sample per one file

source_name: CA1 pyramidal Neurons

growth_protocol: Breeder pairs (female B6EiC3Sn-Rb(12.Ts171665Dn)2Cje/CjeDnJ, strain ID 004850, herein Ts2 and male B6EiC3SnF1/J, strain ID 001875, mice) were purchased from Jackson Laboratories (Bar Harbor, ME, USA) and mated at the Nathan Kline Institute.

extract_protocol:
Brain tissues were accessed from Ts2 and normal disomic female mice. Mice were transcardially perfused with ice-cold 0.15 M phosphate buffer as previously described. Brains were immediately flash frozen and 20 μm-thick tissue sections were cryostat cut (−25°C) in the coronal plane and mounted on polyethylene naphthalate (PEN) membrane slides. Slides were kept under desiccant at -80°C until used for immunohistochemistry. RNase-free precautions were employed, and solutions were made with 18.2 mega Ohm RNase-free water. RNA from Batch 1 and Batch 2 for each CA1 subregion was purified using miRNeasy Micro kit (Qiagen) according to manufacturers’ specifications. A DNase digestion was performed twice sequentially before the final washes and RNA purification. LCM was performed on tissue to isolate the entire rostral CA1 pyramidal neuron layer from 3-6 sections (CA1A, ~500-600 pyramidal neurons) and compared that to the pyramidal neurons from the distal deep layer CA1 region(CA1D; 6-12 sections; ~500-1500 cells) and to the distal superficial layer CA1 region (CA1D; 6-12 sections; ~500-1500 cells).  Deep and superficial neurons were selected from the same tissue sections.  Batch 2 collected CA1A, CA1D and CA1S from adjacent sections within each slide (2-4 slides per brain to collect at least ~500 neurons per region), whereas batch 1 was collected with CA1D and CA1S in adjacent slides to the CA1A slide.
The SMARTer Stranded Total RNA-Seq kit-Pico input Mammalian (Takara Bio, Mountain View, CA) was employed with minor modifications to normalize neuronal numbers for each sample.
RNA-seq samples were quantified (Agilent 2100 HS DNA kit), with all samples reading a minimum of 2nM of the library for QC. For each sample run, library samples were pooled in equimolar concentrations and assayed on an Illumina HiSeq-4000 using a single read 50-cycle protocol at NYUGSOM Genome Technology Center.  Samples were run in multiple batches utilizing the same protocol for each batch.

treatment_protocol: Breeder pairs were on normal chow diet and had ad libitum access to food and water, with enrichment in each cage of t-tube, nestlets, bedding and igloos. Pups were weaned at postnatal 21, given ad libitum food and water, with nestlets and igloos in each cage for enrichment.

tag: region

value: Superficial

value: CA1 pyramidal Neurons


title: P0.5_soma, rep2

value: Spinal cord

value: ChAT-Cre;Ribo-tag

tag: age

value: HA-tag purified RNA


title: 16-179_Superficial,TS,Batch_2

value: TS


title: P0.5_soma, rep3


title: P0.5_soma, rep1


title: Brain.1

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_20

value: E

value: No cells; Brain positive control

tag: dose


title: 15-75_Superficial,2N,Batch_2


title: E15.5_soma, rep3


title: E15.5_soma, rep2


title: DMSO.1a

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_18

value: Dimethyl sulfoxide Treatment Control


title: 15-50_Superficial,2N,Batch_2


title: E15.5_soma, rep1


title: AP, rep1

source_name: Synapse-enriched muscle

value: Synapse-enriched muscle


title: AP, rep3


title: AN, rep2

value: Ribo-tag


title: BCPS.1_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_17

value: BCPS-CTL

value: 4,4′-Dichlorodiphenyl sulfone


title: 16-207_Superficial,TS,Batch_2


title: 16-154_Superficial,TS,Batch_2


title: 16-208_Superficial,2N,Batch_2


title: 2,4BPA.4_10uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_16

value: 2,4'-Bisphenol A


title: AN, rep1


title: P10P, rep3


title: 16-189_Superficial,2N,Batch_2


title: BPBP.1_0.5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_13

value: Bisphenol BP


title: P10P, rep2


title: P10P, rep1


title: P10N, rep3


title: 16-177_Superficial,TS,Batch_2


title: MeOH.1a

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_19

value: Methanol Treatment Control


title: P0.5P, rep2


title: AP, rep2


title: 16-319_Superficial,TS,Batch_1


title: 16-268_Superficial,2N,Batch_1


title: P0.5P, rep3


title: BPBP.1_10uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_10


title: 15-70_Superficial,TS,Batch_2


title: P10N, rep1


title: P0.5N, rep3


title: AN, rep3


title: 16-267_Superficial,TS,Batch_1


title: BPBP.1_5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_11


title: P0.5N, rep1


title: 16-257_Superficial,2N,Batch_1


title: BPBP.4_10uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_15


title: P10N, rep2


title: BPBP.1_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_9


title: 16-178_Superficial,TS,Batch_1


title: RA.1_5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_14

value: Rosalic Acid


title: E15.5P, rep3


title: P0.5N, rep2


title: E15.5P, rep2


title: BPBP.1_1uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_12


title: 15-72_Superficial,TS,Batch_1

value: Superficial


title: 2,4BPA.4_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_8


title: E15.5P, rep1


title: P0.5P, rep1


title: E15.5N, rep3


title: E15.5N, rep2


title: BPBP.4_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_7


title: Liver, fasted, S155A, biol rep4

city: Iowa City

first: Michael

middle: S

last: Chimenti

state: IA

address: 431 Newton Road

department: Iowa Institute of Human Genetics

institute: University of Iowa

data_processing:
Barcoded samples were pooled and sequenced using an Illumina NovaSeq 6000 in the Iowa Institute of Human Genetics (IIHG) Genomics Core Facility.
Paired-end reads were demultiplexed and converted from the native Illumina BCL format to FASTQ format using a custom python workflow wrapper to Illumina’s ‘bcl2fastq’ conversion utility.
FASTQ data were processed with nf-core/rnaseq (v3.12), a best-practices pipeline available at the open-source ‘nf-core’ project (https://nf-co.re/, Nextflow version 22.10).
Reads from the samples were aligned against the mouse reference genome ‘GRCm38’ using the STAR aligner (ref ) and quantified with ‘salmon’.
Samtools was used in conjunction with Qualimap and MultiQC to inspect alignment results. Length-normalized gene-level counts from the STAR/salmon pipeline were used for differential gene expression analysis with DESeq2
Assembly: GRCm38
Supplementary files format and content: flat file counts

source_name: Liver

growth_protocol: Mice were housed 5/cage in 12 h light-dark cycle at 22 °C, allowed free access to water, and fed regular rodent chow (7319 Teklad global diet) prior to fasting experiments.

extract_protocol:
RNA was extracted from the liver using TRIzol Reagent (Invitrogen) according to the user guide and then cleaned using RNA Clean & Concentrator-5 (Zymo Research) kit with DNase I treatment according to manufacturer instructions.
500 ng of DNase I-treated total RNA was used to prepare sequencing libraries using the Illumina TruSeq stranded mRNA library preparation kit (Cat. #RS-122-2101, Illumina, Inc., San Diego, CA).  The molar concentrations of the resulting indexed libraries were measured using the 2100 Agilent Bioanalyzer (Agilent Technologies, Santa Clara, CA) and combined equally into a pool for sequencing. The concentrations of the library pools were measured using the Illumina Library Quantification Kit (KAPA Biosystems, Wilmington, MA) and sequenced on the Illumina NovaSeq 6000 genome sequencer using 100 bp paired-end SBS chemistry.

treatment_protocol: Mice were fasted overnight prior to harvet.

value: Liver

value: Hepatocytes

tag: strain

value: PLIN5 S155A

tag: Sex

value: Female

value: 4 months

value: Overnight fast


title: RA.1_10uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_6

value: E


title: 15-51_Superficial,TS,Batch_1

value: CA1 pyramidal Neurons

value: TS


title: E15.5N, rep1

value: Synapse-enriched muscle

value: Ribo-tag

tag: fraction

value: HA-tag purified RNA


title: Liver, fasted, S155A, biol rep2


title: Liver, fasted, S155A, biol rep1


title: Liver, fasted, S155A, biol rep3


title: 15-224_Superficial,TS,Batch_1


title: 15-49_Superficial,2N,Batch_1


title: BPA.1_0.5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_5

value: Bisphenol A


title: Liver, fasted, WT, biol rep4

value: WT


title: Liver, fasted, WT, biol rep2


title: BPA.1_10uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_2


title: Liver, fasted, WT, biol rep3


title: 15-223_Superficial,2N,Batch_1


title: 15-131_Superficial,2N,Batch_1


title: BPA.1_5uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_3


title: BPA.1_1uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_4


title: NVF_E16-5_Rep1 [ATAC-Seq]

city: Neuherberg

first: Heiko

last: Lickert

address: Ingolstaedter Landstraße 1

department: Institute of Diabetes and Regeneration Research

country: Germany

institute: Helmholtz Zentrum München German Research Center for Environmental Health

description: 10x Genomics single cell multiome (ATAC)

data_processing:
Raw reads from both experiments were jointly aligned to the GRCm38 mouse genome with Ensembl release 102 annotations and pre-processed using the 10X Genomics CellRangerARC pipeline (v 2.0.2) for downstream analyses.
Assembly: ensembl mus musculus v102
Supplementary files format and content: Tab-separated values files and matrix files

source_name: embryonic pancreas

extract_protocol:
Embryonic pancreata from NVF homozygous mice were collected and pooled together (8 pancreata from E14.5 and 11 pancreata from E15.5 for the first experiment ("Exp-1"), and 10 pancreata from E15.5 and 10 pancreata from E16.5 for the second experiment ("Exp-2")). 0.25 % Trypsin was added to the samples for 5 min on ice and following incubation at 37 °C for 10 min. Next, the single-cell samples were centrifuged at 1700 rpm (290 x g) for 5 min at 4 °C. After removing the supernatant, cells were counted. rat 5 μ𝑙 IgG2a K isotype control (eBioscience, 12-4321-42) and anti-mouse CD326 (EpCAM) PE (eBioscience, 12-5791-81) were used for 1×106 cells (100 μ𝑙 total volume). Sample were stained for 30 min at 4 °C following staining with DAPI to detect dead cells. After washing twice and resuspended in FACS buffer (PBS, 1 % BSA, 0.5 mM EDTA), the single-cell samples were loaded for FACS sorting. The following gating strategy was used: main population > single-cells > living cells (DAPI negative) > EpCAM+ (PE+) and Ngn3+ (FITC+) / Ngn3- (FITC-) cells. Sorted cells were pooled in a 2:1 (EpCAM+/Ngn3+ : EpCAM+/Ngn3-) ratio and immediately used for nuclei isolation.
For nuclei isolation, a low input nuclei isolation protocol adapted from 10X Genomics was performed. In brief, sorted cells were washed once with 1 mL PBS + 1% BSA, counted with Trypan blue staining, centrifuged. Subsequently, the washed cell pellet was resuspended in chilled lysis buffer (50 μL per sample) and placed on ice for 4 min. Then wash buffer (500 μL per sample) was added and nuclei were centrifuged. To gradually change from wash to diluted nuclei buffer, cells were washed once in a 1:1 mixture of wash buffer and diluted nuclei buffer and subsequently once with pure diluted nuclei buffer. The washed isolated nuclei were then resuspended in 7-10 μL diluted nuclei buffer and were, after quality control and counting, immediately used for single-cell Multiome library preparation with a target recovery of 10000 cells. Libraries were prepared using the Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Bundle (10x Genomics, 1000283) according to the manufacturer’s instructions. Libraries were and sequenced on an Illumina NovaSeq6000 platform following the recommendations from 10X Genomics.
scRNA-seq/ATAC-seq

value: embryonic pancreas

tag: enrichment


title: 15-71_Superficial,2N,Batch_1


title: NVF_E15-5_Rep2 [ATAC-Seq]


title: NVF_E16-5_Rep1 [RNA-Seq]

description: 10x Genomics single cell multiome (RNA)

library_source: transcriptomic single cell

molecule: nuclear RNA


title: Liver, fasted, WT, biol rep1


title: BPA.1_25uM

description:
Templated oligo sequencing with the Human Whole Transcriptome 2.1 kit (TempO-Seq, BioSpyder, Carlsbad, CA)
22P_1


title: NVF_E15-5_Rep2 [RNA-Seq]

description: 10x Genomics single cell multiome (RNA)


title: E15_5 [ATAC-Seq]


title: 16-154_Deep,TS,Batch_2

value: Deep


title: human VGLL4 rep3

city: Seattle

first: Sonali

last: Arora

state: WA

address: 1100 Fairview Ave N,

institute: FHCRC

description: human VGLL4 rep3

data_processing:
Sequencing was performed using an Illumina NextSeq 2000 employing a paired-end, 50 base read length (PE50) sequencing strategy.
RNA-seq reads were aligned to the UCSC hg38 assembly using STAR2 and counted for gene associations against the UCSC genes database with HTSeq
Assembly: hg38
Supplementary files format and content: tab-delimited text files include raw values for each Sample.

source_name: human VGLL4

growth_protocol: Ben-Men-1 cells were grown in DMEM (11995-073), 20% FBS, 1% PenStrep and lentivirally transduced with pLJM1-GFP or pLJM1-VGLL4. Two days after transduction positive clones were selected with Puromycin for three days. After three days of selection cells were continously cultured in growth media without puromycin. Cells were seeded into 6 well plates in triplicates (200,000 cells per well) and RNA was extracted using the Qiagen RNAeasy Mini kit.

extract_protocol:
RNA-seq libraries were prepared from total RNA using the TruSeq Stranded mRNA kit (Illumina, Inc., San Diego, CA, USA). Library size distribution was validated using an Agilent 4200 TapeStation (Agilent Technologies, Santa Clara, CA, USA). Additional library QC, blending of pooled indexed libraries, and cluster optimization was performed using Life Technologies’ Invitrogen Qubit® 2.0 Fluorometer (Life Technologies-Invitrogen, Carlsbad, CA, USA).
RNA-seq libraries were pooled (70-plex) and clustered onto an P2-100 flow cell.

treatment_protocol: untreated

value: human VGLL4


title: E15_5 [RNA-Seq]

description: 10x Genomics single cell multiome (RNA)


title: E14_5 [ATAC-Seq]


title: E14_5 [RNA-Seq]

description: 10x Genomics single cell multiome (RNA)


title: 15-75_Deep,2N,Batch_2


title: human VGLL4 rep1

description: human VGLL4 rep1


title: F420 lung met: control

city: Columbus

first: Matt

last: Cannon

state: OH

address: 700 Children's Dr.

department: Center for Childhood Cancer

institute: Nationwide Children's Hospital

description: F420 lung met: control

data_processing:
BCL files were converted to fastq files using cellranger mkfastq
Fastq files were processed using cellranger count
Further processing and analyses performed are outlined on GitHub (https://github.com/kidcancerlab/epithelial)
Assembly: mm10
Supplementary files format and content: Tar files for each sample include the contents of the filtered_feature_bc_matrix folder output by Cellranger count (barcodes.tsv.gz  features.tsv.gz  matrix.mtx.gz)

molecule: polyA RNA

source_name: lung tumor

extract_protocol:
Following dissociation into single nuclei, single-nuclei 3’ RNA-Seq libraries were prepared using Chromium Single Cell 3′RNA-sequencing system (10x Genomics) with the Reagent Kit v3.1 (10XGenomics, PN-1000121) according to the manufacturer's instructions
Libraries were constructed per 10x protocols

value: lung tumor

value: Control


title: human GFP rep3

description: human GFP rep3

source_name: human GFP

value: human GFP


title: F420 lung met: nintedanib treated at day 1

value: Mice were treated with nintedanib on day 1 following F420 injection


title: human GFP rep2

description: human GFP rep2


title: 15-70_Deep,TS,Batch_2


title: F420 lung met: nintedanib treated at day 28

value: Mice were treated with nintedanib on day 28 following F420 injection


title: Mouse brain, 4sU-, rep1

city: Shanghai

first: Yang

last: Yang

address: 393 Middle Huaxia Road, Pudong, Shanghai

institute: ShanghaiTech University

description:
Z8L
counts_4L_5L_13L_8L_b01_b03.outs.txt

data_processing:
Raw data from RNA-Seq were preprocessed using Cutadapt v3.1 for adapter sequence/short sequence removal (parameters - minimum length: 15 nucleotides).
Preprocessed data were aligned to the NCBI reference mouse genome (mm10) using Hisat2 v2.1.0
The resulting alignment data in SAM format was converted to BAM format using Samtools v1.9.
The T>C mutation counts for each sample were computed using the software SLAM-DUNK v0.2.4, designed for SLAM-Seq analysis. Annotation data for 3' UTR regions were obtained from the RefSeq and Ensembl databases (parameters - readlength: 150).
Gene expression quantification was performed using FeatureCounts v2.0.1, utilizing the mm10 reference gene GTF file from the UCSC Genome Browser (parameters: Is paired-end, Check for chimeric fragments, Feature type: exon, Attribute type: transcript_id).
Assembly: mm10
Supplementary files format and content: tab-delimited text file includes raw T>C mutation values and T coverage for each Sample.
Supplementary files format and content: tab-delimited text file includes raw counts for each Sample

source_name: auditory cortex

extract_protocol:
RNA was extracted from the amygdala and auditory cortex of mice using the Trizol method, followed by alkylation treatment of the RNA.
RNA libraries for RNA-seq were prepared using VAHTS® Universal V8 RNA-seq Library Prep Kit following manufacturer's protocols.

treatment_protocol: 4sU (100mM desolved in DMSO) or DMSO was injected stereotaxically into mouse amygdala and auditory cortex 24h prior to RNA extraction.

value: auditory cortex


title: Mouse brain, 4sU-, rep2

description:
S1
counts_4L_5L_13L_8L_b01_b03.outs.txt


title: 16-179_Deep,TS,Batch_2


title: Mouse brain, 4sU-, rep3

description:
S3
counts_4L_5L_13L_8L_b01_b03.outs.txt


title: 15-50_Deep,2N,Batch_2


title: 16-208_Deep,2N,Batch_2


title: human GFP rep1

description: human GFP rep1


title: Mouse brain, 4sU, rep1

description:
Z4L
counts_4L_5L_13L_8L_b01_b03.outs.txt

value: 4-thiouridine injection


title: 16-207_Deep,TS,Batch_2

value: CA1 pyramidal Neurons


title: Malignant serous effusions, baseline RNA-seq, batch 5

city: Zurich

first: Berend

last: Snijder

address: Otto-Stern-Weg 3

department: D-BIOL

country: Switzerland

institute: ETH Zurich

description: Read 1 contains the transcript seqeunce, read 2 the well barcode and UMI.

data_processing:
Read 1 and Read 2 were combined in a single bam file using Picard FastqToSam (v.2.18.14)
Demultiplexing, mapping and UMI counting was performed using code available from github: https://github.com/RebekkaWegmann/drugseq_toolbox
Assembly: GRCh38, v.93
Supplementary files format and content: "_DGE_matrix_UMIs.txt: Tab-delimited file containing digital gene expression (DGE) matrix.  Rows correspond to genes (labeled by the HGNC symbol), columns correspond to individual well barcodes.
Supplementary files format and content: "_barcodes.csv": Comma-separated file providing a mapping from well barcode to Sample ID.

source_name: Malignant serous effusion

extract_protocol:
Cell pelets were lysed in TriZol and RNA was extracted using a Zymo Direct-zol-96 extraction kit for larger batches, and a Zymo Direct-zol kit for batch 1 and 6.
RNA-seq libraries were prepared using a customized version of the DRUG-seq protocol (Ye et al., 2018, PMID  PMC9207813) with the following modifications: 1) we changed the reverse transcription (RT) primer sequences to make them compatible with standard Illumina sequencing primers, 2) we start from purified RNA rather than a cell lysate, in our hands this resulted in less gDNA contamination of the sequencing libraries and simplified sample storage, 3) we optimized the protocol for ultra-low input samples by using a Zymo IC column to purify cDNA after RT and eluting in only 10 μl of hot (80C) water, and by using the Nextera XT kit (Illumina FC-131-1096) for library preparation; 4) we optimized the unique mapping rates of our libraries by performing a stringent size selection with one 0.5X cleanup to remove large fragments followed by two times 0.6X cleanup using KAPA pure beads (KAPA KK8000) resulting in fragment sizes between 500-800bp.
RNA-seq  (3' digital gene expression)

value: Malignant serous effusion


title: 16-189_Deep,2N,Batch_2


title: 16-04_Deep,2N,Batch_2


title: Fmr1 knock out, female, 2

city: Tallahassee

first: Cynthia

last: Vied

state: Florida

address: 1115 W. Call St

department: Translational Science Laboratory

institute: Florida State University

data_processing:
Adapters were trimmed during demultiplexing by the core facility and trimmed reads were aligned to the genome using RNA-Seq Alignment version 1.1.1 (Illumina BaseSpace software application). The application uses STAR for alignment to the mouse genome (UCSC mm10) and to generate count tables.
Differential expression analysis of count tables was performed using the default settings of DESeq2 (Benjamini-Hochberg FDR correction).
Assembly: mm10 (GRCm38)
Supplementary files format and content: Microsoft Excel Worksheet (.xlsx). Contains normalized counts generated by DESeq2.

source_name: Ventral Striatum

growth_protocol: Eight-week-old male and female C57BL/6J (https://www.jax.org/strain/000664) and Fmr1 KO (https://www.jax.org/strain/003025) mice were purchased from Jackson Laboratories (Bar Harbor, ME). Mice were maintained in a temperature-controlled environment with a 12/12hr light/dark cycle with food and water available ad libitum

extract_protocol:
Mice were anesthetized (Isoflurane, Pivetal, Liberty, MO), decapitated, and the ventral striatum was collected by microdissection. Samples were snap-frozen in liquid nitrogen.and stored at -80 degrees. Total RNA was extracted from the ventral striatum from each animal using  a RNeasy mini kit (Qiagen, 74104) according to manufacturer’s directions.
A cDNA library was prepared for each sample using the Illumina TruSeq stranded mRNA library kit. Libraries were barcoded for multiplexing with IDT for Illumina 8 nucleotide i7 and i5 unique dual indexes. The multiplexed sample was quantified with qPCR (Kapa Biosystems) specific for Illumina sequencing primers and the average fragment size was determined with a Bioanalyzer high sensitivity DNA chip (Agilent Technologies). The pooled sample was sequenced, with paired-end, 150 base reads on an Illumina NovaSeq 6000 located in the Translational Science Laboratory at the College of Medicine, Florida State University.  The pooled data were demultiplexed into individual sample data.

treatment_protocol: The line was maintained by breeding hemizygous male Fmr1 mice with homozygous female Fmr1 mice. Wild-type (WT) mice of the same strain were used as controls. Adult mice (2-3month old)  completed a battery of behavioral tests. Following completion of behavior, tissue was collected.

value: Ventral Striatum

value: Female

value: Fmr1 KO


title: Fmr1 knock out, female, 1


title: Fmr1 knock out, female, 4


title: human VGLL4 rep2

description: human VGLL4 rep2


title: Fmr1 knock out, male, 3

value: Male


title: 16-177_Deep,TS,Batch_2

value: TS


title: 16-319_Deep,TS,Batch_1


title: Malignant serous effusions, baseline RNA-seq, batch 4


title: Malignant serous effusions, baseline RNA-seq, batch 2


title: 16-268_Deep,2N,Batch_1


title: Wild type, female, 3

value: WT


title: Wild type, female, 4


title: Wild type, female, 1


title: 16-267_Deep,TS,Batch_1

value: Deep


title: Mouse brain, 4sU, rep3

description:
Z13L
counts_4L_5L_13L_8L_b01_b03.outs.txt


title: Drug treated cells from sample FB216 (LUAD pleural effusion), multiplexed RNA-seq, plate 2

description: Read 1 contains the transcript sequence, read 2 the well barcode and UMI

data_processing:
Read 1 and Read 2 were combined in a single bam file using Picard FastqToSam (v.2.18.14)
Demultiplexing, mapping and UMI counting was performed using code available from github: https://github.com/RebekkaWegmann/drugseq_toolbox
Assembly: GRCh38, v.93
Supplementary files format and content: "_DGE_matrix_UMIs.txt: Tab-delimited file containing digital gene expression (DGE) matrix.  Rows correspond to genes (labeled by the HGNC symbol), columns correspond to individual well barcodes.
Supplementary files format and content: "_barcodes.csv": Comma-separated file providing a mapping from well barcode to perturbation.

source_name: Malignant pleural effusion of lung adenocarcinoma

extract_protocol:
Cells after drug perturbation were lysed in a 96-well plate using TriZol-LS. RNA was extracted using a Zymo Direct-zol-96 extraction kit.
RNA-seq libraries were prepared using a customized version of the DRUG-seq protocol (Ye et al., 2018, PMID  PMC9207813) with the following modifications: 1) we changed the reverse transcription (RT) primer sequences to make them compatible with standard Illumina sequencing primers, 2) we start from purified RNA rather than a cell lysate, in our hands this resulted in less gDNA contamination of the sequencing libraries and simplified sample storage, 3) we optimized the protocol for ultra-low input samples by using a Zymo IC column to purify cDNA after RT and eluting in only 10 μl of hot (80C) water, and by using the Nextera XT kit (Illumina FC-131-1096) for library preparation; 4) we optimized the unique mapping rates of our libraries by performing a stringent size selection with one 0.5X cleanup to remove large fragments followed by two times 0.6X cleanup using KAPA pure beads (KAPA KK8000) resulting in fragment sizes between 500-800bp.
RNA-seq  (3' digital gene expression)


title: Mouse brain, 4sU, rep2

description:
Z5L
counts_4L_5L_13L_8L_b01_b03.outs.txt


title: Fmr1 knock out, female, 3


title: Wild type, female, 2


title: Wild type, male, 4


title: 16-257_Deep,2N,Batch_1


title: Drug treated cells from sample FB216 (LUAD pleural effusion), multiplexed RNA-seq, plate 1

value: Malignant pleural effusion of lung adenocarcinoma


title: Fmr1 knock out, male, 4


title: Wild type, male, 3


title: Drug treated cells from sample FB212 (LUAD pleural effusion), multiplexed RNA-seq, plate 2


title: 16-178_Deep,TS,Batch_1


title: Malignant serous effusions, baseline RNA-seq, batch 3


title: Wild type, male, 1


title: Fmr1 knock out, male, 1


title: Wild type, male, 2


title: 15-72_Deep,TS,Batch_1


title: Drug treated cells from sample FB209 (LUAD pleural effusion), multiplexed RNA-seq, plate 2


title: Drug treated cells from sample FB212 (LUAD pleural effusion), multiplexed RNA-seq, plate 1


title: C2AKT1F-4-4_1

city: Incheon

first: Byung-Chul

last: Oh

address: 155 Gaetbeol-ro, Yeonsu-gu

department: Physiology

country: South Korea

institute: Lee Gil Ya Cancer and Diabetes Institute,Gachon University

data_processing:
Illumina bcl2fastq2-v2.20.0 software used for basecalling.
Trimmomatic program is used to remove adapter sequences and bases with base quality lower than three from the ends. Also using sliding window method, bases of reads that does not qualify for window size 4, and mean quality 15 are trimmed. Afterwards, reads with length shorter than 36bp are dropped to produce trimmed data.
Sequenced reads were mapped to GRCm38 (mm10) whole genome by using HISAT2 version 2.1.0 with parameter -p 20 -dta --rna-strandness RF --novel-splicesite-outfile -x -1 -2
Fragment Per Kilobase of exon per Megabase of library size (FPKM) were calculated by using StringTie version 2.1.3b, with parameter --rf -p 20 -l -o -G -A -C -e
Assembly: GRCm38 (mm10)
Supplementary files format and content: tab-delimited text files include FPKM values for each Sample

growth_protocol: Mice were maintained on a 12-hour light/dark cycle and allowed free access to a standard diet and water for 8 weeks of age.

extract_protocol:
RNA was harvested using Trizol reagent.
RNA libraries were prepared for sequencing using standard Illumina protocols

treatment_protocol: At 8 weeks of age, we fed 8-week-old male C2-AKT mice or control mice with a high-fat diet (HFD) for 10 weeks.

value: Liver

value: male

value: HFD for 10 weeks


title: C2AKT1F-4-5_2


title: Drug treated cells from sample FB209 (LUAD pleural effusion), multiplexed RNA-seq, plate 1


title: C2AKTWT-2-3_1

value: wild type; C2-AKT (-/-)


title: C2AKTWT-2-4_1


title: C2AKTWT-2-5


title: Drug treated cells from sample FB184 (LUAD pleural effusion), multiplexed RNA-seq, plate 1


title: 15-49_Deep,2N,Batch_1


title: 15-51_Deep,TS,Batch_1


title: Drug treated cells from sample FB184 (LUAD pleural effusion), multiplexed RNA-seq, plate 2


title: C2AKT1F-4-2


title: C2AKTWT-2-2_1


title: 15-224_Deep,TS,Batch_1


title: 15-223_Deep,2N,Batch_1


title: Drug treated cells from sample FB170 (LUAD pleural effusion), multiplexed RNA-seq, plate 2


title: CHIRP Lac_Odd

city: Stockholm

first: Ning

last: Xu Landén

address: K2 Medicin, Solna, K2 Derm o Ven Xu Landén N

country: Sweden

institute: Karolinska Institutet

description:
SNHG26 CHIRP-Seq
CHIRP_peaks_filtered_anno_Pval001.xlsx

data_processing:
ChIP-seq and CHIRP-seq: Raw sequencing data were quality assessed using Trimmomatic v0.36 by trimming reads of low qualities and with a length less than 20 nucleotides. Clean reads were mapped to GRCh38 human reference genome using Bowtie2 followed by removing PCR duplicates using Picard (v2.20.4) tools. The mapped reads were converted into bigwig format for IGV viewer using deepTools bamCoverage function. Peaks were called using MACS2 (v2.2.6) with the default parameter. Peaks from two biological replicates were merged and filtered using Bedtools.
scRNA-seq: Raw single cell sequencing data were processed using the standard 10X Cell Ranger (v5.0.1) analysis workflow, including demultiplexing, aligning to the GRCm39 (mm39) mouse reference genome, barcode counting and unique molecular identifier (UMI) quantification.
Assembly: GRCh38 for human;GRCm39 for mouse
Supplementary files format and content: Excel file contains the peaks of CHIP-seq.
Supplementary files format and content: tar files contain the raw count data of mouse scRNA-seq.
Supplementary files format and content: Excel file contains the peaks of CHIRP-seq.
Library strategy: ChIRP-seq

source_name: Keratinocytes

growth_protocol: Human adult primary keratinocytes (C0055C; Thermo Fisher) were cultured in Epilife medium supplement with Human Keratinocyte Growth Supplement (HKGS, S0015, Thermo Fisher) and 1X Penicillin-Streptomycin. Snhg26-Ko mice and WT mice were breed in specific pathogen-free conditions in Comparative Medicine Biomedicum (KMB) animal facility in Karolinska Institutet. All the mouse experiments were approved by committee on animal experimentation of Swedish Board of Agriculture (Jordbruksverket).

extract_protocol:
The genomic DNA was isolated by using the DNA Purification Magnetic Beads.
The DNA was purified by phenol–chloroform–isoamyl alcohol and subjected to NEBNext ChIP-seq libraries preparation. The Paired-end sequencing libraries are sequenced by Illumina high-throughput sequencer (DNBSEQ PE100).

value: Keratinocytes

tag: gender

value: not determined

value: LacZ Odd

tag: chip antibody


title: Drug treated cells from sample FB170 (LUAD pleural effusion), multiplexed RNA-seq, plate 1

tag: tissue


title: C2AKT1F-4-3_1


title: 11719GE0006TXP-N__DI_PF

city: Coventry

first: Erin

last: Greaves

address: Gibbet Hill Campus

department: Division of Biomedical Sciences

country: United Kingdom

institute: University of Warwick

data_processing:
Initial processing was performed using Cellranger (v2.1.1) mkfastq and count (aligned to mouse assembly mm10). For each dataset (filtered data from Cell Ranger 27 pipeline), potentially low-quality cells were filtered out using dataset-specific thresholds
Assembly: mm10
Supplementary files format and content: Tab-separated values and matrix files

source_name: Peritoneal lavage cells

extract_protocol:
Lesions were dissected, pooled from each mouse and placed in 2ml ice-cold DMEM. Tissues were cut into small pieces using a scalpel and digested. Peritoneal lavage cells were recovered by injected 7ml ice cold RPMI into the peritoneal cavoity, followed by gentle massage and recovery. RBCs were lysed and cell were blocked. 200,000 CD45+ cells were FACS sorted into 1ml PBS + 2% FBS using a FACS Fusion.
Cells were barcoded using a 10X Genomics Chromium ControllerTM using established pipelines

value: Peritoneal lavage cells

tag: cell type

value: Leukocytes

value: Experimental endometriois (intact)


title: 11719GE0007TXP-N__Na_ve_PF

value: Naïve PF (intact)


title: 11325GEPool04-N__Lesions

source_name: Endometriosis lesions

value: Endometriosis lesions

value: Experimental endometriosis


title: 11325GEPool03-N__Endo_PF


title: 15-131_Deep,2N,Batch_1

value: CA1 pyramidal Neurons

tag: genotype


title: 15-71_Deep,2N,Batch_1


title: CHIRP Input

tag: treatment

value: Input


title: Drug treated bulk MSE and FACS purified cell populations from MSE, RNA-seq, multiplexed

source_name: Malignant serous effusions (various samples)


title: 16-179_ALL,TS,Batch_2

value: ALL

value: TS

tag: batch


title: CHIRP Lac_Even

value: LacZ Even


title: 11325GEPool02-N__Sham_PF

value: Sham


title: Malignant serous effusions, baseline RNA-seq, batch 6

value: Malignant serous effusion


title: 11325GEPool01-N__Decid_tissue

source_name: Menses-like' endometrium

value: Menses-like' endometrium

value: Donor endometrium


title: x.laevis_rep1_ivt

city: Barcelona

first: Gregor

last: Diensthuber

state: Barcelona

address: C/ Dr. Aiguader, 88

department: Genome Biology

country: Spain

institute: Center for Genomic Regulation (CRG)

data_processing:
Processing of raw fast5 files was peformed using the MasterOfPores nextflow pipeline. Reads were basecalled using guppy (v6.0.6) with each of the three tested basecalling models (default, ivt and sup).
Alignments to both reference genome ( -uf -ax splice -k14)  and transcriptome (-ax map-ont -k14) were performed using minimap2 (v2.17). Highly modified synthetic constructs and the corresponding unmodified control were mapped using graphmap (v0.5.2) with sensitive settings (--rebuild-index -v 1 --double-index --mapq -1 -x sensitive -z 1 -K fastq --min-read-len 0 -A 7 -k 5)
To extract error signature on a per-nucleotide basis, EpinanoRMS (v1.1) was run. For in vivo m6A analyisis, pairwise-comparisons between wildtype and knockout samples were performed using eligos2(v2.1.0). Cross-species comparison of read accuacies was performed on reads uniquely mapped to the reference transcriptome (-F 3840) with a mapping quality > 15 (-q 15). To balance reads for strongly varying expression profiles at most the 10 longest reads for each transcript were extracted (bam2select.py). Read identis were extracted using identity_density.py
Assembly: Reference files include curlcake_constructs_EcoRV_BamHI_digestion.fasta, riboswitches.fa and 2021-8_eGFP_mRNA.v2.fa for curlcakes, riboswitches and the syntehtic eGFP vaccine, respectively. We used the following genomic reference for H. sapiens : Homo_sapiens.GRCh38.dna_sm.primary_assembly.fa. And transcriptomic references Homo_sapiens.GRCh38.cdna.all.fa, Mus_musculus.GRCm39.cdna.all.fa, xlaevisMRNA.fasta, Saccharomyces_cerevisiae.R64-1-1.cdna.all.fa and Arabidopsis_thaliana.TAIR10.cdna.all.fa for H. sapiens, M. musculus, X. laevis, S. cervisiae and A. thaliana, respecitvely.
Supplementary files format and content: Files ending in <model>_per_position.csv, contain EpinanoRMS results on a per position basis. Files ending in <model>_per_5mer.tsv contain EpinanoRMS results on a 5-mer basis generated by converting per position results using epinano_to_kmer.R.
Supplementary files format and content: Files ending in _eligos2.txt contain results generated by eligos2 for pairwise comparisons which were already filtered to remove  non-A-positions and homopolymer sequences.
Supplementary files format and content: Files starting with <species>.tsv contain read accuracies per aligned read (rows) for all three models (column 1).

organism: Xenopus laevis

source_name: embyo

extract_protocol:
In vitro constructs were artifically poly-A-tailed using PAP (NEB, M0276S), and cleaned up using the RNEasy MinElute kit (Quiagen,  74204) for curlcake constructs and Zymo Clean and Concentrator kit (Zymo Resarch, R1013). Concentrations were determined using a Qubit 2.0 Fluorometer. For libraries that were not generated as part of this work please refer to the respective publications
Direct RNA-seqeuncing libraries were generated using the SQK-RNA002 according to the manufacturer's protocol (version : DRS_9080_v2_revR_14Aug2019-minion). For curlcakes a final library of 800ng war generated (200ng per curlcake construct) while 750ng for the UNM-S libraries was used (250ng per riboswitch). The reverse transcription step was peformed using SuperScript III R(Thermo Fisher Scientific, 18080044) in the case of ac4C, m5U-containing curlcake constructs and UNM-S, and Superscript IV (Thermo Fisher Scientific, 18090010) in the case of m1Ψ. For libraries that were not generated as part of this work please refer to the respective publications.

value: embyo


title: x.laevis_rep1_sup


title: CHIRP AC2_Odd

value: Keratinocytes

value: not determined

value: AC2  Odd


title: 16-154_ALL,TS,Batch_2

source_name: CA1 pyramidal Neurons


title: 16-04_ALL,2N,Batch_2


title: CHIRP AC2_Even

value: AC2 Even


title: x.laevis_rep1_default


title: As4.1 cells, Input

city: Charlottesville

first: Jason

middle: Paul

last: Smith

state: VA

address: 409 Lane Rd

department: Pediatrics

institute: University of Virginia

data_processing:
Raw sequencing reads were processed using the `nf-core/chipseq` pipeline (version 2.0.0) with alignment to the GRCm38/mm10 mouse genome. Enriched genomic regions for both H3K27ac and p300 were determined using MACS2 (Model-based analysis of ChIP-seq 2) with default settings by comparing ChIP to input samples.  Differential binding analysis was then performed using the `DiffBind` R package. Differentially enriched regions were annotated using the `ChIPpeakAnno` and `clusterProfiler` R packages.
Assembly: GRCm38/mm10
Supplementary files format and content: bigWig - merged signal tracks, BED - consensus or enriched peaks (except input)

source_name: kidney

growth_protocol: The renin-expressing As4.1 cell line (ATCC, CRL-2193) was maintained in high glucose DMEM (supplemented with 10% fetal bovine serum (both from Thermo Fisher Scientific) at 37°C in a humidified incubator containing 5% CO2.

extract_protocol:
Cells were then fixed according to manufacturer protocol.
Library construction and sequencing were performed by Active Motif (Carlsbad, CA) following manufacturer protocols.

treatment_protocol: Cells were treated with 10 μM H89 or equivalent volume DMSO at 0, 24, and 48 hours.

value: kidney

tag: cell line


title: 15-75_ALL,2N,Batch_2


title: CHIRP AC1_Odd

value: AC1 Odd


title: 15-70_ALL,TS,Batch_2


title: As4.1 cells, H89, p300, rep2


title: CHIRP AC1_Even

value: AC1 Even


title: 15-50_ALL,2N,Batch_2

value: CA1 pyramidal Neurons


title: Ndrg3 knockout CD8SP, biol rep 4

city: Freiburg

first: Jeremy

last: Swann

address: Stübeweg 51

department: Developmental Immunology

institute: Max Planck Institute of Immunobiology and Epigenetics

data_processing:
The RNAseq libraries were analysed using the mRNAseq snakepipes pipeline (v.2.5.3)
Reads were trimmed with cutadapt and then aligned to the mouse genome (GRCm38.p4, release M9) using STAR mapper
The gene-level count matrix was generated using featureCounts
Assembly: GRCm38.p4, M9
Supplementary files format and content: Tab-delimited text file containing raw counts for all samples

source_name: thymus

extract_protocol:
RNA was extracted from sorted cells using the RNeasy Plus Micro Kit (Qiagen)
Libraries were constructed with the Illumina Stranded Total RNA w/ Ribo-Zero Plus Kit. Library preparation was carried out by the Deep Sequencing core facility at the Max Planck Institute for Immunobiology and Epigenetics according to their standard protocols.

value: thymus

value: CD8SP TCRb+ MHC-I high

value: Ndrg3 fl/fl;Lck-cre+

value: 6-7 weeks old


title: Ndrg3 control CD8SP, biol rep 3

value: Ndrg3 +/fl;Lck-cre+


title: Ndrg3 control CD8SP, biol rep 2


title: CHIP Input

description:
ILF2 CHIP-Seq
s0 raw Peaks annotation ChIPseq_Jul2021.xlsx

data_processing:
ChIP-seq and CHIRP-seq: Raw sequencing data were quality assessed using Trimmomatic v0.36 by trimming reads of low qualities and with a length less than 20 nucleotides. Clean reads were mapped to GRCh38 human reference genome using Bowtie2 followed by removing PCR duplicates using Picard (v2.20.4) tools. The mapped reads were converted into bigwig format for IGV viewer using deepTools bamCoverage function. Peaks were called using MACS2 (v2.2.6) with the default parameter. Peaks from two biological replicates were merged and filtered using Bedtools.
scRNA-seq: Raw single cell sequencing data were processed using the standard 10X Cell Ranger (v5.0.1) analysis workflow, including demultiplexing, aligning to the GRCm39 (mm39) mouse reference genome, barcode counting and unique molecular identifier (UMI) quantification.
Assembly: GRCh38 for human;GRCm39 for mouse
Supplementary files format and content: Excel file contains the peaks of CHIP-seq.
Supplementary files format and content: tar files contain the raw count data of mouse scRNA-seq.
Supplementary files format and content: Excel file contains the peaks of CHIRP-seq.

extract_protocol:
The genomic DNA was isolated by using the DNA Purification Magnetic Beads.
After electrophoresis, DNA fragments of desired length are gel purified. Adapter ligation and DNA cluster preparation are performed. The DNA was then subjected to NEBNext ChIP-seq libraries preparation (PE 100).

value: Input

value: none (input)


title: Ndrg3 knockout CD8SP, biol rep 3


title: 16-208_ALL,2N,Batch_2

value: ALL


title: CHIP Ctr-ASO2

value: Ctrl


title: Ndrg3 knockout CD8SP, biol rep 2


title: 16-207_ALL,TS,Batch_2

value: TS


title: CHIP Ctr-ASO1

value: Keratinocytes

value: not determined


title: As4.1 cells, DMSO, p300, rep1

value: DMSO


title: Ndrg3 control CD8SP, biol rep 4


title: Ndrg3 knockout CD8SP, biol rep 1


title: 16-189_ALL,2N,Batch_2


title: As4.1 cells, H89, H3K27ac, rep2

value: kidney

value: H3K27ac (Abcam, #ab4729)


title: Ndrg3 control CD8SP, biol rep 1


title: CHIP SNHG26-ASO2

value: TNF


title: As4.1 cells, H89, p300, rep1


title: As4.1 cells, H89, H3K27ac, rep1


title: 16-177_ALL,TS,Batch_2

value: CA1 pyramidal Neurons


title: ChIP SNHG26-ASO1


title: 16-268_ALL,2N,Batch_1


title: As4.1 cells, DMSO, H3K27ac, rep2


title: As4.1 cells, DMSO, H3K27ac, rep1


title: 16-267_ALL,TS,Batch_1


title: As4.1 cells, DMSO, p300, rep2


title: heterozygous_mouse_male [703]

city: London

first: Nazif

last: Alic

address: Gower St

institute: UCL

data_processing:
Read quality assessed with FastQC, and trimming has been done using cutadapt
Aligned to mouse genome and counted with Salmon
Analysed with DESeq2
Supplementary files format and content: containing read counts for genes analysed in corresponding samples

source_name: duodenum

extract_protocol:
Trizol
Glasgow Polyomics

treatment_protocol: size selection of RNAs less than 200 nucleotides in length using mirVANA RNA isolation kit AND demethylated using rtStar™ tRF&tiRNA Pretreatment Kit

value: duodenum

value: male


title: heterozygous_mouse_male [702]


title: heterozygous_mouse_male [705]


title: 16-257_ALL,2N,Batch_1

data_processing:
The reads with adapter contamination/ low quality were trimmed using Trimmomatic 0.39.
Trimmed sequence reads were aligned to the reference  genome (GRCm39-mm10) using STAR Aligner. Gene counts were extracted using RSEM.
Downstream Analysis was performed using R with the gene counts obtained from the RSEM. Genes with over 0.1 counts per million were retained followed by TMM normalization.
Assembly: mm10
Supplementary files format and content: tab delimited text files with gene counts of all samples in a region per one file
Supplementary files format and content: tab delimited text files with gene counts of one sample per one file

extract_protocol:
Brain tissues were accessed from Ts2 and normal disomic female mice. Mice were transcardially perfused with ice-cold 0.15 M phosphate buffer as previously described. Brains were immediately flash frozen and 20 μm-thick tissue sections were cryostat cut (−25°C) in the coronal plane and mounted on polyethylene naphthalate (PEN) membrane slides. Slides were kept under desiccant at -80°C until used for immunohistochemistry. RNase-free precautions were employed, and solutions were made with 18.2 mega Ohm RNase-free water. RNA from Batch 1 and Batch 2 for each CA1 subregion was purified using miRNeasy Micro kit (Qiagen) according to manufacturers’ specifications. A DNase digestion was performed twice sequentially before the final washes and RNA purification. LCM was performed on tissue to isolate the entire rostral CA1 pyramidal neuron layer from 3-6 sections (CA1A, ~500-600 pyramidal neurons) and compared that to the pyramidal neurons from the distal deep layer CA1 region(CA1D; 6-12 sections; ~500-1500 cells) and to the distal superficial layer CA1 region (CA1D; 6-12 sections; ~500-1500 cells).  Deep and superficial neurons were selected from the same tissue sections.  Batch 2 collected CA1A, CA1D and CA1S from adjacent sections within each slide (2-4 slides per brain to collect at least ~500 neurons per region), whereas batch 1 was collected with CA1D and CA1S in adjacent slides to the CA1A slide.
The SMARTer Stranded Total RNA-Seq kit-Pico input Mammalian (Takara Bio, Mountain View, CA) was employed with minor modifications to normalize neuronal numbers for each sample.
RNA-seq samples were quantified (Agilent 2100 HS DNA kit), with all samples reading a minimum of 2nM of the library for QC. For each sample run, library samples were pooled in equimolar concentrations and assayed on an Illumina HiSeq-4000 using a single read 50-cycle protocol at NYUGSOM Genome Technology Center.  Samples were run in multiple batches utilizing the same protocol for each batch.


title: CHIP IgG

value: IgG


title: 15-72_ALL,TS,Batch_1


title: 15-71_ALL,2N,Batch_1


title: 16-319_ALL,TS,Batch_1


title: 16-178_ALL,TS,Batch_1


title: wt1_mouse_male [758]


title: 15-49_ALL,2N,Batch_1


title: heterozygous_mouse_female [715]

value: female


title: 15-224_ALL,TS,Batch_1


title: 15-223_ALL,2N,Batch_1


title: heterozygous_mouse_female [712]


title: wt3_mouse_male [760]


title: 15-131_ALL,2N,Batch_1


title: wt3_mouse_female [752]


title: 15-51_ALL,TS,Batch_1


title: s24_BMSC36_rep3

city: Jacksonville

first: Yingxue

last: Ren

state: FL

address: 4500 San Pablo Rd S

institute: Mayo Clinic

data_processing:
mRNA samples were sequenced using an Illumina HiSeq 4000 platform. Reads were mapped to the human reference genome hg38. Raw gene read counts, along with sequencing quality control, were generated using the Mayo Clinic RNA-Seq analytic pipeline MAP-RSeq Version 3.0
Conditional quantile normalization (CQN) was performed on the raw gene counts to correct for gene length differences, GC bias, and global technical variations and to obtain similar quantile-by-quantile distributions of gene expression levels across samples
Based on the bimodal distribution of the CQN-normalized and log2-transformed reads per kilobase per million (RPKM) gene expression values, genes with an average log2 RPKM ≥ 2 in at least one group were considered expressed. Using this selection threshold, 16,603 genes were included in the downstream analysis.
Assembly: hg38
Supplementary files format and content: tab delimited text file with raw gene counts of all samples

source_name: iPSC

growth_protocol: Human iPSCs were generated from two healthy individuals (MC0039 and MC0063) and cultured in TeSR-E7 complete medium (STEMCELL Technologies, Vancouver, Canada) on Matrigel (Corning, Corning, NY, USA)-coated dishes, as reported previously

extract_protocol:
RNA was isolated using a Direct-zol™ RNA Miniprep kit (Zymo Research, R2053).
The samples were processed according to the manufacturer’s protocol.

value: iPSC

tag: group


title: wt2_mouse_female [751]


title: s23_BMSC36_rep2


title: rpc3_worm

data_processing:
Read quality assessed with FastQC, and trimming has been done using cutadapt
Aligned to DC. elegans genome and counted with Salmon
Analysed with DESeq2
Supplementary files format and content: containing read counts for genes analysed in corresponding samples

organism: Caenorhabditis elegans

source_name: whole worm

extract_protocol:
Direct-ZolRNAMiniPrepKit (ZymoResearch)
Glasgow Polyomics

value: whole worm

value: worm with rpc-1 RNAi


title: BNSC-3_(Rat230_2)

type: RNA

city: Beijing

first: Sumei

last: Liu

address: Nanyitiao 3, zhongguancun

institute: Chinese Academy of Sciences

hyb_protocol: Array hybridization and wash was performed using GeneChip® Hybridization, Wash and Stain Kit (Cat#900720, Affymetrix, Santa Clara, CA, US)in Hybridization Oven 645 (Cat#00-0331-220V, Affymetrix, Santa Clara, CA, US)and Fluidics Station 450 (Cat#00-0079, Affymetrix, Santa Clara, CA, US) followed the manufacturer’s instructions.

scan_protocol: Slides were scanned by GeneChip® Scanner 3000 (Cat#00-00212, Affymetrix, Santa Clara, CA, US) and Command Console Software 4.0 (Affymetrix, Santa Clara, CA, US) with default settings.

data_processing: Raw data were normalized by MAS 5.0 algorithm, Affy packages in R.

label: biotin

organism: Rattus norvegicus

source_name: Brain of postnatal 3-5 days  Sprague Dawley rat

label_protocol: Total RNA were amplified, labeled and purified by using GeneChip 3' IVT PLUS Reagent Kit (Cat#902416, Affymetrix, Santa Clara, CA, US) followed the manufacturer’s instructions to obtain biotin labeled cRNA.

growth_protocol: The NSCs samples were harvested in liquid nitrogen at once.

extract_protocol: Total RNA was extracted using TRIZOL Reagent (Cat#15596-018，Life technologies, Carlsbad, CA, US)following the manufacturer’s instructions and checked for a RIN number to inspect RNA integrity by an Agilent Bioanalyzer 2100 (Agilent technologies, Santa Clara, CA, US).Qualified total RNA was further purified by RNeasy micro kit (Cat#74004, QIAGEN, GmBH, Germany) and RNase-Free DNase Set (Cat#79254, QIAGEN, GmBH, Germany).

treatment_protocol: After grinded on ice, the samples were collected for further RNA extraction.

value: Postnatal day 3-5

value: brain

value: Sprague Dawley rat


title: rpc1_worm


title: wt1_mouse_female [750]


title: m.musculus_rep1_sup

source_name: brain


title: SNSCfenhua-1_(Rat230_2)

source_name: Spinal cord of postnatal 3-5 days  Sprague Dawley rat

value: spinal cord


title: SNSCfenhua-2_(Rat230_2)


title: SNSCfenhua-3_(Rat230_2)


title: heterozygous_mouse_female [709]


title: BNSCfenhua-3_(Rat230_2)


title: wt3_worm

value: worm with empty-vector control


title: m.musculus_rep1_ivt


title: SNSC-3_(Rat230_2)


title: s20_MC0039-3_rep2


title: SNSC-2_(Rat230_2)


title: SNSC-1_(Rat230_2)


title: BNSCfenhua-1_(Rat230_2)


title: BNSCfenhua-2_(Rat230_2)


title: wt2_mouse_male [759]


title: BNSC-1_(Rat230_2)


title: m.musculus_rep1_default


title: rpc2_worm


title: s21_MC0039-3_rep3


title: wt2_worm


title: s19_MC0039-3_rep1


title: s18_MC0039-2_rep3


title: Shield IgG rep 2

city: Pittsburgh

first: Miler

middle: T

last: Lee

state: PA

address: 4249 Fifth Ave

department: Biological Sciences

institute: University of Pittsburgh

data_processing:
Paired-end reads were mapped to the GRCz11 zebrafish genome assembly using bowtie2 v2.4.2 (Langmead and Salzberg, 2012) (--no-mixed --no-discordant). High-quality alignments (MAPQ ≥ 30) were retained, after additional filtering to also exclude reads mapping chrM, or to satellite DNA or rRNA as annotated by RepeatMasker.
For bigWigs, only mononucleosome sized fragments (140-250 bp) were retained. Each read pair was joined end-to-end then tag-centered to 73 bp. BigWigs normalized to total number of mononucleosome fragments per million were made using bedtools v2.30.0 genomecov and wigToBigWig. For triptolide and DMSO treatments, coverage was normalized to total yeast aligned reads per million
Assembly: Zebrafish GRCz11
Supplementary files format and content: bigWig files of mononucleosome coverage pooled across replicates. Fragment per million normalized; or yeast spike-in normalized (labeled with "spikein")
Library strategy: CUT&RUN

organism: Danio rerio

source_name: Whole embryos

extract_protocol:
The CUT&RUN procedure was adapted from Hainer et al (Hainer and Fazzio, 2019), which incorporates optimizations of the method of Skene and Henikoff (Skene and Henikoff, 2017). Embryos were subjected to either mechanical yolk separation or chemical yolk lysis. For chemical yolk lysis, embryos were transferred to 1.5 mL Eppendorf tubes removing excess liquid with a P200 pipettor, then yolk lysis buffer added (55 mM NaCl,1.8 mM KCl, 1.25 mM NaHCO3). Tubes were shaken at 1100 RPM for 5’ at room temperature (Eppendorf ThermoMixer C), centrifuged at 300xg for 30s to pellet, yolk lysis buffer drawn off, and 1 mL Yolk Lysis Wash Buffer was added (110 mM NaCl, 3.5 mM KCl, 2.7 mM CaCl2, 10 mM Tris pH 8.5). Tubes were shaken at 1100g for 2 minutes at room temperature, centrifuged at 300g to pellet, and supernatant was again removed and replaced with Nuclear Extraction Buffer (20 mM HEPES-KOH, pH 7.9, 10 mM KCl, 500 µM spermidine, 0.1% Triton X-100, 20% glycerol). For manually de-yolking, yolks were pinched off using Dumont #5 forceps in Ca-free Ringer’s Solution (116 mM NaCl, 2.9 mM KCl, 5 mM HEPES, ph 7.2). Deyolked embryonic cells were transferred via Pasteur pipette into Nuclear Extraction Buffer taking care to transfer as little liquid as possible, mixed with gentle pipetting with a P200 pipettor, and then centrifuged at 600xg at 4ºC for 3 min. The free nuclei were then bound to 150 µL of activated concanavalin A beads (Polysciences #86057) at RT for 10mins. Nuclei were blocked for 5 min (5 mL Wash Buffer with 2 mM EDTA) at RT then incubated in 1:100 dilution of primary antibody for 2 hr at 4ºC, washed with Wash Buffer, incubated in a 1:200 dilution of pAG-MNase (gift from Sarah Hainer) for 1 hr at 4ºC, and washed again with Wash Buffer. The bound pAG-MNase was activated with 2 mM CaCl2 and allowed to digest for 30 mins at 0ºC, then stopped using 2x STOP buffer (200 mM NaCl, 20 mM EDTA, 4 mM EGTA, 50 µg/mL RNase A, 40 µg/mL glycogen). Yeast mononucleosomes were added to 2x STOP buffer at 10 pg/mL (untreated samples) or 20 pg/mL (triptolide/dmso samples) as a spike-in. Nuclei were incubated at 37ºC for 20 min followed by centrifuging for 5 min at 16,000xg, drawing off the DNA fragments with the supernatant. The extracted fragments were treated with SDS and proteinase K at 70ºC for 10 min followed by phenol chloroform extraction. Purified DNA was resuspended in 50 µL of water and verified by Qubit dsDNA high sensitivity and Fragment Analyzer.
CUT&RUN libraries were constructed using the NEB Ultra II DNA library prep kit (NEB #E7645) according to manufacturer’s protocol. DNA was end repaired and then ligated to sequencing adaptors diluted 1:100. Ligated DNA was purified with 0.9x AmpureXP beads and PCR amplified for 15 cycles, then purified again with 0.9x AmpureXP beads. Libraries were run on a 1.5% TBE agarose gel, and a band corresponding to 175 - 650 bp was cut out and gel purified using the NEB Monarch DNA gel extraction kit (#T1020). Concentration was verified by Qubit dsDNA high sensitivity and Fragment Analyzer.

treatment_protocol: For Triptolide experiments, embryos were bathed in blue water (0.06 g/L Instant Ocean + methylene blue 0.1% w/v) with 2uM final Triptolide made from 4mM stock in DMSO. Control DMSO embryos were bathed in 0.05% DMSO final.

value: Whole embryos

value: TUAB

value: WT

tag: developmental stage

value: shield

tag: antibody

value: Invitrogen #10500C


title: BNSC-2_(Rat230_2)

tag: age


title: Shield IgG rep 1


title: s17_MC0039-2_rep2


title: s22_BMSC36_rep1


title: Malignant serous effusions, baseline RNA-seq, batch 1

source_name: Malignant serous effusion

value: Malignant serous effusion


title: Shield H3K4me3 rep 3

value: Invitrogen #711958


title: Shield H3K4me3 rep 2


title: s13_MC0039-1_rep1


title: wt1_worm


title: s12_MC0192-3_rep3


title: Shield H3K4me1 rep 2

value: Invitrogen #710795


title: s.cerevisiae_rep1_sup


title: s16_MC0039-2_rep1


title: Shield H3K4me1 rep 1


title: s14_MC0039-1_rep2


title: Shield H3K4me3 rep 1


title: s.cerevisiae_rep1_default


title: s.cerevisiae_rep1_ivt


title: s15_MC0039-1_rep3


title: Dome IgG Triptolide rep 2

value: dome

value: Triptolide


title: s10_MC0192-3_rep1

value: iPSC


title: Shield H3K4me2 rep 2

value: Invitrogen #710796


title: s8_MC0192-2_rep2


title: s11_MC0192-3_rep2


title: Shield H3K4me2 rep 1


title: unc-week 4 (week0-2 treatment), replicate 1, scRNA-seq

city: Boston

first: Marcelo

last: Falchetti

state: Massachusetts

address: 300 Longwood Ave Karp Family Building, 7th Floor

institute: Boston Children's Hospital

data_processing:
The demultiplexed FASTQ files were processed with the Cell Ranger pipeline.
Assembly: GRCh38
Supplementary files format and content: h5 files containing filtered counts matrices.

source_name: iPSC-derived T cells

extract_protocol:
Cells were pelleted by centrifugate at 300 x g for 10 mins and resuspended in 0.04% BSA in PBS.
Single-cell RNA-sequencing samples were prepared using the Chromium Next GEM Single Cell 3’ Reagent Kits (v3.1 Chemistry Dual Index) following the manufacturer’s user guide protocols from 10x Genomics.

value: iPSC-derived T cells

value: iPSCs


title: unc-week 4 (week0-4 treatment), replicate 2, scRNA-seq


title: unc-week 4 (week0-4 treatment), replicate 1, scRNA-seq


title: Dome IgG DMSO rep 2

value: DMSO


title: s7_MC0192-2_rep1


title: Dome IgG DMSO rep 1


title: unc-week 4 (week0-2 treatment), replicate 2, scRNA-seq


title: unc-week 2, replicate 2, scRNA-seq


title: unc iT unactivated, replicate 2, scRNA-seq

value: UNC0224, no activation


title: unc iT unactivated, replicate 1, scRNA-seq


title: s5_MC0192-1_rep2


title: con-week 4, replicate 1, scRNA-seq


title: s3_ASC29_rep3


title: Dome IgG Triptolide rep 1


title: con-week 2, replicate 2, scRNA-seq


title: s9_MC0192-2_rep3


title: unc-week 2, replicate 1, scRNA-seq


title: s6_MC0192-1_rep3


title: Dome H3K4me2 Triptolide rep 1


title: Dome H3K4me2 Triptolide rep 4


title: s2_ASC29_rep2


title: con-week 4, replicate 2, scRNA-seq


title: YY1 ChIP-seq cold stress

city: Nanjing

first: Kai

last: Li

address: #818, East Tianyuan Road

institute: Nanjing Medical University

data_processing:
Alignment by BWA
Filtering by Samtools
Narrow peaks by Macs2
BigWig files by Deeptools bamCoverage
Deseq2 for differential analysis
Assembly: mm10
Supplementary files format and content: BigWig files
Library strategy: CUT&TAG

source_name: subcutaneous adipose

extract_protocol:
Mature adipocytes were obtained from adipose tissue in the inguinal position of male mice at 8-10 weeks old. Type I collagenase (2mg/ml, Sigma, SCR103) was preconfigured using DMEM basic medium (Gibco, C11995500BT) containing 2% volume of FBS (Gibco), 100 U/ml penicillin, 100 μg/ml streptomycin, and stored in the dark after removal of bacteria by filtration using a 0.22 μM filter. The volume ratio of collagenase to adipose tissue was maintained at 2:1. Then the adipose pad was cut into pieces to 1mm³ and digested in 37℃ water bath for 1h, during which the tube was reversed and mixed every five minutes. The digested adipocytes were filtered (70 μm), then centrifuged at 1000 rpm for 10 min. Up layer mature adipocyte were collected and treated according to the manufacturer’s instructions by using in situ DNA Binding Library for Illumina Cut&Tag kits (Yeasen, 12598E12).
Native nuclei were purified from adipocytes, and washed twice gently with wash buffer (20 mM HEPES pH 7.5; 150 mM NaCl; 0.5 mM Spermidine; 1× Protease inhibitor cocktail). For CUT&Tag, 1x10^5 nuclei bound to Concanavalin A-coated beads were incubated with indicated YY1 primary antibody and second antibody. Transposase pA-Tn5 was used to induce DNA tagmentation, followed by Proteinase K digestion and PCI extraction. The ethanol precipitated DNA was immediately subjected to PCR amplification by Universal and barcoded i7 primer using NEBNext HiFi 2x PCR Master mix. The amplified library was purified without size selection. ChIP-seq libraries were prepared from 3-5ng DNA using NEBNext® Ultra™ II DNA Library Prep Kit (NEB, E7645S), according to the manufacturer’s instructions. Post-amplification libraries were size selected at 250–450bp in length using AMPure XP beads ((Beckman Counter)) and were quantified using the Library Quantification Kit for Illiumina (Kapa Biosystems, KK4603). The CUT&Tag Library was pooled to a final concentration of 10nM and sequenced paired-end using the Illumina NovaSeq 6000.

value: subcutaneous adipose

value: primary mature adipocyte seperated from fat pad

value: adipocyte

value: wild type

value: Mature adipocyte were seperated from iWAT of mice pre-treated with 7-days cold stress.


title: Dome H3K4me2 Triptolide rep 3

value: Whole embryos

value: TUAB

value: WT


title: s1_ASC29_rep1


title: s4_MC0192-1_rep1


title: Dome H3K4me2 Triptolide rep 2


title: unc iT activated, replicate 2, scRNA-seq


title: Dome H3K4me2 DMSO rep 3


title: unc iT activated, replicate 1, scRNA-seq


title: YY1 ChIP-seq room temperature

value: Mature adipocyte were seperated from iWAT of mice raised at room temperature.


title: CO79 DMSO R2 [A11]

first: Marianne

last: Collard

state: MA

address: 609 Albany Street, Department of Dermatology, 5th Floor, J508

department: Dermatology

institute: Boston University

data_processing:
Sequence reads were trimmed to remove possible adapter sequences and nucleotides with poor quality using Trimmomatic v.0.36.
The trimmed reads were mapped to the Homo sapiens GRCh38 reference genome available on ENSEMBL using the STAR aligner v.2.5.2b.
Unique gene hit counts were calculated by using featureCounts from the Subread package v.1.5.2. Only unique reads that fell within exon regions were counted.
Assembly: Homo sapiens GRCh38 reference genome
Supplementary files format and content: Text files include raw counts for each sample

source_name: tumor

growth_protocol: cell lines were cultured in Dulbecco’s Modified Eagle Medium (Gibco, ThermoFisher Scientific, Grand Island, NY, USA) supplemented with 10% bovine calf serum (Gibco, ThermoFisher Scientific, Grand Island, NY, USA), 1% penicillin/streptomycin (Gibco, ThermoFisher Scientific, Grand Island, NY, USA), and 2mM L-glutamine

extract_protocol:
RNA was then extracted using the RNeasy (Qiagen, Germantown, MD, USA) kit according to manufacturer instructions.
Library prepartion was done by Azenta Life Science LLC: RNA samples were quantified using Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA) and RNA integrity was checked using Agilent TapeStation 4200 (Agilent Technologies, Palo Alto, CA, USA). RNA sequencing libraries were prepared using the NEBNext Ultra II RNA Library Prep for Illumina using manufacturer’s instructions (NEB, Ipswich, MA, USA). Briefly, mRNAs were initially enriched with Oligod(T) beads. Enriched mRNAs were fragmented for 15 minutes at 94°C. First strand and second strand cDNA were subsequently synthesized. cDNA fragments were end repaired and adenylated at 3’ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by PCR with limited cycles. The sequencing libraries were validated on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and quantified by using Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA) as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA).

treatment_protocol: Cells were treated with 1:2000 DMSO or 5 uM A-485

value: tumor

value: melanoma

value: DMSO control


title: patient 12

city: Buffalo

first: Jianxin

last: Wang

address: 665 Elm Street

department: Molecular and Cellular Biology

institute: Roswell Park Comprehensive Cancer Center

description: archival colorectal cancer tissue

data_processing:
Raws fastq files are processed with nf-core/rnaseq: 3.13.2 pipeline under Nexflow 23.10.1.
Assembly: GRCh38
Supplementary files format and content: comma separated text file

source_name: colorectal

extract_protocol:
SureSelect XT HS2 RNA Target Enrichment System (Agilent Inc)
200ng of RNA was fragmented and converted to cDNA followed by end repair, duplex UMI adaptor ligation, and 11 cycles of PCR to complete the dual- index adapters. Unique dual-indexed, molecular-barcoded libraries were purified with AMPureXP beads (Beckman Coulter) and validated for appropriate size (225-275bp) on a Tapestation 4200 DNA1000 screentape (Agilent Inc.). 200ng of purified library was then hybridized to the SureSelectXT Human All Exon V8+UTR Capture library (Agilent Inc.). The hybridized regions were then bound to streptavidin magnetic beads and washed to remove any non-specific bound products. Eluted library underwent a second 13 cycle PCR amplification to generate enough material for sequencing. Final libraries were purified, measured by Tapestation 4200 DNA1000 screentape (250-350bp), and quantitated using KAPA qPCR (KAPA Biosystems). Individual libraries were pooled in equimolar 2nM final concentration. Each pool is denatured and diluted to 200pM with 1% PhiX control library added. The resulting pool is then loaded into the appropriate NovaSeq Reagent cartridge, as determined by the number of sequencing cycles desired, and sequenced on a NovaSeq6000 following the manufacturer’s recommended protocol (Illumina Inc.).

value: colorectal


title: Dome H3K4me2 DMSO rep 4

value: dome

value: Invitrogen #710796


title: CO79 5 uM A-485, 24 h R2 [A12]


title: con-week 2, replicate 1, scRNA-seq

value: iPSC-derived T cells

value: iPSCs

value: DMSO


title: WT, id 8, treated

city: Milano

first: Stefano

last: de Pretis

address: Via Olgettina, 58

department: Center for Omics Sciences

country: Italy

institute: IRCCS San Raffaele Institute

data_processing:
FASTQ sequencing reads were adaptor-trimmed and quality-filtered with Trimmomatic (Bolger et al., 2014)
Filtered reads were mapped to the mm10 mouse reference genome (https://www.gencodegenes.org/mouse/) with STAR (Dobin & Gingeras, 2016).
Gene counts were obtained using featureCounts (Liao et al., 2014).
Normalization and differential gene expression analysis (DEG) have been performed with DESeq2 (Love et al., 2014).
KO vs. WT DGE analysis was adjusted for cell culture and evaluated both in treated and untreated samples (“~cell_culture + genotype*treatment”). Treatment effect was evaluated independently for WT and KO samples and adjusted for the donor mouse (“mouse_id + treatment”).
Assembly: mm10
Supplementary files format and content: tab delimited text files including raw counts for each sample

source_name: Primary cortical neurons

growth_protocol: Primary cortical neurons were prepared from WT and Mecp2 KO male embryos at embryonic day 15.5 (E15.5). The pregnant dam was sacrificed, the embryos collected and sacrificed by decapitation. A tail biopsy was taken from each embryo to perform genotyping and sex determination. Brains were exposed, meninges removed and the cerebral cortex from both hemispheres was rapidly dissected and kept in ice-cold Hank's Buffered Salt Solution (HBSS; ThermoFisher, 14175095), a single embryo per tube. Tissues were washed in HBSS, incubated with 0.25% trypsin/EDTA (ThermoFisher, 25200056) for 10 min at 37°C, washed in HBSS and mechanically dissociated in Neurobasal Plus medium (ThermoFisher, A3582901) supplemented with 2% B-27TM Plus (ThermoFisher, A3582801), 2mM GlutaMAX-I (ThermoFisher, 35050038) and 1% Pen/Strep (Sigma-Aldrich, P0781). Neurons were plated in the same medium on plasticware coated with 0.01 mg/ml poly-D-lysine hydrobromide (Sigma-Aldrich, P7886), at a density of 300.000 cells/well in 6-well plates or 1.5 million cells/dish in 6-cm dishes. For immunofluorescence experiments, neurons were plated on 13-mm or 24-mm poly-D-lysine-coated (0.1 mg/ml) glass coverslips, at a density of 25.000 or 60.000 cells/coverslip.

extract_protocol:
After a rapid wash in PBS (Sigma D8537) to remove cellular debris, total RNA was extracted fromprimary cortical neurons using PureZOL (BioRad 7326890). In details, 0.3 mL of PureZOL (12wells plate) was added on cell culture and left 5 min at room temperature to allow the completedissociation of nucleoprotein complexes. 200L of chloroform was then added for 1 ml of PureZOLaccording to the manufacturer’s protocol. Following, samples were inverted for 15-20 seconds togently mix the phenol:chloroform mixture and incubated 3 min at RT. After centrifugation at12000g for 15 min at 4°C, the upper aqueous phase was transferred into RNAse free tube andprecipitated with 10 ug of RNA-grade glycogen and 100% isopropanol 1:2. Samples were storedo.n. at -20°C. The day after, the RNA was centrifuged at 12,000g for 10 min at 4°C and pellets werethen washed in 500ul of 70% ethanol (7,500 x g for 5 min at 4°C). To remove genomic DNA,sample pellets were treated with DNAse (Sigma-Aldrich, AMPD1) at 37°C for 15 min. A secondpurification with PureZOL was executed on RNAs and their pellets were finally resuspended in 10uL of RNAse-free H2O.
Total RNA was extracted from eight WT and eight KO cortical neurons (14 DIV) treated with trehalose and eight WT and eight KO untreated controls as described above for a total of 32 samples. Only high-quality RNA with an RNA Integrity Number (RIN) of 8 or higher was used. Library preparation and sequencing have been performed with TruSeq RNA Library Preparation Kit v2 (Illumina) by Genewiz. One WT untreated and corresponding treated sample and two KO untreated and related treated samples were found outliers and removed from the dataset.

treatment_protocol: Neurons were treated at 12 DIV with 25 mM trehalose (Sigma-Aldrich, T9531) in cell medium. After 48 hours of treatment, at 14 DIV, neurons were quickly washed in PBS and either lysed in hot SDS lysis buffer (50 mM Tris HCl pH 8, 10 mM EDTA, 1% SDS supplemented with 1:200 protease inhibitor cocktail [Sigma-Aldrich, P8340]) for western blotting, or scraped in -20°C-cold methanol for lipidomics, or fixed in the appropriate fixing solution for immunocytochemistry or electron microscopy experiments. Neurons were treated at 13 DIV neurons with 20 uM Leupeptin (Sigma- Aldrich, L2884) in cell medium. After 24 hours of treatment, at 14 DIV, neurons were lysed in hot SDS lysis buffer. Human fibroblasts at 70–80% confluency were treated with 25 mM trehalose for 48 hours, and cells were lysed in hot SDS lysis buffer. In vivo treatment was performed by intraperitoneal injection of 2g/kg of trehalose (Sigma-Aldrich, T9531) resuspended in sterile saline solution, every other day from postnatal day P18 to P40. Control mice were injected with the same volume of sterile saline solution. At the end of the treatment, mice were sacrificed. Cortices were collected, frozen in dry ice and stored at -80°C for subsequent lysis in SDS lysis buffer.

value: Primary cortical neurons

value: trehalose


title: patient 10

description: archival colorectal cancer tissue


title: Dome H3K4me2 DMSO rep 1


title: Dome H3K4me1 Triptolide rep 2

value: Triptolide

value: Invitrogen #710795


title: Dome H3K4me1 Triptolide rep 1

value: Whole embryos


title: patient 9

description: archival colorectal cancer tissue


title: patient 11

description: archival colorectal cancer tissue


title: WT, id 4, treated


title: IPC-298 5 uM A-485, 24 h R2 [A10]


title: IPC-298 DMSO R2 [A9]


title: Dome H3K4me1 DMSO rep 1

value: TUAB


title: WT, id 4

value: WT


title: A375 5 uM A-485, 24 h R2 [A8]


title: Dome H3K4me2 DMSO rep 2


title: CO79 5 uM A-485, 24 h R1 [A6]


title: WT, id 8


title: CO79 DMSO R1 [A5]


title: IPC-298 5 uM A-485, 24 h R1 [A4]


title: Dome H3K4me1 DMSO rep 2


title: patient 5

description: archival colorectal cancer tissue


title: patient 7

description: archival colorectal cancer tissue


title: Dome H2BK16ac (N-term acetyl) rep 2

value: Abcam #ab177427


title: patient 6

description: archival colorectal cancer tissue


title: WT, id 1, treated


title: WT, id 2, treated


title: A375 DMSO R2 [A7]

value: tumor


title: WT, id 1


title: Dome IgG rep 2

value: Invitrogen #10500C


title: patient 8

description: archival colorectal cancer tissue


title: IPC-298 DMSO R1 [A3]

value: melanoma

value: DMSO control


title: patient 4

description: archival colorectal cancer tissue

value: colorectal


title: patient 3

description: archival colorectal cancer tissue


title: WT, id 17, treated


title: patient 2

description: archival colorectal cancer tissue


title: Dome IgG rep 1

value: dome


title: WT, id 11_8


title: Dome H2BK16ac (N-term acetyl) rep 1