Analysis of preliminary phytochemical screening of Typhonium flagelliforme

Typhonium flagelliforme (Araceae) is a medicinal herb which is endowed with curative properties against a variety of illness including injuries, oedema, coughs, pulmonary ailments, bleeding and cancer. In order to assess its phytochemical components, an experiment was conducted on one to six month old ex vitro and in vitro extracts of T. flagelliforme. The active (ex vitro and in vitro) extracts of T. flagelliforme were screened for phytochemicals components such as alkaloids, flavonlids, terpenoids and steroids. Alkaloids and flavonoids are the main phytochemical constituents of T. flagelliformewhich are found to be in the highest amount in two and four month old of ex vitro plants. High amounts of main phytochemical constituents were observed during the flowering process which started in two month old plant and finished at the end of the three month old plant

In vitro mass propagation of Typhonium flagelliforme as affected by plant growth regulators

Tubers were used as explants in in vitro mass propagation of Rodent Tuber (Typhonium flagelliforme). The explants were obtained from sterile plantlets and placed in shoot induction medium containing basal salts of Murashige and Skoog (MS) and various concentrations of 6-benzylaminopurine (BAP) and -naphthaleneacetic acid (NAA). Treatment containing 5 mg/l (w/v) of BAP with 1 mg/l (w/v) of NAA produced the highest number of shoots per explant (29.17) after 12 weeks of culture and also the highest mean fresh weight of shoots formed in treatment containing 5 mg/l (w/v) of BAP with 1 mg/l (w/v) of NAA. For ex vitro establishment, well- rooted plantlets were transferred in potting medium containing peatmoss, perlite and vermiculite (3:1:1)

In vitro plant regeneration from protocorms-like bodies (PLBs) and callus of Phalaenopsis gigantea (Epidendroideae: Orchidaceae)

Phalaenopsis, with long arching sprays of flowers, are among the most beautiful flowers in the world. Phalaenopsis is an important genus and one of the most popular epiphytic monopodial orchids, grown commercially for the production of cut flowers and potted plants. Most of them have different and interesting morphological characteristics which have different value to the breeders. Phalaenopsis gigantea is one of the most difficult to grow and has the potential of producing beautiful hybrids. An efficient and reproducible method for large-scale propagation of Ph. gigantea using leaf sections has been developed. Leaf sections from in vitro young plants were cultured on New Dogashima Medium (NDM) supplemented with cytokinins (6-Benzylaminopurine (BAP), Thidiazuron (TDZ), and Kinetin (KIN), each at 0.01, 0.1, 0.5 and 1.0 mg/L) alone and in combinations with (auxins a-naphthaleneacetic acid (NAA), at 0.01, 0.1, 0.5 and 1.0 mg/L). The explants developed calli and protocorm-like-bodies (PLBs) within 6 weeks of culture. Treatment TDZ in combination with auxins was found to be the best for the induction of callus and PLBs. In vitro regeneration of Ph. gigantea PLB was achieved by exposure to light and transferring to hormone free NDM solid medium.Key words: Phalaenopsis, PLBs, new Dogashima medium, regeneration

In vitro adventitious shoot regeneration and acclimatisation of Brassica oleracea subsp. italica cv. Green Marvel

Cotyledonary explants of Brassica oleracea subsp. italica (broccoli) cv. Green Marvel were cultured on Murashige and Skoog (MS) medium containing different combinations of the growth regulators 6- benzylaminopurine (BAP) and α-naphthalene acetic acid (NAA) for shoot regeneration. The optimal medium for inducing shoots contained 3 mgl-1 BAP and 1 mgl-1 NAA, which produced a shoot induction percentage of 53.33% and a mean number of 0.43 shoot per explant. The shoots were subsequently rooted in MS medium that contained 0.2 mgl-1 of indol-3-butyric acid (IBA). Different potting media were assessed during plantlet acclimatization. The highest percentage of plant survival (83.33%) was on the medium that contained sand and soil (1:1), while maximum root length (4.37 cm) and plant height (7.87 cm) were attained in potting medium that consisted peat moss, perlite and vermiculite (3:1:1).Key words: Brassica oleracea, broccoli, 6-benzylaminopurine, α-naphthalene acetic acid, indole-3- butyric acid

Rapid multiplication of Safed musli (Chlorophytum borivilianum) through shoot proliferation

Young shoot buds were used as explants for rapid multiplication of Safed musli (Chlorophytum borivilianum). The explants were cultured onto medium containing basal salts of Murashige and Skoog (MS) and various  concentrations of 6-benzylaminopurine (BAP) and kinetin (KIN) for shoot induction. Treatment containing 3.0 mg/l BAP produced the highest mean number of shoots per explants (18.90) and a mean length of shoots (6.0 cm) after 28 days of culture. Regenerated shoots were successfully rooted on MS medium supplemented with 1.0 mg/l indole-3-butyric acid (IBA) and 30 g/l sucrose. For ex vitro establishment, well-rooted plantlets were transferred in potting medium containing vermiculite : organic matters (1:1)

In vitro performances of hypocotyl and cotyledon explants of tomato cultivars under sodium chloride stress

A plant tissue culture technique is a good method for the evaluation and screening of plant genotypes for salt tolerance. In vitro evaluations of sodium chloride (NaCl) effects on two tomato cultivars (Pearl and Beril) were investigated with four NaCl levels (0, 25, 50 and 75 mM) using hypocotyl and cotyledon explants. The explants were cultured in MS media having 2.0 mg/l BAP along with different concentrations of NaCl. Sodium chloride stress negatively affected the growth traits and chlorophyll content. Significant differences were noticed between the cultivars followed by different NaCl levels, where the Beril responded superior than that of Pearl. The type of explant showed a difference in their response to shoots regeneration under NaCl stress, where the cotyledon explants achieved best results than hypocotyl explants.Key words: Cotyledons, hypocotyls, In vitro, salt stress, tomato

Molecular characterization and phylogenetic relationships among and within species of Phalaenopsis (Epidendroideae: Orchidaceae) based on RAPD analysis

Random amplified polymorphic DNA (RAPD) analysis for 20 species of Phalaenopsis was conducted to determine their genetic distances and relationships. Among 20 different primers used for RAPD analysis, 10 primers showed polymorphism, and according to the primer type, 26 to 54 DNA fragments were amplified. A total of 414 polymorphic fragments were generated by 10 primers and used for correlation group analysis. The highest value of Similarity index was 0.28 between Ph. violaceamalaysia and Ph. violacea witte. The dendrogram resulting from UPGMA (Unweighted Pair Group Method using Arithmetic average) hierarchical cluster analysis separated the original species into threegroups: The first group had five species of Ph. violacea blue, Ph. belina, Ph. violacea malaysia, Ph. violacea witte, and Ph. gigantea; the second group included Ph. lamelligera, Ph. amabilis, Ph. parishii, Ph. labbi nepal, Ph. speciosa, Ph. lobbi yellow, Ph. venosa, Ph. hieroglyphica, and Ph. maculata; the third group consisted of Ph. minho princess, Ph. leopard prince, Ph. mannii, Ph. modesta, Ph. cornucervi and Ph. pantherina. RAPD markers can thus be successfully applied in this economicallyimportant group of orchids for the study of molecular characterization and relationships. The data acquired from this study could be used for identification and classification of other orchid genera andoriental Phalaenopsis

RAPD analysis of colchicine induced variation of the Dendrobium Serdang beauty

Variation was detected in Dendrobium Serdang Beauty V (DSB V) plantlets regenerated from protocorm like bodies (plbs) induced by various concentration levels of colchicines in the Murashige and Skoog media (MS) supplemented with 1.5 mg/L IBA. RAPD analysis detected 6 - 26% variation in the regenerants from the mother plant. The highest variation was obtained in regenerates treated with 25 mg/L colchicine, which also exhibited reduced regeneration rates from plbs and mean plantlet fresh weight. RAPD analysis also showed high polymorphism between the mutated regenerant DSB V, and 13 species of the Dendrobium genera, and 13 orchids across generas. However, despite the 26% colchicine induced variation in the regenerants, all RAPD analysis revealed that DSB V was closely related to the mother plant. Thus, the RAPD technique is favourable for variation detection as it was sensitive enough to detect variations at species level and among somaclonal variants in this study

Establishment of a plant regeneration system from callus of Dendrobium cv. Serdang Beauty

An in vitro propagation protocol was established for the Dendrobium Serdang Beauty orchid. The propagation protocol utilized calli tissues that were successfully initiated from protocorm-like bodies(PLBs) explants, while the leaf and root tip explants died. The percentage of protocorm-like bodies explants responding to calli formation was 100% in all tested levels of IAA, IBA and NAA auxintreatments. The highest amount of calli (49.59 gram) proliferated on MS medium containing 1.5 mg/L IBA. These calli successfully regenerated on media supplemented with either KIN or BAP cytokinins and combined treatments of KIN and IAA (4 mg/L) or NAA (1.5 mg/L). However, media supplemented with only 1 mg/L KIN was sufficient to produce significantly high percentage of plantlet formation (80%),high number of planlets per explant (4-5 plantlets) and high mean fresh weight per plantlet (11.128 g). These plantlets were acclimatized on all tested media and obtained satisfactory rate of plantlet survival(80-100%), mean number of leaves per plant (4-6 leaves), and mean leaf length (4 - 5 cm). Among these media, charcoal was considered the most economical and available material in the local market. Duringthe development of this protocol, substantial necrosis of calli were observed when cultures were treated with 2,4-D and BAP. It was proposed that the presence of ethylene within the cultures, which isknown to be emitted by plant growth regulators into the micro-climate of in vitro culture vessels, is the determining factor of a suitable plant growth regulator for the survival and growth of the DendrobiumSerdang Beauty calli cultures in our study

Detection of somaclonal variation by random amplified polymorphic DNA analysis during micropropagation of Phalaenopsis bellina (Rchb.f.) Christenson

Phalaenopsis bellina (Rchb.f.) Christenson orchid species are known for their beautiful flower shape, graceful inflorescence and fragrance. Protocorm-like bodies (PLBs) of P. bellina were induced from leaf segments. The PLBs were then subjected to proliferation using ½ strength Murashige and Skoog (MS) media with two subcultures at three months intervals. Twelve decamer random amplified polymorphic DNA (RAPD) primers were used to study somaclonal variation among the mother plant, the initially induced PLBs and proliferated PLBs after 3 and 6 months in culture. Eight out of twelve primers produced 172 bands with 18 polymorphic bands in all the treatments. The amplified products varied between 125 to 8000 bp. Among the primers used, P 16 produced the highest number of bands (29), while primer OPU 10 produced the lowest number (15). The range of similarity coefficient was from 0.83 to 1.0 among the different sub-cultures and mother plant (MP). It was found that minimal or no changes occurred between the MP and the PLBs produced after 3 months of induction. The induced PLBs were then subcultured for six months for proliferation and this resulted in about 17% dissimilarity with MP. It is reported that micropropagation of P. bellina can be carried out successfully using ½ strength MS media for 6 months but further proliferation may result in somaclonal variation which might change the prolific characteristic of this orchids.Key word: Moth orchid, somaclonal variation, random amplified polymorphic DNA, protocorm-like bodies