Abstract
BackgroundJuniperus procera Hoechst. ex Endl. is a medicinal tree in Saudi Arabia, primarily in the Enemas region, but it is locally threatened due to die-back disease and difficulties regarding seed reproduction (seed dormancy and underdeveloped embryonic anatomy, and germination rate < 40%). Hence, the alternative methods for reproduction of Juniperus procera are really needed for conservation and getting mass propagation for pharmaceutical uses.ResultsIn this manuscript, we articulated the successful in vitro shoot multiplication and callus induction of J. procera by using young seedling as explants and detected an important antibacterial and antitumor product. Explants were grown on different types of media with the supplement of different combinations of Plant Growth Regulators (PGRs) at different concentrations. The best media for shoot multiplication was Woody Plant Media (WPM) supplemented with PGRs (0.5 μM of IAA and 0.5 μM BAP or 0.5 μM IBA and 0.5 μM BAP). Whereas for callus induction and formation Woody Plant Media (WPM) with the addition of PGRs (0.5 μM 2,4-D and 0.5 μM BAP) was better than the Chu Basal Salt Mixture (N6), Gamborg’s B-5 Basal Medium (B5), and Murashige and Skoog media. The possibility of multiplication of J. procera in vitro creates significant advantages to overcome the difficulties of seeds dormancy for the reproduction of plants, conservation of trees, and getting mass propagation material for pharmaceutical studies. The shoot and callus extract of J. procera was detected using gas chromatography-mass spectrometry analysis and revealed more than 20 compounds related to secondary metabolites, which contained antibacterial and antitumor agents, such as ferruginol, Retinol, and Quinolone as well as confirmed by Direct Analysis in Real Time, Time of Flight Mass Spectrometry (DART-ToF-MS). Podophyllotoxin (PTOX) was detected in callus material by HPLC with sigma standard and confirmed by DART-ToF-MS and UV spectra.ConclusionWe successfully conducted in vitro shoot multiplication and callus induction from J. procera seedlings using WPM and a different combination of PGRs and, detected an important antibacterial and antitumor product such as ferruginol and podophyllotoxin. According to our findings, J. procera has become a new natural source of novel bioactive compounds.
Highlights
The genus Juniperus L. is the second most prevalent group of conifer s on Earth [1]
Juniperus procera is indigenous to the mountains of eastern Africa from east Sudan to Zimbabwe, and southwest of the Arabian Peninsula, and it is widely spread throughout southern part of Saudi Arabia [3]
Media and plant growth regulators for shoot multiplication Four types of media were tested to evaluate the effects of Plant Growth Regulators (PGRs) and media compositions on shoot multiplication: (1) Woody Plant Medium (WPM) [35], (2) Murashige and Skoog (MS) medium [36], (3) et al [37] (Gamborg)’s B-5 Basal Medium (B5 medium) [37], and (4) Chu Basal Salt Mixture (N6 medium) [38] (Fig. 1)
Summary
The genus Juniperus L. is the second most prevalent group of conifer s on Earth [1]. Juniperus procera is coniferous evergreen tree or shrub of the Cupressaceae family. On the other hand, seed dormancy is a major hurdle for artificial regeneration of J. procera for economic and ecological importance [16] In this context, it was reported that, some species of Juniperus have produced very small number of viable seeds or anatomically undeveloped [17]. In vitro propagation of juniper species should be exaggerated because, the micropropagation maybe the only alternative methods of the reproduction of this group of plants [1]. It should be prioritized for the possibility of conservation [33], and mass propagation for pharmaceutical uses. The alternative methods for reproduction of Juniperus procera are really needed for conservation and getting mass propagation for pharmaceutical uses
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