characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2021-10-14T00:28:40Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2021-10-14T00:28:40Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA10147638
characteristics.characteristic: External Id
characteristics.text: UNIVERSITY OF CAMBRIDGE
characteristics.characteristic: INSDC center name
characteristics.text: 2021-10-14T00:28:40Z
characteristics.characteristic: INSDC first public
characteristics.text: 2021-10-14T00:28:40Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS7804140
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-10893:OC023
characteristics.characteristic: Submitter Id
characteristics.text: embryonic day 6.5
characteristics.unit: day
characteristics.characteristic: age
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: rbKN3
characteristics.characteristic: cell line
characteristics.text: pluripotent stem cell
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0002248
characteristics.characteristic: cell type
characteristics.text: rabbit
characteristics.characteristic: common name
characteristics.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.
characteristics.characteristic: description
characteristics.text: blastocyst
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/UBERON_0000358
characteristics.characteristic: developmental stage
characteristics.text: normal
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/PATO_0000461
characteristics.characteristic: disease
characteristics.text: low
characteristics.characteristic: fraction
characteristics.text: wild type genotype
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0005168
characteristics.characteristic: genotype
characteristics.text: cell culture
characteristics.characteristic: growth condition
characteristics.text: Oryctolagus cuniculus
characteristics.characteristic: organism
characteristics.text: Oryctolagus cuniculus
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9986
characteristics.characteristic: organism
characteristics.text: embryo
characteristics.characteristic: organism part
characteristics.text: OK
characteristics.characteristic: post analysis well quality
characteristics.text: Oryctolagus cuniculus
characteristics.characteristic: scientific_name
characteristics.text: pass
characteristics.characteristic: single cell well quality
characteristics.text: OC023
characteristics.characteristic: title