Zohreh Chenarani; Farhad Shokouhifar; Mojtaba Mamarabadi
Abstract
Introduction: Iran is the third producer of Melon (Cucumismelo L.) after China and Turkey in the World.Melon is one of the main crops in Khorasan province in the term of cultivation area. The cultivar of Khatooni, among many varieties of melon, can be considered as a breeding cultivar because of its ...
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Introduction: Iran is the third producer of Melon (Cucumismelo L.) after China and Turkey in the World.Melon is one of the main crops in Khorasan province in the term of cultivation area. The cultivar of Khatooni, among many varieties of melon, can be considered as a breeding cultivar because of its great qualitative traits including sweetness and flavor. This cultivar is also economically important and has many attractions for export. However, this cultivar is susceptible to some of plant diseases caused by different bacterial, viral and fungal pathogens which among them vascular wilt caused by Fusariumoxysporum f. sp. melonis (Leach &Currenu) Snyder & Hansen (FOM) is one of the most destructive soil-borne diseases in melon in Iran particularly in temperate and cold regions. The control of melon diseases has depended increasingly on the extensive use of toxic chemicals (pesticides). Many of these chemicals have been shown to be toxic to non-target microorganisms and animals and may be toxic to humans. Another problem with using chemicals to control plant pathogens is that a pathogen may become resistant to the chemicals.
The most promising control approaches are included conventional breeding and genetic engineering of disease-resistant plants. However, the conventional breeding method of melon is very complicated therefore; genetic engineering could be an effective and quick tool for producing new cultivars. In vitro direct regeneration is one of the most crucial step in gene transferring programs (20). In the present study, the effects of explant type and medium were considered in order to optimize the regeneration condition for melon (cv. Khatooni).
Material and Methods: The Khatoonicv.was provided by the Seed Bank of Plant Sciences Research Center from Ferdowsi University of Mashhad. The seeds were disinfected by NaClO in 1 % concentration, washed three times by distilled water and cultivated on MS medium (Murashigo and Skoog) for germination. The explant types were; lateral buds of cotyledon and the true leaves and hypocotyls. The selected media were: M1(0.1 mg.l-1 BAP (6-Benzyl amino purine)), M2 (0.25 mg.l-1 BAP), M3 (0.5 mg.l-1 BAP), M4 (0.75 mg.l-1 BAP),M5 (1 mg.l-1 BAP)and M6(including 0.5 mg.l-1 BAP and 0.01 mg.l-1 NAA (1-Naphthalene acetic acid))The experiment was repeated three times. The experimental –was conducted factorial based on completely randomized design and statistical analysis was performed using the SAS and JMP software. The corresponded graphs were drawn by Microsoft excel 2007.
Results and Discussion: The greatest amount of shoot regeneration in cotyledons was observed in M5 with 1 mg.l-1 BAP while minimum shoot regeneration was observed in M1with 0.1 mg.l-1 BAP. Cotyledon leaves showed the best regeneration efficiency among the other evaluated explants as it has also been reported by many other researchers (12, 8, 20, 21, and 2). The observations showed that in all BAP concentration, hypocotyls only formed callus tissue and did not produce any stem. In all three explants (true leaves, cotyledon and hypocotyls) rooting was observed in a treatment combination including 0.5 mg.l-1BAP and 0.01 mg.l-1 NAA 20 days after cultivation while no stem was formed at the same time. In three weeks after exposure , a weak root system was formed on the rooting mediumin without hormonetreatment . In MS medium enriched with 0.01 and 0.05 mg.l-1 NAA an appropriate root system was formed after 8 to 10 days. The medium containing 0.01mg.l-1 NAA was more appropriate for root regeneration (rooting).
Conclusion: Many studies have been shown that, the manipulation of different phytohormones ratio such as auxin and cytokinin in cultural medium is a suitable strategy to manage differentiation and organogenesis programs in plant (9). In the present study, the effects of explant type and medium were investigated in order to optimize the regeneration condition for melon (cv. Khatooni). As a result, in all BAP concentration, hypocotyls only formed callus tissue and did not produce any stem. However, by increasing of BAP hormone concentration in the cultivated media, shoot regeneration will also increase. The result of our study showed that, cotyledon leaves were more effective on shoot regeneration compared to the true leaves and hypocotyls as it has also been reported by other researchers.
Hassan Hasani Jifroudi; Mehdi Mohebodini; Behrooz Esmaielpour; Esmaeil Chamani
Abstract
Introduction: Fenugreek (Trigonella foenum- graecum) is a medicinal plant extensively distributed in most regions of the world. Fenugreek is an annual plant from the family of papilionaceae, leguminosae. Fenugreek leaves and seeds have been used extensively to prepare extracts and powders for medicinal ...
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Introduction: Fenugreek (Trigonella foenum- graecum) is a medicinal plant extensively distributed in most regions of the world. Fenugreek is an annual plant from the family of papilionaceae, leguminosae. Fenugreek leaves and seeds have been used extensively to prepare extracts and powders for medicinal uses. Its root, leaf and seed contain a number of important medicinal compounds such as polysaccharide, galactomannan, different saponins such as diosgenin, yamogenin, mucilage, volatile oil and alkaloids such as choline and trigonelline. Plant tissue culture is fundamental to most aspects of biotechnology of plants. Establishment of an efficient callus induction and direct regeneration protocol is an essential prerequisite in harnessing the advantage of cell and tissue culture for genetic improvement. For the successful application of the tissue culture technique in plant breeding, callus induction and plant regeneration potential of each plant must be determined. The present study was performed in order to determine the optimum concentration of plant growth regulators (IBA + TDZ) for producing of in vitro plantlet using cotyledon and hypocotyl as an explant for two different Iranian genotypes (Ardestani and Neyshabouri).
Materials and Methods: In this investigation, Ardestani and Neyshabouri genotypes were used for callus induction and direct shoot regeneration. The medium used in this investigation was MS (Murashige and Skoog) basal medium. Then seeds were germinated on MS medium. For callus induction and direct shoot regeneration, cotyledon and hypocotyl explants were excised from 8-day-old sterile seedlings and cultured on MS medium containing various concentrations of IBA and TDZ. In this experiment, two combinations (TDZ + IBA) were used. In the first composition, IBA had four levels (0, 0.1, 0.3, 0.5 mg l-1) and TDZ had five levels (0, 0.2, 0.4, 0.6, 0.8 mg l-1) and in the second composition, IBA had four levels (0, 0.05, 0.1, 0.15 mg l-1) and TDZ had seven levels (0, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45 mg l-1).The experimental designs were factorial based on completely randomized design with four replications. Cultures were incubated at 25° C ± 2 with a 16/8 hour (day/night) photoperiod and an irradiance of 1500 LUX using Sylvania cool white fluorescent tubes. The percentage of callus induction, direct shoot regeneration and average weight of callus were calculated for cotyledon and hypocotyl explants. All Data were analyzed using SPSS16, and mean comparisons were performed with duncan’s multiple range test (P < 0.05).
Results and Discussion: According to our results, explants cultured on MS basal medium without plant growth regulators (control) produced no callus. However, after two weeks, callus formed in both of Ardestani and Neyshabouri genotypes from cotyledon and hypocotyl explants in the presence of IBA + TDZ plant growth regulators in most of the combinations. In hypocotyl explants of Neyshabouri genotype, the highest callus induction was obtained from the medium containing 0.15 mg l-1 IBA + 0.45 mg l-1 TDZ (96.87%). Various important factors such as genotype, source of explants and plant growth regulators significantly influence direct regeneration. Direct regeneration was obtained from hypocotyl explants for Neyshabouri genotype in combination IBA + TDZ. The highest percentage of direct shoot regeneration was observed in MS medium containing 0.05 mg l-1 IBA + 0.35 mg l-1 TDZ in hypocotyl explants of Neyshabouri genotype (37.5%). Direct shoot regeneration requires plant cells to undergo dedifferentiation which is known to be affected by not only exogenous plant growth regulators but also endogenous content of the hormones. Different tissues may have different levels of endogenous hormones and, therefore, the type of explant source would have a critical impact on the regeneration success. In our study, when cotyledon and hypocotyl explants were compared, it was clear that hypocotyl explants were much more productive for direct shoot regeneration than cotyledon explants.
Conclusions: Callus induction and direct shoot regeneration are as in vitro tissue culture methods. Plant growth regulators and types of explant and genotype are the most important factors for callus induction and direct shoot regeneration phases. Therefore, optimization of these factors is essential to establish a high frequency of callus induction, direct shoot regeneration and gene transferring to this plant. According to the results of this investigation, it is recommended to apply plant growth regulators that were used in this study for other landraces of fenugreek cultured in Iran and select the best genotypes in response to tissue culture conditions for using in future studies.
