Developing a homology-driven gene silencing system in Iris japonica: a versatile tool for comparative functional studies

Iris japonica, an evergreen ornamental species known for its beautiful flowers and year-round evergreen foliage, is difficult to genetically manipulate due to the lack of an efficient genetic transformation system. This limitation hinders the functional verification of key genes in I. japonica. To address this, a Virus-Induced Gene Silencing (VIGS) system based on Tobacco rattle virus (TRV) was developed, enabling efficient, transient gene silencing in I. japonica. In this system, the phytoene desaturase (PDS) gene, a common reporter gene used for VIGS, was selected to assess the silencing efficiency. The recombinant vector pTRV2-IjPDS was used to infect I. japonica plants. The PDS gene silencing resulted in distinct photobleaching symptoms, demonstrating the successful silencing of the target gene. The presence of the TRV vector was verified by using a GFP-tagged pTRV2-GFP vector, and GFP expression was detected using fluorescence visualization under ultraviolet (UV) light and confirmed by laser confocal microscopy. Real-time PCR was used to quantify the reduction in IjPDS gene expression, confirming the successful silencing. The study further optimized the VIGS system by evaluating the impact of seedling ages on silencing efficiency, identifying one-year-old seedlings as the most effective for gene silencing (36.67%). This TRV-based VIGS system provides a robust tool for functional gene analysis in I. japonica and offers a new approach to studying the rules of key genes in its biological processes, with potential applications in ornamental plant breeding and genomics research.