Agrobacterium-mediated transformation systems are extensively applied in japonica rice varieties. However, the adaptability of local rice varieties to existing transformation systems remains limited, owing to their complex genotypes, posing a substantial challenge to transformation. In this study, four local rice varieties were selected to optimize the effects of different culture media on callus induction, browning resistance, contamination resistance, callus tolerance, differentiation, regeneration, and root development, and then two varieties were selected to improve plant architecture and tiller development by CRISPR/Cas9-mediated gene editing, based on constructive transformation systems. The goal was to enhance the transformation efficiency of local varieties and innovate germplasms. The results demonstrated that japonica rice varieties XG293 and WD68 exhibited higher induction rates under the treatment of 2 mg/L 2,4-D (2,4-Dichlorophenoxyacetic acid) + 1 mg/L NAA (Naphthaleneacetic acid), whereas indica rice varieties H128 and E33 performed the best under 3 mg/L 2,4-D + 1 mg/L NAA. Severe browning in H128 was effectively mitigated by a carbon source of 20 g/L maltose supplemented with 40 mg/L ascorbic acid. Contamination after Agrobacterium infection was controlled by 300 mg/L Tmt (Timentin). Under a treatment of 200 µM/L acetosyringone +10 min infection duration, XG293 and WD68 exhibited higher callus tolerance, differentiation rates, and GUS staining rates, achieving transformation efficiencies of 43.24% and 52.38%, respectively. In contrast, H128 and E33 performed better under the treatment of 200 µM/L Acetosyringone + 5 min, with transformation efficiencies of 40.00% and 40.74%, respectively. The mutants after OsCCD7 gene editing in WD68 and H128 displayed a dwarfness of plant height, a significant increase in tiller numbers, and compact architecture. These findings demonstrate that an optimized combination of plant growth regulators and infection durations effectively improves transformation efficiency for local varieties, and the OsCCD7 gene regulates plant architecture and tiller development with variable effects, depending on the rice complex genotypes. This study provides a theoretical basis for the efficient transformation of local rice varieties and germplasm innovation.

Genetic transformation is a core technology for wheat molecular breeding. Transformation efficiency and genotype compatibility determine the pace of targeted genetic improvement. Mature embryos are readily accessible but are associated with low transformation efficiency and strong genotype dependence. Using mature wheat embryos as explants, we screened eight wheat varieties representing three ecological types to develop genotype-adapted tissue culture systems. Treatments A5 (4 mg/L 2,4-D + 2 mg/L Dicamba), A7 (4 mg/L 2,4-D + 6 mg/L Dicamba), and A8 (4 mg/L 2,4-D + 8 mg/L Dicamba) were optimal for callus induction in winter, semi-winter, and spring wheat, respectively. WF3 (a 1/2 MS-based medium) was broadly applicable to callus differentiation, whereas IBA produced the highest rooting efficiency in regenerated plantlets. The optimal transformation conditions comprised ultrasound treatment (2.5 min) followed by vacuum infiltration (5 min), supplementation of the infection solution with PEG₆₀₀₀ (250 mg/L) and Tween20 (0.01%), and an OD₆₀₀ of 0.6–0.8. Transformation efficiencies using mature embryos of the semi-winter wheat varieties YX288 and ZM578 were 2.5% and 1.8%, respectively. Using this transformation platform, the dwarfing genes Rht12 in ZM578 and Rht8 in YX288 were targeted by CRISPR/Cas9-mediated editing. Multiple single-base substitutions at the target sites were detected in ZM578 rht12 mutants, which exhibited significantly reduced height, increased tiller number, and a compact plant architecture. However, no successfully edited rht8 lines were obtained in YX288. These findings indicate that this study established a reliable mature-embryo transformation system for wheat and successfully validated gene editing in the semi-winter variety ZM578. The system provides a technical platform for the functional analysis of key genes and molecular breeding in bread wheat.