Science.gov

Sample records for agrobacterium-mediated transformation system

  1. An efficient Agrobacterium-mediated transformation system for poplar.

    PubMed

    Movahedi, Ali; Zhang, Jiaxin; Amirian, Rasoul; Zhuge, Qiang

    2014-06-13

    Poplar is a model system for the regeneration and genetic transformation of woody plants. To shorten the time required for studies of transgenic poplar, efforts have been made to optimize transformation methods that use Agrobacterium tumefaciens. In this study, an Agrobacterium infective suspension was treated at 4 °C for at least 10 h before infecting explants. By transforming the Populus hybrid clone "Nanlin895" (Populus deltoides×P. euramericana) with Agrobacterium harboring the PBI121:CarNAC6 binary vector, we showed that the transformation efficiency was improved significantly by multiple independent factors, including an Agrobacterium infective suspension with an OD600 of 0.7, an Agrobacterium infection for 120 min, an Agrobacterium infective suspension at a pH of 5.0, an acetosyringone concentration of 200 µM, a cocultivation at 28 °C, a cocultivation for 72 h and a sucrose concentration of 30 g/L in the cocultivation medium. We also showed that preculture of wounded leaf explants for two days increased the regeneration rate. The integration of the desired gene into transgenic poplars was detected using selective medium containing kanamycin, followed by southern blot analysis. The expression of the transgene in the transgenic lines was confirmed by northern blot analysis.

  2. An efficient Agrobacterium-mediated transformation of strawberry cv. Camarosa by a dual plasmid system.

    PubMed

    Haddadi, Fatemeh; Aziz, Maheran Abd; Abdullah, Siti Nor Akmar; Tan, Soon Guan; Kamaladini, Hossein

    2015-02-23

    An Agrobacterium-mediated transformation method was applied to introduce the luciferase reporter gene under the control of the CaMV35S promoter in the pGreen0049 binary vector into strawberry cv. Camarosa. The in vitro regeneration system of strawberry leaves to be used in the transformation was optimized using different TDZ concentrations in MS medium. TDZ at 16 µM showed the highest percentage (100%) of shoot formation and the highest mean number of shoots (24) produced per explant. Studies on the effects of different antibiotics, namely timentin, cefotaxime, carbenicillin and ampicillin, on shoot regeneration of strawberry leaf explants showed the best shoot regeneration in the presence of 300 mg/L timentin and 150 mg/L cefotaxime. Assessment of the different factors affecting Agrobacterium mediated-transformation of strawberry with the luciferase gene showed the highest efficiency of putative transformant production (86%) in the treatment with no preculture, bacterial OD600 of 0.6 and the addition of 150 mg/L cefotaxime in the pre-selection and selection media. The presence of the luciferase gene in the plant genome was verified by the luciferase reporter gene assay, nested PCR amplification and dot blot of genomic DNA isolated from the young leaves of each putatively transformed plantlet.

  3. A high-efficiency Agrobacterium-mediated transformation system of rice (Oryza sativa L.).

    PubMed

    Ozawa, Kenjirou

    2012-01-01

    Agrobacterium-mediated transformation of rice has been routinely performed according to the protocol reported by Hiei et al. (Plant J. 6:271-282, 1994). However, several elite japonica and many indica varieties cannot be efficiently transformed by Agrobacterium system. Also a large number of transformants are required to generate T-DNA insertion and FOX libraries as well as gene-targeting studies. To overcome these challenges, we established a high-efficiency transformation system in rice by cocultivating rice calli with Agrobacterium on filter papers moistened with enriched liquid media instead of using solid media (Ozawa, Plant Sci. 176:522-527, 2009; Ozawa and Takaiwa, Plant Sci. 179:333-337, 2010). In this system, the transformation efficiency of the calli is almost 100% in many varieties.

  4. Barley Transformation Using Agrobacterium-Mediated Techniques

    NASA Astrophysics Data System (ADS)

    Harwood, Wendy A.; Bartlett, Joanne G.; Alves, Silvia C.; Perry, Matthew; Smedley, Mark A.; Leyland, Nicola; Snape, John W.

    Methods for the transformation of barley using Agrobacterium-mediated techniques have been available for the past 10 years. Agrobacterium offers a number of advantages over biolistic-mediated techniques in terms of efficiency and the quality of the transformed plants produced. This chapter describes a simple system for the transformation of barley based on the infection of immature embryos with Agrobacterium tumefaciens followed by the selection of transgenic tissue on media containing the antibiotic hygromycin. The method can lead to the production of large numbers of fertile, independent transgenic lines. It is therefore ideal for studies of gene function in a cereal crop system.

  5. Agrobacterium-mediated transformation of Brachypodium distachyon.

    PubMed

    Thole, Vera; Vain, Philippe

    2012-01-01

    Brachypodium distachyon is an attractive genomics and biological model system for grass research. Recently, the complete annotated genome sequence of the diploid line Bd21 has been released. Genetic transformation technologies are critical for the discovery and validation of gene function in Brachypodium. Here, we describe an efficient procedure enabling the Agrobacterium-mediated transformation of a range of diploid and polyploid genotypes of Brachypodium. The procedure relies on the transformation of compact embryogenic calli derived from immature embryos using either chemical selection alone or a combination of chemical and visual screening of transformed tissues and plants. Transformation efficiencies of around 20% can routinely be achieved using this protocol. In the context of the BrachyTAG programme (BrachyTAG.org), this procedure made possible the mass production of Bd21T-DNA mutant plant lines.

  6. Agrobacterium-mediated transformation of cotton.

    PubMed

    Zhang, Baohong

    2013-01-01

    There are many methods and techniques that can be used to transfer foreign genes into cells. In plant biotechnology, Agrobacterium-mediated transformation is a widely used traditional method for inserting foreign genes into plant genome and obtaining transgenic plants, particularly for dicot plant species. Agrobacterium-mediated transformation of cotton involves several important and also critical steps, which includes coculture of cotton explants with Agrobacterium, induction and selection of stable transgenic cell lines, recovery of plants from transgenic cells majorly through somatic embryogenesis, and detection and expression analysis of transgenic plants. In this chapter, we describe a detailed step-by-step protocol for obtaining transgenic cotton plants via Agrobacterium-mediated transformation.

  7. An improved Agrobacterium-mediated transformation system for the functional genetic analysis of Penicillium marneffei.

    PubMed

    Kummasook, Aksarakorn; Cooper, Chester R; Vanittanakom, Nongnuch

    2010-12-01

    We have developed an improved Agrobacterium-mediated transformation (AMT) system for the functional genetic analysis of Penicillium marneffei, a thermally dimorphic, human pathogenic fungus. Our AMT protocol included the use of conidia or pre-germinated conidia of P. marneffei as the host recipient for T-DNA from Agrobacterium tumefaciens and co-cultivation at 28°C for 36 hours. Bleomycin-resistant transformants were selected as yeast-like colonies following incubation at 37°C. The efficiency of transformation was approximately 123 ± 3.27 and 239 ± 13.12 transformants per plate when using 5 × 10(4) conidia and pre-germinated conidia as starting materials, respectively. Southern blot analysis demonstrated that 95% of transformants contained single copies of T-DNA. Inverse PCR was employed for identifying flanking sequences at the T-DNA insertion sites. Analysis of these sequences indicated that integration occurred as random recombination events. Among the mutants isolated were previously described stuA and gasC defective strains. These AMT-derived mutants possessed single T-DNA integrations within their particular coding sequences. In addition, other morphological and pigmentation mutants possessing a variety of gene-specific defects were isolated, including two mutants having T-DNA integrations within putative promoter regions. One of the latter integration events was accompanied by the deletion of the entire corresponding gene. Collectively, these results indicated that AMT could be used for large-scale, functional genetic analyses in P. marneffei. Such analyses can potentially facilitate the identification of those genetic elements related to morphogenesis, as well as pathogenesis in this medically important fungus.

  8. An embryogenic suspension cell culture system for Agrobacterium-mediated transformation of citrus.

    PubMed

    Dutt, M; Grosser, J W

    2010-11-01

    A method for the genetic transformation of several citrus cultivars is described, including cultivars observed to be recalcitrant to conventional epicotyl-mediated transformation. Embryogenic cell suspension cultures, established from unfertilized ovules were used as target tissues for Agrobacterium-mediated transformation. Several modifications were made to the culture environment to investigate factors required for efficient transfer of the T-DNA and the subsequent regeneration of transgenic citrus plants. It was determined that co-cultivation of citrus cells and Agrobacterium in EME medium supplemented with maltose (EME-M) and 100 μM acetosyringone for 5 days at 25°C was optimum for transformation of each of the citrus cultivars. Efficient selection was obtained and escapes were prevented when the antibiotic hygromycin B was used as a selection antibiotic following transformation with an Agrobacterium strain containing hptII in the T-DNA region. Transgenic embryo regeneration and development was enhanced in medium that contained a liquid overlay consisting of a 1:2 mixture of 0.6 M BH3 and 0.15 M EME-M media. PCR and Southern blot analyses confirmed the presence of the T-DNA and the stable integration into the genome of regenerated plants, while RT-PCR demonstrated variable amounts of RNA being transcribed in different transgenic lines. This protocol can create an avenue for insertion of useful traits into any polyembryonic citrus cultivar that can be established as embryogenic cell suspension cultures, including popular specialty mandarins and seedless cultivars.

  9. Agrobacterium-mediated transformation of Fusarium proliferatum.

    PubMed

    Bernardi-Wenzel, J; Quecine, M C; Azevedo, J L; Pamphile, J A

    2016-01-01

    Fusarium proliferatum is an important pathogen that is associated with plant diseases and primarily affects aerial plant parts by producing different mycotoxins, which are toxic to humans and animals. Within the last decade, this fungus has also been described as one of the causes of red root rot or sudden death syndrome in soybean, which causes extensive damage to this crop. This study describes the Agrobacterium tumefaciens-mediated transformation of F. proliferatum as a tool for the disruption of pathogenicity genes. The genetic transformation was performed using two binary vectors (pCAMDsRed and pFAT-GFP) containing the hph (hygromycin B resistance) gene as a selection marker and red and green fluorescence, respectively. The presence of acetosyringone and the use of filter paper or nitrocellulose membrane were evaluated for their effect on the transformation efficiency. A mean processing rate of 94% was obtained with 96 h of co-cultivation only in the presence of acetosyringone and the use of filter paper or nitrocellulose membrane did not affect the transformation process. Hygromycin B resistance and the presence of the hph gene were confirmed by PCR, and fluorescence due to the expression of GFP and DsRed protein was monitored in the transformants. A high rate of mitotic stability (95%) was observed. The efficiency of Agrobacterium-mediated transformation of F. proliferatum allows the technique to be used for random insertional mutagenesis studies and to analyze fungal genes involved in the infection process. PMID:27323127

  10. Agrobacterium-mediated transformation of Fusarium proliferatum.

    PubMed

    Bernardi-Wenzel, J; Quecine, M C; Azevedo, J L; Pamphile, J A

    2016-06-03

    Fusarium proliferatum is an important pathogen that is associated with plant diseases and primarily affects aerial plant parts by producing different mycotoxins, which are toxic to humans and animals. Within the last decade, this fungus has also been described as one of the causes of red root rot or sudden death syndrome in soybean, which causes extensive damage to this crop. This study describes the Agrobacterium tumefaciens-mediated transformation of F. proliferatum as a tool for the disruption of pathogenicity genes. The genetic transformation was performed using two binary vectors (pCAMDsRed and pFAT-GFP) containing the hph (hygromycin B resistance) gene as a selection marker and red and green fluorescence, respectively. The presence of acetosyringone and the use of filter paper or nitrocellulose membrane were evaluated for their effect on the transformation efficiency. A mean processing rate of 94% was obtained with 96 h of co-cultivation only in the presence of acetosyringone and the use of filter paper or nitrocellulose membrane did not affect the transformation process. Hygromycin B resistance and the presence of the hph gene were confirmed by PCR, and fluorescence due to the expression of GFP and DsRed protein was monitored in the transformants. A high rate of mitotic stability (95%) was observed. The efficiency of Agrobacterium-mediated transformation of F. proliferatum allows the technique to be used for random insertional mutagenesis studies and to analyze fungal genes involved in the infection process.

  11. [Agrobacterium-mediated transformation of Cymbidium sinensis].

    PubMed

    Xie, Li; Wang, Fen; Zeng, Ruizhen; Guo, Herong; Zhou, Yuliang; Zhang, Zhisheng

    2015-04-01

    Genetic transformation is an effective method to improve breeding objective traits of orchids. However, there is little information about genetic transformation of Cymbidium sinensis. Rhizomes from shoot-tip culture of C. sinensis cv. 'Qijianbaimo' were used to establish a practical transformation protocol of C. sinensis. Pre-culture time, concentration and treating methods of acetosyringone, concentration of infection bacteria fluid (OD600), infection time, and co-culture time had significant effects on β-glucuronidase (GUS) transient expression rate of C. sinensis cv. 'Qijianbaimo' rhizome. The GUS transient expression rate of rhizome was the highest (11.67%) when rhizomes pre-cultured for 39 d were soaked in bacterium suspension (OD600 = 0.9) supplemented with 200 μmol/L acetosyringone for 35 min, followed by culturing on co-culture medium supplemented with 200 μmol/L acetosyringone for 7 d. Under this transformation conditions, 3 transgenic plantlets, confirmed by GUS histochemical assay and PCR, were obtained from 400 regenerated plantlets, and the genetic transformation rate was 0.75%. This proved that it was feasible to create new cultivars by the use of Agrobacterium-mediated genetic transformation in C. sinense.

  12. Development of a phosphomannose isomerase-based Agrobacterium-mediated transformation system for chickpea (Cicer arietinum L.).

    PubMed

    Patil, Gunvant; Deokar, Amit; Jain, P K; Thengane, R J; Srinivasan, R

    2009-11-01

    To develop an alternative genetic transformation system that is not dependent on an antibiotic selection strategy, the phosphomannose isomerase gene (pmi) system was evaluated for producing transgenic plants of chickpea (Cicer arietinum L.). A shoot morphogenesis protocol based on the thidiazuron (TDZ)-induced shoot morphogenesis system was combined with Agrobacterium-mediated transformation of the pmi gene and selection of transgenic plants on mannose. Embryo axis explants of chickpea cv. C-235 were grown on a TDZ-supplemented medium for shoot proliferation. Embryo axis explants from which the first and second flush of shoots were removed were transformed using Agrobacterium carrying the pmi gene, and emerging shoots were allowed to regenerate on a zeatin-supplemented medium with an initial selection pressure of 20 g l(-1) mannose. Rooting was induced in the selected shoots on an indole-3-butyric acid (IBA)-supplemented medium with a selection pressure of 15 g l(-1) mannose. PCR with marker gene-specific primers and chlorophenol red (CPR) assay of the shoots indicated that shoots had been transformed. RT-PCR and Southern analysis of selected regenerated plants further confirmed integration of the transgene into the chickpea genome. These positive results suggest that the pmi/mannose selection system can be used to produce transgenic plants of chickpea that are free from antibiotic resistance marker genes.

  13. Establishment of a high efficiency Agrobacterium-mediated transformation system of rice (Oryza sativa L.).

    PubMed

    Ozawa, Kenjirou

    2009-04-01

    Technologies for transformation of rice have been developed to meet the requirements of functional genomics in order to enable the production of transgenic rice plants with useful agricultural characters. However, many rice varieties are not efficiently transformed by Agrobacterium. We have succeeded in establishing a highly efficient transformation system in rice by co-cultivating rice calli with Agrobacterium on three filter papers moistened with enriched N6 or DKN media instead of using solid media. Rice calli immersed in Agrobacterium suspension (EHA101, Agrobacterium concentration of OD600=0.04) were co-cultured on three pieces of filter paper (9cm in diameter) moistened with 5.5mL of N6 or DKN liquid co-cultivation medium supplemented with 2,4-d (2mg/L), proline (10mM), casein hydrolysate (300mg/L), sucrose (30g/L), glucose (5g/L), l-cysteine (100mg/L) and acetosyringone (15mg/L) at 25°C for 3 days in the dark. Compared with the transformation efficiency of calli co-cultivated on solid media, transformation efficiency was increased by about fivefold by using the filter paper method for many varieties of rice, including those that previously yielded much poor transformation rates.

  14. Biolistic- and Agrobacterium-mediated transformation protocols for wheat.

    PubMed

    Tamás-Nyitrai, Cecília; Jones, Huw D; Tamás, László

    2012-01-01

    After rice, wheat is considered to be the most important world food crop, and the demand for high-quality wheat flour is increasing. Although there are no GM varieties currently grown, wheat is an important target for biotechnology, and we anticipate that GM wheat will be commercially available in 10-15 years. In this chapter, we summarize the main features and challenges of wheat transformation and then describe detailed protocols for the production of transgenic wheat plants both by biolistic and Agrobacterium-mediated DNA-delivery. Although these methods are used mainly for bread wheat (Triticum aestivum L.), they can also be successfully applied, with slight modifications, to tetraploid durum wheat (T. turgidum L. var. durum). The appropriate size and developmental stage of explants (immature embryo-derived scutella), the conditions to produce embryogenic callus tissues, and the methods to regenerate transgenic plants under increasing selection pressure are provided in the protocol. To illustrate the application of herbicide selection system, we have chosen to describe the use of the plasmid pAHC25 for biolistic transformation, while for Agrobacterium-mediated transformation the binary vector pAL156 (incorporating both the bar gene and the uidA gene) has been chosen. Beside the step-by-step methodology for obtaining stably transformed and normal fertile plants, procedures for screening and testing transgenic wheat plants are also discussed.

  15. Agrobacterium-mediated transformation of maize (Zea mays) immature embryos.

    PubMed

    Lee, Hyeyoung; Zhang, Zhanyuan J

    2014-01-01

    Agrobacterium tumefaciens-mediated transformation is one of the most efficient and simple gene delivery systems for genetic improvement and biology studies in maize. This system has become more widely used by both public and private laboratories. However, transformation efficiencies vary greatly from laboratory to laboratory for the same genotype. Here, we illustrate our advanced Agrobacterium-mediated transformation method in Hi-II maize using simple binary vectors. The protocol utilizes immature embryos as starting explants and the bar gene as a selectable marker coupled with bialaphos as a selective agent. The protocol offers efficient transformation results with high reproducibility, provided that some experimental conditions are well controlled. This transformation method, with minor modifications, can be also employed to transform certain maize inbreds.

  16. Agrobacterium-mediated transformation: rice transformation.

    PubMed

    Slamet-Loedin, Inez H; Chadha-Mohanty, Prabhjit; Torrizo, Lina

    2014-01-01

    Agrobacterium is a common soil bacterium with natural capacity for trans-kingdom transfer of genetic information by transferring its T-DNA into the eukaryotic genome. In agricultural plant biotechnology, combination of non-phytopathogenic strain of Agrobacterium tumefaciens with modified T-DNA and vir-genes in a binary vector system is the most widely utilized system for genetic improvement in diverse plant species and for gene function validation. Here we have described a highly efficient A. tumefaciens-mediated transformation system for indica and japonica rice cultivars based on an immature embryo system.

  17. Efficient production of transgenic melon via Agrobacterium-mediated transformation.

    PubMed

    Bezirganoglu, I; Hwang, S Y; Shaw, J F; Fang, T J

    2014-04-25

    Oriental melon (Cucumis melo L. var. makuwa) is an important fruit for human consumption. However, this plant species is one of the most recalcitrant to genetic transformation. The lack of an efficient in vitro system limits the development of a reproducible genetic transformation protocol for Oriental melon. In this study, an efficient transgenic production method for Agrobacterium-mediated transformation using cotyledon explants of Oriental melon was developed. Cotyledon explants were pre-cultivated for two days in the dark, and the optimal conditions for transformation of melon were determined to be a bacteria concentration of OD600 0.6, inoculation for 30 min, and two days of co-cultivation. Transgenic melon plants were produced from kanamycin-resistant shoots. A total of 11 independent transgenic plants were regenerated with a transformation efficiency of 0.8% of the inoculated explants. The transgenic plants were phenotypically normal and fully fertile, which might be a consequence of the co-cultivation time.

  18. A highly efficient in vitro plant regeneration system and Agrobacterium-mediated transformation in Plumbago zeylanica.

    PubMed

    Wei, Xiaoping; Gou, Xiaoping; Yuan, Tong; Russell, Scott D

    2006-06-01

    Plumbago zeylanica is a unique model for studying flowering plant gametogenesis, heterospermy, and preferential fertilization, yet understanding the control of related molecular mechanisms is impossible without efficient and reproducible regeneration and stable genetic transformation. We found three key factors for enhancing successful regeneration: (1) tissue source of explants, (2) combination and concentration of growth regulators, and (3) culture conditions. The highest frequency of shoot regeneration was achieved using hypocotyl segments cultured on MS basal medium supplemented with BA 2.0 mg/l, NAA 0.75 mg/l, adenine 50 mg/l and 10% (v/v) coconut milk under subdued light at 25+/-2 degrees C; under these conditions, each hypocotyl segment produced over 30 shoots, arising primarily through direct organogenesis after 3 weeks of culture. Regenerated shoots rooted easily on half-strength basal MS medium and were successfully established in the greenhouse. Using this tissue culture protocol, reporter gene GUS under the constitutive CaMV 35S promoter was introduced into P. zeylanica cells of petiole, cotyledon and hypocotyl with A. tumefaciens strains AGL1 and LBA4404. Transient expression was observed in all recipient tissues. Stable transgenic calli originating from petiole were obtained. PMID:16470412

  19. Agrobacterium-mediated transformation of three freshwater microalgal strains.

    PubMed

    Sanitha, Mary; Radha, Sudhakar; Fatima, Anwar Aliya; Devi, Selvaraju Gayathri; Ramya, Mohandass

    2014-01-01

    Microalgal transformation has gained interest in recent years. Agrobacterium-mediated transformation remains as the most efficient method for the development of transgenic plants and microalgae due to its wide host range, inexpensive procedure and transfer of large segments of DNA. In the present study, three different microalgal species were isolated from freshwater environment and identified based on the morphological characteristics and ITS-2 region amplification. Agrobacterium-mediated transformation was successful for the isolates Chlorella sp., Ankistrodesmus sp and Scenedesmus bajacalifornicus. Gene integration and expression was confirmed by PCR amplification of hptII and GUS histochemical assay. A. tumifaciens contamination was checked by amplification of npt II gene (kanamycin resistant) which lies outside the T-border. Based on GUS assay, transformation efficiencies were found to be 12.25% for Chlorella sp. 2.96% for Scenedesmus bajacalifornicus and 3.5% for Ankistrodesmus sp.

  20. Agrobacterium-mediated transformation of barley (Hordeum vulgare L.).

    PubMed

    Ismagul, Ainur; Mazonka, Iryna; Callegari, Corinne; Eliby, Serik

    2014-01-01

    Barley biotechnology requires efficient genetic engineering tools for producing transgenic plants necessary for conducting reverse genetics analyses in breeding and functional genomics research. Agrobacterium-mediated genetic transformation is an important technique for producing barley transgenics with simple low-copy number transgenes. This chapter reports a refined protocol for the systematic high-throughput transformation of the advanced Australian spring barley breeding line WI4330.

  1. VIP1: linking Agrobacterium-mediated transformation to plant immunity?

    PubMed

    Liu, Yukun; Kong, Xiangpei; Pan, Jiaowen; Li, Dequan

    2010-08-01

    Agrobacterium tumefaciens is the most efficient vehicle used today for the production of transgenic plants and plays an essential role in basic scientific research and in agricultural biotechnology. Previously, plant VirE2-interacting protein 1 (VIP1) was shown to play a role in Agrobacterium-mediated transformation. Recent reports demonstrate that VIP1, as one of the bZIP transcription factors, is also involved in plant immunity responses. Agrobacterium is able to activate and abuse VIP1 for transformation. These findings highlight Agrobacterium-host interaction and unveil how Agrobacterium hijacks host cellular mechanism for its own benefit. This review focuses on the roles played by VIP1 in Agrobacterium-mediated transformation and plant immunity. PMID:20473505

  2. VIP1: linking Agrobacterium-mediated transformation to plant immunity?

    PubMed

    Liu, Yukun; Kong, Xiangpei; Pan, Jiaowen; Li, Dequan

    2010-08-01

    Agrobacterium tumefaciens is the most efficient vehicle used today for the production of transgenic plants and plays an essential role in basic scientific research and in agricultural biotechnology. Previously, plant VirE2-interacting protein 1 (VIP1) was shown to play a role in Agrobacterium-mediated transformation. Recent reports demonstrate that VIP1, as one of the bZIP transcription factors, is also involved in plant immunity responses. Agrobacterium is able to activate and abuse VIP1 for transformation. These findings highlight Agrobacterium-host interaction and unveil how Agrobacterium hijacks host cellular mechanism for its own benefit. This review focuses on the roles played by VIP1 in Agrobacterium-mediated transformation and plant immunity.

  3. Is VIP1 important for Agrobacterium-mediated transformation?

    PubMed

    Shi, Yong; Lee, Lan-Ying; Gelvin, Stanton B

    2014-09-01

    Agrobacterium genetically transforms plants by transferring and integrating T-(transferred) DNA into the host genome. This process requires both Agrobacterium and host proteins. VirE2 interacting protein 1 (VIP1), an Arabidopsis bZIP protein, has been suggested to mediate transformation through interaction with and targeting of VirE2 to nuclei. We examined the susceptibility of Arabidopsis vip1 mutant and VIP1 overexpressing plants to transformation by numerous Agrobacterium strains. In no instance could we detect altered transformation susceptibility. We also used confocal microscopy to examine the subcellular localization of Venus-tagged VirE2 or Venus-tagged VIP1, in the presence or absence of the other untagged protein, in different plant cell systems. We found that VIP1-Venus localized in both the cytoplasm and the nucleus of Arabidopsis roots, agroinfiltrated Nicotiana benthamiana leaves, Arabidopsis mesophyll protoplasts and tobacco BY-2 protoplasts, regardless of whether VirE2 was co-expressed. VirE2 localized exclusively to the cytoplasm of tobacco and Arabidopsis protoplasts, whether in the absence or presence of VIP1 overexpression. In transgenic Arabidopsis plants and agroinfiltrated N. benthamina leaves we could occasionally detect small aggregates of the Venus signal in nuclei, but these were likely to be imagining artifacts. The vast majority of VirE2 remained in the cytoplasm. We conclude that VIP1 is not important for Agrobacterium-mediated transformation or VirE2 subcellular localization.

  4. Aboveground insect infestation attenuates belowground Agrobacterium-mediated genetic transformation.

    PubMed

    Song, Geun Cheol; Lee, Soohyun; Hong, Jaehwa; Choi, Hye Kyung; Hong, Gun Hyong; Bae, Dong-Won; Mysore, Kirankumar S; Park, Yong-Soon; Ryu, Choong-Min

    2015-07-01

    Agrobacterium tumefaciens causes crown gall disease. Although Agrobacterium can be popularly used for genetic engineering, the influence of aboveground insect infestation on Agrobacterium induced gall formation has not been investigated. Nicotiana benthamiana leaves were exposed to a sucking insect (whitefly) infestation and benzothiadiazole (BTH) for 7 d, and these exposed plants were inoculated with a tumorigenic Agrobacterium strain. We evaluated, both in planta and in vitro, how whitefly infestation affects crown gall disease. Whitefly-infested plants exhibited at least a two-fold reduction in gall formation on both stem and crown root. Silencing of isochorismate synthase 1 (ICS1), required for salicylic acid (SA) synthesis, compromised gall formation indicating an involvement of SA in whitefly-derived plant defence against Agrobacterium. Endogenous SA content was augmented in whitefly-infested plants upon Agrobacterium inoculation. In addition, SA concentration was three times higher in root exudates from whitefly-infested plants. As a consequence, Agrobacterium-mediated transformation of roots of whitefly-infested plants was clearly inhibited when compared to control plants. These results suggest that aboveground whitefly infestation elicits systemic defence responses throughout the plant. Our findings provide new insights into insect-mediated leaf-root intra-communication and a framework to understand interactions between three organisms: whitefly, N. benthamiana and Agrobacterium. PMID:25676198

  5. Agrobacterium-mediated transformation of Brassica napus and Brassica oleracea.

    PubMed

    Bhalla, Prem L; Singh, Mohan B

    2008-01-01

    Agrobacterium-mediated transformation is widely used for gene delivery in plants. However, commercial cultivars of crop plants are often recalcitrant to transformation because the protocols established for model varieties are not directly applicable to them. The genus Brassica includes the oil seed crop, canola (B. napus), and vegetable crop varieties of Brassica oleracea, including cauliflower, broccoli and cabbage. Here, we describe an efficient protocol for Agrobacterium-mediated transformation using seedling explants that is applicable to various Brassica varieties; this protocol has been used to genetically engineer commercial cultivars of canola and cauliflower in our laboratory. Young seedling explants are inoculated with Agrobacterium on the day of explant preparation. Explants are grown for 1 week in the absence of a selective agent before being transferred to a selective medium to recover transgenic shoots. Transgenic shoots are subjected to an additional round of selection on medium containing higher levels of the selective agent and a low-carbohydrate source; this helps to eliminate false-positive plants. Use of seedling explants offers flexible experiment planning and a convenient explant source. Using this protocol, transgenic plants can be obtained in 2.5 to 3.5 months.

  6. Development of an efficient Agrobacterium-mediated transformation system and production of herbicide-resistant transgenic plants in garlic (Allium sativum L.).

    PubMed

    Ahn, Yul-Kyun; Yoon, Moo-Kyoung; Jeon, Jong-Seong

    2013-08-01

    The genetic improvement of garlic plants (Allium sativum L.) with agronomical beneficial traits is rarely achieved due to the lack of an applicable transformation system. Here, we developed an efficient Agrobacterium-mediated transformation procedure with Danyang, an elite Korean garlic cultivar. Examination of sGFP (synthetic green fluorescence protein) expression revealed that treatment with 2-(N-morpholino) ethanesulfonic acid (MES), L-cysteine and/or dithiothreitol (DTT) gives the highest efficiency in transient gene transfer during Agrobacterium co-cultivation with calli derived from the roots of in vitro plantlets. To increase stable transformation efficiency, a two-step selection was employed on the basis of hygromycin resistance and sGFP expression. Of the hygromycin-resistant calli initially produced, only sGFP-expressing calli were subcultured for selection of transgenic calli. Transgenic plantlets produced from these calli were grown to maturity. The transformation efficiency increased up to 10.6% via our optimized procedure. DNA and RNA gel-blot analysis indicated that transgenic garlic plants stably integrated and expressed the phosphinothricin acetyltransferase (PAT) gene. A herbicide spraying assay demonstrated that transgenic plants of garlic conferred herbicide resistance, whilst nontransgenic plants and weeds died. These results indicate that our transformation system can be efficiently utilized to produce transgenic garlic plants with agronomic benefits.

  7. Role of bacterial virulence proteins in Agrobacterium-mediated transformation of Aspergillus awamori.

    PubMed

    Michielse, C B; Ram, A F J; Hooykaas, P J J; Hondel, C A M J J van den

    2004-05-01

    The Agrobacterium-mediated transformation of Aspergillus awamori was optimized using defined co-cultivation conditions, which resulted in a reproducible and efficient transformation system. Optimal co-cultivation conditions were used to study the role of Agrobacterium tumefaciens virulence proteins in T-DNA transfer. This study revealed that inactivation of either of the regulatory proteins (VirA, VirG), any of the transport pore proteins (VirB), proteins involved in generation of the T-strand (VirD, VirC) or T-strand protection and targeting (VirE2) abolishes or severely reduces the formation of transformants. The results indicate that the Agrobacterium-mediated transformation of A. awamori requires an intact T-DNA machinery for efficient transformation; however, the plant host range factors, like VirE3, VirH, and VirF, are not important. PMID:15050546

  8. Efficient Agrobacterium-mediated transformation of the liverwort Marchantia polymorpha using regenerating thalli.

    PubMed

    Kubota, Akane; Ishizaki, Kimitsune; Hosaka, Masashi; Kohchi, Takayuki

    2013-01-01

    The thallus, the gametophyte body of the liverwort Marchantia polymorpha, develops clonal progenies called gemmae that are useful in the isolation and propagation of isogenic plants. Developmental timing is critical to Agrobacterium-mediated transformation, and high transformation efficiency has been achieved only with sporelings. Here we report an Agrobacterium-mediated transformation system for M. polymorpha using regenerating thalli. Thallus regeneration was induced by cutting the mature thallus across the apical-basal axis and incubating the basal portion of the thallus for 3 d. Regenerating thalli were infected with Agrobacterium carrying binary vector that contained a selection marker, the hygromycin phosphotransferase gene, and hygromycin-resistant transformants were obtained with an efficiency of over 60%. Southern blot analysis verified random integration of 1 to 4 copies of the T-DNA into the M. polymorpha genome. This Agrobacterium-mediated transformation system for M. polymorpha should provide opportunities to perform genetic transformation without preparing spores and to generate a sufficient number of transformants with isogenic background. PMID:23291762

  9. Efficient Agrobacterium-mediated transformation of the liverwort Marchantia polymorpha using regenerating thalli.

    PubMed

    Kubota, Akane; Ishizaki, Kimitsune; Hosaka, Masashi; Kohchi, Takayuki

    2013-01-01

    The thallus, the gametophyte body of the liverwort Marchantia polymorpha, develops clonal progenies called gemmae that are useful in the isolation and propagation of isogenic plants. Developmental timing is critical to Agrobacterium-mediated transformation, and high transformation efficiency has been achieved only with sporelings. Here we report an Agrobacterium-mediated transformation system for M. polymorpha using regenerating thalli. Thallus regeneration was induced by cutting the mature thallus across the apical-basal axis and incubating the basal portion of the thallus for 3 d. Regenerating thalli were infected with Agrobacterium carrying binary vector that contained a selection marker, the hygromycin phosphotransferase gene, and hygromycin-resistant transformants were obtained with an efficiency of over 60%. Southern blot analysis verified random integration of 1 to 4 copies of the T-DNA into the M. polymorpha genome. This Agrobacterium-mediated transformation system for M. polymorpha should provide opportunities to perform genetic transformation without preparing spores and to generate a sufficient number of transformants with isogenic background.

  10. Comparative analysis of transgenic tall fescue (Festuca arundinacea Schreb.) plants obtained by Agrobacterium-mediated transformation and particle bombardment.

    PubMed

    Gao, Caixia; Long, Danfeng; Lenk, Ingo; Nielsen, Klaus Kristian

    2008-10-01

    Agrobacterium-mediated transformation and particle bombardment are the two most widely used methods for genetically modifying grasses. Here, these two systems are compared for transformation efficiency, transgene integration and transgene expression when used to transform tall fescue (Festuca arundinacea Schreb.). The bar gene was used as a selectable marker and selection during tissue culture was performed using 2 mg/l bialaphos in both callus induction and regeneration media. Average transformation efficiency across the four callus lines used in the experiments was 10.5% for Agrobacterium-mediated transformation and 11.5% for particle bombardment. Similar transgene integration patterns and co-integration frequencies of bar and uidA were observed in both gene transfer systems. However, while GUS activity was detected in leaves of 53% of the Agrobacterium transformed lines, only 20% of the bombarded lines showed GUS activity. Thus, Agrobacterium-mediated transformation appears to be the preferred method for producing transgenic tall fescue plants.

  11. In planta Agrobacterium-mediated transformation by vacuum infiltration.

    PubMed

    Tague, Brian W; Mantis, Joanna

    2006-01-01

    In planta Agrobacterium-mediated transformation using vacuum infiltration results in transgenic Arabidopsis thaliana without the use of sterile conditions or plant regeneration. Plants are grown in pots, in standard potting mix. Agrobacterium tumefaciens, carrying an appropriate plant transformation vector, is suspended in an infiltration medium that contains, at a minimum, sucrose and the surfactant Silwet L-77. Flower buds are immersed in the suspension of A. tumefaciens. The application of a vacuum drives the bacteria into the intercellular air spaces. A portion of the Agrobacterium Ti plasmid known as the T-DNA region, which has been engineered to carry a selectable marker, becomes integrated into the plant genomic DNA. Plants are allowed to set seed. Seeds are germinated in selective conditions to recover transformants. PMID:16739579

  12. The Agrobacterium-mediated transformation of common wheat (Triticum aestivum L.) and triticale (x Triticosecale Wittmack): role of the binary vector system and selection cassettes.

    PubMed

    Bińka, Agnieszka; Orczyk, Wacław; Nadolska-Orczyk, Anna

    2012-02-01

    The influence of two binary vector systems, pGreen and pCAMBIA, on the Agrobacterium-mediated transformation ability of wheat and triticale was studied. Both vectors carried selection cassettes with bar or nptII driven by different promoters. Two cultivars of wheat, Kontesa and Torka, and one cultivar of triticale, Wanad, were tested. The transformation rates for the wheat cultivars ranged from 0.00 to 3.58% and from 0.00 to 6.79% for triticale. The best values for wheat were 3.58% for Kontesa and 3.14% for Torka, and these were obtained after transformation with the pGreen vector carrying the nptII selection gene under the control of 35S promoter. In the case of the bar selection system, the best transformation rates were, respectively, 1.46 and 1.79%. Such rates were obtained when the 35S::bar cassette was carried by the pCAMBIA vector; they were significantly lower with the pGreen vector. The triticale cultivar Wanad had its highest transformation rate after transformation with nptII driven by 35S in pCAMBIA. The bar selection system for the same triticale cultivar was better when the gene was driven by nos and the selection cassette was carried by pGreen. The integration of the transgenes was confirmed with at least three pairs of specific starters amplifying the fragments of nptII, bar, or gus. The expression of selection genes, measured by reverse transcriptase polymerase chain reaction (RT-PCR) in relation to the actin gene, was low, ranging from 0.00 to 0.63 for nptII and from 0.00 to 0.33 for bar. The highest relative transcript accumulation was observed for nptII driven by 35S and expressed in Kontesa that had been transformed with pGreen.

  13. Lox-dependent gene expression in transgenic plants obtained via Agrobacterium-mediated transformation.

    PubMed

    Shcherbak, N; Kishchenko, O; Sakhno, L; Komarnytsky, I; Kuchuk, M

    2013-01-01

    Lox sites of the Cre/lox recombination system from bacteriophage P1 were analyzed for their ability to affect on transgene expression when inserted upstream from a gene coding sequence adjacent to the right border (RB) of T-DNA. Wild and mutated types of lox sites were tested for their effect upon bar gene expression in plants obtained via Agrobacterium-mediated and biolistic transformation methods. Lox-mediated expression of bar gene, recognized by resistance of transgenic plants to PPT, occurred only in plants obtained via Agrobacterium-mediated transformation. RT-PCR analysis confirms that PPT-resistant phenotype of transgenic plants obtained via Agrobacterium-mediated transformation was caused by activation of bar gene. The plasmid with promoterless gus gene together with the lox site adjacent to the RB was constructed and transferred to Nicotiana tabacum as well. Transgenic plants exhibited GUS activity and expression of gus gene was detected in plant leaves. Expression of bar gene from the vectors containing lox site near RB allowed recovery of numerous PPT-resistant transformants of such important crops as Beta vulgaris, Brassica napus, Lactuca sativa and Solanum tuberosum. Our results demonstrate that the lox site sequence adjacent to the RB can be used to control bar gene expression in transgenic plants. PMID:23821951

  14. Lox-dependent gene expression in transgenic plants obtained via Agrobacterium-mediated transformation.

    PubMed

    Shcherbak, N; Kishchenko, O; Sakhno, L; Komarnytsky, I; Kuchuk, M

    2013-01-01

    Lox sites of the Cre/lox recombination system from bacteriophage P1 were analyzed for their ability to affect on transgene expression when inserted upstream from a gene coding sequence adjacent to the right border (RB) of T-DNA. Wild and mutated types of lox sites were tested for their effect upon bar gene expression in plants obtained via Agrobacterium-mediated and biolistic transformation methods. Lox-mediated expression of bar gene, recognized by resistance of transgenic plants to PPT, occurred only in plants obtained via Agrobacterium-mediated transformation. RT-PCR analysis confirms that PPT-resistant phenotype of transgenic plants obtained via Agrobacterium-mediated transformation was caused by activation of bar gene. The plasmid with promoterless gus gene together with the lox site adjacent to the RB was constructed and transferred to Nicotiana tabacum as well. Transgenic plants exhibited GUS activity and expression of gus gene was detected in plant leaves. Expression of bar gene from the vectors containing lox site near RB allowed recovery of numerous PPT-resistant transformants of such important crops as Beta vulgaris, Brassica napus, Lactuca sativa and Solanum tuberosum. Our results demonstrate that the lox site sequence adjacent to the RB can be used to control bar gene expression in transgenic plants.

  15. Development of an Agrobacterium-mediated transformation system for the cold-adapted fungi Pseudogymnoascus destructans and P. pannorum.

    PubMed

    Zhang, Tao; Ren, Ping; Chaturvedi, Vishnu; Chaturvedi, Sudha

    2015-08-01

    The mechanisms of cold adaptation by fungi remain unknown. This topic is of high interest due to the emergence of white-nose syndrome (WNS), a skin infection of hibernating bats caused by Pseudogymnoascus destructans (Pd). Recent studies indicated that apart from Pd, there is an abundance of other Pseudogymnoascus species in the hibernacula soil. We developed an Agrobacterium tumefaciens-mediated transformation (ATMT) system for Pd and a related fungus Pseudogymnoascus pannorum (Pp) to advance experimental studies. URE1 gene encoding the enzyme urease was used as an easy to screen marker to facilitate molecular genetic analyses. A Uracil-Specific Excision Reagent (USER) Friendly pRF-HU2 vector containing Pd or Pp ure1::hygromycin (HYG) disruption cassette was introduced into A. tumefaciens AGL-1 cells by electroporation and the resulting strains were co-cultivated with conidia of Pd or Pp for various durations and temperatures to optimize the ATMT system. Overall, 680 Pd (0.006%) and 1800 Pp (0.018%) transformants were obtained from plating of 10(7) conidia; their recoveries were strongly correlated with the length of the incubation period (96h for Pd; 72h for Pp) and with temperature (15-18°C for Pd; 25°C for Pp). The homologous recombination in transformants was 3.1% for Pd and 16.7% for Pp. The availability of a standardized ATMT system would allow future molecular genetic analyses of Pd and related cold-adapted fungi. PMID:26051491

  16. Development of an Agrobacterium-mediated transformation system for the cold-adapted fungi Pseudogymnoascus destructans and P. pannorum.

    PubMed

    Zhang, Tao; Ren, Ping; Chaturvedi, Vishnu; Chaturvedi, Sudha

    2015-08-01

    The mechanisms of cold adaptation by fungi remain unknown. This topic is of high interest due to the emergence of white-nose syndrome (WNS), a skin infection of hibernating bats caused by Pseudogymnoascus destructans (Pd). Recent studies indicated that apart from Pd, there is an abundance of other Pseudogymnoascus species in the hibernacula soil. We developed an Agrobacterium tumefaciens-mediated transformation (ATMT) system for Pd and a related fungus Pseudogymnoascus pannorum (Pp) to advance experimental studies. URE1 gene encoding the enzyme urease was used as an easy to screen marker to facilitate molecular genetic analyses. A Uracil-Specific Excision Reagent (USER) Friendly pRF-HU2 vector containing Pd or Pp ure1::hygromycin (HYG) disruption cassette was introduced into A. tumefaciens AGL-1 cells by electroporation and the resulting strains were co-cultivated with conidia of Pd or Pp for various durations and temperatures to optimize the ATMT system. Overall, 680 Pd (0.006%) and 1800 Pp (0.018%) transformants were obtained from plating of 10(7) conidia; their recoveries were strongly correlated with the length of the incubation period (96h for Pd; 72h for Pp) and with temperature (15-18°C for Pd; 25°C for Pp). The homologous recombination in transformants was 3.1% for Pd and 16.7% for Pp. The availability of a standardized ATMT system would allow future molecular genetic analyses of Pd and related cold-adapted fungi.

  17. Efficient method for Agrobacterium mediated transformation of Artemisia annua L.

    PubMed

    Alam, Pravej; Mohammad, Anis; Ahmad, M M; Khan, Mather Ali; Nadeem, Mohd; Khan, Riyazuddeen; Akmal, Mohd; Ahlawat, Seema; Abdin, M Z

    2014-01-01

    Artemisinin, a potent antimalarial natural products isolated from aerial parts of Artemisia annua L. Many patents have been reported that the demand for artemisinin is exponentially increasing year after year due to increased incidences of drug resistant malaria throughout the world. Leaf explants were used frequently as target tissue to generate transgenic of Artemisia. annua L. However, obtaining a large number of transgenic lines through out the year is a laborious and delicate process. To circumvent this, we have developed a highly efficient leaf explant based Agrobacterium mediated transformation of A. annua L. plant. The gus gene was used as screenable marker to assess and optimize the performance of T-DNA delivery. The age of explant, kind of bacterial inoculation, suspension duration, infection times and co-culture conditions were optimized. The co-culture was carried out with Agrobacterium tumefaciens strain EHA105 under desiccation condition in the dark at 25-28 0C for 2-4 days. Complete analysis of transgene insertion demonstrated that the optimized method of transformation from leaf explants of A. annua L. was efficient and highly reproducible.

  18. Agrobacterium-mediated transformation of finger millet (Eleusine coracana (L.) Gaertn.) using shoot apex explants.

    PubMed

    Ceasar, S Antony; Ignacimuthu, S

    2011-09-01

    A new Agrobacterium-mediated transformation system was developed for finger millet using shoot apex explants. The Agrobacterium strain LBA4404 harboring binary vector pCAMBIA1301, which contained hygromycin phosphotransferase (hptII) as selectable marker gene and β-glucuronidase (GUS) as reporter gene, was used for optimization of transformation conditions. Two finger millet genotypes, GPU 45 and CO 14, were used in this study. The optimal conditions for the Agrobacterium-mediated transformation of finger millet were found to be the co-cultivation of explants obtained on the 16th day after callus induction (DACI), exposure of explants for 30 min to agrobacterial inoculum and 3 days of co-cultivation on filter paper placed on medium supplemented with 100 μM acetosyringone (AS). Addition of 100 μM L: -cysteine in the selection medium enhanced the frequency of transformation and transgenic plant recovery. Both finger millet genotypes were transformed by Agrobacterium. A frequency of 19% transient expression with 3.8% stable transformation was achieved in genotype GPU 45 using optimal conditions. Five stably transformed plants were fully characterized by Southern blot analysis. A segregation analysis was also performed in four R(1) progenies, which showed normal Mendelian pattern of transgene segregation. The inheritance of transgenes in R(1) progenies was also confirmed by Southern blot analysis. This is the first report on Agrobacterium-mediated transformation of finger millet. This study underpins the introduction of numerous agronomically important genes into the genome of finger millet in the future.

  19. Highly efficient Agrobacterium-mediated transformation of banana cv. Rasthali (AAB) via sonication and vacuum infiltration.

    PubMed

    Subramanyam, Kondeti; Subramanyam, Koona; Sailaja, K V; Srinivasulu, M; Lakshmidevi, K

    2011-03-01

    A reproducible and efficient transformation method was developed for the banana cv. Rasthali (AAB) via Agrobacterium-mediated genetic transformation of suckers. Three-month-old banana suckers were used as explant and three Agrobacterium tumefaciens strains (EHA105, EHA101, and LBA4404) harboring the binary vector pCAMBIA1301 were used in the co-cultivation. The banana suckers were sonicated and vacuum infiltered with each of the three A. tumefaciens strains and co-cultivated in the medium containing different concentrations of acetosyringone for 3 days. The transformed shoots were selected in 30 mg/l hygromycin-containing selection medium and rooted in rooting medium containing 1 mg/l IBA and 30 mg/l hygromycin. The presence and integration of the hpt II and gus genes into the banana genome were confirmed by GUS histochemical assay, polymerase chain reaction, and southern hybridization. Among the different combinations tested, high transformation efficiency (39.4 ± 0.5% GUS positive shoots) was obtained when suckers were sonicated and vacuum infiltered for 6 min with A. tumefaciens EHA105 in presence of 50 μM acetosyringone followed by co-cultivation in 50 μM acetosyringone-containing medium for 3 days. These results suggest that an efficient Agrobacterium-mediated transformation protocol for stable integration of foreign genes into banana has been developed and that this transformation system could be useful for future studies on transferring economically important genes into banana.

  20. An improved Agrobacterium-mediated transformation of recalcitrant indica rice (Oryza sativa L.) cultivars.

    PubMed

    Shri, Manju; Rai, Arti; Verma, Pankaj Kumar; Misra, Prashant; Dubey, Sonali; Kumar, Smita; Verma, Sikha; Gautam, Neelam; Tripathi, Rudra Deo; Trivedi, Prabodh Kumar; Chakrabarty, Debasis

    2013-04-01

    Agrobacterium-mediated transformation of indica rice varieties has been quite difficult as these are recalcitrant to in vitro responses. In the present study, we established a high-efficiency Agrobacterium tumefaciens-mediated transformation system of rice (Oryza sativa L. ssp. indica) cv. IR-64, Lalat, and IET-4786. Agrobacterium strain EHA-101 harboring binary vector pIG121-Hm, containing a gene encoding for β-glucuronidase (GUS) and hygromycin resistance, was used in the transformation experiments. Manipulation of different concentrations of acetosyringone, days of co-culture period, bacterial suspension of different optical densities (ODs), and the concentrations of L-cysteine in liquid followed by solid co-culture medium was done for establishing the protocol. Among the different co-culture periods, 5 days of co-culture with bacterial cells (OD600 nm = 0.5-0.8) promoted the highest frequency of transformation (83.04 %) in medium containing L-cysteine (400 mg l(-1)). Putative transformed plants were analyzed for the presence of a transgene through genomic PCR and GUS histochemical analyses. Our results also suggest that different cultural conditions and the addition of L-cysteine in the co-culture medium improve the Agrobacterium-mediated transformation frequencies from an average of 12.82 % to 33.33 % in different indica rice cultivars.

  1. An efficient regeneration protocol for Agrobacterium-mediated transformation of melon (Cucumis melo L.).

    PubMed

    Zhang, H J; Gao, P; Wang, X Z; Luan, F S

    2014-01-08

    An efficient selection and plant regeneration protocol for Agrobacterium-mediated transformation, using cotyledon node zone-stem connection region of melon, has been developed. The new Agrobacterium-mediated transformation methodology, independent of organ culture, used the entire germinated seed as explants. The transformation system was maximized to maintain the integrity of melon itself, thus avoiding the limitations of traditional tissue culture methods. The transformation was carried out under a non-sterile environment. The incorporation of a selectable marker (neomycin phosphotransferase II) into the genome of transgenic plants was confirmed by PCR and Southern blot analyses. The transformation frequency based on the PCR was 13%. Transgenic melon plants were usually detected by PCR in less than 1 month after Agrobacterium inoculation, and seeds could be harvested in 3 months. The growth characteristics and morphology of the transgenic plants were identical to the untransformed wild-type plants. This method would be beneficial for facilitating the characteristics of gene functions and for boosting the manipulation of melon transformation for commercial purposes.

  2. Okadaic acid and trifluoperazine enhance Agrobacterium-mediated transformation in eastern white pine.

    PubMed

    Tang, Wei; Lin, Jinxing; Newton, Ronald J

    2007-05-01

    Mature zygotic embryos of recalcitrant Christmas tree species eastern white pine (Pinus strobus L.) were used as explants for Agrobacterium tumefaciens strain GV3101-mediated transformation using the uidA (beta-Glucuronidase) gene as a reporter. Influence of the time of sonication and the concentrations of protein phosphatase inhibitor (okadaic acid) and kinase inhibitor (trifluoperazine) on Agrobacterium-mediated transformation have been evaluated. A high transformation frequency was obtained after embryos were sonicated for 45-50 s, or treated with 1.5-2.0 microM okadaic acid or treated with 100-200 microM trifluoperazine, respectively. Protein phosphatase and kinase inhibitors enhance Agrobacterium-mediated transformation in eastern white pine. A 2-3.5-fold higher rate of hygromycin-resistant callus was obtained with an addition of 2 microM okadaic acid or 150 microM trifluoperazine or sonicated embryos for 45 s. Stable integration of the uidA gene in the plant genome of eastern white pine was confirmed by polymerase chain reaction (PCR), Southern and northern blot analyses. These results demonstrated that a stable and enhanced transformation system has been established in eastern white pine and this system would provide an opportunity to transfer economically important genes into this Christmas tree species.

  3. Development of an efficient in vitro plant regeneration system amenable to Agrobacterium- mediated transformation of a recalcitrant grain legume blackgram (Vigna mungo L. Hepper).

    PubMed

    Sainger, Manish; Chaudhary, Darshna; Dahiya, Savita; Jaiwal, Ranjana; Jaiwal, Pawan K

    2015-10-01

    An efficient, rapid and direct multiple shoot regeneration system amenable to Agrobacterium-mediated transformation from primary leaf with intact petiole of blackgram (Vigna mungo) is established for the first time. The effect of the explant type and its age, type and concentration of cytokinin and auxin either alone or in combination and genotype on multiple shoot regeneration efficiency and frequency was optimized. The primary leaf explants with petiole excised from 4-day-old seedlings directly developed multiple shoots (an average of 10 shoots/ explant) from the cut ends of the petiole in 95 % of the cultures on MSB (MS salts and B5 vitamins) medium containing 1.0 μM 6-benzylaminopurine. Elongated (2-3 cm) shoots were rooted on MSB medium with 2.5 μM indole-butyric acid and resulted plantlets were hardened and established in soil, where they resumed growth and reached maturity with normal seed set. The regenerated plants were morphologically similar to seed-raised plants and required 8 weeks time from initiation of culture to establish them in soil. The regeneration competent cells present at the cut ends of petiole are fully exposed and are, thus, easily accessible to Agrobacterium, making this plant regeneration protocol amenable for the production of transgenic plants. The protocol was further successfully used to develop fertile transgenic plants of blackgram using Agrobacterium tumefaciens strain EHA 105 carrying a binary vector pCAMBIA2301 that contains a neomycin phosphotransferase gene (nptII) and a β-glucuronidase (GUS) gene (uidA) interrupted with an intron. The presence and integration of transgenes in putative T0 plants were confirmed by polymerase chain reaction (PCR) and Southern blot hybridization, respectively. The transgenes were inherited in Mendelian fashion in T1 progeny and a transformation frequency of 1.3 % was obtained. This protocol can be effectively used for transferring new traits in blackgram and other legumes for their

  4. Development of an efficient in vitro plant regeneration system amenable to Agrobacterium- mediated transformation of a recalcitrant grain legume blackgram (Vigna mungo L. Hepper).

    PubMed

    Sainger, Manish; Chaudhary, Darshna; Dahiya, Savita; Jaiwal, Ranjana; Jaiwal, Pawan K

    2015-10-01

    An efficient, rapid and direct multiple shoot regeneration system amenable to Agrobacterium-mediated transformation from primary leaf with intact petiole of blackgram (Vigna mungo) is established for the first time. The effect of the explant type and its age, type and concentration of cytokinin and auxin either alone or in combination and genotype on multiple shoot regeneration efficiency and frequency was optimized. The primary leaf explants with petiole excised from 4-day-old seedlings directly developed multiple shoots (an average of 10 shoots/ explant) from the cut ends of the petiole in 95 % of the cultures on MSB (MS salts and B5 vitamins) medium containing 1.0 μM 6-benzylaminopurine. Elongated (2-3 cm) shoots were rooted on MSB medium with 2.5 μM indole-butyric acid and resulted plantlets were hardened and established in soil, where they resumed growth and reached maturity with normal seed set. The regenerated plants were morphologically similar to seed-raised plants and required 8 weeks time from initiation of culture to establish them in soil. The regeneration competent cells present at the cut ends of petiole are fully exposed and are, thus, easily accessible to Agrobacterium, making this plant regeneration protocol amenable for the production of transgenic plants. The protocol was further successfully used to develop fertile transgenic plants of blackgram using Agrobacterium tumefaciens strain EHA 105 carrying a binary vector pCAMBIA2301 that contains a neomycin phosphotransferase gene (nptII) and a β-glucuronidase (GUS) gene (uidA) interrupted with an intron. The presence and integration of transgenes in putative T0 plants were confirmed by polymerase chain reaction (PCR) and Southern blot hybridization, respectively. The transgenes were inherited in Mendelian fashion in T1 progeny and a transformation frequency of 1.3 % was obtained. This protocol can be effectively used for transferring new traits in blackgram and other legumes for their

  5. Cinnamic acid, coumarin and vanillin: Alternative phenolic compounds for efficient Agrobacterium-mediated transformation of the unicellular green alga, Nannochloropsis sp.

    PubMed

    Cha, Thye-San; Chen, Chin-Fong; Yee, Willy; Aziz, Ahmad; Loh, Saw-Hong

    2011-03-01

    The use of acetosyringone in Agrobacterium-mediated gene transfer into plant hosts has been favored for the past few decades. The influence of other phenolic compounds and their effectiveness in Agrobacterium-mediated plant transformation systems has been neglected. In this study, the efficacy of four phenolic compounds on Agrobacterium-mediated transformation of the unicellular green alga Nannochloropsis sp. (Strain UMT-M3) was assessed by using β-glucuronidase (GUS) assay. We found that cinnamic acid, vanillin and coumarin produced higher percentages of GUS positive cells as compared to acetosyringone. These results also show that the presence of methoxy group in the phenolic compounds may not be necessary for Agrobacterium vir gene induction and receptor binding as suggested by previous studies. These findings provide possible alternative Agrobacterium vir gene inducers that are more potent as compared to the commonly used acetosyringone in achieving high efficiency of Agrobacterium-mediated transformation in microalgae and possibly for other plants.

  6. Mature seed-derived callus of the model indica rice variety Kasalath is highly competent in Agrobacterium-mediated transformation.

    PubMed

    Saika, Hiroaki; Toki, Seiichi

    2010-12-01

    We previously established an efficient Agrobacterium-mediated transformation system using primary calli derived from mature seeds of the model japonica rice variety Nipponbare. We expected that the shortened tissue culture period would reduce callus browning--a common problem with the indica transformation system during prolonged tissue culture in the undifferentiated state. In this study, we successfully applied our efficient transformation system to Kasalath--a model variety of indica rice. The Luc reporter system is sensitive enough to allow quantitative analysis of the competency of rice callus for Agrobacterium-mediated transformation. We unexpectedly discovered that primary callus of Kasalath exhibits a remarkably high competency for Agrobacterium-mediated transformation compared to Nipponbare. Southern blot analysis and Luc luminescence showed that independent transformation events in primary callus of Kasalath occurred successfully at ca. tenfold higher frequency than in Nipponbare, and single copy T-DNA integration was observed in ~40% of these events. We also compared the competency of secondary callus of Nipponbare and Kasalath and again found superior competency in Kasalath, although the identification and subsequent observation of independent transformation events in secondary callus is difficult due to the vigorous growth of both transformed and non-transformed cells. An efficient transformation system in Kasalath could facilitate the identification of QTL genes, since many QTL genes are analyzed in a Nipponbare × Kasalath genetic background. The higher transformation competency of Kasalath could be a useful trait in the establishment of highly efficient systems involving new transformation technologies such as gene targeting.

  7. The role of RAR1 in Agrobacterium-mediated plant transformation

    PubMed Central

    Anand, Ajith; Mysore, Kirankumar S

    2013-01-01

    RAR1 is identified as a critical protein involved in plant innate immunity. We investigated the role of RAR1 in Agrobacterium-mediated plant transformation based on the previous findings that accessory proteins associated with the E3 ligase complex such as SGT1, which tightly interacts with RAR1, play a role in the transformation process. RAR1 gene silencing in Nicotiana benthamiana and Arabidopsis rar1 mutant analysis suggested that RAR1 is required for early stages of Agrobacterium-mediated plant transformation. This finding further illustrates that RAR1, along with SGT1, that serve as a HSP90 co-chaperone is important for Agrobacterium-mediated plant transformation.

  8. Agrobacterium-mediated transformation in Alpinia galanga (Linn.) Willd. for enhanced acetoxychavicol acetate production.

    PubMed

    Rao, Kiranmayee; Chodisetti, Bhuvaneswari; Mangamoori, Lakshmi Narasu; Giri, Archana

    2012-09-01

    Agrobacterium-mediated transformations ensure elevated amounts of secondary metabolite accumulation with genetic and biosynthetic stability. In the present study, Alpinia galanga rich in bioactive compounds was genetically transformed using different strains of Agrobacterium rhizogenes viz. LBA 9402, A(4), 532, 2364 and PRTGus. Even though a higher growth rate was obtained with the LBA 9402 strain, maximum acetoxychavicol acetate accumulation (ACA) was seen in the PRTGus transformant. PRTGus root line has shown 10.1 fold higher ACA content in comparison to the control roots. The lowest ACA production was shown by the A(4) transformant (4.9 fold). The quantification of ACA in the transformed roots was carried out by using HPLC, which was found to be in the order of PRTGus > LBA 9402 > 2364 > 532 > A(4). The fast growth rate of hairy roots, genetic stability and their ability to synthesize more than one metabolite offer a promising system for the production of valuable secondary metabolites.

  9. Agrobacterium-mediated transformation leads to improved gene replacement efficiency in Aspergillus awamori.

    PubMed

    Michielse, C B; Arentshorst, M; Ram, A F J; van den Hondel, C A M J J

    2005-01-01

    In this study, the efficiency of gene replacement in Aspergillus awamori between Agrobacterium-mediated transformation and CaCl(2)/PEG-mediated transformation was compared. For the genes, pyrG and gfaA, it was found that the homologous recombination frequencies obtained by Agrobacterium-mediated transformation were 3- to 6-fold higher than the frequencies obtained with CaCl(2)/PEG protoplast transformation. For the pyrG gene, it was found that Agrobacterium-mediated transformation allowed an efficient homologous recombination with shorter DNA flanks than CaCl(2)/PEG protoplast transformation. Finally, the addition of the dominant amdS marker as a second selection marker to the gene replacement cassette led to a further 2-fold enrichment in transformants with gene replacement events, resulting in a gene replacement frequency of 55%. Based on the data it can be concluded that Agrobacterium-mediated transformation is an efficient tool for gene replacement and that the amdS gene can be successfully used as a second selection marker to select transformants with putative gene replacement.

  10. Sequential monitoring of transgene expression following Agrobacterium-mediated transformation of rice.

    PubMed

    Saika, Hiroaki; Nonaka, Satoko; Osakabe, Keishi; Toki, Seiichi

    2012-11-01

    Although Agrobacterium-mediated transformation technology is now used widely in rice, many varieties of indica-type rice are still recalcitrant to Agrobacterium-mediated transformation. It was reported recently that T-DNA integration into the rice genome could be the limiting step in this method. Here, we attempted to establish an efficient sequential monitoring system for stable transformation events by visualizing stable transgene expression using a non-destructive and highly sensitive visible marker. Our results demonstrate that click beetle luciferase (ELuc) is an excellent marker allowing the observation of transformed cells in rice callus, exhibiting a sensitivity >30-fold higher than that of firefly luciferase. Since we have previously shown that green fluorescent protein (GFP) is a useful visual marker with which to follow transient and/or stable expression of transgenes in rice, we constructed an enhancer trap vector using both the gfbsd2 (GFP fused to the N-terminus of blasticidin S deaminase) and eluc genes. In this vector, the eluc gene is under the control of the Cauliflower mosaic virus 35S minimal promoter, while the gfbsd2 gene is under the control of the full-length rice elongation factor gene promoter. Observation of transformed callus under a dissecting microscope demonstrated that the level of ELuc luminescence reflected exclusively stable transgene expression, and that both transient and stable expression could be monitored by the level of GFP fluorescence. Moreover, we show that our system enables sequential quantification of transgene expression via differential measurement of ELuc luminescence and GFP fluorescence.

  11. A Fruiting Body Tissue Method for Efficient Agrobacterium-Mediated Transformation of Agaricus bisporus

    PubMed Central

    Chen, Xi; Stone, Michelle; Schlagnhaufer, Carl; Romaine, C. Peter

    2000-01-01

    We describe a modified Agrobacterium-mediated method for the efficient transformation of Agaricus bisporus. Salient features of this procedure include cocultivation of Agrobacterium and fruiting body gill tissue and use of a vector with a homologous promoter. This method offers new prospects for the genetic manipulation of this commercially important mushroom species. PMID:11010906

  12. Agrobacterium-Mediated Transformation of Bread and Durum Wheat Using Freshly Isolated Immature Embryos

    NASA Astrophysics Data System (ADS)

    Huixia, Wu; Angela, Doherty; Jones, Huw D.

    Agrobacterium-mediated transformation of wheat is becoming a viable alternative to the more established biolistic protocols. It offers advantages in terms of simple, low-copy-number integrations and can be applied with similar efficiencies to specific durum wheat and spring and winter bread wheat types varieties.

  13. Agrobacterium-mediated transformation of the haploid liverwort Marchantia polymorpha L., an emerging model for plant biology.

    PubMed

    Ishizaki, Kimitsune; Chiyoda, Shota; Yamato, Katsuyuki T; Kohchi, Takayuki

    2008-07-01

    Agrobacterium-mediated transformation has not been practical in pteridophytes, bryophytes and algae to date, although it is commonly used in model plants including Arabidopsis and rice. Here we present a rapid Agrobacterium-mediated transformation system for the haploid liverwort Marchantia polymorpha L. using immature thalli developed from spores. Hundreds of hygromycin-resistant plants per sporangium were obtained by co-cultivation of immature thalli with Agrobacterium carrying the binary vector that contains a reporter, the beta-glucuronidase (GUS) gene with an intron, and a selection marker, the hygromycin phosphotransferase (hpt) gene. In this system, individual gemmae, which arise asexually from single initial cells, were analyzed as isogenic transformants. GUS activity staining showed that all hygromycin-resistant plants examined expressed the GUS transgene in planta. DNA analyses verified random integration of 1-5 copies of the intact T-DNA between the right and the left borders into the M. polymorpha genome. The efficient and rapid Agrobacterium-mediated transformation of M. polymorpha should provide molecular techniques to facilitate comparative genomics, taking advantage of this unique model plant that retains many features of the common ancestor of land plants.

  14. Multiple host-cell recombination pathways act in Agrobacterium-mediated transformation of plant cells.

    PubMed

    Mestiri, Imen; Norre, Frédéric; Gallego, Maria E; White, Charles I

    2014-02-01

    Using floral-dip, tumorigenesis and root callus transformation assays of both germline and somatic cells, we present here results implicating the four major non-homologous and homologous recombination pathways in Agrobacterium-mediated transformation of Arabidopsis thaliana. All four single mutant lines showed similar mild reductions in transformability, but knocking out three of four pathways severely compromised Agrobacterium-mediated transformation. Although integration of T-DNA into the plant genome is severely compromised in the absence of known DNA double-strand break repair pathways, it does still occur, suggesting the existence of other pathways involved in T-DNA integration. Our results highlight the functional redundancy of the four major plant recombination pathways in transformation, and provide an explanation for the lack of strong effects observed in previous studies on the roles of plant recombination functions in transformation.

  15. Regeneration and Agrobacterium-mediated transformation of hop (Humulus lupulus L.).

    PubMed

    Horlemann, C; Schwekendiek, A; Höhnle, M; Weber, G

    2003-10-01

    An efficient procedure for direct organogenesis and regeneration of hop (Humulus lupulus L.) was established. For the first time Agrobacterium-mediated genetic transformation of hop (cv. "Tettnanger") was achieved. Shoot internodes from in vitro cultures were identified as the most suitable type of explant for regeneration. Using this type of explant, a shoot-inducing medium was developed that supported direct organogenesis of approximately 50% of the explants. Plantlets were successfully rooted and transferred to the greenhouse. Overall, in less than 6 months hop cultures propagated in vitro were regenerated to plants in the greenhouse. Agrobacterium-mediated genetic transformation was performed with the reporter gene GUS (beta-glucuronidase). The presence and function of transgenes in plants growing in the greenhouse was verified by PCR (polymerase chain reaction) and enzyme assay for GUS activity, respectively. We have obtained 21 transgenic plants from 1,440 explants initially transformed, yielding an overall transformation efficiency of 1.5%. PMID:12898178

  16. Enhanced targeted integration mediated by translocated I-SceI during the Agrobacterium mediated transformation of yeast.

    PubMed

    Rolloos, Martijn; Hooykaas, Paul J J; van der Zaal, Bert J

    2015-02-09

    Agrobacterium mediated transformation (AMT) has been embraced by biotechnologists as the technology of choice to introduce or alter genetic traits of plants. However, in plants it is virtually impossible to predetermine the integration site of the transferred T-strand unless one is able to generate a double stranded break (DSB) in the DNA at the site of interest. In this study, we used the model organism Saccharomyces cerevisiae to investigate whether the Agrobacterium mediated translocation of site-specific endonucleases via the type IV secretion system (T4SS), concomitantly with T-DNA transfer is possible and whether this can improve the gene targeting efficiency. In addition to that, the effect of different chromatin states on targeted integration, was investigated. It was found that Agrobacterium mediated translocation of the homing endonuclease I-SceI has a positive effect on the integration of T-DNA via the homologous repair (HR) pathway. Furthermore, we obtained evidence that nucleosome removal has a positive effect on I-SceI facilitated T-DNA integration by HR. Reversely; inducing nucleosome formation at the site of integration removes the positive effect of translocated I-SceI on T-DNA integration.

  17. Screening Chinese soybean genotypes for Agrobacterium-mediated genetic transformation suitability*

    PubMed Central

    Song, Zhang-yue; Tian, Jing-luan; Fu, Wei-zhe; Li, Lin; Lu, Ling-hong; Zhou, Lian; Shan, Zhi-hui; Tang, Gui-xiang; Shou, Hui-xia

    2013-01-01

    The Agrobacterium-mediated transformation system is the most commonly used method in soybean transformation. Screening of soybean genotypes favorable for Agrobacterium-infection and tissue regeneration is the most important step to establish an efficient genetic transformation system. In this study, twenty soybean genotypes that originated from different soybean production regions in China were screened for transient infection, regeneration capacity, and stable transgenic efficiency. Three genotypes, Yuechun 04-5, Yuechun 03-3, and Tianlong 1, showed comparable stable transgenic efficiencies with that of the previously reported American genotypes Williams 82 and Jack in our experimental system. For the Tianlong 1, the average stable transformation efficiency is 4.59%, higher than that of control genotypes (Jack and Williams 82), which is enough for further genomic research and genetic engineering. While polymerase chain reaction (PCR), LibertyLink strips, and β-glucuronidase (GUS) staining assays were used to detect the insertion and expression of the transgene, leaves painted with 135 mg/L Basta could efficiently identify the transformants. PMID:23549846

  18. Improved Agrobacterium-mediated transformation of cowpea via sonication and vacuum infiltration.

    PubMed

    Bakshi, Souvika; Sadhukhan, Ayan; Mishra, Sagarika; Sahoo, Lingaraj

    2011-12-01

    An improved method of Agrobacterium-mediated transformation of cowpea was developed employing both sonication and vacuum infiltration treatments. 4 day-old cotyledonary nodes were used as explants for co-cultivation with Agrobacterium tumefaciens strain EHA105 harbouring the binary vector pSouv-cry1Ac. Among the different injury treatments, vacuum infiltration and their combination treatments tested, sonication for 20 s followed by vacuum infiltration for 5 min with A. tumefaciens resulted in highest transient GUS expression efficiency (93% explants expressing GUS at regenerating sites). After 3 days of co-cultivation, the explants were cultured in 150 mg/l kanamycin-containing selection medium and putative transformed plants were recovered. The presence, integration and expression of nptII and cry1Ac genes in T0 transgenic plants were confirmed by polymerase chain reaction (PCR), genomic Southern and qualitative reverse transcription (RT)-PCR analysis. Western blot hybridization and enzyme-linked immunosorbent assay (ELISA) detected and demonstrated the accumulation of Cry1Ac protein in transgenic plants. The cry1Ac gene transmitted in a Mendelian fashion. The stable transformation efficiency increased by 88.4% using both sonication-assisted Agrobacterium-mediated transformation (SAAT) and vacuum infiltration than simple Agrobacterium-mediated transformation in cowpea.

  19. Agrobacterium-mediated genetic transformation of yam (Dioscorea rotundata): an important tool for functional study of genes and crop improvement.

    PubMed

    Nyaboga, Evans; Tripathi, Jaindra N; Manoharan, Rajesh; Tripathi, Leena

    2014-01-01

    Although genetic transformation of clonally propagated crops has been widely studied as a tool for crop improvement and as a vital part of the development of functional genomics resources, there has been no report of any existing Agrobacterium-mediated transformation of yam (Dioscorea spp.) with evidence of stable integration of T-DNA. Yam is an important crop in the tropics and subtropics providing food security and income to over 300 million people. However, yam production remains constrained by increasing levels of field and storage pests and diseases. A major constraint to the development of biotechnological approaches for yam improvement has been the lack of an efficient and robust transformation and regeneration system. In this study, we developed an Agrobacterium-mediated transformation of Dioscorea rotundata using axillary buds as explants. Two cultivars of D. rotundata were transformed using Agrobacterium tumefaciens harboring the binary vectors containing selectable marker and reporter genes. After selection with appropriate concentrations of antibiotic, shoots were developed on shoot induction and elongation medium. The elongated antibiotic-resistant shoots were subsequently rooted on medium supplemented with selection agent. Successful transformation was confirmed by polymerase chain reaction, Southern blot analysis, and reporter genes assay. Expression of gusA gene in transgenic plants was also verified by reverse transcription polymerase chain reaction analysis. Transformation efficiency varied from 9.4 to 18.2% depending on the cultivars, selectable marker genes, and the Agrobacterium strain used for transformation. It took 3-4 months from Agro-infection to regeneration of complete transgenic plant. Here we report an efficient, fast and reproducible protocol for Agrobacterium-mediated transformation of D. rotundata using axillary buds as explants, which provides a useful platform for future genetic engineering studies in this economically important

  20. Agrobacterium-mediated genetic transformation of yam (Dioscorea rotundata): an important tool for functional study of genes and crop improvement

    PubMed Central

    Nyaboga, Evans; Tripathi, Jaindra N.; Manoharan, Rajesh; Tripathi, Leena

    2014-01-01

    Although genetic transformation of clonally propagated crops has been widely studied as a tool for crop improvement and as a vital part of the development of functional genomics resources, there has been no report of any existing Agrobacterium-mediated transformation of yam (Dioscorea spp.) with evidence of stable integration of T-DNA. Yam is an important crop in the tropics and subtropics providing food security and income to over 300 million people. However, yam production remains constrained by increasing levels of field and storage pests and diseases. A major constraint to the development of biotechnological approaches for yam improvement has been the lack of an efficient and robust transformation and regeneration system. In this study, we developed an Agrobacterium-mediated transformation of Dioscorea rotundata using axillary buds as explants. Two cultivars of D. rotundata were transformed using Agrobacterium tumefaciens harboring the binary vectors containing selectable marker and reporter genes. After selection with appropriate concentrations of antibiotic, shoots were developed on shoot induction and elongation medium. The elongated antibiotic-resistant shoots were subsequently rooted on medium supplemented with selection agent. Successful transformation was confirmed by polymerase chain reaction, Southern blot analysis, and reporter genes assay. Expression of gusA gene in transgenic plants was also verified by reverse transcription polymerase chain reaction analysis. Transformation efficiency varied from 9.4 to 18.2% depending on the cultivars, selectable marker genes, and the Agrobacterium strain used for transformation. It took 3–4 months from Agro-infection to regeneration of complete transgenic plant. Here we report an efficient, fast and reproducible protocol for Agrobacterium-mediated transformation of D. rotundata using axillary buds as explants, which provides a useful platform for future genetic engineering studies in this economically important

  1. Agrobacterium-Mediated Transformation of Subterranean Clover (Trifolium subterraneum L.).

    PubMed Central

    Khan, MRI.; Tabe, L. M.; Heath, L. C.; Spencer, D.; Higgins, TJV.

    1994-01-01

    We have developed a rapid and reproducible transformation system for subterranean clover (Trifolium subterraneum L.) using Agrobacterium tumefaciens-mediated gene delivery. Hypocotyl segments from seeds that had been allowed to imbibe were used as explants, and regeneration was achieved via organogenesis. Glucose and acetosyringone were required in the co-cultivation medium for efficient gene transfer. DNA constructs containing four genes encoding the enzymes phosphinothricin acetyl transferase, [beta]-glucuronidase (GUS), neomycin phosphotransferase, and an [alpha]-amylase inhibitor were used to transform subterranean clover. Transgenic shoots were selected on a medium containing 50 mg/L of phosphinothricin. Four commercial cultivars of subterranean clover (representing all three subspecies) have been successfully transformed. Southern analysis revealed the integration of T-DNA into the subterranean clover genome. The expression of the introduced genes has been confirmed by enzyme assays and northern blot analyses. Transformed plants grown in the glasshouse showed resistance to the herbicide Basta at applications equal to or higher than rates recommended for killing subterranean clover in field conditions. In plants grown from the selfed seeds of the primary transformants, the newly acquired gene encoding GUS segregated as a dominant Mendelian trait. PMID:12232188

  2. Strategies to improve low copy transgenic events in Agrobacterium-mediated transformation of maize.

    PubMed

    Sivamani, Elumalai; Li, Xianggan; Nalapalli, Samson; Barron, Yoshimi; Prairie, Anna; Bradley, David; Doyle, Michele; Que, Qiudeng

    2015-12-01

    Transgenic plants containing low copy transgene insertion free of vector backbone are highly desired for many biotechnological applications. We have investigated two different strategies for increasing the percentage of low copy events in Agrobacterium-mediated transformation experiments in maize. One of the strategies is to use a binary vector with two separate T-DNAs, one T-DNA containing an intact E.coli manA gene encoding phosphomannose isomerase (PMI) as selectable marker gene cassette and another T-DNA containing an RNAi cassette of PMI sequences. By using this strategy, low copy transgenic events containing the transgenes were increased from 43 to 60 % in maize. An alternate strategy is using selectable marker gene cassettes containing regulatory or coding sequences derived from essential plant genes such as 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) or MADS box transcription factor. In this paper we demonstrate that higher percentage of low copy transgenic events can be obtained in Agrobacterium-mediated maize transformation experiments using both strategies. We propose that the above two strategies can be used independently or in combination to increase transgenic events that contain low copy transgene insertion in Agrobacterium-mediated transformation experiments.

  3. Agrobacterium-mediated genetic transformation of pineapple (Ananas comosus L., Merr.).

    PubMed

    Mhatre, Minal

    2013-01-01

    Pineapple (Ananas comosus L., Merr.) is a commercially important crop, grown in the tropical and subtropical regions. However, the crop is faced with postharvest damage and poor varietal and nutritional improvement. Being a vegetatively propagated crop, conventional breeding programs take longer time for genetic improvement, which may not necessarily successfully develop an improved cultivar. Hence, the genetic modification of pineapple is an alternative handy approach to improve pineapple. We have established an Agrobacterium-mediated transformation system using leaf bases from in vitro-grown pineapple plants. Being a monocot, acetosyringone is added to the culture medium for overnight growth of Agrobacterium and transformation to transfer a gene of interest MSI99 soybean ferritin. Leaf bases isolated from in vitro shoot cultures are treated with Agrobacterium suspension at two dilutions, 10× and 20×, for 30 min. Explants are subsequently blot dried and cultured on gelrite solidified hormone-free Pin1 medium for 2 days (cocultivation). Periodic transfer is first done to the regeneration medium (Pin1) containing cefotaxime for the suppression of Agrobacterium growth. The transformants are selected by culturing on Pin1 medium containing cefotaxime and kanamycin. Multiple shoots, regenerated in leaf bases, are further multiplied and individually rooted in the liquid RM medium amended with antibiotics to recover plants. Putative transformants are analyzed for transgene integration and expression using standard molecular biological methods of PCR, RT-PCR, and genomic Southern.

  4. Agrobacterium-mediated genetic transformation of pineapple (Ananas comosus L., Merr.).

    PubMed

    Mhatre, Minal

    2013-01-01

    Pineapple (Ananas comosus L., Merr.) is a commercially important crop, grown in the tropical and subtropical regions. However, the crop is faced with postharvest damage and poor varietal and nutritional improvement. Being a vegetatively propagated crop, conventional breeding programs take longer time for genetic improvement, which may not necessarily successfully develop an improved cultivar. Hence, the genetic modification of pineapple is an alternative handy approach to improve pineapple. We have established an Agrobacterium-mediated transformation system using leaf bases from in vitro-grown pineapple plants. Being a monocot, acetosyringone is added to the culture medium for overnight growth of Agrobacterium and transformation to transfer a gene of interest MSI99 soybean ferritin. Leaf bases isolated from in vitro shoot cultures are treated with Agrobacterium suspension at two dilutions, 10× and 20×, for 30 min. Explants are subsequently blot dried and cultured on gelrite solidified hormone-free Pin1 medium for 2 days (cocultivation). Periodic transfer is first done to the regeneration medium (Pin1) containing cefotaxime for the suppression of Agrobacterium growth. The transformants are selected by culturing on Pin1 medium containing cefotaxime and kanamycin. Multiple shoots, regenerated in leaf bases, are further multiplied and individually rooted in the liquid RM medium amended with antibiotics to recover plants. Putative transformants are analyzed for transgene integration and expression using standard molecular biological methods of PCR, RT-PCR, and genomic Southern. PMID:23179718

  5. Regeneration and Agrobacterium-mediated transformation of the apomictic species Eulaliopsis binata.

    PubMed

    Ma, Kai; Hu, Chun Gen; Xu, Bing; Yao, Jia Ling

    2013-09-01

    Protocols for regeneration and Agrobacterium-mediated transformation of the apomictic species Eulaliopsis binata were developed. Initially, seeds of four genotypes of E. binata were incubated on a callus induction Murashige and Skoog (MS) basal medium supplemented with three concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D). It was found that 36.2 % of explants developed highly friable callus on medium containing 3.0 mg l(-1) 2,4-D. Based on frequency of callus induction, the genotype Neixiang was selected for regeneration and transformation. Callus incubated on MS basal medium supplemented with 0.2 mg l(-1) α-naphthalene acetic acid and 6.0 mg l(-1) 6-furfuryl-aminopurine developed shoots. Subsequently, Agrobacterium tumefaciens strain EHA105-harboring a plasmid pCAMBIA1381 carrying a hygromycin phosphotransferase (hpt) resistance gene and a synthetic green fluorescent protein (GFP) gene, both driven by the cauliflower mosaic virus 35S promoter-was used for transformation system. Putative transgenic callus was obtained following two cycles of hygromycin selection. Expression of the transgene(s) in putative transgenic callus was analyzed using the GFP detection. Molecular identification of putative transformed shoots was performed by polymerase chain reaction and Southern blot analysis to confirm presence and integration of the hpt gene.

  6. Agrobacterium-mediated transformation of peanut (Arachis hypogaea L.) embryo axes and the development of transgenic plants.

    PubMed

    McKently, A H; Moore, G A; Doostdar, H; Niedz, R P

    1995-08-01

    Transgenic peanut (Arachis hypogaea L.) plants have been produced using an Agrobacterium-mediated transformation system. Zygotic embryo axes from mature seed were cocultured with Agrobacterium tumefaciens strain EHA101 harboring a binary vector that contained the genes for the scorable marker B-glucuronidase (GUS) and the selectable marker neomycin phosphotransferase II. Nine percent of the germinated seedlings were GUS+. Polymerase chain reaction analysis confirmed that GUS+ shoots and T1 progeny contained T-DNA. Molecular characterization of one primary transformant and its T1 and T2 progeny plants established that T-DNA was integrated into the host genome. PMID:24186625

  7. Plant Cell Division Analyzed by Transient Agrobacterium-Mediated Transformation of Tobacco BY-2 Cells.

    PubMed

    Buschmann, Henrik

    2016-01-01

    The continuing analysis of plant cell division will require additional protein localization studies. This is greatly aided by GFP-technology, but plant transformation and the maintenance of transgenic lines can present a significant technical bottleneck. In this chapter I describe a method for the Agrobacterium-mediated genetic transformation of tobacco BY-2 cells. The method allows for the microscopic analysis of fluorescence-tagged proteins in dividing cells in within 2 days after starting a coculture. This transient transformation procedure requires only standard laboratory equipment. It is hoped that this rapid method would aid researchers conducting live-cell localization studies in plant mitosis and cytokinesis.

  8. Transgenic sugar beet tolerant to imidazolinone obtained by Agrobacterium-mediated transformation.

    PubMed

    Kishchenko, E M; Komarnitskii, I K; Kuchuk, N V

    2011-01-01

    Sugar beet is highly sensitive to imidazolinone herbicides thus rotational restrictions exist. In order to develop imidazolinone tolerant sugar beets als gene from Arabidopsis thaliana encoding acetolactate synthase with S653N mutation was used for genetic transformation. Transgenic sugar beet plants were obtained by Agrobacterium-mediated transformation of aseptic seedlings using vacuum-infiltration. The efficiency of genetic transformation was 5.8%. RT-PCR analysis of obtained plants revealed accumulation of specific als transcript. The resistance to imidazolinone was proved for developed transgenic sugar beet plants in vitro and in greenhouse conditions after spraying with imazethapyr (Pursuit, BASF).

  9. Agrobacterium-mediated genetic transformation of Prunus salicina.

    PubMed

    Urtubia, Carolina; Devia, Jessica; Castro, Alvaro; Zamora, Pablo; Aguirre, Carlos; Tapia, Eduardo; Barba, Paola; Dell Orto, Paola; Moynihan, Michael R; Petri, César; Scorza, Ralph; Prieto, Humberto

    2008-08-01

    We report Agrobacterium tumefaciens-mediated transformation of two Prunus salicina varieties, 'Angeleno' and 'Larry Anne', using a modification of the hypocotyl slice technique previously described for P. domestica. Regeneration rates on thidiazuron (TDZ) and indole-3-butyric acid (IBA) supplemented Murashige and Skoog (MS) media reached 11% for 'Angeleno' and 19% for 'Larry Anne' hypocotyl slices. Transformation using Agrobacterium tumefaciens GV3101 harboring a plasmid with the neomycin phosphotransferase II (nptII) and the green fluorescent protein (gfp) genes produced ten independent lines, six from 'Angeleno' and four from 'Larry Anne', representing transformation efficiencies of 0.8 and 0.3%, respectively, relative to the initial number of hypocotyl slices. Plants of six lines were found to produce the transgene encoded mRNAs. DNA blotting demonstrated the presence of transgene sequences in trees from five lines after 18 months of growth in the greenhouse.

  10. Agrobacterium-mediated genetic transformation of Prunus salicina

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We report Agrobacterium tumefaciens-mediated transformation from hypocotyls slices of two Prunus salicina varieties, 'Angeleno' and 'Larry Anne', using a modification of the technique previously described for P. domestica. Regeneration rates on thidiazuron (TDZ) and indole-3-butyric acid (IBA) supp...

  11. Agrobacterium-mediated transformation of Ruta graveolens L.

    PubMed

    Lièvre, Karine; Tran, Thi Lê Minh; Doerper, Sébastien; Hehn, Alain; Lacoste, Paul; Thomasset, Brigitte; Bourgaud, Frédéric; Gontier, Eric

    2009-01-01

    Agrobacterium tumefaciens is used to develop a genetic transformation method for a medicinal plant Ruta graveolens. The direct plant regeneration strategy is preferred to callus line establishment. In vitro seedlings, 2- -to 3-wk-old, are used to excise hypocotyls and co-cultivated for 3 d with A. tumefaciens strain C58C1Rif containing plasmid pTDE4 harbouring neomycin phosphotransferase (npt II, kanamycin resistance) and beta-glucuronidase encoding genes. The Southern blot analysis has shown that 78% kanamycin resistant plants contain gene encoding beta-glucuronidase. The GUS histochemical assay shows that 67% transgenic plants exhibit the corresponding enzymatic activity. Routine transformation efficiency of R. graveolens L. is 11% and could reach up to 22%. Transgenic plants are grown in the greenhouse within 4 months after the initial seedlings.

  12. Agrobacterium-mediated transformation of maize (Zea mays) with Cre-lox site specific recombination cassettes in BIBAC vectors.

    PubMed

    Vega, Juan M; Yu, Weichang; Han, Fangpu; Kato, Akio; Peters, Eric M; Zhang, Zhanyuan J; Birchler, James A

    2008-04-01

    The Cre/loxP site-specific recombination system has been applied in various plant species including maize (Zea mays) for marker gene removal, gene targeting, and functional genomics. A BIBAC vector system was adapted for maize transformation with a large fragment of genetic material including a herbicide resistance marker gene, a 30 kb yeast genomic fragment as a marker for fluorescence in situ hybridization (FISH), and a 35S-lox-cre recombination cassette. Seventy-five transgenic lines were generated from Agrobacterium-mediated transformation of a maize Hi II line with multiple B chromosomes. Eighty-four inserts have been localized among all 10 A chromosome pairs by FISH using the yeast DNA probe together with a karyotyping cocktail. No inserts were found on the B chromosomes; thus a bias against the B chromosomes by the Agrobacterium-mediated transformation was revealed. The expression of a cre gene was confirmed in 68 of the 75 transgenic lines by a reporter construct for cre/lox mediated recombination. The placement of the cre/lox site-specific recombination system in many locations in the maize genome will be valuable materials for gene targeting and chromosome engineering.

  13. Genetic transformation of wheat via Agrobacterium-mediated DNA delivery.

    PubMed

    Sparks, Caroline A; Doherty, Angela; Jones, Huw D

    2014-01-01

    The method described involves an initial incubation of wheat immature embryos in a liquid culture of Agrobacterium tumefaciens. The Agrobacterium strain is engineered to contain a binary vector with a gene of interest and a selectable marker gene placed between the T-DNA borders; the T-DNA is the region transferred to the plant cells, thus harnessing the bacterium's natural ability to deliver specific DNA into host cells. Following the initial inoculation with the Agrobacterium, the embryos are co-cultivated for several days after which the Agrobacterium is selectively destroyed using an antibiotic. Tissue culture of the embryos on plant media with a correct balance of hormones allows embryogenic callus formation followed by regeneration of plantlets, and in the later stages of tissue culture a selectable marker (herbicide) is included to minimize the incidence of non-transformed plants. This protocol has been used successfully to generate transformed plants of a wide range of wheat varieties, both spring and winter bread wheats (T. aestivum L.) and durum wheats (T. turgidum L.). PMID:24243208

  14. Efficient gene targeting in Penicillium chrysogenum using novel Agrobacterium-mediated transformation approaches.

    PubMed

    de Boer, Paulo; Bronkhof, Jurian; Dukiќ, Karolina; Kerkman, Richard; Touw, Hesselien; van den Berg, Marco; Offringa, Remko

    2013-12-01

    The industrial production of β-lactam antibiotics by Penicillium chrysogenum has increased tremendously over the last decades, however, further optimization via classical strain and process improvement has reached its limits. The availability of the genome sequence provides new opportunities for directed strain improvement, but this requires the establishment of an efficient gene targeting (GT) system. Recently, mutations affecting the non-homologous end joining (NHEJ) pathway were shown to increase GT efficiencies following PEG-mediated DNA transfer in P. chrysogenum from 1% to 50%. Apart from direct DNA transfer many fungi can efficiently be transformed using the T-DNA transfer system of the soil bacterium Agrobacterium tumefaciens, however, for P. chrysogenum no robust system for Agrobacterium-mediated transformation was available. We obtained efficient AMT of P. chrysogenum spores with the nourseothricin acetyltransferase gene as selection marker, and using this system we investigated if AMT in a NHEJ mutant background could further enhance GT efficiencies. In general, AMT resulted in higher GT efficiencies than direct DNA transfer, although the final frequencies depended on the Agrobacterium strain and plasmid backbone used. Providing overlapping and complementing fragments on two different plasmid backbones via the same Agrobacterium host was shown to be most effective. This so-called split-marker or bi-partite method resulted in highly efficient GT (>97%) almost exclusively without additional ectopic T-DNA insertions. As this method provides for an efficient GT method independent of protoplasts, it can be applied to other fungi for which no protoplasts can be generated or for which protoplast transformation leads to varying results.

  15. Agrobacterium-mediated transformation of tomato with the ICE1 transcription factor gene.

    PubMed

    Juan, J X; Yu, X H; Jiang, X M; Gao, Z; Zhang, Y; Li, W; Duan, Y D; Yang, G

    2015-01-30

    ICE1 genes play a very important role in plants in cold conditions. To improve the cold resistance of tomato, the ICE1 gene of Arabidopsis thaliana was used to construct the plant expression vector p3301-ICE1, and was overexpressed in tomato through Agrobacterium-mediated transformation. Five strains of resistant plants were obtained. PCR and half-quantitative results showed that the ICE1 gene was transferred to tomato; three strains tested positive. After low-temperature stress treatment, praline content and peroxide and catalase activities in the transgenic tomato plants were higher compared with non-transgenic controls, while malondialdehyde content was clearly lower.

  16. Agrobacterium-mediated transformation of the recalcitrant Vanda Kasem's Delight orchid with higher efficiency.

    PubMed

    Gnasekaran, Pavallekoodi; Antony, Jessica Jeyanthi James; Uddain, Jasim; Subramaniam, Sreeramanan

    2014-01-01

    The presented study established Agrobacterium-mediated genetic transformation using protocorm-like bodies (PLBs) for the production of transgenic Vanda Kasem's Delight Tom Boykin (VKD) orchid. Several parameters such as PLB size, immersion period, level of wounding, Agrobacterium density, cocultivation period, and concentration of acetosyringone were tested and quantified using gusA gene expression to optimize the efficiency of Agrobacterium-mediated genetic transformation of VKD's PLBs. Based on the results, 3-4 mm PLBs wounded by scalpel and immersed for 30 minutes in Agrobacterium suspension of 0.8 unit at A 600 nm produced the highest GUS expression. Furthermore, cocultivating infected PLBs for 4 days in the dark on Vacin and Went cocultivation medium containing 200 μM acetosyringone enhanced the GUS expression. PCR analysis of the putative transformants selected in the presence of 250 mg/L cefotaxime and 30 mg/L geneticin proved the presence of wheatwin1, wheatwin2, and nptII genes.

  17. Optimization of Agrobacterium-mediated transformation conditions in mature embryos of elite wheat.

    PubMed

    Ding, Liping; Li, Shengchun; Gao, Jianming; Wang, Yuesheng; Yang, Guangxiao; He, Guangyuan

    2009-01-01

    Immature embryos have been used frequently as target tissues in the genetical transformation of wheat. However, obtaining a large number of high quality immature embryos throughout the year is a laborious and delicate process, because of the need to cultivate the plants under controlled conditions. To circumvent this, we have employed mature embryos rather than immature ones as starter explants for Agrobacterium-mediated transformation of an elite wheat (Triticum aestivum L.) cultivar EM12. The neomycin phosphotransferase II, (npt II) and beta-glucuronidase (gus) genes were used as selectable and screenable marker genes, respectively, to assess and optimize the performance of T-DNA delivery. With the aid of an orthogonal design, the effect of four factors in combination on transfer DNA (T-DNA) delivery was studied. These factors were preculture duration, different kinds of inoculation, length of inoculation and co-culture condition. Optimal conditions for T-DNA delivery were obtained for mature embryos precultured for 14 days, followed by immersing in inoculation suspension with full strength Murashige and Skoog (MS) salts in darkness at 23-25 degrees C for 3 h, and then co-culturing with Agrobacterium under desiccating condition in the dark at 23-24 degrees C for 2-3 days. Complete analysis of transgene insertion demonstrated that the optimized method for Agrobacterium-mediated transformation of mature embryos of wheat was efficient and practicable.

  18. Agrobacterium-mediated transformation of friable embryogenic calli and regeneration of transgenic cassava.

    PubMed

    Bull, S E; Owiti, J A; Niklaus, M; Beeching, J R; Gruissem, W; Vanderschuren, H

    2009-01-01

    Agrobacterium-mediated transformation of friable embryogenic calli (FEC) is the most widely used method to generate transgenic cassava plants. However, this approach has proven to be time-consuming and can lead to changes in the morphology and quality of FEC, influencing regeneration capacity and plant health. Here we present a comprehensive, reliable and improved protocol, taking approximately 6 months, that optimizes Agrobacterium-mediated transformation of FEC from cassava model cultivar TMS60444. We cocultivate the FEC with Agrobacterium directly on the propagation medium and adopt the extensive use of plastic mesh for easy and frequent transfer of material to new media. This minimizes stress to the FEC cultures and permits a finely balanced control of nutrients, hormones and antibiotics. A stepwise increase in antibiotic concentration for selection is also used after cocultivation with Agrobacterium to mature the transformed FEC before regeneration. The detailed information given here for each step should enable successful implementation of this technology in other laboratories, including those being established in developing countries where cassava is a staple crop.

  19. Agrobacterium-mediated transformation of apricot (Prunus armeniaca L.) leaf explants.

    PubMed

    Petri, César; Wang, Hong; Alburquerque, Nuria; Faize, Mohamed; Burgos, Lorenzo

    2008-08-01

    A protocol for Agrobacterium-mediated stable transformation for scored, whole leaf explants of the apricot (Prunus armeniaca) cultivar Helena was developed. Regenerated shoots were selected using a two-step increased concentrations of paromomycin sulphate. Different factors affecting survival of transformed buds, including possible toxicity of green fluorescent protein (GFP) and time of exposure to high cytokine concentration in the regeneration medium, were examined. Transformation efficiency, based on PCR analysis of individual putative transformed shoots from independent lines was 5.6%, when optimal conditions for bud survival were provided. Southern blot analysis on four randomly chosen PCR-positive shoots confirmed the presence of the nptII transgene. This is the first time that stable transformation of an apricot cultivar is reported and constitutes also one of the few reports on the transformation of Prunus cultivars.

  20. Agrobacterium-mediated transformation of two Serbian potato cultivars (Solanum tuberosum L. cv. Dragacevka and cv. Jelica)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An efficient protocol for Agrobacterium-mediated transformation of Serbian potato cultivars Dragacevka and Jelica, enabling the introduction of oryzacystatin genes OCI and OCII, was established. Starting with leaf explants a two-stage transformation protocol combining procedures of Webb and Wenzler...

  1. Agrobacterium-mediated transformation of tomato (Solanum lycopersicum L.) using the expansin 10 (CsEXP10) gene.

    PubMed

    Sun, Y D; Luo, W R; Sun, S Y; Ni, L

    2015-12-08

    The cucumber expansin 10 (CsEXP10) gene was previously cloned from young cucumber fruits but its role has not been defined. To determine the role of this gene in plant growth and development, a CsEXP10 gene transformation system was established. The open reading frame of the gene was inserted behind the CaMV35S promoter of vector pCAMBIA1301, and the construct was introduced into tomato plants by Agrobacterium-mediated transformation. In total, 19 kanamycin-positive lines were produced and nine independent transgenic lines were identified by β-glucuronidase and polymerase chain reaction (PCR) analysis. Quantitative real-time PCR analysis showed that levels of the CsEXP10 transcript were higher in transgenic lines than in a non-transgenic line.

  2. Efficient and high-throughput vector construction and Agrobacterium-mediated transformation of Arabidopsis thaliana suspension-cultured cells for functional genomics.

    PubMed

    Ogawa, Yoichi; Dansako, Tomoko; Yano, Kentaro; Sakurai, Nozomu; Suzuki, Hideyuki; Aoki, Koh; Noji, Masaaki; Saito, Kazuki; Shibata, Daisuke

    2008-02-01

    We established a large-scale, high-throughput protocol to construct Arabidopsis thaliana suspension-cultured cell lines, each of which carries a single transgene, using Agrobacterium-mediated transformation. We took advantage of RIKEN Arabidopsis full-length (RAFL) cDNA clones and the Gateway cloning system for high-throughput preparation of binary vectors carrying individual full-length cDNA sequences. Throughout all cloning steps, multiple-well plates were used to treat 96 samples simultaneously in a high-throughput manner. The optimal conditions for Agrobacterium-mediated transformation of 96 independent binary vector constructs were established to obtain transgenic cell lines efficiently. We evaluated the protocol by generating transgenic Arabidopsis T87 cell lines carrying individual 96 metabolism-related RAFL cDNA fragments, and showed that the protocol was useful for high-throughput and large-scale production of gain-of-function lines for functional genomics.

  3. Overexpression of several Arabidopsis histone genes increases agrobacterium-mediated transformation and transgene expression in plants.

    PubMed

    Tenea, Gabriela N; Spantzel, Joerg; Lee, Lan-Ying; Zhu, Yanmin; Lin, Kui; Johnson, Susan J; Gelvin, Stanton B

    2009-10-01

    The Arabidopsis thaliana histone H2A-1 is important for Agrobacterium tumefaciens-mediated plant transformation. Mutation of HTA1, the gene encoding histone H2A-1, results in decreased T-DNA integration into the genome of Arabidopsis roots, whereas overexpression of HTA1 increases transformation frequency. To understand the mechanism by which HTA1 enhances transformation, we investigated the effects of overexpression of numerous Arabidopsis histones on transformation and transgene expression. Transgenic Arabidopsis containing cDNAs encoding histone H2A (HTA), histone H4 (HFO), and histone H3-11 (HTR11) displayed increased transformation susceptibility, whereas histone H2B (HTB) and most histone H3 (HTR) cDNAs did not increase transformation. A parallel increase in transient gene expression was observed when histone HTA, HFO, or HTR11 overexpression constructs were cotransfected with double- or single-stranded forms of a gusA gene into tobacco (Nicotiana tabacum) protoplasts. However, these cDNAs did not increase expression of a previously integrated transgene. We identified the N-terminal 39 amino acids of H2A-1 as sufficient to increase transient transgene expression in plants. After transfection, transgene DNA accumulates more rapidly in the presence of HTA1 than with a control construction. Our results suggest that certain histones enhance transgene expression, protect incoming transgene DNA during the initial stages of transformation, and subsequently increase the efficiency of Agrobacterium-mediated transformation.

  4. A rapid and stable Agrobacterium-mediated transformation method of a medicinal plant Chelone glabra L.

    PubMed

    Gao, Zhenrui; Li, Ying; Chen, Jinhua; Chen, Zhixing; Cui, Min-Long

    2015-03-01

    Transformation approach is a useful tool for the study of gene function, the mechanism of molecular regulation, and increase usefulness of components by reverse genetic approach in plants. In this study, we developed a stable and rapid method for Agrobacterium-mediated transformation of a medicinal plant Chelone glabra L. using leaf explants. Stable transformants were obtained using Agrobacterium tumefaciens strains GV2260 and GV3101 that harbored the binary vector pBI121 and contained the neomycin phosphotransferase gene (NPT II) as a selectable marker and a reporter gene β-glucuronidase (GUS). Putative transformants were identified by kanamycin selection and a histochemical assay. PCR and Southern blot analysis confirmed the integration of the GUS gene into transformed genomes as well as detected stable expression of the β-glucuronidase gene (GUS) by RT-PCR. Resulting transformed plants had morphologically normal phenotypes. This method requires two changes of medium and few leaf explants as well as the transformation efficiency of 2-8 % after 2-3 months of inoculation. This method can provide a quick and economical transformation method for reverse genetic approach to change the secondary metabolic pathway to increase useful components in C. glabra.

  5. Stable Agrobacterium-Mediated Transformation of Maritime Pine Based on Kanamycin Selection

    PubMed Central

    Alvarez, José M.; Ordás, Ricardo J.

    2013-01-01

    An efficient transformation protocol based on kanamycin selection was developed for Agrobacterium-mediated transformation of maritime pine embryonal masses. The binary vector pBINUbiGUSint, which contained neomycin phosphotransferase II (nptII) as a selectable marker gene and β-glucuronidase (uidA) as a reporter gene, was used for transformation studies. Different factors, such as embryogenic line, bacterial strain, bacterial concentration, and coculture duration, were examined and optimized. For selection of transformants, 15 mgL−1 kanamycin was used. The highest transformation efficiency (11.4 events per gram of fresh mass) was achieved when a vigorously growing embryonal mass (embryogenic line L01) was cocultivated with Agrobacterium strain AGL1 at the optical density (OD600 nm) of 0.3 for 72 h. Evidence of the stable transgene integration was obtained by polymerase chain reaction for the nptII and uidA genes and expression of the uidA gene. Maturation capacity of the transgenic lines was negatively affected by the transformation process. Induction of axillary shoots by preculturing the embryos with benzyladenine allowed overcoming the low maturation rates of some transformed lines. The transgenic embryos were germinated and the axillar shoots were rooted. Transgenic plants were transferred to potting substrate showing normal growth. PMID:24376383

  6. Stable Agrobacterium-mediated transformation of maritime pine based on kanamycin selection.

    PubMed

    Alvarez, José M; Ordás, Ricardo J

    2013-01-01

    An efficient transformation protocol based on kanamycin selection was developed for Agrobacterium-mediated transformation of maritime pine embryonal masses. The binary vector pBINUbiGUSint, which contained neomycin phosphotransferase II (nptII) as a selectable marker gene and β -glucuronidase (uidA) as a reporter gene, was used for transformation studies. Different factors, such as embryogenic line, bacterial strain, bacterial concentration, and coculture duration, were examined and optimized. For selection of transformants, 15 mgL(-1) kanamycin was used. The highest transformation efficiency (11.4 events per gram of fresh mass) was achieved when a vigorously growing embryonal mass (embryogenic line L01) was cocultivated with Agrobacterium strain AGL1 at the optical density (OD(600 nm)) of 0.3 for 72 h. Evidence of the stable transgene integration was obtained by polymerase chain reaction for the nptII and uidA genes and expression of the uidA gene. Maturation capacity of the transgenic lines was negatively affected by the transformation process. Induction of axillary shoots by preculturing the embryos with benzyladenine allowed overcoming the low maturation rates of some transformed lines. The transgenic embryos were germinated and the axillar shoots were rooted. Transgenic plants were transferred to potting substrate showing normal growth.

  7. Agrobacterium-mediated transformation of Vitis Cv. Monastrell suspension-cultured cells: Determination of critical parameters.

    PubMed

    Chu, Mingyu; Quiñonero, Carmen; Akdemir, Hülya; Alburquerque, Nuria; Pedreño, María Ángeles; Burgos, Lorenzo

    2016-05-01

    Although some works have explored the transformation of differentiated, embryogenic suspension-cultured cells (SCC) to produce transgenic grapevine plants, to our knowledge this is one of the first reports on the efficient transformation of dedifferentiated Vitis vinifera cv Monastrell SCC. This protocol has been developed using the sonication-assisted Agrobacterium-mediated transformation (SAAT) method. A construct harboring the selectable nptII and the eyfp/IV2 marker genes was used in the study and transformation efficiencies reached over 50 independent transformed SCC per gram of infected cells. Best results were obtained when cells were infected at the exponential phase. A high density plating (500 mg/dish) gave significantly better results. As selective agent, kanamycin was inefficient for the selection of Monastrell transformed SCC since wild type cells were almost insensitive to this antibiotic whereas application of paromomycin resulted in very effective selection. Selected eyfp-expressing microcalli were grown until enough tissue was available to scale up a new transgenic SCC. These transgenic SCC lines were evaluated molecularly and phenotypically demonstrating the presence and integration of both transgenes, the absence of Agrobacterium contamination and the ability of the transformed SCC to grow in highly selective liquid medium. The methodology described here opens the possibility of improving the production of valuable metabolites. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:725-734, 2016.

  8. Optimization of factors affecting Agrobacterium-mediated transformation of Micro-Tom tomatoes.

    PubMed

    Guo, M; Zhang, Y L; Meng, Z J; Jiang, J

    2012-03-16

    Micro-Tom is the smallest known variety of tomatoes. An orthogonal experimental design L(16) (4(5)) was used to optimize Agrobacterium-mediated transformation of cotyledon explants of Lycopersicon esculentum cv. Micro-Tom. Four parameters were investigated to determine their effect on transformation frequency: the concentration of bacterial suspension, time of dip in bacterial suspension, co-cultivation time, and concentration of carbenicillin. We also examined the effect of these parameters on contamination rate, necrosis rate, mortality, cut-surface browning rate, and undamaged explant rate. Both the bacterial and carbenicillin concentrations had a significant influence on the rate of infected explants. The time of co-cultivation also had a significant influence on the transformation parameters. The optimal transformation protocol consisted of an Agrobacterium suspension of 0.5 × 10(8) cells/mL (OD(600) = 0.5) and an infection time of 5 min, one day of co-cultivation and 500 mg/L carbenicillin. Under these conditions, the transformation efficiency of the shoots reached 5.1%; the mean transformation frequency was 3.9% (N = 838).

  9. Enhanced Agrobacterium-mediated transformation efficiencies in monocot cells is associated with attenuated defense responses.

    PubMed

    Zhang, Wan-Jun; Dewey, Ralph E; Boss, Wendy; Phillippy, Brian Q; Qu, Rongda

    2013-02-01

    Plant defense responses can lead to altered metabolism and even cell death at the sites of Agrobacterium infection, and thus lower transformation frequencies. In this report, we demonstrate that the utilization of culture conditions associated with an attenuation of defense responses in monocot plant cells led to highly improved Agrobacterium-mediated transformation efficiencies in perennial ryegrass (Lolium perenne L.). The removal of myo-inositol from the callus culture media in combination with a cold shock pretreatment and the addition of L-Gln prior to and during Agrobacterium-infection resulted in about 84 % of the treated calluses being stably transformed. The omission of myo-inositol from the callus culture media was associated with the failure of certain pathogenesis related genes to be induced after Agrobacterium infection. The addition of a cold shock and supplemental Gln appeared to have synergistic effects on infection and transformation efficiencies. Nearly 60 % of the stably transformed calluses regenerated into green plantlets. Calluses cultured on media lacking myo-inositol also displayed profound physiological and biochemical changes compared to ones cultured on standard growth media, such as reduced lignin within the cell walls, increased starch and inositol hexaphosphate accumulation, enhanced Agrobacterium binding to the cell surface, and less H(2)O(2) production after Agrobacterium infection. Furthermore, the cold treatment greatly reduced callus browning after infection. The simple modifications described in this report may have broad application for improving genetic transformation of recalcitrant monocot species.

  10. AgarTrap: a simplified Agrobacterium-mediated transformation method for sporelings of the liverwort Marchantia polymorpha L.

    PubMed

    Tsuboyama, Shoko; Kodama, Yutaka

    2014-01-01

    The liverwort Marchantia polymorpha L. is being developed as an emerging model plant, and several transformation techniques were recently reported. Examples are biolistic- and Agrobacterium-mediated transformation methods. Here, we report a simplified method for Agrobacterium-mediated transformation of sporelings, and it is termed Agar-utilized Transformation with Pouring Solutions (AgarTrap). The procedure of the AgarTrap was carried out by simply exchanging appropriate solutions in a Petri dish, and completed within a week, successfully yielding sufficient numbers of independent transformants for molecular analysis (e.g. characterization of gene/protein function) in a single experiment. The AgarTrap method will promote future molecular biological study in M. polymorpha.

  11. Efficient soybean regeneration and Agrobacterium-mediated transformation using a whole cotyledonary node as an explant.

    PubMed

    Zhang, Fuli; Chen, Can; Ge, Honglian; Liu, Jinmei; Luo, Yunling; Liu, Kun; Chen, Long; Xu, Kedong; Zhang, Yi; Tan, Guangxuan; Li, Chengwei

    2014-01-01

    An optimized regeneration and Agrobacterium-mediated transformation protocol based on whole cotyledonary node explants was developed in soybean (Glycine max) cultivar Zhong Huang 13. Adding 6-benzylaminopurine (BAP) in a germinating medium could significantly increase regeneration efficiency; the optimal BAP concentration for shoot formation was 0.5 mg/L. The concentrations of plant growth regulators in a shoot induction medium were optimized by the orthogonal test [L9 (3(3))]. The best combination for shoot regeneration was a medium of Murashige & Skoog salts with B5 vitamins (MSB) supplemented with 3.5 mg/L BAP, 0.2 mg/L indole-3-butyric acid (IBA), and 0.2 mg/L kinetin (KT). Under this favorable condition, one node could regenerate 28-30 shoots. Soybean whole cotyledonary nodes were transformed by inoculation with A. tumefaciens strain EHA105 harboring a vector pBI121 containing a β-glucuronidase gene (gus). GUS assay, polymerase chain reaction, and Southern blot analysis indicated that the gus gene was transformed into soybean plants with 23.1% transformation efficiency. Transgenic plants could be obtained within 5-6 weeks, which was about 4 weeks less than that of a traditional single cotyledonary node method.

  12. Deletion of host histone acetyltransferases and deacetylases strongly affects Agrobacterium-mediated transformation of Saccharomyces cerevisiae.

    PubMed

    Soltani, Jalal; van Heusden, Gerard Paul H; Hooykaas, Paul J J

    2009-09-01

    Agrobacterium tumefaciens is a plant pathogen that genetically transforms plant cells by transferring a part of its Ti-plasmid, the T-strand, to the host cell. Under laboratory conditions, it can also transform cells from many different nonplant organisms, including the yeast Saccharomyces cerevisiae. Collections of S. cerevisiae strains have been developed with systematic deletion of all coding sequences. Here, we used these collections to identify genes involved in the Agrobacterium-mediated transformation (AMT) of S. cerevisiae. We found that deletion of genes (GCN5, NGG1, YAF9 and EAF7) encoding subunits of the SAGA, SLIK, ADA and NuA4 histone acetyltransferase complexes highly increased the efficiency of AMT, while deletion of genes (HDA2, HDA3 and HST4) encoding subunits of histone deacetylase complexes decreased AMT. These effects are specific for AMT as the efficiency of chemical (lithium acetate) transformation was not or only slightly affected by these deletions. Our data are consistent with a positive role of host histone deacetylation in AMT.

  13. [Agrobacterium-mediated transformation of LJAMP2 gene into 'Red Sun' kiwifruit and its molecular identification].

    PubMed

    Zhou, Yue; Zhao, Xupeng; Wu, Xiuhua; Zhang, Yanling; Zhang, Lin; Luo, Keming; Tang, Shaohu

    2014-06-01

    Bacterial canker caused by Pseudomonas syringae pv. Actinidiae is one of the most important diseases of kiwifruit (Actinidia chinensis) and leads to considerable yield losses. In order to obtain transgenic plants with resistance for 'Red Sun' kiwifruit to canker disease, a non-specific lipid transfer protein-like antimicrobial protein gene (LJAMP2) from motherwort (Leonurus japonicus) was introduced into 'Red Sun' kiwifruit through Agrobacterium-mediated transformation. After two days of co-cultivation with A. tumefaciens strain LBA4404 harboring 35S:LJAMP2, the transformed explants were transferred to the selection medium containing 25 mg/L kanamycin+3.0 mg/L BA+1.0 mg/L NAA. The regeneration efficiency of kanamycin-resistant shoots reached to 85%. All (100%) of kanamycin-resistant shoots rooted on half-strength MS medium supplemented with 0.8 mg/L IBA and a total of 40 regenerated plantlets were obtained. PCR and histochemical GUS activity analysis show that 23 of 40 lines (57.50%) were positive, suggesting that the LJAMP2 gene was integrated into the genome of 'Red Sun' kiwifruit. Taken together, we established an efficient genetic transformation method for 'Red Sun' kiwifruit using A. tumefaciens and the transformation frequency reached 5.11%. This protocol will be useful for the genetic breeding of 'Red Sun' kiwifruit for improvement of disease resistance.

  14. An improved plant regeneration and Agrobacterium - mediated transformation of red pepper (Capsicum annuum L.).

    PubMed

    Kumar, R Vinoth; Sharma, V K; Chattopadhyay, B; Chakraborty, S

    2012-10-01

    Capsicum annuum (red pepper) is an important spice cum vegetable crop in tropical and subtropical countries. Here, we report an effective and reproducible auxin free regeneration method for six different red pepper cultivars (ACA-10, Kashi Anmol, LCA-235, PBC-535, Pusa Jwala and Supper) using hypocotyl explants and an efficient Agrobacterium-mediated transformation protocol. The explants (hypocotyls, cotyledonary leaves and leaf discs) collected from axenic seedlings of six red pepper cultivars were cultured on either hormone free MS medium or MS medium supplemented with BAP alone or in combination with IAA. Inclusion of IAA in the regeneration medium resulted in callus formation at the cut ends of explants, formation of rosette leaves and ill defined shoot buds. Regeneration of shoot buds could be achieved from hypocotyls grown in MS medium supplemented with different concentrations of BAP unlike other explants which failed to respond. Incorporation of GA3 in shoot elongation medium at 0.5 mg/l concentration enhanced the elongation in two cultivars, LCA-235 and Supper, while other cultivars showed no significant response. Chilli cultivar, Pusa Jwala was transformed with βC1 ORF of satellite DNA β molecule associated with Chilli leaf curl Joydebpur virus through Agrobacterium tumefaciens. Transgene integration in putative transformants was confirmed by PCR and Southern hybridization analysis.

  15. Agrobacterium-mediated transformation of oat (Avena sativa L.) cultivars via immature embryo and leaf explants.

    PubMed

    Gasparis, Sebastian; Bregier, Cezary; Orczyk, Waclaw; Nadolska-Orczyk, Anna

    2008-11-01

    This paper reports on the successful Agrobacterium-mediated transformation of oat, and on some factors influencing this process. In the first step of the experiments, three cultivars, two types of explant, and three combinations of strain/vectors, which were successfully used for transformation of other cereals were tested. Transgenic plants were obtained from the immature embryos of cvs. Bajka, Slawko and Akt and from leaf base explants of cv. Bajka after transformation with A. thumefaciens strain LBA4404(pTOK233). The highest transformation rate (12.3%) was obtained for immature embryos of cv. Bajka. About 79% of the selected plants proved to be transgenic; however, only 14.3% of the T(0) plants and 27.5% of the T(1) showed GUS expression. Cell competence of both types of explant differed in terms of their transformation ability and transgene expression. The next step of the study was to test the suitability for oat transformation of the pGreen binary vector combined with different selection cassettes: nptII or bar under the nos or 35S promoter. Transgenic plants were selected in combinations transformed with nos::nptII, 35S::nptII and nos::bar. The highest transformation efficiency (5.3%) was obtained for cv. Akt transformed with nos::nptII. A detailed analysis of the T(0) plants selected from a given callus line and their progeny revealed that they were the mixture of transgenic, chimeric-transgenic and non-transgenic individuals. Southern blot analysis of T(0) and T(1) showed simple integration pattern with the low copy number of the introduced transgenes.

  16. Improved Agrobacterium-mediated transformation and high efficiency of root formation from hypocotyl meristem of spring Brassica napus 'Precocity' cultivar.

    PubMed

    Liu, X X; Lang, S R; Su, L Q; Liu, X; Wang, X F

    2015-01-01

    Rape seed (Brassica napus L.) is one of the most important oil seed crops in the world. Genetic manipulation of rapeseed requires a suitable tissue culture system and an efficient method for plant regeneration, as well as an efficient transformation procedure. However, development of transgenic B. napus has been problematic, and current studies are limited to cultivated varieties. In this study, we report a protocol for regeneration of transgenic rape after Agrobacterium-mediated transformation of hypocotyls from the spring B. napus 'Precocity' cultivar. We analyzed the effects of plant growth regulators in the medium on regeneration. Additionally, factors affecting the transformation efficiency, including seedling age, Agrobacterium concentration, infection time, and co-cultivation time, were assessed by monitoring GUS expression. Results from these experiments revealed that transformation was optimized when the meristematic parts of the hypocotyls were taken from 8 day-old seedlings, cultured on Murashinge and Skoog basal media containing 0.1 mg/L 1-naphthaleneacetic acid and 2.5 mg/L 6-benzylaminopurine, and incubated in Agrobacterium suspension (OD600 = 0.5) for 3 to 5 min, followed by 2 days of co-cultivation. Integration of T-DNA into the plant genome was confirmed by polymerase chain reaction (PCR), b-glucuronidase histochemical staining, and quantitative real-time PCR. The protocols developed for regeneration, transformation, and rooting described in this study could help to accelerate the development of transgenic spring rape varieties with novel features. PMID:26681030

  17. Improved Agrobacterium-mediated transformation and high efficiency of root formation from hypocotyl meristem of spring Brassica napus 'Precocity' cultivar.

    PubMed

    Liu, X X; Lang, S R; Su, L Q; Liu, X; Wang, X F

    2015-12-14

    Rape seed (Brassica napus L.) is one of the most important oil seed crops in the world. Genetic manipulation of rapeseed requires a suitable tissue culture system and an efficient method for plant regeneration, as well as an efficient transformation procedure. However, development of transgenic B. napus has been problematic, and current studies are limited to cultivated varieties. In this study, we report a protocol for regeneration of transgenic rape after Agrobacterium-mediated transformation of hypocotyls from the spring B. napus 'Precocity' cultivar. We analyzed the effects of plant growth regulators in the medium on regeneration. Additionally, factors affecting the transformation efficiency, including seedling age, Agrobacterium concentration, infection time, and co-cultivation time, were assessed by monitoring GUS expression. Results from these experiments revealed that transformation was optimized when the meristematic parts of the hypocotyls were taken from 8 day-old seedlings, cultured on Murashinge and Skoog basal media containing 0.1 mg/L 1-naphthaleneacetic acid and 2.5 mg/L 6-benzylaminopurine, and incubated in Agrobacterium suspension (OD600 = 0.5) for 3 to 5 min, followed by 2 days of co-cultivation. Integration of T-DNA into the plant genome was confirmed by polymerase chain reaction (PCR), b-glucuronidase histochemical staining, and quantitative real-time PCR. The protocols developed for regeneration, transformation, and rooting described in this study could help to accelerate the development of transgenic spring rape varieties with novel features.

  18. Development of an Agrobacterium-Mediated Stable Transformation Method for the Sensitive Plant Mimosa pudica

    PubMed Central

    Mano, Hiroaki; Fujii, Tomomi; Sumikawa, Naomi; Hiwatashi, Yuji; Hasebe, Mitsuyasu

    2014-01-01

    The sensitive plant Mimosa pudica has long attracted the interest of researchers due to its spectacular leaf movements in response to touch or other external stimuli. Although various aspects of this seismonastic movement have been elucidated by histological, physiological, biochemical, and behavioral approaches, the lack of reverse genetic tools has hampered the investigation of molecular mechanisms involved in these processes. To overcome this obstacle, we developed an efficient genetic transformation method for M. pudica mediated by Agrobacterium tumefaciens (Agrobacterium). We found that the cotyledonary node explant is suitable for Agrobacterium-mediated transformation because of its high frequency of shoot formation, which was most efficiently induced on medium containing 0.5 µg/ml of a synthetic cytokinin, 6-benzylaminopurine (BAP). Transformation efficiency of cotyledonary node cells was improved from almost 0 to 30.8 positive signals arising from the intron-sGFP reporter gene by using Agrobacterium carrying a super-binary vector pSB111 and stabilizing the pH of the co-cultivation medium with 2-(N-morpholino)ethanesulfonic acid (MES) buffer. Furthermore, treatment of the explants with the detergent Silwet L-77 prior to co-cultivation led to a two-fold increase in the number of transformed shoot buds. Rooting of the regenerated shoots was efficiently induced by cultivation on irrigated vermiculite. The entire procedure for generating transgenic plants achieved a transformation frequency of 18.8%, which is comparable to frequencies obtained for other recalcitrant legumes, such as soybean (Glycine max) and pea (Pisum sativum). The transgene was stably integrated into the host genome and was inherited across generations, without affecting the seismonastic or nyctinastic movements of the plants. This transformation method thus provides an effective genetic tool for studying genes involved in M. pudica movements. PMID:24533121

  19. Agrobacterium-mediated transformation of tomatillo (Physalis ixocarpa) and tissue specific and developmental expression of the CaMV 35S promoter in transgenic tomatillo plants.

    PubMed

    Assad-García, N; Ochoa-Alejo, N; García-Hernández, E; Herrera-Estrella, L; Simpson, J

    1992-10-01

    A protocol for the Agrobacterium-mediated transformation of tomatillo was developed. Up to 40 transgenic plants could be obtained in experiments using 60 cotyledon expiants. The transformed nature of the regenerated plants was confirmed by NPT II and Southern blot hybridization analysis. Using the b-glucuronidase system the tissue specific and developmental patterns of expression of the Cauliflower Mosaic Virus 35S promoter were determined in transgenic tomatillo plants. It was found that this promoter is developmentally regulated during fruit and seed formation. PMID:24213286

  20. High frequency regeneration via direct somatic embryogenesis and efficient Agrobacterium-mediated genetic transformation of tobacco.

    PubMed

    Pathi, Krishna Mohan; Tula, Suresh; Tuteja, Narendra

    2013-06-01

    A direct somatic embryogenesis protocol was developed for four cultivars of Nicotiana species, by using leaf disc as an explant. Direct somatic embryogenesis of Nicotiana by using BAP and IAA has not been investigated so far. This method does not require formation of callus tissues which leads to somaclonal variations. The frequency of somatic embryogenesis was strongly influenced by the plant growth hormones. The somatic embryos developing directly from explant tissue were noticed after 6 d of culture. Somatic embryogenesis of a high frequency (87-96%) was observed in cultures of the all four genotypes (Nicotiana tabacum, N. benthamiyana, N. xanthi, N. t cv petihavana). The results showed that the best medium for direct somatic embryogenesis was MS supplemented with 2.5 mg/l, 0.2 mg/l IAA and 2% sucrose. Subculture of somatic embryos onto hormone free MS medium resulted in their conversion into plants for all genotypes. About 95% of the regenerated somatic embryos germinated into complete plantlets. The plants showed morphological and growth characteristics similar to those of seed-derived plants. Explants were transformed using Agrobacterium tumifacious LBA4404 plasmid pCAMBIA1301 harboring the GUS gene. The regenerated transgenic plants were confirmed by PCR analysis and histochemical GUS assay. The transformation efficiency obtained by using the Agrobacterium- mediated transformation was more than 95%. This method takes 6 wk to accomplish complete transgenic plants through direct somatic embryogenesis. The transgenic plantlets were acclimatized successfully with 98% survival in greenhouse and they showed normal morphological characteristics and were fertile. The regeneration and transformation method described herein is very simple, highly efficient and fast for the introduction of any foreign gene directly in tobacco through direct somatic embryogenesis. PMID:23518589

  1. A rapid, highly efficient and economical method of Agrobacterium-mediated in planta transient transformation in living onion epidermis.

    PubMed

    Xu, Kedong; Huang, Xiaohui; Wu, Manman; Wang, Yan; Chang, Yunxia; Liu, Kun; Zhang, Ju; Zhang, Yi; Zhang, Fuli; Yi, Liming; Li, Tingting; Wang, Ruiyue; Tan, Guangxuan; Li, Chengwei

    2014-01-01

    Transient transformation is simpler, more efficient and economical in analyzing protein subcellular localization than stable transformation. Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living and transparent cells in a monolayer, is suitable to visualize fluorescent fusion proteins. The often used transient transformation methods included particle bombardment, protoplast transfection and Agrobacterium-mediated transformation. Particle bombardment in onion epidermis was successfully established, however, it was expensive, biolistic equipment dependent and with low transformation efficiency. We developed a highly efficient in planta transient transformation method in onion epidermis by using a special agroinfiltration method, which could be fulfilled within 5 days from the pretreatment of onion bulb to the best time-point for analyzing gene expression. The transformation conditions were optimized to achieve 43.87% transformation efficiency in living onion epidermis. The developed method has advantages in cost, time-consuming, equipment dependency and transformation efficiency in contrast with those methods of particle bombardment in onion epidermal cells, protoplast transfection and Agrobacterium-mediated transient transformation in leaf epidermal cells of other plants. It will facilitate the analysis of protein subcellular localization on a large scale.

  2. A comparison of the phenotypic and genetic stability of recombinant Trichoderma spp. generated by protoplast- and Agrobacterium-mediated transformation.

    PubMed

    Cardoza, Rosa Elena; Vizcaino, Juan Antonio; Hermosa, Maria Rosa; Monte, Enrique; Gutiérrez, Santiago

    2006-08-01

    Four different Trichoderma strains, T. harzianum CECT 2413, T. asperellum T53, T. atroviride T11 and T. longibrachiatum T52, which represent three of the four sections contained in this genus, were transformed by two different techniques: a protocol based on the isolation of protoplasts and a protocol based on Agrobacterium-mediated transformation. Both methods were set up using hygromycin B or phleomycin resistance as the selection markers. Using these techniques, we obtained phenotypically stable transformants of these four different strains. The highest transformation efficiencies were obtained with the T. longibrachiatum T52 strain: 65-70 transformants/microg DNA when transformed with the plasmid pAN7-1 (hygromycin B resistance) and 280 transformants/107 spores when the Agrobacterium-mediated transformation was performed with the plasmid pUR5750 (hygromycin B resistance). Overall, the genetic analysis of the transformants showed that some of the strains integrated and maintained the transforming DNA in their genome throughout the entire transformation and selection process. In other cases, the integrated DNA was lost.

  3. A rapid, highly efficient and economical method of Agrobacterium-mediated in planta transient transformation in living onion epidermis.

    PubMed

    Xu, Kedong; Huang, Xiaohui; Wu, Manman; Wang, Yan; Chang, Yunxia; Liu, Kun; Zhang, Ju; Zhang, Yi; Zhang, Fuli; Yi, Liming; Li, Tingting; Wang, Ruiyue; Tan, Guangxuan; Li, Chengwei

    2014-01-01

    Transient transformation is simpler, more efficient and economical in analyzing protein subcellular localization than stable transformation. Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living and transparent cells in a monolayer, is suitable to visualize fluorescent fusion proteins. The often used transient transformation methods included particle bombardment, protoplast transfection and Agrobacterium-mediated transformation. Particle bombardment in onion epidermis was successfully established, however, it was expensive, biolistic equipment dependent and with low transformation efficiency. We developed a highly efficient in planta transient transformation method in onion epidermis by using a special agroinfiltration method, which could be fulfilled within 5 days from the pretreatment of onion bulb to the best time-point for analyzing gene expression. The transformation conditions were optimized to achieve 43.87% transformation efficiency in living onion epidermis. The developed method has advantages in cost, time-consuming, equipment dependency and transformation efficiency in contrast with those methods of particle bombardment in onion epidermal cells, protoplast transfection and Agrobacterium-mediated transient transformation in leaf epidermal cells of other plants. It will facilitate the analysis of protein subcellular localization on a large scale. PMID:24416168

  4. Agrobacterium-mediated transformation of Australian rice varieties and promoter analysis of major pollen allergen gene, Ory s 1.

    PubMed

    Azria, Diah; Bhalla, Prem L

    2011-09-01

    A simple protocol for Agrobacterium-mediated transformation of Australian rice using mature embryos is described. Transgenic plants of two commercial genotypes of Australian rice, Amaroo and Millin, were produced. Transgenic plants were obtained by applying selection pressure to callus and to the regenerated shoots. Exclusion of the selective agent (hygromycin) during plant regeneration was found to be critical for recovery of transgenic plants from these commercial varieties. Transgenic plants were produced after 3 months. The developed system was also used to study spatial and temporal expression of a rice pollen-specific gene, Ory s 1. Expression of pOry s 1::uidA in transgenic rice demonstrated GUS expression in mature pollen, hence indicating potential use of this promoter to direct pollen-specific gene expression. Further Ory s 1 5' deletion study indicated that the pollen-specificity element may reside within -405 bp to the start of the transcription, while the region upstream of -405 contained a cis-acting regulatory element(s) responsible for quantitative expression of this gene.

  5. Mitochondrial Porin Isoform AtVDAC1 Regulates the Competence of Arabidopsis thaliana to Agrobacterium-Mediated Genetic Transformation.

    PubMed

    Kwon, Tackmin

    2016-09-01

    The efficiency of Agrobacterium-mediated transformation in plants depends on the virulence of Agrobacterium strains, the plant tissue culture conditions, and the susceptibility of host plants. Understanding the molecular interactions between Agrobacterium and host plant cells is crucial when manipulating the susceptibility of recalcitrant crop plants and protecting orchard trees from crown gall disease. It was discovered that Arabidopsis voltage-dependent anion channel 1 (atvdac1) mutant has drastic effects on Agrobacterium-mediated tumorigenesis and growth developmental phenotypes, and that these effects are dependent on a Ws-0 genetic background. Genetic complementation of Arabidopsis vdac1 mutants and yeast porin1-deficient strain with members of the AtVDAC gene family revealed that AtVDAC1 is required for Agrobacterium-mediated transformation, and there is weak functional redundancy between AtVDAC1 and AtVDAC3, which is independent of porin activity. Furthermore, atvdac1 mutants were deficient in transient and stable transformation by Agrobacterium, suggesting that AtVDAC1 is involved in the early stages of Agrobacterium infection prior to transferred-DNA (T-DNA) integration. Transgenic plants overexpressing AtVDAC1 not only complemented the phenotypes of the atvdac1 mutant, but also showed high efficiency of transient T-DNA gene expression; however, the efficiency of stable transformation was not affected. Moreover, the effect of phytohormone treatment on competence to Agrobacterium was compromised in atvdac1 mutants. These data indicate that AtVDAC1 regulates the competence of Arabidopsis to Agrobacterium infection. PMID:27643450

  6. Mitochondrial Porin Isoform AtVDAC1 Regulates the Competence of Arabidopsis thaliana to Agrobacterium-Mediated Genetic Transformation

    PubMed Central

    Kwon, Tackmin

    2016-01-01

    The efficiency of Agrobacterium-mediated transformation in plants depends on the virulence of Agrobacterium strains, the plant tissue culture conditions, and the susceptibility of host plants. Understanding the molecular interactions between Agrobacterium and host plant cells is crucial when manipulating the susceptibility of recalcitrant crop plants and protecting orchard trees from crown gall disease. It was discovered that Arabidopsis voltage-dependent anion channel 1 (atvdac1) mutant has drastic effects on Agrobacterium-mediated tumorigenesis and growth developmental phenotypes, and that these effects are dependent on a Ws-0 genetic background. Genetic complementation of Arabidopsis vdac1 mutants and yeast porin1-deficient strain with members of the AtVDAC gene family revealed that AtVDAC1 is required for Agrobacterium-mediated transformation, and there is weak functional redundancy between AtVDAC1 and AtVDAC3, which is independent of porin activity. Furthermore, atvdac1 mutants were deficient in transient and stable transformation by Agrobacterium, suggesting that AtVDAC1 is involved in the early stages of Agrobacterium infection prior to transferred-DNA (T-DNA) integration. Transgenic plants overexpressing AtVDAC1 not only complemented the phenotypes of the atvdac1 mutant, but also showed high efficiency of transient T-DNA gene expression; however, the efficiency of stable transformation was not affected. Moreover, the effect of phytohormone treatment on competence to Agrobacterium was compromised in atvdac1 mutants. These data indicate that AtVDAC1 regulates the competence of Arabidopsis to Agrobacterium infection. PMID:27643450

  7. Generation of Marker- and/or Backbone-Free Transgenic Wheat Plants via Agrobacterium-Mediated Transformation

    PubMed Central

    Wang, Gen-Ping; Yu, Xiu-Dao; Sun, Yong-Wei; Jones, Huw D.; Xia, Lan-Qin

    2016-01-01

    Horizontal transfer of antibiotic resistance genes to animals and vertical transfer of herbicide resistance genes to the weedy relatives are perceived as major biosafety concerns in genetically modified (GM) crops. In this study, five novel vectors which used gusA and bar as a reporter gene and a selection marker gene, respectively, were constructed based on the pCLEAN dual binary vector system. Among these vectors, 1G7B and 5G7B carried two T-DNAs located on two respective plasmids with 5G7B possessing an additional virGwt gene. 5LBTG154 and 5TGTB154 carried two T-DNAs in the target plasmid with either one or double right borders, and 5BTG154 carried the selectable marker gene on the backbone outside of the T-DNA left border in the target plasmid. In addition, 5BTG154, 5LBTG154, and 5TGTB154 used pAL154 as a helper plasmid which contains Komari fragment to facilitate transformation. These five dual binary vector combinations were transformed into Agrobacterium strain AGL1 and used to transform durum wheat cv Stewart 63. Evaluation of the co-transformation efficiencies, the frequencies of marker-free transgenic plants, and integration of backbone sequences in the obtained transgenic lines indicated that two vectors (5G7B and 5TGTB154) were more efficient in generating marker-free transgenic wheat plants with no or minimal integration of backbone sequences in the wheat genome. The vector series developed in this study for generation of marker- and/or backbone-free transgenic wheat plants via Agrobacterium-mediated transformation will be useful to facilitate the creation of “clean” GM wheat containing only the foreign genes of agronomic importance. PMID:27708648

  8. High-efficiency Agrobacterium-mediated transformation of Norway spruce (Picea abies) and loblolly pine (Pinus taeda)

    NASA Technical Reports Server (NTRS)

    Wenck, A. R.; Quinn, M.; Whetten, R. W.; Pullman, G.; Sederoff, R.; Brown, C. S. (Principal Investigator)

    1999-01-01

    Agrobacterium-mediated gene transfer is the method of choice for many plant biotechnology laboratories; however, large-scale use of this organism in conifer transformation has been limited by difficult propagation of explant material, selection efficiencies and low transformation frequency. We have analyzed co-cultivation conditions and different disarmed strains of Agrobacterium to improve transformation. Additional copies of virulence genes were added to three common disarmed strains. These extra virulence genes included either a constitutively active virG or extra copies of virG and virB, both from pTiBo542. In experiments with Norway spruce, we increased transformation efficiencies 1000-fold from initial experiments where little or no transient expression was detected. Over 100 transformed lines expressing the marker gene beta-glucuronidase (GUS) were generated from rapidly dividing embryogenic suspension-cultured cells co-cultivated with Agrobacterium. GUS activity was used to monitor transient expression and to further test lines selected on kanamycin-containing medium. In loblolly pine, transient expression increased 10-fold utilizing modified Agrobacterium strains. Agrobacterium-mediated gene transfer is a useful technique for large-scale generation of transgenic Norway spruce and may prove useful for other conifer species.

  9. An efficient method for Agrobacterium-mediated genetic transformation and plant regeneration in cumin (Cuminum cyminum L.).

    PubMed

    Pandey, Sonika; Mishra, Avinash; Patel, Manish Kumar; Jha, Bhavanath

    2013-09-01

    Cumin is an annual herbaceous medicinally important plant having diverse applications. An efficient and reproducible method of Agrobacterium-mediated genetic transformation was herein established for the first time. A direct regeneration method without callus induction was optimised using embryos as explant material in Gamborg's B5 medium supplemented with 0.5-μM 6-benzyladenine and 2.0-μM α-naphthalene acetic acid. About 1,020 embryos (a mean of 255 embryos per batch) were used for the optimisation of transformation conditions. These conditions were an Agrobacterium cell suspension of 0.6 OD600, a co-cultivation time of 72 h, 300-μM acetosyringone and wounding of explants using a razor blade. Pre-cultured elongated embryos were treated using optimised conditions. About 720 embryos (a mean of 180 embryos per batch) were used for transformation and 95 % embryos showed transient β-glucuronidase expression after co-cultivation. Putative transformed embryos were cultured on B5 medium for shoot proliferation and 21 regenerated plants were obtained after selection and allowed to root. T0 plantlets showed β-glucuronidase expression and gene integration was confirmed via PCR amplification of 0.96 and 1.28 kb fragments of the hygromycin-phosphotransferase II and β-glucuronidase genes, respectively. In this study, a transformation efficiency of 1.5 % was demonstrated and a total of 11 transgenic plants were obtained at the hardening stage, however, only four plants acclimatised during hardening. Gene copy number was analysed by Southern blot analysis of hardened plants and single-copy gene integration was observed. This is the first successful attempt of Agrobacterium-mediated genetic transformation of cumin.

  10. An efficient method for Agrobacterium-mediated genetic transformation and plant regeneration in cumin (Cuminum cyminum L.).

    PubMed

    Pandey, Sonika; Mishra, Avinash; Patel, Manish Kumar; Jha, Bhavanath

    2013-09-01

    Cumin is an annual herbaceous medicinally important plant having diverse applications. An efficient and reproducible method of Agrobacterium-mediated genetic transformation was herein established for the first time. A direct regeneration method without callus induction was optimised using embryos as explant material in Gamborg's B5 medium supplemented with 0.5-μM 6-benzyladenine and 2.0-μM α-naphthalene acetic acid. About 1,020 embryos (a mean of 255 embryos per batch) were used for the optimisation of transformation conditions. These conditions were an Agrobacterium cell suspension of 0.6 OD600, a co-cultivation time of 72 h, 300-μM acetosyringone and wounding of explants using a razor blade. Pre-cultured elongated embryos were treated using optimised conditions. About 720 embryos (a mean of 180 embryos per batch) were used for transformation and 95 % embryos showed transient β-glucuronidase expression after co-cultivation. Putative transformed embryos were cultured on B5 medium for shoot proliferation and 21 regenerated plants were obtained after selection and allowed to root. T0 plantlets showed β-glucuronidase expression and gene integration was confirmed via PCR amplification of 0.96 and 1.28 kb fragments of the hygromycin-phosphotransferase II and β-glucuronidase genes, respectively. In this study, a transformation efficiency of 1.5 % was demonstrated and a total of 11 transgenic plants were obtained at the hardening stage, however, only four plants acclimatised during hardening. Gene copy number was analysed by Southern blot analysis of hardened plants and single-copy gene integration was observed. This is the first successful attempt of Agrobacterium-mediated genetic transformation of cumin. PMID:23813408

  11. Efficient Agrobacterium-mediated transformation of commercial hybrid poplar Populus nigra L. x P. maximowiczii A. Henry.

    PubMed

    Yevtushenko, Dmytro P; Misra, Santosh

    2010-03-01

    Many economically important species of Populus, especially those in sections Aigeiros and Tacamahaca, remain recalcitrant to genetic transformation. In this study, a simple and reliable protocol was developed for the efficient Agrobacterium-mediated transformation of a difficult-to-transform, but commercially viable, hybrid poplar Populus nigra L. x P. maximowiczii A. Henry (NM6). A plant transformation vector designed to express the beta-glucuronidase (GUS) gene was used to detect transformation events at early stages of plant regeneration and to optimize parameters affecting poplar transformation. The use of zeatin riboside in shoot-induction medium, regeneration of shoots via indirect organogenesis, and early selection pressure were the major modifications that drastically improved the efficiency of poplar transformation and minimized the number of untransformed regenerants. Transgenic shoots were routinely obtained 4-10 weeks after co-culture with A. tumefaciens, with a greater than 90% rate of plant recovery. Stable transgene integration, ranging from a single insertion to ten copies per genome, was confirmed by Southern blot analysis. The mean transformation frequency was 36.3% and about two-thirds of the lines had 1-2 transgene copies. Among the explants, petioles and leaves had a higher transformation frequency than did stem segments. Growth characteristics and the morphology of transgenic poplar plants were identical to untransformed controls. These findings will accelerate the development of P. nigra x P. maximowiczii plants with novel traits, and may also be useful to improve transformation procedures for other Populus species.

  12. Agrobacterium-mediated transformation in chickpea (Cicer arietinum L.) with an insecticidal protein gene: optimisation of different factors.

    PubMed

    Indurker, Shivani; Misra, Hari S; Eapen, Susan

    2010-07-01

    Agrobacterium-mediated transformation in chickpea was developed using strain LBA4404 carrying nptII, uidA and cryIAc genes and transformants selected on Murashige and Skoog's basal medium supplemented with benzyladenine, kinetin and kanamycin. Integration of transgenes was demonstrated using polymerase chain reaction and Southern blot hybridization of T0 plants. The expression of CryIAc delta endotoxin and GUS enzyme was shown by enzyme linked immunosorbent assay and histochemical assay respectively. The transgenic plants (T0) showed more tolerance to infection by Helicoverpa armigera compared to control plants. Various factors such as explant source, cultivar type, different preculture treatment period of explants, co-cultivation period, acetosyringone supplementation, Agrobacterium harboring different plasmids, vacuum infiltration and sonication treatment were tested to study the influence on transformation frequency. The results indicated that use of epicotyl as explant, cultivar ICCC37, Agrobacterium harboring plasmid pHS102 as vector, preculture of explant for 48 h, co-cultivation period of 2 days at 25°C and vacuum infiltration for 15 min produced the best transformation results. Sonication treatment of explants with Agrobacteria for 80 s was found to increase the frequency of transformation.

  13. Design, Construction, and Validation of Artificial MicroRNA Vectors Using Agrobacterium-Mediated Transient Expression System.

    PubMed

    Bhagwat, Basdeo; Chi, Ming; Han, Dianwei; Tang, Haifeng; Tang, Guiliang; Xiang, Yu

    2016-01-01

    Artificial microRNA (amiRNA) technology utilizes microRNA (miRNA) biogenesis pathway to produce artificially selected small RNAs using miRNA gene backbone. It provides a feasible strategy for inducing loss of gene function, and has been applied in functional genomics study, improvement of crop quality and plant virus disease resistance. A big challenge in amiRNA applications is the unpredictability of silencing efficacy of the designed amiRNAs and not all constructed amiRNA candidates would be expressed effectively in plant cells. We and others found that high efficiency and specificity in RNA silencing can be achieved by designing amiRNAs with perfect or almost perfect sequence complementarity to their targets. In addition, we recently demonstrated that Agrobacterium-mediated transient expression system can be used to validate amiRNA constructs, which provides a simple, rapid and effective method to select highly expressible amiRNA candidates for stable genetic transformation. Here, we describe the methods for design of amiRNA candidates with perfect or almost perfect base-pairing to the target gene or gene groups, incorporation of amiRNA candidates in miR168a gene backbone by one step inverse PCR amplification, construction of plant amiRNA expression vectors, and assay of transient expression of amiRNAs in Nicotiana benthamiana through agro-infiltration, small RNA extraction, and amiRNA Northern blot.

  14. Agrobacterium-mediated genetic transformation of Coffea arabica (L.) is greatly enhanced by using established embryogenic callus cultures

    PubMed Central

    2011-01-01

    Background Following genome sequencing of crop plants, one of the main challenges today is determining the function of all the predicted genes. When gene validation approaches are used for woody species, the main obstacle is the low recovery rate of transgenic plants from elite or commercial cultivars. Embryogenic calli have frequently been the target tissue for transformation, but the difficulty in producing or maintaining embryogenic tissues is one of the main problems encountered in genetic transformation of many woody plants, including Coffea arabica. Results We identified the conditions required for successful long-term proliferation of embryogenic cultures in C. arabica and designed a highly efficient and reliable Agrobacterium tumefaciens-mediated transformation method based on these conditions. The transformation protocol with LBA1119 harboring pBin 35S GFP was established by evaluating the effect of different parameters on transformation efficiency by GFP detection. Using embryogenic callus cultures, co-cultivation with LBA1119 OD600 = 0.6 for five days at 20 °C enabled reproducible transformation. The maintenance conditions for the embryogenic callus cultures, particularly a high auxin to cytokinin ratio, the age of the culture (optimum for 7-10 months of proliferation) and the use of a yellow callus phenotype, were the most important factors for achieving highly efficient transformation (> 90%). At the histological level, successful transformation was related to the number of proembryogenic masses present. All the selected plants were proved to be transformed by PCR and Southern blot hybridization. Conclusion Most progress in increasing transformation efficiency in coffee has been achieved by optimizing the production conditions of embryogenic cultures used as target tissues for transformation. This is the first time that a strong positive effect of the age of the culture on transformation efficiency was demonstrated. Our results make Agrobacterium-mediated

  15. Agrobacterium-mediated genetic transformation using cotyledons in Japanese pear (Pyrus pyrifolia)

    PubMed Central

    Nakajima, Ikuko; Sato, Yoshihiko; Saito, Toshihiro; Moriguchi, Takaya; Yamamoto, Toshiya

    2013-01-01

    Genetic transformation was successfully established producing both transformed adventitious shoots and calli in Japanese pear (Pyrus pyrifolia Nakai) by using cotyledons as explants. Cotyledons of five cultivars were co-cultivated with Agrobacterium tumefaciens strain LBA4404 carrying the pBIN19-sgfp, which contained a green fluorescent protein gene and the neomycin phosphotransferase gene. In order to increase transformation efficiency, sonication and ethylenedioxybis (ethylamine)-N,N,N′,N′-tetraacetic acid (EGTA) treatments were applied, which could produce physical wounds across the tissue and prevent plant defense reaction, respectively. Green fluorescent protein (GFP) fluorescence was evaluated two weeks and five months after Agrobacterium inoculation as measures of transient and stable transformations, respectively. As a result, sonication significantly increased both transient and stable expression of GFP fluorescence, whereas EGTA treatment did not show a positive effect on either. Out of 18 regenerated plantlets obtained, one plant regenerated from ‘Agenosho Shinanashi’ showed stable GFP fluorescence. This plant was confirmed as a transformant by PCR and genomic Southern blotting. Three other transformed regenerated shoots by myb gene showed red color, which were derived from ‘Imamuraaki’ by the same transformation method. Transformation system in this study was shown to be reproducible since plural transformants were obtained. PMID:24273422

  16. An efficient Agrobacterium-mediated transformation method for the edible mushroom Hypsizygus marmoreus.

    PubMed

    Zhang, Jin jing; Shi, Liang; Chen, Hui; Sun, Yun qi; Zhao, Ming wen; Ren, Ang; Chen, Ming jie; Wang, Hong; Feng, Zhi yong

    2014-01-01

    Hypsizygus marmoreus is one of the major edible mushrooms in East Asia. As no efficient transformation method, the molecular and genetics studies were hindered. The glyceraldehyde-3-phosphate dehydrogenase (GPD) gene of H. marmoreus was isolated and its promoter was used to drive the hygromycin B phosphotransferase (HPH) and enhanced green fluorescent protein (EGFP) in H. marmoreus. Agrobacterium tumefaciens-mediated transformation (ATMT) was successfully applied in H. marmoreus. The transformation parameters were optimized, and it was found that co-cultivation of bacteria with protoplast at a ratio of 1000:1 at a temperature of 26 °C in medium containing 0.3 mM acetosyringone resulted in the highest transformation efficiency for Agrobacterium strain. Besides, three plasmids, each carrying a different promoter (from H. marmoreus, Ganoderma lucidum and Lentinula edodes) driving the expression of an antibiotic resistance marker, were also tested. The construct carrying the H. marmoreus gpd promoter produced more transformants than other constructs. Our analysis showed that over 85% of the transformants tested remained mitotically stable even after five successive rounds of subculturing. Putative transformants were analyzed for the presence of hph gene by PCR and Southern blot. Meanwhile, the expression of EGFP in H. marmoreus transformants was detected by fluorescence imaging. This ATMT system increases the transformation efficiency of H. marmoreus and may represent a useful tool for molecular genetic studies in this mushroom species.

  17. Female Reproductive Tissues Are the Primary Target of Agrobacterium-Mediated Transformation by the Arabidopsis Floral-Dip Method1

    PubMed Central

    Desfeux, Christine; Clough, Steven J.; Bent, Andrew F.

    2000-01-01

    The floral-dip method for Agrobacterium-mediated transformation of Arabidopsis allows efficient plant transformation without need for tissue culture. To facilitate use with other plant species, we investigated the mechanisms that underlie this method. In manual outcrossing experiments, application of Agrobacterium tumefaciens to pollen donor plants did not produce any transformed progeny, whereas application of Agrobacterium to pollen recipient plants yielded transformants at a rate of 0.48%. Agrobacterium strains with T-DNA carrying gusA (encoding β-glucuronidase [GUS]) under the control of 35S, LAT52, or ACT11 promoters revealed delivery of GUS activity to developing ovules, whereas no GUS staining of pollen or pollen tubes was observed. Transformants derived from the same seed pod contained independent T-DNA integration events. In Arabidopsis flowers, the gynoecium develops as an open, vase-like structure that fuses to form closed locules roughly 3 d prior to anthesis. In correlation with this fact, we found that the timing of Agrobacterium infection was critical. Transformants were obtained and GUS staining of ovules and embryo sacs was observed only if the Agrobacterium were applied 5 d or more prior to anthesis. A 6-fold higher rate of transformation was obtained with a CRABS-CLAW mutant that maintains an open gynoecium. Our results suggest that ovules are the site of productive transformation in the floral-dip method, and further suggest that Agrobacterium must be delivered to the interior of the developing gynoecium prior to locule closure if efficient transformation is to be achieved. PMID:10889238

  18. Genetic Transformation of Metroxylon sagu (Rottb.) Cultures via Agrobacterium-Mediated and Particle Bombardment

    PubMed Central

    Ibrahim, Evra Raunie

    2014-01-01

    Sago palm (Metroxylon sagu) is a perennial plant native to Southeast Asia and exploited mainly for the starch content in its trunk. Genetic improvement of sago palm is extremely slow when compared to other annual starch crops. Urgent attention is needed to improve the sago palm planting material and can be achieved through nonconventional methods. We have previously developed a tissue culture method for sago palm, which is used to provide the planting materials and to develop a genetic transformation procedure. Here, we report the genetic transformation of sago embryonic callus derived from suspension culture using Agrobacterium tumefaciens and gene gun systems. The transformed embryoids cells were selected against Basta (concentration 10 to 30 mg/L). Evidence of foreign genes integration and function of the bar and gus genes were verified via gene specific PCR amplification, gus staining, and dot blot analysis. This study showed that the embryogenic callus was the most suitable material for transformation as compared to the fine callus, embryoid stage, and initiated shoots. The gene gun transformation showed higher transformation efficiency than the ones transformed using Agrobacterium when targets were bombarded once or twice using 280 psi of helium pressure at 6 to 8 cm distance. PMID:25295258

  19. Agrobacterium-mediated genetic transformation and plant regeneration of the hardwood tree species Fraxinus profunda.

    PubMed

    Stevens, Micah E; Pijut, Paula M

    2014-06-01

    This transformation and regeneration protocol provides an integral framework for the genetic improvement of Fraxinus profunda (pumpkin ash) for future development of plants resistant to the emerald ash borer. Using mature hypocotyls as the initial explants, an Agrobacterium tumefaciens-mediated genetic transformation system was successfully developed for pumpkin ash (Fraxinus profunda). This transformation protocol is an invaluable tool to combat the highly aggressive, non-native emerald ash borer (EAB), which has the potential to eliminate native Fraxinus spp. from the natural landscape. Hypocotyls were successfully transformed with Agrobacterium strain EHA105 harboring the pq35GR vector, containing an enhanced green fluorescent protein (EGFP) as well as a fusion gene between neomycin phosphotransferase (nptII) and gusA. Hypocotyls were cultured for 7 days on Murashige and Skoog (MS) medium with 22.2 μM 6-benzyladenine (BA), 4.5 μM thidiazuron (TDZ), 50 mg L(-1) adenine hemisulfate (AS), and 10 % coconut water (CW) prior to transformation. Hypocotyls were transformed using 90 s sonication plus 10 min vacuum infiltration after Agrobacterium was exposed to 100 μM acetosyringone for 1 h. Adventitious shoots were regenerated on MS medium with 22.2 μM BA, 4.5 μM TDZ, 50 mg L(-1) AS, 10 % CW, 400 mg L(-1) timentin, and 20 mg L(-1) kanamycin. Timentin at 400 and 20 mg L(-1) kanamycin were most effective at controlling Agrobacterium growth and selecting for transformed cells, respectively. The presence of nptII, GUS (β-glucuronidase), and EGFP in transformed plants was confirmed using polymerase chain reaction (PCR), while the expression of EGFP was also confirmed through fluorescent microscopy and reverse transcription-PCR. This transformation protocol provides an integral foundation for future genetic modifications of F. profunda to provide resistance to EAB. PMID:24493252

  20. Functional validation of Capsicum frutescens aminotransferase gene involved in vanillylamine biosynthesis using Agrobacterium mediated genetic transformation studies in Nicotiana tabacum and Capsicum frutescens calli cultures.

    PubMed

    Gururaj, Harishchandra B; Padma, Mallaya N; Giridhar, Parvatam; Ravishankar, Gokare A

    2012-10-01

    Capsaicinoid biosynthesis involves the participation of two substrates viz. vanillylamine and C(9)-C(11) fatty acid moieties. Vanillylamine which is a derivative of vanillin is synthesized through a transaminase reaction in the phenylpropanoid pathway of capsaicinoid synthesis. Here we report the functional validation of earlier reported putative aminotransferase gene for vanillylamine biosynthesis in heterologous system using Agrobacterium mediated genetic transformation studies in Nicotiana tabacum and Capsicum frutescens calli cultures. Molecular analysis tools comprising PCR and Southern blot analysis have shown the integration of the foreign gene in N. tabacum and C. frutescens calli cultures. The study shows the production of vanillylamine in transformed N. tabacum callus cultures and also the reduction of vanillylamine production when whole gene based antisense binary vector construct was used in transformation of C. frutescens callus cultures. Vanillylamine production, aminotransferase assay with Western blot analysis for crude proteins of transformants established the production of putative aminotransferase (pAMT) protein in alternate plant. The result is a clear evidence of involvement of the reported putative aminotransferase responsible for vanillylamine biosynthesis in capsaicinoid biosynthesis pathway, confirming the gene function through functional validation.

  1. In vitro regeneration and Agrobacterium mediated genetic transformation of Artemisia aucheri Boiss.

    PubMed

    Sharafi, Ali; Sohi, Haleh Hashemi; Mirzaee, Hooman; Azadi, Pejman

    2014-10-01

    In the present study, we developed an efficient protocol for in vitro plant regeneration and genetically transformed root induction in medicinal plant Artemisia aucheri Boiss. Leaf explants were cultivated in MS medium supplemented by combination of plant growth regulators including α-naphthalene-acetic acid, 6-benzyl-aminopurine, indole-3-acetic acid and 2, 4-dichlorophenoxyaceticacid. The highest frequency of shoot organogenesis occurred on MS medium supplemented with 0.05 mg/l NAA plus 2 mg/l BA (96.3 %) and MS medium supplemented with 0.5 mg/l IAA plus 2 mg/l BA (88.3 %). Root induction was obtained on MS medium supplemented with 0.5 mg/l IBA. This is a simple, reliable, rapid and high efficient regeneration system for A. aucheri Boiss in short period via adventitious shoot induction approach. Also, an efficient genetically transformed root induction for A. aucheri was developed through Agrobacterium rhizogenes-mediated transformation by four bacterial strains, A4, ATCC15834, MSU440, and A13 (MAFF-02-10266). The maximum frequency of hairy root induction was obtained using MSU440 (93 %) and ATCC15834 (89 %) bacterial strains. Hairy root lines were confirmed by PCR using the rolB gene specific primers and Southern blot analysis.

  2. A mutation in negative regulator of basal resistance WRKY17 of Arabidopsis increases susceptibility to Agrobacterium-mediated genetic transformation.

    PubMed

    Lacroix, Benoît; Citovsky, Vitaly

    2013-01-01

    Agrobacterium is a phytopathogenic bacterium that induces crown gall disease in many plant species by transferring and integrating a segment of its own DNA (T-DNA) into its host genome. Whereas Agrobacterium usually does not trigger an extensive defense response in its host plants, it induces the expression of several defense-related genes and activates plant stress reactions. In the complex interplay between Agrobacterium and its host plant, Agrobacterium has evolved to take advantage of these plant defense pathways for its own purpose of advancement of the infection process. For example, Agrobacterium utilizes the host stress response transcriptional regulator VIP1 to facilitate nuclear import and proteasomal uncoating of its T-DNA during genetic transformation of the host cell. In Arabidopsis, the VIP1 gene expression is repressed by WRKY17, a negative regulator of basal resistance to Pseudomonas. Thus, we examined whether WRKY17 is also involved in plant susceptibility to genetic transformation by Agrobacterium. Using reverse genetics, we showed that a wrky17 mutant displays higher expression of the VIP1 gene in roots, but not in shoots. In a root infection assay, the wrky17 mutant plants were hyper-susceptible to Agrobacterium compared to wild type plants. WRKY17, therefore, may act as a positive regulator of Arabidopsis resistance to Agrobacterium. This notion is important for understanding the complex regulation of Agrobacterium-mediated genetic transformation; thus, although this paper reports a relatively small set of data that we do not plan to pursue further in our lab, we believe it might be useful for the broad community of plant pathologists and plant biotechnologists. PMID:24358874

  3. A mutation in negative regulator of basal resistance WRKY17 of Arabidopsis increases susceptibility to Agrobacterium-mediated genetic transformation

    PubMed Central

    Lacroix, Benoît; Citovsky, Vitaly

    2013-01-01

    Agrobacterium is a phytopathogenic bacterium that induces crown gall disease in many plant species by transferring and integrating a segment of its own DNA (T-DNA) into its host genome. Whereas Agrobacterium usually does not trigger an extensive defense response in its host plants, it induces the expression of several defense-related genes and activates plant stress reactions. In the complex interplay between Agrobacterium and its host plant, Agrobacterium has evolved to take advantage of these plant defense pathways for its own purpose of advancement of the infection process. For example, Agrobacterium utilizes the host stress response transcriptional regulator VIP1 to facilitate nuclear import and proteasomal uncoating of its T-DNA during genetic transformation of the host cell. In Arabidopsis, the VIP1 gene expression is repressed by WRKY17, a negative regulator of basal resistance to Pseudomonas. Thus, we examined whether WRKY17 is also involved in plant susceptibility to genetic transformation by Agrobacterium. Using reverse genetics, we showed that a wrky17 mutant displays higher expression of the VIP1 gene in roots, but not in shoots. In a root infection assay, the wrky17 mutant plants were hyper-susceptible to Agrobacterium compared to wild type plants. WRKY17, therefore, may act as a positive regulator of Arabidopsis resistance to Agrobacterium. This notion is important for understanding the complex regulation of Agrobacterium-mediated genetic transformation; thus, although this paper reports a relatively small set of data that we do not plan to pursue further in our lab, we believe it might be useful for the broad community of plant pathologists and plant biotechnologists. PMID:24358874

  4. Development of Agrobacterium-mediated transformation of highly valued hill banana cultivar Virupakshi (AAB) for resistance to BBTV disease.

    PubMed

    Elayabalan, Sivalingam; Kalaiponmani, Kalaimughilan; Subramaniam, Sreeramanan; Selvarajan, Ramasamy; Panchanathan, Radha; Muthuvelayoutham, Ramlatha; Kumar, Krish K; Balasubramanian, Ponnuswami

    2013-04-01

    One of the most severe viral diseases of hill banana is caused by banana bunchy top virus (BBTV), a nanovirus transmitted by the aphid Pentalonia nigronervosa. In this study, we reported the Agrobacterium-mediated transformation on a highly valued hill banana cultivar Virupakshi (AAB) for resistance to BBTV disease. The target of the RNA interference (RNAi) is the rep gene, encoded by the BBTV-DNA1. In order to develop RNAi construct targeting the BBTV rep gene, the full-length rep gene of 870 bp was polymerase chain reaction amplified from BBTV infected hill banana sample DNA, cloned and confirmed by DNA sequencing. The partial rep gene fragment was cloned in sense and anti sense orientation in the RNAi intermediate vector, pSTARLING-A. After cloning in pSTARLING-A, the cloned RNAi gene cassette was released by NotI enzyme digestion and cloned into the NotI site of binary vector, pART27. Two different explants, embryogenic cells and embryogenic cell suspension derived microcalli were used for co-cultivation. Selection was done in presence of 100 mg/L kanamycin. In total, 143 putative transgenic hill banana lines were generated and established in green house condition. The presence of the transgenes was confirmed in the selected putative transgenic hill banana lines by PCR and reverse transcription PCR analyses. Transgenic hill banana plants expressing RNAi-BBTV rep were obtained and shown to resist infection by BBTV. The transformed plants are symptomless, and the replication of challenge BBTV almost completely suppressed. Hence, the RNAi mediating resistances were shown to be effective management of BBTV in hill banana.

  5. Highly Efficient Agrobacterium-Mediated Transformation of Wheat Via In Planta Inoculation

    NASA Astrophysics Data System (ADS)

    Risacher, Thierry; Craze, Melanie; Bowden, Sarah; Paul, Wyatt; Barsby, Tina

    This chapter details a reproducible method for the transformation of spring wheat using Agrobacterium tumefaciens via the direct inoculation of bacteria into immature seeds in planta as described in patent WO 00/63398(1. Transformation efficiencies from 1 to 30% have been obtained and average efficiencies of at least 5% are routinely achieved. Regenerated plants are phenotypically normal with 30-50% of transformation events carrying introduced genes at single insertion sites, a higher rate than is typically reported for transgenic plants produced using biolistic transformation methods.

  6. Agrobacterium-Mediated Stable Genetic Transformation of Populus angustifolia and Populus balsamifera.

    PubMed

    Maheshwari, Priti; Kovalchuk, Igor

    2016-01-01

    The present study demonstrates Agrobacterium tumefaciens-mediated stable genetic transformation of two species of poplar - Populus angustifolia and Populus balsamifera. The binary vector pCAMBIA-Npro-long-Luc containing the luciferase reporter gene was used to transform stem internode and axillary bud explants. Putative transformants were regenerated on selection-free medium using our previously established in vitro regeneration method. Explant type, genotype, effect of pre-culture, Agrobacterium concentration, a time period of infection and varying periods of co-culture with bacteria were tested for the transformation frequency. The highest frequency of transformation was obtained with stem internode explants pre-cultured for 2 days, infected with Agrobacterium culture at the concentration of OD600 = 0.5 for 10 min and co-cultivated with Agrobacterium for 48 h. Out of the two genotypes tested, P. balsamifera exhibited a higher transformation rate in comparison to P. angustifolia. The primary transformants that exhibited luciferase activity in a bioluminescence assay under the CCD camera when subjected to polymerase chain reaction and Southern blot analysis revealed a stable single-copy integration of luc in their genomes. The reported protocol is highly reproducible and can be applied to other species of poplar; it will also be useful for future genetic engineering of one of the most important families of woody plants for sustainable development. PMID:27014319

  7. Agrobacterium-Mediated Stable Genetic Transformation of Populus angustifolia and Populus balsamifera

    PubMed Central

    Maheshwari, Priti; Kovalchuk, Igor

    2016-01-01

    The present study demonstrates Agrobacterium tumefaciens-mediated stable genetic transformation of two species of poplar – Populus angustifolia and Populus balsamifera. The binary vector pCAMBIA-Npro-long-Luc containing the luciferase reporter gene was used to transform stem internode and axillary bud explants. Putative transformants were regenerated on selection-free medium using our previously established in vitro regeneration method. Explant type, genotype, effect of pre-culture, Agrobacterium concentration, a time period of infection and varying periods of co-culture with bacteria were tested for the transformation frequency. The highest frequency of transformation was obtained with stem internode explants pre-cultured for 2 days, infected with Agrobacterium culture at the concentration of OD600 = 0.5 for 10 min and co-cultivated with Agrobacterium for 48 h. Out of the two genotypes tested, P. balsamifera exhibited a higher transformation rate in comparison to P. angustifolia. The primary transformants that exhibited luciferase activity in a bioluminescence assay under the CCD camera when subjected to polymerase chain reaction and Southern blot analysis revealed a stable single-copy integration of luc in their genomes. The reported protocol is highly reproducible and can be applied to other species of poplar; it will also be useful for future genetic engineering of one of the most important families of woody plants for sustainable development. PMID:27014319

  8. Agrobacterium-Mediated Stable Genetic Transformation of Populus angustifolia and Populus balsamifera.

    PubMed

    Maheshwari, Priti; Kovalchuk, Igor

    2016-01-01

    The present study demonstrates Agrobacterium tumefaciens-mediated stable genetic transformation of two species of poplar - Populus angustifolia and Populus balsamifera. The binary vector pCAMBIA-Npro-long-Luc containing the luciferase reporter gene was used to transform stem internode and axillary bud explants. Putative transformants were regenerated on selection-free medium using our previously established in vitro regeneration method. Explant type, genotype, effect of pre-culture, Agrobacterium concentration, a time period of infection and varying periods of co-culture with bacteria were tested for the transformation frequency. The highest frequency of transformation was obtained with stem internode explants pre-cultured for 2 days, infected with Agrobacterium culture at the concentration of OD600 = 0.5 for 10 min and co-cultivated with Agrobacterium for 48 h. Out of the two genotypes tested, P. balsamifera exhibited a higher transformation rate in comparison to P. angustifolia. The primary transformants that exhibited luciferase activity in a bioluminescence assay under the CCD camera when subjected to polymerase chain reaction and Southern blot analysis revealed a stable single-copy integration of luc in their genomes. The reported protocol is highly reproducible and can be applied to other species of poplar; it will also be useful for future genetic engineering of one of the most important families of woody plants for sustainable development.

  9. Agrobacterium-produced and exogenous cytokinin-modulated Agrobacterium-mediated plant transformation.

    PubMed

    Hwang, Hau-Hsuan; Wang, Ming-Hsuan; Lee, Ying-Ling; Tsai, Yun-Long; Li, Yi-Ho; Yang, Fong-Jhih; Liao, Yu-Chen; Lin, Shao-Kai; Lai, Erh-Min

    2010-09-01

    Agrobacterium tumefaciens is a plant pathogenic bacterium that causes neoplastic growths, called 'crown gall', via the transfer and integration of transferred DNA (T-DNA) from the bacterium into the plant genome. We characterized an acetosyringone (AS)-induced tumour-inducing (Ti) plasmid gene, tzs (trans-zeatin synthesizing), that is responsible for the synthesis of the plant hormone cytokinin in nopaline-type A. tumefaciens strains. The loss of Tzs protein expression and trans-zeatin secretions by the tzs frameshift (tzs-fs) mutant is associated with reduced tumorigenesis efficiency on white radish stems and reduced transformation efficiencies on Arabidopsis roots. Complementation of the tzs-fs mutant with a wild-type tzs gene restored wild-type levels of trans-zeatin secretions and transformation efficiencies. Exogenous application of cytokinin during infection increased the transient transformation efficiency of Arabidopsis roots infected by strains lacking Tzs, which suggests that the lower transformation efficiency resulted from the lack of Agrobacterium-produced cytokinin. Interestingly, although the tzs-fs mutant displayed reduced tumorigenesis efficiency on several tested plants, the loss of Tzs enhanced tumorigenesis efficiencies on green pepper and cowpea. These data strongly suggest that Tzs, by synthesizing trans-zeatin at early stage(s) of the infection process, modulates plant transformation efficiency by A. tumefaciens. PMID:20696005

  10. Agrobacterium-mediated transformation of promising oil-bearing marine algae Parachlorella kessleri.

    PubMed

    Rathod, Jayant Pralhad; Prakash, Gunjan; Pandit, Reena; Lali, Arvind M

    2013-11-01

    Parachlorella kessleri is a unicellular alga which grows in fresh as well as marine water and is commercially important as biomass/lipid feedstock and in bioremediation. The present study describes the successful transformation of marine P. kessleri with the help of Agrobacterium tumefaciens. Transformed marine P. kessleri was able to tolerate more than 10 mg l(-1) hygromycin concentration. Co-cultivation conditions were modulated to allow the simultaneous growth of both marine P. kessleri and A. tumefaciens. For co-cultivation, P. kessleri was shifted from Walne's to tris acetate phosphate medium to reduce the antibiotic requirement during selection. In the present study, the transfer of T-DNA was successful without using acetosyringone. Biochemical and genetic analyses were performed for expression of transgenes by GUS assay and PCR in transformants. Establishment of this protocol would be useful in further genetic modification of oil-bearing Parachlorella species.

  11. Yeast Actin-Related Protein ARP6 Negatively Regulates Agrobacterium-Mediated Transformation of Yeast Cell

    PubMed Central

    Luo, Yumei; Chen, Zikai; Zhu, Detu; Tu, Haitao; Pan, Shen Quan

    2015-01-01

    The yeasts, including Saccharomyces cerevisiae and Pichia pastoris, are single-cell eukaryotic organisms that can serve as models for human genetic diseases and hosts for large scale production of recombinant proteins in current biopharmaceutical industry. Thus, efficient genetic engineering tools for yeasts are of great research and economic values. Agrobacterium tumefaciens-mediated transformation (AMT) can transfer T-DNA into yeast cells as a method for genetic engineering. However, how the T-DNA is transferred into the yeast cells is not well established yet. Here our genetic screening of yeast knockout mutants identified a yeast actin-related protein ARP6 as a negative regulator of AMT. ARP6 is a critical member of the SWR1 chromatin remodeling complex (SWR-C); knocking out some other components of the complex also increased the transformation efficiency, suggesting that ARP6 might regulate AMT via SWR-C. Moreover, knockout of ARP6 led to disruption of microtubule integrity, higher uptake and degradation of virulence proteins, and increased DNA stability inside the cells, all of which resulted in enhanced transformation efficiency. Our findings have identified molecular and cellular mechanisms regulating AMT and a potential target for enhancing the transformation efficiency in yeast cells. PMID:26425545

  12. Yeast Actin-Related Protein ARP6 Negatively Regulates Agrobacterium-Mediated Transformation of Yeast Cell.

    PubMed

    Luo, Yumei; Chen, Zikai; Zhu, Detu; Tu, Haitao; Pan, Shen Quan

    2015-01-01

    The yeasts, including Saccharomyces cerevisiae and Pichia pastoris, are single-cell eukaryotic organisms that can serve as models for human genetic diseases and hosts for large scale production of recombinant proteins in current biopharmaceutical industry. Thus, efficient genetic engineering tools for yeasts are of great research and economic values. Agrobacterium tumefaciens-mediated transformation (AMT) can transfer T-DNA into yeast cells as a method for genetic engineering. However, how the T-DNA is transferred into the yeast cells is not well established yet. Here our genetic screening of yeast knockout mutants identified a yeast actin-related protein ARP6 as a negative regulator of AMT. ARP6 is a critical member of the SWR1 chromatin remodeling complex (SWR-C); knocking out some other components of the complex also increased the transformation efficiency, suggesting that ARP6 might regulate AMT via SWR-C. Moreover, knockout of ARP6 led to disruption of microtubule integrity, higher uptake and degradation of virulence proteins, and increased DNA stability inside the cells, all of which resulted in enhanced transformation efficiency. Our findings have identified molecular and cellular mechanisms regulating AMT and a potential target for enhancing the transformation efficiency in yeast cells. PMID:26425545

  13. DNA METHYLATION ANALYSIS DURING THE OPTIMIZATION OF Agrobacterium-MEDIATED TRANSFORMATION OF SOYBEAN.

    PubMed

    Jiang, J; Wing, V; Xiet, T; Shi, X; Wang, Y P; Sokolov, V

    2016-01-01

    Soybean is recognized as one of the plants which are very difficult to be transformed. Considering the low transformation efficiency of soybean, we aimed to determine the effect of 6-benzylaminopurine (6-BA), shoot induction time, and infection time of Agrobacterium on the clonal propagation of Glycine max. Results showed that 1.6 mg/L 6-BA could be optimal to promote the induction of adventitious shoots. An induction time of 15 d was considered optimal for the actual experiment involving soybean shoot induction. Agrobacterium was cultured until an OD600 = 0.8 was reached for an infection time of 30 min; this infection time may be optimal to promote soybean transformation. Whole genome DNA methylation was analyzed by high-performance liquid chromatography (HPLC)-assisted quantification, and DNA methylation result is consistent with the phenotypic data of shoot development. In addition, two methylation-related genes (Decrease in DNA methylation 1 and DNA methyltransferases chromomethylase 2) were analyzed to determine expression differences by qRT-PCR in the shoots that were developed under different experimental conditions. In general, the expression values of these genes were normally downregulated under the recommended experimental conditions of soybean regeneration. This study showed the overall methylation changes in the in vitro culture of soybean, as affected by several variable parameters, which is useful to promote the transformation efficiency of soybean. PMID:27183794

  14. Agrobacterium-mediated transformation of the white-rot fungus Physisporinus vitreus.

    PubMed

    Schubert, M; Stührk, C; Fuhr, M J; Schwarze, F W M R

    2013-11-01

    The biotechnologically important white-rot fungus Physisporinus vitreus was co-cultivated with Agrobacterium tumefaciens AGL-1 carrying plasmids with nourseothricin resistance as the selectable marker gene and red fluorescence protein as a visual marker. Mitotically stable transformed isolates were obtained showing red fluorescence protein activity.

  15. DNA METHYLATION ANALYSIS DURING THE OPTIMIZATION OF Agrobacterium-MEDIATED TRANSFORMATION OF SOYBEAN.

    PubMed

    Jiang, J; Wing, V; Xiet, T; Shi, X; Wang, Y P; Sokolov, V

    2016-01-01

    Soybean is recognized as one of the plants which are very difficult to be transformed. Considering the low transformation efficiency of soybean, we aimed to determine the effect of 6-benzylaminopurine (6-BA), shoot induction time, and infection time of Agrobacterium on the clonal propagation of Glycine max. Results showed that 1.6 mg/L 6-BA could be optimal to promote the induction of adventitious shoots. An induction time of 15 d was considered optimal for the actual experiment involving soybean shoot induction. Agrobacterium was cultured until an OD600 = 0.8 was reached for an infection time of 30 min; this infection time may be optimal to promote soybean transformation. Whole genome DNA methylation was analyzed by high-performance liquid chromatography (HPLC)-assisted quantification, and DNA methylation result is consistent with the phenotypic data of shoot development. In addition, two methylation-related genes (Decrease in DNA methylation 1 and DNA methyltransferases chromomethylase 2) were analyzed to determine expression differences by qRT-PCR in the shoots that were developed under different experimental conditions. In general, the expression values of these genes were normally downregulated under the recommended experimental conditions of soybean regeneration. This study showed the overall methylation changes in the in vitro culture of soybean, as affected by several variable parameters, which is useful to promote the transformation efficiency of soybean.

  16. Yeast Actin-Related Protein ARP6 Negatively Regulates Agrobacterium-Mediated Transformation of Yeast Cell.

    PubMed

    Luo, Yumei; Chen, Zikai; Zhu, Detu; Tu, Haitao; Pan, Shen Quan

    2015-01-01

    The yeasts, including Saccharomyces cerevisiae and Pichia pastoris, are single-cell eukaryotic organisms that can serve as models for human genetic diseases and hosts for large scale production of recombinant proteins in current biopharmaceutical industry. Thus, efficient genetic engineering tools for yeasts are of great research and economic values. Agrobacterium tumefaciens-mediated transformation (AMT) can transfer T-DNA into yeast cells as a method for genetic engineering. However, how the T-DNA is transferred into the yeast cells is not well established yet. Here our genetic screening of yeast knockout mutants identified a yeast actin-related protein ARP6 as a negative regulator of AMT. ARP6 is a critical member of the SWR1 chromatin remodeling complex (SWR-C); knocking out some other components of the complex also increased the transformation efficiency, suggesting that ARP6 might regulate AMT via SWR-C. Moreover, knockout of ARP6 led to disruption of microtubule integrity, higher uptake and degradation of virulence proteins, and increased DNA stability inside the cells, all of which resulted in enhanced transformation efficiency. Our findings have identified molecular and cellular mechanisms regulating AMT and a potential target for enhancing the transformation efficiency in yeast cells.

  17. Agrobacterium-mediated transformation of Guignardia citricarpa: an efficient tool to gene transfer and random mutagenesis.

    PubMed

    Rodrigues, Maria Beatriz Calderan; Fávaro, Léia Cecília de Lima; Pallu, Ana Paula de Souza; Ferreira, Anderson; Sebastianes, Fernanda de Souza; Rodrigues, Maria Juliana Calderan; Spósito, Marcel Bellato; de Araújo, Welington Luiz; Pizzirani-Kleiner, Aline Aparecida

    2013-01-01

    Guignardia citricarpa is the causal agent of Citrus Black Spot (CBS), an important disease in Citriculture. Due to the expressive value of this activity worldwide, especially in Brazil, understanding more about the functioning of this fungus is of utmost relevance, making possible the elucidation of its infection mechanisms, and providing tools to control CBS. This work describes for the first time an efficient and successful methodology for genetic transformation of G. citricarpa mycelia, which generated transformants expressing the gene encoding for the gfp (green fluorescent protein) and also their interaction with citrus plant. Mycelia of G. citricarpa were transformed via Agrobacterium tumefaciens, which carried the plasmid pFAT-gfp, contains the genes for hygromycin resistance (hph) as well as gfp. The optimization of the agrotransformation protocol was performed testing different conditions (type of membrane; inductor agent concentration [acetosyringone - AS] and cocultivation time). Results demonstrated that the best condition occurred with the utilization of cellulose's ester membrane; 200 μM of AS and 96 h as cocultivation time. High mitotic stability (82 %) was displayed by transformants using Polymerase Chain Reaction (PCR) technique to confirm the hph gene insertion. In addition, the presence of gfp was observed inside mycelia by epifluorescence optical microscopy. This technique easy visualization of the behaviour of the pathogen interacting with the plant for the first time, allowing future studies on the pathogenesis of this fungus. The establishment of a transformation method for G. citricarpa opens a range of possibilities and facilitates the study of insertional mutagenesis and genetic knockouts, in order to identify the most important genes involved in the pathogenesis mechanisms and plant-pathogen interaction.

  18. A simple shoot multiplication procedure using internode explants, and its application for particle bombardment and Agrobacterium-mediated transformation in watercress.

    PubMed

    Ogita, Shinjiro; Usui, Miki; Shibutani, Nanae; Kato, Yasuo

    2009-07-01

    A shoot multiplication system derived from internode explants was investigated with the aim of improving genetic characteristics of watercress (Nasturtium officinale R. Br.). Internodes of ca. 1 cm excised from in vitro stock shoot culture were placed on half-strength Murashige and Skoog (MS) medium supplemented with 3 muM 2,4-dichlorophenoxyacetic acid as a pre-treatment. Laser scanning microscopy indicated clearly that the first sign of meristematic cell division could be seen after 1-2 days of pre-culture, and meristematic tissues multiplied along the vascular cambium of the internode segment during 7 days of culture. Multiple shoots could be obtained from more than 90% of the pre-treated explants when they were subsequently transferred to MS medium supplemented with 1 muM thidiazuron for 3 weeks. These findings indicate that pre-treatment of the internodes for 7 days promoted their capacity for organogenesis. Using this pre-treatment, frequent generation of transgenic watercress plants was achieved by adapting particle bombardment and Agrobacterium-mediated transformation techniques with a construct expressing a synthetic green florescent protein gene. PMID:19308313

  19. A simple shoot multiplication procedure using internode explants, and its application for particle bombardment and Agrobacterium-mediated transformation in watercress.

    PubMed

    Ogita, Shinjiro; Usui, Miki; Shibutani, Nanae; Kato, Yasuo

    2009-07-01

    A shoot multiplication system derived from internode explants was investigated with the aim of improving genetic characteristics of watercress (Nasturtium officinale R. Br.). Internodes of ca. 1 cm excised from in vitro stock shoot culture were placed on half-strength Murashige and Skoog (MS) medium supplemented with 3 muM 2,4-dichlorophenoxyacetic acid as a pre-treatment. Laser scanning microscopy indicated clearly that the first sign of meristematic cell division could be seen after 1-2 days of pre-culture, and meristematic tissues multiplied along the vascular cambium of the internode segment during 7 days of culture. Multiple shoots could be obtained from more than 90% of the pre-treated explants when they were subsequently transferred to MS medium supplemented with 1 muM thidiazuron for 3 weeks. These findings indicate that pre-treatment of the internodes for 7 days promoted their capacity for organogenesis. Using this pre-treatment, frequent generation of transgenic watercress plants was achieved by adapting particle bombardment and Agrobacterium-mediated transformation techniques with a construct expressing a synthetic green florescent protein gene.

  20. Introduction of a citrus blight-associated gene into Carrizo citrange [Citrus sinensis (L.) Osbc. x Poncirus trifoliata (L.) Raf.] by Agrobacterium-mediated transformation.

    PubMed

    Kayim, M; Ceccardi, T L; Berretta, M J G; Barthe, G A; Derrick, K S

    2004-11-01

    The protein p12 accumulates in leaves of trees with citrus blight (CB), a serious decline of unknown cause. The function of p12 is not known, but sequence analysis indicates it may be related to expansins. In studies to determine the function of p12, sense and antisense constructs were used to make transgenic Carrizo citrange using an Agrobacterium-mediated transformation system. Homogeneous beta-glucuronidase+ (GUS+) sense and antisense transgenic shoots were regenerated using kanamycin as a selective agent. Twenty-five sense and 45 antisense transgenic shoots were in vivo grafted onto Carrizo citrange for further analyses. In addition, 20 sense and 18 antisense shoots were rooted. The homogeneous GUS+ plants contained either the p12 sense or antisense gene (without the intron associated with the gene in untransformed citrus) as shown by PCR and Southern blotting. Northern blots showed the expected RNA in the sense and antisense plants. A protein of identical size and immunoreactivity was observed in seven of nine sense plants but not in nine antisense or non-transgenic plants. At the current stage of growth, there are no visual phenotypic differences between the transgenic and non-transgenic plants. Selected plants will be budded with sweet orange for field evaluation for resistance or susceptibility to CB and general rootstock performance.

  1. IMPa-4, an Arabidopsis importin alpha isoform, is preferentially involved in agrobacterium-mediated plant transformation.

    PubMed

    Bhattacharjee, Saikat; Lee, Lan-Ying; Oltmanns, Heiko; Cao, Hongbin; Veena; Cuperus, Joshua; Gelvin, Stanton B

    2008-10-01

    Successful transformation of plants by Agrobacterium tumefaciens requires that the bacterial T-complex actively escorts T-DNA into the host's nucleus. VirD2 and VirE2 are virulence proteins on the T-complex that have plant-functional nuclear localization signal sequences that may recruit importin alpha proteins of the plant for nuclear import. In this study, we evaluated the involvement of seven of the nine members of the Arabidopsis thaliana importin alpha family in Agrobacterium transformation. Yeast two-hybrid, plant bimolecular fluorescence complementation, and in vitro protein-protein interaction assays demonstrated that all tested Arabidopsis importin alpha members can interact with VirD2 and VirE2. However, only disruption of the importin IMPa-4 inhibited transformation and produced the rat (resistant to Agrobacterium transformation) phenotype. Overexpression of six importin alpha members, including IMPa-4, rescued the rat phenotype in the impa-4 mutant background. Roots of wild-type and impa-4 Arabidopsis plants expressing yellow fluorescent protein-VirD2 displayed nuclear localization of the fusion protein, indicating that nuclear import of VirD2 is not affected in the impa-4 mutant. Somewhat surprisingly, VirE2-yellow fluorescent protein mainly localized to the cytoplasm of both wild-type and impa-4 Arabidopsis cells and to the cytoplasm of wild-type tobacco (Nicotiana tabacum) cells. However, bimolecular fluorescence complementation assays indicated that VirE2 could localize to the nucleus when IMPa-4, but not when IMPa-1, was overexpressed. PMID:18836040

  2. Enhanced Agrobacterium-mediated transformation of embryogenic calli of upland cotton.

    PubMed

    Zhang, Tianzhen; Wu, Shen-Jie

    2012-01-01

    Agrobacterium tumefaciens-mediated transformation of cotton embryogenic calli (EC) was enhanced by choosing appropriate EC and improving efficiency of coculture, selection cultivation, and plant regeneration. The binary vector pBI121 (containing a neomycin phosphotransferase II gene npt-II as a selection marker and a uidA gene as a reporter gene) was used to research transformation efficiency. After 48 h cocultivation, the number of β-glucuronidase (GUS)-positive calli characterized by yellow, loose, and fine-grained EC was twofold greater than that of gray, brown, and coarse granule EC. It indicated that the efficiency of transient transformation was affected by EC morphology. Transient transformation efficiency also was improved by cocultivation on the medium by adding 50 mg/L acetosyringone at 19°C for 48 h. Subculturing EC on the selection medium with low cell density increased the production of kanamycin-resistant (Km-R) calli lines. From an original 0.3 g EC, an average of 20 Km-R calli lines were obtained from a selection dish, and the GUS-positive rate of Km-R clones was 81.97%. A large number of normal plants were rapidly regenerated on the differentiation medium with dehydration treatments, and the GUS-positive rate of regeneration plants was about 72.6%. Polymerase chain reaction analysis of GUS-positive plantlets revealed a 100% positive detection rate for neomycin phosphotransferase II gene and gus gene. Southern blot of transgenic plants regenerated from different Km-R calli lines demonstrated that the target gene, mostly with the low copy number, was integrated into the cotton genome. PMID:22351014

  3. Agrobacterium-mediated transformation of Eucalyptus globulus using explants with shoot apex with introduction of bacterial choline oxidase gene to enhance salt tolerance.

    PubMed

    Matsunaga, Etsuko; Nanto, Kazuya; Oishi, Masatoshi; Ebinuma, Hiroyasu; Morishita, Yoshihiko; Sakurai, Nozomu; Suzuki, Hideyuki; Shibata, Daisuke; Shimada, Teruhisa

    2012-01-01

    Eucalyptus globulus is one of the most economically important plantation hardwoods for paper making. However, its low transformation frequency has prevented genetic engineering of this species with useful genes. We found the hypocotyl section with a shoot apex has the highest regeneration ability among another hypocotyl sections, and have developed an efficient Agrobacterium-mediated transformation method using these materials. We then introduced a salt tolerance gene, namely a bacterial choline oxidase gene (codA) with a GUS reporter gene, into E. globulus. The highest frequency of transgenic shoot regeneration from hypocotyls with shoot apex was 7.4% and the average frequency in four experiments was 4.0%, 12-fold higher than that from hypocotyls without shoot apex. Using about 10,000 explants, over 250 regenerated buds were confirmed as transformants by GUS analysis. Southern blot analysis of 100 elongated shoots confirmed successful generation of stable transformants. Accumulation of glycinebetaine was investigated in 44 selected transgenic lines, which showed 1- to 12-fold higher glycinebetaine levels than non-transgenic controls. Rooting of 16 transgenic lines was successful using a photoautotrophic method under enrichment with 1,000 ppm CO(2). The transgenic whole plantlets were transplanted into potting soil and grown normally in a growth room. They showed salt tolerance to 300 mM NaCl. The points of our system are using explants with shoot apex as materials, inhibiting the elongation of the apex on the selection medium, and regenerating transgenic buds from the side opposite to the apex. This approach may also solve transformation problems in other important plants.

  4. Expression and genomic integration of transgenes after Agrobacterium-mediated transformation of mature barley embryos.

    PubMed

    Uçarlı, C; Tufan, F; Gürel, F

    2015-01-01

    Mature embryos in tissue cultures are advantageous because of their abundance and rapid germination, which reduces genomic instability problems. In this study, 2-day-old isolated mature barley embryos were infected with 2 Agrobacterium hypervirulent strains (AGL1 and EHA105), followed by a 3-day period of co-cultivation in the presence of L-cystein amino acid. Chimeric expression of the b-glucuronidase gene (gusA) directed by a viral promoter of strawberry vein banding virus was observed in coleoptile epidermal cells and seminal roots in 5-day-old germinated seedlings. In addition to varying infectivity patterns in different strains, there was a higher ratio of transient b-glucuronidase expression in developing coleoptiles than in embryonic roots, indicating the high competency of shoot apical meristem cells in the mature embryo. A total of 548 explants were transformed and 156 plants developed to maturity on G418 media after 18-25 days. We detected transgenes in 74% of the screened plant leaves by polymerase chain reaction, and 49% of these expressed neomycin phosphotransferase II gene following AGL1 transformation. Ten randomly selected T0 transformants were analyzed using thermal asymmetric interlaced polymerase chain reaction and 24 fragments ranged between 200-600 base pairs were sequenced. Three of the sequences flanked with transferred-DNA showed high similarity to coding regions of the barley genome, including alpha tubulin5, homeobox 1, and mitochondrial 16S genes. We observed 70-200-base pair filler sequences only in the coding regions of barley in this study. PMID:25730049

  5. Expression and genomic integration of transgenes after Agrobacterium-mediated transformation of mature barley embryos.

    PubMed

    Uçarlı, C; Tufan, F; Gürel, F

    2015-02-06

    Mature embryos in tissue cultures are advantageous because of their abundance and rapid germination, which reduces genomic instability problems. In this study, 2-day-old isolated mature barley embryos were infected with 2 Agrobacterium hypervirulent strains (AGL1 and EHA105), followed by a 3-day period of co-cultivation in the presence of L-cystein amino acid. Chimeric expression of the b-glucuronidase gene (gusA) directed by a viral promoter of strawberry vein banding virus was observed in coleoptile epidermal cells and seminal roots in 5-day-old germinated seedlings. In addition to varying infectivity patterns in different strains, there was a higher ratio of transient b-glucuronidase expression in developing coleoptiles than in embryonic roots, indicating the high competency of shoot apical meristem cells in the mature embryo. A total of 548 explants were transformed and 156 plants developed to maturity on G418 media after 18-25 days. We detected transgenes in 74% of the screened plant leaves by polymerase chain reaction, and 49% of these expressed neomycin phosphotransferase II gene following AGL1 transformation. Ten randomly selected T0 transformants were analyzed using thermal asymmetric interlaced polymerase chain reaction and 24 fragments ranged between 200-600 base pairs were sequenced. Three of the sequences flanked with transferred-DNA showed high similarity to coding regions of the barley genome, including alpha tubulin5, homeobox 1, and mitochondrial 16S genes. We observed 70-200-base pair filler sequences only in the coding regions of barley in this study.

  6. Agrobacterium-mediated transformation of Phalaenopsis by targeting protocorms at an early stage after germination.

    PubMed

    Mishiba, Kei-ichiro; Chin, Dong Poh; Mii, Masahiro

    2005-07-01

    A transformation procedure for phalaenopsis orchid established by using immature protocorms for Agrobacterium infection was aimed at the introduction of target genes into individuals with divergent genetic backgrounds. Protocorms obtained after 21 days of culture on liquid New Dogashima medium were inoculated with Agrobacterium strain EHA101(pIG121Hm) harboring both beta-glucuronidase (GUS) and hygromycin resistance genes. Subculture of the protocorms on acetosyringone-containing medium 2 days before Agrobacterium inoculation gave the highest transformation efficiencies (1.3-1.9%) based on the frequency of hygromycin-resistant plants produced. Surviving protocorms obtained 2 months after Agrobacterium infection on selection medium containing 20 mg l(-1) hygromycin were cut transversely into two pieces before transferring to recovery medium without hygromycin. Protocorm-like bodies (PLBs) proliferated from pieces of protocorms during a 1-month culture on recovery medium followed by transfer to selection medium. Hygromycin-resistant phalaenopsis plants that regenerated after the re-selection culture of PLBs showed histochemical blue staining due to GUS. Transgene integration of the hygromycin-resistant plants was confirmed by Southern blot analysis. A total of 88 transgenic plants, each derived from an independent protocorm, was obtained from ca. 12,500 mature seeds 6 months after infection with Agrobacterium. Due to the convenient protocol for Agrobacterium infection and rapid production of transgenic plants, the present procedure could be utilized to assess expression of transgenes under different genetic backgrounds, and for the molecular breeding of phalaenopsis.

  7. Establishment of an Agrobacterium-mediated Inoculation System for Cucumber Green Mottle Mosaic Virus.

    PubMed

    Kang, Minji; Seo, Jang-Kyun; Song, Dami; Choi, Hong-Soo; Kim, Kook-Hyung

    2015-12-01

    The infectious full-length cDNA clones of Cucumber green mottle mosaic virus (CGMMV) isolates KW and KOM, which were isolated from watermelon and oriental melon, respectively, were constructed under the control of the cauliflower mosaic virus 35S promoter. We successfully inoculated Nicotiana benthamiana with the cloned CGMMV isolates KW and KOM by Agrobacterium-mediated infiltration. Virulence and symptomatic characteristics of the cloned CGMMV isolates KW and KOM were tested on several indicator plants. No obvious differences between two cloned isolates in disease development were observed on the tested indicator plants. We also determined full genome sequences of the cloned CGMMV isolates KW and KOM. Sequence comparison revealed that only four amino acids (at positions 228, 699, 1212, and 1238 of the replicase protein region) differ between the cloned isolates KW and KOM. A previous study reported that the isolate KOM could not infect Chenopodium amaranticolor, but the cloned KOM induced chlorotic spots on the inoculated leaves. When compared with the previously reported sequence of the original KOM isolate, the cloned KOM contained one amino acid mutation (Ala to Thr) at position 228 of the replicase protein, suggesting that this mutation might be responsible for induction of chlorotic spots on the inoculated leaves of C. amaranticolor. PMID:26674677

  8. A new high-frequency Agrobacterium-mediated transformation technique for Sesamum indicum L. using de-embryonated cotyledon as explant.

    PubMed

    Chowdhury, Supriyo; Basu, Arpita; Kundu, Surekha

    2014-09-01

    In spite of the economic importance of sesame (Sesamum indicum L.) and the recent availability of its genome sequence, a high-frequency transformation protocol is still not available. The only two existing Agrobacterium-mediated transformation protocols that are available have poor transformation efficiencies of less than 2%. In the present study, we report a high-frequency, simple, and reproducible transformation protocol for sesame. Transformation was done using de-embryonated cotyledons via somatic embryogenic stages. All the critical parameters of transformation, like incubation period of explants in pre-regeneration medium prior to infection by Agrobacterium tumefaciens, cocultivation period, concentrations of acetosyringone in cocultivation medium, kanamycin concentration, and concentration of plant hormones, including 6-benzylaminopurine, have been optimized. This protocol is superior to the two existing protocols in its high regeneration and transformation efficiencies. The transformed sesame lines have been tested by PCR, RT-PCR for neomycin phosphotransferase II gene expression, and β-glucuronidase (GUS) assay. The regeneration frequency and transformation efficiency are 57.33 and 42.66%, respectively. T0 and T1 generation transgenic plants were analyzed, and several T1 plants homozygous for the transgenes were obtained.

  9. Agrobacterium-mediated genetic transformation of commercially elite rice restorer line using nptII gene as a plant selection marker.

    PubMed

    Chakraborty, M; Sairam Reddy, P; Laxmi Narasu, M; Krishna, Gaurav; Rana, Debashis

    2016-01-01

    Transformation of commercially important indica cultivars remains challenging for the scientific community even though Agrobacterium-mediated transformation protocols for a few indica rice lines have been well established. We report successful transformation of a commercially important restorer line JK1044R of indica rice hybrid JKRH 401. While following existing protocol, we optimized several parameters for callusing, regeneration and genetic transformation of JK1044R. Calli generated from the rice scutellum tissue were used for transformation by Agrobacterium harboring pCAMBIA2201. A novel two tire selection scheme comprising of Geneticin (G418) and Paramomycin were deployed for selection of transgenic calli as well as regenerated plantlets that expressed neomycin phosphotransferase-II gene encoded by the vector. One specific combination of G418 (30 mg l(-1)) and Paramomycin (70 mg l(-1)) was very effective for calli selection. Transformed and selected calli were detected by monitoring the expression of the reporter gene uidA (GUS). Regenerated plantlets were confirmed through PCR analysis of nptII and gus genes specific primers as well as dot blot using gus gene specific as probe. PMID:27186018

  10. Agrobacterium-mediated transformation of leaf base derived callus tissues of popular indica rice (Oryza sativa L. sub sp. indica cv. ADT 43).

    PubMed

    Karthikeyan, Alagarsamy; Pandian, Shunmugiah Karutha; Ramesh, Manikandan

    2011-09-01

    A simple and efficient protocol for the Agrobacterium-mediated transformation of an agronomically useful abiotic sensitive popular indica rice cv. ADT 43 has been developed. Initiation of calli were best achieved from the leaf bases of 4 days old rice seedlings on LS medium supplemented with 2.5mg/L 2,4-D and 1.0mg/L thiamine-HCl. Rice calli immersed in Agrobacterium suspension (strain EHA 105, OD(600)=0.8) were co-cultured on LS30-AsPC medium for 2 days at 25±2°C in the dark. Based on GUS expression analysis, 10min co-cultivation time with 100μM acetosyringone was found optimum for the delivery of gus gene. Calli were proved to be very sensitive to Agrobacterium infection and we found that the level of necrotic response can be minimized after co-cultivation with 30% LS, 10g/L PVP, 10% coconut water and 250mg/L timentin which improved the final transformation efficiency to 9.33%. Molecular and genetic analysis of transgenic plants reveals the integration, expression and inheritance of transgene in the progeny (T(1)) of these plants. The copy number of transgenes has been found to vary from 1 to 2 in transgenic plants (T(0) and T(1)).

  11. High-efficiency Agrobacterium-mediated transformation of chickpea (Cicer arietinum L.) and regeneration of insect-resistant transgenic plants.

    PubMed

    Mehrotra, Meenakshi; Sanyal, Indraneel; Amla, D V

    2011-09-01

    To develop an efficient genetic transformation system of chickpea (Cicer arietinum L.), callus derived from mature embryonic axes of variety P-362 was transformed with Agrobacterium tumefaciens strain LBA4404 harboring p35SGUS-INT plasmid containing the uidA gene encoding β-glucuronidase (GUS) and the nptII gene for kanamycin selection. Various factors affecting transformation efficiency were optimized; as Agrobacterium suspension at OD(600) 0.3 with 48 h of co-cultivation period at 20°C was found optimal for transforming 10-day-old MEA-derived callus. Inclusion of 200 μM acetosyringone, sonication for 4 s with vacuum infiltration for 6 min improved the number of GUS foci per responding explant from 1.0 to 38.6, as determined by histochemical GUS assay. For introducing the insect-resistant trait into chickpea, binary vector pRD400-cry1Ac was also transformed under optimized conditions and 18 T(0) transgenic plants were generated, representing 3.6% transformation frequency. T(0) transgenic plants reflected Mendelian inheritance pattern of transgene segregation in T(1) progeny. PCR, RT-PCR, and Southern hybridization analysis of T(0) and T(1) transgenic plants confirmed stable integration of transgenes into the chickpea genome. The expression level of Bt-Cry protein in T(0) and T(1) transgenic chickpea plants was achieved maximum up to 116 ng mg(-1) of soluble protein, which efficiently causes 100% mortality to second instar larvae of Helicoverpa armigera as analyzed by an insect mortality bioassay. Our results demonstrate an efficient and rapid transformation system of chickpea for producing non-chimeric transgenic plants with high frequency. These findings will certainly accelerate the development of chickpea plants with novel traits.

  12. Evaluation of agrobacterium-mediated transformation of Agaricus bisporus using a range of promoters linked to hygromycin resistance.

    PubMed

    Burns, C; Leach, K M; Elliott, T J; Challen, M P; Foster, G D; Bailey, A

    2006-02-01

    There is interest in establishing genetic modification technologies for the cultivated mushroom Agaricus bisporus, both for improved crop characteristics and for molecular pharming. For these methods to be successful, it is necessary to establish a set of transformation systems that include robust and reliable vectors for gene manipulation. In this article, we report the evaluation of a series of promoters for driving expression of the Escherichia coli hph gene encoding hygromycin phosphotransferase. This was achieved using the Aspergillus nidulans gpdA and the A. bisporus gpdII and trp2 promoters. The Coprinus cinereus beta-tubulin promoter gave contrasting results depending on the size of promoter used, with a 393-bp region being effective, whereas the longer 453-bp fragment failed to yield any hygromycin-resistant transformants. The C. cinereus trp1 and the A. bisporus lcc1 promoters both failed to yield transformants. We also show that transformation efficiency may be improved by careful selection of both appropriate Agrobacterium strains, with AGL-1 yielding more than LBA1126 and by the choice of the binary vectors used to mobilize the DNA, with pCAMBIA vectors appearing to be more efficient than either pBIN19- or pGREEN-based systems.

  13. Improved cotyledonary node method using an alternative explant derived from mature seed for efficient Agrobacterium-mediated soybean transformation.

    PubMed

    Paz, Margie M; Martinez, Juan Carlos; Kalvig, Andrea B; Fonger, Tina M; Wang, Kan

    2006-03-01

    The utility of transformation for soybean improvement requires an efficient system for production of stable transgenic lines. We describe here an improved cotyledonary node method using an alternative explant for Agrobacterium tumefaciens-mediated soybean transformation. We use the term "half-seed" to refer to this alternative cotyledonary explant that is derived from mature seed of soybean following an overnight imbibition and to distinguish it from cotyledonary node derived from 5-7-day-old seedlings. Transformation efficiencies using half-seed explants ranged between 1.4 and 8.7% with an overall efficiency of 3.8% based on the number of transformed events that have been confirmed in the T1 generation by phenotypic assay using the herbicide Liberty (active ingredient glufosinate) and by Southern analysis. This efficiency is 1.5-fold higher than the cotyledonary node method used in our laboratory. Significantly, the half-seed system is simple and does not require deliberate wounding of explants, which is a critical and technically demanding step in the cotyledonary node method. PMID:16249869

  14. Evaluation of agrobacterium-mediated transformation of Agaricus bisporus using a range of promoters linked to hygromycin resistance.

    PubMed

    Burns, C; Leach, K M; Elliott, T J; Challen, M P; Foster, G D; Bailey, A

    2006-02-01

    There is interest in establishing genetic modification technologies for the cultivated mushroom Agaricus bisporus, both for improved crop characteristics and for molecular pharming. For these methods to be successful, it is necessary to establish a set of transformation systems that include robust and reliable vectors for gene manipulation. In this article, we report the evaluation of a series of promoters for driving expression of the Escherichia coli hph gene encoding hygromycin phosphotransferase. This was achieved using the Aspergillus nidulans gpdA and the A. bisporus gpdII and trp2 promoters. The Coprinus cinereus beta-tubulin promoter gave contrasting results depending on the size of promoter used, with a 393-bp region being effective, whereas the longer 453-bp fragment failed to yield any hygromycin-resistant transformants. The C. cinereus trp1 and the A. bisporus lcc1 promoters both failed to yield transformants. We also show that transformation efficiency may be improved by careful selection of both appropriate Agrobacterium strains, with AGL-1 yielding more than LBA1126 and by the choice of the binary vectors used to mobilize the DNA, with pCAMBIA vectors appearing to be more efficient than either pBIN19- or pGREEN-based systems. PMID:16444014

  15. Several components of SKP1/Cullin/F-box E3 ubiquitin ligase complex and associated factors play a role in Agrobacterium-mediated plant transformation.

    PubMed

    Anand, Ajith; Rojas, Clemencia M; Tang, Yuhong; Mysore, Kirankumar S

    2012-07-01

    • Successful genetic transformation of plants by Agrobacterium tumefaciens requires the import of bacterial T-DNA and virulence proteins into the plant cell that eventually form a complex (T-complex). The essential components of the T-complex include the single stranded T-DNA, bacterial virulence proteins (VirD2, VirE2, VirE3 and VirF) and associated host proteins that facilitate the transfer and integration of T-DNA. The removal of the proteins from the T-complex is likely achieved by targeted proteolysis mediated by VirF and the plant ubiquitin proteasome complex. • We evaluated the involvement of the host SKP1/culin/F-box (SCF)-E3 ligase complex and its role in plant transformation. Gene silencing, mutant screening and gene expression studies suggested that the Arabidopsis homologs of yeast SKP1 (suppressor of kinetochore protein 1) protein, ASK1 and ASK2, are required for Agrobacterium-mediated plant transformation. • We identified the role for SGT1b (suppressor of the G2 allele of SKP1), an accessory protein that associates with SCF-complex, in plant transformation. We also report the differential expression of many genes that encode F-box motif containing SKP1-interacting proteins (SKIP) upon Agrobacterium infection. • We speculate that these SKIP genes could encode the plant specific F-box proteins that target the T-complex associated proteins for polyubiquitination and subsequent degradation by the 26S proteasome. PMID:22486382

  16. Functional analysis of autophagy genes via Agrobacterium-mediated transformation in the vascular Wilt fungus Verticillium dahliae.

    PubMed

    Zhou, Lei; Zhao, Jun; Guo, Wangzhen; Zhang, Tianzhen

    2013-08-20

    Autophagy is a widely conserved intracellular process for degradation and recycling of proteins, organelles and cytoplasm in eukaryotic organisms and is now emerging as an important process in foliar infection by many plant pathogenic fungi. However, the role of autophagy in soil-borne fungal physiology and infection biology is poorly understood. Here, we report the establishment of an Agrobacterium tumefaciens-mediated transformation (ATMT) system and its application to investigate two autophagy genes, VdATG8 and VdATG12, by means of targeted gene replacement and complementation. Transformation of a cotton-infecting Verticillium dahliae strain Vd8 with a novel binary vector pCOM led to the production of 384 geneticin-resistant transformants per 1 × 10(6) conidia. V. dahliae mutants lacking either VdATG8 or VdATG12 exhibited reduced conidiation and impaired aerial hyphae production. Disease development on Arabidopsis plants was slightly delayed when inoculated with VdATG8 or VdATG12 gene deletion mutants, compared with the wild-type and gene complemented strains. Surprisingly, in vitro inoculation with unimpaired roots revealed that the abilities of root invasion were not affected in gene deletion mutants. These results indicate that autophagy is necessary for aerial hyphae development and plant colonization but not for root infection in V. dahliae.

  17. An efficient method of agrobacterium-mediated genetic transformation and regeneration in local Indian cultivar of groundnut (Arachis hypogaea) using grafting.

    PubMed

    Tiwari, Vivekanand; Chaturvedi, Amit Kumar; Mishra, Avinash; Jha, Bhavanath

    2015-01-01

    Groundnut (Arachis hypogaea L.) is an industrial crop used as a source of edible oil and nutrients. In this study, an efficient method of regeneration and Agrobacterium-mediated genetic transformation is reported for a local cultivar GG-20 using de-embryonated cotyledon explant. A high regeneration 52.69 ± 2.32 % was achieved by this method with 66.6 μM 6-benzylaminopurine (BAP), while the highest number of shoot buds per explant, 17.67 ± 3.51, was found with 20 μM BAP and 10 μM 2,4-dichlorophenoxyacetic acid (2,4-D). The bacterial culture OD, acetosyringone and L-cysteine concentration were optimized as 1.8, 200 μM and 50 mg L(-1), respectively, in co-cultivation media. It was observed that the addition of 2,4-D in co-cultivation media induced accumulation of endogenous indole-3-acetic acid (IAA). The optimized protocol exhibited 85 % transformation efficiency followed by 14.65 ± 1.06 % regeneration, of which 3.82 ± 0.6 % explants were survived on hygromycin after selection. Finally, 14.58 ± 2.95 % shoots (regenerated on survived explants) were rooted on rooting media (RM3). In grafting method, regenerated shoots (after hygromycin selection) were grafted on the non-transformed stocks with 100 % survival and new leaves emerged in 3 weeks. The putative transgenic plants were then confirmed by PCR, Southern hybridization, reverse transcriptase PCR (RT-PCR) and β-glucuronidase (GUS) histochemical assay. The reported method is efficient and rapid and can also be applied to other crops which are recalcitrant and difficult in rooting.

  18. Assessment of factors influencing the Agrobacterium-mediated in planta seed transformation of brinjal (Solanum melongena L.).

    PubMed

    Subramanyam, Kondeti; Rajesh, Manoharan; Jaganath, Balusamy; Vasuki, Amirthalingam; Theboral, Jeevaraj; Elayaraja, Dhandapani; Karthik, Sivabalan; Manickavasagam, Markandan; Ganapathi, Andy

    2013-09-01

    An efficient and reproducible in planta transformation method was developed for brinjal using seed as an explant. The brinjal seeds were infected with Agrobacterium tumefaciens EHA 105 harbouring pCAMBIA 1301-bar plasmid, and the transformants were selected against BASTA®. Several parameters influencing the in planta seed transformation such as pre-culture duration, acetosyringone concentration, surfactants, duration of sonication, vacuum pressure and vacuum duration have been evaluated. The putatively transformed (T 0) brinjal plants were screened by GUS histochemical analysis. Among the different combinations and concentrations tested, when the 18-h pre-cultured brinjal seeds were sonicated for 20 min and vacuum infiltered for 3 min at 500 mm of Hg in Agrobacterium suspension containing 100 μM acetosyringone, 0.2 % Silwett L-77 favoured the Agrobacterium infection and showed maximum transformation efficiency. Among the five brinjal varieties evaluated, Arka Samhitha showed maximum transformation efficiency at 45.66 %. The transgene was successfully transmitted to progeny plants (T 1) which was evidenced by GUS histochemical analysis, polymerase chain reaction and Southern hybridisation. The in planta protocol developed in the present study would be beneficial to transfer the economically and nutritionally important genes into different varieties of brinjal, and the transgenic brinjal plants can be produced in less time (approximately 27 days).

  19. Transient down-regulation of the RNA silencing machinery increases efficiency of Agrobacterium-mediated transformation of Arabidopsis.

    PubMed

    Bilichak, Andriy; Yao, Youli; Kovalchuk, Igor

    2014-06-01

    Agrobacterium tumefaciens is a plant pathogen that is widely used in plant transformation. As the process of transgenesis includes the delivery of single-stranded T-DNA molecule, we hypothesized that transformation rate may negatively correlate with the efficiency of the RNA-silencing machinery. Using mutants compromised in either the transcriptional or post-transcriptional gene-silencing pathways, two inhibitors of stable transformation were revealed-AGO2 and NRPD1a. Furthermore, an immunoprecipitation experiment has shown that NRPD1, a subunit of Pol IV, directly interacts with Agrobacterium T-DNA in planta. Using the Tobacco rattle virus (TRV)--based virus-induced gene silencing (VIGS) technique, we demonstrated that the transient down-regulation of the expression of either AGO2 or NRPD1a genes in reproductive organs of Arabidopsis, leads to an increase in transformation rate. We observed a 6.0- and 3.5-fold increase in transformation rate upon transient downregulation of either AGO2 or NRPD1a genes, respectively. This is the first report demonstrating the increase in the plant transformation rate via VIGS-mediated transient down-regulation of the components of epigenetic machinery in reproductive tissue.

  20. Transgene stacking and marker elimination in transgenic rice by sequential Agrobacterium-mediated co-transformation with the same selectable marker gene.

    PubMed

    Ramana Rao, Mangu Venkata; Parameswari, Chidambaram; Sripriya, Rajasekaran; Veluthambi, Karuppannan

    2011-07-01

    Rice chitinase (chi11) and tobacco osmotin (ap24) genes, which cause disruption of fungal cell wall and cell membrane, respectively, were stacked in transgenic rice to develop resistance against the sheath blight disease. The homozygous marker-free transgenic rice line CoT23 which harboured the rice chi11 transgene was sequentially re-transformed with a second transgene ap24 by co-transformation using an Agrobacterium tumefaciens strain harbouring a single-copy cointegrate vector pGV2260::pSSJ1 and a multi-copy binary vector pBin19∆nptII-ap24 in the same cell. pGV2260::pSSJ1 T-DNA carried the hygromycin phosphotransferase (hph) and β-glucuronidase (gus) genes. pBin19∆nptII-ap24 T-DNA harboured the tobacco osmotin (ap24) gene. Co-transformation of the gene of interest (ap24) with the selectable marker gene (SMG, hph) occurred in 12 out of 18 T(0) plants (67%). Segregation of hph from ap24 was accomplished in the T(1) generation in one (line 11) of the four analysed co-transformed plants. The presence of ap24 and chi11 transgenes and the absence of the hph gene in the SMG-eliminated T(1) plants of the line 11 were confirmed by DNA blot analyses. The SMG-free transgenic plants of the line 11 harboured a single copy of the ap24 gene. Homozygous, SMG-free T(2) plants of the transgenic line 11 harboured stacked transgenes, chi11 and ap24. Northern blot analysis of the SMG-free plants revealed constitutive expression of chi11 and ap24. The transgenic plants with stacked transgenes displayed high levels of resistance against Rhizoctonia solani. Thus, we demonstrate the development of transgene-stacked and marker-free transgenic rice by sequential Agrobacterium-mediated co-transformation with the same SMG.

  1. Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14

    PubMed Central

    Nyaboga, Evans N.; Njiru, Joshua M.; Tripathi, Leena

    2015-01-01

    Routine production of large numbers of transgenic plants is required to fully exploit advances in cassava biotechnology and support development of improved germplasm for deployment to farmers. This article describes an improved, high-efficiency transformation protocol for recalcitrant cassava cultivar TME14 preferred in Africa. Factors that favor production of friable embryogenic calli (FEC) were found to be use of DKW medium, crushing of organized embryogenic structures (OES) through 1–2 mm sized metal wire mesh, washing of crushed OES tissues and short exposure of tyrosine to somatic embryos; and transformation efficiency was enhanced by use of low Agrobacterium density during co-cultivation, co-centrifugation of FEC with Agrobacterium, germination of paramomycin resistant somatic embryos on medium containing BAP with gradual increase in concentration and variations of the frequency of subculture of cotyledonary-stage embryos on shoot elongation medium. By applying the optimized parameters, FEC were produced for cassava cultivar TME14 and transformed using Agrobacterium strain LBA4404 harboring the binary vector pCAMBIA2301. About 70–80 independent transgenic lines per ml settled cell volume (SCV) of FEC were regenerated on selective medium. Histochemical GUS assays confirmed the expression of gusA gene in transformed calli, somatic embryos and transgenic plants. The presence and integration of the gusA gene were confirmed by PCR and Southern blot analysis, respectively. RT-PCR analysis of transgenic plants confirmed the expression of gusA gene. This protocol demonstrates significantly enhanced transformation efficiency over existing cassava transformation protocols and could become a powerful tool for functional genomics and transferring new traits into cassava. PMID:26113851

  2. Factors influencing somatic embryogenesis, regeneration, and Agrobacterium-mediated transformation of cassava (Manihot esculenta Crantz) cultivar TME14.

    PubMed

    Nyaboga, Evans N; Njiru, Joshua M; Tripathi, Leena

    2015-01-01

    Routine production of large numbers of transgenic plants is required to fully exploit advances in cassava biotechnology and support development of improved germplasm for deployment to farmers. This article describes an improved, high-efficiency transformation protocol for recalcitrant cassava cultivar TME14 preferred in Africa. Factors that favor production of friable embryogenic calli (FEC) were found to be use of DKW medium, crushing of organized embryogenic structures (OES) through 1-2 mm sized metal wire mesh, washing of crushed OES tissues and short exposure of tyrosine to somatic embryos; and transformation efficiency was enhanced by use of low Agrobacterium density during co-cultivation, co-centrifugation of FEC with Agrobacterium, germination of paramomycin resistant somatic embryos on medium containing BAP with gradual increase in concentration and variations of the frequency of subculture of cotyledonary-stage embryos on shoot elongation medium. By applying the optimized parameters, FEC were produced for cassava cultivar TME14 and transformed using Agrobacterium strain LBA4404 harboring the binary vector pCAMBIA2301. About 70-80 independent transgenic lines per ml settled cell volume (SCV) of FEC were regenerated on selective medium. Histochemical GUS assays confirmed the expression of gusA gene in transformed calli, somatic embryos and transgenic plants. The presence and integration of the gusA gene were confirmed by PCR and Southern blot analysis, respectively. RT-PCR analysis of transgenic plants confirmed the expression of gusA gene. This protocol demonstrates significantly enhanced transformation efficiency over existing cassava transformation protocols and could become a powerful tool for functional genomics and transferring new traits into cassava.

  3. Sonication, Vacuum Infiltration and Thiol Compounds Enhance the Agrobacterium-Mediated Transformation Frequency of Withania somnifera (L.) Dunal

    PubMed Central

    Sivanandhan, Ganeshan; Kapil Dev, Gnajothi; Theboral, Jeevaraj; Selvaraj, Natesan; Ganapathi, Andy; Manickavasagam, Markandan

    2015-01-01

    In the present study, we have established a stable transformation protocol via Agrobacterium tumafacines for the pharmaceutically important Withania somnifera. Six day-old nodal explants were used for 3 day co-cultivation with Agrobacterium tumefaciens strain LBA4404 harbouring the vector pCAMIBA2301. Among the different injury treatments, sonication, vacuum infiltration and their combination treatments tested, a vacuum infiltration for 10 min followed by sonication for 10 sec with A. tumefaciens led to a higher transient GUS expression (84% explants expressing GUS at regenerating sites). In order to improve gene integration, thiol compounds were added to co-cultivation medium. A combined treatment of L-Cys at 100 mg/l, STS at 125 mg/l, DTT at 75 mg/l resulted in a higher GUS expression (90%) in the nodal explants. After 3 days of co-cultivation, the explants were subjected to three selection cycles with increasing concentrations of kanamycin [100 to 115 mg/l]. The integration and expression of gusA gene in T0 and T1 transgenic plants were confirmed by polymerase chain reaction (PCR), and Southern blott analysis. These transformed plants (T0 and T1) were fertile and morphologically normal. From the present investigation, we have achieved a higher transformation efficiency of (10%). Withanolides (withanolide A, withanolide B, withanone and withaferin A) contents of transformed plants (T0 and T1) were marginally higher than control plants. PMID:25927703

  4. Sonication, Vacuum Infiltration and Thiol Compounds Enhance the Agrobacterium-Mediated Transformation Frequency of Withania somnifera (L.) Dunal.

    PubMed

    Sivanandhan, Ganeshan; Kapil Dev, Gnajothi; Theboral, Jeevaraj; Selvaraj, Natesan; Ganapathi, Andy; Manickavasagam, Markandan

    2015-01-01

    In the present study, we have established a stable transformation protocol via Agrobacterium tumafacines for the pharmaceutically important Withania somnifera. Six day-old nodal explants were used for 3 day co-cultivation with Agrobacterium tumefaciens strain LBA4404 harbouring the vector pCAMIBA2301. Among the different injury treatments, sonication, vacuum infiltration and their combination treatments tested, a vacuum infiltration for 10 min followed by sonication for 10 sec with A. tumefaciens led to a higher transient GUS expression (84% explants expressing GUS at regenerating sites). In order to improve gene integration, thiol compounds were added to co-cultivation medium. A combined treatment of L-Cys at 100 mg/l, STS at 125 mg/l, DTT at 75 mg/l resulted in a higher GUS expression (90%) in the nodal explants. After 3 days of co-cultivation, the explants were subjected to three selection cycles with increasing concentrations of kanamycin [100 to 115 mg/l]. The integration and expression of gusA gene in T0 and T1 transgenic plants were confirmed by polymerase chain reaction (PCR), and Southern blott analysis. These transformed plants (T0 and T1) were fertile and morphologically normal. From the present investigation, we have achieved a higher transformation efficiency of (10%). Withanolides (withanolide A, withanolide B, withanone and withaferin A) contents of transformed plants (T0 and T1) were marginally higher than control plants.

  5. IMPa-4, an Arabidopsis Importin α Isoform, Is Preferentially Involved in Agrobacterium-Mediated Plant Transformation[W

    PubMed Central

    Bhattacharjee, Saikat; Lee, Lan-Ying; Oltmanns, Heiko; Cao, Hongbin; Veena; Cuperus, Joshua; Gelvin, Stanton B.

    2008-01-01

    Successful transformation of plants by Agrobacterium tumefaciens requires that the bacterial T-complex actively escorts T-DNA into the host's nucleus. VirD2 and VirE2 are virulence proteins on the T-complex that have plant-functional nuclear localization signal sequences that may recruit importin α proteins of the plant for nuclear import. In this study, we evaluated the involvement of seven of the nine members of the Arabidopsis thaliana importin α family in Agrobacterium transformation. Yeast two-hybrid, plant bimolecular fluorescence complementation, and in vitro protein–protein interaction assays demonstrated that all tested Arabidopsis importin α members can interact with VirD2 and VirE2. However, only disruption of the importin IMPa-4 inhibited transformation and produced the rat (resistant to Agrobacterium transformation) phenotype. Overexpression of six importin α members, including IMPa-4, rescued the rat phenotype in the impa-4 mutant background. Roots of wild-type and impa-4 Arabidopsis plants expressing yellow fluorescent protein–VirD2 displayed nuclear localization of the fusion protein, indicating that nuclear import of VirD2 is not affected in the impa-4 mutant. Somewhat surprisingly, VirE2–yellow fluorescent protein mainly localized to the cytoplasm of both wild-type and impa-4 Arabidopsis cells and to the cytoplasm of wild-type tobacco (Nicotiana tabacum) cells. However, bimolecular fluorescence complementation assays indicated that VirE2 could localize to the nucleus when IMPa-4, but not when IMPa-1, was overexpressed. PMID:18836040

  6. Increased Agrobacterium-mediated transformation and rooting efficiencies in canola (Brassica napus L.) from hypocotyl segment explants

    NASA Technical Reports Server (NTRS)

    Cardoza, V.; Stewart, C. N.

    2003-01-01

    An efficient protocol for the production of transgenic Brassica napus cv. Westar plants was developed by optimizing two important parameters: preconditioning time and co-cultivation time. Agrobacterium tumefaciens-mediated transformation was performed using hypocotyls as explant tissue. Two variants of a green fluorescent protein (GFP)-encoding gene--mGFP5-ER and eGFP--both under the constitutive expression of the cauliflower mosaic virus 35S promoter, were used for the experiments. Optimizing the preconditioning time to 72 h and co-cultivation time with Agrobacterium to 48 h provided the increase in the transformation efficiency from a baseline of 4% to 25%. With mGFP5-ER, the transformation rate was 17% and with eGFP it was 25%. Transgenic shoots were selected on 200 mg/l kanamycin. Rooting efficiency was 100% on half-strength Murashige and Skoog medium with 10 g/l sucrose and 0.5 mg/l indole butyric acid in the presence of kanamycin.

  7. Agrobacterium-mediated transformation of the β-subunit gene in 7S globulin protein in soybean using RNAi technology.

    PubMed

    Qu, J; Liu, S Y; Wang, P W; Guan, S Y; Fan, Y G; Yao, D; Zhang, L; Dai, J L

    2016-01-01

    The objective of this study was to use RNA interference (RNAi) to improve protein quality and decrease anti-nutritional effects in soybean. Agrobacterium tumefaciens-mediated transformation was conducted using RNAi and an expression vector containing the 7S globulin β-subunit gene. The BAR gene was used as the selective marker and cotyledonary nodes of soybean genotype Jinong 27 were chosen as explant material. Regenerated plants were detected by molecular biology techniques. Transformation of the β-subunit gene in the 7S protein was detected by PCR, Southern blot, and q-PCR. Positive plants (10 T0, and 6 T1, and 13 T2) were tested by PCR. Hybridization bands were detected by Southern blot analysis in two of the T1 transgenic plants. RNAi expression vectors containing the soybean 7S protein β-subunit gene were successfully integrated into the genome of transgenic plants. qRT-PCR analysis in soybean seeds showed a clear decrease in expression of the soybean β-subunit gene. The level of 7S protein β-subunit expression in transgenic plants decreased by 77.5% as compared to that of the wild-type plants. This study has established a basis for the application of RNAi to improve the anti-nutritional effects of soybean. PMID:27173254

  8. Agrobacterium-mediated transformation of the β-subunit gene in 7S globulin protein in soybean using RNAi technology.

    PubMed

    Qu, J; Liu, S Y; Wang, P W; Guan, S Y; Fan, Y G; Yao, D; Zhang, L; Dai, J L

    2016-04-26

    The objective of this study was to use RNA interference (RNAi) to improve protein quality and decrease anti-nutritional effects in soybean. Agrobacterium tumefaciens-mediated transformation was conducted using RNAi and an expression vector containing the 7S globulin β-subunit gene. The BAR gene was used as the selective marker and cotyledonary nodes of soybean genotype Jinong 27 were chosen as explant material. Regenerated plants were detected by molecular biology techniques. Transformation of the β-subunit gene in the 7S protein was detected by PCR, Southern blot, and q-PCR. Positive plants (10 T0, and 6 T1, and 13 T2) were tested by PCR. Hybridization bands were detected by Southern blot analysis in two of the T1 transgenic plants. RNAi expression vectors containing the soybean 7S protein β-subunit gene were successfully integrated into the genome of transgenic plants. qRT-PCR analysis in soybean seeds showed a clear decrease in expression of the soybean β-subunit gene. The level of 7S protein β-subunit expression in transgenic plants decreased by 77.5% as compared to that of the wild-type plants. This study has established a basis for the application of RNAi to improve the anti-nutritional effects of soybean.

  9. Agrobacterium-mediated transformation of kabocha squash (Cucurbita moschata Duch) induced by wounding with aluminum borate whiskers.

    PubMed

    Nanasato, Yoshihiko; Konagaya, Ken-ichi; Okuzaki, Ayako; Tsuda, Mai; Tabei, Yutaka

    2011-08-01

    An efficient genetic transformation method for kabocha squash (Cucurbita moschata Duch cv. Heiankogiku) was established by wounding cotyledonary node explants with aluminum borate whiskers prior to inoculation with Agrobacterium. Adventitious shoots were induced from only the proximal regions of the cotyledonary nodes and were most efficiently induced on Murashige-Skoog agar medium with 1 mg/L benzyladenine. Vortexing with 1% (w/v) aluminum borate whiskers significantly increased Agrobacterium infection efficiency in the proximal region of the explants. Transgenic plants were screened at the T(0) generation by sGFP fluorescence, genomic PCR, and Southern blot analyses. These transgenic plants grew normally and T(1) seeds were obtained. We confirmed stable integration of the transgene and its inheritance in T(1) generation plants by sGFP fluorescence and genomic PCR analyses. The average transgenic efficiency for producing kabocha squashes with our method was about 2.7%, a value sufficient for practical use.

  10. In vitro regeneration and optimization of factors affecting Agrobacterium mediated transformation in Artemisia Pallens, an important medicinal plant.

    PubMed

    Alok, Anshu; Shukla, Vishnu; Pala, Zarna; Kumar, Jitesh; Kudale, Subhash; Desai, Neetin

    2016-04-01

    Artemisia pallens is an important medicinal plant. In-vitro regeneration and multiplication of A. pallens have been established using attached cotyledons. Different growth regulators were considered for regeneration of multiple shoots. An average of 36 shoots per explants were obtained by culturing attached cotyledons on Murashige and Skoog's medium containing 2 mg/L BAP and 0.1 mg/L NAA, after 45 days. The shoots were rooted best on half Murashige and Skoog's medium with respect to media containing 1 mg/L IBA or 1 mg/L NAA. Different parameters such as type of bacterial strains, OD600 of bacterial culture, co-cultivation duration, concentration of acetosyringone and explants type were optimized for transient expression of the reporter gene. Agrobacterium tumefaciens harbouring pCambia1301 plasmid carrying β-glucuronidase as a reporter gene and hygromycin phosphotransferase as plant selectable marker genes were used for genetic transformation of A. pallens. Hygromycin lethality test showed concentration of 15 mg/L were sufficient to inhibit the growth of attached cotyledons and multiple shoot buds of nontransgenics in selection media. Up to 83 % transient transformation was found when attached cotyledons were co-cultivated with Agrobacterium strain AGL1 for 2 days at 22 °C on shoot induction medium. The bacterial growth was eliminated by addition of cefotaxime (200 mg/L) in selection media. T0 transgenic plants were confirmed by GUS histochemical assay and further by polymerase chain reaction (PCR) using uidA and hpt gene specific primers. The study is useful in establishing technological improvement in A. pallens by genetic engineering. PMID:27436917

  11. Identifying a Carotenoid Cleavage Dioxygenase 4a Gene and Its Efficient Agrobacterium-Mediated Genetic Transformation in Bixa orellana L.

    PubMed

    Sankari, Mohan; Hemachandran, Hridya; Anantharaman, Amirtha; Babu, Subramanian; Madrid, Renata Rivera; C, George Priya Doss; Fulzele, Devanand P; Siva, Ramamoorthy

    2016-07-01

    Carotenoids are metabolized to apocarotenoids through the pathway catalysed by carotenoid cleavage oxygenases (CCOs). The apocarotenoids are economically important as it is known to have therapeutic as well as industrial applications. For instance, bixin from Bixa orellana and crocin from Crocus sativus are commercially used as a food colourant and cosmetics since prehistoric time. In our present study, CCD4a gene has been identified and isolated from leaves of B. orellana for the first time and named as BoCCD4a; phylogenetic analysis was carried out using CLUSTAL W. From sequence analysis, BoCCD4a contains two exons and one intron, which was compared with the selected AtCCD4, RdCCD4, GmCCD4 and CmCCD4a gene. Further, the BoCCD4a gene was cloned into pCAMBIA 1301, transformed into Agrobacterium tumefaciens EHA105 strain and subsequently transferred into hypocotyledons and callus of B. orellana by agro-infection. Selection of stable transformation was screened on the basis of PCR detection by using GUS and hptII specific primer, which was followed by histochemical characterization. The percent transient GUS expression in hypocotyledons and callus was 84.4 and 80 %, respectively. The expression of BoCCD4a gene in B. orellana was confirmed through RT-PCR analysis. From our results, the sequence analysis of BoCCD4a gene of B. orellana was closely related to the CsCCD4 gene of C. sativus, which suggests this gene may have a role in various processes such as fragrance, insect attractant and pollination. PMID:26922728

  12. In vitro regeneration and optimization of factors affecting Agrobacterium mediated transformation in Artemisia Pallens, an important medicinal plant.

    PubMed

    Alok, Anshu; Shukla, Vishnu; Pala, Zarna; Kumar, Jitesh; Kudale, Subhash; Desai, Neetin

    2016-04-01

    Artemisia pallens is an important medicinal plant. In-vitro regeneration and multiplication of A. pallens have been established using attached cotyledons. Different growth regulators were considered for regeneration of multiple shoots. An average of 36 shoots per explants were obtained by culturing attached cotyledons on Murashige and Skoog's medium containing 2 mg/L BAP and 0.1 mg/L NAA, after 45 days. The shoots were rooted best on half Murashige and Skoog's medium with respect to media containing 1 mg/L IBA or 1 mg/L NAA. Different parameters such as type of bacterial strains, OD600 of bacterial culture, co-cultivation duration, concentration of acetosyringone and explants type were optimized for transient expression of the reporter gene. Agrobacterium tumefaciens harbouring pCambia1301 plasmid carrying β-glucuronidase as a reporter gene and hygromycin phosphotransferase as plant selectable marker genes were used for genetic transformation of A. pallens. Hygromycin lethality test showed concentration of 15 mg/L were sufficient to inhibit the growth of attached cotyledons and multiple shoot buds of nontransgenics in selection media. Up to 83 % transient transformation was found when attached cotyledons were co-cultivated with Agrobacterium strain AGL1 for 2 days at 22 °C on shoot induction medium. The bacterial growth was eliminated by addition of cefotaxime (200 mg/L) in selection media. T0 transgenic plants were confirmed by GUS histochemical assay and further by polymerase chain reaction (PCR) using uidA and hpt gene specific primers. The study is useful in establishing technological improvement in A. pallens by genetic engineering.

  13. AGROBEST: an efficient Agrobacterium-mediated transient expression method for versatile gene function analyses in Arabidopsis seedlings

    PubMed Central

    2014-01-01

    Background Transient gene expression via Agrobacterium-mediated DNA transfer offers a simple and fast method to analyze transgene functions. Although Arabidopsis is the most-studied model plant with powerful genetic and genomic resources, achieving highly efficient and consistent transient expression for gene function analysis in Arabidopsis remains challenging. Results We developed a highly efficient and robust Agrobacterium-mediated transient expression system, named AGROBEST (Agrobacterium-mediated enhanced seedling transformation), which achieves versatile analysis of diverse gene functions in intact Arabidopsis seedlings. Using β-glucuronidase (GUS) as a reporter for Agrobacterium-mediated transformation assay, we show that the use of a specific disarmed Agrobacterium strain with vir gene pre-induction resulted in homogenous GUS staining in cotyledons of young Arabidopsis seedlings. Optimization with AB salts in plant culture medium buffered with acidic pH 5.5 during Agrobacterium infection greatly enhanced the transient expression levels, which were significantly higher than with two existing methods. Importantly, the optimized method conferred 100% infected seedlings with highly increased transient expression in shoots and also transformation events in roots of ~70% infected seedlings in both the immune receptor mutant efr-1 and wild-type Col-0 seedlings. Finally, we demonstrated the versatile applicability of the method for examining transcription factor action and circadian reporter-gene regulation as well as protein subcellular localization and protein–protein interactions in physiological contexts. Conclusions AGROBEST is a simple, fast, reliable, and robust transient expression system enabling high transient expression and transformation efficiency in Arabidopsis seedlings. Demonstration of the proof-of-concept experiments elevates the transient expression technology to the level of functional studies in Arabidopsis seedlings in addition to previous

  14. [Agrobacterium-mediated sunflower transformation (Helianthus annuus L.) in vitro and in Planta using strain of LBA4404 harboring binary vector pBi2E with dsRNA-suppressor proline dehydrogenase gene].

    PubMed

    Tishchenko, E N; Komisarenko, A G; Mikhal'skaia, S I; Sergeeva, L E; Adamenko, N I; Morgun, B V; Kochetov, A V

    2014-01-01

    To estimate the efficiency of proline dehydrogenase gene suppression towards increasing of sunflower (Helianthus annuus L.) tolerance level to water deficit and salinity, we employed strain LBA4404 harboring pBi2E with double-stranded RNA-suppressor, which were prepared on basis arabidopsis ProDH1 gene. The techniques of Agrobacterium-mediated transformation in vitro and in planta during fertilization sunflower have been proposed. There was shown the genotype-depended integration of T-DNA in sunflower genome. PCR-analysis showed that ProDH1 presents in genome of inbred lines transformed in planta, as well as in T1- and T2-generations. In trans-genic regenerants the essential accumulation of free L-proline during early stages of in vitro cultivation under normal conditions was shown. There was established the essential accumulation of free proline in transgenic regenerants during cultivation under lethal stress pressure (0.4 M mannitol and 2.0% sea water salts) and its decline upon the recovery period. These data are declared about effectiveness of suppression of sunflower ProDH and gene participation in processes connected with osmotolerance.

  15. A Dark Incubation Period Is Important for Agrobacterium-Mediated Transformation of Mature Internode Explants of Sweet Orange, Grapefruit, Citron, and a Citrange Rootstock

    PubMed Central

    Marutani-Hert, Mizuri; Bowman, Kim D.; McCollum, Greg T.; Mirkov, T. Erik; Evens, Terence J.; Niedz, Randall P.

    2012-01-01

    Background Citrus has an extended juvenile phase and trees can take 2–20 years to transition to the adult reproductive phase and produce fruit. For citrus variety development this substantially prolongs the time before adult traits, such as fruit yield and quality, can be evaluated. Methods to transform tissue from mature citrus trees would shorten the evaluation period via the direct production of adult phase transgenic citrus trees. Methodology/Principal Findings Factors important for promoting shoot regeneration from internode explants from adult phase citrus trees were identified and included a dark incubation period and the use of the cytokinin zeatin riboside. Transgenic trees were produced from four citrus types including sweet orange, citron, grapefruit, and a trifoliate hybrid using the identified factors and factor settings. Significance The critical importance of a dark incubation period for shoot regeneration was established. These results confirm previous reports on the feasibility of transforming mature tissue from sweet orange and are the first to document the transformation of mature tissue from grapefruit, citron, and a trifoliate hybrid. PMID:23082165

  16. Agrobacterium-mediated transformation of tomato with rolB gene results in enhancement of fruit quality and foliar resistance against fungal pathogens.

    PubMed

    Arshad, Waheed; Haq, Ihsan-ul-; Waheed, Mohammad Tahir; Mysore, Kirankumar S; Mirza, Bushra

    2014-01-01

    Tomato (Solanum lycopersicum L.) is the second most important cultivated crop next to potato, worldwide. Tomato serves as an important source of antioxidants in human diet. Alternaria solani and Fusarium oxysporum cause early blight and vascular wilt of tomato, respectively, resulting in severe crop losses. The foremost objective of the present study was to generate transgenic tomato plants with rolB gene and evaluate its effect on plant morphology, nutritional contents, yield and resistance against fungal infection. Tomato cv. Rio Grande was transformed via Agrobacterium tumefaciens harbouring rolB gene of Agrobacterium rhizogenes. rolB. Biochemical analyses showed considerable improvement in nutritional quality of transgenic tomato fruits as indicated by 62% increase in lycopene content, 225% in ascorbic acid content, 58% in total phenolics and 26% in free radical scavenging activity. Furthermore, rolB gene significantly improved the defence response of leaves of transgenic plants against two pathogenic fungal strains A. solani and F. oxysporum. Contrarily, transformed plants exhibited altered morphology and reduced fruit yield. In conclusion, rolB gene from A. rhizogenes can be used to generate transgenic tomato with increased nutritional contents of fruits as well as improved foliar tolerance against fungal pathogens. PMID:24817272

  17. Agrobacterium-mediated transformation of tomato with rolB gene results in enhancement of fruit quality and foliar resistance against fungal pathogens.

    PubMed

    Arshad, Waheed; Haq, Ihsan-ul-; Waheed, Mohammad Tahir; Mysore, Kirankumar S; Mirza, Bushra

    2014-01-01

    Tomato (Solanum lycopersicum L.) is the second most important cultivated crop next to potato, worldwide. Tomato serves as an important source of antioxidants in human diet. Alternaria solani and Fusarium oxysporum cause early blight and vascular wilt of tomato, respectively, resulting in severe crop losses. The foremost objective of the present study was to generate transgenic tomato plants with rolB gene and evaluate its effect on plant morphology, nutritional contents, yield and resistance against fungal infection. Tomato cv. Rio Grande was transformed via Agrobacterium tumefaciens harbouring rolB gene of Agrobacterium rhizogenes. rolB. Biochemical analyses showed considerable improvement in nutritional quality of transgenic tomato fruits as indicated by 62% increase in lycopene content, 225% in ascorbic acid content, 58% in total phenolics and 26% in free radical scavenging activity. Furthermore, rolB gene significantly improved the defence response of leaves of transgenic plants against two pathogenic fungal strains A. solani and F. oxysporum. Contrarily, transformed plants exhibited altered morphology and reduced fruit yield. In conclusion, rolB gene from A. rhizogenes can be used to generate transgenic tomato with increased nutritional contents of fruits as well as improved foliar tolerance against fungal pathogens.

  18. Agrobacterium-mediated transformation of Mexican lime (Citrus aurantifolia Swingle) using optimized systems for epicotyls and cotelydons

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Epicotyl and internodal stem segments provide the predominantly used explants for regeneration of transgenic citrus plants following co-cultivation with Agrobacterium. Previous reports using epicotyls segments from Mexican lime have shown low affinity for Agrobacterium tumefaciens infection which re...

  19. Transformation of Vicia narbonensis via Agrobacterium-mediated gene transfer.

    PubMed

    Pickardt, T; Meixner, M; Schade, V; Schieder, O

    1991-02-01

    Shoot tips and epicotyl-segments of Vicia narbonensis were co-cultivated with Agrobacterium tumefaciens strain C58C1 pGV 3850 HPT, carrying a plasmid coding for hygromycin-phosphotransferase. On callus-induction medium containing 60 mg/l hygromycin for selection, approximately 18% of the explants produced hygromycin-resistant callus. After transfer to regeneration-medium these calluses produced hygromycin-resistant and nopaline-positive somatic embryos which could be regenerated to plantlets. The integration of the T-DNA into the plant genome was confirmed by Southern analysis.

  20. Transformation of somatic embryos of Prunus incisa ‘February Pink’ with a visible reporter gene

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An Agrobacterium-mediated transformation system was developed for the ornamental cherry species Prunus incisa. This system uses both an antibiotic resistance gene (NPTII) and a visible selectable marker, the green fluorescent protein (GFP), to select plants. Cells from leaf and root explants were tr...

  1. Environment Control to Improve Recombinant Protein Yields in Plants Based on Agrobacterium-Mediated Transient Gene Expression.

    PubMed

    Fujiuchi, Naomichi; Matoba, Nobuyuki; Matsuda, Ryo

    2016-01-01

    Agrobacterium-mediated transient expression systems enable plants to produce a wide range of recombinant proteins on a rapid timescale. To achieve economically feasible upstream production and downstream processing, two yield parameters should be considered: (1) recombinant protein content per unit biomass and (2) recombinant protein productivity per unit area-time at the end of the upstream production. Because environmental factors in the upstream production have impacts on these parameters, environment control is important to maximize the recombinant protein yield. In this review, we summarize the effects of pre- and postinoculation environmental factors in the upstream production on the yield parameters and discuss the basic concept of environment control for plant-based transient expression systems. Preinoculation environmental factors associated with planting density, light quality, and nutrient supply affect plant characteristics, such as biomass and morphology, which in turn affect recombinant protein content and productivity. Accordingly, environment control for such plant characteristics has significant implications to achieve a high yield. On the other hand, postinoculation environmental factors, such as temperature, light intensity, and humidity, have been shown to affect recombinant protein content. Considering that recombinant protein production in Agrobacterium-mediated transient expression systems is a result of a series of complex biological events starting from T-DNA transfer from Agrobacterium tumefaciens to protein biosynthesis and accumulation in leaf tissue, we propose that dynamic environment control during the postinoculation process, i.e., changing environmental conditions at an appropriate timing for each event, may be a promising approach to obtain a high yield. Detailed descriptions of plant growth conditions and careful examination of environmental effects will significantly contribute to our knowledge to stably obtain high recombinant

  2. Environment Control to Improve Recombinant Protein Yields in Plants Based on Agrobacterium-Mediated Transient Gene Expression

    PubMed Central

    Fujiuchi, Naomichi; Matoba, Nobuyuki; Matsuda, Ryo

    2016-01-01

    Agrobacterium-mediated transient expression systems enable plants to produce a wide range of recombinant proteins on a rapid timescale. To achieve economically feasible upstream production and downstream processing, two yield parameters should be considered: (1) recombinant protein content per unit biomass and (2) recombinant protein productivity per unit area–time at the end of the upstream production. Because environmental factors in the upstream production have impacts on these parameters, environment control is important to maximize the recombinant protein yield. In this review, we summarize the effects of pre- and postinoculation environmental factors in the upstream production on the yield parameters and discuss the basic concept of environment control for plant-based transient expression systems. Preinoculation environmental factors associated with planting density, light quality, and nutrient supply affect plant characteristics, such as biomass and morphology, which in turn affect recombinant protein content and productivity. Accordingly, environment control for such plant characteristics has significant implications to achieve a high yield. On the other hand, postinoculation environmental factors, such as temperature, light intensity, and humidity, have been shown to affect recombinant protein content. Considering that recombinant protein production in Agrobacterium-mediated transient expression systems is a result of a series of complex biological events starting from T-DNA transfer from Agrobacterium tumefaciens to protein biosynthesis and accumulation in leaf tissue, we propose that dynamic environment control during the postinoculation process, i.e., changing environmental conditions at an appropriate timing for each event, may be a promising approach to obtain a high yield. Detailed descriptions of plant growth conditions and careful examination of environmental effects will significantly contribute to our knowledge to stably obtain high recombinant

  3. Micro-shock Wave Assisted Plant Transformation

    NASA Astrophysics Data System (ADS)

    Gnanadhas, Divya Prakash; Datey, Akshay; Chakravortty, Dipshikha; Gopalan, Jagadeesh

    Genetically modified (GM) crops are developed by transforming the desired DNA to plant. There are various methods employed to achieve the required transformation in plants. Agrobacterium mediated transformation and Biolistics or particle bombardment method are the most commonly used methods.

  4. Reliable transformation system for Microbotryum lychnidis-dioicae informed by genome and transcriptome project.

    PubMed

    Toh, Su San; Treves, David S; Barati, Michelle T; Perlin, Michael H

    2016-10-01

    Microbotryum lychnidis-dioicae is a member of a species complex infecting host plants in the Caryophyllaceae. It is used as a model system in many areas of research, but attempts to make this organism tractable for reverse genetic approaches have not been fruitful. Here, we exploited the recently obtained genome sequence and transcriptome analysis to inform our design of constructs for use in Agrobacterium-mediated transformation techniques currently available for other fungi. Reproducible transformation was demonstrated at the genomic, transcriptional and functional levels. Moreover, these initial proof-of-principle experiments provide evidence that supports the findings from initial global transcriptome analysis regarding expression from the respective promoters under different growth conditions of the fungus. The technique thus provides for the first time the ability to stably introduce transgenes and over-express target M. lychnidis-dioicae genes. PMID:27215216

  5. Improved genetic transformation of cork oak (Quercus suber L.).

    PubMed

    Alvarez-Fernández, Rubén; Ordás, Ricardo-Javier

    2012-01-01

    An Agrobacterium-mediated transformation system for selected mature Quercus suber L. trees has been established. Leaf-derived somatic embryos in an early stage of development are inoculated with an AGL1 strain harboring a kanamycin-selectable plasmid carrying the gene of interest. The transformed embryos are induced to germinate and the plantlets transferred to soil. This protocol, from adult cork oak to transformed plantlet, can be completed in about one and a half years. Transformation efficiencies (i.e., percentage of inoculated explants that yield independent transgenic embryogenic lines) vary depending on the cork oak genotype, reaching up to 43%.

  6. Exploration of new perspectives and limitations in Agrobacterium mediated gene transfer technology. Progress report, [June 1, 1992-- May 31, 1994

    SciTech Connect

    Marton, L.

    1994-12-31

    This report describes progress aimed at constructing gene-transfer technology for Nicotiana plumbaginifolia. Most actual effort as described herein has so far been directed at exploring new perspectives and limitations in Agrobacterium mediated gene transfer. Accomplishments are described using a core homologous gene targeting vector.

  7. Agrobacterium Mediated Transient Gene Silencing (AMTS) in Stevia rebaudiana: Insights into Steviol Glycoside Biosynthesis Pathway

    PubMed Central

    Guleria, Praveen; Yadav, Sudesh Kumar

    2013-01-01

    Background Steviol glycoside biosynthesis pathway has emerged as bifurcation from ent-kaurenoic acid, substrate of methyl erythritol phosphate pathway that also leads to gibberellin biosynthesis. However, the genetic regulation of steviol glycoside biosynthesis has not been studied. So, in present study RNA interference (RNAi) based Agrobacterium mediated transient gene silencing (AMTS) approach was followed. SrKA13H and three SrUGTs (SrUGT85C2, SrUGT74G1 and SrUGT76G1) genes encoding ent-kaurenoic acid-13 hydroxylase and three UDP glycosyltransferases of steviol glycoside biosynthesis pathway were silenced in Stevia rebaudiana to understand its molecular mechanism and association with gibberellins. Methodology/Principal Findings RNAi mediated AMTS of SrKA13H and three SrUGTs has significantly reduced the expression of targeted endogenous genes as well as total steviol glycoside accumulation. While gibberellins (GA3) content was significantly enhanced on AMTS of SrUGT85C2 and SrKA13H. Silencing of SrKA13H and SrUGT85C2 was found to block the metabolite flux of steviol glycoside pathway and shifted it towards GA3 biosynthesis. Further, molecular docking of three SrUGT proteins has documented highest affinity of SrUGT76G1 for the substrates of alternate pathways synthesizing steviol glycosides. This could be a plausible reason for maximum reduction in steviol glycoside content on silencing of SrUGT76G1 than other genes. Conclusions SrKA13H and SrUGT85C2 were identified as regulatory genes influencing carbon flux between steviol glycoside and gibberellin biosynthesis. This study has also documented the existence of alternate steviol glycoside biosynthesis route. PMID:24023961

  8. Barley Transformation Using Biolistic Techniques

    NASA Astrophysics Data System (ADS)

    Harwood, Wendy A.; Smedley, Mark A.

    Microprojectile bombardment or biolistic techniques have been widely used for cereal transformation. These methods rely on the acceleration of gold particles, coated with plasmid DNA, into plant cells as a method of directly introducing the DNA. The first report of the generation of fertile, transgenic barley plants used biolistic techniques. However, more recently Agrobacterium-mediated transformation has been adopted as the method of choice for most cereals including barley. Biolistic procedures are still important for some barley transformation applications and also provide transient test systems for the rapid checking of constructs. This chapter describes methods for the transformation of barley using biolistic procedures and also highlights the use of the technology in transient assays.

  9. Advances in Agrobacterium tumefaciens-mediated genetic transformation of graminaceous crops.

    PubMed

    Singh, Roshan Kumar; Prasad, Manoj

    2016-05-01

    Steady increase in global population poses several challenges to plant science research, including demand for increased crop productivity, grain yield, nutritional quality and improved tolerance to different environmental factors. Transgene-based approaches are promising to address these challenges by transferring potential candidate genes to host organisms through different strategies. Agrobacterium-mediated gene transfer is one such strategy which is well known for enabling efficient gene transfer in both monocot and dicots. Due to its versatility, this technique underwent several advancements including development of improved in vitro plant regeneration system, co-cultivation and selection methods, and use of hyper-virulent strains of Agrobacterium tumefaciens harbouring super-binary vectors. The efficiency of this method has also been enhanced by the use of acetosyringone to induce the activity of vir genes, silver nitrate to reduce the Agrobacterium-induced necrosis and cysteine to avoid callus browning during co-cultivation. In the last two decades, extensive efforts have been invested towards achieving efficient Agrobacterium-mediated transformation in cereals. Though high-efficiency transformation systems have been developed for rice and maize, comparatively lesser progress has been reported in other graminaceous crops. In this context, the present review discusses the progress made in Agrobacterium-mediated transformation system in rice, maize, wheat, barley, sorghum, sugarcane, Brachypodium, millets, bioenergy and forage and turf grasses. In addition, it also provides an overview of the genes that have been recently transferred to these graminaceous crops using Agrobacterium, bottlenecks in this technique and future possibilities for crop improvement. PMID:26660352

  10. Exploration of new perspectives and limitations in Agrobacterium-mediated gene transfer technology. Final report, June 1, 1992--May 31, 1995

    SciTech Connect

    Marton, L.

    1996-02-01

    Genetic manipulation of plants often involves the introduction of homologous or partly homologous genes. Ectropic introduction of homologous sequences into plant genomes may trigger epigenetic changes, making expression of the genes unpredictable. The main project objective was to examine the feasibility of using Agrobacterium-mediated gene transfer for homologous gene targeting in plants.

  11. Development of three full-length infectious cDNA clones of distinct brassica yellows virus genotypes for agrobacterium-mediated inoculation.

    PubMed

    Zhang, Xiao-Yan; Dong, Shu-Wei; Xiang, Hai-Ying; Chen, Xiang-Ru; Li, Da-Wei; Yu, Jia-Lin; Han, Cheng-Gui

    2015-02-01

    Brassica yellows virus is a newly identified species in the genus of Polerovirus within the family Luteoviridae. Brassica yellows virus (BrYV) is prevalently distributed throughout Mainland China and South Korea, is an important virus infecting cruciferous crops. Based on six BrYV genomic sequences of isolates from oilseed rape, rutabaga, radish, and cabbage, three genotypes, BrYV-A, BrYV-B, and BrYV-C, exist, which mainly differ in the 5' terminal half of the genome. BrYV is an aphid-transmitted and phloem-limited virus. The use of infectious cDNA clones is an alternative means of infecting plants that allows reverse genetic studies to be performed. In this study, full-length cDNA clones of BrYV-A, recombinant BrYV5B3A, and BrYV-C were constructed under control of the cauliflower mosaic virus 35S promoter. An agrobacterium-mediated inoculation system of Nicotiana benthamiana was developed using these cDNA clones. Three days after infiltration with full-length BrYV cDNA clones, necrotic symptoms were observed in the inoculated leaves of N. benthamiana; however, no obvious symptoms appeared in the upper leaves. Reverse transcription-PCR (RT-PCR) and western blot detection of samples from the upper leaves showed that the maximum infection efficiency of BrYVs could reach 100%. The infectivity of the BrYV-A, BrYV-5B3A, and BrYV-C cDNA clones was further confirmed by northern hybridization. The system developed here will be useful for further studies of BrYV, such as host range, pathogenicity, viral gene functions, and plant-virus-vector interactions, and especially for discerning the differences among the three genotypes. PMID:25499296

  12. Recent advances in soybean transformation and their application to molecular breeding and genomic analysis

    PubMed Central

    Yamada, Tetsuya; Takagi, Kyoko; Ishimoto, Masao

    2012-01-01

    Herbicide-resistant transgenic soybean plants hold a leading market share in the USA and other countries, but soybean has been regarded as recalcitrant to transformation for many years. The cumulative and, at times, exponential advances in genetic manipulation have made possible further choices for soybean transformation. The most widely and routinely used transformation systems are cotyledonary node–Agrobacterium-mediated transformation and somatic embryo–particle-bombardment-mediated transformation. These ready systems enable us to improve seed qualities and agronomic characteristics by transgenic approaches. In addition, with the accumulation of soybean genomic resources, convenient or promising approaches will be requisite for the determination and use of gene function in soybean. In this article, we describe recent advances in and problems of soybean transformation, and survey the current transgenic approaches for applied and basic research in Japan. PMID:23136488

  13. Transformation based endorsement systems

    NASA Technical Reports Server (NTRS)

    Sudkamp, Thomas

    1988-01-01

    Evidential reasoning techniques classically represent support for a hypothesis by a numeric value or an evidential interval. The combination of support is performed by an arithmetic rule which often requires restrictions to be placed on the set of possibilities. These assumptions usually require the hypotheses to be exhausitive and mutually exclusive. Endorsement based classification systems represent support for the alternatives symbolically rather than numerically. A framework for constructing endorsement systems is presented in which transformations are defined to generate and update the knowledge base. The interaction of the knowledge base and transformations produces a non-monotonic reasoning system. Two endorsement based reasoning systems are presented to demonstrate the flexibility of the transformational approach for reasoning with ambiguous and inconsistent information.

  14. Development of a transgenic hairy root system in jute (Corchorus capsularis L.) with gusA reporter gene through Agrobacterium rhizogenes mediated co-transformation.

    PubMed

    Chattopadhyay, Tirthartha; Roy, Sheuli; Mitra, Adinpunya; Maiti, Mrinal K

    2011-04-01

    Transgenic hairy root system is important in several recalcitrant plants, where Agrobacterium tumefaciens-mediated plant transformation and generation of transgenic plants are problematic. Jute (Corchorus spp.), the major fibre crop in Indian subcontinent, is one of those recalcitrant plants where in vitro tissue culture has provided a little success, and hence, Agrobacterium-mediated genetic transformation remains to be a challenging proposition in this crop. In the present work, a system of transgenic hairy roots in Corchorus capsularis L. has been developed through genetic transformation by Agrobacterium rhizogenes harbouring two plasmids, i.e. the natural Ri plasmid and a recombinant binary vector derived from the disarmed Ti plasmid of A. tumefaciens. Our findings indicate that the system is relatively easy to establish and reproducible. Molecular analysis of the independent lines of transgenic hairy roots revealed the transfer of relevant transgenes from both the T-DNA parts into the plant genome, indicating the co-transformation nature of the event. High level expression and activity of the gusA reporter gene advocate that the transgenic hairy root system, thus developed, could be applicable as gene expression system in general and for root functional genomics in particular. Furthermore, these transgenic hairy roots can be used in future as explants for plantlet regeneration to obtain stable transgenic jute plants.

  15. Mapping the subgenomic RNA promoter of the Citrus leaf blotch virus coat protein gene by Agrobacterium-mediated inoculation.

    PubMed

    Renovell, Agueda; Gago, Selma; Ruiz-Ruiz, Susana; Velázquez, Karelia; Navarro, Luis; Moreno, Pedro; Vives, Mari Carmen; Guerri, José

    2010-10-25

    Citrus leaf blotch virus has a single-stranded positive-sense genomic RNA (gRNA) of 8747 nt organized in three open reading frames (ORFs). The ORF1, encoding a polyprotein involved in replication, is translated directly from the gRNA, whereas ORFs encoding the movement (MP) and coat (CP) proteins are expressed via 3' coterminal subgenomic RNAs (sgRNAs). We characterized the minimal promoter region critical for the CP-sgRNA expression in infected cells by deletion analyses using Agrobacterium-mediated infection of Nicotiana benthamiana plants. The minimal CP-sgRNA promoter was mapped between nucleotides -67 and +50 nt around the transcription start site. Surprisingly, larger deletions in the region between the CP-sgRNA transcription start site and the CP translation initiation codon resulted in increased CP-sgRNA accumulation, suggesting that this sequence could modulate the CP-sgRNA transcription. Site-specific mutational analysis of the transcription start site revealed that the +1 guanylate and the +2 adenylate are important for CP-sgRNA synthesis.

  16. Genetic transformation of Brassica nigra by agrobacterium based vector and direct plasmid uptake.

    PubMed

    Gupta, V; Lakshmi Sita, G; Shaila, M S; Jagannathan, V

    1993-05-01

    Genetic transformation systems have been established for Brassica nigra (cv. IC 257) by using an Agrobacterium binary vector as well as by direct DNA uptake of a plasmid vector. Both the type of vectors carried nptII gene and gus gene. For Agrobacterium mediated transformation, hypocotyl tissue explants were used, and up to 33% of the explants produced calli on selection medium. All of these expressed B-glucuronidase gene on histochemical staining. Protoplasts isolated from hypocotyl tissues of seedlings could be transformed with a plasmid vector by FEG mediated uptake of vector DNA. A number of fertile kanamycin resistant plants were obtained using both the methods, and their transformed nature was confirmed by Southern blot analysis and histochemical staining for GUS. Backcrossed and selfed progenies of these transformed plants showed the presence of npt and gus genes. PMID:24197344

  17. Fourier Transform Spectrometer System

    NASA Technical Reports Server (NTRS)

    Campbell, Joel F. (Inventor)

    2014-01-01

    A Fourier transform spectrometer (FTS) data acquisition system includes an FTS spectrometer that receives a spectral signal and a laser signal. The system further includes a wideband detector, which is in communication with the FTS spectrometer and receives the spectral signal and laser signal from the FTS spectrometer. The wideband detector produces a composite signal comprising the laser signal and the spectral signal. The system further comprises a converter in communication with the wideband detector to receive and digitize the composite signal. The system further includes a signal processing unit that receives the composite signal from the converter. The signal processing unit further filters the laser signal and the spectral signal from the composite signal and demodulates the laser signal, to produce velocity corrected spectral data.

  18. Transformation systems in insects.

    PubMed

    O'Brochta, David A; Atkinson, Peter W

    2004-01-01

    Genetic transformation is an important technology that provides unique opportunities to find, isolate, and analyze genes, as well as to create organisms with unique functional characteristics. Insect biologists have been developing genetic transformation technologies that rely extensively on transposable elements. A number of class II transposable elements isolated originally from insects have been converted into broad host range insect gene vectors. Class II transposable elements are particularly amenable to gene vector development, although they suffer from some limitations such as low rates of recombination. Use of these gene vectors requires the physical introduction of the vectors into developing insect embryos by microinjection. Microinjection methods vary to accommodate the unique physical and developmental characteristics of the target insects. All methods rely on the use of fine glass needles in conjunction with micromanipulators and a microscope. A serious constraint on the use of existing systems can be the inefficiency of successfully delivering the gene vectors to the germ cells of the developing embryo. The general method for vector delivery to insect germ cells is described, as well as variations that are useful under some conditions.

  19. Im"plant"ing of Mammalian Glycosyltransferase Gene into Plant Suspension-Cultured Cells Using Agrobacterium-Mediated Transformation.

    PubMed

    Kajiura, Hiroyuki; Fujiyama, Kazuhito

    2015-01-01

    Enzymatic activity assay of exogenous glycosyltransferase (GT) and glycosylhydrolase (GH) expressed in plants is an important analysis for determination of the expression of the gene of interest. However, generations and establishment of in planta transgenic lines are time-consuming. Furthermore, the expression levels and the activities of the exogenous GTs and GHs are quite low and weak, the radiolabeled donor substrate had to be used to analyze the enzymatic activity. Here, we describe a protocol for the generation of transgenic plants using suspension-cultured cells and a high sensitive assay for GT, especially β1,4-galactosyltransferase, using microsomal fraction from plant cells and fluorescent-labeled sugar chains as an acceptor substrate. This method enables less-time-consuming preparation of stable transgenic plants, non-radiolabeled, high-throughput detail analysis which includes mass spectrometric analysis and exo-glycosidase digestions.

  20. Use of Agrobacterium rhizogenes strain 18r12v and paromomycin selection for transformation of Brachypodium distachyon and Brachypodium sylvaticum

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The genetic transformation of monocot grasses is a resource intensive process, the quality and efficiency of which is dependent in part upon the method of DNA introduction, as well as the ability to effectively separate transformed from wildtype tissue. Agrobacterium-mediated transformation of Brac...

  1. Transformation Systems at NASA Ames

    NASA Technical Reports Server (NTRS)

    Buntine, Wray; Fischer, Bernd; Havelund, Klaus; Lowry, Michael; Pressburger, TOm; Roach, Steve; Robinson, Peter; VanBaalen, Jeffrey

    1999-01-01

    In this paper, we describe the experiences of the Automated Software Engineering Group at the NASA Ames Research Center in the development and application of three different transformation systems. The systems span the entire technology range, from deductive synthesis, to logic-based transformation, to almost compiler-like source-to-source transformation. These systems also span a range of NASA applications, including solving solar system geometry problems, generating data analysis software, and analyzing multi-threaded Java code.

  2. Agrobacterium-mediated inoculation of chrysanthemum (Chrysanthemum morifolium) plants with chrysanthemum stunt viroid.

    PubMed

    Nabeshima, Tomoyuki; Doi, Motoaki; Hosokawa, Munetaka

    2016-08-01

    Agroinfiltration was tested as a method of inoculation of chrysanthemum plants with chrysanthemum stunt viroid (CSVd). Binary vectors harboring dimeric CSVd sequences in sense and antisense orientations were constructed, and Agrobacterium transfected with these binary vectors was infiltrated into chrysanthemum leaves. Northern blotting and reverse transcription polymerase chain reaction analysis showed that local infection was established within 7 days and systemic infection within 20 days. CSVd polarities showed no difference in infectivity. This study showed that agroinfiltration of chrysanthemum plants is an easy, rapid, and cost-effective method for CSVd inoculation. PMID:27155239

  3. Agrobacterium-mediated infection of whole plants by yellow dwarf viruses.

    PubMed

    Yoon, Ju-Yeon; Choi, Seung-Kook; Palukaitis, Peter; Gray, Stewart M

    2011-09-01

    Barley yellow dwarf virus-PAV (BYDV-PAV) and cereal yellow dwarf virus-RPV (CYDV-RPV) are only transmitted between host plants by aphid vectors and not by mechanical transmission. This presents a severe limitation for the use of a reverse genetics approach to analyze the effects of mutations in these viruses on plant infection and aphid transmission. Here we describe the use of agroinfection to infect plants with BYDV-PAV and CYDV-RPV. The cDNAs corresponding to the complete RNA genomes of BYDV-PAV and CYDV-RPV were cloned into a binary vector under the control of the cauliflower mosaic virus 35S promoter and the nopaline synthase transcription termination signal. The self-cleaving ribozyme from hepatitis virus D was included to produce a transcript in planta with a 3' terminus identical to the natural viral RNA. ELISA and RT-PCR analysis showed that the replicons of BYDV-PAV and CYDV-RPV introduced by Agrobacterium into Nicotiana benthamiana and N. clevelandii gave rise to a local infection in the infiltrated mesophyll cells. After several weeks systemic infection of phloem tissue was detected, although no systemic symptoms were observed. Three heterologous virus silencing suppressors increased the efficiency of agroinfection and accumulation of BYDV-PAV and CYDV-RPV in the two Nicotiana species. The progeny viruses purified from infiltrated tissues were successfully transmitted to oat plants by aphids, and typical yellow dwarf symptoms were observed. This study reports the first agroinfection of eudicot plants using BYDV-PAV and CYDV-RPV. PMID:21763366

  4. High transformation efficiency in plum (Prunus domestica L.): a new tool for functional genomics studies in Prunus spp.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    An improved Agrobacterium-mediated transformation protocol in plum (Prunus domestica L.) cv 'Bluebyrd' using hypocotyl slices as source of explants is described. The addition of 2, 4-D to the regeneration media during co-culture allowed us to increase transformation efficiency up to 10 times over p...

  5. Genetic Transformation of Switchgrass

    NASA Astrophysics Data System (ADS)

    Xi, Yajun; Ge, Yaxin; Wang, Zeng-Yu

    Switchgrass (Panicum virgatum L.) is a highly productive warm-season C4 species that is being developed into a dedicated biofuel crop. This chapter describes a protocol that allows the generation of transgenic switchgrass plants by Agrobacterium tumefaciens-mediated transformation. Embryogenic calluses induced from caryopses or inflorescences were used as explants for inoculation with A. tumefaciens strain EHA105. Hygromycin phosphotransferase gene (hph) was used as the selectable marker and hygromycin was used as the selection agent. Calluses resistant to hygromycin were obtained after 5-6 weeks of selection. Soil-grown switchgrass plants were regenerated about 6 months after callus induction and Agrobacterium-mediated transformation.

  6. Energy systems transformation.

    PubMed

    Dangerman, A T C Jérôme; Schellnhuber, Hans Joachim

    2013-02-12

    The contemporary industrial metabolism is not sustainable. Critical problems arise at both the input and the output side of the complex: Although affordable fossil fuels and mineral resources are declining, the waste products of the current production and consumption schemes (especially CO(2) emissions, particulate air pollution, and radioactive residua) cause increasing environmental and social costs. Most challenges are associated with the incumbent energy economy that is unlikely to subsist. However, the crucial question is whether a swift transition to its sustainable alternative, based on renewable sources, can be achieved. The answer requires a deep analysis of the structural conditions responsible for the rigidity of the fossil-nuclear energy system. We argue that the resilience of the fossil-nuclear energy system results mainly from a dynamic lock-in pattern known in operations research as the "Success to the Successful" mode. The present way of generating, distributing, and consuming energy--the largest business on Earth--expands through a combination of factors such as the longevity of pertinent infrastructure, the information technology revolution, the growth of the global population, and even the recent financial crises: Renewable-energy industries evidently suffer more than the conventional-energy industries under recession conditions. Our study tries to elucidate the archetypical traits of the lock-in pattern and to assess the respective importance of the factors involved. In particular, we identify modern corporate law as a crucial system element that thus far has been largely ignored. Our analysis indicates that the rigidity of the existing energy economy would be reduced considerably by the assignment of unlimited liabilities to the shareholders. PMID:23297208

  7. Energy systems transformation.

    PubMed

    Dangerman, A T C Jérôme; Schellnhuber, Hans Joachim

    2013-02-12

    The contemporary industrial metabolism is not sustainable. Critical problems arise at both the input and the output side of the complex: Although affordable fossil fuels and mineral resources are declining, the waste products of the current production and consumption schemes (especially CO(2) emissions, particulate air pollution, and radioactive residua) cause increasing environmental and social costs. Most challenges are associated with the incumbent energy economy that is unlikely to subsist. However, the crucial question is whether a swift transition to its sustainable alternative, based on renewable sources, can be achieved. The answer requires a deep analysis of the structural conditions responsible for the rigidity of the fossil-nuclear energy system. We argue that the resilience of the fossil-nuclear energy system results mainly from a dynamic lock-in pattern known in operations research as the "Success to the Successful" mode. The present way of generating, distributing, and consuming energy--the largest business on Earth--expands through a combination of factors such as the longevity of pertinent infrastructure, the information technology revolution, the growth of the global population, and even the recent financial crises: Renewable-energy industries evidently suffer more than the conventional-energy industries under recession conditions. Our study tries to elucidate the archetypical traits of the lock-in pattern and to assess the respective importance of the factors involved. In particular, we identify modern corporate law as a crucial system element that thus far has been largely ignored. Our analysis indicates that the rigidity of the existing energy economy would be reduced considerably by the assignment of unlimited liabilities to the shareholders.

  8. Energy systems transformation

    PubMed Central

    Dangerman, A. T. C. Jérôme; Schellnhuber, Hans Joachim

    2013-01-01

    The contemporary industrial metabolism is not sustainable. Critical problems arise at both the input and the output side of the complex: Although affordable fossil fuels and mineral resources are declining, the waste products of the current production and consumption schemes (especially CO2 emissions, particulate air pollution, and radioactive residua) cause increasing environmental and social costs. Most challenges are associated with the incumbent energy economy that is unlikely to subsist. However, the crucial question is whether a swift transition to its sustainable alternative, based on renewable sources, can be achieved. The answer requires a deep analysis of the structural conditions responsible for the rigidity of the fossil-nuclear energy system. We argue that the resilience of the fossil-nuclear energy system results mainly from a dynamic lock-in pattern known in operations research as the “Success to the Successful” mode. The present way of generating, distributing, and consuming energy—the largest business on Earth—expands through a combination of factors such as the longevity of pertinent infrastructure, the information technology revolution, the growth of the global population, and even the recent financial crises: Renewable-energy industries evidently suffer more than the conventional-energy industries under recession conditions. Our study tries to elucidate the archetypical traits of the lock-in pattern and to assess the respective importance of the factors involved. In particular, we identify modern corporate law as a crucial system element that thus far has been largely ignored. Our analysis indicates that the rigidity of the existing energy economy would be reduced considerably by the assignment of unlimited liabilities to the shareholders. PMID:23297208

  9. Set Equation Transformation System.

    2002-03-22

    Version 00 SETS is used for symbolic manipulation of Boolean equations, particularly the reduction of equations by the application of Boolean identities. It is a flexible and efficient tool for performing probabilistic risk analysis (PRA), vital area analysis, and common cause analysis. The equation manipulation capabilities of SETS can also be used to analyze noncoherent fault trees and determine prime implicants of Boolean functions, to verify circuit design implementation, to determine minimum cost fire protectionmore » requirements for nuclear reactor plants, to obtain solutions to combinatorial optimization problems with Boolean constraints, and to determine the susceptibility of a facility to unauthorized access through nullification of sensors in its protection system. Two auxiliary programs, SEP and FTD, are included. SEP performs the quantitative analysis of reduced Boolean equations (minimal cut sets) produced by SETS. The user can manipulate and evaluate the equations to find the probability of occurrence of any desired event and to produce an importance ranking of the terms and events in an equation. FTD is a fault tree drawing program which uses the proprietary ISSCO DISSPLA graphics software to produce an annotated drawing of a fault tree processed by SETS. The DISSPLA routines are not included.« less

  10. Maize transformation technology development for commercial event generation

    PubMed Central

    Que, Qiudeng; Elumalai, Sivamani; Li, Xianggan; Zhong, Heng; Nalapalli, Samson; Schweiner, Michael; Fei, Xiaoyin; Nuccio, Michael; Kelliher, Timothy; Gu, Weining; Chen, Zhongying; Chilton, Mary-Dell M.

    2014-01-01

    Maize is an important food and feed crop in many countries. It is also one of the most important target crops for the application of biotechnology. Currently, there are more biotech traits available on the market in maize than in any other crop. Generation of transgenic events is a crucial step in the development of biotech traits. For commercial applications, a high throughput transformation system producing a large number of high quality events in an elite genetic background is highly desirable. There has been tremendous progress in Agrobacterium-mediated maize transformation since the publication of the Ishida et al. (1996) paper and the technology has been widely adopted for transgenic event production by many labs around the world. We will review general efforts in establishing efficient maize transformation technologies useful for transgenic event production in trait research and development. The review will also discuss transformation systems used for generating commercial maize trait events currently on the market. As the number of traits is increasing steadily and two or more modes of action are used to control key pests, new tools are needed to efficiently transform vectors containing multiple trait genes. We will review general guidelines for assembling binary vectors for commercial transformation. Approaches to increase transformation efficiency and gene expression of large gene stack vectors will be discussed. Finally, recent studies of targeted genome modification and transgene insertion using different site-directed nuclease technologies will be reviewed. PMID:25140170

  11. Efficient transformation and artificial miRNA gene silencing in Lemna minor

    PubMed Central

    Cantó-Pastor, Alex; Mollá-Morales, Almudena; Ernst, Evan; Dahl, William; Zhai, Jixian; Yan, Yiheng; Meyers, Blake; Shanklin, John; Martienssen, Robert

    2015-01-01

    Lack of genetic tools in the Lemnaceae (duckweed) has impeded full implementation of this organism as model for biological research, despite its rapid doubling time, simple architecture and unusual metabolic characteristics. Here we present technologies to facilitate high-throughput genetic studies in duckweed. We developed a fast and efficient method for producing Lemna minor stable transgenic fronds via agrobacterium-mediated transformation and regeneration from tissue culture. Additionally, we engineered an artificial microRNA (amiRNA) gene silencing system. We identified a Lemna gibba endogenous miR166 precursor and used it as a backbone to produce amiRNAs. As a proof of concept we induced the silencing of CH42, a Magnesium Chelatase subunit, using our amiRNA platform. Expression of CH42 in transgenic Lemna minor fronds was significantly reduced, which resulted in reduction of chlorophyll pigmentation. The techniques presented here will enable tackling future challenges in the biology and biotechnology of Lemnaceae. PMID:24989135

  12. Efficient genetic transformation of red raspberry, Rubus ideaus L.

    PubMed

    Mathews, H; Wagoner, W; Cohen, C; Kellogg, J; Bestwick, R

    1995-05-01

    We have developed an efficient transformation system for red raspberry (Rubus ideaus L.) using Agrobacterium mediated gene transfer. Using this system we have successfully introduced a gene that encodes an enzyme, S-adenosylmethionine hydrolase (SAMase), in raspberry cultivars Meeker (MK), Chilliwack (CH) and Canby (CY). Leaf and petiole expiants were inoculated with disarmed Agrobacterium tumefaciens strain EHA 105 carrying either of two binary vectors, pAG1452 or pAG1552, encoding gene sequences for SAMase under the control of the wound and fruit specific tomato E4 promoter. Primary shoot regenerants on selection medium were chimeral containing both transformed and non-transformed cells. Non-chimeral transgenic clones were developed by iterative culture of petiole, node and leaf explants, on selection medium, from successive generations of shoots derived from the primary regenerants. Percent recovery of transformants was higher with the selection marker gene hygromycin phosphotransferase (hpt), than with neomycin phosphotransferase (nptII). Transformation frequencies of 49.6%, 0.9% and 8.1% were obtained in cultivars Meeker, Chilliwack and Canby respectively from petiole expiants using hygromycin selection. Genomic integration of transgenes was confirmed by Southern hybridization. Transgenic plants from a total of 218 independent transformation events (161 MK, 4 CH, 53 CY) have been successfully established in soil.

  13. Development of Agrobacterium-Mediated Virus-Induced Gene Silencing and Performance Evaluation of Four Marker Genes in Gossypium barbadense

    PubMed Central

    Pang, Jinhuan; Zhu, Yue; Li, Qing; Liu, Jinzhi; Tian, Yingchuan; Liu, Yule; Wu, Jiahe

    2013-01-01

    Gossypiumbarbadense is a cultivated cotton species and possesses many desirable traits, including high fiber quality and resistance to pathogens, especially Verticilliumdahliae (a devastating pathogen of Gossypium hirsutum, the main cultivated species). These elite traits are difficult to be introduced into G. hirsutum through classical breeding methods. In addition, genetic transformation of G. barbadense has not been successfully performed. It is therefore important to develop methods for evaluating the function and molecular mechanism of genes in G. barbadense. In this study, we had successfully introduced a virus-induced gene silencing (VIGS) system into three cultivars of G. barbadense by inserting marker genes into the tobacco rattle virus (TRV) vector. After we optimized the VIGS conditions, including light intensity, photoperiod, seedling age and Agrobacterium strain, 100% of plants agroinfiltrated with the GaPDS silencing vector showed white colored leaves. Three other marker genes, GaCLA1, GaANS and GaANR, were employed to further test this VIGS system in G. barbadense. The transcript levels of the endogenous genes in the silenced plants were reduced by more than 99% compared to control plants; these plants presented phenotypic symptoms 2 weeks after inoculation. We introduced a fusing sequence fragment of GaPDS and GaANR gene silencing vectors into a single plant, which resulted in both photobleaching and brownish coloration. The extent of silencing in plants agroinfiltrated with fusing two-gene-silencing vector was consistent with plants harboring a single gene silencing vector. The development of this VIGS system should promote analysis of gene function in G. barbadense, and help to contribute desirable traits for breeding of G. barbadense and G. hirsutum. PMID:24023833

  14. A versatile binary vector system with a T-DNA organisational structure conducive to efficient integration of cloned DNA into the plant genome.

    PubMed

    Gleave, A P

    1992-12-01

    A versatile gene expression cartridge and binary vector system was constructed for use in Agrobacterium-mediated plant transformation. The expression cartridge of the primary cloning vector, pART7, comprises of cauliflower mosaic virus Cabb B-JI isolate 35S promoter, a multiple cloning site and the transcriptional termination region of the octopine synthase gene. The entire cartridge can be removed from pART7 as a Not I fragment and introduced directly into the binary vector, pART27, recombinants being selected by blue/white screening for beta-galactosidase. pART27 carries the RK2 minimal replicon for maintenance in Agrobacterium, the ColE1 origin of replication for high-copy maintenance in Escherichia coli and the Tn7 spectinomycin/streptomycin resistance gene as a bacterial selectable marker. The organisational structure of the T-DNA of pART27 has been constructed taking into account the right to left border, 5' to 3' model of T-DNA transfer. The T-DNA carries the chimaeric kanamycin resistance gene (nopaline synthase promoter-neomycin phosphotransferase-nopaline synthase terminator) distal to the right border relative to the lacZ' region. Utilisation of these vectors in Agrobacterium-mediated transformation of tobacco demonstrated efficient T-DNA transfer to the plant genome.

  15. Agrobacterium-Mediated Gene Transfer Results Mainly in Transgenic Plants Transmitting T-DNA as a Single Mendelian Factor

    PubMed Central

    Budar, F.; Thia-Toong, L.; Van Montagu, M.; Hernalsteens, J.-P.

    1986-01-01

    Forty-four independent transformed tobacco plants were obtained from a cocultivation experiment with Agrobacterium tumefaciens strains carrying modified Ti-plasmids. The transformed plants were either self-fertilized or crossed with nontransformed plants or with other transformed plants. The segregation of a phenotypic marker (kanamycin resistance) in the progenies of these plants was determined. In 40 cases out of 44, the segregation of the kanamycin resistance marker is consistent with Mendelian genetics. Among these 40 clones, 35 contain a single kanamycin resistance locus. The five others segregate two independent resistance loci. In two of the single insert clones, the segregation ratio after selfing indicates that the T-DNA insertion may have caused a recessive lethal mutation. PMID:17246346

  16. A set of modular binary vectors for transformation of cereals.

    PubMed

    Himmelbach, Axel; Zierold, Uwe; Hensel, Götz; Riechen, Jan; Douchkov, Dimitar; Schweizer, Patrick; Kumlehn, Jochen

    2007-12-01

    Genetic transformation of crop plants offers the possibility of testing hypotheses about the function of individual genes as well as the exploitation of transgenes for targeted trait improvement. However, in most cereals, this option has long been compromised by tedious and low-efficiency transformation protocols, as well as by the lack of versatile vector systems. After having adopted and further improved the protocols for Agrobacterium-mediated stable transformation of barley (Hordeum vulgare) and wheat (Triticum aestivum), we now present a versatile set of binary vectors for transgene overexpression, as well as for gene silencing by double-stranded RNA interference. The vector set is offered with a series of functionally validated promoters and allows for rapid integration of the desired genes or gene fragments by GATEWAY-based recombination. Additional in-built flexibility lies in the choice of plant selectable markers, cassette orientation, and simple integration of further promoters to drive specific expression of genes of interest. Functionality of the cereal vector set has been demonstrated by transient as well as stable transformation experiments for transgene overexpression, as well as for targeted gene silencing in barley.

  17. (Analysis of proteins essential for Agrobacterium mediated DNA transfer to plant cells). [Single-stranded DNA binding proteins

    SciTech Connect

    Not Available

    1989-12-14

    The tumor inducing (Ti) plasmid of Agrobacterium contains two regions important for infection and transformation of plant cells. One region, the T-DNA, is transferred as a single strand into the plant cell, while the virulence (vir) region is responsible for recognition of susceptible cells, synthesis of the T-DNA single strand (T-strand), formation of a T-strand protein complex and transfer of this complex into susceptible cells. A DNA binding protein, VirE2, was identified as a product of the vir region. Sequencing of the 9000 kilobase pair virB region has been completed. Expression of 10 of the predicted 11 open reading frames (ORFs) was demonstrated in Escherichia coli. Translational coupling was demonstrated for 5 ORFs. Hydropathy analysis indicates that 9 of 11 ORFs have hydrophobic regions that could permit membrane channel formation. In related work, analysis of protons that potentiate movement of plant viruses was discussed, with indications that the tobacco mosaic virus (TMV) protein P30 may mediate transfer of TMV RNA through plasmadesmata. Also, using the T-DNA element as a marker, genes responsible for abnormal flower development are being cloned and isolated. 3 refs. (MHB)

  18. Investigating plasmodesmata genetics with virus-induced gene silencing and an agrobacterium-mediated GFP movement assay.

    PubMed

    Brunkard, Jacob O; Burch-Smith, Tessa M; Runkel, Anne M; Zambryski, Patricia

    2015-01-01

    Plasmodesmata (PD) are channels that connect the cytoplasm of adjacent plant cells, permitting intercellular transport and communication. PD function and formation are essential to plant growth and development, but we still know very little about the genetic pathways regulating PD transport. Here, we present a method for assaying changes in the rate of PD transport following genetic manipulation. Gene expression in leaves is modified by virus-induced gene silencing. Seven to ten days after infection with Tobacco rattle virus carrying a silencing trigger, the gene(s) of interest is silenced in newly arising leaves. In these new leaves, individual cells are then transformed with Agrobacterium to express GFP, and the rate of GFP diffusion via PD is measured. By measuring GFP diffusion both within the epidermis and between the epidermis and mesophyll, the assay can be used to study the effects of silencing a gene(s) on PD transport in general, or transport through secondary PD specifically. Plant biologists working in several fields will find this assay useful, since PD transport impacts plant physiology, development, and defense.

  19. Targeted Mutagenesis in Rice Using TALENs and the CRISPR/Cas9 System.

    PubMed

    Endo, Masaki; Nishizawa-Yokoi, Ayako; Toki, Seiichi

    2016-01-01

    Sequence-specific nucleases (SSNs), such as zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the clustered regularly interspersed short palindromic repeats (CRISPR)/CRISPR-associated protein 9 nuclease (Cas9) system, are powerful tools for understanding gene function and for developing novel traits in plants. In plant species for which transformation and regeneration systems using protoplasts are not yet established, direct delivery to nuclei of SSNs either in the form of RNA or protein is difficult. Thus, Agrobacterium-mediated transformation of SSN expression constructs in cultured cells is a practical means of delivering targeted mutagenesis in some plant species including rice. Because targeted mutagenesis occurs stochastically in transgenic cells and SSN-mediated targeted mutagenesis often leads to no selectable phenotype, identification of highly mutated cell lines is a critical step in obtaining regenerated plants with desired mutations. PMID:27557690

  20. Efficient sweet pepper transformation mediated by the BABY BOOM transcription factor.

    PubMed

    Heidmann, Iris; de Lange, Brenda; Lambalk, Joep; Angenent, Gerco C; Boutilier, Kim

    2011-06-01

    Pepper (Capsicum L.) is a nutritionally and economically important crop that is cultivated throughout the world as a vegetable, condiment, and food additive. Genetic transformation using Agrobacterium tumefaciens (agrobacterium) is a powerful biotechnology tool that could be used in pepper to develop community-based functional genomics resources and to introduce important agronomic traits. However, pepper is considered to be highly recalcitrant for agrobacterium-mediated transformation, and current transformation protocols are either inefficient, cumbersome or highly genotype dependent. The main bottleneck in pepper transformation is the inability to generate cells that are competent for both regeneration and transformation. Here, we report that ectopic expression of the Brassica napus BABY BOOM AP2/ERF transcription factor overcomes this bottleneck and can be used to efficiently regenerate transgenic plants from otherwise recalcitrant sweet pepper (C. annuum) varieties. Transient activation of BABY BOOM in the progeny plants induced prolific cell regeneration and was used to produce a large number of somatic embryos that could be converted readily to seedlings. The data highlight the utility of combining biotechnology and classical plant tissue culture approaches to develop an efficient transformation and regeneration system for a highly recalcitrant vegetable crop. PMID:21305301

  1. Recent advances in plant transformation.

    PubMed

    Barampuram, Shyamkumar; Zhang, Zhanyuan J

    2011-01-01

    Plant genetic engineering has become one of the most important molecular tools in the modern molecular breeding of crops. Over the last decade, significant progress has been made in the development of new and efficient transformation methods in plants. Despite a variety of available DNA delivery methods, Agrobacterium- and biolistic-mediated transformation remain the two predominantly employed approaches. In particular, progress in Agrobacterium-mediated transformation of cereals and other recalcitrant dicot species has been quite remarkable. In the meantime, other transgenic-enabling technologies have emerged, including generation of marker-free transgenics, gene targeting, and chromosomal engineering. Although transformation of some plant species or elite germplasm remains a challenge, further advancement in transformation technology is expected because the mechanisms of governing the regeneration and transformation processes are now better understood and are being creatively applied to designing improved transformation methods or to developing new enabling technologies. PMID:21181522

  2. The School System Transformation (SST) Protocol

    ERIC Educational Resources Information Center

    Duffy, Francis M.; Reigeluth, Charles M., Ed.

    2008-01-01

    This is the second in a series of articles about creating and sustaining systemic transformational change in school districts. The first article described the "AECT FutureMinds: Transforming America's School Systems" initiative for helping state education agencies (SEAs) facilitate paradigm change in their school districts. This article describes…

  3. Complementary DNA cloning of the pear 1-aminocyclopropane-1-carboxylic acid oxidase gene and agrobacterium-mediated anti-sense genetic transformation.

    PubMed

    Qi, Jing; Dong, Zhen; Zhang, Yu-Xing

    2015-12-01

    The aim of the present study was to genetically modify plantlets of the Chinese yali pear to reduce their expression of ripening-associated 1-aminocyclopropane-1-carboxylic acid oxidase (ACO) and therefore increase the shelf-life of the fruit. Primers were designed with selectivity for the conserved regions of published ACO gene sequences, and yali complementary DNA (cDNA) cloning was performed by reverse transcription quantitative polymerase chain reaction (PCR). The obtained cDNA fragment contained 831 base pairs, encoding 276 amino acid residues, and shared no less than 94% nucleotide sequence identity with other published ACO genes. The cDNA fragment was inversely inserted into a pBI121 expression vector, between the cauliflower mosaic virus 35S promoter and the nopaline synthase terminator, in order to construct the anti‑sense expression vector of the ACO gene; it was transfected into cultured yali plants using Agrobacterium LBA4404. Four independent transgenic lines of pear plantlets were obtained and validated by PCR analysis. A Southern blot assay revealed that there were three transgenic lines containing a single copy of exogenous gene and one line with double copies. The present study provided germplasm resources for the cultivation of novel storage varieties of pears, therefore providing a reference for further applications of anti‑sense RNA technology in the genetic improvement of pears and other fruit.

  4. Transformation of radish (Raphanus sativus L.) via sonication and vacuum infiltration of germinated seeds with Agrobacterium harboring a group 3 LEA gene from B. napus.

    PubMed

    Park, Byong-Jin; Liu, Zaochang; Kanno, Akira; Kameya, Toshiaki

    2005-10-01

    A protocol for producing transgenic radish (Raphanus sativus) was obtained by using both ultrasonic and vacuum infiltration assisted, Agrobacterium-mediated transformation. The Agrobacterium strain LBA4404 contained the binary vector pBI121-LEA (late embyogenesis abundant), which carried a Group 3 LEA gene, from Brassica napus. Among six combinations, Agrobacterium-mediated transformation assisted by a combination of 5-min sonication with 5-min vacuum infiltration resulted in the highest transformation frequency. The existence, integration and expression of transferred LEA gene in transgenic T(1) plants were confirmed by PCR, genomic Southern and Western blot analysis. Transgenic radish demonstrated better growth performance than non-transformed control plants under osmotic and salt stress conditions. Accumulation of Group 3 LEA protein in the vegetative tissue of transgenic radish conferred increased tolerance to water deficit and salt stress.

  5. The non-homologous end-joining pathway is involved in stable transformation in rice

    PubMed Central

    Saika, Hiroaki; Nishizawa-Yokoi, Ayako; Toki, Seiichi

    2014-01-01

    Stable transformation with T-DNA needs the coordinated activities of many proteins derived from both host plant cells and Agrobacterium. In dicot plants, including Arabidopsis, it has been suggested that non-homologous end-joining (NHEJ)—one of the main DNA double-strand break repair pathways—is involved in the T-DNA integration step that is crucial to stable transformation. However, how this pathway is involved remains unclear as results with NHEJ mutants in Arabidopsis have given inconsistent results. Recently, a system for visualization of stable expression of genes located on T-DNA has been established in rice callus. Stable expression was shown to be reduced significantly in NHEJ knock-down rice calli, suggesting strongly that NHEJ is involved in Agrobacterium-mediated stable transformation in rice. Since rice transformation is now efficient and reproducible, rice is a good model plant in which to elucidate the molecular mechanisms of T-DNA integration. PMID:25368624

  6. Canonical transformations and Hamiltonian evolutionary systems

    SciTech Connect

    Al-Ashhab, Samer

    2012-06-15

    In many Lagrangian field theories, one has a Poisson bracket defined on the space of local functionals. We find necessary and sufficient conditions for a transformation on the space of local functionals to be canonical in three different cases. These three cases depend on the specific dimensions of the vector bundle of the theory and the associated Hamiltonian differential operator. We also show how a canonical transformation transforms a Hamiltonian evolutionary system and its conservation laws. Finally, we illustrate these ideas with three examples.

  7. [Economics of health system transformation].

    PubMed

    González Pier, Eduardo

    2012-01-01

    Health conditions in Mexico have evolved along with socioeconomic conditions. As a result, today's health system faces several problems characterized by four overlapping transitions: demand, expectations, funding and health resources. These transitions engender significant pressures on the system itself. Additionally, fragmentation of the health system creates disparities in access to services and generates problems in terms of efficiency and use of available resources. To address these complications and to improve equity in access and efficiency, thorough analysis is required in how the right to access health care should be established at a constitutional level without differentiating across population groups. This should be followed by careful discussion about what rules of health care financing should exist, which set of interventions ought to be covered and how services must be organized to meet the health needs of the population.

  8. [Economics of health system transformation].

    PubMed

    González Pier, Eduardo

    2012-01-01

    Health conditions in Mexico have evolved along with socioeconomic conditions. As a result, today's health system faces several problems characterized by four overlapping transitions: demand, expectations, funding and health resources. These transitions engender significant pressures on the system itself. Additionally, fragmentation of the health system creates disparities in access to services and generates problems in terms of efficiency and use of available resources. To address these complications and to improve equity in access and efficiency, thorough analysis is required in how the right to access health care should be established at a constitutional level without differentiating across population groups. This should be followed by careful discussion about what rules of health care financing should exist, which set of interventions ought to be covered and how services must be organized to meet the health needs of the population. PMID:23254712

  9. Toward transformational carbon capture systems

    SciTech Connect

    Miller, David C.; Litynski, John T.; Brickett, Lynn A.; Morreale, Bryan D.

    2015-10-28

    This paper will briefly review the history and current state of Carbon Capture and Storage (CCS) research and development and describe the technical barriers to carbon capture. it will argue forcefully for a new approach to R&D, which leverages both simulation and physical systems at the laboratory and pilot scales to more rapidly move the best technoogies forward, prune less advantageous approaches, and simultaneously develop materials and processes.

  10. pORE: a modular binary vector series suited for both monocot and dicot plant transformation.

    PubMed

    Coutu, Catherine; Brandle, James; Brown, Dan; Brown, Kirk; Miki, Brian; Simmonds, John; Hegedus, Dwayne D

    2007-12-01

    We present a series of 14 binary vectors suitable for Agrobacterium-mediated transformation of dicotyledonous plants and adaptable for biolistic transformation of monocotyledonous plants. The vector size has been minimized by eliminating all non-essential elements from the vector backbone and T-DNA regions while maintaining the ability to replicate independently. The smallest of the vector series is 6.3 kb and possesses an extensive multiple cloning site with 21 unique restriction endonuclease sites that are compatible with common cloning, protein expression, yeast two-hybrid and other binary vectors. The T-DNA region was engineered using a synthetic designer oligonucleotide resulting in an entirely modular system whereby any vector element can be independently exchanged. The high copy number ColE1 origin of replication has been included to enhance plasmid yield in Escherichia coli. FRT recombination sites flank the selectable marker cassette regions and allow for in planta excision by FLP recombinase. The pORE series consists of three basic types; an 'open' set for general plant transformation, a 'reporter' set for promoter analysis and an 'expression' set for constitutive expression of transgenes. The sets comprise various combinations of promoters (P (HPL), P (ENTCUP2) and P (TAPADH)), selectable markers (nptII and pat) and reporter genes (gusA and smgfp). PMID:17273915

  11. Construction of transformation system in Penicillium purpurogenum.

    PubMed

    Kojima, Ryo; Arai, Teppei; Kasumi, Takafumi; Ogihara, Jun

    2015-03-01

    Penicillium purpurogenum attracts attention in the food industry and biomass degradation. We expressed green fluorescent protein (GFP) with pBPE, a novel vector, and constructed a transformation system for P. purpurogenum. The accumulation of GFP was confirmed by fluorescence microscopy. In future, this system may prove useful for the genetic modification of P. purpurogenum.

  12. SETS. Set Equation Transformation System

    SciTech Connect

    Worrell, R.B.

    1992-01-13

    SETS is used for symbolic manipulation of Boolean equations, particularly the reduction of equations by the application of Boolean identities. It is a flexible and efficient tool for performing probabilistic risk analysis (PRA), vital area analysis, and common cause analysis. The equation manipulation capabilities of SETS can also be used to analyze noncoherent fault trees and determine prime implicants of Boolean functions, to verify circuit design implementation, to determine minimum cost fire protection requirements for nuclear reactor plants, to obtain solutions to combinatorial optimization problems with Boolean constraints, and to determine the susceptibility of a facility to unauthorized access through nullification of sensors in its protection system.

  13. A protocol for sonication-assisted Agrobacterium rhizogenes-mediated transformation of haploid and diploid sugar beet (Beta vulgaris L.) explants.

    PubMed

    Klimek-Chodacka, Magdalena; Baranski, Rafal

    2014-01-01

    Hairy root cultures obtained after Agrobacterium rhizogenes-mediated genetic transformation can serve as a model system for studying plant metabolism and physiology, or can be utilized for the production of secondary metabolites. So far no efficient protocol of hairy root development in sugar beet has been publically released. In this work, two A. rhizogenes strains (A4T and LBA1334) carrying a binary vector pBIN-m-gfp5-ER or pCAMBIA1301 possessing gfp and uidA reporter genes were used to transform petiole explants of haploid and diploid sugar beet genotypes. Five treatment combinations of sonicated-assisted Agrobacterium-mediated transformation were compared. Hairy roots appeared on 0% to 54% of explants depending on the treatment combination used. The highest frequency was achieved when explants of a diploid genotype were sonicated for 15 s in the inoculum containing A. rhizogenes of OD600=0.5 and then co-cultured for three days. Using the same treatment combinations the explants of haploid genotypes developed hairy roots with the frequency ranging from 10% to 36%. Transformation efficiency was independent on the bacterial strain used. The results indicate that haploid sugar beet explants are amenable to transformation using A. rhizogenes, and that the efficiency of that process can be increased by applying short ultrasound treatment.

  14. HERCULES: A Pattern Driven Code Transformation System

    SciTech Connect

    Kartsaklis, Christos; Hernandez, Oscar R; Hsu, Chung-Hsing; Ilsche, Thomas; Joubert, Wayne; Graham, Richard L

    2012-01-01

    New parallel computers are emerging, but developing efficient scientific code for them remains difficult. A scientist must manage not only the science-domain complexity but also the performance-optimization complexity. HERCULES is a code transformation system designed to help the scientist to separate the two concerns, which improves code maintenance, and facilitates performance optimization. The system combines three technologies, code patterns, transformation scripts and compiler plugins, to provide the scientist with an environment to quickly implement code transformations that suit his needs. Unlike existing code optimization tools, HERCULES is unique in its focus on user-level accessibility. In this paper we discuss the design, implementation and an initial evaluation of HERCULES.

  15. Agrobacterium-mediated disruption of a nonribosomal peptide synthetase gene in the invertebrate pathogen Metarhizium anisopliae reveals a peptide spore factor

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Numerous secondary metabolites have been isolated from the insect pathogenic fungus Metarhizium anisopliae, but the roles of these compounds as virulence factors in disease development are poorly understood. We targeted for disruption by Agrobacterium tumefaciens-mediated transformation a putative n...

  16. A High-Throughput Regeneration and Transformation Platform for Production of Genetically Modified Banana

    PubMed Central

    Tripathi, Jaindra N.; Oduor, Richard O.; Tripathi, Leena

    2015-01-01

    Banana (Musa spp.) is an important staple food as well as cash crop in tropical and subtropical countries. Various bacterial, fungal, and viral diseases and pests such as nematodes are major constraints in its production and are currently destabilizing the banana production in sub-Saharan Africa. Genetic engineering is a complementary option used for incorporating useful traits in banana to bypass the long generation time, polyploidy, and sterility of most of the cultivated varieties. A robust transformation protocol for farmer preferred varieties is crucial for banana genomics and improvement. A robust and reproducible system for genetic transformation of banana using embryogenic cell suspensions (ECS) has been developed in this study. Two different types of explants (immature male flowers and multiple buds) were tested for their ability to develop ECS in several varieties of banana locally grown in Africa. ECS of banana varieties “Cavendish Williams” and “Gros Michel” were developed using multiple buds, whereas ECS of “Sukali Ndiizi” was developed using immature male flowers. Regeneration efficiency of ECS was about 20,000–50,000 plantlets per ml of settled cell volume (SCV) depending on variety. ECS of three different varieties were transformed through Agrobacterium-mediated transformation using gusA reporter gene and 20–70 independent transgenic events per ml SCV of ECS were regenerated on selective medium. The presence and integration of gusA gene in transgenic plants was confirmed by PCR, dot blot, and Southern blot analysis and expression by histochemical GUS assays. The robust transformation platform was successfully used to generate hundreds of transgenic lines with disease resistance. Such a platform will facilitate the transfer of technologies to national agricultural research systems (NARS) in Africa. PMID:26635849

  17. A High-Throughput Regeneration and Transformation Platform for Production of Genetically Modified Banana.

    PubMed

    Tripathi, Jaindra N; Oduor, Richard O; Tripathi, Leena

    2015-01-01

    Banana (Musa spp.) is an important staple food as well as cash crop in tropical and subtropical countries. Various bacterial, fungal, and viral diseases and pests such as nematodes are major constraints in its production and are currently destabilizing the banana production in sub-Saharan Africa. Genetic engineering is a complementary option used for incorporating useful traits in banana to bypass the long generation time, polyploidy, and sterility of most of the cultivated varieties. A robust transformation protocol for farmer preferred varieties is crucial for banana genomics and improvement. A robust and reproducible system for genetic transformation of banana using embryogenic cell suspensions (ECS) has been developed in this study. Two different types of explants (immature male flowers and multiple buds) were tested for their ability to develop ECS in several varieties of banana locally grown in Africa. ECS of banana varieties "Cavendish Williams" and "Gros Michel" were developed using multiple buds, whereas ECS of "Sukali Ndiizi" was developed using immature male flowers. Regeneration efficiency of ECS was about 20,000-50,000 plantlets per ml of settled cell volume (SCV) depending on variety. ECS of three different varieties were transformed through Agrobacterium-mediated transformation using gusA reporter gene and 20-70 independent transgenic events per ml SCV of ECS were regenerated on selective medium. The presence and integration of gusA gene in transgenic plants was confirmed by PCR, dot blot, and Southern blot analysis and expression by histochemical GUS assays. The robust transformation platform was successfully used to generate hundreds of transgenic lines with disease resistance. Such a platform will facilitate the transfer of technologies to national agricultural research systems (NARS) in Africa.

  18. Efficient transformation and artificial miRNA gene silencing in Lemna minor.

    PubMed

    Cantó-Pastor, A; Mollá-Morales, A; Ernst, E; Dahl, W; Zhai, J; Yan, Y; Meyers, B C; Shanklin, J; Martienssen, R

    2015-01-01

    Despite rapid doubling time, simple architecture and ease of metabolic labelling, a lack of genetic tools in the Lemnaceae (duckweed) has impeded the full implementation of this organism as a model for biological research. Here, we present technologies to facilitate high-throughput genetic studies in duckweed. We developed a fast and efficient method for producing Lemna minor stable transgenic fronds via Agrobacterium-mediated transformation and regeneration from tissue culture. Additionally, we engineered an artificial microRNA (amiRNA) gene silencing system. We identified a Lemna gibba endogenous miR166 precursor and used it as a backbone to produce amiRNAs. As a proof of concept we induced the silencing of CH42, a magnesium chelatase subunit, using our amiRNA platform. Expression of CH42 in transgenic L. minor fronds was significantly reduced, which resulted in reduction of chlorophyll pigmentation. The techniques presented here will enable tackling future challenges in the biology and biotechnology of Lemnaceae.

  19. Efficient transformation and artificial miRNA gene silencing in Lemna minor.

    PubMed

    Cantó-Pastor, A; Mollá-Morales, A; Ernst, E; Dahl, W; Zhai, J; Yan, Y; Meyers, B C; Shanklin, J; Martienssen, R

    2015-01-01

    Despite rapid doubling time, simple architecture and ease of metabolic labelling, a lack of genetic tools in the Lemnaceae (duckweed) has impeded the full implementation of this organism as a model for biological research. Here, we present technologies to facilitate high-throughput genetic studies in duckweed. We developed a fast and efficient method for producing Lemna minor stable transgenic fronds via Agrobacterium-mediated transformation and regeneration from tissue culture. Additionally, we engineered an artificial microRNA (amiRNA) gene silencing system. We identified a Lemna gibba endogenous miR166 precursor and used it as a backbone to produce amiRNAs. As a proof of concept we induced the silencing of CH42, a magnesium chelatase subunit, using our amiRNA platform. Expression of CH42 in transgenic L. minor fronds was significantly reduced, which resulted in reduction of chlorophyll pigmentation. The techniques presented here will enable tackling future challenges in the biology and biotechnology of Lemnaceae. PMID:24989135

  20. Transient plant transformation mediated by Agrobacterium tumefaciens: Principles, methods and applications.

    PubMed

    Krenek, Pavel; Samajova, Olga; Luptovciak, Ivan; Doskocilova, Anna; Komis, George; Samaj, Jozef

    2015-11-01

    Agrobacterium tumefaciens is widely used as a versatile tool for development of stably transformed model plants and crops. However, the development of Agrobacterium based transient plant transformation methods attracted substantial attention in recent years. Transient transformation methods offer several applications advancing stable transformations such as rapid and scalable recombinant protein production and in planta functional genomics studies. Herein, we highlight Agrobacterium and plant genetics factors affecting transfer of T-DNA from Agrobacterium into the plant cell nucleus and subsequent transient transgene expression. We also review recent methods concerning Agrobacterium mediated transient transformation of model plants and crops and outline key physical, physiological and genetic factors leading to their successful establishment. Of interest are especially Agrobacterium based reverse genetics studies in economically important crops relying on use of RNA interference (RNAi) or virus-induced gene silencing (VIGS) technology. The applications of Agrobacterium based transient plant transformation technology in biotech industry are presented in thorough detail. These involve production of recombinant proteins (plantibodies, vaccines and therapeutics) and effectoromics-assisted breeding of late blight resistance in potato. In addition, we also discuss biotechnological potential of recombinant GFP technology and present own examples of successful Agrobacterium mediated transient plant transformations.

  1. Spike-dip transformation of Setaria viridis.

    PubMed

    Saha, Prasenjit; Blumwald, Eduardo

    2016-04-01

    Traditional method of Agrobacterium-mediated transformation through the generation of tissue culture had limited success for Setaria viridis, an emerging C4 monocot model. Here we present an efficient in planta method for Agrobacterium-mediated genetic transformation of S. viridis using spike dip. Pre-anthesis developing spikes were dipped into a solution of Agrobacterium tumefaciens strain AGL1 harboring the β-glucuronidase (GUS) reporter gene driven by the cauliflower mosaic virus 35S (CaMV35S) promoter to standardize and optimize conditions for transient as well as stable transformations. A transformation efficiency of 0.8 ± 0.1% was obtained after dipping of 5-day-old S3 spikes for 20 min in Agrobacterium cultures containing S. viridis spike-dip medium supplemented with 0.025% Silwet L-77 and 200 μm acetosyringone. Reproducibility of this method was demonstrated by generating stable transgenic lines expressing β-glucuronidase plus (GUSplus), green fluorescent protein (GFP) and Discosoma sp. red fluorescent protein (DsRed) reporter genes driven by either CaMV35S or intron-interrupted maize ubiquitin (Ubi) promoters from three S. viridis genotypes. Expression of these reporter genes in transient assays as well as in T1 stable transformed plants was monitored using histochemical, fluorometric GUS activity and fluorescence microscopy. Molecular analysis of transgenic lines revealed stable integration of transgenes into the genome, and inherited transgenes expressed in the subsequent generations. This approach provides opportunities for the high-throughput transformation and potentially facilitates translational research in a monocot model plant. PMID:26932666

  2. Transformers and the Electric Utility System

    ERIC Educational Resources Information Center

    Roman, Harry T.

    2005-01-01

    For electric energy to get from the generating station to a home, it must pass through a transformer, a device that can change voltage levels easily. This article describes how transformers work, covering the following topics: (1) the magnetism-electricity link; (2) transformer basics; (3) the energy seesaw; (4) the turns ratio rule; and (5)…

  3. Citrus transformation using juvenile tissue explants.

    PubMed

    Orbović, Vladimir; Grosser, Jude W

    2015-01-01

    The most frequently used method for production of citrus transgenic plants is via Agrobacterium-mediated transformation of tissues found on explants obtained from juvenile seedlings. Within the last decade and especially within the last 5-6 years, this robust method was employed to produce thousands of transgenic plants. With the newly applied screening methods that allow easier and faster detection of transgenic shoots, estimates of transformation rate for some cultivars have gone up making this approach even more attractive. Although adjustments have to be made regarding the (varietal) source of the starting material and Agrobacterium strain used in each experiment preformed, the major steps of this procedure have not changed significantly if at all. Transgenic citrus plants produced this way belong to cultivars of rootstocks, sweet oranges, grapefruits, mandarins, limes, and lemons.

  4. Unlocking the potential of tropical root crop biotechnology in east Africa by establishing a genetic transformation platform for local farmer-preferred cassava cultivars

    PubMed Central

    Nyaboga, Evans; Njiru, Joshua; Nguu, Edward; Gruissem, Wilhelm; Vanderschuren, Herve; Tripathi, Leena

    2013-01-01

    Cassava genetic transformation capacity is still mostly restricted to advanced laboratories in the USA, Europe and China; and its implementation and maintenance in African laboratories has remained scarce. The impact of transgenic technologies for genetic improvement of cassava will depend largely on the transfer of such capabilities to researchers in Africa, where cassava has an important socioeconomic niche. A major constraint to the development of genetic transformation technologies for cassava improvement has been the lack of an efficient and robust transformation and regeneration system. Despite the success achieved in genetic modification of few cassava cultivars, including the model cultivar 60444, transgenic cassava production remains difficult for farmer-preferred cultivars. In this study, a protocol for cultivar 60444 developed at ETH Zurich was successfully implemented and optimized to establish transformation of farmer-preferred cassava cultivars popular in east Africa. The conditions for production and proliferation of friable embryogenic calli (FEC) and Agrobacterium-mediated transformation were optimized for three east African farmer-preferred cultivars (Ebwanatereka, Kibandameno and Serere). Our results demonstrated transformation efficiencies of about 14–22 independent transgenic lines per 100 mg of FEC for farmer-preferred cultivars in comparison to 28 lines per 100 mg of the model cultivar 60444. The presence, integration and expression of the transgenes were confirmed by PCR, Southern blot analysis and histochemical GUS assay. This study reports the establishment of a cassava transformation platform at International Institute of Tropical Agriculture (IITA) hosted by Biosciences eastern and central Africa (BecA) hub in Kenya and provides the basis for transferring important traits such as virus resistance and prolonged shelf-life to farmer-preferred cultivars in east Africa. We anticipate that such platform will also be instrumental to transfer

  5. [Social security for health system transformation].

    PubMed

    Echevarría-Zuno, Santiago

    2012-01-01

    The aim of this article is to present the management and medical components within the public health institutions that can contribute to the transformation of the National Health System (NHS). It is expected that these will have an impact in the extent of the medical coverage and will improve the health care services delivered to the Mexican population. A diagnostic study revealed the existence of fragmentation in the NHS. The health institutions are vertically established and operate under isolated mechanisms of financing and administration. Additionally, it is pointed out the problematic derived from the multiplicity in the public insurance conditions among individuals and the lack of census of the insured population within the institutions. As part of the universalization of health services, it is necessary to integrate the health care system; accordingly, a variety of mechanisms for the partial and total integration are arise, such as the exchange of the health care services and the portability and convergence of the institutions. Particularly, we listed the actions carried out by the Mexican Institute of Social Security for the integration of the NHS such as, the independent management medical areas, diagnosis-related groups (DRG), the performing evaluation of the medical units, and the preventive and curative strategies in the implemented programs. Finally, is dealt some reflections in order to improve the public health care.

  6. Postreplication targeting of transformants by bacterial immune systems?

    PubMed

    Johnston, Calum; Martin, Bernard; Polard, Patrice; Claverys, Jean-Pierre

    2013-10-01

    Bacteria are constantly challenged by foreign genetic elements such as bacteriophages and plasmids. Several defense systems provide immunity against such attackers, including restriction-modification (R-M) systems and clustered, regularly interspaced short palindromic repeats (CRISPRs). These systems target attacking DNA and thus antagonize natural transformation, which relies on uptake of exogenous DNA to promote acquisition of new genetic traits. It is unclear how this antagonization occurs, because transforming DNA is single stranded, and thus resistant to these immune systems. Here, we propose a simple model whereby these systems limit transformation by attack of transformed chromosomes once double strandedness is restored by chromosomal replication.

  7. Transforming Research Management Systems at Mayo Clinic

    ERIC Educational Resources Information Center

    Smith, Steven C.; Gronseth, Darren L.

    2011-01-01

    In order for research programs at academic medical centers and universities to survive and thrive in the increasingly challenging economic, political and regulatory environment, successful transformation is extremely important. Transformation and quality management techniques are increasingly well established in medical practice organizations. In…

  8. Optimization of in vitro regeneration and Agrobacterium tumefaciens-mediated transformation with heat-resistant cDNA in Brassica oleracea subsp. italica cv. Green Marvel.

    PubMed

    Ravanfar, Seyed Ali; Aziz, Maheran Abdul; Saud, Halimi Mohd; Abdullah, Janna Ong

    2015-11-01

    An efficient system for shoot regeneration and Agrobacterium tumefaciens-mediated transformation of Brassica oleracea cv. Green Marvel cultivar is described. This study focuses on developing shoot regeneration from hypocotyl explants of broccoli cv. Green Marvel using thidiazuron (TDZ), zeatin, and kinetin, the optimization of factors affecting Agrobacterium-mediated transformation of the hypocotyl explants with heat-resistant cDNA, followed by the confirmation of transgenicity of the regenerants. High shoot regeneration was observed in 0.05-0.1 mg dm(-3) TDZ. TDZ at 0.1 mg dm(-3) produced among the highest percentage of shoot regeneration (96.67 %) and mean number of shoot formation (6.17). The highest percentage (13.33 %) and mean number (0.17) of putative transformant production were on hypocotyl explants subjected to preculture on shoot regeneration medium (SRM) with 200 µM acetosyringone. On optimization of bacterial density and inoculation time, the highest percentage and mean number of putative transformant production were on hypocotyl explants inoculated with a bacterial dilution of 1:5 for 30 min. Polymerase chain reaction (PCR) assay indicated a transformation efficiency of 8.33 %. The luciferase assay showed stable integration of the Arabidopsis thaliana HSP101 (AtHSP101) cDNA in the transgenic broccoli regenerants. Three out of five transgenic lines confirmed through PCR showed positive hybridization bands of the AtHSP101 cDNA through Southern blot analysis. The presence of AtHSP101 transcripts in the three transgenic broccoli lines indicated by reverse transcription-PCR (RT-PCR) confirmed the expression of the gene. In conclusion, an improved regeneration system has been established from hypocotyl explants of broccoli followed by successful transformation with AtHSP101 for resistance to high temperature.

  9. Transformational System Concepts and Technologies for Future Space Applications

    NASA Technical Reports Server (NTRS)

    Mankins, John C.; Howell, Joe T.

    2004-01-01

    The President's VIsion for Space Exploration offers opportunities to seek new paradigms and develop transformational space infrastructures. First steps are being taken to develop transformational system concepts such as modular reconfigurable systems, cryogenic propellant depots to preposition propellants, etc., and to develop other critical technologies.

  10. Polynomial Transformations For Discrete-Time Linear Systems

    NASA Technical Reports Server (NTRS)

    Baram, Yoram

    1991-01-01

    Transformations based on polynomial matrices of finite degree developed for use in computing functions for compensation, inversion, and approximation of discrete-time, multivariable, linear systems. Method derived from z-transform transfer-function form of matrices. Applicable to cascade-compensation problems in design of control systems.

  11. RNA viral vectors for improved Agrobacterium-mediated transient expression of heterologous proteins in Nicotiana benthamiana cell suspensions and hairy roots

    PubMed Central

    2012-01-01

    Background Plant cell suspensions and hairy root cultures represent scalable protein expression platforms. Low protein product titers have thus far limited the application of transient protein expression in these hosts. The objective of this work was to overcome this limitation by harnessing A. tumefaciens to deliver replicating and non-replicating RNA viral vectors in plant tissue co-cultures. Results Replicating vectors derived from Potato virus X (PVX) and Tobacco rattle virus (TRV) were modified to contain the reporter gene β-glucuronidase (GUS) with a plant intron to prevent bacterial expression. In cell suspensions, a minimal PVX vector retaining only the viral RNA polymerase gene yielded 6.6-fold more GUS than an analogous full-length PVX vector. Transient co-expression of the minimal PVX vector with P19 of Tomato bushy stunt virus or HC-Pro of Tobacco etch virus to suppress post-transcriptional gene silencing increased GUS expression by 44 and 83%, respectively. A non-replicating vector containing a leader sequence from Cowpea mosaic virus (CPMV-HT) modified for enhanced translation led to 70% higher transient GUS expression than a control treatment. In hairy roots, a TRV vector capable of systemic movement increased GUS accumulation by 150-fold relative to the analogous PVX vector. Histochemical staining for GUS in TRV-infected hairy roots revealed the capacity for achieving even higher productivity per unit biomass. Conclusions For the first time, replicating PVX vectors and a non-replicating CPMV-HT vector were successfully applied toward transient heterologous protein expression in cell suspensions. A replicating TRV vector achieved transient GUS expression levels in hairy roots more than an order of magnitude higher than the highest level previously reported with a viral vector delivered by A. tumefaciens. PMID:22559055

  12. Development of an efficient transformation method by Agrobacterium tumefaciens and high throughput spray assay to identify transgenic plants for woodland strawberry (Fragaria vesca) using NPTII selection.

    PubMed

    Pantazis, Christopher J; Fisk, Sarah; Mills, Kerri; Flinn, Barry S; Shulaev, Vladimir; Veilleux, Richard E; Dan, Yinghui

    2013-03-01

    KEY MESSAGE : We developed an efficient Agrobacterium -mediated transformation method using an Ac/Ds transposon tagging construct for F. vesca and high throughput paromomycin spray assay to identify its transformants for strawberry functional genomics. Genomic resources for Rosaceae species are now readily available, including the Fragaria vesca genome, EST sequences, markers, linkage maps, and physical maps. The Rosaceae Genomic Executive Committee has promoted strawberry as a translational genomics model due to its unique biological features and transformability for fruit trait improvement. Our overall research goal is to use functional genomic and metabolic approaches to pursue high throughput gene discovery in the diploid woodland strawberry. F. vesca offers several advantages of a fleshy fruit typical of most fruit crops, short life cycle (seed to seed in 12-16 weeks), small genome size (206 Mbb/C), small plant size, self-compatibility, and many seeds per plant. We have developed an efficient Agrobacterium tumefaciens-mediated strawberry transformation method using kanamycin selection, and high throughput paromomycin spray assay to efficiently identify transgenic strawberry plants. Using our kanamycin transformation method, we were able to produce up to 98 independent kanamycin resistant insertional mutant lines using a T-DNA construct carrying an Ac/Ds transposon Launchpad system from a single transformation experiment involving inoculation of 22 leaf explants of F. vesca accession 551572 within approx. 11 weeks (from inoculation to soil). Transgenic plants with 1-2 copies of a transgene were confirmed by Southern blot analysis. Using our paromomycin spray assay, transgenic F. vesca plants were rapidly identified within 10 days after spraying.

  13. Multi-view coordinate system transformation based on robot

    NASA Astrophysics Data System (ADS)

    Song, Li-mei; Wang, Peng-qiang; Xi, Jiang-tao; Guo, Qing-hua; Tang, Huan; Li, Jing; Li, Xiao-jie; Zhu, Teng-da

    2015-11-01

    The registration of point cloud is important for large object measurement. A measurement method for coordinate system transformation based on robot is proposed in this paper. Firstly, for obtaining extrinsic parameters, the robot moves to three different positions to capture the images of three targets. Then the transformation matrix X between camera and tool center point (TCP) coordinate systems can be calculated by using the known parameters of robot and the extrinsic parameters, and finally the multi-view coordinate system can be transformed into robot coordinate system by the transformation matrix X. With the help of robot, the multi-view point cloud can be easily transformed into a unified coordinate system. By using robot, the measurement doesn't need any mark. Experimental results show that the method is effective.

  14. Genetic Transformation System for the Fungal Soybean Pathogen Cercospora kikuchii.

    PubMed

    Upchurch, R G; Ehrenshaft, M; Walker, D C; Sanders, L A

    1991-10-01

    An altered beta-tubulin gene that confers resistance to the fungicide benomyl was isolated from a genomic library of a UV-induced mutant of Cercospora kikuchii and used as a selectable marker for transformation. The level of benomyl resistance conferred to the transformants was at least 150-fold greater than the intrinsic resistance of the C. kikuchii recipient protoplasts. In the majority of cases, the tubulin fragment was integrated at the native beta-tubulin locus, apparently by gene replacement or gene conversion. The frequency of transformation ranged from 0.2 to 6 transformants per mug of DNA, depending on the recipient strain. Transformation with linearized plasmid resulted in a higher frequency, without changing the type of integration event. Transformants were phenotypically stable after eight consecutive transfers on medium without benomyl. This is the first report of a genetic transformation system for a Cercospora species.

  15. Genetic Transformation System for the Fungal Soybean Pathogen Cercospora kikuchii

    PubMed Central

    Upchurch, R. G.; Ehrenshaft, M.; Walker, D. C.; Sanders, L. A.

    1991-01-01

    An altered β-tubulin gene that confers resistance to the fungicide benomyl was isolated from a genomic library of a UV-induced mutant of Cercospora kikuchii and used as a selectable marker for transformation. The level of benomyl resistance conferred to the transformants was at least 150-fold greater than the intrinsic resistance of the C. kikuchii recipient protoplasts. In the majority of cases, the tubulin fragment was integrated at the native β-tubulin locus, apparently by gene replacement or gene conversion. The frequency of transformation ranged from 0.2 to 6 transformants per μg of DNA, depending on the recipient strain. Transformation with linearized plasmid resulted in a higher frequency, without changing the type of integration event. Transformants were phenotypically stable after eight consecutive transfers on medium without benomyl. This is the first report of a genetic transformation system for a Cercospora species. Images PMID:16348566

  16. Information retrieval system utilizing wavelet transform

    DOEpatents

    Brewster, Mary E.; Miller, Nancy E.

    2000-01-01

    A method for automatically partitioning an unstructured electronically formatted natural language document into its sub-topic structure. Specifically, the document is converted to an electronic signal and a wavelet transform is then performed on the signal. The resultant signal may then be used to graphically display and interact with the sub-topic structure of the document.

  17. Transforming the structure of a health system.

    PubMed

    Reilly, Patrick

    2012-06-01

    In starting the planning process for an organization's transformation or restructuring, healthcare finance leaders should: Identify strategic imperatives for the organization and physicians, Remember the organization's core area of business, Define the starting point and create clear objectives, Develop a strategy that engages front-line employees to change the culture of the organization. PMID:22734323

  18. Barley (Hordeum vulgare L.) transformation using embryogenic pollen cultures.

    PubMed

    Otto, Ingrid; Müller, Andrea; Kumlehn, Jochen

    2015-01-01

    The temperate cereal barley is grown as a source of food, feed, and malt. The development of a broad range of genetic resources and associated technologies in this species has helped to establish barley as the prime model for the other Triticeae cereals. The specific advantage of the transformation method presented here is that transgene homozygosity is attained in the same generation as the transgenic event occurred through the coupling of haploid technology with Agrobacterium-mediated transformation. Pollen is haploid and, following transformation, can be induced to regenerate into haploid plantlets, which can subsequently subjected to colchicine treatment to obtain diploid, genetically fixed plants. The routine application of the method based on the winter-type barley cultivar 'Igri' over a period of over 10 years has achieved an average yield of about two transgenic plants per donor spike. The whole procedure from pollen isolation to non-segregating transgenic, mature grain takes less than 12 months.

  19. Stable Nuclear Transformation System for the Coccolithophorid Alga Pleurochrysis carterae

    PubMed Central

    Endo, Hirotoshi; Yoshida, Megumi; Uji, Toshiki; Saga, Naotsune; Inoue, Koji; Nagasawa, Hiromichi

    2016-01-01

    Of the three dominant marine microalgal groups, dinoflagellates and diatoms can undergo genetic transformation; however, no transformation method has been established for haptophytes to date. Here, we report the first stable genetic transformation of a coccolithophore, Pleurochrysis carterae, by means of polyethylene glycol (PEG)-mediated transfer of a bacterial hygromycin B-resistance gene. Together with the novel transient green fluorescent protein (GFP) expression system, this approach should facilitate further molecular-based research in this phylum. PMID:26947136

  20. Model Transformation for a System of Systems Dependability Safety Case

    NASA Technical Reports Server (NTRS)

    Murphy, Judy; Driskell, Stephen B.

    2010-01-01

    Software plays an increasingly larger role in all aspects of NASA's science missions. This has been extended to the identification, management and control of faults which affect safety-critical functions and by default, the overall success of the mission. Traditionally, the analysis of fault identification, management and control are hardware based. Due to the increasing complexity of system, there has been a corresponding increase in the complexity in fault management software. The NASA Independent Validation & Verification (IV&V) program is creating processes and procedures to identify, and incorporate safety-critical software requirements along with corresponding software faults so that potential hazards may be mitigated. This Specific to Generic ... A Case for Reuse paper describes the phases of a dependability and safety study which identifies a new, process to create a foundation for reusable assets. These assets support the identification and management of specific software faults and, their transformation from specific to generic software faults. This approach also has applications to other systems outside of the NASA environment. This paper addresses how a mission specific dependability and safety case is being transformed to a generic dependability and safety case which can be reused for any type of space mission with an emphasis on software fault conditions.

  1. Efficient transformation of Kalanchoe blossfeldiana and production of male-sterile plants by engineered anther ablation.

    PubMed

    García-Sogo, Begoña; Pineda, Benito; Castelblanque, Lourdes; Antón, Teresa; Medina, Mónica; Roque, Edelín; Torresi, Claudia; Beltrán, José Pío; Moreno, Vicente; Cañas, Luis Antonio

    2010-01-01

    Engineered male sterility in ornamental plants has many applications such as facilitate hybrid seed production, eliminate pollen allergens, reduce the need for deadheading to extend the flowering period, redirect resources from seeds to vegetative growth, increase flower longevity and prevent gene flow between genetically modified and related native plants. We have developed a reliable and efficient Agrobacterium-mediated protocol for the genetic transformation of different Kalanchoe blossfeldiana commercial cultivars. Transformation efficiency for cv. 'Hillary' was 55.3% whereas that of cv. 'Tenorio' reached 75.8%. Selection was carried out with the nptII gene and increasing the kanamycin concentration from 25 to 100 mg l(-1) allowed to reduced escapes from 50 to 60% to virtually 0%. This method was used to produce male-sterile plants through engineered anther ablation. In our approach, we tested a male sterility chimaeric gene construct (PsEND1::barnase) to evaluate its effectiveness and effect on phenotype. No significant differences were found in the growth patterns between the transgenic lines and the wild-type plants. No viable pollen grains were observed in the ablated anthers of any of the lines carrying the PsEND1::barnase construct, indicating that the male sterility was complete. In addition, seed set was completely abolished in all the transgenic plants obtained. Our engineered male-sterile approach could be used, alone or in combination with a female-sterility system, to reduce the invasive potential of new ornamentals, which has become an important environmental problem in many countries.

  2. Code orange: Towards transformational leadership of emergency management systems.

    PubMed

    Caro, Denis H J

    2015-09-01

    The 21(st) century calls upon health leaders to recognize and respond to emerging threats and systemic emergency management challenges through transformative processes inherent in the LEADS in a caring environment framework. Using a grounded theory approach, this qualitative study explores key informant perspectives of leaders in emergency management across Canada on pressing needs for relevant systemic transformation. The emerging model points to eight specific attributes of transformational leadership central to emergency management and suggests that contextualization of health leadership is of particular import.

  3. Code orange: Towards transformational leadership of emergency management systems.

    PubMed

    Caro, Denis H J

    2015-09-01

    The 21(st) century calls upon health leaders to recognize and respond to emerging threats and systemic emergency management challenges through transformative processes inherent in the LEADS in a caring environment framework. Using a grounded theory approach, this qualitative study explores key informant perspectives of leaders in emergency management across Canada on pressing needs for relevant systemic transformation. The emerging model points to eight specific attributes of transformational leadership central to emergency management and suggests that contextualization of health leadership is of particular import. PMID:26135294

  4. Phase transformation kinetics in finite inhomogeneously nucleated systems

    NASA Technical Reports Server (NTRS)

    Weinberg, Michael; Kapral, Raymond

    1989-01-01

    Phase transformation kinetics that occur by a nucleation and growth process are investigated. A simple discrete space and time model is used for the dynamics and analytical results are obtained for the volume fraction of the material transformed for both finite systems and a special example of an inhomogeneously nucleated system. The theory is developed for two cases, initial nucleation, and continuous nucleation. The results are compared with simulations of the model.

  5. Using systems thinking to support clinical system transformation.

    PubMed

    Best, Allan; Berland, Alex; Herbert, Carol; Bitz, Jennifer; van Dijk, Marlies W; Krause, Christina; Cochrane, Douglas; Noel, Kevin; Marsden, Julian; McKeown, Shari; Millar, John

    2016-05-16

    Purpose - The British Columbia Ministry of Health's Clinical Care Management initiative was used as a case study to better understand large-scale change (LSC) within BC's health system. Using a complex system framework, the purpose of this paper is to examine mechanisms that enable and constrain the implementation of clinical guidelines across various clinical settings. Design/methodology/approach - Researchers applied a general model of complex adaptive systems plus two specific conceptual frameworks (realist evaluation and system dynamics mapping) to define and study enablers and constraints. Focus group sessions and interviews with clinicians, executives, managers and board members were validated through an online survey. Findings - The functional themes for managing large-scale clinical change included: creating a context to prepare clinicians for health system transformation initiatives; promoting shared clinical leadership; strengthening knowledge management, strategic communications and opportunities for networking; and clearing pathways through the complexity of a multilevel, dynamic system. Research limitations/implications - The action research methodology was designed to guide continuing improvement of implementation. A sample of initiatives was selected; it was not intended to compare and contrast facilitators and barriers across all initiatives and regions. Similarly, evaluating the results or process of guideline implementation was outside the scope; the methods were designed to enable conversations at multiple levels - policy, management and practice - about how to improve implementation. The study is best seen as a case study of LSC, offering a possible model for replication by others and a tool to shape further dialogue. Practical implications - Recommended action-oriented strategies included engaging local champions; supporting local adaptation for implementation of clinical guidelines; strengthening local teams to guide implementation; reducing

  6. Enhanced salt tolerance of alfalfa (Medicago sativa) by rstB gene transformation.

    PubMed

    Zhang, Wan-Jun; Wang, Tao

    2015-05-01

    Generating salt tolerance forage plant is essential for use of the land affected by high salinity. A salt tolerance gene rstB was used as a selectable marker gene in Agrobacterium-mediated transformation of tobacco under a selective regime of 170mM NaCl. The transgenic plants showed clear improvement in salt tolerance. To improve salt tolerance of alfalfa (Medicago sativa L.), rstB gene was introduced into alfalfa genome by Agrobacterium-mediated transformation. No abnormal phenotype was observed among the transgenic plants when compared with wild type (wt) plants. Significant enhancement of resistance to salt-shock treatment was noted on the rstB transgenic (T0) plants. Transgenic second-generation (T1) seeds showed improved germination rate and seedling growth under salt-stress condition. Hindered Na(+) accumulation, but enhanced Ca(2+) accumulation was observed on the rstB T1 plants when subjected to salt-stresses. Enhanced calcium accumulation in transgenic plants was also verified by cytohistochemical localization of calcium. Under salt-stress of 50mM NaCl, about 15% of the transgenic plants finished their life-cycle but the wt plants had no flower formation. The results demonstrated that the expression of rstB gene improved salt tolerance in transgenic alfalfa.

  7. Optimization of genetic transformation of Artemisia annua L. Using Agrobacterium for Artemisinin production

    PubMed Central

    Elfahmi; Suhandono, Sony; Chahyadi, Agus

    2014-01-01

    Background: Artemisinin, a sesquiterpene lactone endoperoxide isolated from the medicinal plant Artemisia annua L., is a choice and effective drug for malaria treatment. Due to the low yield of artemisinin in plants, there is a need to enhance the production of artemisinin from A. annua and biotechnological technique may be one of the methods that can be used for the purpose. Aim: To study the transformation efficiency of Agrobacterium tumefaciens in A. annua that could be applied to enhance the production of artemisinin by means of transgenic plants. Setting and Designs: The factors influencing Agrobacterium-mediated transformation of A. annua were explored to optimize the transformation system, which included A. tumefaciens strain and effect of organosilicone surfactants. Three strains of A. tumefaciens, that is, LBA4404, GV1301, and AGL1 harboring the binary vector pCAMBIA 1303 have been used for transformation. The evaluation was based on transient β-glucuronidase (GUS). Materials and Methods: Plant cell cultures were inniatiated from the seeds of A. annua using the germination Murashige and Skoog medium. A. tumefaciens harboring pCAMBIA were tranformed into the leaves of A.annua cultures from 2-week-old-seedling and 2-month-old-seedling for 15 min by vacuum infiltration. Transformation efficiency was determinated by measuring of blue area (GUS expression) on the whole leaves explant using ImageJ 1.43 software. Two organosilicon surfactants, that is, Silwet L-77 and Silwet S-408 were used to improve the transformation efficiency. Results: The transformation frequency with AGL1 strain was higher than GV3101 and LBA4404 which were 70.91, 49.25, and 45.45%, respectively. Effect of organosilicone surfactants, that is, Silwet L-77 and Silwet S-408 were tested on A. tumefaciens AGL1 and GV3101 for their level of transient expression, and on A. rhizogenes R1000 for its hairy root induction frequency. For AGL1, Silwet S-408 produced higher level of expression than

  8. A simple Gateway-assisted construction system of TALEN genes for plant genome editing

    PubMed Central

    Kusano, Hiroaki; Onodera, Hitomi; Kihira, Miho; Aoki, Hiromi; Matsuzaki, Hikaru; Shimada, Hiroaki

    2016-01-01

    TALEN is an artificial nuclease being applied for sequence-specific genome editing. For the plant genome editing, a pair of TALEN genes is expressed in the cells, and a binary plasmid for Agrobacterium-mediated transformation should be assembled. We developed a novel procedure using the Gateway-assisted plasmids, named Emerald–Gateway TALEN system. We constructed entry vectors, pPlat plasmids, for construction of a desired TALEN gene using Platinum Gate TALEN kit. We also created destination plasmid, pDual35SGw1301, which allowed two TALEN genes to both DNA strands to recruit using Gateway technology. Resultant TALEN genes were evaluated by the single-strand annealing (SSA) assay in E. coli cells. By this assay, the TALENs recognized the corresponding targets in the divided luciferase gene, and induced a specific recombination to generate an active luciferase gene. Using the TALEN genes constructed, we created a transformant potato cells in which a site-specific mutation occurred at the target site of the GBSS gene. This suggested that our system worked effectively and was applicable as a convenient tool for the plant genome editing. PMID:27452606

  9. A simple Gateway-assisted construction system of TALEN genes for plant genome editing.

    PubMed

    Kusano, Hiroaki; Onodera, Hitomi; Kihira, Miho; Aoki, Hiromi; Matsuzaki, Hikaru; Shimada, Hiroaki

    2016-01-01

    TALEN is an artificial nuclease being applied for sequence-specific genome editing. For the plant genome editing, a pair of TALEN genes is expressed in the cells, and a binary plasmid for Agrobacterium-mediated transformation should be assembled. We developed a novel procedure using the Gateway-assisted plasmids, named Emerald-Gateway TALEN system. We constructed entry vectors, pPlat plasmids, for construction of a desired TALEN gene using Platinum Gate TALEN kit. We also created destination plasmid, pDual35SGw1301, which allowed two TALEN genes to both DNA strands to recruit using Gateway technology. Resultant TALEN genes were evaluated by the single-strand annealing (SSA) assay in E. coli cells. By this assay, the TALENs recognized the corresponding targets in the divided luciferase gene, and induced a specific recombination to generate an active luciferase gene. Using the TALEN genes constructed, we created a transformant potato cells in which a site-specific mutation occurred at the target site of the GBSS gene. This suggested that our system worked effectively and was applicable as a convenient tool for the plant genome editing. PMID:27452606

  10. TRANSFORMATION

    SciTech Connect

    LACKS,S.A.

    2003-10-09

    Transformation, which alters the genetic makeup of an individual, is a concept that intrigues the human imagination. In Streptococcus pneumoniae such transformation was first demonstrated. Perhaps our fascination with genetics derived from our ancestors observing their own progeny, with its retention and assortment of parental traits, but such interest must have been accelerated after the dawn of agriculture. It was in pea plants that Gregor Mendel in the late 1800s examined inherited traits and found them to be determined by physical elements, or genes, passed from parents to progeny. In our day, the material basis of these genetic determinants was revealed to be DNA by the lowly bacteria, in particular, the pneumococcus. For this species, transformation by free DNA is a sexual process that enables cells to sport new combinations of genes and traits. Genetic transformation of the type found in S. pneumoniae occurs naturally in many species of bacteria (70), but, initially only a few other transformable species were found, namely, Haemophilus influenzae, Neisseria meningitides, Neisseria gonorrheae, and Bacillus subtilis (96). Natural transformation, which requires a set of genes evolved for the purpose, contrasts with artificial transformation, which is accomplished by shocking cells either electrically, as in electroporation, or by ionic and temperature shifts. Although such artificial treatments can introduce very small amounts of DNA into virtually any type of cell, the amounts introduced by natural transformation are a million-fold greater, and S. pneumoniae can take up as much as 10% of its cellular DNA content (40).

  11. The classical Korteweg capillarity system: geometry and invariant transformations

    NASA Astrophysics Data System (ADS)

    Rogers, C.; Schief, W. K.

    2014-08-01

    A class of invariant transformations is presented for the classical Korteweg capillarity system. The invariance is an extension of a kind originally introduced in an anisentropic gasdynamics context. In a particular instance, application of the invariant transformation leads to a deformed one-parameter class of Kármán-Tsien-type capillarity laws associated with a deformation of an integrable nonlinear Schrödinger-type equation which incorporates a de Broglie-Bohm potential. The latter and another integrable case associated with the classical Boussinesq equation may be linked to the motion of curves in Euclidean and projective space so that both the invariant transformation and the Galilean invariance of the capillarity system may be interpreted in a geometric and soliton-theoretic manner. The work is set in the broader context of other connections of invariant transformations in gasdynamics with soliton theory.

  12. Suitability of non-lethal marker and marker-free systems for development of transgenic crop plants: present status and future prospects.

    PubMed

    Manimaran, P; Ramkumar, G; Sakthivel, K; Sundaram, R M; Madhav, M S; Balachandran, S M

    2011-01-01

    Genetically modified crops are one of the prudent options for enhancing the production and productivity of crop plants by safeguarding from the losses due to biotic and abiotic stresses. Agrobacterium-mediated and biolistic transformation methods are used to develop transgenic crop plants in which selectable marker genes (SMG) are generally deployed to identify 'true' transformants. The commonly used SMG obtained from prokaryotic sources when employed in transgenic plants pose risks due to their lethal nature during selection process. In the recent past, some non-lethal SMGs have been identified and used for selection of transformants with increased precision and high selection efficiency. Considering the concerns related to bio-safety of the environment, it is desirable to remove the SMG in order to maximize the commercial success through wide adoption and public acceptance of genetically modified (GM) food crops. In this review, we examine the availability, and the suitability of wide range of non-lethal selection markers and elimination of SMG methods to develop marker-free transgenics for achieving global food security. As the strategies for marker-free plants are still in proof-of-concept stage, adaptation of new genomics tools for identification of novel non-lethal marker systems and its application for developing marker-free transgenics would further strengthen the crop improvement program. PMID:21672619

  13. Suitability of non-lethal marker and marker-free systems for development of transgenic crop plants: present status and future prospects.

    PubMed

    Manimaran, P; Ramkumar, G; Sakthivel, K; Sundaram, R M; Madhav, M S; Balachandran, S M

    2011-01-01

    Genetically modified crops are one of the prudent options for enhancing the production and productivity of crop plants by safeguarding from the losses due to biotic and abiotic stresses. Agrobacterium-mediated and biolistic transformation methods are used to develop transgenic crop plants in which selectable marker genes (SMG) are generally deployed to identify 'true' transformants. The commonly used SMG obtained from prokaryotic sources when employed in transgenic plants pose risks due to their lethal nature during selection process. In the recent past, some non-lethal SMGs have been identified and used for selection of transformants with increased precision and high selection efficiency. Considering the concerns related to bio-safety of the environment, it is desirable to remove the SMG in order to maximize the commercial success through wide adoption and public acceptance of genetically modified (GM) food crops. In this review, we examine the availability, and the suitability of wide range of non-lethal selection markers and elimination of SMG methods to develop marker-free transgenics for achieving global food security. As the strategies for marker-free plants are still in proof-of-concept stage, adaptation of new genomics tools for identification of novel non-lethal marker systems and its application for developing marker-free transgenics would further strengthen the crop improvement program.

  14. TRANSFORMER

    DOEpatents

    Baker, W.R.

    1959-08-25

    Transformers of a type adapted for use with extreme high power vacuum tubes where current requirements may be of the order of 2,000 to 200,000 amperes are described. The transformer casing has the form of a re-entrant section being extended through an opening in one end of the cylinder to form a coaxial terminal arrangement. A toroidal multi-turn primary winding is disposed within the casing in coaxial relationship therein. In a second embodiment, means are provided for forming the casing as a multi-turn secondary. The transformer is characterized by minimized resistance heating, minimized external magnetic flux, and an economical construction.

  15. School Finance in a Transformed Education System.

    ERIC Educational Resources Information Center

    Pipho, Chris

    1992-01-01

    Discussion of the impact of systemic educational change on the school funding mechanism addresses the forces driving school finance and examines new outcome-driven funding models. The implications of decentralization of school districts are considered; and advantages, disadvantages, and implementation of voucher and incentive models are presented.…

  16. Efficient Agrobacterium tumefaciens-mediated transformation and regeneration of garlic (Allium sativum) immature leaf tissue.

    PubMed

    Kenel, Fernand; Eady, Colin; Brinch, Sheree

    2010-03-01

    Transgenic garlic (Allium sativum) plants have been recovered directly from immature leaf material by selective culture following Agrobacterium-mediated transformation. This method involved the use of a binary vector containing the mgfp-ER reporter gene and hpt selectable marker, and followed a similar protocol developed previously for the transformation of immature onion embryos. The choice of tissue and post-transformation selection procedure resulted in a large increase in recovery of transgenic plants compared with previously confirmed allium transformation protocols. The presence of transgenes in the genome of the plants was confirmed using Southern analysis. This improvement in frequency and the use of clonal commercial "Printanor" germplasm now makes possible the integration of useful agronomic and quality traits into this crop. PMID:20099065

  17. Digital interface of electronic transformers based on embedded system

    NASA Astrophysics Data System (ADS)

    Shang, Qiufeng; Qi, Yincheng

    2008-10-01

    Benefited from digital interface of electronic transformers, information sharing and system integration in substation can be realized. An embedded system-based digital output scheme of electronic transformers is proposed. The digital interface is designed with S3C44B0X 32bit RISC microprocessor as the hardware platform. The μCLinux operation system (OS) is transplanted on ARM7 (S3C44B0X). Applying Ethernet technology as the communication mode in the substation automation system is a new trend. The network interface chip RTL8019AS is adopted. Data transmission is realized through the in-line TCP/IP protocol of uClinux embedded OS. The application result and character analysis show that the design can meet the real-time and reliability requirements of IEC60044-7/8 electronic voltage/current instrument transformer standards.

  18. Main transformation tools in ANALITIK-93 programming system

    SciTech Connect

    Gorovoi, V.D.; Klimenko, V.P.; Fishman, Yu.S.

    1995-09-01

    This article reviews the main functions incorporated in the latest version of the source language for the ANALITIK programming system. These functions are intended as transformation tools for nonnumerical models and problems, which are usually represented by systems of symbolic expressions of arbitrary length with a hierarchical structure. Such problems are often connected with nonlinear dependences and oscillating functions, which require multiple integration and differentiation, opening of parentheses, reduction of simmilar terms, various simplifications, etc. Such transformations are available in the languages of the ANALITIK family, REDUCE, Formac, Lisp, MACSYMA, Mathematica, and others. Languages of the ANALITIK family, and especially ANALITIK-93, are distinguished by greater generality and a higher level of intelligence, which is determined by an advanced automatic recognition system of functional properties of the objects being transformed and by a developed interactive system.

  19. Energy transformation in molecular electronic systems

    SciTech Connect

    Kasha, M.

    1985-07-25

    Our new optical pumping spectroscopy (steady state, and double-laser pulse) allows the production and study of the unstable rare tautomer in its ground and excited states, including picosecond dynamic studies. Molecules under study here included 7-azaindole (model for biological purines), 3-hydroxyflavone (model for plant flavones), lumichrome, and other heterocyclics. New detailed molecular mechanisms for proton transfer are derived, especially with catalytic assisting molecules. A new proton-transfer laser of extraordinary efficiency has become a side dividend, possibly worth of industrial development. The excited and highly reactive singlet molecular oxygen species /sup 1/..delta../sub g/) has proven to be ubiquitous in chemical peroxide systems and in physically excited sensitizer-oxygen systems. Hyperbaric oxygen mechanisms in biology probably involve singlet oxygen. We have undertaken a spectroscopic study of tris - dibenzoylmethane chelates of Al, Gd, Eu, and Yb trivalent ions. These chelates offer a variety of electronic behaviors, from Z-effects on ..pi..-electron spin-orbital coupling (Al, Gd) to Weissman intramolecular energy transfer to 4f mestable levels (Eu, Gd). Elegant new spectroscopic resolution at 77K permits separation of tautomeric, parasitic self-absorption, dissociation, and cage effects to be resolved. 18 refs., 4 figs.

  20. Energy transformation in molecular electronic systems

    NASA Astrophysics Data System (ADS)

    Kasha, M.

    1985-07-01

    Our new optical pumping spectroscopy allows the production and study of the unstable rate tautomer in its ground and excited states, including picosecond dynamic studies. Molecules under study here included 7-azaindole 3-hydroxyflavone, lumichrome, and other heterocyclics. New detailed molecular mechanisms for proton transfer are derived, especially with catalytic assisting molecules. A new proton-transfer laser of extraordinary efficiency has become a side dividend, possibly worthy of industrial development. The excited and highly reactive singlet molecular oxygen species (1) DELTA sub g has proven to be ubiquitous in chemical peroxide systems and in physically excited sensitizer-oxygen systems. Hyperbaric oxygen mechanisms in biology probably involve singlet oxygen. We have undertaken a spectroscopic study of trisdibenzoylmethane chelates of Al, Gd, Eu, and Yb trivalent ions. These chelates offer a variety of electronic behaviors, from Z-effects on (PI)--electron spin-orbital coupling (Al, Gd) to Weissman intramolecular energy transfer to 4f mestable levels (Eu, Gd). Elegant new spectroscopic resolution at 77K permits separation of tautomeric, parasitic self-absorption, dissociation, and cage effects to be resolved.

  1. Mechanisms of iron oxide transformation in hydrothermal systems.

    SciTech Connect

    Otake, Tsubasa; Wesolowski, David J; Anovitz, Lawrence {Larry} M

    2010-11-01

    Coexistence of magnetite and hematite in hydrothermal systems has often been used to constrain the redox potential of fluids, assuming that the redox equilibrium is attained among all minerals and aqueous species. However, as temperature decreases, disequilibrium mineral assemblages may occur due to the slow kinetics of reaction involving the minerals and fluids. In this study, we conducted a series of experiments in which hematite or magnetite was reacted with an acidic solution under H{sub 2}-rich hydrothermal conditions (T = 100-250 C, P{sub H{sub 2}} = 0.05-5 MPa) to investigate the kinetics of redox and non-redox transformations between hematite and magnetite, and the mechanisms of iron oxide transformation under hydrothermal conditions. The formation of euhedral crystals of hematite in 150 and 200 C experiments, in which magnetite was used as the starting material, indicates that non-redox transformation of magnetite to hematite occurred within 24 h. The chemical composition of the experimental solutions was controlled by the non-redox transformation between magnetite and hematite throughout the experiments. While solution compositions were controlled by the non-redox transformation in the first 3 days in a 250 C experiment, reductive dissolution of magnetite became important after 5 days and affected the solution chemistry. At 100 C, the presence of maghemite was indicated in the first 7 days. Based on these results, equilibrium constants of non-redox transformation between magnetite and hematite and those of non-redox transformation between magnetite and maghemite were calculated. Our results suggest that the redox transformation of hematite to magnetite occurs in the following steps: (1) reductive dissolution of hematite to Fe{sub (aq)}{sup 2+} and (2) non-redox transformation of hematite and Fe{sub (aq)}{sup 2+} to magnetite.

  2. Mechanisms of iron oxide transformations in hydrothermal systems.

    SciTech Connect

    Otake, Tsubasa; Wesolowski, David J; Anovitz, Lawrence {Larry} M; Allard Jr, Lawrence Frederick; Ohmoto, Hiroshi

    2010-01-01

    Coexistence of magnetite and hematite in hydrothermal systems has often been used to constrain the redox potential of fluids, assuming that the redox equilibrium is attained among all minerals and aqueous species. However, as temperature decreases, disequilibrium mineral assemblages may occur due to the slow kinetics of reaction involving the minerals and fluids. In this study, we conducted a series of experiments in which hematite or magnetite was reacted with an acidic solution under H2-rich hydrothermal conditions (T = 100 250 C,) to investigate the kinetics of redox and non-redox transformations between hematite and magnetite, and the mechanisms of iron oxide transformation under hydrothermal conditions. The formation of euhedral crystals of hematite in 150 and 200 C experiments, in which magnetite was used as the starting material, indicates that non-redox transformation of magnetite to hematite occurred within 24 h. The chemical composition of the experimental solutions was controlled by the non-redox transformation between magnetite and hematite throughout the experiments. While solution compositions were controlled by the non-redox transformation in the first 3 days in a 250 C experiment, reductive dissolution of magnetite became important after 5 days and affected the solution chemistry. At 100 C, the presence of maghemite was indicated in the first 7 days. Based on these results, equilibrium constants of non-redox transformation between magnetite and hematite and those of non-redox transformation between magnetite and maghemite were calculated. Our results suggest that the redox transformation of hematite to magnetite occurs in the following steps: (1) reductive dissolution of hematite to and (2) non-redox transformation of hematite and to magnetite.

  3. In planta transformation method for T-DNA transfer in orchids

    NASA Astrophysics Data System (ADS)

    Semiarti, Endang; Purwantoro, Aziz; Mercuriani, Ixora S.; Anggriasari, Anida M.; Jang, Seonghoe; Suhandono, Sony; Machida, Yasunori; Machida, Chiyoko

    2014-03-01

    Transgenic plant technology is an efficient tool to study the function of gene(s) in plant. The most popular and widely used technique is Agrobacterium-mediated transformation in which cocultivation was done by immersing the plant tissues/organ in overnight bacterial cultured for about 30 minutes to one hour under in vitro condition. In this experiment, we developed more easier technique that omitted the in vitro step during cocultivation with Agrobacterium, namely in planta transformation method. Pollinaria (compact pollen mass of orchid) of Phalaenopsis amabilis and Spathoglottis plicata orchids were used as target explants that were immersed into bacterial culture for 30 minutes, then dried up the pollinaria, the transformed pollinaria was used to pollinate orchid flowers. The T-DNA used for this experiments were Ubipro∷PaFT/A. tumefaciens GV3101 for P. amabilis and MeEF1α2 pro∷GUS/ A. tumefaciens LBA 4404 for S.plicata. Seeds that were produced from pollinated flowers were grown onto 10 mg/l hygromicin containing NP (New Phalaenopsis) medium. The existance of transgene in putative transformant protocorm (developing orchid embryo) genome was confirmed using PCR with specific primers of either PaFT or GUS genes. Histochemical GUS assay was also performed to the putative transformants. The result showed that transformation frequencies were 2.1 % in P. amabilis, and 0,53% in S. plicata. These results indicates that in planta transformation method could be used for Agrobacterium-mediated genetic transformation, with advantage easier and more secure work from contaminants than that of the in vitro method.

  4. Alternative variable transformation for simulation of multibody dynamic systems

    NASA Astrophysics Data System (ADS)

    Yang, Li-Farn

    1995-03-01

    An alternative variable transformation is proposed and analyzed for the simulation of multibody dynamic systems. The developed alternative variable transformation matrix intended to work directly with the multibody equations of motion without altering the inherent dynamic characteristics also eliminates the need for expensive computation of inversion of a large mass matrix required for the simulation of multibody dynamic systems. From the analytical derivation of an articulated multibody model, it was found that the computation can be saved (3n + 1/4)-fold due to the computational merits associated with the band matrix.

  5. SOFA and the algorithms for transformations between scales & between systems

    NASA Astrophysics Data System (ADS)

    Hohenkerk, C. Y.

    2012-12-01

    The SOFA Collection provides users with a myriad of routines. Examples will be given of the various transformations between the time scales TAI, TT, UTC, UT1, TDB, TCB and TCG, which represent the latest set of routines added to the SOFA collection. In particular this paper will also illustrate the combinations of routines that provide transformations between the Geocentric Celestial Reference System (GCRS) and the International Terrestrial Reference System (ITRS). Both the Celestial Intermediate Origin (CIO) and the equinox-based methods are shown, and those supporting International Astronomical Union (IAU) resolutions will be highlighted.

  6. Transformations in Air Transportation Systems For the 21st Century

    NASA Technical Reports Server (NTRS)

    Holmes, Bruce J.

    2004-01-01

    Globally, our transportation systems face increasingly discomforting realities: certain of the legacy air and ground infrastructures of the 20th century will not satisfy our 21st century mobility needs. The consequence of inaction is diminished quality of life and economic opportunity for those nations unable to transform from the 20th to 21st century systems. Clearly, new thinking is required regarding business models that cater to consumers value of time, airspace architectures that enable those new business models, and technology strategies for innovating at the system-of-networks level. This lecture proposes a structured way of thinking about transformation from the legacy systems of the 20th century toward new systems for the 21st century. The comparison and contrast between the legacy systems of the 20th century and the transformed systems of the 21st century provides insights into the structure of transformation of air transportation. Where the legacy systems tend to be analog (versus digital), centralized (versus distributed), and scheduled (versus on-demand) for example, transformed 21st century systems become capable of scalability through technological, business, and policy innovations. Where air mobility in our legacy systems of the 20th century brought economic opportunity and quality of life to large service markets, transformed air mobility of the 21st century becomes more equitable available to ever-thinner and widely distributed populations. Several technological developments in the traditional aircraft disciplines as well as in communication, navigation, surveillance and information systems create new foundations for 21st thinking about air transportation. One of the technological developments of importance arises from complexity science and modern network theory. Scale-free (i.e., scalable) networks represent a promising concept space for modeling airspace system architectures, and for assessing network performance in terms of robustness

  7. Transformers: Shape-Changing Space Systems Built with Robotic Textiles

    NASA Technical Reports Server (NTRS)

    Stoica, Adrian

    2013-01-01

    Prior approaches to transformer-like robots had only very limited success. They suffer from lack of reliability, ability to integrate large surfaces, and very modest change in overall shape. Robots can now be built from two-dimensional (2D) layers of robotic fabric. These transformers, a new kind of robotic space system, are dramatically different from current systems in at least two ways. First, the entire transformer is built from a single, thin sheet; a flexible layer of a robotic fabric (ro-fabric); or robotic textile (ro-textile). Second, the ro-textile layer is foldable to small volume and self-unfolding to adapt shape and function to mission phases.

  8. [ENHANCEMENT OF AGROBACTERIAL TRANSFORMATION OF PLANTS USING PROTEIN KINASE INHIBITORS TRIFLUOPERAZINE AND GENISTEIN].

    PubMed

    Yemets, A I; Fedorchuk, V V; Blume, Ya B

    2016-01-01

    The effect of different concentrations of protein tyrosine kinase inhibitor, genistein and serine/threonine protein kinase inhibitor, trifluoperazine, on the frequency of Agrobacterium-mediated transformation of leaf explants of N. tabacum was investigated. The influence of different concentrations of trifluoperazine in the range from 10 to 300 μM was investigated. It was found that 10 μM trifluoperazine provoked the increase of the frequency of agrobacterial transformation of tobacco leaf disks on 25%. In parallel, the influence of different concentrations of genistein in the range from 10 to 100 μM was investigated. It was found 100 μM genistein provoked the increase of the frequency of agrobacterial transformation of tobacco leaf disks on 12%.

  9. [ENHANCEMENT OF AGROBACTERIAL TRANSFORMATION OF PLANTS USING PROTEIN KINASE INHIBITORS TRIFLUOPERAZINE AND GENISTEIN].

    PubMed

    Yemets, A I; Fedorchuk, V V; Blume, Ya B

    2016-01-01

    The effect of different concentrations of protein tyrosine kinase inhibitor, genistein and serine/threonine protein kinase inhibitor, trifluoperazine, on the frequency of Agrobacterium-mediated transformation of leaf explants of N. tabacum was investigated. The influence of different concentrations of trifluoperazine in the range from 10 to 300 μM was investigated. It was found that 10 μM trifluoperazine provoked the increase of the frequency of agrobacterial transformation of tobacco leaf disks on 25%. In parallel, the influence of different concentrations of genistein in the range from 10 to 100 μM was investigated. It was found 100 μM genistein provoked the increase of the frequency of agrobacterial transformation of tobacco leaf disks on 12%. PMID:27266180

  10. Transforming Domain Knowledge: A Systemic View at the School Curriculum

    ERIC Educational Resources Information Center

    Hameyer, Uwe

    2007-01-01

    "What happens to domain knowledge on its way into the school curriculum?" The article explores this question from a systems view of the knowledge transformation process. This process is not only bound to "contextual knowledge" about "social forces" but also to the school's own "educational knowledge" framework. Further, utilizing knowledge depends…

  11. A Superconducting transformer system for high current cable testing

    SciTech Connect

    Godeke, A.; Dietderich, D. R.; Joseph, J. M.; Lizarazo, J.; Prestemon, S. O.; Miller, G.; Weijers, H. W.

    2010-02-15

    This article describes the development of a direct-current (dc) superconducting transformer system for the high current test of superconducting cables. The transformer consists of a core-free 10 464 turn primary solenoid which is enclosed by a 6.5 turn secondary. The transformer is designed to deliver a 50 kA dc secondary current at a dc primary current of about 50 A. The secondary current is measured inductively using two toroidal-wound Rogowski coils. The Rogowski coil signal is digitally integrated, resulting in a voltage signal that is proportional to the secondary current. This voltage signal is used to control the secondary current using a feedback loop which automatically compensates for resistive losses in the splices to the superconducting cable samples that are connected to the secondary. The system has been commissioned up to 28 kA secondary current. The reproducibility in the secondary current measurement is better than 0.05% for the relevant current range up to 25 kA. The drift in the secondary current, which results from drift in the digital integrator, is estimated to be below 0.5 A/min. The system's performance is further demonstrated through a voltage-current measurement on a superconducting cable sample at 11 T background magnetic field. The superconducting transformer system enables fast, high resolution, economic, and safe tests of the critical current of superconducting cable samples.

  12. The AECT FutureMinds Initiative: Transforming America's School Systems

    ERIC Educational Resources Information Center

    Reigeluth, Charles M., Ed.; Duffy, Francis M.

    2008-01-01

    This article opens by discussing what paradigm change is and why it is needed. Then it describes the new AECT initiative, "FutureMinds: Transforming America's School Systems," beginning with its purpose (to help state departments of education to facilitate paradigm change in school districts), the fundamental ideas underlying the initiative (e.g.,…

  13. Transformation and Transport Processes of Nitrogen in Agricultural Systems

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The transformation and transport processes of nitrogen (N) in agricultural systems are discussed and information is provided on overall reservoir sizes for N. Nitrogen is ubiquitous in the environment and is required for the survival of all living things. It is also one of the most important essen...

  14. Model Transformation for a System of Systems Dependability Safety Case

    NASA Technical Reports Server (NTRS)

    Murphy, Judy; Driskell, Steve

    2011-01-01

    The presentation reviews the dependability and safety effort of NASA's Independent Verification and Validation Facility. Topics include: safety engineering process, applications to non-space environment, Phase I overview, process creation, sample SRM artifact, Phase I end result, Phase II model transformation, fault management, and applying Phase II to individual projects.

  15. Fourier Transform for Fermionic Systems and the Spectral Tensor Network

    NASA Astrophysics Data System (ADS)

    Ferris, Andrew J.

    2014-07-01

    Leveraging the decomposability of the fast Fourier transform, I propose a new class of tensor network that is efficiently contractible and able to represent many-body systems with local entanglement that is greater than the area law. Translationally invariant systems of free fermions in arbitrary dimensions as well as 1D systems solved by the Jordan-Wigner transformation are shown to be exactly represented in this class. Further, it is proposed that these tensor networks be used as generic structures to variationally describe more complicated systems, such as interacting fermions. This class shares some similarities with the Evenbly-Vidal branching multiscale entanglement renormalization ansatz, but with some important differences and greatly reduced computational demands.

  16. Linear transformation and oscillation criteria for Hamiltonian systems

    NASA Astrophysics Data System (ADS)

    Zheng, Zhaowen

    2007-08-01

    Using a linear transformation similar to the Kummer transformation, some new oscillation criteria for linear Hamiltonian systems are established. These results generalize and improve the oscillation criteria due to I.S. Kumari and S. Umanaheswaram [I. Sowjaya Kumari, S. Umanaheswaram, Oscillation criteria for linear matrix Hamiltonian systems, J. Differential Equations 165 (2000) 174-198], Q. Yang et al. [Q. Yang, R. Mathsen, S. Zhu, Oscillation theorems for self-adjoint matrix Hamiltonian systems, J. Differential Equations 190 (2003) 306-329], and S. Chen and Z. Zheng [Shaozhu Chen, Zhaowen Zheng, Oscillation criteria of Yan type for linear Hamiltonian systems, Comput. Math. Appl. 46 (2003) 855-862]. These criteria also unify many of known criteria in literature and simplify the proofs.

  17. Fourier transform for fermionic systems and the spectral tensor network.

    PubMed

    Ferris, Andrew J

    2014-07-01

    Leveraging the decomposability of the fast Fourier transform, I propose a new class of tensor network that is efficiently contractible and able to represent many-body systems with local entanglement that is greater than the area law. Translationally invariant systems of free fermions in arbitrary dimensions as well as 1D systems solved by the Jordan-Wigner transformation are shown to be exactly represented in this class. Further, it is proposed that these tensor networks be used as generic structures to variationally describe more complicated systems, such as interacting fermions. This class shares some similarities with the Evenbly-Vidal branching multiscale entanglement renormalization ansatz, but with some important differences and greatly reduced computational demands.

  18. Fourier transform digital holographic adaptive optics imaging system

    PubMed Central

    Liu, Changgeng; Yu, Xiao; Kim, Myung K.

    2013-01-01

    A Fourier transform digital holographic adaptive optics imaging system and its basic principles are proposed. The CCD is put at the exact Fourier transform plane of the pupil of the eye lens. The spherical curvature introduced by the optics except the eye lens itself is eliminated. The CCD is also at image plane of the target. The point-spread function of the system is directly recorded, making it easier to determine the correct guide-star hologram. Also, the light signal will be stronger at the CCD, especially for phase-aberration sensing. Numerical propagation is avoided. The sensor aperture has nothing to do with the resolution and the possibility of using low coherence or incoherent illumination is opened. The system becomes more efficient and flexible. Although it is intended for ophthalmic use, it also shows potential application in microscopy. The robustness and feasibility of this compact system are demonstrated by simulations and experiments using scattering objects. PMID:23262541

  19. Stability and instability in an evolving oceanic transform fault system

    NASA Astrophysics Data System (ADS)

    Forsyth, D. W.; Harmon, N.; Pickle, R. C.; Saal, A.

    2006-12-01

    The Quebrada/Discovery/Gofar fracture zone system at 4 S on the East Pacific Rise currently comprises 9 active en echelon transform traces separated by 8 intratransform spreading centers. A detailed multi-beam bathymetry, side-scan sonar, magnetic and gravity anomaly survey in spring 2006 allows us to reconstruct the history of the transform/ridge geometry over the last 1 million years. Dredging of fresh basaltic glasses and the distribution of transform seismicity helps to unambiguously identify the current spreading centers. Some parts of the system are surprisingly stable; the length and offset between the spreading centers in the Quebrada fracture zone have been nearly constant for more than 700 kyr despite one of the spreading centers being only 5 km long. Other transform traces have died and at least two new ones have started within the last 1 Myr. Rather than being a response to the changing direction of plate motion, the specific changes in geometry seem to be triggered by local changes in volcanic activity. For example, one new spreading center and an associated transform trace began at the site of a rare (in this area), off-axis seamount that may have weakened the lithosphere. Some ridge segments become more "robust" and extend in length, eliminating transforms. The region between the Quebrada and Discovery fracture zones has had a very complex, recent history. The current 30-km-long spreading center has a median valley, unlike typical East Pacific Rise spreading centers. Spreading on that center appears to be slower than average due to an unusual form of overlapping spreading. Although the center is clearly bounded by transforms, at the western end of the Discovery transform, occasional dikes apparently extend north from the Discovery/Gofar spreading center into the adjacent, older seafloor, accommodating some of the spreading that would normally take place on the Quebrada/Discovery center. Overlapping spreading has trapped and rotated crustal blocks

  20. An accurate continuous calibration system for high voltage current transformer

    NASA Astrophysics Data System (ADS)

    Tong, Yue; Li, Bin Hong

    2011-02-01

    A continuous calibration system for high voltage current transformers is presented in this paper. The sensor of this system is based on a kind of electronic instrument current transformer, which is a clamp-shape air core coil. This system uses an optical fiber transmission system for its signal transmission and power supply. Finally the digital integrator and fourth-order convolution window algorithm as error calculation methods are realized by the virtual instrument with a personal computer. It is found that this system can calibrate a high voltage current transformer while energized, which means avoiding a long calibrating period in the power system and the loss of power metering expense. At the same time, it has a wide dynamic range and frequency band, and it can achieve a high accuracy measurement in a complex electromagnetic field environment. The experimental results and the on-site operation results presented in the last part of the paper, prove that it can reach the 0.05 accuracy class and is easy to operate on site.

  1. Transformational System Concepts and Technologies for Our Future in Space

    NASA Technical Reports Server (NTRS)

    Howell, Joe T.; Mankins, John C.

    2004-01-01

    Continued constrained budgets and growing national and international interests in the commercialization and development of space requires NASA to be constantly vigilant, to be creative, and to seize every opportunity for assuring the maximum return on space infrastructure investments. Accordingly, efforts are underway to forge new and innovative approaches to transform our space systems in the future to ultimately achieve two or three or five times as much with the same resources. This bold undertaking can be achieved only through extensive cooperative efforts throughout the aerospace community and truly effective planning to pursue advanced space system design concepts and high-risk/high-leverage research and technology. Definitive implementation strategies and roadmaps containing new methodologies and revolutionary approaches must be developed to economically accommodate the continued exploration and development of space. Transformation can be realized through modular design and stepping stone development. This approach involves sustainable budget levels and multi-purpose systems development of supporting capabilities that lead to a diverse amy of sustainable future space activities. Transformational design and development requires revolutionary advances by using modular designs and a planned, stepping stone development process. A modular approach to space systems potentially offers many improvements over traditional one-of-a-kind space systems comprised of different subsystem element with little standardization in interfaces or functionality. Modular systems must be more flexible, scaleable, reconfigurable, and evolvable. Costs can be reduced through learning curve effects and economies of scale, and by enabling servicing and repair that would not otherwise be feasible. This paper briefly discusses achieving a promising approach to transforming space systems planning and evolution into a meaningful stepping stone design, development, and implementation process

  2. On the Hilbert-Huang Transform Data Processing System Development

    NASA Technical Reports Server (NTRS)

    Kizhner, Semion; Flatley, Thomas P.; Huang, Norden E.; Cornwell, Evette; Smith, Darell

    2003-01-01

    One of the main heritage tools used in scientific and engineering data spectrum analysis is the Fourier Integral Transform and its high performance digital equivalent - the Fast Fourier Transform (FFT). The Fourier view of nonlinear mechanics that had existed for a long time, and the associated FFT (fairly recent development), carry strong a-priori assumptions about the source data, such as linearity and of being stationary. Natural phenomena measurements are essentially nonlinear and nonstationary. A very recent development at the National Aeronautics and Space Administration (NASA) Goddard Space Flight Center (GSFC), known as the Hilbert-Huang Transform (HHT) proposes a novel approach to the solution for the nonlinear class of spectrum analysis problems. Using the Empirical Mode Decomposition (EMD) followed by the Hilbert Transform of the empirical decomposition data (HT), the HHT allows spectrum analysis of nonlinear and nonstationary data by using an engineering a-posteriori data processing, based on the EMD algorithm. This results in a non-constrained decomposition of a source real value data vector into a finite set of Intrinsic Mode Functions (IMF) that can be further analyzed for spectrum interpretation by the classical Hilbert Transform. This paper describes phase one of the development of a new engineering tool, the HHT Data Processing System (HHTDPS). The HHTDPS allows applying the "T to a data vector in a fashion similar to the heritage FFT. It is a generic, low cost, high performance personal computer (PC) based system that implements the HHT computational algorithms in a user friendly, file driven environment. This paper also presents a quantitative analysis for a complex waveform data sample, a summary of technology commercialization efforts and the lessons learned from this new technology development.

  3. Transformational Systems Concepts and Technologies for Our Future in Space

    NASA Technical Reports Server (NTRS)

    Howell, J. T.; George,P.; Mankins, J. C. (Editor); Christensen, C. B.

    2004-01-01

    NASA is constantly searching for new ideas and approaches yielding opportunities for assuring maximum returns on space infrastructure investments. Perhaps the idea of transformational innovation in developing space systems is long overdue. However, the concept of utilizing modular space system designs combined with stepping-stone development processes has merit and promises to return several times the original investment since each new space system or component is not treated as a unique and/or discrete design and development challenge. New space systems can be planned and designed so that each builds on the technology of previous systems and provides capabilities to support future advanced systems. Subsystems can be designed to use common modular components and achieve economies of scale, production, and operation. Standards, interoperability, and "plug and play" capabilities, when implemented vigorously and consistently, will result in systems that can be upgraded effectively with new technologies. This workshop explored many building-block approaches via way of example across a broad spectrum of technology discipline areas for potentially transforming space systems and inspiring future innovation. Details describing the workshop structure, process, and results are contained in this Conference Publication.

  4. Classification of Transient Phenomena in Distribution System using wavelet Transform

    NASA Astrophysics Data System (ADS)

    Sedighi, Alireza

    2014-05-01

    An efficient procedure for classification of transient phenomena in distribution systems is proposed in this paper. The proposed method has been applied to classify some transient phenomena such as inrush current, load switching, capacitor switching and single phase to ground fault. The new scheme is based on wavelet transform algorithm. All of the events for feature extraction and test are simulated using Electro Magnetic Transient Program (EMTP). Results show high accuracy of proposed method.

  5. Transformation of the Adult Education System in Slovakia

    NASA Astrophysics Data System (ADS)

    Švec, Štefan

    1998-07-01

    This article examines trends in adult education in Slovakia since it became a separate republic in 1993. Economic and social transformations during this period have led to a re-thinking of the adult education system. The author describes four basic modalities for providing adult education in Slovakia: (1) schools and colleges; (2) cultural centres and similar institutions; (3) institutions for vocational training; (4) voluntary organizations such as trade unions, political parties and ethnic minority groups.

  6. Comparison of Soybean Transformation Efficiency and Plant Factors Affecting Transformation during the Agrobacterium Infection Process

    PubMed Central

    Jia, Yuying; Yao, Xingdong; Zhao, Mingzhe; Zhao, Qiang; Du, Yanli; Yu, Cuimei; Xie, Futi

    2015-01-01

    The susceptibility of soybean genotype to Agrobacterium infection is a key factor for the high level of genetic transformation efficiency. The objective of this study is to evaluate the plant factors related to transformation in cotyledonary nodes during the Agrobacterium infection process. This study selected three genotypes (Williams 82, Shennong 9 and Bert) with high transformation efficiency, which presented better susceptibility to Agrobacterium infection, and three low transformation efficiency genotypes (General, Liaodou 16 and Kottman), which showed a relatively weak susceptibility. Gibberellin (GA) levels and soybean GA20ox2 and CYP707A2 transcripts of high-efficiency genotypes increased and were higher than those of low-efficiency genotypes; however, the opposite performance was shown in abscisic acid (ABA). Higher zeatin riboside (ZR) content and DNA quantity, and relatively higher expression of soybean IPT5, CYCD3 and CYCA3 were obtained in high-efficiency genotypes. High-efficiency genotypes had low methyl jasmonate (MeJA) content, polyphenol oxidase (PPO) and peroxidase (POD) activity, and relatively lower expression of soybean OPR3, PPO1 and PRX71. GA and ZR were positive plant factors for Agrobacterium-mediated soybean transformation by facilitating germination and growth, and increasing the number of cells in DNA synthesis cycle, respectively; MeJA, PPO, POD and ABA were negative plant factors by inducing defence reactions and repressing germination and growth, respectively. PMID:26262617

  7. Comparison of Soybean Transformation Efficiency and Plant Factors Affecting Transformation during the Agrobacterium Infection Process.

    PubMed

    Jia, Yuying; Yao, Xingdong; Zhao, Mingzhe; Zhao, Qiang; Du, Yanli; Yu, Cuimei; Xie, Futi

    2015-08-07

    The susceptibility of soybean genotype to Agrobacterium infection is a key factor for the high level of genetic transformation efficiency. The objective of this study is to evaluate the plant factors related to transformation in cotyledonary nodes during the Agrobacterium infection process. This study selected three genotypes (Williams 82, Shennong 9 and Bert) with high transformation efficiency, which presented better susceptibility to Agrobacterium infection, and three low transformation efficiency genotypes (General, Liaodou 16 and Kottman), which showed a relatively weak susceptibility. Gibberellin (GA) levels and soybean GA20ox2 and CYP707A2 transcripts of high-efficiency genotypes increased and were higher than those of low-efficiency genotypes; however, the opposite performance was shown in abscisic acid (ABA). Higher zeatin riboside (ZR) content and DNA quantity, and relatively higher expression of soybean IPT5, CYCD3 and CYCA3 were obtained in high-efficiency genotypes. High-efficiency genotypes had low methyl jasmonate (MeJA) content, polyphenol oxidase (PPO) and peroxidase (POD) activity, and relatively lower expression of soybean OPR3, PPO1 and PRX71. GA and ZR were positive plant factors for Agrobacterium-mediated soybean transformation by facilitating germination and growth, and increasing the number of cells in DNA synthesis cycle, respectively; MeJA, PPO, POD and ABA were negative plant factors by inducing defence reactions and repressing germination and growth, respectively.

  8. Leadership and transformational change in healthcare organisations: a qualitative analysis of the North East Transformation System.

    PubMed

    Erskine, Jonathan; Hunter, David J; Small, Adrian; Hicks, Chris; McGovern, Tom; Lugsden, Ed; Whitty, Paula; Steen, Nick; Eccles, Martin Paul

    2013-02-01

    The research project 'An Evaluation of Transformational Change in NHS North East' examines the progress and success of National Health Service (NHS) organisations in north east England in implementing and embedding the North East Transformation System (NETS), a region-wide programme to improve healthcare quality and safety, and to reduce waste, using a combination of Vision, Compact, and Lean-based Method. This paper concentrates on findings concerning the role of leadership in enabling tranformational change, based on semi-structured interviews with a mix of senior NHS managers and quality improvement staff in 14 study sites. Most interviewees felt that implementing the NETS requires committed, stable leadership, attention to team-building across disciplines and leadership development at many levels. We conclude that without senior leader commitment to continuous improvement over a long time scale and serious efforts to distribute leadership tasks to all levels, healthcare organisations are less likely to achieve positive changes in managerial-clinical relations, sustainable improvements to organisational culture and, ultimately, the region-wide step change in quality, safety and efficiency that the NETS was designed to deliver.

  9. Leadership and transformational change in healthcare organisations: a qualitative analysis of the North East Transformation System.

    PubMed

    Erskine, Jonathan; Hunter, David J; Small, Adrian; Hicks, Chris; McGovern, Tom; Lugsden, Ed; Whitty, Paula; Steen, Nick; Eccles, Martin Paul

    2013-02-01

    The research project 'An Evaluation of Transformational Change in NHS North East' examines the progress and success of National Health Service (NHS) organisations in north east England in implementing and embedding the North East Transformation System (NETS), a region-wide programme to improve healthcare quality and safety, and to reduce waste, using a combination of Vision, Compact, and Lean-based Method. This paper concentrates on findings concerning the role of leadership in enabling tranformational change, based on semi-structured interviews with a mix of senior NHS managers and quality improvement staff in 14 study sites. Most interviewees felt that implementing the NETS requires committed, stable leadership, attention to team-building across disciplines and leadership development at many levels. We conclude that without senior leader commitment to continuous improvement over a long time scale and serious efforts to distribute leadership tasks to all levels, healthcare organisations are less likely to achieve positive changes in managerial-clinical relations, sustainable improvements to organisational culture and, ultimately, the region-wide step change in quality, safety and efficiency that the NETS was designed to deliver. PMID:25594999

  10. Extreme makeover: Transformation of the veterans health care system.

    PubMed

    Kizer, Kenneth W; Dudley, R Adams

    2009-01-01

    The veterans health care system administered by the U.S. Department of Veterans Affairs (VA) was established after World War I to provide health care for veterans who suffered from conditions related to their military service. It has grown to be the nation's largest integrated health care system. As the system grew, a number of factors contributed to its becoming increasingly dysfunctional. By the mid-1990s, VA health care was widely criticized for providing fragmented and disjointed care of unpredictable and irregular quality, which was expensive, difficult to access, and insensitive to individual needs. Between 1995 and 1999, the VA health care system was reengineered, focusing especially on management accountability, care coordination, quality improvement, resource allocation, and information management. Numerous systemic changes were implemented, producing dramatically improved quality, service, and operational efficiency. VA health care is now considered among the best in America, and the VA transformation is viewed as a model for health care reform. PMID:19296778

  11. Large-System Transformation in Health Care: A Realist Review

    PubMed Central

    Best, Allan; Greenhalgh, Trisha; Lewis, Steven; Saul, Jessie E; Carroll, Simon; Bitz, Jennifer

    2012-01-01

    Context An evidence base that addresses issues of complexity and context is urgently needed for large-system transformation (LST) and health care reform. Fundamental conceptual and methodological challenges also must be addressed. The Saskatchewan Ministry of Health in Canada requested a six-month synthesis project to guide four major policy development and strategy initiatives focused on patient- and family-centered care, primary health care renewal, quality improvement, and surgical wait lists. The aims of the review were to analyze examples of successful and less successful transformation initiatives, to synthesize knowledge of the underlying mechanisms, to clarify the role of government, and to outline options for evaluation. Methods We used realist review, whose working assumption is that a particular intervention triggers particular mechanisms of change. Mechanisms may be more or less effective in producing their intended outcomes, depending on their interaction with various contextual factors. We explain the variations in outcome as the interplay between context and mechanisms. We nested this analytic approach in a macro framing of complex adaptive systems (CAS). Findings Our rapid realist review identified five “simple rules” of LST that were likely to enhance the success of the target initiatives: (1) blend designated leadership with distributed leadership; (2) establish feedback loops; (3) attend to history; (4) engage physicians; and (5) include patients and families. These principles play out differently in different contexts affecting human behavior (and thereby contributing to change) through a wide range of different mechanisms. Conclusions Realist review methodology can be applied in combination with a complex system lens on published literature to produce a knowledge synthesis that informs a prospective change effort in large-system transformation. A collaborative process engaging both research producers and research users contributes to local

  12. Transformational derivation of programs using the Focus system

    NASA Technical Reports Server (NTRS)

    Reddy, Uday S.

    1988-01-01

    A program derivation support system called Focus is being constructed. It will formally derive programs using the paradigm of program transformation. The following issues are discussed: (1) the integration of validation and program derivation activities in the Focus system; (2) its tree-based user interface; (3) the control of search spaces in program derivation; and (4) the structure and organization of program derivation records. The inference procedures of the system are based on the integration of functional and logic programming principles. This brings about a synthesis of paradigms that were heretofore considered far apart, such as logical and executable specifications and constructive and transformational approaches to program derivation. A great emphasis has been placed, in the design of Focus, on achieving small search spaces during program derivation. The program manipulation operations such as expansion, simplification and rewriting were designed with this objective. The role of operations that are expensive in search spaces, such as folding, has been reduced. Program derivations are documented in Focus in a way that the high level descriptions of derivations are expressed only using program level information. All the meta-level information, together with dependencies between derivations of program components, is automatically recorded by the system at a lower level of description for its own use in replay.

  13. Initiating nuclear-chemical transformations in native systems: Phenomenology

    NASA Astrophysics Data System (ADS)

    Timashev, S. F.

    2016-10-01

    A possible mechanism of nuclear transformations in biological systems in vivo is proposed. Reasons why there is no ionizing radiation that could be detrimental to native systems during the corresponding nuclear reactions are given. It is established that the initial stage of these processes is associated with that of ATP hydrolysis, which initiates the action of the inner-shell electron of an atom participating in the reaction on its nucleus according to the mechanism of weak nuclear interaction. This results in the formation of a nucleus in a metastable state with a disturbed nucleon structure and a charge one unit lower than that of the initial nucleus. It is also assumed that the atom participating in the reaction is adsorbed near the mouth of one of the transport ATPases in the cell's cytoplasmic membrane, and the reason for the initiating impact the electron has on the nucleus is due to the emergence of a local electric field formed during ATP hydrolysis near the ion channel of a donor-acceptor pair of charges that is opposite to the direction of the average membrane field. It is concluded that as a result of the key role of weak nuclear interaction in these processes, the energy of nuclear transformations in biological systems in vivo is released through the emission of neutrino-antineutrino pairs that are harmless to living organisms.

  14. Module Ten: Transformers; Basic Electricity and Electronics Individualized Learning System.

    ERIC Educational Resources Information Center

    Bureau of Naval Personnel, Washington, DC.

    The module introduces a very important electrical device, the transformer. The module is divided into six lessons: transformer construction, transformer theory and operation, turns and voltage ratios, power and current, transformer efficiency, and semiconductor rectifiers. Each lesson consists of an overview, a list of study resources, lesson…

  15. [Influence of collimation system on static Fourier transform spectrometer].

    PubMed

    Jiang, Cheng-Zhi; Liang, Jing-Qiu; Liang, Zhong-Zhu; Sun, Qiang; Wang, Wei-Biao

    2014-01-01

    Collimation system provides collimated light for the static Fourier-transform spectroscopy (SFTS). Its quality is crucial to the signal to noise ratio (SNR) of SFTS. In the present paper, the physical model of SFTS was established based on the Fresnel diffraction theory by means of numerical software. The influence of collimation system on the SFTS was discussed in detail focusing on the aberrations of collimation lens and the quality of extended source. The results of simulation show that the influences of different kinds of aberrations on SNR take on obvious regularity, and in particular, the influences of off-axis aberrations on SNR are closely related to the location of off-axis point source. Finally the extended source's maximum radius allowed was obtained by simulation, which equals to 0.65 mm. The discussion results will be used for the design of collimation system.

  16. Status Report on Power System Transformation: A 21st Century Power Partnership Report

    SciTech Connect

    Miller, Mackay; Martinot, Eric; Cox, Sadie; Speer, Bethany; Zinaman, Owen; Booth, Sam; Zissler, Romain; Cochran, Jaquelin; Soonee, S. K.; Audinet, Pierre; Munuera, Luis; Arent, Doug

    2015-05-27

    This report has three primary goals: (1) to articulate the concept of power system transformation; (2) to explore the current global landscape of ‘innovations’ that constitute power system transformation and provide evidence of how these innovations are emerging; and (3) to suggest an analytical framework for assessing the status of power system transformation on an on-going basis.

  17. The Gradual Transformation of the Polish Public Science System.

    PubMed

    Heinecke, Steffi

    2016-01-01

    This paper investigates institutional change in the Polish public science system (PPSS) in the past twenty years. Employing macro-statistical data, the paper argues that this change process has unfolded stepwise and relatively late despite major political and economic transformations in post-socialist Poland. Using a historical-institutionalist perspective, the paper focuses on processes of institutional change, including layering, displacement, and dismantling. One major finding is that the speed and depth of the gradual transformation differs considerably between the three research performing sectors of the Polish public science system. As the Polish Academy of Sciences was reproduced institutionally, the former governmental units for applied R&D were partly dismantled and displaced by private sector R&D units. In contrast, the Higher Education sector underwent a strong expansion and, thus, layering of new research activities and fields. Since policy shifts within the PPSS occurred relatively late, the more than two decades following the collapse of communism are of special interest to scholars of incremental, yet cumulative, institutional change.

  18. Quantitative transformation for implementation of adder circuits in physical systems.

    PubMed

    Jones, Jeff; Whiting, James G H; Adamatzky, Andrew

    2015-08-01

    Computing devices are composed of spatial arrangements of simple fundamental logic gates. These gates may be combined to form more complex adding circuits and, ultimately, complete computer systems. Implementing classical adding circuits using unconventional, or even living substrates such as slime mould Physarum polycephalum, is made difficult and often impractical by the challenges of branching fan-out of inputs and regions where circuit lines must cross without interference. In this report we explore whether it is possible to avoid spatial propagation, branching and crossing completely in the design of adding circuits. We analyse the input and output patterns of a single-bit full adder circuit. A simple quantitative transformation of the input patterns which considers the total number of bits in the input string allows us to map the respective input combinations to the correct outputs patterns of the full adder circuit, reducing the circuit combinations from a 2:1 mapping to a 1:1 mapping. The mapping of inputs to outputs also shows an incremental linear progression, suggesting its implementation in a range of physical systems. We demonstrate an example implementation, first in simulation, inspired by self-oscillatory dynamics of the acellular slime mould P. polycephalum. We then assess the potential implementation using plasmodium of slime mould itself. This simple transformation may enrich the potential for using unconventional computing substrates to implement digital circuits.

  19. The Gradual Transformation of the Polish Public Science System.

    PubMed

    Heinecke, Steffi

    2016-01-01

    This paper investigates institutional change in the Polish public science system (PPSS) in the past twenty years. Employing macro-statistical data, the paper argues that this change process has unfolded stepwise and relatively late despite major political and economic transformations in post-socialist Poland. Using a historical-institutionalist perspective, the paper focuses on processes of institutional change, including layering, displacement, and dismantling. One major finding is that the speed and depth of the gradual transformation differs considerably between the three research performing sectors of the Polish public science system. As the Polish Academy of Sciences was reproduced institutionally, the former governmental units for applied R&D were partly dismantled and displaced by private sector R&D units. In contrast, the Higher Education sector underwent a strong expansion and, thus, layering of new research activities and fields. Since policy shifts within the PPSS occurred relatively late, the more than two decades following the collapse of communism are of special interest to scholars of incremental, yet cumulative, institutional change. PMID:27077386

  20. Quantitative transformation for implementation of adder circuits in physical systems.

    PubMed

    Jones, Jeff; Whiting, James G H; Adamatzky, Andrew

    2015-08-01

    Computing devices are composed of spatial arrangements of simple fundamental logic gates. These gates may be combined to form more complex adding circuits and, ultimately, complete computer systems. Implementing classical adding circuits using unconventional, or even living substrates such as slime mould Physarum polycephalum, is made difficult and often impractical by the challenges of branching fan-out of inputs and regions where circuit lines must cross without interference. In this report we explore whether it is possible to avoid spatial propagation, branching and crossing completely in the design of adding circuits. We analyse the input and output patterns of a single-bit full adder circuit. A simple quantitative transformation of the input patterns which considers the total number of bits in the input string allows us to map the respective input combinations to the correct outputs patterns of the full adder circuit, reducing the circuit combinations from a 2:1 mapping to a 1:1 mapping. The mapping of inputs to outputs also shows an incremental linear progression, suggesting its implementation in a range of physical systems. We demonstrate an example implementation, first in simulation, inspired by self-oscillatory dynamics of the acellular slime mould P. polycephalum. We then assess the potential implementation using plasmodium of slime mould itself. This simple transformation may enrich the potential for using unconventional computing substrates to implement digital circuits. PMID:26007225

  1. The Gradual Transformation of the Polish Public Science System

    PubMed Central

    Heinecke, Steffi

    2016-01-01

    This paper investigates institutional change in the Polish public science system (PPSS) in the past twenty years. Employing macro-statistical data, the paper argues that this change process has unfolded stepwise and relatively late despite major political and economic transformations in post-socialist Poland. Using a historical-institutionalist perspective, the paper focuses on processes of institutional change, including layering, displacement, and dismantling. One major finding is that the speed and depth of the gradual transformation differs considerably between the three research performing sectors of the Polish public science system. As the Polish Academy of Sciences was reproduced institutionally, the former governmental units for applied R&D were partly dismantled and displaced by private sector R&D units. In contrast, the Higher Education sector underwent a strong expansion and, thus, layering of new research activities and fields. Since policy shifts within the PPSS occurred relatively late, the more than two decades following the collapse of communism are of special interest to scholars of incremental, yet cumulative, institutional change. PMID:27077386

  2. Establishment of invasive and non-invasive reporter systems to investigate American elm-Ophiostoma novo-ulmi interactions.

    PubMed

    Sherif, S; Jones, A M P; Shukla, M R; Saxena, P K

    2014-10-01

    Dutch elm disease (DED), caused by ascomycete fungi in the Ophiostoma genus, is the most devastating disease of American elm (Ulmus americana) trees. Cerato ulmin (CU), a hydrophobin secreted by the fungus, has been implicated in the development of DED, but its role in fungal pathogenicity and virulence remains uncertain and controversial. Here, we describe reporter systems based on the CU promoter and three reporter proteins (GFP, GUS and LUC), developed as research tools for quantitative and qualitative studies of DED in vitro, in vivo and in planta. A strain of the aggressive species Ophiostoma novo-ulmi was transformed with the reporter constructs using Agrobacterium-mediated transformation and the fungal transformants, namely M75-GFP, M75-GUS and M75-LUC, were examined for mitotic stability after repeated subcultures. The intensity of GFP fluorescence was strong in M75-GFP spores and hyphae, allowing microscopic investigations of spore structure, fungal morphogenesis and fungal development. The interaction of M75-GFP and U. americana callus cells was explored with scanning laser confocal microscopy facilitating qualitative studies on fungal strategies for the invasion and penetration of elm cells. M75-GUS was generated to provide an invasive, yet quantitative approach to study fungal-plant interactions in vitro and in planta. The generation of M75-LUC transformants was aimed at providing a non-destructive quantitative approach to study the role of CU in vivo. The sensitivity, low background signal and linearity of LUC assays all predict a very reliable approach to investigate and re-test previously claimed roles of this CU in fungal pathogenicity. These reporter systems provide new tools to investigate plant-pathogen interactions in this complex pathosystem and may aid in better understanding the development of DED. PMID:25139300

  3. Optimization of beam transformation system for laser-diode bars.

    PubMed

    Yu, Junhong; Guo, Linhui; Wu, Hualing; Wang, Zhao; Gao, Songxin; Wu, Deyong

    2016-08-22

    An optimized beam transformation system (BTS) is proposed to improve the beam quality of laser-diode bars. Through this optimized design, the deterioration of beam quality after the BTS can be significantly reduced. Both the simulation and experimental results demonstrate that the optimized system enables the beam quality of a mini-bar (9 emitters) approximately equal to 5.0 mm × 3.6 mrad in the fast-axis and slow-axis. After beam shaping by the optimized BTS, the laser-diode beam can be coupled into a 100 μm core, 0.15 numerical aperture (NA) fiber with an output power of over 100 W and an electric-optical efficiency of 46.8%.

  4. Optimization of beam transformation system for laser-diode bars.

    PubMed

    Yu, Junhong; Guo, Linhui; Wu, Hualing; Wang, Zhao; Gao, Songxin; Wu, Deyong

    2016-08-22

    An optimized beam transformation system (BTS) is proposed to improve the beam quality of laser-diode bars. Through this optimized design, the deterioration of beam quality after the BTS can be significantly reduced. Both the simulation and experimental results demonstrate that the optimized system enables the beam quality of a mini-bar (9 emitters) approximately equal to 5.0 mm × 3.6 mrad in the fast-axis and slow-axis. After beam shaping by the optimized BTS, the laser-diode beam can be coupled into a 100 μm core, 0.15 numerical aperture (NA) fiber with an output power of over 100 W and an electric-optical efficiency of 46.8%. PMID:27557249

  5. Towards a Unifying Framework for Pattern Transformation in Swarm Systems

    NASA Astrophysics Data System (ADS)

    Varghese, Blesson; McKee, Gerard

    2009-03-01

    This paper identifies the major challenges in the area of pattern formation. The work is also motivated by the need for development of a single framework to surmount these challenges. A framework based on the control of macroscopic parameters is proposed. The issue of transformation of patterns is specifically considered. A definition for transformation and four special cases, namely elementary and geometrical transformations by repositioning all or some robots in the pattern are provided. Two feasible tools for pattern transformation namely, a macroscopic parameter method and a mathematical tool—Moebius transformation also known as the linear fractional transformation are introduced. The realization of the unifying framework considering planning and communication is reported.

  6. Optimal PMU placement using topology transformation method in power systems.

    PubMed

    Rahman, Nadia H A; Zobaa, Ahmed F

    2016-09-01

    Optimal phasor measurement units (PMUs) placement involves the process of minimizing the number of PMUs needed while ensuring the entire power system completely observable. A power system is identified observable when the voltages of all buses in the power system are known. This paper proposes selection rules for topology transformation method that involves a merging process of zero-injection bus with one of its neighbors. The result from the merging process is influenced by the selection of bus selected to merge with the zero-injection bus. The proposed method will determine the best candidate bus to merge with zero-injection bus according to the three rules created in order to determine the minimum number of PMUs required for full observability of the power system. In addition, this paper also considered the case of power flow measurements. The problem is formulated as integer linear programming (ILP). The simulation for the proposed method is tested by using MATLAB for different IEEE bus systems. The explanation of the proposed method is demonstrated by using IEEE 14-bus system. The results obtained in this paper proved the effectiveness of the proposed method since the number of PMUs obtained is comparable with other available techniques. PMID:27489729

  7. Optimal PMU placement using topology transformation method in power systems.

    PubMed

    Rahman, Nadia H A; Zobaa, Ahmed F

    2016-09-01

    Optimal phasor measurement units (PMUs) placement involves the process of minimizing the number of PMUs needed while ensuring the entire power system completely observable. A power system is identified observable when the voltages of all buses in the power system are known. This paper proposes selection rules for topology transformation method that involves a merging process of zero-injection bus with one of its neighbors. The result from the merging process is influenced by the selection of bus selected to merge with the zero-injection bus. The proposed method will determine the best candidate bus to merge with zero-injection bus according to the three rules created in order to determine the minimum number of PMUs required for full observability of the power system. In addition, this paper also considered the case of power flow measurements. The problem is formulated as integer linear programming (ILP). The simulation for the proposed method is tested by using MATLAB for different IEEE bus systems. The explanation of the proposed method is demonstrated by using IEEE 14-bus system. The results obtained in this paper proved the effectiveness of the proposed method since the number of PMUs obtained is comparable with other available techniques.

  8. Transformation of legacy network management system to service oriented architecture

    NASA Astrophysics Data System (ADS)

    Sathyan, Jithesh; Shenoy, Krishnananda

    2007-09-01

    Service providers today are facing the challenge of operating and maintaining multiple networks, based on multiple technologies. Network Management System (NMS) solutions are being used to manage these networks. However the NMS is tightly coupled with Element or the Core network components. Hence there are multiple NMS solutions for heterogeneous networks. Current network management solutions are targeted at a variety of independent networks. The wide spread popularity of IP Multimedia Subsystem (IMS) is a clear indication that all of these independent networks will be integrated into a single IP-based infrastructure referred to as Next Generation Networks (NGN) in the near future. The services, network architectures and traffic pattern in NGN will dramatically differ from the current networks. The heterogeneity and complexity in NGN including concepts like Fixed Mobile Convergence will bring a number of challenges to network management. The high degree of complexity accompanying the network element technology necessitates network management systems (NMS) which can utilize this technology to provide more service interfaces while hiding the inherent complexity. As operators begin to add new networks and expand existing networks to support new technologies and products, the necessity of scalable, flexible and functionally rich NMS systems arises. Another important factor influencing NMS architecture is mergers and acquisitions among the key vendors. Ease of integration is a key impediment in the traditional hierarchical NMS architecture. These requirements trigger the need for an architectural framework that will address the NGNM (Next Generation Network Management) issues seamlessly. This paper presents a unique perspective of bringing service orientated architecture (SOA) to legacy network management systems (NMS). It advocates a staged approach in transforming a legacy NMS to SOA. The architecture at each stage is detailed along with the technical advantages and

  9. System for electronic transformation and geographic correlation of satellite television information. [cloud cover photography

    NASA Technical Reports Server (NTRS)

    Dubenskiy, V. P.; Nemkovskiy, B. L.; Rodionov, B. N.

    1974-01-01

    An electronic transformation and correlation system has been developed for the Meteor space weather system which provides transformation and scaling of the original picture, accounts for satellite flight altitude and inclinations of the optical axes of the transmitting devices, and simultaneously superposes the geographical coordinate grid on the transformed picture.

  10. Stability of linear systems in second-order form based on structure preserving similarity transformations

    SciTech Connect

    Stoustrup, Jakob; Pommer, Christian; Kliem, Wolfhard

    2015-10-31

    This paper deals with two stability aspects of linear systems of the form I ¨ x +B˙ x +Cx = 0 given by the triple (I;B;C). A general transformation scheme is given for a structure and Jordan form preserving transformation of the triple. We investigate how a system can be transformed by suitable choices of the transformation parameters into a new system (I;B1;C1) with a symmetrizable matrix C1. This procedure facilitates stability investigations. We also consider systems with a Hamiltonian spectrum which discloses marginal stability after a Jordan form preserving transformation.

  11. Structure and topography of the Siqueiros transform fault system: Evidence for the development of intra-transform spreading centers

    NASA Astrophysics Data System (ADS)

    Fornari, Daniel J.; Gallo, David G.; Edwards, Margo H.; Madsen, John A.; Perfit, Michael R.; Shor, Alexander N.

    1989-12-01

    The Siqueiros transform fault system, which offsets the East Pacific Rise between 8°20'N 8°30'N, has been mapped with the Sea MARC II sonar system and is found to consist of four intra-transform spreading centers and five strike-slip faults. The bathymetric and side-looking sonar data define the total width of the transform domain to be ≈20km. The transform domain includes prominent topographic features that are related to either seafloor spreading processes at the short spreading centers or shearing along the bounding faults. The spreading axes and the seafloor on the flanks of each small spreading center comprise morphological and structural features which suggest that the two western spreading centers are older than the eastern spreading centers. Structural data for the Clipperton, Orozco and Siqueiros transforms, indicate that the relative plate motion geometry of the Pacific-Cocos plate boundary has been stable for the past ≈1.5 Ma. Because the seafloor spreading fabric on the flanks of the western spreading centers is ≈500 000 years old and parallels the present EPR abyssal hill trend (350°) we conclude that a small change in plate motion was not the cause for intra-transform spreading center development in Siqueiros. We suggest that the impetus for the development of intra-transform spreading centers along the Siqueiros transform system was provided by the interaction of small melt anomalies in the mantle (SMAM) with deepseated, throughgoing lithospheric fractures within the shear zone. Initially, eruption sites may have been preferentially located along strike-slip faults and/or along cross-faults that eventually developed into pull-apart basins. Spreading centers C and D in the eastern portion of Siqueiros are in this initial pull-apart stage. Continued intrusion and volcanism along a short ridge within a pull-apart basin may lead to the formation of a stable, small intra-transform spreading center that creates a narrow swath of ridge

  12. PHELIX: design of transformer-driven linear implosion system

    SciTech Connect

    Turchi, Peter J; Atchison, Walter L; Rousculp, Chris L; Reinovsky, Robert E

    2008-01-01

    Experiments involving electromagnetically-imploded, solid-density liners can be achieved at reduced cost and energy if we start with a scale-size based on diagnostic resolution, rather than on the largest capacitor bank or generator we could bring to bear. For minimum resolution of 100 microns or less, many useful experiments could be performed with initial liner diameters that are factors of two to three smaller than used on high-energy systems, such as Atlas, thereby reducing energy requirements to sub-megajoule levels. Reduction in scale-size, however, also decreases the inductance change associated with liner motion relative to other inductances in the circuit. To improve coupling efficiency to liner kinetic energy, we invoke a current step-up transformer. Scaling relations have been developed for reducing the size and energy of such systems and compared with detailed numerical simulations. We discuss these calculations and describe the engineering embodiment of the resulting design for a system called PHELIX (Precision High Energy-density Liner Implosion eXperiment).

  13. Is public health ready to participate in the transformation of the healthcare system?

    PubMed

    Millar, John; Bruce, Ted; Cheng, Siu Mee; Masse, Richard; McKeown, David

    2013-01-01

    The healthcare system in Canada is undergoing significant transformation in response to three major interrelated pressures: the overall burden of illness is rising, patients are getting poor quality of care and healthcare costs are inexorably rising. One idea to guide this change is to transform the primary care system into a community-based primary healthcare (CBPH) system. This paper discusses, in particular, the readiness of public health to participate in the transformation to a CBPH system.

  14. Photometric transformation from RGB Bayer filter system to Johnson-Cousins BVR filter system

    NASA Astrophysics Data System (ADS)

    Park, Woojin; Pak, Soojong; Shim, Hyunjin; Le, Huynh Anh N.; Im, Myungshin; Chang, Seunghyuk; Yu, Joonkyu

    2016-01-01

    The RGB Bayer filter system consists of a mosaic of R, G , and B filters on the grid of the photo sensors which typical commercial DSLR (Digital Single Lens Reflex) cameras and CCD cameras are equipped with. Lot of unique astronomical data obtained using an RGB Bayer filter system are available, including transient objects, e.g. supernovae, variable stars, and solar system bodies. The utilization of such data in scientific research requires that reliable photometric transformation methods are available between the systems. In this work, we develop a series of equations to convert the observed magnitudes in the RGB Bayer filter system (RB,GB , and BB) into the Johnson-Cousins BVR filter system (BJ,VJ , and RC). The new transformation equations derive the calculated magnitudes in the Johnson-Cousins filters (BJcal,VJcal , and RCcal) as functions of RGB magnitudes and colors. The mean differences between the transformed magnitudes and original magnitudes, i.e. the residuals, are Δ (BJ -BJcal) = 0.064 mag, Δ (VJ -VJcal) = 0.041 mag, and Δ (RC -RCcal) = 0.039 mag. The calculated Johnson-Cousins magnitudes from the transformation equations show a good linear correlation with the observed Johnson-Cousins magnitudes.

  15. Rethinking health systems strengthening: key systems thinking tools and strategies for transformational change.

    PubMed

    Swanson, R Chad; Cattaneo, Adriano; Bradley, Elizabeth; Chunharas, Somsak; Atun, Rifat; Abbas, Kaja M; Katsaliaki, Korina; Mustafee, Navonil; Mason Meier, Benjamin; Best, Allan

    2012-10-01

    While reaching consensus on future plans to address current global health challenges is far from easy, there is broad agreement that reductionist approaches that suggest a limited set of targeted interventions to improve health around the world are inadequate. We argue that a comprehensive systems perspective should guide health practice, education, research and policy. We propose key 'systems thinking' tools and strategies that have the potential for transformational change in health systems. Three overarching themes span these tools and strategies: collaboration across disciplines, sectors and organizations; ongoing, iterative learning; and transformational leadership. The proposed tools and strategies in this paper can be applied, in varying degrees, to every organization within health systems, from families and communities to national ministries of health. While our categorization is necessarily incomplete, this initial effort will provide a valuable contribution to the health systems strengthening debate, as the need for a more systemic, rigorous perspective in health has never been greater. PMID:23014154

  16. High-Performance Work Systems: American Models of Workplace Transformation.

    ERIC Educational Resources Information Center

    Appelbaum, Eileen; Batt, Rosemary

    Rising competition in world and domestic markets for the past 2 decades has necessitated that U.S. companies undergo significant transformations to improve their performance with respect to a wide array of efficiency and quality indicators. Research on the transformations recently undertaken by some U.S. companies to boost performance revealed two…

  17. Floral-dip transformation of flax (Linum usitatissimum) to generate transgenic progenies with a high transformation rate.

    PubMed

    Bastaki, Nasmah K; Cullis, Christopher A

    2014-01-01

    Agrobacterium-mediated plant transformation via floral-dip is a widely used technique in the field of plant transformation and has been reported to be successful for many plant species. However, flax (Linum usitatissimum) transformation by floral-dip has not been reported. The goal of this protocol is to establish that Agrobacterium and the floral-dip method can be used to generate transgenic flax. We show that this technique is simple, inexpensive, efficient, and more importantly, gives a higher transformation rate than the current available methods of flax transformation. In summary, inflorescences of flax were dipped in a solution of Agrobacterium carrying a binary vector plasmid (T-DNA fragment plus the Linum Insertion Sequence, LIS-1) for 1 - 2 min. The plants were laid flat on their side for 24 hr. Then, plants were maintained under normal growth conditions until the next treatment. The process of dipping was repeated 2 - 3 times, with approximately 10 - 14 day intervals between dipping. The T1 seeds were collected and germinated on soil. After approximately two weeks, treated progenies were tested by direct PCR; 2 - 3 leaves were used per plant plus the appropriate T-DNA primers. Positive transformants were selected and grown to maturity. The transformation rate was unexpectedly high, with 50 - 60% of the seeds from treated plants being positive transformants. This is a higher transformation rate than those reported for Arabidopsis thaliana and other plant species, using floral-dip transformation. It is also the highest, which has been reported so far, for flax transformation using other methods for transformation.

  18. Genetic transformation of cotton with a harpin-encoding gene hpaXoo confers an enhanced defense response against Verticillium dahliae Kleb.

    PubMed

    Miao, Weiguo; Wang, Jingsheng

    2013-01-01

    The soil-borne fungal pathogen Verticillium dahliae Kleb causes Verticillium wilt in a wide range of crops including cotton (Gossypium hirsutum). To date, most upland cotton varieties are susceptible to V. dahliae and the breeding for cotton varieties with the resistance to Verticillium wilt has not been successful. Hpa1Xoo is a harpin protein from Xanthomonas oryzae pv. oryzae which induces the hypersensitive cell death in plants. When hpa1Xoo was transformed into the susceptible cotton line Z35 through Agrobacterium-mediated transformation, the transgenic cotton line (T-34) with an improved resistance to Verticillium dahliae was obtained. Here, we describe the related research approach, such as Western blot, Southern blot, immuno-gold labeling, evaluation of resistance to Verticillium dahliae, and how to detect the micro-hypersensitive response and oxidative burst elicited by harpin(Xoo) in plant tissue.

  19. Genetic transformation of cotton with a harpin-encoding gene hpaXoo confers an enhanced defense response against Verticillium dahliae Kleb.

    PubMed

    Miao, Weiguo; Wang, Jingsheng

    2013-01-01

    The soil-borne fungal pathogen Verticillium dahliae Kleb causes Verticillium wilt in a wide range of crops including cotton (Gossypium hirsutum). To date, most upland cotton varieties are susceptible to V. dahliae and the breeding for cotton varieties with the resistance to Verticillium wilt has not been successful. Hpa1Xoo is a harpin protein from Xanthomonas oryzae pv. oryzae which induces the hypersensitive cell death in plants. When hpa1Xoo was transformed into the susceptible cotton line Z35 through Agrobacterium-mediated transformation, the transgenic cotton line (T-34) with an improved resistance to Verticillium dahliae was obtained. Here, we describe the related research approach, such as Western blot, Southern blot, immuno-gold labeling, evaluation of resistance to Verticillium dahliae, and how to detect the micro-hypersensitive response and oxidative burst elicited by harpin(Xoo) in plant tissue. PMID:23143497

  20. An Overview of HATS: A Language Independent High Assurance Transformation System

    SciTech Connect

    Winter, V.L.

    1998-10-16

    Transformations that are based on syntax directed rewriting systems can have a significant impact on the construction of high assurance systems. However, in order for a transformational approach to be useful to a particular problem domain, a (general) transformation system must be adapted to the notation of that particular domain. A transformation system that can be easily adapted to various domain notations has the potential of having a wide range of applicability. In this paper we dissus why transforrmtion is attractive horn a high assurance perspective, as well as some issues surrounding automated transformation within specific problem domains. We then give an overview of a language independent High Assurance Transformation System (HATS) that is being developed at Sandia National Laboratories.

  1. Mechanism for the primary transformation of acetaminophen in a soil/water system.

    PubMed

    Liang, Chuanzhou; Lan, Zhonghui; Zhang, Xu; Liu, Yingbao

    2016-07-01

    The transformation of acetaminophen (APAP) in a soil/water system was systematically investigated by a combination of kinetic studies and a quantitative analysis of the reaction intermediates. Biotransformation was the predominant pathway for the elimination of APAP, whereas hydrolysis or other chemical transformation, and adsorption processes made almost no contribution to the transformation under a dark incubation. Bacillus aryabhattai strain 1-Sj-5-2-5-M, Klebsiella pneumoniae strain S001, and Bacillus subtilis strain HJ5 were the main bacteria identified in the biotransformation of APAP. The soil-to-water ratio and soil preincubation were able to alter the transformation kinetic pattern. Light irradiation promoted the overall transformation kinetics through enhanced biotransformation and extra photosensitized chemical reactions. The transformation pathways were strongly dependent on the initial concentration of APAP. The main primary transformation products were APAP oligomers and p-aminophenol, with the initial addition of 26.5 and 530 μM APAP, respectively. APAP oligomers accounted for more than 95% of transformed APAP, indicating that almost no bound residues were generated through the transformation of APAP in the soil/water system. The potential environmental risks of APAP could increase following the transformation of APAP in the soil/water system because of the higher toxicity of the transformation intermediates. PMID:27107139

  2. Efficient genetic transformation of Sorghum using a visual screening marker.

    PubMed

    Gao, Zhensheng; Jayaraj, J; Muthukrishnan, S; Claflin, Larry; Liang, G H

    2005-04-01

    To transform grain sorghum (Sorghum bicolor (L.) Moench) with a visual reporter gene (gfp) and a target gene (tlp), three genotypes (two inbreds, Tx 430 and C401, and a commercial hybrid, Pioneer 8505) were used. We obtained a total of 1011 fertile transgenic plants from 61 independent callus lines, which were produced from 2463 zygotic immature embryos via Agrobacterium-mediated transformation. The reporter gene, gfp, encoding green fluorescent protein (GFP), was used as a visual screening marker, and the target gene, tlp, encoding thaumatin-like protein (TLP), was chosen for enhancing resistance to fungal diseases and drought. Both genes were under the control of the maize ubi 1 promoter in the binary vector pPZP201. A total of 320 plants showing GFP expression, derived from 45 calli, were selected and analyzed by Southern blot analysis. There was a 100% correlation between the GFP expression and the presence of the target gene, tlp, in these plants. Transgenic plants showing strong TLP expression were confirmed by Western blotting with antiserum specific for TLP. The transgene segregated in various ratios among progeny, which was confirmed by examining seedlings showing GFP fluorescence. The progeny also showed different copy numbers of transgenics. This report describes the successful use of GFP screening for efficient production of stably transformed sorghum plants without using antibiotics or herbicides as selection agents.

  3. Darboux transformation of the Drinfeld–Sokolov–Satsuma–Hirota system and exact solutions

    SciTech Connect

    Geng, Xianguo; Li, Ruomeng

    2015-10-15

    A Darboux transformation for the Drinfeld–Sokolov–Satsuma–Hirota system of coupled equations is constructed with the aid of gauge transformations between the Lax pairs. As an application, several types of solutions of the Drinfeld–Sokolov–Satsuma–Hirota system are obtained, including soliton solutions, periodic solutions, rational solutions and others.

  4. Applications to aeronautics of the theory of transformations of nonlinear systems

    NASA Technical Reports Server (NTRS)

    Meyer, G.; Su, R.; Hunt, L. R.

    1982-01-01

    The development of the transformation theory is discussed. Results and applications concerning the use of this design technique for automatic flight control of aircraft are presented. The theory examines the transformation of nonlinear systems to linear systems. The tracking of linear models by nonlinear plants is discussed. Results of manned simulation are also presented.

  5. Phase Transformation Hysteresis in a Plutonium Alloy System: Modeling the Resistivity during the Transformation

    SciTech Connect

    Haslam, J J; Wall, M A; Johnson, D L; Mayhall, D J; Schwartz, A J

    2001-11-14

    We have induced, measured, and modeled the {delta}-{alpha}' martensitic transformation in a Pu-Ga alloy by a resistivity technique on a 2.8-mm diameter disk sample. Our measurements of the resistance by a 4-probe technique were consistent with the expected resistance obtained from a finite element analysis of the 4-point measurement of resistivity in our round disk configuration. Analysis by finite element methods of the postulated configuration of {alpha}' particles within model {delta} grains suggests that a considerable anisotropy in the resistivity may be obtained depending on the arrangement of the {alpha}' lens shaped particles within the grains. The resistivity of these grains departs from the series resistance model and can lead to significant errors in the predicted amount of the {alpha}' phase present in the microstructure. An underestimation of the amount of {alpha}' in the sample by 15%, or more, appears to be possible.

  6. Lie algebraic similarity transformed Hamiltonians for lattice model systems

    NASA Astrophysics Data System (ADS)

    Wahlen-Strothman, Jacob M.; Jiménez-Hoyos, Carlos A.; Henderson, Thomas M.; Scuseria, Gustavo E.

    2015-01-01

    We present a class of Lie algebraic similarity transformations generated by exponentials of two-body on-site Hermitian operators whose Hausdorff series can be summed exactly without truncation. The correlators are defined over the entire lattice and include the Gutzwiller factor ni ↑ni ↓ , and two-site products of density (ni ↑+ni ↓) and spin (ni ↑-ni ↓) operators. The resulting non-Hermitian many-body Hamiltonian can be solved in a biorthogonal mean-field approach with polynomial computational cost. The proposed similarity transformation generates locally weighted orbital transformations of the reference determinant. Although the energy of the model is unbound, projective equations in the spirit of coupled cluster theory lead to well-defined solutions. The theory is tested on the one- and two-dimensional repulsive Hubbard model where it yields accurate results for small and medium sized interaction strengths.

  7. Transformations of inorganic coal constituents in combustion systems

    SciTech Connect

    Helble, J.J.; Srinivasachar, S.; Wilemski, G.; Boni, A.A. ); Kang, Shin-Gyoo; Sarofim, A.F.; Beer, J.M. ); Peterson, T.W.; Wendt, J.O.L.; Gallagher, N.B.; Bool, L. ); Shah, N.; Huggins, F.E.; Huffman, G.P. )

    1991-09-01

    The technical objectives of this project are: (1) To define the partitioning of inorganic constituents associated with raw coal particles among products (including vapors, aerosols, and residual char/ash particles) formed under conditions representative of pulverized coal flames as a function of the specific (intrinsic and extrinsic) characteristics of the raw coal and the environment in which the transformations occur; and to characterize the resultant spectrum of products in detail. (2) To elucidate and quantify the fundamental processes (involving basic principles of physics, chemistry, thermodynamics) by which transformations of the inorganic constituents occur; and (3) to develop, based on the information required in (1) and (2), a tractable process'' model capable of predicting the significant features of the transformation process, most importantly, the nature and distribution of products. 26 refs., 151 figs., 51 tabs.

  8. Fate and transformation of silver nanoparticles in urban wastewater systems.

    PubMed

    Kaegi, Ralf; Voegelin, Andreas; Ort, Christoph; Sinnet, Brian; Thalmann, Basilius; Krismer, Jasmin; Hagendorfer, Harald; Elumelu, Maline; Mueller, Elisabeth

    2013-08-01

    Discharge of silver nanoparticles (Ag-NP) from textiles and cosmetics, todays major application areas for metallic Ag-NP, into wastewater is inevitable. Transformation and removal processes in sewers and wastewater treatment plants (WWTP) will determine the impact of Ag-NP on aquatic and terrestrial environments, via the effluents of the WWTP and via the use of digested sludge as fertilizer. We thus conducted experiments addressing the behavior of Ag-NP in sewers and in WWTP. We spiked Ag-NP to a 5 km long main trunk sewer and collected 40 wastewater samples after 500 m, 2400 m and 5000 m each according to the expected travel times of the Ag-NP. Excellent mass closure of the Ag derived by multiplying the measured Ag concentrations times the volumetric flow rates indicate an efficient transport of the Ag-NP without substantial losses to the sewer biofilm. Ag-NP reacted with raw wastewater in batch experiments were sulfidized to roughly 15% after 5 h reaction time as revealed by X-ray absorption spectroscopy (XAS). However, acid volatile sulfide (AVS) concentrations were substantially higher in the sewer channel (100 μM) compared to the batch experiments (3 μM; still sufficient to sulfidize spiked 2 μM Ag) possibly resulting in a higher degree of sulfidation in the sewer channel. We further investigated the removal efficiency of 10 nm and 100 nm Ag- and gold (Au)-NP coated with citrate or polyvinylpyrrolidone in activated sludge batch experiments. We obtained very high removal efficiencies (≈ 99%) irrespective of size and coating for Ag- and Au-NP, the latter confirming that the particle type was of minor importance with respect to the degree of NP removal. We observed a strong size dependence of the sulfidation kinetics. We conclude that Ag-NP discharged to the wastewater stream will become sulfidized to various degrees in the sewer system and are efficiently transported to the WWTP. The sulfidation of the Ag-NP will continue in the WWTP, but primarily

  9. Rapid Business Transformations in Health Care: A Systems Approach

    ERIC Educational Resources Information Center

    Shulaiba, Refaat A.

    2011-01-01

    The top two priorities of health care business leaders are to constantly improve the quality of health care while striving to contain and reduce the high cost of health care. The Health Care industry, similar to all businesses, is motivated to deliver innovative solutions that accelerate business transformation and increase business capabilities. …

  10. Transforming Teacher Education to Reform America's P-20 Education System

    ERIC Educational Resources Information Center

    Futrell, Mary Hatwood

    2010-01-01

    Moving beyond "reform", teacher education programs need to be "transformed" to better prepare teachers, school counselors, and administrators to teach, guide, and lead in this complex society. Educators must have the skills, knowledge, and commitment to be part of the leadership team that assumes the responsibility for carrying the mantra of the…

  11. Transformations of inorganic coal constituents in combustion systems

    SciTech Connect

    Helble, J.J.; Srinivasachar, S.; Wilemski, G.; Boni, A.A. ); Kang, Shin-Gyoo; Sarofim, A.F.; Beer, J.M. ); Peterson, T.W.; Wendt, J.O.L.; Gallagher, M.N.B. ); Shah, N.; Huggins, F.E. Huffman, G.P. )

    1991-05-01

    Objectives of this project are: (1) define the partitioning of inorganic constituents associated with raw coal particles among products (including vapors, aerosols, and residual char/ash particles) formed under conditions representative of pulverized coal flames as a function of the specific (intrinsic and extrinsic) characteristics of the raw coal and the environment in which the transformations occur; and (2) characterize the resultant spectrum of products in detail; To elucidate and quantify the fundamental processes and by which transformations of the inorganic constituents occur; and To develop, based on the information required in the above, a tractable process'' model capable of predicting the significant features of the transformation process, most importantly, the nature and distribution of products. The work discussed herein highlights recent accomplishments: at the University of Kentucky (UK), in analyzing recently obtained XAFS data; at the Massachusetts Institute of Technology (MIT), in modeling the char fragmentation process, allowing for pore interactions to produce clustered pore arrangements; at MIT, in analyzing experiments with chars of varying porosity to quantify the effect on char fragmentation; at the University of Arizona (UA), in examining the behavior of alkali metals during combustion; at UA, in examining the capture of vaporized sodium by extraneous kaolinite additive; at PSI Technology Company (PSIT), in examining the transformations and stickiness of nepheline and kaolinite/montmorillonite mixtures, and conducting initial experiments with the SOAP and Loy Yang 1953 coals; and at PSIT, in continuing development of the engineering model for ash particle formation. 96 figs., 4 tabs.

  12. Simulation Test System of Non-Contact D-dot Voltage Transformer

    NASA Astrophysics Data System (ADS)

    Yang, Jie; Wang, Jingang; Luo, Ruixi; Gao, Can; Songnong, Li; Kongjun, Zhou

    2016-04-01

    The development trend of future voltage transformer in smart grid is non-contact measurement, miniaturization and intellectualization. This paper proposes one simulation test system of non-contact D-dot transformer for voltage measurement. This simulation test system consists of D-dot transformer, signal processing circuit and ground PC port. D-dot transformer realizes the indirect voltage measurement by measuring the change rate of electric displacement vector, a non-contact means (He et al. 2004, Principles and experiments of voltage transformer based on self-integrating D-dot probe. Proc CSEE 2014;15:2445-51). Specific to the characteristics of D-dot transformer signals, signal processing circuits with strong resistance to interference and distortion-free amplified sensor output signal are designed. WIFI wireless network is used to transmit the voltage detection to LabVIEW-based ground collection port and LabVIEW technology is adopted for signal reception, data processing and analysis and other functions. Finally, a test platform is established to simulate the performance of the whole test system of single-phase voltage transformer. Test results indicate that this voltage transformer has sound real-time performance, high accuracy and fast response speed and the simulation test system is stable and reliable and can be a new prototype of voltage transformers.

  13. Development of a Simplified Calculation Tool for Voltage Drop caused by Transformer Inrush in Distribution System

    NASA Astrophysics Data System (ADS)

    Nakachi, Yoshiki; Fukae, Takayuki; Sugahara, Toshinori; Nakamura, Hayato; Koyama, Mitsuaki; Ueda, Fukashi

    It is well known that the voltage drop due to the inrush current at energizing transformer, may at times, interrupt electrical equipment. Generally, the voltage drop is calculated by using the sophisticated tool such as EMTP, so the transformer saturation phenomenon has been properly represented. However, it is not practical for distribution system engineers to calculate the voltage drop with transformer inrush by using EMTP, because there are a lot of network access requests needing such calculations with many kinds of transformers. Therefore, in this paper, a simplified and easy-to-use calculation tool for voltage drop caused by transformer inrush in distribution system is developed. In order to grasp the voltage drop by inrush current during the planning stage, it is formulated by considering the transformer saturation/unsaturation periods in each winding type, the developed tool is based on the versatile spreadsheet software such as Microsoft Excel®. It can be used with accuracy almost similar to the EMTP.

  14. An intensive understanding of vacuum infiltration transformation of pakchoi (Brassica rapa ssp. chinensis).

    PubMed

    Xu, Hengjian; Wang, Xiufeng; Zhao, Hong; Liu, Fan

    2008-08-01

    Pakchoi (Brassica rapa L. ssp. chinensis), a kind of Chinese cabbage, is an important vegetable in Asian countries. Agrobacterium mediated in planta vacuum infiltration transformation has been performed in pakchoi since 1998, but a detailed study on this technique was lacking. Pakchoi plants 40-50 days old with inflorescences were vacuum infiltrated with Agrobacterium tumefaciens strain C58C1 harboring the binary vector pBBBast-gus-intron. The transformation frequency in the harvested seeds mainly varied from 1 x 10(-4) to 3 x 10(-4) over several years, and it was lower than the frequency in Arabidopsis thaliana. Transformants were obtained from both the upper and the lower parts of the infiltrated plants with or without an elongated inflorescence. Stained ovules and pollen grains were found in the unopened flower 13 days post-infiltration, which was about 0.5-1 mm in diameter at infiltration time with an open ovary as revealed by paraffin sections. Histochemical assays revealed that Agrobacteria were more abundant in the flower tissue than in stem and leaf tissues at all times after infiltration despite the sharp decrease of live Agrobacteria in plant 14 days post infiltration as revealed by the colony forming units on the Agrobacteria culture medium. The results of vacuum infiltration transformation of pakchoi and Arabidopsis thaliana were compared and a strategy to optimize the transformation conditions to increase the transformation frequency in pakchoi was discussed.

  15. Efficient Transformation of Oil Palm Protoplasts by PEG-Mediated Transfection and DNA Microinjection

    PubMed Central

    Masani, Mat Yunus Abdul; Noll, Gundula A.; Parveez, Ghulam Kadir Ahmad; Sambanthamurthi, Ravigadevi; Prüfer, Dirk

    2014-01-01

    Background Genetic engineering remains a major challenge in oil palm (Elaeis guineensis) because particle bombardment and Agrobacterium-mediated transformation are laborious and/or inefficient in this species, often producing chimeric plants and escapes. Protoplasts are beneficial as a starting material for genetic engineering because they are totipotent, and chimeras are avoided by regenerating transgenic plants from single cells. Novel approaches for the transformation of oil palm protoplasts could therefore offer a new and efficient strategy for the development of transgenic oil palm plants. Methodology/Principal Findings We recently achieved the regeneration of healthy and fertile oil palms from protoplasts. Therefore, we focused on the development of a reliable PEG-mediated transformation protocol for oil palm protoplasts by establishing and validating optimal heat shock conditions, concentrations of DNA, PEG and magnesium chloride, and the transfection procedure. We also investigated the transformation of oil palm protoplasts by DNA microinjection and successfully regenerated transgenic microcalli expressing green fluorescent protein as a visible marker to determine the efficiency of transformation. Conclusions/Significance We have established the first successful protocols for the transformation of oil palm protoplasts by PEG-mediated transfection and DNA microinjection. These novel protocols allow the rapid and efficient generation of non-chimeric transgenic callus and represent a significant milestone in the use of protoplasts as a starting material for the development of genetically-engineered oil palm plants. PMID:24821306

  16. Harmonic overvoltages in industrial power system with capacitors and saturated transformers

    SciTech Connect

    Tran-Quoc, T.; Pierrat, L. |; Montmeat, A.; Giard, A.

    1995-12-31

    This paper presents a new method for determination of instantaneous saturation and hysteresis loops of transformers with a sinusoidal or nonsinusoidal voltage excitation by taking into account only the rms values and no-load losses. This model is used to study harmonic overvoltages in an industrial power system with saturated transformers and power factor correction capacitors. Influences of short-circuit power source and switching orders of transformer and capacitor on overvoltages are investigated.

  17. Joint-transform correlator systems using deformable-mirror spatial light modulators

    NASA Technical Reports Server (NTRS)

    Florence, James M.

    1989-01-01

    Two joint-transform correlator systems based on deformable-mirror-device (DMD) spatial light modulators and CCD video cameras are described. The system designs provide for real-time correlation between video-formatted input scenes and stored reference images. The DMD light modulators are used exclusively for inputting data in the optical stages of these systems, and the CCD cameras are used to provide the critical square-law detection required in joint-transform achitectures. Experimental verification of the functionality of these DMD/CCD joint-transform systems is also presented.

  18. Chloroplast transformation.

    PubMed

    Lu, Xiao-Mei; Yin, Wei-Bo; Hu, Zan-Min

    2006-01-01

    In this chapter we briefly review the developmental history and current research status of chloroplast transformation and introduce the merits of chloroplast transformation as compared with the nuclear genome transformation. Furthermore, according to the chloroplast transformation achieved in oilseed rape (Brassica napus), we introduce the preparation of explants, transformation methods, system selection, identification methods of the transplastomic plants, and experimental results. The technical points, the bottleneck, and the further research directions of the chloroplast transformation are discussed in the notes.

  19. Progress of cereal transformation technology mediated by Agrobacterium tumefaciens.

    PubMed

    Hiei, Yukoh; Ishida, Yuji; Komari, Toshihiko

    2014-01-01

    Monocotyledonous plants were believed to be not transformable by the soil bacterium Agrobacterium tumefaciens until two decades ago, although convenient protocols for infection of leaf disks and subsequent regeneration of transgenic plants had been well established in a number of dicotyledonous species by then. This belief was reinforced by the fact that monocotyledons are mostly outside the host range of crown gall disease caused by the bacterium and by the failures in trials in monocotyledons to mimic the transformation protocols for dicotyledons. However, a key reason for the failure could have been the lack of active cell divisions at the wound sites in monocotyledons. The complexity and narrow optimal windows of critical factors, such as genotypes of plants, conditions of the plants from which explants are prepared, tissue culture methods and culture media, pre-treatments of explants, strains of A. tumefaciens, inducers of virulence genes, transformation vectors, selection marker genes and selective agents, kept technical hurdles high. Eventually it was demonstrated that rice and maize could be transformed by co-cultivating cells of callus cultures or immature embryos, which are actively dividing or about to divide, with A. tumefaciens. Subsequently, these initial difficulties were resolved one by one by many research groups, and the major cereals are now transformed quite efficiently. As many as 15 independent transgenic events may be regenerated from a single piece of immature embryo of rice. Maize transformation protocols are well established, and almost all transgenic events deregulated for commercialization after 2003 were generated by Agrobacterium-mediated transformation. Wheat, barley, and sorghum are also among those plants that can be efficiently transformed by A. tumefaciens. PMID:25426132

  20. Progress of cereal transformation technology mediated by Agrobacterium tumefaciens

    PubMed Central

    Hiei, Yukoh; Ishida, Yuji; Komari, Toshihiko

    2014-01-01

    Monocotyledonous plants were believed to be not transformable by the soil bacterium Agrobacterium tumefaciens until two decades ago, although convenient protocols for infection of leaf disks and subsequent regeneration of transgenic plants had been well established in a number of dicotyledonous species by then. This belief was reinforced by the fact that monocotyledons are mostly outside the host range of crown gall disease caused by the bacterium and by the failures in trials in monocotyledons to mimic the transformation protocols for dicotyledons. However, a key reason for the failure could have been the lack of active cell divisions at the wound sites in monocotyledons. The complexity and narrow optimal windows of critical factors, such as genotypes of plants, conditions of the plants from which explants are prepared, tissue culture methods and culture media, pre-treatments of explants, strains of A. tumefaciens, inducers of virulence genes, transformation vectors, selection marker genes and selective agents, kept technical hurdles high. Eventually it was demonstrated that rice and maize could be transformed by co-cultivating cells of callus cultures or immature embryos, which are actively dividing or about to divide, with A. tumefaciens. Subsequently, these initial difficulties were resolved one by one by many research groups, and the major cereals are now transformed quite efficiently. As many as 15 independent transgenic events may be regenerated from a single piece of immature embryo of rice. Maize transformation protocols are well established, and almost all transgenic events deregulated for commercialization after 2003 were generated by Agrobacterium-mediated transformation. Wheat, barley, and sorghum are also among those plants that can be efficiently transformed by A. tumefaciens. PMID:25426132

  1. Transformation of rice mediated by Agrobacterium tumefaciens.

    PubMed

    Hiei, Y; Komari, T; Kubo, T

    1997-09-01

    Agrobacterium tumefaciens has been routinely utilized in gene transfer to dicotyledonous plants, but monocotyledonous plants including important cereals were thought to be recalcitrant to this technology as they were outside the host range of crown gall. Various challenges to infect monocotyledons including rice with Agrobacterium had been made in many laboratories, but the results were not conclusive until recently. Efficient transformation protocols mediated by Agrobacterium were reported for rice in 1994 and 1996. A key point in the protocols was the fact that tissues consisting of actively dividing, embryonic cells, such as immature embryos and calli induced from scutella, were co-cultivated with Agrobacterium in the presence of acetosyringonc, which is a potent inducer of the virulence genes. It is now clear that Agrobacterium is capable of transferring DNA to monocotyledons if tissues containing 'competent' cells are infected. The studies of transformation of rice suggested that numerous factors including genotype of plants, types and ages of tissues inoculated, kind of vectors, strains of Agrobacterium, selection marker genes and selective agents, and various conditions of tissue culture, are of critical importance. Advantages of the Agrobacterium-mediated transformation in rice, like on dicotyledons, include the transfer of pieces of DNA with defined ends with minimal rearrangements, the transfer of relatively large segments of DNA, the integration of small numbers of copies of genes into plant chromosomes, and high quality and fertility of transgenic plants. Delivery of foreign DNA to rice plants via A. tumefaciens is a routine technique in a growing number of laboratories. This technique will allow the genetic improvement of diverse varieties of rice, as well as studies of many aspects of the molecular biology of rice. PMID:9291974

  2. Efficient transformations from geodetic to UTM coordinate systems

    SciTech Connect

    Toms, R.M.

    1996-08-07

    The problem of efficiently performing transformations from geocentric to geodetic coordinates has been addressed at previous DIS (Distributed Interactive Simulation) workshops. This paper extends the work presented at the 14th DIS Workshop. As a consequence of the new algorithm for geocentric to geodetic coordinate conversion, a subsequent conversion to Universal Transverse Mercator coordinates is made considerably more efficient. No additional trigonometric or square root evaluations are required and accuracy is not degraded.

  3. Schrieffer-Wolff transformation for quantum many-body systems

    SciTech Connect

    Bravyi, Sergey; DiVincenzo, David P.; Loss, Daniel

    2011-10-15

    The Schrieffer-Wolff (SW) method is a version of degenerate perturbation theory in which the low-energy effective Hamiltonian H{sub eff} is obtained from the exact Hamiltonian by a unitary transformation decoupling the low-energy and high-energy subspaces. We give a self-contained summary of the SW method with a focus on rigorous results. We begin with an exact definition of the SW transformation in terms of the so-called direct rotation between linear subspaces. From this we obtain elementary proofs of several important properties of H{sub eff} such as the linked cluster theorem. We then study the perturbative version of the SW transformation obtained from a Taylor series representation of the direct rotation. Our perturbative approach provides a systematic diagram technique for computing high-order corrections to H{sub eff}. We then specialize the SW method to quantum spin lattices with short-range interactions. We establish unitary equivalence between effective low-energy Hamiltonians obtained using two different versions of the SW method studied in the literature. Finally, we derive an upper bound on the precision up to which the ground state energy of the nth-order effective Hamiltonian approximates the exact ground state energy. - Highlights: > The Schrieffer-Wolff transformation is specialized to quantum spin lattices with short-range interactions. > We provide a diagram technique for computing high-order corrections to the effective low-energy Hamiltonian. > We derive a rigorous bound on the error up to which the nth-order effective low-energy dynamics approximates the exact dynamics.

  4. Transformations of inorganic coal constituents in combustion systems

    SciTech Connect

    Boni, A.A.; Helble, J.J.; Srinivasachar, S.; Wilemski, G. ); Sarofim, A.F.; Beer, J.M. ); Peterson, T.W.; Wendt, J.O.L. ); Huffman, G.P.; Huggins, F.E. )

    1990-11-01

    Objectives of this project are: (1) define the partitioning of inorganic constituents associated with raw coal particles among products (including vapors, aerosols, and residual char/ash particles) formed under conditions representative of pulverized coal flames as a function of the specific (intrinsic and extrinsic) characteristics of the raw coal and the environment in which the transformations occur; and (2) characterize the resultant spectrum of products in detail; elucidate and quantify the fundamental processes by which transformations of the inorganic constituents occur; and develop, based on the information required in a. and b. above, a tractable process'' model capable of predicting the significant features of the transformation process, most importantly, the nature and distribution of products. The work discussed highlights recent accomplishments in modeling synthetic char particle combustion times and temperatures with allowance for oxygen dissociation in the char boundary layer; in developing a continuum model for residual ash information and char fragmentation, permitting a more detailed time-resolved study of ash coalescence within a burning char particle; and in continuing development of the engineering model for ash particle formation. 40 refs., 25 figs., 10 tabs.

  5. Coordinate transformation between rotating and inertial systems under the constant two-way speed of light

    NASA Astrophysics Data System (ADS)

    Choi, Yang-Ho

    2016-09-01

    An observation system consists of the world lines of rest observers in the system. Recently a coordinate transformation between an isotropic and a rotating observation system has been presented which was derived through a relativistic circular approach based on the Lorentz transformation. It was formulated such that the relative speeds between the two systems are the same, but the two-way speed of light is not constant in the rotating observation system. The constancy of the two-way speed of light in inertial frames has been known to be experimentally verified. This paper presents the transformation that holds the constancy in the rotating system as well. Though the rotating system is in motion with acceleration, it can be regarded as locally inertial. Thus, in the limit, a transformation into a rotating system should be reduced to a transformation into an inertial systems. The transformation presented is consistent with the one between inertial systems so that the latter can be derived from the former in the limit. Moreover it allows us to theoretically analyze the generalized Sagnac effect, which involves rectilinear motion as well as circular motion. The theoretical analysis corresponds to the experimental results.

  6. A system-wide innovation in transition services: transforming the home care liaison role.

    PubMed

    Meadows, Carl A; Fraser, Julie; Camus, Susann; Henderson, Kathy

    2014-02-01

    This article describes how a provincial health authority in Canada improved patient care and staff satisfaction by transforming the role of home care (HC) liaison. The transformation focused on clearly defining the role, function and reporting structure, and identifying which healthcare providers could fill the liaison role. The transformation included adoption of transition best practices, leveraging an electronic referral system, creation of an interprofessional team, standardization of tools/orientation, and strong evaluation metrics, centralizing decision making, and developing a process for streaming referrals. The authors identify key success factors that made the transformation possible, as well as challenges and work that remains to sustain the change. PMID:24492265

  7. A system-wide innovation in transition services: transforming the home care liaison role.

    PubMed

    Meadows, Carl A; Fraser, Julie; Camus, Susann; Henderson, Kathy

    2014-02-01

    This article describes how a provincial health authority in Canada improved patient care and staff satisfaction by transforming the role of home care (HC) liaison. The transformation focused on clearly defining the role, function and reporting structure, and identifying which healthcare providers could fill the liaison role. The transformation included adoption of transition best practices, leveraging an electronic referral system, creation of an interprofessional team, standardization of tools/orientation, and strong evaluation metrics, centralizing decision making, and developing a process for streaming referrals. The authors identify key success factors that made the transformation possible, as well as challenges and work that remains to sustain the change.

  8. Transient expression systems for plant-derived biopharmaceuticals.

    PubMed

    Komarova, Tatiana V; Baschieri, Selene; Donini, Marcello; Marusic, Carla; Benvenuto, Eugenio; Dorokhov, Yuri L

    2010-08-01

    In the molecular farming area, transient expression approaches for pharmaceutical proteins production, mainly recombinant monoclonal antibodies and vaccines, were developed almost two decades ago and, to date, these systems basically depend on Agrobacterium-mediated delivery and virus expression machinery. We survey here the current state-of-the-art of this research field. Several vectors have been designed on the basis of DNA- and RNA-based plant virus genomes and viral vectors are used both as single- and multicomponent expression systems in different combinations depending on the protein of interest. The obvious advantages of these systems are ease of manipulation, speed, low cost and high yield of proteins. In addition, Agrobacterium-mediated expression also allows the production in plants of complex proteins assembled from subunits. Currently, the transient expression methods are preferential over any other transgenic system for the exploitation of large and unrestricted numbers of plants in a contained environment. By designing optimal constructs and related means of delivery into plant cells, the overall technology plan considers scenarios that envisage high yield of bioproducts and ease in monitoring the whole spectrum of upstream production, before entering good manufacturing practice facilities. In this way, plant-derived bioproducts show promise of high competitiveness towards classical eukaryotic cell factory systems. PMID:20673010

  9. Molecular transformation, gene cloning, and gene expression systems for filamentous fungi

    USGS Publications Warehouse

    Gold, Scott E.; Duick, John W.; Redman, Regina S.; Rodriguez, Rusty J.

    2001-01-01

    This chapter discusses the molecular transformation, gene cloning, and gene expression systems for filamentous fungi. Molecular transformation involves the movement of discrete amounts of DNA into cells, the expression of genes on the transported DNA, and the sustainable replication of the transforming DNA. The ability to transform fungi is dependent on the stable replication and expression of genes located on the transforming DNA. Three phenomena observed in bacteria, that is, competence, plasmids, and restriction enzymes to facilitate cloning, were responsible for the development of molecular transformation in fungi. Initial transformation success with filamentous fungi, involving the complementation of auxotrophic mutants by exposure to sheared genomic DNA or RNA from wt isolates, occurred with low transformation efficiencies. In addition, it was difficult to retrieve complementing DNA fragments and isolate genes of interest. This prompted the development of transformation vectors and methods to increase efficiencies. The physiological studies performed with fungi indicated that the cell wall could be removed to generate protoplasts. It was evident that protoplasts could be transformed with significantly greater efficiencies than walled cells.

  10. A generalized Hirota-Satsuma coupled KdV system: Darboux transformations and reductions

    NASA Astrophysics Data System (ADS)

    Xue, Lingling; Liu, Q. P.; Wang, Dengshan

    2016-08-01

    A Darboux transformation is constructed for the generalized Hirota-Satsuma coupled KdV system and the result is compared with the recent work of Geng and his collaborators [X. G. Geng et al., Phys. Rev. E 79, 056602 (2009) and X. G. Geng and G. L. He, J. Math. Phys. 51, 033514 (2010)]. It is shown that our Darboux transformation may be applied to three interesting reductions of the general system. In addition, the iteration of this Darboux transformation is worked out, and some solutions to the associated systems are obtained.

  11. Drought-tolerant rice germplasm developed from an Oryza officinalis transformation-competent artificial chromosome clone.

    PubMed

    Liu, R; Zhang, H H; Chen, Z X; Shahid, M Q; Fu, X L; Liu, X D

    2015-10-29

    Oryza officinalis has proven to be a natural gene reservoir for the improvement of domesticated rice as it carries many desirable traits; however, the transfer of elite genes to cultivated rice by conventional hybridization has been a challenge for rice breeders. In this study, the conserved sequence of plant stress-related NAC transcription factors was selected as a probe to screen the O. officinalis genomic transformation-competent artificial chromosome library by Southern blot; 11 positive transformation-competent artificial chromosome clones were subsequently detected. By Agrobacterium-mediated transformation, an indica rice variety, Huajingxian 74 (HJX74), was transformed with a TAC clone harboring a NAC gene-positive genomic fragment from O. officinalis. Molecular analysis revealed that the O. officinalis genomic fragment was integrated into the genome of HJX74. The transgenic lines exhibited high tolerance to drought stress. Our results demonstrate that the introduction of stress-related transformation-competent artificial chromosome clones, coupled with a transgenic validation approach, is an effective method of transferring agronomically important genes from O. officinalis to cultivated rice.

  12. Co-transformation of canola by chimeric chitinase and tlp genes towards improving resistance to Sclerotinia sclerotiorum.

    PubMed

    Aghazadeh, Rustam; Zamani, Mohammadreza; Motallebi, Mostafa; Moradyar, Mehdi; Moghadassi Jahromi, Zahra

    2016-09-01

    Canola (Brassica napus) plants were co-transformed with two pathogenesis-related protein genes expressing a Trichoderma atroviride chitinase with a chitin-binding domain (chimeric chitinase) and a thaumatin-like protein (tlp) from Oryza sativa conferring resistance to phytopatogenic fungi by Agrobacterium-mediated transformation. The putative transgenic plants were confirmed by PCR. After measuring the specific activity of the chimeric chitinase and glucanase activity for tlp genes, transgenic plants with high specific activity were selected for southern blot analysis to confirm the copy number of the genes. In vitro assays, the antifungal activity of crude extracted protein against Sclerotinia sclerotiorum showed that the inhibition percentage in double transgenic plants was between 55 and 62, whereas the inhibition percentage in single-gene transformants (chimeric chitinase) ranged from 35 to 45 percent. Importantly, in greenhouse conditions, the double transgenic plants showed significant resistance than the single-gene transformant and wild type plants. The results in T2 generation using the intact leaf inoculation method showed that the average lesion diameters were 10, 14.7 and 29 mm for the double transformant, single-gene transformant and non-transgenic plants, respectively. Combined expression of chimeric chitinase and tlp in transgenic plants showed significantly enhanced resistance against S. sclerotiorum than the one that express single-gene transformant plants. These results suggest that the co-expression of chimeric chitinase and tlp can confer enhanced disease resistance in canola plant. PMID:27430511

  13. Co-transformation of canola by chimeric chitinase and tlp genes towards improving resistance to Sclerotinia sclerotiorum.

    PubMed

    Aghazadeh, Rustam; Zamani, Mohammadreza; Motallebi, Mostafa; Moradyar, Mehdi; Moghadassi Jahromi, Zahra

    2016-09-01

    Canola (Brassica napus) plants were co-transformed with two pathogenesis-related protein genes expressing a Trichoderma atroviride chitinase with a chitin-binding domain (chimeric chitinase) and a thaumatin-like protein (tlp) from Oryza sativa conferring resistance to phytopatogenic fungi by Agrobacterium-mediated transformation. The putative transgenic plants were confirmed by PCR. After measuring the specific activity of the chimeric chitinase and glucanase activity for tlp genes, transgenic plants with high specific activity were selected for southern blot analysis to confirm the copy number of the genes. In vitro assays, the antifungal activity of crude extracted protein against Sclerotinia sclerotiorum showed that the inhibition percentage in double transgenic plants was between 55 and 62, whereas the inhibition percentage in single-gene transformants (chimeric chitinase) ranged from 35 to 45 percent. Importantly, in greenhouse conditions, the double transgenic plants showed significant resistance than the single-gene transformant and wild type plants. The results in T2 generation using the intact leaf inoculation method showed that the average lesion diameters were 10, 14.7 and 29 mm for the double transformant, single-gene transformant and non-transgenic plants, respectively. Combined expression of chimeric chitinase and tlp in transgenic plants showed significantly enhanced resistance against S. sclerotiorum than the one that express single-gene transformant plants. These results suggest that the co-expression of chimeric chitinase and tlp can confer enhanced disease resistance in canola plant.

  14. Early Anthropogenic Transformation of the Danube-Black Sea System

    PubMed Central

    Giosan, Liviu; Coolen, Marco J. L.; Kaplan, Jed O.; Constantinescu, Stefan; Filip, Florin; Filipova-Marinova, Mariana; Kettner, Albert J.; Thom, Nick

    2012-01-01

    Over the last century humans have altered the export of fluvial materials leading to significant changes in morphology, chemistry, and biology of the coastal ocean. Here we present sedimentary, paleoenvironmental and paleogenetic evidence to show that the Black Sea, a nearly enclosed marine basin, was affected by land use long before the changes of the Industrial Era. Although watershed hydroclimate was spatially and temporally variable over the last ~3000 years, surface salinity dropped systematically in the Black Sea. Sediment loads delivered by Danube River, the main tributary of the Black Sea, significantly increased as land use intensified in the last two millennia, which led to a rapid expansion of its delta. Lastly, proliferation of diatoms and dinoflagellates over the last five to six centuries, when intensive deforestation occurred in Eastern Europe, points to an anthropogenic pulse of river-borne nutrients that radically transformed the food web structure in the Black Sea. PMID:22937219

  15. An efficient Foxtail mosaic virus vector system with reduced environmental risk

    PubMed Central

    2010-01-01

    Background Plant viral vectors offer high-yield expression of pharmaceutical and commercially important proteins with a minimum of cost and preparation time. The use of Agrobacterium tumefaciens has been introduced to deliver the viral vector as a transgene to each plant cell via a simple, nonsterile infiltration technique called "agroinoculation". With agroinoculation, a full length, systemically moving virus is no longer necessary for excellent protein yield, since the viral transgene is transcribed and replicates in every infiltrated cell. Viral genes may therefore be deleted to decrease the potential for accidental spread and persistence of the viral vector in the environment. Results In this study, both the coat protein (CP) and triple gene block (TGB) genetic segments were eliminated from Foxtail mosaic virus to create the "FECT" vector series, comprising a deletion of 29% of the genome. This viral vector is highly crippled and expresses little or no marker gene within the inoculated leaf. However, when co-agroinoculated with a silencing suppressor (p19 or HcPro), FECT expressed GFP at 40% total soluble protein in the tobacco host, Nicotiana benthamiana. The modified FoMV vector retained the full-length replicase ORF, the TGB1 subgenomic RNA leader sequence and either 0, 22 or 40 bases of TGB1 ORF (in vectors FECT0, FECT22 and FECT40, respectively). As well as N. benthamiana, infection of legumes was demonstrated. Despite many attempts, expression of GFP via syringe agroinoculation of various grass species was very low, reflecting the low Agrobacterium-mediated transformation rate of monocots. Conclusions The FECT/40 vector expresses foreign genes at a very high level, and yet has a greatly reduced biohazard potential. It can form no virions and can effectively replicate only in a plant with suppressed silencing. PMID:21162736

  16. Computer transformation of partial differential equations into any coordinate system

    NASA Technical Reports Server (NTRS)

    Sullivan, R. D.

    1977-01-01

    The use of tensors to provide a compact way of writing partial differential equations in a form valid in all coordinate systems is discussed. In order to find solutions to the equations with their boundary conditions they must be expressed in terms of the coordinate system under consideration. The process of arriving at these expressions from the tensor formulation was automated by a software system, TENSR. An allied system that analyzes the resulting expressions term by term and drops those that are negligible is also described.

  17. On Cyclical Phase Transformations in Driven Alloy Systems

    NASA Astrophysics Data System (ADS)

    Lee, Jong K.

    2008-05-01

    Cyclical phase transformations occurring in driven materials syntheses such as ball milling are described in terms of a free energy minimization process of participant phases. The oscillatory flow behavior of metals with low stacking fault energies during hot working is taken as a prototype in which a ductile crystalline phase sustains undulation in its free energy, due to the alternate succession of work-hardening and work-softening mechanisms. A time-dependent, oscillatory free energy function is then obtained by solving a delay differential equation (DDE), which accounts for a time lag due to diffusion. To understand cyclical transitions on an atomistic scale, work is extended to molecular dynamics simulations. Under shear deformation, a two-dimensional nanocrystal shows cyclical transitions between an equilibrium rhombus and a nonequilibrium square phase. Three-dimensional simulations show crystalline-to-glass transitions at high strain rates, but very high shear strain rates are found to lead to a latticelike network structure in the plane perpendicular to the shear direction, with strings of atoms parallel to the shear direction.

  18. Darboux transformations with tetrahedral reduction group and related integrable systems

    NASA Astrophysics Data System (ADS)

    Berkeley, George; Mikhailov, Alexander V.; Xenitidis, Pavlos

    2016-09-01

    In this paper, we derive new two-component integrable differential difference and partial difference systems by applying a Lax-Darboux scheme to an operator formed from an 𝔰𝔩3(ℂ)-based automorphic Lie algebra. The integrability of the found systems is demonstrated via Lax pairs and generalised symmetries.

  19. Surprises of the Transformer as a Coupled Oscillator System

    ERIC Educational Resources Information Center

    Silva, J. P.; Silvestre, A. J.

    2008-01-01

    We study a system of two RLC oscillators coupled through a variable mutual inductance. The system is interesting because it exhibits some peculiar features of coupled oscillators: (i) there are two natural frequencies; (ii) in general, the resonant frequencies do not coincide with the natural frequencies; (iii) the resonant frequencies of both…

  20. Application of Input-State of the System Transformation for Linearization of Selected Electrical Circuits

    NASA Astrophysics Data System (ADS)

    Zawadzki, Andrzej; Różowicz, Sebastian

    2016-05-01

    The paper presents a transformation of nonlinear electric circuit into linear one through changing coordinates (local diffeomorphism) with the use of closed feedback loop. The necessary conditions that must be fulfilled by nonlinear system to enable carrying out linearizing procedures are presented. Numerical solutions of state equations for the nonlinear system and equivalent linearized system are included.

  1. Uncoupling of the functions of the Arabidopsis VIP1 protein in transient and stable plant genetic transformation by Agrobacterium

    PubMed Central

    Li, Jianxiong; Krichevsky, Alexander; Vaidya, Manjusha; Tzfira, Tzvi; Citovsky, Vitaly

    2005-01-01

    Agrobacterium-mediated genetic transformation of plants, a unique example of transkingdom DNA transfer, requires the presence of several proteins encoded by the host cell. One such cellular factor is VIP1, an Arabidopsis protein proposed to interact with and facilitate import of the bacterial DNA–protein transport (T) complexes into the plant cell nucleus. Thus, VIP1 is required for transient expression of the bacterial DNA, an early step in the transformation process. However, the role of VIP1 in subsequent transformation events leading to the stable expression of bacterial DNA was unexplored. Here, we used reverse genetics to dissect VIP1 functionally and demonstrate its involvement in the stable genetic transformation of Arabidopsis plants by Agrobacterium. Our data indicate that the ability of VIP1 to interact with the VirE2 protein component of the T-complex and localize to the cell nucleus is sufficient for transient genetic transformation, whereas its ability to form homomultimers and interact with the host cell H2A histone in planta is required for tumorigenesis and, by implication, stable genetic transformation. PMID:15824315

  2. Development of a transformation system for the flavinogenic yeast Candida famata.

    PubMed

    Voronovsky, Andriy A; Abbas, Charles A; Fayura, Lyubov R; Kshanovska, Barbara V; Dmytruk, Kostyantyn V; Sybirna, Kateryna A; Sibirny, Andriy A

    2002-08-01

    Riboflavin-overproducing mutants of the flavinogenic yeast Candida famata are used for industrial riboflavin production. This paper describes the development of an efficient transformation system for this species. Leucine-deficient mutants have been isolated from C. famata VKM Y-9 wild-type strain. Among them leu2 mutants were identified by transformation to leucine prototrophy with plasmids YEp13 and PRpL2 carrying the Saccharomyces cerevisiae LEU2 gene. DNA fragments (called CfARSs) conferring increased transformation frequencies and extrachromosomal replication were isolated from a C. famata gene library constructed on the integrative vector containing the S. cerevisiae LEU2 gene as a selective marker. The smallest cloned fragment (CfARS16) has been sequenced. This one had high adenine plus thymine (A+T) base pair content and a sequence homologous to the S. cerevisiae ARS Consensus Sequence. Methods for spheroplast transformation and electrotransformation of the yeast C. famata were optimized. They conferred high transformation frequencies (up to 10(5) transformants per microg DNA) with a C. famata leu2 mutant using replicative plasmids containing the S. cerevisiae LEU2 gene as a selective marker. Riboflavin-deficient mutants were isolated from the C. famata leu2 strain and their biochemical identification was carried out. Using the developed transformation system, several C. famata genomic fragments complementing mutations of structural genes for riboflavin biosynthesis (coding for GTP cyclohydrolase, reductase, dihydroxybutanone phosphate synthase and riboflavin synthase, respectively) have been cloned. PMID:12702288

  3. Bäcklund transformations for the elliptic Gaudin model and a Clebsch system

    NASA Astrophysics Data System (ADS)

    Zullo, Federico

    2011-07-01

    A two-parameters family of Bäcklund transformations for the classical elliptic Gaudin model is constructed. The maps are explicit, symplectic, preserve the same integrals as for the continuous flows, and are a time discretization of each of these flows. The transformations can map real variables into real variables, sending physical solutions of the equations of motion into physical solutions. The starting point of the analysis is the integrability structure of the model. It is shown how the analogue transformations for the rational and trigonometric Gaudin model are a limiting case of this one. An application to a particular case of the Clebsch system is given.

  4. Linear-algebraic bath transformation for simulating complex open quantum systems

    NASA Astrophysics Data System (ADS)

    Huh, Joonsuk; Mostame, Sarah; Fujita, Takatoshi; Yung, Man-Hong; Aspuru-Guzik, Alán

    2014-12-01

    In studying open quantum systems, the environment is often approximated as a collection of non-interacting harmonic oscillators, a configuration also known as the star-bath model. It is also well known that the star-bath can be transformed into a nearest-neighbor interacting chain of oscillators. The chain-bath model has been widely used in renormalization group approaches. The transformation can be obtained by recursion relations or orthogonal polynomials. Based on a simple linear algebraic approach, we propose a bath partition strategy to reduce the system-bath coupling strength. As a result, the non-interacting star-bath is transformed into a set of weakly coupled multiple parallel chains. The transformed bath model allows complex problems to be practically implemented on quantum simulators, and it can also be employed in various numerical simulations of open quantum dynamics.

  5. "NONLINEAR DYNAMIC SYSTEMS RESPONSE TO NON-STATIONARY EXCITATION USING THE WAVELET TRANSFORM"

    SciTech Connect

    SPANOS, POL D.

    2006-01-15

    The objective of this research project has been the development of techniques for estimating the power spectra of stochastic processes using wavelet transform, and the development of related techniques for determining the response of linear/nonlinear systems to excitations which are described via the wavelet transform. Both of the objectives have been achieved, and the research findings have been disseminated in papers in archival journals and technical conferences.

  6. Educating a New Majority: Transforming America's Educational System for Diversity.

    ERIC Educational Resources Information Center

    Rendon, Laura I., Ed.; Hope, Richard O., Ed.

    This book presents 20 papers on the current status and future needs of disadvantaged minority students in the elementary, secondary, and higher education systems. Papers are grouped into four sections: current challenges to minority education; restructuring schools to foster minority student success; reforming higher education; and leadership…

  7. New! Improved? The Transformation of the Global Agrifood System

    ERIC Educational Resources Information Center

    Busch, Lawrence; Bain, Carmen

    2004-01-01

    The last decade has witnessed a dramatic rise in global trade in food and agricultural products. While much analysis has focused on the role of the world Trade Organization (WTO) in this process, we argue that other forms of regulation are of far greater consequence. In this paper, we examine changes in the agrifood system made possible by the…

  8. Note: An improved calibration system with phase correction for electronic transformers with digital output

    SciTech Connect

    Cheng, Han-miao Li, Hong-bin

    2015-08-15

    The existing electronic transformer calibration systems employing data acquisition cards cannot satisfy some practical applications, because the calibration systems have phase measurement errors when they work in the mode of receiving external synchronization signals. This paper proposes an improved calibration system scheme with phase correction to improve the phase measurement accuracy. We employ NI PCI-4474 to design a calibration system, and the system has the potential to receive external synchronization signals and reach extremely high accuracy classes. Accuracy verification has been carried out in the China Electric Power Research Institute, and results demonstrate that the system surpasses the accuracy class 0.05. Furthermore, this system has been used to test the harmonics measurement accuracy of all-fiber optical current transformers. In the same process, we have used an existing calibration system, and a comparison of the test results is presented. The system after improvement is suitable for the intended applications.

  9. A polarized digital shearing speckle pattern interferometry system based on temporal wavelet transformation.

    PubMed

    Feng, Ziang; Gao, Zhan; Zhang, Xiaoqiong; Wang, Shengjia; Yang, Dong; Yuan, Hao; Qin, Jie

    2015-09-01

    Digital shearing speckle pattern interferometry (DSSPI) has been recognized as a practical tool in testing strain. The DSSPI system which is based on temporal analysis is attractive because of its ability to measure strain dynamically. In this paper, such a DSSPI system with Wollaston prism has been built. The principles and system arrangement are described and the preliminary experimental result of the displacement-derivative test of an aluminum plate is shown with the wavelet transformation method and the Fourier transformation method. The simulations have been conducted with the finite element method. The comparison of the results shows that quantitative measurement of displacement-derivative has been realized.

  10. Combining discrete cosine transform with clipping for PAPR reduction in intensity-modulated OFDM systems

    NASA Astrophysics Data System (ADS)

    Wang, Zhong-peng; Chen, Shou-fa; Zhou, Yang; Chen, Ming; Tang, Jin; Chen, Lin

    2014-09-01

    In this paper, the peak-to-average power ratio (PAPR) of orthogonal frequency division multiplexing (OFDM) signal is reduced by combining the discrete cosine transform (DCT) with clipping in optical intensity-modulated direct-detection (IM/DD) OFDM systems. First, the data are transformed into new modified data by DCT. Second, the proposed scheme utilizes the clipping technique to further reduce the PAPR of OFDM signal. We experimentally demonstrate that the optical OFDM transmission system with this proposed scheme can achieve significant performance improvement in terms of PAPR and bit error rate (BER) compared with the original optical OFDM systems.

  11. A polarized digital shearing speckle pattern interferometry system based on temporal wavelet transformation.

    PubMed

    Feng, Ziang; Gao, Zhan; Zhang, Xiaoqiong; Wang, Shengjia; Yang, Dong; Yuan, Hao; Qin, Jie

    2015-09-01

    Digital shearing speckle pattern interferometry (DSSPI) has been recognized as a practical tool in testing strain. The DSSPI system which is based on temporal analysis is attractive because of its ability to measure strain dynamically. In this paper, such a DSSPI system with Wollaston prism has been built. The principles and system arrangement are described and the preliminary experimental result of the displacement-derivative test of an aluminum plate is shown with the wavelet transformation method and the Fourier transformation method. The simulations have been conducted with the finite element method. The comparison of the results shows that quantitative measurement of displacement-derivative has been realized. PMID:26429424

  12. Carbon Nanotube Computer: Transforming Scientific Discoveries into Working Systems

    NASA Astrophysics Data System (ADS)

    Mitra, Subhasish

    2014-03-01

    The miniaturization of electronic devices has been the principal driving force behind the semiconductor industry, and has brought about major improvements in computational power and energy efficiency. Although advances with silicon-based electronics continue to be made, alternative technologies are being explored. Digital circuits based on transistors fabricated from carbon nanotubes (CNTs) have the potential to outperform silicon by improving the energy- delay product, a metric of energy efficiency, by more than an order of magnitude. Hence, CNTs are an exciting complement to existing semiconductor technologies. However, carbon nanotubes (CNTs) are subject to substantial inherent imperfections that pose major obstacles to the design of robust and very large-scale CNFET digital systems: (i) It is nearly impossible to guarantee perfect alignment and positioning of all CNTs. This limitation introduces stray conducting paths, resulting in incorrect circuit functionality. (ii) CNTs can be metallic or semiconducting depending on chirality. Metallic CNTs cause shorts resulting in excessive leakage and incorrect circuit functionality. A combination of design and processing technique overcomes these challenges by creating robust CNFET digital circuits that are immune to these inherent imperfections. This imperfection-immune design paradigm enables the first experimental demonstration of the carbon nanotube computer, and, more generally, arbitrary digital systems that can be built using CNFETs. The CNT computer is capable of performing multitasking: as a demonstration, we perform counting and integer-sorting simultaneously. In addition, we emulate 20 different instructions from the commercial MIPS instruction set to demonstrate the generality of our CNT computer. This is the most complex carbon-based electronic system yet demonstrated. It is a considerable advance because CNTs are prominent among a variety of emerging technologies that are being considered for the next

  13. The Transforming Earth System Science Education (TESSE) program

    NASA Astrophysics Data System (ADS)

    Graham, K. J.; Bryce, J. G.; Brown, D.; Darwish, A.; Finkel, L.; Froburg, E.; Furman, T.; Guertin, L.; Hale, S. R.; Johnson, J.; Porter, W.; Smith, M.; Varner, R.; von Damm, K.

    2007-12-01

    A partnership between the University of New Hampshire (UNH), Dillard University, Elizabeth City State University, and Pennsylvania State University has been established to prepare middle and high school teachers to teach Earth and environmental sciences from a processes and systems approach. Specific project goals include: providing Earth system science content instruction; assisting teachers in implementing Earth system science in their own classrooms; and creating opportunities for pre-service teachers to experience authentic research with Earth scientists. TESSE programmatic components comprise (1) a two-week intensive summer institutes for current and future teachers; (2) eight-week research immersion experiences that match preservice teachers with Earth science faculty mentors; and (3) a science liaison program involving the pairing of inservice teachers with graduate students or future teachers. The first year of the program supported a total of 49 participants (42 inservice and preservice teachers, as well as 7 graduate fellows). All participants in the program attended an intensive two-week summer workshop at UNH, and the academic-year science liaison program is underway. In future summers, all partnering institutions will hold similar two-week summer institutes. UNH will offer a more advanced course geared towards "hot topics" and research techniques in the Earth and environmental sciences.

  14. Speckle-based metrology systems and linear canonical transform photography in mixed domains

    NASA Astrophysics Data System (ADS)

    Ward, Jennifer E.; Kelly, Damien P.; Patten, Robert F.; Hennelly, Bryan M.; Liu, Ying; Sheridan, John T.

    2006-04-01

    It has been shown that complex paraxial optical systems, consisting of various lens and distances of free space propagation, can be described using the Linear Canonical Transform (LCT). Indeed it can be shown that many well know optical transforms such as the Optical Fourier Transform (OFT), Optical Fractional Fourier Transform (OFRT), the effect of a lens or Chirp Modulation Transform (CMT) are all subsets of the more general LCT. Using the ABCD Collins matrix formula it is possible to represent these integral transforms in a simpler form, which facilitates system analysis and design. Speckle Photography (SP) in combination with an optical LCT can be used to measure surface motion of an optically rough body. It has previously been shown that Optical FRT's (OFRT) can be used in speckle based metrology systems to vary the range and sensitivity of a metrology system and also to determine both, the magnitude and direction, of tilting (rotation) and translation motion simultaneously, provided that the motion is captured in two separate OFRT domains. In this paper we extend the OFRT analysis to more general LCT systems and demonstrate how simultaneous tilt and translation measurements can be discerned from the speckle images captured prior to, and after motion. A spherical wavefront can be conveniently described using the Collin's matrix notation. By changing the wavefront of the illuminating light we show that we effectively change the domain of the LCT system without changing the bulk elements in the optical system. Thus the complete motion (in-plane translation and small surface tilting) of a rigid body can be determined using one optical LCT system and illuminating fields of varying curvature.

  15. The Relationship between Chief Information Officer Transformational Leadership and Computing Platform Operating Systems

    ERIC Educational Resources Information Center

    Anderson, George W.

    2010-01-01

    The purpose of this study was to relate the strength of Chief Information Officer (CIO) transformational leadership behaviors to 1 of 5 computing platform operating systems (OSs) that may be selected for a firm's Enterprise Resource Planning (ERP) business system. Research shows executive leader behaviors may promote innovation through the use of…

  16. The Transformation of Ergonomic Affordances into Cultural Affordances: The Case of the Alnuset System

    ERIC Educational Resources Information Center

    Chiappini, Giampaolo

    2012-01-01

    Is it possible to study the ergonomic affordances offered by a system designed for educational aims and their transformation into cultural affordances? To this purpose, what references can we adopt? This work describes the theoretical framework used to realise this study referring to AlNuSet, a system realised within the EC ReMath project to…

  17. Step-up-to-Excellence: A Change Navigation Protocol for Transforming School Systems

    ERIC Educational Resources Information Center

    Duffy, Francis M.

    2006-01-01

    This article presents a protocol change leaders can use to navigate whole-system change in their school districts. The information describing the protocol will help change leaders in school districts and policymakers interested in whole-district change answer the question, "How do we transform our entire school system"? The protocol is…

  18. Transformation formulas relating geodetic coordinates to a tangent to Earth, plane coordinate system

    NASA Technical Reports Server (NTRS)

    Credeur, L.

    1981-01-01

    Formulas and their approximation were developed to map geodetic position to an Earth tangent plane with an airport centered rectangular coordinate system. The transformations were developed for use in a terminal area air traffic model with deterministic aircraft traffic. The exact configured vehicle's approximation equations used in their precision microwave landing system navigation experiments.

  19. Concordium 2015: Strategic Uses of Evidence to Transform Delivery Systems

    PubMed Central

    Holve, Erin; Weiss, Samantha

    2016-01-01

    In September 2015 the EDM Forum hosted AcademyHealth’s newest national conference, Concordium. The 11 papers featured in the eGEMs “Concordium 2015” special issue successfully reflect the major themes and issues discussed at the meeting. Many of the papers address informatics or methodological approaches to natural language processing (NLP) or text analysis, which is indicative of the importance of analyzing text data to gain insights into care coordination and patient-centered outcomes. Perspectives on the tools and infrastructure requirements that are needed to build learning health systems were also recurrent themes.

  20. Concordium 2015: Strategic Uses of Evidence to Transform Delivery Systems

    PubMed Central

    Holve, Erin; Weiss, Samantha

    2016-01-01

    In September 2015 the EDM Forum hosted AcademyHealth’s newest national conference, Concordium. The 11 papers featured in the eGEMs “Concordium 2015” special issue successfully reflect the major themes and issues discussed at the meeting. Many of the papers address informatics or methodological approaches to natural language processing (NLP) or text analysis, which is indicative of the importance of analyzing text data to gain insights into care coordination and patient-centered outcomes. Perspectives on the tools and infrastructure requirements that are needed to build learning health systems were also recurrent themes. PMID:27683671

  1. Concordium 2015: Strategic Uses of Evidence to Transform Delivery Systems.

    PubMed

    Holve, Erin; Weiss, Samantha

    2016-01-01

    In September 2015 the EDM Forum hosted AcademyHealth's newest national conference, Concordium. The 11 papers featured in the eGEMs "Concordium 2015" special issue successfully reflect the major themes and issues discussed at the meeting. Many of the papers address informatics or methodological approaches to natural language processing (NLP) or text analysis, which is indicative of the importance of analyzing text data to gain insights into care coordination and patient-centered outcomes. Perspectives on the tools and infrastructure requirements that are needed to build learning health systems were also recurrent themes.

  2. [Casalud: innovations for the transformation of health systems].

    PubMed

    Tapia-Conyer, Roberto; Gallardo-Rincón, Héctor; Saucedo-Martínez, Rodrigo

    2012-01-01

    The increasing prevalence of non-communicable diseases (NCDs) has impacted the health systems by demanding unbearable costs of care and diminishing people's quality of life. The Instituto Carlos Slim de la Salud, a non-profit organization, has developed the model Casalud to convene partnerships with state governments to establish a reengineering process of primary care clinics, improving the delivery of services and leveraging on the different technological platforms to connect them with the households. The model Casalud strengthens the delivery of services and increases the timely demand of care. It is actually implemented in seven states seeking a progressive scale-up. PMID:23254716

  3. Transforming the NAS: The Next Generation Air Traffic Control System

    NASA Technical Reports Server (NTRS)

    Erzberger, Heinz

    2004-01-01

    The next-generation air traffic control system must be designed to safely and efficiently accommodate the large growth of traffic expected in the near future. It should be sufficiently scalable to contend with the factor of 2 or more increase in demand expected by the year 2020. Analysis has shown that the current method of controlling air traffic cannot be scaled up to provide such levels of capacity. Therefore, to achieve a large increase in capacity while also giving pilots increased freedom to optimize their flight trajectories requires a fundamental change in the way air traffic is controlled. The key to achieving a factor of 2 or more increase in airspace capacity is to automate separation monitoring and control and to use an air-ground data link to send trajectories and clearances directly between ground-based and airborne systems. In addition to increasing capacity and offering greater flexibility in the selection of trajectories, this approach also has the potential to increase safety by reducing controller and pilot errors that occur in routine monitoring and voice communication tasks.

  4. Use of Agrobacterium rhizogenes strain 18r12v and paromomycin selection for transformation of Brachypodium distachyon and Brachypodium sylvaticum

    DOE PAGES

    Collier, Ray; Bragg, Jennifer; Hernandez, Bryan T.; Vogel, John P.; Thilmony, Roger

    2016-05-24

    In this study, the genetic transformation of monocot grasses is a resource intensive process, the quality and efficiency of which is dependent in part upon the method of DNA introduction, as well as the ability to effectively separate transformed from wildtype tissue. Agrobacterium-mediated transformation of Brachypodium has relied mainly on Agrobacterium tumefaciens strain AGL1. Currently the antibiotic hygromycin B has been the selective agent of choice for robust identification of transgenic calli in Brachypodium distachyon and Brachypodium sylvaticum but few other chemicals have been shown to work as well for selection of transgenic Brachypodium cells in tissue culture. This studymore » demonstrates that Agrobacterium rhizogenes strain 18r12v and paromomycin selection can be successfully used for the efficient generation of transgenic B. distachyon and B. sylvaticurn. Additionally we observed that the transformation rates were similar to or higher than those obtained with A. turnefaciens strain AGL1 and hygromycin selection. The A. rhizogenes strain 18r12v harboring the pARS1 binary vector and paromomycin selection is an effective means of generating transgenic Brachypodium plants. This novel approach will facilitate the transgenic complementation of T-DNA knockout mutants of B. distachyon which were created using hygromycin selection, as well as aid the implementation of more complex genome manipulation strategies which require multiple rounds of transformation.« less

  5. Use of Agrobacterium rhizogenes Strain 18r12v and Paromomycin Selection for Transformation of Brachypodium distachyon and Brachypodium sylvaticum

    PubMed Central

    Collier, Ray; Bragg, Jennifer; Hernandez, Bryan T.; Vogel, John P.; Thilmony, Roger

    2016-01-01

    The genetic transformation of monocot grasses is a resource intensive process, the quality and efficiency of which is dependent in part upon the method of DNA introduction, as well as the ability to effectively separate transformed from wildtype tissue. Agrobacterium-mediated transformation of Brachypodium has relied mainly on Agrobacterium tumefaciens strain AGL1. Currently the antibiotic hygromycin B has been the selective agent of choice for robust identification of transgenic calli in Brachypodium distachyon and Brachypodium sylvaticum but few other chemicals have been shown to work as well for selection of transgenic Brachypodium cells in tissue culture. This study demonstrates that Agrobacterium rhizogenes strain 18r12v and paromomycin selection can be successfully used for the efficient generation of transgenic B. distachyon and B. sylvaticum. Additionally we observed that the transformation rates were similar to or higher than those obtained with A. tumefaciens strain AGL1 and hygromycin selection. The A. rhizogenes strain 18r12v harboring the pARS1 binary vector and paromomycin selection is an effective means of generating transgenic Brachypodium plants. This novel approach will facilitate the transgenic complementation of T-DNA knockout mutants of B. distachyon which were created using hygromycin selection, as well as aid the implementation of more complex genome manipulation strategies which require multiple rounds of transformation. PMID:27252729

  6. Use of Agrobacterium rhizogenes Strain 18r12v and Paromomycin Selection for Transformation of Brachypodium distachyon and Brachypodium sylvaticum.

    PubMed

    Collier, Ray; Bragg, Jennifer; Hernandez, Bryan T; Vogel, John P; Thilmony, Roger

    2016-01-01

    The genetic transformation of monocot grasses is a resource intensive process, the quality and efficiency of which is dependent in part upon the method of DNA introduction, as well as the ability to effectively separate transformed from wildtype tissue. Agrobacterium-mediated transformation of Brachypodium has relied mainly on Agrobacterium tumefaciens strain AGL1. Currently the antibiotic hygromycin B has been the selective agent of choice for robust identification of transgenic calli in Brachypodium distachyon and Brachypodium sylvaticum but few other chemicals have been shown to work as well for selection of transgenic Brachypodium cells in tissue culture. This study demonstrates that Agrobacterium rhizogenes strain 18r12v and paromomycin selection can be successfully used for the efficient generation of transgenic B. distachyon and B. sylvaticum. Additionally we observed that the transformation rates were similar to or higher than those obtained with A. tumefaciens strain AGL1 and hygromycin selection. The A. rhizogenes strain 18r12v harboring the pARS1 binary vector and paromomycin selection is an effective means of generating transgenic Brachypodium plants. This novel approach will facilitate the transgenic complementation of T-DNA knockout mutants of B. distachyon which were created using hygromycin selection, as well as aid the implementation of more complex genome manipulation strategies which require multiple rounds of transformation. PMID:27252729

  7. Use of Agrobacterium rhizogenes Strain 18r12v and Paromomycin Selection for Transformation of Brachypodium distachyon and Brachypodium sylvaticum.

    PubMed

    Collier, Ray; Bragg, Jennifer; Hernandez, Bryan T; Vogel, John P; Thilmony, Roger

    2016-01-01

    The genetic transformation of monocot grasses is a resource intensive process, the quality and efficiency of which is dependent in part upon the method of DNA introduction, as well as the ability to effectively separate transformed from wildtype tissue. Agrobacterium-mediated transformation of Brachypodium has relied mainly on Agrobacterium tumefaciens strain AGL1. Currently the antibiotic hygromycin B has been the selective agent of choice for robust identification of transgenic calli in Brachypodium distachyon and Brachypodium sylvaticum but few other chemicals have been shown to work as well for selection of transgenic Brachypodium cells in tissue culture. This study demonstrates that Agrobacterium rhizogenes strain 18r12v and paromomycin selection can be successfully used for the efficient generation of transgenic B. distachyon and B. sylvaticum. Additionally we observed that the transformation rates were similar to or higher than those obtained with A. tumefaciens strain AGL1 and hygromycin selection. The A. rhizogenes strain 18r12v harboring the pARS1 binary vector and paromomycin selection is an effective means of generating transgenic Brachypodium plants. This novel approach will facilitate the transgenic complementation of T-DNA knockout mutants of B. distachyon which were created using hygromycin selection, as well as aid the implementation of more complex genome manipulation strategies which require multiple rounds of transformation.

  8. High reliability transformation of the wheat pathogen Bipolaris sorokiniana using Agrobacterium tumefaciens.

    PubMed

    Nizam, Shadab; Verma, Sandhya; Singh, Kunal; Aggarwal, Rashmi; Srivastava, Krishna Dutt; Verma, Praveen K

    2012-03-01

    Bipolaris sorokiniana, the causal agent of spot blotch of wheat, significantly reduces grain yield worldwide. In order to study pathogenic mechanisms of the fungus, conditions for efficient transformation using Agrobacterium-mediated transformation were investigated. To study different stages of hyphal fusion and pathogenic mechanisms of the fungus, two fluorescence markers viz. the red fluorescent protein (DsRed-Express) and the green fluorescent protein (EGFP1) were constitutively expressed. Southern hybridizations confirmed the presence of T-DNA in all hygromycin B or geneticin resistant transformants, and also showed random and single copy integration. Fluorescence microscopy suggested the high level expression of both DsRed and EGFP fluorescent proteins in spores and mycelia. The results signify that DsRed and EGFP can be used as efficient reporter gene for monitoring B. sorokiniana hyphal fusion as well as colonization in the host tissues. This work will be useful to develop methodologies for understanding the mechanisms of Bipolaris-wheat interaction and functional genomics of B. sorokiniana for various applications including insertional mutagenesis, targeted disruption of specific genes, ectopic complementation of loss-of-function strains and over-expression.

  9. MHD spectra and coordinate transformations in toroidal systems

    NASA Astrophysics Data System (ADS)

    Predebon, I.; Momo, B.; Terranova, D.; Innocente, P.

    2016-09-01

    A fully geometric approach is used to study the mutual spectral properties of different classes of MHD equilibria with symmetries defined by action-angle coordinates. We refer mostly to the helical equilibria of tokamak and reversed field pinch plasmas compared to the axisymmetric counterpart. Based on the existence of different coordinate systems, we show how the magnetic field and the vector potential are correspondingly evaluated, and how their spectral decompositions may largely vary. Most notably, a monochromatic perturbation in a reference frame can appear with a rich spectrum in another frame. The consequences on the interpretation of the measurements and the implications on the effectiveness of the externally applied magnetic fields on the plasma are discussed with practical examples.

  10. Optimized use of Hough transform in an ultrasound measurement system for human signature analysis

    NASA Astrophysics Data System (ADS)

    Ricci, R.; Sona, A.

    2013-05-01

    Human motion analysis is a task of increasing importance in several modern application fields, such as in medicine, avionics, security, and disaster recovery. In this paper, the use of Hough transform is considered and discussed in the scenario of human motion analysis. In particular, the influence of some transform parameters is investigated with the aim of improving Hough transform set-up when used in a measurement approach for human motion analysis. To this purpose, Hough transform has been applied to a set of results obtained by exploiting a suitable measurement system developed by the same authors, in the specific case of ultrasound waves. Such results have been obtained by using some reference images in the form of spectrograms achieved by using the system along with a purposely developed reference target emulating some human body movements. The results show that the considered measurement system, and more generally the human motion analysis and detection system, can be optimized by a proper set-up and use of Hough transform algorithm.

  11. Mechanism of Occurring Over-Voltage Phenomena in Distributed Power System on Energization of Transformers

    NASA Astrophysics Data System (ADS)

    Nakachi, Yoshiki; Ueda, Fukashi; Kajikawa, Takuya; Amau, Tooru; Kameyama, Hirokazu; Ito, Hisanori

    This paper verifies the mechanism of occurring over voltage phenomena in the distributed power system on energizing the transformer. This over-voltage, which is observed at the actual distributed power system, with heavy inrush current is found to occur at about 0.1-0.2sec after the energizing and continue for a duration of more than 0.1[sec]. There is a concern that this over-voltage may operate the protection relay and deteriorate the insulation of apparatus. It is basically caused by the resonance between the shunt capacitors and saturated/unsaturated magnetizing inductance of transformer, system inductance. By using analytical formulation of a simple equivalent circuit, its mechanism has been verified through simulations carried out by using EMTP. Moreover, the sympathetic interaction between transformers is prolonged the duration of the over-voltage by the field test data is discussed in this paper.

  12. Children's behavioral health system transformation: one state's context and strategies for sustained change.

    PubMed

    Harburger, Deborah S; Stephan, Sharon H; Kaye, Sarah

    2013-10-01

    The purpose of this paper was to examine the State of Maryland as a case study of sustained change efforts in the service delivery system for children with significant behavioral health needs and their families. A punctuated equilibrium paradigm is introduced to describe Maryland's behavioral health system transformation over the course of three decades. The context and specific strategies that characterized Maryland's execution of its recent Mental Health Transformation State Incentive Grant are highlighted. There is a discussion of one of the pinnacle achievements of Maryland's transformation efforts, the recent statewide establishment of care management entities for children with behavioral health challenges, and its implications for behavioral health in the context of health care reform changes. This case study illustrates how a state can systematically and incrementally develop systems of care for children and families that are values-based, sustainable, and flexible.

  13. Spatial instability of the rift in the St. Paul multifault transform fracture system, Atlantic Ocean

    NASA Astrophysics Data System (ADS)

    Sokolov, S. Yu.; Zaraiskaya, Yu. A.; Mazarovich, A. O.; Efimov, V. N.; Sokolov, N. S.

    2016-05-01

    The structure of the acoustic basement of the eastern part of the St. Paul multifault transform fracture system hosts rift paleovalleys and a paleonodal depression that mismatch the position of the currently active zones. This displacement zone, which is composed of five fault troughs, is unstable in terms of the position of the rift segments, which jumped according to redistribution of stresses. The St. Paul system is characterized by straightening of the transform transition between two remote segments of the Mid-Atlantic Ridge (MAR). The eastern part of the system contains anomalous bright-spot-like reflectors on the flattened basement, which is a result of atypical magmatism, that forms the standard ridge relief of the acoustic basement. Deformations of the acoustic basement have a presedimentation character. The present-day deformations with lower amplitude in comparison to the basement are accompanied by acoustic brightening of the sedimentary sequence. The axial Bouguer anomalies in the east of the system continue to the north for 120 km from the active segments of the St. Paul system. Currently seismically active segments of the spreading system are characterized by increasing amplitudes of the E-W displacement along the fault troughs. Cross-correlation of the lengths of the active structural elements of the MAR zone (segments of the ridge and transform fracture zones of displacement) indicates that, statistically, the multifault transform fracture system is a specific type of oceanic strike-slip faults.

  14. Fuel salt and container material studies for MOSART transforming system

    SciTech Connect

    Ignatiev, V.; Feynberg, O.; Merzlyakov, A.; Surenkov, A.; Zagnitko, A.; Afonichkin, V.; Bovet, A.; Khokhlov, V.; Subbotin, V.; Gordeev, M.; Panov, A.; Toropov, A.

    2013-07-01

    A study is under progress to examine the feasibility of single stream Molten Salt Actinide Recycling and Transmuting system without and with Th support (MOSART) fuelled with different compositions of actinide tri-fluorides (AnF{sub 3}) from used LWR fuel. New fast-spectrum design options with homogeneous core and fuel salts with high enough solubility for AnF{sub 3} are being examined because of new goals. The flexibility of single fluid MOSART concept with Th support is underlined, particularly, possibility of its operation in self-sustainable mode (Conversion Ratio: CR=1) using different loadings and make up. The paper summarizes the most current status of fuel salt and container material data for the MOSART concept received within ISTC-3749 and ROSATOM-MARS projects. Key physical and chemical properties of various fluoride fuel salts are reported. The issues like salt purification, the electroreduction of U(IV) to U(III) in LiF-ThF{sub 4} and the electroreduction of Yb(III) to Yb(II) in LiF-NaF are detailed.

  15. Transforming mental health and substance abuse data systems in the United States.

    PubMed

    Coffey, Rosanna M; Buck, Jeffrey A; Kassed, Cheryl A; Dilonardo, Joan; Forhan, Carol; Marder, William D; Vandivort-Warren, Rita

    2008-11-01

    State efforts to improve mental health and substance abuse service systems cannot overlook the fragmented data systems that reinforce the historical separateness of systems of care. These separate systems have discrete approaches to treatment, and there are distinct funding streams for state mental health, substance abuse, and Medicaid agencies. Transforming mental health and substance abuse services in the United States depends on resolving issues that underlie separate treatment systems--access barriers, uneven quality, disjointed coordination, and information silos across agencies and providers. This article discusses one aspect of transformation--the need for interoperable information systems. It describes current federal and state initiatives for improving data interoperability and the special issue of confidentiality associated with mental health and substance abuse treatment data. Some achievable steps for states to consider in reforming their behavioral health data systems are outlined. The steps include collecting encounter-level data; using coding that is compliant with the Health Insurance Portability and Accountability Act, including national provider identifiers; forging linkages with other state data systems and developing unique client identifiers among systems; investing in flexible and adaptable data systems and business processes; and finding innovative solutions to the difficult confidentiality restrictions on use of behavioral health data. Changing data systems will not in itself transform the delivery of care; however, it will enable agencies to exchange information about shared clients, to understand coordination problems better, and to track successes and failures of policy decisions.

  16. CGH calculation with the ray tracing method for the Fourier transform optical system.

    PubMed

    Ichikawa, Tsubasa; Yoneyama, Takuo; Sakamoto, Yuji

    2013-12-30

    Computer-generated holograms (CGHs) are usually displayed on electronic devices. However, the resolution of current output devices is not high enough to display CGHs, so the visual field is very narrow. A method using a Fourier transform optical system has been proposed, to enlarge the size of reconstructed images. This paper describes a method of CGH calculations for the Fourier transform optical system to enlarge the visual field and reconstruct realistic images by using the ray tracing method. This method reconstructs images at arbitrary depths and also eliminates unnecessary light including zero-th order light.

  17. Electromagnetic wave manipulation by layered systems using the transformation media concept

    NASA Astrophysics Data System (ADS)

    Chen, Huanyang; Chan, C. T.

    2008-08-01

    We show that the optical properties of an oblique layered system with two kinds of isotropic materials can be described using the concept of transformation media, as long as the thickness of the layers is much smaller than the wavelength. Once the connection with transformation media is established, we then show that oblique layered system can serve as a universal element to build a variety of interesting functional optical components such as wave splitters, wave combiners, one-dimensional cloaking devices, and reflectionless field rotators.

  18. Two-dimensional phase transformation probed by second harmonic generation: Oscillatory transformation of the K/Al(111) system

    SciTech Connect

    Ying, Z.C.; Plummer, E.W. |

    1995-12-31

    The technique of optical second harmonic generation is used to study phase transformations at two-dimensional surfaces and interfaces. Examples are given to illustrate that changes in surface symmetry, adsorption configuration, and electronic structure can be detected by this nonlinear optical technique. An oscillatory phase transformation of potassium adsorbed atoms on Al(111) probed by second harmonic generation is analyzed in detail.

  19. Color information verification system based on singular value decomposition in gyrator transform domains

    NASA Astrophysics Data System (ADS)

    Abuturab, Muhammad Rafiq

    2014-06-01

    A new color image security system based on singular value decomposition (SVD) in gyrator transform (GT) domains is proposed. In the encryption process, a color image is decomposed into red, green and blue channels. Each channel is independently modulated by random phase masks and then separately gyrator transformed at different parameters. The three gyrator spectra are joined by multiplication to get one gray ciphertext. The ciphertext is separated into U, S, and V parts by SVD. All the three parts are individually gyrator transformed at different transformation angles. The three encoded information can be assigned to different authorized users for highly secure verification. Only when all the authorized users place the U, S, and V parts in correct multiplication order in the verification system, the correct information can be obtained with all the right keys. In the proposed method, SVD offers one-way asymmetrical decomposition algorithm and it is an optimal matrix decomposition in a least-square sense. The transformation angles of GT provide very sensitive additional keys. The pre-generated keys for red, green and blue channels are served as decryption (private) keys. As all the three encrypted parts are the gray scale ciphertexts with stationary white noise distributions, which have camouflage property to some extent. These advantages enhance the security and robustness. Numerical simulations are presented to support the viability of the proposed verification system.

  20. A new inducible expression system in a transformed green alga, Chlorella vulgaris.

    PubMed

    Niu, Y F; Zhang, M H; Xie, W H; Li, J N; Gao, Y F; Yang, W D; Liu, J S; Li, H Y

    2011-01-01

    Genetic transformation is useful for basic research and applied biotechnology. However, genetic transformation of microalgae is usually quite difficult due to the technical limitations of existing methods. We cloned the promoter and terminator of the nitrate reductase gene from the microalga Phaeodactylum tricornutum and used them for optimization of a transformation system of the microalga Chlorella vulgaris. This species has been used for food production and is a promising candidate as a bioreactor for large-scale production of value-added proteins. A construct was made containing the CAT (chloramphenicol acetyltransferase) reporter gene driven by the nitrate reductase promoter. This construct was transferred into the C. vulgaris genome by electroporation. Expression of CAT in transgenic Chlorella conferred resistance to the antibiotic chloramphenicol and enabled growth in selective media. Overall efficiency for the transformation was estimated to be approximately 0.03%, which is relatively high compared with other available Chlorella transformation systems. Expression of CAT was induced in the presence of nitrate and inhibited in the presence of ammonium as a sole nitrogen source. This study presented an inducible recombinant gene expression system, also providing more gene regulation elements with potential for biotechnological applications.

  1. Mental Health System Transformation: Drivers for Change, Organizational Preparation, Engaging Partners and Outcomes.

    PubMed

    Corring, Deborah J; Whittall, Sandy; MustinPowell, Jill; Jarmain, Sarah; Chapman, Patty; Sussman, Sam

    2016-01-01

    St. Joseph's Health Care London (hereafter referred to as St. Joseph's) is a publicly funded hospital that has led mental health (MH) service system transformation in south west Ontario following directives from the Health Services Restructuring Commission (HSRC) (Sinclair 2000). This paper documents how provincial policy; HSRC directives; use of change management activities; organizational planning; and partnerships with other hospitals, community agencies and LHINs drove, shaped and accomplished the transformational change. The transformation included divestment of beds and related ambulatory services to four other hospitals, closure of beds and employment services and the construction of two state-of-the-art facilities. This paper documents the tracking of system performance measures and the outcomes that resulted.

  2. Mental Health System Transformation: Drivers for Change, Organizational Preparation, Engaging Partners and Outcomes.

    PubMed

    Corring, Deborah J; Whittall, Sandy; MustinPowell, Jill; Jarmain, Sarah; Chapman, Patty; Sussman, Sam

    2016-01-01

    St. Joseph's Health Care London (hereafter referred to as St. Joseph's) is a publicly funded hospital that has led mental health (MH) service system transformation in south west Ontario following directives from the Health Services Restructuring Commission (HSRC) (Sinclair 2000). This paper documents how provincial policy; HSRC directives; use of change management activities; organizational planning; and partnerships with other hospitals, community agencies and LHINs drove, shaped and accomplished the transformational change. The transformation included divestment of beds and related ambulatory services to four other hospitals, closure of beds and employment services and the construction of two state-of-the-art facilities. This paper documents the tracking of system performance measures and the outcomes that resulted. PMID:26854541

  3. AgarTrap-mediated genetic transformation using intact gemmae/gemmalings of the liverwort Marchantia polymorpha L.

    PubMed

    Tsuboyama-Tanaka, Shoko; Kodama, Yutaka

    2015-03-01

    The dioecious liverwort, Marchantia polymorpha L., is an emerging model plant. Various molecular biological techniques have been optimized for M. polymorpha for the past several years, and recently we reported a simplified Agrobacterium-mediated transformation method using sporelings (immature thalli from spores) of M. polymorpha. This method, termed AgarTrap (Agar-utilized Transformation with Pouring Solutions), completed by exchanging appropriate solutions on a single Petri dish to produce a sufficient number of independent transgenic sporelings. However, because spores are produced by crosses between males and females, the genetic backgrounds of resulting transgenic sporelings are not uniform. To easily produce transgenic liverworts with a uniform genetic background using AgarTrap, we developed an AgarTrap-mediated transformation method using intact gemmae/gemmalings produced by asexual reproduction. Using AgarTrap with male and female gemmae/gemmalings produced a sufficient number of independent transgenic gemmalings with uniform genetic backgrounds. The optimized transformation efficiencies were approximately 30 and 50 % in males and females, respectively. As with AgarTrap using sporelings, AgarTrap using intact gemmae/gemmalings will be useful in promoting studies of the molecular biology of M. polymorpha. PMID:25663453

  4. Cloning of TPS gene from eelgrass species Zostera marina and its functional identification by genetic transformation in rice.

    PubMed

    Zhao, Feng; Li, Qiuying; Weng, Manli; Wang, Xiuliang; Guo, Baotai; Wang, Li; Wang, Wei; Duan, Delin; Wang, Bin

    2013-12-01

    The full-length cDNA sequence (2613 bp) of the trehalose-6-phosphate synthase (TPS) gene of eelgrass Zostera marina (ZmTPS) was identified and cloned. Z. marina is a kind of seed-plant growing in sea water during its whole life history. The open reading frame (ORF) region of ZmTPS gene encodes a protein of 870 amino acid residues and a stop codon. The corresponding genomic DNA sequence is 3770 bp in length, which contains 3 exons and 2 introns. The ZmTPS gene was transformed into rice variety ZH11 via Agrobacterium-mediated transformation method. After antibiotic screening, molecular characterization, salt-tolerance and trehalose content determinations, two transgenic lines resistant to 150 mM NaCL solutions were screened. Our study results indicated that the ZmTPS gene was integrated into the genomic DNA of the two transgenic rice lines and could be expressed well. Moreover, the detection of the transformed ZmTPS gene in the progenies of the two transgenic lines was performed from T1 to T4 generations; and results suggested that the transformed ZmTPS gene can be transmitted from parent to the progeny in transgenic rice.

  5. A novel gateway-compatible binary vector series (PC-GW) for flexible cloning of multiple genes for genetic transformation of plants.

    PubMed

    Dalal, Jyoti; Yalamanchili, Roopa; La Hovary, Christophe; Ji, Mikyoung; Rodriguez-Welsh, Maria; Aslett, Denise; Ganapathy, Sowmya; Grunden, Amy; Sederoff, Heike; Qu, Rongda

    2015-09-01

    The rapidly advancing field of plant synthetic biology requires transforming plants with multiple genes. This has sparked a growing interest in flexible plant transformation vectors, which can be used for multi-gene transformations. We have developed a novel binary vector series, named the PC-GW series (GenBank: KP826769-KP826773), for Agrobacterium-mediated plant transformation. The PC-GW vectors use the pCAMBIA vector backbone, and contain NPTII, hpt, bar, mCherry or egfp genes as selectable markers for plant transformation. In a modified multiple cloning site (MCS) of the T-DNA region, we have placed the attR1, attR2 and ccdB sequences for rapid cloning of one to four genes by Gateway™-assisted recombination. In addition, we have introduced four meganuclease sites, and other restriction sites for multi-gene vector construction. Finally, we have placed a CaMV 35S promoter and a 35S terminator on the 5' and 3' ends of the MCS. The CaMV 35S promoter is flanked by PstI restriction sites that can be used to replace it with another promoter sequence if needed. The PC-GW vectors provide choices for selectable markers, cloning methods, and can accommodate up to eight gene constructs in a single T-DNA, thereby significantly reducing the number of transformations or crosses needed to generate multi-transgene expressing plants.

  6. Conceptualizing Teacher Identity as a Complex Dynamic System: The Inner Dynamics of Transformations during a Practicum

    ERIC Educational Resources Information Center

    Henry, Alastair

    2016-01-01

    Currently, the inner dynamics of teacher identity transformations remain a "black box." Conceptualizing preservice teacher identity as a complex dynamic system, and the notion of "being someone who teaches" in dialogical terms as involving shifts between different teacher voices, the study investigates the dynamical processes…

  7. Chemical Transformation System: Cloud Based Cheminformatic Services to Support Integrated Environmental Modeling

    EPA Science Inventory

    Integrated Environmental Modeling (IEM) systems that account for the fate/transport of organics frequently require physicochemical properties as well as transformation products. A myriad of chemical property databases exist but these can be difficult to access and often do not co...

  8. Postdispersion system for astronomical observations with Fourier transform spectrometers in the thermal infrared

    NASA Technical Reports Server (NTRS)

    Wiedermann, Guenter; Jennings, D. E.; Hanel, R. H.; Kunde, V. G.; Moseley, S. H.

    1989-01-01

    A postdispersion system for astronomical observations with Fourier transform spectrometers in the thermal infrared has been developed which improves the sensitivity of radiation noise limited observations by reducing the spectral range incident on the detector. Special attention is given to the first-generation blocked impurity band detector. Planetary, solar, and stellar observations are reported.

  9. Laplace and Z transforms of linear dynamical systems and conic sections

    NASA Astrophysics Data System (ADS)

    Rodrigo, Marianito R.

    2016-06-01

    We consider the solution trajectories of linear continuous and discrete dynamical systems and show that in the Laplace and Z transform spaces, respectively, they lie on the intersections of hypersurfaces described by second-degree polynomial equations. In particular, in two dimensions, these intersections are conic sections whose types are determined by the eigenvalues of the coefficient matrices.

  10. Transforming to a computerized system for nursing care: organizational success within Magnet idealism.

    PubMed

    Lindgren, Carolyn L; Elie, Leslie G; Vidal, Elizabeth C; Vasserman, Alex

    2010-01-01

    In reaching the goal for standardized, quality care, a not-for-profit healthcare system consisting of seven institutional entities is transforming nursing practice guidelines, patient care workflow, and patient documents into electronic, online, real-time modalities for use across departments and all healthcare delivery entities of the system. Organizational structure and a strategic plan were developed for the 2-year Clinical Transformation Project. The Siemens Patient Care Document System was adopted and adapted to the hospitals' documentation and information needs. Two fast-track sessions of more than 100 nurses and representatives from other health disciplines were held to standardize assessments, histories, care protocols, and interdisciplinary plans of care for the top 10 diagnostic regulatory groups. Education needs of the users were addressed. After the first year, a productive, functional system is evidenced. For example, the bar-coded Medication Administration Check System is in full use on the clinical units of one of the hospitals, and the other institutional entities are at substantial stages of implementation of Patient Care Documentation System. The project requires significant allocation of personnel and financial resources for a highly functional informatics system that will transform clinical care. The project exemplifies four of the Magnet ideals and serves as a model for others who may be deciding about launching a similar endeavor. PMID:20182156

  11. Transforming to a computerized system for nursing care: organizational success within Magnet idealism.

    PubMed

    Lindgren, Carolyn L; Elie, Leslie G; Vidal, Elizabeth C; Vasserman, Alex

    2010-01-01

    In reaching the goal for standardized, quality care, a not-for-profit healthcare system consisting of seven institutional entities is transforming nursing practice guidelines, patient care workflow, and patient documents into electronic, online, real-time modalities for use across departments and all healthcare delivery entities of the system. Organizational structure and a strategic plan were developed for the 2-year Clinical Transformation Project. The Siemens Patient Care Document System was adopted and adapted to the hospitals' documentation and information needs. Two fast-track sessions of more than 100 nurses and representatives from other health disciplines were held to standardize assessments, histories, care protocols, and interdisciplinary plans of care for the top 10 diagnostic regulatory groups. Education needs of the users were addressed. After the first year, a productive, functional system is evidenced. For example, the bar-coded Medication Administration Check System is in full use on the clinical units of one of the hospitals, and the other institutional entities are at substantial stages of implementation of Patient Care Documentation System. The project requires significant allocation of personnel and financial resources for a highly functional informatics system that will transform clinical care. The project exemplifies four of the Magnet ideals and serves as a model for others who may be deciding about launching a similar endeavor.

  12. The 3D representation of the new transformation from the terrestrial to the celestial system.

    NASA Astrophysics Data System (ADS)

    Dehant, V.; de Viron, O.; Capitaine, N.

    2006-08-01

    To study the sky from the Earth or to use navigation satellites, we need two reference systems, a celestial reference system, as fixed as possible with respect to the inertial frame, and a terrestrial reference system, rotating with the Earth. Additionally, we need a way to go from one reference system to the other. This transformation involves the Earth rotation rate, the polar motion, and the precession-nutation. This transformation is done using an intermediate system, in which the Earth rotation it-self is corrected for. Previously one used an intermediate system related to the equinox; the new paradigm involved a point, denoted the Celestial Intermediate Origin (CIO), which, due to its kinematical property of "Non Rotating Origin", allows better describing the length-of-day of the Earth. The use or not of the CIO only affects this intermediate frame. The new transformation system involving the CIO is additionally much simpler. Moreover, the use of the CIO allows an elegant separation between the polar motion, the precession nutation and the rotation rate variation. In this presentation we will show 3D representations that explain all this.

  13. Setting the Foundation for Transforming the U.S. Energy System

    SciTech Connect

    Davis, Jon M.; Virden, Jud W.; Walton, Terry L.; Brog, Terrence K.; Stiles, Dennis L.; Alderson, Karis P.; Melland, Jodi C.; Hobbs, Lori L.; Martinez, Sam J.; Quadrel, Marilyn J.; Quinn, Rod K.

    2008-12-29

    Setting the Foundation for Transforming the U.S. Energy System: Three Federal Action Plans for Near- and Long-term Success is a short document written for the incoming U.S. Federal Administration's Transistion Team. The document represents what we believe to be foundational elements of a broad campaign to make rapid progress towards these goals for the longterm transformation necessary to meet our national economic, security, and environmental goals. The three immediate action plans include: * Creating Tomorrow’s Electricity Infrastructure * Decarbonizing the Energy Economy *Enabling Science-Based Policy

  14. [Study on transformation of snowdrop lectin gene to chrysanthemum and aphid resistance of the transgenic plants].

    PubMed

    Wang, Guan-Lin; Liu, Yan-Hong; Guo, Shao-Hua; Wang, Yu; Ji, Yan; Fang, Hong-Jun

    2004-12-01

    Agrobacterium-mediated transformation in chrysanthemum was studied to prevent the insect pest of aphid (Mizus persicae). The gna gene was successfully transferred into chrysanthemum by leaf dish, and 93 transgenic clones were obtained. The highest transformation frequency 11.21% was achieved on the optimization facts, which were medium YEB with pH5.6, bacterial concentration OD600 = 0.4, precultivation for one day, cocultivation for four days, the cocultivation media supplemented with GA3 0.5 mg/L and leaf explants growed for 45 days. The results from PCR and FQ-PCR analysis confirmed that gna gene was integrated into the genome of chrysanthemum plants. The insect bioassay with aphid showed that the aphid resistance of different transgenic plants was difference, and the rate of aphid population inhibition of them were from 10% to 84% with an average rate of 39.4%. The leaf-extracts from different transgenic plants showed varying actinties in red-blood cell bioassay.

  15. A leaf-based regeneration and transformation system for maize (Zea mays L.).

    PubMed

    Ahmadabadi, Mohammad; Ruf, Stephanie; Bock, Ralph

    2007-08-01

    Efficient methods for in vitro propagation, regeneration, and transformation of plants are of pivotal importance to both basic and applied research. While being the world's major food crops, cereals are among the most difficult-to-handle plants in tissue culture which severely limits genetic engineering approaches. In maize, immature zygotic embryos provide the predominantly used material for establishing regeneration-competent cell or callus cultures for genetic transformation experiments. The procedures involved are demanding, laborious and time consuming and depend on greenhouse facilities. We have developed a novel tissue culture and plant regeneration system that uses maize leaf tissue and thus is independent of zygotic embryos and greenhouse facilities. We report here: (i) a protocol for the efficient induction of regeneration-competent callus from maize leaves in the dark, (ii) a protocol for inducing highly regenerable callus in the light, and (iii) the use of leaf-derived callus for the generation of stably transformed maize plants.

  16. Technical Note: The Normal Quantile Transformation and its application in a flood forecasting system

    NASA Astrophysics Data System (ADS)

    Bogner, K.; Pappenberger, F.; Cloke, H. L.

    2011-10-01

    The Normal Quantile Transform (NQT) has been used in many hydrological and meteorological applications in order to make the Cumulated Density Function (CDF) of the observed, simulated and forecast river discharge, water level or precipitation data Gaussian. It is also the heart of the meta-Gaussian model for assessing the total predictive uncertainty of the Hydrological Uncertainty Processor (HUP) developed by Krzysztofowicz. In the field of geo-statistics this transformation is better known as Normal-Score Transform. In this paper some possible problems caused by small sample sizes for the applicability in flood forecasting systems will be discussed and illustrated by examples. For the practical implementation commands and examples from the freely available and widely used statistical computing language R (R Development Core Team, 2011) will be given (represented in Courier font) and possible solutions are suggested by combining extreme value analysis and non-parametric regression methods.

  17. Performance analysis of a finite radon transform in OFDM system under different channel models

    SciTech Connect

    Dawood, Sameer A.; Anuar, M. S.; Fayadh, Rashid A.; Malek, F.; Abdullah, Farrah Salwani

    2015-05-15

    In this paper, a class of discrete Radon transforms namely Finite Radon Transform (FRAT) was proposed as a modulation technique in the realization of Orthogonal Frequency Division Multiplexing (OFDM). The proposed FRAT operates as a data mapper in the OFDM transceiver instead of the conventional phase shift mapping and quadrature amplitude mapping that are usually used with the standard OFDM based on Fast Fourier Transform (FFT), by the way that ensure increasing the orthogonality of the system. The Fourier domain approach was found here to be the more suitable way for obtaining the forward and inverse FRAT. This structure resulted in a more suitable realization of conventional FFT- OFDM. It was shown that this application increases the orthogonality significantly in this case due to the use of Inverse Fast Fourier Transform (IFFT) twice, namely, in the data mapping and in the sub-carrier modulation also due to the use of an efficient algorithm in determining the FRAT coefficients called the optimal ordering method. The proposed approach was tested and compared with conventional OFDM, for additive white Gaussian noise (AWGN) channel, flat fading channel, and multi-path frequency selective fading channel. The obtained results showed that the proposed system has improved the bit error rate (BER) performance by reducing inter-symbol interference (ISI) and inter-carrier interference (ICI), comparing with conventional OFDM system.

  18. Identification of material parameters for continuum modeling of phase transformations in multicomponent systems

    NASA Astrophysics Data System (ADS)

    Umantsev, Alex

    2007-01-01

    The continuum (field theoretic) method has become the method of choice for multiscale structure-formation modeling of very different phase transformations in the past decade. One of the challenges in application of the method to transformations in real materials is to obtain the mesoscopic parameters, which characterize the thermodynamic system of interest. Significant progress has been made in the case of pure systems; however, one would like to know what changes need to be made in the case of binary or multicomponent systems. We consider an exactly solvable case of the linear multicomponent system undergoing a phase transformation and derive equations that relate parameters of the continuum method, like barrier height, gradient energy, and relaxation coefficients, to the measurable quantities, like interface energy, interfacial thickness, and kinetic coefficient. We find that the contribution of chemical interactions in the system can be expressed as the renormalization of the barrier-height parameter of the continuum method and replacement of the latent heat with the chemical modulus. Atomic-scale simulations data for a solid/liquid transition in a binary Cu-Ni system were chosen for comparison with the theory and the fitting yields the estimates for the continuum-method parameters. Analysis of the temperature dependence of the interfacial energy allowed us to shed light on the magnitudes of the internal energy and entropy contributions into the solid/liquid interface.

  19. Classification of the quantum two-dimensional superintegrable systems with quadratic integrals and the Staeckel transforms

    SciTech Connect

    Daskaloyannis, C. Tanoudis, Y.

    2008-05-15

    The two-dimensional quantum superintegrable systems with quadratic integrals of motion on a manifold are classified by using the quadratic associative algebra of the integrals of motion. There are six general fundamental classes of quantum superintegrable systems corresponding to the classical ones. Analytic formulas for the involved integrals are calculated in all the cases. All the known quantum superintegrable systems with quadratic integrals are classified as special cases of these six general classes. The coefficients of the quadratic associative algebra of integrals are calculated and they are compared to the coefficients of the corresponding coefficients of the Poisson quadratic algebra of the classical systems. The quantum coefficients are similar to the classical ones multiplied by a quantum coefficient -{h_bar}{sup 2} plus a quantum deformation of order {h_bar}{sup 4} and {h_bar}{sup 6}. The systems inside the classes are transformed using Staeckel transforms in the quantum case as in the classical case. The general form of the Staeckel transform between superintegrable systems is discussed.

  20. Improved decryption quality and security of a joint transform correlator-based encryption system

    NASA Astrophysics Data System (ADS)

    Vilardy, Juan M.; Millán, María S.; Pérez-Cabré, Elisabet

    2013-02-01

    Some image encryption systems based on modified double random phase encoding and joint transform correlator architecture produce low quality decrypted images and are vulnerable to a variety of attacks. In this work, we analyse the algorithm of some reported methods that optically implement the double random phase encryption in a joint transform correlator. We show that it is possible to significantly improve the quality of the decrypted image by introducing a simple nonlinear operation in the encrypted function that contains the joint power spectrum. This nonlinearity also makes the system more resistant to chosen-plaintext attacks. We additionally explore the system resistance against this type of attack when a variety of probability density functions are used to generate the two random phase masks of the encryption-decryption process. Numerical results are presented and discussed.

  1. Quantum memory effects in noninteracting cold-atom systems: Hysteresis loop and lattice transformation

    NASA Astrophysics Data System (ADS)

    Chien, Chihchun; Metcalf, Mekena; Lai, Chenyen

    2016-05-01

    Memory effects are observable in magnetization, rechargeable batteries, and many systems exhibiting history-dependent states. Quantum memory effects are observable, for instance, in atomic superfluids. A counter-intuitive question is whether quantum memory effects can exist in noninteracting systems. Here we present two examples of cold-atom systems demonstrating memory effects in noninteracting systems. The first example is a ring-shaped potential loaded with noninteracting fermions. An artificial vector potential drives a current and with a tunable dissipative background, the current lags behind the driving and exhibits hysteresis loops. The dissipative energy can be controlled by the coupling between the fermions and the background. In the second example, cold atoms loaded in a tunable optical lattice transformed from the triangular to the kagome geometry. The kagome lattice supports a flat-band consisting of degenerate localized states. Quantum memory effects are observable after a lattice transformation as the steady-state density depends on the rate of the transformation. The versatility of memory effects in cold-atom systems promises novel applications in atomtronics.

  2. Transport and transformation of de-icing urea from airport runways in a constructed wetland system.

    PubMed

    Thorén, A K; Legrand, C; Herrmann, J

    2003-01-01

    Urea, NH2-CO-NH2, is used as a de-icing agent at Kalmar Airport, southeast Sweden. During 1998-2001, urea contributed on average 30% of the yearly nitrogen (N) transport of 41,000 kg via Törnebybäcken stream to the coastal zone of the Baltic Sea. In order to reduce stream transport of N from airport, agricultural and other diffuse sources, a wetland was constructed in 1996. Annual wetland retention of total-N varied in the range of 2,500-8,100 kg (6-36% of influent) during 1998-2001, according to mass balances calculated from monthly sampling. During airport de-icing, January-March 2001,660 kg urea-N out of 2,600 kg applied urea-N reached the wetland according to daily sampling. This indicated that 75% of the urea was transformed before entering the wetland. Urea was found to be only a minor part (8%) of total-N in the wetland influent. Calculations of cumulative urea-N loads at the wetland inlet and outlet respectively, showed a significant urea transformation during February 2001 with approximately 40% of the incoming urea-N being transformed in the wetland system. These results show that significant amounts of urea can be transformed in a wetland system at air temperatures around 0 degree C. PMID:14621175

  3. Agrobacterium tumefaciens-mediated transformation of taro (Colocasia esculenta (L.) Schott) with a rice chitinase gene for improved tolerance to a fungal pathogen Sclerotium rolfsii.

    PubMed

    He, Xiaoling; Miyasaka, Susan C; Fitch, Maureen M M; Moore, Paul H; Zhu, Yun J

    2008-05-01

    Taro (Colocasia esculenta) is one of the most important crops in the Pacific Islands, however, taro yields have been declining in Hawaii over the past 30 years partly due to diseases caused by oomycete and fungal pathogens. In this study, an efficient Agrobacterium tumefaciens-mediated transformation method for taro is first reported. In total, approximately 200 pieces (8 g) of embryogenic calluses were infected with the super-virulent A. tumefaciens strain EHA105 harboring the plant transformation plasmid pBI121/ricchi11 that contains the rice chitinase gene ricchi11. The presence and expression of the transgene ricchi11 in six independent transgenic lines was confirmed using polymerase chain reaction (PCR) and reverse transcription-PCR (RT-PCR). Southern blot analysis of the six independent lines indicated that three out of six (50%) had integrated a single copy of the transgene, and the other three lines had two or three copies of the transgene. Compared to the particle bombardment transformation of taro method, which was used in the previous studies, the Agrobacterium-mediated transformation method obtained 43-fold higher transformation efficiency. In addition, these six transgenic lines via Agrobacterium may be more effective for transgene expression as a result of single-copy or low-copy insertion of the transgene than the single line with multiple copies of the transgene via particle bombardment. In a laboratory bioassay, all six transgenic lines exhibited increased tolerance to the fungal pathogen Sclerotium rolfsii, ranging from 42 to 63% reduction in lesion expansion.

  4. Dynamic interferometer alignment and its utility in UV Fourier transform spectrometer systems

    NASA Technical Reports Server (NTRS)

    Dorval, Rick K.; Engel, James R.; Wyntjes, Geert J.

    1993-01-01

    Dynamic alignment has been demonstrated as a practical approach to alignment maintenance for systems in the infrared region of the spectrum. On the basis of work done by OPTRA, this technique was introduced in commercial Fourier transform spectrometer systems in 1982 and in various forms is now available from a number of manufacturers. This paper reports on work by OPTRA to extend the basic technique to systems operating in the ultraviolet. In addition, this paper reports the preliminary results of the development of an alignment system using a laser diode in place of a gas laser normally found in dynamic alignment systems. A unique optical system and spatial heterodyne technique allows for achievement of a metrology system with characteristics that fully satisfy the requirements of an ultraviolet spectrometer system.

  5. The effects of harmonics generated by an over-excited transformer on a multimachine system

    SciTech Connect

    Jalali, J.; Mokhtari, S.

    1995-09-01

    An electric power system consisting of three generators and nine buses is considered. The transformer connected to bus seven is considered as a source of harmonics, and it is modeled accordingly. The harmonic source generates harmonics with 6K {+-} 1 orders for K = 1, 2, 3 ..., n. The harmonics of order 6K {+-} 1 with significant effects on different buses of the power system network are evaluated during the steady state operation of the system by using SKM System Software for HIWAVE. Frequency scan and distortion calculations are studied to determine how the harmonics vary the operation of the power system network. The frequency spectrum of the harmonic source indicates that 5th order harmonic affects the operation of the power system. The effects of harmonics on the system are reduced by modeling a single-tuned shunt filter connected to bus seven parallel to the harmonic source to improve the system operation.

  6. Establishment of an overall transformation system for an oil-producing filamentous fungus, Mortierella alpina 1S-4.

    PubMed

    Takeno, Seiki; Sakuradani, Eiji; Murata, Shoichi; Inohara-Ochiai, Misa; Kawashima, Hiroshi; Ashikari, Toshihiko; Shimizu, Sakayu

    2004-09-01

    Oil-producing fungus Mortierella alpina 1S-4 is an industrial strain. To determine its physiological properties and to clarify the biosynthetic pathways for polyunsaturated fatty acids, a transformation system for this fungus was established using a derivative of it, i.e., a ura5- mutant lacking orotate phosphoribosyl transferase (OPRTase, EC.2.4.2.10) activity. Transformation with a vector containing the homologous ura5 gene as a marker was successfully performed using microprojectile bombardment, other methods frequently used for transformation, such as the protoplasting, lithium acetate, or electroporation methods, not giving satisfactory results. As a result, two types of transformants were obtained: a few stable transformants overexpressing the ura5 gene, and many unstable transformants showing OPRTase activity comparable to that of the wild-type strain. The results of quantitative PCR indicated that the stable transformants could retain the ura5 genes originating from the transformation vector regardless of the culture conditions. On the other hand, unstable transformants easily lost the marker gene under uracil-containing conditions, as expected. In this paper, we report that an overall transformation system for this fungus was successfully established, and propose how to select useful transformants as experimental and industrial strains. PMID:15138730

  7. Theory of the amplitude-phase retrieval in any linear-transform system and its applications

    NASA Astrophysics Data System (ADS)

    Yang, Guozhen; Gu, Ben-Yuan; Dong, Bi-Zhen

    1992-12-01

    This paper is a summary of the theory of the amplitude-phase retrieval problem in any linear transform system and its applications based on our previous works in the past decade. We describe the general statement on the amplitude-phase retrieval problem in an imaging system and derive a set of equations governing the amplitude-phase distribution in terms of the rigorous mathematical derivation. We then show that, by using these equations and an iterative algorithm, a variety of amplitude-phase problems can be successfully handled. We carry out the systematic investigations and comprehensive numerical calculations to demonstrate the utilization of this new algorithm in various transform systems. For instance, we have achieved the phase retrieval from two intensity measurements in an imaging system with diffraction loss (non-unitary transform), both theoretically and experimentally, and the recovery of model real image from its Hartley-transform modulus only in one and two dimensional cases. We discuss the achievement of the phase retrieval problem from a single intensity only based on the sampling theorem and our algorithm. We also apply this algorithm to provide an optimal design of the phase-adjusted plate for a phase-adjustment focusing laser accelerator and a design approach of single phase-only element for implementing optical interconnect. In order to closely simulate the really measured data, we examine the reconstruction of image from its spectral modulus corrupted by a random noise in detail. The results show that the convergent solution can always be obtained and the quality of the recovered image is satisfactory. We also indicated the relationship and distinction between our algorithm and the original Gerchberg- Saxton algorithm. From these studies, we conclude that our algorithm shows great capability to deal with the comprehensive phase-retrieval problems in the imaging system and the inverse problem in solid state physics. It may open a new way to

  8. Kernel functions and Baecklund transformations for relativistic Calogero-Moser and Toda systems

    SciTech Connect

    Hallnaes, Martin; Ruijsenaars, Simon

    2012-12-15

    We obtain kernel functions associated with the quantum relativistic Toda systems, both for the periodic version and for the nonperiodic version with its dual. This involves taking limits of previously known results concerning kernel functions for the elliptic and hyperbolic relativistic Calogero-Moser systems. We show that the special kernel functions at issue admit a limit that yields generating functions of Baecklund transformations for the classical relativistic Calogero-Moser and Toda systems. We also obtain the nonrelativistic counterparts of our results, which tie in with previous results in the literature.

  9. Phase equilibria and transformations in the Ti-Al-Nb system

    NASA Astrophysics Data System (ADS)

    Mishurda, Joseph Constantine

    The phase equilibria and transformations in the Nb-Ti-Al system in the vicinity of the Sigma phase field have been examined with respect to the Liquidus Surface (Phase 1), the Phase Equilibria and Equilibrium Transformations (Phase II), and the Phase Transformations, Mechanisms and Kinetics (Phase III). Eight alloy compositions were produced by arc melting. The alloys were characterized by differential thermal analysis, metallography, X-ray diffraction, scanning electron microscopy (BSEI), electron probe microanalysis and transmission electron microscopy. The liquidus examination shed new light on previous microstructural interpretations, opening up new possibilities for microstructural development and control of multiphase alloys. Differential thermal analysis has identified the existence of a beta to sigma + gamma transformation in an alloy where it was not previously thought to exist. The results differed from the calculated diagram by higher titanium solubility in the sigma and delta phases than predicted at lower temperatures and a lower solubility of alpha2 and gamma. The high temperature betao transforms to gamma + sigma in a eutectoid fashion resulting in a desirable lamellar structure of sigma and gamma. The existence of a new body centered tetragonal crystal structure ao = 5.11A and co 28.12A with a point group symmetry of P4/mmm, at 700°C was discovered. A plethel section was found for the sigma + beta two phase alloys. A betao + O + sigma three phase field passes through the alloys between 981 and 1000°C. The plethel section at the transformation has an eutectoid characteristic, however, the nature of the transformation changes to a peritectoid. At temperatures below 970°C, the first transformation to occur is the decomposition of the metastable betao phase to an intermediate metastable phase O'. Reasonable values for Q were obtained, applicable to the diffusion limited region of the TTT-curve. The microstructure evolution of the sigma + beta

  10. Streptomyces lipmanii expresses two restriction systems that inhibit plasmid transformation and bacteriophage plaque formation.

    PubMed

    Matsushima, P; Baltz, R H

    1989-06-01

    Bacteriophage host range studies suggested that several beta-lactam-producing streptomycetes express similar restriction-modification systems. Streptomyces lipmanii LE32 expressed two restriction-modification systems, designated SliI and SliII. A mutant strain, PM87, was defective only in SliI restriction but expressed both SliI and SliII modification. Streptomyces sp. strain A57986, a natural isolate partially deficient in the expression of SliI and SliII restriction, nevertheless modified bacteriophage DNA for both SliI and SliII specificities. Protoplasts of PM87 and A57986 were transformed by several plasmids, and the modified plasmids isolated from these strains transformed wild-type S. lipmanii efficiently.

  11. Active health system based on wavelet transform analysis of diffracted Lamb waves

    NASA Astrophysics Data System (ADS)

    Lemistre, Michel B.; Osmont, Daniel L.; Balageas, Daniel L.

    2000-08-01

    In composite materials, delaminations are discontinuities producing mode conversion processes generating various out-going modes. The Discrete Wavelet Transform allows isolating various propagation modes and extracting them in order to measure the time delay between the arrivals of the main burst and a specific out-going mode, for various propagation paths. This process permits, with a good accuracy, to localize a damage and to estimate its extension. An active health monitoring system composed of integrated disc-shaped, 100 (mu) m-thick and 5 mm-dia PZT transducers working sequentially as actuators and receives is presented. The diagnostic is based on multiresolution process by wavelet transform applied on recorded Lamb wave signals obtained before and after damage. The robustness and portability of this technique is demonstrated by the fact that, after validation in our laboratory it was successfully applied to data coming from an experiment conducted in an other Laboratory using its own Health Monitoring system.

  12. Transform and relax sampling for highly anisotropic systems: Application to protein domain motion and folding

    NASA Astrophysics Data System (ADS)

    Kitao, Akio

    2011-07-01

    Transform and relax sampling (TRS) is proposed as a conformational sampling method to enhance "soft" fluctuation in highly anisotropic systems using molecular dynamics simulation. This method consists of three stages; transform, relax, and sampling. In the transform stage, molecular dynamics simulation is performed with randomly assigned force bias to enhance the fluctuations along relatively soft collective movements, as expected from the linear response theory. After relaxing the heated system to equilibrium without force bias in the relax stage, Monte Carlo-type determination is made as to whether the generated state is accepted or not. The sampling stage is then conducted for conformational sampling by conventional molecular dynamics simulation. TRS is first applied for the idealized multidimensional double-well Cα model to mimic protein open-close transition. Subsequently, it is applied to three different all-atom protein systems in an explicit solvent model; T4 lysozyme, glutamine binding protein, and a mini-protein chignolin. Investigation of structural variations in the hinge angle of T4 lysozyme in crystals is demonstrated by TRS. The liganded close structure of the glutamine binding protein is sampled starting from the unliganded open form. Chignolin is shown to fold into a native structure multiple times starting from highly extended structures within 100 ns. It is concluded that TRS sampled a reasonable conformational space within a relatively short simulation time in these cases. Possible future extensions of TRS are also discussed.

  13. Calibration of 3D laser measurement system based on projective transformation

    NASA Astrophysics Data System (ADS)

    Guo, Yang; Du, Yue-yang; Du, Zheng-chun; Yao, Zhen-qiang

    2010-08-01

    This paper presents a planar projective transformation based method for fully automated exterior and interior calibration of a three-dimensional laser scanning system. The calibration is crucial for applications that attempt to produce accurately registered or fused three-dimensional sensor data. A key contribution of the method lies in the derivation of transformation relations that describe the same point in three defined coordinate systems with respect to the rotating characteristic of two scanning planes and its calibration target object whose geometric feature can be reliably recognized from a single observation. The transformation relationship can be converted to the closed-form solution to the constraint equations of the system parameters in the form of intrinsic and extrinsic matrices. By deriving the relationship between a single two-dimensional range scan and the point location presentation in the absolute frame, the interior and exterior calibration can be accomplished simultaneously and the algorithm of the 6 DOF pose improves the identification precision. Finally, this paper reports the performance and stability of this method on real data sets, and demonstrates the accuracy within +/-0.1 degree of the orientation precision and 8mm of position precision in a realistic configuration.

  14. Tele-Operation of a Mobile Haptic System Using Dynamical Modal Transformation

    NASA Astrophysics Data System (ADS)

    Yamanouchi, Wataru; Katsura, Seiichiro

    Recent advances in control technology have contributed to the development of robot systems for communication. Robot systems recognize their environment on the basis of audio-visual information. Recognition methods based on audio-visual feedback have been developed by many researchers. Apart from auditory and visual information, haptic information has recently attracted attention as the third type of multimedia information. The sense of touch is useful for remote manipulation. Feedback of haptic information is realized by bilateral control. In conventional research, most systems are constructed using a master-slave system in which the master-slave systems have the same mechanical structure. However, very few studies have been carried out on force feedback systems with different mechanical structures. This paper proposes a novel control method for mobile-hapto, which involves force feedback using mobility systems. In this study, the mobile-hapto consists of a mobile robot that can move in an infinite area for motion and a joystick that is fixed at a given position and can be operated manually. To realize of force feedback in the mobile-hapto, a modal transformation matrix for bilateral control is proposed. The proposed modal transformation matrix is able to change the dimension of the controlled value. The joystick is treated as a pedal by changing two dimensions of the mobile robot position. The mobile-hapto is possible to be subjected to intuitive manipulation. The validity of the proposed method is experimentally verified.

  15. Assessment of simple marker-free genetic transformation techniques in alfalfa.

    PubMed

    Ferradini, Nicoletta; Nicolia, Alessandro; Capomaccio, Stefano; Veronesi, Fabio; Rosellini, Daniele

    2011-11-01

    Methods to avoid the presence of selectable marker genes (SMG) in transgenic plants are available but not implemented in many crop species. We assessed the efficiency of simple marker-free Agrobacterium-mediated transformation techniques in alfalfa: regeneration without selection, or marker-less, and co-transformation with two vectors, one containing the SMG and one containing a non-selected gene. To easily estimate the efficiency of marker-less transformation, the nptII and the GUS markers were used as non-selected genes. After Agrobacterium treatment, somatic embryos were regenerated without selection. The percentage of transgenic embryos was determined by a second cycle of regeneration using the embryos as starting material, in the presence of kanamycin, by PCR screening of T1 progenies, and by the GUS test. In two experiments, from 0 to 1.7% of the somatic embryos were transgenic. Co-transformation was performed with two vectors, one with the hemL SMG and one with the unselected nptII gene, each carried by a different culture of Agrobacterium. Only 15 putative co-transformed plants were regenerated from two experiments, with an average co-transformation percentage of 3.7. Southern blot hybridizations and/or T(1) progeny segregation were used to confirm transgene integration, and qPCR was also used to estimate the T-DNA copy number. In the T(1) progenies obtained by crossing with a non-transgenic pollinator, marker-free segregants were obtained. Both marker-free approaches showed very low efficiency. PMID:21691741

  16. Bivariate distribution, correlation, and transformation properties of two-color infrared systems.

    PubMed

    Clow, R; McNolty, F

    1974-05-01

    A two-dimensional (two-color) statistical structure is formulated that is applicable to pattern recognition, discrimination, and detection problems occurring in infrared signal-processing systems. The methodology relates physical quantities such as the temperature T of an object, its projected area A, emissivity , range R from the sensor, and noise equivalent flux density (NEFD) to the geometry of a local orthogonal coordinate system where the coordinate axes correspond to the apparent radiant intensity J in each micron bandwidth. The bivariate distribution, correlation, and transformation properties attendant to this framework are discussed in detail. Additional insight into the structure of the problem is achieved by investigating the two-color system in terms of a nonorthogonal local coordinate system. The various results presented in the paper may be extended to three-, four-, or five-color systems by direct analogies.

  17. Micro-Tom Tomato as an Alternative Plant Model System: Mutant Collection and Efficient Transformation.

    PubMed

    Shikata, Masahito; Ezura, Hiroshi

    2016-01-01

    Tomato is a model plant for fruit development, a unique feature that classical model plants such as Arabidopsis and rice do not have. The tomato genome was sequenced in 2012 and tomato is becoming very popular as an alternative system for plant research. Among many varieties of tomato, Micro-Tom has been recognized as a model cultivar for tomato research because it shares some key advantages with Arabidopsis including its small size, short life cycle, and capacity to grow under fluorescent lights at a high density. Mutants and transgenic plants are essential materials for functional genomics research, and therefore, the availability of mutant resources and methods for genetic transformation are key tools to facilitate tomato research. Here, we introduce the Micro-Tom mutant database "TOMATOMA" and an efficient transformation protocol for Micro-Tom.

  18. Output feedback control of linear fractional transformation systems subject to actuator saturation

    NASA Astrophysics Data System (ADS)

    Ban, Xiaojun; Wu, Fen

    2016-11-01

    In this paper, the control problem for a class of linear parameter varying (LPV) plant subject to actuator saturation is investigated. For the saturated LPV plant depending on the scheduling parameters in linear fractional transformation (LFT) fashion, a gain-scheduled output feedback controller in the LFT form is designed to guarantee the stability of the closed-loop LPV system and provide optimised disturbance/error attenuation performance. By using the congruent transformation, the synthesis condition is formulated as a convex optimisation problem in terms of a finite number of LMIs for which efficient optimisation techniques are available. The nonlinear inverted pendulum problem is employed to demonstrate the effectiveness of the proposed approach. Moreover, the comparison between our LPV saturated approach with an existing linear saturated method reveals the advantage of the LPV controller when handling nonlinear plants.

  19. Enhanced iterative learning control for a piezoelectric actuator system using wavelet transform filtering

    NASA Astrophysics Data System (ADS)

    Chien, Chiang-Ju; Lee, Fu-Shin; Wang, Jhen-Cheng

    2007-01-01

    For trajectory tracking of a piezoelectric actuator system, an enhanced iterative learning control (ILC) scheme based on wavelet transform filtering (WTF) is proposed in this research. The enhanced ILC scheme incorporates a state compensation in the ILC formula. Combining state compensation with iterative learning, the scheme enhances tracking accuracies substantially, in comparison to the conventional D-type ILC and a proportional control-aided D-type ILC. The wavelet transform is adopted to filter learnable tracking errors without phase shift. Based on both a time-frequency analysis of tracking errors and a convergence bandwidth analysis of ILC, a two-level WTF is chosen for ILC in this study. The enhanced ILC scheme using WTF was applied to track two desired trajectories, one with a single frequency and the other with multiple frequencies, respectively. Experimental results validate the efficacy of the enhanced ILC in terms of the speed of convergence and the level of long-term tracking errors.

  20. Stable transformation of Theobroma cacao L. and influence of matrix attachment regions on GFP expression.

    PubMed

    Maximova, S; Miller, C; Antúnez de Mayolo, G; Pishak, S; Young, A; Guiltinan, M J

    2003-06-01

    We describe a protocol for Agrobacterium-mediated genetic transformation of Theobroma cacao L. using cotyledonary explants from primary somatic embryos (SEs) and A. tumefaciens strain AGL1. Transgenic plants carrying the visible marker, gene green fluorescent protein ( EGFP), the selectable marker gene neomycin phosphotransferase II ( NPTII), the class I chitinase gene from cacao ( Chi), and tobacco nuclear matrix attachment regions (MARs) in different combinations were successfully produced via regeneration of secondary SEs. The presence of the Chi gene or MARs did not influence the number of transgenic plants produced compared to the marker genes alone. However, the inclusion of MARs contributed to increased mean GFP expression in the population of transgenics. Additionally, the presence of MARs reduced the occurrence of gene silencing and stabilized high levels of GFP expression in lines of transgenic plants multiplied via reiterative somatic embryogenesis. Ninety-four transgenic plants were acclimated in a greenhouse and grown to maturity. Detailed growth analysis indicated that there were no differences in various growth parameters between transgenic and non-transgenic SE-derived plants. Seeds produced from two genetic crosses with one of the transgenic lines were analyzed for EGFP expression-a near-perfect 1:1 segregation was observed, indicating that this line resulted from the insertion of a single locus of T-DNA. PMID:12789505

  1. Stock, turnover and functions of carbon in heavily weathered soils under lowland rainforest transformation systems

    NASA Astrophysics Data System (ADS)

    Guillaume, Thomas; Kuzyakov, Yakov

    2013-04-01

    Tropical rainforest are experiencing worldwide a strong lost through deforestation and transformation into agricultural systems. Land use changes in such ecosystems leads to major modifications of soils properties and processes. One indication of it are the losses of organic carbon content (Corg); an important soil fertility parameter in heavily weathered soil. Between 1985 and 2007, Sumatra (Indonesia) has lost half of his remaining natural rainforest, which currently covers only 30% the island. The deforestation is still ongoing and the main drivers of deforestation are oil palm, rubber and timber industries. Our study aims to identify and quantify the impacts of lowland rainforest transformation systems (TS): oil palm, rubber and jungle rubber plantations on soil organic carbon (SOC) and nitrogen (SON) quality, turnover and stocks and so, on soil fertility and functions. We hypothesize that transformation of natural lowland rainforest changes not only C stock and budget throughout quantity and quality of C input, but also DOM production and water consumption by vegetation, leading to a relocation of C in the subsoil. This should be reflected in C and N content in soil profile horizons as well as their δ13C and δ15N isotopic signatures. We will evaluate also C stability through biological, thermal and chemical stability in bulk soil and aggregate fractions. The TS investigated, including lowland rainforest as reference sites, are located in Jambi Province (Sumatra). Soil has been described and sampled per horizon on 4 replicates of each TS in 2 different regions (32 sites) in Autumn 2012. As hypothesized, first results show strong effects of forest transformation on C and N content, as well as on isotopic signatures of soil. Those results will also be used further to select DOM sampling depths and adequate horizons to perform sorption and incubation experiments.

  2. Propagation of a general-type beam through a truncated fractional Fourier transform optical system.

    PubMed

    Zhao, Chengliang; Cai, Yangjian

    2010-03-01

    Paraxial propagation of a general-type beam through a truncated fractional Fourier transform (FRT) optical system is investigated. Analytical formulas for the electric field and effective beam width of a general-type beam in the FRT plane are derived based on the Collins formula. Our formulas can be used to study the propagation of a variety of laser beams--such as Gaussian, cos-Gaussian, cosh-Gaussian, sine-Gaussian, sinh-Gaussian, flat-topped, Hermite-cosh-Gaussian, Hermite-sine-Gaussian, higher-order annular Gaussian, Hermite-sinh-Gaussian and Hermite-cos-Gaussian beams--through a FRT optical system with or without truncation. The propagation properties of a Hermite-cos-Gaussian beam passing through a rectangularly truncated FRT optical system are studied as a numerical example. Our results clearly show that the truncated FRT optical system provides a convenient way for laser beam shaping.

  3. Thermodynamic Modelling of Phase Transformation in a Multi-Component System

    SciTech Connect

    Vala, J.

    2007-09-06

    Diffusion in multi-component alloys can be characterized by the vacancy mechanism for substitutional components, by the existence of sources and sinks for vacancies and by the motion of atoms of interstitial components. The description of diffusive and massive phase transformation of a multi-component system is based on the thermodynamic extremal principle by Onsager; the finite thickness of the interface between both phases is respected. The resulting system of partial differential equations of evolution with integral terms for unknown mole fractions (and additional variables in case of non-ideal sources and sinks for vacancies), can be analyzed using the method of lines and the finite difference technique (or, alternatively, the finite element one) together with the semi-analytic and numerical integration formulae and with certain iteration procedure, making use of the spectral properties of linear operators. The original software code for the numerical evaluation of solutions of such systems, written in MATLAB, offers a chance to simulate various real processes of diffusional phase transformation. Some results for the (nearly) steady-state real processes in substitutional alloys have been published yet. The aim of this paper is to demonstrate that the same approach can handle both substitutional and interstitial components even in case of a general system of evolution.

  4. A Preliminary Application of Ensemble Transform Kalman Filter System to Rainfall Forecast

    NASA Astrophysics Data System (ADS)

    Tie, Wang

    2014-05-01

    WangTie(1,2),YangXiaofeng(1),ZangXin(1),GengLining(1),ZhouYan(1) (1)NanJing China-Spacenet Satellite Telecom CO., Ltd, Nanjing, 210061, China,(2)Institute of Atmospheric Physics, Chinese Academy of Sciences, LASG, Beijing, 100029, China The Ensemble Transform Kalman Filter(ETKF) method is applied to WRF model(Weather Research & Forecast Model system) Version 3.5.1, and the ETKF based data assimilation system is constructed. The accurate of forecast result and the sensitivity of initial data error on a heavy rainfall from 5th to 7th July 2003 in Jiangsu province China are investigated. The numerical result indicated that the ETKF method could decrease the forecast error and improves the forecast result either the synoptic scale system and the meso-scale system, the RMSEs of wind decrease less than 18 percent, while the temperature and humidity decrease about 6 precent. The forecast of precipitation, especially the 12h accumulated precipitation is improved significant after ETKF assimilation. Keywords: intense rainfall, prediction error, Ensemble Transform Kalman Filter(ETKF), WRF model

  5. Nitrogen transformations in intensive aquaculture system and its implication to climate change through nitrous oxide emission.

    PubMed

    Hu, Zhen; Lee, Jae Woo; Chandran, Kartik; Kim, Sungpyo; Sharma, Keshab; Brotto, Ariane Coelho; Khanal, Samir Kumar

    2013-02-01

    The rapid development of aquaculture could result in significant environmental concerns such as eutrophication and climate change. However, to date, very few studies have been conducted to investigate nitrogen transformations in aquaculture systems; and specifically the emission of nitrous oxide (N(2)O), which is an important greenhouse gas and ozone-depleting substance. In this study, nitrogen transformations in intensive laboratory-scale Chinese catfish (Clarias fuscus) aquaculture systems were investigated by identifying and quantifying N(2)O emissions. Results indicated that about 1.3% of the nitrogen input was emitted as N(2)O gas. Dissolved oxygen (DO) concentrations and feeding rates had significant effects on N(2)O emissions. Higher N(2)O emissions were obtained in aquaculture systems with lower DO concentrations and higher feeding rates. Both nitrification and denitrification appeared to be responsible for the emissions of N(2)O. Key factors which correlated with the N(2)O emission rate in aquaculture systems were NO(2)(-), DO and total ammonia nitrogen concentrations.

  6. The explanation of the twin paradox using Poincare transformation and computer algebra system REDUCE

    NASA Astrophysics Data System (ADS)

    Hermanto, Arief

    2012-06-01

    We explain the twin (A and B) paradox using Poincare transformation (as the generalization of Lorentz transformation) in Special Relativity. We want to emphasize the fact that the paradox can really be explained in the context of Special Relativity. The twin A stays at home whereas B makes a round trip. We can still stay in Special Relativity if the non-inertial reference frame of B is in the form of a set of two inertial frames (K1 and K2) moving with different velocities with respect to a fixed inertial reference frame (K0) of A. K1 and K2 are each connected to K0 with Poincare Transformation. We use the CAS (computer algebra system) REDUCE to assist the computation. To make the discussion realistic and simpler we use rational numbers (so that we will get exact computational results) instead of symbols. The important point is that we will show how the fact can be understood by both parties (A and B) by simulating numerically the trip from the points of view of each A and B. A will accept the fact that B is younger and B will also accept the fact that A is older at the reunion. We hope the paradox will thus be explained away satisfactorily.

  7. Transformation of ground vibration signal for debris-flow monitoring and detection in alarm systems.

    PubMed

    Abancó, Clàudia; Hürlimann, Marcel; Fritschi, Bruno; Graf, Christoph; Moya, José

    2012-01-01

    Debris flows are fast mass movements formed by a mix of water and solid materials, which occur in steep torrents, and are a source of high risks for human settlements. Geophones are widely used to detect the ground vibration induced by passing debris flows. However, the recording of geophone signals usually requires storing a huge amount of data, which leads to problems in storage capacity and power consumption. This paper presents a method to transform and simplify the signals measured by geophones. The key input parameter is the ground velocity threshold, which removes the seismic noise that is not related to debris flows. A signal conditioner was developed to implement the transformation and the ground velocity threshold was set by electrical resistors. The signal conditioner was installed at various European monitoring sites to test the method. Results show that data amount and power consumption can be greatly reduced without losing much information on the main features of the debris flows. However, the outcome stresses the importance of choosing a ground vibration threshold, which must be accurately calibrated. The transformation is also suitable to detect other rapid mass movements and to distinguish among different processes, which points to a possible implementation in alarm systems.

  8. The parallel implementation of the one-dimensional Fourier transformed Vlasov Poisson system

    NASA Astrophysics Data System (ADS)

    Eliasson, Bengt

    2005-08-01

    A parallel implementation of an algorithm for solving the one-dimensional, Fourier transformed Vlasov-Poisson system of equations is documented, together with the code structure, file formats and settings to run the code. The properties of the Fourier transformed Vlasov-Poisson system is discussed in connection with the numerical solution of the system. The Fourier method in velocity space is used to treat numerical problems arising due the filamentation of the solution in velocity space. Outflow boundary conditions in the Fourier transformed velocity space removes the highest oscillations in velocity space. A fourth-order compact Padé scheme is used to calculate derivatives in the Fourier transformed velocity space, and spatial derivatives are calculated with a pseudo-spectral method. The parallel algorithms used are described in more detail, in particular the parallel solver of the tri-diagonal systems occurring in the Padé scheme. Program summaryTitle of program:vlasov Catalogue identifier:ADVQ Program summary URL:http://cpc.cs.qub.ac.uk/summaries/ADVQ Program obtainable from: CPC Program Library, Queen's University of Belfast, N. Ireland Operating system under which the program has been tested: Sun Solaris; HP-UX; Read Hat Linux Programming language used: FORTRAN 90 with Message Passing Interface (MPI) Computers: Sun Ultra Sparc; HP 9000/785; HP IPF (Itanium Processor Family) ia64 Cluster; PCs cluster Number of lines in distributed program, including test data, etc.:3737 Number of bytes in distributed program, including test data, etc.:18 772 Distribution format: tar.gz Nature of physical problem: Kinetic simulations of collisionless electron-ion plasmas. Method of solution: A Fourier method in velocity space, a pseudo-spectral method in space and a fourth-order Runge-Kutta scheme in time. Memory required to execute with typical data: Uses typically of the order 10 5-10 6 double precision numbers. Restriction on the complexity of the problem: The program uses

  9. Genetic transformation of Fusarium avenaceum by Agrobacterium tumefaciens mediated transformation and the development of a USER-Brick vector construction system

    PubMed Central

    2014-01-01

    Background The plant pathogenic and saprophytic fungus Fusarium avenaceum causes considerable in-field and post-field losses worldwide due to its infections of a wide range of different crops. Despite its significant impact on the profitability of agriculture production and a desire to characterize the infection process at the molecular biological level, no genetic transformation protocol has yet been established for F. avenaceum. In the current study, it is shown that F. avenaceum can be efficiently transformed by Agrobacterium tumefaciens mediated transformation. In addition, an efficient and versatile single step vector construction strategy relying on Uracil Specific Excision Reagent (USER) Fusion cloning, is developed. Results The new vector construction system, termed USER-Brick, is based on a limited number of PCR amplified vector fragments (core USER-Bricks) which are combined with PCR generated fragments from the gene of interest. The system was found to have an assembly efficiency of 97% with up to six DNA fragments, based on the construction of 55 vectors targeting different polyketide synthase (PKS) and PKS associated transcription factor encoding genes in F. avenaceum. Subsequently, the ΔFaPKS3 vector was used for optimizing A. tumefaciens mediated transformation (ATMT) of F. avenaceum with respect to six variables. Acetosyringone concentration, co-culturing time, co-culturing temperature and fungal inoculum were found to significantly impact the transformation frequency. Following optimization, an average of 140 transformants per 106 macroconidia was obtained in experiments aimed at introducing targeted genome modifications. Targeted deletion of FaPKS6 (FA08709.2) in F. avenaceum showed that this gene is essential for biosynthesis of the polyketide/nonribosomal compound fusaristatin A. Conclusion The new USER-Brick system is highly versatile by allowing for the reuse of a common set of building blocks to accommodate seven different types of genome

  10. A new Watermarking System based on Discrete Cosine Transform (DCT) in color biometric images.

    PubMed

    Dogan, Sengul; Tuncer, Turker; Avci, Engin; Gulten, Arif

    2012-08-01

    This paper recommend a biometric color images hiding approach An Watermarking System based on Discrete Cosine Transform (DCT), which is used to protect the security and integrity of transmitted biometric color images. Watermarking is a very important hiding information (audio, video, color image, gray image) technique. It is commonly used on digital objects together with the developing technology in the last few years. One of the common methods used for hiding information on image files is DCT method which used in the frequency domain. In this study, DCT methods in order to embed watermark data into face images, without corrupting their features.

  11. A new Watermarking System based on Discrete Cosine Transform (DCT) in color biometric images.

    PubMed

    Dogan, Sengul; Tuncer, Turker; Avci, Engin; Gulten, Arif

    2012-08-01

    This paper recommend a biometric color images hiding approach An Watermarking System based on Discrete Cosine Transform (DCT), which is used to protect the security and integrity of transmitted biometric color images. Watermarking is a very important hiding information (audio, video, color image, gray image) technique. It is commonly used on digital objects together with the developing technology in the last few years. One of the common methods used for hiding information on image files is DCT method which used in the frequency domain. In this study, DCT methods in order to embed watermark data into face images, without corrupting their features. PMID:21537852

  12. [The role of educational institutions in the transformation of the Mexican health system].

    PubMed

    Graue-Wiechers, Enrique

    2012-01-01

    The vocation of any medical school is educate the physicians that the country needs to address current and future health challenges, and able to work effectively in the gradual transformation of the health system of our country. The reason for the inclusion of this topic in this symposium is to ask whether this has been happening and if has been appropriate, and if so, to what extent medical education institutions establish actions and directions for the better and more quickly achieve the desired goal.

  13. Iterative clipping and filtering based on discrete cosine transform/inverse discrete cosine transform for intensity modulator direct detection optical orthogonal frequency division multiplexing system

    NASA Astrophysics Data System (ADS)

    Mangone, Fall; Tang, Jin; Chen, Ming; Xiao, Jiangnan; Fan, Li; Chen, Lin

    2013-06-01

    Iterative clipping and filtering (ICF) is a useful technique to reduce the peak-to-average power ratio (PAPR) of orthogonal frequency division multiplexing (OFDM) signals. However, the classical ICF with Fast Fourier Transform/Inverse Fast Fourier Transform requires much iteration to approach a specified PAPR threshold in the complementary cumulative distribution function. To reduce the nonlinear distortion in both electrical and optical devices and in the optical fiber, we propose a novel ICF based on discrete cosine transform/inverse discrete cosine transform to reduce the PAPR in an intensity modulator and direct detection (IM/DD) optical OFDM system. Furthermore, the new technique considerably improves bit error rate (BER) and reduces the PAPR with just few iterations. The experimental results show that the receiver sensitivity at a BER of 1 for a 2.5-Gbytes/s OFDM signal and after 200-km standard single-mode fiber transmission has been improved by 1.1, 2.3, and 3.6 dBm with launch powers of 6, 8, and 12 dBm respectively.

  14. A FPGA system for QRS complex detection based on Integer Wavelet Transform

    NASA Astrophysics Data System (ADS)

    Stojanović, R.; Karadaglić, D.; Mirković, M.; Milošević, D.

    2011-01-01

    Due to complexity of their mathematical computation, many QRS detectors are implemented in software and cannot operate in real time. The paper presents a real-time hardware based solution for this task. To filter ECG signal and to extract QRS complex it employs the Integer Wavelet Transform. The system includes several components and is incorporated in a single FPGA chip what makes it suitable for direct embedding in medical instruments or wearable health care devices. It has sufficient accuracy (about 95%), showing remarkable noise immunity and low cost. Additionally, each system component is composed of several identical blocks/cells what makes the design highly generic. The capacity of today existing FPGAs allows even dozens of detectors to be placed in a single chip. After the theoretical introduction of wavelets and the review of their application in QRS detection, it will be shown how some basic wavelets can be optimized for easy hardware implementation. For this purpose the migration to the integer arithmetic and additional simplifications in calculations has to be done. Further, the system architecture will be presented with the demonstrations in both, software simulation and real testing. At the end, the working performances and preliminary results will be outlined and discussed. The same principle can be applied with other signals where the hardware implementation of wavelet transform can be of benefit.

  15. Efficient Temperature-Dependent Green's Functions Methods for Realistic Systems: Compact Grids for Orthogonal Polynomial Transforms.

    PubMed

    Kananenka, Alexei A; Phillips, Jordan J; Zgid, Dominika

    2016-02-01

    The Matsubara Green's function that is used to describe temperature-dependent behavior is expressed on a numerical grid. While such a grid usually has a couple of hundred points for low-energy model systems, for realistic systems with large basis sets the size of an accurate grid can be tens of thousands of points, constituting a severe computational and memory bottleneck. In this paper, we determine efficient imaginary time grids for the temperature-dependent Matsubara Green's function formalism that can be used for calculations on realistic systems. We show that, because of the use of an orthogonal polynomial transform, we can restrict the imaginary time grid to a few hundred points and reach micro-Hartree accuracy in the electronic energy evaluation. Moreover, we show that only a limited number of orthogonal polynomial expansion coefficients are necessary to preserve accuracy when working with a dual representation of the Green's function or self-energy and transforming between the imaginary time and frequency domain.

  16. Robust colour calibration of an imaging system using a colour space transform and advanced regression modelling.

    PubMed

    Jackman, Patrick; Sun, Da-Wen; Elmasry, Gamal

    2012-08-01

    A new algorithm for the conversion of device dependent RGB colour data into device independent L*a*b* colour data without introducing noticeable error has been developed. By combining a linear colour space transform and advanced multiple regression methodologies it was possible to predict L*a*b* colour data with less than 2.2 colour units of error (CIE 1976). By transforming the red, green and blue colour components into new variables that better reflect the structure of the L*a*b* colour space, a low colour calibration error was immediately achieved (ΔE(CAL) = 14.1). Application of a range of regression models on the data further reduced the colour calibration error substantially (multilinear regression ΔE(CAL) = 5.4; response surface ΔE(CAL) = 2.9; PLSR ΔE(CAL) = 2.6; LASSO regression ΔE(CAL) = 2.1). Only the PLSR models deteriorated substantially under cross validation. The algorithm is adaptable and can be easily recalibrated to any working computer vision system. The algorithm was tested on a typical working laboratory computer vision system and delivered only a very marginal loss of colour information ΔE(CAL) = 2.35. Colour features derived on this system were able to safely discriminate between three classes of ham with 100% correct classification whereas colour features measured on a conventional colourimeter were not.

  17. Like daughter, like son? Fertility decline and the transformation of gender systems in the family

    PubMed Central

    Allendorf, Keera

    2016-01-01

    BACKGROUND An important question for population research is whether fertility decline transforms gender systems. OBJECTIVE This paper contributes to answering this broad question by examining how fertility decline may change the relative value and roles of daughters and sons in families. First, I outline theoretical pathways, suggesting that a key factor is the gender composition of families. As fertility declines, the proportion of families with children of only one gender increases, which may facilitate greater gender symmetry between daughters and sons. Second, I explore how fertility decline may have contributed to the transformation of the relative value and roles of sons and daughters in practice in one place. METHODS The analysis draws primarily on semi-structured interviews with 30 respondents living in one Indian village. This village is located in a district where fertility has declined to at least the replacement level. RESULTS Respondents perceive changes in the gender system, including less son preference, more equal schooling for sons and daughters, more freedom in marriage and premarital relationships, and perhaps greater daughter support of parents in old age. CONCLUSIONS The results describe changes in the relative value, treatment, and behavior of sons and daughters that are consistent with the theorized effects of fertility decline. Future research is needed, however, to determine whether fertility decline makes a causal contribution to changes in the gender system. PMID:27147902

  18. Evaluation of a FTIR (Fourier Transform Infrared Spectrometer) mobile source measurement system

    NASA Astrophysics Data System (ADS)

    Sigsby, John E., Jr.; McArver, Alex; Snow, Richard

    1989-04-01

    An initial evaluation was made of a prototype Fourier transform infrared spectrometer on its ability to measure mobile source emissions. This prototype represents the commercialization of research technology developed by the Ford Motor Co. The system utilizes a Mattson Instruments Co. interferometer coupled to a Masscomp computer. The required software is still incomplete. This has resulted in the inability to correct errors or optimize results. Unreliability of both hardware and software is a major shortcoming of the system. When operating properly, the system measures carbon dioxide, nitric oxide, and formaldehyde accurately, both when compared to bag analysis and on direct point-by-point comparisons with real-time data. Completion of the evaluation awaits the final software translations and improvements in reliability.

  19. Engagement in system redesign: A wellness example to enable a cultural transformation.

    PubMed

    Anderson, Bradley; Hansson, Wendy Kai

    2016-09-01

    For the past several years, Interior Health (IH) has worked collaboratively with Aboriginal leaders to build strong relationships and develop an environment of reciprocal accountability and knowledge exchange. All partners are committed to working together to change the healthcare system so that it responds appropriately and effectively to the needs of a population of people with the poorest health outcomes. The development of the IH Aboriginal Health and Wellness Strategy is an example of meaningful engagement with First Nations communities in the IH region at the system level. The strategy was built by IH, led by First Nations, and developed in collaboration with First Nations, Urban, and Métis partners. It aims to address the shared long-term goal of improving the overall health and wellness of Aboriginal people and enable a cultural transformation to overcome the barriers that are stopping Aboriginal people from seeking care from a shared healthcare system. PMID:27576857

  20. Collusion-resistant audio fingerprinting system in the modulated complex lapped transform domain.

    PubMed

    Garcia-Hernandez, Jose Juan; Feregrino-Uribe, Claudia; Cumplido, Rene

    2013-01-01

    Collusion-resistant fingerprinting paradigm seems to be a practical solution to the piracy problem as it allows media owners to detect any unauthorized copy and trace it back to the dishonest users. Despite the billionaire losses in the music industry, most of the collusion-resistant fingerprinting systems are devoted to digital images and very few to audio signals. In this paper, state-of-the-art collusion-resistant fingerprinting ideas are extended to audio signals and the corresponding parameters and operation conditions are proposed. Moreover, in order to carry out fingerprint detection using just a fraction of the pirate audio clip, block-based embedding and its corresponding detector is proposed. Extensive simulations show the robustness of the proposed system against average collusion attack. Moreover, by using an efficient Fast Fourier Transform core and standard computer machines it is shown that the proposed system is suitable for real-world scenarios.

  1. Collusion-Resistant Audio Fingerprinting System in the Modulated Complex Lapped Transform Domain

    PubMed Central

    Garcia-Hernandez, Jose Juan; Feregrino-Uribe, Claudia; Cumplido, Rene

    2013-01-01

    Collusion-resistant fingerprinting paradigm seems to be a practical solution to the piracy problem as it allows media owners to detect any unauthorized copy and trace it back to the dishonest users. Despite the billionaire losses in the music industry, most of the collusion-resistant fingerprinting systems are devoted to digital images and very few to audio signals. In this paper, state-of-the-art collusion-resistant fingerprinting ideas are extended to audio signals and the corresponding parameters and operation conditions are proposed. Moreover, in order to carry out fingerprint detection using just a fraction of the pirate audio clip, block-based embedding and its corresponding detector is proposed. Extensive simulations show the robustness of the proposed system against average collusion attack. Moreover, by using an efficient Fast Fourier Transform core and standard computer machines it is shown that the proposed system is suitable for real-world scenarios. PMID:23762455

  2. Automated cervical precancerous cells screening system based on Fourier transform infrared spectroscopy features

    NASA Astrophysics Data System (ADS)

    Jusman, Yessi; Mat Isa, Nor Ashidi; Ng, Siew-Cheok; Hasikin, Khairunnisa; Abu Osman, Noor Azuan

    2016-07-01

    Fourier transform infrared (FTIR) spectroscopy technique can detect the abnormality of a cervical cell that occurs before the morphological change could be observed under the light microscope as employed in conventional techniques. This paper presents developed features extraction for an automated screening system for cervical precancerous cell based on the FTIR spectroscopy as a second opinion to pathologists. The automated system generally consists of the developed features extraction and classification stages. Signal processing techniques are used in the features extraction stage. Then, discriminant analysis and principal component analysis are employed to select dominant features for the classification process. The datasets of the cervical precancerous cells obtained from the feature selection process are classified using a hybrid multilayered perceptron network. The proposed system achieved 92% accuracy.

  3. Nanowire humidity optical sensor system based on fast Fourier transform technique

    NASA Astrophysics Data System (ADS)

    Rota-Rodrigo, S.; Pérez-Herrera, R.; Lopez-Aldaba, A.; López Bautista, M. C.; Esteban, O.; López-Amo, M.

    2015-09-01

    In this paper, a new sensor system for relative humidity measurements based on its interaction with the evanescent field of a nanowire is presented. The interrogation of the sensing head is carried out by monitoring the fast Fourier transform phase variations of one of the nanowire interference frequencies. This method is independent of the signal amplitude and also avoids the necessity of tracking the wavelength evolution in the spectrum, which can be a handicap when there are multiple interference frequency components with different sensitivities. The sensor is operated within a wide humidity range (20%-70% relative humidity) with a maximum sensitivity achieved of 0.14rad/% relative humidity. Finally, due to the system uses an optical interrogator as unique active element, the system presents a cost-effective feature.

  4. Collusion-resistant audio fingerprinting system in the modulated complex lapped transform domain.

    PubMed

    Garcia-Hernandez, Jose Juan; Feregrino-Uribe, Claudia; Cumplido, Rene

    2013-01-01

    Collusion-resistant fingerprinting paradigm seems to be a practical solution to the piracy problem as it allows media owners to detect any unauthorized copy and trace it back to the dishonest users. Despite the billionaire losses in the music industry, most of the collusion-resistant fingerprinting systems are devoted to digital images and very few to audio signals. In this paper, state-of-the-art collusion-resistant fingerprinting ideas are extended to audio signals and the corresponding parameters and operation conditions are proposed. Moreover, in order to carry out fingerprint detection using just a fraction of the pirate audio clip, block-based embedding and its corresponding detector is proposed. Extensive simulations show the robustness of the proposed system against average collusion attack. Moreover, by using an efficient Fast Fourier Transform core and standard computer machines it is shown that the proposed system is suitable for real-world scenarios. PMID:23762455

  5. Stable reduced-order models of generalized dynamical systems using coordinate-transformed Arnoldi algorithms

    SciTech Connect

    Silveira, L.M.; Kamon, M.; Elfadel, I.; White, J.

    1996-12-31

    Model order reduction based on Krylov subspace iterative methods has recently emerged as a major tool for compressing the number of states in linear models used for simulating very large physical systems (VLSI circuits, electromagnetic interactions). There are currently two main methods for accomplishing such a compression: one is based on the nonsymmetric look-ahead Lanczos algorithm that gives a numerically stable procedure for finding Pade approximations, while the other is based on a less well characterized Arnoldi algorithm. In this paper, we show that for certain classes of generalized state-space systems, the reduced-order models produced by a coordinate-transformed Arnoldi algorithm inherit the stability of the original system. Complete Proofs of our results will be given in the final paper.

  6. Transformation from proper time on earth to coordinate time in solar system barycentric space-time frame of reference

    NASA Technical Reports Server (NTRS)

    Moyer, T. D.

    1976-01-01

    An expression was derived for the time transformation t - tau, where t is coordinate time in the solar system barycentric space-time frame of reference and tau is proper time obtained from a fixed atomic clock on earth. This transformation is suitable for use in the computation of high-precision earth-based range and Doppler observables of a spacecraft or celestial body located anywhere in the solar system; it can also be used in obtaining computed values of very long baseline interferometry data types. The formulation for computing range and Doppler observables, which is an explicit function of the transformation t - tau, is described briefly.

  7. Verifying end-to-end system performance with the transformational information extraction model

    NASA Astrophysics Data System (ADS)

    Mauck, Alisha; Roszyk, Greg

    2006-05-01

    In the intelligence community, the volume of imagery data threatens to overwhelm the traditional process of information extraction. Satellite systems are capable of producing large quantities of imagery data every day. Traditionally, intelligence analysts have the arduous task of manually reviewing satellite imagery data and generating information products. In a time of increasing imagery data, this manual approach is not consistent with the goal of a timely and highly responsive system. These realities are key factors in Booz Allen Hamilton's transformational approach to information extraction. This approach employs information services and value added processes (VAP) to reduce the amount of data being manually reviewed. Booz Allen has utilized a specialization/generalization hierarchy to aggregate hundreds of thousands of imagery intelligence needs into sixteen information services. Information Services are automated by employing value added processes, which extract the information from the imagery data and generate information products. While the intelligence needs and information services remain relatively static in time, the VAP's have the ability to evolve rapidly with advancing technologies. The Booz Allen Transformational Information Extraction Model validates this automated approach by simulating realistic system parameters. The functional flow model includes image formation, three information services, six VAP's, and reduced manual intervention. Adjustable model variables for VAP time, VAP confidence, number of intelligence analyst, and time for analyst review provide a flexible framework for modeling different system cases. End-to-End system metrics such as intelligence need satisfaction, end-to-end timeliness, and sensitivity to number of analyst and VAP variables quantify the system performance.

  8. Bäcklund transformations, Lax system, conservation laws and multisoliton solutions for Jimbo-Miwa equation with Bell-polynomials

    NASA Astrophysics Data System (ADS)

    Singh, Manjit; Gupta, R. K.

    2016-08-01

    Based on binary Bell polynomial approach, the bilinear equation and B a ¨cklund transformations for (3+1)-dimensional Jimbo-Miwa equation are obtained. By virtue of Cole-Hopf transformation, Lax system is constructed by direct linearization of coupled system of binary Bell polynomials. Furthermore, infinite conservation laws are obtained from two field condition in quick and natural way. Finally, a test function of extended three wave method is used to construct multisoliton solutions via bilinear equation.

  9. Co-transforming bar and CsLEA enhanced tolerance to drought and salt stress in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Zhang, Jiyu; Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Wang, Yanrong

    2016-03-25

    Drought and high salinity are two major abiotic factors that restrict alfalfa productivity. A dehydrin protein, CsLEA, from the desert grass Cleistogenes songorica was transformed into alfalfa (Medicago sativa L.) via Agrobacterium-mediated transformation using the bar gene as a selectable marker, and the drought and salt stress tolerances of the transgenic plants were assessed. Thirty-nine of 119 transformants were positive, as screened by Basta, and further molecularly authenticated using PCR and RT-PCR. Phenotype observations revealed that the transgenic plants grew better than the wild-type (WT) plants after 15d of drought stress and 10d of salt stress: the leaves of WT alfalfa turned yellow, whereas the transgenic alfalfa leaves only wilted; after rewatering, the transgenic plants returned to a normal state, though the WT plants could not be restored. Evaluation of physiologic and biochemical indices during drought and salt stresses showed a relatively lower Na(+) content in the leaves of the transgenic plants, which would reduce toxic ion effects. In addition, the transgenic plants were able to maintain a higher relative water content (RWC), higher shoot biomass, fewer photosystem changes, decreased membrane injury, and a lower level of osmotic stress injury. These results demonstrate that overexpression of the CsLEA gene can enhance the drought and salt tolerance of transgenic alfalfa; in addition, carrying the bar gene in the genome may increase herbicide resistance.

  10. Co-transforming bar and CsLEA enhanced tolerance to drought and salt stress in transgenic alfalfa (Medicago sativa L.).

    PubMed

    Zhang, Jiyu; Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Wang, Yanrong

    2016-03-25

    Drought and high salinity are two major abiotic factors that restrict alfalfa productivity. A dehydrin protein, CsLEA, from the desert grass Cleistogenes songorica was transformed into alfalfa (Medicago sativa L.) via Agrobacterium-mediated transformation using the bar gene as a selectable marker, and the drought and salt stress tolerances of the transgenic plants were assessed. Thirty-nine of 119 transformants were positive, as screened by Basta, and further molecularly authenticated using PCR and RT-PCR. Phenotype observations revealed that the transgenic plants grew better than the wild-type (WT) plants after 15d of drought stress and 10d of salt stress: the leaves of WT alfalfa turned yellow, whereas the transgenic alfalfa leaves only wilted; after rewatering, the transgenic plants returned to a normal state, though the WT plants could not be restored. Evaluation of physiologic and biochemical indices during drought and salt stresses showed a relatively lower Na(+) content in the leaves of the transgenic plants, which would reduce toxic ion effects. In addition, the transgenic plants were able to maintain a higher relative water content (RWC), higher shoot biomass, fewer photosystem changes, decreased membrane injury, and a lower level of osmotic stress injury. These results demonstrate that overexpression of the CsLEA gene can enhance the drought and salt tolerance of transgenic alfalfa; in addition, carrying the bar gene in the genome may increase herbicide resistance. PMID:26906624

  11. Effectiveness of dereverberation, feature transformation, discriminative training methods, and system combination approach for various reverberant environments

    NASA Astrophysics Data System (ADS)

    Tachioka, Yuuki; Narita, Tomohiro; Watanabe, Shinji

    2015-12-01

    The recently released REverberant Voice Enhancement and Recognition Benchmark (REVERB) challenge includes a reverberant automatic speech recognition (ASR) task. This paper describes our proposed system based on multi-channel speech enhancement preprocessing and state-of-the-art ASR techniques. For preprocessing, we propose a single-channel dereverberation method with reverberation time estimation, which is combined with multichannel beamforming that enhances direct sound compared with the reflected sound. In addition, this paper also focuses on state-of-the-art ASR techniques such as discriminative training of acoustic models including the Gaussian mixture model, subspace Gaussian mixture model, and deep neural networks, as well as various feature transformation techniques. Although, for the REVERB challenge, it is necessary to handle various acoustic environments, a single ASR system tends to be overly tuned for a specific environment, which degrades the performance in the mismatch environments. To overcome this mismatch problem with a single ASR system, we use a system combination approach using multiple ASR systems with different features and different model types because a combination of various systems that have different error patterns is beneficial. In particular, we use our discriminative training technique for system combination that achieves better generalization by making systems complementary with the modified discriminative criteria. Experiments show the effectiveness of these approaches, reaching 6.76 and 18.60 % word error rates on the REVERB simulated and real test sets. These are 68.8 and 61.5 % relative improvements over the baseline.

  12. Dynamic visuomotor transformation involved with remote flying of a plane utilizes the 'Mirror Neuron' system.

    PubMed

    Callan, Daniel E; Gamez, Mario; Cassel, Daniel B; Terzibas, Cengiz; Callan, Akiko; Kawato, Mitsuo; Sato, Masa-aki

    2012-01-01

    Brain regions involved with processing dynamic visuomotor representational transformation are investigated using fMRI. The perceptual-motor task involved flying (or observing) a plane through a simulated Red Bull Air Race course in first person and third person chase perspective. The third person perspective is akin to remote operation of a vehicle. The ability for humans to remotely operate vehicles likely has its roots in neural processes related to imitation in which visuomotor transformation is necessary to interpret the action goals in an egocentric manner suitable for execution. In this experiment for 3(rd) person perspective the visuomotor transformation is dynamically changing in accordance to the orientation of the plane. It was predicted that 3(rd) person remote flying, over 1(st), would utilize brain regions composing the 'Mirror Neuron' system that is thought to be intimately involved with imitation for both execution and observation tasks. Consistent with this prediction differential brain activity was present for 3(rd) person over 1(st) person perspectives for both execution and observation tasks in left ventral premotor cortex, right dorsal premotor cortex, and inferior parietal lobule bilaterally (Mirror Neuron System) (Behaviorally: 1(st)>3(rd)). These regions additionally showed greater activity for flying (execution) over watching (observation) conditions. Even though visual and motor aspects of the tasks were controlled for, differential activity was also found in brain regions involved with tool use, motion perception, and body perspective including left cerebellum, temporo-occipital regions, lateral occipital cortex, medial temporal region, and extrastriate body area. This experiment successfully demonstrates that a complex perceptual motor real-world task can be utilized to investigate visuomotor processing. This approach (Aviation Cerebral Experimental Sciences ACES) focusing on direct application to lab and field is in contrast to standard

  13. System Efficiency of a Tap Transformer Based Grid Connection Topology Applied on a Direct Driven Generator for Wind Power

    PubMed Central

    2014-01-01

    Results from experiments on a tap transformer based grid connection system for a variable speed vertical axis wind turbine are presented. The tap transformer based system topology consists of a passive diode rectifier, DC-link, IGBT inverter, LCL-filter, and tap transformer. Full range variable speed operation is enabled by using the different step-up ratios of a tap transformer. Simulations using MATLAB/Simulink have been performed in order to study the behavior of the system. A full experimental set up of the system has been used in the laboratory study, where a clone of the on-site generator was driven by an induction motor and the system was connected to a resistive load to better evaluate the performance. Furthermore, the system is run and evaluated for realistic wind speeds and variable speed operation. For a more complete picture of the system performance, a case study using real site Weibull parameters is done, comparing different tap selection options. The results show high system efficiency at nominal power and an increase in overall power output for full tap operation in comparison with the base case, a standard transformer. In addition, the loss distribution at different wind speeds is shown, which highlights the dominant losses at low and high wind speeds. Finally, means for further increasing the overall system efficiency are proposed. PMID:25258733

  14. System efficiency of a tap transformer based grid connection topology applied on a direct driven generator for wind power.

    PubMed

    Apelfröjd, Senad; Eriksson, Sandra

    2014-01-01

    Results from experiments on a tap transformer based grid connection system for a variable speed vertical axis wind turbine are presented. The tap transformer based system topology consists of a passive diode rectifier, DC-link, IGBT inverter, LCL-filter, and tap transformer. Full range variable speed operation is enabled by using the different step-up ratios of a tap transformer. Simulations using MATLAB/Simulink have been performed in order to study the behavior of the system. A full experimental set up of the system has been used in the laboratory study, where a clone of the on-site generator was driven by an induction motor and the system was connected to a resistive load to better evaluate the performance. Furthermore, the system is run and evaluated for realistic wind speeds and variable speed operation. For a more complete picture of the system performance, a case study using real site Weibull parameters is done, comparing different tap selection options. The results show high system efficiency at nominal power and an increase in overall power output for full tap operation in comparison with the base case, a standard transformer. In addition, the loss distribution at different wind speeds is shown, which highlights the dominant losses at low and high wind speeds. Finally, means for further increasing the overall system efficiency are proposed.

  15. Evaluating transformational solvent systems for post-combustion CO2 separations

    SciTech Connect

    Heldebrant, David J.; Glezakou, Vassiliki Alexandra; Koech, Phillip K.; Mathias, Paul M.; Cantu Cantu, David; Rousseau, Roger J.; Malhotra, Deepika; Bhakta, Mukund; Bearden, Mark D.; Freeman, Charles J.; Zheng, Feng

    2014-01-06

    Broad research is underway on developing transformational solvents that can capture of CO2 from flue gas with lower energy compared to aqueous amines. Water-lean, or non-aqueous, solvents are being considered as a class of transformational solvents due to the prospect of lower energy duties by not having to heat and condense water. To date, little is known about the real world performance of water-lean solvent systems compared to commercial aqueous amine technologies, and whether or not they can utilize existing or at least similar processing infrastructure. This paper provides the key results from a comprehensive three-year study of the water-lean CO2-Binding Organic Liquids (CO2BOL) solvent platform coupled with Polarity-Swing Assisted Regeneration (PSAR). We present here thermodynamic, kinetic, and bench-scale data, followed by Aspen Plus projections of full-scale process performance for three CO2BOL/PSAR cases. This paper also provides discussions on materials performance and identifies viscosity as a critical property that most greatly limits the viability of water-lean solvent platforms. We provide results from a new effort spanning molecular modeling and synthesis and experimental testing to decipher the critical material properties needed to address this challenge. We conclude with implications for development of other water-lean solvent systems

  16. Transformation of Personal Computers and Mobile Phones into Genetic Diagnostic Systems

    PubMed Central

    2014-01-01

    Molecular diagnostics based on the polymerase chain reaction (PCR) offer rapid and sensitive means for detecting infectious disease, but prohibitive costs have impeded their use in resource-limited settings where such diseases are endemic. In this work, we report an innovative method for transforming a desktop computer and a mobile camera phone—devices that have become readily accessible in developing countries—into a highly sensitive DNA detection system. This transformation was achieved by converting a desktop computer into a de facto thermal cycler with software that controls the temperature of the central processing unit (CPU), allowing for highly efficient PCR. Next, we reconfigured the mobile phone into a fluorescence imager by adding a low-cost filter, which enabled us to quantitatively measure the resulting PCR amplicons. Our system is highly sensitive, achieving quantitative detection of as little as 9.6 attograms of target DNA, and we show that its performance is comparable to advanced laboratory instruments at approximately 1/500th of the cost. Finally, in order to demonstrate clinical utility, we have used our platform for the successful detection of genomic DNA from the parasite that causes Chagas disease, Trypanosoma cruzi, directly in whole, unprocessed human blood at concentrations 4-fold below the clinical titer of the parasite. PMID:25223929

  17. Transformation of personal computers and mobile phones into genetic diagnostic systems.

    PubMed

    Walker, Faye M; Ahmad, Kareem M; Eisenstein, Michael; Soh, H Tom

    2014-09-16

    Molecular diagnostics based on the polymerase chain reaction (PCR) offer rapid and sensitive means for detecting infectious disease, but prohibitive costs have impeded their use in resource-limited settings where such diseases are endemic. In this work, we report an innovative method for transforming a desktop computer and a mobile camera phone--devices that have become readily accessible in developing countries--into a highly sensitive DNA detection system. This transformation was achieved by converting a desktop computer into a de facto thermal cycler with software that controls the temperature of the central processing unit (CPU), allowing for highly efficient PCR. Next, we reconfigured the mobile phone into a fluorescence imager by adding a low-cost filter, which enabled us to quantitatively measure the resulting PCR amplicons. Our system is highly sensitive, achieving quantitative detection of as little as 9.6 attograms of target DNA, and we show that its performance is comparable to advanced laboratory instruments at approximately 1/500th of the cost. Finally, in order to demonstrate clinical utility, we have used our platform for the successful detection of genomic DNA from the parasite that causes Chagas disease, Trypanosoma cruzi, directly in whole, unprocessed human blood at concentrations 4-fold below the clinical titer of the parasite. PMID:25223929

  18. STARTS--a stable root transformation system for rapid functional analyses of proteins of the monocot model plant barley.

    PubMed

    Imani, Jafargholi; Li, Liang; Schäfer, Patrick; Kogel, Karl-Heinz

    2011-08-01

    Large data sets are generated from plants by the various 'omics platforms. Currently, a limiting step in data analysis is the assessment of protein function and its translation into a biological context. The lack of robust high-throughput transformation systems for monocotyledonous plants, to which the vast majority of crop plants belong, is a major restriction and impedes exploitation of novel traits in agriculture. Here we present a stable root transformation system for barley, termed STARTS, that allows assessment of gene function in root tissues within 6 weeks. The system is based on the finding that a callus, produced on root induction medium from the scutellum of the immature embryo, is able to regenerate roots from single transformed cells by concomitant suppression of shoot development. Using Agrobacterium tumefaciens-mediated transfer of genes involved in root development and pathogenesis, we show that those calli regenerate large amounts of uniformly transformed roots for in situ functional analysis of newly expressed proteins.

  19. Transforming Education to Strengthen Health Systems in the Sultanate of Oman

    PubMed Central

    White, Gillian

    2012-01-01

    Conspicuous gaps demonstrate a collective global failure in the world’s health systems as they struggle to manage complex and expensive demands. The Lancet Commission recently took a global interdisciplinary perspective and systematic approach to consider alliances between education for health professionals and health systems in order to address these problems. They concluded that positive outcomes require new instructional and institutional designs. Findings from the Lancet Commission have implications for the development of health professional education in Oman, particularly with regard to the call for integrative and transformative education for the next generation of health professionals. Education in the Omani health sector must keep up with increasing challenges in both the health and education sectors. PMID:23275838

  20. Quantum Color Image Encryption Algorithm Based on A Hyper-Chaotic System and Quantum Fourier Transform

    NASA Astrophysics Data System (ADS)

    Tan, Ru-Chao; Lei, Tong; Zhao, Qing-Min; Gong, Li-Hua; Zhou, Zhi-Hong

    2016-09-01

    To improve the slow processing speed of the classical image encryption algorithms and enhance the security of the private color images, a new quantum color image encryption algorithm based on a hyper-chaotic system is proposed, in which the sequences generated by the Chen's hyper-chaotic system are scrambled and diffused with three components of the original color image. Sequentially, the quantum Fourier transform is exploited to fulfill the encryption. Numerical simulations show that the presented quantum color image encryption algorithm possesses large key space to resist illegal attacks, sensitive dependence on initial keys, uniform distribution of gray values for the encrypted image and weak correlation between two adjacent pixels in the cipher-image.