Regeneration features and leaf anatomical struc- ture of Actinidia polygama under in vitro conditions
https://doi.org/10.35102/cbg.2025.40.28.004
Abstract
Actinidia polygama is an unusual berry crop whose fruits are rich in carotenoids (including βcarotene), ascorbic acid, and other valuable bioactive compounds. This study investigated the regeneration of promising A. polygama cultivars during micropropagation. We assessed the morphometric responses of explants to different growth regulators, including 6-benzylaminopurine (6-BAP), metaTopolin (mT), and 2-isopentenyladenine (2iP). Cultivation on a medium supplemented with 0,5 mg L⁻¹ mT stimulated the formation of adventitious microshoots at the explant base and induced microshoot bud formation. This treatment increased the micropropagation efficiency of A. polygama cultivars by 1,1–1,3 times compared to media containing 0,5 mg L⁻¹ 6-BAP or 0,5 mg L⁻¹ 2iP. A synergistic effect was observed from the combined application of mT and 6-BAP (both at 0,5 mg L⁻¹), which increased micropropagation rates by 1,2–1,3 times in most cultivars. The study also examined the anatomical structure of the leaf assimilation tissues and stomatal apparatus during the transition from in vitro to ex vitro conditions. All A. polygama specimens had hypostomatous leaves with anomocytic stomata, regardless of culture conditions. Under in vitro conditions, leaves featured thin blades and mesophyll composed of three to four layers (one palisade layer and two to three spongy chlorenchyma layers) with large intercellular spaces, as well as numerous large, rounded stomata. During ex vitro acclimatization, the leaf tissues expanded and differentiated, the stomatal density and area decreased, and the stomatal shape changed from rounded to elliptical. This anatomical analysis demonstrated that leaf development in A. polygama cultivars is influenced by both culture conditions and genetic characteristics.
About the Authors
I. L. Krakhmaleva
Tsitsin Main Botanical Garden of the Russian Academy of Sciences
Russian Federation
Russian Federation
Moscow
O. I. Molkanova
Tsitsin Main Botanical Garden of the Russian Academy of Sciences
Russian Federation
Russian Federation
Moscow
References
1. Butenko R.G. 1991. Biology of cultivated cells and plant biotechnology. Moscow, Nauka. 279 p. (In Russian)
2. Deb C.R., Gangmei P.K. 2020. In vitro morphogenesis of foliar explants and plant regeneration of Actinidia deliciosa A. Chev. – a horticultural important plant. Plant Cell Biotechnology and Molecular Biology. 21(15–16): 114–123.
3. Debenham M.C., Seelye J.F., Mullan A.C. 2016. An in vitro repository for clonal kiwifruit. Acta Horticulturae. 1113: 93–98. DOI: 10.17660/ActaHortic.2016.1113.13.
4. Ferguson A.R., Huang H. 2007. Genetic resources of kiwifruit: Domestication and breeding. In: Jules Janick, ed. Horticultural reviews. Vol. 33. Hoboken, New Jersey, USA, John Wiley and Sons, Inc. P. 1–121.
5. Gins M.S., Gins V.K., Baykov А.А., Pivovarov V.F., Kozak N.V., Imamkulova Z.А., Kulikov I.M. et al. 2018. Foliar biomass of the vegetable and small fruit plants source of the antioxidants. Vestnik of the Russian agricultural science. 1: 39–44. (In Russian)
6. Hameg, R., Arteta, T.A., Landin, M., Gallego, P.P., Barreal, M.E. 2020. Modeling and optimizing culture medium mineral composition for in vitro propagation of Actinidia arguta. Frontiers in Plant Science. 11: 554905. DOI: 10.3389/fpls.2020.554905.
7. Jagiełło-Kubiec K., Nowakowska K., Łukaszewska A.J., Pacholczak A. 2021. Acclimation to ex vitro conditions in ninebark. Agronomy. 11: 612. DOI: 10.3390/agronomy11040612.
8. Jayaprakash K., Manokari M., Badhepuri M.K., Raj M.C., Dey A., Shekhawat M.S. 2021. Influence of meta-topolin on in vitro propagation and foliar micro-morpho-anatomical developments of Oxystelma esculentum (L.f.) Sm. Plant Cell, Tissue and Organ Culture. 147: 325–337. DOI: 10.1007/s11240-021-02126-y.
9. Kamelin R.V. 2022. Flora of East Asia and its genesis. Turczaninowia. 25(3): 5–16. (In Russian).
10. Khanam M.N., Javed S.B., Anis M., Alatar A.A. 2020. meta-Topolin induced in vitro regeneration and metabolic profiling in Allamanda cathartica L. Industrial Crops and Products. 145: 111944. DOI: 10.1016/j.indcrop.2019.111944.
11. Kozak N.V., Imamkulova Z.A., Kulikov I.M., Vlasova E.V., Medvedev S.M., Ilinova L.N. 2020. Rare berry crops: morphology, biochemistry, ecology. Moscow, FSBSO ARHCBAN. 72 p. (In Russian).
12. Krakhmaleva I.L., Molkanova O.I., Orlova N.D., Koroleva O.V., Mitrofanova I.V. 2024. In vitro morpho-anatomical and regeneration features of cultivars of Actinidia kolomikta (Maxim.) Maxim. Horticulturae. 10(12): 1335. DOI: 10.3390/horticulturae10121335.
13. Kucharska D., Orlikowska T., Maciorowski R., Kunka M., Wójcik D., Pluta S. 2020. Application of meta-Topolin for improving micropropagation of gooseberry (Ribes grossularia). Scientia Horticulturae. 272: 109529. DOI: 10.1016/j.scienta.2020.109529.
14. Lalthafamkimi L., Bhattacharyya P., Bhau B.S., Wann S.B., Banik D. 2021. Direct organogenesis mediated improvised mass propagation of Pogostemon cablin: A natural reserve of pharmaceutical biomolecules. South African Journal of Botany. 140: 375–384. DOI: 10.1016/j.sajb.2020.08.018.
15. Levchyk N., Skrypchenko N., Dziuba O., Gajdosova A., Liubinska A., Zaimenko N. 2022. Features of morphogenesis of Actinidia arguta leaf tissues at microclonal propagation. Journal of Microbiology, Biotechnology and Food Sciences. 12(1): e4667. DOI: 10.55251/jmbfs.4667.
16. Ma J.T., Li D.W., Liu J.K., He, J. 2021. Advances in research on chemical constituents and their biological activities of the genus Actinidia. Natural Products and Bioprospecting. 11: 573–609. DOI: 10.1007/s13659-021-00319-8.
17. Malaeva E.V. 2008 Biological and molecular-genetic features of Far Eastern species of the genus Actinidia Lindl. Cand. Biol. Sci. diss. Moscow. 136 p.
18. Malaeva E.V. 2020. Theoretical and practical aspects of clonal propagation of small fruit cultures. Problems of botany of south Siberia and Mongolia. 19(2): 19–23 (In Russian)
19. Mitrofanova I.V. (ed.) 2018. Fundamentals of in vitro genebank of species, cultivars and forms in ornamental, aromatic and fruit crops: A collective monograph. Simferopol, Arial. 260 p. (In Russian)
20. Mitrofanova I., Lesnikova-Sedoshenko N., Tsiupka V., Smykov A., Mitrofanova O. 2021. Use of biotechnological methods to support the production of new peach hybrids. Horticulturae. 7: 533. DOI: 10.3390/horticulturae7120533.
21. Molkanova O.I., Koroleva O.V., Stacheeva T.S., Krakhmaleva I.L., Meleshchuk E.A. 2018. Improvement of clonal micropropagation technology of valuable fruit and berry crops varieties for commercial conditions. Dostizheniya nauki i tekhniki APK. 32(9): 66–69. (In Russian)
22. Motyleva S., Kozak N., Kulikov I., Medvedev S., Imamkulova Z. 2017. The peculiarities of Actinidia species leaves micromorphology. Agrobiodiversity for Improving Nutrition, Health and Life Quality. 1: 342–346. http://dx.doi.org/10.15414/agrobiodiversity.2017.2585-8246.342-346
23. Muratova S.A., Solovyh N.V., Terekhova V.I. 2011. Induction of Morphogenesis from Isolated Somatic Tissues of Plants. Michurinsk, Michurinsk State Agrarian University Publishing House. 107 p. (In Russian)
24. Plaksina T.V., Borodulina I.D. 2016. The effect of growth regulators on the clonal micropropagation of Actinidia genus. Acta Biologica Sibirica. 2(3): 54–60. (In Russian)
25. POWO. Plants of the World Online, [online] Available at: https://powo.science.kew.org/taxon/ urn:lsid:ipni.org:names:38999-1 (Accessed 10.02.2025).
26. Prado M.J., Herrera M.T. 2005. Micropropagation of two selected male kiwifruit and analysis of genetic variation with AFLP Markers. HortScience. 40(3): 740–746. DOI: 10.21273/HORTSCI.40.3.740.
27. Prozina M.N. 1960. Botanical Microtechniques. Moscow, Vyshaya Shkola. 206 p. (In Russian)
28. Quoirin M., Lepoivre P. 1977. Improved medium for in vitro culture of Prunus sp. Acta Horticulturae. 78: 437–442. DOI: 10.17660/ActaHortic.1977.78.54.
29. Raeva-Bogoslovskaya E., Vinogradova Y., Molkanova O., Hussien M. 2023. Anatomical structures of Saskatoon berry (Amelanchier Medik.) leaves under different cultivation conditions. Bangladesh Journal of Plant Taxonomy. 30(2): 185–193. DOI: 10.3329/bjpt.v30i2.70495.
30. Saeiahagh H., Mousavi M., Wiedow C., Bassett H.B., Pathirana R. 2019. Effect of cytokinins and sucrose concentration on the efficiency of micropropagation of ‘Zes006’ Actinidia chinensis var. chinensis, a red-fleshed kiwifruit cultivar. Plant Cell Tissue and Organ Culture. 138: 1– 10. DOI: 10.1007/s11240-019-01597-4.
31. Semenova D.A., Krakhmaleva I.L., Mishanova E.V., Molkanova O.I., Mitrofanova I.V. 2023. Features of regeneration in vitro in promising Actinidia arguta cultivars. Taurida Herald of the Agrarian Sciences. 1(32): 93–103 (In Russian)
32. Shaheen A., Dewir Y.H., Kher M., Khan M., El-Banna A.N., Alaizari A. 2022. Synergistic effect of benzylaminopurine and meta-Topolin combination for micropropagation of gerbera ‘Pink Melody’. Ciênciae Agrotecnologia. 46:e017521. http://dx.doi.org/10.1590/1413-7054202246017521.
33. Shekhawat J.K., Rai M.K., Shekhawat N.S., Kataria V. 2021. Synergism of m-topolin with auxin and cytokinin enhanced micropropagation of Maytenus emarginata. In Vitro Cellular & Developmental Biology – Plant. 57: 418–426. DOI: 10.1007/s11627-020-10132-6.
34. Shoyama Y., Chen S., Tanaka H., Sasaki Y., Sashida Y. 1998. Actinidia polygama (Japanese name Matatabi): In vitro culture, micropropagation, and the production of monoterpenes and triterpenoids. In: Bajaj Y.P.S., ed. Biotechnology in Agriculture and Forestry. Vol. 41 Medicinal and Aromatic Plants X. Berlin, Springer. P. 1–13.
35. de Souza L.M., Barbosa M.R., Zárate-Salazar J.R., Lozano-Isla F., Camara T.R. 2019. Use of metaTopolin, an unconventional cytokinin in the in vitro multiplication of Opuntia stricta Haw. Biotecnología Vegetal. 19(2): 85–97.
36. State register of varieties and hybrids of agricultural plants approved for use in 2024
37. Sugawara F., Yamamoto N., Tanaka O. 1994. Plant regeneration in in vitro culture of leaf, stem and petiole segments of Actinidia polygama Miq. Plant tissue culture letters. 11(1): 14–18. DOI: 10.5511/plantbiotechnology1984.11.14.
38. Tsiupka V., Zhdanova I.V., Bulavin I.V., Tsiupka S., Mitrofanova I.V. 2022. Ex vitro acclimatization of Lavandula angustifolia Mill. plants. Acta Horticulturae. 1339: 363–370. DOI: 10.17660/ActaHortic.2022.1339.45.
39. Vinogradova Y., Grygorieva O., Vergun O. 2019. Stomatal structure in Solidago L. species as the index of their adaptation opportunities. Agrobiodiversity for Improving Nutrition, Health and Life Quality. 3: 101–110. DOI: 10.15414/agrobiodiversity.2019.2585-8246.101-110.
40. Vorobev V.N., Nevmerzhickaya Y.Y., Khusnetdinova L.Z., Yakushenkova T.P. 2013. Practicum on Plant Physiology: Textbook. Kazan, Kazan University. 80 p. (In Russian)
41. Wang F., Xin X., Wei H., Qiu X., Liu B. 2020. In vitro regeneration, ex vitro rooting and foliar stoma studies of Pseudostellaria heterophylla (Miq.) Pax. Agronomy. 10: 949. DOI: 10.3390/agronomy10070949.
42. Wang S., Qiu Y., Zhu F. 2021. Kiwifruit (Actinidia spp.): A review of chemical diversity and biological activities. Food Chemistry. 350: 128469. DOI: 10.1016/j.foodchem.2020.128469.
43. Werner E.T., Milanez C.R.D., Gontijo A.B.P.L., Soares T.C.B., Amaral J.A.T. 2018. Leaf anatomy changes related to cultivate in vivo and in vitro and during pre-acclimatization of Crambe abyssinica Hochst. Plant Cell Culture & Micropropagation. 14(1): 10–17.
44. Zaytseva Y.G., Ambros E.V., Novikova T.I. 2021. Meta-topolin: Advantages and disadvantages for in vitro propagation. In: Ahmad N., Strnad M., eds. Meta-topolin: a growth regulator for plant biotechnology and agriculture. Singapore, Springer. P. 119–141. DOI: 10.1007/978-981-15-9046-7_11.
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For citations:
Krakhmaleva I.L., Molkanova O.I. Regeneration features and leaf anatomical struc- ture of Actinidia polygama under in vitro conditions. New Bull. MBG. 2025;1(1-2):48-64. (In Russ.) https://doi.org/10.35102/cbg.2025.40.28.004
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