title: Sample 2
organism: Mus musculus
description: Protocols: Ear skin from 10-12-week-old mice (n=2 per strain) was split into dorsal and ventral parts and incubated with 2.5 μg/μL Dispase II (Roche, Basel, Switzerland) for 1 h at 37°C. Epidermis and dermis were separated with closed forceps. The epidermis was further homogenized and digested with 1 μg/μL of collagenase D (Roche, Basel, Switzerland) and 0.2 μg/μL of DNase, (Roche, Basel, Switzerland) for 1 h at 37°C. Digested samples were passed through a 0.7 μm filter (BD Falcon, San Jose, USA) to generate a single cell suspension for staining and sorting. Single cell suspensions of digested epidermis samples were incubated with Fc Block (5 μg, rat anti-mouse CD19/CD32, clone 93, eBioscience) diluted in PBS for 30 min on ice. Samples were subsequently incubated with APC-conjugated rat anti-mouse CD45.2 antibodies (0.5 μg, Clone 104, Biolegend) diluted in PBS + 2% serum + 2 mM EDTA for 30 min on ice. Prior to sorting, cells were labelled with 7-AAD (0.25 μg, eBioscience). Live CD45- cells (7-AAD- and APC- or PE-Cy7-) were sorted using the BD ARIA Fusion sorter at 12 psi with a 100 μm nozzle. ~300,000-500,000 events/cells were collected per sample. scRNA-seq was performed in duplicate for wild-type and K14E7 transgenic animals. The 10X Genomics Chromium instrument (10X Genomics) was used to partition viable CD45- cells with barcoded beads, and cDNA from each cell was prepared using the Single Cell 3' Library, Gel Bead and Multiplex Kit (v1; 10X Genomics; PN-120233) as per the manufacturer's instructions. Library construction performed as per 10X Genomic Single Cell 3' RNA-seq protocol version 1.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2018-10-24T17:03:35Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2018-10-24T11:14:13Z
attributes.tag: External Id
attributes.value: SAMEA5043065
attributes.tag: INSDC center name
attributes.value: Division of Genomics of Disease and Development, Institute for Molecular Bioscience, The University of Queensland, Brisbane, Queensland, Australia, 4072
attributes.tag: INSDC first public
attributes.value: 2018-10-24T17:03:35Z
attributes.tag: INSDC last update
attributes.value: 2018-10-24T11:14:13Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-6429_2:Sample 2
attributes.tag: age
attributes.value: 10 to 12
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell type
attributes.value: keratinocyte
attributes.tag: common name
attributes.value: house mouse
attributes.tag: genotype
attributes.value: KRT14-HPV16E7
attributes.tag: organism part
attributes.value: epidermis
attributes.tag: phenotype
attributes.value: HPV16 E7 oncogene expression in keratinocytes
attributes.tag: sample name
attributes.value: E-MTAB-6429_2:Sample 2
attributes.tag: sampling site
attributes.value: ear
attributes.tag: scientific_name
attributes.value: Mus musculus
attributes.tag: sex
attributes.value: female
attributes.tag: strain
attributes.value: C57BL/6