GSM3069596: Epi_Control rep3; Mus musculus; RNA-Seq
Newborn P0 back skins were collected and subjected to 0.25% collagenase digestion for 60 minutes at 37ºC with 80 rpm shaking, followed by 0.25% trypsin treatment for 15 minutes at 37ºC. The cell suspension was washed twice with 1x PBS, stained with 1:200 Sca1-PerCP-Cy5.5 (Biolegend), 1:100 α6-integrin-FITC (eBiosciences), and 1:200 EpCAM-APC (Biolegend) for 30 minutes on ice, and washed twice with 1x HBSS prior to cell sorting. Interfollicular epidermis, enriched for epidermal progenitors, was sorted as EpCAM(+), Sca1(+), and α6-integrin(high). FACS-purified cells were collected directly into RLT Plus buffer (QIAGEN), and total RNA was isolated with the RNeasy Plus Micro Kit (QIAGEN). 10ng of total RNA were reverse transcribed and amplified using the Ovation RNA-seq System V2 (Nugen). Libraries were constructed from 100ng of sonicated cDNA (Covaris) using the Ovation Ultra Low DR Multiplex system (Nugen). The concentration and quality of the libraries were determined using Qubit (Invitrogen) and Bioanalyzer (Agilent). Constructed RNA-seq libraries were sequenced on the Illumina HiSeq 2500 platform, obtaining 100-nucleotide single reads.
GEO Accession
GSM3069596