Search results for: callus.

Authors: K. Uteulin, S. Mukhambetzhanov, I. Rakhimbaev

Abstract:

This experiment was performed to optimize the medium for tissue culture of Taraxacum kok-saghyz Rodin. Different tissue culture approaches such as shoot regeneration from seed, callus formation from leaf explants and plant regeneration from callus were investigated in this study. All the explants were cultured on MS basal medium supplemented with 20g/l sucrose, 7g/l agar and different plant growth regulators. Seeds of Taraxacum kok-saghyzwere cultured on media containing different levels of BA and 2,4-D (0.5, 1.0 and 3.0mg/L) to direct shoot regeneration study. Leaf explants were cultured in different combination of BA (at three levels: 0.5, 1.0 and 3.0mg/L) and zeatin (at two levels: 0.5 and 1.0mg/L) to examine callus formation. After the callus formation the formed calli were cultured on different combinations of BA and NAA for shoot regeneration. BA at three levels (0.5 and 1.0 and 3.0mg/L) and NAA at two levels (0.5 and 1.0mg/L) in all possible combinations were used for shoot regeneration from callus. The results showed that the treatment containing 1.0mg/L 2,4-D in combination with 1.0mg/L BA was found to be the best one for shoot regeneration from seeds. The treatment with 1.0mg/L BA in combination with 1.0mg/L zeatin were found to be suitable treatments for callus production from leaf explants, as well. Moreover, 0.5mg/L BA alone or in combination with 1.0mg/L NAA were found to be the best treatments for shoot regeneration from callus.

Keywords: Taraxacum kok-saghyz Rodin., shoot regeneration, callus.

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Authors: Masoume Amirkhani, Kambiz Mashayekhi, Maurizio Lambardi

Abstract:

Agropyron cristatum L. Gaertn. is a native grass of semiarid region in Iran which is quit resistant to cool and drought climate and withstand heavy grazing. This species has close phylogenetic relationship with Triticum and Hordeum. In this research, the effect of seven different concentrations of growth regulator 2,4-D on callus production and somatic embryogenesis of A. cristatum was investigated on Murashige and Skoog medium. The results showed that the rate of callus, embryo and neomorph were highest in 1 mg L-1 2,4-D. Callus production was increased in 1 mg L-1 2,4-D but dramatically decreased at 5.5 and 9 mg L-1 2,4-D. The somatic embryos were observed at 1 and 4 mg L-1 2,4-D but matured embryos and plantlet were only occurred at 1 mg L-1 2,4-D. There were significant differences between 1 mg L-1 2,4-D and other treatments for producing globular and torpedo embryos, plantlet, rooted callus and number of roots (p<0.05) and there was not any callus production and embryogenesis in control treatment without growth regulator.

Keywords: 2, 4-D, callus production, somatic embryogenesis, Agropyron cristatum.

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Authors: H.K. Luk, Y.M. Lai, L. Qin, C.W. Chan, Z. Liu, Y.P. Huang, Y.P. Zheng

Abstract:

Ultrasound is useful in demonstrating bone mineral density of regenerating osseous tissue as well as structural alterations. A proposed ultrasound method, which included ultrasonography and acoustic parameters measurement, was employed to evaluate its efficacy in monitoring the bone callus changes in a rabbit tibial distraction osteogenesis (DO) model. The findings demonstrated that ultrasonographic images depicted characteristic changes of the bone callus, typical of histology findings, during the distraction phase. Follow-up acoustic parameters measurement of the bone callus, including speed of sound, reflection and attenuation, showed significant linear changes over time during the distraction phase. The acoustic parameters obtained during the distraction phase also showed moderate to strong correlation with consolidated bone callus density and micro-architecture measured by micro-computed tomography at the end of the consolidation phase. The results support the preferred use of ultrasound imaging in the early monitoring of bone callus changes during DO treatment.

Keywords: Bone Callus Growth, Rabbit Tibial DistractionOsteogenesis, Ultrasonography, Ultrasonometry

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Authors: Anuar, N., Markom, M., Khairedin, S., Johari, N. A.

Abstract:

Quercetin and (+)-catechin are metabolites present in Phyllanthus niruri plant, have potential in medicinal uses as anticancer and antioxidant agents. Studies on production of quercetin and (+)-catechin from P. niruri callus culture via in vitro technique were carried out and the results were compared to the intact plant. P. niruri explants were cultured on Murashige and Skoog (MS) solidified media supplemented with several phytohormone combinations for one month. The metabolites were extracted from P. niruri callus and intact plant by using carbon dioxide supercritical fluid extraction (SFE) with ethanol as modifier and solvent extraction techniques. The extracts were analyzed by means of HPLC method. Results showed that P. niruri callus culture was successfully established. The highest content of quercetin (1.72%) was found from P. niruri callus grown in media supplemented with 0.8mg/L kinetin and 0.2mg/L 2,4-dicholophenoxyacetic acid (2,4-D), which was 1.2 fold higher than intact plant. Meanwhile, the highest amounts of (+)-catechin (0.63%) was found from P. niruri callus grown in media with addition of 0.2mg/L 1-naphthalene acetic acid (NAA) and 0.8mg/L 2,4-D. The SFE condition in this study showed better extraction efficiency when higher contents of selected metabolites were found in all SFE extracts compared to the common solvent extracts.

Keywords: Callus culture, Phyllanthus niruri, secondary metabolite, supercritical fluid extraction.

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Authors: Münüre Tanur Erkoyuncu, Mustafa Yorgancılar

Abstract:

Crop improvement through genetic engineering depends on effective and reproducible plant regeneration systems. Immature embryos are the most widely used explant source for in vitro regeneration in barley (Hordeum vulgare L.). However, immature embryos require the continuous growth of donor plants and the suitable stage for their culture is also certainly limited. On the other hand, mature embryos can be procured and stored easily; they can be studied throughout the year. In this study, an effective callus induction and plant regeneration were aimed to develop from mature embryos of different barley genotypes. The effect of medium (MS₁ and MS₂), auxin type (2,4-D, dicamba, picloram and 2,4,5-T) and concentrations (2, 4, 6 mg/l) on callus formation and effect of cytokinin type (TDZ, BAP) and concentrations (0.2, 0.5, 1.0 mg/l) on green plant regeneration were evaluated in mature embryo culture of barley. Callus and shoot formation was successful for all genotypes. By depending on genotype, MS₁ is the best medium, 4 mg/l dicamba is the best growth regulator in the callus induction and MS₁ is the best medium, 1 mg/l BAP is the best growth regulator in the shoot formation were determined.

Keywords: Barley, callus, embryo culture, mature embryo.

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Authors: Iriawati, R. R. Esyanti, W. Natalia, N. Zahya

Abstract:

In vitro plant regeneration has been successfully obtained from basal shoot explant of Vetiveria zizanioides through indirect organogenesis. The explant was cultured in Murashige & Skoog’s (MS) media supplemented with 2,4-D, IAA, and kinetin in various concentrations. Callus was well induced in media supplemented with 2 ppm 2,4-D, 1 ppm IAA, and 1 ppm kinetin. This callus was then transferred to MS media supplemented with 1 - 5 ppm of BAP for shoot regeneration. The media supplemented with 3 ppm BAP was a suitable medium for shoot induction, as well as for shoot multiplication. Rooting was well developed in shoot following transferred to half MS media containing 0.2 ppm IBA. Plantlet was then transferred to husk charcoal for acclimatization, and almost all (90%) of plantlets were survived during acclimatization.

Keywords: Callus, plantlet regeneration, shoot induction, Vetiveria zizanioides.

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Authors: A. B. A. Ahmed, Amar, D. I., R. M. Taha

Abstract:

This research aims to investigate callus induction, somatic embryogenesis and indirect plant regeneration of Crassula ovata (Mill.) Druce – the famous ornamental plant. Experiment no.1: Callus induction was obtained from leaf and stem explants on Murashige and Skoog (MS) medium supplemented with various plant growth regulators (PGRs). Effects of different PGRs, plant regeneration and subsequent plantlet conversion were also assessed. Indirect plant regeneration was achieved from the callus of stem explants by the addition of 1.5 mg/L Kinetin (KN) alone. Best shoot induction was achieved (6.5 shoots/per explant) after 60 days. For successful rooting, regenerated plantlets were sub-cultured on the same MS media supplemented with 1.5 mg/L KN alone. The rooted plantlets were acclimatized and the survival rate was 90%. Experiment no.2: Results revealed that 0.5 mg/L 2,4-D alone and in combination with 1.0 mg/L 6-Benzyladenine (BA) gave 89.8% callus from the stem explants as compared to leaf explants. Callus proliferation and somatic embryo formation were also evaluated by ‘Double Staining Method’ and different stages of somatic embryogenesis were revealed by scanning electron microscope. Full Strength MS medium produced the highest number (49.6%) of cotyledonary stage somatic embryos (SEs). Mature cotyledonary stage SEs developed into plantlets after 12 weeks of culture. Wellrooted plantlets were successfully acclimatized at the survival rate of 85%. Indirectly regenerated plants did not show any detectable variation in morphological and growth characteristics when compared with the donor plant.

Keywords: Callus induction, Crassula ovata, Double Staining, Indirect plant regeneration, Somatic embryogenesis.

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Authors: Pratibha Gupta, Satyawati Sharma, Sanjay Saxena

Abstract:

Stevia rebaudiana Bertoni (natural sweetener) belongs to Asteraceae family and can be used as substitute of artificial sweeteners for diabetic patients. Conventionally, it is cultivated by seeds or stem cutting, but seed viability rate is poor. A protocol for callus induction and multiplication was developed to produce large no. of calli in short period. Surface sterilized nodal, leaf and root explants were cultured on Murashige and Skoog (MS) medium with different concentrations of plant hormone like, IBA, kinetin, NAA, 2,4-D, and NAA in combination with 2,4-D. 100% callusing was observed from leaf explants cultured on combination of NAA and 2,4-D after three weeks while with 2,4-D, only 10% callusing was observed. Calli obtained from leaf and root explants were shiny green while with nodal explants it was hard and brown. The present findings deal with induction of callusing in Stevia to achieve the rapid callus multiplication for study of steviol glycosides in callus culture.

Keywords: 2, 4-D, Callusing, NAA, Stevia, Steviol glycosides

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Authors: Naira Sahakyan, Margarit Petrosyan, Armen Trchounian

Abstract:

One of the tasks in contemporary biotechnology, pharmacology and other fields of human activities is to obtain biologically active substances from plants. They are very essential in the treatment of many diseases due to their actually high therapeutic value without visible side effects. However, sometimes the possibility of obtaining the metabolites is limited due to the reduction of wild-growing plants. That is why the plant cell cultures are of great interest as alternative sources of biologically active substances. Besides, during the monitored cultivation, it is possible to obtain substances that are not synthesized by plants in nature. Isolated culture of Ajuga genevensis with high growth activity and ability of regeneration was obtained using MS nutrient medium. The agar-diffusion method showed that aqueous extracts of callus culture revealed high antimicrobial activity towards various gram-positive (Bacillus subtilis A1WT; B. mesentericus WDCM 1873; Staphylococcus aureus WDCM 5233; Staph. citreus WT) and gram-negative (Escherichia coli WKPM M-17; Salmonella typhimurium TA 100) microorganisms. The broth dilution method revealed that the minimal and half maximal inhibitory concentration values against E. coli corresponded to the 70 μg/mL and 140 μg/mL concentration of the extract respectively. According to the photochemiluminescent analysis, callus tissue extracts of leaf and root origin showed higher antioxidant activity than the same quantity of A. genevensis intact plant extract. A. genevensis intact plant and callus culture extracts showed no cytotoxic effect on K-562 suspension cell line of human chronic myeloid leukemia. The GC-MS analysis showed deep differences between the qualitative and quantitative composition of callus culture and intact plant extracts. Hexacosane (11.17%); n-hexadecanoic acid (9.33%); and 2-methoxy-4-vinylphenol (4.28%) were the main components of intact plant extracts. 10-Methylnonadecane (57.0%); methoxyacetic acid, 2-tetradecyl ester (17.75%) and 1-Bromopentadecane (14.55%) were the main components of A. genevensis callus culture extracts. Obtained data indicate that callus culture of A. genevensis can be used as an alternative source of biologically active substances.

Keywords: Ajuga genevensis, antibacterial activity, antioxidant activity, callus cultures.

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Authors: A. Pourjabar, S.A. Mohammadi, R. Ghahramanzadeh, Gh. Salimi

Abstract:

The extract of milk thistle contains a mix of flavonolignans termed silymarine.. In order to analysis influence of growth regulators, genotype, explant and subculture on the accumulation of flavonolignans, a study was carried out by using two genotype (Budakalszi and Noor abad moghan cultivars), cotyledon and hypocotyle explants, solid media of MS supplemented by different combinations of two growth regulators; Kinetin (0.1, 1 mg/l) and 2,4-D (1, 2 mg/l). Seeds of the plant were germinated in MS media whitout growth regulators in growth chamber at 26°C and darkness condition. In order to callus induction, the culture media was supplemented whit different concentrations of 2,4-D and kinetin. Calli obtained from explants were sub-cultured four times into the fresh media of the first experiment. flavonoides was extracted from calli in four subcultures. The flavonoid components were determined by high- performance liquid choromatography (HPLC) and separated into Taxifolin, Silydianin+Silychristin, Silybin A+B and Isosilybin A+B. Results showed that with increasing callus age, increased accumulation of silybin A+B, but reduced Isosilybin A+B content. Highest accumulation of Taxifolin was observed at first calli. Calli produced from cotyledon explant of Budakalszi cultivar were superior for Silybin A+B, where calli from hypocotyl explant produced higher amount of Taxifolin and Silydianin+Silychristin. The best cultivar for Silymarin production in this study was Budakalszi cultivar. High amount of SBN A+B and TXF were obtained from hypocotil explant.

Keywords: Callus culture, Flavonolignans, Silimarine

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Authors: Shyama Ranjani Weerakoon

Abstract:

Direct and indirect somatic embryogenesis (SE) from petiole and leaf explants of Scaevola aemula R. Br. cv. 'Purple Fanfare' was achieved. High frequency of somatic embryos was obtained directly from petiole and leaf explants using an inductive plant growth regulator signal thidiazuron (TDZ). Petiole explants were more responsive to SE than leaves. Plants derived from somatic embryos of petiole explants germinated more readily into plants. SE occurred more efficiently in half-strength Murashige and Skoog (MS) medium than in full-strength MS medium. Non-embryogenic callus induced by 2, 4-dichlorophenoxyacetic acid was used to investigate the feasibility of obtaining SE with TDZ as a secondary inductive plant growth regulator (PGR) signal. Non-embryogenic callus of S. aemula was able to convert into an “embryogenic competent mode" with PGR signal. Protocol developed for induction of direct and indirect somatic embryogenesis in S. aemula can improve the large scale propagation system of the plant in future.

Keywords: Petiole and leaf explants, Scaevola aemula, Somaticembryogenesis

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Authors: R. Razavizadeh, F. Adabavazeh, M. Rezaee Chermahini

Abstract:

Salinity is one of the most widespread agricultural problems in arid and semi-arid areas that limits the plant growth and crop productivity. In this study, the salt stress effects on protein, reducing sugar, proline contents and antioxidant enzymes activities of Carum copticum L. under in vitro conditions were studied. Seeds of C. copticum were cultured in Murashige and Skoog (MS) medium containing 0, 25, 50, 100 and 150 mM NaCl and calli were cultured in MS medium containing 1 μM 2, 4-dichlorophenoxyacetic acid, 4 μM benzyl amino purine and different levels of NaCl (0, 25, 50, 100 and 150 mM). After NaCl treatment for 28 days, the proline and reducing sugar contents of shoots, roots and calli increased significantly in relation to the severity of the salt stress. The highest amount of proline and carbohydrate were observed at 150 and 100 mM NaCl, respectively. The reducing sugar accumulation in shoots was the highest as compared to roots, whereas, proline contents did not show any significant difference in roots and shoots under salt stress. The results showed significant reduction of protein contents in seedlings and calli. Based on these results, proteins extracted from the shoots, roots and calli of C. copticum treated with 150 mM NaCl showed the lowest contents. The positive relationships were observed between activity of antioxidant enzymes and the increase in stress levels. Catalase, ascorbate peroxidase and superoxide dismutase activity increased significantly under salt concentrations in comparison to the control. These results suggest that the accumulation of proline and sugars, and activation of antioxidant enzymes play adaptive roles in the adaptation of seedlings and callus of C. copticum to saline conditions.

Keywords: Antioxidant enzymes, Carum copticum, organic solutes, salt stress.

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