补充尿素可减少较短和较长的 RNA 副产物,从而改善 mRNA 的体外转录。

IF 3.6 3区 生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY
RNA Biology Pub Date : 2024-01-01 Epub Date: 2024-02-27 DOI:10.1080/15476286.2024.2321764
Combes Francis, Pettersson Frida J, Bui Thanh-Huong, Molska Alicja, Komissarov Artem, Parot Jérémie, Borgos Sven Even
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引用次数: 0

摘要

这封致编辑的信描述了通过毛细管凝胶电泳、非对称流场流动分馏、免疫印迹、无细胞翻译检测以及在 IFN 报告细胞中评估的小规模体外转录(IVT)反应中 RNA 副产物的存在情况。我们将基于标准 T7 RNA 聚合酶(RNAP)的 IVT 反应与最近描述的采用尿素补充或使用 VSW3 RNAP 的两种方案进行了比较。结果表明,补充尿素可大大减少 RNA 副产物的产生,并对全长转录本的总体数量产生积极影响。相比之下,VSW3 IVT 反应的产量较低,产生的截短转录本比例较高。最后,尿素 mRNA 和 VSW3 mRNA 转染小鼠巨噬细胞后引起的 IFN 反应都要少得多。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Urea supplementation improves mRNA in vitro transcription by decreasing both shorter and longer RNA byproducts.

The current letter to the editor describes the presence of RNA byproducts in small-scale in vitro transcription (IVT) reactions as evaluated by capillary gel electrophoresis, asymmetric flow field flow fractionation, immunoblotting, cell-free translation assays, and in IFN reporter cells. We compare standard T7 RNA polymerase (RNAP) based IVT reactions to two recently described protocols employing either urea supplementation or using the VSW3 RNAP. Our results indicate that urea supplementation yields considerably less RNA byproducts and positively affects the overall number of full-length transcripts. In contrast, VSW3 IVT reactions demonstrated a low yield and generated a higher fraction of truncated transcripts. Lastly, both urea mRNA and VSW3 mRNA elicited considerably less IFN responses after transfection in mouse macrophages.

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来源期刊
RNA Biology
RNA Biology 生物-生化与分子生物学
CiteScore
8.60
自引率
0.00%
发文量
82
审稿时长
1 months
期刊介绍: RNA has played a central role in all cellular processes since the beginning of life: decoding the genome, regulating gene expression, mediating molecular interactions, catalyzing chemical reactions. RNA Biology, as a leading journal in the field, provides a platform for presenting and discussing cutting-edge RNA research. RNA Biology brings together a multidisciplinary community of scientists working in the areas of: Transcription and splicing Post-transcriptional regulation of gene expression Non-coding RNAs RNA localization Translation and catalysis by RNA Structural biology Bioinformatics RNA in disease and therapy
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