N2O induces mitotic polyploidization in anther somatic cells and restores fertility in sterile interspecific hybrid lilies

Abstract

Fertile plants undergoing male gametogenesis can be treated with nitrous oxide (N2O) gas to obtain 2n male gametes. N2O treatment is also expected to restore the fertility of interspecific hybrids through meiotic restitution or mitotic amphidiploidization. However, this technique has few applications to date, and it is un-known how N2O treatment restores fertility in sterile hybrids. To establish optimal N2O treatment conditions and determine its cytological mechanism of action, we treated various sized floral buds with N2O gas at different anther developmental stages from fertile and sterile hybrid lilies. N2O treatment using the optimal 1–4 mm floral buds induced mitotic polyploidization of male archesporial cells to produce 2n pollen in fertile hybrid lilies. In sterile hybrid lilies, N2O treatment doubled the chromosome number in male archesporial cells followed by homologous chromosome pairing and normal meiosis in pollen mother cells (PMC), resulting in restoration of pollen fertility. Backcrossing the resultant fertile pollen to Lilium × formolongi produced many triploid BC1 plants. Thus N2O treatment at the archesporial cell proliferating stage effectively overcame pollen sterility in hybrid lilies, resulting in fertile, 2n pollen grains that could produce progeny. The procedure presented here will promote interspecific or interploidy hybridization of lilies.

Breeding of lilies and tulips—Interspecific hybridization and genetic background—

“…In such a sterile hybrid (allopolyploid), e.g. , Oriental Trumpet hybrid ‘Yelloween’, the N 2 O treatment can be optimally applied to anthers in 1–6 mm buds that contain the proliferating PMCs ( Nukui et al . 2011 ).…”

The importance of reproductive barriers and the effect of allopolyploidization on crop breeding

“…Hybrid sterility can often be restored by zygotic or somatic chromosome doubling through cell cycle disruption by antimitotic agents such as colchicine, oryzalin, or trifluralin ( Dhooghe et al 2011 ). Nitrous oxide (N 2 O) is also applied as a polyploidizing agent in lilies ( Nukui et al 2011 ).…”

Multiple Mechanisms and Challenges for the Application of Allopolyploidy in Plants

“…[ 64 ]. Nitrous oxide (N 2 O) has been applied to zygotes [ 69 ], seedlings [ 70 ], pollen mother cells [ 71 , 72 ], and anther somatic cells [ 73 ], as a polyploidizing agent in lieu of colchicine treatment. Nitrous oxide may act through depolymerization of microtubules [ 74 ].…”

“…Additionally, the gas is expected to be rapidly dissipated from treated tissues when the pressure is released, thereby preventing further harmful after-effects. For these reason nitrous oxide has been applied as a polyploidizing agent to overcome hybrid sterility in lilies [ 73 , 75 ].…”