Induction of 2n pollen formation in Begonia by trifluralin and N2O treatments

Abstract

In this study, treatments of both trifluralin (at 10, 100 and 1000 mu M) and N2O (in the form of gas under pressure) were applied to Begonia flower buds to induce the formation of 2n pollen. Three male fertile species (B. cucullata, B. subvillosa var. leptotricha and B. fischeri) and two male sterile hybrids (B. schmidtiana x B. cucullata and B. subvillosa var. leptotricha x B. cucullata) were treated. Pollen size, which is related to pollen DNA content, increased after both N2O and trifluralin treatments, but the induction of large pollen was genotype dependent. Trifluralin induced large pollen only in the male fertile species, while N2O treatments induced fertile 2n pollen in the male sterile B. schmidtiana x B. cucullata. Cytological studies showed that trifluralin induced multinuclear monads that resulted in 4n gametes in stead of 2n gametes. In general, large pollen obtained after trifluralin treatments showed low germination capability, while large pollen obtained after N2O treatments retained high germination capability. Seedlings with raised ploidy level could only be obtained after crosses were performed with large pollen obtained from N2O treatments. Hence, N2O treatments are preferable to the use of trifluralin to induce 2n gametes in Begonia

Use of 2n Gametes in Plant Breeding

“…Recently different attempts, with variable success, were made to induce these gametes in any genotype of interest. An overview of the methods used to induce 2n gametes is given in Dewitte et al, 2010b;Okazaki et al, 2005). N 2 O is suitable for treating organs inside tissues as the gas simply permeates the tissue, thereby protecting the tissues from harmful after-effects as soon as the gas is released (Kato & Geiger, 2002;Östergren, 1954).…”

“…N 2 O treatments are performed in a pressure tolerant vessel at 6 bar for 24 or 48h on bulbs (when microsporogenesis occurs inside the bulb) or flower buds at the stage of meiosis. In the latter case, the stage of meiosis can be estimated by the size of the flower bud (Akutsu et al, 2007;Barba-Gonzalez et al, 2006;Dewitte et al, 2010b). Consequently, bud size has been used as a criterium to start N 2 O treatments.…”

“…Akutsu et al (2007) showed that effects were optimal when treatments started during pollen mother cell progression to metaphase I. By applying this technique, male sterile hybrids may produce fertile 2n gametes after treatments, but the efficiency of the treatment seems to be genotype specific (Barba-Gonzalez et al, 2006;Dewitte et al, 2010b). In Begonia for example, viable 2n pollen could be induced in the male sterile hybrid B. schmidtiana x B.cucullata in 14 of the 49 treated flowers, while no 2n pollen were induced in B. fischeri with a similar number of flowers treated.…”

“…Furthermore, Kitamura et al (2009) showed in Lilium that microtubules were depolymerised during metaphase I, which prevented chromosomes from moving to the poles. Dewitte et al (2010b) tried to induce 2n pollen by submerging flower buds of Begonia in a trifluralin solution. Their treatments resulted in a disturbed meiosis and finally in 4n gametes but no progeny with raised ploidy level could be obtained after crosses with these gametes.…”

Phenotypic and genome size changes (variation) in synthetic tetraploids of daylily (Hemerocallis) in relation to their diploid counterparts

“…In meiotic (sexual) polyploidisation, 2n gametes are induced (usually 2n pollen) by arresting the mitotic process. 2n pollen (induced with nitrous oxide gas) has recently been used for crossing with diploid cultivars in order to obtain triploids of tulip, lily, and begonia (Okazaki et al 2005;BarbaGonzalez et al 2006;Dewitte et al 2010). The natural sexual polyploids (mainly triploids) that originate through the functioning of 2n gametes (produced by almost all species in some frequencies) have been considered an important factor in flowering plant evolution (Osborn et al 2003;Ramanna and Jacobsen 2003;Younis et al 2014).…”