MicroTom番茄( Solanum lycopersicum ‘MicroTom’)是一种矮化模式植物, 因其具有较短的生命周期和清晰的遗传背景而广泛用于植物基因功能研究。传统基于组培的MicroTom番茄遗传转化体系存在效率低、转化周期长及操作复杂等缺陷, 限制了该体系的应用。该研究以2周龄MicroTom实生苗为材料, 在切除顶端子叶与真叶后, 用根癌农杆菌( Agrobacterium tumefaciens)菌液浸润下胚轴切口并诱导再生芽, 成功构建了一套高效的MicroTom番茄非组培遗传转化体系。结果显示, 切口处再生芽阳性率为28.6%, 4–5个月即可收获T ₀代番茄种子, T ₁代植株中外源基因表达比例达73.5%。与传统组培遗传转化体系相比, MicroTom番茄非组培转化体系转化效率显著提高, 转化周期缩短且操作流程简单, 为番茄果实性状基因功能解析提供了高效的技术平台, 对加快番茄分子育种进程具有重要意义。

INTRODUCTION: Genetic transformation, combined with genome editing strategies, has provided essential insights into plant biology and revolutionized crop improvement. MicroTom ( Solanum lycopersicum ‘MicroTom’) is widely used for functional characterization due to its short life cycle and clear genetic background. However, traditional tissue culture-dependent genetic transformation systems for MicroTom are constrained by low efficiency, long durations, and highly trained individuals. Therefore, developing a rapid and efficient tissue culture-independent genetic transformation system is necessary. RATIONALE: Agrobacterium-mediated transformation is the most widely used method for gene transfer in plants. During this process, wounded plant cells secrete phenolic compounds that induce Agrobacterium cells to transfer and integrate foreign DNA into plant chromosomes. In this study, we established a novel transformation protocol using two-week-old MicroTom seedlings as recipients. After removing the apical cotyledons and true leaves, the wounded hypocotyls were directly inoculated with an A. tumefaciens suspension. Based on the somatic cell reprogramming mechanism triggered by wounding signaling, the wound hypocotyls formed calli and regenerated adventitious shoots, accompanied by the integration of foreign DNA. RESULTS: Two-week-old seedlings with hypocotyl diameters exceeding 1.5 mm were optimal for Agrobacterium infection. After a 2-day preculture in darkness, the hypocotyls were infected with A. tumefaciens suspension (OD ₆₀₀=0.6) for 10 min and then cocultured in darkness at 90% relative humidity for 3 d. Callus differentiation was observed at the hypocotyl ends at 10 days post-inoculation (dpi), and adventitious shoots regenerated at 20 dpi. Mature T ₀-generation seeds could be harvested within 4–5 months post-inoculation. Approximately 87.6% of the wounded hypocotyls regenerated adventitious shoots at 20 dpi, and PCR analysis confirmed that 28.6% of the regenerated shoots contained the foreign gene. Through antibiotic screening combined with an EGFP reporter system, the stable expression rate of the foreign gene in T ₁-generation lines reached 73.5%. CONCLUSION: Compared to conventional tissue culture-dependent systems, the established in planta transformation in this study offers improved transformation efficiency, a shorter transformation cycle, and simplified nonsterile operational procedures. This system provides a robust platform for functional genomics studies and significantly lowers technical barriers in tomato molecular breeding.