title: Sort2_HFneg
organism: Mus musculus
description: Protocols: Keratinocytes were isolated from three-week-old Lgr6-EGFP-CreETR2 mice as described previously (Jaks et al., 2008). Briefly, dorsal skin was harvested, cut in strips and digested in trypsin. Epidermal cells were peeled off the dermis and singularized by magnetic stirring. Cells from 2 - 3 mice were pooled per experiment. The collected cells were stained with AlexaFluor647-conjugated anti-CD49f antibody and PE-Cy7-conjugated anti-Sca-1 antibody (both from BD Biosciences), and filtered before sorting. Keratinocytes stained for CD49f, Sca-1 and 7-AAD were live sorted using a FACSAria (BD). Lgr6+ IFE cells were defined as the CD49fhigh, Sca-1high and EGFPhigh fraction. Lgr6+ HF cells were defined as the CD49fhigh+dim, Sca-1neg and EGFPhigh fraction. One hundred thousand cells were collected for each population. RNA was isolated from sorted keratinocytes by combining the RNA-containing aqueous phase extract using RNA-Bee (Amsbio) with the RNAaqueous-Micro Kit (Ambion). RNA integrity was measured using an Agilent 2100 Bioanalyzer Pico Chip or a 2200 TapeStation High Sensitivity R6K Screen Tape. The RNA integrity number of the used samples was higher than 8.5. The sequencing library was generated according to the STRT protocol, described in (Islam et al., 2012), using 1 ng total RNA as input for each sample. For each sample two technical replicates were sequenced (suffix _1 and _2). Islam, S., Kjällquist, U., Moliner, A., Zajac, P., Fan, J.B., Lönnerberg, P. and Linnarsson, S. (2012). Highly multiplexed and strand-specific single-cell RNA 5' end sequencing. Nat Protoc 7, 813-828.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2015-10-22T17:01:25Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2018-03-09T00:02:41Z
attributes.tag: External Id
attributes.value: SAMEA3367435
attributes.tag: INSDC center name
attributes.value: Karolinska Institutet Department of Biosciences and Nutrition
attributes.tag: INSDC first public
attributes.value: 2015-10-22T17:01:25Z
attributes.tag: INSDC last update
attributes.value: 2018-03-09T00:02:41Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-3533:Sort2_HFneg
attributes.tag: age
attributes.value: 21 day
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell type
attributes.value: Lgr6-negative hair follicle/sebaceous gland cell
attributes.tag: common name
attributes.value: house mouse
attributes.tag: genotype
attributes.value: Lgr6cre-EGFP-IRES-CreERT2 heterozygous
attributes.tag: organism part
attributes.value: dorsal skin
attributes.tag: phenotype
attributes.value: CD49f hi/Sca1 neg/EGFP neg
attributes.tag: sample name
attributes.value: E-MTAB-3533:Sort2_HFneg
attributes.tag: scientific_name
attributes.value: Mus musculus
attributes.tag: sex
attributes.value: male
attributes.tag: strain
attributes.value: C57BL/6