title: NextSeq 500 sequencing; SmallRNAs profile in 60 Neuroendocrine Neoplasms (NEN) and 3 sera derived from NEN patients
design: SmallRNAs profile in 60 Neuroendocrine Neoplasms (NEN) and 3 sera derived from NEN patients
library_construction_protocol: Out of 60 RNAs, 16 derived from FFPE sections (Formalin fixed paraffin embedded), 29 from frozen sections and 15 were previously extracted RNAs from MTC patients. 3 sera were obtained from whole blood samples by centrifugation at 1900 x g for 10 min at 4°C. The supernatant was further centrifuged at 16000 x g for 10 min at 4°C and stored in aliquots of 0.5 mL at -80°C until analysis. RNA isolations were performed from both FFPE and frozen sections using FFPE DNA Purification Kit (Cat. 47400, Norgen Biotek Corp) and RNA/DNA Purification Kit (Cat. 48700, Norgen Biotek Corp.) respectively, according to the manufacturers protocol.The extraction of total RNA from 200 µL of serum was performed within 1 year of storage at -80°C using miRNeasy Serum/Plasma Kit (Qiagen, Hilden, Germany) according to the manufacturers instructions. Small-RNA libraries were prepared with NEXTFLEX Small RNA-Seq Kit v3 (Cat NOVA-5132-06, Perkin Elmer) starting from 50ng of RNA as input.