Efficient Regeneration of Hedychium coronarium through Protocorm-Like Bodies

Hedychium coronarium J. Koenig is a multipurpose plant with significant economic value, but it has been overexploited and listed as a vulnerable, near threatened or endangered species. In vitro culture methods have been used for propagating disease-free propagules for its conservation and production. However, explant contamination has been a bottleneck in in vitro propagation due to the use of rhizomes as the explant source. Plants in the family Zingiberaceae have pseudostems that support inflorescences, while rhizomes are considered true stems. The present study, for the first time, reported that the pseudostem bears nodes and vegetative buds and could actually be true stems. The evaluation of different sources of explants showed that mature node explants derived from the stem were the most suitable ones for in vitro culture because of the lowest contamination and the highest bud break rates. Culture of mature node explants on MS medium supplemented with 13.32, 17.76, and 22.20 μM 6-benzylaminopurine (BA), each in combination with 9.08 μM thidiazurin (TDZ) and 0.05 μM α-naphthaleneacetic acid (NAA) induced the conversion of buds to micro-rhizomes in six weeks. More than 96% of the micro-rhizomes cultured on MS medium supplemented with 17.76 μM BA, 6.81 μM TDZ, and 2.46 μM indole-3-butyric acid (IBA) were converted to globular-shaped clumps with protocorm-like bodies (PLBs). Further culture of a piece of the clumps induced more than 15 adventitious shoots. Adventitious roots were produced at the base of adventitious shoots, and plantlets were readily transplanted to a substrate for acclimatization in a shaded greenhouse. The survival rate of the plants in the greenhouse was up to 90%. Plants grew vigorously, and there were no off-types from the regenerated 11,100 plants. Our study also, for the first time, shows that H. coronarium can be regenerated via PLBs, which may represent a new way of the in vitro propagation of H. coronarium. The established protocol could be used for the increased propagation of H. coronarium for conservation or commercial production.

Identification of Best Surface Sterilization Treatment and Control of Endophytic Bacterial Contamination in Annona squamosa L.

Surface sterilization is most important step in plant tissue culture protocol. In the present investigation, an attempt was made to eliminate microbial and fungal contaminants from the surface and interior of plant material, thus obtaining axenic culture with highest survival rate. Sequential surface sterilizations of hypocotyl, leaf, shoot tip and mature node were carried out to investigate its effectiveness in controlling surface contamination with satisfactory survival of explants. Combination of different surfactant were used for surface sterilization treatments. The least contamination was obtained when hypocotyl explants were treated with 200 ppm cefotaxime and 500 ppm carbendazim along with 0.1% HgCl2 with best survival percentage. Treatments consisting of alcohol treatment, carbendazim (2000 ppm) followed by 1000 ppm cefotaxime, 500 ppm kanamycin, 2% sodium hypochloride and 0.1% HgCl2 sequentially resulted in complete elimination of surface contaminants from shoot tip, soft node and hard node obtained from field grown mature tree. Optimal elimination of bioburden from young leaf (77.38%) were obtained using 1000 ppm carbendazim, 500 ppm cefotaxime, 500 ppm kanamycin and 0.1% HgCl2. Gentamicin used in the medium was able to control the endophytic bacterial bioburden completely in the first cycle of 15 days itself at higher concentration of 96 mol/l to remove endophytic bacterial contamination with out effecting plant growth.

A primary requirement for nodal in the formation and maintenance of the primitive streak in the mouse

The 413.d insertional mutation arrests mouse development shortly after gastrulation. nodal, a novel TGF beta-related gene, is closely associated with the locus. The present study provides direct evidence that the proviral insertion causes a loss of function mutation. nodal RNA is initially detected at day 5.5 in the primitive ectoderm. Concomitant with gastrulation, expression becomes restricted to the proximal posterior regions of the embryonic ectoderm. nodal RNA is also expressed in the primitive endoderm overlying the primitive streak. A few hours later, expression is strictly confined to the periphery of the mature node. Interestingly 413.d mutant embryos show no morphological evidence for the formation of a primitive streak. Nonetheless, about 25% of mutant embryos do form randomly positioned patches of cells of a posterior mesodermal character. Data presented in this report demonstrate the involvement of a TGF beta-related molecule in axis formation in mammals.

Membrane Specializations in Developing Nodes of Ranvier

Mature myelinated nerve fibers exhibit distinctive structural features at nodes of Ranvier and the adjacent paranodal regions. In order to obtain information about the interrelationships between these specializations during development, thin sections and freeze-fracture replicas of immature peripheral nerve fibers from grass frog tadpole hind legs were examined during the period of myelinogenesis. Early in myelination axons are enwrapped individually by a few loose Schwann cell layers whose edges overhang each other forming "terminal loops" against the axolemma. Unlike those of the mature node, these loops are widely sepa-rated and irregularly spaced (Fig.l), and similarly the presumptive nodal region between successive developing myelin segments is usually much longer than adult nodes of Ranvier. The presumptive nodal axolemma may exhibit a cytoplasmic "undercoating." However, the density of this coating is highly variable. Usually it is much lower than at adult nodes, and in some cases the undercoating is not distinguishable. The outermost layers of the Schwann cell are usually the first to form axoglial junctional specializations character¬ized by the presence of "transverse bands" and ER cisternae applied to the junctional Schwann cell membrane. In some instances the outermost layer con¬tacts the axon over an extensive area and forms multiple small junctional specializations at widely separated intervals along the length of the axolemma.

Aspects of Vascular Anatomy and Differentiation of Vascular Tissues and Transfer Cells in Vegetative Nodes of Wheat

Xylem transfer cells are strongly developed in the departing leaf traces of the mature wheat node. Their differentiation is initiated soon after the appearance of the first tracheary elements in these bundles. and wall ingrowth development reaches its peak just as the leaf to which the bundle belongs becomes fully expanded. It is suggested that the xylem transfer cells play an important role in redirecting solutes travelling in the xylem of the mature leaf to the developing leaves at the shoot apex. It is further suggested that they form an integral part of the normal xylem transpiration pathway, compensating for xylem restrictions and discontinuities in the mature node. Phloem transfer cells also appear very early in the differentiation of the nodal vasculature, although their function remains obscure.