Abstract
In recent years, the awareness of pomegranate health benefits has grown exponentially; nonetheless the existing propagation methods remain a challenge to supply adequate suitable planting materials needed for commercial production. Micropropagation can lead to mass production of plantlets and callus-mediated in vitro regeneration can open avenues for the use of genetic engineering to improve this crop. The aim of this study was to evaluate appropriate conditions for pomegranate micropropagation, callogenesis and use Simple Sequence Repeat markers to screen for somaclonal variation. Cytokinins (Benzylaminopurine, Kinetin and Thiadiazol-5ylurea) were tested for shoot induction from nodal explants while auxins (1-Naphthaleneacetic acid, Indole-3-butyric acid and Indole-3-acetic acid) were tested for root induction of in vitro regenerated shoots. 1-Naphthaleneacetic acid combined with Benzylaminopurine was assessed for their ability to induce callus from cotyledon and leaf explants. Genetic integrity between mother plant, callus and in vitro regenerated shoots were assessed using eight Simple Sequence Repeat markers. Maximum number of shoots and leaves were obtained on full strength Murashige and Skoog media with 6.9 µM kinetin. The highest number of roots was achieved on half strength Murashige and Skoog media with 4.9 µM Indole-3-butyric acid and the longest root was got on half strength Murashige and Skoog media with 5.3 µM Indole-3-acetic acid. Leaves and cotyledons demonstrated to be potential explants for callus formation at all hormonal combination levels tested. Eight out of 13 amplified alleles were polymorphic. A wider genetic variation was found with similarity coefficient range of 0.46-0.92. More somaclonal variation was in regenerated shoots compared to callus.
Highlights
Pomegranate (P. granatum L.) is a native plant in Iran and the Himalayas of northern India
The maximum mean number of shoots formed per explant was two, achieved on Murashige and Skoog (MS) media supplemented with KN in all treatments (Figure 1)
Further increase in KN concentration above 6.9 μM and BAP concentration above 8.8 μM led to a decrease in number of leaves (Figure 1).There was no significant difference in number of leaves per explant among TDZ treatments (Figure 1) at P ≤ 0.05
Summary
Pomegranate (P. granatum L.) is a native plant in Iran and the Himalayas of northern India It has been cultivated and naturalized over the Mediterranean and the Caucasus region of Asia since ancient times. Cuttings have poor root development and this propagation method is season dependent, labour intensive, time consuming and results in slow establishment (Kaji, Ershadi, & Tohidfar, 2013a; Kaji, Ershadi, & Tohidfar, 2013b; Singh, Patel, & Kadam, 2013). This is a setback for commercial pomegranate production. An in vitro regeneration protocol can be used to produce a suitable pomegranate planting material en mass which will make commercialization achievable
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