Interspecific hybrids between Diospyros glandulosa (2n = 2x = 30) and D. kaki cv. Jiro (2n = 6x = 90) were produced by electrofusion of protoplasts. Protoplasts were isolated from calli derived from leaf primordia, fused electrically, and cultured by agarose-bead culture using modified KM8p medium. Relative nuclear DNA contents of calli derived from fusion-treated protoplasts were determined by flow cytometry. One-hundred-forty-nine of 166 calli obtained had the nuclear DNA content of the sum of those of D. glandulosa and D. kaki cv. Jiro. RAPD analysis showed that the 149 callus lines yielded specific bands for both D. glandulosa and D. kaki cv. Jiro and they appeared to be interspecific somatic hybrid calli. Shoots were regenerated from 63 of the 149 interspecific hybrid calli. PCR-RFLP of chloroplast DNA analysis, flow cytometric determination of nuclear DNA content, and RAPD analysis revealed that the 63 interspecific hybrid shoot lines contained nuclear genome from both the parents but only chloroplast genome from D. glandulosa. Microscopic observation of root tip cells confirmed that somatic chromosome numbers of the interspecific hybrids were 2n = 8x = 120.
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