CRISPR/Cas9-mediated editing of carotenoid biosynthesis genes alters carotenoid concentrations in kiwifruit

CRISPR/Cas9 technology has garnered increasing attention for its simplicity and precision in genome editing, making it an indispensable tool for gene function research and crop genetic improvement. However, the inefficiency and time-consuming nature of genetic transformation continue to pose substantial challenges to its widespread application in woody plants. In this study, we developed a rapid and efficient Agrobacterium-mediated transformation system using petioles as explants for kiwifruit. Positive resistant seedlings were obtained within three months by inoculating on MS medium supplemented with 2.0 mg·L-1 6-benzylaminopurine (6-BA), 0.2 mg·L-1 naphthaleneacetic acid (NAA), and 10 mg·L-1 hygromycin, which was faster than using leaves as explants. Using this system, CRISPR/Cas9-mediated editing of phytoene desaturase (AcPDS) and ζ-carotene desaturase (AcZDS) achieved an editing efficiency of 20%. Transgenic kiwifruit lines with edited AcZDS exhibited a significant reduction in carotenoid content. Overall, we established an efficient Agrobacterium-mediated transformation system using petioles as explants, which is applicable for CRISPR/Cas9-mediated gene editing in kiwifruit, thereby facilitating functional gene studies and genetic improvement.