在 mRNA 模板指导合成过程中去除双链 RNA 污染物。

Removal of Double-Stranded RNA Contaminants During Template-Directed Synthesis of mRNA.

机构信息

State Research Center of Virology and Biotechnology "VECTOR", Federal Service for Surveillance on Consumer Rights Protection and Human Wellbeing, Koltsovo, Novosibirsk region, Russia.

Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, Novosibirsk, Russia.

出版信息

Bull Exp Biol Med. 2024 Apr;176(6):751-755. doi: 10.1007/s10517-024-06102-2. Epub 2024 Jun 19.

DOI:10.1007/s10517-024-06102-2

PMID:38896322

Abstract

The removal of double-stranded RNA (dsRNA) contaminants during in vitro mRNA synthesis is one of the technological problems to be solved. Apparently, these contaminants are the result of the T7 RNA polymerase side activity. In this study, we used a modified method of mRNA purification based on the selective binding of dsRNA to cellulose in ethanol-containing buffer. It was shown both in vivo and in vitro that the cellulose-purified mRNA preparation leads neither to activation of the lymphocyte inflammatory marker CD69 nor to increased release of IFNα in mice, and does not contain impurities detectable by antibodies to dsRNA.

摘要

在体外 mRNA 合成过程中去除双链 RNA (dsRNA) 污染物是需要解决的技术问题之一。显然，这些污染物是 T7 RNA 聚合酶的副产物。在本研究中，我们使用了一种改良的基于 dsRNA 在含乙醇缓冲液中选择性结合纤维素的 mRNA 纯化方法。无论是在体内还是体外实验，均表明经纤维素纯化的 mRNA 制剂既不会导致小鼠淋巴细胞炎症标志物 CD69 的激活，也不会导致 IFNα 的释放增加，且不含有可被 dsRNA 抗体检测到的杂质。