μM Kin Research Articles

Micropropagation and Acclimatization of Common Oregano (<i>Origanum vulgare</i> L. Subsp. <i>vulgare</i>) by Shoot Tip Culture

Origanum vulgare L. is a commercially valued species with remarkable biological properties. It is subject to over-exploitation practices that seriously threaten its sustainability for future generations. Thus, micropropagation serves as a tool for the protection and domestication of this species. In this study, we established an in vitro vegetative propagation protocol for Origanum vulgare. This is done through the axillary bud technique by carrying out various tests. Six culture media (MS, MSm, N30K, SD, SH and B5) were tested. Therefore, SD was chosen for the following experiments. Seven cytokinins (adenine (Ad), N6-(2-isopentenyl) (2ip), zeatin (Zeat), kinetin (Kin), benzyladenine (BAP), 1,3-diphenylurea (DPU) and thidiazuron (TDZ) at 5 concentrations (0.44, 1.33, 2.22, 3.11 and 4.44 μM/L) were evaluated. Thus, Kin at 3.11 μM allowed high regeneration of vitroplants, optimal elongation, total rooting of explants, maximum bud multiplication, and absence of hyperhydric explants. In fact, the integration of auxins (indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), and 1-naphthaleneacetic acid (NAA)) into the culture medium and their combinations with 3.11 μM Kinetin contributed to the optimization of the root part. Thus, it was improved in particular in the case of 3.11 μM Kin and 6.27 μM IBA. Three polyamines (Putrescine, Spermidine and Spermine) at different concentrations (1.134, 3.402, 5.67, 7.938 and 11.34 μM/L) combined at 3.11 μM Kin and 6.27 μM IBA were tested. In fact, 1.304 μM putrescine was considered to be the most suitable for in vitro culture of explants, since it allowed optimal propagation of buds and roots, also a high rate of regeneration and rhizogenesis. GA3 at 1.15 μM combined with 3.11 μM Kin and 6.27 μM IBA permitted maximum bud multiplication. The acclimatization was carried out successfully using vitroplants showing good foliar and root development. Thus, three months after acclimatization, the seedlings were transferred into large pots under natural light and temperature conditions. Almost all acclimatized plants developed flowers in the first year between May and July.

Interaction Effect of Plant Growth Regulators on Shoot Micropropagation of Aromatic Plant <i>Origanum elongatum</i> (Bonnet) Emberger & Maire

Origanum elongatum (Bonnet) Emb. & Maire, is a medicinal, aromatic and endemic plant of Morocco, characterized by its pharmacological effects, and is commonly used for the production of essential oils and aromas, resulting in high harvest and overexploitation pressure. This is why the present study aims to implement the in vitro micropropagation of Origanum elongatum for optimal vitroplant production. Six macroelements were tested (SH, SD, N30K, MS, MSm and B5) and the SD medium was selected for vegetative propagation of the explants. Seven cytokinins: adenine (Ad), N6-(2-Isopentenyl) adenine, zeatin (Zeat), kinetin (Kin), benzyladenine (BAP), 1,3-diphenylurea (DPU) and thidiazuron (TDZ) were then evaluated at five concentrations (0.44, 1.33, 2.22, 3.11 and 4.44 μM/L) on growth, development, budding, rooting and hyperhydricity. 0.44 μM Kin was selected and combined with three auxins: indole-3-acetic acid (IAA), indole-3-butyric acid (AIB), and 1-naphthaleneacetic acid (NAA) at four concentrations (1.14, 2.85, 4.56 and 6.27 μM/L) to improve rooting and association with 1.14 μM IAA was shown to be efficient for roots development. Different concentrations of gibberellic acid (0.29, 1.5, 2.60 and 2.89 μM/L), combined with 0.44 μM/L Kin and 1.14 μM/L IAA, were tested and 2.60 μM/L GA3 gave maximum buds and shoots. Then, the combination of three polyamines at five concentrations (1.134, 3.402, 5.67, 7.938 and 11.34 μM/L) with 0.44 μM Kin and 1.14 μM/L IAA showed an increase in the number of buds and shoots for 7.938 μM/L putrescine and 3.402 μM/L spermine. Finally, seedlings with good foliar and root development were acclimatized.

Dynamics of Polyphenol Biosynthesis by Calli Cultures, Suspension Cultures and Wild Specimens of the Medicinal Plant Ligaria cuneifolia (Ruiz & Pav.) Tiegh. (Loranthaceae). Analysis of Their Biological Activity.

Ligaria cuneifolia (R. et P.) Tiegh. (Loranthaceae) is a South American hemiparasitic species with antioxidant, antitumoral, antimicrobial, and antilipidemic activities attributed to its polyphenolic content. We studied the polyphenolic pattern of L. cuneifolia during different phenological stages: flowering, fruiting, and post-fruiting. The highest total phenolic content was found in stems at post-fruiting (214 ± 12.1 mg gallic acid eq·g−1 DW) and fruiting (209 ± 13.7 mg gallic acid eq·g−1 DW), followed by post-fruiting leaves (207 ± 17.5 mg gallic acid eq·g−1 DW). Flavonoids accumulated at higher levels in leaves and hydroxycinnamic acids in leaves at flowering and post-fruiting. The polyphenolic pattern was similar between organs from wild plants and in vitro cultures, although at a significantly lower level in the latter ones. The performance of calli growing under a 16 h photoperiod in a modified White medium with 1-naphthalene acetic acid (2.50 μM) and Kinetin (9.20 μM) was better than in the dark. When calli grew in media only with auxins (IAA, NAA, and 2,4-D, all at 2.50 µM concentration), its growth and polyphenolic content improved. Cell suspensions with 2.50 µM NAA and 9.20 µM KIN grew slowly and produced very small amounts of polyphenols. As for the antioxidant activity, it was detected in all samples (approximately 1000 µmol trolox eq·g−1 DW) except fruits, where a lower value was found (328 µmol trolox eq·g−1 DW). In vitro cultures have the lowest antioxidant activity when compared to methanolic extracts from organs of wild specimens. Finally, antimutagenic or mutagenic activity in wild plants and in vitro culture extracts was not detected by the Ames test.

Propagation of plum (Prunus salicina L.) cultivar Frontier in vitro through control of shoot tip necrosis (STN) and validation of genetic integrity using ISSR markers

The present investigation was carried out to for developing a competent protocol for micropropagation of commercial plum cultivar Frontier, through control of shoot tip necrosis (STN) observed during in vitro shoot multiplication. Apical and axillary buds were collected from elite high yielding tree of Frontier and inoculated on Murashige and Skoog (MS) medium fortified with phytohormones at different concentrations. MS medium containing 0.5 µM BA, 0.5 µM Kin and 0.2 µM IBA resulted in highest shoot bud establishment (58.95%) after 4 weeks interval. However, all the shoots showed necrosis and died after 1st subculture. Necrosis appeared with yellowing of shoot tips within 7 days of first subculture and gradually increased downwards resulting in yellowing of leaves, followed by death of the shoots. Among various reported methods tested to control STN, combination of Fructose (0.50 mM) and Calcium chloride (1.0 mM) proved 100% effective. Highest in vitro shoot multiplication (fivefold) was recorded on medium enriched with 2.0 µM BA, 0.2 µM GA3, 0.2 µM IBA, 0.50 mM Fructose and 1.0 mM Calcium chloride. For rooting, two step rooting procedure was followed which resulted in 85.00% rooting when microshoots were given a dark treatment by dipping in ½ MS broth supplemented with 2.5 µM IBA for 48 h before transferring to semisolid ½ MS basal medium. Rooted plantlets when transferred in vivo for acclimatization, showed 70–80% survival after 10 weeks and no mortality on field transfer. Molecular analysis of the plants using ISSR primers proved 100% similarity between them and mother tree.

Influence of Plant Growth Regulators on Callogenesis and the Biomass of Cell Suspensions in Lily (Lilium ledebourii and Lilium regal)

Introduction: Lily is often described as one of the most widespread, commercial crops in the floriculture industry. Commercially grown cultivars are mostly propagated by bulb scales which it cost effective and uniformity in tissue culture conditions. Plant tissue culture techniques can effectively provide far-reaching implications in micro-propagation. Materials and Methods: This study was conducted to investigate the effects of various concentrations of cytokines and auxins on callogenesis and Biomass under in vitro conditions. In this experiment, the evaluations were aimed at measuring different characteristics in two lily cultivars, namely, Lilium ledebourii and Lilium regal. Results: The results showed significant values in all of the measured characteristics. The highest percentage of callogenesis was caused by 2 µm PIC plus 1 µm KIN in L. ledebourii (88.66%) and L. regal (88.66%). Also, the callus weight in both cultivars was obtained by applying the same combination of treatments. In the second experiment, the highest fresh biomass of the cell suspension occurred by applying 2 µm PIC plus 4 µm KIN. The maximum amount of fresh biomass in L. ledebourii (88.93 g/L) and L. regal (41 g/L) occurred on the 24th and 20th day of the culture, respectively. Conclusions: An efficient and fragile callus induction was developed by NAA, which nonetheless made the condition more suitable for cell suspension culture. These lily cultivars need high amounts of PIC as auxin to grow well in cell suspensions. By increasing the PIC level, the biomass accumulates more. These results can moderately optimize large-scale production of both fragile calli and cell suspension biomass.

Tissue culture of Physalis angulata L. (Solanaceae): techniques for micropropagation and germplasm long-term preservation

Physalis angulata is considered an important crop in Mexico, established for commercial production due to its nutritional and medicinal properties. This study’s objective was to develop an efficient method for the mass multiplication of P. angulata and cryopreservation of apical meristems by droplet-vitrification. In vitro propagation was given in MS basal culture medium supplemented with 2.22, 4.43, and 6.65 μM 6-Benziladenine (BA) in combination with 2.32, 4.64, and 6.96 μM Kinetin (Kin). Root induction was obtained by dipping shoots in an auxin medium 1.07, 2.68, 5.37, and 8.05 μM 1-Naphthaleneacetic acid (NAA) or 1.41, 2.85, 5.70, and 8.55 μM Indoleacetic acid (IAA). The plantlets were acclimatized and transferred to the greenhouse. Results showed an efficient proliferation with an offshoot number of 6.57 ± 0.46 with 4.43 µM BA in combination with 2.32 µM Kin. Efficient rooting was obtained with 1.07 µM NAA with an average of 30.51 ± 0.94 roots. For cryopreservation, the effect of dehydration was evaluated by exposing shoot-tips to plant vitrification solution 2 at different times; 15, 30, 45, 60, and 75 min were tested before and after immersion to liquid nitrogen, and the highest percentage of regrowth (90%) was observed at 30 min. Experimental micropropagation by shoot proliferation system with cytokinins interaction (BA/Kin), rooting stimulation with auxins (NAA or IAA), successful acclimation, and cryopreservation by the droplet-vitrification procedure of Physalis angulata apical meristems.

Micropropagtion and DNA-barcoding of the endangered endemic Phlomis aurea plant of Saint Katherine

Saint Katherine is considered a “biodiversity hotspot” because of the high level of endemism of reported plant species. In this study, conservation of the endangered endemic plant; Phlomis aurea of Saint Katherine, Southern Sinai, Egypt, was carried out through micropropagation and DNA barcoding. The first efficient micropropagation protocol for Phlomis aurea was established as a mean of ex situ conservation of the plant. Shoot tips and nodal segments of in vitro germinated seedlings were established on Murashige and Skoog medium supplemented with 0.54 µM β-naphthalene acetic acid (NAA) and 2.46 µM N6-(2-isopentenyl) adenine (2iP) in combination with 6-benzylaminopurine (BA) or kinetin (Kin). The medium supplemented with 3.48 µM Kin considered optimum for both explants. For multiplication, BA was the most efficient cytokinin. The percentage of rooted explants reached 100% at the concentration of 14.7 µM indolebutyric acid (IBA), whereas the highest number of roots was recorded for 4.90 µM, which considered the optimum concentration with a percentage of 80% of rooting. Rooted plantlets were transplanted in the greenhouse with 75% survival rate. The present study also aimed to carry out DNA barcoding of Phlomis aurea for accurate identification to provide a database for establishing an efficient conservation program for the plant. Three chloroplast DNA markers were used [ribulose-1,5-bisphosphate carboxylase/oxygenase large subunit (rbcL), maturase K (matK) and RNA polymerase C1 (rpoC1)] and all were successful in amplifying target regions, however the performance of both rbcL and matK markers seemed to be species‐specific. The similarity percentage was maximum for rbcL (99.81%) and matK (100%) compared to the database of the same species.

Micropropagation of the Moroccan Endemic Plant <i>Thymus broussonetii</i> Boiss. with Aromatic-Medicinal Value and Conservation Concern

Micropropagation from shoot tips and nodal segments was carried out for the conservation and domestication of spontaneous Moroccan thyme, Thymus broussonetii Boiss. subsp. broussonetii (endemic threatened). The mineral composition of the culture medium, as well as the succession of different growth regulators, influenced the in vitro growth of this species. Sterilized achenes of T. broussonetii were able to germinate on an agar medium containing Gautheret macronutrients with a rate of 25% and a degree of contamination of less than 4%. Shoot apices of 15-day seedlings (two cotyledon leaves) were cultivated on SD + 0.46 μM Kin medium and the explants obtained were transplanted every month. Six macronutrients (MS, B5, SH, SD, MSm and N30K) were tested and N30K was chosen for the following experiments. Seven cytokinins (Kin, BAP, 2iP, DPU, adenine, Zeat and TDZ) at 0.46, 0.93 and 2.32 μM/l were evaluated and the addition of 0.93 μM adenine to N30K medium favored significantly the induction of buds and the elongation of explants. Three polyamines (putrescine, spermidine and spermine) at 2, 5, 10 and 20 μM/l were tested. A better multiplication of buds, shoots and roots was noted for N30K + 10 μM spermine. Cytokinin-auxin combinations led to better root multiplication and an increase in the number of buds and the length of explants, particularly for 0.46 μM Kin + 2.85 μM IAA. Acclimatization was successfully carried out using vitroplants developing a good root system. One month after the start of acclimatization, 97% of T. broussonetii plantlets were healthy. Three months later, they were transplanted into larger pots. 100% of the acclimatized plants developed flowers in the 2nd year between June and August. Re-initiation of the in vitro culture was carried out from sterilized twig segments collected from the acclimatized plants of T. broussonetii with 1 - 2 nodes on the medium N30K + 0.46 μM Kin, and 52.1% of the explants healthily proliferated. Finally, two micropropagation prototypes were developed: shoot tip culture from seedlings obtained after germination of achenes and node culture from acclimatized plants.

Assessment of genetic fidelity, biochemical and physiological characteristics of in vitro grown date palm cv. Al-Fayda

The effects of various culture conditions on adventitious bud multiplication and plantlet regeneration in date palm cv. Al-Fayda were investigated. Adventitious buds were cultured on semi-solid Murashige and Skoog (MS) medium supplemented with three different plant growth regulator (PGR) combinations. The effects of various strengths and textures of the multiplication medium were evaluated and the adventitious buds were cultured on media supplemented with various types and concentrations of carbon sources. It was found that semi-solid 1/2MS medium supplemented with 2.4 µM NOA, 2.3 µM KIN and 40 g l−1 sucrose is the most effective for shoot bud multiplication with 30 shoot buds per explant. On the other hand, liquid media resulted in very high levels of hyperhydricity (up to 100%). Peroxidase activity ranged from 372 to 709 U g−1 FW and was higher in buds cultured on liquid media, while the total protein concentration ranged from 16.3 to 46.7 mg g−1 FW. Shoot elongation and rooting were successfully achieved on semi-solid 1/2MS medium supplemented with 2.4 µM NOA, 2.3 µM KIN and 40 g l−1 sucrose. Regarding the physiological characteristics of developed shoots, the chlorophyll content ranged from 6.50 to 14.40 CCI, the chlorophyll fluorescence from 0.682 to 0.750, the foliar surface from 6.85 to 9.88 cm2 and the stomatal conductance from 8.97 to 12.51 mmol m−2 s−1. The rooted plantlets were successfully acclimatized in the glasshouse with a survival rate of 70%. The genetic conformity of regenerants was assessed by inter-simple sequence repeat (ISSR) markers, and a high similarity with the mother plant was observed.

Somatic embryogenesis from stamen filaments of Aesculus flava Sol. and peroxidase activity during the transition from friable to embryogenic callus

Species of the genus Aesculus are among the most attractive ornamental woody plants. Conventional propagation methods of these species are either inefficient (stem cuttings) or unsuitable for clonal propagation (seeds). The aim of the present study was to develop an efficient protocol for clonal propagation of elite specimens of yellow buckeye (Aesculus flava) by somatic embryogenesis. For this purpose, stamen filaments of yellow buckeye were cultivated on media supplemented with 1, 5 or 10 μM 2,4-dichlorophenoxyacetic acid (2,4-D) combined with 0, 1, 5 or 10 μM 6-furfurylaminopurine (Kin), either under light or dark conditions, for 8 weeks, and then subcultivated on plant growth regulator (PGR)-free medium with 400 mg/l of glutamine. The highest somatic embryo (SE) initiation rates were achieved for the explants cultivated in darkness on medium containing 1 μM 2,4-D + 10 μM Kin during callus induction (CI) phase. Embryogenic calli (EC) were initiated from friable calli, starting from the 7th week of culture initiation, while SEs appeared two weeks later, following a week of subcultivation of the explants on PGR-free medium. EC and SEs were observed only in the explants grown in darkness during CI phase. Minimal duration of CI phase and darkness necessary for SE induction was four weeks, while the highest embryogenic response was achieved when each lasted for 8–10 weeks. Obtained SEs were efficiently multiplied on medium supplemented with 0.05 μM 2,4-D + 5 μM Kin by recurrent somatic embryogenesis. SEs at globular stage of development exhibited the highest capacity for secondary SE regeneration. High germination and conversion rates were attained in cotyledonary-stage SEs cultivated on medium with 0.05 μM 2,4-D + 5 μM Kin, but this phase needs to be further optimised, since the obtained plants failed to acclimatize to greenhouse conditions. During the transition of calli from friable to embryogenic state, total peroxidase (POX) activity significantly decreased, indicating their involvement in the acquisition of embryogenic capacity. The presented protocol is suitable for clonal propagation and genetic transformation of this ornamental species, and POX activity may be used as a marker for SE initiation.

An efficient in vitro regeneration protocol to generate stable transgenic lines of Black gram (Vigna mungo)

An efficient and quick in vitro regeneration protocol was developed for black gram (Vigna mungo) using wounded embryonic axis with cotyledon as explant. Murashige and Skoog (MS) medium supplemented with 4.44 μM BAP and 2.32 μM Kinetin was found to be effective in producing maximum number (mean 7.80) of multiple shoots. The individual shoots elongated to 4.5 cm when MS medium was supplemented with 2.89 μM GA3 along with 0.44 μM BAP and 0.46 μM KIN. A novel in vitro rooting technique was also optimized for black gram using half-strength liquid MS medium supplemented with 1.34 μM NAA. The shoots in this medium produced the highest number (mean 7.50) of roots with root length of 6.02 cm. The plantlets were transferred to soil mixture and placed in greenhouse where more than 80% successfully grew to maturity. The same protocol was successfully used to generate transgenic black gram lines carrying Bt-Cry2Aa gene through Agrobacteriummediated transformation with a transformation efficiency of 0.42%. The rooted T0 plants grew to maturity and produced T1 seeds with the presence and expression of transgene in T1 plants. Thus, we have standardized an in vitro regeneration protocol suitable for generation of stable transgenic black gram plants.

Growth, Metabolite Profile, Oxidative Status, and Phytohormone Levels in the Green Alga Acutodesmus obliquus Exposed to Exogenous Auxins and Cytokinins

The effects of auxins and cytokinins at the range of concentrations 0.0001–100 µM on Acutodesmus obliquus (Chlorophyceae) cultures were studied. Microalga exhibited sensitivity to cytokinins in the following order: 0.01 µM tZ > 0.1 µM Kin > 1 µM DPU, whereas the hierarchy of auxin activity was: 0.01 µM IAA > 0.1 µM IBA > 0.1 µM PAA. Cytokinins possessed higher stimulating properties on the cell number, whereas auxins increased the size of cells. Differences in the metabolite profiles of the cultures treated with phytohormones were observed. Auxins and cytokinins had a positive effect on the photosynthetic apparatus enhancing the level of chlorophylls, carotenes, and xanthophylls. In comparison with auxins, cytokinins more effectively delayed oxidative damage by increasing the level of non-enzymatic antioxidants (ascorbate, glutathione) and the activity of enzymes scavenging reactive oxidative species (catalase, glutathione reductase, ascorbate peroxidase). On the other hand, auxins stimulated superoxide dismutase activity and provoked hydrogen peroxide generation, which may be involved in cell enlargement. All phytohormones reduced the content of abscisic acid and controlled the level of endogenous auxin and cytokinins suggesting complex interactions. Different dynamics of A. obliquus responses to auxins and cytokinins clearly demonstrated their diverse roles in algal growth and metabolism.

Callus induction and establishment of cell suspension cultures of the halophyte Armeria maritima (Mill.) Willd.

Abstract The objective of this work was to establish a cell suspension culture from Armeria maritima (Mill.) Willd., an halophyte so far only valuable for horticultural purposes, as a pre-requisite step towards the production of bio-actives secondary metabolites in vitro. For the initiation of callogenesis, five types of explants were tested: leaves from mature plants; leaves, cotyledons and roots from axenic seedlings; seeds. Three criteria either from a quantitative (percentage of callus induction) or a qualitative basis (aspect and friability) were used to evaluate the effectiveness of callus formation. Seeds-derived calli grown in the dark on MS medium supplemented with 4.5 μM 2.4D and 0.93 μM KIN were successfully selected for the establishment of cell suspension cultures, characterized by a growth index of 3.4 after 14 days of culture.

English

Curcuma longa L., from the Zingiberaceae family, generally reproduces through its rhizomes, which are also utilized for therapeutic purposes because they are rich in terpenoids. Its conventional propagation has low efficiency due to the small number of seedlings and their contamination by pathogens. Therefore, this study aimed to evaluate the influence of growth regulators on the development of in vitro-cultivated C. longa as well as to determine their influence on DNA content and foliar anatomy. Shoots were inoculated in MS culture medium with the addition of 30 g/L of sucrose and 6.5 g/L of agar, and a pH adjusted to 5.8. Two assays were built to study the multiplication and rooting phases of growth. The first assay evaluated the influence of eight concentrations of cytokinins and auxins on the multiplication phase. Leaf samples were analyzed for DNA content through flow cytometry, utilizing two reference standards, green pea, and tomato. Characteristics of leaf anatomy were also measured in four time periods. The second assay analyzed the influence of six auxin concentrations on the rooting phase. The first assay showed that the root systems grew more in treatment 3 (4.44 µM benzylaminopurine [BAP], 0.46 µM kinetin [KIN]) and reached greater dry mass in T8 (8.88 µM BAP, 0.92 µM KIN, 2.16 µM naphthalene acetic acid [NAA]). The largest fresh matter of the main shoot was found in T2 (4.44 µM BAP). The estimated DNA content varied depending on the presence of supplemental growth regulators, from 2.38 to 2.77 pg, and was greater in T4 (4.44 µM BAP, 1.08 µM NAA) and T5 (4.44 µM BAP, 0.46 µM KIN, 1.08 µM NAA). Results from the latter two treatments were not significantly different. Estimates of DNA content were precise, as indicated by coefficients of variation that were much lower than 5%. The results also showed quantitative variation of evaluated anatomical traits. In general, there was a thin epidermis layer with rectangular cells, followed by parenchyma with octahedral cells and differentiated xylem and phloem. In leaf parenchyma, the presence of idioblasts containing phenolic compounds was observed in all growth stages. In the rooting phase, the supplementary auxins affected the dry matter of the aerial part and roots. The highest averages were observed in treatments with 2.0 µM NAA. Key words: Turmeric, micropropagation, flow cytometry, vegetal anatomy.

In vitro rapid multiplication of Coldenia procumbens L. from shoot tip explants

An efficient protocol for in vitro rapid multiplication of Coldenia procumbens L. has been developed. The shoot tip explants were cultured on MS basal medium supplemented with different concentrations of BAP and KIN. The two cytokinins are tested, BAP was found to develop higher number of shoots from the shoot tip explants when compared to KIN. higher number of shoots was produced from all the concentrations of BAP and KIN. The highest frequency of 100% shoot induction was observed on MS basal medium supplemented with 6 μM BAP and 10 μM KIN. The number of shoots observed on the MS basal medium supplemented with BAP obtained 14 as well as 11 in KIN. The data's are recorded after 30days of inoculation.

Establishment of the Biosorptive Properties of Ulva Lactuca Subjecting Eleusine Coracana (L) Gaertn to Heavy metal stress By Cobalt


 An efficient protocol for in vitro rapid multiplication of Coldenia procumbens L. has been developed. The shoot tip explants were cultured on MS basal medium supplemented with different concentrations of BAP and KIN. The two cytokinins are tested, BAP was found to develop higher number of shoots from the shoot tip explants when compared to KIN. higher number of shoots was produced from all the concentrations of BAP and KIN. The highest frequency of 100% shoot induction was observed on MS basal medium supplemented with 6 μM BAP and 10 μM KIN. The number of shoots observed on the MS basal medium supplemented with BAP obtained 14 as well as 11 in KIN. The data's are recorded after 30days of inoculation.

In vitro Shoot multiplication of Physalis minima L. - an important Medicinal Herb

The present study was to develop an efficient protocol for shoot multiplication of Physalis minima L. Shoots were cultured using nodal segments from 20 days old field grown young plant. The nodal explants were cultured on MS medium supplemented with different concentrations of BAP and KIN. The two cytokinins tested, KIN was found to respond well in shoot multiplication and number of shoots from the nodal explants when compared to BAP. Large number of shoots was induced from all the concentrations of both BAP and KIN. BAP was found to develop in shoot multiplication and higher number of shoots from the nodal explants when compared to KIN. The highest frequency of 100% shoot induction was observed on MS basal medium supplemented with 8μM BAP and 10 μM KIN. Matured shoots are isolated and then transferred to the MS basal medium supplemented with different concentration of NAA and IBA for root induction. The higher number of roots were produced in 30 days on MS basal medium supplemented with 10 μM IBA and 6 μM NAA. The well rooted plantlets were isolated and transplanted to the paper cup for hardening and the well established plants were transferred to the field for acclimatization.

Cytokinins differentially affect regeneration, plant growth and antioxidative enzymes activity in chive (Allium schoenoprasum L.)

Unlike garlic and onion, the regeneration of chive (Allium schoenoprasum L.), cultivated both for culinary and ornamental purposes, has not been intensively studied. The effects of the eight cytokinins and the plant basal section thickness on regeneration efficiency and subsequent plant growth were studied. Representatives of all cytokinin structural groups: isoprenoide side chain (trans-zeatin) and aromatic side chain (benziladenine, kinetin, meta-topolin) adenine derivatives, and phenylurea derivatives (thidiazuron and N-(2-chloro-4-pyridyl)-N′-phenylurea) at 0, 1, 5 or 10 μM were used. Histological analysis revealed adventitious buds formation from the leaf sheaths’ bases and the basal plate. The highest regeneration frequency (100 %) and the mean bud number per explant (20.0) were achieved with 10 μM thidiazuron (TDZ), and 5 mm-thick basal sections were the most responsive explants. Inferior shoot and root growth characteristics of plants regenerated by this treatment was avoided by exclusion or replacement of 10 μM TDZ with 5 μM kinetin (Kin) after a 4-week bud induction period, without consequences on the regeneration efficiency. In addition, a positive correlation between peroxidase, catalase and superoxide dismutase activity and the regeneration capacity was observed. All antioxidative enzymes activity changed much faster with 10 μM TDZ than with 1 μM Kin, which provoked the weakest regeneration response. Moreover, a unique peroxidase isoform was observed only in TDZ-treated explants after 3rd day of treatment. This work is useful for genetic engineering and virus-free plant production advancement, and for the knowledge expansion regarding the role of antioxidative enzymes in plant organogenesis.

English

Prunus africana (Kanda stick) is a valuable medicinal tree classified as priority species for domestication in Cameroon. To measure the ability for developing axillary buds and multiple shoots in relation with the origin and state of explants, some biochemical parameters were assessed in eight types of single-node cuttings. These cuttings were treated with different concentrations of kinetin (kin) indole butyric acid (IBA) and thidiazuron (TDZ) and cultured in Murashige and Skoog (MS) medium. The glutelin content (1.13 ± 0.201 µg/mg FW) was higher compared to those of total albumin, globulin and prolamin and most representative at mature vegetative apical stage (MVA). The highest (15.60 ± 1.966 µg/mg FW) and lower (2.82 ± 0.072 µg/mg FW) amount of phenol compound was observed in the mature dormant stages and immature vegetative distal stage (IVD), respectively. Regarding the peroxidases, highest activity (0.453 ± 0.055 OD/mg/ FW.5 min -1 ) was obtained in mature dormant apical stage. Apart from IVA state, there was no reactive explant cultured in vitro. The high percentage of budding and proliferation of budding (100%) and rooting (100%) were achieved with 12 μM Kin and 24 μM IBA, respectively. In the presence of 24 μM TDZ, multiple shoot was induced with a maximum of 5 shoots/explants. Micropropagation success of P. africana is influenced by cytokinin, physiological and biochemical status of single node cutting. Key words : Biochemical profile, Prunus africana, budding, multiple shoots, rooting.

Root explant produces multiple shoot from pericycle in Psoralea corylifolia – a leprosy destroyer medicinal plant

Babchi (Psoralea corylifolia L.) is a rare medicinal plant. It has been considered to be efficacious in leprosy and hence given the name ‘Kushtanashini’ (leprosy destroyer) in Ayurveda. The high death rate in wild along with aimless and unlawful gathering and destruction of habitat put the plant in the category of endangered list. In this study a highly reproducible in vitro regeneration system in P. corylifolia using root segments was developed. The effect of different PGRs (Kin BA, and NAA) on the percentage of adventitious shoot induction and the number of shoot buds per root explant was assessed. The maximum response (65.57%) was observed in the media supplemented with 2.22μM BAP and 6.98μM Kin. The treatment had in average 39.67 number of shoot buds and 3.37 elongated shoots. Elongation response was best (92.0%) in MS+2.22μM BAP+6.98μM Kin media fortified with 2.89μM GA3 and 39.7μMPG, producing a mean of 9.3 elongated shoots with an average of 4.6cm length. An anatomical study showed that the origin of shoot buds were directly from pericycle of root. Elongated shoots were carefully excised and rooted on basal MS+4.92μM IBA+158.6μMPG, with 92 per cent rooting. Rooted explants survived in soil: vermicompost: vermiculite (1:1:1) with 100% success.