Annual Report 2003

Plant regeneration system through morphogenesisto develop the genetic transformationmethod in Japanese commercialvariety of wheatTakashi HagioPlant Biotechnology DepartmentMore than 90% frequency of plant regenerationwas achieved from tissue cultures ofwheat immature embryos on revised culturemedia using wheat lines selected for favorabletissue culture response.Twenty-three Japanese commercial varietiesof wheat were examined for theirresponse in tissue culture, and the tissue culturesystem was optimized. The cultureswere initiated from immature embryos takenapproximately two weeks after flowering.Regeneration occurred at higher frequencieswhen cytokinins (kinetin or BAP) had beenadded in the callus induction medium (modifiedMS medium), followed by regenerationmedium devoid of growth regulators. Theaddition of maltose, proline glutamine andasparagine also enhanced shoot formation,but the addition of cytokinins to regenerationmedia did not improve shoot formation. Inmany cases, shoots were formed as adventitiousbuds and distinct somatic embryogenesiswas not observed (Fig. 1). Five varieties(Norin No. 35, Norin No. 40, Norin No.67,Norin No.81, Norin No. 83) showed a highfrequency of callus formation and plantregeneration. Self-fertilized seeds of “NorinNo.67” were obtained and advanced to thenext two generations (R1 and R2). TissueFig. 2Frequency change of shoot regeneration from cultured immatureembryos in the R0, R1 and R2 generations of wheat.Fig. 1Plant regeneration of wheat calli on the revised medium.Fig. 3Transgenic wheat plants which showed positive response in Southernanalysis.Annual Report 2003 45