The Study on the Effect of Anther Orientation, Type and Composition of Culture Medium and Ovary Co-culture on Callus Induction from Cultured Anthers of Pumpkin (Cucurbita pepo var. Styriaca)

The production of doubled haploid cucurbit plants can accelerate the breeding programs of these species. In this research, anther culture of Cucurbita pepo var. styriaca was studied to produce callus and haploid embryos. Cytological studies showed that 15 mm male buds containing 9-10 mm anthers in size and microspores at mid to late-uninucleate were desirable for anther culture experiments. Three experiments were conducted as factorial based on completely Randomized Design (CRD) with three replications. In first experiment, the effect of anther orientation on culture medium as well as type and concentration of the plant growth regulators of NAA (0, 0.5 and 1 mgl-1) and 2,4-D (0, 1 and 2 mgl-1) on callogenesis percentage of pumpkin anther cultures were studied. Results showed that the culturing of anthers from convex side in a culture medium supplemented with 2 mgl-1 2,4-D and 1 mgl-1 NAA resulted in the highest percentage of callogenesis. In second experiment, the interaction effects of medium type (B5 and E20) and combination treatments of 2,4-D (0, 2.5 and 5 mgl-1) and BAP (0, 0.5 and 1 mgl-1) on callogenesis percentage were studied. E20 culture medium supplemented with 2.5 mgl-1 2,4-D and 1 mgl-1 BAP produced the highest percentage of callogenesis compared to other treatments. In third experiment, the effect of pumpkin and wheat ovary conditioned medium on callogenesis percentage from pumpkin anthers were studied in two E20 culture media (Solid and liquid). In this experiment, co-culturing of pumpkin anthers with wheat ovaries in solid culture medium resulted in the highest percentage of callogenesis compared to other treatments.