Indução de plantas haplóides in vitro de Allium cepa L. a partir de ovários não fertilizados.

Abstract

Hybrid production is considered one of the main objectives of onion breeding programs. However, the time required to produce hybrids from inbred lines by conventional methods is a limiting factor. One way to overcome this problem, efficiently, has been done using the doublehaploid (DH) technique. The haploid embryos regenerated by gynogenesis, are converted into double-haploid plants, and later, are crossed to produce hybrids. Thus, the objective was to induce the formation of haploid plants in vitro of Allium cepa L. from unfertilized ovaries. Initially, unfertilized ovaries of the IPA-12 cultivar were inoculated in BDS culture medium (Dunstan and Short, 1977), supplemented with 100 gL-1 of sucrose, 200 mg.L-1 of proline, 500 mg.L-1 of myoinositol, 10 mg.L-1 of adenine sulfate and vitamins B5, and under different combinations of growth regulators 2,4-D (0, 1,0 and 2,0 mg.L-1) and BAP (0 , 1.0 and 2.0 mg.L-1). The pH of the medium was adjusted to 5.8 before adding 6 g.L-1 of agar, then autoclaved for 15 minutes at 121 ° C. Twenty unfertilized ovaries were inoculated into each Petri dish, and the calluses and embryos formed were collected and transferred to medium strength MS medium, supplemented with 1.0 mg.L-1 of IBA + 1.0 mg.L-1 of KIN. After 3-4 weeks, the seedlings were transferred to commercial PlantMax® substrate for acclimatization. The experiment was conducted in a completely randomized design, in a factorial scheme 32, the treatments consisted of the combination of doses of 2,4-D and BAP, with 9 treatments and 25 repetitions each. The variables analyzed were the number of explants that formed calluses; percentage of callus formation (%); number of explants that formed embryos and percentage of embryo formation (%). The data were subjected to analysis of variance and when there were significant differences, the means were compared by the Tukey test at the level of 5% probability. Callus production was observed from the fourth week of ovary inoculation. Altogether 411 calli and 69 embryos were formed, corresponding to 22.83% and 5.4%, respectively. Higher number of haploid seedlings were obtained in BDS medium free of growth regulator, while the highest percentage of calluses was obtained when the BDS medium was supplemented with 1.0 mg.L-1 of 2.4-D + 2.0 mg .L-1 of BAP. The IPA-12 genotype has a high gynogenic capacity, and it is not necessary to use growth regulators to induce haploids. The ploidy level will be determined by flow cytometry.