Consideration of adenine hemi-sulfate effect on increasing the direct regeneration rate of carnation cultivars’ leaf explants

Document Type : Research Paper


1 Ph.D. Candidate, Department of Horticultural Science, Agriculture and Natural Resources, University of Hormozgan, Bandar Abbas, Iran

2 Associate Professor, Department of Biotechnology Engineering, Faculty of Agriculture, Shahid Bahonar University of Kerman, Kerman, Iran

3 Department of Genetic Engineering, Agricultural Biotechnology Research Institute of Iran (ABRII), Karaj, Agricultural Research, Education and Extension Organization (AREEO).

4 rnamental Plants Research Center (OPRC), Horticultural Sciences Research Institute (HSRI), Agricultural Research, Education and Extension Organization (AREEO), Mahallat, Iran.


Purpose The genetic improvement of carnation (Dianthus Caryophyllus), as one of the most important cut flowers in the world, is of utmost significance. For this purpose, the use of in vitro breeding techniques is very important. One of the most crucial in vitro breeding needs of carnation is accessibility to a regeneration method with maximum productivity and minimum genetic modification probability. The present study aimed to determine an optimal and fast method for the direct regeneration of carnation cultivars by employing various regulators and adenine hemi-sulfate for the first time in these cultivars. Materials and Methods For the determination of the optimal level of the culture medium compositions toward the direct regeneration of carnation cultivars (Skimo, Tibor, and Labret), an experiment with a completely randomized design was designed and implemented in an MS culture medium, in different concentrations of growth regulators, including four levels of TDZ (0.5, 1, 2, and 3mg/L), two levels of IAA (0.5 and 1mg/L), and four levels of adenine hemi-sulfate (0. 20, 40, and 80mg/l), were used. Results The model used for this experiment was significant at the level of 1%, where R2=96% and CV=22, indicating the acceptable accuracy of the analysis. With respect to the results, it was observed that, in the leaf explants of different carnation cultivars, the utilization of the MS culture medium in light conditions with TDZ 2 mg/L+ IAA 0.5 mg/L occurred the direct regeneration of the leaf explants. It is worth mentioning that the addition of 40mg/L of adenine hemi-sulfate to the culture medium increased the regeneration mean from 66% to 94% among different cultivars. In addition, the mean shoot number per explant increased from 12 to 36 shoots. Conclusion Generally, the results of the experiments showed that the mixing of the MS base medium, leaf explants in light conditions, and the TDZ 2 mg/L+ IAA 0.5 mg/L+As 40 mg/L regulator composition significantly led to the leaf regeneration of carnation cultivars. Briefly, the results of this study proved that the use of adenine hemi-sulfate significantly impacted the upsurge of the direct regeneration rate of carnation cultivars.


Agnieszka, W, Edyta S, Eleonora G (2016) Morphological and biochemical responses to gibberellic acid in Magnolia × ‘Spectrum’ In vitro. Acta Biol Crac Ser Bot58 (1), 103-111.
Arif M, Rauf S, Ud Din A et al. (2010) High Frequency Plant Regeneration from Leaf Derived Callus of High 9-tetrahydrocannabinol Yielding Cannabis Sativa L. Planta Med 76(14), 1629–1633.
Azadi P, Kermani MJ, Samiei L (2018) Somatic Embryogenesis in Rosa Hybrida. In: Step Wise Protocols for Somatic Embryogenesis of Important Woody Plants, Forestry Sciences 85, (1 st. edn). Springer International publishing, part of Springer Nature. Cham. pp. 161-170.
Datta SK (2014) Induced mutagenesis: basic knowledge for technological success. In: Mutagenesis, exploring novel genes and pathways (1 st edn). Wageningen Academic Publisher, Wageningen, Netherland. pp. 97–140.
Dyaberi A, Dhananjaya MV, Kumar R et al. (2015) Floral Biology and Seed Setting in Standard Carnation (Dianthus caryophyllus). Indian J Agric Sci 8(59), 1175–1180.
Gatica AAM, Munoz VJ, Ramírez FP et al. (2010) In vitro plant regeneration system for common bean (Phaseolus vulgaris): effect of N 6 -benzylaminopurine and adenine sulphate. Electron J Biotechnol 13(1), 0717-3458.
Hegazy A, Nasr MI, Ibrahim IA et al. (2009). Micropropagation of Date Palm cv. malakaby through somatic embryogenesis (Effect of adenine hemisulfate, glutamine and glutathione). J Agric Sci Mansoura Uni 34 (3), 1613–1627.
Iantcheva A (2016). Somatic Embryogenesis and Genetic Transformation of Carnation (Dianthus caryophyllus L.). In Somatic Embryogenesis in Ornamentals and Its Applications. Springer India, India. pp, 107–120.
Jaberi M, Azadi P, Gharehyazi B et al. (2017) Silver Nitrate and Adenine Sulphate Induced High Regeneration Frequency in the Recalcitrant Plant Cosmos Bipinnatus Using cotyledon explants. J Hortic Sci Biotechnol 1–5.
Jain A, Kantia A, Kothari SL (2001) De Novo Differentiation of Shoot Buds from Leaf-callus of Dianthus Caryophyllus L. and Control of Hyperhydricity. S Sci Hortic 8(74), 319–326.
Kantia A, Kothari SL (2002) High Efficiency Adventitious Shoot Bud Formation and Plant Regeneration from Leaf Explants of Dianthus Chinensis L. Sci Hortic 96(1–4), 205–212.
Kanwar JK, Kumar S (2009) Influence of growth regulators and explants on shoot regeneration in carnation. Hortic Sci 36, 140–146.
Karami O, Deljou A, Kordestani, GK (2008) Secondary Somatic Embryogenesis of Carnation (Dianthus caryophyllus L.). Plant Cell Tissue Organ Cult 92(3), 273–280.
Kumari P, Baskaran J, Van S (2017) In vitro regeneration of Begonia homonyma — A threatened plant. S A fr J Bot 109, 174-177.
Lukatkin AS, Mokshin EV, Teixeira da Silva JA (2017) Use of Alternative Plant Growth Regulators and Carbon Sources to Manipulate Dianthus caryophyllus L. Shoot Induction in Vitro. Rend Lincei 28 (3), 583–588.
Mujib A (2015) Somatic Embryogenesis in Ornamentals and its Applications. Somatic Embryogenesis in Ornamentals and Its Applications, 1–267.
Naaz A, Shahzad A, Anis, M. (2014). Effect of Adenine Sulphate Interaction on Growth and Development of Shoot Regeneration and Inhibition of Shoot Tip Necrosis Under in Vitro Condition in Adult Syzygium cumini L. A Multipurpose Tree. Appl Biochem Biotechnon 173 (1), 90–102.
Phillips GC, Garda M (2019) Plant Tissue Culture Media and Practices: an overview. In: In Vitro Cellular and Developmental Biology - Plant. Springer New York LLC, USA. pp. 242–257.
Prasad HM, Mythili JB, Anand L et al. (2016) Optimization of Regeneration Protocol and Agrobacterium Mediated Transformation in Carnation (Dianthus caryophyllus L.). Int J Hortic. Sci (4)2, 120–127.
Shen X, Kane ME, Chen J (2008) Effects of Genotype, Explant Source, and Plant Growth Regulators on Indirect Shoot Organogenesis in Dieffenbachia Cultivars. In Vitro Cell Dev Biol - Plant 44(4), 282–288.
Siddique I, Yadav, V (2021) Cytokinin Influence on Micropropagation System of Dianthus caryophyllus L. In Propagation and Genetic Manipulation of Plants. Springer Singapore pp, 33–41.
Silue O, Kouassi M. K, Koffi EK et al. (2017) Effect of Adenine Sulphate, Casein Hydrolysate and Spermidine on in Vitro Shoot Multiplication of Two Banana Varieties. Afr J Biotechnol 46, 2152–2159.
Thakur K, Kanwar, K (2018) In Vitro Plant Regeneration by Organogenesis from Leaf Callus of Carnation, Dianthus caryophyllus L. cv. Proceedings of the National Academy of Sciences India Section B - Biological Sciences (88)3, 1147–1155.
Varshney A, Anis M, Aref IM (2013). Potential Role of Cytokinin-auxin Synergism, Antioxidant Enzymes Activities and Appraisal of Genetic Stability in Dianthus caryophyllus L.-An Important Cut Flower Crop. In Vitro Cell Dev Biol Plant (49)2, 166–174.
Zia M, Yaqoob K, Mannan, A. et al. (2020). Regeneration Response of Carnation Cultivars in Response of Silver Nanoparticles under in Vitro Conditions. Vegetos 1(33), 11–20.