title: Comparison of Two Available RNA Extraction Protocols for microRNA Amplification in Serum Samples.
abstract: BACKGROUND
  microRNAs play a critical role in many biological processes such as cell proliferation and maturation, apoptosis, regulation of chronic inflammation and development of cancer.
  
  
  METHODS
  In this study is described a protocol for the isolation of RNA from serum and subsequent determination of miRNA expression levels using TaqMan-based MGB Real-Time PCR detection. RNA was extracted using two different isolation methods including available kits RNAzol and a modified RNAzol protocol. In all cases, RNA was eluted in RNase free H2 O, kept frozen until analysis and the presence of contaminants assessed by NanoDrop spectrophotometry.
  
  
  RESULTS
  Higher RNA quantity was observed in RNAzol (378.8 ng/μl) vs RNAzol modified protocol (226.5 ng/μl) and a better performance in terms of RNA extraction yield and purity. Subsequently, measurements of endogenous miRNAs (RNU43), cellular miRNAs (mir155 and mir146a) and EBV miRNAs (mirBART2-5p, mirBART15 and mirBART22) were performed by RT-qPCR.
  
  
  CONCLUSIONS
  In contrast to the findings in terms of purity and quantity, the amplifiable RNA was more abundant using RNAzol modified protocol compared to not modified protocol.
authors: Department of Public Health and Pediatric Sciences, Medical School, University of Turin, Turin, Italy.
authors: Massimiliano
authors: M
authors: Bergallo
authors: Department of Public Health and Pediatric Sciences, Medical School, University of Turin, Turin, Italy.
authors: Stefano
authors: S
authors: Gambarino
authors: Department of Public Health and Pediatric Sciences, Medical School, University of Turin, Turin, Italy.
authors: Silvana
authors: S
authors: Martino
authors: Department of Public Health and Pediatric Sciences, Medical School, University of Turin, Turin, Italy.
authors: Davide
authors: D
authors: Montin
authors: Department of Public Health and Pediatric Sciences, Medical School, University of Turin, Turin, Italy.
authors: Paola
authors: P
authors: Montanari
authors: Department of Public Health and Pediatric Sciences, Medical School, University of Turin, Turin, Italy.
authors: Ilaria
authors: I
authors: Galliano
authors: Department of Public Health and Pediatric Sciences, Medical School, University of Turin, Turin, Italy.
authors: Pier-Angelo
authors: PA
authors: Tovo
mesh_terms: D006801:Humans; D035683:MicroRNAs / Q000097:blood* / Q000737:chemistry / Q000302:isolation & purification*; D009690:Nucleic Acid Conformation; D012313:RNA / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*
chemical_list: D035683:MicroRNAs; C034477:RNA primers; D012313:RNA
keywords: MGB probe; extraction; microRNA; real-time PCR; retrotrascription; serum; stem-loop primers