Agrobacterium-mediated Transformation of Kentucky Bluegrass
CHAI Bao-Feng, LIANG Ai-Hua, WANG Wei, Hu Wei ( Institute ofHydrobiology, The Chinese Academy of Sciences, Wuhan 430072, China; ( Institute of Biotechnology, Shanxi University, Taiyuan 030006, China )
Embryogenic calli of Kentucky bluegrass, named Md, were induced from mature seeds and embryos, and proliferated on medium K3 containing 2,4-dichlorophenoxyacetic acid (2,4-D, 10.0μmol/L), 6-benzylaminopurine ( BAP, 0.5μmol/L) and K5 which was the K3 medium supplemented with cupric sulfa (0.5 μmol/L) under dim-light condition ( 20-30 μmol·m-2·s-1 , 16 h light) at 24℃. Embryogenic calli were transformed with plasmids pDM805 Carring bar and gus genes, Which was mediated by an Agrobacterium strain AGL1, four transgenic lines were obtained. The important factors that affect the transformation efficiency and obtain desirable number of transgenic plants included: (1) the quality of embryogenic calli; (2) light condition and time of co-cultivation; (3) concentration of antibiotics used for suppressing the overgrowth of Agrobacterium in the course of transformed plant regeneration; (4) selection pressure, etc. The micro nutrient of cupric had significant influence on the quality of embryogenic calli. This presentation is the first successful protocol of Kentucky bluegrass transformation mediated by Agrobacterium.