Abstract
Agrobacterium-mediated transformation factors for sweet potato embryogenic calli were optimized using β-glucuronidase (GUS) as a reporter. The binary vector pTCK303 harboring the modified GUS gene driven by the CaMV 35S promoter was used. Transformation parameters were optimized including bacterial concen-tration, pre-culture period, co-cultivation period, immersion time, acetosyringone (AS) concentration and mannitol treated time. Results were obtained based on the percentage of GUS expression. Agrobacterium tumefaciens strain EHA105 at concentration OD600 nm = 0.8 showed the highest virulence on sweet potato embryogenic callus. Four days of pre-culture, four days of co-cultivation, 10 min of immersion, 200 μM acetosyringone and 60 min of mannitol-treated embryogenic callus gave the highest percentage of GUS positive transformants. Key words: Agrobacterium-mediated, transformation parameters, sweet potato embryogenic callus, β-glucuronidase.
Highlights
The sweet potato (Ipomoea batatas (L) Lam.) is the sixth most important crop in the world, right after wheat, rice, corn, white potato and barley (Vietmeyer, 1986)
Agrobacterium- mediated is the most commonly used system which involves the delivering of DNA from Agrobacterium plasmid into cells of a wide variety of dicotyledonous and some monocotyledonous plants, and the Agrobacterium-based transformation method possess several advantages over other forms of transformation, including: (1) the ability to transfer large segments of DNA with minimal rearrangement, (2) the precise insertion of transgenes resulting in fewer copies of inserted genes, and (3) simple technology with lower cost (Binns, 1990)
Cell transformation was influenced by several factors such as bacterial concentration, pre-culture period, co-cultivation period, immersion time, AS concentration and mannitol treated time
Summary
The sweet potato (Ipomoea batatas (L) Lam.) is the sixth most important crop in the world, right after wheat, rice, corn, white potato and barley (Vietmeyer, 1986). Sweet potato has some advantage, such as its clonal propagation, ease of cultivation and high productivity of storage roots and foliage, male sterility, incompatibility and a hexaploid genome make it difficult to improve by conventional. There are several methods vailable for transferring desired genes into plant genome such as Agrobacterium-mediated method, micro-projectile bombardment and electroporation Among these methods, Agrobacterium- mediated is the most commonly used system which involves the delivering of DNA from Agrobacterium plasmid into cells of a wide variety of dicotyledonous and some monocotyledonous plants, and the Agrobacterium-based transformation method possess several advantages over other forms of transformation, including: (1) the ability to transfer large segments of DNA with minimal rearrangement, (2) the precise insertion of transgenes resulting in fewer copies of inserted genes, and (3) simple technology with lower cost (Binns, 1990)
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