The effect of 2,4-dichlorophenoxyacetic acid and 6-benzylaminopurine on callus induction and plant regeneration of allamanda cathartica - a valuable medicinal plant

Allamanda cathartica is a potential medicinal plant which starts to achieve awareness in society due to its value to treat various types of diseases, especially in the treatment of jaundice, malaria and cancer. In the present study, the effects of 2,4-dichlorophenoxyacetic acid (2,4-D) and 6-benzylaminopurine (BAP) on the callus induction from the leaf and stem explants were investigated. Plant regeneration from the nodal explants was achieved. Surface sterilization by mercuric chloride (HgCl2) was utilized in order to surface sterilize the leaf (0.1), stem and nodal (0.2) explants. The leaf and stem explants were cultured on full-strength Murashige and Skoog (MS) medium supplemented with different concentrations of 2,4-D alone (0.5 and 1.0 mg/L) or in combinations of 2,4-D (0.5, 1.0 and 1.5 mg/L) with BAP (0.5, 1.0 and 1.5 mg/L). In the study of plant regeneration, the nodal explants were cultured on MS medium supplemented with BAP at 1.0, 3.0 or 5.0 mg/L for shoot multiplication. MS basal medium was used as a control and also used for shoot elongation. All the cultures were incubated under a photoperiod of 16 hours light and 8 hours darkness. For callus induction, the leaf and stem explants cultured on 1.0 mg/L 2,4-D and 1.0 mg/L BAP gave the best callus response (100) with yellow-white, greenish friable callus (0.0707 +/- 0.0549 g with callus initiated after 6 days) and brown-white, greenish friable callus (0.0207 +/- 0.0009 g with callus initiated after 5 days), respectively. For shoot multiplication, MS supplemented with 5 mg/L BAP gave the best response (100) with multiple shoots formed (6 +/- 0.6 shoots per explant) from the nodal explants. In the plant growth regulator (PGR)-free medium, the elongated shoots were developed (1.01 +/- 0.07 cm) with white hairy roots

Production of artificial seeds derived from encapsulated in vitro micro shoots of cauliflower, Brassica oleracea var. botrytis

A high number of micro shoots (21 +/- 2.31) of Brassica oleracea var. botrytis (cauliflower) were obtained when hypocotyl explants from 2-week-old aseptic seedlings were cultured on MS medium supplemented with 0.1 mg/L NAA and 5 mg/L BAP. Artificial or synthetic seeds were formed when the micro shoots were encapsulated in 4 (w/v) sodium alginate with 100 mM CaCl2 as complexing solution. The artificial seeds took 12 days (after 7 days storage) and 14 days (after 30 days storage) to germinate on MS basal medium. The germination percentage of artificial seeds was enhanced by the inclusion of 0.3 mg/L NAA and 3.0 mg/L BAP in the encapsulation matrix after 7 and 30 days of pre-germination storage. The time taken for germination was also faster (5 days after 7 days of storage and 11 days after 30 days of storage) when MS fortified with 0.3 mg/L NAA and 3.0mg/L BAP were used. Isolated micro shoots encapsulated in MS supplemented with 0.3 mg/L NAA and 3.0mg/L BAP gave high germination percentages (70 +/- 5.43 and 63.33 +/- 4.17) after 7 and 30 days of pre-germination storage period, respectively. Complete plant regeneration was achieved from the germination of these artificial seeds derived from the micro shoots after 3-4 weeks in culture

Studies on seed and forage yield in 10 populations of sainfoin (Onobrychis saliva) grown as spaced plants and swards

In order to study seed and dry matter (DM) yield and relationships between yield and morphological traits, 10 populations of sainfoin (Onobrychis saliva), were assessed in two separate experiments under sward and spaced plant conditions using complete block design with two replications, in Karaj, Iran, during 2004-2007. The data were collected for seed yield, DM yield, plant height, stem number per m(2), flowering date, inflorescences length and 1000-grain weight. The data were analyzed for individual experiment and combined over two experiments. Results showed significant differences among populations for all of traits except 1000-grain weight in sward and DM yield and stem number in spaced plants. Results of combined analysis showed no significant differences among 2 conditions for both seed and DM yield. However, the average values of 7.3 and 6.2 ton ha(-1) DM yield and 337 and 375 kg ha(-1) seed yield were obtained for sward and spaced plants, respectively. Results showed that, populations of Shahrkord and Kashan with average values of 8.08 and 8.06 ton ha(-1) DM yield and Shahrkord with average values 551 kg ha(-1) seed yield had higher production under sward conditions. For spaced plants, Orumieh and Bijar with average values of 7.98 and 7.67 ton ha(-1) DM yield and Bijar and Esfahan with average values of 510 kg ha(-1) seed yield had higher production. Therefore, the populations of Shahrkord and Bijar were introduced as the best ones for both DM and seed production under swards and spaced plants, respectively. DM yield was positively correlated with seed yield, plant height, stem number and inflorescences length, while seed yield had positive and significant relationships with 1000-grain weight, plant height, stem number and inflorescences length. The relationship between spaced plants and swards as measured by phenotypic correlation was poor for all traits except 1000-grain weight. This suggests that response to selection based on spaced plant data would not be reflected in swards, so, for all characters selection should be based on sward yield

Effects of benzylaminopurine and naphthalene acetic acid on proliferation and shoot growth of pineapple (Ananas comosus L. Merr) in vitro

This study was conducted to evaluate the pineapple regeneration and shoot growth as affected by 6-benzylaminopurine (BAP) at 2.0 mg/l and naphthalene acetic acid (NAA) at 0.2 mg/l in vitro. BAP and NAA at the concentration of 2.0 and 0.2 mg/l were used in this study. BAP at 2.0 mg/l significantly affected the production of shoots per explant, shoot length and weight. Total shoot length was higher in BAP (2 mg/l) than in control (MS medium without hormone) and NAA (0.2mg/l) after 10, 20, 30, 40, 50 and 60 days incubation period. Total shoot length was highest in BAP in all incubation periods. Total shoot weight was higher in BAP (2 mg/l) and lower in NAA (0.2 mg/l) as compared to MS medium without hormone. The results showed that BAP at the concentration of 2 mg/l was effective for pineapple shoot growth and development

Different wavelengths light to induce physiological changes callus for the biosynthesis of gymnemic acid in Gymnema sylvestre

A callus culture of Gymnema sylvestre was cultivated under different light conditions (blue, white, red and green light). Green-white coloured gymnemic acid containing cell groups were formed on the surface of the callus culture under blue light respectively. The phytohormone kinetin enhanced the gymnemic acid formation in blue light, but was unable to induce gymnemic acid in darkness. The phytohormone 2,4-D induced the formation gymnemic acid under all light conditions studied. Analysis of the callus growth phase under all treatments revealed that gymnemic acid accumulation was maximum (12.22 mg/g d.w) in the callus during stationary phase of 45 days, grown in MS medium with 2,4-D (1.5 mg/l) and KN 0.5 mg/l under white fluorescent light. Blue light enhanced the gymnemic acid accumulation up to 4.4 fold of that found under white fluorescent light and 2.8 fold of that found in intact leaves. Present findings concluded that blue light can be used as tool for enhancing pentacyclic tri-terpenoids in batch culture of G. sylvestre

Physiological responses of callus from gerbera jamesonii Bolus ex. Hook f. to Gamma Irradiation

In the present study, in vitro mutagenesis techniques were applied to investigate the effects of gamma irradiation at 0, 10, 20, 30, 40, 50 and 60 Gy on physiological changes in callus of Gerbera jamesonii Bolus ex. Hook f. Biochemical changes in chlorophyll and soluble protein content of pre- and post- irradiated Gerbera callus were studied. Non-irradiated callus demonstrated the highest amount of chlorophyll content as compared to callus irradiated at 10, 20, 30, 40, 50 and 60 Gy. In addition, the amount of chlorophyll b was relatively higher than chlorophyll a in both the irradiated and non-irradiated callus, except for callus irradiated at 10 Gy. Biochemical differentiation based on total soluble protein content revealed gradual reduction after day 9 of exposure to gamma irradiation. Reduction of soluble protein content was observed in all the treatments as the increase of incubation period

Tissue Culture Studies on Fortunella polyandra ‘Nagami’ and ‘Meiwa’

Studies of Fortunella polyandra ‘Nagami’ and ‘Meiwa’ were carried out to observe their responses in tissue culture systems. Explant sources investigated included roots, stems, leaves and cotyledons which were cultured on Murashige and Skoog (MS) medium supplemented with different combinations and concentrations of various hormones. The hormones used were naphthalene acetic acid (NAA) and benzylaminopurine (BAP) at pH 5.8 with a temperature of 23-26°C and a photoperiod of 16 hours light and 8 hours dark. The pH of the media was also altered to include 4.8, 5.8, 6.8 and 7.8. The various explants were subjected to light and dark treatments in order to study the morphogenesis of this species. From the results, it was observed that stem and leaf explants were more responsive than other explants. The best media for regeneration and callus formation was MS supplemented with 0.5 mg/L NAA and 0.5 mg/L BAP at pH 5.8 for ‘Nagami’ and pH 7.8 for ‘Meiwa’. Regeneration was achieved via direct organogenesis and also via callus formation. The highest percentage of shoot formation (40%) was obtained from stem explants (‘Meiwa’) cultured on MS supplemented with 0.5 mg/L NAA and 0.5 mg/L BAP at pH 7.8

Tissue Culture, Anatomical and Morphological Studies of Triphasia trifolia (Burm. f.) P. Wilson

Comparative anatomical studies were carried out on in vivo plants of Triphasia trifolia (Burm. f.) P. Wilson and in vitro plantlets of the same age. To get the in vitro plantlets, explants were cultured on MS (Murashige and Skoog) media supplemented with different concentrations and combinations of hormones. The explant sources of Triphasia trifolia (Burm. f.) P. Wilson were obtained from cotyledons, leaves, stems, roots and shoot tips that were placed under conditions of 16 hours light and 8 hours dark. The optimum media for regeneration was MS supplemented with 1.0 mg/L BAP and 1.0 mg/L NAA. Cotyledon explants were found to be the most responsive. Regeneration of complete plantlets was achieved from cotyledon explants after about 4 months in culture. Sectioning was done to study the characteristics of the respective vascular bundles, shape of cells, palisade cell layers, presence of oil glands, druse and cuticle layers. Vascular bundles of in vivo leaves were extremely well developed compared to those in in vitro leaves. The vascular bundle of the in vivo leaf showed well developed xylem. However, the xylem and phloem cells of the in vitro leaf were very poorly developed which is one of the features of in vitro plants. Scanning electron microscope (SEM) studies were also carried out on the in vivo and in vitro plantlets to observe differences on the leaf surface

Callus induction, plant regeneration and somaclonal variation in in vivo and in vitro grown White shrimp plant (Justicia betonica Linn.)

Justicia betonica Linn. is traditionally used to treat various illness like cough, scaly skin and as an erosion control plant. This paper reports on callus induction and in vitro propagation of J. betonica from 3-week-old petiole and internode explants of this species. Callus was readily induced from petiole explants when cultured on Murashige and Skoog's medium (MS) fortified with NAA (0.5-2.0 mg L-1), BAP (0.5-2.0 mg L-1), Kinetin (1.0-2.0 mg L-1) and Zeatin (1.0-2.0 mg L-1), while internode explants only showed formation of callus on MS basal and when 0.5 mg L-1 BAP or 1.5 mg L-1 Kinetin were added. Optimum in vitro regeneration in J. betonica had been successfully achieved using internode explants cultured on MS medium supplemented with 1.5 mg L-1 NAA and 0.5 mg L-1 BAP. Rhizogenesis was observed from petiole cultures supplemented with 1.0 mg L-1 NAA and 1.0 mg L-1 BAP as well as on MS with 1.5 mg L-1 NAA and 0.5 mg L-1 BAP, where the latter had shown the most optimum response, with percentage of root formation of 66.67 ± 5.36. More hormone treatments were observed to yield rhizogenesis from internode explants, especially when cultures were fortified with 0.5 mg L-1 NAA and 1.0 mg L-1 BAP. Genetic stability in in vitro J. betonica plants was observed by studying the variation in mitotic index and chromosome numbers using cytological data. Similar values of mean Mitotic index (MI) and mean chromosome numbers (2n = 14) indicated that the transfer from in vivo to in vitro environment had no significant effect on genetic stability of this plant. However, the mean nuclear size was found to increase, while the mean cell size was reduced in in vitro J. betonica compared to in vivo plants. In vitro regenerated J. betonica plants were clonally uniform and showed no distinct morphological abnormalities, indicating the lack of somaclonal variation