GSM4844223: Eed Control rep3; Mus musculus; RNA-Seq
P71 adult back skins were collected and subjected to 0.25% trypsin digestion for 60 minutes at 37ºC with 80 rpm shaking. 1:200 Sca1-PerCP-Cy5.5 (Biolegend), 1:200 α6-integrin-FITC (eBiosciences), and 1:100 EpCAM-APC (Biolegend) for 30 minutes on ice and washed twice with 1x DPBS before 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 Ultralow system V2 (NuGEN).  The concentration and quality of the libraries were determined using Qubit (Invitrogen) and Bioanalyzer (Agilent). Constructed RNA-seq libraries were sequenced at GENEWIZ on the Illumina HiSeq platform, obtaining 150 nucleotide paired-end reads.