Agrobacterium-mediated genetic transformation of California poppy, Eschscholzia californica Cham., via somatic embryogenesis

An efficient Agrobacterium-mediated protocol for the stable genetic transfo rmation of Eschscholzia californica Cham. (California poppy) via somatic em bryogenesis is reported. Excised cotyledons were co-cultivated with A. tume faciens strain GV3101 carrying the pBI121 binary vector. Except for the co- cultivation medium, all formulations included 50 mg 1(-1) paromomycin as th e selective agent and 200 mg 1(-1) timentin to eliminate the Agrobacterium. Four to five weeks after infection, paromomycin-resistant calli grew on 80 % of explants in the presence of 2.0 mg 1(-1) l-naphthaleneacetic acid (NA A) and 0.1 mg 1(-1) 6-benzylaminopurine (BAP). Calli were cultured on somat ic embryogenesis induction medium containing 1.0 mg 1(-1) NAA and 0.5 mg 1( -1) BAP, and somatic embryos were visible on 30 % of the paromomycin-resist ant calli within 3-4 weeks. Three to four weeks after the somatic embryos w ere transferred to phytohormone-free plant regeneration medium, 32 % conver ted to paromomycin-resistant plants. Detection of the neomycin phosphotrans ferase gene and high levels of beta -glucuronidase (GUS) mRNA and enzyme ac tivity, and the cytohistochemical localization of GUS activity in all plant tissues confirmed the integrative transformation of the regenerated plants . The normal alkaloid profile of California poppy was unaffected by the tra nsformation process; thus, the reported protocol could serve as a valuable tool to investigate the molecular and metabolic regulation of the benzophen anthridine alkaloid pathway.