Atefeh Ghoochani Khorasani; Iman Roohollahi; Porandokht Golkar
Abstract
Introduction: Zamifolia, Zamioculcas zamiifolia, belongs to the family Araceae, In vitro propagation is suggested for increase the propagation rate and disease-free production of explants because of vegetative propagation of Zamifolia. Micropropagation is one of the commercial aspects of indoor cultivation ...
Read More
Introduction: Zamifolia, Zamioculcas zamiifolia, belongs to the family Araceae, In vitro propagation is suggested for increase the propagation rate and disease-free production of explants because of vegetative propagation of Zamifolia. Micropropagation is one of the commercial aspects of indoor cultivation and has many advantages over conventional methods of vegetative propagation. Nitrogen is an important component in the structure of molecules and metabolic compounds in plant cells. It is also a major constituent of amino acids in the structure of proteins. Therefore, nitrogen source and its type are important factors in the application of nutrient solutions, controlling the amount of nitrogen, which is divided into two factors: concentration and type of nitrogen source. In this study, the effects of 2.4, D as plant growth regulator, different concentrations of total nitrogen and nitrate to ammonium ratios on growth of Zamifolia were investigated in separate experiments. Materials and Methods: According to the results of previous studies, the best explants for achieving full callus and seedling are petiole and main vein leaf. The leaves were rinsed with running water for 20 minutes. The leaves were then immersed in 3.5% sodium hypochlorite under laminar hood for 20 minutes. Next, they were rinsed with sterile distilled water three times and then immersed in 70% alcohol for 60 seconds. Finally, the samples were extracted from alcohol and cultured in a suitable culture medium vertically. In the first experiment, callus production from whole leaf of Zamifolia was studied in MS and 1/2 MS medium with two concentrations of 2,4-D )9.05 and 18.1 M(. In the second experiment, the effect of two concentrations of total nitrogen at 60 and 30 μM and two nitrate to ammonium ratio (1: 3 and 1: 1 ) in presence of naphthalene acetic acid (2.69 μM ) and benzyl amino purine (2.22 μM) were studied on whole plantlet production in the shortest growth period. After 16 weeks of continuous culture in the subculture medium, entire explants with tubers, roots and 1-2 expanded leaves were transferred to ex vitro condition, in peat, peat: perlite (1:1) and sand and then on a shaded greenhouse bench. Statistical analysis of the results was factorial based on completely randomized design with 20 replications, and plantlet production acclimatization test, factorial experiment based on completely randomized design with four replications, were performed and analyzed with SAS software version 9.3 Results and Discussion: The results of the first experiment showed that the largest calli were produced in MS 1.2 and 2,4-D 9.5 μM. After 16 weeks, largest embryo-like size (4.5 cm2) can be seen in the same treatment under nitrogen concentration (30 μM) and nitrate to ammonium ratio (1: 3). On the other hand, the smallest embryo-like structure size (3.5 cm2) is related to interaction of nitrogen concentration (30 μM) and nitrate to ammonium ratio (1: 1). Overall, the results of the first part of this study showed the best callus induction and growth of Zamifolia callus under culture conditions under modified M.S. 1.2 and 2, 4-D 9.5 μM. Secondly, culture medium NAA 2.69 μM + BAP 2.22 μM and modified M.S. media and nitrogen concentration of 30 μM and nitrate to ammonium ratio (1: 3). The best medium for embryo like structure growth in the early regeneration and seedling production weeks, increased leaf number in the last weeks compared to other treatments. According to this protocol, small number of leaves can be propagated to over a period of about 16 weeks to a large number of Zamifolia plants, which is comparable to other methods of reproduction of this plant, and similar studies that it has been economically justified to do this for about 30 weeks. Conclusion: Based on our results, we can conclude Zamifolia tissue culture can be an effective way to improve it growth conditions which can get acclimatized in ex vitro conditions on peat medium.
Seyyed Mehran Alavi; Asad Masoumiasl; Naser Zare; Rasul Asghari Zakaria; Parisa Sheikhzade Mosaddegh
Abstract
Introduction: The main habitat of Chavil, Ferulago angulata, in Iran is Zagros area. This plant has a rejuvenating effect and is used to treat digestive diseases and intestinal worms. Because the different explants show different amounts of callogenesis under the effect of different growth regulators, ...
Read More
Introduction: The main habitat of Chavil, Ferulago angulata, in Iran is Zagros area. This plant has a rejuvenating effect and is used to treat digestive diseases and intestinal worms. Because the different explants show different amounts of callogenesis under the effect of different growth regulators, selection of an optimal explant and suitable plant growth regulators combination has a significant effect on the production of callus and their suspension culture. There is no reports on Ferulago angulata callogenesis and its cell suspension culture. Therefore, this study was designed and implemented to optimize callus production and cell suspension culture in this important medicinal plant.
Materials and Methods: Seeds of Chavil were collected from four different habitats in Kohgilooyeh and BoyerAhmad Province in Southwest of Iran include Abenahr, Guayoune, Vezg and Sisakhat. Seedlings obtained from embryo culture were used to prepare the explants. Various explants (leaf, root and stem) were cultured on MS medium supplemented with different concentrations (0, 0.5, 1 and 2 mgl-1) of NAA and BAP. Callus traits were evaluated and from the best culture medium, the best explants and the best PGRs composition for callogenesis of each ecotype were used to cell suspension culture. In order to study the growth rate of cells in suspension culture and plotting the curve of cells growth, two cell density indices and packed cell volume index were evaluated. To determine the cell density index, every 3 days, 10 ml of cell suspension were transferred to the graded falcon and centrifuged at 5000 g for 5 minutes, and the percentage of sediment cells was calculated as the total volume. To determine the packed cell volume index, also every 3 days, 10 ml of culture medium containing cells were transferred to the graded falcon and stored for 30 minutes to precipitate cells and cell masses. Finally, the cell volume was recorded and was calculated as percentage of the total collected medium.
Results and Discussion: According to the callogenesis percentage, the best ecotype is Abenahr and best explant is leaf explant. The highest level of NAA is 2 mgl-1, and the best level of BAP is 2 mgl-1, which causes 100 callogenesis percentage. The best medium for cell suspension culture is MS medium containing 2 mgl-1 NAA and 0.5 mgl-1 BAP for callus was obtained from leaf explant of Abenahr ecotype. Along with these plant growth regulators, 2,4-D was used in combination with BAP to form suspension culture. The results also showed that 2 mgl-1 2,4-D plus 0.5 mgl-1 BAP were useful in producing suspensions. The difference between 2,4-D +BAP and NAA + BAP combinations more cell volumes were observed, and cell suspension was created at a faster rate and in less time, which is an advantage in research work. Growth rate of cell suspension originated from the leaf explant was higher than root explant. In terms of culturing cell suspension, the Abenahr ecotype was favorable compared to other ecotypes. During cell suppression culture of Cyperus aromaticus by applying different levels of NAA, cell growth was increased up to 3 weeks after application, and then decreased. By applying 2,4-D, cell growth also increased until the third week, and after the third week, cell growth declined, which was very low growth rate compared with the NAA. In cell suspension culture of sugar beet, using 2,4-D was much more effective than NAA on all explants. In the present study, 2,4-D was also more effective than NAA for cell suspension culture of Chavil.
Conclusion: In general, the Abenahr was the best ecotype among of investigated. The explants in both callus culture and the suspension culture, and the best combination of plant growth regulator in both culture was 2 mgl-1 NAA plus 0.5 mgl-1 BAP.
Esmaeil Chamani; Zahra Eftekhari; Alireza Ghanbari; Hamid Reza Heydari; Mousa Arshad
Abstract
Introduction: Fritillaria imperialis L. is an ornamental and medicinal plant native to mountainous regions of Iran. This plant genetic resource is in danger of extinction, Because of grazing livestock and pest outbreaks. Therefore, micro propagation of Fritillaria through in vitro regeneration is essential ...
Read More
Introduction: Fritillaria imperialis L. is an ornamental and medicinal plant native to mountainous regions of Iran. This plant genetic resource is in danger of extinction, Because of grazing livestock and pest outbreaks. Therefore, micro propagation of Fritillaria through in vitro regeneration is essential for conservation and commercial production. Thymol and Carvacrol are one of the main essential oil compounds in family Lamiaceae.
Material and Methods: Fritillariaimperialis L. bulbs in dormancy stage obtained from mountainous regions of Lorestan in Iran and were placed in cold room at +4 °C for 4-6 weeks. Then, Bulbs were surface-sterilized with 70% ethanol for 45 s followed by immersion in 5% (v/v) NaOCl solution for 20 min with gentle agitation, and then rinsed three times in sterile double distilled water. Present study was conducted in two separate experiments. In first experiment, effect of different concentration of Thymol and Carvacrol and in second experiment, different concentration of NAA and BA on in vitro characteristics of Fritillaria was evaluated. Explants (1× 1 cm) prepared from the lower third of scales with basal plate and were placed in MS basal medium supplemented with different concentrations of Thymol (50, 100, 150 and 300 ppm), Carvacrol (10, 100, 500 and 100 ppm), BA (1, 2 and 4 mg/l) and NAA (1, 2 and 4 mg/l).All cultures were incubated in a growth chamber at 24±2°C, and a photosynthetic photon flux of 40-60 μmol m–2 s–1 was provided by cool white fluorescent lamps with a 16-h photoperiod. This experiment wascarried out in completely randomized designs with fivereplications.
Results and Discussion: Analysis of variance showed that Thymol and Carvacrol were not effective on number of new bulblets but had significant effects on bulb diameter, number and length or roots, number and length leaves and callus induction and diameter of callus obtained from scales (P< 0.05). The highest rate (3 bulblets) of bulblets formation was obtained fromMS medium supplemented with 50 ppm Thymol that showed significantly difference from other treatments. Medium containing 10 ppm Carvacrol gave the highest Bulblet formation (2.5 bulblets) between Carvacrol treatments. Investigation of rooting was done by assessment of the number and length of roots. Mean comparison of the effect of cultivar type on root number showed that the largest number of roots per explant was obtained fromMS medium containing 50 ppm Thymol. Lowest number of roots observed in mediums supplemented with 300 ppm Thymol and 100 ppm Carvacrol. The best medium for increasing the root length per explant (10.90 cm) was MS medium supplemented with 100 ppm Carvacrol, while the least increasing in root length per explant observed from culture mediums contained 300 ppm Thymol and 100 ppm Carvacrol. Also, the largest number of leave formation obtained from culture medium supplemented with 50 ppm Thymol that significantly higher than other treatments. Statistical analysis (ANOVA) of the data showed that high frequency callus induction and formation occurred in MS mediums contained 50, 100 and 150 ppm Thymol and 10 ppm Carvacrol and culture mediums supplemented with 300 ppm Thymol and 1000 ppm Carvacrol showed least callus induction. In contrast, largest callus diameter observed in culture mediums supplemented with 300 ppm Thymol and 500, 100 ppm Carvacrol.
Statistical analysis of results showed that different concentrations of BA and NAA had significant effects on bulblets number and bulblets diameter (P
Ahmad Noroozi; Abdolreza Bagheri; Nasrin Moshtaghi; Ahmad Sharifi
Abstract
Introduction: Anthurium is a popular genus of the Araceae (order Spathiflorae).The flower consists of a protruding spadix containing numerous florets, subtended by a brightly colored modified leaf, the spathe. Anthuriums are bisexual and protogynous.Anthuriumscherzerianum as the most important species ...
Read More
Introduction: Anthurium is a popular genus of the Araceae (order Spathiflorae).The flower consists of a protruding spadix containing numerous florets, subtended by a brightly colored modified leaf, the spathe. Anthuriums are bisexual and protogynous.Anthuriumscherzerianum as the most important species ofAnthurium genus is a potted perennial plant. Due to having beautiful, attractive and long-life flowers, A. scherzerianum can be used for the production of pot and cut flowers. Tissue culture is suggested as the most commonly method in order to rapid propagation and removing disease in a short period of time. This method also recommended for Anthuriumbecause of problems in classical propagation method of this flower..The three basic propagation methods for Anthuriumare propagation by seed, traditional vegetative and tissue culture.Micropropagation of Anthurium is using forcommercial production.
Materials and Methods: In this study, the effect of plant growth regulators and explants on indirect regeneration of A. scherzerianumdetermined in separate experiments. In the first experiment, callogenesis was done by leaf explants on MS medium containing growth regulators, BA in three concentrations (0.5, 1.25 and 2 mg/l) in combination with
2, 4-D (0.5, 1.25 and 2 mg/l) or NAA (0.5, 1.25 and 2 mg/l) and the combinations of TDZ (0.5, 1.25 and 2 mg/l) with
2, 4-D (0, 0.5 mg/l). In the second experiment, regeneration was done on MS medium containing 0.75 mg/l BA with 0.05 mg/l 2, 4-D and 0.1 mg/l NAA and also in combination with TDZ (0.75mg/l). For rooting, MS medium containing different concentrations of IBA and IAA (0, 0.2 and 1 mg/l) were used. Callus induction, regeneration and rooting experiments were done based on completely randomized design, with 12, 6 and 6 replications, respectively.Data from all the schemes used in this study were analyzed with SAS statistical software. The comparison of means using Duncan's multiple range test was evaluated at the 5% level.
Results and Discussion: Analysis of variance showed that the effect of explant type and hormone combinations was significant on the percentage of callogenesis, callus volume and survival percentage. The interaction effect of explant type and combination of hormones was also significant on percentage of callogenesisand the volume of callus. Means comparisons showed that the highest callogenesis, viability and callus volume were achieved on MS medium containing 2 mg/l of BA and 0.5 mg/l of 2, 4-D. Petiole explants, also produced the highest percentage of callus (95%), survival rate (96%) and callus with dimensions of 6 mm2. Callus formation in leaf vein explants was higher than others. The effect of explant type and hormone combinations on regeneration, number of branches, number of leaves and leaf length was significant.The interaction of explant and hormone combinations on regeneration, number of branches, number of leaves and leaf length was also significant. Moreover, results of regeneration experiment indicated that the maximum number of shoots (6.9) and the maximum shoot length (5 cm), number of leaves (18) and the leaf length (2.8 mm) were achieved in 0.75 mg/l BA mg/l of and 0.05 mg/l 2, 4-D. In this study, petiole explants were also regenerated earlier than leaf explants.The effect of hormone combinations and concentrations was significant on rooting specially on the number of roots and root length.Furthermore, results of rooting experiment revealed that the highest rooting percentage (95%), the maximum number of roots (4.5 per plantlet) and the longest roots (3.5 cm) were produced in the medium containing 0.2 mg/l of IBA. Finally, the rooted plantlets were adapted (90%) in vivo condition by placing them on a mixture of cocopeat and perlite (2:1) substrate.
Conclusion: In this study callugensis, regeneration and rooting of A.scherzerianum’s petiole and leaf explants were studied and different levels of plant growth regulators used for callugensis and regeneration. In this study petiole explants showed the highest callugenesis and regeneration. MS medium containing BA (2 mg/l) and 2, 4-D (0.5 mg/l), was the best for callugenesis. Also the highest percentage of regeneration was observed in medium containing BA (0.75 mg/l) and 2, 4-D (0.05 mg/l). Moreover low concentration (0.2 mg/l) of auxin has a better effect on rooting than high levels (1mg/l) so that the highest rooting percentage was produced in medium containing IBA (0.2 mg/l) and the lowest rooting percentage was produced in medium containing IAA (1 mg/l). Anthurium plantlets acclimized is cocopeat and perlite substrate (2: 1) with 90% acclimation.
