Illumina HiSeq 2500 paired end sequencing; Identification of long non-coding RNA expression patterns useful for molecular-based classification of type I endometrial cancers
Identification of long non-coding RNA expression patterns useful for molecular-based classification of type I endometrial cancers
Patients and tissue collection. Six patients, who underwent type A radical hysterectomy with or without pelvic lymphadenectomy for EC, were enrolled in the present study at the University of Salerno. The study protocol received institutional review board (IRB) approval before the beginning of the study, in accordance with The Code of Ethics of the Declaration of Helsinki, and informed consent was obtained from all patients.  From each patient a sample of neoplastic tissue and corresponding normal endometrium were collected, washed and stored at -80° until analysis. The histology of each EC was classified according to World Health Organization criteria, whereas surgical staging followed International Federation of Gynecology and Obstetrics (FIGO) standards.  RNA isolation and quality controls. Tissue specimen were disrupted and homogenized using TissueLyser LT (Qiagen, Germany). Total RNA was extracted with miRVana RNA isolation Kit (Thermo Fisher Scientific, MA, USA) according to the manufacturer's protocol. Before use, RNA concentration in each sample was assayed with a NanoDrop ND-2000c (Thermo Fisher Scientific) and its quality assessed with the Agilent 2100 Bioanalyzer using Agilent RNA 6000 nano kit (Agilent Technologies, CA, USA). Briefly, 1 μg of total RNA was used in a library preparation according to the Illumina TruSeq stranded total RNA sample preparation protocol (Illumina, CA, USA). Each library was sequenced on HiSeq 2500 (Illumina) at a concentration of 8pM for 200 cycles plus 7 additional cycles for indexes sequencing in paired-mode (2x100 base pair).
Experimental Factor: sampling site
neoplasm