Hoda Zare Mirakabad; Mohammad Farsi; Saeid Malekzadeh Shafaroudi; Mehrdad Iranshahi; Abdolreza Bagheri; Nasrin Moshtaghi
Abstract
Introduction: There is a growing body of the literature that recognizes the importance of ferutinin (C22H30O4) as one of the natural phytoestrogens with potency to treat osteoporosis and some kind of cancers. One of the greatest challenges is availability of ferutinin that is found just in root of some ...
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Introduction: There is a growing body of the literature that recognizes the importance of ferutinin (C22H30O4) as one of the natural phytoestrogens with potency to treat osteoporosis and some kind of cancers. One of the greatest challenges is availability of ferutinin that is found just in root of some plants of the genus Ferula (Apiaceae), which is the reason of high price of it in global markets. Ferula ovina , an endemic plant of Iran, is known as one of the greatest sources of ferutinin. Unfortunately, access to ferutinin requires uprooting Ferula ovina especially older plants with more secondary metabolites content. There is a large volume of published studies describing the importance of tissue culture in propagation of endangered plants including secondary metabolites without possibility of chemical production and with deep dormancy seed exactly like the characters of F. ovina. Up to now, far too little attention has been paid to importance of tissue culture in accessing ferutinin without degrading the germplasm resources of it. The main aim of this study was to find an approach to access ferutinin associated F. ovina germplasm conservation.
Materials and Methods: In this experiment, the aerial parts of F. ovina were collected from Zoshk area (Mashhad, Iran). The plants were recognized as F. ovina by the Institute of Plant Sciences (Ferdowsi University of Mashhad). Tissue culture part was performed with preparation of sterilized media and explants. Therefore, the MS salts and vitamins was applied as basic medium, however MS salt was decreased to half strength for rooting of shoots. The node (root junction) explants were cultured on 24 callus induction/shooting media with different combinations of plant growth regulators (BAP, IAA, Kin, NAA and IBA). The shoots from direct and indirect regeneration were then transferred to media with different combinations of PGRs (BAP, IBA, IAA and NAA) in order to find rooting medium. Following these treatments, unbalanced ANOVA for analysis of data were performed using IBM SPSS 19. The final stage of the study comprised a TLC test for the purpose of finding ferutinin in samples which resulted from tissue culture. For this purpose, air-dried parts of samples were powdered and extraction was done after being in 3-5 times dichloromethane for 24hours. Then after optimizing solvent system was done with selection the ratio that presented bands in middle of TLC paper.
Results and Discussions: The results indicated that from 24 tested media, just 8 of them had potency of callus formation, but just L6 on MS medium containing 1 mg/l BAP and 1.5 mg/l IAA showed significant difference for percentage of callus induction at 5% level with compact green and the heaviest calluses. Although direct and indirect shoot regeneration was observed in this study, L9 on MS medium along with 1.5 mg/l IAA and 3 mg/l kin demonstrated significant difference for percentage of shooting at 5% level. Moreover, R6 on ½MS with 1 mg/l IBA showed significant difference for percentage of rooting of shoots at 5% level. The most surprising observation to emerge from the data comparison was L24 on half strength MS medium containing 0.2 mg/l BAP and 3 mg/l IBA with potency of callus induction, shooting and rooting of shoots. Regarding to the results of TLC test, ferutinin positive bands were observed in some samples.
Conclusions: The aim of the present research was to examine possibility of achieving ferutinin without need to uproot F. ovina because there are several problems to achieve this valuable sesquiterpene: 1) Chemical production of ferutinin is impossible, 2) It could be accessible just from roots of genus Ferula, 3) Propagation with seeds of F. ovina is limited because of morphophysiological dormancy of them, 4) Natural habitat of this plant in Iran is going to be destructed, 5) Access to natural habitat is difficult, 6) Time of access is limited with short growth season, 7) Maintaining F. ovina in greenhouse condition is impossible. The results of this research support the idea that producing ferutinin in Iran without any harmful effect on germplasm resources of F. ovina is possible. This is the first study of high scale commercial production of ferutinin which examine associations between tissue culture and ferutinin production.
Saba Nejatie Zadeh; Saeid Malekzadeh Shafaroudi; Ali Reza Astaraei; Nasrin Moshtaghi
Abstract
Introduction: An emerging field of nanotechnology in recent years is the use of nanoparticles and nanomaterials in agricultural systems which is due to their excellent mechanical, electrical, optical, surface properties, crop protection and nano-fertilizers. Titanium dioxide (TiO2) is a class of nanoparticles ...
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Introduction: An emerging field of nanotechnology in recent years is the use of nanoparticles and nanomaterials in agricultural systems which is due to their excellent mechanical, electrical, optical, surface properties, crop protection and nano-fertilizers. Titanium dioxide (TiO2) is a class of nanoparticles which widely used in the food industry, cosmetics, papers, pharmaceuticals, plastics and industrial raw materials. The widespread industrial application of TiO2 is due to its white pigment, ultraviolet blocking property, and chemical features commonly used to alleviate pollutants concentration in water, soil and air. Owing to its increasing use in the industry, a large part of TiO2 residues are released into the environment, and currently, TiO2 nanoparticles are being considered an emerging environmental contaminant. However, there have been a number of studies reporting beneficial effects of TiO2 on growth and physiological traits of crops. It has been postulated that the TiO2-induced improvement of crop growth is not merely related to the promotion of photosynthesis; other biochemical processes especially nitrogen metabolism are also involved in this event. Ethylene diamine tetraacetic acid (EDTA) is a widely used as a chelating agent, i.e., the chemical is able to sequester metal ions such as Ca2+ and Fe3+. EDTA is used as nitrogen source for doping of TiO2 nanoparticles which improves TiO2 photocatalytic features. The present study was conducted to investigate the effects of TiO2 nanoparticles and EDTA on growth indices and biochemical parameters in spinach (Spinacia oleracea). For detailed evaluation of treatment effects, different concentrations of TiO2 nanoparticles were sprayed on spinach leaves and the samples were collected in a time course.
Materials and Methods: A factorial experiment was carried out in the form of completely randomized design (CRD) with three replications. Soil samples were taken before cultivation of spinach (S. oleracea) seeds (Var VIROFLAY) and analyzed for nutrients’ concentration. Treatments include different levels of TiO2 (T1=0, T2=0.05mg/l and T3=0.1mg/l) and two concentrations of EDTA (E1=0 and E2=130mg/l) sprayed on spinach plants in research greenhouse of agriculture faculty, Ferdowsi University of Mashhad. Aqueous solutions of nanoparticles were treated by ultrasound for 10 min to enhance homogeneity. The solutions were sprayed on the plant at six- leaves stage. The plant samples were taken before reproductive phase for measurement of biochemical parameters. Nitrogen content of plant samples was measured by PDV 500 Macro- Kjeldahl device; Potassium content was determined by 310c flame photometer; phosphorus concentration in plant samples was measured by spectrophotometer model 2100. Chlorophyll and carotenoid contents were measured by the method proposed by Lichtenthaler (1978). For analysis of growth parameters, plant samples were taken a week after TiO2 treatments and leaf area, shoot fresh/dry weight, stem length, internode length, root area, root fresh/dry weight and total root diameter were measured.
Results and Discussion: Application of 0.05mg/l of TiO2 nanoparticles without EDTA resulted in 13.5% and 9.48% increase in nitrogen and protein; respectively, however by increasing nanoparticles to 0.1mg/l, nitrogen and protein content in the treated plants were respectively reduced to 21% and 19.57% of those of control group (p
Hajar Shayesteh; Saeid Malekzadeh Shafaroudi; Kamal Ghous; Farajollah Shahriari
Abstract
Introduction Jujube (Zizyphus jujuba Mill.) as a valuable medicinal plant and adapted to different climatic conditions is widespread in many parts of Iran. Nowadays, beside the export of its fruit, jujube is also used as an herbal medicine to treat the diseases, so it has a high economic value. Study ...
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Introduction Jujube (Zizyphus jujuba Mill.) as a valuable medicinal plant and adapted to different climatic conditions is widespread in many parts of Iran. Nowadays, beside the export of its fruit, jujube is also used as an herbal medicine to treat the diseases, so it has a high economic value. Study on genetic diversity is the first step to identify and preservation of germplasm. It is also considered as the basic principles of plant breeding. DNA markers seem to be the best way in determination of the genetic diversity. Inter simple sequence repeats (ISSR) markers are highly polymorphic and combine most benefits of Simple Sequence Repeats (SSRs) and amplified fragment length polymorphism (AFLP) to the generality of random amplified polymorphic DNA (RAPD).
Materials and Methods In order to study of the genetic diversity among 31 ecotypes collected from eight Jujube-rich provinces, including South Khorasan, Razavi Khorasan, Mazandaran, Golestan, Qom, Isfahan, Lorestan and Fars. Genomic DNA was extracted by CTAB method and polymerase chain reaction (PCR) was performed with 13 ISSR primers in which six most efficient primers were selected. Cluster analysis based on Dice similarity coefficient and Unweighed Pair Group Method with Arithmetic Mean (UPGMA) was carried out and POPGENe.3.2 software was used to determine the similarity of populations with each other.
Results and Discussion 84 loci were amplified and 70 of them (83%) revealed a proper polymorphism with the size between 200 and 2000 base pair. The average number of amplified and polymorphic bands per primer was 14 and 11.6 respectively. Primers with di-nucleotide repeats produced more polymorphic bands than ones with tri-nucleotide repeats. It seems that this is due to a higher frequency of sequences containing di-nucleotide repeats in intergenic regions and higher possibility of mutation revealed in more diversity in comparison to gene coding regions. Anchored primers with 1 or 2 nucleotides at the 5’ end make sure annealing only to the ends of SSRs in template DNA, so avoiding internal priming and smear formation. In addition, the anchor lets only a subset of the microsatellites to serve as priming sites. Primers (AC)8YT and (GA)8A with the higher percentage of polymorphism is recommended for further analysis. According to the cluster analysis, the ecotypes could be classified into seven main groups at the 0.85 level of genetic similarity. The most genetic similarity (0.95) was observed between ecotypes from Kalaleh and Doroh and also Noghab and Dustiran and the least genetic similarity (0.48) observed between Kangan and Borzaderan. POPGENe.3.2 software data indicated that populations of Isfahan and South Khorasan had the slightest difference while populations of Isfahan and Razavi Khorasan showed the most difference.
Conclusions In general results demonstrated that the total diversity of jujube ecotypes in Iran is summarized in the area of South Khorasan province. Given data showed that South Khorasan has been an original place of cultivation of this medicinal plant, this area could be considered as one of the important centers of jujube diversity. In addition, significant levels of diversity were observed among ecotypes belonging to Isfahan and Mazandaran provinces.
Seyedeh Zeinab Attari; Mahmood Shoor; Mahmoud Ghorbanzadeh Neghab; Ali Tehranifar; Saeid Malekzadeh Shafaroudi
Abstract
Introduction: Some of Iris species are growing in different parts of the Iran as wild species. Iris species have important medicinal and horticultural properties. Understanding of the genetic variation within and between populations is essential for the establishment of effective and efficient methods ...
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Introduction: Some of Iris species are growing in different parts of the Iran as wild species. Iris species have important medicinal and horticultural properties. Understanding of the genetic variation within and between populations is essential for the establishment of effective and efficient methods for conservation of the plants. Genetic variation studies are fundamental for the management and conservation of this species. The use of molecular markers is a powerful tool in the genetic study of populations. The use of DNA marker, such as AFLP, SSR, RAPD and ISSR represents an alternative method in detection of polymorphism. ISSRs are highly variable, require less investment in time, money and labor than other methods. ISSR can generate higher percentages of polymorphic loci than other PCR methods. These can serve as an efficient tool for phylogenetic studies. ISSRs had reported that used in studies of cultivated species to produce genetic linkage maps and to determine the relatedness of lines of agriculturally important species. ISSR analysis involves the PCR amplification of regions between adjacent, inversely oriented microsatellites, using a single simple sequence repeat (SSR) motifs (dinucleotide, trinucleotide, tetranucleotide or penta nucleotides). Therefore, little is known about the genetic variability of the Iranian Iris ssp .The objectives of this study were to evaluate genetic diversity among genotypes using ISSR markers and the degree of polymorphism generated from ISSR technique as a pre-requisite for their applicability to population genetics studies in Iris ssp.
Materials and Methods: To evaluate genetic variations in some wild Iris genotypes, Iris kopetdaghensis ،Iris songarica and Iris fosteriana were collected from some parts of Khorasan province. Genomic DNA was extracted from young leaves following the cetyltrimethylammonium bromide (CTAB) procedure. Extracted DNA concentration was quantified by using the spectrophotometer and qualified using agarose gel electrophoresis. A total of 16 primers were initially screened against two plants selected from different regions and finally six primers for final analysis was selected based on consistent (CA)8G ،(CT)8RG ،(TC)8C ،(TG)8G ، (AC)8YG and (AG)8YT, strong amplification products, production of polymorph, reproducible fragments between replicate Polymerase Chain Reaction (PCR). The ISSR amplification reactions contained 30-50 ηg of genomic DNA, 2.5 μL 1 × buffer, 2 mM MgCl2, 200 μM of each dNTP (Fermentas), 10 μM primers and 0.2 U Taq DNA polymerase (Fermentas), with the final volume adjusted to 25μL with H2O bidest. ISSR reaction products were separated on 1.5% horizontal agarose gels, in TBE buffer and visualized under ultraviolet light after staining in 0.5μg/mL ethidium bromide. Digital photo was taken with gel documentation system. The 100 bp DNA ladder plus molecular weight marker was used to compare the molecular weight of amplified products. Amplified products were scored for the presence (1) or absence (0) of bands and binary matrices were assembled for the ISSR markers. The binary matrices were subjected to statistical analyses using NTSYS-pc software version 2.02.
Results and Discussion: Six ISSR primers produced 126 bands across the 16 genotypes, of which 119 were polymorphic. The number of amplified fragments varied from 16 [primer (CA)8G)] to 24 [primer (TC)8C and (AC)8YG)] across the genotypes. The average polymorphic bands per primer were 19.4. The percentage of polymorphism for primers ranged from 76 to 100, with an average of 94.4.The amplified bands genotypes related to a species the same banding pattern was observed but there was lower similarity between the species. Our data indicated that ISSR technology can detect considerable polymorphisms (76.4 %) in our genotypes, suggesting that it will be useful in characterization and fingerprinting of Iris germplasm. The results of this study also provide fundamental evidence demonstrate that ISSR marker is a simple, informative, reproducible and suitable approach to evaluation of molecular diversity and phylogenetic relationships in Iris spp. The highest genetic similarity was between species Iris kopetdaghensis and Iris fosteriana. This study revealed a significant variation especially between Iris kopetdaghensis and Iris songarica.
Conclusions: The results of cluster analysis showed that molecular markers able to identify the species and genotypes within a species from each other. Results of this study showed that the use of molecular markers in breeding programs, especially fingerprinting is useful for lily. ISSR molecular markers have proved to be an efficient tool for studying genetic diversity and management of lily germplasm. . Also the result showed these genotypes have high genetic diversity, and the success in Iris breeding programs use to recommend Iranian local Iris.
A. Khazaei; N. Moshtaghi; Saeid Malekzadeh Shafaroudi; Kamal Ghous
Abstract
Introduction: Jujube (Ziziphus jujuba) is one of the most important fruit trees in Asia which has been planted from 3,000 years ago in China for medicinal purposes. Jujube belongs to the Rhamnaceae family. The Jujube fruit is used in fresh and dry forms. The fruit is full of vitamin C and has anticancer ...
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Introduction: Jujube (Ziziphus jujuba) is one of the most important fruit trees in Asia which has been planted from 3,000 years ago in China for medicinal purposes. Jujube belongs to the Rhamnaceae family. The Jujube fruit is used in fresh and dry forms. The fruit is full of vitamin C and has anticancer and medicinal effects. This tree can grow on salty and dry lands in Iran. Therefore, increasing the cultivation area of Jujube can be effective for soil conservation. In the last 20years, cultivation of Jujube is is considerable in Iran specially in the South Khorasan Province and 98 % of total production of Jujube in Iran belongs to this province. The low rate of seed germination and low production of shootlets are the most important problems in Jujube proliferation, so micropropagation of this plant through tissue culture was considered.
Materials and methods: In this study, Cangan ecotype of Jujube was used for multiple shoot regeneration. At the end of May, apical buds of shoots were cut from mature trees of the Research Collection of Jujube at Sarbishe, Birjand, South Khorasan Province in Iran. The buds were disinfected with 70% ethanol for 1 min and 2% sodium hypoclorite for 25 min. Then the buds were rinsed with distilled water for 25 min completely. Apical buds were placed on Murashige and Skoog (MS) medium supplemented with different concentrations of BA (0.5, 1, 1.5, 2 mg/L) in combination with IBA or NAA (0, 0.1, 0.2, 0.4 mg/L). After one month, the shoots with 3-5 cm length were transferred to rooting media (1/2 MS + IBA or IAA : 0.5, 2, 5, 10 mg/L). The data were recorded after shooting and rooting and were analysed in the facorial experiment.
Results and Discussion: The results of variance analysis and mean comparisons showed that there are differences between different levels of IBA and BA alone for the number of shoots and their length (P
Saeid Malekzadeh Shafaroudi; Askar Ghani; Maziyar Habibi; Akram Amiri
Abstract
Abstract
In order to induce autopolyploidy in basil (Ocimum basilicum) two separate experiments (seed treatment and cotton plug method on apical meristem) was conducted in a factorial experiment based on completely randomized design including colchicines concentrations (0, 0.05%, 0.1% and 0.2%) and ...
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Abstract
In order to induce autopolyploidy in basil (Ocimum basilicum) two separate experiments (seed treatment and cotton plug method on apical meristem) was conducted in a factorial experiment based on completely randomized design including colchicines concentrations (0, 0.05%, 0.1% and 0.2%) and the treatment time (6, 12 and 24 h) with four replications. On seed treatment all plants except control were perished. Morphological characteristics and flowcytometry data were used to detect and confirm polyploidization. The morphological and microscopic results showed that colchicines concentrations significantly affect autoploidy induction and the most cases (3.63%) observed on 0.05% concentration. Treatment time and interaction between concentration and time did not show a significant effect on this character. However the simple effects of concentration and treatment time were significantly different on survival percentage. Also concentration X time interaction showed a significant effect on this trait. Among the colchicines concentration levels, the second level (0.05%) showed the maximum survival percentage (47.7%) after the control. Higher concentrations caused more death in plants. Also the highest survival percentage (60.5%) appears in 12h duration of treatment. Generally, the best results to induce polyploidy obtained in 0.05% colchicines concentration for six hours when the treatment was treated using cotton plug.
Keywords: Ocimum basilicum, Ploidy level, Flowcytometry, Colchicines
