characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

characteristic: INSDC center name

text: public

tag: Namespace:Center for Biotechnology, CeBiTec, University of Bielefeld, Universitatsstrasse 27, 33699 Bielefeld, Germany

characteristic: Submitter Id

text: ArrayExpress

characteristic: broker name

text: Protocols: 1 mL culture was added to 250 μL pure ethanol supplemented with 5 % phenol, flash-frozen in liquid nitrogen and stored at −80°C . Synechocystis strains were cultivated in BG11 medium with appropriate antibiotics at 30°C and aeration with 0.5% CO2 in all experiments. For the endpoint measurements cultures were grown in Erlenmeyer flask in an Infors HT multitron chamber, shaken at 150 rpm, and light intensity was 80 umol/m2/sec. For the time series 1 L culture was grown in a Lambda Photobioreactor ca. 90 umol/m2/sec per OD (manually adjusted to OD) with pH kept stable at pH 8. Inducers for gyrase knock-down and topoisomerase I overexpression were added at inoculation after pre-culturing without inducer. For the time series experiment the inducer was added at time point t=0 h, 1 h after SAMPLE_10 was taken. Frozen samples were centrifuged ( 10 min , 4°C , maximum speed). The pellet was resuspended in 1 mL PGTX and incubated ( 5 min , 95°C ). After cooling on ice for 2 min , 700 μL chloroform:isoamyl alcohol (24:1) was added and the mixture incubated (10 min , room temperature) while shaking gently. The mixture was centrifuged (10 min , 4°C , maximal speed). The upper phase was transferred to a fresh tube and 1 volume chloroform:isoamyl alcohol was added. After repeating the centrifugation step, the upper phase was transferred and precipitated with 3 volumes of 99.5 % ethanol and 1/2 volume 7.5 M ammonium acetate and (time series only) 1 μL RNA-grade glycogen at −20°C over night. The RNA was pelleted by centrifugation (30 min , 4°C , maximum speed), washed twice with 70 % ethanol and resuspended in 30 μL RNase-free water. Volumes were adjusted to contain 2 μg of nucleic acid (Nanodrop), and DNA was removed by DNaseI (ThermoFisher: EN0525) according to the manufacturer’s specifications but at 2x reaction buffer concentration. RNA was extracted as above but using 1/10 volume of 3 M sodium acetate (pH 5.3) instead of ammonium acetate for precipitation. The Illumina Ribo-Zero Plus rRNA Depletion Kit was used to remove the ribosomal RNA molecules from the isolated total RNA. Removal of rRNA was evaluated with the RNA Pico 6000 kit on the Agilent 2100 Bioanalyzer. RNA was free of detectable rRNA. Preparation of cDNA libraries was performed according to the manufacturer’s instructions for the TruSeq stranded mRNA kit (Illumina, San Diego, CA, United States). Subsequently,

characteristic: description

text: wild type genotype

characteristic: genotype

text: Synechocystis sp. PCC 6803

text: Synechocystis sp. PCC 6803

text: whole organism

characteristic: organism part

characteristic: replicate

text: Synechocystis sp. PCC 6803

text: wildtype control strain EVC

characteristic: strain

characteristic: title


status: PUBLIC

tag: attribute

text: Center for Biotechnology, CeBiTec, University of Bielefeld, Universitatsstrasse 27, 33699 Bielefeld, Germany

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: ArrayExpress

text: wild type genotype

text: Synechocystis sp. PCC 6803

characteristic: organism

text: Synechocystis sp. PCC 6803

text: whole organism

text: Synechocystis sp. PCC 6803

text: wildtype control strain EVC


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

text: public

text: ArrayExpress

text: Crispr-mediated gyrA knockdown

text: Synechocystis sp. PCC 6803

text: Synechocystis sp. PCC 6803

text: whole organism

text: Synechocystis sp. PCC 6803

text: gyrA-kd


characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: public

text: ArrayExpress

text: Crispr-mediated gyrB knockdown

text: Synechocystis sp. PCC 6803

text: Synechocystis sp. PCC 6803

text: whole organism

text: Synechocystis sp. PCC 6803

text: gyrB-kd


characteristic: INSDC center name

text: public

characteristic: Submitter Id

text: ArrayExpress

text: topA overexpression

text: Synechocystis sp. PCC 6803

text: Synechocystis sp. PCC 6803

text: whole organism

text: Synechocystis sp. PCC 6803

text: topA-OX


text: public

text: ArrayExpress

characteristic: broker name

characteristic: description

text: topA overexpression

text: Synechocystis sp. PCC 6803

text: Synechocystis sp. PCC 6803

text: whole organism

text: Synechocystis sp. PCC 6803

text: topA-OX


Costus vinosus R2952

text: Cornell University

text: public

text: Costus vinosus R2952

tag: Namespace:Cornell University

text: Dave Skinner

characteristic: collected by

text: Cornell University

characteristic: collecting institution

characteristic: collection date

text: freshly collected leaf material

text: Panama

characteristic: geographic location (country and/or sea)

unit: DD

characteristic: geographic location (latitude)

characteristic: geographic location (longitude)

text: Rio Guanche

characteristic: geographic location (region and locality)

text: neotropical forest

characteristic: habitat

text: vegetative structure

characteristic: lifestage

text: Costus vinosus

text: leaf

characteristic: organism part

text: ORG.one

characteristic: project name

text: Costus vinosus

text: hermaphroditic

characteristic: sex

characteristic: specimen voucher

text: leaf material Costsus vinosus


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-86241632152501:Negative_control_rep1

tag: Namespace:European Research Institute for the Biology of Ageing, University of Groningen, UMC Groningen

text: ArrayExpress

text: hTERT RPE-1

characteristic: cell line

text: epithelial cell

characteristic: cell type

text: human

characteristic: common name

text: Protocols: Cell culture hTERT immortalized human retinal pigmented epithelial cells (hTERT RPE-1, ATCC ®, CRL-4000™) were grown in T175 flasks at 37 °C with 5 % CO2 with DMEM, high glucose, GlutaMax™, pyruvate (Gibco) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin (10,000 units penicillin and 10 mg streptomycin/mL, Gibco). Cells were passaged with Trypsin-EDTA at 70-80% confluency and used for a maximum of 20 passages. Cells were regularly tested for mycoplasma infection. For Double-Click-seq experiments DMEM containing azidohomoalanine (AHA) and 5-ethynyl-2'-deoxyuridine (EdU) was used. This 100% AHA medium was made as follows: DMEM, high glucose, no glutamine, no methionine, no cysteine (Gibco) was supplemented with 10% fetal bovine serum, 1% penicillin-streptomycin (10,000 units penicillin and 10 mg streptomycin/mL, Gibco), 4 mM GlutaMax™ (Gibco), 0.2 mM L-Cystine 2HCl, 1 mM sodium pyruvate, 0.2 mM azidohomoalanine (AHA) and 20 µM 5-ethynyl-2'-deoxyuridine (EdU). Double-click-seq Incubation and harvest of cells. Five T175 flask were each seeded with approximately 3 x 106 cells per experiment. After overnight incubation, culture medium was removed and replaced by pre-mixed medium containing 0.95 x volume 100% AHA medium (see recipe above) and 0.05 x volume of DMEM (see recipe above). For induction of replication stress either 100 nM curaxin (CBL0137), 150 or 200 µM hydroxyurea (HU), 0.5 µM ATR inhibitor (VE-821, Axon Medchem) or a combination was added to the culture medium. Cells were incubated for 24 h, after which the culture medium was removed and cells were washed twice with ice-cold Phosphate-Buffered Saline (PBS). Cells were harvested with Trypsin-EDTA (Gibco) for 5 minutes at 37 °C. Trypsinization was quenched by the addition of medium and cells were collected by centrifugation at 200 g for 5 minutes. The pellet was resuspended in ice-cold PBS and centrifuged again at 200 g for 5 minutes. The cell pellet was stored at -80 °C until further use. Nuclei isolation. The cell pellet was resuspended in 1 mL of ice-cold PBS containing 0.1% NP-40 supplemented with proteinase inhibitor cocktail (PIC) (cOmplete™, Mini, EDTA-free, Roche). The cell suspension was vortexed for five seconds at maximum speed. The lysed cell suspension was centrifuged for 10 seconds at 9000 g. The supernatant was discarded and the nuclei were resuspended in 1 mL of ice-cold PBS containing 0.1% NP-40 supplemented with PIC. The washed nuclei were centrifuged for 10 seconds at 9000 g and the supernatant was discarded. First click reaction. The nuclei were resuspended in 250 µL nuclei buffer (320 mM sucrose, 5 mM MgCl2, 10 mM HEPES at pH 7.4) supplemented with PIC and 10 µL 100 mM (in methanol) biotin-PEG4-alkyne (Merck) was added. Next, a pre-mixed solution of 5 µL 50 mM CuSO4 with 5 µL 250 mM THPTA (Merck) was added, followed by 10 µL of a freshly prepared solution of 100 mM sodium ascorbate. The nuclei were placed in an end-over-end rotator at room temperature for 30 minutes. After that, the click reaction was repeated as described above using fresh reagents. Nuclei were pelleted for 10 seconds at 9000 g (to remove excess biotin, which would interfere with streptavidin bead pull-down later on) and the supernatant was discarded. MNAse digestion. Nuclei were resuspended in 100 µL of MNAse buffer (50 mM Tris.HCl, 5 mM CaCl2, pH 7.9, supplemented with 100 µg/mL BSA) and warmed to 37 °C in a water bath. Then, 60 U micrococcal nuclease (New England Biolabs, MNase, 1U/µL in MNAse buffer) was added and cells were incubated at 37 °C for 5 minutes. MNase reactions were stopped by adding EDTA to a final concentration of 5 mM and by cooling on ice. Next, 100 µL PBS supplemented with 0.1% BSA and 0.01% Tween was added and the nuclei were pelleted by centrifugation for 10 seconds at 9000 g. Streptavidin enrichment of labeled nucleosomes. The supernatant (containing the nucleosomes) was added to 30 µL streptavidin magnetic beads (Dynabeads™ MyOne™ Streptavidin C1, Thermo Scientific, pre-washed three times with PBS according to the instruction manual) and incubated at room temperature for 1 hour in an end-to-end rotator. The supernatant was removed by placing it on a magnet for 3-4 minutes. The beads were washed five times by resuspending them in 200 µL PBS supplemented with 0.1% BSA and 0.01% Tween and placing them back on the magnet. After washing, the DNA bound to the captured nucleosomes was released. This was done by resuspending the beads in 100 µL PBS supplemented with 1 mg/mL proteinase K and incubating the suspension for 30 minutes at 50 °C (pipet up and down after 15 minutes to ensure a homogenous mixture). Then, 100 µL 5 M NaCl was added and the mixture was left rotating for 15 minutes at room temperature. Finally, 360 µL (1.8 x volume of sample) Ampure beads (Beckman) was added directly to the suspension to purify the released DNA according to the protocol of the manufacturer. The DNA was eluted in 80 µL water. Second click reaction. 1 µL 100 mM (in methanol) biotin-PEG4-azide was added to the eluted DNA, followed by a pre-mixed solution of 5 µL 50 mM CuSO4 with 5 µL 250 mM THPTA and 10 µL of a freshly prepared solution of 100 mM sodium ascorbate. The sample was placed in an end-over-end rotator at room temperature for 45 minutes. To discard DNA fragments larger than approximately 200 bp 70 µL (0.7 x volume of sample) Ampure beads were added and after 5 minutes incubation at room temperature the beads were discarded and the supernatant was transferred to a new tube. Then 180 µL (1.8 x original sample volume) Ampure beads were added and purification was completed according to the manufacturer’s protocol. The DNA was eluted in 55.5 µL water. End prep and adaptor ligation. End repair, 5’ phosphorylation, dA-tailing and adaptor ligation was performed using the NEBNext® Ultra™ DNA library prep kit for Illumina®. After adaptor ligation the sample was added to 25 µL streptavidin magnetic beads (Dynabeads™ MyOne™ Streptavidin T1, Thermo, pre-washed three times with B&W buffer (5 mM Tris HCl pH 7.5, 0.5 mM EDTA, 1 M NaCl ) according to the instruction manual). Isolation of replicated double stranded DNA fragments. After 15 minutes incubation at room temperature in an end-over-end rotator, beads were washed four times with 200 μl 1x B&WT buffer (5 mM Tris HCl pH 7.5, 0.5 mM EDTA, 1 M NaCl, 0.05 % Tween 20) and once with 2 x B&WT buffer. Isolation of replicated parental strands. The parental strands were then eluted from the beads by incubation with 100 μl alkaline buffer (0.1 M NaOH, 0,05 % Tween 20) for 1 minute. Elution of the parental strand was repeated twice for a total of three times and the pH of the combined alkaline supernatants was neutralized by adding acetic acid to a final concentration of 0.1 M and 2 mM EDTA pH 8.0. The sample was diluted to 1 mL with water and concentrated to approximately 20 uL with a Vivaspin® 2 5K according to the manufacturer’s instruction (15 minutes centrifuging at 4000 g in a swing bucket). The DNA was recovered by performing a reverse spin at 3000 g for 2 minutes. Library amplification and sequencing. Libraries were amplified using the NEBNext® Ultra™ DNA library prep kit for Illumina® using NEBNext® multiplex oligos for Illumina® (index primer set 1 or 2) with 13 cycles of PCR. Purification was done as described in the manual (with 0.9 x Ampure beads) and eluted in 30 μl water. Libraries were quantified with Qubit™ dsDNA high sensitivity assay kit (Thermo) and further analyzed with the high sensitivity D1000 ScreenTape system (Agilent).

text: adult

characteristic: developmental stage

text: normal

characteristic: disease

text: wild type genotype

characteristic: genotype

text: Homo sapiens

characteristic: organism

text: Homo sapiens

text: retinal pigmented epithelium

text: Homo sapiens

characteristic: title


characteristic: ENA-CHECKLIST

text: UNIVERSITY OF CAMBRIDGE

text: public

tag: Namespace:UNIVERSITY OF CAMBRIDGE

characteristic: Submitter Id

text: embryonic day 6.5

unit: day

characteristic: age

text: ArrayExpress

characteristic: broker name

text: pluripotent stem cell

text: rabbit

text: Protocols: Rabbit embryos at E6-7 were trypsinized with 0.25% trypsin/EDTA for 5min. Then, single cells were sorted by SH800 into wells of 384 well plates filled with 2.875 ul of lysis buffer (Takara Bio Inc.). We had also included additional samples; cells from posterior epiblast at E7 containing rbPGCs, in vitro rbPSCs, rbPSCs cultured for 2 days with lower concentration (5ng/ml) of FGF (termed as rbPSC_F5), and NT positive d2 rbPGCLCs. To enrich rbPGCs, 3-4 posterior epiblast dissected from E7 (st.3-4) rabbit embryos were dissociated into single cell by TrypLE Express (Thermo Fisher Scientific) at 37℃ for 8 min. After inactivation of the TrypLE Express, dissociated cells were incubated with anti-c-KIT antibody labeled by Biotin Labeling Kit (Dojindo), and subsequently incubated with Alexa Fluor® 647 Streptavidin (Biolegend, San Diego, CA). For rbPSC rbPSC_F5 and rbPGCLC samples, cells dissociated with 0.25% trypsin/EDTA (Thermo Fisher Scientific) were incubated with anti-feeder cell antibody (Miltenyi Biotec) for 15 min at 4℃ to exclude residual MEF by FACS. To prepare rbPSCs (rbKN3) the epiblast was isolated from the bilaminar disc-embryo at E6 for seeding on feeder cells. Epiblast cells proliferated uniformly as a homogeneous epithelial layer. After passaging, the cells maintained constant monolayer colonies without detectable signs of differentiation and continued to proliferate robustly while retaining a normal karyotype in most cell lines. PSCs were cultured on a medium consisting of Essential 8, supplemented with two WNT inhibitors, XAV939 and IWP-2. PSC_low cell were cultured in reduced FGF (5ng/ml). cells were directly sorted into PCR tubes filled with 10 ul of the lysis buffer by SH800 or MA900 cell sorter. Single-cell transcriptomes of sorted cells were assayed as follows: for E6-7 embryos, reverse transcription and cDNA amplification was performed using the miniaturized version of the SMART-seq HT protocol using the low-volume liquid-handling robot Mosquito HV (SPT Lab Tech). For additional samples, reverse transcription and cDNA amplification was performed according to manufacturer’s protocol of Smart-seq HT kit (Takara Bio Inc.). Libraries were prepared using the Illumina Nextera XT DNA preparation kit.

characteristic: description

text: blastocyst

text: normal

text: wild type genotype

text: cell culture

characteristic: growth condition

text: Oryctolagus cuniculus

text: Oryctolagus cuniculus

text: embryo

characteristic: organism part

text: OK

characteristic: post analysis well quality

text: Oryctolagus cuniculus

text: pass

characteristic: single cell well quality


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: UNIVERSITY OF CAMBRIDGE

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: embryonic day 6.5

text: ArrayExpress

characteristic: cell line

text: pluripotent stem cell

text: rabbit

text: blastocyst

text: normal

text: low

characteristic: fraction

text: wild type genotype

text: cell culture

text: Oryctolagus cuniculus

text: Oryctolagus cuniculus

text: embryo

text: OK

text: Oryctolagus cuniculus

text: pass


characteristic: ENA-CHECKLIST

text: UNIVERSITY OF CAMBRIDGE

text: public

characteristic: Submitter Id

text: embryonic day 7

characteristic: age

text: ArrayExpress

text: primordial germ cell

characteristic: cell type

text: rabbit

characteristic: common name

text: blastocyst

characteristic: developmental stage

text: normal

characteristic: disease

text: PGCLC

text: wild type genotype

characteristic: genotype

text: cell culture

text: Oryctolagus cuniculus

characteristic: organism

text: Oryctolagus cuniculus

text: epiblast

text: OK

text: posterior epiblast

characteristic: sampling site

text: Oryctolagus cuniculus

text: pass

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: UNIVERSITY OF CAMBRIDGE

characteristic: INSDC center name

characteristic: INSDC first public

text: public

text: embryonic day 4

text: ArrayExpress

characteristic: broker name

text: pluripotent stem cell

text: rabbit

text: Protocols: Rabbit embryos at E6-7 were trypsinized with 0.25% trypsin/EDTA for 5min. Then, single cells were sorted by SH800 into wells of 384 well plates filled with 2.875 ul of lysis buffer (Takara Bio Inc.). We had also included additional samples; cells from posterior epiblast at E7 containing rbPGCs, in vitro rbPSCs, rbPSCs cultured for 2 days with lower concentration (5ng/ml) of FGF (termed as rbPSC_F5), and NT positive d2 rbPGCLCs. To enrich rbPGCs, 3-4 posterior epiblast dissected from E7 (st.3-4) rabbit embryos were dissociated into single cell by TrypLE Express (Thermo Fisher Scientific) at 37℃ for 8 min. After inactivation of the TrypLE Express, dissociated cells were incubated with anti-c-KIT antibody labeled by Biotin Labeling Kit (Dojindo), and subsequently incubated with Alexa Fluor® 647 Streptavidin (Biolegend, San Diego, CA). For rbPSC rbPSC_F5 and rbPGCLC samples, cells dissociated with 0.25% trypsin/EDTA (Thermo Fisher Scientific) were incubated with anti-feeder cell antibody (Miltenyi Biotec) for 15 min at 4℃ to exclude residual MEF by FACS. PSCs were maintained on mitomycin-C-treated mouse embryonic fibroblasts (MEF) in Essential 8 medium (Thermo Fisher Scientific) containing 100 nM IWP2 (Toc ris Bioscience, Bristol, UK) at 38.5 °C and 5% CO2. trypsinized rbPSCs were seeded on vitronectin-coated dish at 200,000 cells/well in 12 well plates and cultured in ME induction medium for 24 h or 6-30 h as indicated in the figures. ME induction medium was composed of aFB27 medium (Advanced DMEM/F12 medium (Thermo Fisher Scientific) supplemented with 1% B27 supplement (Thermo Fisher Scientific), 0.1 mM NEAA, 100 U/ml Penicillin- 0.1 mg/ml Streptomycin, 2 mM L-Glutamine and 0.1% PVA) supplemented with 100 ng/ml Activin-A (Peprotech, Cranbury, NJ), 3 uM GSK3i (CHIR99021; Axon Medchem, Reston, VA), 50 nM PI3Ki (PI103; Tocris Bioscience) and 10 uM of ROCKi. rbPSCs trypsinized into single cells and harvested into EZSPHERE®SP microwell 96 well plate (IWAKI) at 2x10e4 cells/well in aFB27 medium containing 100 ng/ml BMP4 (Peprotech), 10 ng/ml human LIF (Peprotech), 100 ng/ml mouse SCF (R&D Systems), 50 ng/ml human EGF (R&D Systems), and 10 uM ROCKi. ME induction medium was replaced with DE induction medium after washing with PBS once and cells were cultured for additional 2 days. DE induction medium was composed of aFB27 medium supplemented with 100 ng/ml Activin-A, 0.5 uM BMPi (LDN193189; Tocris Bioscience) and 2.5 uM IWR1 (Tocris Bioscience). cells were directly sorted into PCR tubes filled with 10 ul of the lysis buffer by SH800 or MA900 cell sorter. Single-cell transcriptomes of sorted cells were assayed as follows: for E6-7 embryos, reverse transcription and cDNA amplification was performed using the miniaturized version of the SMART-seq HT protocol using the low-volume liquid-handling robot Mosquito HV (SPT Lab Tech). For additional samples, reverse transcription and cDNA amplification was performed according to manufacturer’s protocol of Smart-seq HT kit (Takara Bio Inc.). Libraries were prepared using the Illumina Nextera XT DNA preparation kit.

characteristic: description

text: embryo

text: normal

text: wild type genotype

text: 100 nM IWP2

characteristic: growth condition

text: Oryctolagus cuniculus

text: Oryctolagus cuniculus

text: embryo

characteristic: organism part

text: embryonic stem cell

characteristic: progenitor cell type

text: Oryctolagus cuniculus


characteristic: ENA-CHECKLIST

text: UNIVERSITY OF CAMBRIDGE

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: embryonic day 6

characteristic: age

text: ArrayExpress

characteristic: cell line

text: pluripotent stem cell

characteristic: cell type

text: rabbit

characteristic: common name

text: embryo

text: normal

text: wild type genotype

text: 100 nM IWP2

text: Oryctolagus cuniculus

text: Oryctolagus cuniculus

text: epiblast

text: epiblast stem cell

text: Oryctolagus cuniculus


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: UNIVERSITY OF CAMBRIDGE

text: public

text: embryonic day 4

text: ArrayExpress

text: mesondoderm like cell

text: rabbit

text: embryo

characteristic: developmental stage

text: normal

characteristic: disease

text: wild type genotype

characteristic: genotype

text: ME induction medium

text: Oryctolagus cuniculus

characteristic: organism

text: Oryctolagus cuniculus

text: embryo

text: pluripotent stem cell

text: Oryctolagus cuniculus

characteristic: title


characteristic: ENA-CHECKLIST

text: UNIVERSITY OF CAMBRIDGE

characteristic: INSDC center name

characteristic: INSDC first public

text: public

text: embryonic day 4

text: ArrayExpress

characteristic: broker name

text: primordial germ cell like cell

text: rabbit

characteristic: description

text: embryo

text: normal

text: wild type genotype

text: aFB27 medium containing 100 ng/ml BMP4 (Peprotech), 10 ng/ml human LIF (Peprotech), 100 ng/ml mouse SCF (R&D Systems), 50 ng/ml human EGF (R&D Systems), and 10 uM ROCKi

characteristic: growth condition

text: Oryctolagus cuniculus

text: Oryctolagus cuniculus

text: embryo

characteristic: organism part

text: pluripotent stem cell

characteristic: progenitor cell type

text: Oryctolagus cuniculus


text: UNIVERSITY OF CAMBRIDGE

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: embryonic day 4

characteristic: age

text: ArrayExpress

characteristic: cell line

text: definitive endoderm like cell

characteristic: cell type

text: rabbit

characteristic: common name

text: embryo

text: normal

text: wild type genotype

text: DE induction medium

text: Oryctolagus cuniculus

text: Oryctolagus cuniculus

text: embryo

text: pluripotent stem cell

text: Oryctolagus cuniculus


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: UNIVERSITY OF CAMBRIDGE

text: public

text: embryonic day 4

text: ArrayExpress

text: somatic cell

text: rabbit

text: epiblast

characteristic: developmental stage

text: normal

characteristic: disease

text: wild type genotype

characteristic: genotype

text: Oryctolagus cuniculus

characteristic: organism

text: Oryctolagus cuniculus

text: epiblast

text: epiblast stem cell

text: Oryctolagus cuniculus

characteristic: title


E-MTAB-10961:Sample 1

characteristic: ENA-CHECKLIST

text: Xi'an Jiaotong University

characteristic: INSDC center name

characteristic: INSDC first public

text: public

text: E-MTAB-10961:Sample 1

tag: Namespace:Xi'an Jiaotong University

unit: year

text: ArrayExpress

characteristic: broker name

text: human

text: Protocols: From September to December 2018, five children with POTS and five healthy children were recruited from the outpatient pediatric department of the Children’s hospital of Zhejiang University School of Medicine, China. All five POTS children underwent a thorough medical history evaluation, physical examination, and laboratory assessment, including electrocardiography (ECG), electroencephalography, blood glucose and blood biochemistry analyses, and cranial computed tomography or magnetic resonance imaging to exclude cardiac, neurologic, metabolic, and psychogenic causes. Five healthy children, matched as far as possible in terms of age, gender, height, weight, and residence location, were recruited as control subjects. The healthy controls were screened by medical history assessment, physical examination, and laboratory assessment, including ECG, Holter ECG, and standing tests. These 10 children were used as sequencing groups. Another 40 POTS patients and 20 healthy children from Shenmu County Hospital and Shaanxi Provincal People’s Hospital were used as a validation subjects. All children were informed about the purposes of the research and agreed to participate. Written informed consent was obtained from the parents or guardians of all the study subjects. The study was approved by the ethics committees of the Children’s Hospital of Zhejiang University School of Medicine, Shenmu County Hospital, and Shaanxi Provincial People’s Hospital. Blood sample collection for whole-blood miRNA sequencing POTS patients and healthy children were required to fast before blood sampling. Blood samples (5 mL) were drawn at 8:00 am from supine subjects into PAXgene Blood RNA Tubes (BD). The tubes were inverted 8-10 times gently immediately after blood collection. Each tube contained 2.5 mL of whole blood and 7 mL of additives. The blood samples were allowed to stand at room temperature for 4 h and then stored at -80 ℃. Total RNA extraction We extracted RNA from the PAXgene Blood RNA tubes and performed quality control. All the quality control results were analyzed centrally at the end of 2018. All the samples were thawed at room temperature, agitated, and observed to determine the presence of any blood clots in the tubes. The PAXgene Blood RNA kit and a QIA cube were used to extract RNA from the samples automatically. RNA was eluted with elution buffer and stored at -80 °C. A total of 2 μg of RNA/sample was used as input material for the miRNA library. The miRNA sequencing libraries were generated using a TruSeq Small RNA Sample Preparation Kit (Illumina, Inc., San Diego, CA, USA), and index codes were added to attribute sequences to each sample. Briefly, an RNA 3′ adaptor was added to the total RNA and the mixture was incubated at 70 °C for 2 min to break the RNA secondary structure and then immediately placed on ice. Then, the RNA 3′ adaptor was specifically ligated to the 3′ end of the miRNA with T4 RNA Ligase 2 and T4 Rnl2 truncated at 28 °C for 1 h. After the 3′ ligation reaction, the ligated fragment was further ligated to an RNA 5′ adapter using T4RNA ligase at 28 °C for 1 h. The first strand cDNA was then synthesized using SuperScript IV (SS IV) according to the manufacturer’s instructions. Polymerase chain reaction (PCR) amplification was performed using 5× Phusion HF buffer, RNA PCR Primer (RP1), RNA PCR Primer Index (RPLX) primer, dNTP, and Phusion DNA polymerase. The PCR products were purified on a 6% polyacrylamide gel. RNA fragments corresponding to 140-160 bp (the length of small noncoding RNA with the 3′ and 5′ adaptors) were recovered and dissolved in 8 μL of elution buffer. Finally, the library concentration was measured using a Qubit Spectrophotometer and the library quality was assessed using an Agilent 2100 Bioanalyzer system with a High Sensitivity DNA Kit (Agilent Technologies). The clustering of the index-coded samples was performed on an Illumina cBot cluster station using a TruSeq PE Cluster Kit v3-cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the library preparations were sequenced on an Illumina HiSeq 2000 platform and 50 bp single-end reads were generated for transcriptome analysis. The quality control of the samples, experiments, and data analysis were completed at Genesky Biotechnologies Inc. (Shanghai, People’s Republic of China).

characteristic: description

text: juvenile stage

text: postural orthostatic tachycardia syndrome

characteristic: individual

text: Homo sapiens

text: Homo sapiens

text: blood

characteristic: organism part

text: Homo sapiens

text: male

characteristic: sex

text: Sample 1


E-MTAB-10961:Sample 2

text: Xi'an Jiaotong University

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10961:Sample 2

characteristic: Submitter Id

characteristic: age

text: ArrayExpress

text: human

characteristic: common name

text: juvenile stage

text: Homo sapiens

text: Homo sapiens

text: blood

text: Homo sapiens

text: male

text: Sample 2


E-MTAB-10961:Sample 3

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: Xi'an Jiaotong University

text: public

text: E-MTAB-10961:Sample 3

text: ArrayExpress

text: human

text: juvenile stage

characteristic: developmental stage

characteristic: disease

text: Homo sapiens

text: Homo sapiens

text: blood

text: Homo sapiens

text: male

text: Sample 3


E-MTAB-10961:Sample 4

characteristic: ENA-CHECKLIST

text: Xi'an Jiaotong University

characteristic: INSDC center name

characteristic: INSDC first public

text: public

text: E-MTAB-10961:Sample 4

text: ArrayExpress

text: human

text: juvenile stage

text: Homo sapiens

characteristic: organism

text: Homo sapiens

text: blood

text: Homo sapiens

text: female

text: Sample 4

characteristic: title


E-MTAB-10961:Sample 5

text: Xi'an Jiaotong University

text: public

text: E-MTAB-10961:Sample 5

text: ArrayExpress

characteristic: broker name

text: human

characteristic: description

text: juvenile stage

characteristic: individual

text: Homo sapiens

text: Homo sapiens

text: blood

characteristic: organism part

text: Homo sapiens

text: female

characteristic: sex

text: Sample 5


E-MTAB-10961:Sample 6

text: Xi'an Jiaotong University

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10961:Sample 6

characteristic: Submitter Id

characteristic: age

text: ArrayExpress

text: human

text: juvenile stage

text: normal

text: Homo sapiens

text: Homo sapiens

text: blood

text: Homo sapiens

text: male

text: Sample 6


E-MTAB-10961:Sample 7

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: Xi'an Jiaotong University

text: public

text: E-MTAB-10961:Sample 7

text: ArrayExpress

text: human

characteristic: common name

text: juvenile stage

characteristic: developmental stage

text: normal

text: Homo sapiens

text: Homo sapiens

text: blood

text: Homo sapiens

text: male

text: Sample 7


E-MTAB-10961:Sample 8

characteristic: ENA-CHECKLIST

text: Xi'an Jiaotong University

characteristic: INSDC center name

characteristic: INSDC first public

text: public

text: E-MTAB-10961:Sample 8

text: ArrayExpress

text: human

text: juvenile stage

text: normal

characteristic: disease

text: Homo sapiens

characteristic: organism

text: Homo sapiens

text: blood

text: Homo sapiens

text: male

text: Sample 8


E-MTAB-10961:Sample 9

text: Xi'an Jiaotong University

text: public

text: E-MTAB-10961:Sample 9

text: ArrayExpress

characteristic: broker name

text: human

characteristic: description

text: juvenile stage

text: normal

characteristic: individual

text: Homo sapiens

text: Homo sapiens

text: blood

characteristic: organism part

text: Homo sapiens

text: female

text: Sample 9

characteristic: title


E-MTAB-10961:Sample 10

text: Xi'an Jiaotong University

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10961:Sample 10

characteristic: Submitter Id

text: ArrayExpress

text: human

text: juvenile stage

text: normal

text: Homo sapiens

text: Homo sapiens

text: blood

text: Homo sapiens

text: female

characteristic: sex

text: Sample 10


E-MTAB-10963:Col-0 SA rep_a

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: public

text: E-MTAB-10963:Col-0 SA rep_a

tag: Namespace:Institute of Molecular Plant Sciences School of Biological Sciences University of Edinburgh

characteristic: age

text: ArrayExpress

text: thale cress

characteristic: common name

text: Protocols: Leaves were collected and placed into a 2 ml screw-cap tube and immediately frozen in liquid nitrogen before storing at -80 degrees celcius. Seeds were germinated on soil in 100% relative humidity. After 10 days seedlings were transplanted to larger pots (6 plants per pot) and grown for an additional 18 days. Plants were watered as required. Growth conditions: 16/8 hours light/dark cycle (120 mol m-2 s-1), 21/18 degrees Celsius day/night cycle, 65% relative humidity. Plants were sprayed with a spray bottle that produced fine mist. Leaves were sprayed with water or 0.5 mM SA until thoroughly covered in fine droplets. After 24 hours leaf tissue was harvested for analysis. 1) Grind tissue very fine in liquid nitrogen. 2) Add 0.5 ml RNA extraction buffer (warmed to ~80ºC) and 0.5 ml phenol:chloroform:isoamylalcohol mixture (25:24:1). Vortex vigorously. 3) Centrifuge max. speed for 5 min. at 4ºC, but do not put samples on ice. 4) Transfer the aqueous phase (upper phase) to a new tube. 5) Add 0.5 ml cold chloroform:isoamylalcohol (24:1) and vortex vigorously. 6) Centrifuge max. speed for 5 min. at 4ºC, but do not put samples on ice. 7) Repeat steps 4, 5, and 6. 8)Transfer the aqueous phase to a new tube on ice and add 1/3 volume of 8 M LiCl. Optional: If protocol was started with small amounts of tissue also add 25 µg glycogen (5 µl of 5 mg/ml stock) to visualize RNA pellet in step 10. 9) Incubate overnight at 4ºC. 10) Centrifuge max. speed for 15 min. at 4ºC. A small clear pellet of total RNA should be visible. From now on keep tubes on ice at all times. 11) Without disturbing the pellet, very carefully remove the supernatant with a pipette. 12) Wash RNA pellet gently with ice-cold (-20ºC) 0.5 ml of 70% ethanol. Do not resuspend pellet. 13) Centrifuge max. speed for 1 min. at 4ºC to make sure the RNA pellet stays in place.Discard supernatant without disturbing the RNA pellet. 14) Repeat step 13 to remove any remaining 70% ethanol. 15) Dry RNA pellet in the SpeedVac for 1 min. without heating. Do not dry longer than this!!! 16) Add 400 μl ddH2O (autoclaved) and keep on ice for 30-60 min. to rehydrate the RNA pellet. 17) Dissolve the RNA pellet by vigorously pipetting the solution up and down. 18) Add 40 μl NaAc pH5.2 and 1 ml of ice-cold (-20ºC) 96% ethanol. Invert to mix (do not vortex!!!). 19) Incubate at least 60 min. at -20ºC. 20) Centrifuge max. speed for 15 min. at 4ºC. A gel-like pellet of pure RNA should now be visible. 21) Without disturbing the pellet, very carefully remove the supernatant with a pipette. 22) Wash RNA pellet gently with ice-cold (-20ºC) 0.5 ml of 70% ethanol. Do not resuspend pellet!!! 23) Centrifuge max. speed for 1 min. at 4ºC to make sure the RNA pellet stays in place. Discard supernatant without disturbing the RNA pellet. 24) Repeat step 23 to remove any remaining 70% ethanol. 25) Dry RNA pellet in the SpeedVac for 1 min. without heating. Do not dry longer than this! 26) Add 25 μl ddH2O (autoclaved) and let sit on ice for 30 min. to rehydrate the RNA pellet. 27) Resuspend RNA pellet by pipetting up and down. 28) RNA was then purified further using an RNeasy Mini Kit (Qiagen) according to the manufacturer’s instructions; Buffers & Solutions: RNA extraction buffer (autoclave) consists of 100 mM Tris-HCl, pH 8.0, 100 mM LiCl, 10 mM EDTA, 1% SDS; Phenol:chloroform:isoamyl alcohol (25:24:1); Chloroform:isoamylalcohol (24:1); 8M LiCl (autoclave); 3M NaAC, pH 5.2 (autoclave); 70% Ethanol; 96% Ethanol (pure ethanol) Strand-specific RNA libraries were constructed by GATC/Eurofins, the protocol of which is proprietary.

characteristic: ecotype

text: wild type genotype

characteristic: genotype

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: Col-0 SA rep_a


E-MTAB-10963:Col-0 SA rep_b

characteristic: ENA-CHECKLIST

text: Institute of Molecular Plant Sciences School of Biological Sciences University of Edinburgh

characteristic: INSDC center name

characteristic: INSDC first public

text: public

text: E-MTAB-10963:Col-0 SA rep_b

text: ArrayExpress

characteristic: broker name

text: thale cress

text: wild type genotype

text: Arabidopsis thaliana

characteristic: organism

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: Col-0 SA rep_b


E-MTAB-10963:npr1-1 SA rep_a

characteristic: INSDC last update

text: public

text: E-MTAB-10963:npr1-1 SA rep_a

text: ArrayExpress

text: thale cress

characteristic: description

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

characteristic: organism part

text: Arabidopsis thaliana

text: npr1-1 SA rep_a

characteristic: title


E-MTAB-10963:npr1-1 SA rep_b

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10963:npr1-1 SA rep_b

characteristic: Submitter Id

text: ArrayExpress

text: thale cress

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: npr1-1 SA rep_b


E-MTAB-10963:upl1 SA rep_a

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: public

text: E-MTAB-10963:upl1 SA rep_a

text: ArrayExpress

text: thale cress

text: upl1 (SALK_063972)

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: upl1 SA rep_a


E-MTAB-10963:upl1 SA rep_b

text: public

text: E-MTAB-10963:upl1 SA rep_b

characteristic: age

text: ArrayExpress

text: thale cress

characteristic: common name

characteristic: ecotype

text: upl1 (SALK_063972)

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: upl1 SA rep_b


E-MTAB-10963:upl5 SA rep_a

characteristic: ENA-CHECKLIST

characteristic: INSDC center name

text: public

text: E-MTAB-10963:upl5 SA rep_a

text: ArrayExpress

text: thale cress

text: upl5 (SALK_116446)

characteristic: genotype

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: upl5 SA rep_a


E-MTAB-10963:upl5 SA rep_b

characteristic: INSDC first public

text: public

text: E-MTAB-10963:upl5 SA rep_b

text: ArrayExpress

characteristic: broker name

text: thale cress

text: upl5 (SALK_116446)

text: Arabidopsis thaliana

characteristic: organism

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: upl5 SA rep_b


E-MTAB-10963:Col-0 H2O rep_a

characteristic: INSDC last update

text: public

text: E-MTAB-10963:Col-0 H2O rep_a

text: ArrayExpress

text: thale cress

characteristic: description

text: wild type genotype

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

characteristic: organism part

text: Arabidopsis thaliana

text: Col-0 H2O rep_a


E-MTAB-10963:Col-0 H2O rep_b

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10963:Col-0 H2O rep_b

text: ArrayExpress

text: thale cress

text: wild type genotype

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: Col-0 H2O rep_b

characteristic: title


E-MTAB-10963:npr1-1 H2O rep_a

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: public

text: E-MTAB-10963:npr1-1 H2O rep_a

characteristic: Submitter Id

text: ArrayExpress

text: thale cress

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: npr1-1 H2O rep_a


E-MTAB-10963:npr1-1 H2O rep_b

characteristic: External Id

text: public

text: E-MTAB-10963:npr1-1 H2O rep_b

characteristic: age

text: ArrayExpress

text: thale cress

characteristic: common name

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: npr1-1 H2O rep_b


E-MTAB-10963:upl1 H2O rep_a

characteristic: ENA-CHECKLIST

characteristic: INSDC center name

text: public

text: E-MTAB-10963:upl1 H2O rep_a

text: ArrayExpress

text: thale cress

characteristic: ecotype

text: upl1 (SALK_063972)

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: upl1 H2O rep_a


E-MTAB-10963:upl1 H2O rep_b

characteristic: INSDC first public

text: public

text: E-MTAB-10963:upl1 H2O rep_b

text: ArrayExpress

characteristic: broker name

text: thale cress

text: upl1 (SALK_063972)

characteristic: genotype

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: upl1 H2O rep_b


E-MTAB-10963:upl5 H2O rep_a

characteristic: INSDC last update

text: public

text: E-MTAB-10963:upl5 H2O rep_a

text: ArrayExpress

text: thale cress

characteristic: description

text: upl5 (SALK_116446)

text: Arabidopsis thaliana

characteristic: organism

text: Arabidopsis thaliana

text: leaf

text: Arabidopsis thaliana

text: upl5 H2O rep_a


E-MTAB-10963:upl5 H2O rep_b

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10963:upl5 H2O rep_b

text: ArrayExpress

text: thale cress

text: upl5 (SALK_116446)

text: Arabidopsis thaliana

text: Arabidopsis thaliana

text: leaf

characteristic: organism part

text: Arabidopsis thaliana

text: upl5 H2O rep_b


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: The Weizmann Institute of Science

text: public

tag: Namespace:The Weizmann Institute of Science

text: 8 to 12

unit: week

text: ArrayExpress

text: house mouse

text: Protocols: Colons were extracted, cut longitudinally and washed with cold PBS. Tumors were excised with scissors and homogenized with a razor blade. Tumorigenesis is induced in MLH1-KO mice by giving drinking water with 1% DSS for 1 week. DNeasy DNA isolation protocol for animal tissue. Illumina paired-end Whole exome capture library preparation using full-length index adapters and KAPA DNA polymerase. DNA is fragmented before library construction and Clusters are generated before sequencing.

text: adult

characteristic: developmental stage

text: hereditary nonpolyposis colon cancer

characteristic: disease

characteristic: individual

text: Mus musculus

text: Mus musculus

text: colon

text: neoplasm

characteristic: sampling site

text: Mus musculus

text: male

characteristic: sex

characteristic: strain

characteristic: title

characteristic: tumor grading


characteristic: ENA-CHECKLIST

characteristic: External Id

text: The Weizmann Institute of Science

characteristic: INSDC center name

characteristic: INSDC first public

text: public

characteristic: Submitter Id

text: 8 to 12

characteristic: age

text: ArrayExpress

characteristic: broker name

text: house mouse

characteristic: common name

text: adult

text: hereditary nonpolyposis colon cancer

characteristic: genotype

text: Mus musculus

text: Mus musculus

text: colon

text: normal tissue adjacent to neoplasm

text: Mus musculus

text: male


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

tag: Namespace:Human embryo and stem cell laboratory, The Francis Crick Institute

text: ArrayExpress

characteristic: cell line

text: embryonic stem cell

characteristic: cell type

text: human

text: Protocols: RNA was isolated using TRI reagent (Sigma) by applying 1 ml reagent in-plate to one 6-well of cells and scraping to collect all material. Doxycycline-inducible overexpression of HA-tagged proteins was achieved using the Lenti-X Tet-On 3G Inducible Expression System (Clontech) following the manufacturer’s protocol, and as outlined previously (Wamaitha et al., 2015). Lentiviral packaging was achieved using 7 μg of transgene-containing plasmid and the Lenti-X Packaging Single Shot reagents. Lentiviral supernatant was harvested after 48hrs and concentrated by ultracentrifugation. To produce stably transduced cells, hESCs were plated under standard conditions and changed into fresh medium the following morning. 24hrs post-plating, 10 μl concentrated virus was added to hESCs for transduction overnight (~16hrs). hESCs were dual selected with 150 μg/ml G418 and 0.5 μg/ml puromycin 48hrs post-transduction. For the RNA-seq experiments, KLF17-inducible hESCs were plated as normal and induction initiated after 24 hours by addition of 1 μg/ml Dox to the culture medium (mTeSR1). At ~30 hours, a day 0 (pre-induction) control sample was collected, then both induced (+Dox) and uninduced (UI) samples were collected at 24hr intervals from 48hrs (day 1 post-induction) until 144hrs (day 5 post-induction). TRI reagent samples were subjected to phenol/chloroform-based extraction, and DNase I-treated (Ambion). cDNA was synthesised using a Maxima first strand cDNA synthesis kit (Fermentas). DNase-treated RNA was supplied to the in-house sequencing facility and subjected to library construction via the KAPA mRNA (polyA) HyperPrep protocol.

characteristic: description

text: embryo stage

characteristic: developmental stage

text: normal

text: Dox-inducible KLF17_HA transgene

text: Homo sapiens

characteristic: organism

text: Homo sapiens

text: blastocyst

characteristic: organism part

text: Homo sapiens

text: doxycycline, 1 microgram per milliliter

characteristic: stimulus


characteristic: ENA-CHECKLIST

characteristic: External Id

text: Human embryo and stem cell laboratory, The Francis Crick Institute

characteristic: INSDC center name

characteristic: INSDC first public

text: public

characteristic: Submitter Id

text: ArrayExpress

text: embryonic stem cell

text: human

text: embryo stage

text: normal

characteristic: disease

text: Dox-inducible KLF17_HA transgene

text: Homo sapiens

text: Homo sapiens

text: blastocyst

text: Homo sapiens

text: doxycycline, 1 microgram per milliliter

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: INSDC last update

text: public

text: ArrayExpress

characteristic: broker name

text: embryonic stem cell

text: human

characteristic: common name

text: embryo stage

text: normal

text: Dox-inducible KLF17_HA transgene

characteristic: genotype

text: Homo sapiens

text: Homo sapiens

text: blastocyst

text: Homo sapiens


text: Haukeland University Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

tag: Namespace:Haukeland University Hospital

text: Spondylodiscitis

characteristic: bio material

characteristic: collection date

text: Norway

characteristic: geographic location (country and/or sea)

text: diseased

characteristic: host health state

text: Homo sapiens

characteristic: host scientific name

characteristic: isolate

text: Hospital

characteristic: isolation source

characteristic: lat lon

text: Escherichia marmotae

text: Escherichia marmotae

text: Clinical E. marmotae


characteristic: ENA-CHECKLIST

characteristic: External Id

text: Haukeland University Hospital

characteristic: INSDC center name

characteristic: INSDC first public

text: public

characteristic: Submitter Id

text: Urine

text: Norway

text: diseased

text: Homo sapiens

text: Hospital

text: Escherichia marmotae

characteristic: organism

text: Escherichia marmotae

text: Clinical E. marmotae

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: Haukeland University Hospital

characteristic: INSDC last update

text: public

characteristic: INSDC status

text: Blood

text: Norway

text: diseased

text: Homo sapiens

text: Hospital

text: Escherichia marmotae

text: Escherichia marmotae

text: Clinical E. marmotae


text: Haukeland University Hospital

text: public

characteristic: SRA accession

text: Pus

characteristic: bio material

characteristic: collection date

text: Norway

characteristic: geographic location (country and/or sea)

text: diseased

text: Homo sapiens

text: Hospital

text: Escherichia marmotae

text: Escherichia marmotae

text: Clinical E. marmotae


text: public

tag: Namespace:Department of Neurological Surgery Washington University in St. Louis

characteristic: age

text: ArrayExpress

characteristic: broker name

text: glioblastoma and infiltrating normal cell

text: human

characteristic: common name

text: Protocols: Cell dissociation and tissue processing: Fresh tumor samples were dissociated using the gentleMACS Octo Dissociator and Brain tumor dissociation kit (Miltenyi). Myelin and red blood cells were removed using the MACS Myelin removal beads II and ACK Red Blood Cell lysis buffer, respectively. Dead cells were removed (MACS Dead cell removal microbeads) and viable cells were counted by Trypan blue exclusion. Fresh tumor samples were dissociated using the gentleMACS Octo Dissociator and Brain tumor dissociation kit (Miltenyi). Myelin and red blood cells were removed using the MACS Myelin removal beads II and ACK Red Blood Cell lysis buffer, respectively. Dead cells were removed (MACS Dead cell removal microbeads) and viable cells were counted by Trypan blue exclusion. Dissociated tumor cells were processed using the 10x Genomics Chromium Controller and the Chromium Single Cell 3′V2 Library & Gel Bead Kit following the manufacturer's protocols (https://tinyurl.com/ybpg2pfz). Dissociated tumor cells were processed using the 10x Genomics Chromium Controller and the Chromium Single Cell 3′V2 Library & Gel Bead Kit following the manufacturer's protocols (https://tinyurl.com/ybpg2pfz).

characteristic: description

text: adult

characteristic: developmental stage

text: glioblastoma multiforme

characteristic: disease

characteristic: immunophenotype

characteristic: individual

text: not applicable

characteristic: isolate

text: Homo sapiens

text: Homo sapiens

text: brain

characteristic: organism part

text: Homo sapiens

text: male

characteristic: sex


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: Department of Neurological Surgery Washington University in St. Louis

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: Submitter Id

text: ArrayExpress

text: human

text: adult

text: glioblastoma multiforme

text: unsorted

text: not applicable

text: Homo sapiens

characteristic: organism

text: Homo sapiens

text: brain

text: Homo sapiens

text: male

characteristic: title


text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: age

text: ArrayExpress

characteristic: broker name

text: human

characteristic: common name

characteristic: description

text: adult

characteristic: developmental stage

text: glioblastoma multiforme

text: unsorted

text: not applicable

text: Homo sapiens

text: Homo sapiens

text: brain

text: Homo sapiens

text: female


E-MTAB-10979:Sample 1

characteristic: ENA-CHECKLIST

text: EMBL HEIDELBERG

text: public

text: E-MTAB-10979:Sample 1

tag: Namespace:EMBL HEIDELBERG

text: ArrayExpress

text: primary cell line

characteristic: cell line

text: mammary epithelial cell

characteristic: cell type

text: house mouse

text: Protocols: The cells for the DNA extraction were harvested from three 3D gels per experimental condition by the digestion of the matrigel with collagenase and liberase for 1.5h at 37oC. Subsequently, the individually digested gels were pooled, washed three times with PBS and STOP medium (Lonza/Amaxa DMEM/F12 1:1 Mixture with HEPES, L-Gln, BE12-719F supplemented with HEPES to the final concentration of 25 mM and 10% Tet System Approved Fetal Bovine Serum, Biowest, S181T) and centrifuged shortly (1000 rpm, 2 min, at room temperature). Primary mammary epithelial cells were obtained through a digestion of mouse mammary glands in 5 mL of digestion medium (Lonza/Amaxa DMEM/F12 1:1 Mixture with HEPES, L-Gln, BE12-719F) supplemented with HEPES to the final concentration of 25 mM, 150 U/mL Collagenase type 3 (Worthington, LS004183), 20 μg/mL Liberase Blendzyme 2 (Roche, 05401020001) and 5 mL of Penicillin/Streptomycin (Gibco Life Technologies, 15140-122). The digestion for 15-16 hours at 37oC in 5% (vol/vol) CO2 atmopshere in loosely capped 50 mL polypropylene conical tubes was followed by a washing step with 45 mL of phosphate-buffered saline (PBS). Upon centrifugation at room temperature, 1000 rpm for 5 min, the interphase between the upper fat layer and the cell pellet was removed and 5 mL of 0.25% trypsin-EDTA (Invitrogen, 25200-056) was added. The suspension was incubated for 40 min at 37oC, 5% CO2 in loosely capped tubes, followed by a wash with 25 mL of STOP medium (Lonza/Amaxa DMEM/F12 1:1 Mixture with HEPES, L-Gln, BE12-719F supplemented with HEPES to the final concentration of 25 mM and 10% Tet System Approved Fetal Bovine Serum, Biowest, S181T) and a treatment with 5-15 mg/mL DNase I (ThermoFisher, 18068015). After another centrifugation step at room temperature, 1000 rpm for 5 min, the dissociated cells were resuspended in MEBM medium (Lonza, Mammary Epithelial Cell Basal Medium CC-3151 with supplements from Mammary Epithelial Cell Medium BulletKit CC- 3150) and plated onto collagen-coated plates (BD Biosciences, 356400) for the selection of epithelial cells. On the next day, the cells were washed with PBS and the remaining cells were treated with 500 μL of 0.25 % trypsin-EDTA until detachment. Trypsin was inactivated with 9 mL of STOP medium (described above), followed by a centrifugation step at room temperature with 1000 rpm for 5 min to obtain the primary mammary epithelial cells for seeding. To establish the 3D organoid culture the cell pellets were resuspended in PBS, counted, and mixed rapidly on ice with the prepared Matrigel-collagen mixture - Cultrex 3D Culture Matrix Basement Membrane Extract (Biozol, TRE-3445-005-01) and 1.5 mg/mL Cultrex 3D Collagen I rat tail (TEMA Ricerca, 3447-020-01). The mixed droplets in a volume of 100 μL, containing 12000 primary mouse mammary epithelial cells, were dispensed into flat bottom wells (Corning CellBIND 12 Well Clear Multiple Well Plates, 3336). After solidifying for 40 min at 37o C, 1.5 ml of MEBM serum-free medium (500 mL supplemented with 2 mL of bovine pituitary extract, 0.5 mL of hEGF, 0.5 mL of hydrocortisone, 0.5 mL of GA-1000, 0.5 mL Insulin from Mammary Epithelial Cell Medium BulletKit CC-3150) was added to each well and changed every second day. Subsequently, the cells were grown in the wells for 7 days until the 3D organoid culture had established including the formation of acini. After 7 days of 3D culture establishment the experiment was started by the addition of doxycycline (200 ng/mL) to the cell culture medium initiating transcription of the oncogenes and rapid cell proliferation. Tumor samples were harvested 5 days after oncogene induction. Subsequently, doxycycline was removed from the cell culture medium, which resulted in a silencing of the oncogenes followed by tumor regression. Residual samples were harvested 7 days after de-induction (12 days in total). Non-induced structures were grown in medium without the addition of doxycycline and sampled in parallel at the same time point as the residual cells (day 12). The medium was changed every second day with the exception of the doxycycline-induced cultures at the days three to eight, during which it was changed daily. The DNA extraction from the harvested cell pellet was done following the Qiagen protocol for cultured cells (Qiagen Blood & Cell Culture DNA Mini kit, 13323). The library for Enzymatic Methyl-seq (EM-seq) was prepared with the NEBNext enzymatic Methyl-seq Kit (NEW ENGLAND BioLabs Inc.) from 100 ng of sheared DNA. According to the protocol’s recommendations 2 ng of lambda phage DNA and 0.1 ng of pUC19 plasmid DNA were spiked-in per 100 ng sample DNA prior to fragmentation. The fragmentation of the DNA mix was done using a Covaris S2 focused-ultrasonicator.

text: adult

text: food pellets TD (Envigo)

characteristic: diet

text: normal

characteristic: disease

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

characteristic: genotype

text: Lonza MEBM serum free media (CC-3151), 500 mL supplemented with 2 mL of bovine pituitary extract, 0.5 mL of hEGF, 0.5 mL of hydrocortisone, 0.5 mL of GA-1000 and 0.5 mL Insulin from Mammary Epithelial Cell Medium BulletKit (CC-3150)

characteristic: growth condition

text: Mouse 283

characteristic: individual

text: Mus musculus

text: Mus musculus

text: mammary gland

characteristic: organism part

text: Mus musculus

text: female

characteristic: sex

text: FVB

characteristic: strain

text: Sample 1


E-MTAB-10979:Sample 2

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: EMBL HEIDELBERG

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10979:Sample 2

characteristic: Submitter Id

text: ArrayExpress

text: primary cell line

text: mammary epithelial cell

text: house mouse

text: adult

text: food pellets TD (Envigo)

text: normal

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

text: Mouse 503

text: Mus musculus

characteristic: organism

text: Mus musculus

text: mammary gland

text: Mus musculus

text: female

text: FVB

text: Sample 2

characteristic: title


E-MTAB-10979:Sample 3

characteristic: ENA-CHECKLIST

text: EMBL HEIDELBERG

text: public

text: E-MTAB-10979:Sample 3

characteristic: age

text: ArrayExpress

characteristic: broker name

text: primary cell line

characteristic: cell line

text: mammary epithelial cell

characteristic: cell type

text: house mouse

characteristic: common name

characteristic: description

text: adult

characteristic: developmental stage

text: food pellets TD (Envigo)

characteristic: diet

text: tumor

characteristic: disease

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

characteristic: genotype

text: Mouse 283

text: Mus musculus

text: Mus musculus

text: mammary gland

text: Mus musculus

text: female

text: FVB

text: Sample 3


E-MTAB-10979:Sample 4

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: EMBL HEIDELBERG

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10979:Sample 4

text: ArrayExpress

text: primary cell line

text: mammary epithelial cell

text: house mouse

text: adult

text: food pellets TD (Envigo)

text: tumor

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

characteristic: growth condition

text: Mouse 503

characteristic: individual

text: Mus musculus

text: Mus musculus

text: mammary gland

characteristic: organism part

text: Mus musculus

text: female

characteristic: sex

text: FVB

characteristic: strain

text: Sample 4


E-MTAB-10979:Sample 5

characteristic: ENA-CHECKLIST

text: EMBL HEIDELBERG

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10979:Sample 5

characteristic: Submitter Id

characteristic: age

text: ArrayExpress

characteristic: broker name

text: primary cell line

characteristic: cell line

text: mammary epithelial cell

characteristic: cell type

text: house mouse

characteristic: common name

text: adult

text: food pellets TD (Envigo)

text: tumor

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

text: Mouse 625

text: Mus musculus

characteristic: organism

text: Mus musculus

text: mammary gland

text: Mus musculus

text: female

text: FVB

text: Sample 5

characteristic: title


E-MTAB-10979:Sample 6

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: EMBL HEIDELBERG

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10979:Sample 6

text: ArrayExpress

text: primary cell line

text: mammary epithelial cell

text: house mouse

characteristic: description

text: adult

characteristic: developmental stage

text: food pellets TD (Envigo)

characteristic: diet

text: residual

characteristic: disease

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

characteristic: genotype

text: Mouse 283

text: Mus musculus

text: Mus musculus

text: mammary gland

text: Mus musculus

text: female

text: FVB

text: Sample 6


E-MTAB-10979:Sample 7

characteristic: ENA-CHECKLIST

text: EMBL HEIDELBERG

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10979:Sample 7

characteristic: Submitter Id

characteristic: age

text: ArrayExpress

characteristic: broker name

text: primary cell line

text: mammary epithelial cell

text: house mouse

text: adult

text: food pellets TD (Envigo)

text: residual

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

characteristic: growth condition

text: Mouse 503

characteristic: individual

text: Mus musculus

text: Mus musculus

text: mammary gland

characteristic: organism part

text: Mus musculus

text: female

characteristic: sex

text: FVB

characteristic: strain

text: Sample 7


E-MTAB-10979:Sample 8

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: EMBL HEIDELBERG

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10979:Sample 8

text: ArrayExpress

text: primary cell line

characteristic: cell line

text: mammary epithelial cell

characteristic: cell type

text: house mouse

characteristic: common name

text: adult

text: food pellets TD (Envigo)

text: residual

text: TetO-cMYC/TetO-Neu/MMTV-rtTA

text: Mouse 625

text: Mus musculus

characteristic: organism

text: Mus musculus

text: mammary gland

text: Mus musculus

text: female

text: FVB

text: Sample 8

characteristic: title


characteristic: ENA-CHECKLIST

text: SLC18A3 negative

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND25_SLC18A3_Negative_Large_Rep1

tag: Namespace:Suzhou Ninth People's Hospital

characteristic: Submitter Id

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

text: Protocols: Cells harvested from FACS were centrifuged at 10,000 g for sheath fluid removal, and suspended by 10 μl lysis buffer (Cat. No. 635013, ClonTech Laboratories, Mountain View, CA, USA) containing with 40 U Recombinant Rnase Inhibitor (Cat. No. 2313Q, ClonTech Laboratories). RNA was then processed for reverse transcription using SMARTer Ultra Low Input RNA Kit for Sequencing v3 (Cat. No. 634852, ClonTech Laboratories). The libraries were constructed using KAPA Hyper Prep Kit (Cat. No. KK8504, Roche, Basel, Switzerland)

characteristic: description

text: infant

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: SLC18A3 negative

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND25_SLC18A3_Negative_Large_Rep2

text: ArrayExpress

text: Norway rat

text: infant

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: large cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 negative

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND35_SLC18A3_Negative_Large_Rep1

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: juvenile stage

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: SLC18A3 negative

text: Suzhou Ninth People's Hospital

text: public

text: E-MTAB-10975:PND35_SLC18A3_Negative_Large_Rep2

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

characteristic: description

text: juvenile stage

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: External Id

text: SLC18A3 negative

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND45_SLC18A3_Negative_Large_Rep1

text: ArrayExpress

text: Norway rat

text: adult

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: large cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 negative

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND45_SLC18A3_Negative_Large_Rep2

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: adult

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: SLC18A3 negative

text: Suzhou Ninth People's Hospital

text: public

text: E-MTAB-10975:PND25_SLC18A3_Negative_Small_Rep1

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

characteristic: description

text: infant

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: External Id

text: SLC18A3 negative

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND25_SLC18A3_Negative_Small_Rep2

text: ArrayExpress

text: Norway rat

text: infant

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: small cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 negative

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND35_SLC18A3_Negative_Small_Rep1

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: juvenile stage

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: SLC18A3 negative

text: Suzhou Ninth People's Hospital

text: public

text: E-MTAB-10975:PND35_SLC18A3_Negative_Small_Rep2

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

characteristic: description

text: juvenile stage

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: External Id

text: SLC18A3 negative

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND45_SLC18A3_Negative_Small_Rep1

text: ArrayExpress

text: Norway rat

text: adult

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: small cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 negative

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND45_SLC18A3_Negative_Small_Rep2

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: adult

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

text: public

text: E-MTAB-10975:PND25_SLC18A3_Positive_Large_Rep1

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

characteristic: description

text: infant

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: External Id

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND25_SLC18A3_Positive_Large_Rep2

text: ArrayExpress

text: Norway rat

text: infant

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: large cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 positive

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND35_SLC18A3_Positive_Large_Rep1

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: juvenile stage

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

text: public

text: E-MTAB-10975:PND35_SLC18A3_Positive_Large_Rep2

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

characteristic: description

text: juvenile stage

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: External Id

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND45_SLC18A3_Positive_Large_Rep1

text: ArrayExpress

text: Norway rat

text: adult

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: large cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 positive

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND45_SLC18A3_Positive_Large_Rep2

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: Protocols: Cells harvested from FACS were centrifuged at 10,000 g for sheath fluid removal, and suspended by 10 μl lysis buffer (Cat. No. 635013, ClonTech Laboratories, Mountain View, CA, USA) containing with 40 U Recombinant Rnase Inhibitor (Cat. No. 2313Q, ClonTech Laboratories). RNA was then processed for reverse transcription using SMARTer Ultra Low Input RNA Kit for Sequencing v3 (Cat. No. 634852, ClonTech Laboratories). The libraries were constructed using KAPA Hyper Prep Kit (Cat. No. KK8504, Roche, Basel, Switzerland)

text: adult

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: large cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

text: public

text: E-MTAB-10975:PND25_SLC18A3_Positive_Small_Rep1

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

characteristic: description

text: infant

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: External Id

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND25_SLC18A3_Positive_Small_Rep2

text: ArrayExpress

text: Norway rat

text: infant

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: small cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 positive

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND35_SLC18A3_Positive_Small_Rep1

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: juvenile stage

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

text: public

text: E-MTAB-10975:PND35_SLC18A3_Positive_Small_Rep2

characteristic: age

text: ArrayExpress

characteristic: broker name

text: Norway rat

characteristic: description

text: juvenile stage

characteristic: developmental stage

text: wild type genotype

characteristic: genotype

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

characteristic: phenotype

text: Rattus norvegicus

text: female

text: Sprague Dawley


characteristic: External Id

text: SLC18A3 positive

text: Suzhou Ninth People's Hospital

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: E-MTAB-10975:PND45_SLC18A3_Positive_Small_Rep1

text: ArrayExpress

text: Norway rat

text: adult

text: wild type genotype

characteristic: individual

text: Rattus norvegicus

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

characteristic: organism part

text: small cell

text: Rattus norvegicus

text: female

characteristic: sex

text: Sprague Dawley

characteristic: strain


characteristic: ENA-CHECKLIST

text: SLC18A3 positive

characteristic: FACs marker

text: Suzhou Ninth People's Hospital

text: public

characteristic: INSDC status

characteristic: SRA accession

text: E-MTAB-10975:PND45_SLC18A3_Positive_Small_Rep2

characteristic: Submitter Id

text: ArrayExpress

text: Norway rat

characteristic: common name

text: adult

text: wild type genotype

text: Rattus norvegicus

characteristic: organism

text: Rattus norvegicus

text: arcuate nucleus of hypothalamus

text: small cell

text: Rattus norvegicus

text: female

text: Sprague Dawley

characteristic: title


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: ETH Zurich, D-BSSE

text: public

tag: Namespace:ETH Zurich, D-BSSE

text: ArrayExpress

characteristic: broker name

characteristic: cell line

text: brain cell

characteristic: cell type

text: human

text: Protocols: Whole organoids were dissociated for generating single cell suspensions by Papain based dissociation (Miltenyi Biotec, Neural tissue dissociation kit (P)). RFP+, GFP+, and BFP+ cells were sorted into separate pools using FACS. Single cells were encapsulated along with beads and reverse transcription reagents into droplets using 10X microfluidic chip system. Reverse transcription of first strand cDNA was performed in droplets. RFP/GFP+ (iTracer-TSC2-targeting iCRISPR) and BFP/GFP+ (iTracer-non-targeting) cells were sorted into separate FACS tubes and plated in 12-cell matrigel-coated plates with mTeSR1. Following cell recovery after FACS, 2000 cells per well in a 96-well plate (10% RFP/GFP+, 10% BFP/GFP+, and 80% wildtype) were seeded and differentiated into cerebral organoids using a whole organoid differentiation protocol by Lancaster et al. (Nature Protocols, 2014). Organoids were scarred by activation of inducible Cas9 at day 5. Scarring was achieved using 8ug doxycycline in NIM for 24 hours. Organoids were incubated in scarring media for 24 hours before returning to base media without doxycycline. Single stranded cDNA was cleaned up and amplified after breaking the droplets generated during the sample collection using the 10X 3' Single cell Kit v3 according to the manufacturer's instructions. Quality and quantity of the resulting cDNA was assessed using Agilent's Bioanalyzer High Sensitivity DNA assay. Libraries were constructed following the 10X 3' library construction protocol v3. Briefly, cDNA was fragmented, end repaired and adaptor sequences were ligated. Barcode and scar regions were further amplified from 60-70ng of cDNA. Afterwards, a PCR was applied to introduce library specific sequencing primers as provided by 10X. Libraries were quantified and checked for size distribution using Agilent Bioanalyzer DNA High Sensitivity Assays.

characteristic: description

text: normal

characteristic: disease

text: iTracer

characteristic: genotype

text: cerebral organoid

characteristic: growth condition

text: BFP+

characteristic: immunophenotype

text: Homo sapiens

text: Homo sapiens

text: induced pluripotent stem cell

characteristic: progenitor cell type

unit: month

characteristic: sampling time point

text: Homo sapiens


characteristic: ENA-CHECKLIST

characteristic: External Id

text: ETH Zurich, D-BSSE

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

text: ArrayExpress

text: brain cell

text: human

text: normal

text: iTracer

text: cerebral organoid

text: GFP+

text: Homo sapiens

text: Homo sapiens

text: induced pluripotent stem cell

text: Homo sapiens


characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

text: ETH Zurich, D-BSSE

text: public

characteristic: Submitter Id

text: ArrayExpress

characteristic: broker name

text: brain cell

text: human

characteristic: common name

text: normal

text: iTracer

text: cerebral organoid

text: RFP+

text: Homo sapiens

characteristic: organism

text: Homo sapiens

text: induced pluripotent stem cell

text: Homo sapiens

characteristic: title


text: University of Oxford

characteristic: GAL

text: NOT PROVIDED

text: EarlhamInstitute

text: public

text: NOT PROVIDED

characteristic: barcoding center

text: COPO

text: Liam Crowley

characteristic: collected by

text: University of Oxford

characteristic: collecting institution

characteristic: collection date

text: United Kingdom

characteristic: geographic location (country and/or sea)

unit: m

characteristic: geographic location (elevation)

characteristic: geographic location (latitude)

characteristic: geographic location (longitude)

text: Berkshire | Wytham woods

characteristic: geographic location (region and locality)

text: Grassland

characteristic: habitat

text: Liam Crowley

characteristic: identified by

text: University of Oxford

text: adult

characteristic: lifestage

text: Dioctria atricapilla

text: WHOLE ORGANISM

characteristic: organism part

text: DTOL

characteristic: project name

text: Dioctria atricapilla

characteristic: scientific name

text: NOT COLLECTED

characteristic: sex

characteristic: specimen id

text: a840e5f6-d66a-4819-a113-fefd287dbb51-DTOL-specimen

characteristic: tolid


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: University of Oxford

characteristic: GAL

text: NOT PROVIDED

text: EarlhamInstitute

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: NOT PROVIDED

characteristic: barcoding center

text: COPO

characteristic: broker name

text: Liam Crowley

characteristic: collected by

text: University of Oxford

text: common hen flea

characteristic: common name

text: United Kingdom

text: Berkshire | Wytham woods

text: Bird's nest

text: Liam Crowley

text: University of Oxford

text: adult

text: Ceratophyllus gallinae

characteristic: organism

text: WHOLE ORGANISM

text: DTOL

text: Ceratophyllus gallinae

text: NOT COLLECTED

text: 634e1ae7-8f59-45c4-b789-7ef43246dcb0-DTOL-specimen

characteristic: title


characteristic: ENA-CHECKLIST

text: University of Oxford

text: NOT PROVIDED

text: EarlhamInstitute

text: public

text: NOT PROVIDED

text: COPO

text: Liam Crowley

text: University of Oxford

characteristic: collecting institution

characteristic: collection date

text: common hen flea

text: United Kingdom

characteristic: geographic location (country and/or sea)

characteristic: geographic location (elevation)

characteristic: geographic location (latitude)

characteristic: geographic location (longitude)

text: Berkshire | Wytham woods

characteristic: geographic location (region and locality)

text: Bird's nest

characteristic: habitat

text: Liam Crowley

characteristic: identified by

text: University of Oxford

text: adult

characteristic: lifestage

text: Ceratophyllus gallinae

text: WHOLE ORGANISM

characteristic: organism part

text: DTOL

characteristic: project name

text: Ceratophyllus gallinae

characteristic: scientific name

text: NOT COLLECTED

characteristic: sex

characteristic: specimen id

text: 632cc6bf-8404-4cd7-a6b8-22f82a1930b4-DTOL-specimen

characteristic: tolid


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: University of Oxford

characteristic: GAL

text: NOT PROVIDED

text: EarlhamInstitute

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: NOT PROVIDED

characteristic: barcoding center

text: COPO

characteristic: broker name

text: Liam Crowley

characteristic: collected by

text: University of Oxford

text: common hen flea

characteristic: common name

text: United Kingdom

text: Berkshire | Wytham woods

text: Bird's nest

text: Liam Crowley

text: University of Oxford

text: adult

text: Ceratophyllus gallinae

characteristic: organism

text: WHOLE ORGANISM

text: DTOL

text: Ceratophyllus gallinae

text: NOT COLLECTED

text: e90f56ba-5a55-4cf4-8734-6600d85fc077-DTOL-specimen

characteristic: title


characteristic: ENA-CHECKLIST

text: University of Oxford

text: NOT PROVIDED

text: EarlhamInstitute

text: public

text: NOT PROVIDED

text: COPO

text: Liam Crowley

text: University of Oxford

characteristic: collecting institution

characteristic: collection date

text: United Kingdom

characteristic: geographic location (country and/or sea)

characteristic: geographic location (elevation)

characteristic: geographic location (latitude)

characteristic: geographic location (longitude)

text: Berkshire | Wytham Farm

characteristic: geographic location (region and locality)

text: Hedgerow

characteristic: habitat

text: Liam Crowley

characteristic: identified by

text: University of Oxford

text: larva

characteristic: lifestage

text: Yponomeuta cagnagella

text: WHOLE ORGANISM

characteristic: organism part

text: DTOL

characteristic: project name

text: Yponomeuta cagnagella

characteristic: scientific name

text: NOT COLLECTED

characteristic: sex

characteristic: specimen id

text: 09866df6-fc98-4673-9642-3a95b2bcb6bd-DTOL-specimen

characteristic: tolid


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: University of Oxford

characteristic: GAL

text: NOT PROVIDED

text: EarlhamInstitute

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: NOT PROVIDED

characteristic: barcoding center

text: COPO

characteristic: broker name

text: Liam Crowley

characteristic: collected by

text: University of Oxford

text: United Kingdom

text: Berkshire | Wytham Farm

text: Hedgerow

text: Liam Crowley

text: University of Oxford

text: larva

text: Yponomeuta cagnagella

characteristic: organism

text: WHOLE ORGANISM

text: DTOL

text: Yponomeuta cagnagella

text: NOT COLLECTED

text: 20756eae-3264-4971-98b5-984bb2ce7628-DTOL-specimen

characteristic: title


text: University of Oxford

text: NOT PROVIDED

text: EarlhamInstitute

text: public

text: NOT PROVIDED

text: COPO

text: Vladimir Blagoderov

text: University of Oxford

characteristic: collecting institution

characteristic: collection date

text: United Kingdom

characteristic: geographic location (country and/or sea)

characteristic: geographic location (elevation)

characteristic: geographic location (latitude)

characteristic: geographic location (longitude)

text: Berkshire | Wytham woods

characteristic: geographic location (region and locality)

text: Woodland

characteristic: habitat

text: Vladimir Blagoderov

characteristic: identified by

text: University of Oxford

text: adult

characteristic: lifestage

text: Dixa nubilipennis

text: WHOLE ORGANISM

characteristic: organism part

text: DTOL

characteristic: project name

text: Dixa nubilipennis

characteristic: scientific name

text: NOT COLLECTED

characteristic: sex

characteristic: specimen id

text: bbfba04d-7718-4807-995f-7752e4638679-DTOL-specimen

characteristic: tolid


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: University of Oxford

characteristic: GAL

text: NOT PROVIDED

text: EarlhamInstitute

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: NOT PROVIDED

characteristic: barcoding center

text: COPO

characteristic: broker name

text: Vladimir Blagoderov

characteristic: collected by

text: University of Oxford

text: United Kingdom

text: Berkshire | Wytham woods

text: Woodland

text: Vladimir Blagoderov

text: University of Oxford

text: adult

text: Macrocera phalerata

characteristic: organism

text: WHOLE ORGANISM

text: DTOL

text: Macrocera phalerata

text: MALE

text: be3d9131-0293-40bc-a56a-3aefb0290be3-DTOL-specimen

characteristic: title


text: University of Oxford

text: NOT PROVIDED

text: EarlhamInstitute

text: public

text: NOT PROVIDED

text: COPO

text: Liam Crowley

text: University of Oxford

characteristic: collecting institution

characteristic: collection date

text: United Kingdom

characteristic: geographic location (country and/or sea)

characteristic: geographic location (elevation)

characteristic: geographic location (latitude)

characteristic: geographic location (longitude)

text: Berkshire | Wytham woods

characteristic: geographic location (region and locality)

text: Woodland

characteristic: habitat

text: Liam Crowley

characteristic: identified by

text: University of Oxford

text: adult

characteristic: lifestage

text: Barypeithes pellucidus

text: WHOLE ORGANISM

characteristic: organism part

text: DTOL

characteristic: project name

text: Barypeithes pellucidus

characteristic: scientific name

text: NOT COLLECTED

characteristic: sex

characteristic: specimen id

text: 779eb3f4-3d5d-4ce0-927c-aea97f22093d-DTOL-specimen

characteristic: tolid


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: University of Oxford

characteristic: GAL

text: NOT PROVIDED

text: EarlhamInstitute

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: NOT PROVIDED

characteristic: barcoding center

text: COPO

characteristic: broker name

text: Liam Crowley

characteristic: collected by

text: University of Oxford

text: United Kingdom

text: Berkshire | Wytham woods

text: Woodland

text: Liam Crowley

text: University of Oxford

text: adult

text: Barypeithes pellucidus

characteristic: organism

text: WHOLE ORGANISM

text: DTOL

text: Barypeithes pellucidus

text: NOT COLLECTED

text: 54a159c9-df1c-4d6d-8563-e1f5b40e6b35-DTOL-specimen

characteristic: title


text: University of Oxford

text: NOT PROVIDED

text: EarlhamInstitute

text: public

text: NOT PROVIDED

text: COPO

text: Liam Crowley

text: University of Oxford

characteristic: collecting institution

characteristic: collection date

text: United Kingdom

characteristic: geographic location (country and/or sea)

characteristic: geographic location (elevation)

characteristic: geographic location (latitude)

characteristic: geographic location (longitude)

text: Berkshire | Wytham woods

characteristic: geographic location (region and locality)

text: Woodland

characteristic: habitat

text: Liam Crowley

characteristic: identified by

text: University of Oxford

text: adult

characteristic: lifestage

text: Hermaeophaga mercurialis

text: WHOLE ORGANISM

characteristic: organism part

text: DTOL

characteristic: project name

text: Hermaeophaga mercurialis

characteristic: scientific name

text: NOT COLLECTED

characteristic: sex

characteristic: specimen id

text: e0ba78ac-8fcd-4af7-a347-ded63d75d250-DTOL-specimen

characteristic: tolid


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: ETH Zurich, D-BSSE

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: ArrayExpress

characteristic: broker name

characteristic: cell line

text: brain cell

characteristic: cell type

text: human

characteristic: common name

text: Protocols: Organoids were embedded in pre-chilled Optimal Cutting Temperature (OCT). The sample was then set into a dry ice bath with isopentane until frozen and stored at -80°C. Cryosections were cut at 10um thickness, adhered to ST slides (10x) and stored at -80°C until the following day. Tissue slices were fixed in cold methanol, before being stained in Hematoxylin and eosin. ST slides were imaged as recommended on a Nikon T2i at 20x using a tile scan over all slice sections. 2000 iTracer+ iCRISPR iPSCs per well in a 96-well plate were seeded and differentiated into cerebral organoids using a whole organoid differentiation protocol by Lancaster et al. (Nature Protocols, 2014). Organoids were scarred by activation of inducible Cas9 at day 15. Scarring was achieved by first selecting the organoid to be scarred and transferring it to a 6-24-well plate (depending on organoid size) filled with 8ug doxycycline scarring media. Organoids were incubated in scarring media for 24 hours before returning to base media without doxycycline. Tissue slices were permeabilized to obtain RNA, optimal permeabilization conditions were determined by using the Tissue Optimization Kit (10x) and was found to be 24 minutes. Spot-captured RNA was reverse transcribed before second strand synthesis and cDNA denaturation. qPCR was used to determine the optimal number of cDNA amplification cycles as recommended by the manufacturer. cDNA was amplified using 17-18 cycles. Libraries were constructed following the 10X Visium library construction protocol v1. Barcode and scar regions were further amplified from 60-70ng of cDNA. Afterwards, a PCR was applied to introduce library specific sequencing primers as provided by 10X. Libraries were quantified and checked for size distribution using Agilent Bioanalyzer DNA High Sensitivity Assays.

characteristic: description

text: normal

characteristic: disease

text: iTracer

characteristic: genotype

text: cerebral organoid

characteristic: growth condition

text: Homo sapiens

characteristic: organism

text: Homo sapiens

text: induced pluripotent stem cell

characteristic: progenitor cell type

characteristic: sampling site

characteristic: sampling time point

text: Homo sapiens

characteristic: title


characteristic: ENA-CHECKLIST

characteristic: ENA-FIRST-PUBLIC

characteristic: ENA-LAST-UPDATE

characteristic: External Id

text: SC

characteristic: INSDC center name

characteristic: INSDC first public

characteristic: INSDC last update

text: public

text: SC

text: Lighthouse Lab in Alderley Park

characteristic: collecting institution

characteristic: collection date

text: Samples prepared at Lighthouse Lab in Alderley Park by Jacquelyn Wynn, Mairead Hyland, The Lighthouse Lab in Alderley Park and Alex Alderton, Roberto Amato, Jeffrey Barrett, Sonia Goncalves, Ewan Harrison, David K. Jackson, Ian Johnston, Dominic Kwiatkowski, Cordelia Langford, John Sillitoe on behalf of the Wellcome Sanger Institute COVID-19 Surveillance Team (https://www.sanger.ac.uk/covid-team)

text: United Kingdom

characteristic: geographic location (country and/or sea)

text: England

characteristic: geographic location (region and locality)

text: Human

characteristic: host scientific name

text: Severe acute respiratory syndrome coronavirus 2

text: active surveillance in response to outbreak

characteristic: sample capture status

characteristic: scientific name


text: SC

text: public

characteristic: INSDC status

characteristic: SRA accession

characteristic: Submitter Id

text: SC

characteristic: broker name

text: Lighthouse Lab in Milton Keynes

text: Samples prepared at Lighthouse Lab in Milton Keynes by The Lighthouse Lab in Milton Keynes and Alex Alderton, Roberto Amato, Jeffrey Barrett, Sonia Goncalves, Ewan Harrison, David K. Jackson, Ian Johnston, Dominic Kwiatkowski, Cordelia Langford, John Sillitoe on behalf of the Wellcome Sanger Institute COVID-19 Surveillance Team (https://www.sanger.ac.uk/covid-team)

characteristic: description

text: United Kingdom

text: England

text: Human

characteristic: organism