Figure 7. Development of microcalli expressing GFP.

Five days after DNA microinjection, the alginate layer was transferred to Y3A liquid medium comprising 5.5% (w/v) sucrose and 8.2% (w/v) glucose supplemented with 10 µM NAA, 2 µM 2,4–D, 2 µM IBA, 2 µM GA₃, 2 µM 2iP and 200 mg/l ascorbic acid and cultured at 28°C for 2 weeks. The medium was then replaced with similar Y3A liquid medium comprising 4% (w/v) sucrose and 7.2% (w/v) glucose to allow the development of microcolonies (A and B, after 2 months). The medium was then replaced with Y3A liquid medium comprising 4% (w/v) sucrose to promote the conversion of microcolonies (C and D, after 4 months) into microcalli (E and F, after 6 months). Finally, the alginate layer containing microcalli (G) was transferred onto Y31N0.1BA solid medium (H) for the regeneration of oil palm plants. Arrows indicate the injected protoplasts. The transformation efficiencies represent the mean of three replicates. Scale bar  = 100 µm in (A)–(F), 1 cm in (G) and (H).