Somatic embryogenesis and transformation of the diploid Rosa chinensis cv Old Blush

  • Philippe Vergne
  • Marion Maene
  • Guillaume Gabant
  • Aurélie Chauvet
  • Thomas Debener
  • Mohammed BendahmaneEmail author
Original Paper


Somatic embryogenesis was induced from in vitro-derived leaf explants of Rosa chinensis cultivar (cv) Old Blush. Calli producing embryos with expanded cotyledons (RcOBType1 embryos) were obtained. Further refinements of the callus maintenance medium generated a more typical rose embryogenic callus (RcOBType2) displaying high levels of secondary embryogenesis and embryos with limited cotyledon expansion Agrobacterium tumefaciens-mediated transformation assays using β-glucuronidase (GUS) reporter gene showed that both types of embryos were competent for transformation. Under selection conditions, transformed RcOBType1 explants produced non chimaeric transformed embryos, from which shoots could be adventitiously regenerated. In contrast to RcOBType1, transformed RcOBType2 embryos directly yielded transformed shoots when repeatedly cultured in selective regeneration conditions. Transformation efficiency ranged between three to nine percent and shoots suitable for rooting were obtained within 6–8 months. Transgenic plants were transferred into the greenhouse and molecularly confirmed. The availability of transformation methods in a diploid rose, R. chinensis cv. Old Blush, will be useful for gene functional studies.


Rosa Diploid Somatic embryogenesis Agrobacterium Genetic transformation 



2,4-Dichlorophenoxy acetic acid




Ethylenediamine di-2-hydroxyphenyl acetate ferric


Gibberellic acid A3




Indole-3-acetic acid


Indole-3-butyric acid


Murashige and Skoog medium


α-Naphthaleneacetic acid



We thank Vincent David for dedicated assistance, Isabelle Desbouchages and Alexis Lacroix for assistance in growing the plants. This work was funded by Région Rhône-Alpes (Programme ‘Essor des Biotechnologies’ 2000–2003), by the German–French collaborative program ‘Procope’ (2004–2005) and by the Plant Biology Department of the National Institute of Agronomic Research (INRA-France). MM was supported by a PhD fellowship ‘Prospective’ from Région Rhône-Alpes (2005–2008).


  1. Chandler SF, Lu CY (2005) Biotechnology in ornamental horticulture. In Vitro Cell Dev Biol-Plant 41:591–601CrossRefGoogle Scholar
  2. Channeliere S, Riviere S, Scalliet G, Szecsi J, Jullien F, Dolle C, Vergne P, Dumas C, Bendahmane M, Hugueney P, Cock JM (2002) Analysis of gene expression in rose petals using expressed sequence tags. FEBS Lett 515:35–38CrossRefPubMedGoogle Scholar
  3. Chen JR, Liu R, Chen SY, Wang HF (2006) Plant regeneration of transgenic China Rose (Rosa chinensis Jacq.) from organogenic callus. For Stud China 8:92–97CrossRefGoogle Scholar
  4. Condliffe PC, Davey MR, Power JB, Koehorst-van Putten H, Visser PB (2003) An optimized protocol for rose transformation applicable to different cultivars. Acta Hort 612:115–120Google Scholar
  5. Crépin F (1866) Etudes sur les roses. Bull Soc Roy Bot Belg 5:13–27Google Scholar
  6. De Vries DP, Dubois L (1996) Rose breeding: past, present, prospects. Acta Hort 424:241–248Google Scholar
  7. Derks FHM, van Dijk AJ, Hänisch ten Cate CH, Florack DEA, Dubois LAM, de Vries DP (1995) Prolongation of vase life of cut roses via introduction of genes coding for antibacterial activity. Somatic embryogenesis and Agrobacterium-mediated transformation. Acta Hort 405:205–209Google Scholar
  8. Dohm A, Ludwig C, Schilling D, Debener T (2001a) Transformation of roses with genes for antifungal proteins. Acta Hort 547:27–33Google Scholar
  9. Dohm A, Ludwig C, Nehring K, Debener T (2001b) Somatic embryogenesis in roses. Acta Hort 547:341–347Google Scholar
  10. Dohm A, Ludwig C, Schilling D, Debener T (2002) Transformation of roses with genes for antifungal proteins to reduce their susceptibility to fungal diseases. Acta Hort 572:105–111Google Scholar
  11. Firoozabady E, Moy Y, Courtney-Gutterson N, Robinson K (1994) Regeneration of transgenic rose (Rosa hybrida) plants from embryogenic tissue. Bio/Technol 12:609–613CrossRefGoogle Scholar
  12. Gudin S (2001) Rose breeding technologies. Acta Hort 547:23–26Google Scholar
  13. Hood EE, Gelvin SB, Melchers LS, Hoekema A (1993) New Agrobacterium helper plasmids for gene transfer to plants. Transgen Res 2:208–218CrossRefGoogle Scholar
  14. Jefferson RA, Kavanagh TA, Bevan MW (1987) GUS fusions: β-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907PubMedGoogle Scholar
  15. Katsumoto Y, Fukuchi-Mizutani M, Fukui Y, Brugliera F, Holton TA, Karan M, Nakamura N, Yonekura-Sakakibara K, Togami J, Pigeaire A, Tao GQ, Nehra NS, Lu CY, Dyson BK, Tsuda S, Ashikari T, Kusumi T, Mason JG, Tanaka Y (2007) Engineering of the rose flavonoid biosynthetic pathway successfully generated blue-hued flowers accumulating delphinidin. Plant Cell Physiol 48:1589–1600CrossRefPubMedGoogle Scholar
  16. Kim CK, Chung JD, Jee SO, Oh JY (2003) Somatic embryogenesis from in vitro grown leaf explants of Rosa hybrida L. J Plant Biotechnol 5:169–172Google Scholar
  17. Kim CK, Chung JD, Park SH, Burrell AM, Kamo KK, Byrne DH (2004) Agrobacterium tumefaciens-mediated transformation of Rosa hybrida using the green fluorescent protein (GFP) gene. Plant Cell Tissue Organ Cult 78:107–111CrossRefGoogle Scholar
  18. Korban SS, Gasic K, Li X (2006) Rose (Rosa hybrida L.). In: Wang K (ed) Methods in molecular biology, vol 344: Agrobacterium protocols, vol 2, 2nd edn. Humana Press, Totowa, pp 351–358Google Scholar
  19. Krüssmann G (1982) Roses. B. T. Batsford, LondonGoogle Scholar
  20. Li X, Krasnyanski SF, Korban SS (2002) Optimization of the uidA gene transfer into somatic embryos of rose via Agrobacterium tumefaciens. Plant Physiol Biochem 40:453–459CrossRefGoogle Scholar
  21. Li X, Gasic K, Cammue B, Broekaert W, Korban SS (2003) Transgenic rose lines harboring an antimicrobial protein gene, Ace-AMP1, demonstrate enhanced resistance to powdery mildew (Spaerotheca pannosa). Planta 218:226–232CrossRefPubMedGoogle Scholar
  22. Maia N, Vénard P (1976) Cytotaxonomie du genre Rosa et origine des rosiers cultivés. In: FN PH (ed) Travaux sur le rosier de serre. INRA Editions, Paris, pp 7–20Google Scholar
  23. Marchant R, Power JB, Lucas JA, Davey MR (1998a) Biolistic transformation of rose (Rosa hybrida L.). Ann Bot 81:109–114CrossRefGoogle Scholar
  24. Marchant R, Davey MR, Lucas JA, Lamb CJ, Dixon RA, Power JB (1998b) Expression of a chitinase transgene in rose (Rosa hybrida L.) reduces development of blackspot disease (Diplocarpon rosae Wolf). Mol Breed 4:187–194CrossRefGoogle Scholar
  25. Martin M, Piola F, Chessel D, Jay M, Heizmann P (2001) The domestication process of the modern rose: genetic structure and allelic composition of the rose complex. Theor Appl Genet 102:398–404CrossRefGoogle Scholar
  26. Rehder A (1940) Manual of cultivated trees and shrubs, hardy in North America, 2nd edn. The McMillan Co., New YorkGoogle Scholar
  27. Reynders-Aloisi S, Bollereau P (1996) Characterisation of genetic diversity in genus Rosa by randomly amplified polymorphic DNA. Acta Hort 424:253–259Google Scholar
  28. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor Laboratory Press, PlainviewGoogle Scholar
  29. Scalliet G, Journot N, Jullien F, Baudino S, Magnard JL, Channeliere S, Vergne P, Dumas C, Bendahmane M, Cock JM, Hugueney P (2002) Biosynthesis of the major scent components 3, 5-dimethoxytoluene and 1, 3, 5-trimethoxybenzene by novel rose O-methyltransferases. FEBS Lett 523:113–118CrossRefPubMedGoogle Scholar
  30. Scalliet G, Piola F, Douady C, Réty S, Raymond O, Baudino S, Bordji K, Bendahmane M, Dumas C, Cock JM, Hugueney P (2008) Evolution of tea scent in roses. Proc Natl Acad Sci USA 105:5927–5932CrossRefPubMedGoogle Scholar
  31. Schum A, Dohm A (2003) In vitro regeneration techniques. In: Roberts A, Debener T, Gudin S (eds) Encyclopedia of rose science, vol 1. Elsevier Academic Press, Amsterdam, pp 76–90Google Scholar
  32. Souq F, Coutos-Thevenot P, Yean H, Delbard G, Maziere Y, Barbe JP, Boulay M (1996) Genetic transformation of roses, 2 examples: one on morphogenesis, the other on anthocyanin biosynthetic pathway. Acta Hort 424:381–388Google Scholar
  33. Tanaka Y, Katsumoto Y, Brugliera F, Mason J (2005) Genetic engineering in floriculture. Plant Cell Tissue Organ Cult 80:1–24CrossRefGoogle Scholar
  34. Temmerman W, Vereecke D, Dreesen R, Van Montagu M, Holsters M, Goethals K (2000) Leafy gall formation is controlled by fasR, an AraC-type regulatory gene in Rhodococcus fascians. J Bact 182:5832–5840CrossRefPubMedGoogle Scholar
  35. van der Salm T, van der Toorn C, Bouwer R, Hanisch ten Cate C, Dons H (1997) Production of ROL gene transformed plants of Rosa hybrida L. and characterization of their rooting ability. Mol Breed 3:39–47CrossRefGoogle Scholar
  36. Vancanneyt G, Schmidt R, O’Connor-Sanchez A, Willmitzer L, Rocha-Sosa M (1990) Construction of an intron-containing marker gene: splicing of the intron in transgenic plants and its use in monitoring early events in Agrobacterium-mediated plant transformation. Mol Gen Genet 220:245–250CrossRefPubMedGoogle Scholar
  37. Wylie AP (1954) The history of garden roses. J Roy Hort Soc 79:555–571; 80:8–24; 80:27–87Google Scholar

Copyright information

© Springer Science+Business Media B.V. 2009

Authors and Affiliations

  • Philippe Vergne
    • 1
  • Marion Maene
    • 1
  • Guillaume Gabant
    • 1
    • 3
  • Aurélie Chauvet
    • 1
  • Thomas Debener
    • 2
  • Mohammed Bendahmane
    • 1
    • 4
    Email author
  1. 1.Reproduction et Développement des PlantesUMR INRA-CNRS-Université Lyon 1-ENSLLyon cedex 07France
  2. 2.Institute for Plant GeneticsLeibniz University HannoverHannoverGermany
  3. 3.CNRS, Centre de Biophysique MoléculaireOrléansFrance
  4. 4.RDP, UMR5667, Ecole Normale SupérieureLyon cedex 07France

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