title: iCount_ctx_embryo_20190320BJ_N15


title: iCount_ctx_embryo_20190320BJ_N05

status: Public on Jul 19 2021

type: SRA

city: Zürich

first: Baptiste

middle: N

last: Jaeger

address: Winterthurerstrasse 190

department: Brain Research Institute

country: Switzerland

institute: University of Zürich

description: RawCount_Mouse_iCount_Embryonic_Cortex.txt

library_source: transcriptomic

data_processing:
Single-end 126nt-long reads were adaptor removed and trimmed using cutadapt v1.16 and sickle v1.33 with default parameters.
Reads were mapped against the mouse GRCm38.90 primary assembly or the human GRCh38.p13 primary assembly using STAR v2.6.0c in `alignReads` mode.
Gene count matrices were quantified at the exon level ignoring multimappers using featurecounts from subread v1.6.2.
The following paramaters were used for QC filtering.In the human dataset, only cells with 3000-7000 genes detected, less than 10% of mitochondrial reads and more than 50’000 reads were kept. In each dataset (i.e. 384 well-plate), cells with more or less than 1.5-fold the median read content per cell were excluded.  In the mouse dataset, only nuclei with 1000-3000 genes detected, less than 5% of mitochondrial reads and more than 50’000 reads were kept. In each dataset (i.e. 384 well-plate), nuclei with more or less than 1.5-fold the median read content per cell were excluded
Seurat v3.1.1 was used to normalize read counts, regress out the library size and mitochondrial reads proportion per cell, find variable features with the `vst` method, integrate multiple plates, dimensionality reduce the data, cluster cells and test for differentially expressed genes.
For pseudotime calculation, independent component analysis was computed with fastICA package v1.2.2.
Genome_build: mouse GRCm38.90 and human GRCh38.p13
Supplementary_files_format_and_content: one single metadata file will both mouse and human data. One rawcout file for mouse single nucleus data. One rawcount file for human single cell data.

molecule: total RNA

organism: Mus musculus

source_name: embryonic cortex

extract_protocol:
singe-cells and single-nulcei
illumina nextera

tag: mouse strain

value: H3.1-iCOUNT x ROSA26:CreERT2

tag: age of the mice/organoids at the time of the tamoxifen injection

tag: time of experiment

value: 38h post tamoxifen injection

tag: sorted material

value: individual iCount+ nuclei

tag: batch number

tag: icount color

value: Yellow


title: iCount_hOrganoids_plateII_20190308BJ_B23

organism: Homo sapiens

source_name: human forebrain organoids

value: 39 days in culture

value: 7 days post Cre electroporation

value: individual iCount+ cells

value: Green


title: iCount_ctx_embryo_20190320BJ_N04


title: iCount_hOrganoids_plateII_20190308BJ_B11

value: Red


title: iCount_ctx_embryo_20190320BJ_N03


title: iCount_hOrganoids_plateII_20190308BJ_B12


title: iCount_hOrganoids_plateII_20190308BJ_B10


title: iCount_ctx_embryo_20190320BJ_M22

value: Orange


title: iCount_ctx_embryo_20190320BJ_M19


title: iCount_hOrganoids_plateII_20190308BJ_B07


title: iCount_ctx_embryo_20190320BJ_M20


title: iCount_ctx_embryo_20190320BJ_N02


title: iCount_hOrganoids_plateII_20190308BJ_B05


title: iCount_hOrganoids_plateII_20190308BJ_B09


title: iCount_hOrganoids_plateII_20190308BJ_B06


title: iCount_hOrganoids_plateII_20190308BJ_B17


title: iCount_hOrganoids_plateII_20190308BJ_B03


title: iCount_hOrganoids_plateII_20190308BJ_B01


title: iCount_hOrganoids_plateII_20190308BJ_B16


title: iCount_ctx_embryo_20190320BJ_M18


title: iCount_hOrganoids_plateII_20190308BJ_A22


title: iCount_ctx_embryo_20190320BJ_M16


title: iCount_ctx_embryo_20190320BJ_M17


title: iCount_hOrganoids_plateII_20190308BJ_A20


title: iCount_ctx_embryo_20190320BJ_M15


title: iCount_hOrganoids_plateII_20190308BJ_A16


title: iCount_ctx_embryo_20190320BJ_M12


title: iCount_ctx_embryo_20190320BJ_M21


title: iCount_hOrganoids_plateII_20190308BJ_A19


title: iCount_ctx_embryo_20190320BJ_M10


title: iCount_ctx_embryo_20190320BJ_M09


title: iCount_hOrganoids_plateII_20190308BJ_A12


title: iCount_ctx_embryo_20190320BJ_M14


title: iCount_hOrganoids_plateII_20190308BJ_A07


title: iCount_hOrganoids_plateII_20190308BJ_A18


title: iCount_hOrganoids_plateII_20190308BJ_A06


title: iCount_ctx_embryo_20190320BJ_M13


title: iCount_hOrganoids_plateII_20190308BJ_A17


title: iCount_hOrganoids_plateII_20190308BJ_A15


title: iCount_ctx_embryo_20190320BJ_M05


title: iCount_hOrganoids_plateII_20190308BJ_A13


title: iCount_ctx_embryo_20190320BJ_M11


title: iCount_hOrganoids_plateII_20190308BJ_A04


title: iCount_ctx_embryo_20190320BJ_M04


title: iCount_hOrganoids_plateII_20190308BJ_A02


title: iCount_ctx_embryo_20190320BJ_L21


title: iCount_hOrganoids_d4_20190226BJ_P21

value: 4 days post Cre electroporation


title: iCount_ctx_embryo_20190320BJ_L20


title: iCount_ctx_embryo_20190320BJ_M06


title: iCount_hOrganoids_plateII_20190308BJ_A05


title: iCount_ctx_embryo_20190320BJ_M03


title: iCount_ctx_embryo_20190320BJ_L19


title: iCount_ctx_embryo_20190320BJ_L22


title: iCount_hOrganoids_d4_20190226BJ_P16


title: iCount_hOrganoids_plateII_20190308BJ_A09


title: iCount_ctx_embryo_20190320BJ_M08


title: iCount_ctx_embryo_20190320BJ_L18


title: iCount_hOrganoids_d4_20190226BJ_P15


title: iCount_ctx_embryo_20190320BJ_L15


title: iCount_hOrganoids_d4_20190226BJ_P10


title: iCount_hOrganoids_d4_20190226BJ_P08


title: iCount_hOrganoids_d4_20190226BJ_P14


title: iCount_ctx_embryo_20190320BJ_L14


title: iCount_hOrganoids_d4_20190226BJ_P07


title: iCount_hOrganoids_d4_20190226BJ_P22


title: iCount_ctx_embryo_20190320BJ_L11


title: iCount_hOrganoids_d4_20190226BJ_P04


title: iCount_hOrganoids_d4_20190226BJ_P06


title: iCount_ctx_embryo_20190320BJ_L12


title: iCount_hOrganoids_d4_20190226BJ_P09


title: iCount_ctx_embryo_20190320BJ_M07


title: iCount_hOrganoids_d4_20190226BJ_P02


title: iCount_ctx_embryo_20190320BJ_L08


title: iCount_ctx_embryo_20190320BJ_L07


title: iCount_hOrganoids_d4_20190226BJ_O24


title: iCount_hOrganoids_d4_20190226BJ_O22


title: iCount_ctx_embryo_20190320BJ_L06


title: iCount_hOrganoids_d4_20190226BJ_O21


title: iCount_ctx_embryo_20190320BJ_L13


title: iCount_ctx_embryo_20190320BJ_L05


title: iCount_ctx_embryo_20190320BJ_L04


title: iCount_ctx_embryo_20190320BJ_L09


title: iCount_hOrganoids_d4_20190226BJ_O20


title: iCount_ctx_embryo_20190320BJ_L03


title: iCount_hOrganoids_d4_20190226BJ_O16


title: iCount_hOrganoids_d4_20190226BJ_O15


title: iCount_ctx_embryo_20190320BJ_L17


title: iCount_hOrganoids_d4_20190226BJ_O18


title: iCount_ctx_embryo_20190320BJ_L02


title: iCount_ctx_embryo_20190320BJ_K23


title: iCount_ctx_embryo_20190320BJ_K22


title: iCount_hOrganoids_d4_20190226BJ_O12


title: iCount_ctx_embryo_20190320BJ_K21


title: iCount_hOrganoids_d4_20190226BJ_O09


title: iCount_ctx_embryo_20190320BJ_K20


title: iCount_hOrganoids_d4_20190226BJ_O08


title: iCount_ctx_embryo_20190320BJ_K19


title: iCount_hOrganoids_d4_20190226BJ_O07


title: iCount_ctx_embryo_20190320BJ_K18


title: iCount_ctx_embryo_20190320BJ_K17


title: iCount_hOrganoids_d4_20190226BJ_O06


title: iCount_hOrganoids_d4_20190226BJ_O05


title: iCount_ctx_embryo_20190320BJ_K15


title: iCount_ctx_embryo_20190320BJ_K14


title: iCount_ctx_embryo_20190320BJ_K13


title: iCount_hOrganoids_d4_20190226BJ_N24


title: iCount_hOrganoids_d4_20190226BJ_O02


title: iCount_hOrganoids_d4_20190226BJ_N21


title: iCount_ctx_embryo_20190320BJ_K11


title: iCount_hOrganoids_d4_20190226BJ_N20


title: iCount_ctx_embryo_20190320BJ_K09


title: iCount_hOrganoids_d4_20190226BJ_N18


title: iCount_ctx_embryo_20190320BJ_K12


title: iCount_hOrganoids_d4_20190226BJ_O11


title: iCount_ctx_embryo_20190320BJ_K08


title: iCount_hOrganoids_d4_20190226BJ_N16


title: iCount_hOrganoids_d4_20190226BJ_N12


title: iCount_hOrganoids_d4_20190226BJ_N13


title: iCount_ctx_embryo_20190320BJ_K04


title: iCount_ctx_embryo_20190320BJ_K05


title: iCount_ctx_embryo_20190320BJ_K03


title: iCount_hOrganoids_d4_20190226BJ_N09


title: iCount_ctx_embryo_20190320BJ_K02


title: iCount_hOrganoids_d4_20190226BJ_O04


title: iCount_ctx_embryo_20190320BJ_K16


title: iCount_ctx_embryo_20190320BJ_J23


title: iCount_hOrganoids_d4_20190226BJ_O03


title: iCount_hOrganoids_d4_20190226BJ_N08


title: iCount_ctx_embryo_20190320BJ_J22


title: iCount_hOrganoids_d4_20190226BJ_N06


title: iCount_ctx_embryo_20190320BJ_J21


title: iCount_hOrganoids_d4_20190226BJ_N05


title: iCount_ctx_embryo_20190320BJ_J20


title: iCount_hOrganoids_d4_20190226BJ_N04


title: iCount_ctx_embryo_20190320BJ_K06


title: iCount_hOrganoids_d4_20190226BJ_N01


title: iCount_ctx_embryo_20190320BJ_J18


title: iCount_ctx_embryo_20190320BJ_J17


title: iCount_hOrganoids_d4_20190226BJ_M19


title: iCount_ctx_embryo_20190320BJ_J15


title: iCount_hOrganoids_d4_20190226BJ_N11


title: iCount_hOrganoids_d4_20190226BJ_M18


title: iCount_hOrganoids_d4_20190226BJ_N10


title: iCount_hOrganoids_d4_20190226BJ_M16


title: iCount_ctx_embryo_20190320BJ_J14


title: iCount_hOrganoids_d4_20190226BJ_M15


title: iCount_hOrganoids_d4_20190226BJ_M14


title: iCount_hOrganoids_d4_20190226BJ_M13


title: iCount_hOrganoids_d4_20190226BJ_M12


title: iCount_hOrganoids_d4_20190226BJ_M11


title: iCount_ctx_embryo_20190320BJ_J12


title: iCount_ctx_embryo_20190320BJ_J10


title: iCount_ctx_embryo_20190320BJ_J11


title: iCount_ctx_embryo_20190320BJ_J09


title: iCount_hOrganoids_d4_20190226BJ_M09


title: iCount_ctx_embryo_20190320BJ_J06


title: iCount_hOrganoids_d4_20190226BJ_M22


title: iCount_ctx_embryo_20190320BJ_J05


title: iCount_ctx_embryo_20190320BJ_J19


title: iCount_ctx_embryo_20190320BJ_J07


title: iCount_hOrganoids_d4_20190226BJ_M08


title: iCount_ctx_embryo_20190320BJ_J03


title: iCount_ctx_embryo_20190320BJ_J02


title: iCount_hOrganoids_d4_20190226BJ_M01


title: iCount_hOrganoids_d4_20190226BJ_M17


title: iCount_ctx_embryo_20190320BJ_J01


title: iCount_hOrganoids_d4_20190226BJ_M05


title: iCount_hOrganoids_d4_20190226BJ_M02


title: iCount_hOrganoids_d4_20190226BJ_L22


title: iCount_hOrganoids_d4_20190226BJ_L24


title: iCount_ctx_embryo_20190320BJ_I22


title: iCount_ctx_embryo_20190320BJ_I23


title: iCount_hOrganoids_d4_20190226BJ_L21


title: iCount_ctx_embryo_20190320BJ_J13


title: iCount_ctx_embryo_20190320BJ_I20


title: iCount_ctx_embryo_20190320BJ_J08


title: iCount_ctx_embryo_20190320BJ_I19


title: iCount_hOrganoids_d4_20190226BJ_L19


title: iCount_ctx_embryo_20190320BJ_I18


title: iCount_hOrganoids_d4_20190226BJ_L14


title: iCount_hOrganoids_d4_20190226BJ_L16


title: iCount_ctx_embryo_20190320BJ_I17


title: iCount_ctx_embryo_20190320BJ_I15


title: iCount_hOrganoids_d4_20190226BJ_L13


title: iCount_ctx_embryo_20190320BJ_I14


title: iCount_hOrganoids_d4_20190226BJ_L09


title: iCount_hOrganoids_d4_20190226BJ_L11


title: iCount_ctx_embryo_20190320BJ_I12


title: iCount_hOrganoids_d4_20190226BJ_L05


title: iCount_hOrganoids_d4_20190226BJ_J24


title: iCount_ctx_embryo_20190320BJ_I13


title: iCount_ctx_embryo_20190320BJ_I11


title: iCount_ctx_embryo_20190320BJ_I09


title: iCount_hOrganoids_d4_20190226BJ_J23


title: iCount_ctx_embryo_20190320BJ_I10


title: iCount_hOrganoids_d4_20190226BJ_J22


title: iCount_hOrganoids_d4_20190226BJ_J20


title: iCount_ctx_embryo_20190320BJ_I07


title: iCount_ctx_embryo_20190320BJ_I08


title: iCount_hOrganoids_d4_20190226BJ_J18


title: iCount_hOrganoids_d4_20190226BJ_J21


title: iCount_ctx_embryo_20190320BJ_I05


title: iCount_ctx_embryo_20190320BJ_I04


title: iCount_ctx_embryo_20190320BJ_I06


title: iCount_hOrganoids_d4_20190226BJ_J15


title: iCount_hOrganoids_d4_20190226BJ_J16


title: iCount_ctx_embryo_20190320BJ_I03


title: iCount_ctx_embryo_20190320BJ_I02


title: iCount_hOrganoids_d4_20190226BJ_J09


title: iCount_hOrganoids_d4_20190226BJ_J07


title: iCount_ctx_embryo_20190320BJ_H22


title: iCount_ctx_embryo_20190320BJ_H21


title: iCount_hOrganoids_d4_20190226BJ_J05


title: iCount_hOrganoids_d4_20190226BJ_J03


title: iCount_ctx_embryo_20190320BJ_H20


title: iCount_hOrganoids_d4_20190226BJ_J01


title: iCount_hOrganoids_d4_20190226BJ_I23


title: iCount_hOrganoids_d4_20190226BJ_J17


title: 3K_PBS_D19_2

city: Cambridge

first: Justin

last: Nicholatos

state: Massachusetts

address: 225 Binney St

department: Neurodegeneration

country: USA

institute: Biogen

data_processing:
RNA-Seq read data was analyzed using a pipeline that used STAR (version 2.5.2b) for read alignment and RSEM (version 1.2.31) transcript read count quantification
All samples passed quality checks on raw reads with FASTQC and read alignment quality (>90% uniquely mapped paired reads).
Counts were normalized by DESeq2 across all samples
Genome_build: Rnor_6.0 (GCA_000001895.4)
Supplementary_files_format_and_content: Matrix table with normalized counts for every gene and every sample

organism: Rattus norvegicus

source_name: Primary cortical neuron cultures

extract_protocol:
RNA was extracted with a Rneasy plus kit with Dnase I digestion from Qiagen.
Full length cDNA was prepared with the SMART-Seq v4 Ultra Low Input RNA Kit (Takara, 634891) according to manufacturer’s instructions from 8 ng of input RNA.
Libraries were prepared from amplified cDNA using the Nextera XT DNA Library Prep Kit (Illumina, FC-131-1096) according to manufacturer’s instructions. 150 pg of cDNA was used as input, 12 cycles of PCR were used to amplify tagmented cDNA with Nextera Index Kit Set A (Illumina, FC-131-2001), and library cleanup was performed as recommended for 300-500bp libraries. Libraries were quantified and QCed on the Lab Chip GX DNA HS Chip (Perkin Elmer, CLS760672). Libraries were normalized to 10 nM, pooled, and sequenced on a HiSeq 2500 in paired end mode 2x50 bp to an average depth of 8.8m reads per sample.

treatment_protocol: Rat cortical neuron cultures were treated with AAV9-EV or AAV9-3K aSyn on DIV 5. RNA was isolated 6-, 12-, and 19-days post AAV treatment

tag: aav treatment

value: AAV-aSyn 3K

tag: days post aav for rna isolation

value: Day 19


title: iCount_ctx_embryo_20190320BJ_H18


title: iCount_ctx_embryo_20190320BJ_H19


title: iCount_hOrganoids_d4_20190226BJ_I22


title: iCount_ctx_embryo_20190320BJ_H15


title: 3K_PBS_D12_3

value: Day 12


title: 3K_PBS_D19_5


title: 3K_PBS_D19_4


title: 3K_PBS_D12_4


title: 3K_PBS_D19_3


title: 3K_PBS_D19_1


title: iCount_hOrganoids_d4_20190226BJ_I20


title: 3K_PBS_D12_2


title: iCount_hOrganoids_d4_20190226BJ_J14


title: 3K_PBS_D12_5


title: 3K_PBS_D12_1


title: iCount_ctx_embryo_20190320BJ_H14


title: 3K_PBS_D6_5

value: Day 6


title: iCount_hOrganoids_d4_20190226BJ_I18


title: iCount_ctx_embryo_20190320BJ_H12


title: 3K_PBS_D6_2


title: iCount_ctx_embryo_20190320BJ_H13


title: 3K_PBS_D6_3


title: 3K_PBS_D6_1


title: iCount_hOrganoids_d4_20190226BJ_I16


title: EV_PBS_D19_5

value: AAV-empty vector


title: iCount_hOrganoids_d4_20190226BJ_I15


title: EV_PBS_D19_4


title: EV_PBS_D19_3


title: EV_PBS_D19_2


title: iCount_hOrganoids_d4_20190226BJ_I14


title: EV_PBS_D19_1


title: EV_PBS_D12_5


title: iCount_hOrganoids_d4_20190226BJ_I12


title: iCount_ctx_embryo_20190320BJ_H10


title: iCount_ctx_embryo_20190320BJ_H08


title: iCount_hOrganoids_d4_20190226BJ_I11


title: 3K_PBS_D6_4


title: EV_PBS_D12_3


title: iCount_ctx_embryo_20190320BJ_H07


title: iCount_hOrganoids_d4_20190226BJ_I09


title: EV_PBS_D12_4


title: iCount_hOrganoids_d4_20190226BJ_I17


title: EV_PBS_D12_2


title: iCount_ctx_embryo_20190320BJ_H05


title: EV_PBS_D6_5


title: EV_PBS_D6_4


title: iCount_hOrganoids_d4_20190226BJ_I07


title: iCount_ctx_embryo_20190320BJ_H11


title: iCount_ctx_embryo_20190320BJ_H04


title: EV_PBS_D6_3


title: iCount_hOrganoids_d4_20190226BJ_I06


title: iCount_ctx_embryo_20190320BJ_H03


title: EV_PBS_D6_2


title: iCount_ctx_embryo_20190320BJ_H09


title: EV_PBS_D12_1


title: EV_PBS_D6_1


title: iCount_ctx_embryo_20190320BJ_G23


title: iCount_hOrganoids_d4_20190226BJ_I01


title: iCount_hOrganoids_d4_20190226BJ_H24


title: iCount_ctx_embryo_20190320BJ_G22


title: iCount_ctx_embryo_20190320BJ_G21


title: glp-1_6

city: Montreal

first: Jeremy

middle: Michael

last: Van Raamsdonk

state: QC

address: 1001 Decarie Boulevard

department: Neurology and Neurosurgery

country: Canada

institute: McGill University

data_processing:
Adapter sequences, other contaminant sequences such as polyA tails and low quality sequences were trimmed from reads using cutadapt
Trimmed reads were aligned to genome using STAR
Quality of alignments was assessed by checking for evenness of coverage, rRNA content, genomic context of alignments (for example, alignments in known transcripts and introns), complexity and other quality check using Qualimap
Expression quantification was performed with Salmon
Transcript-level abundance estimates  collapsed down to the gene-level using the R Bioconductor package tximport
Genome_build: Ensembl build WBcel235 (release 90)
Supplementary_files_format_and_content: Csv file contains the gene-level  raw counts for each sample

molecule: polyA RNA

organism: Caenorhabditis elegans

source_name: Whole worm

growth_protocol: NGM plates, 20 degrees Celsius, OP50 bacteria

extract_protocol:
Day 1 adult worms from a limited lay were collected in M9 buffer and washed 3 times to remove bacteria. After supernatant M9 was re-moved worms were resuspended in TRIZOL reagent. Worms were vortexed, frozen in liquid nitrogen and thawed a total of three times and then left to sit for 15 min. After adding chloroform, shaking vigor-ously and sitting at room temperature for 3 min, tubes were centrifugedat 12,000 g for 15 min at 4 °C. The RNA-containing upper phase was transferred to a new tube and precipitated with an equal volume of isopropanol. After sitting 10 min at room temperature, RNA was pelleted by centrifugingat 12,000 g for 10 min.Finally, RNA was washed with 75% ethanol, re-pelleted and then dissolved in RNAse free water.
The 36 samples were run over 3 flowcells, all samples were contained on all flowcells.

tag: developmental stage

value: prefertile young adult

tag: temperature

tag: genotype

tag: tissue

value: whole worm


title: iCount_hOrganoids_d4_20190226BJ_H23


title: glp-1_4


title: WT25_6

value: WT


title: glp-1_1


title: glp-1_2


title: glp-1_5


title: glp-1_3


title: WT25_5


title: iCount_ctx_embryo_20190320BJ_H24

value: H3.1-iCOUNT x ROSA26:CreERT2

value: 38h post tamoxifen injection

value: individual iCount+ nuclei


title: iCount_hOrganoids_d4_20190226BJ_H19

value: 39 days in culture


title: iCount_ctx_embryo_20190320BJ_G20

value: Yellow


title: WT25_4


title: iCount_ctx_embryo_20190320BJ_G19


title: iCount_hOrganoids_d4_20190226BJ_H16

value: individual iCount+ cells


title: WT25_2


title: WT25_1


title: osm-5_6


title: iCount_hOrganoids_d4_20190226BJ_I02


title: iCount_ctx_embryo_20190320BJ_G17


title: osm-5_5


title: iCount_hOrganoids_d4_20190226BJ_H14


title: iCount_hOrganoids_d4_20190226BJ_H15


title: osm-5_4


title: iCount_ctx_embryo_20190320BJ_G14


title: iCount_hOrganoids_d4_20190226BJ_H13


title: iCount_ctx_embryo_20190320BJ_G13


title: osm-5_3


title: osm-5_2


title: osm-5_1


title: iCount_hOrganoids_d4_20190226BJ_H10


title: iCount_ctx_embryo_20190320BJ_G12


title: ife-2_4


title: ife-2_3


title: ife-2_6


title: WT25_3


title: ife-2_2


title: ife-2_5


title: iCount_ctx_embryo_20190320BJ_G18


title: ife-2_1


title: iCount_ctx_embryo_20190320BJ_G09


title: iCount_hOrganoids_d4_20190226BJ_H01


title: eat-2_5


title: eat-2_6


title: iCount_hOrganoids_d4_20190226BJ_H02


title: eat-2_4


title: eat-2_3


title: iCount_hOrganoids_d4_20190226BJ_G24


title: iCount_ctx_embryo_20190320BJ_G07


title: eat-2_2


title: iCount_ctx_embryo_20190320BJ_G11


title: iCount_hOrganoids_d4_20190226BJ_H05


title: eat-2_1


title: iCount_hOrganoids_d4_20190226BJ_H04


title: WT_6


title: iCount_ctx_embryo_20190320BJ_G06


title: iCount_hOrganoids_d4_20190226BJ_G22


title: WT_4


title: WT_5


title: iCount_ctx_embryo_20190320BJ_G05


title: iCount_hOrganoids_d4_20190226BJ_G19


title: WT_2


title: iCount_ctx_embryo_20190320BJ_G04


title: iCount_hOrganoids_d4_20190226BJ_G18


title: iCount_ctx_embryo_20190320BJ_G10


title: WT_3


title: WT_1


title: iCount_ctx_embryo_20190320BJ_G08


title: AW

city: Brescia

first: Chiara

last: Magri

state: Italy

address: Viale Europa

department: DMMT

country: Italy

institute: Università degli Studi di Brescia

data_processing:
base-calling, demultiplexing and barcode trimming were performed automatically by the Torrent Suite Software (version 5.12.0) after each sequencing run and data from multiple runs were merged in order to create a unique FASTQ file for each sample.
cutadapt v1.14  (with paramiters cutadapt -a "A{25}" -u 5 -q 10 -m 30 ) was used to cut poly-A tails and five bases from the 5’ prime end and to discard all reads with length below 30bp and quality trimming below Q10.
Sequenced reads were mapped to mm10 whole genome using STAR aligner v. 2.6 with the following paramiter:  --outFilterMultimapNmax 10 --alignIntronMin 20 --alignIntronMax 1000000 --outSAMtype BAM SortedByCoordinate --quantMode GeneCounts
Genome_build: mm10
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample (only genes with >=10 reads in at least 3 samples are reported)

source_name: whole blood

growth_protocol: The animals were housed under a 12-h light-dark cycle at a temperature of 22 - 23 °C and a humidity of 60-75%

extract_protocol:
Total RNA was isolated from peripheral blood with the Quick DNA/RNA miniprep plus kit (ZymoResearch) according to the manufacturer’s instruction. The RNA purity was evaluated with NanoDrop Spectrophotometer 2000 (Thermo Fisher Scientific) and RNA integrity number (RIN) was determined with Agilent 2100 Bioanalyzer (Agilent Technologies).  Alpha and beta globin were depleted from total RNA with the GLOBINclear Kit (Invitrogen by Thermo Fisher Scientific) and sequin synthetic RNA spike-in controls were added to mRNA as sequencing internal contro
Libraries were produced with QuantSeq 3’ mRNA-Seq Library Prep Kit for Ion Torrent (Lexogen), according to the manufacturer’s instruction.

treatment_protocol: For tDCS stimulation, we adopted a unilateral epicranial electrode arrangement

tag: strain

tag: age

value: 3 months old

value: hippocampus

tag: stimulation

value: sham

value: whole blood


title: AX


title: iCount_hOrganoids_d4_20190226BJ_G17

value: 4 days post Cre electroporation

value: Orange


title: iCount_hOrganoids_d4_20190226BJ_G16


title: AS


title: iCount_ctx_embryo_20190320BJ_G03


title: AV


title: iCount_hOrganoids_d4_20190226BJ_G14


title: I


title: iCount_ctx_embryo_20190320BJ_G02


title: A2


title: iCount_hOrganoids_d4_20190226BJ_G08


title: BO

value: 6 months old

value: prefrontal cortex

value: tdcs


title: FF


title: FD


title: FE


title: iCount_ctx_embryo_20190320BJ_F22


title: iCount_ctx_embryo_20190320BJ_F21


title: FC


title: iCount_hOrganoids_d4_20190226BJ_G06


title: FB


title: iCount_hOrganoids_d4_20190226BJ_G05


title: FA


title: BN


title: iCount_hOrganoids_d4_20190226BJ_G04


title: iCount_ctx_embryo_20190320BJ_F18


title: DU


title: iCount_ctx_embryo_20190320BJ_F17


title: DR


title: DS


title: iCount_ctx_embryo_20190320BJ_G01


title: iCount_hOrganoids_d4_20190226BJ_G03


title: iCount_ctx_embryo_20190320BJ_F16


title: DT


title: iCount_hOrganoids_d4_20190226BJ_G02


title: tdcs2


title: tdcs7


title: iCount_hOrganoids_d4_20190226BJ_G01


title: BA


title: iCount_hOrganoids_d4_20190226BJ_G07


title: AZ


title: iCount_ctx_embryo_20190320BJ_F19


title: AY


title: iCount_hOrganoids_d4_20190226BJ_F24


title: iCount_ctx_embryo_20190320BJ_F13


title: iCount_ctx_embryo_20190320BJ_F14


title: AN


title: AQ


title: iCount_ctx_embryo_20190320BJ_F12


title: iCount_hOrganoids_d4_20190226BJ_F22


title: iCount_ctx_embryo_20190320BJ_F11


title: AM


title: AA


title: iCount_hOrganoids_d4_20190226BJ_F21


title: AL


title: iCount_hOrganoids_d4_20190226BJ_F20


title: Z


title: iCount_ctx_embryo_20190320BJ_F15


title: iCount_ctx_embryo_20190320BJ_F10


title: Y


title: W


title: X


title: U


title: iCount_ctx_embryo_20190320BJ_F09


title: V


title: iCount_ctx_embryo_20190320BJ_F07


title: iCount_ctx_embryo_20190320BJ_F08


title: iCount_hOrganoids_d4_20190226BJ_F19


title: iCount_hOrganoids_d4_20190226BJ_F17


title: T


title: iCount_hOrganoids_d4_20190226BJ_F18


title: S


title: iCount_hOrganoids_d4_20190226BJ_F16


title: P


title: iCount_ctx_embryo_20190320BJ_F05


title: Q


title: iCount_ctx_embryo_20190320BJ_F03


title: O


title: N


title: iCount_hOrganoids_d4_20190226BJ_F13


title: M


title: R


title: iCount_hOrganoids_d4_20190226BJ_F14


title: iCount_ctx_embryo_20190320BJ_E23


title: iCount_ctx_embryo_20190320BJ_E21


title: iCount_hOrganoids_d4_20190226BJ_E24


title: iCount_ctx_embryo_20190320BJ_F02


title: iCount_ctx_embryo_20190320BJ_F06


title: SUZ12dTAG_NucRNA_UNT_rep2

city: Oxford

first: Paula

last: Dobrinić

address: South Parks Road

department: Department of Biochemistry

country: United Kingdom

institute: University of Oxford

data_processing:
Paired-end reads were first aligned using Bowtie 2 (“–very-fast,” “–no-mixed” and “–no-discordant” options) (Langmead and Salzberg, 2012) against the concatenated mm10 and dm6 rDNA genomic sequence (GenBank: BK000964.3 and M21017.1) in order to discard the reads mapping to rDNA. The remaining unmapped reads were then aligned against the concatenated mm10 and dm6 genome using STAR (Dobin et al., 2013). Reads which failed to map using STAR were further aligned against the concatenated mm10 and dm6 genome using Bowtie 2 (“–sensitive-local,” “–no-mixed” and “–no-discordant” options), to improve the overall mapping of intronic sequences of nascent transcripts present in nuclear RNA fraction. Uniquely mapped reads from the two alignment steps were combined and PCR duplicates were removed using Sambamba (Tarasov et al., 2015).
To internally calibrate RNA-seq, we spiked-in a fixed number of Drosophila SG4 to each experimental sample. For data visualisation, mm10 reads were randomly subsampled using factors that reflected the total number of dm6 reads in each sample.
For differential gene expression analysis, we obtained read counts from the original bam files prior to spike-in normalization for a non-redundant mouse gene set using a SAMtools-based custom Perl script. The non-redundant mm10 gene set (n = 20633) was derived from mm10 refGene genes by filtering out very short genes with poor sequence mappability and highly similar transcripts.
To identify significant gene expression changes following auxin or dTAG-13 treatment, we used a custom R script that incorporates spike-in calibration into DESeq2 analysis (Love et al., 2014) and uses “apeglm” method for LFC shrinkage (Zhu et al., 2019). Spike-in calibration was incorporated by using read counts from a set of unique dm6 refGene genes to calculate DESeq2 size factors for normalization of raw mm10 read counts, as previously described in (Fursova et al., 2019; Taruttis et al., 2017). For a change to be considered significant, we applied a threshold of p-adj < 0.05 and fold change > 1.5.
Genome_build: mm10
Supplementary_files_format_and_content: Stranded bigWig files generated with genomeCoverageBed from BEDTools representing genome coverage for merged biological replicates which have been pre-normalised using spike-in downsampling factors
Supplementary_files_format_and_content: DESeq2 results tables with log2 fold changes shrunken by “apeglm” method and statistical parameters of differential expression gene analysis for comparisons between treated and untreated cells.

organism:
Drosophila melanogaster
Mus musculus

source_name: Mouse embryonic stem cells with Drosophila SG4 spike-in, untreated control

growth_protocol: Mouse embryonic stem cells were grown on gelatin-coated plates, in Dulbecco’s Modified Eagle Medium (DMEM, Life Technologies) supplemented with 15% fetal bovine serum (FBS, Labtech), 0.5 mM beta-mercaptoethanol (Life Technologies), 2 mM L-glutamine (Life Technologies), 1x penicillin-streptomycin (Life Technologies), 1x non-essential amino acids (Life Technologies) and 10 ng/mL leukemia inhibitory factor. Cells were cultured at 37°C with 5% CO2. For calibration of genomic experiments, we used either Drosophila S2 (SG4) cells, cultured at 25°C in Schneider’s Drosophila Medium (Life Technologies), supplemented with 10% heat-inactivated FBS and 1x penicillin-streptomycin.

extract_protocol:
2x107 mouse ESCs (untreated or treated with auxin or dTAG-13 for indicated times) were mixed with 8x106 Drosophila SG4 cells in PBS. Nuclei were released in 1 mL HS Lysis buffer (50 mM KCl, 10 mM MgSO4.7H20, 5 mM HEPES, 0.05% NP40, 1 mM PMSF, 3 mM DTT, 1x PIC) for 1 min at room temperature, and recovered by centrifugation at 1000 g for 5 min at 4°C, followed by three washes with ice-cold RSB buffer (10 mM NaCl, 10 mM Tris pH 7.4, 3 mM MgCl2). Pelleted nuclei were resuspended in 1 mL of TRIzol reagent and RNA was extracted according to the manufacturer’s protocol, followed by treatment with the TURBO DNA-free Kit. Quality of RNA was assessed using 2100 Bioanalyzer RNA 6000 Pico kit (Agilent), followed by rRNA depletion using the NEBNext rRNA Depletion kit (NEB).
RNA-seq libraries were prepared using the NEBNext Ultra (for PRC1deg) or Ultra II (for PRC2deg) Directional RNA Library Prep kit (NEB), indexed using NEBNext Multiplex Oligos (NEB), polled and sequenced as 80 bp (PRC1deg) or 40 bp (PRC2deg) paired-end reads on the Illumina NextSeq 500 in biological triplicates.

treatment_protocol: To induce AID-RING1B degradation, water-dissolved auxin (indole-3-acetic acid (IAA), Sigma) was mixed with cell medium to the final concentration of 500 µM and added to PRC1deg cells at designated times before harvesting by trypsinisation. To induce dTAG-SUZ12 degradation, PRC2deg cells were treated with 100 nM dTAG-13 compound for 2 hours.

tag: cell type

value: Mouse embryonic stem cells

value: E14 Pcgf2-HaloTag; dTAG-Suz12

tag: replicate

tag: treatment

tag: treatment duration

value: 0 hours

tag: spike-in reference organism

value: Drosophila melanogaster

tag: spike-in cell line

tag: molecular subtype

value: nuclear RNA


title: SUZ12dTAG_NucRNA_dTAG_2h_rep3

source_name: Mouse embryonic stem cells with Drosophila SG4 spike-in, dTAG-13 treated (2h dTAG)

value: 2 hours


title: SUZ12dTAG_NucRNA_UNT_rep1


title: SUZ12dTAG_NucRNA_UNT_rep3


title: SUZ12dTAG_NucRNA_dTAG_2h_rep1


title: iCount_hOrganoids_d4_20190226BJ_F15

tag: age of the mice/organoids at the time of the tamoxifen injection

value: 39 days in culture

tag: time of experiment

tag: sorted material

tag: batch number

tag: icount color


title: SUZ12dTAG_NucRNA_dTAG_2h_rep2

tag: genotype


title: iCount_hOrganoids_d4_20190226BJ_E23

value: individual iCount+ cells


title: SUZ12dTAG_NativeInput_dTAG_2h_rep3

library_source: genomic

data_processing:
Paired-end reads were aligned to the concatenated mouse (mm10) and spike-in (dm6 for native, hg19 for cross-linked cChIP-seq) genome sequences using Bowtie 2 (“–no-mixed” and “–no-discordant” options). Only uniquely mapped reads were kept for downstream analysis, after removal of PCR duplicates with Sambamba.
To internally calibrate cChIP-seq experiments, we spiked-in a fixed number of control cells (Drosophila SG4 cells for native cChIP-seq and human HEK293T cells for cross-linked cChIP-seq) to each experimental sample
For visualization of cChIP-seq sequencing data and annotation of genomic regions with read counts, uniquely mapped mouse reads were normalized using dm6 or hg19 spike-in as described previously (Furosva et al., 2019). Briefly, mouse reads were randomly subsampled based on the total number of spike-in (dm6 or hg19) reads in each sample. Additionally, to account for possible minor variations in spike-in cell mixing between cChIP-seq replicates, we corrected the subsampling factors by using the ratio of spike-in to mouse total read counts in the corresponding input samples. Prior to merging normalized replicates for visualization and analysis, read coverage across regions of interest (RING1B peaks for RING1B, SUZ12, H2AK119ub1 and H3K27me3, or TSS ± 2.5 kb for total Pol II, Ser5P-Pol II and H3K4me3, or gene bodies for Ser2P) was analyzed using multiBamSummary and plotCorrelation functions from deepTools , confirming a high correlation between replicates (Pearson's correlation coefficient > 0.9). Genome coverage tracks for cChIP-seq were generated using the pileup function from MACS2.
To identify genomic regions bound by PRC1 we performed peak calling using MACS2 (“BAMPE” and “–broad” options) on the RING1B cChIP-seq data with corresponding input samples for background normalization. A set of peaks identified in all biological replicates of untreated PRC1deg cells was used for further analysis, after filtering out several peaks identified following 4 hours of auxin treatment (i.e. following loss of RING1B) and manually removing sequencing artefacts. In total, 7240 stringent RING1B peaks were identified.
Genome_build: mm10
Supplementary_files_format_and_content: bigWig files representing genome coverage for merged replicates of spike-in normalised cChIP-seq
Supplementary_files_format_and_content: bed file for a set of enrichment peaks for RING1B

molecule: genomic DNA

growth_protocol: Mouse ESCs were grown on gelatin-coated plates, in Dulbecco’s Modified Eagle Medium (DMEM, Life Technologies) supplemented with 15% fetal bovine serum (FBS, Labtech), 0.5 mM beta-mercaptoethanol (Life Technologies), 2 mM L-glutamine (Life Technologies), 1x penicillin-streptomycin (Life Technologies), 1x non-essential amino acids (Life Technologies) and 10 ng/mL leukemia inhibitory factor. Cells were cultured at 37°C with 5% CO2. For calibration of genomic experiments, we used either Drosophila S2 (SG4) cells, cultured at 25°C in Schneider’s Drosophila Medium (Life Technologies), supplemented with 10% heat-inactivated FBS and 1x penicillin-streptomycin, or human HEK293T cells, cultured at 37°C with 5% CO2, in DMEM supplemented with 10% FBS, 2 mM L-glutamine, 1x penicillin-streptomycin and 0.5 mM beta-mercaptoethanol.

extract_protocol:
For RING1B and SUZ12 cChIP-seq, 5x107 mouse ESCs were first crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Scientific) while rotating for 45 min at 25°C, and then with 1% formaldehyde (methanol-free, Thermo Scientific) for further 15 min. Reactions were quenched by addition of 125 mM glycine. Mouse ESCs were then mixed with 2x106 double-crosslinked HEK293T cells and incubated in lysis buffer (50 mM HEPES pH 7.9, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100, 1x PIC) for 10 min at 4°C. The released nuclei were washed (10 mM Tris-HCl pH 8, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1x PIC) for 5 min at 4°C. Chromatin was then resuspended in 1 mL of sonication buffer (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% sodium deoxycholate, 0.5% N-lauroylsarcosine, 1x PIC) and sonicated for 30 min using the BioRuptor Pico (Diagenode). Following sonication, Triton X-100 was added to a final concentration of 1%. The supernatant was collected after centrifugation at 20,000 g for 10 min at 4°C. For ChIP, chromatin was diluted 10-fold in ChIP dilution buffer (1% Triton X-100, 1 mM EDTA, 20 mM Tris-HCl pH 8, 150 mM NaCl, 1x PIC) and pre-cleared for 1 h with either protein A agarose beads (Repligen, for RING1B ChIP) or protein A magnetic Dynabeads (Invitrogen, for SUZ12 ChIP) blocked with 1 mg/mL bovine serum albumin (BSA) and 1 mg/mL yeast tRNA. For each ChIP reaction, 1 mL of diluted and pre-cleared chromatin was incubated overnight with the appropriate antibody, either anti-RING1B (CST, D22F2, 3 μl), or anti-SUZ12 (CST, D39F6, 3 μl). Antibody-bound chromatin was captured using blocked protein A beads (agarose for RING1B, magnetic Dynabeads for SUZ12) for at least 2 h at 4°C and collected by centrifugation/on a magnetic rack. ChIP washes were performed as described previously (Farcas et al. 2013). ChIP DNA was eluted in elution buffer (1% SDS, 0.1 M NaHCO3) and cross-linking was reversed overnight at 65°C with 200 mM NaCl and 2 μL RNase A (Sigma). A matched input sample (1/10 of original ChIP reaction) was treated identically. The following day, ChIP samples and inputs were incubated with Proteinase K (Sigma) for 1.5 h at 56°C and purified using ChIP DNA Clean and Concentrator Kit (Zymo Research). For Pol II cChIP-seq, 5x107 mouse ESCs were crosslinked in 1% formaldehyde for 10 min at 25°C and then quenched with 125 mM glycine. Mouse ECSs were mixed with 2x106 single-crosslinked HEK293T cells and incubated in FA-lysis buffer (50 mM HEPES pH 7.9, 150 mM NaCl, 2 mM EDTA, 0.5 mM EGTA, 0.5% NP-40, 0.1% sodium deoxycholate, 0.1% SDS, 10 mM NaF, 1 mM AEBSF, 1x PIC) for 10 min. Chromatin was sonicated for 30 min using the BioRuptor Pico, centrifuged at 20,000 g for 10 min at 4°C, and supernatant taken as chromatin. For ChIP, 300 ug of chromatin was diluted to 1 ml in FA-lysis buffer and pre-cleared for 1 h with protein A agarose beads blocked with 1 mg/mL BSA and 1 mg/mL yeast tRNA. For each ChIP reaction, diluted and pre-cleared chromatin was incubated overnight with the appropriate antibody, anti-Rbp1-NTD (CST, D8L4Y, 15 μl) to detect total Pol II, anti-Rbp1-CTD-Ser5P (CST, D9N5I, 12.5 μl), or anti-Rbp1-CTD-Ser2P (CST, E1Z3G, 12.5 μl). Antibody-bound chromatin was isolated using blocked protein A agarose beads for 3 h at 4°C. Washes were performed with FA-Lysis buffer, FA-Lysis buffer containing 500 mM NaCl, DOC buffer (250 mM LiCl, 0.5% NP-40, 0.5% sodium deoxycholate, 2 mM EDTA, 10 mM Tris–HCl pH 8), followed by two washes with TE buffer pH 8. ChIP DNA was eluted, de-crosslinked and purified as described above, along with the matched input samples. For H2AK119ub1, H3K27me3 and H3K4me3 cChIP-seq, 5x107 mouse ESCs were mixed with 2x107 Drosophila SG4 cells in PBS. The cells were pelleted and nuclei were released by resuspending in ice-cold lysis buffer (10 mM Tris-HCl pH 8, 10 mM NaCl, 3 mM MgCl2, 0.1% NP-40, 5 mM NEM). Nuclei were then washed, and resuspended in 1 mL of MNase digestion buffer (10 mM Tris-HCl pH 8, 10 mM NaCl, 3 mM MgCl2, 0.25 M sucrose, 3 mM CaCl2, 10 mM NEM, 1x PIC). Samples were digested with 150 units of MNase (Fermentas) for 5 min at 37°C, stopped by 4 mM EDTA. The supernatant (S1) was collected following centrifugation at 1500 g for 5 min at 4°C. The remaining pellet was incubated with 300 μl of nucleosome release buffer (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 0.2 mM EDTA, 10 mM NEM, 1x PIC) at 4°C for 1 h, passed five times through a 27G needle, and spun at 1500 g for 5 min at 4°C. The second supernatant (S2) was collected and combined with corresponding S1 supernatant, and stored at -80°C until use. Digestion to predominantly mono-nucleosomal fragments was confirmed by agarose gel electrophoresis of purified DNA. Native chromatin was diluted 10-fold in native ChIP incubation buffer (70 mM NaCl, 10 mM Tris–HCl pH 7.5, 2 mM MgCl2, 2 mM EDTA, 0.1% TritonX-100, 10 mM NEM, 1x PIC). For each ChIP reaction, 1 ml of diluted nucleosomes was incubated overnight at 4°C with the appropriate antibody, anti-H2AK119ub1 (CST, D27C4, 5 μL), anti-H3K27me3 (in-house, 5 μL) or anti-H3K4me3 (in-house, 3 μL). Antibody-bound nucleosomes were captured by incubation for 1 h at 4°C with protein A agarose beads, pre-blocked overnight in native ChIP incubation buffer supplemented with 1 mg/ml BSA and 1 mg/ml yeast tRNA. The beads were then washed four times with native ChIP wash buffer (20 mM Tris–HCl pH 7.5, 2 mM EDTA, 125 mM NaCl, 0.1% TritonX-100) and once with TE buffer pH 8. Immunoprecipitated DNA was eluted using 100 μl of elution buffer (1% SDS, 0.1 M NaHCO3) and purified using ChIP DNA Clean and Concentrator Kit. DNA from a matched input sample (corresponding to 10% of the original ChIP reaction) purified in the same way.
cChIP-seq libraries for both ChIP and input samples were prepared using NEBNext Ultra DNA Library Prep Kit for Illumina (double-crosslinked and native ChIP-seq) or NEBNext Ultra II DNA Library Prep Kit for Illumina (Pol II ChIP-seq), following manufacturer’s guidelines. Samples were indexed using NEBNext Multiplex Oligos. The average size and concentration of all libraries was analysed using the 2100 Bioanalyzer High Sensitivity DNA Kit (Agilent) followed by qPCR using SensiMix SYBR (Bioline, UK) and KAPA Illumina DNA standards (Roche). Libraries were sequenced as 40 bp paired-end reads on the Illumina NextSeq 500.

tag: genotype/variation

tag: chip antibody

tag: antibody info


title: SUZ12dTAG_NativeInput_UNT_rep3


title: E14_wt_ctrl_NativeInput_rep3


title: iCount_ctx_embryo_20190320BJ_E20

tag: mouse strain

value: H3.1-iCOUNT x ROSA26:CreERT2

value: 38h post tamoxifen injection

value: individual iCount+ nuclei

value: Yellow


title: iCount_ctx_embryo_20190320BJ_E19


title: iCount_hOrganoids_d4_20190226BJ_E20

value: 4 days post Cre electroporation


title: E14_wt_ctrl_NativeInput_rep2

tag: cell type


title: iCount_hOrganoids_d4_20190226BJ_F03

value: Orange


title: SUZ12dTAG_NativeInput_dTAG_2h_rep2

tag: replicate


title: iCount_hOrganoids_d4_20190226BJ_E21


title: iCount_hOrganoids_d4_20190226BJ_F08


title: SUZ12dTAG_NativeInput_UNT_rep2

tag: treatment


title: iCount_ctx_embryo_20190320BJ_E18

value: Green


title: SUZ12dTAG_NativeInput_dTAG_2h_rep1

tag: treatment duration


title: iCount_ctx_embryo_20190320BJ_E22


title: iCount_hOrganoids_d4_20190226BJ_E19


title: SUZ12dTAG_H3K27me3_dTAG_2h_rep3

tag: spike-in reference organism

value: In house (Rose, 2016)


title: iCount_ctx_embryo_20190320BJ_E17


title: SUZ12dTAG_H3K27me3_UNT_rep3

tag: spike-in cell line


title: iCount_hOrganoids_d4_20190226BJ_E14


title: iCount_ctx_embryo_20190320BJ_E16


title: iCount_ctx_embryo_20190320BJ_E15


title: iCount_hOrganoids_d4_20190226BJ_E13


title: iCount_ctx_embryo_20190320BJ_E14


title: SUZ12dTAG_H3K27me3_dTAG_2h_rep2


title: E14_wt_ctrl_H3K27me3_rep3


title: SUZ12dTAG_H3K27me3_UNT_rep2


title: iCount_hOrganoids_d4_20190226BJ_E12


title: E14_wt_ctrl_H3K27me3_rep2


title: E14_wt_ctrl_H3K27me3_rep1


title: SUZ12dTAG_H3K27me3_dTAG_2h_rep1


title: SUZ12dTAG_H3K27me3_UNT_rep1


title: RING1AKO.RING1BAID_NucRNA_IAA_24h_rep3

source_name: Mouse embryonic stem cells with Drosophila SG4 spike-in, auxin-treated (24h IAA)

value: E14 Rosa26::Tir1; Ring1A-/-; AID-Ring1B

value: auxin (IAA)

value: 24 hours

tag: molecule subtype


title: RING1AKO.RING1BAID_NucRNA_IAA_24h_rep2

value: Mouse embryonic stem cells


title: iCount_ctx_embryo_20190320BJ_E13

tag: age of the mice/organoids at the time of the tamoxifen injection


title: E14_wt_ctrl_NativeInput_rep1

value: 0 hours

value: Drosophila melanogaster


title: SUZ12dTAG_NativeInput_UNT_rep1

value: E14 Pcgf2-HaloTag; dTAG-Suz12


title: RING1AKO.RING1BAID_NucRNA_IAA_24h_rep1

value: nuclear RNA


title: iCount_hOrganoids_d4_20190226BJ_E04

value: 39 days in culture

tag: time of experiment

value: individual iCount+ cells


title: iCount_hOrganoids_d4_20190226BJ_E16

tag: sorted material


title: RING1AKO.RING1BAID_NucRNA_IAA_8h_rep3

source_name: Mouse embryonic stem cells with Drosophila SG4 spike-in, auxin-treated (8h IAA)

value: 8 hours


title: RING1AKO.RING1BAID_NucRNA_IAA_8h_rep2


title: iCount_ctx_embryo_20190320BJ_E10

value: H3.1-iCOUNT x ROSA26:CreERT2

value: 38h post tamoxifen injection

value: individual iCount+ nuclei

tag: batch number

value: Red


title: iCount_hOrganoids_d4_20190226BJ_D20

value: 4 days post Cre electroporation

tag: icount color

value: Orange


title: RING1AKO.RING1BAID_NucRNA_IAA_4h_rep3

source_name: Mouse embryonic stem cells with Drosophila SG4 spike-in, auxin-treated (4h IAA)

tag: genotype

value: E14 Rosa26::Tir1; Ring1A-/-; AID-Ring1B

value: 4 hours


title: iCount_hOrganoids_d4_20190226BJ_E01


title: iCount_hOrganoids_d4_20190226BJ_D24


title: iCount_ctx_embryo_20190320BJ_E09


title: RING1AKO.RING1BAID_NucRNA_IAA_4h_rep1

value: auxin (IAA)


title: iCount_ctx_embryo_20190320BJ_E12


title: RING1AKO.RING1BAID_NucRNA_IAA_4h_rep2

value: Mouse embryonic stem cells


title: RING1AKO.RING1BAID_NucRNA_IAA_2h_rep3

source_name: Mouse embryonic stem cells with Drosophila SG4 spike-in, auxin-treated (2h IAA)

value: 2 hours


title: RING1AKO.RING1BAID_NucRNA_IAA_8h_rep1

value: Drosophila melanogaster


title: iCount_ctx_embryo_20190320BJ_E07


title: iCount_hOrganoids_d4_20190226BJ_D19


title: iCount_hOrganoids_d4_20190226BJ_E07


title: iCount_hOrganoids_d4_20190226BJ_D17


title: RING1AKO.RING1BAID_NucRNA_IAA_2h_rep1


title: RING1AKO.RING1BAID_NucRNA_UNT_rep2

value: 0 hours


title: RING1AKO.RING1BAID_NucRNA_IAA_2h_rep2

value: nuclear RNA


title: iCount_ctx_embryo_20190320BJ_E04


title: iCount_ctx_embryo_20190320BJ_E06


title: iCount_ctx_embryo_20190320BJ_E11


title: RING1AKO.RING1BAID_NucRNA_UNT_rep1


title: RING1AKO.RING1BAID_NucRNA_UNT_rep3


title: iCount_hOrganoids_d4_20190226BJ_D16

value: 39 days in culture


title: iCount_ctx_embryo_20190320BJ_E03


title: iCount_hOrganoids_d4_20190226BJ_D12

value: individual iCount+ cells

value: Green


title: iCount_hOrganoids_d4_20190226BJ_D10


title: RING1AKO.RING1BAID_NativeInput_IAA_24h_rep2

description: RING1AKO_RING1BAID_Ninput_AUX_24h_rep4_S12

data_processing:
Paired-end reads were aligned to the concatenated mouse (mm10) and spike-in (dm6 for native, hg19 for cross-linked cChIP-seq) genome sequences using Bowtie 2 (“–no-mixed” and “–no-discordant” options). Only uniquely mapped reads were kept for downstream analysis, after removal of PCR duplicates with Sambamba.
To internally calibrate cChIP-seq experiments, we spiked-in a fixed number of control cells (Drosophila SG4 cells for native cChIP-seq and human HEK293T cells for cross-linked cChIP-seq) to each experimental sample
For visualization of cChIP-seq sequencing data and annotation of genomic regions with read counts, uniquely mapped mouse reads were normalized using dm6 or hg19 spike-in as described previously (Furosva et al., 2019). Briefly, mouse reads were randomly subsampled based on the total number of spike-in (dm6 or hg19) reads in each sample. Additionally, to account for possible minor variations in spike-in cell mixing between cChIP-seq replicates, we corrected the subsampling factors by using the ratio of spike-in to mouse total read counts in the corresponding input samples. Prior to merging normalized replicates for visualization and analysis, read coverage across regions of interest (RING1B peaks for RING1B, SUZ12, H2AK119ub1 and H3K27me3, or TSS ± 2.5 kb for total Pol II, Ser5P-Pol II and H3K4me3, or gene bodies for Ser2P) was analyzed using multiBamSummary and plotCorrelation functions from deepTools , confirming a high correlation between replicates (Pearson's correlation coefficient > 0.9). Genome coverage tracks for cChIP-seq were generated using the pileup function from MACS2.
To identify genomic regions bound by PRC1 we performed peak calling using MACS2 (“BAMPE” and “–broad” options) on the RING1B cChIP-seq data with corresponding input samples for background normalization. A set of peaks identified in all biological replicates of untreated PRC1deg cells was used for further analysis, after filtering out several peaks identified following 4 hours of auxin treatment (i.e. following loss of RING1B) and manually removing sequencing artefacts. In total, 7240 stringent RING1B peaks were identified.
Genome_build: mm10
Supplementary_files_format_and_content: bigWig files representing genome coverage for merged replicates of spike-in normalised cChIP-seq.
Supplementary_files_format_and_content: bed file for a set of enrichment peaks for RING1B

extract_protocol:
For RING1B and SUZ12 cChIP-seq, 5x107 mouse ESCs were first crosslinked with 2 mM disuccinimidyl glutarate (DSG, Thermo Scientific) while rotating for 45 min at 25°C, and then with 1% formaldehyde (methanol-free, Thermo Scientific) for further 15 min. Reactions were quenched by addition of 125 mM glycine. Mouse ESCs were then mixed with 2x106 double-crosslinked HEK293T cells and incubated in lysis buffer (50 mM HEPES pH 7.9, 140 mM NaCl, 1 mM EDTA, 10% glycerol, 0.5% NP-40, 0.25% Triton X-100, 1x PIC) for 10 min at 4°C. The released nuclei were washed (10 mM Tris-HCl pH 8, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1x PIC) for 5 min at 4°C. Chromatin was then resuspended in 1 mL of sonication buffer (10 mM Tris-HCl pH 8, 100 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 0.1% sodium deoxycholate, 0.5% N-lauroylsarcosine, 1x PIC) and sonicated for 30 min using the BioRuptor Pico (Diagenode). Following sonication, Triton X-100 was added to a final concentration of 1%. The supernatant was collected after centrifugation at 20,000 g for 10 min at 4°C. For ChIP, chromatin was diluted 10-fold in ChIP dilution buffer (1% Triton X-100, 1 mM EDTA, 20 mM Tris-HCl pH 8, 150 mM NaCl, 1x PIC) and pre-cleared for 1 h with either protein A agarose beads (Repligen, for RING1B ChIP) or protein A magnetic Dynabeads (Invitrogen, for SUZ12 ChIP) blocked with 1 mg/mL bovine serum albumin (BSA) and 1 mg/mL yeast tRNA. For each ChIP reaction, 1 mL of diluted and pre-cleared chromatin was incubated overnight with the appropriate antibody, either anti-RING1B (CST, D22F2, 3 μl), or anti-SUZ12 (CST, D39F6, 3 μl). Antibody-bound chromatin was captured using blocked protein A beads (agarose for RING1B, magnetic Dynabeads for SUZ12) for at least 2 h at 4°C and collected by centrifugation/on a magnetic rack. ChIP washes were performed as described previously (Farcas et al. 2013). ChIP DNA was eluted in elution buffer (1% SDS, 0.1 M NaHCO3) and cross-linking was reversed overnight at 65°C with 200 mM NaCl and 2 μL RNase A (Sigma). A matched input sample (1/10 of original ChIP reaction) was treated identically. The following day, ChIP samples and inputs were incubated with Proteinase K (Sigma) for 1.5 h at 56°C and purified using ChIP DNA Clean and Concentrator Kit (Zymo Research). For Pol II cChIP-seq, 5x107 mouse ESCs were crosslinked in 1% formaldehyde for 10 min at 25°C and then quenched with 125 mM glycine. Mouse ECSs were mixed with 2x106 single-crosslinked HEK293T cells and incubated in FA-lysis buffer (50 mM HEPES pH 7.9, 150 mM NaCl, 2 mM EDTA, 0.5 mM EGTA, 0.5% NP-40, 0.1% sodium deoxycholate, 0.1% SDS, 10 mM NaF, 1 mM AEBSF, 1x PIC) for 10 min. Chromatin was sonicated for 30 min using the BioRuptor Pico, centrifuged at 20,000 g for 10 min at 4°C, and supernatant taken as chromatin. For ChIP, 300 ug of chromatin was diluted to 1 ml in FA-lysis buffer and pre-cleared for 1 h with protein A agarose beads blocked with 1 mg/mL BSA and 1 mg/mL yeast tRNA. For each ChIP reaction, diluted and pre-cleared chromatin was incubated overnight with the appropriate antibody, anti-Rbp1-NTD (CST, D8L4Y, 15 μl) to detect total Pol II, anti-Rbp1-CTD-Ser5P (CST, D9N5I, 12.5 μl), or anti-Rbp1-CTD-Ser2P (CST, E1Z3G, 12.5 μl). Antibody-bound chromatin was isolated using blocked protein A agarose beads for 3 h at 4°C. Washes were performed with FA-Lysis buffer, FA-Lysis buffer containing 500 mM NaCl, DOC buffer (250 mM LiCl, 0.5% NP-40, 0.5% sodium deoxycholate, 2 mM EDTA, 10 mM Tris–HCl pH 8), followed by two washes with TE buffer pH 8. ChIP DNA was eluted, de-crosslinked and purified as described above, along with the matched input samples. For H2AK119ub1, H3K27me3 and H3K4me3 cChIP-seq, 5x107 mouse ESCs were mixed with 2x107 Drosophila SG4 cells in PBS. The cells were pelleted and nuclei were released by resuspending in ice-cold lysis buffer (10 mM Tris-HCl pH 8, 10 mM NaCl, 3 mM MgCl2, 0.1% NP-40, 5 mM NEM). Nuclei were then washed, and resuspended in 1 mL of MNase digestion buffer (10 mM Tris-HCl pH 8, 10 mM NaCl, 3 mM MgCl2, 0.25 M sucrose, 3 mM CaCl2, 10 mM NEM, 1x PIC). Samples were digested with 150 units of MNase (Fermentas) for 5 min at 37°C, stopped by 4 mM EDTA. The supernatant (S1) was collected following centrifugation at 1500 g for 5 min at 4°C. The remaining pellet was incubated with 300 μl of nucleosome release buffer (10 mM Tris-HCl pH 7.5, 10 mM NaCl, 0.2 mM EDTA, 10 mM NEM, 1x PIC) at 4°C for 1 h, passed five times through a 27G needle, and spun at 1500 g for 5 min at 4°C. The second supernatant (S2) was collected and combined with corresponding S1 supernatant, and stored at -80°C until use. Digestion to predominantly mono-nucleosomal fragments was confirmed by agarose gel electrophoresies of purified DNA. Native chromatin was diluted 10-fold in native ChIP incubation buffer (70 mM NaCl, 10 mM Tris–HCl pH 7.5, 2 mM MgCl2, 2 mM EDTA, 0.1% TritonX-100, 10 mM NEM, 1x PIC). For each ChIP reaction, 1 ml of diluted nucleosomes was incubated overnight at 4°C with the appropriate antibody, anti-H2AK119ub1 (CST, D27C4, 5 μL), anti-H3K27me3 (in-house, 5 μL) or anti-H3K4me3 (in-house, 3 μL). Antibody-bound nucleosomes were captured by incubation for 1 h at 4°C with protein A agarose beads, pre-blocked overnight in native ChIP incubation buffer supplemented with 1 mg/ml BSA and 1 mg/ml yeast tRNA. The beads were then washed four times with native ChIP wash buffer (20 mM Tris–HCl pH 7.5, 2 mM EDTA, 125 mM NaCl, 0.1% TritonX-100) and once with TE buffer pH 8. Immunoprecipitated DNA was eluted using 100 μl of elution buffer (1% SDS, 0.1 M NaHCO3) and purified using ChIP DNA Clean and Concentrator Kit. DNA from a matched input sample (corresponding to 10% of the original ChIP reaction) purified in the same way.
cChIP-seq libraries for both ChIP and input samples were prepared using NEBNext Ultra DNA Library Prep Kit for Illumina (double-crosslinked and native ChIP-seq) or NEBNext Ultra II DNA Library Prep Kit for Illumina (Pol II ChIP-seq), following manufacturer’s guidelines. Samples were indexed using NEBNext Multiplex Oligos. The average size and concentration of all libraries was analysed using the 2100 Bioanalyzer High Sensitivity DNA Kit (Agilent) followed by qPCR using SensiMix SYBR (Bioline, UK) and KAPA Illumina DNA standards (Roche). Libraries were sequenced as 40 bp paired-end reads on the Illumina NextSeq 500.

tag: genotype/variation

value: 24 hours

tag: antibody info.


title: RING1AKO.RING1BAID_NativeInput_IAA_24h_rep1

description: RING1AKO_RING1BAID_Ninput_AUX_24h_rep2_S12

tag: chip antibody


title: RING1AKO.RING1BAID_NativeInput_IAA_24h_rep3

description: RING1AKO_RING1BAID_Ninput_AUX_24h_rep5_S22


title: RING1AKO.RING1BAID_NativeInput_IAA_8h_rep2

description: RING1AKO_RING1BAID_Ninput_AUX_8h_rep4_S19

value: 8 hours


title: iCount_ctx_embryo_20190320BJ_D23

tag: mouse strain

value: H3.1-iCOUNT x ROSA26:CreERT2

value: 38h post tamoxifen injection

value: individual iCount+ nuclei


title: iCount_hOrganoids_d4_20190226BJ_D09

value: 4 days post Cre electroporation


title: iCount_hOrganoids_d4_20190226BJ_D08


title: iCount_ctx_embryo_20190320BJ_D22


title: RING1AKO.RING1BAID_NativeInput_for_H3K4me3_IAA_8h_rep1

description: RING1AKO_RING1BAID_Ninput_AUX_8h_rep3_S9

tag: cell type

value: E14 Rosa26::Tir1; Ring1A-/-; AID-Ring1B


title: RING1AKO.RING1BAID_NativeInput_for_H3K27me3_IAA_8h_rep1

description: RING1AKO_RING1BAID_Ninput_AUX_8h_rep2_S9

tag: replicate


title: iCount_hOrganoids_d4_20190226BJ_D06


title: RING1AKO.RING1BAID_NativeInput_for_H3K27me3_IAA_4h_rep1

description: RING1AKO_RING1BAID_Ninput_AUX_4h_rep2_S8

tag: treatment

value: 4 hours


title: iCount_ctx_embryo_20190320BJ_D20


title: iCount_ctx_embryo_20190320BJ_D21


title: RING1AKO.RING1BAID_NativeInput_IAA_4h_rep2

description: RING1AKO_RING1BAID_Ninput_AUX_4h_rep4_S21

value: auxin (IAA)

tag: treatment duration


title: RING1AKO.RING1BAID_NativeInput_IAA_4h_rep3

description: RING1AKO_RING1BAID_Ninput_AUX_4h_rep5_S13

value: Mouse embryonic stem cells

tag: spike-in reference organism


title: iCount_hOrganoids_d4_20190226BJ_D05


title: RING1AKO.RING1BAID_NativeInput_for_H2AK119ub1_H3K4me3_IAA_4h_rep1

description: RING1AKO_RING1BAID_Ninput_AUX_4h_rep3_S14

tag: spike-in cell line


title: iCount_ctx_embryo_20190320BJ_D18


title: iCount_hOrganoids_d4_20190226BJ_D04


title: iCount_ctx_embryo_20190320BJ_E02

value: Orange


title: RING1AKO.RING1BAID_NativeInput_IAA_2h_rep2

description: RING1AKO_RING1BAID_Ninput_AUX_2h_rep4_S22

value: 2 hours

value: Drosophila melanogaster


title: RING1AKO.RING1BAID_NativeInput_IAA_2h_rep3

description: RING1AKO_RING1BAID_Ninput_AUX_2h_rep5_S4


title: RING1AKO.RING1BAID_NativeInput_for_H3K27me3_IAA_2h_rep1

description: RING1AKO_RING1BAID_Ninput_AUX_2h_rep2_S7


title: iCount_hOrganoids_d4_20190226BJ_D03


title: RING1AKO.RING1BAID_NativeInput_for_H2AK119ub1_H3K4me3_IAA_2h_rep1

description: RING1AKO_RING1BAID_Ninput_AUX_2h_rep3_S15


title: RING1AKO.RING1BAID_NativeInput_UNT_rep3

value: 0 hours


title: RING1AKO.RING1BAID_NativeInput_IAA_8h_rep3

description: RING1AKO_RING1BAID_Ninput_AUX_8h_rep5_S21


title: iCount_ctx_embryo_20190320BJ_D16


title: RING1AKO.RING1BAID_NativeInput_UNT_rep2


title: iCount_hOrganoids_d4_20190226BJ_D01

value: 39 days in culture


title: RING1AKO.RING1BAID_NativeInput_for_H3K27me3_UNT_rep1


title: iCount_hOrganoids_d4_20190226BJ_C22

value: individual iCount+ cells


title: iCount_hOrganoids_d4_20190226BJ_C18


title: RING1AKO.RING1BAID_H3K4me3_IAA_8h_rep3

description: RING1AKO_RING1BAID_H3K4me3_AUX_8h_rep5_S14

value: In house (Farcas, 2012)


title: iCount_ctx_embryo_20190320BJ_D14

description: RawCount_Mouse_iCount_Embryonic_Cortex.txt

value: Green


title: RING1AKO.RING1BAID_NativeInput_for_H2AK119ub1_H3K4me3_UNT_rep1


title: iCount_ctx_embryo_20190320BJ_D15


title: RING1AKO.RING1BAID_H3K4me3_IAA_8h_rep2

description: RING1AKO_RING1BAID_H3K4me3_AUX_8h_rep4_S9

value: 8 hours


title: RING1AKO.RING1BAID_H3K4me3_IAA_4h_rep3

description: RING1AKO_RING1BAID_H3K4me3_AUX_4h_rep5_S13


title: iCount_ctx_embryo_20190320BJ_D13

value: H3.1-iCOUNT x ROSA26:CreERT2


title: iCount_ctx_embryo_20190320BJ_D03

value: 38h post tamoxifen injection


title: iCount_ctx_embryo_20190320BJ_D17

value: individual iCount+ nuclei


title: iCount_hOrganoids_d4_20190226BJ_C16

value: 4 days post Cre electroporation


title: RING1AKO.RING1BAID_H3K4me3_IAA_2h_rep3

description: RING1AKO_RING1BAID_H3K4me3_AUX_2h_rep5_S12

value: E14 Rosa26::Tir1; Ring1A-/-; AID-Ring1B


title: RING1AKO.RING1BAID_H3K4me3_IAA_4h_rep2

description: RING1AKO_RING1BAID_H3K4me3_AUX_4h_rep4_S8

value: 4 hours


title: RING1AKO.RING1BAID_H3K4me3_IAA_4h_rep1

description: RING1AKO_RING1BAID_H3K4me3_AUX_4h_rep3_S3

value: auxin (IAA)


title: iCount_hOrganoids_d4_20190226BJ_C13


title: RING1AKO.RING1BAID_H3K4me3_IAA_2h_rep1

description: RING1AKO_RING1BAID_H3K4me3_AUX_2h_rep3_S2

value: Mouse embryonic stem cells


title: RING1AKO.RING1BAID_H3K4me3_IAA_8h_rep1

description: RING1AKO_RING1BAID_H3K4me3_AUX_8h_rep3_S4


title: iCount_ctx_embryo_20190320BJ_C24


title: iCount_ctx_embryo_20190320BJ_D02

value: Orange


title: RING1AKO.RING1BAID_H3K4me3_IAA_2h_rep2

description: RING1AKO_RING1BAID_H3K4me3_AUX_2h_rep4_S7

value: 2 hours


title: RING1AKO.RING1BAID_H3K4me3_UNT_rep3

value: Drosophila melanogaster


title: iCount_hOrganoids_d4_20190226BJ_C11


title: iCount_ctx_embryo_20190320BJ_C23


title: iCount_hOrganoids_d4_20190226BJ_C10


title: RING1AKO.RING1BAID_H3K4me3_UNT_rep2

value: 0 hours


title: iCount_ctx_embryo_20190320BJ_C21


title: iCount_ctx_embryo_20190320BJ_C20


title: RING1AKO.RING1BAID_H3K27me3_IAA_24h_rep2

description: RING1AKO_RING1BAID_H3K27me3_AUX_24h_rep4_S18

value: 24 hours

value: In house (Rose, 2016)


title: iCount_hOrganoids_d4_20190226BJ_C08

value: 39 days in culture

value: individual iCount+ cells


title: iCount_hOrganoids_d4_20190226BJ_C15


title: RING1AKO.RING1BAID_H3K27me3_IAA_8h_rep3

description: RING1AKO_RING1BAID_H3K27me3_AUX_8h_rep5_S19

value: 8 hours


title: iCount_hOrganoids_d4_20190226BJ_C07

value: Green


title: RING1AKO.RING1BAID_H3K27me3_IAA_8h_rep2

description: RING1AKO_RING1BAID_H3K27me3_AUX_8h_rep4_S1


title: iCount_ctx_embryo_20190320BJ_C19

value: H3.1-iCOUNT x ROSA26:CreERT2

value: 38h post tamoxifen injection


title: RING1AKO.RING1BAID_H3K27me3_IAA_8h_rep1

description: RING1AKO_RING1BAID_H3K27me3_AUX_8h_rep2_S4


title: iCount_hOrganoids_d4_20190226BJ_C05

value: 4 days post Cre electroporation


title: iCount_ctx_embryo_20190320BJ_C18

value: individual iCount+ nuclei


title: RING1AKO.RING1BAID_H3K27me3_IAA_2h_rep3

description: RING1AKO_RING1BAID_H3K27me3_AUX_2h_rep5_S17

value: E14 Rosa26::Tir1; Ring1A-/-; AID-Ring1B


title: iCount_hOrganoids_d4_20190226BJ_C02


title: iCount_ctx_embryo_20190320BJ_C17


title: RING1AKO.RING1BAID_H3K27me3_IAA_4h_rep1

description: RING1AKO_RING1BAID_H3K27me3_AUX_4h_rep2_S3

value: auxin (IAA)

value: 4 hours


title: iCount_ctx_embryo_20190320BJ_C15


title: RING1AKO.RING1BAID_H3K27me3_IAA_4h_rep3

description: RING1AKO_RING1BAID_H3K27me3_AUX_4h_rep5_S18

value: Mouse embryonic stem cells


title: iCount_hOrganoids_d4_20190226BJ_C04


title: RING1AKO.RING1BAID_H3K27me3_IAA_4h_rep2

description: RING1AKO_RING1BAID_H3K27me3_AUX_4h_rep4_S10


title: RING1AKO.RING1BAID_H3K4me3_UNT_rep1

value: In house (Farcas, 2012)


title: RING1AKO.RING1BAID_H3K27me3_IAA_2h_rep2

description: RING1AKO_RING1BAID_H3K27me3_AUX_2h_rep4_S2

value: 2 hours

value: Drosophila melanogaster


title: RING1AKO.RING1BAID_H3K27me3_IAA_2h_rep1

description: RING1AKO_RING1BAID_H3K27me3_AUX_2h_rep2_S2


title: iCount_hOrganoids_d4_20190226BJ_C01


title: RING1AKO.RING1BAID_H3K27me3_IAA_24h_rep3

description: RING1AKO_RING1BAID_H3K27me3_AUX_24h_rep5_S20

value: 24 hours


title: RING1AKO.RING1BAID_H3K27me3_UNT_rep3

description: RING1AKO_RING1BAID_H3K27me3_UNT_rep5_S16

value: 0 hours

value: In house (Rose, 2016)


title: iCount_ctx_embryo_20190320BJ_C14

description: RawCount_Mouse_iCount_Embryonic_Cortex.txt


title: RING1AKO.RING1BAID_H3K27me3_UNT_rep2


title: iCount_hOrganoids_d4_20190226BJ_B24

value: 39 days in culture

value: individual iCount+ cells

value: Orange


title: RING1AKO.RING1BAID_H3K27me3_IAA_24h_rep1

description: RING1AKO_RING1BAID_H3K27me3_AUX_24h_rep2_S5


title: iCount_ctx_embryo_20190320BJ_C12

value: H3.1-iCOUNT x ROSA26:CreERT2

value: Red


title: iCount_hOrganoids_d4_20190226BJ_B21

value: 4 days post Cre electroporation


title: RING1AKO.RING1BAID_H3K27me3_UNT_rep1


title: RING1AKO.RING1BAID_H2AK119ub1_IAA_4h_rep3

value: Cell Signaling Technology (#D27C4 )


title: iCount_ctx_embryo_20190320BJ_C11

value: 38h post tamoxifen injection

value: individual iCount+ nuclei


title: RING1AKO.RING1BAID_H2AK119ub1_IAA_4h_rep2

value: E14 Rosa26::Tir1; Ring1A-/-; AID-Ring1B

value: auxin (IAA)

value: 4 hours


title: RING1AKO.RING1BAID_H2AK119ub1_IAA_4h_rep1

value: Mouse embryonic stem cells


title: RING1AKO.RING1BAID_H2AK119ub1_IAA_2h_rep3

value: 2 hours

value: Drosophila melanogaster


title: iCount_hOrganoids_d4_20190226BJ_B20


title: iCount_hOrganoids_d4_20190226BJ_B17


title: iCount_ctx_embryo_20190320BJ_C10


title: iCount_ctx_embryo_20190320BJ_C07


title: iCount_hOrganoids_d4_20190226BJ_B14


title: iCount_ctx_embryo_20190320BJ_C08


title: iCount_ctx_embryo_20190320BJ_C05


title: iCount_hOrganoids_d4_20190226BJ_B13


title: RING1AKO.RING1BAID_H2AK119ub1_IAA_2h_rep2


title: RING1AKO.RING1BAID_H2AK119ub1_IAA_2h_rep1


title: iCount_ctx_embryo_20190320BJ_C06


title: RING1AKO.RING1BAID_H2AK119ub1_UNT_rep2

value: 0 hours


title: RING1AKO.RING1BAID_H2AK119ub1_UNT_rep1


title: iCount_hOrganoids_d4_20190226BJ_B11

value: Green


title: iCount_ctx_embryo_20190320BJ_C03


title: iCount_ctx_embryo_20190320BJ_C02


title: RING1AKO.RING1BAID_H2AK119ub1_UNT_rep3


title: iCount_hOrganoids_d4_20190226BJ_B15

value: 39 days in culture


title: iCount_hOrganoids_d4_20190226BJ_B09

value: individual iCount+ cells


title: iCount_ctx_embryo_20190320BJ_C01


title: iCount_ctx_embryo_20190320BJ_B23


title: iCount_hOrganoids_d4_20190226BJ_B06


title: iCount_ctx_embryo_20190320BJ_B22


title: iCount_hOrganoids_d4_20190226BJ_B05


title: iCount_ctx_embryo_20190320BJ_B20


title: iCount_hOrganoids_d4_20190226BJ_B04


title: iCount_hOrganoids_d4_20190226BJ_B10


title: Lymph Node_Donor_LN08_CFSE_neg

type: RNA

city: Cleveland

first: Mark

last: Cameron

state: OH

address: 10900 Euclid Ave

department: Population and Quantitative Health Sciences

institute: Case Western Reserve University

description:
PAXgene Sample
9610356004_G

hyb_protocol: Standard Illumina hybridization protocol for HumanHT-12_V4 BeadChips.

scan_protocol: Standard Illumina scanning protocol

data_processing: The data were normalised with the normalizeBetweenArrays function using the quantile method in limma. After this,  normalized values less than 2^0.1 or missing were replaced with that value and log2 transformed.  Detection Pvalues are not available.

label: biotin

source_name: Lymph Node

label_protocol: Biotinylated cRNA was oligo-dT primed and prepared with the Illumina TotalPrep RNA amplification kits (Ambion).

extract_protocol: Cells were sorted directly into RLT lysis buffer with beta mercaptoethanol added and then flash frozen on dry ice. Total RNA was isolated using Qiagen's RNeasy Micro kit (cat# 74004)

tag: tissue

value: lymph node

tag: hiv status

value: neg

tag: donor id

tag: cfse status


title: Lymph Node_Donor_LN04_CFSE_neg

description:
PAXgene Sample
9610356004_K


title: Lymph Node_Donor_LNNY_CFSE_neg

description:
PAXgene Sample
9610356004_L

value: pos

value: LNNY


title: Lymph Node_Donor_LN102_CFSE_neg

description:
PAXgene Sample
9610356004_A


title: iCount_hOrganoids_d4_20190226BJ_B08

tag: age of the mice/organoids at the time of the tamoxifen injection

tag: time of experiment

value: 4 days post Cre electroporation

tag: sorted material

tag: batch number


title: iCount_hOrganoids_d4_20190226BJ_B03

tag: icount color


title: Lymph Node_Donor_LN07_CFSE_neg

description:
PAXgene Sample
9610356004_H


title: iCount_ctx_embryo_20190320BJ_B19

value: H3.1-iCOUNT x ROSA26:CreERT2

value: 38h post tamoxifen injection


title: Lymph Node_Donor_LN04_CFSE_pos

description:
PAXgene Sample
9610356004_D


title: Lymph Node_Donor_LNJS_CFSE_pos

description:
PAXgene Sample
9610356004_F

value: LNJS


title: Lymph Node_Donor_LNJB_CFSE_neg

description:
PAXgene Sample
9610356004_B

value: LNJB


title: Lymph Node_Donor_LNJB_CFSE_pos

description:
PAXgene Sample
9610356004_J


title: iCount_ctx_embryo_20190320BJ_B18

value: individual iCount+ nuclei


title: iCount_hOrganoids_d4_20190226BJ_B02


title: iCount_ctx_embryo_20190320BJ_B15


title: Lymph Node_Donor_LNSA_CFSE_pos

description:
PAXgene Sample
9477874117_L

value: LNSA


title: iCount_hOrganoids_d4_20190226BJ_B01


title: Lymph Node_Donor_LN7_26_12_CFSE_neg

description:
PAXgene Sample
9477874117_F


title: iCount_ctx_embryo_20190320BJ_B14


title: Lymph Node_Donor_LNNY_CFSE_pos

description:
PAXgene Sample
9477874117_H


title: Lymph Node_Donor_LNWR_CFSE_pos

description:
PAXgene Sample
9610356004_C

value: LNWR


title: CD40L_KO_R3

city: Belfast

first: Dessi

last: Malinova

address: 97 Lisburn Road

department: WWIEM

institute: Queen's University Belfast

data_processing:
Illumina software used for basecalling
sequenced reads trimmed for adaptor sequence (cutadapt version 1.9.1), low quality and low complexity reads excluded
mapping and subsequent gene-level counting of sequenced reads was performed using the RSEM package (version 1.3.0) and the STAR alignment algorithm (version 2.5.2a)
Genome_build: Ensembl genome browser (assembly GRCm38, release 89)
Supplementary_files_format_and_content: expected_count, TPM and FPKM values for every sample

source_name: CD40L-treated follicular B cells

extract_protocol:
RNA extracted using Qiagen RNeasy mini kit according to manufacturer instructions
stranded mRNA libraries were prepared using polyA_KAPA_mRNA_Hyper_Prep (Roche)

tag: strain

tag: genotype

value: CD40L treatment for 14hours


title: CD40L_KO_R1


title: CD40L_KO_R2


title: Lymph Node_Donor_LN07_CFSE_pos

description:
PAXgene Sample
9610356004_E


title: Lymph Node_Donor_LNJS_CFSE_neg

description:
PAXgene Sample
9477874117_D


title: Lymph Node_Donor_LN08_CFSE_pos

description:
PAXgene Sample
9477874117_E


title: Lymph Node_Donor_LNSA_CFSE_neg

description:
PAXgene Sample
9477874117_K


title: Lymph Node_Donor_LNEK_CFSE_pos

description:
PAXgene Sample
9477874117_A

value: LNEK


title: Lymph Node_Donor_LNEK_CFSE_neg

description:
PAXgene Sample
9477874117_C


title: iCount_ctx_embryo_20190320BJ_B12

tag: mouse strain

value: Orange


title: iCount_hOrganoids_d4_20190226BJ_A19

value: 39 days in culture


title: iCount_hOrganoids_d4_20190226BJ_A15

value: individual iCount+ cells


title: CD40L_WT_R2

value: wildtype


title: CD40L_WT_R1


title: FO_KO_R3

source_name: follicular B cells

value: untreated


title: iCount_hOrganoids_d4_20190226BJ_A14


title: FO_KO_R1


title: CD40L_WT_R3


title: iCount_ctx_embryo_20190320BJ_B11


title: Lymph Node_Donor_LNWR_CFSE_neg

description:
PAXgene Sample
9477874117_G

value: lymph node

value: pos

value: neg


title: FO_WT_R2


title: FO_WT_R3


title: iCount_hOrganoids_d4_20190226BJ_A12

value: 4 days post Cre electroporation

value: Green


title: iCount_ctx_embryo_20190320BJ_B08

value: H3.1-iCOUNT x ROSA26:CreERT2

value: 38h post tamoxifen injection


title: FO_WT_R1


title: iCount_hOrganoids_d4_20190226BJ_A11


title: TRANS_KO_R2

source_name: transitional B cells


title: TRANS_KO_R1


title: iCount_ctx_embryo_20190320BJ_B06

value: individual iCount+ nuclei

value: Red


title: FO_KO_R2


title: TRANS_WT_R3


title: iCount_hOrganoids_d4_20190226BJ_A09


title: TRANS_WT_R2


title: iCount_ctx_embryo_20190320BJ_B05


title: iCount_ctx_embryo_20190320BJ_B10


title: iCount_hOrganoids_d4_20190226BJ_A08


title: TRANS_WT_R1


title: iCount_ctx_embryo_20190320BJ_A20


title: iCount_hOrganoids_d4_20190226BJ_A06


title: iCount_hOrganoids_d4_20190226BJ_A07


title: iCount_hOrganoids_d4_20190226BJ_A13


title: iCount_ctx_embryo_20190320BJ_A18


title: TRANS_KO_R3


title: iCount_hOrganoids_d4_20190226BJ_A05


title: iCount_ctx_embryo_20190320BJ_A15


title: iCount_hOrganoids_d4_20190226BJ_A02


title: iCount_ctx_embryo_20190320BJ_A14


title: iCount_ctx_embryo_20190320BJ_B09


title: RR107e_00126_Peds

city: San Francisco

first: Charles

last: Langelier

state: CA

address: HSE 401, 513 Parnassus Ave

department: Medicine

institute: University of California San Francisco

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00126

data_processing:
Alignment against human genome- Kallisto v 0.46.1
Gene-level counts were generated from the transcript-level abundance estimates using the R package tximport
Fitering to retain samples with 400,000 gene counts
Differential expression with R package limma
Genome_build: HG-38, ENSEMBL v. 99
Supplementary_files_format_and_content: .csv files with genecounts

source_name: NP/OP swab

growth_protocol: Clinical naso/pharyngeal swab specimens

extract_protocol:
Zymo DNA/RNA magbead
NEBnext with Qiagen Fastselect rRNA depletion
Random hexamer RNAseq

treatment_protocol: DNAse

tag: gender

value: M

tag: age

tag: infection

tag: sample type

value: naso/pharyngeal swab


title: RR107e_00095_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00095


title: RR107e_00091_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00091


title: RR107e_00078_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00078


title: RR107e_00100_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00100


title: 9_primary_tenocyte

city: Kyoto

first: Masayoshi

last: Tsukahara

state: Kyoto

address: Shogoin kawahara-cho, sakyo-ku,

department: Research and Develpoment center

country: Japan

institute: CiRA foundation

data_processing:
Illumina bcl2fastq-2.20 used for basecalling
10x genomics cellranger-v3.1.0 used for demultiplex, mapping and read count.
Genome_build: GRCh38
Supplementary_files_format_and_content: barcodes.tsv, features.tsv, matrix.mtx

source_name: tenocyte

growth_protocol: Human iPS cell line, 1231A3, was cultivated on iMartix-511 (0.5 μg/cm2)-coated dish with StemFit AK03 medium, and passaged via dissociation into single cells using Accutase.

extract_protocol: Dissociated cells were applied to 10x genomics chip. 10x genomics chromium single cell 3' library & Gel Bead kit v3 were used.

treatment_protocol: iPSCs were cultivated in the medium containing transforming growth factor-βinhibitor SB431542 (SB), an WNT activator CHIR99021 (CHIR), a bone morphologic protein (BMP) inhibitor DMH1, and fibroblast growth factor-2 recombinant protein for first 4 days, followed by a medium containing only SB and CHIR so that differentiate iPSCs into somite through presomitic mesoderm stage. At day 8, smoothened agonist SAG and a BMP inhibitor LDN193189 were added to the medium to generate sclerotome. Then, the cells were treated with FGF8 and TGFβ3 for 2 days followed by BMP7 and TGFβ3 for 6 days.

tag: cell type

value: primary tenocyte


title: iCount_ctx_embryo_20190320BJ_A22

source_name: embryonic cortex


title: 8_SYN_day8

source_name: syndetome

value: iPS-derived syndetome


title: RR107e_00125_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00125

value: F


title: RR107e_00073_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00073


title: iCount_ctx_embryo_20190320BJ_A13


title: iCount_ctx_embryo_20190320BJ_A11

value: Orange


title: RR107e_00070_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00070


title: RR107e_00063_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00063


title: iCount_ctx_embryo_20190320BJ_A09


title: RR107e_00066_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00066


title: 7_SYN_day6


title: 6_SYN_day4


title: RR107e_00059_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00059


title: gfp_static_V

city: Mannheim

first: Carsten

last: Sticht

address: Theodor-Kutzer-Ufer

department: ZMF

country: Germany

institute: University Heidelberg

hyb_protocol: Hybridization (16h x 45°C) was processed according to the standard Affymetrix protocol.

scan_protocol: Affymetrix GeneArray Scanner 3000.

data_processing: The data were analyzed with a commercial software called JMP Genomics, version 6, from SAS. Gene expression profiling was performed using arrays of human Hugene-2_0-type from Affymetrix. A Custom CDF Version 20 with Entrez based gene definitions was used to annotate the arrays. The Raw fluorescence intensity values were normalized applying quantile normalization, RMA background correction and Medianpolish Probeset Summary.

source_name: HUASMCs - treated with AdGFP adenovirus

label_protocol: Biotinylated cRNA were prepared according to the standard Affymetrix protocol.

growth_protocol: Human umbilical artery smooth muscle cells (HUASMCs) were cultured in DMEM +15% FCS and propagated 1:3.

extract_protocol: RNA was extracted from VSMCs by utilizing the Qiagen mini RNA extraction kit following man. instr.

treatment_protocol: HUASMCs were exposed to adenovirus (AdGFP - control GFP overexpression, AdRGS5 - RGS5 overexpression) before RNA extraction.

tag: treatment

value: Control - GFP overexpression

tag: status

value: static


title: rgs5_static_VI

source_name: HUASMCs - treated with AdRGS5 adenovirus

value: Human umbilical artery smooth muscle cells

value: RGS5 overexpression


title: RR107e_00058_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00058


title: rgs5_static_V


title: RR107e_00056_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00056


title: 5_SYN_day2


title: gfp_static_IV


title: rgs5_static_IV


title: gfp_static_VI


title: RR107e_00055_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00055


title: 4_SCL

source_name: sclerotome

value: iPS-derived sclerotome


title: RR107e_00051_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00051


title: RR107e_00043_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00043


title: naive-2_S1 [WGBS]

city: La Jolla

first: Anjana

last: Rao

state: California

address: 9420 Athena Cir

department: Signaling and Gene Expression

institute: La Jolla Institute for Allergy and Immunology

description:
processed data file:
Group.common.5.annot.cov.gz
Sample.common.5.annot.cov.gz
WT_naive.minCov.5.bw.gz

data_processing:
Bcl2fastq v2.20.0.442 was used to obtain fastq files from Illumina
WGBS FASTQ files were quality and adapter trimmed using Trim Galore (v0.4.2) prior to mapping to the in silico bisulfite converted UCSC mouse genome mm9 using Bismark (v0.19.0) to call Bowtie2 (v2.2.6).
Mapped BAM files were sorted with Samtools (v1.7) and CpG methylation calls were extracted using bespoke code as previously described.
CpG methylation calls were collapsed to one strand and DNA methylation coverage was determined as CpGs with ≥5x coverage per group resulting in DNA methylation data at 18,531,012 CpGs with an average coverage of 14.9x per CpG per sample (range 11.7x – 17.8x).
Genome_build: mm9
Supplementary_files_format_and_content: *common.5.annot.cov.gz: Tab delimited file with the methylated and unmethylated raw counts in CpGs that had at least 5x coverage in at least one group or samples
Supplementary_files_format_and_content: *bw.gz: Bigwig file with the WGBS replicates in CpGs with 5x minimum coverage

extract_protocol:
Genomic DNA was extracted using either Purelink genomic DNA mini kit (Invitrogen) or FlexiGene DNA Kit (Qiagen)
Unmethylated lambda DNA (Promega, D1521) was spiked into the genomic DNA samples at a ratio of 1:200 to monitor the bisulfite conversion efficiency. The samples were then sheared using Covaris S2, purified with Ampure XP beads (Beckman Coulter) and processed with NEBNext End Repair and dA-Tailing Modules (NEB), and ligated to methylated Illumina Adaptors using NEBNext Quick Ligation Module (NEB). DNA with ligated adaptors was then treated with sodium bisulfite (MethylCode, Thermo Fisher Scientific) for 4hrs and amplified with index primers using KAPA HiFi HotStart Uracil+ Ready Mix (KAPA Biosystems).
Whole genome bisulfite sequencing (WGBS)

treatment_protocol: Cells were sorted directly from the mice or differentiated from CD4+ naïve T cells into  iTreg cells under the indicated conditions

value: 5-6 weeks

tag: genotype/variation

value: Foxp3-eGFP reporter mice

tag: tissue

value: Spleen and lymph nodes

tag: sorting/differentiation strategy

value: CD4+eGFP-CD25-CD62LhiCD44lo naïve T cells

tag: molecule subtype

value: Bisulfite-treated genomic DNA


title: Treg-2_S3 [WGBS]

description:
processed data file:
Group.common.5.annot.cov.gz
Sample.common.5.annot.cov.gz
WT_Treg.minCov.5.bw.gz

value: CD4+eGFP+CD25+ Treg cells


title: 3_SM

source_name: somitic mesoderm

value: iPS-derived somitic mesoderm


title: naive-3_S2 [WGBS]


title: RR107e_00044_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00044

value: M

value: naso/pharyngeal swab


title: 2_PSM

source_name: presomitic mesoderm

value: iPS-derived presomitic mesoderm


title: RR101e_00092_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00092

value: F


title: dko-3_S6 [WGBS]

description:
processed data file:
Group.common.5.annot.cov.gz
Sample.common.5.annot.cov.gz
DKO_Treg.minCov.5.bw.gz

value: 3 weeks

value: Tet2fl/fl Tet3fl/fl Foxp3-eGFP CD4-Cre


title: 1_iPS

source_name: induced pluripotent stem cell

tag: cell line

value: Human iPS cell line, 1231A3


title: Treg-3_S4 [WGBS]


title: dko-2_S5 [WGBS]


title: RR101e_00090_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00090


title: RR101e_00091_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00091


title: RR101e_00088_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00088


title: RR101e_00087_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00087


title: plate_9_scRNAseq: mbs8-22-23

city: Beijing

first: Xiangxiang

last: Jiang

state: Beijing

address: 1 Beichen West Road, Chaoyang District

country: China

institute: Institute of Zoology, Chinese Academy of Sciences

description: 3' end of mRNA

data_processing:
The paired-end reads Smart-seq2 data was processed using the standard pipeline of Drop-seq_tools-2.0.0 (http://mccarrolllab.com/dropseq/) with modifications.
For read2, bases 1 to 8 were tagged with cell-barcode “XC,”and bases 8 to 16 were tagged with UMI “XM”.
Read1 was trimmed at the 5' end to remove adaptor and TSO sequence, and the 3' end was trimmed to remove poly(A) sequences of length 6 or more and then aligned to Macaca_fascicularis_5.0 genome reference using STAR aligner.
Gene expression matrix for each cell was then generated with DigitalExpression command.
Genome_build: Mus_musculus.GRCm38.84
Supplementary_files_format_and_content: csv (Comma Separate Values) files include UMI (unique molecular identifiers ) count values for each cell

source_name: Blastocyst

growth_protocol: The embryos at the early two-, four- or eight-cell stages were obtained from the oviduct and then moved to drops of EmbryoMax KSOM medium covered with paraffin oil after washing several times in KSOM medium in an incubator at 37°C and 5% CO2.

extract_protocol:
The blastocysts were exposed to acid Tyrode’s solution to remove the zona pellucida and washed thoroughly in PBS with 0.1% BSA. The denuded embryos were treated with 0.5% trypsin for 10 min at 37°C dispersing them using thorough pipetting. Single GFP-positive cells were deposited into 2.5 μl lysis buffer with a mouth pipette
The linearized RNA molecules were releasein a thermocycler at 72 °C for 3 min at first. Then, transfer the reactions immediately on ice. Add 2.85 µl RT mixture into every reaction with component as SuperScript II reverse transcriptase, RNase Inhibitor, 5× Superscript II first-strand buffer, dithiothreitol (DTT), betaine, MgCl2, and template switch oligo (TSO) primer. Spin down the 5.4 µl samples in the thermocycler at 25 °C for 5 min, 42 °C for 60 min, 50 °C for 30 min, and then 70 °C for 10 min, 4 °C hold. Prepare 7.5 µl PCR mixture (6.25 μL 2× KAPA HiFi HotStart ReadyMix, ISPCR oligo, and 3′ Anchored oligo) to add in every tube reaction, and amplify samples for 19 cycles. Pool together samples with different barcodes (up to 96 barcodes) and purify DNA. Then prepare the second round of amplification with bio-tinylated index primer and ISPCR oligo for an additional five cycles. Subsequently, purify the biotinylated cDNAs further with 0.8× Ampure XP beads and elute cDNA in 50 µl EB. Next, shear the cDNAs into approximately 300-bp fragments with sonicator.Enrich the cDNA fragments  by Dynabeads® MyOneTM Streptavi- din C1. Libraries were prepared with KAPA Hyper Prep Kits. Use the NEB U-shaped adapter for ligation. Finally, amplify the adapter-ligated fragments using an Illumina QP2 primer and a short universal primer for 9 cycles.

treatment_protocol: Theembryos must be washed several times in M2 medium before culture in KSOM.

tag: condittion

value: in vitro

tag: developmental stage

value: Blastocyst

value: embryo


title: RR101e_00086_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00086


title: RR101e_00084_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00084


title: plate_8_scRNAseq: mbs8-16-21

description: 3' end of mRNA


title: RR101e_00083_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00083


title: plate_6_scRNAseq: mbs4-31-36

description: 3' end of mRNA


title: plate_7_scRNAseq: mbs8-01-09

description: 3' end of mRNA


title: TGF-3-5_S3 [WGBS]

description:
processed data file:
Group.common.5.annot.cov.gz
Sample.common.5.annot.cov.gz
TGF.minCov.5.bw.gz

tag: strain

value: 5-6 weeks

value: iTregs differentiated in the presence of TGFβ


title: RA-VC-3-5_S4 [WGBS]

description:
processed data file:
Group.common.5.annot.cov.gz
Sample.common.5.annot.cov.gz
RA_VC.minCov.5.bw.gz

value: Foxp3-eGFP reporter mice

value: Spleen and lymph nodes

value: iTregs differentiated in the presence of TGFβ+RA+VitC


title: RR101e_00078_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00078


title: TGF-1-16_S1 [WGBS]

value: Bisulfite-treated genomic DNA


title: RR107e_00039_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR107e_00039


title: RR101e_00077_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00077


title: RR101e_00080_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00080


title: naive2-Input

data_processing:
Bcl2fastq v2.20.0.442 was used to obtain fastq files from Illumina
CMS-IP FASTQ files were quality and adapter trimmed using Trim Galore (v0.4.2).
Trimmed FASTQ files were mapped to the in silico bisulfite converted mm9 genome with the unmethylated and 5hmC amplicons added as artificial chromosomes.
Mapped SAM files were converted to BAM and putative PCR duplicates were marked with Samtools (v1.7 ).
Normalization was performed using the 5hmC spike-ins.
Regions of enriched 5hmC were determined using MACS2 (v2.1.0.20151222) with the following settings “-g mm --broad -q 0.01”.
Genome_build: mm9
Supplementary_files_format_and_content: MACS2_broad_q_0.01.union.annot.clean.cts.gz: Raw counts, each column represent a sample
Supplementary_files_format_and_content: MACS2_broad_q_0.01.union.annot.clean.rpm.spikein.cov.gz: RPM normalized counts, each column represent a sample

extract_protocol:
Genomic DNA was extracted using either Purelink genomic DNA mini kit (Invitrogen) or FlexiGene DNA Kit (Qiagen)
genomic DNA was sheared using Covaris S2 and was spiked with two 210-base-pair PCR amplicons (C amplicon and 5hmC amplicon) at a ratio of around 1:20,000. DNA was purified with Ampure XP beads (Beckman Coulter) and processed with NEBNext End Repair and dA-Tailing Modules (NEB), and ligated to methylated Illumina Adaptors using NEBNext Quick Ligation Module (NEB). DNA with ligated adaptors was then treated with sodium bisulfite (MethylCode, Thermo Fisher Scientific) for 4hrs, denatured and immunoprecipitated with anti-CMS serum (input samples were reserved as 1% of total DNA before immunoprecipitation). Samples for immunoprecipitated DNA and input DNA were then purified using Phenol/Chloroform and amplified with index primers using KAPA HiFi HotStart Uracil+ Ready Mix (KAPA Biosystems).
Sodium bisulfite treatment converts 5hmC into CMS (cytosine-5-methylenesulfonate), which is then recognized by anti-CMS serum with high sensitivity and selectivity to pull down 5hmC enriched fragments

value: CD4+eGFP-CD25-CD62LhiCD44lo naïve T cells

value: Input for 5hmC-pulldown by CMSIP


title: RA-VC-1-16_S2 [WGBS]


title: RR101e_00074_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00074

value: naso/pharyngeal swab


title: plate_5_scRNAseq: mbs4-26-30

description: 3' end of mRNA


title: plate_4_scRNAseq: mbs4-24-25

description: 3' end of mRNA


title: naive2-IP

value: 5hmC-enriched by CMSIP


title: RR101e_00068_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00068

value: F


title: RR101e_00069_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00069

value: M


title: plate_3_scRNAseq: mbs4-14-23

description: 3' end of mRNA


title: RR101e_00066_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00066


title: naive1-Input


title: RR101e_00065_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00065


title: naive1-IP


title: RR101e_00064_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00064


title: RR101e_00082_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00082


title: plate_2_scRNAseq: mbs4-07-12

description: 3' end of mRNA


title: RR101e_00063_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00063


title: plate_1_scRNAseq: mbs4-01-06

description: 3' end of mRNA


title: RR101e_00062_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00062


title: Treg2-Input

value: CD4+eGFP+ Treg cells


title: RR101e_00071_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00071


title: RR101e_00061_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00061


title: RR101e_00058_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00058


title: RR101e_00056_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00056


title: RR101e_00053_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00053


title: RR101e_00055_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00055


title: Treg1-Input


title: RR101e_00060_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00060


title: Treg2-IP


title: Treg1-IP


title: RR101e_00051_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00051


title: TGF-3-5-Input


title: RR101e_00046_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00046


title: RR101e_00047_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00047


title: TGF-3-5-IP


title: RR101e_00045_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00045


title: LNA-non-target_1

city: Rochester

first: Thanh Thanh

middle: Le

last: Nguyen

state: Minnesota

address: 200 1st St. SW

institute: Mayo Clinic

data_processing:
Raw read quality was checked with FASTQC and adapter was trimmed with Trimmomatic
Trimmed RNA sequencing reads were aligned to hg38 using STAR
Raw counts were generated with FeatureCount
Genome_build: hg38
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample

source_name: iPSC-derived Astrocytes

growth_protocol: A human induced pluripotent stem cell (hiPSC)-derived astrocyte progenitor kit (cat#: ax0081; discontinued) was purchased from Axol Bioscience Ltd. (Cambridge, United Kingdom), and hiPSC-derived hepatocytes (Cellartis® Enhanced hiPS-HEPv2 from ChiPSC12, cat#: Y10133). Cells were cultured by following the manufacturers’ protocols.

extract_protocol:
RNAeasy mini kit per manufacturer’s instruction (Qiagen)
poly(A) enrichment; Illumina TruSeq RNA Library Prep Kit v2

treatment_protocol: KD experiments were conducted with siRNA and antisense oligos.

value: non-target control  antisense oligo

tag: replicate

value: replicate 1


title: Astrocytes_TCF2

value: TCF7L2 siRNA

value: replicate 2


title: LNA-non-target_2


title: RR101e_00039_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00039


title: RR101e_00030_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00030


title: Astrocytes_SNC2

value: non-target control siRNA


title: LNA_T3KD_2

value: T208 lncRNA antisense oligo


title: TGF-3-20-Input

value: 5-6 weeks

value: Foxp3-eGFP reporter mice

value: iTregs differentiated in the presence of TGFβ


title: RR101e_00029_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00029


title: LNA_T3KD_1


title: Astrocytes_SNC1


title: RR101e_00022_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00022


title: RR101e_00021_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00021


title: RR101e_00017_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00017


title: RR101e_00052_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00052


title: RA-VC-3-5-IP

value: Spleen and lymph nodes

value: iTregs differentiated in the presence of TGFβ+RA+VitC

value: Bisulfite-treated genomic DNA


title: RA-VC-3-5-Input

value: Input for 5hmC-pulldown by CMSIP


title: TCF7L2

description: TCF7L2 antibody

data_processing:
Raw read quality was checked with FASTQC and adapter was trimmed with Trimmomatic
Trimmed DNA sequencing reads were aligned to hg38 using BWA
PCR duplicates were removed with Picard
Peaks were called with MACS2
Genome_build: hg38
Supplementary_files_format_and_content: peak bed files

growth_protocol: A human induced pluripotent stem cell (hiPSC)-derived astrocyte progenitor kit (cat#: ax0081; discontinued) was purchased from Axol Bioscience Ltd. (Cambridge, United Kingdom, cat#: Y10133). Cells were cultured by following the manufacturers’ protocols.

extract_protocol: ThruPLEX® DNA-seq Kit V2

tag: chip antibody


title: RR057i_00185_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057i_00185

tag: gender

value: naso/pharyngeal swab


title: RA-VC-3-20-Input

tag: age


title: RA-VC-3-20-IP

value: 5hmC-enriched by CMSIP


title: RR057e_00924_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00924

value: F

tag: infection


title: monolayer-d19

city: East Lansing

first: Aitor

last: Aguirre

state: MI

address: 775 Woodlot Dr

department: Biomedical Engineering

institute: Michigan State University

description: >200 bp RNA

data_processing:
Qualiy control using FASTQC
alignment was performed with HISAT2
Reads were counted with featurecounts
Differential expression analysis was performed with EdgeR
Genome_build: hg38
Supplementary_files_format_and_content: Excel files containing counts per million

growth_protocol: Specific differentiation protocol conditions can be found in Israeli et al., 2020

extract_protocol:
Qiagen RNAeasy Kit
Libraries were prepared according to Illumina Truseq protocols

treatment_protocol: Each sample represents a mean pool of 3 6-wells of a 6-well plate for monolayer samples, and 5-8 organoids for organoid samples

value: human induced pluripotent stem cells derived from a healthy patient

tag: experimental condition

value: cells differentiated using monolayer differentiation


title: RR057e_00912_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00912

value: M

tag: sample type


title: RR057e_00920_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00920


title: Astrocytes_TCF1

tag: treatment

value: TCF7L2 siRNA

value: replicate 1


title: RR101e_00033_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00033


title: RR057e_00906_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00906


title: TGF-3-20-IP


title: RR057e_00908_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00908


title: RA+vitC.Ad2.9 [ATAC-seq]

data_processing:
Bcl2fastq v2.20.0.442 was used to obtain fastq files from Illumina
ATAC-seq FASTQ files were quality and adapter trimmed using Trim Galore (v0.4.2) prior to mapping to the UCSC mouse genome mm9 using Bowtie2 (v2.2.6) with default parameters.
Mapped SAM files were converted to BAM and putative PCR duplicates were marked with Samtools (v1.7).
Regions of chromatin accessibility were determined using MACS2 (v2.1.0.20151222) with the following settings “-g mm -q 0.01”.
The union of accessible regions in all samples were determined using the ‘GenomicRanges’ package (v1.36.1) in R and data was normalized to reads in peaks per million (RPPM) as previously described.
Genome_build: mm9
Supplementary_files_format_and_content: MACS2_q_0.05.union.annot.clean.cts.gz: Raw counts, each column represent a sample
Supplementary_files_format_and_content: MACS2_q_0.05.union.annot.clean.rppm.cov.gz: RPPM normalized counts, each column represent a sample

extract_protocol:
50,000 cells were sorted and directly used for the library preparation
50,000 cells were washed once with 120 μl ice-cold 1 X PBS and spin down at 500g for 10mins at 4°C. Cell pellets were resuspended in 50 μl cold lysis buffer (10mM Tris-HCl pH7.4, 10mM NaCl, 3mM MgCl2, 0.1% NP-40) and immediately spin down at 500g for 15mins at 4°C. The supernatant was discarded and the nuclei pellets was resuspended in 25 μl of transposition mix (12.5 μl 2 X Nextera Tagment DNA Buffer, 1.25 μl Nextera Tagment DNA Enzyme, 11.25 μl H2O), incubated at 37°C for 30mins. DNA was purified using Zymo DNA Clean and Concentrator Kit (Zymo Research). Samples were then amplified using KAPA HiFi HotStart ReadyMix (KAPA Biosystems) for 11 cycles and purified using Ampure XP Beads (Beckman Coulter).

treatment_protocol: Cells were sorted directly from the mice or differentiated from CD4+ naïve T cells into Th0 cells or iTreg cells under the indicated conditions


title: monolayer-d11

description: >200 bp RNA

tag: cell type


title: RR057e_00898_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00898


title: RR057e_00894_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00894


title: RR057e_00897_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00897


title: monolayer-d9

description: >200 bp RNA


title: monolayer-d15

description: >200 bp RNA


title: RR057e_00893_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00893


title: vitC.Ad2.7 [ATAC-seq]

tag: genotype/variation

value: iTregs differentiated in the presence of TGFβ+VitC


title: RA.Ad2.8 [ATAC-seq]

tag: tissue

value: iTregs differentiated in the presence of TGFβ+RA


title: RR101e_00010_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR101e_00010


title: monolayer-d7

description: >200 bp RNA


title: monolayer-d5

description: >200 bp RNA


title: RR057e_00556_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00556


title: TGF.Ad2.6 [ATAC-seq]

value: 5-6 weeks

tag: sorting/differentiation strategy


title: Input

description: Input


title: Th0.Ad2.5 [ATAC-seq]

value: Foxp3-eGFP reporter mice

value: Th0 cells differentiated with anti-CD3/CD28


title: RR057e_00544_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00544


title: monolayer-d3

description: >200 bp RNA


title: RR057e_00495_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00495


title: RR057e_00518_Peds

description:
all_adult_and_ped_samples_combined_counts_unfiltered.csv
RR057e_00518


title: monolayer-d1

description: >200 bp RNA


title: RA.Ad2.3 [ATAC-seq]

value: Spleen and lymph nodes


title: monolayer-d0

description: >200 bp RNA