characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2022-02-03T00:29:41Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2022-02-04T00:14:26Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA12547457
characteristics.characteristic: External Id
characteristics.text: Department of Cell and Developmental Biology
characteristics.characteristic: INSDC center name
characteristics.text: 2022-02-03T00:29:41Z
characteristics.characteristic: INSDC first public
characteristics.text: 2022-02-04T00:14:26Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS10154011
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-11347:h9_HIO_Suspension_EGF_28days
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: H9
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0003045
characteristics.characteristic: cell line
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: To dissociate human intestinal organoids to single cells, organoids were mechanically isolated from Matrigel droplets and then tissue minced into small fragments using a scalpel in a petri dish filled with ice-cold 1X HBSS (with Mg2+, Ca2). This tissue was then transferred to a 15 mL conical tube. Dissociation enzymes and reagents from the Neural Tissue Dissociation Kit (Miltenyi, cat. no. 130-092-628) were used, and all incubation steps were carried out in a 37ºC incubator unless otherwise stated. All tubes and pipette tips used to handle cell suspensions were pre-washed with 1% BSA in HBSS to prevent adhesion of cells to the plastic. Tissue was treated for 15 minutes at 37ºC with Mix 1. Mix 2 was added to the digestion, and tissue was incubated for 10 minute increments at 37ºC until digestion was complete. After each 10 minute incubation, tissue was agitated using a P1000, and tissue digestion was visually assessed under a stereo microscope. This process continued until the tissue was fully digested. Cells were filtered through a 70 µm filter coated with 1% BSA in 1X HBSS, spun down at 500g for 5 minutes at 4ºC and resuspended in 500µl 1X HBSS (with Mg2+, Ca2+). Cells were counted using a hemocytometer, then spun down and resuspended (if necessary) to reach a concentration of 1000 cells/µL and kept on ice. Single cell droplets were immediately prepared on the 10x Chromium according to manufacturer instructions at the University of Michigan The Advanced Genomics Core, with a target of capturing 5,000 cells. Single cell libraries were prepared using the Chromium Next GEM Single Cell 3’ Library Construction Kit v3 according to manufacturer instructions. Low attachment plates were generated using previously published methods (Choi et al., 2011). In summary, poly (2-hydroxyethyl methacrylate) (pHEMA) coating solution was prepared by dissolving 4g of pHEMA (Thermo Fisher) into 40 mL of 95% ethanol with 10 mM NaOH. The solution was shaken immediately upon addition of pHEMA to avoid precipitation and rotated continuously overnight until fully dissolved. 1 mL of pHEMA coating solution was applied to each well of a 6 well plate inside a biosafety cabinet to generate a low attachment culture plate. The plate was rocked side to side to ensure distribution of pHEMA, and then pHEMA was collected for re-use. The low attachment plate was left in the biosafety cabinet overnight with the plate lid open to allow evaporation of excess coating solution, and UV was turned on for 15 minutes to sterilize the plate. Low attachment plates were rinsed twice with 1XPBS prior to addition of spheroids to remove excess pHEMA. Approximately 200 spheroids were transferred to 1 well of a 6-well low-attachment plate containing 5mL of media for suspension HIO culture. To change media without aspirating organoids in suspension, media was collected under a stereomicroscope using a P1000 after allowing organoids to settle to the bottom of the plate. Immediately after tissue dissociation, single cell libraries were prepared on the 10x Chromium with a target of 5000 cells. Chromium Single Cell 3’ Library & Gel Bead Kit v3, 16 rxns PN-120237 was used Immediately after tissue dissociation, single cell libraries were prepared on the 10x Chromium with a target of 5000 cells. Chromium Single Cell 3’ Library & Gel Bead Kit v3, 16 rxns PN-120237 was used
characteristics.characteristic: description
characteristics.text: intestinal organoid culture
characteristics.characteristic: growth condition
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: organism
characteristics.text: embryonic stem cell
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0002322
characteristics.characteristic: progenitor cell type
characteristics.text: 28
characteristics.unit: day
characteristics.characteristic: sampling time point
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: minigut media supplemented with epidermal growth factor
characteristics.characteristic: stimulus
characteristics.text: h9_HIO_Suspension_EGF_28days
characteristics.characteristic: title