Induction of 2n pollen in tulips by arresting the meiotic process with nitrous oxide gas

Abstract

Triploid tulips have agronomically desirable traits such as vigorous growth and large flower size, but only a portion of all cultivated tulips is triploid. To apply 2n pollen to polyploid breeding of tulips, the polyploidizing agent, nitrous oxide gas (N 2 O), was applied to bulbs. In tulips, meiosis in anthers occurs inside the bulbs from mid-to late-October. When meiosis in anthers (excised from bulbs) reached metaphase I, we treated other bulbs of the same clones with N 2 O for 24-48 h. Most of the treated plants produced pollen grains with a wide-ranging or bimodal size distribution, indicating a mixture of n, 2n and aneuploid pollen grains. The use of pollen containing a relatively high proportion of giant pollen grains tended to yield larger numbers of triploids in the progeny. The number of giant pollen grains could be increased when N 2 O-treated pollen grains were suspended in 10% sucrose and then sieved through a nylon mesh. Very few polyploids were observed in some cross combinations, even those involving pollen with a relatively high proportion of giant grains. Even so, this low polyploid yield most likely is due to a triploid block, because the capsules obtained in the crosses of the diploid × N 2 O-treated plants contained some abnormal seeds, which were mostly triploid. Embryo culture was useful in rescuing abnormal embryos. The present study reveals that 2n pollen can be produced at high frequency using N 2 O during tulip breeding.

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).…”

“…Detailed studies on the exact mechanism of N 2 O mediated chromosome doubling during meiosis are also limited, but it has been suggested that N 2 O disrupts the spindle mechanism in both mitosis (Dvorak et al, 1973;Kato & Birchler, 2006;Kihari & Tsunewaki, 1960;Östergren, 1954) and meiosis (Akutsu et al, 2007;Barba-Gonzalez et al, 2006;Okazaki et al, 2005;). Consequently, aneuploidy is frequently reported after treatments.…”

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).…”

Occurrence of a Spontaneous Triploid Progeny from Crosses between Diploid Asparagus (Asparagus officinalis L.) Plants and Its Origin Determined by SSR Markers

“…The occurrence of spontaneous triploids from crosses between diploids was reported in some plant species. Triploids of Citrus (Chen et al, 2008) and carnation (Dianthus caryophyllus L.) (Nimura et al, 2008) were derived from the fertilization of 2n (female) and n (male) gametes, whereas those of tulip (Okazaki et al, 2005) and poplar (Populus tomentosa Carr.) (Zhang et al, 2007) were from the fertilization of n (female) and 2n (male) gametes.…”