Journal of Plant Biology

, 50:24 | Cite as

Direct and indirect regeneration of plants from internodal and leaf expiants ofHypencum bupleuroides gris

  • Cüneyt çirak
  • Ali Kemal Ayan
  • Kudret KevseroĞlu


Species of the genusHypencum are of considerable interest worldwide because of their medicinal properties.In- vitro culture is a useful tool for both multiplication of the genus and studying its economically important secondary metabolites. Here, we present an effectivein- vitro propagation method forH. bupleuroides. Leaf and internodal expiants excised from 9-week-old,in vitro-germinated seedlings were cultured on a Murashige and Skoog (MS) medium supplemented with benzyladenine (BA; 1.0 or 0.1 mg L-1) and 2,4-dichlorophenoxyacetic acid (2,4-D; 1.0 or 0.1 mg L1). Depending on the BA and 2,4-D combination used, these cultures produced adventitious shoot buds directly on the surfaces of both types of explants as well as excessive calli. Numerous shoots were obtained when the calli from both expiant types were cultured on an MS medium supplemented with 2 mg L-1 BA. Internodal expiants were more responsive than leaf tissues to direct and indirect plant regeneration. After shoots that regenerated from either the calli or the expiant surface were excised, rooting was best on an MS medium lacking any growth hormones. These rooted plants were then acclimatized under greenhouse conditions, and 90% of regenerants had survived. Ours is the first report ofin- vitro plant regeneration fromH. bupleuroides.


callus induction Hypencum bupleuroides plant growth regulators plant regeneration rooting shoot induction 

Literature Cited

  1. Abreu IN, Azevedo MTA, Solferini MY Mazzafera P (2003)In vitro propagation and isoenzyme polymorphism of the medicinal plantHypericum brasiliense. Biol Plant47: 629–632CrossRefGoogle Scholar
  2. AHS(1980) Shrubs and Hedges: The American Horticultural Society Illustrated Encyclopedia of Gardening. Franklin Library, Franklin Center, PennsylvaniaGoogle Scholar
  3. Al-Juboory KH, Skirvin RM, Williams DJ (1998) Callus induction and adventitious shoot regeneration of gardenia (Gardenia jasminoides Ellis) leaf expiants. Sci Hort72: 171–178CrossRefGoogle Scholar
  4. Ayan AK, çlrak C (2005) Callus induction in someHypericum species growing in Turkey. Turk J Field Crops10: 21–29Google Scholar
  5. Ayan AK, çlrak C (2006)In vitro multiplication ofHypericum heterophyllum, an endemic Turkish species. Amer J Plant Physiol1: 76–81Google Scholar
  6. Ayan AK, çlrak C, Kevseroglu K, Sökmen A (2005) The effectsof expiant types and culture constituents on plant regeneration and hypericin content inHypericum perforatum L. Turk J Agric For29: 197–204Google Scholar
  7. Ayan AK, Cirak C, Yanar O (2006) Variations in total phenolics during ontogenetic, morphogenetic, and diurnal cycles inHypericum species from Turkey. J Plant Biol49: 432–439Google Scholar
  8. Bais HP, Walker TS, McGrew JJ, Vivanco JM (2002) Factors affecting growth of suspension culture ofHypericum perforatum L. and production of hypericin. In Vitro Cell Dev-Plant38: 58–65CrossRefGoogle Scholar
  9. Baytop T (1999) Therapy with Medicinal Plants in Turkey. Istanbul University press, Istanbul, pp 66–167Google Scholar
  10. Bezo M, Stefunova V (2001) Indirect regeneration ofHypericum perforatum L. underin vitro conditions. Acta Fytotech Zootech4: 277–279Google Scholar
  11. Camloha M, Gogala N, Rode J (1994) Plant regeneration from leaf expiants of the fernPlatycerium bifurcatum in vitro. Sci Hort56: 257–266CrossRefGoogle Scholar
  12. Campbell MH, Delfosse ES (1984) The biology of Australian weeds 13. Hypericumperforatum L. J Aust Inst Agric Sci50: 50–63Google Scholar
  13. Cardoso MA, Oliviera DE (1996) Tissue culture ofHypericum brasiliense Choosy, shoot multiplication and callus induction. Plant Cell Tiss Org44: 91–94CrossRefGoogle Scholar
  14. Cellarova E, Kimakova K (1999) Morphoregulatory effect of plant growth regulators onHypericum perforatum L seedlings. Acta Biotechnol19: 163–169CrossRefGoogle Scholar
  15. Cellarova E, Kimakova K, Brutovska R (1992) Multiple shoot for mation inHypericum perforation L. and variability of Ro. Theor Appl Genet101: 46–50Google Scholar
  16. çlrak C (2005) The effects of different tissue culture applications on regeneration ability and the hypericin and total phenolic accumulation in two St. John’s wort species (Hypericum perforatum andHypericum bupleuroides). Ph.D. thesis, University of Ondokuz Mayls, SamsunGoogle Scholar
  17. Davis PH (1988) Flora of Turkey and the East Aegean Islands. Edinburgh University Press, EdinburghGoogle Scholar
  18. Dias ACP, Francisco A, Barberan T, Ferreria FM, Ferreres F (1998) Unusual flavanoids produced by callus ofHypericum perforatum L. Phytochemistry48: 1165–1168CrossRefGoogle Scholar
  19. Georges D, ChenieuxJC, Ochatt SJ (1993) Plant regeneration from aged-callus of the woody ornamental speciesLonicera japonica cv. Hall’s Prolific. Plant Cell Rep13: 91–94CrossRefGoogle Scholar
  20. Holz J, Ostrowiski E (1987) St. John’s wort HPLC analysis of main components and their variability in the populations. Deuts Apothekertz127: 1227–1230Google Scholar
  21. Hong MH, Kim OT, Park JI, Hwang B (2004) Micro propagation ofSchizandra ch’snensis BAILLON using glucose from cotyledonary nodes. J Plant Biol47: 270–274Google Scholar
  22. Hwang SJ (2006) Rapidin vitro propagation and enhanced stevioside accumulation inStevia rebaudiana Bert. J Plant Biol49: 267–270Google Scholar
  23. Ibrahim R, Debergh PC (2001) Factors controlling high effciency adventitious bud formation and plant regeneration fromin vitro leaf expiants of roses (Rosahybrida L.). Sci Hort88: 41–57CrossRefGoogle Scholar
  24. Ishiguro K, Nagareya N, Fukomoto H (1998) A phloroglucinol derivative from cell suspension cultures ofHypericum perforaturn L. Phytochemistry47: 347–369CrossRefGoogle Scholar
  25. Kantia A, Kothari SL (2002) High efficiency adventitious shoot bud formation and plant regeneration from leaf expiants ofDianthus chinensis L. Sci Hort96: 205–212CrossRefGoogle Scholar
  26. Kartnig T, Brantner A (1990) Secondary constituents in cell cultures ofHypericum perforatum andHypericum macuiatum. Planta Med56: 634–637CrossRefGoogle Scholar
  27. Kirakosyan A, Hayashi H, Inoue K, Charchoglyan A, Vardapetyan H (2000) Stimulation of the production of hypericins by mannan inHypericum perforatum L shoot cultures. Phytochemistry53: 345–348PubMedCrossRefGoogle Scholar
  28. Kirakosyan A, Hayashi H, Inoue K, Charchoglyan A, Vardapetyan H, YammotoH (2001) The effect of cork pieces on pseudohypericin production in cells ofHypericum perforatum L. shoots. Russ J Plant Physiol48: 816–819CrossRefGoogle Scholar
  29. Lee T, Huang MEE, Pua EC (1997) High frequency shoot regeneration from leaf disc expiants of garland chrysanthemum (Chrysanthemum coronarium L.)in vitro. Plant Sci126: 219–226CrossRefGoogle Scholar
  30. Leng TC, Haw AB, Lai-Keng C (2004) Effect of reduced N6-benzyladenine, explant type, expiant orientation, culture temperature and culture vessel type on regeneration of adventitious shoot andin vitro plantlets ofSpilanthes acmella. J Plant Biol47: 15–20CrossRefGoogle Scholar
  31. Maciel GB, Moura M (2000) Multiplication strategies forHypericum foliosum aiton, an endemic Azorean species. In Vitro Cell Dev-Plant36: 3CrossRefGoogle Scholar
  32. Mederos MS (1991)in vitro growth and multiplication ofHyperi cum canariense L. Acta Hortic289: 133–135Google Scholar
  33. Moura M (1998) Conservation ofHypericum foliosum aiton, an endemic Azorean species, by micropropagation. In Vitro Cell Dev-Piant34: 244–248CrossRefGoogle Scholar
  34. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant15: 473–497CrossRefGoogle Scholar
  35. Murch SJ, Choffe KL, Victor JMR, Slimmon TY, Krishna R, Saxena PK (2000) Thidiazuron induced plant regeneration from hypocotyl cultures of St. John’s wort (Hypericum perforatum cv “Anthos”). Plant Cell Rep19: 576–581CrossRefGoogle Scholar
  36. Nehra NS, Stushnoff C, Kartha KK (1990) Regeneration of plants from immature leaf-derived callus of strawberry (Fragariax ananassa). Plant Sci66: 119–126CrossRefGoogle Scholar
  37. Nhut DT, Jaime A, da Silva T, Huyen PX, Paek KY (2004) The importance of expiant source on regeneration and micropropagation ofGladiolus by liquid shake culture. Sci Hort102: 407–414CrossRefGoogle Scholar
  38. Özen T, Ayan AK, çlrak C, Kevseroğlu K (2005) Total phenol content of someHypericum species growing in Turkey. Chem Natl Compd41: 232–233CrossRefGoogle Scholar
  39. Patocka J (2003) The chemistry pharmacology, and toxicology of the biologically active constituents of the herbHypericum perforatum L. J Appl Biomed1: 61–73Google Scholar
  40. Pretto FR, Santarem ER (2000) Callus formation and plant regeneration fromHypericum perforatum L. leaves. Plant Cell Tiss Org67: 107–113CrossRefGoogle Scholar
  41. Sahoo Y, Pattnaik SK, Chand PK (1997) Plant regeneration from callus cultures ofMorus indica L. derived from seedlings and mature plants. Sci Hort69: 85–98CrossRefGoogle Scholar
  42. Santarem ER, Astarita LV (2003) Multiple shoot formation inHypericum perforatum L. and hypericin production. Braz J Plant Physiol15: 43–47CrossRefGoogle Scholar
  43. Sirvent T, Gibson D (2002) Induction of hypericins and hyperforin inHypericum perforatum L. in response to biotic and chemical elicitors. Physiol Mol Plant60: 311–320Google Scholar
  44. Steel RG, Torrie JH (1980) Principles and Procedures of Statistics. McGraw-Hill, New YorkGoogle Scholar
  45. Te-chato S, Lim M (2000) Improvement of mangosteen (Garcinia mangostana L.) micropropagation through meristematic nodular callus formation fromin vitro-derived leaf expiants. Sci Hort86: 291–298CrossRefGoogle Scholar
  46. Walker TS, Bais HR Vivanco JM (2002) Jasmonic acid-induced hypericin production in cell suspension cultures ofHypericum perforatum L. (St. John’s wort). Phytochemistry60: 289–293PubMedCrossRefGoogle Scholar
  47. Yazaki K, Okuda T (1990) Procyanins in callus and multiple shoots ofHypericum erectum. Planta Med56: 490–491PubMedCrossRefGoogle Scholar
  48. Zobayed SMA, Murch SJ, Rupasinghe HPV, Saxena PK (2004)In vitro production and chemical characterization of St. John’s wort (Hypericum perforatum L. cv ‘New Stem’) Plant Sci166: 333–340CrossRefGoogle Scholar
  49. Zobayed SMA, Saxena PK (2003) In vitro-grown roots, a superior expiant for prolific shoot regeneration of St. John’s wort (Hypericum perforatum L. cv “New Stem”) in a temporary immersion bioreactor. Plant Sci165: 463–470CrossRefGoogle Scholar

Copyright information

© The Botanical Society of Korea 2007

Authors and Affiliations

  • Cüneyt çirak
    • 1
  • Ali Kemal Ayan
    • 2
  • Kudret KevseroĞlu
    • 1
  1. 1.Faculty of Agriculture, Department of AgronomyUniversity of Ondokuz MayisSamsun/Turkey
  2. 2.the High School of Profession of BafraUniversity of Ondokuz MayisSamsun/Turkey

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