RNA isolation
Total RNA was isolated from placental tissue with the use of Gene MATRIX
Universal RNA Purification Kit (EurX, Gdansk, Poland) supplemented with
TRI-reagent (phenol equilibrated, stabilized: chloroform: isoamyl
alcohol 25:24:1) (AppliChem GmbH, Darmstadt, Germany) and
β-mercaptoethanol (Acros Organics, New Jersey, USA). The procedure was
conducted according to the manufacturer’s protocol. 3 mg of each sample
were homogenized using gentleMACS Tubes (Miltenyi Biotec GmbH, Bergisch
Gladbach, Germany) and gentleMACS Dissociator (Miltenyi Biotec GmbH,
Bergisch Gladbach, Germany). Total RNA quantity and concentration were
measured by Nanodrop 2000 spectrophotometer (Thermofisher Scientific,
Massachusetts, USA) and stored at -80°C.