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Somatic hybrids were obtained by the symmetric fusion of embryogenic callus cells from tetraploid 'Mame' kumquat [Fortunella hindsii (Champ.) Swing.] and mesophyll cells from diploid trifoliate orange [Poncirus trifoliata (L.) Raf.]. Southern blot analysis of three regenerants revealed that they carried specific rDNA fragments from both fusion partners, thereby confirming their hybridity. In contrast, mitochondrial DNA (mtDNA) and chloroblast DNA (cpDNA) were unidirectionally transmitted from the callus parent without any evidence of recombination. No differences in the restriction fragment patterns of rDNA, mtDNA or cpDNA could be detected among the regenerants. Flow cytometry showed that two regenerants were hexaploids, as expected, but that one was pentaploid, probably due to elimination of chromosomes prior to the regeneration of this plant.

Protoplast fusion in higher plants is a useful technique for production of novel plants which cannot be obtained by conventional breeding methods. For more than a decade, successful somatic hybridization was limited to herbaceous plants. In Citrus, the use of nucellar callus obtained from ovules, possessing high regeneration ability, made it possible to regenerate whole plants from protoplasts. Then, somatic hybridization in the Rutaceae family was established, and many intergeneric or interspecific amphidiploid somatic hybrid plants were produced. Recently developed molecular biology techniques have provided more direct methods for the identification of somatic hybrids in both the nucleus and cytoplasm, and the results are expected to provide useful information about nuclear‐cytoplasmie interaction. Some somatic hybrids have flowered and produced fruit, and their male and female fertility has been confirmed. Somatic hybridization in Citrus will undoubtedly be a useful technique for overcoming ba...

Citrus and Poncirus belong to the family Rutaceae. Citrus includes many commercially important fruit species such as sweet orange (C. sinensis), mandarin (C. reticulata), lemon (C. limon), grapefruit (C. paradisi), and is grown in over 100 countries in tropical and subtropical areas spreading over approximately 40° latitude. On the other hand, Poncirus includes only one species of trifoliate orange. (P. trifoliata), which is a deciduous shrub widely grown in Japan and China. It is an important rootstock which possesses cold hardiness, and is resistant to phytophthora, nematode, and tristeza virus (Saunt 1990). The morphology of Citrus and Poncirus is very different; however, their genomes are similar and the chromosome number is 2n = 18. These two genera are sexually compatible and the most widely grown sexual hybrid is Troyer citrange, which has become a very important rootstock in many countries.

Somatic hybrid plants of Rutaceae were obtained by protoplast fusion between Citrus sinensis Osb. ('Trovita' orange) and Poncirus trifoliata. Protoplasts isolated from embryogenic cells of C. sinensis and from leaves of P. trifoliata, and the culture of fusion products in the presence of high concentrations of sucrose were essential requirements for the selection of hybrids. Green globular embryoids derived from protoplasts resulted in the regeneration of trifoliate plants. Other morphological characters of these plants were intermediate between both parents. The chromosome number in one of the hybrid plants was 36, which was the sum of C. sinensis (2n=18) and P. trifoliata (2n=18). EcoRI restriction analysis of rDNA confirmed the presence of parental nuclear DNAs in the hybrid.