title: GSM3475605: BS IFE 1; Mus musculus; RNA-Seq
library_construction_protocol: Newborn P0 back skins and paw skin 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) and 1:200 EpCAM-APC (Biolegend) for 30 minutes on ice, and washed twice with 1x HBSS prior to cell sorting. We sorted IFE as GFP(-), EPCAM(+), and SCA1(+). We sorted Merkel cells as GFP(+), EPCAM(+), and SCA(-). 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 150-nucleotide paired-end reads.
attributes.tag: GEO Accession
attributes.value: GSM3475605