3-D human epidermal tissue cultures bulk RNA-Seq expression

This dataset corresponds to gene expression changed occurring in the formation of 3-D human epidermal raft cultures. Normal human epidermal keratinocytes were isolated from epidermis (n=3) and grown using J2-3T3 mouse fibroblasts as a feeder layer originally described by Rheinwald and Green53. 3-D human epidermal raft cultures seeded in collagen hydrogels were prepared using three distinct donor pools as described previously54 and grown at an air-liquid interface for 12 days in E-Medium (DMEM/DMEM-F12 (1:1), 5% Fetal Bovine Serum, adenine (180µM), Bovine pancreatic insulin (5µg/ml), Human apo- transferrin (5µg/ml), triiodothyronine (5µg/ml), L-Glutamine (4mM), Cholera toxin (10ng/ml), Gentamicin (10µg/ml), Amphotericin B (0.25µg/ml)). At day 9 at an air-liquid-interface to allow for epidermal maturation, the epidermal rafts (RHE) were treated with 0.1% BSA/phosphate-buffered saline (Sigma Aldrich, St Louis, MO) for 72 Hrs. Epidermal tissues were separated at the stages from Sub-confluent stage to 3-D raft on day 12 (Sub-confluent, Day 0-Confluent, Day 3-Confluent, Day 3-Raft, Day 6-Raft, Day 9-Raft, Day 12-Raft) from the collagen scaffold and lysed in QIAzol for RNA isolation. RNA samples were sent to the University of Michigan Advanced Genomics Core for RNA sequencing. Libraries for RNA-Seq were generated from polyadenylated RNA and sequenced at six libraries per lane on the Illumina Genome Analyzer IIx. We used Tophat2 to align RNA-seq reads to the human genome, using annotations of GENCODE as gene model. HTSeq was used to quantify gene expression levels. Normalization was performed by DESeq2.