Fìzìol. rosl. genet. 2025, vol. 57, no. 2, 95-116, doi: https://doi.org/10.15407/frg2025.02.095

Problems of experimental design in research on drought resistance of plants

Kolupaev Y.E.1,2,3, Lugova G.A.2, Rozhkov R.V.2, Karpets Y.V.2

  1. Yuryev Plant Production Institute, National Academy of Agrarian Sciences of Ukraine  142 Heroiv Kharkova Ave., 61060 Kharkiv, Ukraine
  2. State Biotechnological University  44 Alchevskikh St., 61002 Kharkiv, Ukraine
  3. Poltava State Agrarian University  1/3 Skovorody St., 36003 Poltava, Ukraine

Drought is now considered the most harmful weather phenomena for the global economy. In view of this, increasing the drought resistance of plants is important not only for agriculture as an industry, but also for society as a whole. The use of classical and new breeding and genetic approaches (including editing plant genomes), as well as biotechnological tools associated with the use of physiologically active substances (resistance inducers), requires improving methodological approaches in the study of drought resistance of plants. The review provides a critical analysis of the main methods for creating experimental drought and choosing integral plant indice that characterize their drought resistance. Experimental approaches to studying the main strategies for survival and functioning of cultivated plants under conditions of moisture deficiency are discussed: the isohydric strategy, which is based on water conservation, and the anisohydric strategy, which is based on resistance to dehydration. In the context of these two strategies, the issues of studying the functioning of the main stress-protective systems in plants are considered: osmoregulatory and antioxidant. Emphasis is placed on the specific features of drought tolerance of cultivated cereals, in particular, Triticum aestivum. Approaches to studying the relationships between integral indice of drought tolerance and changes in markers of oxidative stress, antioxidant activity, and increased accumulation of multifunctional low-molecular protective compounds are discussed. The feasibility of using mathematical methods, in particular, meta-analysis for objective generalization of experimental data in the field of plant drought tolerance is noted.

Keywords: drought resistance of plants, experimental approaches, adaptive strategies of plants, osmotic regulation, oxidative stress, antioxidant system

Fìzìol. rosl. genet.
2025, vol. 57, no. 2, 95-116

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References

1. Boyer, J.S. (1982). Plant productivity and environment. Science, 218(4571), pp. 443-448. https://doi.org/10.1126/science.218.4571.443

2. Dietz, K.J., ZШrb, C. & Geilfus, C.M. (2021). Drought and crop yield. Plant Biol., 23(6), pp. 881-893. https://doi.org/10.1111/plb.13304

3. Ilyas, M., Nisar, M., Khan, N., Hazrat, A., Khan, A.H., Hayat, K., Fahad, S., Khan, A. & Ullah, A. (2021). Drought tolerance strategies in plants: A mechanistic approach. J. Plant Growth Regul., 40, pp. 926-944. https://doi.org/10.1007/s00344-020-10174-5

4. Feng, D., Liu, W., Chen, K., Ning, S., Gao, Q., Chen, J., Liu, J., Sun, X. & Xu, W. (2024). Exogenous substances used to relieve plants from drought stress and their associated underlying mechanisms, Int. J. Mol. Sci., 25, 9249. https://doi.org/10.3390/ijms25179249

5. Lau, S.-E., Lim, L.W.T., Hamdan, M.F., Chan, C., Saidi, N.B., Ong-Abdullah, J. & Tan, B.C. (2025). Enhancing plant resilience to abiotic stress: The power of biostimulants, Phyton-Int. J. Exp. Bot., 94(1), pp. 1-31. https://doi.org/10.32604/phyton.2025.059930

6. Plessis, A. (2023). Abiotic stress experiments need a reality check to improve translation to the field. J. Exp. Bot., 74(6), pp. 1741-1744. https://doi.org/10.1093/jxb/erac509

7. Bhardwaj, A., Devi, P., Chaudhary, S., Rani, A., Jha, U.C., Kumar, S., Bindumadhava, H., Prasad, P.V.V., Sharma, K.D., Siddique, K.H.M. & Nayyar, H. (2022). 'Omics' approaches in developing combined drought and heat tolerance in food crops. Plant Cell Rep., 41(3), pp. 699-739. https://doi.org/10.1007/s00299-021-02742-0

8. Nick, P. (2024). Towards a grammar of plant stress: modular signalling conveys meaning. Theor. Exp. Plant Physiol., 36(3), pp. 503-521. https://doi.org/10.1007/s40626-023-00292-2

9. Bandurska, H. (2022). Drought stress responses: coping strategy and resistance. Plants, 11, 922. https://doi.org/10.3390/plants11070922

10. Wang, X., Li, X., Zhao, W., Hou, X. & Dong, S (2024). Current views of drought research: experimental methods, adaptation mechanisms and regulatory strategies. Front. Plant Sci. 15, 1371895. https://doi.org/10.3389/fpls.2024.1371895

11. Blum, A. (2011). Drought resistance - is it really a complex trait?. Funct. Plant Biol., 38(10), pp. 753-757. https://doi.org/10.1071/FP11101

12. Fang, Y. & Xiong, L. (2015). General mechanisms of drought response and their application in drought resistance improvement in plants. Cell. Mol. Life Sci., 72(4), pp. 673-689. https://doi.org/10.1007/s00018-014-1767-0

13. Rojas, O. (2020). Agricultural extreme drought assessment at global level using FAO-Agricultural stress index system (ASIS). Weather Clim. Extrem., 27, 100184. https://doi.org/10.1016/j.wace.2018.09.001

14. Lawlor, D.W. (2013). Genetic engineering to improve plant performance under drought: physiological evaluation of achievements, limitations, and possibilities. J. Exp. Bot., 64, pp. 83-108. https://doi.org/10.1093/jxb/ers326

15. Pandolfi, C., Mancuso, S. & Shabala, S. (2012). Physiology of acclimation to salinity stress in pea (Pisum sativum). Environ. Exp. Bot., 84, pp. 44-51. https://doi.org/10.1016/j.envexpbot.2012.04.015

16. Galviz, Y., Souza, G.M. & Lтttge, U. (2022). The biological concept of stress revisited: relations of stress and memory of plants as a matter of space-time. Theor. Exp. Plant Physiol., 34, pp. 239-264. https://doi.org/10.1007/s40626-022-00245-1

17. Kolupaev, Yu.E. (2010). Fundamentals of the Physiology of Plant Resistance. Kharkiv [in Ukrainian].

18. Tardieu, F. & Simonneau, T. (1998). Variability among species of stomatal control under fluctuating soil water status and evaporative demand: Modelling isohydric and anisohydric behaviours. J. Exp. Bot., 49, pp. 419-432. https://doi.org/10.1093/jxb/

19. Blum, A. (2011). Drought Resistance and Its Improvement. In: Plant Breeding for Water-Limited Environments. Springer, New York, pp. 53-152. https://doi.org/10.1007/978-1-4419-7491-4_3

20. Takahashi, F., Kuromori, T., Urano, K., Yamaguchi-Shinozaki, K. & Shinozaki, K. (2020). Drought stress responses and resistance in plants: From cellular responses to long-distance intercellular communication. Front. Plant Sci., 11, 556972. https://doi.org/10.3389/fpls.2020.556972

21. Mahmood, T., Khalid, S., Abdullah, M., Ahmed, Z., Shah, M.K.N., Ghafoor, A. & Du, X. (2019). Insights into drought stress signaling in plants and the molecular genetic basis of cotton drought tolerance. Cells, 2019, 9(1), 105. https://doi.org/10.3390/cells9010105

22. Rane, J., Singh, A.K., Tiwari, M., Prasad, P.V. & Jagadish, S.K. (2022). Effective use of water in crop plants in dryland agriculture: implications of reactive oxygen species and antioxidative system. Front Plant Sci., 12. https://doi.org/10.3389/fpls.2021.778270

23. Kolupaev, Yu.E., Yastreb, T.O., Ryabchun, N.I., Kokorev, A.I., Kolomatska, V.P. & Dmitriev, A.P., (2023). Redox homeostasis of cereals during acclimation to drought. Theor. Exp. Plant Physiol., 35(2), pp. 133-168. https://doi.org/10.1007/s40626-023-00271-7

24. Spollen, W.G., LeNoble, M.E., Samuels, T.D., Bernstein, N. & Sharp, R.E. (2000). Abscisic acid accumulation maintains maize primary root elongation at low water potentials by restricting ethylene production. Plant Physiol., 122(3), pp. 967-976. https://doi.org/10.1104/pp.122.3.967

25. Ogura, T., Goeschl, C., Filiault, D., Mirea, M., Slovak, R., Wolhrab, B., Satbhai, S.B. & Busch, W. (2019). Root system depth in arabidopsis is shaped by EXOCYST70A3 via the dynamic modulation of auxin transport. Cell, 178(2), pp. 400-412.e16. https://doi.org/10.1016/j.cell.2019.06.021

26. Rajasheker, G., Jawahar, G., Jalaja, N., Kumar, S.A., Kumari, P.H., Punita, D.L., Karumanchi, A.R., Palakolanu, S.R., Polavarapu, R., Sreenivasulu, N. & Kavi Kishor, P.B. (2019). Role and Regulation of Osmolytes and ABA Interaction in Salt and Drought Stress Tolerance. In: Khan, M.I.R., Reddy, P.S., Ferrante, A., Khan, N.A. (eds.) Plant Signaling Molecules. Woodhead Publishing: Sawston, UK, 2019; pp. 417-436. https://doi.org/10.1016/B978-0-12-816451-8.00026-5

27. Vandeleur, R.K., Mayo, G., Shelden, M.C., Gilliham, M., Kaiser, B.N. & Tyerman, S.D. (2009). The role of plasma membrane intrinsic protein aquaporins in water transport through roots: diurnal and drought stress responses reveal different strategies between isohydric and anisohydric cultivars of grapevine. Plant Physiol., 149(1), pp. 445-460. https://doi.org/10.1104/pp.108.128645

28. Juenger, T.E. & Verslues, P.E. (2023). Time for a drought experiment: Do you know your plants' water status?. Plant Cell, 35(1), pp. 10-23. https://doi.org/10.1093/plcell/koac324

29. Gangola, M.P. & Ramadoss, B.R. (2018). Sugars play a critical role in abiotic stress tolerance in plants. In: Wani, S.H. (ed.) Biochemical, physiological and molecular avenues for combating abiotic stress tolerance in plants, Academic Press, pp. 17-38. https://doi.org/10.1016/B978-0-12-813066-7.00002-4

30. Ende, W.V. & Peshev, D. (2013). Sugars as antioxidants in plants. In: Tuteja N., Gill S. (eds). Crop improvement under adverse conditions. Springer, New York, pp. 285-307. https://doi.org/10.1007/978-1-4614-4633-0_13

31. Caffery, M., Tonseca, V. & Carl Leopold, A. (1988). Lipid-sugar interaction: Relevance to anhydrous biology. Plant Physiol., 86(3), pp. 754-758. https://doi.org/10.1104/pp.86.3.754

32. Kosar, F., Akram, N.A., Sadiq, M., Al-Qurainy, F. & Ashraf, M. (2019). Trehalose: A key organic osmolyte effectively involved in plant abiotic stress tolerance. J. Plant Growth Regul., 38, pp. 606-618. https://doi.org/10.1007/s00344-018-9876-x

33. Per, T.S., Khan, N.A., Reddy, P.S., Masood, A., Hasanuzzaman, M., Khan, M.I.R. & Anjum, N.A. (2017). Approaches in modulating proline metabolism in plants for salt and drought stress tolerance: Phytohormones, mineral nutrients and transgenics. Plant Physiol. Biochem., 115, pp. 126-140. https://doi.org/10.1016/j.plaphy.2017.03.018

34. Zarattini, M. & Forlani, G. (2017). Toward unveiling the mechanisms for transcriptional regulation of proline biosynthesis in the plant cell response to biotic and abiotic stress conditions. Front. Plant Sci., 8, 927. https://doi.org/10.3389/fpls.2017.00927

35. Moukhtari, A., Cabassa-Hourton, C., Farissi, M. & Savoure, A. (2020). How does proline treatment promote salt stress tolerance during crop plant development? Front. Plant Sci., 11, 1127. https://doi.org/10.3389/fpls.2020.01127

36. Signorelli, S., Dans, P.D., CoitiФo, E.L., Borsani, O. & Monza, J. (2015). Connecting proline and g-aminobutyric acid in stressed plants through non-enzymatic reactions. PLoS ONE, 10(3), e0115349. https://doi.org/10.1371/journal.pone.0115349

37. de Carvalho, K., Campos, M.K., Domingues, D.S., Pereira, L.F. & Vieira, L.G. (2013). The accumulation of endogenous proline induces changes in gene expression of several antioxidant enzymes in leaves of transgenic Swingle citrumelo. Mol. Biol. Rep., 40, pp. 3269-3279. https://doi.org/10.1007/s11033-012-2402-5

38. Dubrovna, O.V., Priadkina, G.O., Mykhalska, S.I. & Komisarenko, A.G. (2022). Drought-tolerance of transgenic winter wheat with partial suppression of the proline dehydrogenase gene. Regul. Mech. Biosyst., 13(4), pp. 385-392. https://doi.org/10.15421/022251

39. Dubrovna, O.V., Stasik, O.O., Priadkina, G.O., Zborivska, O.V. & Sokolovska-Sergiienko, O.G. (2020). Resistance of genetically modified wheat plants, containing a doublestranded RNA suppressor of the proline dehydrogenase gene. Agricul. Sci. Pract., 7(2), pp. 24-34. https://doi.org/10.15407/agrisp7.02.024

40. Mansour MMF. & Salama KHA. (2020). Proline and abiotic stresses: Responses and adaptation. In: Hasanuzzaman M. (ed.). Plant Ecophysiology and Adaptation under Climate Change: Mechanisms and Perspectives II. Springer, Singapore, pp. 357-397. https://doi.org/10.1007/978-981-15-2172-0_12

41. Rajendrakumar, C.S., Reddy, B.V. & Reddy, A.R. (1994). Proline-protein interactions: protection of structural and functional integrity of M4 lactate dehydrogenase. Biochem. Biophys. Res. Commun., 201(2), pp. 957-963. https://doi.org/10.1006/bbrc.1994.1795

42. Sun, Z., Li, S., Chen, W., Zhang, J., Zhang, L., Sun, W. & Wang, Z. (2021). Plant dehydrins: Expression, regulatory networks, and protective roles in plants challenged by abiotic stress. Int. J. Mol. Sci., 22, 12619. https://doi.org/10.3390/ijms222312619

43. Szlachtowska, Z. & Rurek, M. (2023). Plant dehydrins and dehydrin-like proteins: characterization and participation in abiotic stress response. Front. Plant Sci., 14, 1213188. https://doi.org/10.3389/fpls.2023.1213188

44. Drira, M., Saibi, W., Brini, F., Gargouri, A., Masmoudi, K. & Hanin, M. (2013). The K-segments of the wheat dehydrin DHN-5 are essential for the protection of lactate dehydrogenase and b-glucosidase activities in vitro. Mol. Biotechnol., 54(2), pp. 643-650. https://doi.org/10.1007/s12033-012-9606-8

45. Pykalo, S., Demydov, O., Yurchenko, T., Khomenko, S., Humeniuk, O., Kharchenko, M. & Prokopik, N. 2020. Methods for evaluation of wheat breeding material for drought tolerance. Visnyk of the Lviv University. Series Biology, 82, pp. 63-79. [in Ukrainian]. https://doi.org/10.30970/vlubs.2020.82.05

46. Frantov«, N., R«bek, M., Elzner, P., Stйeda, T., Jovanoviє, I., Holkov«, L., Martinek, P., Smutn«, P. & Pr«лil, I.T. (2022). Different drought tolerance strategy of wheat varieties in spike architecture. Agronomy, 12(10), 2328. https://doi.org/10.3390/agronomy12102328

47. Zhang, Y.M., Hu, H.Y., Bai, X.M., Cory, M., Javier, G.F. & Iv«n, P.O. (2022). Effects of soil water restriction on root growth and root morphology of perennial Rye grass and pasture brome. Chin. J. Eco-Agriculture, 30, pp. 1784-1794. https://doi.org/ 10.12357/cjea.20220336

48. He, Z., Wang, F. & Bo, X. (2019). A comparative study on the effects of drought stress simulated by PEG6000 and mannitol. Anhui Agric. Sci. Bull., (10), pp. 19-40. https://doi.org/10.16377/j.cnki.issn1007-7731.2019.10.009

49. Borysova, O.V. & Ruzhitska, O.M. (2014). Physiological and biochemical parameters of Triticum aestivum L. and Triticum spelta L. wheats under water deficit modeling J V.N.Karazin Kharkiv Nat. Univ., Ser. Biol., 23(1129), pp. 81-88 [in Ukrainian].

50. Bulavka, N.V., Yurchenko, T.V., Kucherenko, O.M. & Pirych, A.V. (2018). Soft winter wheat varieties with resistance to negative environmental factors. Plant Var. Stud. Protect., 14(3), 255-261 [in Ukrainian]. https://doi.org/10.21498/2518-1017.14.3.2018.145285

51. Kolupaev, Yu.E., Yastreb, T.O., Salii, A.M., Kokorev, A.I., Ryabchun, N.I., Zmiievska, O.A. & Shkliarevskyi, M.A. (2022). State of antioxidant and osmoprotective systems in etiolated winter wheat seedlings of different cultivars due to their drought tolerance. Zemdirbyste-Agriculture, 109(4), pp. 313-322. https://doi.org/10.13080/z-a.2022.109.040

52. Kolupaev, Yu.E., Ryabchun, N.I., Leonov, O.Yu., Kokorev, A.I., Taraban, D.A., Shakhov, I.V., Shkliarevskyi, M.A. & Yastreb, T.O. (2024). Functioning of the antioxidant and osmoprotective systems of Triticum aestivum cultivars growing under soil drought conditions. Botanica, 30(3), pp. 102-116. https://doi.org/10.35513/botlit

53. Karpets, Yu.V., Taraban, D.A., Kokorev, A.I., Yastreb, T.O., Kobyzeva, L.N. & Kolupaev, Yu.E. (2023). Response of wheat seedlings with different drought tolerance to melatonin action under osmotic stress. Agriculture and Forestry, 69(5), pp. 53-69. https://doi.org/10.17707/AgricultForest.69.4.05

54. Kolupaev, Yu.E., Ryabchun, N.I., Yastreb, T.O., Kokorev, O.I. & Shakhov I.V. (2023). Assessment of the stress-protective effect of physiologically active substances on grain crops. Scientific and methodological recommendations. Kharkiv [in Ukrainian].

55. Chowdhury, M.K., Hasan, M.A., Bahadur, M.M., Islam, M.R., Hakim, M.A., Iqbal, M.A., Javed, T., Raza, A., Shabbir, R., Sorour, S., Elsanafawy, N.E.M., Anwar, S., Alamri, S., Sabagh, A.E. & Islam, M.S. (2021). Evaluation of drought tolerance of some wheat (Triticum aestivum L.) genotypes through phenology, growth, and physiological indices. Agronomy, 11(9), 1792. https://doi.org/10.3390/agronomy11091792

56. Ahmad, A., Aslam, Z., Javed, T., Hussain, S., Raza, A., Shabbir, R., Mora-Poblete, F., Saeed, T., Zulfiqar, F., Ali, M.M., Nawaz, M., Rafiq, M., Osman, H.S., Albaqami, M., Ahmed, M.A.A. & Tauseef, M. (2022). Screening of wheat (Triticum aestivum L.) genotypes for drought tolerance through agronomic and physiological response. Agronomy, 12, 287. https://doi.org/10.3390/agronomy12020287

57. Laxa, M., Liebthal, M., Telman, W., Chibani, K. & Dietz, K.J. (2019). The role of the plant antioxidant system in drought tolerance. Antioxidants, 8(4), 94. https://doi.org/10.3390/antiox8040094

58. Barrs, H. & Weatherley, P. (1962). A re-examination of the relative turgidity technique for estimating water deficits in leaves. Aust. J. Biol. Sci., 15(3), pp. 413-428. https://doi.org/10.1071/BI9620413

59. Bajji, M., Kinet, J.-M. & Lutts, S. (2002). The use of electrolyte leakage method for assessing cell membrane stability as a water stress tolerance test in durum wheat. Plant Growth Regul., 36, pp. 61-70. https://doi.org/10.1023/A:1014732714549

60. Blum, A. & Tuberosa, R. (2018). Dehydration survival of crop plants and its measurement. J. Exp. Bot., 69(5), pp. 975-981. https://doi.org/10.1093/jxb/erx445

61. Sun, Y., Wang, C., Chen, H.Y.H. & Ruan, H. (2020). Response of plants to water stress: a meta-analysis. Front. Plant Sci., 11, 978. https://doi.org/10.3389/fpls.2020.00978

62. Khatun, M., Sarkar, S., Era, F.M., Islam, A.K.M.M., Anwar, M.P., Fahad, S., Datta, R. & Islam, A.K.M.A. (2021). Drought Stress in Grain Legumes: Effects, Tolerance Mechanisms and Management. Agronomy, 11, 2374. https://doi.org/10.3390/agronomy11122374

63. Bao, X., Hou, X., Duan, W., Yin, B., Ren, J., Wang, Y., Liu, X., Gu, L. & Zhen, W. (2023). Screening and evaluation of drought resistance traits of winter wheat in the North China. Plain. Front. Plant Sci., 14, 1194759. https://doi.org/10.3389/fpls.2023.1194759

64. Kirova, E.,·Moskova, I.,·Geneva, M.·& Kocheva, K. (2022) Antioxidant potential of tolerant and susceptible wheat varieties under drought and recovery. Cereal Res. Commun., 50, pp. 841-849. https://doi.org/10.1007/s42976-021-00222-5

65. Amoah, J.N., Ko, C.S., Yoon, J.S. & Weon, S.Y. Effect of drought acclimation on oxidative stress and transcript expression in wheat (Triticum aestivum L.). J. Plant Int., 2019, 14, No. 1, pp. 492-505. https://doi.org/10.1080/17429145.2019.1662098

66. Moloi, S.J., Alqarni, A.O., Brown, A.P., Goche, T., Shargie, N.G., Moloi, M.J., Gokul, A., Chivasa, S. & Ngara, R. (2024). Comparative physiological, biochemical, and leaf proteome responses of contrasting wheat varieties to drought stress. Plants, 13(19), 2797. https://doi.org/10.3390/plants13192797

67. Mar№ek, T., Hamow, K.A., Vegh, B., Janda, T. & Darko, E. (2019). Metabolic response to drought in six winter wheat genotypes. PLoS ONE, 14(2), e0212411. https://doi.org/10.1371/journal.pone.0212411

68. Upadhyay, D., Budhlakoti, N., Singh, A.K., Bansal, R., Kumari, J., Chaudhary, N., Padaria, J.C., Sareen, S. & Kumar, S. (2020). Drought tolerance in Triticum aestivum L. genotypes associated with enhanced antioxidative protection and declined lipid pe­roxidation. 3 Biotech., 10(6), 281. https://doi.org/10.1007/s13205-020-02264-8

69. Zadoks, J.C., Chang, T.T. & Konzak, C.F. (1974). A decimal code for the growth stages of cereals. Weed Res., 14, pp. 415-421. https://doi.org/10.1111/j.1365-3180.1974.tb01084.x

70. Cheng, L., Wang, Y., He, Q., Li, H., Zhang, X. & Zhang, F. (2016). Comparative proteomics illustrates the complexity of drought resistance mechanisms in two wheat (Triticum aestivum L.) cultivars under dehydration and rehydration. BMC Plant Biol., 16, pp. 1-23. https://doi.org/10.1186/s12870-016-0871-8

71. Qayyum, A., Razzaq, A., Bibi, Y., Khan, S.U., Abbasi, K.S., Sher, A., Mehmood, A., Ahmed, W., Mahmood, I., Manaf, A., Khan, A., Farid, A. & Jenks, M.A. (2018). Water stress effects on biochemical traits and antioxidant activities of wheat (Triticum aestivum L.) under in vitro conditions. Acta Agricult. Scandinav. Sect. B - Soil Plant Sci., 68(4), pp. 283-290. https://doi.org/10.1080/09064710.2017.1395064

72. Nasirzadeh, L., Sorkhilaleloo, B., Majidi Hervan, E. & Fatehi, F. (2021). Changes in antioxidant enzyme activities and gene expression profiles under drought stress in tole­rant, intermediate, and susceptible wheat genotypes. Cereal Res. Commun., 49(1), pp. 83-89. https://doi.org/10.1007/s42976-020-00085-2

73. Simova-Stoilova, L., Demirevska, K., Petrova, T., Tsenov, N. & Feller U. (2009). Antioxidative protection and proteolytic activity in tolerant and sensitive wheat (Triticum aestivum L.) varieties subjected to long-term field drought. Plant Growth Regul., 58(1), pp. 107-117. https://doi.org/10.1007/s10725-008-9356-6

74. Ford, K.L., Cassin, A. & Bacic, A.F. (2011). Quantitative proteomic analysis of wheat cultivars with differing drought stress tolerance. Front. Plant Sci., 2, 44. https://doi.org/10.3389/fpls.2011.00044

75. Harb, A., Awad, D. & Samarah, N. (2015). Gene expression and activity of antioxidant enzymes in barley (Hordeum vulgare L.) under controlled severe drought. J. Plant Interact., 10(1), pp. 109-116. https://doi.org/10.1080/17429145.2015.1033023

76. Marok, M.A., Tarrago, L., Ksas, B., Henri. P., Abrous-Belbachir, O., Havaux, M. & Rey, P. (2013). A drought-sensitive barley variety displays oxidative stress and strongly increased contents in low-molecular weight antioxidant compounds during water deficit compared to a tolerant variety. J. Plant Physiol., 170(7), pp. 633-645. https://doi.org/10.1016/j.jplph.2012.12.008

77. Gurrieri, L., Merico, M., Trost, P., Forlani, G. & Sparla, F. (2020). Impact of drought on soluble sugars and free proline content in selected Arabidopsis mutants. Biology, 9(11), 367. https://doi.org/10.3390/biology9110367

78. Faisal, S., Mujtaba, S.M., Asma & Mahboob, W. (2019). Polyethylene glycol mediated osmotic stress impacts on growth and biochemical aspects of wheat (Triticum aestivum L.). J. Crop Sci. Biotechnol., 22, pp. 213-223. https://doi.org/10.1007/s12892-018-0166-0

79. Ghaffar, A., Hussain, N., Ajaj, R., Shahin, S.M., Bano, H., Javed, M., Khalid, A., Yasmin, M., Shah, K.H., Zaheer, M., Iqbal, M., Zafar, Z.U. & Athar, H-u-R. (2023). Photosynthetic activity and metabolic profiling of bread wheat cultivars contrasting in drought tolerance. Front. Plant Sci., 14, 1123080. https://doi.org/10.3389/fpls.2023.1123080

80. Singh, S., Prakash, P. & Singh, A.K. (2021). Salicylic acid and hydrogen peroxide improve antioxidant response and compatible osmolytes in wheat (Triticum aestivum L.) under water deficit. Agric. Res., 10, pp. 175-186. https://doi.org/10.1007/s40003-020-00490-3

81. MarciXska, I., CzyczyYo-Mysza, I., Skrzypek, E., Filek, M., Grzesiak, S., Grzesiak, M.T., Janowiak, F., Hura, T., Dziurka, M., Dziurka, K., Nowakowska, A. & Quarrie, S.A. (2013). Impact of osmotic stress on physiological and biochemical characteristics in drought-susceptible and drought-resistant wheat genotypes. Acta Physiol. Plant., 35, pp. 451-461. https://doi.org/10.1007/s11738-012-1088-6

82. Abid, M., Ali, S., Qi, L.K., Zahoor, R., Tian, Z., Jiang, D., Snider, J.L. & Dai, T. (2018). Physiological and biochemical changes during drought and recovery periods at tillering and jointing stages in wheat (Triticum aestivum L.). Sci. Rep., 8, 4615. https://doi.org/10.1038/s41598-018-21441-7

83. Schneider, J.R., Wurlitzer, W.B., Ferla, N.J., Huzar-Novakowiski, J. & Chavarria, G. (2025). A metanalytic study: does water deficit always increase soybean proline concentration?. Theor. Exp. Plant Physiol., 37, 4. https://doi.org/10.1007/s40626-024-00346-z

84. Anjum, S.A., Ashraf, U,. Tanveer, M., Khan, I., Hussain, S., Shahzad, B., Zohaib, A., Abbas, F., Saleem, M.F., Ali, I. & Wang, L.C. (2017). Drought induced changes in growth, osmolyte accumulation and antioxidant metabolism of three maize hybrids. Front. Plant Sci., 8, 69. https://doi.org/10.3389/fpls.2017.00069

85. Ayub, M., Ashraf, M.Y., Kausar, A., Saleem, S., Anwar, S., Altay, V. & Munir, O. (2021). Growth and physio-biochemical responses of maize (Zea mays L.) to drought and heat stresses. Plant Biosystems, 155(3), pp. 535-542. https://doi.org/10.1080/11263504.2020.1762785

86. Samota, M.K., Sasi, M. & Singh, A. (2017). Impact of seed priming on proline content and antioxidant enzymes to mitigate drought stress in rice genotype. Int. J. Curr. Microbiol. App. Sci., 6(5), pp. 2459-2466. https://doi.org/10.20546/ijcmas.2017.605.275

87. Panda, D., Rath, C., Behera, P.K. & Lenka, S.K. (2021). Physiological introspection of leaf photochemical activity and antioxidant metabolism in selected indigenous finger millet genotypes in relation to drought stress. Cereal Res. Commun., 49(4), pp. 607-618. https://doi.org/10.1007/s42976-021-00132-6

88. Cheng, L., Wang, Y., He, Q., Li, H., Zhang, X. & Zhang, F. (2016). Comparative proteomics illustrates the complexity of drought resistance mechanisms in two wheat (Triticum aestivum L.) cultivars under dehydration and rehydration. BMC Plant Biol., 16, pp. 1-23. https://doi.org/10.1186/s12870-016-0871-8

89. Kaur, G. & Asthir, B.J.B.P. (2015). Proline: a key player in plant abiotic stress tolerance. Biol. Plant, 59(4), pp. 609-619. https://doi.org/10.1007/s10535-015-0549-3

90. Nasirzadeh, L, Sorkhilaleloo, B., Majidi Hervan, E. & Fatehi, F. (2021). Changes in antioxidant enzyme activities and gene expression profiles under drought stress in tole­rant, intermediate, and susceptible wheat genotypes. Cereal Res. Commun., 49(1), pp. 83-89. https://doi.org/10.1007/s42976-020-00085-2

91. Kolupaev, Yu.E. & Shkliarevskyi, M.A. (2025). Accumulation of proline in vegetative organs of Triticum aestivum (L.) under drought conditions in the early phases of development: a meta-analysis of data. Ukr. Bot. J., 82(4) [in Ukrainian].

92. Wu H. & Yang Z. (2024). Effects of drought stress and postdrought rewatering onwinter wheat: A meta-analysis. Agronomy, 14, 298. https://doi.org/10.3390/agronomy14020298

93. Rymar, Yu.Yu., Pronina, O.V., Kiriziy, D.A., Duplij, V.P., Morgun, B.V., Stasik, O.O. (2025). Characteristics of flag leaf stomata in relation to gas exchange rate and drought tolerance in related spring wheat species. Fiziol. rosl. genet., 57(1), pp. 64-82 [in Ukrainian]. https://doi.org/10.15407/frg2025.01.064