关键词: 橙, 原位转化, 苗龄, 外植体, 转化效率

Abstract: By using the Valencia sweet orange（Citrus sinensis‘Valencia’）seedlings as plant materials，the in planta transformation system was established and optimized. The influence of infection sites on in planta transformation efficiency was firstly investigated. Agrobacterium carrying the PBI121 vector was used to infect the epicotyl incision or apical bud incision（with/without true leaves）of the 5-week-old Valencia sweet orange seedling for 2 times. After three days’ co-culture，resistance screening and dark treatments were performed. Seven weeks later，leaf DNA of the candidate transgenic plants was isolated and was used as template for PCR detection to calculate the transformation rate. The effect of different seedling ages on the in planta transformation efficiency was further studied by using the epicotyl incision as the infection site. Results showed that the regeneration rate of 5-week-old seedling epicotyl incision was 93.10%，and the transformation rate was 20.68%. The apical bud regeneration rate for with/without true leaf was 81.74% or 94.55%，and the transformation rate was 19.09% or 17.82%. The regeneration rates of 4- and 6-week-old Valencia sweet orange seedling epicotyl incisions were 94.59% and 91.50%，respectively，and the transformation rates were 25.42% and 19.36%，respectively. By infecting the epicotyl incision of 4-week-old Valencia sweet orange seedlings，high in planta transformation rate could be obtained. The in planta transformation method is easier to be applied，less time consuming and more efficient. Thus，it is of great potential to be applied in sweet orange genetic breeding.

Key words: orange, in planta transformation, seedling age, explant, transformation efficiency