characteristics: ERC000011
characteristics: ENA-CHECKLIST
characteristics: 2022-10-15T00:24:54Z
characteristics: ENA-FIRST-PUBLIC
characteristics: 2022-10-15T00:24:54Z
characteristics: ENA-LAST-UPDATE
characteristics: SAMEA10630644
characteristics: External Id
characteristics: UNIVERSITY OF MICHIGAN
characteristics: INSDC center name
characteristics: 2022-10-15T00:24:54Z
characteristics: INSDC first public
characteristics: 2022-10-15T00:24:54Z
characteristics: INSDC last update
characteristics: public
characteristics: INSDC status
characteristics: ERS8282509
characteristics: SRA accession
characteristics: E-MTAB-11089:SE HIO-3
characteristics: Submitter Id
characteristics: ArrayExpress
characteristics: broker name
characteristics: H9
characteristics: cell line
characteristics: intestinal cell
characteristics: cell type
characteristics: monoculture
characteristics: coculture
characteristics: human
characteristics: common name
characteristics: Protocols: 5 organoids were collected per replicate with 4 replicate for each condition at each time point. HIOs were generated by the In Vivo Animal and Human StudiesCore at the University of Michigan Center for Gastrointestinal Research. Human ES cell line WA09 (H9) was obtained from Wicell International Stem Cell Bank and cultured on Matrigel (BD Biosciences)-coated 6-well plates in mTeSR1 medium (Stem Cell Technologies) at 37°C in 5% CO2. Cells were seeded onto Matrigel-coated 24-well plates in fresh mTeSR1 medium and grown until 85 to 90% confluence. Definitive endoderm differentiation was induced by washing the cells with PBS and culturing in endoderm differentiation medium (RPMI 1640, 2% fetal bovine serum [FBS], 2 mM L-glutamine, 100 ng/ml activin A, and 100 U/ml penicillin/streptomycin) for 3 days, and media were exchanged each day. Cells were then washed with endoderm differentiation medium without activin A and cultured in mid/hindgut differentiation medium (RPMI 1640, 2% FBS, 2 mM L-glutamine, 500 ng/ml FGF4, 500 ng/ml WNT3A, and 100 U/ml penicillin-streptomycin) for 4 days until spheroids were present. Spheroids were collected, mixed with ice-cold Matrigel (50ml of Matrigel plus 25ml of media plus 50 spheroids), placed in the center of each well of a 24-well plate, and incubated at 37°C for 10 min to allow Matrigel to solidify. Matrigel embedded spheroids were grown in ENR medium (Dulbecco’s modified Eagle medium [DMEM]-F12, 1 B27 supplement, 2 mM L-glutamine, 100 ng/ml epidermal growth factor [EGF], 100 ng/ml Noggin, 500 ng/ml Rspondin1, and 15 mM HEPES) for 14 days, and medium was replaced every 4 days. Spheroids growing into organoids (HIOs) were dissociated from Matrigel by pipetting using a cut wide-tip pipette (2 to 3 mm). HIOs were mixed with Matrigel (6 HIOs plus 25ml of media plus 50ml of Matrigel) and placed in the center of each well of 24-well plates and incubated at 37°C for 10 min. HIOs were further grown for 14 days in ENR medium with medium exchanged every 4 days. Before use in experiments, HIOs were carved out of Matrigel, washed with DMEM-F12, and replated with 5 HIO/well in 50ml of Matrigel in ENR medium with medium exchanged every 2 to 3 days for 7 days prior to microinjection. Organoids were microinjected with 10^5 CFU/HIO of each Salmonella strain and samples were harvested at 8h post injection. Transcript reads were pseudoaligned to the human genome (GRCh38.p12) using kallisto software RNA was isolated using the mirVana miRNA isolation kit cDNA libraries were prepared by the University of Michigan DNA Sequencing Core using the TruSeq Stranded mRNA Kit (Illumina)
characteristics: description
characteristics: embryo
characteristics: developmental stage
characteristics: normal
characteristics: disease
characteristics: organoid culture
characteristics: growth condition
characteristics: mixed Salmonella enterica subsp. enterica serovar Typhimurium and Salmonella enterica subsp. enterica serovar Enteritidis
characteristics: infect
characteristics: Homo sapiens
characteristics: organism
characteristics: Homo sapiens
characteristics: organism
characteristics: inner cell mass
characteristics: organism part
characteristics: embryonic stem cell
characteristics: progenitor cell type
characteristics: Homo sapiens
characteristics: scientific_name
characteristics: SE HIO-3
characteristics: title