EXPERT Expands Prime Editing Efficiency and Range of Large Fragment Edits

Overview

Journal Nat Commun

Specialty Biology

Date 2025 Feb 12

PMID 39939583

Authors

Youcai Xiong

Yinyu Su

Ruigao He

Xiaosong Han

Sheng Li

Minghuan Liu

Xiaoning Xi

Zijia Liu

Heng Wang

Shengsong Xie

Xuewen Xu

Kui Li

Jifeng Zhang

Jie Xu

Xinyun Li

Shuhong Zhao

Jinxue Ruan

Affiliations

Soon will be listed here.

Abstract

Prime editing systems (PEs) hold great promise in modern biotechnology. However, their editing range is limited as PEs can only modify the downstream sequences of the pegRNA nick. Here, we report the development of the extended prime editor system (EXPERT) to overcome this limitation by using an extended pegRNA (ext-pegRNA) with modified 3' extension, and an additional sgRNA (ups-sgRNA) targeting the upstream region of the ext-pegRNA. We demonstrate that EXPERT can efficiently perform editing on both sides of the ext-pegRNA nick, a task that is unattainable by canonical PEs. EXPERT exhibits prominent capacity in replacing sequences up to 88 base pairs and inserting sequences up to 100 base pairs within the upstream region of the ext-pegRNA nick. Compared to canonical PEs such as PE2, the utilization of the EXPERT strategy significantly enhances the editing efficiency for large fragment edits with an average improvement of 3.12-fold, up to 122.1 times higher. Safety wise, the use of ups-sgRNA does not increase the rates of undesirable insertions and deletions (indels), as the two nicks are on the same strand. Moreover, we do not observe increased off-target editing rates genome-wide. Our work introduces EXPERT as a PE tool with significant potential in life sciences.

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