An immuno-northern technique to measure the size of dsRNA byproducts in in vitro transcribed RNA.
Electrophoresis
; 45(17-18): 1546-1554, 2024 Sep.
Article
em En
| MEDLINE
| ID: mdl-38785136
ABSTRACT
Double-stranded RNA is an immunogenic byproduct present in RNA synthesized with in vitro transcription. dsRNA byproducts engage virus-sensing innate immunity receptors and cause inflammation. Removing dsRNA from in vitro transcribed messenger RNA (mRNA) reduces immunogenicity and improves protein translation. Levels of dsRNA are typically 0.1%-0.5% of total transcribed RNA. Because they form such a minor fraction of the total RNA in transcription reactions, it is difficult to confidently identify discrete bands on agarose gels that correspond to the dsRNA byproducts. Thus, the sizes of dsRNA byproducts are largely unknown. Total levels of dsRNA are typically assayed with dsRNA-specific antibodies in ELISA and immuno dot-blot assays. Here we report a dsRNA-specific immuno-northern blot technique that provides a clear picture of the dsRNA size distributions in transcribed RNA. This technique could complement existing dsRNA analytical methods in studies of dsRNA byproduct synthesis, dsRNA removal, and characterization of therapeutic RNA drug substances.
Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Transcrição Gênica
/
RNA de Cadeia Dupla
/
Northern Blotting
Idioma:
En
Revista:
Electrophoresis
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Estados Unidos