organism: Mus musculus
description: Protocols: Mice were treated by peroral colonization with 50 microliters of cecal content from SFB monoassociated animals at 21 and 22 days of age or left untreated. Mice were euthanized by cervical dislocation, their small intestine was dissected. Epithelial cells were isolated from small intestines by serial incubation in HBSS + 3% FBS + 2mM EDTA followed by digestion in TrypLE Express enzyme. Intraepithelial cells were isolated in the same manner, but due to their lower frequency in the epithelial fraction they were enriched using 40%/80% percol gradient centrifugation. Lamina propria cells were isolated by the same protocol, but the epithelial fraction was discarded and remaining tissue was used further. This tissue was then minced, digested in collagenase D and DNAse I and immune cells were enriched using 40%/80% percol gradient centrifugation. Single cell suspension of intestinal epithelial cells, lamina propria cells or intraepithelial cells was sorted by flow cytometry as live, single cells. On the top of that, epithelial cells were sorted as CD45- EpCAM+, lamina propria cells and intraepithelial cells were sorted as CD45+. First-strand cDNA was extracted from cell suspension during library preparation routine using Chromium next gem single-cell 3′ reagent kit (version 3.1). Single-cell RNA-seq libraries were prepared using Chromium controller instrument and Chromium Next Gem single-cell 3' reagent kit version 3.1 (both 10X Genomics, Pleasanton, CA, USA) according to the manufacturer's protocol targeting 4000 cells per sample.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2023-01-01
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2023-01-01
attributes.tag: External Id
attributes.value: SAMEA110377196
attributes.tag: INSDC center alias
attributes.value: Institute of Molecular Genetics of the Czech Academy of Sciences
attributes.tag: INSDC center name
attributes.value: Institute of Molecular Genetics of the Czech Academy of Sciences
attributes.tag: INSDC first public
attributes.value: 2023-01-01T00:28:18Z
attributes.tag: INSDC last update
attributes.value: 2023-01-01T00:28:18Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-11982:LP_JAX_SFB
attributes.tag: age
attributes.value: 5
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell_type
attributes.value: small intestinal lamina propria CD45+ cell
attributes.tag: common name
attributes.value: house mouse
attributes.tag: developmental stage
attributes.value: juvenile
attributes.tag: genotype
attributes.value: wild type genotype
attributes.tag: individual
attributes.value: F
attributes.tag: isolate
attributes.value: not applicable
attributes.tag: organism part
attributes.value: small intestine
attributes.tag: replicate
attributes.value: 1
attributes.tag: sample name
attributes.value: E-MTAB-11982:LP_JAX_SFB
attributes.tag: sex
attributes.value: female
attributes.tag: strain
attributes.value: C57BL/6