近日，美国密苏里大学R. Keith Slotkin及其团队提发现，转座酶辅助的靶点整合可用于高效植物基因组工程。相关论文于2024年6月26日在线发表在《自然》杂志上。

研究人员开发了一种基因组工程工具，它能控制转座元件（TE）的插入位点和输送的货物，利用TE的天然能力精确切除并插入基因组。受在细菌中以可编程方式定向转座的CRISPR相关转座酶的启发，研究人员将水稻Pong转座酶蛋白与Cas9或Cas12a可编程核酸酶融合。研究人员证明了增强子元件、开放阅读框和基因表达盒的序列特异性定向插入（由CRISPR gRNA引导）到模式植物拟南芥的基因组中。然后，研究人员将这一系统应用于大豆——一种需要定向插入技术的全球主要作物。研究人员已经将TE“寄生虫”设计成了一个可用、易用的工具包，可以将定制的DNA按特定序列定向插入植物基因组。

据介绍，目前将新DNA整合入植物基因组特定位置的技术频率低且容易出错，这种低效率阻碍了开发改良作物的基因组编辑方法。TE通常被认为是基因组的“寄生虫”，它们在演化过程中将自己的DNA无缝插入基因组。真核生物的TE根据对染色质环境的偏好选择插入位点，每种TE的环境都不同。

附：英文原文

Title: Transposase-assisted target-site integration for efficient plant genome engineering

Author: Liu, Peng, Panda, Kaushik, Edwards, Seth A., Swanson, Ryan, Yi, Hochul, Pandesha, Pratheek, Hung, Yu-Hung, Klaas, Gerald, Ye, Xudong, Collins, Megan V., Renken, Kaili N., Gilbertson, Larry A., Veena, Veena, Hancock, C. Nathan, Slotkin, R. Keith

Issue&Volume: 2024-06-26

Abstract: The current technologies to place new DNA into specific locations in plant genomes are low frequency and error-prone, and this inefficiency hampers genome-editing approaches to develop improved crops1,2. Often considered to be genome ‘parasites’, transposable elements (TEs) evolved to insert their DNA seamlessly into genomes3,4,5. Eukaryotic TEs select their site of insertion based on preferences for chromatin contexts, which differ for each TE type6,7,8,9. Here we developed a genome engineering tool that controls the TE insertion site and cargo delivered, taking advantage of the natural ability of the TE to precisely excise and insert into the genome. Inspired by CRISPR-associated transposases that target transposition in a programmable manner in bacteria10,11,12, we fused the rice Pong transposase protein to the Cas9 or Cas12a programmable nucleases. We demonstrated sequence-specific targeted insertion (guided by the CRISPR gRNA) of enhancer elements, an open reading frame and a gene expression cassette into the genome of the model plant Arabidopsis. We then translated this system into soybean—a major global crop in need of targeted insertion technology. We have engineered a TE ‘parasite’ into a usable and accessible toolkit that enables the sequence-specific targeting of custom DNA into plant genomes.

DOI: 10.1038/s41586-024-07613-8

Source: https://www.nature.com/articles/s41586-024-07613-8