Agrobacterium-mediated genetic transformation and plant regeneration of sweetpotato (Ipomoea batatas) = Agrobacterium 매개에 의한 고구마 형질전환 및 식물체 재분화

Abstract

An efficient and rapid Agrobacterium-mediated transformation method based on de novo (via callus) organogenesis has been developed from petioles with leaf for sweetpotato (Ipomoea batatas (L.) Lam). Stable transgenic sweetpotato plants cv. Jewel were obtained in six to ten weeks after infection with Agrobacterium tumefaciens hyper-virulent strain EHA105 harboring a binary vector pCIP45 bearing the nptII gene conferring resistance to kanamycin and a gene of interest. PCR and Southern analyses confirmed stable integration of both genes into the sweetpotato genome. The expression of the nptII gene was assessed by reverse-transcribed PCR and callus development in a high kanamycin medium. A two-step organogenesis regeneration using media containing 4-fluorophenoxyacetic acid (4-FA) and zeatin was used in two independent transformation experiments yielding 20% and 10% transformation efficiency, respectively. When using indolacetic acid (IAA) in regeneration media, the transformation efficiency dropped to 4.0%. It indicated an auxin to cytokinin treatment could improve the regeneration of transgenic calluses. This rapid organogenesis-based transformation strategy represents an important improvement over existing methods and will facilitate producing large-scale transgenic sweetpotato plants the genetic improvement of a crop that is reputed to be difficult to transform.