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. CONCLUSION: 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.
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. CONCLUSION: 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: Etienne Moussay; Kai Wang; Ji-Hoon Cho; Kris van Moer; Sandrine Pierson; Jérôme Paggetti; Petr V Nazarov; Valérie Palissot; Leroy E Hood; Guy Berchem; David J Galas Journal: Proc Natl Acad Sci U S A Date: 2011-04-01 Impact factor: 11.205
Authors: Jason D Arroyo; John R Chevillet; Evan M Kroh; Ingrid K Ruf; Colin C Pritchard; Donald F Gibson; Patrick S Mitchell; Christopher F Bennett; Era L Pogosova-Agadjanyan; Derek L Stirewalt; Jonathan F Tait; Muneesh Tewari Journal: Proc Natl Acad Sci U S A Date: 2011-03-07 Impact factor: 11.205
Authors: Yu Li; Jiangning Li; Sarah Belisle; Carole R Baskin; Terrence M Tumpey; Michael G Katze Journal: Virology Date: 2011-10-13 Impact factor: 3.616
Authors: Katharina L Kopp; Ulrik Ralfkiaer; Lise Mette R Gjerdrum; Rikke Helvad; Ida H Pedersen; Thomas Litman; Lars Jønson; Peter H Hagedorn; Thorbjørn Krejsgaard; Robert Gniadecki; Charlotte M Bonefeld; Lone Skov; Carsten Geisler; Mariusz A Wasik; Elisabeth Ralfkiaer; Niels Ødum; Anders Woetmann Journal: Cell Cycle Date: 2013-05-15 Impact factor: 4.534
Authors: Patricia Rodil-Garcia; Elvira Del Carmen Arellanes-Licea; Angélica Montoya-Contreras; Luis A Salazar-Olivo Journal: Int J Mol Sci Date: 2017-11-28 Impact factor: 5.923
Authors: Antonio Rodríguez; Hans Duyvejonck; Jonas D Van Belleghem; Tessa Gryp; Leen Van Simaey; Stefan Vermeulen; Els Van Mechelen; Mario Vaneechoutte Journal: PLoS One Date: 2020-02-21 Impact factor: 3.240