Application of steric exclusion chromatography on monoliths for separation and purification of RNA molecules

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Levanova , A & Poranen , M M 2018 , ' Application of steric exclusion chromatography on monoliths for separation and purification of RNA molecules ' , Journal of Chromatography. A , vol. 1574 , pp. 50-59 . https://doi.org/10.1016/j.chroma.2018.08.063

Title: Application of steric exclusion chromatography on monoliths for separation and purification of RNA molecules
Author: Levanova, Alesia; Poranen, Minna Marjetta
Contributor: University of Helsinki, Molecular and Translational Virology
University of Helsinki, Molecular and Integrative Biosciences Research Programme
Date: 2018-11-02
Language: eng
Number of pages: 10
Belongs to series: Journal of Chromatography. A
ISSN: 0021-9673
URI: http://hdl.handle.net/10138/259031
Abstract: Steric exclusion chromatography (SXC) is a method for separation of large target solutes based on their association with a hydrophilic stationary phase through mutual steric exclusion of polyethylene glycol (PEG). Selectivity in SXC is determined by the size or shape (or both) of the solutes alongside the size and concentration of PEG molecules. Elution is achieved by decreasing the PEG concentration. In this study, SXC applicability for the separation and purification of single-stranded (ss) and double-stranded (ds) RNA molecules was evaluated for the first time. The retention of ssRNA and dsRNA molecules of different lengths on convective interaction media (CIM) monolithic columns was systematically studied under variable PEG-6000 and NaCl concentrations. We determined that over 90% of long ssRNAs (700-6374 nucleotides) and long dsRNAs (500-6374 base pairs) are retained on the stationary phase in 15% PEG-6000 and >= 0.4 M NaCl. dsDNA and dsRNA molecules of the same length were partially separated by SXC. Separation of RNA molecules below 100 nucleotides from longer RNA species is easily achieved by SXC. Furthermore, SXC has the potential to separate dsRNAs from ssRNAs of the same length. We also demonstrated that SXC is suitable for the enrichment of ssRNA (PRR1 bacteriophage) and dsRNA (Phi6 bacteriophage) viral genomes from contaminating cellular RNA species. In summary, SXC on CIM monolithic columns is an appropriate tool for rapid RNA separation and concentration. (C) 2018 The Authors. Published by Elsevier B.V.
Subject: Nucleic acids
RNA
Steric exclusion chromatography
CIM monolithic columns
Polyethylene glycol (PEG)
DOUBLE-STRANDED-RNA
PHENOL-CHLOROFORM EXTRACTION
SINGLE-STEP METHOD
BACTERIOPHAGE PHI-6
POLYETHYLENE-GLYCOL
AFFINITY-CHROMATOGRAPHY
LIQUID-CHROMATOGRAPHY
VIRUS
PROTEINS
DSRNA
116 Chemical sciences
1182 Biochemistry, cell and molecular biology
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