Skip to main content
SpringerLink
Log in

Somatic embryogenesis of Gentiana genus III. Characterization of three-year-old embryogenic suspensions of G. pannonica originated from various seedling explants

  • Published:
Acta Physiologiae Plantarum Aims and scope Submit manuscript

Abstract

Experiments were carried out with three-year-old embryogenic suspension culture of Gentiana pannonica Scop. The initial explant for the suspension determinated both the embryogenic character and embryo production. Cultures were initiated by culture of hypocotyl, cotyledon and root explants on MS (Murashige and Skoog 1962) medium supplemented with 1.0 mg·l−1 Kinetin and 0.5 mg·l−1 2,4-D, later transferred and maintained in liquid MS medium with 1.0 mg·l−1 Dicamba, 0.1 mg·l−1 NAA, 2.0 mg·l−1 BAP and 80.0 mg·l−1 AS. Regeneration medium included 0.0–1.0 mg·l−1 GA3+0.0−2.0 mg·l−1 Kin.+0.0−160 mg·l−1 AS.

In these culture conditions, the effect of the explant was found to be the most important factor. The curve of growth, growth coefficient and % of participation of various size aggregates differed in the studied suspensions. Flow cytometry revealed various DNA content in nuclei from praembryogenic mass depending on the explant origin. To complete embryogenesis the medium was changed from liquid to solidified in the presence of the same plant growth regulators combination required. The most embryogenic culture appeared hypocotyl-derived and it yielded the highest number of somatic embryos. The suspension culture originating from root proliferated the highest number of embryogenic cell clusters but did not produce embryos for fraction 120–450 µm. One hundred mg of suspension of the fraction that was larger than 450 µm yielded 309, 175, 123 embryos for the following suspensions: root-, cotyledon-, hypocotyl-derived, respectively. Almost 50 % of non-deformed fully developed embryos from all studied suspensions passed conversion into germling stage and finally plants were regenerated.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

AS:

adenine sulphate

BAP:

benzylaminopurine

Dic:

dicamba (2,methoxy-3,6 dichlorobenzoic acid)

GA3 :

gibberellic acid

Kin:

kinetin (furfurylaminopurine)

MES:

methylethylsulphuroxide

NAA:

naphthaleneacetic acid

DAPI:

4′,6-diamidino-2-phenylindole

IM:

initiation medium

RM:

regeneration medium

ECM:

embryo conversion medium

MS:

Murashige and Skoog medium

PGR:

plant growth regulators

PEM:

praembryogenic mass

References

  • Bach A., Pawłowska B., Micor J. 1994. Somatic embryogenesis of some genus of mono- and dicotyledonous ornamentals (in Polish). Prace Ogrodu Botanicznego PAN 5/6: 433–437.

    Google Scholar 

  • Doleżel J. 1991. Flow cytometric analysis of nuclear DNA content in higher plants. Analysis 2: 143–154.

    Google Scholar 

  • Freason E.M., Power J.B., Cocking E.C. 1973. The isolation, culture and regeneration of Petunia leaf protoplasts. Developmental Biology 33: 130–137.

    Article  Google Scholar 

  • Fujimura T., Komamine A. 1975. Effects of various growth regulators on the embryogenesis in carrot cell suspension culture. Plant Science Letters 5: 359–364.

    Article  CAS  Google Scholar 

  • Fujimura T., Komamine A. 1980. Synchronization of somatic embryogenesis in a carrot cell suspension culture. Plant Physiology 64: 162–164.

    Article  Google Scholar 

  • Hosokawa K., Nakano M., Oikawa Y., Yamamura S. 1996. Adventitious shoot regeneration from leaf, stem and root explants of commercial cultivars of Gentiana. Plant Cell Reports 15: 578–581.

    Article  CAS  Google Scholar 

  • Hosokawa K., Matsuki R., Oikawa Y., Yamamura S. 1997. Genetic transformation of gentian using wild type Agrobacterium rhizogenes. Plant Cell Tissue and Organ Culture 51: 137–140.

    Article  Google Scholar 

  • Hosokawa K., Matsuki R., Oikawa Y., Yamammura S. 2000. Production of transgenic gentian plants by particle bombardment. Plant Cell Reports 19: 454–458.

    Article  CAS  Google Scholar 

  • Jomori H., Takahata Y., Kaizuma N. 1995. Plant regeneration from leaf-derived calli of Gentians and their protoplast culture. Acta Horticulturae 392: 81–86.

    CAS  Google Scholar 

  • Lamproye A., Crevecoeur M., Kevers C., Gaspar Th. 1987. Multiplication vegetative in vitro de Gentiana lutea et de Gentiana pneumonanthe. Med. Fac. Landbouww. Rijksuniv. Gent. 52: 1255–1257.

    Google Scholar 

  • Meng Y., Gao Y., Jia J. 1996. Plant regeneration from protoplasts isolated from callus of Gentiana crassicaulis. Plant Cell Reports 16: 88–91.

    CAS  Google Scholar 

  • Mikuła A., Tykarska T., Kuraś M., Iwanowska A., Rybczyński J.J. 1996a. Studies on the morphogenic potential of Gentians in cell suspension. Proceedings International Symposium Breeding Research on Medicinal and Aromatic Plants, June 30–July 4, Quedlinburg, Germany, pp: 290–295.

  • Mikuła A., Wesołowska M., Kapusta J., Skrzypczak L., Rybczyński J.J. 1996b. Cytomorphological studies of somatic embryogenesis of Gentiana tibetica (King) and G. cruciata (L.). Acta Soc. Bot. Pol. 65: 47–51.

    Google Scholar 

  • Mikuła A., Rybczyński J.J. 2001. Somatic embryogenesis of Gentiana genus. I. The effect of the preculture treatment and primary explant origin on somatic embryogenesis of: Gentiana cruciata (L), G. pannonica (Scop.) and G. tibetica (King). Acta Physiol. Plant. 23: 15–25.

    Article  Google Scholar 

  • Mikuła A., Rybczyński J.J., Tykarska T., Kuraś M. 2001. Somatic embryogenesis of Gentiana genus. II Scanning and ultrastructural analysis of early stages of somatic embryogenesis in liquid medium. Biol. Bull. Poznań 38: 47–53.

    Google Scholar 

  • Momčilović I., Grubišič D., Kojič M., Nešković M. 1997a. Agrobacterium rhisogenes — mediated transformation and plant regeneration of four Gentiana species. Plant Cell Tissue and Organ Culture 50:1–6.

    Article  Google Scholar 

  • Momčilović I., Grubišič D., Nešković M. 1997b. Micropropagation of four Gentiana species (G. lutea, G. cruciata, G. purpurea and G. acaulis). Plant Cell Tissue and Organ Culture 49: 141–144.

    Article  Google Scholar 

  • Murashige T., Skoog F. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15: 473–497.

    Article  CAS  Google Scholar 

  • Nakano M., Hosokawa K., Oomiya T., Yamamura S. 1995. Plant regeneration from protoplast of Gentiana by embedding protoplats in gellan gum. Plant Cell Tissue and Organ Culture 41: 221–227.

    Article  Google Scholar 

  • Nuti Ronchi V. 1990. Cytogenetics of plant cell cultures. (Ed.:) S. S. Bhojwani In: Plant Tissue Culture: Applications and Limitations. Elsevier, Amsterdam, Oxford, New York, Tokyo, 276–300.

    Google Scholar 

  • Ruffoni B., Massabó F. 1996. Plant production by somatic embryogenesis in cell suspension cultures of Lisianthus russellianus Hook. Plant Tissue Culture and Biotechnology 2: 194–198

    Google Scholar 

  • Ørnstrup H., Mølgaard J.P., Farestveit B. 1993. Somatic embryogenesis and plant regeneration from cell suspensions of Exacum affine. Plant Cell Tissue and Organ Culture 35: 37–41.

    Article  Google Scholar 

  • Sharma N., Chandel K.P.S., Paul A. 1993. In vitro propagation of Gentiana kurroo — an indigenous threatened plant of medicinal importance. Plant Cell Tissue and Organ Culture 34: 307–309.

    Article  CAS  Google Scholar 

  • Skrzypczak L., Wesołowska M., Skrzypczak E. 1993. Gentiana species: in vitro culture, regeneration and production of secoiridoid glucosides. In: Bajaj Y.P.S. (ed) Biotechnology in Agriculture and Forestry, vol. 21. Medicinal and Aromatic Plants IV. Springer, Berlin, Heidelberg, New York, 172–186.

    Google Scholar 

  • Skrzypczak-Pietraszek E., Skrzypczak L., Wesołowska M. 1993. Gentiana tibetica King from tissue culture. Scientia Pharmaceutica 61: 287–296.

    Google Scholar 

  • Tabira H. 1994. Development of micropropagation system of Gentiana triflora. Abstracts of XXIVth International Horticultural Congress. 21–27 August, Kyoto Japan, p. 186.

  • Vinterhalter B., Orbovič P., Vinterhalter D. 1999. Transgenic root culture of Gentiana punctata L. Acta Soc. Bot. Pol. 68: 275–280.

    Google Scholar 

  • Wesołowska M., Kapusta J., Skrzypczak L. 1994. Micropropagation via somatic embriogenesis of Gentiana cruciata L. Abstracts of VIIIth International Congress of Plant Tissue and Cell Culture, June 12–17, Florentia, Italy, p. 176.

  • Wesołowska M., Skrzypczak L., Dudzińska R. 1985. In vitro cultures of Gentiana species plants (in Polish). Acta Polonica Pharmaceutica 42: 79–83.

    Google Scholar 

  • Yamada Y., Shoyama Y., Nishioka I., Kohda H., Namera A., Okamoto T. 1991. Clonal micropropagation of Gentiana scabra Bunge var. buergeri Maxim. and examination of the homogenity concerning the gentiopicroside content. Chemical and Pharmaceutical Bulletin 39: 204–206.

    CAS  Google Scholar 

  • Yuan Y.-M., Küpfer P., Doyle J. 1996. Infrageneric phylogeny of the genus Gentiana (Gentianaceae) inferred from nucleotide sequences of the internal transcribed spacers (ITS) of nuclear ribosomal DNA. American J. Bot. 83: 641–652.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Botanical Garden - Centre for Biological Diversity Conservation of the Polish Academy of Sciences, Prawdziwka 2, 02-973, Warsaw

    Anna Mikuła & Jan J. Rybczyński

  2. Drug Institute, Flow Cytometry Laboratory, Chełmska 30/34, 00-725, Warsaw

    Janusz Skierski

Authors
  1. Anna Mikuła
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Janusz Skierski
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jan J. Rybczyński
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Mikuła, A., Skierski, J. & Rybczyński, J.J. Somatic embryogenesis of Gentiana genus III. Characterization of three-year-old embryogenic suspensions of G. pannonica originated from various seedling explants. Acta Physiol Plant 24, 311–322 (2002). https://doi.org/10.1007/s11738-002-0057-x

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11738-002-0057-x

Key words

Search

Navigation