Plant Cell, Tissue and Organ Culture (PCTOC)

, Volume 112, Issue 1, pp 101–108 | Cite as

Efficiency of direct and indirect shoot organogenesis in different genotypes of Rosa hybrida

  • Leila Pourhosseini
  • Maryam Jafarkhani KermaniEmail author
  • Ali Akbar Habashi
  • Ahmad Khalighi
Research Note


To optimize indirect regeneration (IR) and direct regeneration (DR) in Rosa hybrida cv. Apollo different explant types and different concentrations of plant growth regulators were investigated. Among the different auxins studied and over all explant types, 10 µM 2,4-dichlorophenoxyacetic acid (2,4-D) promoted the highest frequency of callus production for IR. The highest frequency of regeneration (60.8 %) was obtained when calli were transferred to Murashige and Skoog medium supplemented with 2.5 µM thidiazuron (TDZ) and 2 µM gibberellic acid. The highest frequency of regeneration (80.2 %) for DR was obtained from leaves cultured on the medium containing 10 µM TDZ. The efficiency of IR and DR were compared in four different rose cultivars including ‘Apollo’, ‘Black Baccara’, ‘Maroussia’ and ‘Amanda’. The frequency of regeneration in all four cultivars was significantly higher in DR compared to IR. Also shoots regenerated by DR appeared earlier than the shoots regenerated by IR. The results of flow cytometry showed that the shoots derived from IR to DR were tetraploid like the original cultivars.


Direct regeneration Indirect regeneration Rosa hybrida TDZ 2,4-D 


MS medium

Murashige and Skoog medium


2,4,5-Trichlorophenoxyacetic acid


2,4-Dichlorophenoxyacetic acid






Gibberellic acid 3


α-Naphthalene acetic acid


Indirect regeneration


Direct regeneration



This work was part of a PhD thesis on “Study of gene transformation and regeneration in roses” in Science and Research Branch, Islamic Azad University, funded by Agricultural Biotechnology Research Institute of Iran (ABRII) (Project number: 12-05-05-01-8704-88003). Authors wish to thank Prof A. V. Roberts from University of East London, UK, for his constructive comments on the manuscript.


  1. Arene L, Pellegrino C, Gudin S (1993) A comparison of the somaclonal variation level of Rosa hybrida L. cv. Meirutral plants regenerated from callus or DR induction from different vegetative and embryonic tissues. Euphytica 71:83–90CrossRefGoogle Scholar
  2. Bakshi S, Roy NK, Sahoo L (2012) Seedling preconditioning in thidiazuron enhances axillary shoot proliferation and recovery of transgenic cowpea plants. Plant Cell Tiss Organ Cult 110:77–91CrossRefGoogle Scholar
  3. Bao Y, Liu G, Shi X, Xing W, Ning G, Liu J, Bao M (2012) Primary and repetitive secondary somatic embryogenesis in Rosa hybrida ‘Samantha’. Plant Cell Tiss Org Cult 109:411–418CrossRefGoogle Scholar
  4. Burger DW, Liu L, Zary KW, Lee CI (1990) Organogenesis and plant regeneration from immature embryos of R. hybrida L. Plant Cell Tiss Org Cult 21:147–152CrossRefGoogle Scholar
  5. De Wit JC, Esendam HF, Honkanen JJ, Tuominen U (1990) Somatic embryogenesis and regeneration of flowering plants in rose. Plant Cell Rep 9:456–458CrossRefGoogle Scholar
  6. Dohm A, Ludwig C, Nehring K, Debener T (2001) Somatic embryogenesis in roses. Acta Hortic 547:341–347Google Scholar
  7. Dubois LAM, de Vries DP (1995) Preliminary report on the direct regeneration of adventitious buds on leaf explants of in vivo grown glasshouse rose cultivars. Gartenbauwissenschaft 60:249–253Google Scholar
  8. Dubois LAM, de Vries DP, Koot A (2000) Direct shoot regeneration in the rose: genetic variation of cultivars. Gartenbauwissenschaft 65(1):45–49Google Scholar
  9. Estabrooks T, Browne R, Dong Z (2007) 2,4,5-Trichlorophenoxyacetic acid promotes somatic embryogenesis in the rose cultivar Livin Easy’ (Rosa sp.). Plant Cell Rep 26:153–160PubMedCrossRefGoogle Scholar
  10. Haghighat Afshar M, Jafarkhani KM, Khalighi A, Habashi A, Mohammadi A (2011) Direct shoot regeneration on three cultivars of Rosa Hybrida using five rexplant. Am Eurasian J Agric Environ Sci 10(6):962–967Google Scholar
  11. Hsia C, Korban S (1996) Organogenesis and somatic embryogenesis in callus cultures of Rosa hybrida and Rosa chinensis minima. Plant Cell Tiss Org Cult 44:1–6CrossRefGoogle Scholar
  12. Huetteman CA, Preece JE (1993) Thidiazuron: a potent cytokinin for woody plant tissue culture. Plant Cell Tissue Organ Cult 33:105–119CrossRefGoogle Scholar
  13. Ibrahim R, Debergh PC (1999) Improvement of adventitious bud formation and plantlet regeneration from in vitro leaf explants of Rosa hybrida L. Biotechnol Breed 4:413Google Scholar
  14. Ishioka N, Tanimoto S (1990) Plant regeneration from bulgarian rose callus. Plant Cell Tissue Organ Cult 22:197–199CrossRefGoogle Scholar
  15. Kermani MJ, Sarasan V, Roberts AV, Yokoya K, Wentworth J, Sieber VK (2003) Oryzalin-induced chromosome doubling in Rosa and its effect on plant morphology and pollen viability. Theor Appl Genet 107:1195–1200PubMedCrossRefGoogle Scholar
  16. Khosravi R, Kermani MJ, Nematzadeh GA, Bihamta MR, Yokoya K (2007) Role of mitotic inhibitors and genotype on chromosome doubling of Rosa. Euphytica 160:267–275CrossRefGoogle Scholar
  17. Kim SW, Oh SC, Liu JR (2003a) Control of direct and indirect somatic embryogenesis by exogenous growth regulators in immature zygotic embryo cultures of rose. Plant Cell Tissue Organ Cult 74:61–66CrossRefGoogle Scholar
  18. Kim SW, Seung CO, Dong SI, Liu JR (2003b) Plant regeneration of rose (Rosa hybrida) from embryogenic cell-derived protoplasts. Plant Cell Tissue Organ Cult 73:15–19CrossRefGoogle Scholar
  19. Kim CH, Oh JY, Chung J-D, Burrell AM, Byrne DH (2004) Somatic embryogenesis and plant regeneration from in vitro-grown leaf explants of rose. Hortic Sci 39(6):1378–1380Google Scholar
  20. Kintzios S, Manos C, Makri O (1999) Somatic embryogenesis from mature leaves of rose (Rosa sp.). Plant Cell Rep 18:467–472CrossRefGoogle Scholar
  21. Korban SS (2007) Rose. In: Pua EC, Davey MR (eds) Biotechnology in agriculture and forestry transgenic crops VI, vol 61. Springer, Berlin, pp 227–239Google Scholar
  22. Krishna Kumar G, Dennis Thomas T (2012) High frequency somatic embryogenesis and synthetic seed production in Clitoria ternatea Linn. Plant Cell Tiss Org Cult 110:141–151CrossRefGoogle Scholar
  23. Kucharska D, Orlikowska T (2009) Enhancement of in vitro organogenic capacity of rose by preculture of donor shoots on the medium with thidiazuron. Acta Physiol Plant 31:495–500CrossRefGoogle Scholar
  24. Kunitake H, Imamizo H, Mii M (1993) Somatic embryogenesis and plant regeneration from immature seed-derived calli of rose (Rosa rugosa Thunb). Plant Sci 90:187–194CrossRefGoogle Scholar
  25. Li X, Krasnyanski S, Korban SS (2002) Somatic embryogenesis, secondary somatic embryogenesis, and shoot organogenesis in Rosa. J Plant Physiol 159:313–319CrossRefGoogle Scholar
  26. Lloyd D, Roberts AV, Short KC (1998) The induction in vitro of adventitious shoots in Rosa. Euphytica 37:31–36CrossRefGoogle Scholar
  27. Magyar-Tábori K, Dobránszki J, da Teixeira Silva JA, Bulley SM, Hudák I (2010) The role of cytokinins in shoot organogenesis in apple. Plant Cell Tiss Org Cult 101:251–267CrossRefGoogle Scholar
  28. Marchant R, Davey MR, Lucas JA, Power JB (1996) Somatic embryogenesis and plant regeneration in floribunda rose (Rosa hybrida L.) cvs trumpeter and glad tidings. Plant Sci 120:95–105CrossRefGoogle Scholar
  29. Matthews D, Mottley J, Horan I, Roberts AV (1991) A protoplast to plant system in roses. Plant Cell Tiss Org Cult 24:173–180CrossRefGoogle Scholar
  30. Murali S, Sreedhar D, Lokeswari TS (1996) Regeneration through somatic embryogenesis from petal-derived calli of Rosa hybrida L. cv. Arizona (hybrid tea). Euphytica 91:271–275CrossRefGoogle Scholar
  31. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  32. Murthy BNS, Murch JS, Saxena PK (1998) Thidiazuron: a potent regulators of in vitro plant morphogenesis in vitro cell. Dev Biol Plant 34:267–275CrossRefGoogle Scholar
  33. Noriega C, Sondahl MR (1991) Somatic embryogenesis in hybrid tea roses. Biotechnology 9:991–993CrossRefGoogle Scholar
  34. Pati PK, Sharma M, Sood A, Ahuja PS (2004) Direct shoot regeneration from leaf explants of R. damascena mill. In Vitro Cell Dev Biol Plant 40(2):192–195CrossRefGoogle Scholar
  35. Pati PK, Rath SP, Sharma MM, Sood A, Ahuja PS (2006) In vitro propagation of rose—a review. Biotechnol Adv 24:94–114PubMedCrossRefGoogle Scholar
  36. Roberts AV, Horan I, Matthews D, Mottley J (1990) Protoplast technology and somatic embryogenesis in Rosa. In: de Jong J (ed) Integration of the in vitro techniques in ornamental plant breeding. CPO Centre for Plant Breeding Research, Wageningen, The Netherlands, pp 100–115Google Scholar
  37. Roberts AV, Yokoya K, Walker S, Mottley J (1995) Somatic embryogenesis in Rosa spp. In: Jain SM, Gupta PK, Newton R (eds) Somatic embryogenesis in woody plants, vol 2. Kluwer, The Netherlands, pp 277–289CrossRefGoogle Scholar
  38. Rosu A, Skirvin RM, Bein A, Norton MA, Kushad M (1995) The development of putative adventitious shoots from a chimeral thornless rose (Rosa multiflora Thunb. ex J. Murr.) in vitro. J Hortic Sci 70:901–907Google Scholar
  39. Rout GR, Debata BK, Das P (1991) Somatic embryogenesis in callus culture of Rosa hybrida L. cv landora. Plant Cell Tiss Org Cult 27:65–69CrossRefGoogle Scholar
  40. Rout GR, Samantaray S, Mottey J, Das P (1999) Biotechnology of the rose: a review of recent progress. Sci Hortic 81:201–228CrossRefGoogle Scholar
  41. Sarasan V, Roberts AV, Rout GR (2001) Methyl laurate and 6-benzyladenine promote the germination of somatic embryos of a hybrid rose. Plant Cell Rep 20:183–186CrossRefGoogle Scholar
  42. Schum A, Hofmann K, Ghalib N, Tawfik A (2001) Factors affecting protoplast isolation and plant regeneration in Rosa spp. Gartenbauwiss 66:115–122Google Scholar
  43. Shabannezhad Mamaghani M, Asareh MH, Omidi M, Ghamari Zare A, Shahrzad SH (2008) Comparison of direct and indirect regeneration in Eucalyptus Microtheca M. Iranian Rangel Forest Plant Breed Genet Res 16(2):229–237Google Scholar
  44. Shirini S, Mahdavi F, Maziah M (2009) Morphological abnormality among regenerated shoots of banana and plantain (Musa spp.) after in vitro multiplication with TDZ and BA from excised shoot tips. Afr J Biotechnol 8(21):5755–5761Google Scholar
  45. Tian C, Chen Y, Zhao X (2008) Plant regeneration through protocorm-like bodies induced from rhizoids using leaf explants of Rosa spp. Plant Cell Rep 27:823–831PubMedCrossRefGoogle Scholar
  46. van der Salm TPM, van der Toorn CJG, Ten Cate Hanisch CH, Dubois LAM, de Vries DP, Dons HJM (1996) Somatic embryogenesis and shoot regeneration from excised adventitious roots of the rootstock Rosa hybrida L., moneyway. Plant Cell Rep 15:522–526CrossRefGoogle Scholar
  47. Vergne P, Maene M, Chauvet A, Debener T, Bendahmane M (2010) Versatile somatic embryogenesis systems and transformation methods for the diploid rose genotype Rosa chinensis cv. Old blush. Plant Cell Tiss Organ Cult 100:73–81CrossRefGoogle Scholar
  48. Visessuwan R, Kawai T, Mii M (1997) Plant regeneration systems from leaf segment culture through embryogenic callus formation of Rosa hybrida and R. canina. Breed Sci 47:217–222Google Scholar
  49. Visser C, Qureshi JA, Gill R, Saxena PK (1992) Morphoregulatory role of thidiazuron. Plant Physiol 99:1704–1707PubMedCrossRefGoogle Scholar
  50. Yokoya K, Roberts AV, Mottley J, Lewis R, Brandham PE (2000) Nuclear DNA amounts in roses. Ann Bot 85:557–561CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2012

Authors and Affiliations

  • Leila Pourhosseini
    • 1
    • 2
  • Maryam Jafarkhani Kermani
    • 1
    • 2
    Email author
  • Ali Akbar Habashi
    • 1
  • Ahmad Khalighi
    • 2
  1. 1.Department of Tissue Culture and Gene TransformationAgricultural Biotechnology Research Institute of Iran (ABRII)KarajIran
  2. 2.Department of Horticultural Science, Science and Research BranchIslamic Azad University (IAU)TehranIran

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