Regeneration and characterization of transgenic kumquat plants containing the Arabidopsis APETALA1 gene

'Meiwa' kumquat (Fortunella crassifolia Swingle.) is famous for its relatively short juvenility, delicious flavor, human health benefits and high resistance to citrus canker. To establish kumquat transformation system and to further shorten its juvenility, Agrobacterium-mediated epicotyledon segment transformation of APETALA1 (AP1 from Arabidopsis) gene was conducted. Transformation efficiency ranged from 1.00 to 4.08% depending on seedling age, and 20 day age seedlings proved to be the best explants for transformation. Five stable transgenic plants were obtained as revealed by GUS assay, and further confirmed by specific PCR and Southern blot analyses. After transfer to the greenhouse, one transgenic line (J3) flowered at the 11th month and continued to flower in the next years, till the third year when all non-transformed and transgenic plants but J66 flowered. Gene expression analysis of AP1 and four endogenous flowering genes CiAP1, CiFT, CiLFY, and CiTFL1 by real-time RT-PCR suggested that CiFT and CiTFL1 played important roles in the regulation of flowering in transgenic AP1 kumquat. Factors influencing kumquat transformation efficiency and the relationship between flowering time in transgenic AP1 kumquat and expression levels of endogenous FT and TFL1 genes were discussed.