引体编辑可以对杨树杂交品种进行精确的基因组修饰

IF 4.6 4区 农林科学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
Jinpeng Zou, Yuhong Li, Kejian Wang, Chun Wang, Renying Zhuo
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引用次数: 0

摘要

基于CRISPR/ cas的基因组编辑已广泛应用于树木的育种和遗传改良,但在这些物种中进行精确编辑仍然具有挑战性。主要编辑(PE)是一项革命性的精确编辑技术,允许任意碱基替换和小片段的插入/删除。本研究以杨树84K (Populus alba × P)为研究对象。glandulosa)。我们利用2 × 35S启动子表达spCas9缺口酶(nCas9)与工程Moloney小鼠白血病病毒(MMLV)的融合蛋白,利用拟南芥AtU6启动子表达工程PE引导RNA (epegRNA)和Nick gRNA,率先在杨树中建立了Prime Editor 3 (PE3)系统。单碱基替换、多碱基替换和小片段插入/缺失被编辑成三个内源性靶基因。在耐湿霉素(转化)愈伤组织的9个目标位点中的7个中鉴定出所需的编辑,平均编辑效率从0.1到3.6%不等。此外,稳定的T0植物在9个目标中的4个包含所需的编辑,编辑效率从3.6到22.2%不等。PE3体系的建立为杨树基因组的精确修饰提供了有力的工具。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Prime editing enables precise genome modification of a Populus hybrid

CRISPR/Cas-based genome editing has been extensively employed in the breeding and genetic improvement of trees, yet precise editing remains challenging in these species. Prime editing (PE), a revolutionary technology for precise editing, allows for arbitrary base substitutions and the insertion/deletion of small fragments. In this study, we focused on the model tree poplar 84K (Populus alba × P. glandulosa). We used the 2 × 35S promoter to express a fusion protein of spCas9 nickase (nCas9) and engineered Moloney murine leukemia virus (MMLV), and the Arabidopsis thaliana AtU6 promoter to express an engineered PE guide RNA (epegRNA) and Nick gRNA, pioneering the establishment of the Prime Editor 3 (PE3) system in dicot poplar. Single-base substitutions, multiple-base substitutions, and small-fragment insertions/deletions were edited into three endogenous target genes. The desired edits were identified in hygromycin-resistant (transformed) calli at seven out of nine target sites, with an average editing efficiency ranging from 0.1 to 3.6%. Furthermore, stable T0 plants contained the desired edits at four out of nine targets, with editing efficiencies ranging from 3.6 to 22.2%. Establishment of the PE3 system provides a powerful tool for the precise modification of the poplar genome.

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来源期刊
CiteScore
7.70
自引率
2.80%
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