Agrobacterium을 이용한 油菜(brassica napus)의 形質轉換

Transformation of brassica napus using agrobacterium

The addition of AgNO₃(ethylene inhibitor) in MS medium remarkably enhanced the shoot regeneration from the cotyledonary petioles. The maximum frequency of shoot regeneration(74.2%) was obtained from the cotyledonary petioles cultured in the medium with 3㎎/ℓ AgNO₃. The presence of AgNO₃(3㎎/ℓ) also enhanced the shoot regeneration from the calli of hypocotyl and cotyledon tissues. The cotyledonary petioles cocultivated with Agrobacterium vectors for 72hours were transferred to MS medium with 0.5㎎/ℓ NAA, 2.0㎎/ℓ BA, 30㎎/ℓ kanamycin, 100㎎/ℓ cefotaxime, 30g/l sucrose, 3㎎/ℓ AgNO₃ and 2g/l gelrite. The cotyledonary petioles transferred to the selection medium formed green shoot with 17.5% efficiency. The putative transgenic plants were also obtained from the hypocotyl- and cotyledon-derived calli of B. napus. Southern blot analysis was done on the putative transformed plants’ genomic DNA digested with EcoR I and Hpa I restruction endonucleous. Agrobacterium-mediated transformation in B. napus was confirmed by hybridization with rol C and NPTⅡ gene(1.0kb and 2.0kb) as probes.