Mesoporous Silica Particle as an RNA Adsorbent for Facile Purification of In Vitro-Transcribed RNA

Messenger RNA vaccines against SARS-CoV-2 hold great promise for the treatment of a wide range of diseases by using mRNA as a tool for generating vaccination antigens as well as therapeutic proteins in vivo. Increasing interest in mRNA preparation warrants reliable methods for in vitro transcription...

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Bibliographic Details
Published in:International journal of molecular sciences 2023-08, Vol.24 (15), p.12408
Main Authors: Cho, Eunbin, Namgung, Jayoung, Lee, Jong Sam, Jang, Jinmin, Jun, Bong-Hyun, Kim, Dong-Eun
Format: Article
Language:English
Subjects:
Acids
Adsorption
Cellulose
Chromatography
COVID-19
COVID-19 Vaccines
Edetic Acid
Ethylenediaminetetraacetic acid
Health aspects
Humans
Hydrogen
Immune response
Messenger RNA
Polyamines
RNA
RNA polymerase
RNA, Messenger
SARS-CoV-2
Silica
Silicon Dioxide - chemistry
Spermidine
Tin
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Summary:Messenger RNA vaccines against SARS-CoV-2 hold great promise for the treatment of a wide range of diseases by using mRNA as a tool for generating vaccination antigens as well as therapeutic proteins in vivo. Increasing interest in mRNA preparation warrants reliable methods for in vitro transcription (IVT) of mRNA, which must entail the elimination of surplus side products such as immunogenic double-stranded RNA (dsRNA). We developed a facile method for the removal of dsRNA from in vitro transcribed RNA with mesoporous silica particles as RNA adsorbents. Various polyamines were tested for the facilitation of RNA adsorption onto mesoporous silica particles in the chromatography. Among the polyamines tested for RNA adsorption, spermidine showed a superior capability of RNA binding to the silica matrix. Mesoporous silica-adsorbed RNA was readily desorbed with elution buffer containing either salt, EDTA, or urea, possibly by disrupting electrostatic interaction and hydrogen bonding between RNA and the silica matrix. Purification of IVT RNA was enabled with the adsorption of RNA to mesoporous silica in a spermidine-containing buffer and subsequent elution with EDTA. By differing EDTA concentration in the eluting buffer, we demonstrated that at least 80% of the dsRNA can be removed from the mesoporous silica-adsorbed RNA. When compared with the cellulose-based removal of dsRNA from IVT RNA, the mesoporous silica-based purification of IVT RNA using spermidine and EDTA in binding and elution, respectively, exhibited more effective removal of dsRNA contaminants from IVT RNA. Thus, mRNA purification with mesoporous silica particles as RNA adsorbents is applicable for the facile preparation of nonimmunogenic RNA suitable for in vivo uses.
ISSN:1422-0067
1661-6596
1422-0067
DOI:10.3390/ijms241512408