Submitted to: Congress on In Vitro Biology
Publication Type: Abstract Only
Publication Acceptance Date: 6/4/1996
Publication Date: N/A
Technical Abstract: Genetic transformation of rice (Oryza sativa L.) mediated by A. tumefaciens has been confirmed for japonicqa varieties and extended to include the more recalcitrant indica varieties. Immature embyos were inoculated with either A. tumefaciens At656 (pCNL56) or LBA4404 (pTOK233). Experimental conditions were developed initially for immature embryos treated with strain At656, based upon both transient and stable GUS activities. However, plant regeneration following selection of G418 (pCNL56 contained the nptII gene) did not occur. Using the same protocol, but inoculating immature embryos of rice with LBA4404 (pTOK233) resulted in efficient (27%) production of transgenic plants of the japonica variety, Radon, and an acceptable efficiency (from 1-5%) for indica varieties IR72 and TCS10. Transformation was based upon resistance to hygromycin, presence of GUS activity (from the gusA gene containing an intron), Southern blots for detection of the integrated gusA gene and transmission of GUS activity to progeny in a Mendelian 3:1 segregation ratio. Southern blots indicated two to three copies of the gene integrated in most transformants. Transgenic plants of both japonica and indica varieties were self fertile and comparable in fertility to seed grown plants. Key factors in the transformation of rice by A. tumefaciens were use of embryos as the explant, use of hygromycin as the selection agent, the presence of extra copies of certain vir genes on the binary vector pTOK233, and maintaining high concentrations of acetosyringone for inducing the vir genes during cocultivation of embryos with Agrobacterium. Preliminary results on wheat transformation using LBA4404 (pTOK233) will be presented.