数字农科院2.0

Efffcient site-speciffc integration of kilobase-length DNA fragments in plant cells via Kp03 recombinase

文献类型： 外文期刊

作者： Daqi Yan;;Yanyan Meng;Nan Zhang;Yali Zhao;Conghui Ning;Lina Zhu;Yuntong Liu;Jing Wang;Chunyu Li;Xuehong Shen;Dan Yang;Tianyu Zhang;Yang Liu; Sujie Zhang; Jingqi Du; Huanbin Zhou;Yanlin Liu;Haodong Chen; Tongda Xu; Xiaoyue Zhu; Mingzhang Wen; Yan Xiong

关键词： Kp03large serine recombinasesynthetic biologylarge DNA insertionriceArabidopsis

期刊名称： Cell Reports

ISSN： 2211-1247

年卷期： 2025 年

页码：

收录情况： SCIE（2025版）

摘要： Targeted insertion of large DNA sequences into plant genomes remains a major challenge in synthetic biology. Here, we evaluate the large serine recombinase Kp03 for site-specific integration of DNA fragments in rice and Arabidopsis. In transient protoplast assays, Kp03 mediates efficient insertion of donor DNA up to 27.3 kilobases (kb), with plasmid integration efficiencies reaching 99.1% for fragments up to 3.4 kb. Truncation experiments reveal that a minimal 15-bp attB sequence is necessary for integration. As a proof of concept, Kp03 successfully incorporates a 3.4-kb donor DNA into the rice genome at a locus containing this minimal attB sequence. Moreover, in rice callus, combining Kp03 with the NM-PE genome editing system to install a 26-bp attB site enables targeted integration of a 3.4-kb donor at the desired genomic locus. These findings establish Kp03 as a versatile tool for plant genome engineering, with broad applications for synthetic biology.

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