Maryam Zare Hasan Abadi; Ali Ganjeali; Mehrdad Lahouti; Nasrin Moshtaghi
Abstract
Introduction: Agrobacterium rhizogenes hairy roots induction is used for secondary metabolite production in plants. A. rhizogenes is a genus of gram-negative soil bacteria belonging to the Rhizobiaceae family that causes hairy roots at the site of infection. Hairy roots have various advantages, including ...
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Introduction: Agrobacterium rhizogenes hairy roots induction is used for secondary metabolite production in plants. A. rhizogenes is a genus of gram-negative soil bacteria belonging to the Rhizobiaceae family that causes hairy roots at the site of infection. Hairy roots have various advantages, including high growth rate, more genetic stability than the callus and suspension cultures, growing well on hormone-free media that have been reported effective for producing high levels of secondary metabolites. Basil (Ocimum basilicum) is a popular herb with important economical applications in food, cosmetic, and pharmaceutical industry. It is a digestive stimulant with anticarcinogenic, antibacterial, and anticonvulsant properties. The main phenolics reported in basil plants are in the classes of phenolic acids and flavonoids, some of which have human health benefits. This study was designed to develop hairy root culture from O. basilicum using different of A. rhizogenes strains for the production of total phenols and introduce the best strain of A. rhizogenes to induce hairy root and growth and production of total phenol.
Materials and Methods: Different A. rhizogenes strains (ATCC-15834, A4, MSU, and R1000) were studied to investigate their effects for the transformation and production of secondary metabolites in O. basilicum. Therefore, shoot and leaf explants and nodes of the seedlings were used for Agrobacterium-mediated transformation. These explants were inoculated with four A. rhizogenes strains and transferred to ½ MS medium. About four weeks after cultivation with A. rhizogenes, hairy roots were excised from the seedlings and subcultured to fresh medium MS liquid culture containing 500 mg/l cefotaxime. After 60 days of inoculation, various parameters, including dry weight, infection percentage, number of hairy roots per explant, and total phenol contents were measured. The growth rate and phenolic contents of the transformed hairy roots were compared with normal ones. Total genomic DNA was isolated from non-transgenic and transgenic hairy root lines using the Cetyl Trimethyl Ammonium Bromide (CTAB) method. Isolated genomic DNA was used to detect the rolC gene through polymerase chain reaction (PCR) analysis. The PCR using specific primers for rolC of T-DNA and virD2 was used to confirm the nature of resulted transgenic hairy roots.
Results and Discussion: Selecting efficient A. rhizogenes strains, as well as the type of explants, are crucial factors for hairy root induction. All used A. rhizogenes strains were able to produce hairy roots. Hairy roots appeared on the nodes at the point of injection, but were not forming on the shoot and leaf explants. So, the choice of the plant material is crucial for successful transformation with A. rhizogenes and usually, transformation of young tissues gives the best results. The transgenic status of the hairy roots was confirmed using PCR with rolC and virD specific forward and reverse primers. All lines showed the presence of 612 bp rolC amplified products, indicating the integration of T-DNA of A. rhizogenes and O. basilicum. Hairy roots could synthesize phenolic compounds, which was significantly higher in hairy roots than non-transformed control. Four hairy root lines were independently evaluated for their content and these lines showed variation in total phenolic contents, with the highest amount (312 mgGAE/ g DW) in hairy roots induced by ATCC-15834 strain and the lowest amount (113.2 mgGAE/ g DW) in hairy roots induced by R1000 strain. The results showed that the strain ATCC-15834 caused the highest infection percentage (68.1%) along with the highest number of hairy roots (4.8) per explant and root length (1.8 cm). The growth rate and phenolics production were investigated in each hairy root of O. basilicum from infection by four different A. rhizogenes strains. The highest growth rate (103.2 mg DW) and production of total phenol (312 mg/g DW) were found in ATCC-15834. The growth rate of transformed hairy roots was more than that of normal ones.Total phenol contents in all hairy roots were also increased significantly compared with non-transformed control plants (4.6 times in hairy roots induced by A. rhizogenes strain ATCC-15834). ATCC-15834 has been reported as the most widely used A. rhizogenes strain owing to its strong induction ability, and the variation in hairy root induction could be due to disparity in the virulence of different A. rhizogenes strains.
Conclusion: The hairy roots of O. basilicum had shown promising results in terms of significant yield of phenolic contents and had the potential for being scaled-up further for phenol production. It could be concluded that A. rhizogenes strains had different abilities in hairy roots induction. Therefore, the selection of an effective A. rhizogenes strain for the production of transformed root cultures is important, highly dependent on the plant species, and must be determined in future experiments.
Azadeh Saffar Yazdi; Ali Ganjeali; Reza Farhoosh; Monireh Cheniany
Abstract
Introduction: Purslane (P. oleracea) is considered as valuable plant due to its high antioxidant compounds and important fatty acids such as omega-3 and 6. Phenolic and flavonoid compounds are one of the most important constituents in the purslane. Phenolics are a large group of natural plant compounds ...
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Introduction: Purslane (P. oleracea) is considered as valuable plant due to its high antioxidant compounds and important fatty acids such as omega-3 and 6. Phenolic and flavonoid compounds are one of the most important constituents in the purslane. Phenolics are a large group of natural plant compounds with antioxidant and Anti-inflammatory properties. Flavonoids, as a subset of phenolic compounds, have a wide range of effects on plants, including antioxidant activity and improve resistance to environmental stresses. Callus culture is one of the important strategies for the production secondary metabolites, which are difficult to produce chemically. Plant growth regulators including auxins and cytokinins play a crucial role in the stages of plant growth. Various combinations of these two hormones are used to make the desired changes in the cultures. Studies suggest that the accumulation of secondary metabolites can be increased by the application of different elicitors in medium. Researchers reported an increase in the content of secondary metabolites such as phenol and flavonoid compounds in calli treated with elicitors such as yeast extract. The purpose of this study was to determine the best explant, medium and hormonal treatment for calli induction of purslane. The effect of different levels of yeast extract on total phenol and flavonoid content and antioxidant capacity of purslane calluses was also investigated.
Materials and Methods: Seeds of purslane plant were cultivated in a solid 1/2MS medium for the preparation of sterile seedlings. The explants from sterile seedlings including to leaves, 1 cm stem specimens and terminal buds, were placed on MS and 1/2MS medium containing 0, 0.1, 0.3, and 0.5 mg L-1 BAP and NAA. After five months, calluses were evaluated for callogenesis and some morphological traits such as color, texture, and size, fresh and dry weight. This experiment was conducted based on completely randomized design with three replications. In the second experiment, the calluses obtained from the previous stage were transferred to MS medium with selected hormone treatment of the first experiment (0.5 mg L-1 NAA and BAP) and different levels of yeast extract (0, 125, 250, and 500 mg L-1). Total phenol and flavonoid contents of the calluses were determined by Folin-Ciocalteau and aluminum chloride methods, respectively. Furthermore, Ferric reducing antioxidant power (FRAP) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) assays were used to determine the antioxidant activities.
Results and Discussion: The results showed that 1/2MS medium was suitable for sterile seedling production from purslane seeds. Based on the present study, only stem explants in a medium containing BAP and NAA, produced durable calluses. The color of the resulting calluses were green and had a constant and firm texture. The highest callus percentage (90.46%), the size (21.6 mm), and fresh (1826.5 mg) and dry weight (75.33 mg) of calluses belong to MS medium containing 0.5 mg L-1 BAP and NAA. Results of the second experiment showed positive and significant effects of yeast extract on the total phenol, flavonoid contents and antioxidant activities. The highest content of total phenol (664.12 mg GAE 100g-1 DW), flavonoid (42.25 mg QE 100g-1 DW) and FRAP data (787 µmol Fe g-1 DW) were obtained from the calli treated with 500 mg l-1 yeast extract. The maximum DPPH IC50 (2.45 mg ml-1) was also observed in control. The formation of callus associated with plant species, hormonal composition, the stage of development, and the type of explants. Auxin and cytokinin as plant growth regulators are key factors for controlling cell division in tissue culture. In most studies, callus formation in purslane plant were induced in medium containing auxin and cytokinin. The presence of green calluses derived from purslane explants can be due to the formation of chloroplastids in the cells of the callus tissue that rapidly produce chloroplasts under light conditions. In the second experiment, increased phenolic and flavonoid compounds with yeast extract treatment probably resulted in increased antioxidant activity.
Conclusion: In the present study, 1/2MS medium is suitable for the production of sterile seedlings from purslane seeds. MS medium containing 0.5 mg l-1 BAP and NAA is the best treatment for calli induction from stem specimens. The concentration of 500 mg L-1 of yeast extract is introduced as the most effective concentration for increasing the phenolic and flavonoid content and antioxidant activity in the purslane calluses.
Simin Irankhah; Ali Ganjeali; Mehrdad Lahouti; Mansour Mashreghi
Abstract
Introduction: Fenugreek (Trigonellafoenum-graecum L.) is a traditional medicinal plant belonging to the legume family Fabaceae. Diverse groups of microorganisms are symbiotic with Fenugreek roots system. This integration leads to significant increases in the development and production by increasing nitrogen ...
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Introduction: Fenugreek (Trigonellafoenum-graecum L.) is a traditional medicinal plant belonging to the legume family Fabaceae. Diverse groups of microorganisms are symbiotic with Fenugreek roots system. This integration leads to significant increases in the development and production by increasing nitrogen fixation, phytohormones production, siderophores and phosphate solubilization. Plant growth-promoting bacteria increase plant growth byimproving nutrientuptake and phytohormones production. In addition, the beneficial effect of these bacteria could be due totheirinteractionwithArbuscularMycorrhizal fungi(VAM). Drought is one of the major limiting factors for crop production in many parts of the world including Iran. Symbiotic microorganisms can enhance plant tolerance to drought. This experiment was carried out to investigate the effect of Vesicular ArbuscularMycorrhiza (VAM) and Plant Growth Promoting Rhizobacteria (PGPR) on morphological and biochemical characteristics of Fenugreek in drought stress conditions.
Materials and Methods: The experiment was carried out in completely random design with 3 replications.There were four treatments including inoculation with Pseudomonas putida, inoculation with Glomusintraradices, combined association of Pseudomonas putida and Glomusintraradices and untreated as a check under drought stress (40% of field capacity) and non-stress conditions (80% of field capacity). In this experiment fiveseeds were sowninplastic pots. Before sowing, seeds were inoculated with microorganisms. In order to inoculation ofseed with Mycorrhizal fungi, for each kilogram of soil, 100 grams of powder containing 10 to 15 thousand spores of fungal soil (produced by the biotech company Toos) was added to three centimeters of soil in the pot. For seed inoculation with Plant Growth Promoting Rhizobacteria, the growth curve of the bacteria was drawn at first and then the best time for the growth of bacteria was determined. The bacteria at the best time and at a dilution of 0.5 McFarland was added to the seed.Pots were placed in a growth chamber (with a temperature of 25 ° C and 16 hours of light and 8 hours of darkness photoperiod).After ten days of planting, the water treatment was applied and continued until the end of the experiment.
Results and Discussion:The results showed that the use of co -inoculation treatment ofP.putidaandG.intraradicesand application of G.intraradices aloneat non-stress conditions did not make a significant difference on the amount of plant biomass. In drought conditions, application of G.intraradice alone had significant difference (P≤0.05) compare with control (no inoculation microorganisms) for biomass.In stress conditions, P.putida inoculation and also in combination with G.intraradicesincreased biomass in compare with control (no inoculation of micro-organisms), but this increasing was not statistically significant.VAMare important ecological symbiotic with roots are important component of the ecosystem and affect the absorption of minerals through the roots. The results of the present study showed that the amount of phosphorus in all of treatments was increased and the greatest increase was related to G.intraradicetreatment.The results also showed that drought stress increased the leaf soluble proteins in compared with non-stress condition. Increasing the concentration of soluble proteins under drought stress can be related to increased protein synthesis that maybe related to adaptation and reprogramming under new situation and itprotect the cells against stress. The results showed that the use of co -inoculation treatment ofP.putidaand G.intraradices, as well as each individual treatment, increased the amount of soluble proteins in leaves.In the case of the Diosgenin percentage, drought stress reduced the amount of Diosgenin percentage. Underdrought stress conditions, use of co -inoculation treatment ofP.putidaand G.intraradicesand application of G.intraradicewithout presence of bacteria made a significant increase in plant Diosgenin concentration.
Conclusion: Theresults revealed that seed treatment with Pseudomonas putida and Glomusintraradices increased the biomass, protein content, phosphorus uptake and diosgenin percent in Fenugreek under drought stress condition. Since Diosgenin is very important medicinal compound, inoculation of fenugreek with these microorganisms can be a way to increase the Diosgenin production.
Masumeh Modarres; Mehrdad Lahouti; Ali Ganjeali; Javad Asili
Abstract
Salvia leriifolia (Lamiaceae) is endemic of Khorasan and Semnan province and an endangered plant. Poor seed germination of this plant has a serious problem with the high production. The first step to improve this precious plant is to produce sterile plantlets in order to prepare explants of appropriate ...
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Salvia leriifolia (Lamiaceae) is endemic of Khorasan and Semnan province and an endangered plant. Poor seed germination of this plant has a serious problem with the high production. The first step to improve this precious plant is to produce sterile plantlets in order to prepare explants of appropriate vigor. In this study,in vitro culture of Salvia leriifolia zygotic embryo was performed through a factorial experiment in the form of completely random design including culture medium, BAP and NAA. The results showed that, MS and 1/2MS media appeared to be more efficient than B5 medium and significant differences were observed. Efficient concentrations of BAP and NAA were 1mgL-1 and had a significant effect in growth and development of embryos. The seedlings were obtained 10 days after planting. Based on these results, the best choice for quick access to strong seedlings, seedling growth and development of embryos is MS and 1/2MS media supplemented with 1mgL-1 BAP and NAA.
Parvaneh Abrishamchi; Ali Ganjeali; Abdollah Beyk Khormizi; Amir Avan
Abstract
Two separate experiments were carried out with the aim of investigating the effect of vermicompost and its aqueous extracts on traits related to germination and growth of tomato varieties, (Mobil and Superorbina). In the first experiment, 0.5, 1, 1.5, 2, 2.5, 5, 7.5, 10 and 100% extracts of vermicompost ...
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Two separate experiments were carried out with the aim of investigating the effect of vermicompost and its aqueous extracts on traits related to germination and growth of tomato varieties, (Mobil and Superorbina). In the first experiment, 0.5, 1, 1.5, 2, 2.5, 5, 7.5, 10 and 100% extracts of vermicompost (V - V) along with control (distilled water) were studied on germination of two varieties of tomato. The number of germinated seeds was counted each day and was sampled after a week. Then, the percentage and rate of germination and length and dry weight of shoot and root were determined. In the second experiment, volume ratios of vermicompost to sand, including to, 0:100; 10:90; 20:80; 40:60; 60:40; 80:20 and 100: 0, on tomato varieties seedling growth were investigated. The seeds were planted in plastic pots and the sampling of seedlings was carried out 22 days after planting. Both experiments were conducted in Completely Random Design with four replications. The results showed that vermicompost extract could not improve the growth of Mobil and Superorbina varieties, whereas application of vermicompost, had a significant effect on plant height, area and dry weight of leaves, stem dry weight, area, diameter and dry weight of roots and the concentration of potassium, calcium and phosphorus in tomato leaves. These results confirm that using low ratios of vermicompost, have more effect on the growth of tomato seedlings rather than high ratios, so that the maximum effect of vermicompost were recorded in the ratios of 20 and 40 % for Mobil and Superorbina varieties, respectively. In this experiment, high vermicompost ratios had a negative effect on growth of tomato seedlings.
Zahra Setayesh-Mehr; Ali Ganjeali
Abstract
In order to study the effect of different water potential on Anethum graveolens L. an experiment was conducted. This study was carried out based on completely randomized design with three replications. In this experiment, morpho-physiological characteristics were assessed under hydroponic. Stress was ...
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In order to study the effect of different water potential on Anethum graveolens L. an experiment was conducted. This study was carried out based on completely randomized design with three replications. In this experiment, morpho-physiological characteristics were assessed under hydroponic. Stress was established with PEG 6000, and drought potentials were: 0 (control), -1.5, -2, -2.5 and -3 bar. The results showed that effect of drought stress on morphological characteristics including: shoot length, root length, surface of leaves and leaves number was significant (p≤0.01). Mean comparison of treatments showed that with increasing drought stress, all parameters decreased. Effect of drought stress on amount of a, b and total chlorophyll, carotenoid, soluble proteins, phenolic compounds and K+, P and Ca+2 concentrations in shoot and root were significant (p≤0.01). Effect of drought stress on chlorophyll a/b ratio was significant (p≤0.05). Mean comparison of treatments showed that with increasing drought stress, amount of chlorophyll, carotenoid, soluble proteins, K+, P and Ca+2 and shoot/root K+ ratio decreased where as amount of phenolic compounds in shoot and root increased. Shoot to root length ratio and shoot/root K+ ratio were not influenced by drought stress (p≤0.05). From the results of this study, it can be deducted that dill medicinal plant respond to drought stress by increasing of phenolic compounds.
Azadeh Saffar Yazdi; Mehrdad Lahouti; Ali Ganjeali
Abstract
Selenium is a metalloid that it is antioxidant characteristics is proved for humans, animals and plants. However, some plants are known as hyper accumulators for selenium and sometimes seems to be useful for growth of some plants. In this paper, we have studied the effect of different selenium concentrations ...
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Selenium is a metalloid that it is antioxidant characteristics is proved for humans, animals and plants. However, some plants are known as hyper accumulators for selenium and sometimes seems to be useful for growth of some plants. In this paper, we have studied the effect of different selenium concentrations on growth and morpho-physiological characteristics of spinach. Seeds of spinach were germinated in germinator and seedlings were transferred to hydroponic cultures. The seedlings were grown in Hogland´s solution with different selenium concentrations (in form of sodium selenite) of 0,1,2,4,6, and 10 mgL-1 SeO3-2. The experiment was conducted based on a completely randomized design with four replications. Four weeks after treatments, morphophysiological characteristics including plant biomass, length of shoot and root, number of leaves, amount of chlorophyll a and b were measured. The results showed that application of different selenium concentrations had significant effect on morphological and physiological characteristics of root and shoot consisting wet and dry weight, root and shoot length and amount of chlorophyll. In this experiment, increasing selenium concentrations (except in 1 mgL-1 SeO3-2 concentration) decreased amounts of all mentioned characters in comparison with control. Morphological symptoms of selenium toxicity on spinach was chlorosis on young leaves and significant reduction in root and shoot growth.
Zahra Khakshoor Moghadam; Mehrdad Lahouti; Ali Ganjeali
Abstract
Abstract
In order to study the effect of different water potential on Anethum graveolens two experiments were conducted for germination and vegetative stages. This study was carried out as a factorial experiment based on completely randomized design with three replications. In the first experiment, ...
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Abstract
In order to study the effect of different water potential on Anethum graveolens two experiments were conducted for germination and vegetative stages. This study was carried out as a factorial experiment based on completely randomized design with three replications. In the first experiment, response of dill seeds germination to levels of drought stress including to drought potentials zero(control), -1.5, -2, -2.5 and -3 bar that were obtained by polyethylene glycol 6000 were investigated. In the second experiment, similar to first, influence of drought stress on dill morphophysiological parameters were investigated. The results showed that effect of drought stress on germination characteristics including to germination percentage, germination rate, radicle and plumule length , radicle and plumule dry weight , plumule to radicle dry weight ratio was significant (p≤ 0.01). Mean camparison of treatments showed that with increasing drought stress, all parameters decreased. Effect of drought stress on amount of proline and soluble carbohydrates in shoot and root was significant (p≤ 0.01). Mean camparison of treatments showed that with increasing stress, amount of proline and soluble carbohydrates in shoot and root, shoot to root proline and soluble carbohydrates ratio increased. Shoot to root proline and soluble carbohydrates ratio were not influenced by drought stress (p≤0.05).
Keywords: PEG, Morphological parameters, Proline, Soluble carbohydrate
Matin Jami; Seyyed Ali Mohammad Modarres; Peyman Keshavarz; Ali Soroushzadeh; Ali Ganjeali
Abstract
Abstract
To evaluate the influence of nitrogen rate and split application method on tuber yield and other quantitative characteristics of different potato cultivars, a 2-yr experiment was conducted at Khorasan Razavi Agricultural and Natural Resources Research Center. Experimental treatments consisted ...
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Abstract
To evaluate the influence of nitrogen rate and split application method on tuber yield and other quantitative characteristics of different potato cultivars, a 2-yr experiment was conducted at Khorasan Razavi Agricultural and Natural Resources Research Center. Experimental treatments consisted of four nitrogen rates (0, 90, 180 and 270 kg N ha-1) with two split applications (emergence and hilling stages) or three split applications (planting, emergence and hilling stages) and six potato cultivars (Fontane, Kuras, Agria, Miriam, Cosmos and Picasso) that were arranged in a randomized complete block split-split plot design. Shoot dry weight, average stem length, tuber number per plant, total tuber yield, marketable tuber yield (
