RNAseq of skin biopsy
rRNA depletion sequencing library was prepared by using QIAGEN FastSelect rRNA HMR Kit (Qiagen, Hilden, Germany). RNA sequencing library preparation uses NEBNext Ultra II RNA Library Preparation Kit for Illumina by following the manufacturers recommendations (NEB, Ipswich, MA, USA). Briefly, enriched RNAs are fragmented for 15 minutes at 94 C. First strand and second strand cDNA are subsequently synthesized. cDNA fragments are end repaired and adenylated at 3ends, and universal adapters are ligated to cDNA fragments, followed by index addition and library enrichment with limited cycle PCR. Sequencing libraries were validated using the Agilent TapeStation 4200 (Agilent Technologies, Palo Alto, CA, USA), and quantified using Qubit 2.0 Fluorometer (ThermoFisher Scientific, Waltham, MA, USA) as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA).