Heavy metals are among the priority pollutants (Sakson et al. 2018). Cu is one of the most enriched elements in soil. Especially high concentrations of copper in the soil are observed in areas located near copper smelter (Radojevic et al. 2017).

Copper is among the contaminant of agricultural soil (Sacristan and Carbo 2016; Napoli et al. 2019; Htwe et al. 2022). Copper is one of the urban soil pollutants. For example, in Copenhagen soils, total Cu concentrations were significantly higher compared to the reference agricultural soil (Li et al. 2014). Copper in urban soils is being accumulated mainly due to anthropogenic activities under rapid urbanisation (Li et al. 2018). Copper limits the spread of urban plants (Gladkov and Gladkova 2021b).

Cell selection has been used to increase plant resistance to copper. Technologies have been developed to obtain copper-resistant Agrostis stolonifera and Chrysanthemum carinatum plants. The detailed description of these technologies is given in Gladkov et al. (2021a). Can the developed technologies be used for other plant species? These technologies have been applied to other plant species.

The objects of our study were ornamental plants: Festuca rubra L., Brachycome iberidifolia Benth. and Linum perenne L.

  • Festuca rubra (red fescue) is used as a lawn grass in ornamental gardening. Linum perenne is a perennial plant with blue flowers.

  • Linum perenne is very effective in joint plantings with other flowering plants.

  • Brachycome iberidifolia features attractive foliage and blooms blue, purple, lilac, pink and white flowers (Fig. 1). Brachycome iberidifolia can be planted in rock gardens and flower beds.

Fig. 1
figure 1

Brachycome iberidifolia

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Different modified mediums were used for each plant species. However, Murashige-Skoog modified medium was the same for two lawn grass species (Festuca rubra and Agrostis stolonifera) (Gladkov and Gladkova 2021a). Sucrose concentration was 30 g/L, casein hydrolysate 500 mg/L for Festuca rubra and Agrostis stolonifera in modified Murashige-Skoog medium. Seeds were used to obtain lawn grass callus.

Callus of Agrostis stolonifera was obtained on Murashige-Skoog modified medium with 3 mg/L 2,4-dichlorophenoxyacetic acid. Callus of Festuca rubra was obtained on Murashige-Skoog modified medium with 3 mg/L 2,4-dichlorophenoxyacetic acid.

Calli of Agrostis stolonifera and Festuca rubra were cultured on Murashige-Skoog medium supplement with 1 mg/L 2,4-dichlorophenoxyacetic acid.

The composition of the mediums for Chrysanthemum carinatum, Brachycome iberidifolia and Linum perenne differed.

Gamborg modified medium was modified to obtain callus of Brachycome iberidifolia and Linum perenne. Callus of Linum perenne was obtained on Gamborg medium with 6 mg/L 2,4-dichlorophenoxyacetic acid. The regeneration of Linum perenne plants was on Gamborg modified medium with 1 mg/L 6-benzylaminopurine and 0.1 mg/L α-naphthaleneacetic acid. Calli (Fig. 2) were cultivated on Gamborg modified medium until shoots were formed. The duration of cultivation was 2–3 passages.

Fig. 2
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Calli of Linum perenne

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Rooting of Linum perenne plants was on Murashige-Skoog modified medium with the addition of naphthaleneacetic acid. Root development on liquid medium was faster than on agar medium. Seedlings were transplanted to liquid medium and mounted on filter paper bridges.

The regeneration of chrysanthemum plants was on Murashige-Skoog modified medium with 0.5 mg/L 6-benzylaminopurine.

To prevent the death of callus Brachycome iberidifolia, the growing cycle was no more than 21–26 days. The regeneration of Brachycome iberidifolia plants was on Murashige-Skoog modified medium with 2 mg/L 6-benzylaminopurine and 0.1 mg/L α-naphthaleneacetic acid.

Calli of Agrostis stolonifera and Chrysanthemum carinatum had different resistance to copper (Gladkov et al. 2021a). Therefore, we used different cell selection schemes.

Copper negatively affects morphogenic potential (Yemets et al. 2013; Ahmad et al. 2015).

However, small concentrations of copper can increase the regenerative capacity of calli (Kintzios et al. 2000; Malik et al. 2021). High copper concentrations always lead to a decrease in morphogenic potential (Gladkov et al. 2019, 2021a, b). Therefore, the use of copper at the stage of regeneration is impractical in the case of very high sensitivity of calli.

Calli of Festuca rubra and Agrostis stolonifera had comparable sensitivity to copper (Gladkov et al. 2021a, b). The selective copper concentration for calli of Festuca rubra and Agrostis stolonifera was 150 mg/L. Therefore, a cell selection scheme for Agrostis stolonifera was used for Festuca rubra (Gladkov et al. 2019, 2021a, b). Calli of Agrostis stolonifera and Festuca rubra showed moderate sensitivity to copper. Therefore, copper was used at all the stages of cultivation. We recommend using the developed scheme of cell selection with an average degree of resistance of calli to copper (Fig. 3).

Fig. 3
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Cell selection scheme for obtaining lawn grasses tolerant to copper

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The sensitivity of Brachycome iberidifolia calli was comparable to that of Chrysanthemum carinatum (Fig. 4). Calli of Brachycome iberidifolia was dead at the selective copper concentration for lawn grass.

Fig. 4
figure 4

Effect of copper concentration (mg/L) on callus survival in the second passage (%)

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The selective copper concentration for calli of Chrysanthemum carinatum, Brachycome iberidifolia and Linum perenne was 20 mg/L. Calli of Chrysanthemum carinatum, Brachycome iberidifolia and Linum perenne were less resistant to copper than calli of lawn grass.

A cell selection scheme for Chrysanthemum carinatum was used for Brachycome iberidifolia and Linum perenne (Fig. 5). The duration of cultivation depended on the species.

Fig. 5
figure 5

Cell selection scheme for obtaining Brachycome iberidifolia and Linum perenne tolerant to copper

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Copper-resistant Linum perenne plants were particularly difficult to obtain. Various modified Gamborg and Murashige-Skoog media were used to obtain resistant plants (Fig. 6).

Fig. 6
figure 6

Cell selection scheme for obtaining Linum perenne tolerant to copper

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The concept of obtaining copper-resistant plants has been developed. This concept consists of two methods. The first method is applicable when calli is highly sensitive to copper. Copper is not used in the stages of regeneration and rooting. The second method is applicable when calli are moderately sensitive to copper. Copper is used at all the stages (cultivation, regeneration and rooting).

We obtained copper-resistant plants Festuca rubra, Brachycome iberidifolia and Linum perenne. Therefore, developed technologies for obtaining copper-resistant plants Agrostis stolonifera and Chrysanthemum carinatum can be applied to other plant species.