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Evgeny Aleksandrovich Gladkov, Yuliya Ivanovna Dolgikh & Olga Victorovna Gladkova

Biotechnological aspects of obtaining salt-tolerant plants

(Volume 91 - Année 2022 — Numéro 1)
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Soil salinization is an important ecological problem in urban ecosystems and agro-ecosystems. Salinization has direct adverse effects on plants. Sodium chloride showed greater toxicity than bischofite to Festuca rubra. We used cell selection to increase the salt tolerance of the plants. We used sodium chloride in all stages of cultivation.

We have obtained salt-tolerant Festuca rubra plants. Most Festuca rubra regenerants showed increased salt tolerance. Thus, biotechnological methods can be used to obtain lawn grasses that are resistant to salinity.

Keywords : soil salinization, Festuca rubra, biotechnological methods

1Manuscrit reçu le 2 octobre 2022 et accepté le 14 novembre 2022

2Article publié selon les termes et les conditions de la licence Creative Commons CC-BY 4.0

1. Introduction

3Soil salinization is an important ecological problem in urban ecosystems and agro-ecosystems. Globally, this environmental factor is one of the main land-degrading threats influencing soil fertility (Hassani et al., 2020). Soil salinity and sodicity, called collectively salt-affected soils, affect approximately 932 million ha of land globally (Wong et al., 2010). 20% of cultivated land in the world, and 33% of irrigated land, are salt-affected and degraded (Machado et al., 2017).

4The use of deicing reagents causes salinisation of urban ecosystems (Gladkov, Gladkova, 2022). Salinization has an adverse effect on plants. Indeed , salinity stress significantly reduced the plant growth, leaf water potential and relative water content in five Coleus species (Kotagiri et al., 2017). The salinity affected both growth and physiological attributes of the maize species (Zahra et al., 2020).

5Crop loss due to soil salinization is an increasing threat to agriculture worldwide (Van Zelm et al., 2020). This environmental factor endangers the development of ecological agriculture (Chen et al., 2022).

6Alternative uses for saline land are proposed. High salinity land may provide an alternative resource for the cultivation of dedicated biomass crops (Stavridou et al. 2017).

7Improving plant resistance to salinity is an important task for many researchers. The use of biotechnological methods to improve salt tolerance is a promising development. These methods are used to increase resistance to salinization and other stresses (Gladkov et al., 2021; Gladkov & Gladkova, 2022). The salt tolerance of some crop plants and cell lines has been increased. Cell lines in Solanum tuberosum resistant to sodium chloride have been obtained (Queiros et al., 2007).

8The regenerants obtained from the salt-treated showed improved growth characteristics(Al-Khateeb et al., 2020).

9An efficient regeneration protocol was applied to regenerate shoots on salt stress-tolerant calli lines of aubergine (Solanum melongena) (Hannachi et al., 2021).

10Biotechnological methods can be used in urban greening (Gladkov, Gladkova, 2021). Cell selection has been used to obtain salt-tolerant Agrostis stolonifera plants. (Gladkov et al., 2014; Gladkov, Gladkova, 2022).

11The aim of the present work was to develop the biotechnological method of increasing the salt tolerance of the lawn grass Festuca rubra.

2. Materials and methods

12The phytotoxicity of sodium chloride and magnesium chloride (deicing reagent bischofite, magnesium chloride hexahydrate (MgCl2·6H2O)) was assessed in water solutions of the toxicants. 10 seeds each were placed in Petri dishes on filter paper moistened with toxicant solutions. Water was used as a control.

13Calli of Festuca rubra was obtained from seeds on Murashige and Skoog modified medium with 3 mg/l 2,4- dichlorophenoxyacetic acid (Gladkov et al., 2014).

14Determination of the sensitivity of callus cultures to sodium chloride was carried out by morphogenesis ability. We counted the percentage of morphogenic calli in relation to all calli. Only surviving calli were evaluated. Calli with green buds were considered morphogenic (Gladkov et al., 2014).

15To select tolerant clones calli of Festuca rubra cultivated on Murashige and Skoog modified medium with 1 mg/l 2,4-diclorophenoxiacetic acid and 1% sodium chloride. The cultures were maintained in a growth room at 26 °C, exposed to a 16-h photoperiod in the light and humidity 70% (Gladkov et al., 2014).

16The tolerance of the obtained regenerants was evaluated in soil. We assessed the increase in plant growth.

3. Results and discussion

17Various deicing reagents are used in Russia. Deicing reagents are among the priority pollutants of the Moscow soils. All reagents used that contain chlorine are hazardous to plants. Sodium chloride is widely used as deicing reagent. It showed greater toxicity than magnesium chloride (bischofite, magnesium chloride hexahydrate) to Festuca rubra (Figure 1). Therefore, obtaining plants resistant to sodium chloride is the most appropriate.

Image 10000201000001BF0000017BE1C86C49B04DFE6E.png

Figure 1. Effect of 1% deicing reagent (sodium chloride; bischofite) on Festuca rubra plants

18We used cell selection to increase the salt tolerance of the plants.

19When developing a cell selection scheme, the sensitivity of calli to toxicant (Gladkov, Gladkova, 2020), the choice of selective concentration and the duration of cultivation are of prime importance. An inhibitory effect on the callus was observed at 1% sodium chloride. 56% of the surviving calli retained the ability for morphogenesis. The concentration of 1% NaCl was chosen as selective because some surviving callus retained their morphogenic ability. Our research has shown that the use of higher concentrations of sodium chloride significantly reduces the possibility of obtaining regenerants. The scheme for cell selection to obtain salt-tolerant plants can vary depending on the duration of cultivation (Figure 2).

Image 1000020100000209000001DC479614D7131BF3C7.png

Figure 2. Cell selection scheme for obtaining Festuca rubra plants tolerant to sodium chloride

20The duration of cultivation is an important biotechnological aspect (Table 1). 300 calli were studied. Prolonged cultivation resulted in the death of most callus. The regenerants obtained after prolonged cultivation were very impaired and slow root formation was observed in the surviving regenerants which were planted again on modified Murashige and Skoog medium for rooting.

21However, most of the regenerants did not take root in the soil after prolonged cultivation. Consequently, The duration of cultivation had an influence on the regeneration capacity of the callus and the obtaining of salt-tolerant plants.

Table 1. Selection of NaCl-tolerant plants

Cultivation duration, number of passages

Number of calli

Regenerants rooted in the soil










22We used two passages to cultivate calli in further studies. In further studies, we also used one passage for the cultivation of calli. The use of one passage is less effective, due to the lower tolerance of many plants to salinity.

23We used sodium chloride in all stages of cultivation.

24We have obtained salt-tolerant plants. Most of the regenerants showed increased salt tolerance. Initial studies showed increased salt tolerance of the regenerants at sodium chloride concentrations of 0.5- 2%. Some regenerants were able to survive at 2% sodium chloride. The original plants did not survive at this concentration. Further studies were for concentrations of 0.5-1% sodium chloride. There was no growth retardation in most of the regenerants studied at 0.5 % sodium chloride. Only one regenerant showed growth like the original plants. The growth of most of the regenerants was more than twice that of the original plants at 1% salinity. One of the most salt-tolerant regenerants showed no growth retardation.

25In perspective, it is of interest to compare the development of the descendants of the regenerants with the original plants.

26The concept developed for obtaining salt-tolerant plants can be used for other plant species. Earlier, salt-tolerant Agrostis stolonifera plants were obtained (Gladkov, Gladkova, 2022). Thus, biotechnological methods can be used to obtain lawn grasses that are resistant to salinity. Lawn grasses will be able to grow at higher levels of salinity in the soil and retain their ornamental qualities.

4. Funding

27Research was carried out within the state assignment of Ministry of Science and Higher Education of the Russian Federation (theme122042600086-7).

5. References

28Al-Khateeb SA, Al-Khateeb AA, Sattar MN, Mohmand AS. 2020. Induced in vitro adaptation for salt tolerance in date palm (Phoenix dactylifera L.) cultivar

29Khalas. Biol Res. Aug 26;53(1):37. doi: 10.1186/s40659-020-00305-3

30Chen Baili, Zheng Hongwei, Geping Luo, Chunbo Chen, Anming Bao, Tie Liu & Xi Chen. 2022. Adaptive estimation of multi-regional soil salinization using extreme gradient boosting with Bayesian TPE optimization. International Journal of Remote Sensing, 43:3, 778-811, 

31doi: 10.1080/01431161.2021.2009589

32Gladkov E.A., Gladkova O.V. 2020. Evaluation of resistance calli cultures of lawn grass for copper, cadmium and use of these results in ecological biotechnology Resources and Technology, 4, 17,107‒113

33Gladkov E.A., Gladkova, O. V. 2021. New directions of biology and biotechnology in urban environmental sciences. Hemijska industrija, 75: 6, 365-368.

34Gladkov E.A., Gladkova, O.V., 2022. Ornamental plants adapted to urban ecosystem pollution: lawn grasses tolerating deicing reagents. Environmental Science and Pollution Research, 29, 22947–22951.

35Gladkov E.A., Dolgikh Y.I., Gladkova O.V. 2014. In vitro selection for tolerance to soil chloride salinization in perennial grasses. Sel’skokhozyaistvennaya Biologiya (Agricultural Biology), 4, 106-111.

36Gladkov, E.A., Tashlieva, I.I. & Gladkova, O.V. 2022. Cell selection for increasing resistance of ornamental plants to copper. Environmental Science and Pollution Research; 25965–25969 .

37Hannachi, S.; Werbrouck, S.; Bahrini, I.; Abdelgadir, A.; Siddiqui, H.A.; Van Labeke, M.C. 2021.Obtaining Salt Stress-Tolerant Eggplant Somaclonal Variants from In Vitro Selection. Plants, 10, 2539.

38Hassani, A., Azapagic A., and Shokri N.. 2020. “Predicting Long-Term Dynamics of Soil Salinity and Sodicity on a Global Scale. Proceedings of the National Academy of Sciences. 117 (52): 33017–33027. doi:

39Kotagiri D, Kolluru V. C. 2017. Effect of Salinity Stress on the Morphology and Physiology of Five Different Coleus Species. Biomed Pharmacol J;10(4).

40Machado RMA, Serralheiro RP. 2017.Soil Salinity: Effect on Vegetable Crop Growth. Management Practices to Prevent and Mitigate Soil Salinization. Horticulturae.; 3(2):30.

41Queirós, F., Fidalgo, F., Santos, I., & Salema, R. 2007. In vitro selection of salt tolerant cell lines in Solanum tuberosum L. Biologia plantarum, 51(4), 728-734. doi: 10.1007/s10535-007-0149-y

42Stavridou, E., Hastings, A., Webster, R.J. and Robson, P.R.H. 2017. The impact of soil salinity on the yield, composition and physiology of the bioenergy grass Miscanthus x giganteus . GCB Bioenergy. 9 92–104

43Van Zelm E, Zhang Y, Testerink C. 2020. Salt Tolerance Mechanisms of Plants.Annual Review of Plant Biology 71:1, 403-433

44Wong V. N., Greene R.S..,Dalal R. C.,, Murphy B. W., 2010. Soil carbon dynamics in saline and sodic soils: A review. Soil Use Manage. 26, 2–11

45Zahra, N.; Raza, Z.A.; Mahmood, S. 2020. Effect of salinity stress on various growth and physiological attributes of two contrasting maize genotypes. Brazilian Arch. Biol. Technol., 63.

Om dit artikel te citeren:

Evgeny Aleksandrovich Gladkov, Yuliya Ivanovna Dolgikh & Olga Victorovna Gladkova, «Biotechnological aspects of obtaining salt-tolerant plants», Bulletin de la Société Royale des Sciences de Liège [En ligne], Volume 91 - Année 2022, Numéro 1, 128 - 133 URL :

Over : Evgeny Aleksandrovich Gladkov

K.А. Timiryazev Institute of Plant Physiology Russian Academy of Sciences, IPP RAS, 35 Botanicheskaya St.,Moscow, 127276 Russia, corresponding author

Over : Yuliya Ivanovna Dolgikh

K.А. Timiryazev Institute of Plant Physiology Russian Academy of Sciences, IPP RAS, 35 Botanicheskaya St.,Moscow, 127276 Russia

Over : Olga Victorovna Gladkova

Independent scientist, Moscow, corresponding author