Efficient Agrobacterium tumefaciens-mediated transformation of Malus zumi (Matsumura) Rehd using leaf explant regeneration system
Yu Zhen Wang
Heng Xia Yin
Xiao Jing Liu*
Financial support: The research was supported by the Knowledge Innovation Program of the Chinese Academy of Sciences (Grant no. 0707013603).
Key words: Agrobacterium tumefaciens, apple rootstock, genetic transformation.
Malus zumi is known as an excellent dwarfing apple rootstock occurring in natural or arid/semiarid soil or salina. Gene manipulation of M. zumi through transgenic technology can modify plant feature for further improvement fruit tree production by grafting the scion on a transgenic rootstock. Here, we report the establishment of an efficient, in vitro, shoot regeneration system and Agrobacterium tumefaciens- mediated transformation from the leaf explants for Malus zumi (Matsumura) Rehd. Leaf explants were infected with Agrobacterium strains containing nptII and gus gene. The highest frequency of shoot regeneration was obtained on MS medium containing 500 mg l-1 Lactalbumin hydrolysate, 30 g l-1 fructose, supplemented with 3.0 mg l-1 BA, 0.2 mg l-1 NAA.Using fructose instead of sucrose significantly increases the shoot regeneration and decreases vitrification. This regeneration procedure was incorporated into an Agrobacterium-mediated transformation procedure in M. zumi. Kanamycin was an efficient selective agent for selection. Pre-selection (5 days after co-cultivation) improved the transformation efficiency. The emergence of expected bands by PCR analysis and Southern blot in transgenic plantlets confirmed the transformation of foreign DNA into plant genome.