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The effect of osmotic stress, lighting spectrum and temperature on growth and gene expression related to anthocyanin biosynthetic pathway in wild strawberry (Fragaria vesca L.) in vitro

Jurgita Vinskienė
Jurgita Vinskienė
, 
Vidmantas Bendokas
Vidmantas Bendokas
, 
Vidmantas Stanys
Vidmantas Stanys
, 
Audrius Sasnauskas
Audrius Sasnauskas
 and 
Rytis Rugienius
Rytis Rugienius
   | Dec 31, 2023
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Article Category: ORIGINAL ARTICLE
Published Online: Dec 31, 2023
Page range: 419 - 431
Received: Oct 26, 2022
Accepted: Oct 24, 2023
DOI: https://doi.org/10.2478/fhort-2023-0030
Keywords
© 2023 Jurgita Vinskienė et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 3.0 License.

Figure 1.

Gene expression in wild strawberry microshoots grown on MS medium with different concentrations of sucrose. MS medium with 3% sucrose was used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHI, chalcone isomerase; CHS, chalcone synthase; MS, Murashige and Skoog; Myb10, MYB domain protein 10; UFGT, UDP glucose: flavonol 3-O-glucosyltransferase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.
Gene expression in wild strawberry microshoots grown on MS medium with different concentrations of sucrose. MS medium with 3% sucrose was used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHI, chalcone isomerase; CHS, chalcone synthase; MS, Murashige and Skoog; Myb10, MYB domain protein 10; UFGT, UDP glucose: flavonol 3-O-glucosyltransferase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.

Figure 2.

Gene expression in wild strawberry microshoots grown on MS medium with different concentrations of PEG. MS medium with 3% sucrose was used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHS, chalcone synthase; DFR, dihydroflavonol reductase; EGL, enhancer of glabra; MS, Murashige and Skoog; PEG, polyethylene glycol; UFGT, UDP glucose: flavonol 3-O-glucosyltransferase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.
Gene expression in wild strawberry microshoots grown on MS medium with different concentrations of PEG. MS medium with 3% sucrose was used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHS, chalcone synthase; DFR, dihydroflavonol reductase; EGL, enhancer of glabra; MS, Murashige and Skoog; PEG, polyethylene glycol; UFGT, UDP glucose: flavonol 3-O-glucosyltransferase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.

Figure 3.

Wild strawberry microshoots grown under different light spectra: fluorescent light (F), LED lighting system: blue light (B), red light (R), blue + red lights (BR), and blue + red + UV lights (BR UV). LED, light-emitting diode.
Wild strawberry microshoots grown under different light spectra: fluorescent light (F), LED lighting system: blue light (B), red light (R), blue + red lights (BR), and blue + red + UV lights (BR UV). LED, light-emitting diode.

Figure 4.

Gene expression in wild strawberry microshoots grown at 15°C for 1-4 weeks. Microshoots grown at 22°C were used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHI, chalcone isomerase; CHS, chalcone synthase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.
Gene expression in wild strawberry microshoots grown at 15°C for 1-4 weeks. Microshoots grown at 22°C were used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHI, chalcone isomerase; CHS, chalcone synthase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.

Figure 5.

Gene expression in wild strawberry microshoots grown at 30°C for 1-4weeks. Microshoots grown at 22°C were used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHI, chalcone isomerase; CHS, chalcone synthase; DFR, dihydroflavonol reductase; F3H, flavanone 3-hydroxylase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.
Gene expression in wild strawberry microshoots grown at 30°C for 1-4weeks. Microshoots grown at 22°C were used as a control. Bars denote the standard error of the mean. Asterisks indicate significant differences compared to the control assessed by Fisher’s test (*p < 0.05, **p < 0.01), n = 3. CHI, chalcone isomerase; CHS, chalcone synthase; DFR, dihydroflavonol reductase; F3H, flavanone 3-hydroxylase; WD40, a tryptophan-aspartic acid (W-D) dipeptide.

Supplementary Figure S1.

Gene expression in wild strawberry microshoots grown on MS medium under different light spectra. Microshoots grown under fluorescent lamps were used as a control. Bars denote the standard error of the mean. CHI, chalcone isomerase; CHS, chalcone synthase; DFR, dihydroflavonol reductase; F3H, flavanone 3-hydroxylase; MS, Murashige and Skoog; Myb10, MYB domain protein 10; WD40, a tryptophan-aspartic acid (W-D) dipeptide.
Gene expression in wild strawberry microshoots grown on MS medium under different light spectra. Microshoots grown under fluorescent lamps were used as a control. Bars denote the standard error of the mean. CHI, chalcone isomerase; CHS, chalcone synthase; DFR, dihydroflavonol reductase; F3H, flavanone 3-hydroxylase; MS, Murashige and Skoog; Myb10, MYB domain protein 10; WD40, a tryptophan-aspartic acid (W-D) dipeptide.

Primers used for anthocyanin biosynthesis gene expression analysis

Gene Accession number Primer sequence Annealing temperature, °C Product size, bp
Myb10 EU155163 F: 5’-CGGAAGATTGCCAGGAAGAAC-3’ 62.4 165
R: 5’-ATGAAGGTTCGTGGTCGAGG-3’ 63.1
WD40-TTG1 XM_004307863 F: 5’-AGCAGGACTTGAGGTACATGG-3’ 63 129
R: 5’-ACGCAATCGCATTCACACTC-3’ 62.8
EGL XM_004308329 F: 5’-GCCTTCGATAAACAAGCGGAAG-3’ 63.1 131
R: 5’-TCTCTATCAGAACCTCCTGCTC-3’ 61.9
UFGT XM_004307828 F: 5’-GCGCATGGTTCAGTTGGAG-3’ 62.8 151
R: 5’-GACCAATCTTCCACACATCCTC-3’ 62
DFR KC894052 F: 5’-GTCTCATTACCGGACTTTCGC-3’ 62.1 131
R: 5’-CTCTGCTTTCGGATGCTCG-3’ 61.9
F3H AB201760 F: 5’-CACAGCAGGTTGTCCATAGC-3’ 62.2 114
R: 5’-AGTGTAAGTCATCGGCTCCTC-3’ 62.6
CHI XM_004307403 5’-AAAGATCAGACCTTCCCACCC-3’ 63 119
R: 5’-TCAATCACCGCATTCCCAAC-3’ 62.4
CHS AB250913 F: 5’-ACTTTTCTGGATTGCACACCC-3’ 62.6 189
R: 5’-GTCTTGTGCCCATTAGCTGC-3’ 62.8
β-actin* XM_004306544 F: 5’-TCAACTATGTTCCCTGGTATTGC-3’ 62 175
R: 5’-CTCCCTTGGAAATCCACATCTG-3’ 62.1

Average weight of wild strawberry microshoots, affected by osmotic components in MS medium, light and growth temperature.

Trait Parameter Average weight of microshoot (g)
Osmotic components Sucrose 15 g ∙ L-1 0.27 ± 0.03
Sucrose 30 g ∙ L-1 0.30 ± 0.03
Sucrose 60 g ∙ L-1 0.13 ± 0.04*
Sucrose 90 g ∙ L-1 0.10 ± 0.04*
Sucrose 30 g ∙ L-1 + PEG 50 g ∙ L-1 0.12 ± 0.10*
Sucrose 30 g ∙ L-1 + PEG 100 g ∙ L-1 0.11 ± 0.11*
Sucrose 30 g ∙ L-1 + PEG 120 g ∙ L-1 0.44 ± 0.60*
Light Fluorescent 0.28 ± 0.01
Blue 0.31 ± 0.02
Red 0.27 ± 0.02
BR 0.34 ± 0.03
BRUV 0.24 ± 0.01
Temperature 15°C 0.13 ± 0.03*
22°C 0.34 ± 0.06
30°C 0.30 ± 0.15
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