Background: Cucumber (Cucumis sativus) is one of the most important vegetable crops in the world. As conventional breeding of cucumber is very challenging, genetic engineering is an alternative option to introduce important traits such as enhanced stress resistance and nutritional value. However, the efficiency of the transformation system depends on genotypes, transformation conditions, selection agents, etc. This study aims to speed up the process of Agrobacterium-mediated transformation of cucumber. ‘Xintaimici’, a very popular and typical north China-type cucumber variety, was transformed with Agrobacterium GV3101. The strain carried the pCAMBIA2300s plasmid, a double vector with the marker gene neomycin phosphotransferase II (npt II).
Results: The research results indicated that cefotaxime sodium was suitable for inhibiting Agrobacterium in the screening and bud elongation stages. Timentin was best used during the rooting stage. Furthermore, 25 mg/L kanamycin was used in the early stage of screening and increased to 50 mg/L for further screening. At the bud elongation and rooting stages, 75 and 100 mg/L kanamycin was used, respectively, to improve the screening efficiency. To obtain the highest regeneration frequency of resistant buds, 50, 150, and 100 μM acetosyringone was added in the pre-culture medium, infection solution, and co-culture medium, respectively. To confirm the presence of the transgenes, DNA from npt II transgenic cucumber plants was analysed by polymerase chain reaction after transplanting resistant regenerated plants.
Conclusions: We finally achieved an 8.1% conversion, which is among the highest values reported to date using the cucumber ‘Xintaimici’. Thus, an effective protocol for Agrobacterium tumefaciens-mediated genetic transformation of cucumber was optimized.