Production of mandarin plus pummelo somatic hybrid citrus rootstocks with potential for improved tolerance/resistance to sting nematode

Sting nematode (Belonolaimus longicaudatus Rau) has become a primary factor limiting citrus production in localized regions of the central Florida sandridge citrus production area, making the development of resistant rootstocks a new breeding objective. In efforts to develop a replacement rootstock for the widely adapted sour orange, our focus has been on somatic hybridization of selected mandarin + pummelo combinations [Grosser, J.W., Gmitter, Jr., F.G., 1990. Protoplast fusion and citrus improvement. Plant Breed. Rev. 8, 339-374; Ananthakrishnan, G., Calovic, M., Serrano, P., Grosser, J.W., 2006. Production of additional allotetraploid somatic hybrids combining mandarins and sweet oranges with pre-selected pummelos as potential candidates to replace sour orange rootstock. In Vitro Cell. Dev.: Plant 42, 367-371], since sour orange is probably an introgression hybrid of mandarin and pummelo as suggested by molecular marker analyses [Nicolosi, E., Deng, Z.N., Gentile, A., La Malfa. S., Tribulato, E., 2000. Citrus phylogeny and genetic origin of important species as investigated by molecular markers. Theor. Appl. Genet. 100. 1155-1166-, Gulsen, O., Roose, M.L., 2001. Lemons: diversity and relationships with selected Citrus genotypes as measured with nuclear genome markers. J. Am. Soc. Hort. Sci. 126, 309-317]. Somatic hybrid plants were produced from four new mandarin (C reticulata Blanco) + pummelo (C. grandis L. Osbeck) parental combinations by fusing embryogenic suspension culture-derived protoplasts isolated from selected mandarins with leaf protoplasts of pummelo seedlings previously selected for tolerance/resi stance to the sting nematode (B. longicaudatus Rau) as follows: Amblycarpa mandarin + 'Liang Ping Yau' (seedling) pummelo seedling SN7; Amblycarpa mandarin + 'Hirado Buntan Pink' (HBP) pummelo seedling SN3; Murcott tangor + pummelo seedling SN3; and Shekwasha mandarin + pummelo seedling SN3. Somatic hybridization was verified by ploidy