RNA was isolated from 300 µL serum using NucleoSpin miRNA Plasma of Macherey Nagel (Dürer, Germany), following the protocol and instructions provided by the manufacturer. This method is a phenol-free protocol based on a silica membrane technology for purification that provides higher miRNA concentrations.14 (link) Carriers or spike-in controls have not been used, and any prior DNase-digestion was performed. Serum samples do not contain significant amounts of DNA; therefore, digestion is not recommended. Total RNA was eluted in 20 µL of RNase-free water. RNA input was normalized by using equal volumes of serum (300 µL).
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Serum miRNA Extraction Protocol
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Corresponding Organization : Centro de Investigación Biomédica en Red
Other organizations : Hospital Universitari i Politècnic La Fe, Sistemas Genómicos
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Variable analysis
independent variables
- RNA isolation method: NucleoSpin miRNA Plasma kit from Macherey Nagel
- MiRNA concentrations
- Serum sample volume: 300 µL
- No use of carriers or spike-in controls
- No prior DNase-digestion
- The authors state that serum samples do not contain significant amounts of DNA, therefore, digestion is not recommended.
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