Engineering a DNA polymerase for modifying large RNA at specific positions.

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Tác giả: Dian Chen, Zhanghui Han, Xiaoge Liang, Yu Liu

Ngôn ngữ: eng

Ký hiệu phân loại: 782.6 +Womens voices

Thông tin xuất bản: England : Nature chemistry , 2025

Mô tả vật lý:

Bộ sưu tập: NCBI

ID: 160262

The synthesis of large RNA with precise modifications at specific positions is in high demand for both basic research and therapeutic applications, but efficient methods are limited. Engineered DNA polymerases have recently emerged as attractive tools for RNA labelling, offering distinct advantages over conventional RNA polymerases. Here, through semi-rational designs, we engineered a DNA polymerase variant and used it to precisely incorporate a diverse range of modifications, including base modifications, 2'-ribose modifications and backbone modifications, into desired positions within RNA. We achieved efficiencies exceeding 85% in the majority of modification cases, demonstrating success in introducing 2'-O-methyl, phosphorothioate, N4-acetylcytidine and a fluorophore to specific sites in eGFP and Firefly luciferase messenger RNA. Our mRNA products with N4-acetylcytidine, 2'-O-methyl and/or phosphorothioate have demonstrated the ability to enhance stability and affect protein production. This method presents a promising tool for the comprehensive functionalization of RNA, enabling the introduction of plentiful modifications irrespective of RNA lengths and sequences.
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