Effect of Microspore Density and Embryo Size on Embryogenesis, Rooting, Shooting and Secondary Embryogenesis in Rapeseed (Brassica napus L. cv. Global)

The aim of this study was production of haploid plants, using microspore culture method, in to be used in rapeseed breeding programs. Donor plants (cv. Global) were grown at a day/night temperature of 15/10˚C (16/8 hours). After 90 days microspores were isolated from buds 2.5-3.5 mm containing microspores at late-uninuclate to early-binucluate stages and cultured in a modified medium (NLN-13). Samples were incubated at 30˚C and in darkness for 14 days then transferred to a shaker at 25˚C for 20 days. In this experiment, the effect of different microspore densities included of: 60000, 40000 and 20000 microspores per ml, was used to study the embryogenesis of microspores. Indeed, effect of embryo size (2-3, 3-4, 4-5, 5-6 and greater than 6 mm) on seedling germination properties like shoot formation, secondary embryogenesis and root formation were also studied. Results showed significant differences between various culture densities (p<0/01). The density of 60000 microspores produced 1031 embryos in each petri dish (containing 12/5 ml culture medium) and was the best density in this study. The size of embryo showed significant effects on shoot and root formation at 1% of probability level. The size of embryo also showed significant differences on secondary embryogenesis at 5%. Probably level in this expriment, it was observed that 73.33 % of embryos greater than 6 mm produced normal shoot had the best form of germination. In this study 43% of embryos were also regenerated.