Real-time monitoring strategies for optimization of in vitro transcription and quality control of RNA

被引：4

作者：

Lee, Kyung Hyun ^{[1
]}

Song, Jaehwi ^{[1
]}

Kim, Seongcheol ^{[1
]}

Han, Seung Ryul ^{[1
]}

Lee, Seong-Wook ^{[1
,2
]}

机构：

[1] Rznomics Inc, R&D Ctr, Seongnam, South Korea

[2] Dankook Univ, Res Inst Adv Om, Dept Bioconvergence Engn, Yongin, South Korea

来源：

FRONTIERS IN MOLECULAR BIOSCIENCES | 2023年 / 10卷

基金：

新加坡国家研究基金会;

关键词：

in vitro transcription; IVT; transcription; real-time monitoring; RNA; NUCLEIC-ACIDS; FLUORESCENCE; SPECTROSCOPY; FLUOROPHORE; TRANSLATION; SELECTION; APTAMERS; COMPLEX; BINDING; GENE;

D O I：

10.3389/fmolb.2023.1229246

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

RNA-based therapeutics and vaccines are opening up new avenues for modern medicine. To produce these useful RNA-based reagents, in vitro transcription (IVT) is an important reaction that primarily determines the yield and quality of the product. Therefore, IVT condition should be well optimized to achieve high yield and purity of transcribed RNAs. To this end, real-time monitoring of RNA production during IVT, which allows for fine tuning of the condition, would be required. Currently, light-up RNA aptamer and fluorescent dye pairs are considered as useful strategies to monitor IVT in real time. Fluorophore-labeled antisense probe-based methods can also be used for real-time IVT monitoring. In addition, a high-performance liquid chromatography (HPLC)-based method that can monitor IVT reagent consumption has been developed as a powerful tool to monitor IVT reaction in near real-time. This mini-review briefly introduces some strategies and examples for real-time IVT monitoring and discusses pros and cons of IVT monitoring methods.

引用

页数：7

共 59 条

[21] Revisiting T7 RNA polymerase transcription in vitro with the Broccoli RNA aptamer as a simplified real-time fluorescent reporter [J].