Cytology and Genetics

, Volume 53, Issue 3, pp 192–201 | Cite as

Effects of Exogenous Cytokinins on Spore Germination and Gametophyte Morphogenesis of Dryopteris filix-mas (L.) Schott in vitro Culture

  • K. O. RomanenkoEmail author
  • I. V. KosakivskaEmail author
  • L. M. BabenkoEmail author
  • O. V. VashekaEmail author
  • P. O. RomanenkoEmail author
  • V. A. NegretskyEmail author
  • V. M. MinarchenkoEmail author


The impact of exogenous cytokinins (kinetin, zeatin, 6-benzylaminopurine, and N6-2-isopentenyladenine) on the pattern of Dryopteris filix-mas (L.) Schott spore germination, gametophyte growth and morphology in vitro have been studied. It was found that all studied cytokinins significantly retarded spore germination, inhibited gametophyte growth, caused deformations and decrease in the thallus size, and suppressed the development of reproductive structures and sporophyte growth at the concentration of 10–5 М. The reduction of the hormone concentration to 10–8 М stimulated the gametophyte development, induced cell divisions, particularly in the apical zone, due to which some of thalli were deformed, promoted the production of rhizoids, affected the formation of antheridia and archegonia, and slowed the sporophyte development.


Dryopteris filix-mas gametophyte spores prothallium thallus cytokinins kinetin zeatin 6-benzylaminopurine N6-2-isopentenyladenine 



  1. 1.
    Kosakivska, I.V., Babenko, L.M., Shcherbatiuk, M.M., Vedenicheva, N.P., Voytenko, L.V., and Vasyuk, V.A., Phytohormones during growth and development of Polypodiophyta, Adv. Biol. Eearth Sci., 2016, vol. 1, pp. 26–44.Google Scholar
  2. 2.
    Plant Hormones: Biosynthesis, Signal Transduction, Action, Davies, P.J., Ed., Netherlands: Springer, 2010. 3rd ed. Google Scholar
  3. 3.
    Fonseca, S., Rosado, A., Vaughan-Hirsch, J., Bishopp, A., and Chini, A., Molecular locks and keys: the role of small molecules in phytohormone research, Front. Plant Sci., 2014, vol. 5, art. 709, pp. 1–16.
  4. 4.
    Wang, Y.H. and Irving, H.R., Developing a model of plant hormone interactions, Plant Signal. Behav., 2011, vol. 6, pp. 494–500. Scholar
  5. 5.
    Wells, D.M., Laplaze, L., Bennett, M.J., and Vernoux, T., Biosensors for phytohormone quantification: challenges, solutions, and opportunities, Trends Plant Sci., 2013, vol. 18, pp. 244–249. Scholar
  6. 6.
    Borghi, L., Kang, J., Ko, D., Lee, Y., and Martinoia, E., The role of ABCG-type ABC transporters in phytohormone transport, Biochem. Soc. Trans., 2015, vol. 43, pp. 924–930. Scholar
  7. 7.
    Raghavan, V., Developmental Biology of Fern Gametophytes, Cambridge University Press, 1989. Scholar
  8. 8.
    Du, H., Li, Y., Li, D., Dai, S., Jiang, C., and Shi, L., Effects of light, temperature and pH on spore germination and early gametophytic development of Alsophila metteniana, Biodiv. Sci., 2009, vol. 17, pp. 182–187. Scholar
  9. 9.
    Juárez-Orozco, S., Orozco-Segovia, A., Mendoza-Ruiz, A., and Pérez-García, B., Spore germination of eight homosporous ferns in a temperature gradient, S. Afr. J. Bot., 2013, vol. 87, pp. 112–117. CrossRefGoogle Scholar
  10. 10.
    Edwards, E.S. and Roux, S.J., Influence of gravity and light on the developmental polarity of Ceratopteris richardii fern spores, Planta, 1998, vol. 205, pp. 553–560. Scholar
  11. 11.
    Wu, H., Liu, X.-Q., Ji, H., and Chen, L.-Q., Effects of light, macronutrients, and sucrose on germination and development of the endangered fern Adiantum reniforme var. sinense (Adiantaceae), Sci. Hortic., 2010, vol. 125, pp. 417–421. Scholar
  12. 12.
    Suo, J., Chen, S., Zhao, Q., Shi, L., and Dai, S., Fern spore germination in response to environmental factors, Front. Biol., 2015, vol. 10, pp. 358–376. CrossRefGoogle Scholar
  13. 13.
    Edwards, M.E., Carbon dioxide and ethylene control of spore germination in Onoclea sensibilis L., Plant Physiol., 1977, vol. 59, pp. 756–758. Scholar
  14. 14.
    Camloh, M., Ravnikar, M., and Zel, J., Jasmonic acid promotes division of fern protoplasts, elongation of rhizoids and early development of gametophytes, Physiol. Plant, 1996, vol. 97, pp. 659–664. Scholar
  15. 15.
    Chia, S.-G.E. and Raghavan, V., Abscisic acid effects on spore germination and protonemal growth in the fern, Mohria caffrorum, New Phytol, 1982, vol. 92, pp. 31–37.CrossRefGoogle Scholar
  16. 16.
    Babenko, L.M., Romanenko, K.O., Shcherbatiuk, M.M., Vasheka, O.V., Romanenko, P.O., Negretsky, V.A., and Kosakivska, I.V., Effects of exogenous phytohormones on spore germination and morphogenesis of Polystichum aculeatum (L.) Roth gametophyte in vitro culture, Cytol. Genet., 2018, vol. 52, pp. 117–126. CrossRefGoogle Scholar
  17. 17.
    Gómez-Garay, A., Galán, J.M.G., Cabezuelo, A., Pintos, B., Prada, C., and Martín, L., Ecological significance of brassinosteroids in three temperate ferns, in Current Advances in Fern Research, Cham: Springer, 2018, pp. 453–466. Scholar
  18. 18.
    Takeno, K. and Furuya, M., Inhibitory effect of gibberellins on archegonial differentiation in Lygodium japonicum, Physiol. Plant., 1977, vol. 39, pp. 135–138. Scholar
  19. 19.
    Swami, P. and Raghavan, V., Control of morphogenesis in the gametophyte of a fern by light and growth hormones, Can. J. Bot., 1980, vol. 58, pp. 1464–1473. Scholar
  20. 20.
    Kazmierczak, A., Induction of cell division and cell expansion at the beginning of gibberellin A3-induced precocious antheridia formation in Anemia phyllitidis gametophytes, Plant Sci., 2003, vol. 165, pp. 933–939. CrossRefGoogle Scholar
  21. 21.
    Castilho, C.V.V., Neto, J.F.F., Leitao, S.G., Bar-reto, S.C.P., and Silva, N.C.B., Anemia tomentosa var. anthriscifolia in vitro culture: sporophyte development and volatile compound profile of an aromatic fern, Plant Cell Tiss. Organ Cult., 2018, pp. 1–13. Scholar
  22. 22.
    Miller, J.H., Fern gametophytes as experimental material, Bot. Rev., 1968, vol. 34, pp. 361–440. Scholar
  23. 23.
    Korpelainen, H., Growth, sex determination and reproduction of Dryopteris filix-mas (L.) Schott gametophytes under varying nutritional conditions, Bot. J. Linn. Soc., 1994, vol. 114, pp. 357–366. Scholar
  24. 24.
    Atallah, N.M. and Banks, J.A., Reproduction and the pheromonal regulation of sex type in fern gametophytes, Front. Plant Sci., 2015, vol. 6, art. 100, pp. 1–6.
  25. 25.
    Hollingsworth, S., Andres, E., and Greery, G., Pheromonal interactions among gametophytes of Osmundastrum cinnamomeum and the origins of antheridiogen systems in leptosporangiate ferns, Int. J. Plant Sci., 2012, vol. 173, pp. 382–390. Scholar
  26. 26.
    Menéndez, V., Revilla, M.A., Bernard, P., Gotor, V., and Fernández, H., Gibberellins and antheridiogen on sex in Blechnum spicant L., Plant Cell Rep., 2006, vol. 25, pp. 1104–1110. Scholar
  27. 27.
    Kwa, S.H., Wee, Y.C., Lim, T.M., and Kumar, P.P., IAA-induced apogamy in Platycerium coronarium (Koenig) Desv. gametophytes cultured in vitro, Plant Cell Rep., 1995, vol. 14, pp. 598–602. Scholar
  28. 28.
    Higuchi, H., Amaki, W., and Suzuki, S., In vitro propagation of Nephrolepis cordifolia Prsel., Sci. Hortic., 1987, vol. 32, pp. 105–113. Scholar
  29. 29.
    Fernández, H. and Revilla, M.A., In vitro culture of ornamental ferns, Plant Cell Tissue Organ Cult., 2003, vol. 73, pp. 1–13. Scholar
  30. 30.
    Bharati, S.K., Manabendra, D.C., and Mazumder, P.B., In vitro propagation in Pteridophytes, Int. J. Res. Ayurveda Pharm., 2013, vol. 4, pp. 297–303. Scholar
  31. 31.
    Somer, M., Arbesū, R., Menéndez, V., Revilla, M.A., and Fernández, H., Sporophyte induction studies in ferns in vitro, Euphytica, 2010, vol. 171, p. 203. Scholar
  32. 32.
    Chen, S.Y. and Read, P.E., Micropropagation of leatherleaf fern (Rumohra adiantiformis), Proc. Fla. State Hort. Soc., 1983, vol. 96, pp. 266–269.Google Scholar
  33. 33.
    Menéndez, V., Abul, Y., Bohanec, B., Lafont, F., and Fernández, H., The effect of exogenous and endogenous phytohormones on the in vitro development of gametophyte and sporophyte in Asplenium nidus L., Acta Physiol. Plant, 2011, vol. 33, pp. 2493–500. Scholar
  34. 34.
    Higuchi, H. and Amaki, W., Effects of 6-benzylaminopurine on the organogenesis of Asplenium nidus L. through in vitro propagation, Sci. Hortic., 1989, vol. 37, pp. 351–359. CrossRefGoogle Scholar
  35. 35.
    Vedenicheva, N.P. and Kosakivska, I.V., Modern aspects of cytokinins studies: evolution and crosstalk with other phytohormones, Fiziol. Rast. Genet., 2016, vol. 48, pp. 3–19.CrossRefGoogle Scholar
  36. 36.
    Vedenicheva, N.P. and Kosakivska, I.V., Cytokinins As Regulators of Plant Ontogenesis Under Different Growth Conditions, Kyiv: Nash Format, 2017.Google Scholar
  37. 37.
    Veselov, D.S., Kudoyarova, G.R., Kudryakova, N.V., and Kusnetsov, V.V., Role of cytokinins in stress resistance of plants, Russ. J. Plant Physiol., 2017, vol. 64, pp. 15–27. Scholar
  38. 38.
    Kudo, T., Makita, No., Kojima, M., Tokunaga, H., and Sakakibara, H., Cytokinin activity of cis-zeatin and phenotypic alterations induced by overexpression of putative cis-zeatin-o-glucosyltransferase in rice, Plant Physiol., 2012, vol. 160, pp. 319–331. Scholar
  39. 39.
    Huang, S., Cerny, R.E., Qi, Y., Bhat, D., Aydt, C.M., Hanson, D.D., Malloy, K.P., and Ness, L.A., Transgenic studies on the involvement of cytokinin and gibberellin in male development, Plant Physiol., 2003, vol. 131, pp. 1270–1282. Scholar
  40. 40.
    Gerashchenkov, G.A. and Rozhnova, N.A., The involvement of phytohormones in the plant sex regulation, Russ. J. Plant Physiol., 2013, vol. 60, pp. 597–610. Scholar
  41. 41.
    Beck, M.J. and Caponetti, J.D., The effects of kinetin and naphthaleneacetic acid on in vitro shoot multiplication and rooting in the fishtail fern, Am. J. Bot, 1983, vol. 70, pp. 1–7.CrossRefGoogle Scholar
  42. 42.
    Hicks, G. and Aderkas, P.V., A tissue culture of the Ostrich fern Matteuccia struthiopteris (L.) Todaro, Plant Cell Tiss. Organ Cult, 1986, vol. 5, pp. 199–204. Scholar
  43. 43.
    Amaki, W. and Higuchi, H., A possible propagation system of Nephrolepis, Asplenium, Pteris, Adiantum and Rumora through tissue culture, Acta Hortic., 1991, vol. 300, pp. 237–243. Scholar
  44. 44.
    Fernández, H., Bertrand, A.M., and Sánchez-Tamés, R., Micropropagation and phase change in Blechnum spicant and Pteris ensiformis, Plant Cell Tiss. Organ Cult., 1996, vol. 44, pp. 261–265. Scholar
  45. 45.
    Fernández, H., Bertrand, A., and Sánchez-Tamés, R., Plantlet regeneration in Asplenium nidus L. and Pteris ensiformis L. by homogenization of BA treated rhizomes, Sci. Hortic., 1997, vol. 68, pp. 243–247. Scholar
  46. 46.
    Menéndez, V., Revilla, M.A., Fal, M.A., and Fernández, H., The effect of cytokinins on growth and sexual organ development in the gametophyte of Blechnum spicant L., Plant Cell Tiss. Organ Cult., 2009, vol. 96, pp. 245–250. Scholar
  47. 47.
    Greer, G.K., Dietrich, M.A., DeVol, J.A., and Rebert, A., The effects of exogenous cytokinin on the morphology and gender expression of Osmunda regalisgametophytes, Am. Fern J., 2012, vol. 102, pp. 32–46. Scholar
  48. 48.
    Bonomo, M.C., Martinez, O.G., Tanco, M.E., Cardozo, R., and Aviles, Z., Spores germination and gametophytes of Alsophila odonelliana (Cyatheaceae) in different sterile media, Phyton (Buenos Aires), 2013, vol. 82, pp. 119–126.Google Scholar
  49. 49.
    Spiro, M.D., Torabi, B., and Cornell, C.N., Cytokinins induce photomorphogenic development in dark-grown gametophytes of Ceratopteris richardii, Plant Cell Physiol., 2004, vol. 45, pp. 1252–1260. Scholar
  50. 50.
    Banks, J.A., Gametophyte development in ferns, Annu. Rev. Plant Physiol. Plant Mol. Biol., 1999, vol. 50, pp. 163–186. Scholar
  51. 51.
    Parajuli, J. and Joshi, S.D., In vitro study of effects of growth hormones on sporophyte development of Cyathea spinulosa, Int. J. Biodivers. Conserv., 2014, vol. 6, pp. 247–255. Scholar
  52. 52.
    Grichuk, V.P. and Monoszon, M.H., The Determinant of Single-Beam Spores of the Ferns from the Family Polypodiaceae R. Br., Growing on the Territory of the USSR, Moscow: Nauka, 1971.Google Scholar
  53. 53.
    Nayar, B.K. and Kaur, S., Gametophytes of homosporous ferns, Bot. Rev., 1971, vol. 37, pp. 295–396. Scholar
  54. 54.
    Tryon, A.F. and Lugardon, B., Dryopteridaceae Herter, in spores of the pteridophyta, springer, New York, 1991. Scholar
  55. 55.
    Gaba, V., Plant growth regulators in plant tissue culture and development, in Plant Tissue Culture, Development, and Biotechnology, Trigiano, R.N. and Gray, D.J., Eds., CRC Press, 2005, pp. 87–99.Google Scholar

Copyright information

© Allerton Press, Inc. 2019

Authors and Affiliations

  1. 1.Kholodny Institute of Botany, National Academy of Sciences of UkraineKyivUkraine
  2. 2.Educational and Scientific Center “Institute of Biology and Medicine,” Taras Shevchenko National University of KyivKyivUkraine
  3. 3.Bogomolets National Medical UniversityKyivUkraine

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