Development of a Low-Cost Stem-Loop Real-Time Quantification PCR Technique for EBV miRNA Expression Analysis
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  • 作者:Massimiliano Bergallo ; Chiara Merlino ; Davide Montin…
  • 刊名:Molecular Biotechnology
  • 出版年:2016
  • 出版时间:September 2016
  • 年:2016
  • 卷:58
  • 期:8-9
  • 页码:540-550
  • 全文大小:1,100 KB
  • 刊物主题:Biotechnology; Biochemistry, general; Cell Biology; Protein Science; Biological Techniques; Human Genetics;
  • 出版者:Springer US
  • ISSN:1559-0305
  • 卷排序:58
文摘
MicroRNAs (miRNAs) are short, single stranded, non-coding RNA molecules. They are produced by many different species and are key regulators of several physiological processes. miRNAs are also encoded by the genomes of multiple virus families, such as herpesvirus family. In particular, miRNAs from Epstein Barr virus were found at high concentrations in different associated pathologies, such as Burkitt’s lymphoma, Hodgkin disease, and nasopharyngeal carcinoma. Thanks to their stability, these molecules could possibly serve as biomarkers for EBV-associated diseases. In this study, a stem-loop real-time PCR for miR-BART2-5p, miR-BART15, and miR-BART22 EBV miRNAs detection and quantification has been developed. Evaluation of these miRNAs in 31 serum samples (12 from patients affected by primary immunodeficiency, 9 from X-linked agammaglobulinemia and 10 from healthy subjects) has been carried out. The amplification performance showed a wide dynamic range (108–102 copies/reaction) and sensibility equal to 102 copies/reaction for all the target tested. Serum samples analysis, on the other hand, showed a statistical significant higher level of miR-BART22 in primary immunodeficiency patients (P = 0.0001) compared to other groups and targets. The results confirmed the potential use of this assay as a tool for monitoring EBV-associated disease and for miRNAs expression profile analysis.KeywordsmiRNAEBVStem-loop real-time PCRMGBEBV-associated pathologies
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