Individual Shoots Research Articles

Assessing the efficiencies and challenges for nutrient uptake by aquatic plants

Aquatic plant meadows are valuable components to the ‘coastal filter’ and it is important to understand the processes that drive their ability to cycle nutrients. However, at present, the field-based evidence for understanding the drivers of nutrient uptake by plants is lacking. This study aimed to investigate how well individual shoots of aquatic plants could meet their nitrogen demands using the sediment nutrient pool (porewater ammonium) and to explore which traits helped to facilitate such uptake. Several species were investigated in shallow, submerged (2–4 m) mixed-species communities in the northern Baltic Sea using incubation experiments with enriched ammonium. After a 3.5 h incubation time, individuals were collected and analysed for nitrogen (% DW) and 15N (at-%) concentrations. Uptake by plants was calculated per unit nitrogen in response to the 15N-labelled source and to overall nitrogen availability. Background porewater ammonium availability was highly variable between individual plants. Species identity did not significantly affect uptake metrics and the effect of ambient porewater availability was weak. As biomass increased there were significant logarithmic declines in the 95th quantiles of nutrient uptake rates, ambient porewater nutrient availability and aboveground nitrogen tissue concentrations (% DW). Such findings suggested that uptake rates of plants were significantly demand-driven and the nutrient conditions of the porewater were significantly driven by the demands of the plant. Findings parameterised the unfulfilled potential for some aquatic plants to cycle nutrients more efficiently and highlighted the potential importance of access to new nutrient sources as a way of enhancing nutrient cycling by aquatic plants. Plant traits and community properties such as the activity of infauna could facilitate such an access and are likely important for nutrient uptake.

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.

Effect of gamma irradiation on the morphological and physiological variation from In vitro individual shoot of banana cv. Tanduk (Musa spp.)

Inducing genetic and morphological variation through conventional method is very difficult. Therefore, mutation induction through in vitro technology brings numerous advantages over the conventional breeding. Thus, the individual shoots (1 ~ 2 cm) were irradiated with gamma rays (10 ~ 70 Gy). The result revealed that the explants treated with higher doses (40, 50, 60, and 70 Gy) showed deleterious effects of ionizing radiation. The highest survival rate among γ treated explants recorded was 71% in 10 Gy treatments while the lowest survivality was 15% in 70 Gy. Lethal dose 50% (LD50) dose was found to be 33 Gy. In the in vitro condition, rooting reponse showed that increase in gamma irradiation dose resulted in the inhibition of root growth. Meanwhile, non-treated explants had the best rooting ability with the maximum number of root per explant (20) within a short period of time (6 days), with the highest root length of (15.1 cm). The longer period in rooting (12 days) and lowest number of root per explant (8) with shortest root length (10.1 cm) were recorded at 30 Gy treatment. The highest shoot length (13.6 cm) was observed at control treatment and the shortest shoot length (10.4 cm) was observed at 30 Gy. In the nursery, lowest leaf number (5) was observed at 30 Gy compared with other treatments. The highest chlorophyll content (49.8) was recorded at 10 Gy treated seedling. Irradiated explants with 10 Gy found to be superior over the control treatment and had positive effects in main growth parameters such as chlorophyll content.

Local Insect Damage Reduces Fluctuating Asymmetry in Next-year’s Leaves of Downy Birch

Insect herbivory imposes stress on host plants. This stress may cause an increase in leaf fluctuating asymmetry (FA), which is defined as the magnitude of the random deviations from a symmetrical leaf shape. We tested the hypothesis that differences in leaf FA among individual shoots of downy birch, Betula pubescens, are at least partly explained by local damage caused by insects in the previous year. Unexpectedly, we found that in the year following the damage imposed by miners, leafrollers and defoliators, damaged birch shoots produced leaves with lower FAs compared to shoots from the same tree that had not been damaged by insects. This effect was consistent among the different groups of insects investigated, but intra-species comparisons showed that statistical significance was reached only in shoots that had been damaged by the birch leaf roller, Deporaus betulae. The detected decrease in leaf FA in the year following the damage agrees with the increases in shoot performance and in antiherbivore defence. The present results indicate that within-plant variation in leaf FA may have its origin in previous-year damage by insects, and that FA may influence the current-year’s distribution of herbivory.

Influence of Sucrose on Growth and [6]-Gingerol Production of In vitro-Grown Ginger (Zingiber officinale Roscoe)

The effect of sucrose on growth and [6]-gingerol production of in vitro- grown Zingiber officinale was investigated. Individual shoots from multiple shoot clumps were cultured in Murashige and Skoog (MS) basal medium supplemented with varying amounts of sucrose, i.e. 30, 60, 90 and 120 g/L. After three months of incubation, the growth and [6]-gingerol production of the cultures were compared. Results showed an increasing number of microrhizomes formed in response to increasing concentration of sucrose from 30-90 g/L except at a higher concentration of 120 g/L which was found to be already inhibitory. Likewise, the highest [6]-gingerol production was observed in medium supplemented with 90 g/L sucrose suggesting a positive correlation of [6]-gingerol production with the number of microrhizomes. These results suggest that sucrose concentration can be manipulated to improve [6]-gingerol production in ginger tissue culture.

Induction of Direct Shoot Organogenesis from Shoot Tip Explants of an Ornamental Aquatic Plant, Cryptocoryne wendtii

Cryptocoryne wendtii is an important amphibious species with a wide range of foliage colors. Although it has a high market demand, the natural propagation of its aquatic species is limited due to the limited production on the number of plants with a long cultivation period, disease, and the requirement for a large space for propagation. Thus, we studied the effects of the plant growth regulators and their concentrations on the induction of direct shoot organogenesis from shoot tip explants of Cryptocoryne wendtii. The shoot tips were sterilized on its surface using 8 % Clorox® (5.25 % sodium hypochlorite, NaOCl) for 15 min followed by rinsing them three times with sterile distilled water. They were again sterilized on the surface for another 4 % Clorox® (5.25 % sodium hypochlorite, NaOCl) for 5 min. Cytokinins played a crucial role in direct shoot organogenesis. Direct shoot organogenesis from shoot tip explants was promoted by incubating these explants on Murashige and Skoog (MS) medium [1] in the presence of two different cytokinins [6-benzyl-aminopurine (BAP) or Kinetin (Kin)], each provided at four different levels. Direct shoot organogenesis was induced in both explants. Shooting occurred 100 % from in vitro shoot tip explants, which was cultured on MS medium and supplemented with 3.0 mg/l BAP. This was significantly different from the other treatments with the highest number of 16.20 shoots per explant and number of leaves at 72.40 leaves per explant after 60 days of culture. Individual shoots, aseptically excised, which produced normal roots within 45 days on the MS medium supplemented with α-Naphthaleneacetic acid (NAA). The highest number of roots per shoot and the longest roots were obtained on MS medium supplemented with 1.0 mg/l NAA (100 % rooting, which was an average of 36 roots per plantlet and root length at 26.02 mm). Rooted plantlets were successfully hardened and established in pots containing a mixture of organic soil and sand (1:1) overlaid with tap water under greenhouse conditions at 90 % survival. This complete study has successfully outlined a rapid, high frequency direct shoot organogenesis induction of an ornamental aquatic plant, Cryptocoryne wendtii from shoot tip explants inclusive of shoot proliferation, rooting and acclimatization. The present in vitro propagation protocol would facilitate an alternative method for rapid, large-scale production and germplasm preservation of this important endangered species C. wendtii.

Ascorbic Acid Controls Lethal Browning and Pluronic F-68 Promotes High-frequency Multiple Shoot Regeneration from Cotyldonary Node Explant of Okra (Abelmoschus esculentus L.)

The regeneration frequency of okra (Abelmoschus esculentus) is greatly influenced by its genetic makeup and recalcitrant nature. Phenolic secretion, in particular, is a major problem in okra tissue culture. This study describes a reproducible, rapid, and more efficient in vitro regeneration method using cotyledonary node explants of okra. Explants were incubated on Murashige and Skoog (MS) medium containing different concentrations and combinations of various plant growth regulators (PGRs) [benzyladenine (BA), thidiazuron (TDZ), and α-naphthylacetic acid (NAA)], and regeneration enhancers [silver nitrate (AgNO3) and Pluronic F-68]. Cut ends of cotyledonary node segments rapidly turned brown and cultures failed to establish. Antibrowning additives, such as activated charcoal (AC), ascorbic acid (AA), and AgNO3 at various concentrations in PGR-free MS basal medium were tested for their ability to control phenolic secretion from explants. Among these additives, 15 mg·L−1 AA was found to be optimal for controlling phenolic secretion, resulting in healthy explants and culture establishment. The highest number of shoots (a mean of 9.3 ± 0.9 shoots per cotyledonary node explant) was obtained on MS media containing 0.5 mg·L−1 NAA + 1 mg·L−1 TDZ + 0.1% Pluronic F-68. Individual shoots were elongated on MS medium + 1 mg·L−1 BA + 0.1 mg·L−1 gibberellic acid (GA3) (shoot length 5.3 ± 0.2 cm) and rooted on ½ MS medium + 1 mg·L−1 indole-3-butyric acid (IBA) and 200 mg·L−1 AC (5.3 ± 0.2 roots per shoot). Rooted plantlets were acclimatized in plastic pots inside a plant growth chamber at 25 ± 2 °C and 70% relative humidity, with an 80% survival rate. This optimized protocol can be used for producing transgenic plants of commercial okra cultivars through genetic transformation.

Genetic Diversity of Sorghum (Sorghum bicolor (L) Moench) for Agronomic Traits and Shoot Fly Tolerance and Suitability for Winter Season

Genetic diversity analysis of sorghum germplasm could provide useful information on selection of parental lines and diversifying the genetic base in winter sorghum wherein yield improvement has been marginal. An experiment was conducted on 4000 germplasm lines belonging to 28 countries of origin in augmented design in two winter sorghum growing regions of Maharashtra. Observations were recorded on yield parameters and traits governing shoot fly tolerance. Promising germplasm lines were identified for individual agronomic traits and shoot fly tolerance. The lines from Rwanda, Burundi and Kenya had more grain yield and the lines from India were tolerant to shoot fly. Though grain yield per plant was more variable among all the traits, the PCA analysis revealed that the panicle length, panicle width and shoot fly dead heart % contributed maximum towards divergence. A hierarchical cluster resulted in 11 clusters of originating countries with the accessions from Yemen, Gambia, Somalia, Mozambique, China and India clustering individually. Grain yield was positively associated with shoot fly resistance indicating possibility of co-evolution.

Improving In Vitro Somatic Embryos Production of Medjool and Khalas Date Palm Cultivars via Modification of Ammonium and Potassium Nitrate

Date palm is one of the most economically powerful fruit tree grown in Egypt. Although Egypt tops the list of date's product in the world. Egyptian date palm industry suffers from many constraints. Low-quality varieties and limited conventional propagation methods are the main obstacles. Extension of date palm plantations in various areas of Egypt and replanting trees to compensate loss due to diseases or human factors are reduced expected to lack adequate planting materials. The knowledge of the interplay between some fundamental medium parameters allows not only the optimization of the micropropagation system but also gives models to investigate and rationalize the process of induction and development of somatic embryogenesis itself. In vitro growth and development of produced embryos and plantlets has been improved by using of full MS salt strength. MS medium modification of some nutrients concentrations, i.e. NH4NO3(1237.5 mg/l) which recorded (13.20 and 9.60 embryos/explant for Medjool and Khalas respectively) at the end of three subculture. KNO3 which gave a higher score of mature somatic embryos (14.40 and 10.00 embryo/culture for Medjool and Khalas respectively) was obtained with 1425 mg/l KNO3 after three subcultures as compared to those obtained from any other treatment combination. The morphological response of somatic embryos production is controlled by some internal factors that fall under the influence of the genetic make-up of the plant and specialized for each genotype that is responsible for the production of somatic embryos (Medjool and Khalas cultivars). The genetic make-up is a decisive factor during somatic embryos production. There are differences between the two cultivars named, Medjool and Khalas. The produced somatic embryos at germination stage were transferred from the two experiments namely ammonium nitrate and potassium nitrate to multiplication and rooting stages for more in vitro growth for 12 weeks at three re-cultures. Individual shoots were cultured on modified MS basal medium in addition to IBA (1.0 mg/l), sucrose (30 g/l) and solidified with phyto-agar (8.0 g/l) for more in vitro growth and development. In vitro plantlets were transferred to acclimatization stage in plastic pots 5*18 cm diameter containing peatmoss, perlite and washed sand at equal volume. In vitro date palm plantlets produced from rooting stage grow well in the greenhouse during acclimatization stage without morphological abnormality. The growth and development of Medjool cultivar were better than Khalas during in vitro culture and ex vitro acclimatization.

Coping with waves: Plasticity in tidal marsh plants as self‐adapting coastal ecosystem engineers

AbstractTidal marsh vegetation is increasingly valued for its role in ecosystem‐based coastal protection due to its wave dissipating capacity. As the efficiency of wave dissipation is known to depend on specific vegetation properties, we quantified how these morphological, biochemical, and biomechanical properties of tidal marsh vegetation are, in turn, affected by wave exposure. This was achieved by field measurements at two locations, with contrasting wave exposure, in the brackish part of the Scheldt Estuary (SW Netherlands), where Scirpus maritimus is the dominant pioneer species. Our results show that shoots from more wave‐exposed conditions developed significantly shorter and thicker stems than the ones growing in more sheltered conditions. Furthermore, we show that the more exposed shoots are more flexible whereas the shoots growing in more sheltered conditions are stiffer. This may indicate plasticity in response to wave exposure following a stress‐avoidance strategy. Increasing stiffness was shown to be related to enhanced biogenic silica and lignin contents of the shoot tissue. These properties might affect the wave‐attenuating capacity of the marsh as stiff plants are known to mitigate waves more effectively than flexible ones. However, we also found higher shoot densities on the exposed site, which may partly explain why higher relative wave attenuation rates were found on the exposed site, despite the presence of more flexible individual shoots. This study highlights that the efficiency of wave attenuation by tidal marsh vegetation ultimately depends on mutual interactions between waves and plasticity in morphological, biochemical, and biomechanical plant properties.

Direct evidence of Hymenoscyphus fraxineus infection pathway through the petiole‐shoot junction

SummaryThe symptoms of ash dieback caused by the fungus Hymenoscyphus fraxineus include wilting of the foliage followed by dieback of shoots, twigs and branches. Necroses in shoots are assumed to develop after infection through leaf petioles; however, clear evidence of this infection pathway has not yet been provided. Considering the multiple pathogen genotypes in dead ash petioles, we aimed to obtain a spatial overview of all H. fraxineus genotypes colonizing individual shoots and their corresponding petioles before leaf shedding to acquire precise information about the infection biology of H. fraxineus and its ability to cross the petiole‐shoot junction. Individual genotypes of H. fraxineus were characterized by the analysis of microsatellites using DNA extracted directly from petiole segments or cultures isolated from the segments. We detected 150 different multilocus genotypes in 10 analysed shoots and their respective petioles; the highest number of genotypes was eight for a single petiole and three for a single shoot. The genotypes of most shoot lesions were identical to particular genotypes from the proximal segments of petioles, implicating the main pathway of shoot infections. To test whether the amount of colonized substrate or intraspecific competition have an effect on successful infection, genotypes that reached the most proximal end of the petioles were scored for the number of invaded petiole segments and for the number of other H. fraxineus genotypes co‐occurring in the segments. However, the extent of colonization of the scored genotypes and intraspecific competition with other H. fraxineus strains did not influence pathogen success in entering the shoot. This study confirms that the majority of ash shoot infections are caused by genotypes of H. fraxineus originating from petioles. Compared to petioles, the frequency of shoot colonization as well as number of H. fraxineus genotypes in shoots was much lower.

Detection of viruses of Bangladeshi and Japanese garlic and their elimination through root meristem culture

A number of viruses cause considerable yield loss and quality deterioration in garlic. Root meristems of virus infected plants are known to be free from detectable viruses. This potentiality could be exploited to obtain virus free clones at a high frequency by culturing excised root meristems in vitro. We have developed efficient methods of direct and somatic embryo derived shoot regeneration from root meristems of garlic. The objectives of this work were to detect viruses infecting Bangladeshi and Japanese garlic clones and find an easy and efficient method of eliminating the viruses for the improvement of both yield and quality of garlic. At first, we confirmed the presence of detectable viruses in three Bangladeshi and one Japanese clones. The clones were infected with four different types of viruses: Garlic viruses (GarVs), Onion yellow dwarf virus (OYDV), Leek yellow stripe virus (LYSV), and Garlic common latent virus (GCLV). To eliminate those viruses, as per our previous method, root meristems were cultured on MS medium supplemented with 1.0 µM NAA and 10.0 µM BA. Shoot primordia developed from the cultured explants within 1 month. The regenerated individual shoot buds (2-5 mm) were separated from the mother explants and transferred to growth regulators free medium. RT-PCR confirmed that the viruses present in the mother garlic plants were absent in the shoots found after two-step culture. The regenerated shoots were rooted on growth regulator free medium and transferred to pots. Results indicated that the plants remained free from LYSV. Virus elimination through root meristem culture emerged as an efficient novel technique for the eradication of multiple viruses as confirmed by RT-PCR in this study. This technique has the potential for the production and supply of virus free propagules (plants/bulblets) for the yield and quality improvement of garlic.Progressive Agriculture 28 (2): 55-63, 2017

Rapid plant regeneration, validation of genetic integrity by ISSR markers and conservation of Reseda pentagyna an endemic plant growing in Saudi Arabia.

Reseda pentagyna is the only endemic species among the seven species of the genera Reseda found in Saudi Arabia. Probably no information is available on regeneration by conventional method of regeneration through seeds or cuttings. Therefore, alternative method of tissue culture was attempted to regenerate and multiply the plant. High shoot regeneration (14.44 shoots/explant) was obtained after four weeks, when shoot cuttings cultured on MS containing BA at 1.0 µM. Other cytokinins e.g., Kn, 2iP and TDZ found to be less effective in bud induction and shoot multiplication. Individual shoots were rooted on MS medium supplemented with various auxins at 0.5–5.0 µM concentrations. The IBA (1.5 µM) supplemented MS media induced maximum (83.3%) rooting. The plantlets were acclimatized and hardened under greenhouse conditions in plastic pots containing soil and farm yard manure with 95.0% success. The protocol developed would help to multiply the plant as well as conserve them in natural habitat. This can also be utilized to obtain active constituents for pharmaceutics and genetic manipulations.

Effect of Altered Canopy:Root Volume Ratio on Grapevine Growth Compensation

Nine-year-old, intensively irrigated, 3.0 x 1.2 m spaced and vertically trellised (5-strand) Chenin blanc/99 Richter vines were converted to double the original cordon length by either removing alternate vines or by implementing a Lyre trellising system. In the former case the root volume was doubled, whereas in the latter case it was kept the same as for the non-converted vines. After five years, a root profile study showed that roots of the single-cordoned, vertically trellised vines with extended cordon were distributed beyond the original soil volume (before conversion); this was not the case for the Lyre system. No difference in root density occurred between vines of converted systems. The available soil volume was apparently better colonised by roots of the converted systems, mainly because of an increase in fine roots. The greatest reduction in individual shoot growth was found for vines trained to the Lyre system. Yield per vine varied according to cane mass. Yield per hectare of vines trained to the vertical trellis with extended cordon space and the Lyre system increased significantly. Yield:cane mass ratios of 11, 15 and 16 were found in the case of the vertical, the vertical with extended cordons and the Lyre systems, respectively. The results indicated that shoot growth was better accommodated and distributed by extending cordon length, particularly when the ratio of cordon length to root volume was increased (as for the Lyre system). Compensatory growth occurred when both above-ground and subterranean growth volumes were increased. Preventing compensatory growth by the root system, resulted in balanced growth and improved microclimatic conditions for pest and disease control and grape ripening. This principle may be implemented as a long-term solution to problems associated with excessive vigour.

A novel technique for Musa acuminata Colla ‘Grand Naine’ (AAA) micropropagation through transverse sectioning of the shoot apex

A new approach is described for in vitro shoot proliferation using transverse sections of individual shoot apices as explants, and its efficiency was compared to conventional shoot apex culture of banana ‘Grand Naine’. Individual shoot apex explants were inoculated on Murashige and Skoog (MS) media supplemented with 0, 10, 20, and 30 μM of 6-benzylaminopurine (BAP) in combination with 1.0 μM α-naphthaleneacetic acid (NAA) and maintained for 6 mo with monthly subculture. Shoot proliferation increased at each subculture to a maximum of 23.65 shoots on 20 μM BAP-supplemented medium. To increase the proliferation rate, individual shoot apices cultured on MS media containing 10 μM BAP were split into three transverse sections, and the middle section produced the highest number of shoots (7.22). These shoots, when cultured on MS medium supplemented with 20 μM BAP, produced 8.26 shoots per explant vs. 6.18 and 7.94, respectively, for media supplemented with BAP at 10 or 30 μM. From just one transverse section, 32.77 shoots could be produced after 6 mo vs. 11.79 shoots via direct regeneration from individual shoot apex explants. MS medium supplemented with 6 μM IBA induced vigorous rooting within 4 wk. Acclimatization was achieved under greenhouse conditions, and the highest survival rate was 88.63%. Randomly amplified polymorphic DNA (RAPD) analysis showed that plants from shoot apex cultures had 23.46% polymorphism, while those from transverse sections showed no genetic variation when compared to the parent plants.

Sodium nitroprusside stimulates growth and shoot regeneration in chrysanthemum

In this study, we demonstrate that the nitric oxide (NO) donor sodium nitroprusside (SNP) improves plant regeneration in chrysanthemums. Internode explants of three different chrysanthemum cultivars, ‘White ND’, ‘White wing × Peach ND’, and ‘Hunt × Lemon ND’, were cultured on shoot induction medium (SIM) containing various concentrations of N6–benzyladenine (2.22 or 4.44 μM BA) and SNP (0.83–6.71 μM) individually and in combination. Most combinations of BA and SNP significantly improved the morphogenetic potential of internode explants and enhanced shoot regeneration in all three chrysanthemum cultivars compared to treatment with BA alone. The cultivar ‘White wing × Peach ND’ displayed the highest regeneration response (98.3%) and shoot regeneration rate (27.3 shoots/explant) in SIM containing optimal BA (4.44 μM) and SNP (0.83 μM) concentrations within 30 days of culture. Individual shoots of ‘White wing × Peach ND’ were transferred to root induction medium (RIM) containing various concentrations of SNP (0.83–6.71 μM) alone. Shoots rooted in the presence of SNP resulted in healthy plantlets within 30 days of culture with improved root (number of roots/shoot, root length, and fresh weight) and shoot (number of leaves, shoot length, and fresh weight) growth characteristics compared to the control. In addition, the regeneration procedure described in this study only requires a short duration (60 days) to obtain rooted plantlets from internode explants of chrysanthemums. Our results suggest that supplementation of chrysanthemum regeneration medium with SNP enhances shoot regeneration and improves plant growth, overcoming problems associated with propagation and genetic transformation.

Direct <i>in vitro</i> Regeneration of <i>Nicotiana plumbaginifolia</i> L. and the Potential for Genetic Transformation

Wild crop relatives are important source of genetic diversity. Plant tissue culture provides proficienct means of conservation of endangered wild plants sustaining biodiversity, improving plant traits by virtue of induced somaclonal variation and trangensis. The objective of study was to establish direct in vitro regeneration and biolistic transformation protocol for wild tobacco N. plumbaginifolia using uidA gene. For direct in vitro regeneration, six combinations of IAA and kinetin were used. Then, biolistic transformation was carried out using BioRad PDS 1 000 (He) gun and 1 100-psi rupture discs. Transformation was confirmed through histochemical gus assay. A proficient and reproducible in vitro regeneration system was standardized for wild tobacco named as Nicotiana plumbaginifolia . In present study 1 mg/L kinetin in combination with 0.25 mg/L IAA proved best for direct in vitro shoot regeneration from mature leaf-based explants. Success in direct in vitro regeneration circumvented the callogensis step, accelerating the process of in vitro regeneration and increasing the chances of obtaining true to type plants. Individual shoots were placed on MS medium with/without IBA for rooting. Individual shoots were successfully rooted on MS medium without IBA. Then, explants were transformed with uidA gene using gene gun. Transient assay of bombarded calli showed gus activity on histochemical assay even after a week. Analysis of gus expression has been found as a principal method of plant transformation optimizations.

Effect of Explants Length and Segmentation and Separated and Cluster of Shoots on the in vitro Shoot Formation and Growth of Pineapple (Ananas comosus L. Merr.)

Two experiments were conducted. In one, multiple shoots complex from Smooth cayenne pineapple stock cultures were cut into cluster of two, three, four shoots and were also separated into individual shoots of different length 5, 10, 15, 20 and 25 mm long and cultured on agar solidified MS medium enriched with benzylaminopurine (BAP) at 2.23 mg/l. In the other one, 25 mm long shoots were cut longitudinally into halves and quarters and cultured at density of one explant and 15 mm long shoots were cultured intact at density of one, two, three and four explants per culture. Longer (25 mm) intact and halved explants produced the highest (10 shoots) while 5 and 10 mm long explants resulted in lowest (6 shoots) and 15 and 20 mm long explants resulted in an intermediate (8 shoots) shoot formation. Shoot formation whether the explants were cultured as separated intact or as cluster of shoots decreased from 10 to 4 shoots per explant as the density increased from one to four. Shoot length was not affected by using of separated intact explants of different length. However, increasing the explants density increased the shoots length particularly when cluster rather than separated shoot were used as explants.

Clonality increases with snow depth in the arctic dwarf shrub Empetrum hermaphroditum.

Vegetative reproduction and spread through clonal growth plays an important role in arctic-alpine ecosystems with short cool growing seasons. Local variation in winter snow accumulation leads to discrete habitat types that may provide divergent conditions for sexual and vegetative reproduction. Therefore, we studied variation in clonal structure of a dominant, evergreen, dwarf shrub (Empetrum nigrum s.l. with the two taxa E. nigrum L. s.s. and E. hermaphroditum Hagerup) along a snow cover gradient and compared clonal diversity and spatial genetic structure between habitats. We studied 374 individual shoots using 105 polymorphic AFLP markers and analyses based on hierarchical clustering, clonal diversity indices, and small-scale spatial genetic structure with pairwise kinship coefficient. We used two approaches to define a threshold of genotypic distance between two samples that are considered the same clone. Clonality was examined among three habitats (exposed ridges, sheltered depressions, birch forest) differing in snow conditions replicated in four study regions in Norway and Sweden. Clonality of E. hermaphroditum differed between habitats with an increase in clonal diversity with decreasing snow depth. Small-scale spatial genetic structure increased with decreasing clonal diversity and increasing clone size. In three study regions, E. hermaphroditum was the only species, whereas in one region E. nigrum also occurred, largely confined to exposed ridges. Our results demonstrated that snow cover in conjunction with associated habitat conditions plays an important role for the mode of propagation of the dwarf shrub E. hermaphroditum.

Revisiting Domestication to Revitalize Crop Improvement: The Florigen Revolution

Most flowering plants have evolved to coordinate proper floral transition in shoot apical meristems at an appropriate time during development, referred to as “flowering time.” In domesticated crops, shoot life time has been manipulated to enhance productivity via synchronizing flowering time to the local environment, thereby creating a balance between vegetative and reproductive plant architecture during the year. Rice, a typical single transitional crop, has acquired balance between prolonged tiller growth and a relatively short reproductive phase on individual shoot, resulting in increasing yield. Crops with sympodial growth, such as tomato, which undergo multiple floral transitions on the main shoot, have been artificially selected for the determination of shoot growth by investigating the dosage sensitivity of florigen activation controlling flowering time on individual shoots. Here, we summarize recent genetic-molecular studies on tomato, which have been subject to genetic manipulation of flowering time in an effort to optimize seed productivity. We also review advanced genetic approaches as potential new tools for the enhancement of crop yield by manipulating flowering time via the dosage effect of florigen.