Plant Cell Reports

, Volume 15, Issue 9, pp 672–676 | Cite as

Further evidence of a cybridization requirement for plant regeneration from citrus leaf protoplasts following somatic fusion

  • J. W. Grosser
  • F. G. GmitterJr.
  • N. Tusa
  • G. Reforgiato Recupero
  • P. Cucinotta


Somatic hybridization experiments in Citrus that involve the fusion of protoplasts of one parent isolated from either nucellus-derived embryogenic callus or suspension cultures with leaf-derived protoplasts of a second parent, often result in the regeneration of diploid plants that phenotypically resemble the leaf parent. In this study, plants of this type regenerated following somatic fusions of the following three parental combinations were analyzed to determine their genetic origin (nuclear and organelle): (embryogenic parent listed first, leaf parent second) (1) calamondin (C. microcarpa Bunge) + ‘Keen’ sour orange (C. aurantium L.), (2) Cleopatra mandarin (C. reticulata Blanco) + sour orange, and (3) ‘Valencia’ sweet orange (C. sinensis (L.) Osbeck) + ‘Femminello’ lemon (C. limon (L.) Burm. f.). Isozyme analyses of PGI, PGM, GOT, and IDH zymograms of putative cybrid plants, along with RFLP analyses using a nuclear genome-specific probe showed that these plants contained the nucleus of the leaf parent. RFLP analyses using mtDNA-specific probes showed that these plants contained the mitochondrial genome of the embryogenic callus donor, thereby confirming cybridization. RFLP analyses using cpDNA-specific probes revealed that the cybrid plants contained the chloroplast genome of either one or the other parent. These results support previous reports indicating that acquisition of the mitochondria of embryogenic protoplasts by leaf protoplasts is a prerequisite for recovering plants with the leaf parent phenotype via somatic embryogenesis following somatic fusion.

Key words

chloroplast mitochondria somatic embryogenesis tissue culture 





glutamateoxaloacetate transaminase


isocitrate dehydrogenase




polyethylene glycol


phosphoglucose isomerase




restriction fragment length polymorphism


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Copyright information

© Springer-Verlag 1996

Authors and Affiliations

  • J. W. Grosser
    • 1
  • F. G. GmitterJr.
    • 1
  • N. Tusa
    • 2
  • G. Reforgiato Recupero
    • 3
  • P. Cucinotta
    • 3
  1. 1.University of Florida, IFAS, Citrus Research and Education CenterLake AlfredUSA
  2. 2.Centro Miglioramento Genetico Degli AgrumiPalermoItaly
  3. 3.Istituto Sperimentale per L'AgrumicolturaAcirealeItaly

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