Plant Cell Reports

, 28:1689 | Cite as

Non-additive phenotypic and transcriptomic inheritance in a citrus allotetraploid somatic hybrid between C. reticulata and C. limon: the case of pulp carotenoid biosynthesis pathway

  • Jean Baptiste Bassene
  • Yann Froelicher
  • Claudie Dhuique-Mayer
  • Waffa Mouhaya
  • Rosa Mar Ferrer
  • Gema Ancillo
  • Raphael Morillon
  • Luis Navarro
  • Patrick OllitraultEmail author
Original Paper


Allopolyploidy is known to induce novel patterns of gene expression and often gives rise to new phenotypes. Here we report on the first attempt to relate phenotypic inheritance in an allotetraploid somatic hybrid with gene expression. Carotenoid compounds in the fruit pulp of the two parental species and the hybrid were evaluated quantitatively by HPLC. Only very low levels of β-carotene and β-cryptoxanthin were observed in Citrus limon, while β-cryptoxanthin was a major component of C. reticulata, which also displayed high levels of phytoene, phytofluene, β-carotene, lutein, zeaxantin and violaxanthin. Total carotenoid content in mandarin juice sacs was 60 times greater than that in lemon. The allotetraploid hybrid produced all the same compounds as mandarin but at very low levels. Transgressive concentration of abscisic acid (ABA) was observed in the somatic hybrid. Real-time RT-PCR of total RNA from juice sacs was used to study expression of seven genes (CitDxs, CitPsy, CitPds, CitZds, CitLcy-b, CitChx-b, and CitZep) of the carotenoid biosynthetic pathway and two genes (CitNced1 and CitNced2) involved in abscisic acid synthesis from carotenoid. Gene expression was significantly higher for mandarin than lemon for seven of the nine genes analyzed. Lemon under expression was partially dominant in the somatic hybrid for three upstream steps of the biosynthetic pathway, particularly for CitDxs. Transgressive over expression was observed for the two CitNced genes. A limitation of the upstream steps of the pathway and a downstream higher consumption of carotenoids may explain the phenotype of the somatic hybrid.


Citrus Somatic hybrid Carotenoid Gene expression Inheritance 



This study was subsided by the Collectivité Territoriale de Corse, the ‘Triploid mandarin breeding project (Proyectos de Investigación Fundamental no orientada; AGL 2008-00596) and the ‘Biotechnologia de citricos’ project (Prometeo 2008/12).


  1. Adams KL, Cronn R, Percifield R, Wendel JF (2003) Genes duplicated by polyploidy show unequal contributions to the transcriptome and organ-specific reciprocal silencing. Proc Natl Acad Sci USA 100:4649–4654CrossRefPubMedGoogle Scholar
  2. Adams KL, Percifield R, Wendel JF (2004) Organ-specific silencing of duplicated genes in a newly synthesized cotton allotetraploid. Genetics 168:2217–2226CrossRefPubMedGoogle Scholar
  3. Albertin W, Brabant P, Catrice O, Eber F, Jenczewski E, Chevre AM, Thiellement H (2005) Autopolyploidy in cabbage (Brassica oleracea L.) does not alter significantly the proteomes of green tissues. Proteomics 5:2131–2139CrossRefPubMedGoogle Scholar
  4. Albertin W, Alix K, Balliau T, Brabant P, Davanture M, Malosse C, Valot B, Thiellement H (2007) Differential regulation of gene products in newly synthesized Brassica napus allotetraploids is not related to protein function nor subcellular localization. BMC Genomics 8:56Google Scholar
  5. Alós E, Cercos M, Rodrigo MJ, Zacarias L, Talon M (2006) Regulation of color break in citrus fruits. Changes in pigment profiling and gene expression induced by gibberellins and nitrate, two ripening retardants. J Agric Food Chem 54:4888–4895CrossRefPubMedGoogle Scholar
  6. Bassene JB, Berti L, Costantino G, Carcouet E, Kamiri M, Tomi F, Dambier D, Ollitrault P, Froelicher Y (2009a) Inheritance of characters involved in fruit quality in a citrus interspecific allotetraploid somatic hybrid. J Agric Food Chem. 57(11):5065–5070CrossRefPubMedGoogle Scholar
  7. Bassene JB, Froelicher Y, Ferrer RM, Dubois C, Navarro L, Ollitrault P, Ancillo G (2009b) Non additive gene regulation in a citrus allotetraploid somatic hybrid between C. reticulata Blanco and C. limon (L.) Burm. Heredity (in press)Google Scholar
  8. Chen ZJ (2007) Genetic and epigenetic mechanisms for gene expression and phenotypic variation in plant polyploids. Annu Rev Plant Biol 58:377–406CrossRefPubMedGoogle Scholar
  9. Chen M, Ha M, Lackey E, Wang J, Chen ZJ (2008) RNAi of met1 reduces DNA methylation and induces genome-specific changes in gene expression and centromeric small RNA accumulation in Arabidopsis allopolyploids. Genetics 178:1845–1858CrossRefPubMedGoogle Scholar
  10. Comai L, Tyagi AP, Winter K, Holmes-Davis R, Reynolds SH, Stevens Y, Byers B (2000) Phenotypic instability and rapid gene silencing in newly formed Arabidopsis allotetraploids. Plant Cell 12:1551–1567CrossRefPubMedGoogle Scholar
  11. Dhuique-Mayer C, Caris-Veyrat C, Ollitrault P, Curk F, Amiot MJ (2005) Varietal and interspecific influence on micronutrient contents in citrus from the Mediterranean area. J Agric Food Chem 53:2140–2145CrossRefPubMedGoogle Scholar
  12. Fanciullino AL (2007) Etude des déterminants génétiques et moléculaires de la variabilité des compositions en caroténoïdes au sein du genre Citrus. UFR des Sciences et Techniques. Université de Corse-Pascal Paoli, Corte, p 212Google Scholar
  13. Fanciullino AL, Dhuique-Mayer C, Luro F, Casanova J, Morillon R, Ollitrault P (2006) Carotenoid diversity in cultivated Citrus is highly influenced by genetic factors. J Agric Food Chem 54:4397–4406CrossRefPubMedGoogle Scholar
  14. Fanciullino AL, Dhuique-Mayer C, Luro F, Morillon R, Ollitrault P (2007) Carotenoid biosynthetic pathway in the Citrus genus: number of copies and phylogenetic diversity of seven genes. J Agric Food Chem 55:7405–7417CrossRefPubMedGoogle Scholar
  15. Fanciullino AL, Cercos M, Dhuique-Mayer C, Froelicher Y, Talon M, Ollitrault P, Morillon R (2008) Changes in carotenoid content and biosynthetic gene expression in juice sacs of four orange varieties (Citrus sinensis) differing in flesh fruit color. J Agric Food Chem 56:3628–3638CrossRefPubMedGoogle Scholar
  16. Flagel L, Udall J, Nettleton D, Wendel J (2008) Duplicate gene expression in allopolyploid Gossypium reveals two temporally distinct phases of expression evolution. BMC Biol 6:16CrossRefPubMedGoogle Scholar
  17. Gancel AL, Ollitrault P, Froelicher Y, Tomi F, Jacquemond C, Luro F, Brillouet JM (2003) Leaf volatile compounds of seven citrus somatic tetraploid hybrids sharing willow leaf mandarin (Citrus deliciosa Ten.) as their common parent. J Agric Food Chem 51:6006–6013CrossRefPubMedGoogle Scholar
  18. Gancel AL, Ollitrault P, Froelicher Y, Tomi F, Jacquemond C, Luro F, Brillouet JM (2005) Leaf volatile compounds of six citrus somatic allotetraploid hybrids originating from various combinations of lime, lemon, citron, sweet orange, and grapefruit. J Agric Food Chem 53:2224–2230CrossRefPubMedGoogle Scholar
  19. Grosser JW, Ollitrault P, Olivares-Fuster O (2000) Somatic hybridization in citrus: an effective tool to facilitate variety improvement. In Vitro Cell Dev Biol Plant 36:434–449CrossRefGoogle Scholar
  20. Hirschberg J (2001) Carotenoid biosynthesis in flowering plants. Curr Opin Plant Biol 4:210–218CrossRefPubMedGoogle Scholar
  21. Kashkush K, Feldman M, Levy AA (2002) Gene loss, silencing and activation in a newly synthesized wheat allotetraploid. Genetics 160:1651–1659PubMedGoogle Scholar
  22. Kato M, Ikoma Y, Matsumoto H, Sugiura M, Hyodo H, Yano M (2004) Accumulation of carotenoids and expression of carotenoid biosynthetic genes during maturation in citrus fruit. Plant Physiol 134:824–837CrossRefPubMedGoogle Scholar
  23. Kato M, Matsumoto H, Ikoma Y, Okuda H, Yano M (2006) The role of carotenoid cleavage dioxygenases in the regulation of carotenoid profiles during maturation in citrus fruit. J Exp Bot 57:2153–2164CrossRefPubMedGoogle Scholar
  24. Kim I, Ko K, Kim C, Chung W (2001) Isolation and characterization of cDNAs encoding beta-carotene hydroxylase in Citrus. Plant Sci 161:1005–1010CrossRefGoogle Scholar
  25. Lai Z, Gross BL, Zou Y, Andrews J, Rieseberg LH (2006) Microarray analysis reveals differential gene expression in hybrid sunflower species. Mol Ecol 15:1213–1227CrossRefPubMedGoogle Scholar
  26. Landry CR, Hartl DL, Ranz JM (2007) Genome clashes in hybrids: insights from gene expression. Heredity 99:483–493CrossRefPubMedGoogle Scholar
  27. Lee HS, Chen ZJ (2001) Protein-coding genes are epigenetically regulated in Arabidopsis polyploids. Proc Natl Acad Sci USA 98:6753–6758CrossRefPubMedGoogle Scholar
  28. Lois LM, Rodriguez-Concepcion M, Gallego F, Campos N, Boronat A (2000) Carotenoid biosynthesis during tomato fruit development: regulatory role of 1-deoxy-d-xylulose 5-phosphate synthase. Plant J 22:503–513CrossRefPubMedGoogle Scholar
  29. Manning K (1991) Isolation of nucleic acids from plants by differential solvent precipitation. Anal Biochem 195:45–50CrossRefPubMedGoogle Scholar
  30. McClintock B (1984) The significance of responses of the genome to challenge. Science 226:792–801CrossRefPubMedGoogle Scholar
  31. Mouhaya W (2008) Caractérisation physiologique et moléculaire de la tolérance au stress salin chez les porte-greffe tétraploïdes d’agrumes. UFR Ciences et techniques. Université de Corse-Pascal Paoli, Corte, p 202Google Scholar
  32. Nicolosi E, Deng ZN, Gentile A, Malfa Sl, Continella G, Tribulato E (2000) Citrus phylogeny and genetic origin of important species as investigated by molecular markers. Theor Appl Genet 100:1155–1166CrossRefGoogle Scholar
  33. Ollitrault P, Dambier D, Froelicher Y, Carreel F, d’Hont A, Luro F, Bruyere S, Cabasson C, Lotfy S, Joumaa A, Vanel F, Maddi F, Treanton K, Grisoni M (2000) Somatic hybridisation potential for Citrus germplasm utilization. Cahiers Agric 9:223–236Google Scholar
  34. Ollitrault P, Guo W, Grosser JW (2007) Somatic hybridization. In: Khan IA (ed) Citrus genetics, breeding and biotechnology. CABI Publishing, Wallingford, pp 235–260Google Scholar
  35. Ollitrault P, Dambier D, Luro F, Froelicher Y (2008) Ploidy manipulation for breeding seedless triploid citrus. Plant Breed Rev 30:323–352CrossRefGoogle Scholar
  36. Osborn TC, Pires JC, Birchler JA, Auger DL, Chen ZJ, Lee HS, Comai L, Madlung A, Doerge RW, Colot V, Martienssen RA (2003) Understanding mechanisms of novel gene expression in polyploids. Trends Genet 19:141–147CrossRefPubMedGoogle Scholar
  37. Rodriguez-Concepcion M, Boronat A (2002) Elucidation of the methylerythritol phosphate pathway for isoprenoid biosynthesis in bacteria and plastids. A metabolic milestone achieved through genomics. Plant Physiol 130:1079–1089CrossRefPubMedGoogle Scholar
  38. Romagosa I, Prada D, Moralejo MA, Sopena A, Munoz P, Casa AM, Swanston JS, Molina-Cano JL (2001) Dormancy, ABA content and sensitivity of a barley mutant to ABA application during seed development and after ripening. J Exp Bot 52:1499–1506CrossRefPubMedGoogle Scholar
  39. Saleh B, Allario T, Dambier D, Ollitrault P, Morillon R (2008) Tetraploid citrus rootstocks are more tolerant to salt stress than diploid. C R Biol 331:703–710CrossRefPubMedGoogle Scholar
  40. Salmon A, Ainouche ML, Wendel JF (2005) Genetic and epigenetic consequences of recent hybridization and polyploidy in Spartina (Poaceae). Mol Ecol 14:1163–1175CrossRefPubMedGoogle Scholar
  41. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory Press, New YorkGoogle Scholar
  42. Sandmann G (2001) Carotenoid biosynthesis and biotechnological application. Arch Biochem Biophys 385:4–12CrossRefPubMedGoogle Scholar
  43. Stupar RM, Bhaskar PB, Yandell BS, Rensink WA, Hart AL, Ouyang S, Veilleux RE, Busse JS, Erhardt RJ, Buell CR, Jiang JM (2007) Phenotypic and transcriptomic changes associated with potato autopolyploidization. Genetics 176:2055–2067CrossRefPubMedGoogle Scholar
  44. Udall JA, Swanson JM, Nettleton D, Percifield RJ, Wendel JF (2006) A novel approach for characterizing expression levels of genes duplicated by polyploidy. Genetics 173:1823–1827CrossRefPubMedGoogle Scholar
  45. Wang J, Tian L, Madlung A, Lee HS, Chen M, Lee JJ, Watson B, Kagochi T, Comai L, Chen ZJ (2004) Stochastic and epigenetic changes of gene expression in Arabidopsis polyploids. Genetics 167:1961–1973CrossRefPubMedGoogle Scholar
  46. Wang JL, Tian L, Lee HS, Wei NE, Jiang HM, Watson B, Madlung A, Osborn TC, Doerge RW, Comai L, Chen ZJ (2006) Genomewide nonadditive gene regulation in Arabidopsis allotetraploids. Genetics 172:507–517CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Jean Baptiste Bassene
    • 1
  • Yann Froelicher
    • 1
  • Claudie Dhuique-Mayer
    • 2
  • Waffa Mouhaya
    • 3
  • Rosa Mar Ferrer
    • 4
  • Gema Ancillo
    • 4
  • Raphael Morillon
    • 3
  • Luis Navarro
    • 4
  • Patrick Ollitrault
    • 3
    • 3
    Email author
  1. 1.UPR Amélioration génétique des espèces à multiplication végétativeCIRAD (Centre de Coopération Internationale en Recherche Agronomique pour le Développement)San GiulianoFrance
  2. 2.UMR QUALISUDCIRAD (Centre de Coope´ration Internationale en Recherche Agronomique pour le Développement)Montpellier Cedex 5France
  3. 3.UPR Amélioration génétique des espèces à multiplication végétativeCIRAD (Centre de Coopération Internationale en Recherche Agronomique pour le Développement), IVIAMoncadaSpain
  4. 4.Centro de Proteccion vegetal y biotechnologiaInstituto Valenciano de Investigaciones Agrarias (IVIA)MoncadaSpain

Personalised recommendations