Sasaki Kentaro, Urano Kaoru, Mimida Naozumi, Nonaka Satoko, Ezura Hiroshi, Imai Ryozo
Genome-Edited Crop Development Group, Institute of Agrobiological Sciences, National Agriculture and Food Research Organization (NARO), Tsukuba, Japan.
Sanatech Life Science Co. Ltd., Minato-ku, Tokyo, Japan.
Front Genome Ed. 2025 Jun 18;7:1623097. doi: 10.3389/fgeed.2025.1623097. eCollection 2025.
Genome editing in melon ( L.) remains a significant challenge due to the inefficiencies associated with conventional cell culture-based transformation methods. In the present study, a novel Particle Bombardment (iPB) approach was developed to enable DNA-free genome editing in melon without the need for cell culture. CRISPR/Cas9 ribonucleoproteins (RNPs) were coated onto gold particles and delivered directly into shoot apical meristem tissue, which harbors potential germline cells, via particle bombardment. This method was applied to enhance fruit shelf-life by targeting an ethylene biosynthesis gene (). The resulting mutant demonstrated a significantly extended shelf-life, attributable to reduced ethylene production during fruit ripening. This delayed ripening phenotype was reversed upon treatment with exogenous ethylene, confirming the functional impact of disruption. Because this strategy bypasses cell culture, the iPB-RNP method offers a solution to common limitations in genome editing, such as genotype dependence and somaclonal variation. Consequently, this technique holds substantial promise for advancing commercial melon breeding efforts and may be broadly applicable to other species within the Cucurbitaceae family.
由于与传统的基于细胞培养的转化方法相关的低效性，甜瓜（L.）的基因组编辑仍然是一项重大挑战。在本研究中，开发了一种新型的粒子轰击（iPB）方法，以在无需细胞培养的情况下实现甜瓜的无DNA基因组编辑。将CRISPR/Cas9核糖核蛋白（RNPs）包被在金颗粒上，并通过粒子轰击直接递送至含有潜在生殖细胞的茎尖分生组织中。该方法通过靶向乙烯生物合成基因（）来延长果实货架期。所得的突变体表现出显著延长的货架期，这归因于果实成熟期间乙烯产量的降低。用外源乙烯处理后，这种延迟成熟的表型得以逆转，证实了破坏的功能影响。由于该策略绕过了细胞培养，iPB-RNP方法为基因组编辑中的常见限制提供了解决方案，如基因型依赖性和体细胞克隆变异。因此，这项技术在推进商业甜瓜育种工作方面具有巨大潜力，并且可能广泛适用于葫芦科的其他物种。
