Showing papers on "Protoplast published in 1999"
TL;DR: In this paper, two types of dual-Pt microdisk electrodes (disk radius, 0.5−4.0 μm; whole tip radius, 5.0−15 μm) were fabricated to simultaneously detect two electroactive species near a single algal protoplast (radius, 25 μm).
Abstract: Two types (A and B; see Figure 1 in the text) of dual-Pt microdisk electrodes (disk radius, 0.5−4.0 μm; whole tip radius, 5.0−15 μm) were fabricated to simultaneously detect two electroactive species near a single algal protoplast (radius, 25 μm). At the tip of type A, the two disks were on the same plane. Two disk planes of electrode type B formed an angle of ∼135° at the tip. Cyclic voltammetric investigation indicated that, compared with type A, one disk in type B only slightly collected the species electrogenerated at the other disk. These dual-microdisk electrodes were applied to electrochemical measurements of a single protoplast in artificial seawater. The topographic profiles of the single protoplast were obtained from the variation of oxidation current for 1.0 mM Fe(CN)64- at one disk. The photosynthetic oxygen generation from the protoplast was also monitored by detecting the reduction current for oxygen at another disk. The electrode of type B was used as a probe for scanning electrochemical mi...
106 citations
TL;DR: Results show that early developmental processes modify water-transport properties of the plasma membrane, and that the transference chamber is adapted to the study of water-Transport mechanisms in native membranes.
Abstract: A transference chamber was developed to measure the osmotic water permeability coefficient (Pos) in protoplasts 40 to 120 μm in diameter. The protoplast was held by a micropipette and submitted to a steep osmotic gradient created in the transference chamber. Pos was derived from the changes in protoplast dimensions, as measured using a light microscope. Permeabilities were in the range 1 to 1000 μm s-1 for the various types of protoplasts tested. The precision for Pos was
80 citations
TL;DR: In both plant and seed morphology, the hybrid (SH6) was distinct from its rice parental cultivar, as well as from the wild donor species, Z. latifolia, which is a highly asymmetric and fertile somatic hybrid plant obtained via protoplast fusion in an intergenric combination.
Abstract: A highly asymmetric and fertile somatic hybrid plant was obtained via protoplast fusion in an intergenric combination. Gamma-ray-irradiated Zizania latifolia (Griseb). Turcz. ex Stapf mesophyll protoplasts were electrofused with idoacetamide-inactivated rice protoplasts derived from a 2-month-old suspension cell culture. Two of the six putative hybrid calli regenerated plants. Cytological observation showed that the somatic chromosome numbers of both plants were the same as the rice parent (2n=24). Nevertheless, the hybrid nature and inter-genomic exchange events of one of the plants, i.e. SH6 (SH for somatic hybrid), were confirmed by Southern analysis using both total genomic DNA and moderate-copy, Z. latifolia-abundant DNA sequences as probes; in both cases, parental specific and/or new intergenomic recombinant hybridization fragments were detected. In both plant and seed morphology, the hybrid (SH6) was distinct from its rice parental cultivar, as well as from the wild donor species, Z. latifolia.
51 citations
47 citations
TL;DR: Protoplasts from a total of thirty-six genotypes of Brassica species – B. napus, B. campestris and Xinjiang wild rape – were analysed for shoot regeneration using a feeder culture system, finding several genotypes with high regeneration ability were elite breeding lines.
Abstract: Protoplasts from a total of thirty-six genotypes of Brassica species – B. napus, B. campestris (syn. B. rapa), B. juncea, and three distant relatives, Orychophragmus violaceus, Isatis indigotica and Xinjiang wild rape – were analysed for shoot regeneration using a feeder culture system. With the exception of B. campestris and Xinjiang wild rape, some genotypes of all the species could regenerate plants with high efficiency (above 20% of isolated calli initiating shoots). Several genotypes with high regeneration ability were elite breeding lines. Culture conditions as well as genotype had a significant impact on shoot regeneration frequency. In particular, silver nitrate added to the regeneration medium at doses of 6 and 30 μM improved shoot regeneration frequency to 25.4% and 52.2% of isolated calli, respectively, compared to 7.3% percent shoot regeneration without silver nitrate in seven responsive genotypes. Addition of silver nitrate to the regeneration medium also induced shoot regeneration in non-responsive genotypes. Intact plants could be obtained within three months from protoplast isolation in the regenerative genotypes using the current culture system. Advantages of mesophyll protoplasts as compared to protoplasts isolated from hypocotyls for genetic manipulation in Brassica species are discussed.
44 citations
TL;DR: Plants of one line had an allohexaploid chromosome complement and, following examination of its vegetative clones by GISH, were confirmed as somatic hybrids containing full chromosome complements of both O. sativa and P. coarctata.
Abstract: Somatic hybrid plants were obtained following the electrofusion of rice (Oryza sativa L. cv ’Taipei 309’, 2n = 2x = 24) cell suspension–derived protoplasts with non-dividing leaf protoplasts of Porteresia coarctata (2n = 4x = 48), a saline-tolerant wild species. Fusion-treated protoplasts were plated on the surface of cellulose nitrate filter membranes, overlaying Lolium multiflorum nurse cells. The nurse cells were embedded in KPR medium containing 0.5 mg l−1 2,4–dichlorophenoxyacetic acid and semi-solidified with SeaPlaque agarose. Putative somatic hybrid cell colonies were selected on the basis of their growth, whereby faster growing colonies were transferred preferentially to MS-based medium with 2.0 mg l−1 kinetin, 0.5 mg l−1α-naphthaleneacetic acid, 30 g l−1 sucrose and 4.0 g l−1 SeaKem agarose to induce shoot regeneration. One hundred and nineteen regenerated plants were micropropagated clonally on MS-based medium containing 2.0 mg l−1 6–benzylaminopurine, 50 g l−1 sucrose and 4.0 g l−1 SeaKem agarose, prior to DNA extraction of plant samples. Putative somatic hybrids were initially identified by RAPD analysis, and 8 plant lines were selected for further investigation by flow cytometric ploidy determination and cytology. Plants of one line had an allohexaploid chromosome complement (2n = 6x = 72) and, following examination of its vegetative clones by GISH, were confirmed as somatic hybrids containing full chromosome complements of both O. sativa and P. coarctata.
41 citations
TL;DR: The stimulation of plant cells isolated from nontarget tissue suggests the activation of a general signal transduction mechanism by an oligopeptide secreted by the nematode, and the secretion of a mitogenic low-molecular-weight peptide(s) by infective juveniles of the potato cyst nematodes could contribute to the redifferentiation of plants cells into such a feeding cell.
Abstract: Naturally induced secretions from infective juveniles of the potato cyst nematode Globodera rostochiensis co-stimulate the proliferation of tobacco leaf protoplasts in the presence of the synthetic phytohormones α-naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BAP). With the use of a protoplast-based bioassay, a low-molecular-weight peptide(s) (<3 kDa) was shown to be responsible for the observed effect. This mitogenic oligopeptide(s) is functionally dissimilar to auxin and cytokinin and, in addition, it does not change the sensitivity of the protoplasts toward these phytohormones. In combination with the mitogen phytohemagglutinin (PHA), cyst nematode secretions also co-stimulated mitogenesis in human peripheral blood mononuclear cells (PBMC). The stimulation of plant cells isolated from nontarget tissue - these nematodes normally invade the roots of potato plants - suggests the activation of a general signal transduction mechanism(s) by an oligopeptide(s) secreted by the nematode. Whether a similar oligopeptide-induced mechanism underlies human PBMC activation remains to be investigated. Reactivation of the cell cycle is a crucial event in feeding cell formation by cyst nematodes. The secretion of a mitogenic low-molecular-weight peptide(s) by infective juveniles of the potato cyst nematode could contribute to the redifferentiation of plant cells into such a feeding cell.
40 citations
TL;DR: Fertile rooted plantlets were recovered from leaf mesophyll protoplasts of Capsella bursa-pastoris, and somatic hybrids rooted easily and produced sterile flowers when transplanted to soil.
Abstract: Fertile rooted plantlets were recovered from leaf mesophyll protoplasts of Capsella bursa-pastoris. Protoplasts cultured over a feeder layer of Brassica napus cells produced 221 colonies, 7 of which regenerated multiple plantlets. The nuclear DNA content of most regenerates varied from 0.89 to 1.0 pg/nucleus, close to the value for seed-grown C. bursa-pastoris (0.94±0.03 pg/nucleus). Two regenerants had a tetraploid DNA content (1.8– 2.0 pg). Plants with a DNA content close to Capsella produced seeds, both in vitro and in soil. Intertribal somatic hybrids were obtained by polyethylene glycol-mediated fusion of untreated C. bursa-pastoris protoplasts with iodoacetate-treated protoplasts of rapid-cycling B. oleracea. Plants were confirmed as somatic hybrids by isozyme and RAPD analysis. The nuclear DNA content of the hybrids ranged from 3.2 to 6.4 pg, higher than the sum of the parental genomes. One of two hybrids tested was resistant to Alternaria brassicicola, like the Capsella fusion partner. Hybrids rooted easily and produced sterile flowers when transplanted to soil.
29 citations
29 citations
TL;DR: The results suggest that selection for high regeneration response may be efficient at early stages because of the lower environmental influence on the characteristic, and because fewer genes are involved at this stage than at advanced stages.
Abstract: In Brassica oleracea L, plant regeneration from protoplasts is genotype-dependent and colony formation can be obtained routinely In order to identify genes for regenerability, we performed a genetic analysis of the characteristic in the F2 generation of a cross between two accessions selected for high and low regenerability Regeneration frequencies were obtained from protoplast culture of 248 individual F2 plants after 5, 10, and 15 weeks of culture on regeneration medium Broad-sense heritability estimate was 0492 at the early stage and lower (0046–0149) at advanced stages The frequency distribution observed during short-term culture can be explained by two independent loci with duplicate dominant genes controlling regeneration In long-term culture, one additional dominant gene can confer regeneration; ie, three independent loci are responsible for regenerability The results suggest that selection for high regeneration response may be efficient at early stages because of the lower environmental influence on the characteristic, and because fewer genes are involved at this stage than at advanced stages The control of regenerability by a few dominant genes facilitates incorporation of the trait into breeding material Molecular markers linked to the genes may assist in the selection of genotypes with high regeneration percentage in the future
28 citations
TL;DR: In this article, the authors used electrochemical microscopy (SECM) for imaging of respiratory and photosynthetic activities of single, living protoplasts, which showed a decrease in photoynthetic activity of the protoplast injected with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) inhibitor.
Abstract: Scanning electrochemical microscopy (SECM) based on the reduction current for oxygen was used for imaging of respiratory and photosynthetic activities of single, living protoplasts. In the dark, the image of the protoplast appeared as a spot with lower oxygen concentration due to consumption of oxygen by respiration. Under light irradiation, the protoplast appeared as the opposite image because it generated oxygen by photosynthesis. SECM images clearly displayed a decrease in photosynthetic activity of the protoplast injected with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), a photosynthetic inhibitor.
TL;DR: Interspecific asymmetric somatic hybrids have been produced between the sexually incompatible annual Medicago species Medicago truncatula and M. scutellata using only one regenerable parent and improved protoplast techniques.
Abstract: Interspecific asymmetric somatic hybrids have been produced between the sexually incompatible annual Medicago species Medicago truncatula and M scutellata using only one regenerable parent and improved protoplast techniques Regenerable M truncatula leaf protoplasts were inactivated by lethal doses of iodoacetamide Cotyledon protoplasts of non-regenerable M scutellata were produced from 9 to 13 day old γ-irradiated cotyledons Polyethylene glycol was used for protoplast fusion, and its successful use depended on a strategy to eliminate toxic effects More than 50 mature regenerated plants were obtained from the fusion products and amplified fragment length polymorphism and chromosome analysis of a sample of 20 plants allowed the identification of two asymmetric hybrid plants This technique is suitable for transferring characters from incompatible annual Medicago species to Medicago truncatula
TL;DR: Production of useful breeding material and new cultivars derived from somatic hybrids are the final proof of the importance of the technique, as is the case for the improved Ogura-cytoplasmic male sterility (CMS)-lines produced.
Abstract: Publisher Summary This chapter describes somatic hybridization. For utilization of protoplast technology to improve the crops within the family of Brassicaceae , substantial progress has been made. Protoplast fusion can be used as a method to bypass barriers restricting sexual hybridization of different species. Although hybrid plants can be obtained in most of the somatic hybridization experiments performed, low fertility severely restricts the likelihood of seed production upon self-fertilization. Seeds can be obtained by pollinating the hybrids with pollen from one of the parents. The inheritance of certain desired traits, transferred to different Brassica crops, has been followed in several investigations. To improve Brassica crops, traits like disease resistance to different pathogens, modifications of fatty acid composition, cytoplasmic male sterility, herbicide tolerance and repair of seedling chlorosis or sensitivity to cold have been modified and stably inherited over several consecutive generations of back-crosses. Thus, several lines have been derived from somatic hybrids that are of great potential value for further breeding. Production of useful breeding material and new cultivars derived from somatic hybrids are the final proof of the importance of the technique, as is the case for the improved Ogura-cytoplasmic male sterility (CMS)-lines produced.
TL;DR: Both JIM8- and JIM13-responsive epitopes were widespread in sugar beet cells of different origin and stage of cell wall synthesis, and may play an important role in cell wall formation and growth under in vitro conditions.
Abstract: L.). The spatial and temporal expression of both antigens was studied in suspension cells used as the source-tissue for protoplast isolation, in suspension- and mesophyll-derived protoplasts, and in cells which developed from both types of protoplast. Immunofluorescence and immunocytochemical-electron microscopic methods revealed that labeling was present in the cell walls of most suspension cells and also in the incipients of cell walls synthesized around the protoplasts. This signal became much more intense as rebuilding of the cell wall progressed during culture. Relatively weaker labeling was observed in the cytoplasm, where it was frequently associated with the vacuolar compartment. Signal intensity varied between individual cells of the same population and in successive stages of development, but was always stronger with JIM13 than with JIM8. The role of JIM13-responsive epitope in the development of suspension-derived protoplasts was further studied by its ability to bind antibody added to cultures of different ages. Both JIM8- and JIM13-responsive epitopes were widespread in sugar beet cells of different origin and stage of cell wall synthesis. These epitopes may play an important role in cell wall formation and growth under in vitro conditions.
TL;DR: Although after self-pollination pollen germinated on the stigma and pollen tubes were visible in the style, the pods did not develop properly without in vitro culture, and the hybrids were morphologically intermediate between both fusion parents.
Abstract: Protoplast fusion between Brassica oleracea and Moricandia nitens, a C3–C4 intermediate wild species, was carried out. Four hundred and twenty five plants were regenerated from 1995 calli. More than 90% of the regenerated plants were verified as true intergeneric hybrids on the basis of morphological observation and molecular-marker analysis. The hybrids were morphologically intermediate between both fusion parents. Variations in flower color and petal number were also observed. The chromosome number and pollen fertility varied across the individual hybrids. Although after self-pollination pollen germinated on the stigma and pollen tubes were visible in the style, the pods did not develop properly without in vitro culture. Measurements of the CO2 compensation point revealed that six out of eight hybrid plants expressed a gas-exchange character that was intermediate between the C3–C4M. nitens and C3B. oleracea parents.
01 Jan 1999
TL;DR: Methods for direct gene transfer into protoplasts via polyethylene glycol treatment and electroporation were developed in early 1980’s and played an important role in basic studies of gene regulation and function, understanding of the transformation process, and in the production of transgenic crops, particularly of many grass species.
Abstract: Methods for direct gene transfer into protoplasts via polyethylene glycol (PEG) treatment and electroporation were developed in early 1980’s. Genetic transformation of protoplasts led to the production of the first transgenic cereals. This paper describes events leading up to the stable transformation of protoplasts via these techniques, the possible mechanisms involved, the improvement of the methods, and their application to the molecular improvement of cereals. While no longer the predominant method for transforming cereal crops, protoplast transformation still plays an important role in basic studies of gene regulation and function, understanding of the transformation process, and in the production of transgenic crops, particularly of many grass species.
01 Jul 1999
TL;DR: A chronology of key events and events leading to and following the publication of the first book of this kind is presented.
Abstract: .................................................................................................................6 LIST OF PUBLICATION .................................................................................................8 LIST OF ABBREVIATION...............................................................................................9
TL;DR: Five-six months after cell fusion, green plants were obtained in the symmetric and asymmetric fusion and in the random amplified polymorphic DNA (RAPD) analysis, the green plants had bands specific to both parental species.
Abstract: Wide hybridization that cannot be attained through conventional sexual crosses, can now be approached by somatic hybridization. Protoplasts of I. ensata and I. germanica were fused by electrofusion. For the selection of somatic hybrids, protoplasts of I. ensata which did not form colonies in protoplast culture and protoplasts of I. germanica which had regeneration ability for only albino shoot were used in symmetric fusion. On the other hand, the protoplasts of I. ensata and I. germanica protoplasts which were inactivated by iodoacetamide (IOA) treatment were used in asymmetric fusion. Five-six months after cell fusion, green plants were obtained in the symmetric and asymmetric fusion. In the random amplified polymorphic DNA (RAPD) analysis, the green plants had bands specific to both parental species. Therefore, these plants were somatic hybrids between I. ensata and I. germanica.
TL;DR: The results demonstrate that protoplasts fusion can be used for transferring maternally inherited traits like cytoplasmic male sterility to the desired nuclear background which can, in turn, be used in hybrid seed production programme of rice in the tropical world.
Abstract: Wild abortive cytoplasmic male sterility has been extensively used in hybrid seed production in the tropics. Using protoplast fusion between cytoplasmic male sterile and fertile maintainer lines; we report here, transfer of wild abortive cytoplasmic male sterility to the nuclear background of RCPL1-2C, an advance breeding line which also served as maintainer of this cytoplasm. In total, 27 putative cybrids between V20A and RCPL1-2C and 23 lines between V20A and V20B were recovered and all of them were sterile. DNA blots prepared from the mitochondrial DNA of the cybrid lines from both the sets were probed with orf155 that is known to exhibit polymorphism between the mitochondrial DNA of the male-sterile and fertile maintainer lines. Hybridization of orf155 to 1.3 kb HindIII-digested mitochondrial DNA fragment of the cybrids showed transfer of mitochondrial DNA from wild abortive cytoplasmic male-sterile line to the maintainers, viz. RCPL 1-2C and V20B. Expression of male sterility was confirmed by the presence of sterile pollen grains and the lack of seed setting due to selfing in all the cybrid lines. These cybrids, on crossing with respective fertile maintainers set seeds that in turn, produced sterile BC1 plants. DNA blots from HindIII-digested mitochondrial DNA of these BC1 plants when probed with orf155 again exhibited localization of orf155 in wild abortive cytoplasm-specific 1.3 kb HindIII-digested mitochondrial DNA fragments. This demonstrated that the cytoplasmic male sterility transferred through protoplast fusion retained intact female fertility and was inherited and expressed in BC1 plants. Fusion-derived CMS lines, on pollination with pollen grains from restorer, showed restoration of fertility in all the lines. The results demonstrate that protoplasts fusion can be used for transferring maternally inherited traits like cytoplasmic male sterility to the desired nuclear background which can, in turn, be used in hybrid seed production programme of rice in the tropical world.
15 Feb 1999
TL;DR: This paper presents an introduction to Plant Cell Culture and its applications for Plant Protoplasts, as well as specific techniques for the introduction of Specific Genes, including Mutagenesis and the Selection of Resistant Mutants.
Abstract: Introduction. An Introduction to Plant Cell Culture: Pointers to Success R. D. Hall. Cell Culture and Plant Regeneration. Callus Initiation Maintenance and Shoot Induction in Rice N. W. Blackhall J. P. Jotham K. Azakanandam J. B. Power K. C. Lowe E. C. Cocking and M. R. Davey. Callus Initiation Maintenance and Shoot Induction in Potato:Monitoring of Spontaneous Genetic Variability In Vitro and In Vivo R. F. Curry and A. C. Cassells. Somatic Embryogenesis in Barley Suspension Cultures M. Kihara H. Funatsuki K. Ito and P. A. Lazzeri. Somatic Embryogenesis in Picea Suspension Cultures U. Egertsdotter. Specialized Techniques. Direct Cyclic Somatic Embryogenesis in Cassava for Mass Production Purposes K. J. J. M. Raemakers E. Jacobsen and R. G. F. Visser. Immature Inflorescence Culture of Cereals S. Rasco-Gaunt and P. Barcelo. Cryopreservation of Rice Tissue Cultures E. E. Benson and P. T. Lynch. Non-Cryogenic Long-Term Germplasm Storage of Potato A. Golmirzaie and J. Toledo. Plant Propagation In Vitro. Microprogation of Strawberry via Axillary Shoot Proliferation P. Boxus. Meristem-Tip Culture for Propagation and Virus Elimination B. W. W. Grout. Specialized Techniques. Clonal Propagation of Orchids B. Tisserat and D. Jones. Clonal Propagation of Cacti J. Gratton and M. F. Fay. Microproragation of Flower Bulbs: Lily and Narcissus M. M. Langens-Gerrits and G-J . M. de Klerk. Clonal Propagation of Conifers I. S. Harry and T. A. Thorpe. The Culture of Ferns M. V. Ford and M. F. Fay. Applications for Plant Protoplasts. Protoplast Isolation Culture and Plant Regeneration from Passiflora Leaves P. Anthony W. Otoni J. B. Power K. C. Lowe and M. R. Davey. Isolation Culture and Plant Regeneration from Suspension-Derived Protoplasts of Lolium M. Folling and A. Olesen. Protoplast Fusion for Symmetric Somatic Hybrid Production in Brassicaceae J. Fahleson and K. Glimelius. Production of Cybrids in Rapeseed (Brassica napus) S. Yarrow. Specialized Techniques. Microprotoplast Isolation and the Production of Monosomic Addition Lines K. S. Ramulu. Guard Cell Protoplasts: Isolation Culture and Regeneration of Plants G. Boorse and G. Tallman. In Vitro Fertilization with Isolated Single Gametes E. Kranz. Protocols for Genomic Manipulation. Anther and Microspore Culture in Barley H. Lorz and A. J"hne-G"rtner. Microspore Embryogenesis and In Vitro Pollen Maturation for the Production of (Double) Haploids in Tobacco E. Heberle-Bors. Embryo Rescue Following Wide Crosses H. C. Sharma. Specialized Techniques. Mutagenesis and the Selection of Resistant Mutants P. J. Dix. The Generation of Plastid Mutants In Vitro P. J. Dix. Protocols for the Introduction of Specific Genes. Agrobacterium-Mediated Transformation of Petunia Leaf Discs I. M. van der Meer. Transformation of Rice Via PEG-Mediated DNA Uptake into Protoplasts K. Datta and S. K. Datta. Transformation of Wheat Via Particle Bombardment I. K. Vasil and V. Vasil. Plant Transformation via Protoplast Electroporation G. W. Bates. Specialized Techniques. Transformation of Maize via Tissue Electroporation K. D'Halluin E. Bonne M. Bossut and R. Le Page. Transformation of Maize Using Silicon Carbide Whiskers J. M. Dunwell. Suspension Culture Initiation and the Accumulation of Metabolites. Directing Anthraquinone Accumulation via Manipulation of Morinda Suspension Cultures M. J. M. Hagendoorn D. C. L. Jamar and L. H. W. van der Plas. Alkaloid Accumulation in Catharanthus roseus Suspension Cultures A. H. Scragg. Betalains: Their Accumulation and Release In Vitro C.S. Hunter and N. J. Kilby. Appendix. Index.
TL;DR: Cytological analysis revealed extensive variation in chromosome number, such as aneuploid, aneusomatic and mixoploid hybrids, andFloral development and fertility were restored in hybrids derived from fusions between non-flowering or sterile dihaploids.
Abstract: Intraspecific somatic hybrids have been produced by protoplast fusion in eight combinations involving 10 dihaploids (2n = 2x = 24) in an attempt to provide new material for potato breeding. Cytological analysis revealed extensive variation in chromosome number, such as aneuploid, aneusomatic and mixoploid hybrids. Most of the hybrids represented the expected chromosome number of 48; however, the frequency of aneuploids reached 50% in some combinations. Some hybrids carried structurally rearranged chromosomes and exhibited a high frequency of aberrant anaphases. Isozyme and random amplified polymorphic DNA (RAPD) patterns of the hybrids from the same fusion combination were uniform. In the field, somatic hybrids showed wide phenotypic variation in 20 morphological characters. There was a significant correlation between certain leaf characters and the ploidy level, which may be used to distinguish the tetraploid hybrids from hexaploids and octoploids. Tuber yield and flowering intensity were highest in tetraploid hybrids (2n = 4x = 48). Eighteen of the 73 hybrids reached higher yields than the standard variety 'Adretta'. Floral development and fertility were restored in hybrids derived from fusions between non-flowering or sterile dihaploids.
TL;DR: The results demonstrate that the expression levels of both clones are correlated with the rate of growth and coordinatively controlled in petunia plant.
TL;DR: The small number of volatile compounds detected from both the fusion calli and from the donor species indicates that the unorganized callus tissue is unable to produce tissue-specific volatile compounds.
Abstract: Pyrethrum (Tanacetum cinerariifolium (Trevir.) Schultz-Bip.) produces environmentally benign pesticides, the pyrethrins, and tansy (Tanacetum vulgare L.) lower terpenes of variable biological effectiveness. As an approach to improve the oil content and composition of tansy for enhanced biological activity, a somatic hybridization technique between tansy and pyrethrum was established. About 1×106 of leaf-mesophyll protoplasts of both species were mixed and fused with a solution containing 15% polyethylene glycol. Light-green and yellowish calli developed from the fusion experiments. The fusion-derived calli grew vigorously on MS medium supplemented with 6.4 mg l-1 of BAP, 0.8 mg l-1 of NAA, and 30–40 g l-1 of glucose. Nuclear DNA content, RAPD patterns, and volatile compounds were analyzed to determine the hybridity of the calli. The nuclear DNA content of the tansy and pyrethrum genotypes, and the protoplast-derived calli of tansy were 6.41, 7.39, 13.84, and 8.11 pg, respectively. The nuclear DNA content of individual calli derived from the protoplast fusion between tansy +tansy ranged from 8.84 (F43A) to 19.59 pg (F43C) while those of the tansy+pyrethrum fusions were 10.66 (F46A) and 31.87 pg (F46B). Using four 10-mer primers a total of 56 RAPD-PCR fragments were produced. The distance matrices of fragments were calculated by average linkage cluster analysis. Two visually separated clusters were observed. One cluster consisted of the two tansy genotypes and the fusion-derived callus F43A; the other consisted of pyrethrum and fusion-derived calli F46B and F46C. Volatile compounds, such as decadienal, artedouglasia oxide, heptadecane, syringaldehyde and coniferyl alcohol, analyzed by gas chromatography mass spectrometry, were found only in the protoplast fusion-derived calli F43A and F46B. Several less volatile compounds were also detected only in fusion calli. Hexadecanoic and linoleic acids were common to fusion-derived calli and tansy, and one unknown compound to fusion-derived calli and pyrethrum. Pyrethrins I and II were detected from pyrethrum, but not from the fusion-derived calli. The additive nuclear DNA content of protoplast fusion-derived calli and the results of the RAPDs suggest that interspecific fusions had occurred. The small number of volatile compounds detected from both the fusion calli and from the donor species indicates that the unorganized callus tissue is unable to produce tissue-specific volatile compounds.
TL;DR: Success in inducing colony formation from T. yunnanensis protoplasts provides an opportunity to obtain cell lines with high paclitaxel productivity from mutagenized protoplast cultures, and a negative correlation existed between pac litaxel accumulation in colonies and their growth to some extent.
Abstract: Viable protoplasts of Taxus yunnanensis were isolated from friable, light yellow callus. Protoplast yield was dependent on callus age, with a maximum from 20-day-old callus. Protoplasts were induced to undergo sustained divisions and to form cell colonies when cultured in medium consisting of B5 salts, KM vitamin and organic components, 0.45 M fructose, 3.0 mg l-1 2,4-dichlorophenoxyacetic acid and 0.1 mg l-1 kinetin. The planting density was 2.5–3.0×105 protoplasts per ml of culture medium. Cell-free extract from callus enhanced protoplast division and the highest plating efficiency was about 7%. Protoplast-derived colonies showed significant variations in both growth and paclitaxel content. A negative correlation existed between paclitaxel accumulation in colonies and their growth to some extent (r = −0.4485). Among 70 colonies isolated from the heterogeneous protoplast cultures, colony TY-7 accumulated the highest paclitaxel content. Paclitaxel accumulation in colony TY-7 was not great enough to produce paclitaxel for commercial purposes, however, success in inducing colony formation from T. yunnanensis protoplasts provides an opportunity to obtain cell lines with high paclitaxel productivity from mutagenized protoplast cultures.
Journal Article•
TL;DR: The hybrid F 1 and F 2 plants grew vigorously, the stalks were strong, the ears and grains were bigger than parent wheat (“Jinan 177”), now a lot of F 2 spike lines are growing and further analysis will be underway.
Abstract: Hybrid plants were obtained from protoplast fusion between Triticum aestivum L. (2n=42) and Agropyron elongatum (2n=70) via PEG method, but they did not produce seeds. The ovaries of hybrid plants were used to induce hybrid calli again from which was followed by plant regeneration. The hybrid characteristics of the calli and plants were determined by chromosome counting and analysis of esterase isozyme. The results revealed that both of them still retained the hybrid nature. Two of these hybrid plants survived and produced seeds after they were transported into soil. The analysis of phenotype, chromosome, isozyme and RAPD of F 1 plants again proved their hybridity thus indicating that fertile hybrid plants were produced. Chromosome fragments appeared in the root cells of F 1 and F 2 plants. The analysis of PMCs of F 2 plants revealed that the range of chromosomal number were 18Ⅱ～22Ⅱ and pairing or segregating chromosomal fragments were observed, confirming that the chromosomal fragments were minichromosomes. The hybrid F 1 and F 2 plants grew vigorously, the stalks were strong, the ears and grains were bigger than parent wheat (“Jinan 177”). Now a lot of F 2 spike lines are growing and further analysis will be underway.
TL;DR: Out of the 31 fusants obtained, two showed antimicrobial activity against Bacillus subtilis PCI 219, without any activity against Escherichia coli K-12 or Candida albicans 3147.
TL;DR: Results revealed that male-sterile cybrid lines were obtained with 38 chromosomes and a relatively high level of chromosome-pairing ability, indicating their potential for establishing a stable male-stersile rapeseed line.
Abstract: Cytogenetic studies were performed on four male-sterile progenies derived from four different cybrids produced between Brassica napus and B. tournefortii using the donor-recipient protoplast fusion method. The objective of these studies was to characterize the nuclear constitution of the plants. Mitotic investigation revealed that three of the four male-sterile lines had 38 chromosomes, which is equal to that of B. napus. The fourth line, C6, had variable chromosome numbers, ranging from 39 to 42 in different plants. The meiotic behavior in each progeny varied distinctly. Of the plants having 38 chromosomes, fairly high chromosome pairing, on average 18.08 bivalents per cell, was detected at metaphase-I. However, univalents with an average of 1.39 per cell, and very low frequencies of trivalents and/or tetravalents, were also observed in the lines. These results revealed that male-sterile cybrid lines were obtained with 38 chromosomes and a relatively high level of chromosome-pairing ability, indicating their potential for establishing a stable male-sterile rapeseed line.