D. Tjokrokusumo et al., Vacuum infiltration of Petunia hybrida pollen with Agrobacterium tumefaciens to achieve plant transformation, PL CELL REP, 19(8), 2000, pp. 792-797
Genetic transformation of Petunia hybrida with a reporter gene and selectab
le marker gene (35S-bar) was achieved in similar frequencies by pollinating
flowers with pollen vacuum-infiltrated with Agrobacterium tumefaciens or a
pplying a drop of Agrobacterium suspension to the stigma immediately prior
to pollination. Nine percent of the T-1, and 5% of the T-2 progeny germinat
ed in nutrient medium with 3 mgl/l Basta (R). Polymerase chain reaction ass
ays indicated that of the Basta (R)-resistant plants, 66% of the T-1 plants
, and 61% of the T-2 plants harboured the GUS gene. Histochemical assays sh
owed that 10% of the putatively transformed T-1 plants and 5% of their prog
eny expressed GUS in leaf tissue, pistils and young anthers. Southern hybri
dization confirmed genomic integration of the bar gene in one to three plac
es in selected T-1 and T-2 progeny.