Cell and Tissue Journal

Abstract Introduction: The anther culture technique is used in vitro for many plant species as an effective tool for obtaining haploid and doubled-haploid lines.
Aims: This study aimed to investigate the influence of different concentrations of iron oxide nanoparticles and benzylaminopurine (BAP) on callus formation, embryogenesis, regeneration, and rooting in bell pepper anther culture
Materials and Methods: The experiment was conducted as a factorial experiment in a completely randomized design under in vitro culture conditions. Flower buds of appropriate size (equal sepal to petal ratio or slightly longer petal) were collected from the greenhouse, and acetocarmine solution was used to determine the growth and development stage of microspores. The results showed that the most suitable stages for embryogenesis induction were the late mononuclear and early binuclear stages. In order to sterilize the flower buds, 70% ethanol for 30 seconds and 5% sodium hypochlorite for 20 minutes were used, and after each stage, they were washed three times with sterile distilled water. Then, the anthers were separated from the flower bud and placed in C medium containing 2 mg/L naphthalene acetic acid (NAA), different concentrations of BAP (0, 0.1, 0.5, and 1 mg/L), and different concentrations of iron oxide nanoparticles (0, 1, 10, and 20 mg/L). After that, the explants cultured in C medium were kept at 35°C in a dark place for 8 days in order to apply heat treatment. Then, they were transferred to 25°C in the light for 4 days. After this period, in order to induce embryogenesis, the explants were transferred from C medium to R medium and were subcultured every three weeks until embryos emerged. For further growth and root development, the embryos were transferred to V medium.
Results: The analysis of variance showed that different concentrations of iron oxide nanoparticles had a significant effect on the percentages of embryogenesis, regeneration, and rooting, but had no significant effect on the percentage of callus formation. The results of the mean comparison showed that among the different concentrations of iron oxide nanoparticles, the 1 mg/L treatment produced the highest embryogenesis percentage (11.11%). Furthermore, the mean comparison results for regeneration indicated that the 1 mg/L treatment resulted in the highest regeneration percentage (16.66%). The results of the interaction effects showed that among the different concentrations of iron oxide nanoparticles and BAP, the highest percentage of embryogenesis was observed in the treatment of 20 mg/L iron oxide nanoparticles and 0 mg/L BAP. Also, the treatment of 20 mg/L iron oxide nanoparticles and 0 mg/L BAP had the highest percentage of regeneration (33.33%). After sufficient growth and root formation, the obtained plants were removed from the glass culture containers and transferred to pots containing sterilized culture medium and watered for adaptation. The tops of the pots were covered with plastic cups, and after three days, the cups were pierced, and the plastic was gradually removed from the plant for further adaptation. Ploidy levels were determined by chromosome counting after staining the root tip cells. The results showed that out of the 23 obtained plants, 21 were diploid and had 2n=2x=24 chromosomes, and 2 were haploid and had n=x=12 chromosomes.
Discussion: Applying different concentrations of iron oxide nanoparticles in medium C had a positive effect on the regeneration of bell pepper plants. Iron is a vital micronutrient for several key cellular processes in plants. In addition, iron is essential for ensuring the structural integrity of proteins. Iron nanoparticles positively affect plant growth in tissue culture by increasing morphological parameters. The beneficial effects of nanoparticles on plants are related to their high solubility and reactivity, which affect how they interact with membranes and other cellular components, due to their large specific surface area. The treatment of 20 mg/L of nanoparticles and 0 mg/L of BAP had the highest regeneration percentage. The culture medium containing nano-iron leads to the plant benefiting from the element iron directly at all different stages of growth and its participation with other nutrients, and therefore the production of many nutrients containing it leads to the transfer of their excess to different parts of the plant. The presence of nano iron in the culture medium has led to a nutritional balance in the plant and, as a result, has improved the plant growth process by increasing the amount of nutrients and elements accumulated in the microsample. Cytokinins act as a nutrient reservoir, which leads to an increase in the percentage of elements in the microsample and an increase in the percentage of sugars in the culture medium, which is important in the structural processes in tissues and affects the accumulation of sugars in it, and is reflected in increased vegetative growth.
Conclusion: In bell pepper anther culture, different concentrations of iron oxide nanoparticles, along with plant growth regulators at different concentrations, showed a great effect on embryogenesis, regeneration, and rooting.

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Abstract

Abstract Aim: The aims of study were to investigate callogenesis and regeneration of the studied cultivars and also exploring the possibility of gene transfer into maize by using complete bud and stem apical meristem explants.
Material and methods: Complete bud and stem apical meristem of six cultivars were used as explants on MS medium + 5 mg.L-¹ 2,4-D. For regeneration, calluses were cultured in MS including 1 mg.L-¹ Kn and 10 mg.L-¹ BAP. According to the results of tissue culture, SC703 and SC704 cultivars were selected for gene transfer. Transformation of explants was done with LBA4404 strain of Agrobacterium tumefaciens containing pBI121 vector. To investigate the transgenic nature of the samples, PCR and histochemical assay were used.
Results: Callus induction was taking place after 30 to 40 days and the frequency of callus induction in six cultivars was 100%, in addition the highest rate of regeneration (90 %) was observed in stem apical meristem explants of SC704. In some cultivars, stem apical meristem explants were more suitable, although in other cultivars, complete bud explants were better. PCR results for NPTII gene and GUS histochemical assay showed that some calli are transgenic and GUS gene is expressed.
Conclusion: The results of callogenesis, regeneration and genetic transformation showed that SC 704 was better than the others.
 

Abstract Aim: In this study, we explored the effect of epigenetic modifications on planarian regeneration process by treating with small molecule epigenetic modifiers
Material and Methods: Planaria worms were cut into 3 sections including head, trunk, and tail regions and treated with 6 small molecules with epigenetic modification effects. All samples were evaluated morphologically for 20 days to monitor the effects on regeneration process.
Results: Morphological analysis of samples treated with selected six small molecules with different epigenetic modification functions revealed that treating with two of them including Sodium butyrate and Valporic acid will result in delayed head, trunk, and tail fragments regeneration, and delayed eye formation in trunk and tail fragments, respectively. In addition, samples treated with other four small molecules including BIX01294, RG108, SAHA, and Tranylcypromine with other epigenetic modification functions, regenerated normally.
Conclusion: These results indicating that different epigenetic levels and activity of histone deacetylases play a key role in planarian regeneration and inhibition of histone deacetylases activity will result in abnormal regeneration. However, further work is needed to determine the exact mechanism of histone deacetylases activity effect on planarian regeneration process. Finally, we hope to understand more about the underlying mechanism of planarian and vertebrate’s regeneration inhibition and activation using these findings.

Abstract Aim: Plant production from tissue culture is of high importance from genetic engineering point of view, so in this study, optimization of tissue culture conditions for callus production and plant regeneration of Salsola arbuscula were examined. Material and Methods: Seeds were planted MS medium and after two months, roots, stems (internodes) and leaves explants were transferred to the MS medium with different concentration of 2,4-D and Kinetin hormones for production callus. Plant regeneration was also examined on the various media. Results: Results showed that the best medium to induce callus production were medium of MS + 2,4-D (1 mg/L) + Kin(1 mg/L) and the best explants were roots. So, direct shoot regeneration produced in the medium MS + Kin(1 mg/l) and MS + Kin (0.5 mg/L). Conclusion: Despite production of callus on medium containing only 2,4-D, combined two hormones, auxin and cytokinin, increased the rate of callus production. Also direct shoot regeneration has occurred in medium without auxin. So, production rate of callus and plant regeneration are dependent on amount of external plant growth regulators, and also, amount of external plant growth regulators significantly dependent to genotype and amount of internal plant hormones.

Abstract Aim: The Pelargonium roseum L. is an ornamental plant with important medical properties. The purpose of this study was to optimized the proliferation and regeneration of Pelargonium roseum L. and investigation of the possible somatic variation in the regenerated plant.
Material and methods: In this study first explants from the terminal buds as well as some small part of the stem were prepared and cultured in MS basal medium. Then some fragments of the cultured explants including parts of stems and leaves were isolated and transferred to mediums containing different growth regulators to stimulate micro-regeneration of the plants. From regenerated plants several lines were selected and their morphological characters as well as genetic diversity were evaluated.
Results: The rate of shoots and roots production was maximum in mediums containing leaf explants supplemented with 2mg/l BAP and 0.1 mg/l NAA. Somatic variation of the regenerated lines in comparison with control group showed differences in leaf shape and number, in addition genetic differences were expressed in form of band addition and band removal in bonding pattern of DNA.
Conclusion: The different percentage of shoots and roots production was an indication of the inductive effect of mediums and explants variation on regeneration of the Pelargonium roseum L. Comparison of genetic variation and morphological diversity of regenerated plants could show the common base of genetic for somatic variation.
 
 

Abstract < p >Aim: The aim of this research is evaluation of auxin changes in plant regenerated from tobacco root with Ri-TDNA Material and methods: In this research leaf segments of tobacco were transformed by Agrobacterium rhizogenes. Transformed hairy root was selected by their ability to grow on medium containing Kanamycin. For confirmation of transformation x-gluc substrate was used. From transformed roots at first callus and then plant was regenerated. Finally auxin content from roots and leaves of regenerated plants were measured. Results: Blue color of roots confirmed the successful transformation of samples. Results showed that auxin content in transgenic plants compare to control was increased 80-90 %. Transgenic plants showed shorted internodes and smaller leaf area than non transgenic plants. Conclusion: Regenerated plants from transgenic roots showed higher level of auxin content than non transgenic plants. Change of auxin content and its interaction with gibberellic acid possibly resulted in shorter length of transgenic plants.