Quantifying miRNA and mRNA Expression

For qPCR, RNA was isolated from tissue or cells using TRIzol (Invitrogen 15596–026) according to the manufacturer's protocol. Subsequently, 200 ng RNA was treated with DNAse I (Invitrogen 18068–015). cDNA was synthesised using Superscript II reverse transcriptase (Invitrogen 18064–071) and diluted 4 times with milliQ water. Quantitative real time PCR was performed on a Lightcycler 480 (Roche) using SYBR green (Roche 04887352001). For miRNA quantification cDNA was synthesized with the miScript reverse transcription kit (Qiagen 218061), diluted 8 times with milliQ and qPCR was performed with High Resolution Melting Master (Roche) on a Lightcycler 480 according to the manufacturers' protocols. To validate miRNA-30c expression levels we also performed qPCR with the Taqman primer assay according to the manufacturers' protocol. Primer sequences are listed in Table S3. Analysis of qPCR data was performed using LinRegPCR analysis software [25] (link), [26] (link).