Intergeneric somatic hybrids of rice [Oryza sativa L.(+) Porteresia coarctata (Roxb.) Tateoka]

Abstract

Somatic hybrid plants were obtained following the electrofusion of rice (Oryza sativa L. cv 'Taipei 309', 2n = 2x = 24) cell suspension-derived protoplasts with non-dividing leaf protoplasts of Porteresia coarctata (2n = 4x = 48), a saline-tolerant wild species. Fusion-treated protoplasts were plated on the surface of cellulose nitrate filter membranes, overlaying Lolium multiflorum nurse cells. The nurse cells were embedded in KPR medium containing 0.5 mg l^-1 2,4-dichlorophenoxyacetic acid and semi-solidified with SeaPlaque agarose. Putative somatic hybrid cell colonies were selected on the basis of their growth, whereby faster growing colonies were transferred preferentially to MS-based medium with 2.0 mg l^-1 kinetin, 0.5 mg l^-1 α-naphthaleneacetic acid, 30 g l^-1 sucrose and 4.0 g l^-1 SeaKem agarose to induce shoot regeneration. One hundred and nineteen regenerated plants were micropropagated clonally on MS-based medium containing 2.0 mg l^-1 6-benzylaminopurine, 50 g l^-1 sucrose and 4.0 g l^-1 SeaKem agarose, prior to DNA extraction of plant samples. Putative somatic hybrids were initially identified by RAPD analysis, and 8 plant lines were selected for further investigation by flow cytometric ploidy determination and cytology. Plants of one line had an allohexaploid chromosome complement (2n = 6x = 72) and, following examination of its vegetative clones by GISH, were confirmed as somatic hybrids containing full chromosome complements of both O. sativa and P. coarctata.