Abstract
The citrus fresh market demands the production of seedless citrus fruits, as seedy fruits are not accepted by consumers. The recovery of triploid plants has proven to be the most promising approach to achieve this goal, since triploids have very low fertility, are generally seedless and do not induce seeds in other cultivars by cross pollination. Triploid plants can be recovered by 2x × 4x sexual hybridization. In this work, we present an effective methodology to recover triploid plants from 2x × 4x hybridizations based on in vitro embryo rescue, ploidy level analysis by flow cytometry and genetic origin of triploid plants. The pollen viability of diploid and tetraploid citrus genotypes was analyzed by comparing the pollen germination rate in vitro. The pollen viability of tetraploid (doubled-diploid) genotypes is generally reduced but sufficient for successful pollination. Triploid embryos were identified in normal and undeveloped seeds that did not germinate under greenhouse conditions. The influence of parents and environmental conditions on obtaining triploid plants was analyzed and a strong interaction was noted between the parents and environmental conditions. The parental effect on the length of the juvenile phase was also demonstrated through observations of a large number of progeny over the last 15 years. The juvenile phase length of the triploid hybrids obtained with ‘Fortune’ mandarin as female parent and tetraploid ‘Orlando’ tangelo as male parent was shorter than the juvenile phase obtained with a clementine as female parent and tetraploids of ‘Nova’, ‘W. Leaf’ and ‘Pineapple’ male parents.
Key message Effective methodology to recover citrus triploid plants from 2x × 4x sexual hybridizations and the parental effect on the length of the juvenile phase.
Similar content being viewed by others
References
Adams S, Vinkenoog R, Spielman M, Dickinson HG, Scott RJ (2000) Parent-of-origin effects on seed development in Arabidopsis thaliana require DNA methylation. Development 127:2493–2502
Aleza P, Juárez J, Ollitrault P, Navarro L (2009) Production of tetraploid plants of non apomictic citrus genotypes. Plant Cell Rep 28:1837–1846
Aleza P, Juárez J, Cuenca J, Ollitrault P, Navarro L (2010a) Recovery of citrus triploid hybrids by embryo rescue and flow cytometry from 2x × 2x sexual hybridisation and its application to extensive breeding programs. Plant Cell Rep 29:1023–1034
Aleza P, Juárez J, Ollitrault P, Navarro L (2010b) Polyembryony in non-apomictic citrus genotypes. Ann Bot 106:533–545
Aleza P, Froelicher Y, Schwarz S, Agustí M, Hernández M, Juárez J, Luro F, Morillon R, Navarro L, Ollitrault P (2011) Tetraploidization events by chromosome doubling of nucellar cells are frequent in apomictic citrus and are dependent on genotype and environment. Ann Bot 108:37–50
Aleza P, Juárez J, Hernández M, Ollitrault P, Navarro L (2012) Implementation of extensive citrus triploid breeding programs based on 4x × 2x sexual hybridisations. Tree Genet Genomes. doi:10.1007/s11295-012-0515-6
Cameron JW, Frost HB (1968) Genetic, breeding and nucellar embryony. In: Reuther W, Batchelor LD, Webber HJ (eds) The citrus industry, vol 1. University of California, Riverside, pp 325–370
Cameron JW, Soost RK (1969) Characters of new populations of citrus polyploids, and the relation between tetraploidy in the pollen parent and hybrid tetraploid progeny. Proc First Int Citrus symp 1:199–205
Carputo D, Frusciante L, Peloquin SJ (2003) The role of 2n gametes and endosperm balance number in the origin and evolution of polyploids in the tuber-bearing Solanums. Genetics 163:287–294
Dellaporta J, Hicks JB (1983) A plant DNA minipreparation: version II. Plant Mol Biol Rep 1:19–21
Esen A, Soost RK (1971a) Unexpected triploids in citrus: their origin, identification and possible use. J Hered 62:329–333
Esen A, Soost RK (1971b) Tetraploid progenies from 2x–4x crosses of Citrus and their origin. J Am Soc Hortic Sci 97:410–414
Fatta Del Bosco S, Matranga G, Geraci G (1992) Micro and macro-sporogenesis of two triploid hybrids of Citrus. In: Proceedings of 7th International Citrus Congress, vol 1. International Society of Citriculture, Acireale, Italy, pp 122–124
Fatta del Bosco S, Tusa N, Conicell C (1999) Microsporogenesis in a citrus interspecific tetraploid somatic hybrid and its fusion parents. Heredity 83:373–377
Froelicher Y, Dambier D, Bassene JB, Costantino G, Lotfy S, Didout C, Beaumont V, Brottier P, Risterucci AM, Luro F, Ollitrault P (2008) Characterization of microsatellite markers in mandarin orange (Citrus reticulata Blanco). Mol Ecol Resour 8(1):119–122
Frost HB, Soost RK (1968) Seed reproduction: development of gametes and embryos. In: Reuther W, Batchelor LD, Webber HB (eds) The citrus industry, vol 2., University of CaliforniaBarkley, USA, pp 290–324
Grosser JW, Gmitter FG (2011) Protoplast fusion for production of tetraploids and triploids: applications for scion and rootstock breeding in citrus. Plant Cell Tiss Organ Cult 104:343–357. doi:10.1007/s11240-010-9823-4
Grosser JW, Ollitrault P, Olivares O (2000) Somatic hybridization in Citrus : an effective tool to facilitate variety improvement. In Vitro Cell Dev Biol Plant 36:434–449
Grosser JW, Hyum JA, Calovic M, Dong HL, Chen C, Vasconcellos M, Gmitter FG (2010) Production of new allotetraploid and autotetraploid citrus breeding parents: focus on Zipperskin mandarins. HortScience 45(8):1160–1163
Hedhly A, Hormaza JL, Herrero M (2005a) Influence of genotype-temperature interaction on pollen performe. J Evol Biol 18:1494–1502
Hedhly A, Hormaza JL, Herrero M (2005b) The effect of temperature on pollen germination, pollen tube growth, and stigmatic receptivity in Peach. Plant Biol 7(5):476–483
Luro F, Maddy F, Jacquemond C, Froelicher Y, Morillon R, Rist D, Ollitrault P (2004) Identification and evaluation of diplogyny in clementine (Citrus clementina) for use in breeding. In: XI Eucarpia Symposium on Fruit Breeding and Genetics. Acta Horticulturae. 663: 841–847
Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–479
Navarro L, Juárez J (2007) Shoot-tip grafting in vitro. In: Khan IA (ed) Citrus Genetics, breeding and biotechnology. CABI Head Office, Wallingford, pp 353–364
Navarro L, Roistacher CN, Murashige T (1975) Improvement of shoot-tip grafting in vitro for virus-free citrus. J Am Soc Hortic Sci 100:471–479
Navarro L, Juárez J, Aleza P, Pina JA (2002a) Recovery of triploid seedless mandarin hybrids from 2n × 2n and 2n × 4n crosses by embryo rescue and flow cytometry. In: Proceedings of the 10th IAPTC&B Congres, Plant Biotechnology 2002 and Beyond. Orlando, Florida, USA, pp 541–544
Navarro L, Pina JA, Juárez J, Ballester-Olmos JF, Arregui JM, Ortega C, Navarro A, Duran-Vila N, Guerri J, Moreno P, Cambra M, Zaragoza S (2002b) The Citrus Variety Improvement Program in Spain in the Period 1975–2001. In: Proceedings of the 15th Conference of the International Organization of Citrus Virologists, IOCV, Riverside, pp 306–316
Navarro L, Juárez J, Aleza P, Pina JA, Olivares-Fuster O, Cuenca J, Julve JM (2005) Programa de obtención de híbridos triploides de mandarino en España. Phytoma 170:36–41
Ollitrault P, Michaux-Ferriere N (1992) Application of flow cytometry for citrus genetic and breeding. In: Proceeding of the 7th International Citrus Congress, vol 1. International Society of Citriculture, Acireale, pp 193–198
Ollitrault P, Dambier D, Luro F, Froelicher Y (2008) Ploidy manipulation for breeding seedless triploid citrus. Plant Breeding Rev 20:323–354
Otto SP, Whitton J (2000) Polyploid incidence and evolution. Ann Rev Genet 34:401–437
Soost RK (1965) Incompatibility alleles in the genus citrus. Proc Am Soc Hortic Sci 87:176–180
Soost RK (1969) The incompatibility gene system in citrus. In: Chapman HD (ed) Proceedings of the first international citrus symposium, vol 1. University of California, Riverside, pp 189–190
Tachikawa T, Tanaka Y, Hara S (1961) Investigation on the breeding of citrus trees. I study on the breeding of triploid citrus varieties. Bul Shizuoka Pref Citrus Exp Sta 4:33–44
Viloria Z, Grosser JW (2005) Acid citrus fruit improvement via interploid hybridization using allotetraploid somatic hybrid and autotetraploid breeding parents. J Am Soc Hortic Sci 130:392–402
Vinkenoog R, Bushell C, Spielman M, Adams S, Dickinson HG, Scott RJ (2003) Genomic imprinting and endosperm development in flowering plants. Mol Biotechnol 25(2):149–184
Williams TE, Roose ML (2004) ‘TDE2’ Mandarin hybrid (Shasta Gold® Mandarin), ‘TDE3’ Mandarin hybrid (Tahoe Gold® Mandarin) and ‘TDE4’ Mandarin hybrid (Yosemite Gold® Mandarin): three new mid and late-season triploid seedless Mandarin hybrids from California. In: Proceedings of 10th International Citrus Congress, vol 1. International Society of Citriculture, Agadir, pp 394–398
Young HJ, Stanton ML (1990) Influence of environmental quality on pollen competitive ability in wild radish. Science 248:1631–1633
Acknowledgments
We thank M. Hernández, J.M. Arregui, C. Ortega, A. Navarro, V. Ortega, and C. Martí for technical assistance in the laboratory, and J.A Pina, V. Lloris, J.M. Conchilla, F. Ahuir, D Conchilla, A. Conchilla, R. López, and F.J. Martí for growing plants in the greenhouse and field. We also thank Dr. F. Luro from INRA (France) for providing unpublished SSR markers. This work was supported by a grant (Prometeo/2008/121) from the Generalitat Valenciana, Spain and by two grants (AGL2008-00596 and AGL2011-26490) from the Ministry of Science and Innovation of Spain-Fondo Europeo de Desarrollo Regional (FEDER).
Author information
Authors and Affiliations
Corresponding author
Additional information
Communicated by W. Harwood.
Electronic supplementary material
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Aleza, P., Juárez, J., Cuenca, J. et al. Extensive citrus triploid hybrid production by 2x × 4x sexual hybridizations and parent-effect on the length of the juvenile phase. Plant Cell Rep 31, 1723–1735 (2012). https://doi.org/10.1007/s00299-012-1286-0
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00299-012-1286-0