Gibberellins (GAs) control many aspects of plant development, including seed germination, shoot growth, flower induction and growth and fruit expansion. Leaf explants of Solanum nigrum (Black Nightshade; Solanaceae) were used for Agrobacterium-mediated delivery of GA-biosynthetic genes to determine the influence of their encoded enzymes on the production of bioactive GAs and plant stature in this species. Constructs were prepared containing the neomycin phosphotransferase (nptII) gene for kanamycin resistance as a selectable marker, and the GA-biosynthetic genes, their expression under the control of the CaMV 35S promoter. The GA-biosynthetic genes comprised AtGA20ox1, isolated from Arabidopsis thaliana, the product from which catalyses the formation of C19-GAs, and MmGA3ox1 and MmGA3ox2, isolated from Marah macrocarpus, which encode functionally different GA 3-oxidases that convert C19-GAs to biologically active forms. Increase in stature was observed in plants transformed with AtGA20ox1, MmGA3ox2 and MmGA3ox1 + MmGA3ox2, their presence and expression being confirmed by PCR and RT-PCR, respectively, accompanied by an increase in GA1 content. Interestingly, MmGA3ox1 alone did not induce a sustained increase in plant height, probably because of only a marginal increase in bioactive GA1 content in the transformed plants. The results are discussed in the context of regulating plant stature, since this strategy would decrease the use of chemicals to promote plant growth.

Podophyllotoxin is obtained from the rhizomes and roots of wild populations of Podophyllum hexandrum Royle. This is a low-growing plant with a long juvenile phase, making the availability of this natural product limited (1). Demand for podophyllotoxin was created with the introduction of its semi-synthetic derivatives in cancer chemotherapy (2). The species is endangered in the Himalayan region (3) through over collecting and lack of organized cultivation (4). Efforts remain to facilitate the in vitro propagation (5, 6) of Podophyllum. In the present investigation on the tissue culture of P. hexandrum, seeds germinated within 35 to 40 days in moist, dark conditions, with in vitro grown seedlings being obtained either on 0.2 normal strength semi-solid B5 medium (7) or full-strength MS medium both lacking growth regulators. Although callus induction from root explants cultured on 0.5 normal strength B5 medium containing 1.0 mgl-1 2,4-D, 1.0 mgl-1 BAP and 1.0 mgl-1 GA3 was slow, tissue became embryogenic after sucessive subcultures. Embryogenic cell suspensions were established in the dark from root-derived callus, cultured in liquid MS medium containing 2,4-D and kinetin, at 2.0 mgl-1 and 0.25 mgl-1, respectively. Differentiation of somatic embryos and subsequent shoot formation occurred on either full-strength or half-strength MS medium with 0.45 mgl-1 BAP. Rooting of somatic embryo-derived plants was stimulated by the inclusion of 10-5 M lipo-oligosaccharide in the culture medium. A robust explant-to-plant micropropagation system for Podophyllum will reduce the pressure on wild resources and may offer an alternative source of podophyllotoxin production.

High frequency direct plant regeneration from leaf and petal explants was accomplished for the first time in Streptocarpus varieties. The shoot induction frequency varied with respect to the benzylaminopurine (BAP) concentration added to the Murashige and Skoog (MS) medium. MS medium with 0.5 mg l−1 BAP exhibited the highest (69.9%) plant regeneration frequency with an average of 186 shoots per explant. A higher concentration of BAP inhibited shoot bud induction and plant regeneration along with necrosis of explants. Petal explants derived from the varieties ‘Branwen’ (pink and white) and ‘Chorus Line’ (violet and white) displayed plant regeneration frequency of 22.2–47.4% (within a total of 12 weeks) on MS medium containing 2.0 mg l−1 α-naphthaleneacetic acid and 0.5 mg l−1 BAP for 8 weeks followed by 4 weeks on MS medium with 1.0 mg l−1 BAP. Scanning electron microscopy confirmed direct plant regeneration without callus. Regenerated plants from leaf explants with well-developed leaves and roots were hardened and successfully transferred to pots in glasshouse exhibiting 86% survival at the end of 4–6 weeks. Whereas, regenerated plants from flower petal explants upon transfer to pots in glasshouse exhibited 75–82% survival at the end of 4–6 weeks.

Plant Protoplasts M.R. Davey, M.R. Davey University of Nottingham, School of Biosciences, Sutton Bonington Campus, Loughborough, United KingdomSearch for more papers by this authorJ.B. Power, J.B. Power University of Nottingham, School of Biosciences, Sutton Bonington Campus, Loughborough, United KingdomSearch for more papers by this authorK.C. Lowe, K.C. Lowe University of Nottingham, School of Life and Environmental Sciences, University Park, Nottingham, United KingdomSearch for more papers by this authorP. Anthony, P. Anthony University of Nottingham, School of Biosciences, Sutton Bonington Campus, Loughborough, United KingdomSearch for more papers by this author M.R. Davey, M.R. Davey University of Nottingham, School of Biosciences, Sutton Bonington Campus, Loughborough, United KingdomSearch for more papers by this authorJ.B. Power, J.B. Power University of Nottingham, School of Biosciences, Sutton Bonington Campus, Loughborough, United KingdomSearch for more papers by this authorK.C. Lowe, K.C. Lowe University of Nottingham, School of Life and Environmental Sciences, University Park, Nottingham, United KingdomSearch for more papers by this authorP. Anthony, P. Anthony University of Nottingham, School of Biosciences, Sutton Bonington Campus, Loughborough, United KingdomSearch for more papers by this author First published: 15 April 2010 https://doi.org/10.1002/9780470054581.eib486 Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Abstract Introduction What are Plant Protoplasts? Isolation of Plant Protoplasts Mechanical and Enzymatic Isolation Procedures Source Material for Protoplast Isolation Culture of Isolated Plant Protoplasts Nutritional Requirements of Protoplasts and Culture Media Experimental Systems for Protoplast Culture Plating Density and the Use of Nurse Cells Innovative Approaches to Protoplast Culture Chemical Supplements for Protoplast Culture Media: Surfactants and Antibiotics Manipulation of Respiratory Gases Physical Procedures to Stimulate Protoplast Growth in Culture Protoplast-to-Plant Systems Exploitation of Protoplast-to-Plant Systems: Somatic and Gametosomatic Hybridisation Transformation by DNA Uptake into Isolated Protoplasts Transformation of Protoplasts by Isolated DNA Induction of DNA Uptake into Protoplasts Factors Influencing Protoplast Transformation Application of DNA Uptake into Protoplasts: Stable and Transient Gene Expression Studies Somaclonal Variation: A Simple Form of Genetic Engineering? Miscellaneous Studies with Plant Protoplasts Concluding Remarks References Encyclopedia of Industrial Biotechnology: Bioprocess, Bioseparation, and Cell TechnologyBrowse other articles of this reference work:BROWSE A-Z RelatedInformation

Global production of ornamental plants is increasing each year with fierce competition between producers, stimulated by increasing consumer demand. Innovations are required in terms of new products at competitive prices to attract consumers. Thus, the industry is under constant pressure to create novel traits. Manipulating the concentration of endogenous growth regulators, such as gibberellins (GAs), in plants can potentially modify shoot architecture. In this investigation, genes from the GA metabolic pathway have been expressed ectopically using the CaMV 35S constitutive promoter in Solanum nigrum and Nicotiana sylvestris. Plants showed statistically significant alteration to their architecture (t-test at 0.01 probability). The feasibility of using tissue-specific promoters was also evaluated in relation to the modification of stature. The technology may lead to decreased dependence on chemical growth regulators, over which there are concerns in relation to human health and potential environmental consequences.

Gibberellins (GAs) are endogenous hormones that play a predominant role in regulating plant stature by increasing cell division and elongation in stem internodes. The product of the GA 2-oxidase gene from Phaseolus coccineus (PcGA2ox1) inactivates C19-GAs, including the bioactive GAs GA1 and GA4, by 2β-hydroxylation, reducing the availability of these GAs in plants. The PcGA2ox1 gene was introduced into Solanum melanocerasum and S. nigrum (Solanaceae) by Agrobacterium-mediated transformation with the aim of decreasing the amounts of bioactive GA in these plants and thereby reducing their stature. The transgenic plants exhibited a range of dwarf phenotypes associated with a severe reduction in the concentrations of the biologically active GA1 and GA4. Flowering and fruit development were unaffected. The transgenic plants contained greater concentrations of chlorophyll b (by 88%) and total chlorophyll (11%), although chlorophyll a and carotenoid contents were reduced by 8 and 50%, respectively. This approach may provide an alternative to the application of chemical growth retardants for reducing the stature of plants, particularly ornamentals, in view of concerns over the potential environmental and health hazards of such compounds.

Widespread soil salinity is an increasing problem in regions of rice cultivation since this major environmental stress limits rice productivity. Numerous studies conducted on biochemical and molecular aspects have shown that the ability of plants to tolerate water stress and salinity is determined by multiple biochemical pathways that facilitate retention and/or acquisition of water, protect chloroplast functions, and maintain ion homeostasis. This abiotic stress is under polygenic control and a combination of approaches is required to achieve the goal of increased tolerance for water stress and salinity. These approaches include the collection of tolerant genotypes to be used as a source of new genes and conventional breeding and selection measures. Also required are novel molecular and biotechnological methodologies to identify stress-related genes and to use them as probes for selecting tolerant genotypes and for producing transgenic plants. Functional genomic studies, QTL analysis and screening, and the use of somatic cell genetics will be involved.

The localization was determined of the triterpenoids, asiaticoside and madecassoside, in different organs of glasshouse-grown plants and cultured material, including transformed roots, of two phenotypes of Centella asiatica (L.) Urban of Malaysian origin. Methanolic extracts of asiaticoside and madecassoside were prepared for gradient HPLC analysis. The two phenotypes of C. asiatica exhibited differences in terpenoid content that were tissue specific and varied between glasshouse-grown plants and tissue culture-derived material. Terpenoid content was highest in leaves, with asiaticoside (0.79 ± 0.03 and 1.15 ± 0.10 % of dry mass) and madecassoside [0.97 ± 0.06 and 1.65 ± 0.01 %(d.m.)] in the fringed (F) and smooth leaf (S) phenotypes, respectively. Roots of the F-phenotype contained the lowest content of asiaticoside [0.12 ± 0.01 %(d.m.)], whereas petioles of S-phenotype plants contained the lowest content of asiaticoside [0.16 ± 0.01 %(d.m.)] and madecassoside [0.18 ± 0.14 %(d.m.)]. Transformed roots were induced using Agrobacterium rhizogens and their growth was maximal on Murashige and Skoog basal medium supplemented with 60 g dm −3 sucrose. However, asiaticoside and madecassoside were undetectable in transformed roots and undifferentiated callus.

A key requirement to enhance our understanding of the response of biological organisms to different levels of gravity is the availability of experimental systems that can simulate microgravity and hypergravity in ground-based laboratories. This paper compares the results obtained from analysing gene expression profiles of Drosophila in space versus those obtained in a random position machine (RPM) and by centrifugation. The correlation found validates the use of the RPM simulation technique to establish the effects of real microgravity on biological systems. This work is being extended to investigate Drosophila development in another gravity modifying instrument, the levitation magnet.

Gravity is an important environmental factor that controls plant growth and development. Studies have shown that the perception of gravity is not only a property of specialized cells, but can also be performed by undifferentiated cultured cells. In this investigation, callus of Arabidopsis thaliana cv. Columbia was used to investigate the initial steps of gravity-related signalling cascades, through altered expression of transcription factors (TFs). TFs are families of small proteins that regulate gene expression by binding to specific promoter sequences. Based on microarray studies, members of the gene families WRKY, MADS-box, MYB, and AP2/EREBP were selected for investigation, as well as members of signalling chains, namely IAA 19 and phosphoinositol-4-kinase. Using qRT-PCR, transcripts were quantified within a period of 30 min in response to hypergravity (8g), clinorotation [2-D clinostat and 3-D random positioning machine (RPM)] and magnetic levitation (ML). The data indicated that (1) changes in gravity induced stress-related signalling, and (2) exposure in the RPM induced changes in gene expression which resemble those of magnetic levitation. Two dimensional clinorotation resulted in responses similar to those caused by hypergravity. It is suggested that RPM and ML are preferable to simulate microgravity than clinorotation.

Pineapple plants of the cv. Phuket transformed with the bar gene for herbicide tolerance were micropropagated, rooted and established in a shade house before transfer to an experimental field plot. Seven months after transfer to the field, plants were tolerant to 1600 ml rai(-1) (240 g rai(-1)) of the herbicide Basta X-(R), this being twice the dose recommended for field application of the herbicide. In contrast, non-transformed pineapple plants became necrotic and died within 21 days of spraying with the herbicide at 800 ml rai(-1). Bar gene stability and expression in clonally-derived plants were assessed by PCR, RT-PCR and Southern analyses at 120, 210 and 380 days following transfer of the plants to the field. The bar gene was stable and expressed in transgenic plants throughout the duration of the trial. Importantly, fruit quality and yield were unaffected by transformation.