Propagation of ashwagandha is done mainly by seed but seed viability is limited to one year.As the viability of seeds is less and because of viral attacks on seedlings, there is overwhelming demand for an alternative to traditional propagation method of cultivations. Tissueculturetechniques can play an important role in conservation, clonal propagation and qualitative improvement of this medicinally important plant (Sivanesan 2007).
Growth and development response of callus segments of I. gabonensis were examined using tissueculturetechniques. Explants were aseptically placed and inoculated on Murashige and Skoog (MS) callus induction medium of full-MS, 1/2 MS and 1/4 MS strengths, each supplemented with the following concentrations of plant growth regulators (PGR); To (control) - treatment without plant growth regulator, T1 - treatment containing 0.1 mg/l kinetin + 2.0 mg/l NAA, T2- treatment containing 0.2 mg/l kinetin + 4.0 mg/l NAA, T3- treatment containing 0.3 mg/l kinetin + 6.0 mg/l NAA, T4- treatment containing 0.4 mg/l kinetin + 8.0 mg/l NAA and T5- treatment containing 0.5 mg/l kinetin + 10.0 mg/l NAA. The fresh weight, dry weight, moisture contents, chlorophyll a and b contents of callus segments of the species significantly (P< 0.05) increased with increase in concentration of plant growth regulators and with decrease in medium strength. The optimum growth and development response were recorded at one quarter strength medium relative to full and half strengths media. Therefore, this study provides baseline information for optimum callus induction protocols and continuous domestication of this over exploited species.
ABSTRACT: Despite extensive research done on nuclear embryogenesis in citrus, a major snag remains unsolved that the nuclear embryo proliferation does not lead to production of even a fraction of the number of globular embryos formed initially. During further proliferation, mostly normal differentiation of embryos gets drowned in the mass of proliferating cotyledonary structures. The present experimentation is mainly aimed to resolve this problem and produce sufficient number of nucellar plantlets per culture. The significance of nucellar embryogenesis, particularly in case of rootstocks can hardly be over emphasized. However, it is also significant in case of C. jambhiri, where commercial use of its nucellars for raising orchards is in practice. Nucellus is a somatic tissue having a genetic makeup similarly to the vegetative plant body. The culture of nucellar tissue and augmentation and induction of nucellar polyembryony have initially been the main focus of research. However, the associated extended juvenile characteristics of nucellar plantlets, if not slightly poor quality of fruit, restrict the application of in vitro nucellar embryogenesis to rootstocks only, since both such characteristics will not be of any consequence while virus-free rootstocks will be produced at a rapid rate throughout the year, which in turn is imperative for all the year round production of micrografts. When critically examined, the nucellar embryonal proliferation, i.e., embryo-to-embryo multiplication is an uncommon phenomenon. Instead, there is cotyledonary proliferation forming resette-like structures having a single radicle with pluricotyly, which is a big snag in production of nucellar plantlets.
cotyledon-like-structures and their fasciation taking over the normal differentiation and proliferation of embryos. The point became more explicit when the bicotyledonary structures were taken out from such cultures and subjected to germination in comparison to a few normal embryos. BAP along with putrescine deteriorated the response of putrescine used alone in respect of normal embryo formation per culture, while Z at any of the 3 concentrations used inhibited the process of normal differentiation of embryos. Details of responses evidenced by proliferating globular nucellar embryos to different concentrations of putrescine used alone and along with 3 cytokinins, viz., 2iP, BAP and Z. In the optimum treatment containing 0.1 mg l -1 2iP, 0.5 mg l -1 putrescine and 0.1 mg l -1 NAA, mostly the proliferating embryos remained isolated as if there had been embryo-to-embryo proliferation. Occasionally, the embryos were also seen in groups, but did maintain their individual entity with cotyledonary and radicular ends. Besides, embryo-to-embryo proliferation, there was another mode of multiplication of embryos, in which the secondary embryos differentiated from the hypocotyls region of earlier formed developed embryos. The deceptive impression given by the cultures of cotyledon-like-structures as if the 2 proximally united cotyledonary structures were destined to produce plantlets was obvious when they were isolated and cultured in the embryo germination medium. From such aberrant embryos, radical like structures were formed but their was no formation of shoots from the so called plumular ends. On the other hand, the normal embryos picked up from amongst proliferating and developing normal embryos readily germinated giving rise to root and shoot from opposite poles.
Embryo culture is the most successful technique of the tissueculturetechniques used by plant breeders. Embryo abortion is one of the common cases that occur after cross breeding between species and sometimes after cross breeding between species. In such cases, hybrid seeds unless the embryo is isolated in early stages and grown on a diet to grow and develop into a plant. In cases where the abortion takes place at very early stages after direct fertilization, in which the embryo cannot be isolated because of its small size, the researchers isolate the whole ovule and fertilize it media appropriate until the development of the embryo. Ovules technology is also used to eliminate incompatibilities, especially in genetically divergent species. Pollen may mature at an early time before mites are ready to be vaccinated. The vaccine tube does not grow on the flowerbed due to the presence of some inhibitors in the flowering season. In vitro pollination and fertilization this problem can be overcome by isolating the oocytes and then fertilizing them with pollen from the desired plant in order to obtain the seeds of live embryos that grow later to seedlings or ovarian implants can be used in studies that relate to the appearance of fruits separation , stages of development and food needs (8).
G. superba L. is a medicinal climber, its seeds and tubers contain valuable alkaloids viz., colchicine and colchicoside as the major constituents, which are used to treat many diseases (Chitra, 2010). G. superba usually multiply by corm and seeds but due to low germination capability it restricts for the regeneration. G. superba is a commercially imperative medicinal plant which has diverse medicinal applications and eventually due to over-exploitation this plant is facing local extinction. Some time plant tissueculturetechniques play a key for conservation of this plant. At present Plant tissueculture offers a valuable to overcome the problem regarding conventional propagation, and obtain disease free healthy plants (Neha Bhagat, 2011). Its seeds have poor germination and low availability, while propagation by corm also a limiting factor, making micropropagation an essential proposition in order to meet the demand for quite a huge amount for raw material by pharmaceutical industries (chaturvedi 2007). Plants have been regenerated from somatic embryogenesis, caulogenesis, direct plantlet formation and regeneration of shoot buds
Even with a sound methodology on rice tissueculture, in vitro regeneration of indica rice is still an interesting task and is genotype-dependent (Kumria et al., 2000; Lin and Zhang 2005, Zhang et al., 2005; Ge et al., 2006; Pazuki and Sohani 2013). The genotype and composition of the nutrient media are the two major factors in the tissueculture of rice (Mikami and Kinoshita 1988; Khanna and Raina 1998; Ogawa et al., 1999; Gul et al., 2000; Al-Forkan et al., 2005). In rice, the scutellum is the best explant for callus induction and plant regeneration (Rashid et al., 2001, 2004; Cho et al., 2004; Khaleda and Al-Forkan 2006). Due to a low rate of embryogenic callus induction and plantlet regeneration, the transformation of indica rice is still difficult (Lee et al., 2002; Kumar et al., 2005; Nishimura et al., 2006). For carrying out successful genetic transformation in rice, identification and screening of useful cultivars and establishment of efficient regeneration protocols are very essential (Hoque and Mansfield 2004; Dabul 2009; Joyia and Khan 2012). This review will summarize various treatments used to carry out a successful tissueculture of diverse rice varieties.
Embryonic cells can activate random motility using various genetic signals, and this seems to be a prerequisite for initiation of directional migration in response to external guidance cues (Aman and Piotrowski, 2010). Random motility results in random patterns of cell migration, which per se can impact cell and tissue morphogenesis. In the context of individual cell migration, random motility has a dispersive and exploratory effect that prompts cells to colonise new territories within the embryo. Such an effect is characteristic of the early migratory phase of the zebrafish endoderm. During gastrulation, zebrafish endodermal cells disperse from the margin towards the animal pole following an intrinsic cell-autonomous ‘ random walk ’ , which is characterised by the formation of short-lived small protrusions on cells in almost all directions (Fig. 5A, top) (Pezeron et al., 2008; Woo et al., 2012). This random migratory behaviour later switches to become persistently directed as endodermal cells start converging towards the midline in response to chemotactic cues (discussed below), while forming broader and more stable dorsally directed protrusions (Fig. 5A, bottom) (Pezeron et al., 2008; Woo et al., 2012). Random walking is induced by Nodal signalling at least in part through the expression of the Rac activator Prex1 (Woo et al., 2012): when Rac1 activity is disrupted, endodermal cells anticipate their dorsal-directed migration and, as a consequence, become mis- localised and in some cases fail to maintain their original fate to become mesoderm (Woo et al., 2012). These observations suggest that, in addition to its role in cell dispersion, random motility might also serve as a control mechanism for the response of the cell to guidance cues, generating a ‘ noise ’ that reduces the ability of cells to respond to weak environmental guidance cues that have the potential to interfere with their normal development (Woo et al., 2012).
al., 2006 and Bao et al., 2006), shoot tip (Sebastian 1983 and Aasim 2009), cotyledon (Brar et al., 1999, Chaudhary et al., 2007, Mao et al., 2006, and Pellegrineschi., 1997) and shoot meristem (Kartha et al., 1981 and Manoharrain 2008) have been employed for plant organogenesis in cowpea. To know the influence of culture medium, Diallo et al., (2008) employed two types of basal media (B5 and MS) to regenerate the cotyledonary node explants. They have reported that MS basal medium is efficient for shoot elongation and MS medium for multiple shoot formation. The most common cytokinins were 6- benzylaminopurine (BAP), Kinetin (KI) and Thidiazuron (TDZ). The effect of different concentrations of BAP 2.5-10µm on the regeneration of cotyledonary node was reported by Chaudhury et al., (2007).
Int 1 De\' 1Iiol 15 2S9 2~3 (1991) 259 Peroxidase as a developmental marker in plant tissue culture MARIJANA KRSNIK RASOL Department of Molecular Biology, Faculty of Science, University of Zagreb, Rep[.]
In a first step we employed mouse fibroblasts (cell line L929) that were cultured for 7 days on specifically tuned nanotube arrays tube diameter (32 ± 3) nm) as model system to address within an environmental scanning electron microscope (ESEM) study cell adherence, lift-off and nanotube regeneration. In fact, the L929 cell line is widely used and due to their easy handling we can ensure almost identical nanotube-cell-samples for investigations of different nanotube regeneration ap- proaches in a well-controllable and reproducible man- ner. Integrity of cells on top of the nanotube arrays was characterized using the ESEM and compared to cells in culture dishes as reference (not shown here). Both cul- ture substrates have the same growth area of 1cm 2 . After 1, 4 and 7 days, nanotube samples were imaged with the ESEM, as shown in Fig. 1. Within 24 h, cells are well stretched, not rounded up and adhered well to the nano- tube array, as can be concluded from formation of la- mellipodia. After 4 days in culture, cells arrange in groups and form an aggregate which makes it difficult to distinguish individual cells from each other. With
microspheres from each set were added to every well of a Costar white polystyrene, 96-well plate. Next, 96 unique, normal human serum samples (Covance, Princeton, NJ) were diluted 1:5 in PBS-BT. Eight of these serum samples were randomly selected and received a serial dilution of LASV antigen (tissueculture supernatant). These 96 samples (88 normal, 8 spiked) were added individually to the 96 wells containing microspheres. The plate was incubated at RT for 1 h with shaking, and then washed three times with PBS-BT. After washing, each well received a cocktail of two biotinylated detector MAbs diluted to 4 g/ml with PBS-BT. One MAb was speciﬁc for nucleocapsid protein (NP; L52-189-13) and the other was the same M13 MAb (27942001; GE Lifesciences) used in microsphere coating. The plate was incubated at RT for 1 h with shaking, and then washed three times with PBS-BT. Lastly, all plates received a streptavidin, R-phycoerythrin conjugate (Life Technologies) diluted to 4 g/ml with PBS-BT, were incubated at RT for 30 min with shaking, and then washed three times with PBS-BT. The signal generated by each well was measured with the Luminex MagPix instrument. For a result to be valid, a minimum of 50 beads/well had to be detected by the system. The LASV-NP/M13 MFI ratio for each sample was obtained. The values are expressed as Z-scores, using the means and standard deviations of all ratios from the experimental population.
Advances in biotechniques, particularly methods for culturing plant cell cultures should provide new means for the commercial processing of even rare plants and the chemicals they provide .The advantage of this method is that it can ultimately provide a continuous, reliable source of natural products .The major advantage of the cell cultures include synthesis of bioactive secondary metabolites, running in controlled environment, independently from climate and soil conditions. The use of in vitro plant cell culture for the production of chemicals and pharmaceuticals has made great strides building an advances in plant science. The increased use of genetic tools and an emerging picture of the structure and regulation of pathways for secondary metabolites will provide the basis for the production of commercially acceptable levels of product. The increased level of natural products for medicinal purposes coupled with the low product yields and supply concerns of plant harvest has renewed interest in large-scale plant culture technology. Knowledge of biosynthetic pathways of desired phytochemicals in plants as well as in culture is often still in its infancy, and consequently strategies needed to develop an information based on a cellular and molecular level. These results show that in vitro plant cell cultures have potential for commercial production of secondary metabolites. The introduction of newer techniques of molecular biology, so as to produce transgenic cultures and to effect the expression and regulation of biosynthetic pathways, is also likely to be a significant step towards making cell cultures more generally applicable to that commercial production of secondary metabolites.
In the present meta-analysis, four studies were pooled to evaluate the diagnostic value of periprosthetic tissue in BCB for PJI to provide further evidence for its clinical use. Among the included studies, sensitivity ranged from 0.66 to 0.75 while specificity ranged from 0.95 to 0.98. Analysis of periprosthetic tissue can be valuable for the diagnosis of PJI. Although the histology provides a higher sensitivity than that of tissueculture [5, 20, 21], it does not isolate microorganisms. Tissueculture is a common method for the microbiological diagnosis of PJI. Recently, the new technique of periprosthetic tissueculture in BCB has been used in the clinical setting. The authors in one study found that periprosthetic tissueculture in BCB offered better culture results than trad- itional medium culture methods. The sensitivity of peri- prosthetic tissueculture in BCB (87%) is higher than direct solid media (agar plates), cooked meat broth, and fastidious anaerobic broth (39, 83, 57%, respectively) . One reason for the lower sensitivity of the conven- tional method, may be due to the incubation period of only 5 days of these growth media. Fink and colleagues showed higher sensitivity of tissueculture inform pa- tients with hip and knee PJI (73 and 78%, respectively) when the incubation was prolonged to 14 days [22, 23]. In another study investigating the arthroscopic tissue bi- opsy in hip PJI, the culture sensitivity was reported even higher (87.5%), along with a specificity of 100% . In a
molecular biology have opened up new vistas for introducing foreign genes into crop plants. These techniques may not only facilitate the transfer of cloned, well-defined single genes or parts of genes (Cocking et al., 1981; Schell et al. 1982), but also allow gene transfer from completely unrelated species. They may also enable the modification or replacement of undesirable genes (Shillito et al 1986), and thus introduce new economic potential to cultivated plants. Foreign genes have been transferred /incorporated through Agrobacterium as well as through direct DNA transfers methods (Uchimiya, et al., 1989).