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.
Homeyra Hadizadeh; Mehdi Mohebodini; Behrooz Esmaeilpour; Esmaeil Chamani
Abstract
Introduction: Chicory (Cichorium intybus L.) belongs to Asteraceae family is commonly known as witloof chicory. The leaves and the roots of this medicinal plant are edible and commonly used as salad. Some varieties are also cultivated as coffee substitute after roasting the roots. All parts of the plant ...
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Introduction: Chicory (Cichorium intybus L.) belongs to Asteraceae family is commonly known as witloof chicory. The leaves and the roots of this medicinal plant are edible and commonly used as salad. Some varieties are also cultivated as coffee substitute after roasting the roots. All parts of the plant contain these volatile oils, with the majority of the toxic components concentrated in the plant's root. In folk medicine, the plant is used for the treatment of diarrhea, spleen enlargement, fever, and vomiting. Antihepatotoxic activity on damaged rat’s liver sections and anti-bacterial activity of this crop has been recently reported. In vitro regeneration from leaf explants with various hormonal combinations has been reported previously. Moreover, in vitro regeneration of Chicory from cotyledon explants using different combinations of plant growth regulators has been studied. Also, a protocol for the regeneration of plantlets from leaf and petiole explants of witloof chicory has been developed. The aim of the present investigation was optimization of callus induction and shoot regeneration from leaf and petiole tissues of Chicory (Esfahan genotype).
Materials and Methods: In this investigation, Esfahan genotype was used for callus induction and direct shoot regeneration. Seeds were first washed with running tap water for 30 min then seeds were surface sterilized by dipping in 70% ethanol for 90 s and rinsed with sterile distilled water, followed by immersing in 5% sodium hypochlorite solution for 25 min and thereafter rinsed for 30 min with sterile distilled water. The basal medium used in this investigation was MS. For shoot regeneration, leaf and petiole explants (5 mm segments) were excised from 4-week-old sterile seedlings and cultured on MS medium containing different combinations of NAA / BA and KIN / BA in two separate experiments. Experiments were performed factorial based on completely randomized design. 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, shoot regeneration and the number of regenerated shoots were calculated for the leaf and petiole explants. Data was subjected for analysis of variance and means were compared in 5% level with Duncan’s multiple range tests.
Results and Discussion: Explants cultured on medium containing either no plant growth regulators (control) or cytokines alone produced no callus. However, after 2 weeks, other concentrations of NAA and BA indicated callus formation from leaf and petiole explants in all hormone combinations. In leaf explants, the highest callus induction were obtained in the medium containing 0.3 mg l-1 NAA with 1 mg l-1 KIN and 0.3 mg l-1 NAA with 1.5 mg l-1 KIN (81.25%). Leaf and petiole explants cultured on medium containing no plant growth regulators (control treatment) and medium containing NAA produced no shoots. The combination of 0.3 mg l-1 NAA and 0.1 mg l-1 BA was the best treatment tested. This treatment produced 2.7 shoots per explant at 71% shoot regeneration frequency in leaf explant and 2.73 shoots per explant at 73% shoot regeneration frequency in petiole explants. The results also showed that the highest percentage of regeneration and the highest number of regenerated shoots were obtained in the medium containing 0.1 mg l-1 NAA and 1 mg l-1 KIN in leaf explants (65.6% regeneration and 1.37 shoots per explant, respectively). The highest number of regenerated shoots was obtained in the medium containing 0.3 mg l-1 NAA and 0.5 mg l-1 KIN in petiole explants (40.6% regeneration and 0.5 shoot per explants, respectively. 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 leaf and petiole explants were compared, it was clear that leaf explants were much more productive for regeneration than petiole explants.
Conclusion: Callus induction and shoot regeneration are in vitro tissue culture methods. Plant growth regulators and types of explant are the most important factors for callus induction and shoot regeneration phases. Therefore, optimization of these factors is essential to establish a high frequency of callus induction, shoot regeneration and gene transfer to this plant.
Esmaeil Chamani; Behrooz Esmaeilpour; Yunos Pourbeyrami Hir; Hassan Maleki Lajayer; Akbar Saadati
Abstract
Two separated experiments were conducted to evaluate the effects of different concentrations of thidiazouron (10, 20, 30, 40 and 50 µM) and humic acid (1, 10, 100, 1000 and 10000 ppm) on vase life of cut Alstroemria flowers. Experiments were carried out based on completely randomized design with 8 replications ...
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Two separated experiments were conducted to evaluate the effects of different concentrations of thidiazouron (10, 20, 30, 40 and 50 µM) and humic acid (1, 10, 100, 1000 and 10000 ppm) on vase life of cut Alstroemria flowers. Experiments were carried out based on completely randomized design with 8 replications in postharvest laboratory of Horticultural Department, Mohaghegh Ardabili University in 2009. The results showed that lower concentrations of humic acid had positive effects on flower vase life, water content of tissue and solution uptake, while didn’t affect relative fresh mass and leaf chlorophyll content. In both experiments results also indicated that humic acid in higher concentrations affected all traits, negatively. Howewer, this compound at 10000 ppm decreased flower vase life, water content, relative fresh mass, solution uptake and chlorophyll content. By increasing the humic acid concentration up to 100 ppm the vase life of cut flowers improved. Compared with control and other treatments, 1000 ppm of humic acid decreased flower vase life, considerably. The highest vase life of flowers devoted to 10 µm of TDZ, however higher concentrations of this compound reduced flowers vase life. Moreover, the highest solution uptake and leaf chlorophyll content obtained by 30 µm TDZ, while the highest relative fresh mass devoted to 40 µm of this compound.
