qRT-PCR data from: miR-27b-5p is associated with better overall survival and a less aggressive character of HNSCC

The raw qRT-PCR data from an article entitled “miR-27b-5p is associated with better overall survival and a less aggressive character of HNSCC” financed by the National Science Centre, Poland, allocated based on decision no.: 2021/41/N/NZ5/02966 The attached files contain the processed data (qRT-PCR_results.xlsx) and the raw data divided into prepared plates for miRNA-based reactions (miRNA_qRT-PCR_raw_data.xlsx) and analyzed genes for mRNA-based reactions (mRNA_qRT-PCR_raw_data.xlsx). Paired normal and cancer samples are numbered 1 to 54, while unpaired cancer samples are numbered 55 to 155. Total RNA from the FFPE normal tissue and cancer samples was isolated using the High Pure FFPE RNA Micro Kit (Roche, Basel, Switzerland) according to the manufacturer’s protocol. Total RNA from fresh frozen samples was isolated according to Chomczynski’s method. RNA was quantified using a NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA), its quality was assessed by 28S and 18S rRNA bands (1.5% agarose gel electrophoresis in TAE (Tris-acetate-EDTA (ethylenediaminetetraacetic acid) buffer), and then stored at -80°C until use. For miRNA detection, complementary DNA (cDNA) was synthesized from 10 ng of the total RNA using the TaqMan™ MicroRNA Reverse Transcription Kit (Thermo Scientific, Waltham, MA, USA). qRT-PCR was performed with LightCycler 480 Probes Master (Roche, Basel, Switzerland) and specific primers from TaqMan™ MicroRNA Assay (Thermo Scientific, Waltham, MA, USA) to detect miR-27b-5p (miRBase accession number: MIMAT0004588, assay ID: 002174), miR-27b-3p (miRBase accession number: MIMAT0000419, assay ID: 000409), and U18 snoRNA as internal reference (NCBI accession number: AB061820, assay ID: 001204). For mRNA expression estimation: cDNA was synthesized from 1000 ng of the total RNA using the iScript (BioRad, Hercules, CA, USA) kit and subsequently diluted 10x. qRT-PCR was performed as described above using SYBR Green Master (Roche, Basel, Switzerland) and specific primers for FOXD1, MMP2, MMP16, TGFBI, VIM, as well as GAPDH and HPRT1 as internal reference. Reactions were carried out in a LightCycler 96 (Roche, Basel, Switzerland), and a melting curve was performed to discriminate between non-specific products of the PCR reaction. All qRT-PCR data were analyzed by calculating the 2-ΔΔCT and normalizing it against the mean of U18 or housekeeping genes expression – HPRT1 and GAPDH, and subsequently against the mean of each analyzed gene expression value in the normal margin samples.