Synergistic effect of benzylaminopurine and meta-Topolin combination for micropropagation of gerbera ‘Pink Melody’

Gerbera (Gerbera jamesonii Bolus ex Hook. F.; Asteraceae), is one of the most economically important ornamental plants due to its aesthetic value. In the present study, we established a micropropagation method for the large-scale production of quality planting material of gerbera ‘Pink Melody’. Eighty-six percent of the capitulum explants produced adventitious shoots (15.44 ± 0.34 shoots per capitulum) on Murashige and Skoogs (MS) medium containing 2 mg L⁻¹ 6-benzylaminopurine (BAP) after six weeks of incubation. The highest shoot multiplication rate (17 shoots per explant) was obtained on MS medium supplemented with BAP and meta-Topolin (each at 2 mg L⁻¹) after eight weeks. The micro-shoots were successfully rooted (91.35%) on half-strength MS medium containing 2 mg L⁻¹ indole-3-butyric acid (IBA) within four weeks. The micropropagated plantlets were acclimatized with a 97.5% survival rate and produced flowers with no visible morphological aberrations.

Index terms:
Acclimatization; Asteraceae; floriculture; tissue culture

Authorship

Aaqib Shaheen

Henan University, College of Agriculture, School of Life Sciences, Kaifeng, Henan, ChinaHenan UniversityChinaKaifeng, Henan, ChinaHenan University, College of Agriculture, School of Life Sciences, Kaifeng, Henan, China

http://orcid.org/0000-0002-8295-7054

Yaser Hassan Dewir ^**Corresponding author: ydewir@ksu.edu.sa

King Saud University, College of Food and Agriculture Sciences, Plant Production Department, Riyadh, Saudi ArabiaKing Saud UniversitySaudi ArabiaRiyadh, Saudi ArabiaKing Saud University, College of Food and Agriculture Sciences, Plant Production Department, Riyadh, Saudi Arabia

http://orcid.org/0000-0002-5174-8137

Mafatlal Kher

GSFC University, School of Science, Vadodara, Gujarat, IndiaGSFC UniversityIndiaVadodara, Gujarat, IndiaGSFC University, School of Science, Vadodara, Gujarat, India

http://orcid.org/0000-0001-5600-0040

Majid Khan

Chinese Academy of Agricultural Sciences, Institute of Cotton Research, Anyang, Henan, ChinaChinese Academy of Agricultural SciencesChinaAnyang, Henan, ChinaChinese Academy of Agricultural Sciences, Institute of Cotton Research, Anyang, Henan, China

http://orcid.org/0000-0001-9014-6107

Antar Nasr El-Banna

Julius Kühn-Institute/JKI, Federal Research Centre for Cultivated Plants, Institute for Epidemiology and Pathogen Diagnostics, Braunschweig, GermanyJulius Kühn-Institute/JKIGermanyBraunschweig, GermanyJulius Kühn-Institute/JKI, Federal Research Centre for Cultivated Plants, Institute for Epidemiology and Pathogen Diagnostics, Braunschweig, Germany

http://orcid.org/0000-0002-7778-3109

Ahmed Alaizari

http://orcid.org/0000-0003-0761-7321

^*Corresponding author: ydewir@ksu.edu.sa

SCIMAGO INSTITUTIONS RANKINGS

GSFC University, School of Science, Vadodara, Gujarat, IndiaGSFC UniversityIndiaVadodara, Gujarat, IndiaGSFC University, School of Science, Vadodara, Gujarat, India

Figures | Tables

Figures (2)
Tables (4)

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Figure 1:
Micropropagation of Gerbera jamesonii ‘Pink Melody.’ A) Capitulum explant cultured on Murashige-Skoog (1962MURASHIGE, T.; SKOOG, F. A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologae Plantarum, 15(3):473-497, 1962. ) medium (MS) with 2 mg L⁻¹ 6-benzylaminopurine (BAP). B) Adventitious shoot regeneration and shoot development (inset) on the same initiation medium. C) Axillary shoot multiplication on MS supplemented with 2 mg L⁻¹ BAP and 2 mg L⁻¹ meta-Topolin after eight weeks, and D) In vitro rooting on half-strength MS medium with 2 mg L⁻¹ indole-3-butyric acid after four weeks.

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Figure 2:
Acclimatization and growth of Gerbera jamesonii ‘Pink Melody’ A) Acclimatization in the growth chamber for three weeks, B) Plantlet with well-developed root system after three weeks in the growth chamber, C and D) Further growth of micropropagated plantlets after five and eight weeks in the greenhouse, respectively, and E) Flowering plant grown in peatmoss and soil (1:1, v/v) after 15 weeks in the greenhouse.

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Table 1:
Percentages shoot regeneration and number of shoots of Gerbera jamesonii ‘Pink Melody’ on MS medium with 2 mg L^-1 6-benzylaminopurine after 6 weeks in culture.

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Table 2:
Effect of different combinations of plant growth regulators on the in vitro shoot multiplication of Gerbera jamesonii ‘Pink Melody’ after an 8-week subculture period (every four week). The medium without cytokinin was served as the control.

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Table 3:
Effect of indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) concentrations and salt strength of Murashige and Skoog’s (MS) medium on in vitro rooting of Gerbera jamesonii ‘Pink Melody’ after a 4-weeks.

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Table 4:
Effect of different growing media on acclimatization and flowering of Gerbera jamesonii ‘Pink Melody’ plantlets after 15 weeks in greenhouse.

Figure 1: Micropropagation of Gerbera jamesonii ‘Pink Melody.’ A) Capitulum explant cultured on Murashige-Skoog (1962MURASHIGE, T.; SKOOG, F. A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiologae Plantarum, 15(3):473-497, 1962. ) medium (MS) with 2 mg L⁻¹ 6-benzylaminopurine (BAP). B) Adventitious shoot regeneration and shoot development (inset) on the same initiation medium. C) Axillary shoot multiplication on MS supplemented with 2 mg L⁻¹ BAP and 2 mg L⁻¹ meta-Topolin after eight weeks, and D) In vitro rooting on half-strength MS medium with 2 mg L⁻¹ indole-3-butyric acid after four weeks.

Figure 2: Acclimatization and growth of Gerbera jamesonii ‘Pink Melody’ A) Acclimatization in the growth chamber for three weeks, B) Plantlet with well-developed root system after three weeks in the growth chamber, C and D) Further growth of micropropagated plantlets after five and eight weeks in the greenhouse, respectively, and E) Flowering plant grown in peatmoss and soil (1:1, v/v) after 15 weeks in the greenhouse.

Table 1: Percentages shoot regeneration and number of shoots of Gerbera jamesonii ‘Pink Melody’ on MS medium with 2 mg L^-1 6-benzylaminopurine after 6 weeks in culture.

Explant type	Survival (%)	Explants forming adventitious shoots (%)	Adventitious shoot (number/explant)
Capitulum	65 ± 3.90	86.42 ± 3.60	15.44 ± 0.34

Table 2: Effect of different combinations of plant growth regulators on the in vitro shoot multiplication of Gerbera jamesonii ‘Pink Melody’ after an 8-week subculture period (every four week). The medium without cytokinin was served as the control.

Growth regulators	Shoot number/explant	Length of the main shoot (cm)
Control	1.0±0.00 g ^z	1.55±0.59 f ^z
1 mg L^-1 BAP +2 mg L^-1 meta-Topolin	10±0.17 b	2.10±0.14 ef
2 mg L^-1 BAP + 2 mg L^-1 meta-Topolin	17±0.45 a	4.38±0.09 a
3 mg L^-1 BAP +2 mg L^-1 meta-Topolin	11±0.33 b	2.26±0.10 de
4 mg L^-1 BAP +2 mg L^-1 meta-Topolin	9±0.24 c	2.84±0.12 bcd
5 mg L^-1 BAP +2 mg L^-1 meta-Topolin	8±0.33 c	3.39±0.23 b
6 mg L^-1 BAP +2 mg L^-1 meta-Topolin	7±0.24 d	2.99±0.04 bc
2 mg L^-1 BAP +2 mg L^-1 2,4-D	7.4±0.24 cd	2.75±0.11 cd
2 mg L^-1 BAP + 2 mg L^-1 TDZ	5.2±0.28 e	2.04±0.05 ef
1 mg L^-1 GA₃	5.1±0.20 e	2.83±0.07 bcd
2 mg L^-1 GA₃	5.3±0.29 e	4.29±0.18 a
3 mg L^-1 GA₃	4±0.17 f	3.04±0.21 bc

^zValues followed by the same letter in the same column are not significantly different at P ? 0.05 level, according to Tukey’s multiple range test. 2,4-D = 2,4-dichlorophenoxyacetic acid, BAP= 6-benzylaminopurine, GA₃ = gibberillic acid, TDZ= thidiazuron.

Table 3: Effect of indole-3-acetic acid (IAA) and indole-3-butyric acid (IBA) concentrations and salt strength of Murashige and Skoog’s (MS) medium on in vitro rooting of Gerbera jamesonii ‘Pink Melody’ after a 4-weeks.

Auxins (mg L^-1)	Rooting (%)	Roots (no./per plantlet)	Length of the main root (cm)	Leaves (no./plantlet)
MS without auxins	16.05±3.27 e^z	2.34±0.17 f^z	2.44±0.15 cd^z	3±0.26 de^z
½ MS without auxins	38.14±1.85 d	2.66±0.17 f	3.26±0.20 bc	4±0.22 cd
½ MS + IAA 1	50.61±1.95 c	6.44±0.18 d	2.97±0.25 c	3±0.26 e
2	56.79±2.23 c	9.66±0.17 b	4.07±0.14 b	6±0.20 b
3	46.91±1.63 c	6.22±0.22 d	3.17±0.35 bc	4±0.18 c
½ MS + IBA 1	53.08±3.60 c	4.88±0.31 e	2.47±0.21 cd	5±0.17 b
2	91.35±2.47 a	11.4±0.24 a	5.63±0.18 a	8±0.11 a
3	80.85±2.62 b	7.88±0.35 c	2.01±0.15 d	3±0.18 de

^zValues followed by the same letter in the same column are not significantly different at P ? 0.05 level, according to Tukey’s multiple range test.

Table 4: Effect of different growing media on acclimatization and flowering of Gerbera jamesonii ‘Pink Melody’ plantlets after 15 weeks in greenhouse.

Potting mixture (by equal volume)	Survival (%)	Days to flower emergence	Flowers (no./plant)	Flower diameter (cm)	Peduncle length (cm)
Soil	46.9±1.63 ef^z	74.667±0.75 cde^z	2±0.39 bcd^z	2.84±0.16 e^z	42.0±1.19 de^z
Soil + silt	56.8±2.90 cde	76.667±0.29 abc	3±0.15 ab	3.9±0.20 cd	38.4±0.44 f
Soil + sand	59.3±1.85 bcd	75.9±0.54 bcd	2±0.18 bcd	3.4±0.18 de	43.1±0.39 d
Silt + sand	50.6±2.69 def	76.8±0.43 abc	2±0.26 bcd	4.3±0.08 bc	39.0±0.47 f
Soil + silt + sand	61.7±1.95 bcd	77.9±0.45 ab	1±0.18 d	4.4±0.09 bc	42.0±0.33 de
Soil + sand + silt + coco coir	69.1±2.47 b	76.4±0.34 bc	3±0.29 abc	4.6±0.07 abc	46.7±0.41 b
Soil + silt + sand + Manure compost	62.9±1.85 bc	78.0±0.60 ab	2±0.36 bcd	3.9±0.23 cd	40.4±0.50 ef
Soil + silt + sand + coco coir + manure compost	56.7±2.23 cde	79.1±0.73 a	2±0.24 bcd	4.2±0.18 bcd	45.9±0.26 bc
Peat moss	96.2±1.85 a	68.0±0.63 f	4±0.32 a	5.3±0.12 a	55.2±0.43 a
Peat moss + soil	97.5±1.63 a	66.0±0.26 f	4±0.35 a	4.9±0.26 ab	56.2±0.57 a
Peat moss + soil + silt + sand	53.0±3.09 cde	73.4±0.67 de	2±0.26 bcd	4.0±0.13 cd	43.9±0.48 cd
Peat moss + silt + sand + manure compost	40.7±2.62 f	72.6±0.24 e	2±0.17 d	3.9±0.21 cd	42.8±0.52 de
Peat moss + soil + silt + sand + coco coir + manure compost	55.5±3.21 cde	79.1±0.48 a	2±0.28 cd	3.4±0.10 de	44.0±0.33 cd

^zValues followed by the same letter in the same column are not significantly different at P ? 0.05 level, according to Tukey’s range test.

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Synergistic effect of benzylaminopurine and meta-Topolin combination for micropropagation of gerbera ‘Pink Melody’

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[1] ^*Corresponding author: ydewir@ksu.edu.sa