Sample records for agrobacterium-based transformation method

  1. 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.

  2. Agrobacterium-mediated transformation of lipomyces

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Dai, Ziyu; Magnuson, Jon K.; Deng, Shuang

    This disclosure provides Agrobacterium-mediated transformation methods for the oil-producing (oleaginous) yeast Lipomyces sp., as well as yeast produced by the method. Such methods utilize Agrobacterium sp. cells that have a T-DNA binary plasmid, wherein the T-DNA binary plasmid comprises a first nucleic acid molecule encoding a first protein and a second nucleic acid molecule encoding a selective marker that permits growth of transformed Lipomyces sp. cells in selective culture media comprising an antibiotic.

  3. 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

  4. Female reproductive tissues are the primary target of Agrobacterium-mediated transformation by the Arabidopsis floral-dip method.

    PubMed

    Desfeux, C; Clough, S J; Bent, A F

    2000-07-01

    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 beta-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.

  5. Optimization of Agrobacterium-Mediated Transformation in Soybean

    PubMed Central

    Li, Shuxuan; Cong, Yahui; Liu, Yaping; Wang, Tingting; Shuai, Qin; Chen, Nana; Gai, Junyi; Li, Yan

    2017-01-01

    High transformation efficiency is a prerequisite for study of gene function and molecular breeding. Agrobacterium tumefaciens-mediated transformation is a preferred method in many plants. However, the transformation efficiency in soybean is still low. The objective of this study is to optimize Agrobacterium-mediated transformation in soybean by improving the infection efficiency of Agrobacterium and regeneration efficiency of explants. Firstly, four factors affecting Agrobacterium infection efficiency were investigated by estimation of the rate of GUS transient expression in soybean cotyledonary explants, including Agrobacterium concentrations, soybean explants, Agrobacterium suspension medium, and co-cultivation time. The results showed that an infection efficiency of over 96% was achieved by collecting the Agrobacterium at a concentration of OD650 = 0.6, then using an Agrobacterium suspension medium containing 154.2 mg/L dithiothreitol to infect the half-seed cotyledonary explants (from mature seeds imbibed for 1 day), and co-cultured them for 5 days. The Agrobacterium infection efficiencies for soybean varieties Jack Purple and Tianlong 1 were higher than the other six varieties. Secondly, the rates of shoot elongation were compared among six different concentration combinations of gibberellic acid (GA3) and indole-3-acetic acid (IAA). The shoot elongation rate of 34 and 26% was achieved when using the combination of 1.0 mg/L GA3 and 0.1 mg/L IAA for Jack Purple and Tianlong 1, respectively. This rate was higher than the other five concentration combinations of GA3 and IAA, with an 18 and 11% increase over the original laboratory protocol (a combination of 0.5 mg/L GA3 and 0.1 mg/L IAA), respectively. The transformation efficiency was 7 and 10% for Jack Purple and Tianlong 1 at this optimized hormone concentration combination, respectively, which was 2 and 6% higher than the original protocol, respectively. Finally, GUS histochemical staining, PCR, herbicide

  6. Optimization of Agrobacterium-Mediated Transformation in Soybean.

    PubMed

    Li, Shuxuan; Cong, Yahui; Liu, Yaping; Wang, Tingting; Shuai, Qin; Chen, Nana; Gai, Junyi; Li, Yan

    2017-01-01

    High transformation efficiency is a prerequisite for study of gene function and molecular breeding. Agrobacterium tumefaciens -mediated transformation is a preferred method in many plants. However, the transformation efficiency in soybean is still low. The objective of this study is to optimize Agrobacterium -mediated transformation in soybean by improving the infection efficiency of Agrobacterium and regeneration efficiency of explants. Firstly, four factors affecting Agrobacterium infection efficiency were investigated by estimation of the rate of GUS transient expression in soybean cotyledonary explants, including Agrobacterium concentrations, soybean explants, Agrobacterium suspension medium, and co-cultivation time. The results showed that an infection efficiency of over 96% was achieved by collecting the Agrobacterium at a concentration of OD 650 = 0.6, then using an Agrobacterium suspension medium containing 154.2 mg/L dithiothreitol to infect the half-seed cotyledonary explants (from mature seeds imbibed for 1 day), and co-cultured them for 5 days. The Agrobacterium infection efficiencies for soybean varieties Jack Purple and Tianlong 1 were higher than the other six varieties. Secondly, the rates of shoot elongation were compared among six different concentration combinations of gibberellic acid (GA 3 ) and indole-3-acetic acid (IAA). The shoot elongation rate of 34 and 26% was achieved when using the combination of 1.0 mg/L GA 3 and 0.1 mg/L IAA for Jack Purple and Tianlong 1, respectively. This rate was higher than the other five concentration combinations of GA 3 and IAA, with an 18 and 11% increase over the original laboratory protocol (a combination of 0.5 mg/L GA 3 and 0.1 mg/L IAA), respectively. The transformation efficiency was 7 and 10% for Jack Purple and Tianlong 1 at this optimized hormone concentration combination, respectively, which was 2 and 6% higher than the original protocol, respectively. Finally, GUS histochemical staining, PCR

  7. 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

  8. 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.

  9. Transformation of medicinal plants using Agrobacterium tumefaciens.

    PubMed

    Bandurska, Katarzyna; Berdowska, Agnieszka; Król, Małgorzata

    2016-12-20

    For many years attempts are made to develop efficient methods for transformation of medicinal plants via Agrobacterium tumefaciens. It is a soil bacteria which possess a natural ability to infect plants in places of injures which results in arise of cancerous growths (crown gall). This is possible thanks a transfer of fragment of Ti plasmid into plant cells and stable integration with a plant genome. Efficiency of medicinal plant transformation depends on many factors for example: Agrobacterium strain, methods and procedures of transformation as well as on plant species, type and age of the explants and regeneration conditions. The main goal of plant transformation is to increase the amount of naturally occurring bioactive compounds and the production of biopharmaceuticals. Genetic plant transformation via bacteria of the genus Agrobacterium is a complex process which requires detailed analysis of incorporated transgene expression and occurs only in the case when the plant cell acquires the ability to regenerate. In many cases, the regeneration efficiency observed in medicinal plants are inefficient after applied transformation procedures. To date there have been attempts of genetic transformation by using A. tumefaciens of medicinal plants belonging to the families: Apocynaceae, Araceae, Araliaceae, Asphodelaceae, Asteraceae, Begoniaceae, Crassulaceae, Fabaceae, Lamiaceae, Linaceae, Papaveraceae, Plantaginaceae, Scrophulariaceae and Solanaceae.

  10. Agrobacterium tumefaciens-mediated transformation of Mucor circinelloides.

    PubMed

    Nyilasi, I; Acs, K; Papp, T; Nagy, E; Vágvölgyi, C

    2005-01-01

    The Agrobacterium tumefaciens-mediated transformation of the zygomycetous fungus Mucor circinelloides is described. A method was also developed for the hygromycin B-based selection of Mucor transformants. Transformation with the hygromycin B phosphotransferase gene of Escherichia coli controlled by the heterologous Aspergillus nidulans trpC promoter resulted in hygromycin B-resistant clones. The presence of the hygromycin resistance gene in the genome of the transformants was verified by polymerase chain reaction and Southern hybridization: the latter analyses revealed integrations in the host genome at different sites in different transformants. The stability of transformants remained questionable during the latter analyses.

  11. 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.

  12. 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

  13. Development of an Agrobacterium-mediated stable transformation method for the sensitive plant Mimosa pudica.

    PubMed

    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.

  14. Agrobacterium tumefaciens-mediated transformation of oleaginous yeast Lipomyces species.

    PubMed

    Dai, Ziyu; Deng, Shuang; Culley, David E; Bruno, Kenneth S; Magnuson, Jon K

    2017-08-01

    Interest in using renewable sources of carbon, especially lignocellulosic biomass, for the production of hydrocarbon fuels and chemicals has fueled interest in exploring various organisms capable of producing hydrocarbon biofuels and chemicals or their precursors. The oleaginous (oil-producing) yeast Lipomyces starkeyi is the subject of active research regarding the production of triacylglycerides as hydrocarbon fuel precursors using a variety of carbohydrate and nutrient sources. The genome of L. starkeyi has been published, which opens the door to production strain improvements through the development and use of the tools of synthetic biology for this oleaginous species. The first step in establishment of synthetic biology tools for an organism is the development of effective and reliable transformation methods with suitable selectable marker genes and demonstration of the utility of the genetic elements needed for expression of introduced genes or deletion of endogenous genes. Chemical-based methods of transformation have been published but suffer from low efficiency. To address these problems, Agrobacterium-mediated transformation was investigated as an alternative method for L. starkeyi and other Lipomyces species. In this study, Agrobacterium-mediated transformation was demonstrated to be effective in the transformation of both L. starkeyi and other Lipomyces species. The deletion of the peroxisomal biogenesis factor 10 gene was also demonstrated in L. starkeyi. In addition to the bacterial antibiotic selection marker gene hygromycin B phosphotransferase, the bacterial β-glucuronidase reporter gene under the control of L. starkeyi translation elongation factor 1α promoter was also stably expressed in six different Lipomyces species. The results from this study demonstrate that Agrobacterium-mediated transformation is a reliable and effective genetic tool for homologous recombination and expression of heterologous genes in L. starkeyi and other Lipomyces

  15. 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. Copyright © 2014 Elsevier GmbH. All rights reserved.

  16. Agrobacterium tumefaciens-mediated transformation of oleaginous yeast Lipomyces species

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Dai, Ziyu; Deng, Shuang; Culley, David E.

    Background: Because of interest in the production of renewable bio-hydrocarbon fuels, various living organisms have been explored for their potential use in producing fuels and chemicals. The oil-producing (oleaginous) yeast Lipomyces starkeyi is the subject of active research regarding the production of lipids using a wide variety of carbon and nutrient sources. The genome of L. starkeyi has been published, which opens the door to production strain improvements using the tools of synthetic biology and metabolic engineering. However, using these tools for strain improvement requires the establishment of effective and reliable transformation methods with suitable selectable markers (antibiotic resistance ormore » auxotrophic marker genes) and the necessary genetic elements (promoters and terminators) for expression of introduced genes. Chemical-based methods have been published, but suffer from low efficiency or the requirement for targeting to rRNA loci. To address these problems, Agrobacterium-mediated transformation was investigated as an alternative method for L. starkeyi and other Lipomyces species. Results: In this study, Agrobacterium-mediated transformation was demonstrated to be effective in the transformation of both L. starkeyi and other Lipomyces species and that the introduced DNA can be reliably integrated into the chromosomes of these species. The gene deletion of Ku70 and Pex10 was also demonstrated in L. starkeyi. In addition to the bacterial antibiotic selection marker gene hygromycin B phosphotransferase, the bacterial -glucuronidase reporter gene under the control of L. starkeyi translation elongation factor 1 promoter was also stably expressed in seven different Lipomyces species. Conclusion: The results from this study clearly demonstrate that Agrobacterium-mediated transformation is a reliable genetic tool for gene deletion and integration and expression of heterologous genes in L. starkeyi and other Lipomyces species.« less

  17. 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.

  18. A Rapid, Highly Efficient and Economical Method of Agrobacterium-Mediated In planta Transient Transformation in Living Onion Epidermis

    PubMed Central

    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

  19. Agrobacterium- and Biolistic-Mediated Transformation of Maize B104 Inbred.

    PubMed

    Raji, Jennifer A; Frame, Bronwyn; Little, Daniel; Santoso, Tri Joko; Wang, Kan

    2018-01-01

    Genetic transformation of maize inbred genotypes remains non-routine for many laboratories due to variations in cell competency to induce embryogenic callus, as well as the cell's ability to receive and incorporate transgenes into the genome. This chapter describes two transformation protocols using Agrobacterium- and biolistic-mediated methods for gene delivery. Immature zygotic embryos of maize inbred B104, excised from ears harvested 10-14 days post pollination, are used as starting explant material. Disarmed Agrobacterium strains harboring standard binary vectors and the biolistic gun system Bio-Rad PDS-1000/He are used as gene delivery systems. The herbicide resistant bar gene and selection agent bialaphos are used for identifying putative transgenic type I callus events. Using the step-by-step protocols described here, average transformation frequencies (number of bialaphos resistant T 0 callus events per 100 explants infected or bombarded) of 4% and 8% can be achieved using the Agrobacterium- and biolistic-mediated methods, respectively. An estimated duration of 16-21 weeks is needed using either protocol from the start of transformation experiments to obtaining putative transgenic plantlets with established roots. In addition to laboratory in vitro procedures, detailed greenhouse protocols for producing immature ears as transformation starting material and caring for transgenic plants for seed production are also described.

  20. Efficient and genotype-independent Agrobacterium--mediated tomato transformation.

    PubMed

    Park, Sung Hun; Morris, Jay L; Park, Jung Eun; Hirschi, Kendal D; Smith, Roberta H

    2003-10-01

    An efficient method to transform five cultivars of tomato (Lycopersicon esculentum), Micro-Tom, Red Cherry, Rubion, Piedmont, and E6203 is reported. A comparison was made of leaf, cotyledon, and hypocotyl explants on 7 different regeneration media without Agrobacterium tumefaciens cocultivation and on 11 different media with cocultivation. Although all cultivars and explants formed callus and regenerated on the initial 7 media, cocultivation with A. tumefaciens significantly reduced the callus induction and regeneration. From these experiments, a transformation methodology using either hypocotyls or cotyledons cultured for one day on BA 1 mgL-1, NAA 0.1 mgL-1 and 3 days cocultivation with the Agrobacterium on this same medium followed by a transfer to a medium with zeatin 2 mgL-1 and IAA 0.1 mgL-1 for 4-6 weeks resulted in a greater than 20% transformation frequency for all five cultivars tested. In this transformation method, no feeder layers of tobacco, petunia or tomato suspension cultures were used, and the subculture media was minimal. Stable integration and transmission of the transgene in T1 generation plants were confirmed by Southern blot analysis. This procedure represents a simple, efficient and general means of transforming tomato.

  1. Improvement of Agrobacterium-mediated transformation and rooting of black cherry

    Treesearch

    Ying Wang; Paula M. Pijut

    2014-01-01

    An improved protocol for Agrobacterium-mediated transformation of an elite, mature black cherry genotype was developed. To increase transformation efficiency, vacuum infiltration, sonication, and a combination of the two treatments were applied during the cocultivation of leaf explants with Agrobacterium tumefaciens strain EHA105...

  2. Agrobacterium-mediated transformation of protocorm-like bodies in Cymbidium.

    PubMed

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

    2007-06-01

    Genetically transformed plants of Cymbidium were regenerated after cocultivating protocorm-like bodies (PLB) with Agrobacterium tumefaciens strain EHA101 (pIG121Hm) that harbored genes for beta-glucuronidase (gus), hygromycin phosphotransferase (hpt) and neomycin phosphotransferase II (nptII). PLB of three genotypes maintained in liquid new Dogashima medium (NDM), were subjected to transformation experiments. The PLB inoculated with Agrobacterium produced secondary PLB, 4 weeks after transfer onto 2.5 g L(-1) gellan gum-solidified NDM containing 10 g L(-1) sucrose, 20 mg L(-1) hygromycin and 40 mg L(-1) meropenem. Transformation efficiency was affected by genotype and the presence of acetosyringone during cocultivation. The highest transformation efficiency was obtained when PLB from the genotype L4 were infected and cocultivated with Agrobacterium on medium containing 100 muM acetosyringone. Transformation of the hygromycin-resistant plantlets regenerated from different sites of inoculated PLB was confirmed by histochemical GUS assay, PCR analysis and Southern blot hybridization.

  3. 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.

  4. Agrobacterium rhizogenes-mediated transformation: root cultures as a source of alkaloids.

    PubMed

    Sevón, Nina; Oksman-Caldentey, Kirsi-Marja

    2002-10-01

    Hairy roots, transformed with Agrobacterium rhizogenes, have been found to be suitable for the production of secondary metabolites because of their stable and high productivity in hormone-free culture conditions. A number of plant species including many medicinal plants have been successfully transformed with Agrobacterium rhizogenes. Transformed root cultures have also been found to be a potential source of high-value pharmaceuticals. In this article the most important alkaloids produced by hairy roots are summarised. Several different methods have been used to increase the alkaloid accumulation in hairy root cultures. The selection of high productive root lines based on somaclonal variation offers an interesting option to enhance the productivity. Elicitors and modification of culture conditions have been shown to increase the growth and the alkaloid production in some cases. Genetic engineering is a modern tool to regulate the secondary metabolism also in hairy roots. However, our knowledge on biosynthesis of many alkaloids is still poor. Only a limited number of enzymes and their respective genes which regulate the biosynthetic pathways are fully characterised.

  5. Novel compounds that enhance Agrobacterium-mediated plant transformation by mitigating oxidative stress.

    PubMed

    Dan, Yinghui; Zhang, Song; Zhong, Heng; Yi, Hochul; Sainz, Manuel B

    2015-02-01

    Agrobacterium tumefaciens caused tissue browning leading to subsequent cell death in plant transformation and novel anti-oxidative compounds enhanced Agrobacterium -mediated plant transformation by mitigating oxidative stress. Browning and death of cells transformed with Agrobacterium tumefaciens is a long-standing and high impact problem in plant transformation and the agricultural biotechnology industry, severely limiting the production of transgenic plants. Using our tomato cv. MicroTom transformation system, we demonstrated that Agrobacterium caused tissue browning (TB) leading to subsequent cell death by our correlation study. Without an antioxidant (lipoic acid, LA) TB was severe and associated with high levels of GUS transient expression and low stable transformation frequency (STF). LA addition shifted the curve in that most TB was intermediate and associated with the highest levels of GUS transient expression and STF. We evaluated 18 novel anti-oxidative compounds for their potential to enhance Agrobacterium-mediated transformation, by screening for TB reduction and monitoring GUS transient expression. Promising compounds were further evaluated for their effect on MicroTom and soybean STF. Among twelve non-antioxidant compounds, seven and five significantly (P < 0.05) reduced TB and increased STF, respectively. Among six antioxidants four of them significantly reduced TB and five of them significantly increased STF. The most efficient compound found to increase STF was melatonin (MEL, an antioxidant). Optimal concentrations and stages to use MEL in transformation were determined, and Southern blot analysis showed that T-DNA integration was not affected by MEL. The ability of diverse compounds with different anti-oxidative mechanisms can reduce Agrobacterium-mediated TB and increase STF, strongly supporting that oxidative stress is an important limiting factor in Agrobacterium-mediated transformation and the limiting factor can be controlled by these

  6. 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.

  7. 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.

  8. SacB-SacR gene cassette as the negative selection marker to suppress Agrobacterium overgrowth in Agrobacterium-mediated plant transformation

    USDA-ARS?s Scientific Manuscript database

    Agrobacterium overgrowth is a common problem in Agrobacterium-mediated plant transformation. To suppress the Agrobacterium overgrowth, various antibiotics have been used during plant tissue culture steps. The antibiotics are expensive and may adversely affect plant cell differentiation and reduce ...

  9. Agrobacterium tumefaciens-mediated transformation of Phellodendron amurense Rupr. using mature-seed explants.

    PubMed

    Yang, Jingli; Zhao, Bo; Kim, Yeon Bok; Zhou, Chenguang; Li, Chunyan; Chen, Yunlin; Zhang, Haizhen; Li, Cheng Hao

    2013-01-01

    An efficient transformation protocol was developed for Agrobacterium-mediated transformation of Phellodendron amurense Rupr. for using explants from mature seeds. The binary vector pCAMBIA1303, which contained hygromycin phosphotransferase (hptII) as a selectable marker gene and β-glucuronidase (GUS) as a reporter gene, was used for transformation studies. Different factors that affect survival of transformed buds, namely Agrobacterium infection method, bacterial strain, pre-culture duration, acetosyringone concentration, co-culture duration, and co-culture temperature were examined and optimized for transformation efficiency on the basis of GUS staining of hygromycin-resistant buds. Polymerase chain reaction (PCR), Southern blot and reverse transcription PCR confirmed the presence of the GUS gene. A transformation frequency of 13.1 % was achieved under optimized conditions for transformation (A. tumefaciens strain EHA105, 4 days co-cultivation at 4 °C, and infection of the pre-cultured mature-seed explants for 2 days). This is the first report of a successful genetic transformation protocol for P. amurense.

  10. 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.

  11. Efficient regeneration and improved sonication-assisted Agrobacterium transformation (SAAT) method for Catharanthus roseus.

    PubMed

    Alam, Pravej; Khan, Zainul Abdeen; Abdin, Malik Zainul; Khan, Jawaid A; Ahmad, Parvaiz; Elkholy, Shereen F; Sharaf-Eldin, Mahmoud A

    2017-05-01

    Catharanthus roseus is an important medicinal plant known for its pharmacological qualities such as antimicrobial, anticancerous, antifeedant, antisterility, antidiabetic activities. More than 130 bioactive compounds like vinblastine, vindoline and vincristine have been synthesized in this plant. Extensive studies have been carried out for optimization regeneration and transformation protocols. Most of the protocol described are laborious and time-consuming. Due to sophisticated protocol of regeneration and genetic transformation, the production of these bioactive molecules is less and not feasible to be commercialized worldwide. Here we have optimized the efficient protocol for regeneration and transformation to minimize the time scale and enhance the transformation frequency through Agrobacterium and sonication-assisted transformation (SAAT) method. In this study, hypocotyl explants responded best for maximal production of transformed shoots. The callus percentage were recorded 52% with 1.0 mg L -1 (BAP) and 0.5 mg L -1 (NAA) while 80% shoot percentage obtained with 4.0 mg L -1 (BAP) and 0.05 mg L -1 (NAA). The microscopic studies revealed that the expression of GFP was clearly localized in leaf tissue of the C. roseus after transformation of pRepGFP0029 construct. Consequently, transformation efficiency was revealed on the basis of GFP localization. The transformation efficiency of SAAT method was 6.0% comparable to 3.5% as conventional method. Further, PCR analysis confirmed the integration of the nptII gene in the transformed plantlets of C. roseus.

  12. An efficient Agrobacterium-mediated transformation method for aflatoxin generation fungus Aspergillus flavus.

    PubMed

    Han, Guomin; Shao, Qian; Li, Cuiping; Zhao, Kai; Jiang, Li; Fan, Jun; Jiang, Haiyang; Tao, Fang

    2018-05-01

    Aspergillus flavus often invade many important corps and produce harmful aflatoxins both in preharvest and during storage stages. The regulation mechanism of aflatoxin biosynthesis in this fungus has not been well explored mainly due to the lack of an efficient transformation method for constructing a genome-wide gene mutant library. This challenge was resolved in this study, where a reliable and efficient Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for A. flavus NRRL 3357 was established. The results showed that removal of multinucleate conidia, to collect a homogenous sample of uninucleate conidia for use as the transformation material, is the key step in this procedure. A. tumefaciens strain AGL-1 harboring the ble gene for zeocin resistance under the control of the gpdA promoter from A. nidulans is suitable for genetic transformation of this fungus. We successfully generated A. flavus transformants with an efficiency of ∼ 60 positive transformants per 10 6 conidia using our protocol. A small-scale insertional mutant library (∼ 1,000 mutants) was constructed using this method and the resulting several mutants lacked both production of conidia and aflatoxin biosynthesis capacity. Southern blotting analysis demonstrated that the majority of the transformants contained a single T-DNA insert on the genome. To the best of our knowledge, this is the first report of genetic transformation of A. flavus via ATMT and our protocol provides an effective tool for construction of genome-wide gene mutant libraries for functional analysis of important genes in A. flavus.

  13. Assessment of factors affecting Agrobacterium-mediated genetic transformation of the unicellular green alga, Chlorella vulgaris.

    PubMed

    Cha, Thye San; Yee, Willy; Aziz, Ahmad

    2012-04-01

    The successful establishment of an Agrobacterium-mediated transformation method and optimisation of six critical parameters known to influence the efficacy of Agrobacterium T-DNA transfer in the unicellular microalga Chlorella vulgaris (UMT-M1) are reported. Agrobacterium tumefaciens strain LBA4404 harbouring the binary vector pCAMBIA1304 containing the gfp:gusA fusion reporter and a hygromycin phosphotransferase (hpt) selectable marker driven by the CaMV35S promoter were used for transformation. Transformation frequency was assessed by monitoring transient β-glucuronidase (GUS) expression 2 days post-infection. It was found that co-cultivation temperature at 24°C, co-cultivation medium at pH 5.5, 3 days of co-cultivation, 150 μM acetosyringone, Agrobacterium density of 1.0 units (OD(600)) and 2 days of pre-culture were optimum variables which produced the highest number of GUS-positive cells (8.8-20.1%) when each of these parameters was optimised individually. Transformation conducted with the combination of all optimal parameters above produced 25.0% of GUS-positive cells, which was almost a threefold increase from 8.9% obtained from un-optimised parameters. Evidence of transformation was further confirmed in 30% of 30 randomly-selected hygromycin B (20 mg L(-1)) resistant colonies by polymerase chain reaction (PCR) using gfp:gusA and hpt-specific primers. The developed transformation method is expected to facilitate the genetic improvement of this commercially-important microalga.

  14. Differential roles of glucosinolates and camalexin at different stages of Agrobacterium-mediated transformation.

    PubMed

    Shih, Po-Yuan; Chou, Shu-Jen; Müller, Caroline; Halkier, Barbara Ann; Deeken, Rosalia; Lai, Erh-Min

    2018-03-02

    Agrobacterium tumefaciens is the causal agent of crown gall disease in a wide range of plants via a unique interkingdom DNA transfer from bacterial cells into the plant genome. Agrobacterium tumefaciens is capable of transferring its T-DNA into different plant parts at different developmental stages for transient and stable transformation. However, the plant genes and mechanisms involved in these transformation processes are not well understood. We used Arabidopsis thaliana Col-0 seedlings to reveal the gene expression profiles at early time points during Agrobacterium infection. Common and differentially expressed genes were found in shoots and roots. A gene ontology analysis showed that the glucosinolate (GS) biosynthesis pathway was an enriched common response. Strikingly, several genes involved in indole glucosinolate (iGS) modification and the camalexin biosynthesis pathway were up-regulated, whereas genes in aliphatic glucosinolate (aGS) biosynthesis were generally down-regulated, on Agrobacterium infection. Thus, we evaluated the impacts of GSs and camalexin during different stages of Agrobacterium-mediated transformation combining Arabidopsis mutant studies, metabolite profiling and exogenous applications of various GS hydrolysis products or camalexin. The results suggest that the iGS hydrolysis pathway plays an inhibitory role on transformation efficiency in Arabidopsis seedlings at the early infection stage. Later in the Agrobacterium infection process, the accumulation of camalexin is a key factor inhibiting tumour development on Arabidopsis inflorescence stalks. In conclusion, this study reveals the differential roles of GSs and camalexin at different stages of Agrobacterium-mediated transformation and provides new insights into crown gall disease control and improvement of plant transformation. © 2018 THE AUTHORS. MOLECULAR PLANT PATHOLOGY PUBLISHED BY BRITISH SOCIETY FOR PLANT PATHOLOGY AND JOHN WILEY & SONS LTD.

  15. Agrobacterium tumefasciens-mediated transformation of the aquatic fungus Blastocladiella emersonii.

    PubMed

    Vieira, André L G; Camilo, César M

    2011-08-01

    Agrobacterium tumefaciens is widely used for plant DNA transformation and more recently, has also been used to transform yeast, filamentous fungi and even human cells. Using this technique, we developed the first transformation protocol for the saprobic aquatic fungus Blastocladiella emersonii, a Blastocladiomycete localized at the base of fungal phylogenetic tree, which has been shown as a promising and interesting model of study of cellular function and differentiation. We constructed binary T-DNA vectors containing hygromycin phosphotransferase (hph) or enhanced green fluorescent protein (egfp) genes, under the control of Aspergillus nidulans trpC promoter and terminator sequences. 24 h of co-cultivation in induction medium (IM) agar plates, followed by transfer to PYG-agar plates containing cefotaxim to kill Agrobacterium tumefsciens and hygromycin to select transformants, resulted in growth and sporulation of resistant transformants. Genomic DNA from the pool o resistant zoospores were shown to contain T-DNA insertion as evidenced by PCR amplification of hph gene. Using a similar protocol we could also evidence the expression of enhanced green fluorescent protein (EGFP) in zoospores derived from transformed cells. This protocol can also open new perspectives for other non-transformable closely related fungi, like the Chytridiomycete class. Copyright © 2011 Elsevier Inc. All rights reserved.

  16. Detection of Oil Palm Root Penetration by Agrobacterium-Mediated Transformed Ganoderma boninense, Expressing Green Fluorescent Protein.

    PubMed

    Govender, Nisha; Wong, Mui-Yun

    2017-04-01

    A highly efficient and reproducible Agrobacterium-mediated transformation protocol for Ganoderma boninense was developed to facilitate observation of the early stage infection of basal stem rot (BSR). The method was proven amenable to different explants (basidiospore, protoplast, and mycelium) of G. boninense. The transformation efficiency was highest (62%) under a treatment combination of protoplast explant and Agrobacterium strain LBA4404, with successful expression of an hyg marker gene and gus-gfp fusion gene under the control of heterologous p416 glyceraldehyde 3-phosphate dehydrogenase promoter. Optimal transformation conditions included a 1:100 Agrobacterium/explant ratio, induction of Agrobacterium virulence genes in the presence of 250 μm acetosyringone, co-cultivation at 22°C for 2 days on nitrocellulose membrane overlaid on an induction medium, and regeneration of transformants on potato glucose agar prepared with 0.6 M sucrose and 20 mM phosphate buffer. Evaluated transformants were able to infect root tissues of oil palm plantlets with needle-like microhyphae during the penetration event. The availability of this model pathogen system for BSR may lead to a better understanding of the pathogenicity factors associated with G. boninense penetration into oil palm roots.

  17. Gene disruption in Trichoderma atroviride via Agrobacterium-mediated transformation.

    PubMed

    Zeilinger, Susanne

    2004-02-01

    A modified Agrobacterium-mediated transformation method for the efficient disruption of two genes encoding signaling compounds of the mycoparasite Trichoderma atroviride is described, using the hph gene of Escherichia coli as selection marker. The transformation vectors contained about 1 kb of 5' and 3' non-coding regions from the tmk1 (encoding a MAP kinase) or tga3 (encoding an alpha-subunit of a heterotrimeric G protein) target loci flanking a selection marker. Transformation of fungal conidia and selection on hygromycin-containing media applying an overlay-based procedure, which overcomes the lack of formation of distinct single colonies by the fungus, led to stable clones for both disruption constructs. Southern and PCR analyses proved gene disruption by single-copy homologous integration with a frequency of approximately 60% for both genes; and the loss of tmk1 and tga3 transcript formation in the disruptants was demonstrated by RT-PCR.

  18. Cytoskeletal dynamics in interphase, mitosis and cytokinesis analysed through Agrobacterium-mediated transient transformation of tobacco BY-2 cells.

    PubMed

    Buschmann, H; Green, P; Sambade, A; Doonan, J H; Lloyd, C W

    2011-04-01

    Transient transformation with Agrobacterium is a widespread tool allowing rapid expression analyses in plants. However, the available methods generate expression in interphase and do not allow the routine analysis of dividing cells. Here, we present a transient transformation method (termed 'TAMBY2') to enable cell biological studies in interphase and cell division. Agrobacterium-mediated transient gene expression in tobacco BY-2 was analysed by Western blotting and quantitative fluorescence microscopy. Time-lapse microscopy of cytoskeletal markers was employed to monitor cell division. Double-labelling in interphase and mitosis enabled localization studies. We found that the transient transformation efficiency was highest when BY-2/Agrobacterium co-cultivation was performed on solid medium. Transformants produced in this way divided at high frequency. We demonstrated the utility of the method by defining the behaviour of a previously uncharacterized microtubule motor, KinG, throughout the cell cycle. Our analyses demonstrated that TAMBY2 provides a flexible tool for the transient transformation of BY-2 with Agrobacterium. Fluorescence double-labelling showed that KinG localizes to microtubules and to F-actin. In interphase, KinG accumulates on microtubule lagging ends, suggesting a minus-end-directed function in vivo. Time-lapse studies of cell division showed that GFP-KinG strongly labels preprophase band and phragmoplast, but not the metaphase spindle. © 2010 The Authors. New Phytologist © 2010 New Phytologist Trust.

  19. Optimization of factors influencing microinjection method for Agrobacterium tumefaciens-mediated transformation of tomato.

    PubMed

    Vinoth, S; Gurusaravanan, P; Jayabalan, N

    2013-02-01

    A simple and efficient protocol for Agrobacterium-mediated genetic transformation of tomato was developed using combination of non-tissue culture and micropropagation systems. Initially, ESAM region of 1-day-old germinated tomato seeds were microinjected for one to five times with Agrobacterium inoculums (OD(600) = 0.2-1.0). The germinated seeds were cocultivated in the MS medium fortified with (0-200 mM) acetosyringone and minimal concentrations of (0-20 mg L(-1)) kanamycin, and the antibiotic concentration was doubled during the second round of selection. Bacterial concentration of OD(600) = 0.6 served as an optimal concentration for infection and the transformation efficiency was significantly higher of about 46.28 %. In another set of experiment, an improved and stable regeneration system was adapted for the explants from the selection medium. Four-day-old double cotyledonary nodal explants were excised from the microinjected seedlings and cultured onto the MS medium supplemented with 1.5 mg L(-1) thidiazuron, 1.5 mg L(-1) indole-3-butyric acid, 30 mg L(-1) kanamycin, and 0-1.5 mg L(-1) adenine sulphate. Maximum of 9 out of 13 micropropagated shoots were shown positive to GUS assay. By this technique, the transformation efficiency was increased from 46.28 to 65.90 %. Thus, this paper reports the successful protocol for the mass production of transformants using microinjection and micropropagation techniques.

  20. 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.

  1. 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

  2. Development of efficient plant regeneration and transformation system for impatiens using Agrobacterium tumefaciens and multiple bud cultures as explants.

    PubMed

    Dan, Yinghui; Baxter, Aaron; Zhang, Song; Pantazis, Christopher J; Veilleux, Richard E

    2010-08-09

    Impatiens (Impatiens walleriana) is a top selling floriculture crop. The potential for genetic transformation of Impatiens to introduce novel flower colors or virus resistance has been limited by its general recalcitrance to tissue culture and transformation manipulations. We have established a regeneration and transformation system for Impatiens that provides new alternatives to genetic improvement of this crop. In a first step towards the development of transgenic INSV-resistant Impatiens, we developed an efficient plant regeneration system using hypocotyl segments containing cotyledonary nodes as explants. With this regeneration system, 80% of explants produced an average of 32.3 elongated shoots per initial explant plated, with up to 167 elongated shoots produced per explant. Rooting efficiency was high, and 100% of shoots produced roots within 12 days under optimal conditions, allowing plant regeneration within approximately 8 weeks. Using this regeneration system, we developed an efficient Agrobacterium-mediated Impatiens transformation method using in vitro multiple bud cultures as explants and a binary plasmid (pHB2892) bearing gfp and nptII genes. Transgenic Impatiens plants, with a frequency up to 58.9%, were obtained within 12 to 16 weeks from inoculation to transfer of transgenic plants to soil. Transgenic plants were confirmed by Southern blot, phenotypic assays and T1 segregation analysis. Transgene expression was observed in leaves, stems, roots, flowers, and fruit. The transgenic plants were fertile and phenotypically normal. We report the development of a simple and efficient Agrobacterium-mediated transformation system for Impatiens. To the best of our knowledge, there have been no reports of Agrobacterium-mediated transformation of Impatiens with experimental evidence of stable integration of T-DNA and of Agrobacterium-mediated transformation method for plants using in vitro maintained multiple bud cultures as explants. This transformation system

  3. 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

  4. 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.

  5. A simple and reliable multi-gene transformation method for switchgrass.

    PubMed

    Ogawa, Yoichi; Shirakawa, Makoto; Koumoto, Yasuko; Honda, Masaho; Asami, Yuki; Kondo, Yasuhiro; Hara-Nishimura, Ikuko

    2014-07-01

    A simple and reliable Agrobacterium -mediated transformation method was developed for switchgrass. Using this method, many transgenic plants carrying multiple genes-of-interest could be produced without untransformed escape. Switchgrass (Panicum virgatum L.) is a promising biomass crop for bioenergy. To obtain transgenic switchgrass plants carrying a multi-gene trait in a simple manner, an Agrobacterium-mediated transformation method was established by constructing a Gateway-based binary vector, optimizing transformation conditions and developing a novel selection method. A MultiRound Gateway-compatible destination binary vector carrying the bar selectable marker gene, pHKGB110, was constructed to introduce multiple genes of interest in a single transformation. Two reporter gene expression cassettes, GUSPlus and gfp, were constructed independently on two entry vectors and then introduced into a single T-DNA region of pHKGB110 via sequential LR reactions. Agrobacterium tumefaciens EHA101 carrying the resultant binary vector pHKGB112 and caryopsis-derived compact embryogenic calli were used for transformation experiments. Prolonged cocultivation for 7 days followed by cultivation on media containing meropenem improved transformation efficiency without overgrowth of Agrobacterium, which was, however, not inhibited by cefotaxime or Timentin. In addition, untransformed escape shoots were completely eliminated during the rooting stage by direct dipping the putatively transformed shoots into the herbicide Basta solution for a few seconds, designated as the 'herbicide dipping method'. It was also demonstrated that more than 90 % of the bar-positive transformants carried both reporters delivered from pHKGB112. This simple and reliable transformation method, which incorporates a new selection technique and the use of a MultiRound Gateway-based binary vector, would be suitable for producing a large number of transgenic lines carrying multiple genes.

  6. Agrobacterium-mediated transformation of Fraxinus pennsylvanica hypocotyls and plant regeneration

    Treesearch

    Ningxia Du; Paula M. Pijut

    2009-01-01

    A genetic transformation protocol for green ash (Fraxinus pennsylvanica) hypocotyl explants was developed. Green ash hypocotyls were transformed using Agrobacterium tumefaciens strain EHA105 harboring binary vector pq35GR containing the neomycin phosphotransferase (nptII) and β-glucuronidase (GUS) fusion...

  7. Genotype-independent and enhanced in planta Agrobacterium tumefaciens-mediated genetic transformation of peanut [Arachis hypogaea (L.)].

    PubMed

    Karthik, Sivabalan; Pavan, Gadamchetty; Sathish, Selvam; Siva, Ramamoorthy; Kumar, Periyasamy Suresh; Manickavasagam, Markandan

    2018-04-01

    Agrobacterium infection and regeneration of the putatively transformed plant from the explant remains arduous for some crop species like peanut. Henceforth, a competent and reproducible in planta genetic transformation protocol is established for peanut cv. CO7 by standardizing various factors such as pre-culture duration, acetosyringone concentration, duration of co-cultivation, sonication and vacuum infiltration. In the present investigation, Agrobacterium tumefaciens strain EHA105 harboring the binary vector pCAMBIA1301- bar was used for transformation. The two-stage selection was carried out using 4 and 250 mg l -1 BASTA ® to completely eliminate the chimeric and non-transformed plants. The transgene integration into plant genome was evaluated by GUS histochemical assay, polymerase chain reaction (PCR), and Southern blot hybridization. Among the various combinations and concentrations analyzed, highest transformation efficiency was obtained when the 2-day pre-cultured explants were subjected to sonication for 6 min and vacuum infiltrated for 3 min in Agrobacterium suspension, and co-cultivated on MS medium supplemented with 150 µM acetosyringone for 3 days. The fidelity of the standardized in planta transformation method was assessed in five peanut cultivars and all the cultivars responded positively with a transformation efficiency ranging from minimum 31.3% (with cv. CO6) to maximum 38.6% (with cv. TMV7). The in planta transformation method optimized in this study could be beneficial to develop superior peanut cultivars with desirable genetic traits.

  8. 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.

  9. Establishment of an efficient genetic transformation method in Dunaliella tertiolecta mediated by Agrobacterium tumefaciens.

    PubMed

    Norzagaray-Valenzuela, Claudia D; Germán-Báez, Lourdes J; Valdez-Flores, Marco A; Hernández-Verdugo, Sergio; Shelton, Luke M; Valdez-Ortiz, Angel

    2018-05-16

    Microalgae are photosynthetic microorganisms widely used for the production of highly valued compounds, and recently they have been shown to be promising as a system for the heterologous expression of proteins. Several transformation methods have been successfully developed, from which the Agrobacterium tumefaciens-mediated method remains the most promising. However, microalgae transformation efficiency by A. tumefaciens is shown to vary depending on several transformation conditions. The present study aimed to establish an efficient genetic transformation system in the green microalgae Dunaliella tertiolecta using the A. tumefaciens method. The parameters assessed were the infection medium, the concentration of the A. tumefaciens and co-culture time. As a preliminary screening, the expression of the gusA gene and the viability of transformed cells were evaluated and used to calculate a novel parameter called Transformation Efficiency Index (TEI). The statistical analysis of TEI values showed five treatments with the highest gusA gene expression. To ensure stable transformation, transformed colonies were cultured on selective medium using hygromycin B and the DNA of resistant colonies were extracted after five subcultures and molecularly analyzed by PCR. Results revealed that treatments which use solid infection medium, A. tumefaciens OD 600  = 0.5 and co-culture times of 72 h exhibited the highest percentage of stable gusA expression. Overall, this study established an efficient, optimized A. tumefaciens-mediated genetic transformation of D. tertiolecta, which represents a relatively easy procedure with no expensive equipment required. This simple and efficient protocol opens the possibility for further genetic manipulation of this commercially-important microalgae for biotechnological applications. Copyright © 2018 Elsevier B.V. All rights reserved.

  10. Agrobacterium-mediated genetic transformation of Fraxinus americana hypocotyls

    Treesearch

    Kaitlin J. Palla; Paula M. Pijut

    2015-01-01

    An Agrobacterium tumefaciens-mediated genetic transformation system was successfully developed for white ash (Fraxinus americana) using hypocotyls as the initial explants. Hypocotyls isolated from mature embryos germinated on Murashige and Skoog (MS) medium supplemented with 22.2 µM 6-benzyladenine (BA) and 0.5 µM...

  11. Factors influencing Agrobacterium-mediated embryogenic callus transformation of Valencia sweet orange (Citrus sinensis) containing the pTA29-barnase gene.

    PubMed

    Li, D D; Shi, W; Deng, X X

    2003-12-01

    Valencia sweet orange (Citrus sinensis (L.) Osbeck) calluses were used as explants to develop a new transformation system for citrus mediated by Agrobacterium tumefaciens. Factors affecting Agrobacterium-mediated transformation efficiency included mode of pre-cultivation, temperature of cocultivation and presence of acetosyringone (AS). The highest transformation efficiency was obtained with a 4-day pre-cultivation period in liquid medium. Transformation efficiency was higher when cocultivation was performed for 3 days at 19 degrees C than at 23 or 28 degrees C. Almost no resistant callus was obtained if the cocultivation medium lacked AS. The transformation procedure yielded transgenic Valencia plants containing the pTA29-barnase gene, as verified by PCR amplification and confirmed by Southern blotting. Because male sterility is a common factor leading to seedlessness in citrus cultivars with parthenocarpic characteristics, production of seedless citrus genotypes by Agrobacterium-mediated genetic transformation is a promising alternative to conventional breeding methods.

  12. Agrobacterium tumefaciens-mediated transformation of Narcissus tazzeta var. chinensis.

    PubMed

    Lu, Gang; Zou, Qingcheng; Guo, Deping; Zhuang, Xiaoying; Yu, Xiaolin; Xiang, Xun; Cao, Jiashu

    2007-09-01

    Phytoene synthase (PSY), as a key regulatory enzyme for carotene biosynthesis, plays an important role in regulating color formation in many species. In the present study, a protocol was developed for the transformation of Narcissus tazzeta var chinensis using Agrobacterium tumefaciens strain LBA4404 harboring a binary vector pCAMBIA1301 plasmid which contained an antisense phytoene synthase gene, a reporter beta-glucuronidase gene and a selectable marker hygromycin phosphotransferase gene. Effects of some factors on efficiency of transformation and regeneration were examined. Preculture of the explants for 6 days before inoculation enhanced the transient GUS expression. The addition of acetosyringone (AS) at 100 micromol l(-1) for inoculation and a period of 3 days co-cultivation yielded efficient transient GUS expression. Transformants were obtained through selection on MS medium containing 5 mg l(-1) 6-benzylaminopurine (BA), 0.1 mg l(-1)alpha-naphthalene acetic acid (NAA) and 40 mg l(-1) hygromycin. The transformation frequency was 1.24% based on PCR analysis of gus gene. One or two copies of transgene were demonstrated in different transformations by Southern blotting analyses. Northern blotting results confirmed that the transcription of the endogenous psy gene in transgenic plants was inhibited or silenced. The method reported here provides new opportunities for improvement of quality traits of Narcissus tazzeta via genetic transformation.

  13. Agrobacterium-medicated transformation of mature Prunus serotina (black cherry) and regeneration of trangenic shoots

    Treesearch

    Xiaomei Liu; Paula Pijut

    2010-01-01

    A protocol for Agrobacterium-mediated transformation was developed for in vitro leaf explants of an elite, mature Prunus serotina tree. Agrobacterium tumefaciens strain EHA105 harboring an RNAi plasmid with the black cherry AGAMOUS (AG) gene was used. Bacteria were induced...

  14. Persistence of Agrobacterium tumefaciens in transformed conifers.

    PubMed

    Charity, Julia A; Klimaszewska, Krystyna

    2005-01-01

    Previous studies have shown that the widely used plant transformation vector Agrobacterium tumefaciens can persist in genetically engineered plants in vitro and in transgenic greenhouse-grown plants, despite the use of counter-selective antibiotics. However, little is known regarding Agrobacterium persistence in tree species. To understand the kinetics of A. tumefaciens decline and persistence in transformation experiments, we assayed for the presence of A. tumefaciens in spruce and pine embryogenic tissue for up to 10 weeks post-transformation. The A. tumefaciens populations declined rapidly in the first five days post-cocultivation but generally declined more slowly in pine, relative to spruce. No bacteria were detected in spruce embryogenic tissue beyond four weeks after cocultivation, however in pine there were -100 colony forming units per g tissue at 10 weeks post-cocultivation. We present evidence that the detection limit for PCR using virD2 primers to detect A. tumefaciens in a background of pine needle DNA was approximately 10(9)-10(10) A. tumefaciens cells per g of tissue. We also assayed for A. tumefaciens in transgenic pine and spruce embryogenic tissue and from needles, branches, stems and roots of transformed plants, up to four years post-inoculation. Occasionally A. tumefaciens was detected in embryogenic tissue up to 12 months post-inoculation. A. tumefaciens was never detected in cultured embryogenic tissue more than twelve months after inoculation, nor in developing somatic embryos or germinating plantlets, nor any of the parts of greenhouse-grown plants. From these data we conclude that if A. tumefaciens persists in transgenic conifers, it does so beneath our ability to detect it.

  15. Transformation of lettuce (Lactuca sativa) mediated by Agrobacterium tumefaciens.

    PubMed

    Michelmore, R; Marsh, E; Seely, S; Landry, B

    1987-12-01

    Lactuca sativa can be routinely transformed using Ti plasmids of Agrobacterium tumefaciens containing a chimeric kanamycin resistance gene (NOS.NPTII.NOS). Critical experimental variables were plant genotype, bacterial concentration, presence of a nurse culture and timing of transfers between tissue culture media. Transformation was confirmed by the ability to callus and root in the presence of kanamycin, nopaline production, and by hybridization in Southern blots. Transformation has been achieved with several Ti vectors. Several hundred transformed plants have been regenerated. Kanamycin resistance was inherited monogenically. Homozygotes can be selected by growing R2 seedlings on media containing G418.

  16. Transformation of the cucurbit powdery mildew pathogen Podosphaera xanthii by Agrobacterium tumefaciens.

    PubMed

    Martínez-Cruz, Jesús; Romero, Diego; de Vicente, Antonio; Pérez-García, Alejandro

    2017-03-01

    The obligate biotrophic fungal pathogen Podosphaera xanthii is the main causal agent of powdery mildew in cucurbit crops all over the world. A major limitation of molecular studies of powdery mildew fungi (Erysiphales) is their genetic intractability. In this work, we describe a robust method based on the promiscuous transformation ability of Agrobacterium tumefaciens for reliable transformation of P. xanthii. The A. tumefaciens-mediated transformation (ATMT) system yielded transformants of P. xanthii with diverse transferred DNA (T-DNA) constructs. Analysis of the resultant transformants showed the random integration of T-DNA into the P. xanthii genome. The integrations were maintained in successive generations in the presence of selection pressure. Transformation was found to be transient, because in the absence of selection agent, the introduced genetic markers were lost due to excision of T-DNA from the genome. The ATMT system represents a potent tool for genetic manipulation of P. xanthii and will likely be useful for studying other biotrophic fungi. We hope that this method will contribute to the development of detailed molecular studies of the intimate interaction established between powdery mildew fungi and their host plants. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

  17. 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.

  18. An improved ternary vector system for Agrobacterium-mediated rapid maize transformation.

    PubMed

    Anand, Ajith; Bass, Steven H; Wu, Emily; Wang, Ning; McBride, Kevin E; Annaluru, Narayana; Miller, Michael; Hua, Mo; Jones, Todd J

    2018-05-01

    A simple and versatile ternary vector system that utilizes improved accessory plasmids for rapid maize transformation is described. This system facilitates high-throughput vector construction and plant transformation. The super binary plasmid pSB1 is a mainstay of maize transformation. However, the large size of the base vector makes it challenging to clone, the process of co-integration is cumbersome and inefficient, and some Agrobacterium strains are known to give rise to spontaneous mutants resistant to tetracycline. These limitations present substantial barriers to high throughput vector construction. Here we describe a smaller, simpler and versatile ternary vector system for maize transformation that utilizes improved accessory plasmids requiring no co-integration step. In addition, the newly described accessory plasmids have restored virulence genes found to be defective in pSB1, as well as added virulence genes. Testing of different configurations of the accessory plasmids in combination with T-DNA binary vector as ternary vectors nearly doubles both the raw transformation frequency and the number of transformation events of usable quality in difficult-to-transform maize inbreds. The newly described ternary vectors enabled the development of a rapid maize transformation method for elite inbreds. This vector system facilitated screening different origins of replication on the accessory plasmid and T-DNA vector, and four combinations were identified that have high (86-103%) raw transformation frequency in an elite maize inbred.

  19. Effect of selection agents to Chrysanthemum (Chrysanthemum morifolium) callus growth after Agrobacterium-mediated genetic transformation

    NASA Astrophysics Data System (ADS)

    Sjahril, R.; Jamaluddin, I.; Nadir, M.; Asman; Dungga, N. E.

    2018-05-01

    Genetic transformation mediated by Agrobacterium tumefaciens requires an efficient selection method for successful progress of transformation. This study aims to determine the concentration and kind of antibiotics and selection agents used during transformation to formulate standard protocol of chrysanthemum in the process of propagating disease resistant Chrysanthemum mediated by Agrobacterium tumefaciens EHA105 (pEKB-WD). The experiments were performed by planting chrysanthemum explants leaf cutting (5 mm diameter on NAA medium 2 mg L-1 BAP 2 mg L-1) with addition of Kanamycin: 25, 50, 100, 150 and 200 (mg L-1); Hygromycin: 5, 10, 25, 50 and 75 (mg L-1); Paromomycin: 10, 25, 50, 75 and 100 (mg L-1). Experiment was arranged in a Completely Randomized Design (CRD). Each treatment was repeated five times thus 75 bottles of culture were used; each bottle consists of 5 pieces of leaf cuttings, resulted in total of 375 pieces. The results showed that selection agent had a critical value for Hygromycin 25 mg L-1 and Kanamycin 100 mg L-1 which can make explant experienced necrosis better than Paromomycin. Paromomycin at 100 mg L-1 was only able to kill explant’s periphery. Remained callus stayed fresh more than 50% so that when used as the selection agent could produce more escape cell. The optimum transformation with concentration of 10% Agrobacterium (vol/vol) with 30 minutes co-cultivation can produce more efficient transformed callus. Considering the high price of Hygromycin, it was best to use Kanamycin as selective agents.

  20. Efficient and rapid Agrobacterium-mediated genetic transformation of durum wheat (Triticum turgidum L. var. durum) using additional virulence genes.

    PubMed

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

    2008-06-01

    Genetic transformation of wheat, using biolistics or Agrobacterium, underpins a range of specific research methods for identifying genes and studying their function in planta. Transgenic approaches to study and modify traits in durum wheat have lagged behind those for bread wheat. Here we report the use of Agrobacterium strain AGL1, with additional vir genes housed in a helper plasmid, to transform and regenerate the durum wheat variety Ofanto. The use of the basic pSoup helper plasmid with no additional vir genes failed to generate transformants, whereas the presence of either virG542 or the 15 kb Komari fragment containing virB, virC and virG542 produced transformation efficiencies of between 0.6 and 9.7%. Of the 42 transgenic plants made, all but one (which set very few seeds) appeared morphologically normal and produced between 100 and 300 viable seeds. The transgene copy number and the segregation ratios were found to be very similar to those previously reported for bread wheat. We believe that this is the first report describing successful genetic transformation of tetraploid durum wheat (Triticum turgidum L. var. durum) mediated by Agrobacterium tumefaciens using immature embryos as the explant.

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

    USDA-ARS?s Scientific Manuscript database

    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...

  2. 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.

  3. 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.

  4. Early events in Agrobacterium-mediated genetic transformation of citrus explants.

    PubMed

    Peña, Leandro; Pérez, Rosa M; Cervera, Magdalena; Juárez, José A; Navarro, Luis

    2004-07-01

    Genetic transformation of plants relies on two independent but concurrent processes: integration of foreign DNA into plant cells and regeneration of whole plants from these transformed cells. Cell competence for regeneration and for transformation does not always fall into the same cell type/developmental stage, and this is one of the main causes of the so-called recalcitrance for transformation of certain plant species. In this study, a detailed examination of the first steps of morphogenesis from citrus explants after co-cultivation with Agrobacterium tumefaciens was performed, and an investigation into which cells and tissues are competent for regeneration and transformation was carried out. Moreover, the role of phytohormones in the co-cultivation medium as possible enhancers of gene transfer was also studied. A highly responsive citrus genotype and well-established culture conditions were used to perform a histological analysis of morphogenesis and cell competence for transformation after co-cultivation of citrus epicotyl segments with A. tumefaciens. In addition, the role of phytohormones as transformation enhancers was investigated by flow cytometry. It is demonstrated that cells competent for transformation are located in the newly formed callus growing from the cambial ring. Conditions conducive to further development of this callus, such as treatment of explants in a medium rich in auxins, resulted in a more pronounced formation of cambial callus and a slower shoot regeneration process, both in Agrobacterium-inoculated and non-inoculated explants. Furthermore, co- cultivation in a medium rich in auxins caused a significant increase in the rate of actively dividing cells in S-phase, the stage in which cells are more prone to integrate foreign DNA. Use of proper co-cultivation medium and conditions led to a higher number of stably transformed cells and to an increase in the final number of regenerated transgenic plants.

  5. Agrobacterium-mediated transformation of Easter lily (Lilium longiflorum cv. Nellie White)

    USDA-ARS?s Scientific Manuscript database

    Conditions were optimized for transient transformation of Lilium longiflorum cv. Nellie White using Agrobacterium tumefaciens. Bulb scale and basal meristem explants were inoculated with A. tumefaciens strain AGL1 containing the binary vector pCAMBIA 2301 which has the uidA gene that codes for ß-gl...

  6. 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

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

    Treesearch

    Micah E. Stevens; Paula M. Pijut

    2014-01-01

    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...

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

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Collier, Ray; Bragg, Jennifer; Hernandez, Bryan T.

    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

  9. 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.; ...

    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

  10. 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.

  11. Crown gall transformation of tobacco callus cells by cocultivation with Agrobacterium tumefaciens

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Muller, A.; Manzara, T.; Lurquin, P.F.

    1984-09-17

    Incubation of cells from squashed tobacco callus tissue with virulent Agrobacterium tumefaciens leads to the production of cells displaying a crown gall phenotype. In vitro crown gall transformation of dicotyledonous plant cells has been demonstrated after cocultivation of cell-wall regenerating mesophyll protoplasts with Agrobacterium tumefaciens cells. In addition, it has been shown that protoplasts freshly isolated from suspension cultures, when treated with A. tumefaciens spheroplasts and a fusogen, also generated cells displaying a typical crown gall phenotype, i.e., phytohormone-independent growth and opine synthesis. Subsequently, both techniques were used to transfer and express foreign genes in plant cells via A. tumefaciensmore » T-DNA integration. For practical purposes, it would be advantageous to be able to perform crown gall transformation of plant cells in tissue culture. The authors report here for the first time the production of Nicotiana tabacum crown gall cells after cocultivation of callus tissue with A. tumefaciens A136 cells. 11 references, 1 figure, 1 table.« less

  12. Genetic transformation of tobacco NT1 cells with Agrobacterium tumefaciens.

    PubMed

    Mayo, Kristin J; Gonzales, Barbara J; Mason, Hugh S

    2006-01-01

    This protocol is used to produce stably transformed tobacco (Nicotiana tabacum) NT1 cell lines, using Agrobacterium tumefaciens-mediated DNA delivery of a binary vector containing a gene encoding hepatitis B surface antigen and a gene encoding the kanamycin selection marker. The NT1 cultures, at the appropriate stage of growth, are inoculated with A. tumefaciens containing the binary vector. A 3-day cocultivation period follows, after which the cultures are rinsed and placed on solid selective medium. Transformed colonies ('calli') appear in approximately 4 weeks; they are subcultured until adequate material is obtained for analysis of antigen production. 'Elite' lines are selected based on antigen expression and growth characteristics. The time required for the procedure from preparation of the plant cell materials to callus development is approximately 5 weeks. Growth of selected calli to sufficient quantities for antigen screening may require 4-6 weeks beyond the initial selection. Creation of the plasmid constructs, transformation of the A. tumefaciens line, and ELISA and Bradford assays to assess protein production require additional time.

  13. 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

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

    USDA-ARS?s Scientific Manuscript database

    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...

  15. Agrobacterium tumefaciens-mediated transformation of the entomopathogenic fungus Nomuraea rileyi.

    PubMed

    Shao, Changwen; Yin, Youping; Qi, Zhaoran; Li, Ren; Song, Zhangyong; Li, Yan; Wang, Zhongkang

    2015-10-01

    An Agrobacterium-mediated genetic transformation system for the entomopathogenic fungus Nomuraea rileyi was established. Three binary T-DNA vectors, pPZP-Hph, pPZP-Hph-RNAi and pPZP-Hph-DsRed2, were constructed. The trpc promoter from Aspergillus nidulans was used as the cis-regulatory element to drive the expression of hygromycin phosphotransferase (hph) gene and DsRed2, which conferred the hygromycin B (Hyg B) resistance and red fluorescence visualization, respectively. The blastospores and conidia were used as the recipients. The blastospores' transformation efficiency reached ∼20-40 transformants per 10(6) blastospores, whereas the conidia were not transformed. Based on an analysis of five generations of subcultures, PCR and Southern blotting assays, the Ptrpc-hph cassette had integrated into the genomes of all transformants, which contained single copy of the hph gene and showed mitotic stability. Abundant altered morphologic phenotypes in colonies, blastospores and hyphae formations were observed in the arbitrary insertional mutants of N. rileyi, which made it possible to study the relationships between the functions and the interrupted genes over the whole genome. The transformation protocol will promote the functional characterization of genes, and the construction of genetically engineered strains of this important entomopathogenic fungus, and potentially of other similar fungal pathogens. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. 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.

  17. Methods for genetic transformation of filamentous fungi.

    PubMed

    Li, Dandan; Tang, Yu; Lin, Jun; Cai, Weiwen

    2017-10-03

    Filamentous fungi have been of great interest because of their excellent ability as cell factories to manufacture useful products for human beings. The development of genetic transformation techniques is a precondition that enables scientists to target and modify genes efficiently and may reveal the function of target genes. The method to deliver foreign nucleic acid into cells is the sticking point for fungal genome modification. Up to date, there are some general methods of genetic transformation for fungi, including protoplast-mediated transformation, Agrobacterium-mediated transformation, electroporation, biolistic method and shock-wave-mediated transformation. This article reviews basic protocols and principles of these transformation methods, as well as their advantages and disadvantages.

  18. Establishment of a simple and efficient Agrobacterium-mediated transformation system for Phytophthora palmivora.

    PubMed

    Wu, Dongliang; Navet, Natasha; Liu, Yingchao; Uchida, Janice; Tian, Miaoying

    2016-09-06

    As an agriculturally important oomycete genus, Phytophthora contains a large number of destructive plant pathogens that severely threaten agricultural production and natural ecosystems. Among them is the broad host range pathogen P. palmivora, which infects many economically important plant species. An essential way to dissect their pathogenesis mechanisms is genetic modification of candidate genes, which requires effective transformation systems. Four methods were developed for transformation of Phytophthora spp., including PEG(polyethylene glycol)/CaCl2 mediated protoplast transformation, electroporation of zoospores, microprojectile bombardment and Agrobacterium-mediated transformation (AMT). Among them, AMT has many advantages over the other methods such as easy handling and mainly generating single-copy integration in the genome. An AMT method previously reported for P. infestans and P. palmivora has barely been used in oomycete research due to low success and low reproducibility. In this study, we report a simple and efficient AMT system for P. palmivora. Using this system, we were able to reproducibly generate over 40 transformants using zoospores collected from culture grown in a single 100 mm-diameter petri dish. The generated GFP transformants constitutively expressed GFP readily detectable using a fluorescence microscope. All of the transformants tested using Southern blot analysis contained a single-copy T-DNA insertion. This system is highly effective and reproducible for transformation of P. palmivora and expected to be adaptable for transformation of additional Phytophthora spp. and other oomycetes. Its establishment will greatly accelerate their functional genomic studies.

  19. A really useful pathogen, Agrobacterium tumefaciens.

    PubMed

    Yuan, Ze-Chun; Williams, Mary

    2012-10-01

    Bacteria of the genus Agrobacterium are very useful and unusual plant pathogens. Through a rare inter-kingdom DNA transfer, the bacteria move some of their genes into their host's genome, thereby inducing the host cells to proliferate and produce opines, nutrients sources for the pathogen. Agrobacterium's ability to transfer DNA makes can be adapted to introduce other genes, such as those encoding useful traits, into plant genomes. The development of Agrobacterium as a tool to transform plants is a landmark event in modern plant biology. This lecture provides an introduction to Agrobacterium tumefaciens and related species, focusing on their modes of pathogenicity, their usefulness as tools for plant transformation, and their use as a model for the study of plant-pathogen interactions.

  20. Natural Transformation of Pseudomonas fluorescens and Agrobacterium tumefaciens in Soil

    PubMed Central

    Demanèche, Sandrine; Kay, Elisabeth; Gourbière, François; Simonet, Pascal

    2001-01-01

    Little information is available concerning the occurrence of natural transformation of bacteria in soil, the frequency of such events, and the actual role of this process on bacterial evolution. This is because few bacteria are known to possess the genes required to develop competence and because the tested bacteria are unable to reach this physiological state in situ. In this study we found that two soil bacteria, Agrobacterium tumefaciens and Pseudomonas fluorescens, can undergo transformation in soil microcosms without any specific physical or chemical treatment. Moreover, P. fluorescens produced transformants in both sterile and nonsterile soil microcosms but failed to do so in the various in vitro conditions we tested. A. tumefaciens could be transformed in vitro and in sterile soil samples. These results indicate that the number of transformable bacteria could be higher than previously thought and that these bacteria could find the conditions necessary for uptake of extracellular DNA in soil. PMID:11375171

  1. 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.

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

    PubMed

    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.

  3. An Improved Single-Step Cloning Strategy Simplifies the Agrobacterium tumefaciens-Mediated Transformation (ATMT)-Based Gene-Disruption Method for Verticillium dahliae.

    PubMed

    Wang, Sheng; Xing, Haiying; Hua, Chenlei; Guo, Hui-Shan; Zhang, Jie

    2016-06-01

    The soilborne fungal pathogen Verticillium dahliae infects a broad range of plant species to cause severe diseases. The availability of Verticillium genome sequences has provided opportunities for large-scale investigations of individual gene function in Verticillium strains using Agrobacterium tumefaciens-mediated transformation (ATMT)-based gene-disruption strategies. Traditional ATMT vectors require multiple cloning steps and elaborate characterization procedures to achieve successful gene replacement; thus, these vectors are not suitable for high-throughput ATMT-based gene deletion. Several advancements have been made that either involve simplification of the steps required for gene-deletion vector construction or increase the efficiency of the technique for rapid recombinant characterization. However, an ATMT binary vector that is both simple and efficient is still lacking. Here, we generated a USER-ATMT dual-selection (DS) binary vector, which combines both the advantages of the USER single-step cloning technique and the efficiency of the herpes simplex virus thymidine kinase negative-selection marker. Highly efficient deletion of three different genes in V. dahliae using the USER-ATMT-DS vector enabled verification that this newly-generated vector not only facilitates the cloning process but also simplifies the subsequent identification of fungal homologous recombinants. The results suggest that the USER-ATMT-DS vector is applicable for efficient gene deletion and suitable for large-scale gene deletion in V. dahliae.

  4. Agrobacterium tumefaciens-mediated transformation of Campanula carpatica: factors affecting transformation and regeneration of transgenic shoots.

    PubMed

    Sriskandarajah, Sridevy; Frello, Stefan; Jørgensen, Kirsten; Serek, Margrethe

    2004-08-01

    An efficient transformation system for Campanula carpatica was developed using Agrobacterium tumefaciens strains LBA4404 (harbouring the plasmid pBI121), and AGL0 (harbouring the plasmid pBEO210). This is the first report on the transformation of C. carpatica. Various factors affecting the transformation efficiency and subsequent regeneration were identified. The age of seedlings from which the explants for transformation studies were taken, and the growth conditions under which the seedlings were grown had a significant influence on the production of transformed shoots. Hypocotyls taken from 12-day-old seedlings grown in the dark were the most productive, with up to 25% of hypocotyls producing transformed shoots. Explants taken from 5-week-old seedlings produced only transformed callus. The medium used for co-cultivation and incubation also had a significant influence on transformation frequency and shoot regeneration. The cultivar "Blue Uniform" was more responsive than "White Uniform". Both bacterial strains and plasmids were equally effective in producing transformed tissue. Transformed shoots were selected on kanamycin medium, and the presence of the uidA and nptII genes in those selected shoots was confirmed by beta-glucuronidase and ELISA analyses, respectively.

  5. 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.

  6. Wheat (Triticum aestivum L.) transformation using mature embryos.

    PubMed

    Medvecká, Eva; Harwood, Wendy A

    2015-01-01

    In most protocols for the Agrobacterium-mediated transformation of wheat, the preferred target tissues are immature embryos. However, transformation methods relying on immature embryos require the growth of plants under controlled conditions to provide a continuous supply of good-quality target tissue. The use of mature embryos as a target tissue has the advantage of only requiring good-quality seed as the starting material. Here we describe a transformation method based on the Agrobacterium-mediated transformation of callus cultures derived from mature wheat embryos of the genotype Bobwhite S56.

  7. 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.

  8. Efficient Agrobacterium-Mediated Transformation of Hybrid Poplar Populus davidiana Dode × Populus bollena Lauche

    PubMed Central

    Han, Xue; Ma, Shurong; Kong, Xianghui; Takano, Tetsuo; Liu, Shenkui

    2013-01-01

    Poplar is a model organism for high in vitro regeneration in woody plants. We have chosen a hybrid poplar Populus davidiana Dode × Populus bollena Lauche. By optimizing the Murashige and Skoog medium with (0.3 mg/L) 6-benzylaminopurine and (0.08 mg/L) naphthaleneacetic acid, we have achieved the highest frequency (90%) for shoot regeneration from poplar leaves. It was also important to improve the transformation efficiency of poplar for genetic breeding and other applications. In this study, we found a significant improvement of the transformation frequency by controlling the leaf age. Transformation efficiency was enhanced by optimizing the Agrobacterium concentration (OD600 = 0.8–1.0) and an infection time (20–30 min). According to transmission electron microscopy observations, there were more Agrobacterium invasions in the 30-day-old leaf explants than in 60-day-old and 90-day-old explants. Using the green fluorescent protein (GFP) marker, the expression of MD–GFP fusion proteins in the leaf, shoot, and root of hybrid poplar P. davidiana Dode × P. bollena Lauche was visualized for confirmation of transgene integration. Southern and Northern blot analysis also showed the integration of T-DNA into the genome and gene expression of transgenic plants. Our results suggest that younger leaves had higher transformation efficiency (~30%) than older leaves (10%). PMID:23354481

  9. Genetic transformation of Fusarium oxysporum f.sp. gladioli with Agrobacterium to study pathogenesis in Gladiolus

    USDA-ARS?s Scientific Manuscript database

    Fusarium rot caused by Fusarium oxysporum f.sp. gladioli (Fog) is one of the most serious diseases of Gladiolus, both in the field and in stored bulbs. In order to study the pathogenesis of this fungus, we have transformed Fog with Agrobacterium tumefaciens binary vectors containing the hygromycin B...

  10. A comparison of Agrobacterium-mediated transformation and protoplast-mediated transformation with CRISPR-Cas9 and bipartite gene targeting substrates, as effective gene targeting tools for Aspergillus carbonarius.

    PubMed

    Weyda, István; Yang, Lei; Vang, Jesper; Ahring, Birgitte K; Lübeck, Mette; Lübeck, Peter S

    2017-04-01

    In recent years, versatile genetic tools have been developed and applied to a number of filamentous fungi of industrial importance. However, the existing techniques have limitations when it comes to achieve the desired genetic modifications, especially for efficient gene targeting. In this study, we used Aspergillus carbonarius as a host strain due to its potential as a cell factory, and compared three gene targeting techniques by disrupting the ayg1 gene involved in the biosynthesis of conidial pigment in A. carbonarius. The absence of the ayg1 gene leads to phenotypic change in conidia color, which facilitated the analysis on the gene targeting frequency. The examined transformation techniques included Agrobacterium-mediated transformation (AMT) and protoplast-mediated transformation (PMT). Furthermore, the PMT for the disruption of the ayg1 gene was carried out with bipartite gene targeting fragments and the recently adapted CRISPR-Cas9 system. All three techniques were successful in generating Δayg1 mutants, but showed different efficiencies. The most efficient method for gene targeting was AMT, but further it was shown to be dependent on the choice of Agrobacterium strain. However, there are different advantages and disadvantages of all three gene targeting methods which are discussed, in order to facilitate future approaches for fungal strain improvements. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. Selection system and co-cultivation medium are important determinants of Agrobacterium-mediated transformation of sugarcane.

    PubMed

    Joyce, Priya; Kuwahata, Melissa; Turner, Nicole; Lakshmanan, Prakash

    2010-02-01

    A reproducible method for transformation of sugarcane using various strains of Agrobacterium tumefaciens (A. tumefaciens) (AGL0, AGL1, EHA105 and LBA4404) has been developed. The selection system and co-cultivation medium were the most important factors determining the success of transformation and transgenic plant regeneration. Plant regeneration at a frequency of 0.8-4.8% occurred only when callus was transformed with A. tumefaciens carrying a newly constructed superbinary plasmid containing neomycin phosphotransferase (nptII) and beta-glucuronidase (gusA) genes, both driven by the maize ubiquitin (ubi-1) promoter. Regeneration was successful in plants carrying the nptII gene but not the hygromycin phosphotransferase (hph) gene. NptII gene selection was imposed at a concentration of 150 mg/l paromomycin sulphate and applied either immediately or 4 days after the co-cultivation period. Co-cultivation on Murashige and Skoog (MS)-based medium for a period of 4 days produced the highest number of transgenic plants. Over 200 independent transgenic lines were created using this protocol. Regenerated plants appeared phenotypically normal and contained both gusA and nptII genes. Southern blot analysis revealed 1-3 transgene insertion events that were randomly integrated in the majority of the plants produced.

  12. Highly efficient transformation system for Malassezia furfur and Malassezia pachydermatis using Agrobacterium tumefaciens-mediated transformation.

    PubMed

    Celis, A M; Vos, A M; Triana, S; Medina, C A; Escobar, N; Restrepo, S; Wösten, H A B; de Cock, H

    2017-03-01

    Malassezia spp. are part of the normal human and animal mycobiota but are also associated with a variety of dermatological diseases. The absence of a transformation system hampered studies to reveal mechanisms underlying the switch from the non-pathogenic to pathogenic life style. Here we describe, a highly efficient Agrobacterium-mediated genetic transformation system for Malassezia furfur and M. pachydermatis. A binary T-DNA vector with the hygromycin B phosphotransferase (hpt) selection marker and the green fluorescent protein gene (gfp) was introduced in M. furfur and M. pachydermatis by combining the transformation protocols of Agaricus bisporus and Cryptococcus neoformans. Optimal temperature and co-cultivation time for transformation were 5 and 7days at 19°C and 24°C, respectively. Transformation efficiency was 0.75-1.5% for M. furfur and 0.6-7.5% for M. pachydermatis. Integration of the hpt resistance cassette and gfp was verified using PCR and fluorescence microscopy, respectively. The T-DNA was mitotically stable in approximately 80% of the transformants after 10 times sub-culturing in the absence of hygromycin. Improving transformation protocols contribute to study the biology and pathophysiology of Malassezia. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  13. An efficient and high-throughput protocol for Agrobacterium-mediated transformation based on phosphomannose isomerase positive selection in Japonica rice (Oryza sativa L.).

    PubMed

    Duan, Yongbo; Zhai, Chenguang; Li, Hao; Li, Juan; Mei, Wenqian; Gui, Huaping; Ni, Dahu; Song, Fengshun; Li, Li; Zhang, Wanggen; Yang, Jianbo

    2012-09-01

    A number of Agrobacterium-mediated rice transformation systems have been developed and widely used in numerous laboratories and research institutes. However, those systems generally employ antibiotics like kanamycin and hygromycin, or herbicide as selectable agents, and are used for the small-scale experiments. To address high-throughput production of transgenic rice plants via Agrobacterium-mediated transformation, and to eliminate public concern on antibiotic markers, we developed a comprehensive efficient protocol, covering from explant preparation to the acquisition of low copy events by real-time PCR analysis before transplant to field, for high-throughput production of transgenic plants of Japonica rice varieties Wanjing97 and Nipponbare using Escherichia coli phosphomannose isomerase gene (pmi) as a selectable marker. The transformation frequencies (TF) of Wanjing97 and Nipponbare were achieved as high as 54.8 and 47.5%, respectively, in one round of selection of 7.5 or 12.5 g/L mannose appended with 5 g/L sucrose. High-throughput transformation from inoculation to transplant of low copy events was accomplished within 55-60 days. Moreover, the Taqman assay data from a large number of transformants showed 45.2% in Wanjing97 and 31.5% in Nipponbare as a low copy rate, and the transformants are fertile and follow the Mendelian segregation ratio. This protocol facilitates us to perform genome-wide functional annotation of the open reading frames and utilization of the agronomically important genes in rice under a reduced public concern on selectable markers. We describe a comprehensive protocol for large scale production of transgenic Japonica rice plants using non-antibiotic selectable agent, at simplified, cost- and labor-saving manners.

  14. Agrobacterium tumefaciens-mediated transformation of Lasiodiplodia theobromae, the causal agent of gummosis in cashew nut plants.

    PubMed

    Muniz, C R; da Silva, G F; Souza, M T; Freire, F C O; Kema, G H J; Guedes, M I F

    2014-02-21

    Lasiodiplodia theobromae is a major pathogen of many different crop cultures, including cashew nut plants. This paper describes an efficient Agrobacterium tumefaciens-mediated transformation (ATMT) system for the successful delivery of T-DNA, transferring the genes of green fluorescent protein (gfp) and hygromycin B phosphotransferase (hph) to L. theobromae. When the fungal pycnidiospores were co-cultured with A. tumefaciens harboring the binary vector with hph-gfp gene, hygromycin-resistant fungus only developed with acetosyringone supplementation. The cashew plants inoculated with the fungus expressing GFP revealed characteristic pathogen colonization by epifluorescence microscopy. Intense and bright green hyphae were observed for transformants in all extensions of mycelium cultures. The penetration of parenchyma cells near to the inoculation site, beneath the epicuticle surface, was observed prior to 25 dpi. Penetration was followed by the development of hyphae within invaded host cells. These findings provide a rapid and reproducible ATMT method for L. theobromae transformation.

  15. 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.

  16. Agrobacterium tumefaciens-mediated transformation for investigating pathogenicity genes of the phytopathogenic fungus Colletotrichum sansevieriae.

    PubMed

    Nakamura, Masayuki; Kuwahara, Hideto; Onoyama, Keisuke; Iwai, Hisashi

    2012-08-01

    Agrobacterium tumefaciens-mediated transformation (AtMT) has become a common technique for DNA transformation of yeast and filamentous fungi. In this study, we first established a protocol of AtMT for the phytopathogenic fungus Colletotrichum sansevieriae. Binary T-DNA vector containing the hygromycin B phosphotransferase gene controlled by the Aspergillus nidulans gpdA promoter and the trpC terminator was constructed with pCAMBIA0380 and used with three different strains LBA4404, GV3101, and GV2260 of A. tumefaciens. Transformants were most effectively obtained when GV2260 and C. sansevieriae Sa-1-2 were co-cultivated; there were about 320 transformants per 10(6) spores. When 1,048 transformants were inoculated on Sansevieria trifasciata, three transformants were found to have completely lost their pathogenicity and two transformants displayed reduced pathogenicity. All of the five transformants had a single copy of T-DNA in their genomes. The three pathogenicity-deficient transformants were subjected to thermal asymmetric interlaced polymerase chain reaction and the reaction allowed us to amplify the sequences flanking the left and/or right borders. The flanking sequences of the two transformants, M154 and M875, showed no homology to any sequences in databases, but the sequences of M678 contained motifs of alpha-1,3-glucan synthase, suggesting that the gene might contribute to the pathogenicity of C. sansevieriae. This study describes a useful method for investigating pathogenicity genes in C. sansevieriae.

  17. Hairy root cultures of butterfly pea (Clitoria ternatea L.): Agrobacterium × plant factors influencing transformation.

    PubMed

    Swain, S S; Sahu, L; Pal, A; Barik, D P; Pradhan, C; Chand, P K

    2012-02-01

    Transformed rhizoclones were developed from Agrobacterium-treated explants of the medicinally important twinning legume Clitoria ternatea L. Several key factors influencing transformation events were optimized. A4T was the most infectious among the strains employed. Internode segments were more responsive than leaves, outdoor-grown explants preferred to those from in vitro cultures. High frequency transformation, resulting in up to 85.8% rhizogenesis, was attained using pre-pricked internodal explants for immersion (10 min) in Agrobacterium rhizogenes suspension grown overnight with acetosyringone (100 μM) to an OD(660) ≅ 0.6, diluted to a density of 10(9) cells ml(-1), followed by 5-day co-cultivation. Roots were individually cultured in MS0 supplemented with the bacteriostatic antibiotic cefotaxime (500 μg ml(-1)). Rhizoclones were renewed through successive subcultures in MS0 under diffused illumination. The T ( L )-DNA rolB and rolC ORF were detected in rhizoclones through PCR amplification. The T ( R )-DNA gene encoding mannopine synthase (man2) was revealed by positive amplification and opine gene expression substantiated by agropine and mannopine biosynthesis in all selected transformed rhizoclones. The implication of such findings is discussed on the context of utilization of such genetically transformed root cultures towards sustainable production of medicinally useful phytocompounds, besides providing a means for plant conservation.

  18. 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.

  19. Improved production of transgenic Dioscorea zingiberensis (Dioscoreaceae) by Agrobacterium tumefaciens-mediated transformation.

    PubMed

    Shi, L; Fan, J Q; Hu, C G; Luo, J; Yao, J L

    2012-02-03

    The establishment of high-efficiency Agrobacterium-mediated transformation techniques could improve the production of Dioscorea zingiberensis, a medicinal species with a high diosgenin content. We co-cultivated embryogenic calli induced from mature seeds with A. tumefaciens strain EHA105. A binary vector, pCAMBIA1381, which contains the gfp and hpt genes under the control of the ubiquitin promoter and the CaMV 35S promoter, respectively, was used for transformation. Pre-culture, basic medium, acetosyringone, and bacterial density were evaluated to establish the most efficient protocol. The optimal conditions consisted of MS medium without CaCl(2) for pre- and co-cultivation, three days for pre-culture, addition of 200 μM AS, and an OD(600) of 0.5. The transgenic plants grown under selection were confirmed by PCR analysis and Southern blot analysis. This protocol produced transgenic D. zingiberensis plants in seven months, with a transformation efficiency of 6%.

  20. A Genetic Transformation Method for Cadmium Hyperaccumulator Sedum plumbizincicola and Non-hyperaccumulating Ecotype of Sedum alfredii

    PubMed Central

    Liu, Huan; Zhao, Haixia; Wu, Longhua; Xu, Wenzhong

    2017-01-01

    The present study demonstrates the development of an Agrobacterium-mediated genetic transformation method for species of the Sedum genus, which includes the Cd/Zn hyperaccumulator Sedum plumbizincicola and the non-hyperaccumulating ecotype of S. alfredii. Multiple shoots were induced from stem nodes of two Sedum plants using Murashige and Skoog (MS) medium containing 0.1 mg/L cytokinin 6-benzyladenine (6-BA) and 1.0 mg/L auxin 1-naphthaleneacetic acid (NAA). The shoot primordia were used as direct targets for Agrobacterium infection. Selection on hygromycin was highly effective in generating Agrobacterium-transformed explants. This callus-free procedure allowed us to obtain transgenic plantlets after rooting hygromycin-resistant shoots on phytohormone-free MS medium containing the antibiotic. The presence and expression of the reporter genes gusA and GFP in transgenic plants were confirmed by a real-time polymerase chain reaction, histochemical GUS assays, and confocal microscopy. This reliable method for genetic transformation of Sedum plants will help us to understand gene functions and the molecular mechanisms underlying Cd hypertolerance and hyperaccumulation in these species. PMID:28670322

  1. An improved method for Agrobacterium rhizogenes-mediated transformation of tomato suitable for the study of arbuscular mycorrhizal symbiosis.

    PubMed

    Ho-Plágaro, Tania; Huertas, Raúl; Tamayo-Navarrete, María I; Ocampo, Juan A; García-Garrido, José M

    2018-01-01

    Solanum lycopersicum , an economically important crop grown worldwide, has been used as a model for the study of arbuscular mycorrhizal (AM) symbiosis in non-legume plants for several years and several cDNA array hybridization studies have revealed specific transcriptomic profiles of mycorrhizal tomato roots. However, a method to easily screen candidate genes which could play an important role during tomato mycorrhization is required. We have developed an optimized procedure for composite tomato plant obtaining achieved through Agrobacterium rhizogenes -mediated transformation. This protocol involves the unusual in vitro culture of composite plants between two filter papers placed on the culture media. In addition, we show that DsRed is an appropriate molecular marker for the precise selection of cotransformed tomato hairy roots . S. lycopersicum composite plant hairy roots appear to be colonized by the AM fungus Rhizophagus irregularis in a manner similar to that of normal roots, and a modified construct useful for localizing the expression of promoters putatively associated with mycorrhization was developed and tested. In this study, we present an easy, fast and low-cost procedure to study AM symbiosis in tomato roots.

  2. High-frequency transformation of Lobelia erinus L. by Agrobacterium-mediated gene transfer.

    PubMed

    Tsugawa, H; Kagami, T; Suzuki, M

    2004-05-01

    A highly efficient transformation procedure was developed for Lobelia erinus. Leaf or cotyledon discs were inoculated with Agrobacterium tumefaciens strain EHA105 harboring the binary vector plasmid pIG121Hm, which contains a beta-glucuronidase gene with an intron as a reporter gene and both the neomycin phosphotransferase II and hygromycin phosphotransferase genes as selectable markers. The hygromycin-resistant calli produced on the selection medium were transferred to MS medium supplemented with 0.5 mg/l benzyladenine and 0.2 mg/l indole-3-acetic acid for regeneration of adventitious shoots. Transgenic plants were obtained as a result of the high regeneration rate of the transformed calli, which was as high as 83%. In contrast, no transgenic plant was obtained by the procedure of direct shoot formation following inoculation with A. tumefaciens. Transgenic plants flowered 3-4 months after transformation. Integration of the transgenes was detected using PCR and Southern blot analysis, which revealed that one to several copies were integrated into the genomes of the host plants. The transformation frequency at the stage of whole plants was very high--45% per inoculated disc. Copyright 2004 Springer-Verlag

  3. Transformation of Morinda citrifolia via simple mature seed imbibition method.

    PubMed

    Lee, J J; Ahmad, S; Roslan, H A

    2013-12-15

    Morinda citrifolia, is a valuable medicinal plant with a wide range of therapeutic properties and extensive transformation study on this plant has yet been known. Present study was conducted to establish a simple and reliable transformation protocol for M. citrifolia utilising Agrobacterium tumefaciens via direct seed exposure. In this study, the seeds were processed by tips clipping and dried and subsequently incubated in inoculation medium. Four different parameters during the incubation such as incubation period, bacterial density, temperature and binary vectors harbouring beta-glucuronidase (GUS) gene (pBI121 and pGSA1131), were tested to examine its effect on transformation efficiency. The leaves from the treated and germinated seedlings were analysed via Polymerase Chain Reaction (PCR), histochemical assay of the GUS gene and reverse transcription-PCR (RT-PCR). Results of the study showed that Agrobacterium strain LBA4404 with optical density of 1.0 and 2 h incubation period were optimum for M. citrifolia transformation. It was found that various co-cultivation temperatures tested and type of vector used did not affect the transformation efficiency. The highest transformation efficiency for M. citrifolia direct seed transformation harbouring pBI121 and pGSA1131 was determined to be 96.8% with 2 h co-cultivation treatment and 80.4% when using bacterial density of 1.0, respectively. The transformation method can be applied for future characterization study of M. citrifolia.

  4. A new and efficient approach for construction of uridine/uracil auxotrophic mutants in the filamentous fungus Aspergillus oryzae using Agrobacterium tumefaciens-mediated transformation.

    PubMed

    Nguyen, Khuyen Thi; Ho, Quynh Ngoc; Do, Loc Thi Binh Xuan; Mai, Linh Thi Dam; Pham, Duc-Ngoc; Tran, Huyen Thi Thanh; Le, Diep Hong; Nguyen, Huy Quang; Tran, Van-Tuan

    2017-06-01

    Aspergillus oryzae is a filamentous fungus widely used in food industry and as a microbial cell factory for recombinant protein production. Due to the inherent resistance of A. oryzae to common antifungal compounds, genetic transformation of this mold usually requires auxotrophic mutants. In this study, we show that Agrobacterium tumefaciens-mediated transformation (ATMT) method is very efficient for deletion of the pyrG gene in different Aspergillus oryzae wild-type strains to generate uridine/uracil auxotrophic mutants. Our data indicated that all the obtained uridine/uracil auxotrophic transformants, which are 5- fluoroorotic acid (5-FOA) resistant, exist as the pyrG deletion mutants. Using these auxotrophic mutants and the pyrG selectable marker for genetic transformation via A. tumefaciens, we could get about 1060 transformants per 10 6 fungal spores. In addition, these A. oryzae mutants were also used successfully for expression of the DsRed fluorescent reporter gene under control of the A. oryzae amyB promoter by the ATMT method, which resulted in obvious red transformants on agar plates. Our work provides a new and effective approach for constructing the uridine/uracil auxotrophic mutants in the importantly industrial fungus A. oryzae. This strategy appears to be applicable to other filamentous fungi to develop similar genetic transformation systems based on auxotrophic/nutritional markers for food-grade recombinant applications.

  5. 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

  6. Development of an Efficient Agrobacterium-Mediated Transformation System and Production of Herbicide-Resistant Transgenic Plants in Garlic (Allium sativum L.)

    PubMed Central

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

    2013-01-01

    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 non-transgenic plants and weeds died. These results indicate that our transformation system can be efficiently utilized to produce transgenic garlic plants with agronomic benefits. PMID:23832764

  7. 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.

  8. Organogenesis from transformed tomato explants.

    PubMed

    Frary, Anne; Van Eck, Joyce

    2005-01-01

    Tomato was one of the first crops for which a genetic transformation system was reported involving regeneration by organogenesis from Agrobacterium-transformed explants. Since the initial reports, various factors have been studied that affect the efficiency of tomato transformation and the technique has been useful for the isolation and identification of many genes involved in plant disease resistance, morphology and development. In this method, cotyledon explants from in vitro-grown seedlings are precultured overnight on a tobacco suspension feeder layer. The explants are then inoculated with Agrobacterium and returned to the feeder layer for a 2-d period of cocultivation. After cocultivation, the explants are transferred to an MS-based selective regeneration medium containing zeatin. Regenerated shoots are then rooted on a separate selective medium. This protocol has been used with several tomato cultivars and routinely yields transformation efficiencies of 10-15%.

  9. VIP1 and Its Homologs Are Not Required for Agrobacterium-Mediated Transformation, but Play a Role in Botrytis and Salt Stress Responses

    PubMed Central

    Lapham, Rachelle; Lee, Lan-Ying; Tsugama, Daisuke; Lee, Sanghun; Mengiste, Tesfaye; Gelvin, Stanton B.

    2018-01-01

    The bZIP transcription factor VIP1 interacts with the Agrobacterium virulence protein VirE2, but the role of VIP1 in Agrobacterium-mediated transformation remains controversial. Previously tested vip1-1 mutant plants produce a truncated protein containing the crucial bZIP DNA-binding domain. We generated the CRISPR/Cas mutant vip1-2 that lacks this domain. The transformation susceptibility of vip1-2 and wild-type plants is similar. Because of potential functional redundancy among VIP1 homologs, we tested transgenic lines expressing VIP1 fused to a SRDX repression domain. All VIP1-SRDX transgenic lines showed wild-type levels of transformation, indicating that neither VIP1 nor its homologs are required for Agrobacterium-mediated transformation. Because VIP1 is involved in innate immune response signaling, we tested the susceptibility of vip1 mutant and VIP1-SRDX plants to Pseudomonas syringae and Botrytis cinerea. vip1 mutant and VIP1-SRDX plants show increased susceptibility to B. cinerea but not to P. syringae infection, suggesting a role for VIP1 in B. cinerea, but not in P. syringae, defense signaling. B. cinerea susceptibility is dependent on abscisic acid (ABA) which is also important for abiotic stress responses. The germination of vip1 mutant and VIP1-SRDX seeds is sensitive to exogenous ABA, suggesting a role for VIP1 in response to ABA. vip1 mutant and VIP1-SRDX plants show increased tolerance to growth in salt, indicating a role for VIP1 in response to salt stress. PMID:29946325

  10. An efficient in planta transformation of Jatropha curcas (L.) and multiplication of transformed plants through in vivo grafting.

    PubMed

    Jaganath, Balusamy; Subramanyam, Kondeti; Mayavan, Subramanian; Karthik, Sivabalan; Elayaraja, Dhandapani; Udayakumar, Rajangam; Manickavasagam, Markandan; Ganapathi, Andy

    2014-05-01

    An efficient and reproducible Agrobacterium-mediated in planta transformation was developed in Jatropha curcas. The various factors affecting J. curcas in planta transformation were optimized, including decapitation, Agrobacterium strain, pin-pricking, vacuum infiltration duration and vacuum pressure. Simple vegetative in vivo cleft grafting method was adopted in the multiplication of transformants without the aid of tissue culture. Among the various parameters evaluated, decapitated plants on pin-pricking and vacuum infiltrated at 250 mmHg for 3 min with the Agrobacterium strain EHA 105 harbouring the binary vector pGA 492 was proved to be efficient in all terms with a transformation efficiency of 62.66%. Transgene integration was evinced by the GUS histochemical analysis, and the GUS positive plants were subjected to grafting. Putatively transformed J. curcas served as "Scion" and the wild type J. curcas plant severed as "Stock". There was no occurrence of graft rejection and the plants were then confirmed by GUS histochemical analysis, polymerase chain reaction (PCR) and Southern hybridization. Genetic stability of the grafted plants was evaluated by using randomly amplified polymorphic DNA (RAPD), marker which showed 100% genetic stability between mother and grafted plants. Thus, an efficient in planta transformation and grafting based multiplication of J. curcas was established.

  11. Evaluation of parameters affecting switchgrass tissue culture: toward a consolidated procedure for Agrobacterium-mediated transformation of switchgrass (Panicum virgatum)

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lin, Chien-Yuan; Donohoe, Bryon S.; Ahuja, Neha

    Switchgrass (Panicum virgatum), a robust perennial C4-type grass, has been evaluated and designated as a model bioenergy crop by the U.S. DOE and USDA. Conventional breeding of switchgrass biomass is difficult because it displays self-incompatible hindrance. Therefore, direct genetic modifications of switchgrass have been considered the more effective approach to tailor switchgrass with traits of interest. Successful transformations have demonstrated increased biomass yields, reduction in the recalcitrance of cell walls and enhanced saccharification efficiency. Several tissue culture protocols have been previously described to produce transgenic switchgrass lines using different nutrient-based media, co-cultivation approaches, and antibiotic strengths for selection. After evaluatingmore » the published protocols, we consolidated these approaches and optimized the process to develop a more efficient protocol for producing transgenic switchgrass. First, seed sterilization was optimized, which led to a 20% increase in yield of induced calluses. Second, we have selected a N 6 macronutrient/B 5 micronutrient (NB)-based medium for callus induction from mature seeds of the Alamo cultivar, and chose a Murashige and Skoog-based medium to regenerate both Type I and Type II calluses. Third, Agrobacterium-mediated transformation was adopted that resulted in 50-100% positive regenerated transformants after three rounds (2 weeks/round) of selection with antibiotic. Genomic DNA PCR, RT-PCR, Southern blot, visualization of the red fluorescent protein and histochemical β-glucuronidase (GUS) staining were conducted to confirm the positive switchgrass transformants. The optimized methods developed here provide an improved strategy to promote the production and selection of callus and generation of transgenic switchgrass lines. The process for switchgrass transformation has been evaluated and consolidated to devise an improved approach for transgenic switchgrass production. With

  12. Evaluation of parameters affecting switchgrass tissue culture: toward a consolidated procedure for Agrobacterium-mediated transformation of switchgrass (Panicum virgatum)

    DOE PAGES

    Lin, Chien-Yuan; Donohoe, Bryon S.; Ahuja, Neha; ...

    2017-12-19

    Switchgrass (Panicum virgatum), a robust perennial C4-type grass, has been evaluated and designated as a model bioenergy crop by the U.S. DOE and USDA. Conventional breeding of switchgrass biomass is difficult because it displays self-incompatible hindrance. Therefore, direct genetic modifications of switchgrass have been considered the more effective approach to tailor switchgrass with traits of interest. Successful transformations have demonstrated increased biomass yields, reduction in the recalcitrance of cell walls and enhanced saccharification efficiency. Several tissue culture protocols have been previously described to produce transgenic switchgrass lines using different nutrient-based media, co-cultivation approaches, and antibiotic strengths for selection. After evaluatingmore » the published protocols, we consolidated these approaches and optimized the process to develop a more efficient protocol for producing transgenic switchgrass. First, seed sterilization was optimized, which led to a 20% increase in yield of induced calluses. Second, we have selected a N 6 macronutrient/B 5 micronutrient (NB)-based medium for callus induction from mature seeds of the Alamo cultivar, and chose a Murashige and Skoog-based medium to regenerate both Type I and Type II calluses. Third, Agrobacterium-mediated transformation was adopted that resulted in 50-100% positive regenerated transformants after three rounds (2 weeks/round) of selection with antibiotic. Genomic DNA PCR, RT-PCR, Southern blot, visualization of the red fluorescent protein and histochemical β-glucuronidase (GUS) staining were conducted to confirm the positive switchgrass transformants. The optimized methods developed here provide an improved strategy to promote the production and selection of callus and generation of transgenic switchgrass lines. The process for switchgrass transformation has been evaluated and consolidated to devise an improved approach for transgenic switchgrass production. With

  13. Agrobacterium rhizogenes-mediated DNA transfer to Aesculus hippocastanum L. and the regeneration of transformed plants.

    PubMed

    Zdravković-Korać, S; Muhovski, Y; Druart, P; Calić, D; Radojević, L

    2004-04-01

    Hairy roots were induced from androgenic embryos of horse chestnut (Aesculus hippocastanum L.) by infection with Agrobacterium rhizogenes strain A4GUS. Single roots were selected according to their morphology in the absence of antibiotic or herbicide resistance markers. Seventy-one putative transformed hairy root lines from independent transformation events were established. Regeneration was induced in MS liquid medium supplemented with 30 microM 6-benzylaminopurine (BA), and the regenerants were multiplied on MS solid medium containing 10 microM BA. Following elongation on MS medium supplemented with 1 microM BA and 500 mg/l polyvinylpyrrolidone, the shoots were subjected to a root-inducing treatment. Stable integration of TL-DNA within the horse chestnut genome was confirmed by Southern hybridization. The copy number of transgenes was estimated to be from two to four.

  14. Factors enhancing Agrobacterium tumefaciens-mediated gene transfer in peanut (Arachis hypogaea L.)

    NASA Technical Reports Server (NTRS)

    Egnin, M.; Mora, A.; Prakash, C. S.; Mortley, D. G. (Principal Investigator)

    1998-01-01

    Parameters enhancing Agrobacterium-mediated transfer of foreign genes to peanut (Arachis hypogaea L.) cells were investigated. An intron-containing beta-glucuronidase uidA (gusA) gene under the transcriptional control of CaMV 35S promoter served as a reporter. Transformation frequency was evaluated by scoring the number of sectors expressing GUS activity on leaf and epicotyl explants. The 'Valencia Select' market type cv. New Mexico was more amenable to Agrobacterium transformation than the 'runner' market type cultivars tested (Florunner, Georgia Runner, Sunrunner, or South Runner). The disarmed Agrobacterium tumefaciens strain EHA101 was superior in facilitating the transfer of uidA gene to peanut cells compared to the disarmed strain C58. Rinsing of explants in half-strength Murashige-Skoog (MS) media prior to infection by Agrobacterium significantly increased the transformation efficiency. The use of cocultivation media containing high auxin [1.0 or 2.5 mg/l (4.53 micromolar or 11.31 micromolar) 2,4-D] and low cytokinin [0.25 or 0.5 mg/l (1.0 micromolar or 2.0 micromolar) BA] promoted higher transformation than either hormone-free or thidiazuron-containing medium. The polarity of the epicotyl during cocultivation was important; explants incubated in an inverted (vertically) manner followed by a vertically upright position resulted in improved transformation and shoot regeneration frequencies. Preculture of explants in MS basal medium or with 2.5 mg thidiazuron per l prior to infection drastically decreased the number of transformed zones. The optimized protocol was used to obtain transient transformation frequencies ranging from 12% to 36% for leaf explants, 15% to 42% for epicotyls. Initial evidence of transformation was obtained by polymerase chain reaction and subsequently confirmed by Southern analysis of regenerated plants.

  15. Global Analysis of Differentially Expressed Genes and Proteins in the Wheat Callus Infected by Agrobacterium tumefaciens

    PubMed Central

    Zhou, Xiaohong; Wang, Ke; Lv, Dongwen; Wu, Chengjun; Li, Jiarui; Zhao, Pei; Lin, Zhishan; Du, Lipu; Yan, Yueming; Ye, Xingguo

    2013-01-01

    Agrobacterium-mediated plant transformation is an extremely complex and evolved process involving genetic determinants of both the bacteria and the host plant cells. However, the mechanism of the determinants remains obscure, especially in some cereal crops such as wheat, which is recalcitrant for Agrobacterium-mediated transformation. In this study, differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) were analyzed in wheat callus cells co-cultured with Agrobacterium by using RNA sequencing (RNA-seq) and two-dimensional electrophoresis (2-DE) in conjunction with mass spectrometry (MS). A set of 4,889 DEGs and 90 DEPs were identified, respectively. Most of them are related to metabolism, chromatin assembly or disassembly and immune defense. After comparative analysis, 24 of the 90 DEPs were detected in RNA-seq and proteomics datasets simultaneously. In addition, real-time RT-PCR experiments were performed to check the differential expression of the 24 genes, and the results were consistent with the RNA-seq data. According to gene ontology (GO) analysis, we found that a big part of these differentially expressed genes were related to the process of stress or immunity response. Several putative determinants and candidate effectors responsive to Agrobacterium mediated transformation of wheat cells were discussed. We speculate that some of these genes are possibly related to Agrobacterium infection. Our results will help to understand the interaction between Agrobacterium and host cells, and may facilitate developing efficient transformation strategies in cereal crops. PMID:24278131

  16. Floral-Dip Transformation of Flax (Linum usitatissimum) to Generate Transgenic Progenies with a High Transformation Rate

    PubMed Central

    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. PMID:25549243

  17. Floral-dip transformation of flax (Linum usitatissimum) to generate transgenic progenies with a high transformation rate.

    PubMed

    Bastaki, Nasmah K; Cullis, Christopher A

    2014-12-19

    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. Highly efficient Agrobacterium-mediated transformation of embryogenic cell suspensions of Musa acuminata cv. Mas (AA) via a liquid co-cultivation system.

    PubMed

    Huang, Xia; Huang, Xue-Lin; Xiao, Wang; Zhao, Jie-Tang; Dai, Xue-Mei; Chen, Yun-Feng; Li, Xiao-Ju

    2007-10-01

    A high efficient protocol of Agrobacterium-mediated transformation of Musa acuminata cv. Mas (AA), a major banana variety of the South East Asia region, was developed in this study. Male-flower-derived embryogenic cell suspensions (ECS) were co-cultivated in liquid medium with Agrobacterium strain EHA105 harboring a binary vector pCAMBIA2301 carrying nptII and gusA gene in the T-DNA. Depending upon conditions and duration of co-cultivation in liquid medium, 0-490 transgenic plants per 0.5 ml packed cell volume (PCV) of ECS were obtained. The optimum duration of inoculation was 2 h, and the highest transformation frequency was achieved when infected ECS were co-cultivated in liquid medium first for 12 h at 40 rpm and then for 156 h at 100 rpm on a rotary shaker. Co-cultivation for a shorter duration (72 h) or shaking constantly at 100 rpm at the same duration gave 1.6 and 1.8 folds lower transformation efficiency, respectively. No transgenic plants were obtained in parallel experiments carried on semi-solid media. Histochemical GUS assay and molecular analysis in several tissues of the transgenic plants demonstrated that foreign genes were stably integrated into the banana genome. Compared to semi-solid co-cultivation transformation in other banana species, it is remarkable that liquid co-cultivation was much more efficient for transformation of the Mas cultivar, and was at least 1 month faster for regenerating transgenic plants.

  19. Agrobacterium rhizogenes-induced cotton hairy root culture as an alternative tool for cotton functional genomics

    USDA-ARS?s Scientific Manuscript database

    Although well-accepted as the ultimate method for cotton functional genomics, Agrobacterium tumefaciens-mediated cotton transformation is not widely used for functional analyses of cotton genes and their promoters since regeneration of cotton in tissue culture is lengthy and labor intensive. In cer...

  20. Beyond Agrobacterium-Mediated Transformation: Horizontal Gene Transfer from Bacteria to Eukaryotes.

    PubMed

    Lacroix, Benoît; Citovsky, Vitaly

    2018-03-03

    Besides the massive gene transfer from organelles to the nuclear genomes, which occurred during the early evolution of eukaryote lineages, the importance of horizontal gene transfer (HGT) in eukaryotes remains controversial. Yet, increasing amounts of genomic data reveal many cases of bacterium-to-eukaryote HGT that likely represent a significant force in adaptive evolution of eukaryotic species. However, DNA transfer involved in genetic transformation of plants by Agrobacterium species has traditionally been considered as the unique example of natural DNA transfer and integration into eukaryotic genomes. Recent discoveries indicate that the repertoire of donor bacterial species and of recipient eukaryotic hosts potentially are much wider than previously thought, including donor bacterial species, such as plant symbiotic nitrogen-fixing bacteria (e.g., Rhizobium etli) and animal bacterial pathogens (e.g., Bartonella henselae, Helicobacter pylori), and recipient species from virtually all eukaryotic clades. Here, we review the molecular pathways and potential mechanisms of these trans-kingdom HGT events and discuss their utilization in biotechnology and research.

  1. Agrobacterium-mediated transient MaFT expression in mulberry (Morus alba L.) leaves.

    PubMed

    Wu, Su-Li; Yang, Xiao-Bing; Liu, Li-Qun; Jiang, Tao; Wu, Hai; Su, Chao; Qian, Yong-Hua; Jiao, Feng

    2015-01-01

    To optimize Agrobacterium-mediated transient transformation assay in mulberry (Morus alba L.), various infiltration methods, Agrobacterium tumefaciens (A. tumefaciens) strains, and bacterial concentrations were tested in mulberry seedlings. Compared with LBA4404, GV3101 harboring pBE2133 plasmids presented stronger GUS signals at 3 days post infiltration using syringe. Recombinant plasmids pBE2133:GFP and pBE2133:GFP:MaFT were successfully constructed. Transient expression of MaFT:GFP protein was found in leaves, petiole (cross section), and shoot apical meristem (SAM) of mulberry according to the GFP signal. Moreover, MaFT:GFP mRNA was also detected in leaves and SAM via RT-PCR and qRT-PCR. An efficient transient transformation system could be achieved in mulberry seedlings by syringe using A. tumefaciens GV3101 at the OD600 of 0.5. The movement of MaFT expression from leaves to SAM might trigger the precocious flowering of mulberry.

  2. Development of Protoporphyrinogen Oxidase as an Efficient Selection Marker for Agrobacterium tumefaciens-Mediated Transformation of Maize

    PubMed Central

    Li, Xianggan; Volrath, Sandy L.; Nicholl, David B.G.; Chilcott, Charles E.; Johnson, Marie A.; Ward, Eric R.; Law, Marcus D.

    2003-01-01

    In this article, we report the isolation of plant protoporphyrinogen oxidase (PPO) genes and the isolation of herbicide-tolerant mutants. Subsequently, an Arabidopsis double mutant (Y426M + S305L) was used to develop a selectable marker system for Agrobacterium tumefaciens-mediated transformation of maize (Zea mays) and to obtain multiple events tolerant to the PPO family of herbicides. Maize transformants were produced via butafenacil selection using a flexible light regime to increase selection pressure. Butafenacil selection per se did not change transgene copy number distribution relative to other selectable marker systems, but the most tolerant events identified in the greenhouse were more likely to contain multiple copies of the introduced mutant PPO gene. To date, more than 2,500 independent transgenic maize events have been produced using butafenacil selection. The high frequency of A. tumefaciens-mediated transformation via PPO selection enabled us to obtain single-copy transgenic maize lines tolerant to field levels of butafenacil. PMID:12972658

  3. Genetic transformation of Bacopa monnieri by wild type strains of Agrobacterium rhizogenes stimulates production of bacopa saponins in transformed calli and plants.

    PubMed

    Majumdar, Sukanya; Garai, Saraswati; Jha, Sumita

    2011-05-01

    We have developed an efficient transformation system for Bacopa monnieri, an important Indian medicinal plant, using Agrobacterium rhizogenes strains LBA 9402 and A4. Transformed roots induced by strain LBA 9402 spontaneously dedifferentiated to callus while excised roots induced by strain A4 spontaneously showed induction of shoot buds within 10 days. PCR and RT-PCR analysis revealed the presence and expression of the rolAB and rolC genes at the transcription level in pRi A4 transformed cultures indicating that the TL-DNA was integrated retained and expressed in the A4-Ri transformed shoots. Transformed calli showed the presence of rolAB or rol A, TR and ags genes. Transformed plants showed morphological features typically seen in transgenic plants produced by A. rhizogenes. Growth and biomass accumulation was significantly higher in the transformed shoots (twofold) and roots (fourfold) than in the non-transformed (WT) plants. In pRi A4-transformed plants, the content of bacopasaponin D, bacopasaponin F, bacopaside II and bacopaside V was enhanced significantly as compared to WT plants of similar age while bacoside A3 and bacopasaponin C content was comparable with that of WT plants. Significant increase in content of five bacopa saponins could be detected in pRi 9402-transformed callus cultures. There is an overall stimulatory effect on accumulation of bacopa saponins in transformed plants and cells of B. monnieri establishing the role of endogenous elicitation by Ri T-DNA of A. rhizogenes.

  4. 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

  5. Agrobacterium tumefaciens-mediated transformation for investigation of somatic recombination in the fungal pathogen Armillaria mellea.

    PubMed

    Baumgartner, Kendra; Fujiyoshi, Phillip; Foster, Gary D; Bailey, Andy M

    2010-12-01

    Armillaria root disease is one of the most damaging timber and fruit tree diseases in the world. Despite its economic importance, many basic questions about the biology of the causal fungi, Armillaria spp., are unanswered. For example, Armillaria undergoes matings between diploid and haploid mycelia, which can result in a recombinant diploid without meiosis. Evidence of such somatic recombination in natural populations suggests that this reproductive mode may affect the pathogen's ecology. Investigations of the mechanisms and adaptive consequences of somatic recombination are, however, hampered by the lack of a method to reliably synthesize somatic recombinants. Here we report the first genetic transformation system for the genus Armillaria. We transformed A. mellea with selective markers for use in diploid-haploid matings to reliably synthesize somatic recombinants. This was accomplished with Agrobacterium tumefaciens carrying pBGgHg, which carries the hygromycin phosphotransferase gene (hph). hph was integrated into transformants, as evidenced by serial transfer to selective media, PCR, reverse transcription-PCR (RT-PCR), and Southern hybridization. Nuclear and mitochondrial markers were developed to genotype synthesized mycelia. In matings between a wild-type diploid and hygromycin-resistant haploids (transgenic), we identified recombinant, hygromycin-resistant diploids and, additionally, hygromycin-resistant triploids, all with the mitochondrial haplotype of the haploid partner. Our approach created no mycelium in which the haploid nucleus was replaced by the diploid nucleus, the typical outcome of diploid-haploid matings in Armillaria. This genetic transformation system, in combination with new markers to track chromosomal and cytoplasmic inheritance in A. mellea, will advance research aimed at characterizing the significance of somatic recombination in the ecology of this important fungus.

  6. A new transformation-regeneration procedure in the model legume Lotus japonicus: root explants as a source of large numbers of cells susceptible to Agrobacterium-mediated transformation.

    PubMed

    Lombari, P; Ercolano, E; El Alaoui, H; Chiurazzi, M

    2003-04-01

    We describe herein a simple and efficient transformation procedure for the production of transgenic Lotus japonicus plants. In this new procedure, dedifferentiated root explants, used as starting material, are the source of a large number of cells that are competent for the regeneration procedure, with a high susceptibility to Agrobacterium infection. The application of this protocol resulted in a tenfold increase in the number of transformants produced by a single plant in comparison to the widely used hypocotyl transformation procedure. Furthermore, our procedure allowed the use of intact plants stored for a long time at 4 degrees C, thus providing a potential continuous supply of explants for transformation experiments. The overall time of incubation under tissue culture conditions required to obtain a plant transferable into soil is 4 months. The transgenic nature of the transformants was demonstrated by the detection of beta-glucuronidase (GUS) activity in the primary transformants and by molecular analysis. Stable transformation was indicated by Mendelian segregation of the hygromycin selectable marker and of the gusA activity after selfing of the transgenic plants.

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

    USDA-ARS?s Scientific Manuscript database

    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...

  8. In planta transformation of pigeon pea: a method to overcome recalcitrancy of the crop to regeneration in vitro.

    PubMed

    Sankara Rao, K; Sreevathsa, Rohini; Sharma, Pinakee D; Keshamma, E; Udaya Kumar, M

    2008-10-01

    Development of transgenics in pigeon pea remains dogged by poor plant regeneration in vitro from transformed tissues and low frequency transformation protocols. This article presents a non-tissue culture-based method of generating transgenic pigeon pea (Cajanus cajan (L.) Millisp.) plants using Agrobacterium-Ti plasmid-mediated transformation system. The protocol involves raising of whole plant transformants (T0 plants) directly from Agrobacterium-infected young seedlings. The plumular and intercotyledonary meristems of the seedling axes are targeted for transformation. The transformation conditions optimized were, pricking of the apical and intercotyledonary region of the seedling axes of two-day old germinating seedlings with a sewing needle, infection with Agrobacterium (LBA4404/pKIWI105 carrying uid A and npt II genes) in Winans' AB medium that was added with wounded tobacco leaf extract, co-cultivation in the same medium for 1h and transfer of seedlings to soilrite for further growth and hardening and subsequent transfer of seedlings to soil in pots in the greenhouse. Out of the 22-25 primary transformants that survived infection-hardening treatments from each of the three experiments, 15 plants on the average established on the soil under greenhouse conditions, showed slow growth initially, nevertheless grew as normal plants, and flowered and set seed eventually. Of the several seeds harvested from all the T0 plants, six hundred were sown to obtain progeny (T1) plants and 350 of these were randomly analysed to determine their transgenic nature. PCR was performed for both gus (uid A) and npt II genes. Forty eight of the 350 T1 plants amplified both transgenes. Southern blot analysis substantiated the integration and transmission of these genes. The protocol ensured generation of pigeon pea transgenic plants with considerable ease in a short time and is applicable across different genotypes/cultivars of the crop and offers immense potential as a supplemental or

  9. Plant cell transformation with Agrobacterium tumefaciens under simulated microgravity

    NASA Astrophysics Data System (ADS)

    Sarnatska, Veresa; Gladun, Hanna; Padalko, Svetlana

    To investigate simulated microgravity (clinorotation) effect on plant cell transformation with Agrobacterium tumefaciens and crown gall formation, the culture of primary explants of potato and Jerusalem artichoke tubers was used. It is found that the efficiency of tumor formation and development in clinorotated explants are considerably reduced. When using the explants isolated from potato tubers clinorotated for 3, 5 and 19 days, drastic reduction of formation and development of crown gall tumors was observed. Conversely, the tumor number and their development increased when potato tubers were clinorotated for one day. As was estimated by us previously, cells of Jerusalem artichoke explants are the most sensitive to agrobacteria on 4-5 h of in vitro culturing and this time corresponds to the certain period of G1-stage of the cell cycle. We have also estimated that this period is characterized by the increase of binding of acridine orange by nuclear chromatin and increase in activity of RNA-polymerase I and II. Inoculation of explants with agrobacteria in this period was the most optimal for transformation and crown gall induction. We estimated that at four - hour clinorotation of explants the intensity of acridine orange binding to nuclei was considerably lower than on 4h in the control. At one-day clinorotation of potato tubers, a considerable increase in template accessibility of chromatin and in activity of RNA-polymerase I and II occurred. These results may serve as an evidence for the ability of plant dormant tissues to respond to microgravity. Another demonstration of dormant tissue response to changed gravity we obtained when investigating pathogenesis-related proteins (PR-proteins). PR-proteins were subjected to nondenaturing PAGE.and we have not found any effect of microgravity on PR-proteins of potato explants with normal or tumorous growth. We may suggest that such response derives from the common effects of two stress factors - wounding and changed

  10. Improvement of tissue culture, genetic transformation, and applications of biotechnology to Brassica.

    PubMed

    Ravanfar, Seyed Ali; Orbovic, Vladimir; Moradpour, Mahdi; Abdul Aziz, Maheran; Karan, Ratna; Wallace, Simon; Parajuli, Saroj

    2017-04-01

    Development of in vitro plant regeneration method from Brassica explants via organogenesis and somatic embryogenesis is influenced by many factors such as culture environment, culture medium composition, explant sources, and genotypes which are reviewed in this study. An efficient in vitro regeneration system to allow genetic transformation of Brassica is a crucial tool for improving its economical value. Methods to optimize transformation protocols for the efficient introduction of desirable traits, and a comparative analysis of these methods are also reviewed. Hence, binary vectors, selectable marker genes, minimum inhibitory concentration of selection agents, reporter marker genes, preculture media, Agrobacterium concentration and regeneration ability of putative transformants for improvement of Agrobacterium-mediated transformation of Brassica are discussed.

  11. Agrobacterium-mediated virus-induced gene silencing assay in cotton.

    PubMed

    Gao, Xiquan; Britt, Robert C; Shan, Libo; He, Ping

    2011-08-20

    Cotton (Gossypium hirsutum) is one of the most important crops worldwide. Considerable efforts have been made on molecular breeding of new varieties. The large-scale gene functional analysis in cotton has been lagged behind most of the modern plant species, likely due to its large size of genome, gene duplication and polyploidy, long growth cycle and recalcitrance to genetic transformation(1). To facilitate high throughput functional genetic/genomic study in cotton, we attempt to develop rapid and efficient transient assays to assess cotton gene functions. Virus-Induced Gene Silencing (VIGS) is a powerful technique that was developed based on the host Post-Transcriptional Gene Silencing (PTGS) to repress viral proliferation(2,3). Agrobacterium-mediated VIGS has been successfully applied in a wide range of dicots species such as Solanaceae, Arabidopsis and legume species, and monocots species including barley, wheat and maize, for various functional genomic studies(3,4). As this rapid and efficient approach avoids plant transformation and overcomes functional redundancy, it is particularly attractive and suitable for functional genomic study in crop species like cotton not amenable for transformation. In this study, we report the detailed protocol of Agrobacterium-mediated VIGS system in cotton. Among the several viral VIGS vectors, the tobacco rattle virus (TRV) invades a wide range of hosts and is able to spread vigorously throughout the entire plant yet produce mild symptoms on the hosts5. To monitor the silencing efficiency, GrCLA1, a homolog gene of Arabidopsis Cloroplastos alterados 1 gene (AtCLA1) in cotton, has been cloned and inserted into the VIGS binary vector pYL156. CLA1 gene is involved in chloroplast development(6), and previous studies have shown that loss-of-function of AtCLA1 resulted in an albino phenotype on true leaves(7), providing an excellent visual marker for silencing efficiency. At approximately two weeks post Agrobacterium infiltration

  12. Agrobacterium-Mediated Virus-Induced Gene Silencing Assay In Cotton

    PubMed Central

    Gao, Xiquan; Britt Jr., Robert C.; Shan, Libo; He, Ping

    2011-01-01

    Cotton (Gossypium hirsutum) is one of the most important crops worldwide. Considerable efforts have been made on molecular breeding of new varieties. The large-scale gene functional analysis in cotton has been lagged behind most of the modern plant species, likely due to its large size of genome, gene duplication and polyploidy, long growth cycle and recalcitrance to genetic transformation1. To facilitate high throughput functional genetic/genomic study in cotton, we attempt to develop rapid and efficient transient assays to assess cotton gene functions. Virus-Induced Gene Silencing (VIGS) is a powerful technique that was developed based on the host Post-Transcriptional Gene Silencing (PTGS) to repress viral proliferation2,3. Agrobacterium-mediated VIGS has been successfully applied in a wide range of dicots species such as Solanaceae, Arabidopsis and legume species, and monocots species including barley, wheat and maize, for various functional genomic studies3,4. As this rapid and efficient approach avoids plant transformation and overcomes functional redundancy, it is particularly attractive and suitable for functional genomic study in crop species like cotton not amenable for transformation. In this study, we report the detailed protocol of Agrobacterium-mediated VIGS system in cotton. Among the several viral VIGS vectors, the tobacco rattle virus (TRV) invades a wide range of hosts and is able to spread vigorously throughout the entire plant yet produce mild symptoms on the hosts5. To monitor the silencing efficiency, GrCLA1, a homolog gene of Arabidopsis Cloroplastos alterados 1 gene (AtCLA1) in cotton, has been cloned and inserted into the VIGS binary vector pYL156. CLA1 gene is involved in chloroplast development6, and previous studies have shown that loss-of-function of AtCLA1 resulted in an albino phenotype on true leaves7, providing an excellent visual marker for silencing efficiency. At approximately two weeks post Agrobacterium infiltration, the albino

  13. Relationship between expression level of hygromycin B-resistant gene and Agrobacterium tumefaciens-mediated transformation efficiency in Beauveria bassiana JEF-007.

    PubMed

    Nai, Y S; Lee, M R; Kim, S; Lee, S J; Kim, J C; Yang, Y T; Kim, J S

    2017-09-01

    Agrobacterium tumefaciens-mediated transformation (AtMT) is an effective method for generation of entomopathogenic Beauveria bassiana transformants. However, some strains grow on the selective medium containing hygromycin B (HygB), which reduces the selection efficiency of the putative transformants. In this work, a relationship between HygB resistance gene promoter and AtMT efficiency was investigated to improve the transformant selection. Ten B. bassiana isolates were grown on 800 μg ml -1 HygB medium, but only JEF-006, -007 and -013 showed susceptibility to the antibiotics. Particularly, JEF-007 showed the most dose-dependent susceptibility. Two different Ti-Plasmids, pCeg (gpdA promoter based) and pCambia-egfp (CaMV 35S promoter based), were constructed to evaluate the promoters on the expression of HygB resistance gene (hph) at 100, 150 and 200 μg ml -1 HygB medium. Eight days after the transformation, wild type, AtMT/pCeg and AtMT/pCambia-egfp colonies were observed on 100 μg ml -1 HygB, but significantly larger numbers of colonies were counted on AtMT/pCeg plates. At higher HygB concentration (150 μg ml -1 ), only AtMT/pCeg colonies were further observed, but very few colonies were observed on the wild type and AtMT/pCambia-egfp plates. Putative transformants were subjected to PCR, RT-PCR and qRT-PCR to investigate the T-DNA insertion rate and gene expression level. Consequently, >80% of colonies showed successful AtMT transformation, and the hph expression level in AtMT/pCeg colonies was higher than that of AtMT/pCambia-egfp colonies. In the HygB-susceptible B. bassianaJEF-007, gpdA promoter works better than CaMV 35S promoter in the expression of HygB resistance gene at 150 μg ml -1 HygB, consequently improving the selection efficiency of putative transformants. These results provide useful information for determining AtMT effectiveness in B. bassiana isolates, particularly antibiotic susceptibility and the role of promoters. © 2017 The

  14. 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.

  15. The construction and use of versatile binary vectors carrying pyrG auxotrophic marker and fluorescent reporter genes for Agrobacterium-mediated transformation of Aspergillus oryzae.

    PubMed

    Nguyen, Khuyen Thi; Ho, Quynh Ngoc; Pham, Thu Ha; Phan, Tuan-Nghia; Tran, Van-Tuan

    2016-12-01

    Aspergillus oryzae is a safe mold widely used in food industry. It is also considered as a microbial cell factory for production of recombinant proteins and enzymes. Currently, genetic manipulation of filamentous fungi is achieved via Agrobacterium tumefaciens-mediated transformation methods usually employing antibiotic resistance markers. These methods are hardly usable for A. oryzae due to its strong resistance to the common antifungal compounds used for fungal transformation. In this study, we have constructed two binary vectors carrying the pyrG gene from A. oryzae as a biochemical marker than an antibiotic resistance marker, and an expression cassette for GFP or DsRed reporter gene under control of the constitutive gpdA promoter from Aspergillus nidulans. All components of these vectors are changeable to generate new versions for specific research purposes. The developed vectors are fully functional for heterologous expression of the GFP and DsRed fluorescent proteins in the uridine/uracil auxotrophic A. oryzae strain. Our study provides a new approach for A. oryzae transformation using pyrG as the selectable auxotrophic marker, A. tumefaciens as the DNA transfer tool and fungal spores as the transformation material. The binary vectors constructed can be used for gene expression studies in this industrially important filamentous fungus.

  16. 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.

  17. Acetosyringone, pH and temperature effects on transient genetic transformation of immature embryos of Brazilian wheat genotypes by Agrobacterium tumefaciens.

    PubMed

    Manfroi, Ernandes; Yamazaki-Lau, Elene; Grando, Magali F; Roesler, Eduardo A

    2015-12-01

    Low transformation efficiency is one of the main limiting factors in the establishment of genetic transformation of wheat via Agrobacterium tumefaciens. To determine more favorable conditions for T-DNA delivery and explant regeneration after infection, this study investigated combinations of acetosyringone concentration and pH variation in the inoculation and co-cultivation media and co-culture temperatures using immature embryos from two Brazilian genotypes (BR 18 Terena and PF 020037). Based on transient expression of uidA, the most favorable conditions for T-DNA delivery were culture media with pH 5.0 and 5.4 combined with co-culture temperatures of 22 °C and 25 °C, and a 400 μM acetosyringone supplement. These conditions resulted in blue foci in 81% of the embryos. Media with more acidic pH also presented reduced A. tumefaciens overgrowth during co-culture, and improved regeneration frequency of the inoculated explants. BR 18 Terena was more susceptible to infection by A. tumefaciens than PF 020037. We found that it is possible to improve T-DNA delivery and explant regeneration by adjusting factors involved in the early stages of A. tumefaciens infection. This can contribute to establishing a stable transformation procedure in the future.

  18. Biolistics Transformation of Wheat

    NASA Astrophysics Data System (ADS)

    Sparks, Caroline A.; Jones, Huw D.

    We present a complete, step-by-step guide to the production of transformed wheat plants using a particle bombardment device to deliver plasmid DNA into immature embryos and the regeneration of transgenic plants via somatic embryogenesis. Currently, this is the most commonly used method for transforming wheat and it offers some advantages. However, it will be interesting to see whether this position is challenged as facile methods are developed for delivering DNA by Agrobacterium tumefaciens or by the production of transformants via a germ-line process (see other chapters in this book).

  19. Hairy Root Transformation Using Agrobacterium rhizogenes as a Tool for Exploring Cell Type-Specific Gene Expression and Function Using Tomato as a Model1[W][OPEN

    PubMed Central

    Ron, Mily; Kajala, Kaisa; Pauluzzi, Germain; Wang, Dongxue; Reynoso, Mauricio A.; Zumstein, Kristina; Garcha, Jasmine; Winte, Sonja; Masson, Helen; Inagaki, Soichi; Federici, Fernán; Sinha, Neelima; Deal, Roger B.; Bailey-Serres, Julia; Brady, Siobhan M.

    2014-01-01

    Agrobacterium rhizogenes (or Rhizobium rhizogenes) is able to transform plant genomes and induce the production of hairy roots. We describe the use of A. rhizogenes in tomato (Solanum spp.) to rapidly assess gene expression and function. Gene expression of reporters is indistinguishable in plants transformed by Agrobacterium tumefaciens as compared with A. rhizogenes. A root cell type- and tissue-specific promoter resource has been generated for domesticated and wild tomato (Solanum lycopersicum and Solanum pennellii, respectively) using these approaches. Imaging of tomato roots using A. rhizogenes coupled with laser scanning confocal microscopy is facilitated by the use of a membrane-tagged protein fused to a red fluorescent protein marker present in binary vectors. Tomato-optimized isolation of nuclei tagged in specific cell types and translating ribosome affinity purification binary vectors were generated and used to monitor associated messenger RNA abundance or chromatin modification. Finally, transcriptional reporters, translational reporters, and clustered regularly interspaced short palindromic repeats-associated nuclease9 genome editing demonstrate that SHORT-ROOT and SCARECROW gene function is conserved between Arabidopsis (Arabidopsis thaliana) and tomato. PMID:24868032

  20. Transformation of pickling cucumber with chitinase-encoding genes using Agrobacterium tumefaciens.

    PubMed

    Raharjo, S H; Hernandez, M O; Zhang, Y Y; Punja, Z K

    1996-04-01

    Transformation of cucumber cv. Endeavor was attempted using three Agrobacterium tumefaciens strains (a supervirulent leucinopine type, an octopine type and a nopaline type), each harbouring one of three binary vectors which contained an acidic chitinase gene from petunia, and basic chitinase genes from tobacco and bean, respectively, driven by the CaMV 35S promoter. Petiole explants were inoculated with a bacterial suspension (10(8) cells·ml(-1)), cocultivated for 48-96 h and placed on Murashige and Skoog (MS) medium with 5.0 μM each of 2,4-D and BA, 50 mg·l(-1) kanamycin and 500 mg·l(-1) carbenicillin. The frequency of embryogenic callus formation ranged from 0 to 12%, depending on strains/vectors used and length of cocultivation, with the highest being obtained using the leucinopine strain with petunia acidic chitinase gene. The kanamycin-resistant embryogenic calli were used to initiate suspension cultures (in liquid MS medium with 1.0/1.0 μM 2,4-D/BA, 50 mg·l(-1) kanamycin) for multiplication of embryogenic cell aggregates. Upon plating of cell aggregates onto solid MS medium with 1.0/1.0 μM NAA/BA and 50 mg·l(-1) kanamycin, calli continued to grow and later differentiated into plantlets. Transformation by the leucinopine strain and all three vectors was confirmed by PCR amplification of the NPT II gene in transgenic calli and plants, in addition to Southern analysis. Expression of the acidic chitinase gene (from petunia) and both basic chitinase genes (from tobacco and bean) in different transgenic cucumber lines was confirmed by Western analyses.

  1. Development of a rapid and simple Agrobacterium tumefaciens mediated transformation system for the fungal pathogen Heterobasidion annosum

    Treesearch

    Nicklas Samils; Malin Elfstrand; Daniel L. Lindner Czederpiltz; Jan Fahleson; Ake Olson; Christina Dixelius; Jan Stenlid

    2006-01-01

    Heterobasidion annosum causes root and butt-rot in trees and is the most serious forest pathogen in the northern hemisphere. We developed a rapid and simple Agrobacterium-mediated method of gene delivery into H. annosum to be used in functional studies of candidate genes and for visualization of mycelial interactions. Heterobasidion annosum TC 32-1 was cocultivated at...

  2. Nopaline-type Ti plasmid of Agrobacterium encodes a VirF-like functional F-box protein.

    PubMed

    Lacroix, Benoît; Citovsky, Vitaly

    2015-11-20

    During Agrobacterium-mediated genetic transformation of plants, several bacterial virulence (Vir) proteins are translocated into the host cell to facilitate infection. One of the most important of such translocated factors is VirF, an F-box protein produced by octopine strains of Agrobacterium, which presumably facilitates proteasomal uncoating of the invading T-DNA from its associated proteins. The presence of VirF also is thought to be involved in differences in host specificity between octopine and nopaline strains of Agrobacterium, with the current dogma being that no functional VirF is encoded by nopaline strains. Here, we show that a protein with homology to octopine VirF is encoded by the Ti plasmid of the nopaline C58 strain of Agrobacterium. This protein, C58VirF, possesses the hallmarks of functional F-box proteins: it contains an active F-box domain and specifically interacts, via its F-box domain, with SKP1-like (ASK) protein components of the plant ubiquitin/proteasome system. Thus, our data suggest that nopaline strains of Agrobacterium have evolved to encode a functional F-box protein VirF.

  3. A Perspective on Hypericum perforatum Genetic Transformation

    PubMed Central

    Hou, Weina; Shakya, Preeti; Franklin, Gregory

    2016-01-01

    Hypericum perforatum (St John's wort) is a reservoir of diverse classes of biologically active and high value secondary metabolites, which captured the interest of both researchers and the pharmaceutical industry alike. Several studies and clinical trials have shown that H. perforatum extracts possess an astounding array of pharmacological properties. These properties include antidepressant, anti-inflammatory, antiviral, anti-cancer, and antibacterial activities; and are largely attributed to the naphtodianthrones and xanthones found in the genus. Hence, improving their production via genetic manipulation is an important strategy. In spite of the presence of contemporary genome editing tools, genetic improvement of this genus remains challenging without robust transformation methods in place. In the recent past, we found that H. perforatum remains recalcitrant to Agrobacterium tumefaciens mediated transformation partly due to the induction of plant defense responses coming into play. However, H. perforatum transformation is possible via a non-biological method, biolistic bombardment. Some research groups have observed the induction of hairy roots in H. perforatum after Agrobacterium rhizogenes co-cultivation. In this review, we aim at updating the available methods for regeneration and transformation of H. perforatum. In addition, we also propose a brief perspective on certain novel strategies to improve transformation efficiency in order to meet the demands of the pharmaceutical industry via metabolic engineering. PMID:27446112

  4. Methods for genetic transformation in Dendrobium.

    PubMed

    da Silva, Jaime A Teixeira; Dobránszki, Judit; Cardoso, Jean Carlos; Chandler, Stephen F; Zeng, Songjun

    2016-03-01

    The genetic transformation of Dendrobium orchids will allow for the introduction of novel colours, altered architecture and valuable traits such as abiotic and biotic stress tolerance. The orchid genus Dendrobium contains species that have both ornamental value and medicinal importance. There is thus interest in producing cultivars that have increased resistance to pests, novel horticultural characteristics such as novel flower colours, improved productivity, longer flower spikes, or longer post-harvest shelf-life. Tissue culture is used to establish clonal plants while in vitro flowering allows for the production of flowers or floral parts within a sterile environment, expanding the selection of explants that can be used for tissue culture or genetic transformation. The latter is potentially the most effective, rapid and practical way to introduce new agronomic traits into Dendrobium. Most (69.4 %) Dendrobium genetic transformation studies have used particle bombardment (biolistics) while 64 % have employed some form of Agrobacterium-mediated transformation. A singe study has explored ovary injection, but no studies exist on floral dip transformation. While most of these studies have involved the use of selector or reporter genes, there are now a handful of studies that have introduced genes for horticulturally important traits.

  5. Agrobacterium tumefaciens-mediated transformation of blueberry (Vaccinium corymbosum L.).

    PubMed

    Song, Guo-Qing; Sink, K C

    2004-12-01

    Transient expression studies using blueberry leaf explants and monitored by beta-glucuronidase (GUS) assays indicated Agrobacterium tumefaciens strain EHA105 was more effective than LBA4404 or GV3101; and the use of acetosyringone (AS) at 100 microM for inoculation and 6 days co-cultivation was optimum compared to 2, 4, 8, 10 or 12 days. Subsequently, explants of the cultivars Aurora, Bluecrop, Brigitta, and Legacy were inoculated with strain EHA105 containing the binary vector pBISN1 with the neomycin phosphotransferase gene (nptII) and an intron-interrupted GUS gene directed by the chimeric super promoter (Aocs)3AmasPmas. Co-cultivation was for 6 days on modified woody plant medium (WPM) plus 100 microM AS. Explants were then placed on modified WPM supplemented with 1.0 mg l(-1) thidiazuron, 0.5 mg l(-1) alpha-naphthaleneacetic, 10 mg l(-1) kanamycin (Km), and 250 mg l(-1) cefotaxime. Selection for Km-resistant shoots was carried out in the dark for 2 weeks followed by culture in the light at 30 microE m(-2) s(-1) at 25 degrees C. After 12 weeks, selected shoots that were both Km resistant and GUS positive were obtained from 15.3% of the inoculated leaf explants of cultivar Aurora. Sixty-eight independent clones derived from such shoots all tested positive by the polymerase chain reaction using a nptII primer. Eight of eight among these 68 clones tested positive by Southern hybridization using a gusA gene derived probe. The transformation protocol also yielded Km-resistant, GUS-positive shoots that were also PCR positive at frequencies of 5.0% for Bluecrop, 10.0% for Brigitta and 5.6% for Legacy.

  6. Horizontal gene transfer from Agrobacterium to plants.

    PubMed

    Matveeva, Tatiana V; Lutova, Ludmila A

    2014-01-01

    Most genetic engineering of plants uses Agrobacterium mediated transformation to introduce novel gene content. In nature, insertion of T-DNA in the plant genome and its subsequent transfer via sexual reproduction has been shown in several species in the genera Nicotiana and Linaria. In these natural examples of horizontal gene transfer from Agrobacterium to plants, the T-DNA donor is assumed to be a mikimopine strain of A. rhizogenes. A sequence homologous to the T-DNA of the Ri plasmid of Agrobacterium rhizogenes was found in the genome of untransformed Nicotiana glauca about 30 years ago, and was named "cellular T-DNA" (cT-DNA). It represents an imperfect inverted repeat and contains homologs of several T-DNA oncogenes (NgrolB, NgrolC, NgORF13, NgORF14) and an opine synthesis gene (Ngmis). A similar cT-DNA has also been found in other species of the genus Nicotiana. These presumably ancient homologs of T-DNA genes are still expressed, indicating that they may play a role in the evolution of these plants. Recently T-DNA has been detected and characterized in Linaria vulgaris and L. dalmatica. In Linaria vulgaris the cT-DNA is present in two copies and organized as a tandem imperfect direct repeat, containing LvORF2, LvORF3, LvORF8, LvrolA, LvrolB, LvrolC, LvORF13, LvORF14, and the Lvmis genes. All L. vulgaris and L. dalmatica plants screened contained the same T-DNA oncogenes and the mis gene. Evidence suggests that there were several independent T-DNA integration events into the genomes of these plant genera. We speculate that ancient plants transformed by A. rhizogenes might have acquired a selective advantage in competition with the parental species. Thus, the events of T-DNA insertion in the plant genome might have affected their evolution, resulting in the creation of new plant species. In this review we focus on the structure and functions of cT-DNA in Linaria and Nicotiana and discuss their possible evolutionary role.

  7. [Transgenic wheat (Triticum aestivum L.) with increased resistance to the storage pest obtained by Agrobacterium tumefaciens--mediated].

    PubMed

    Bi, Rui-Ming; Jia, Hai-Yan; Feng, De-Shun; Wang, Hong-Gang

    2006-05-01

    The transgenic wheat of improved resistance to the storage pest was production. We have introduced the cowpea trypsin inhibitor gene (CpTI) into cultured embryonic callus cells of immature embryos of wheat elite line by Agrobacterium-mediated method. Independent plantlets were obtained from the kanamycin-resistant calli after screening. PCR and real time PCR analysis, PCR-Southern and Southern blot hybridization indicated that there were 3 transgenic plants viz. transformed- I, II and III (T- I, T-II and T-III). The transformation frequencies were obviously affected by Agrobacterium concentration, the infection duration and transformation treatment. The segregations of CpTI in the transgenic wheat progenies were not easily to be elucidated, and some transgenic wheat lines (T- I and T-III) showed Mendelian segregations. The determinations of insect resistance to the stored grain insect of wheat viz. the grain moth (Sitotroga cerealella Olivier) indicated that the 3 transgenic wheat progeny seeds moth-resistance was improved significantly. The seed moth-eaten ratio of T- I, T-II, T-III and nontransformed control was 19.8%, 21.9%, 32.9% and 58.3% respectively. 3 transgenic wheat T1 PCR-positive plants revealed that the 3 transgenic lines had excellent agronomic traits. They supplied good germplasm resource of insect-resistance for wheat genetic improvement.

  8. Protoplast isolation, transient transformation of leaf mesophyll protoplasts and improved Agrobacterium-mediated leaf disc infiltration of Phaseolus vulgaris: tools for rapid gene expression analysis.

    PubMed

    Nanjareddy, Kalpana; Arthikala, Manoj-Kumar; Blanco, Lourdes; Arellano, Elizabeth S; Lara, Miguel

    2016-06-24

    Phaseolus vulgaris is one of the most extensively studied model legumes in the world. The P. vulgaris genome sequence is available; therefore, the need for an efficient and rapid transformation system is more imperative than ever. The functional characterization of P. vulgaris genes is impeded chiefly due to the non-amenable nature of Phaseolus sp. to stable genetic transformation. Transient transformation systems are convenient and versatile alternatives for rapid gene functional characterization studies. Hence, the present work focuses on standardizing methodologies for protoplast isolation from multiple tissues and transient transformation protocols for rapid gene expression analysis in the recalcitrant grain legume P. vulgaris. Herein, we provide methodologies for the high-throughput isolation of leaf mesophyll-, flower petal-, hypocotyl-, root- and nodule-derived protoplasts from P. vulgaris. The highly efficient polyethylene glycol-mannitol magnesium (PEG-MMG)-mediated transformation of leaf mesophyll protoplasts was optimized using a GUS reporter gene. We used the P. vulgaris SNF1-related protein kinase 1 (PvSnRK1) gene as proof of concept to demonstrate rapid gene functional analysis. An RT-qPCR analysis of protoplasts that had been transformed with PvSnRK1-RNAi and PvSnRK1-OE vectors showed the significant downregulation and ectopic constitutive expression (overexpression), respectively, of the PvSnRK1 transcript. We also demonstrated an improved transient transformation approach, sonication-assisted Agrobacterium-mediated transformation (SAAT), for the leaf disc infiltration of P. vulgaris. Interestingly, this method resulted in a 90 % transformation efficiency and transformed 60-85 % of the cells in a given area of the leaf surface. The constitutive expression of YFP further confirmed the amenability of the system to gene functional characterization studies. We present simple and efficient methodologies for protoplast isolation from multiple P

  9. Adapting rice anther culture to gene transformation and RNA interference.

    PubMed

    Chen, Caiyan; Xiao, Han; Zhang, Wenli; Wang, Aiju; Xia, Zhihui; Li, Xiaobing; Zhai, Wenxue; Cheng, Zhukuan; Zhu, Lihuang

    2006-10-01

    Anther culture offers a rapid method of generating homozygous lines for breeding program and genetic analysis. To produce homozygous transgenic lines of rice (Oryza sativa L.) in one step, we developed an efficient protocol of anther-callus-based transformation mediated by Agrobacterium after optimizing several factors influencing efficient transformation, including callus induction and Agrobacterium density for co-cultivation. Using this protocol, we obtained 145 independent green transformants from five cultivars of japonica rice by transformation with a binary vector pCXK1301 bearing the rice gene, Xa21 for resistance to bacterial blight, of which 140 were further confirmed by PCR and Southern hybridization analysis, including haploids (32.1%), diploids (62.1%) and mixoploids (7.5%). Fifteen diploids were found to be doubled haploids, which accounted for 10.7% of the total positive lines. Finally, by including 28 from colchicine induced or spontaneous diploidization of haploids later after transformation, a total of 43 doubled haploids (30.7%) of Xa21 transgenic lines were obtained. We also generated two RNAi transgenic haploids of the rice OsMADS2 gene, a putative redundant gene of OsMADS4 based on their sequence similarity, to investigate its possible roles in rice flower development by this method. Flowers from the two OsMADS2 RNAi transgenic haploids displayed obvious homeotic alternations, in which lodicules were transformed into palea/lemma-like tissues, whereas identities of other floral organs were maintained. The phenotypic alternations were proved to result from specific transcriptional suppression of OsMADS2 gene by the introduced RNAi transgene. The results confirmed that OsMADS2 is involved in lodicule development of rice flower and functionally redundant with OsMADS4 gene. Our results demonstrated that rice anther culture could be adapted to gene transformation and RNAi analysis in rice.

  10. Selection of transformation-efficient barley genotypes based on TFA (transformation amenability) haplotype and higher resolution mapping of the TFA loci.

    PubMed

    Hisano, Hiroshi; Meints, Brigid; Moscou, Matthew J; Cistue, Luis; Echávarri, Begoña; Sato, Kazuhiro; Hayes, Patrick M

    2017-04-01

    The genetic substitution of transformation amenability alleles from 'Golden Promise' can facilitate the development of transformation-efficient lines from recalcitrant barley cultivars. Barley (Hordeum vulgare) cv. 'Golden Promise' is one of the most useful and well-studied cultivars for genetic manipulation. In a previous report, we identified several transformation amenability (TFA) loci responsible for Agrobacterium-mediated transformation using the F 2 generation of immature embryos, derived from 'Haruna Nijo' × 'Golden Promise,' as explants. In this report, we describe higher density mapping of these TFA regions with additional SNP markers using the same transgenic plants. To demonstrate the robustness of transformability alleles at the TFA loci, we genotyped 202 doubled haploid progeny from the cross 'Golden Promise' × 'Full Pint.' Based on SNP genotype, we selected lines having 'Golden Promise' alleles at TFA loci and used them for transformation. Of the successfully transformed lines, DH120366 came the closest to achieving a level of transformation efficiency comparable to 'Golden Promise.' The results validate that the genetic substitution of TFA alleles from 'Golden Promise' can facilitate the development of transformation-efficient lines from recalcitrant barley cultivars.

  11. Component identification of electron transport chains in curdlan-producing Agrobacterium sp. ATCC 31749 and its genome-specific prediction using comparative genome and phylogenetic trees analysis.

    PubMed

    Zhang, Hongtao; Setubal, Joao Carlos; Zhan, Xiaobei; Zheng, Zhiyong; Yu, Lijun; Wu, Jianrong; Chen, Dingqiang

    2011-06-01

    Agrobacterium sp. ATCC 31749 (formerly named Alcaligenes faecalis var. myxogenes) is a non-pathogenic aerobic soil bacterium used in large scale biotechnological production of curdlan. However, little is known about its genomic information. DNA partial sequence of electron transport chains (ETCs) protein genes were obtained in order to understand the components of ETC and genomic-specificity in Agrobacterium sp. ATCC 31749. Degenerate primers were designed according to ETC conserved sequences in other reported species. DNA partial sequences of ETC genes in Agrobacterium sp. ATCC 31749 were cloned by the PCR method using degenerate primers. Based on comparative genomic analysis, nine electron transport elements were ascertained, including NADH ubiquinone oxidoreductase, succinate dehydrogenase complex II, complex III, cytochrome c, ubiquinone biosynthesis protein ubiB, cytochrome d terminal oxidase, cytochrome bo terminal oxidase, cytochrome cbb (3)-type terminal oxidase and cytochrome caa (3)-type terminal oxidase. Similarity and phylogenetic analyses of these genes revealed that among fully sequenced Agrobacterium species, Agrobacterium sp. ATCC 31749 is closest to Agrobacterium tumefaciens C58. Based on these results a comprehensive ETC model for Agrobacterium sp. ATCC 31749 is proposed.

  12. Morphogenetic and chemical stability of long-term maintained Agrobacterium-mediated transgenic Catharanthus roseus plants.

    PubMed

    Verma, Priyanka; Sharma, Abhishek; Khan, Shamshad Ahmad; Mathur, Ajay Kumar; Shanker, Karuna

    2015-01-01

    Transgenic Catharanthus roseus plants (transgenic Dhawal [DT] and transgenic Nirmal [NT]) obtained from the Agrobacterium tumefaciens and Agrobacterium rhizognenes-mediated transformations, respectively, have been maintained in vitro for 5 years. Plants were studied at regular intervals for various parameters such as plant height, leaf size, multiplication rate, alkaloid profile and presence of marker genes. DT plant gradually lost the GUS gene expression and it was not detected in the fifth year while NT plant demonstrated the presence of genes rolA, rolB and rolC even in the fifth year, indicating the more stable nature of Ri transgene. Vindoline content in the DT was two times more than in non-transformed control plants. Alkaloid and tryptophan profiles were almost constant during the 5 years. The cluster analysis revealed that the DT plant is more close to the control Nirmal plant followed by NT plant.

  13. High Efficiency Transformation of Cultured Tobacco Cells 1

    PubMed Central

    An, Gynheung

    1985-01-01

    Tobacco calli were transformed at levels up to 50% by cocultivation of tobacco cultured cells with Agrobacterium tumefaciens harboring the binary transfer-DNA vector, pGA472, containing a kanamycin resistance marker. Transformation frequency was dependent on the physiological state of the tobacco cells, the nature of Agrobacterium strain and, less so, on the expression of the vir genes of the tumor-inducing plasmid. Maximum transformation frequency was obtained with exponentially growing plant cells, suggesting that rapid growth of plant cells is an essental factor for efficient transformation of higher plants. Images Fig. 1 PMID:16664453

  14. 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. Copyright © 2015. Published by Elsevier Inc.

  15. Mapping of the Interaction Between Agrobacterium tumefaciens and Vanda Kasem's Delight Orchid Protocorm-Like Bodies.

    PubMed

    Gnasekaran, Pavallekoodi; Subramaniam, Sreeramanan

    2015-09-01

    Physical contact between A. tumefaciens and the target plant cell walls is essential to transfer and integrate the transgene to introduce a novel trait. Chemotaxis response and attachment of Agrobacterium towards Vanda Kasem's Delight (VKD) protocorm-like bodies (PLBs) were studied to analyse the interaction between Agrobacterium and PLB during the transformation event. The study shows that initially A. tumefaciens reversibly attached to PLB surface via polar and lateral mode of adherence followed by the irreversible attachment which involved the production of cellulosic fibril by A. tumefaciens. Cellulosic fibril allows formation of biofilm at the tip of trichome. Contrarily, attachment mutant Escherichia coli strain DH5α was significantly deficient in the attachment process. Spectrophotometric GUS assay showed the mean value of attachment by A. tumefaciens was 8.72 % compared to the negative control E. coli strain DH5α that produced 0.16 %. A. tumefaciens swarmed with sharper and brighter edge when severe wounding was applied to the PLBs producing the highest swarming ratio of 1.46 demonstrating the positive effect of the plant exudates on bacterial movement. The study shows that VKD's PLBs are the suitable explants for Agrobacterium-mediated transformation since the bacteria expressed higher competency rate.

  16. High-efficiency Agrobacterium rhizogenes-mediated transformation of heat inducible sHSP18.2-GUS in Nicotiana tabacum.

    PubMed

    Chen, Shih-Cheng; Liu, Hui-Wen; Lee, Kung-Ta; Yamakawa, Takashi

    2007-01-01

    The chimerical gene, Arabidopsis thaliana sHSP18.2 promoter fused to E. coli gusA gene, was Agrobacterium rhizogenes-mediated transformed into Nicotiana tabacum as a heat-regulatable model, and the thermo-inducible expression of GUS activity in N. tabacum transgenic hairy roots was profiled. An activation of A. rhizogenes with acetosyringone (AS) before cocultured with tobacco's leaf disc strongly promoted transgenic hairy roots formation. Transgenic hairy roots formation efficiency of A. rhizogenes precultured with 200 microM AS supplementation was 3.1-fold and 7.5-fold, respectively, compared to the formation efficiency obtained with and without AS supplementation in coculture. Transgenic hairy roots transformed with different AS concentration exhibited a similar pattern of thermo-inducibility after 10 min to 3 h heat treatments detected by GUS expression. The peak of expressed GUS specific activity, 399,530 pmol MUG per mg total protein per min, of the transgenic hairy roots was observed at 48 h after 3 h of 42 degrees C heat treatment, and the expressed GUS specific activity was 7-26 times more than that reported in A. thaliana, tobacco BY-2 cells and Nicotiana plumbaginifolia. Interference caused by AS supplementation on the growth of transgenic hairy roots, time-course of GUS expression and its expression level were not observed.

  17. 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.

  18. Genetic transformation of switchgrass.

    PubMed

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

    2009-01-01

    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.

  19. An Efficient Plant Regeneration and Transformation System of Ma Bamboo (Dendrocalamus latiflorus Munro) Started from Young Shoot as Explant

    PubMed Central

    Ye, Shanwen; Cai, Changyang; Ren, Huibo; Wang, Wenjia; Xiang, Mengqi; Tang, Xiaoshan; Zhu, Caiping; Yin, Tengfei; Zhang, Li; Zhu, Qiang

    2017-01-01

    Genetic engineering technology has been successfully used in many plant species, but is limited in woody plants, especially in bamboos. Ma bamboo (Dendrocalamus latiflorus Munro) is one of the most important bamboo species in Asia, and its genetic improvement was largely restricted by the lack of an efficient regeneration and transformation method. Here we reported a plantlet regeneration and Agrobacterium-mediated transformation protocol by using Ma bamboo young shoots as explants. Under our optimized conditions, embryogenic calluses were successfully induced from the excised young shoots on callus induction medium and rapidly grew on callus multiplication medium. Shoots and roots were regenerated on shoot induction medium and root induction medium, respectively, with high efficiency. An Agrobacterium-mediated genetic transformation protocol of Ma bamboo was established, verified by PCR and GUS staining. Furthermore, the maize Lc gene under the control of the ubiquitin promoter was successfully introduced into Ma bamboo genome and generated an anthocyanin over-accumulation phenotype. Our methods established here will facilitate the basic research as well as genetic breeding of this important bamboo species. Key achievements: A stable and high efficiency regeneration and Agrobacterium-mediated transformation protocol for Ma bamboo from vegetative organ is established. PMID:28798758

  20. Soybean (Glycine max) transformation using mature cotyledonary node explants.

    PubMed

    Olhoft, Paula M; Donovan, Christopher M; Somers, David A

    2006-01-01

    Agrobacterium tumefaciens-mediated transformation of soybeans has been steadily improved since its development in 1988. Soybean transformation is now possible in a range of genotypes from different maturity groups using different explants as sources of regenerable cells, various selectable marker genes and selective agents, and different A. tumefaciens strains. The cotyledonary-node method has been extensively investigated and across a number of laboratories yields on average greater than 1% transformation efficiency (one Southern-positive, independent event per 100 cotyledonary-node explants). Continued improvements in the cotyledonary-node method concomitant with further increases in transformation efficiency will enhance broader adoption of this already productive transformation method for use in crop improvement and functional genomics research efforts.

  1. Effect of Medium Supplements on Agrobacterium rhizogenes Mediated Hairy Root Induction from the Callus Tissues of Camellia sinensis var. sinensis

    PubMed Central

    Rana, Mohammad M.; Han, Zhuo-Xiao; Song, Da-Peng; Liu, Guo-Feng; Li, Da-Xiang; Wan, Xiao-Chun; Karthikeyan, Alagarsamy; Wei, Shu

    2016-01-01

    Tea (Camellia sinensis L.) is recalcitrant to Agrobacterium-mediated genetic transformation largely due to the bactericidal effects of tea polyphenols and phenolics oxidation induced by necrosis of explant tissue over the process of transformation. In this study, different antioxidants/adsorbents were added as supplements to the co-cultivation and post co-cultivation media to overcome these problems for the transformation improvement. Tea-cotyledon-derived calli were used as explants and Agrobacterium rhizognes strain ATCC 15834 was used as a mediator. Results showed that Agrobacterium growth, virulence (vir) gene expression and browning of explant tissue were greatly influenced by different supplements. Murashige and Skoog (MS) basal salts medium supplemented with 30 g·L−1 sucrose, 0.1 g·L−1 l-glutamine and 5 g·L−1 polyvinylpolypyrrolidone (PVPP) as co-cultivation and post co-cultivation media could maintain these parameters better that ultimately led to significant improvement of hairy root generation efficiency compared to that in the control (MS + 30 g·L−1 sucrose). Additionally, the reporter genes β-glucuronidase (gusA) and cyan fluorescent protein (cfp) were also stably expressed in the transgenic hairy roots. Our study would be helpful in establishing a feasible approach for tea biological studies and genetic improvement of tea varieties. PMID:27428960

  2. Effect of Medium Supplements on Agrobacterium rhizogenes Mediated Hairy Root Induction from the Callus Tissues of Camellia sinensis var. sinensis.

    PubMed

    Rana, Mohammad M; Han, Zhuo-Xiao; Song, Da-Peng; Liu, Guo-Feng; Li, Da-Xiang; Wan, Xiao-Chun; Karthikeyan, Alagarsamy; Wei, Shu

    2016-07-15

    Tea (Camellia sinensis L.) is recalcitrant to Agrobacterium-mediated genetic transformation largely due to the bactericidal effects of tea polyphenols and phenolics oxidation induced by necrosis of explant tissue over the process of transformation. In this study, different antioxidants/adsorbents were added as supplements to the co-cultivation and post co-cultivation media to overcome these problems for the transformation improvement. Tea-cotyledon-derived calli were used as explants and Agrobacterium rhizognes strain ATCC 15834 was used as a mediator. Results showed that Agrobacterium growth, virulence (vir) gene expression and browning of explant tissue were greatly influenced by different supplements. Murashige and Skoog (MS) basal salts medium supplemented with 30 g·L(-1) sucrose, 0.1 g·L(-1) l-glutamine and 5 g·L(-1) polyvinylpolypyrrolidone (PVPP) as co-cultivation and post co-cultivation media could maintain these parameters better that ultimately led to significant improvement of hairy root generation efficiency compared to that in the control (MS + 30 g·L(-1) sucrose). Additionally, the reporter genes β-glucuronidase (gusA) and cyan fluorescent protein (cfp) were also stably expressed in the transgenic hairy roots. Our study would be helpful in establishing a feasible approach for tea biological studies and genetic improvement of tea varieties.

  3. Renilla luciferase-based quantitation of Potato virus A infection initiated with Agrobacterium infiltration of N. benthamiana leaves.

    PubMed

    Eskelin, K; Suntio, T; Hyvärinen, S; Hafren, A; Mäkinen, K

    2010-03-01

    A quantitation method based on the sensitive detection of Renilla luciferase (Rluc) activity was developed and optimized for Potato virus A (PVA; genus Potyviridae) gene expression. This system is based on infections initiated by Agrobacterium infiltration and subsequent detection of the translation of PVA::Rluc RNA, which is enhanced by viral replication, first within the cells infected initially and later by translation and replication within new cells after spread of the virus. Firefly luciferase (Fluc) was used as an internal control to normalize the Rluc activity. An approximately 10-fold difference in the Rluc/Fluc activity ratio between a movement-deficient and a replication-deficient mutant was observed starting from 48h post Agrobacterium infiltration (h.p.i.). The Rluc activity derived from wild type (wt) PVA increased significantly between 48 and 72h.p.i. and the Rluc/Fluc activity deviated clearly from that of the mutant viruses. Quantitation of the Rluc and Fluc mRNAs by semi-quantitative RT-PCR indicated that increases and decreases in the Renillareniformis luciferase (rluc) mRNA levels coincided with changes in Rluc activity. However, a subtle increase in the mRNA level led to pronounced changes in Rluc activity. PVA CP accumulation was quantitated by enzyme-linked immunosorbent assay. The increase in Rluc activity correlated closely with virus accumulation. Copyright (c) 2009 Elsevier B.V. All rights reserved.

  4. Genetic transformation of carnation (Dianthus caryophylus L.).

    PubMed

    Nontaswatsri, Chalermsri; Fukai, Seiichi

    2010-01-01

    This chapter describes a rapid and efficient protocol for explant preparation and genetic transformation of carnation. Node explants from greenhouse-grown plants and leaf explants from in vitro plants are infected with Agrobacterium tumefaciens AGL0 harboring pKT3 plasmid, consisting of GUS and NPTII genes. Explant preparation is an important factor to obtain the transformed plants. The GUS-staining area was located only on the cut end of explants and only explants with a cut end close to the connecting area between node and leaf, produced transformed shoots. The cocultivation medium is also an important factor for the successful genetic transformation of carnation node and leaf explants. High genetic transformation efficiency of node and leaf explants cocultured with Agrobacterium tumefaciens was achieved when the explants were cocultivated on a filter paper soaked with water or water and acetosyringone mixture (AS).

  5. 3-D surface profilometry based on modulation measurement by applying wavelet transform method

    NASA Astrophysics Data System (ADS)

    Zhong, Min; Chen, Feng; Xiao, Chao; Wei, Yongchao

    2017-01-01

    A new analysis of 3-D surface profilometry based on modulation measurement technique by the application of Wavelet Transform method is proposed. As a tool excelling for its multi-resolution and localization in the time and frequency domains, Wavelet Transform method with good localized time-frequency analysis ability and effective de-noizing capacity can extract the modulation distribution more accurately than Fourier Transform method. Especially for the analysis of complex object, more details of the measured object can be well remained. In this paper, the theoretical derivation of Wavelet Transform method that obtains the modulation values from a captured fringe pattern is given. Both computer simulation and elementary experiment are used to show the validity of the proposed method by making a comparison with the results of Fourier Transform method. The results show that the Wavelet Transform method has a better performance than the Fourier Transform method in modulation values retrieval.

  6. Generation of marker-free transgenic hexaploid wheat via an Agrobacterium-mediated co-transformation strategy in commercial Chinese wheat varieties.

    PubMed

    Wang, Ke; Liu, Huiyun; Du, Lipu; Ye, Xingguo

    2017-05-01

    Genotype specificity is a big problem lagging the development of efficient hexaploid wheat transformation system. Increasingly, the biosecurity of genetically modified organisms is garnering public attention, so the generation of marker-free transgenic plants is very important to the eventual potential commercial release of transgenic wheat. In this study, 15 commercial Chinese hexaploid wheat varieties were successfully transformed via an Agrobacterium-mediated method, with efficiency of up to 37.7%, as confirmed by the use of Quickstix strips, histochemical staining, PCR analysis and Southern blotting. Of particular interest, marker-free transgenic wheat plants from various commercial Chinese varieties and their F 1 hybrids were successfully obtained for the first time, with a frequency of 4.3%, using a plasmid harbouring two independent T-DNA regions. The average co-integration frequency of the gus and the bar genes located on the two independent T-DNA regions was 49.0% in T 0 plants. We further found that the efficiency of generating marker-free plants was related to the number of bar gene copies integrated in the genome. Marker-free transgenic wheat plants were identified in the progeny of three transgenic lines that had only one or two bar gene copies. Moreover, silencing of the bar gene was detected in 30.7% of T 1 positive plants, but the gus gene was never found to be silenced in T 1 plants. Bisulphite genomic sequencing suggested that DNA methylation in the 35S promoter of the bar gene regulatory region might be the main reason for bar gene silencing in the transgenic plants. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  7. Setaria viridis floral-dip: A simple and rapid Agrobacterium-medicated transformation method

    USDA-ARS?s Scientific Manuscript database

    Setaria viridis was recently described as a new monocotyledonous model species for C4 photosynthesis research and genetic transformation. It has biological attributes (rapid life cycle, small genome, diploid, short stature and simple growth requirements) that make it suitable for use as a model plan...

  8. Rapid deletion plasmid construction methods for protoplast and Agrobacterium based fungal transformation systems

    USDA-ARS?s Scientific Manuscript database

    Increasing availability of genomic data and sophistication of analytical methodology in fungi has elevated the need for functional genomics tools in these organisms. Gene deletion is a critical tool for functional analysis. The targeted deletion of genes requires both a suitable method for the trans...

  9. Agrobacterium-delivered virulence protein VirE2 is trafficked inside host cells via a myosin XI-K–powered ER/actin network

    PubMed Central

    Yang, Qinghua; Li, Xiaoyang; Tu, Haitao; Pan, Shen Q.

    2017-01-01

    Agrobacterium tumefaciens causes crown gall tumors on various plants by delivering transferred DNA (T-DNA) and virulence proteins into host plant cells. Under laboratory conditions, the bacterium is widely used as a vector to genetically modify a wide range of organisms, including plants, yeasts, fungi, and algae. Various studies suggest that T-DNA is protected inside host cells by VirE2, one of the virulence proteins. However, it is not clear how Agrobacterium-delivered factors are trafficked through the cytoplasm. In this study, we monitored the movement of Agrobacterium-delivered VirE2 inside plant cells by using a split-GFP approach in real time. Agrobacterium-delivered VirE2 trafficked via the endoplasmic reticulum (ER) and F-actin network inside plant cells. During this process, VirE2 was aggregated as filamentous structures and was present on the cytosolic side of the ER. VirE2 movement was powered by myosin XI-K. Thus, exogenously produced and delivered VirE2 protein can use the endogenous host ER/actin network for movement inside host cells. The A. tumefaciens pathogen hijacks the conserved host infrastructure for virulence trafficking. Well-conserved infrastructure may be useful for Agrobacterium to target a wide range of recipient cells and achieve a high efficiency of transformation. PMID:28242680

  10. Evaluation on the effectiveness of 2-deoxyglucose-6-phosphate phosphatase (DOGR1) gene as a selectable marker for oil palm (Elaeis guineensis Jacq.) embryogenic calli transformation mediated by Agrobacterium tumefaciens

    PubMed Central

    Izawati, Abang Masli Dayang; Masani, Mat Yunus Abdul; Ismanizan, Ismail; Parveez, Ghulam Kadir Ahmad

    2015-01-01

    DOGR1, which encodes 2-deoxyglucose-6-phosphate phosphatase, has been used as a selectable marker gene to produce transgenic plants. In this study, a transformation vector, pBIDOG, which contains the DOGR1 gene, was transformed into oil palm embryogenic calli (EC) mediated by Agrobacterium tumefaciens strain LBA4404. Transformed EC were exposed to 400 mg l-1 2-deoxyglucose (2-DOG) as the selection agent. 2-DOG resistant tissues were regenerated into whole plantlets on various regeneration media containing the same concentration of 2-DOG. The plantlets were later transferred into soil and grown in a biosafety screenhouse. PCR and subsequently Southern blot analyses were carried out to confirm the integration of the transgene in the plantlets. A transformation efficiency of about 1.0% was obtained using DOGR1 gene into the genome of oil palm. This result demonstrates the potential of using combination of DOGR1 gene and 2-DOG for regenerating transgenic oil palm. PMID:26442041

  11. Establishment of an efficient transformation system for Pleurotus ostreatus.

    PubMed

    Lei, Min; Wu, Xiangli; Zhang, Jinxia; Wang, Hexiang; Huang, Chenyang

    2017-11-21

    Pleurotus ostreatus is widely cultivated worldwide, but the lack of an efficient transformation system regarding its use restricts its genetic research. The present study developed an improved and efficient Agrobacterium tumefaciens-mediated transformation method in P. ostreatus. Four parameters were optimized to obtain the most efficient transformation method. The strain LBA4404 was the most suitable for the transformation of P. ostreatus. A bacteria-to-protoplast ratio of 100:1, an acetosyringone (AS) concentration of 0.1 mM, and 18 h of co-culture showed the best transformation efficiency. The hygromycin B phosphotransferase gene (HPH) was used as the selective marker, and EGFP was used as the reporter gene in this study. Southern blot analysis combined with EGFP fluorescence assay showed positive results, and mitotic stability assay showed that more than 75% transformants were stable after five generations. These results showed that our transformation method is effective and stable and may facilitate future genetic studies in P. ostreatus.

  12. Chemical modification of L-glutamine to alpha-amino glutarimide on autoclaving facilitates Agrobacterium infection of host and non-host plants: A new use of a known compound

    PubMed Central

    2011-01-01

    Background Accidental autoclaving of L-glutamine was found to facilitate the Agrobacterium infection of a non host plant like tea in an earlier study. In the present communication, we elucidate the structural changes in L-glutamine due to autoclaving and also confirm the role of heat transformed L-glutamine in Agrobacterium mediated genetic transformation of host/non host plants. Results When autoclaved at 121°C and 15 psi for 20 or 40 min, L-glutamine was structurally modified into 5-oxo proline and 3-amino glutarimide (α-amino glutarimide), respectively. Of the two autoclaved products, only α-amino glutarimide facilitated Agrobacterium infection of a number of resistant to susceptible plants. However, the compound did not have any vir gene inducing property. Conclusions We report a one pot autoclave process for the synthesis of 5-oxo proline and α-amino glutarimide from L-glutamine. Xenobiotic detoxifying property of α-amino glutarimide is also proposed. PMID:21624145

  13. A Rapid Coordinate Transformation Method Applied in Industrial Robot Calibration Based on Characteristic Line Coincidence.

    PubMed

    Liu, Bailing; Zhang, Fumin; Qu, Xinghua; Shi, Xiaojia

    2016-02-18

    Coordinate transformation plays an indispensable role in industrial measurements, including photogrammetry, geodesy, laser 3-D measurement and robotics. The widely applied methods of coordinate transformation are generally based on solving the equations of point clouds. Despite the high accuracy, this might result in no solution due to the use of ill conditioned matrices. In this paper, a novel coordinate transformation method is proposed, not based on the equation solution but based on the geometric transformation. We construct characteristic lines to represent the coordinate systems. According to the space geometry relation, the characteristic line scan is made to coincide by a series of rotations and translations. The transformation matrix can be obtained using matrix transformation theory. Experiments are designed to compare the proposed method with other methods. The results show that the proposed method has the same high accuracy, but the operation is more convenient and flexible. A multi-sensor combined measurement system is also presented to improve the position accuracy of a robot with the calibration of the robot kinematic parameters. Experimental verification shows that the position accuracy of robot manipulator is improved by 45.8% with the proposed method and robot calibration.

  14. A Rapid Coordinate Transformation Method Applied in Industrial Robot Calibration Based on Characteristic Line Coincidence

    PubMed Central

    Liu, Bailing; Zhang, Fumin; Qu, Xinghua; Shi, Xiaojia

    2016-01-01

    Coordinate transformation plays an indispensable role in industrial measurements, including photogrammetry, geodesy, laser 3-D measurement and robotics. The widely applied methods of coordinate transformation are generally based on solving the equations of point clouds. Despite the high accuracy, this might result in no solution due to the use of ill conditioned matrices. In this paper, a novel coordinate transformation method is proposed, not based on the equation solution but based on the geometric transformation. We construct characteristic lines to represent the coordinate systems. According to the space geometry relation, the characteristic line scan is made to coincide by a series of rotations and translations. The transformation matrix can be obtained using matrix transformation theory. Experiments are designed to compare the proposed method with other methods. The results show that the proposed method has the same high accuracy, but the operation is more convenient and flexible. A multi-sensor combined measurement system is also presented to improve the position accuracy of a robot with the calibration of the robot kinematic parameters. Experimental verification shows that the position accuracy of robot manipulator is improved by 45.8% with the proposed method and robot calibration. PMID:26901203

  15. Expression of anti-tumor necrosis factor alpha (TNFα) single-chain variable fragment (scFv) in Spirodela punctata plants transformed with Agrobacterium tumefaciens.

    PubMed

    Balaji, Parthasarathy; Satheeshkumar, P K; Venkataraman, Krishnan; Vijayalakshmi, M A

    2016-05-01

    Therapeutic antibodies against tumor necrosis factor alpha (TNFα) have been considered effective for some of the autoimmune diseases such as rheumatoid arthritis, Crohn's diseases, and so on. But associated limitations of the current therapeutics in terms of cost, availability, and immunogenicity have necessitated the need for alternative candidates. Single-chain variable fragment (scFv) can negate the limitations tagged with the anti-TNFα therapeutics to a greater extent. In the present study, Spirodela punctata plants were transformed with anti-TNFα through in planta transformation using Agrobacterium tumefaciens strain, EHA105. Instead of cefotaxime, garlic extract (1 mg/mL) was used to remove the agrobacterial cells after cocultivation. To the best of our knowledge, this report shows for the first time the application of plant extracts in transgenic plant development. 95% of the plants survived screening under hygromycin. ScFv cDNA integration in the plant genomic DNA was confirmed at the molecular level by PCR. The transgenic protein expression was followed up to 10 months. Expression of scFv was confirmed by immunodot blot. Protein expression levels of up to 6.3% of total soluble protein were observed. β-Glucuronidase and green fluorescent protein expressions were also detected in the antibiotic resistant plants. The paper shows the generation of transgenic Spirodela punctuata plants through in planta transformation. © 2015 International Union of Biochemistry and Molecular Biology, Inc.

  16. Agrobacterium tumefaciens supports DNA replication of diverse geminivirus types.

    PubMed

    Selth, Luke A; Randles, John W; Rezaian, M Ali

    2002-04-10

    We have previously shown that the soil-borne plant pathogen Agrobacterium tumefaciens supports the replication of tomato leaf curl geminivirus (Australian isolate) (TLCV) DNA. However, the reproducibility of this observation with other geminiviruses has been questioned. Here, we show that replicative DNA forms of three other geminiviruses also accumulate at varying levels in Agrobacterium. Geminiviral DNA constructs that lacked the ability to replicate in Agrobacterium were rendered replication-competent by changing their configuration so that two copies of the viral ori were present. Furthermore, we report that low-level replication of TLCV DNA can occur in Escherichia coli containing a dimeric TLCV construct in a high copy number plasmid. These findings were reinforced by expression studies using beta-glucuronidase which revealed that all six TLCV promoters are active in Agrobacterium, and two are functional in E. coli.

  17. Method for hull-less barley transformation and manipulation of grain mixed-linkage beta-glucan.

    PubMed

    Lim, Wai Li; Collins, Helen M; Singh, Rohan R; Kibble, Natalie A J; Yap, Kuok; Taylor, Jillian; Fincher, Geoffrey B; Burton, Rachel A

    2018-05-01

    Hull-less barley is increasingly offering scope for breeding grains with improved characteristics for human nutrition; however, recalcitrance of hull-less cultivars to transformation has limited the use of these varieties. To overcome this limitation, we sought to develop an effective transformation system for hull-less barley using the cultivar Torrens. Torrens yielded a transformation efficiency of 1.8%, using a modified Agrobacterium transformation method. This method was used to over-express genes encoding synthases for the important dietary fiber component, (1,3;1,4)-β-glucan (mixed-linkage glucan), primarily present in starchy endosperm cell walls. Over-expression of the HvCslF6 gene, driven by an endosperm-specific promoter, produced lines where mixed-linkage glucan content increased on average by 45%, peaking at 70% in some lines, with smaller increases in transgenic HvCslH1 grain. Transgenic HvCslF6 lines displayed alterations where grain had a darker color, were more easily crushed than wild type and were smaller. This was associated with an enlarged cavity in the central endosperm and changes in cell morphology, including aleurone and sub-aleurone cells. This work provides proof-of-concept evidence that mixed-linkage glucan content in hull-less barley grain can be increased by over-expression of the HvCslF6 gene, but also indicates that hull-less cultivars may be more sensitive to attempts to modify cell wall composition. © 2017 Institute of Botany, Chinese Academy of Sciences.

  18. Nuclear transformation of eukaryotic microalgae: historical overview, achievements and problems.

    PubMed

    León, Rosa; Fernández, Emilio

    2007-01-01

    Transformation of microalgae is a first step in their use for biotechnological applications involving foreign protein production or molecular modifications of specific cell metabolic pathways. Since the first reliable achievements of nuclear transformation in Chlamydomonas, other eukaryotic microalgae have become transformed with molecular markers that allow a direct selection. Different methods--glass beads, electroporation, particle bombardment, or Agrobacterium--and constructions have been set up in several organisms and successfully used. However, some problems associated with efficiency, integration, or stability of the transgenes still persist and are analysed herein. Though the number of microalgae species successfully transformed is not very high, prospects for transformation of many more are good enough on the basis of what has been achieved so far.

  19. Development of a Plant Transformation Selection System Based on Expression of Genes Encoding Gentamicin Acetyltransferases

    PubMed Central

    Hayford, Maria B.; Medford, June I.; Hoffman, Nancy L.; Rogers, Stephen G.; Klee, Harry J.

    1988-01-01

    The development of selectable markers for transformation has been a major factor in the successful genetic manipulation of plants. A new selectable marker system has been developed based on bacterial gentamicin-3-N-acetyltransferases [AAC(3)]. These enzymes inactivate aminoglycoside antibiotics by acetylation. Two examples of AAC(3) enzymes have been manipulated to be expressed in plants. Chimeric AAC(3)-III and AAC(3)-IV genes were assembled using the constitutively expressed cauliflower mosaic virus 35S promoter and the nopaline synthase 3′ nontranslated region. These chimeric genes were engineered into vectors for Agrobacterium-mediated plant transformation. Petunia hybrida and Arabidopsis thaliana tissue transformed with these vectors grew in the presence of normally lethal levels of gentamicin. The transformed nature of regenerated Arabidopsis plants was confirmed by DNA hybridization analysis and inheritance of the selectable phenotype in progeny. The chimeric AAC(3)-IV gene has also been used to select transformants in several additional plant species. These results show that the bacterial AAC(3) genes will serve as useful selectable markers in plant tissue culture. Images Fig. 3 Fig. 4 Fig. 5 PMID:16666057

  20. Genome sequence of Ensifer adhaerens OV14 provides insights into its ability as a novel vector for the genetic transformation of plant genomes.

    PubMed

    Rudder, Steven; Doohan, Fiona; Creevey, Christopher J; Wendt, Toni; Mullins, Ewen

    2014-04-07

    Recently it has been shown that Ensifer adhaerens can be used as a plant transformation technology, transferring genes into several plant genomes when equipped with a Ti plasmid. For this study, we have sequenced the genome of Ensifer adhaerens OV14 (OV14) and compared it with those of Agrobacterium tumefaciens C58 (C58) and Sinorhizobium meliloti 1021 (1021); the latter of which has also demonstrated a capacity to genetically transform crop genomes, albeit at significantly reduced frequencies. The 7.7 Mb OV14 genome comprises two chromosomes and two plasmids. All protein coding regions in the OV14 genome were functionally grouped based on an eggNOG database. No genes homologous to the A. tumefaciens Ti plasmid vir genes appeared to be present in the OV14 genome. Unexpectedly, OV14 and 1021 were found to possess homologs to chromosomal based genes cited as essential to A. tumefaciens T-DNA transfer. Of significance, genes that are non-essential but exert a positive influence on virulence and the ability to genetically transform host genomes were identified in OV14 but were absent from the 1021 genome. This study reveals the presence of homologs to chromosomally based Agrobacterium genes that support T-DNA transfer within the genome of OV14 and other alphaproteobacteria. The sequencing and analysis of the OV14 genome increases our understanding of T-DNA transfer by non-Agrobacterium species and creates a platform for the continued improvement of Ensifer-mediated transformation (EMT).

  1. A new QRT-PCR assay designed for the differentiation between elements provided from Agrobacterium sp. in GMOs plant events and natural Agrobacterium sp. bacteria.

    PubMed

    Nabi, Nesrine; Chaouachi, Maher; Zellama, Mohamed Salem; Ben Hafsa, Ahmed; Mrabet, Besma; Saïd, Khaled; Fathia, Harzallah Skhiri

    2016-04-01

    The question asked in the present work was how to differentiate between contamination of field samples with and GM plants contained sequences provided from this bacterium in order to avoid false positives in the frame of the detection and the quantification of GMO. For this, new set of primers and corresponding TaqMan Minor Groove Binder (MGB) probes were designed to target Agrobacterium sp. using the tumor-morphology-shooty gene (TMS1). Final standard curves were calculated for each pathogen by plotting the threshold cycle value against the bacterial number (log (colony forming units) per milliliter) via linear regression. The method designed was highly specific and sensitive, with a detection limit of 10CFU/ml. No significant cross-reaction was observed. Results from this study showed that TaqMan real-time PCR, is potentially an effective method for the rapid and reliable quantification of Agrobacterium sp. in samples containing GMO or non GMO samples. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. Agrobacterium-derived cytokinin influences plastid morphology and starch accumulation in Nicotiana benthamiana during transient assays

    PubMed Central

    2014-01-01

    Background Agrobacterium tumefaciens-based transient assays have become a common tool for answering questions related to protein localization and gene expression in a cellular context. The use of these assays assumes that the transiently transformed cells are observed under relatively authentic physiological conditions and maintain ‘normal’ sub-cellular behaviour. Although this premise is widely accepted, the question of whether cellular organization and organelle morphology is altered in Agrobacterium-infiltrated cells has not been examined in detail. The first indications of an altered sub-cellular environment came from our observation that a common laboratory strain, GV3101(pMP90), caused a drastic increase in stromule frequency. Stromules, or ‘stroma-filled-tubules’ emanate from the surface of plastids and are sensitive to a variety of biotic and abiotic stresses. Starting from this observation, the goal of our experiments was to further characterize the changes to the cell resulting from short-term bacterial infestation, and to identify the factor responsible for eliciting these changes. Results Using a protocol typical of transient assays we evaluated the impact of GV3101(pMP90) infiltration on chloroplast behaviour and morphology in Nicotiana benthamiana. Our experiments confirmed that GV3101(pMP90) consistently induces stromules and alters plastid position relative to the nucleus. These effects were found to be the result of strain-dependant secretion of cytokinin and its accumulation in the plant tissue. Bacterial production of the hormone was found to be dependant on the presence of a trans-zeatin synthase gene (tzs) located on the Ti plasmid of GV3101(pMP90). Bacteria-derived cytokinins were also correlated with changes to both soluble sugar level and starch accumulation. Conclusion Although we have chosen to focus on how transient Agrobacterium infestation alters plastid based parameters, these changes to the morphology and position of a single

  3. Description of recovery method used for curdlan produced by Agrobacterium sp. IFO 13140 and its relation to the morphology and physicochemical and technological properties of the polysaccharide

    PubMed Central

    Mangolim, Camila Sampaio; da Silva, Thamara Thaiane; Fenelon, Vanderson Carvalho; Koga, Luciana Numata; Ferreira, Sabrina Barbosa de Souza; Bruschi, Marcos Luciano; Matioli, Graciette

    2017-01-01

    Curdlan is a linear polysaccharide considered a dietary fiber and with gelation properties. This study evaluated the structure, morphology and the physicochemical and technological properties of curdlan produced by Agrobacterium sp. IFO 13140 recovered by pre-gelation and precipitation methods. Commercial curdlan submitted or otherwise to the pre-gelation process was also evaluated. The data obtained from structural analysis revealed a similarity between the curdlan produced by Agrobacterium sp. IFO 13140 (recovered by both methods) and the commercial curdlans. The results showed that the curdlans evaluated differed significantly in terms of dispersibility and gelation, and only the pre-gelled ones had significant potential for food application, because this method influence on the size of the particles and in the presence of NaCl. In terms of technological properties, the curdlan produced by Agrobacterium sp. IFO 13140 (pre-gelation method) had a greater water and oil holding capacity (64% and 98% greater, respectively) and a greater thickening capacity than the pre-gelled commercial curdlan. The pre-gelled commercial curdlan displayed a greater gelling capacity at 95°C than the others. When applied to food, only the pre-gelled curdlans improved the texture parameters of yogurts and reduced syneresis. The curdlan gels, which are rigid and stable in structure, demonstrated potential for improving the texture of food products, with potential industrial use. PMID:28245244

  4. Description of recovery method used for curdlan produced by Agrobacterium sp. IFO 13140 and its relation to the morphology and physicochemical and technological properties of the polysaccharide.

    PubMed

    Mangolim, Camila Sampaio; Silva, Thamara Thaiane da; Fenelon, Vanderson Carvalho; Koga, Luciana Numata; Ferreira, Sabrina Barbosa de Souza; Bruschi, Marcos Luciano; Matioli, Graciette

    2017-01-01

    Curdlan is a linear polysaccharide considered a dietary fiber and with gelation properties. This study evaluated the structure, morphology and the physicochemical and technological properties of curdlan produced by Agrobacterium sp. IFO 13140 recovered by pre-gelation and precipitation methods. Commercial curdlan submitted or otherwise to the pre-gelation process was also evaluated. The data obtained from structural analysis revealed a similarity between the curdlan produced by Agrobacterium sp. IFO 13140 (recovered by both methods) and the commercial curdlans. The results showed that the curdlans evaluated differed significantly in terms of dispersibility and gelation, and only the pre-gelled ones had significant potential for food application, because this method influence on the size of the particles and in the presence of NaCl. In terms of technological properties, the curdlan produced by Agrobacterium sp. IFO 13140 (pre-gelation method) had a greater water and oil holding capacity (64% and 98% greater, respectively) and a greater thickening capacity than the pre-gelled commercial curdlan. The pre-gelled commercial curdlan displayed a greater gelling capacity at 95°C than the others. When applied to food, only the pre-gelled curdlans improved the texture parameters of yogurts and reduced syneresis. The curdlan gels, which are rigid and stable in structure, demonstrated potential for improving the texture of food products, with potential industrial use.

  5. 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.

  6. Research on the winding losses based on finite element method for transformer

    NASA Astrophysics Data System (ADS)

    Li, Wenpeng; Lai, Wenqing; Ye, Ligang; Luo, Hanwu; Luo, Changjiang; Cui, Shigang; Wang, Yongqiang

    2018-04-01

    Transformer loss can cause the transformer to overheat. Under the action of high frequency current, the loss of transformer windings will be aggravated due to proximity effect and skin effect. In this paper, a three-dimensional model of high frequency transformer windings is established. Considering of the proximity effect and skin effect, the eddy current effects loss in the transformer windings are simulated based on finite element method. And the winding losses of the transformer windings are obtained under different arrangements. The influence of the winding layout on the winding losses is given. Finally, the trend of winding loss with current frequency, winding thickness and inter layer spacing is obtained through calculation. The winding loss initially decreases as the thickness of the winding increases, but when it reaches a certain level, this reduction becomes insignificant.

  7. Application of succulent plant leaves for Agrobacterium infiltration-mediated protein production

    USDA-ARS?s Scientific Manuscript database

    Infiltration of tobacco leaves with a suspension of Agrobacterium tumefaciens harboring a binary plant expression plasmid provides a convenient method for laboratory scale protein production. When expressing plant cell wall degrading enzymes in the widely used tobacco (Nicotiana benthamiana), diffic...

  8. Overexpression of VpPR10.1 by an efficient transformation method enhances downy mildew resistance in V. vinifera.

    PubMed

    Su, Hang; Jiao, Yun-Tong; Wang, Fang-Fang; Liu, Yue-E; Niu, Wei-Li; Liu, Guo-Tian; Xu, Yan

    2018-05-01

    Putrescine and spermidine increase the transformation efficiency of Vitis vinifera L. cv. Thompson seedless. Accumulation of VpPR10.1 in transgenic V. vinifera Thompson seedless, likely increases its resistance to downy mildew. A more efficient method is described for facilitating Agrobacterium-mediated transformation of Vitis vinifera L. cv. Thompson Seedless somatic embryogenesis using polyamines (PAs). The efficacies of putrescine, spermidine and spermine are identified at a range of concentrations (10 µM, 100 µM and 1 mM) added to the culture medium during somatic embryo growth. Putrescine (PUT) and spermidine (SPD) promote the recovery of proembryonic masses (PEM) and the development of somatic embryos (SE) after co-cultivation. Judging from the importance of the time-frame in genetic transformation, PAs added at the co-cultivation stage have a stronger effect than delayed selection treatments, which are superior to antibiotic treatments in the selection stage. Best embryogenic responses are with 1 mM PUT and 100 µM SPD added to the co-culture medium. Using the above method, a pathogenesis-related gene (VpPR10.1) from Chinese wild Vitis pseudoreticulata was transferred into Thompson Seedless for functional evaluation. The transgenic line, confirmed by western blot analysis, was inoculated with Plasmopara viticola to test for downy mildew resistance. Based on observed restrictions of hyphal growth and increases in H 2 O 2 accumulation in the transgenic plants, the accumulation of VpPR10.1 likely enhanced the transgenic plants resistance to downy mildew.

  9. Missing texture reconstruction method based on error reduction algorithm using Fourier transform magnitude estimation scheme.

    PubMed

    Ogawa, Takahiro; Haseyama, Miki

    2013-03-01

    A missing texture reconstruction method based on an error reduction (ER) algorithm, including a novel estimation scheme of Fourier transform magnitudes is presented in this brief. In our method, Fourier transform magnitude is estimated for a target patch including missing areas, and the missing intensities are estimated by retrieving its phase based on the ER algorithm. Specifically, by monitoring errors converged in the ER algorithm, known patches whose Fourier transform magnitudes are similar to that of the target patch are selected from the target image. In the second approach, the Fourier transform magnitude of the target patch is estimated from those of the selected known patches and their corresponding errors. Consequently, by using the ER algorithm, we can estimate both the Fourier transform magnitudes and phases to reconstruct the missing areas.

  10. Estimation of phase derivatives using discrete chirp-Fourier-transform-based method.

    PubMed

    Gorthi, Sai Siva; Rastogi, Pramod

    2009-08-15

    Estimation of phase derivatives is an important task in many interferometric measurements in optical metrology. This Letter introduces a method based on discrete chirp-Fourier transform for accurate and direct estimation of phase derivatives, even in the presence of noise. The method is introduced in the context of the analysis of reconstructed interference fields in digital holographic interferometry. We present simulation and experimental results demonstrating the utility of the proposed method.

  11. Interpolating seismic data via the POCS method based on shearlet transform

    NASA Astrophysics Data System (ADS)

    Jicheng, Liu; Yongxin, Chou; Jianjiang, Zhu

    2018-06-01

    A method based on shearlet transform and the projection onto convex sets with L0-norm constraint is proposed to interpolate irregularly sampled 2D and 3D seismic data. The 2D directional filter of shearlet transform is constructed by modulating a low-pass diamond filter pair to minimize the effect of additional edges introduced by the missing traces. In order to abate the spatial aliasing and control the maximal gap between missing traces for a 3D data cube, a 2D separable jittered sampling strategy is discussed. Finally, numerical experiments on 2D and 3D synthetic and real data with different under-sampling rates prove the validity of the proposed method.

  12. Genome sequence of Ensifer adhaerens OV14 provides insights into its ability as a novel vector for the genetic transformation of plant genomes

    PubMed Central

    2014-01-01

    Background Recently it has been shown that Ensifer adhaerens can be used as a plant transformation technology, transferring genes into several plant genomes when equipped with a Ti plasmid. For this study, we have sequenced the genome of Ensifer adhaerens OV14 (OV14) and compared it with those of Agrobacterium tumefaciens C58 (C58) and Sinorhizobium meliloti 1021 (1021); the latter of which has also demonstrated a capacity to genetically transform crop genomes, albeit at significantly reduced frequencies. Results The 7.7 Mb OV14 genome comprises two chromosomes and two plasmids. All protein coding regions in the OV14 genome were functionally grouped based on an eggNOG database. No genes homologous to the A. tumefaciens Ti plasmid vir genes appeared to be present in the OV14 genome. Unexpectedly, OV14 and 1021 were found to possess homologs to chromosomal based genes cited as essential to A. tumefaciens T-DNA transfer. Of significance, genes that are non-essential but exert a positive influence on virulence and the ability to genetically transform host genomes were identified in OV14 but were absent from the 1021 genome. Conclusions This study reveals the presence of homologs to chromosomally based Agrobacterium genes that support T-DNA transfer within the genome of OV14 and other alphaproteobacteria. The sequencing and analysis of the OV14 genome increases our understanding of T-DNA transfer by non-Agrobacterium species and creates a platform for the continued improvement of Ensifer-mediated transformation (EMT). PMID:24708309

  13. [A wavelet-transform-based method for the automatic detection of late-type stars].

    PubMed

    Liu, Zhong-tian; Zhao, Rrui-zhen; Zhao, Yong-heng; Wu, Fu-chao

    2005-07-01

    The LAMOST project, the world largest sky survey project, urgently needs an automatic late-type stars detection system. However, to our knowledge, no effective methods for automatic late-type stars detection have been reported in the literature up to now. The present study work is intended to explore possible ways to deal with this issue. Here, by "late-type stars" we mean those stars with strong molecule absorption bands, including oxygen-rich M, L and T type stars and carbon-rich C stars. Based on experimental results, the authors find that after a wavelet transform with 5 scales on the late-type stars spectra, their frequency spectrum of the transformed coefficient on the 5th scale consistently manifests a unimodal distribution, and the energy of frequency spectrum is largely concentrated on a small neighborhood centered around the unique peak. However, for the spectra of other celestial bodies, the corresponding frequency spectrum is of multimodal and the energy of frequency spectrum is dispersible. Based on such a finding, the authors presented a wavelet-transform-based automatic late-type stars detection method. The proposed method is shown by extensive experiments to be practical and of good robustness.

  14. Remote Sensing Image Fusion Method Based on Nonsubsampled Shearlet Transform and Sparse Representation

    NASA Astrophysics Data System (ADS)

    Moonon, Altan-Ulzii; Hu, Jianwen; Li, Shutao

    2015-12-01

    The remote sensing image fusion is an important preprocessing technique in remote sensing image processing. In this paper, a remote sensing image fusion method based on the nonsubsampled shearlet transform (NSST) with sparse representation (SR) is proposed. Firstly, the low resolution multispectral (MS) image is upsampled and color space is transformed from Red-Green-Blue (RGB) to Intensity-Hue-Saturation (IHS). Then, the high resolution panchromatic (PAN) image and intensity component of MS image are decomposed by NSST to high and low frequency coefficients. The low frequency coefficients of PAN and the intensity component are fused by the SR with the learned dictionary. The high frequency coefficients of intensity component and PAN image are fused by local energy based fusion rule. Finally, the fused result is obtained by performing inverse NSST and inverse IHS transform. The experimental results on IKONOS and QuickBird satellites demonstrate that the proposed method provides better spectral quality and superior spatial information in the fused image than other remote sensing image fusion methods both in visual effect and object evaluation.

  15. Scale-Up of Agrobacterium rhizogenes-Mediated Hairy Root Cultures of Rauwolfia serpentina: A Persuasive Approach for Stable Reserpine Production.

    PubMed

    Mehrotra, Shakti; Srivastava, Vikas; Goel, Manoj K; Kukreja, Arun K

    2016-01-01

    Roots of Rauwolfia serpentina, also known as "Sarpagandha" possess high pharmaceutical value due to the presence of reserpine and other medicinally important terpene indole alkaloids. Ever increasing commercial demand of R. serpentina roots is the major reason behind the unsystematic harvesting and fast decline of the species from its natural environment. Considering Agrobacterium rhizogenes-mediated hairy root cultures as an alternative source for the production of plant-based secondary metabolites, the present optimized protocol offers a commercially feasible method for the production of reserpine, the most potent alkaloid from R. serpentina roots. This end-to-end protocol presents the establishment of hairy root culture from the leaf explants of R. serpentina through the infection of A. rhizogenes strain A4 in liquid B5 culture medium and its up-scaling in a 5 L bench top, mechanically agitated bioreactor. The transformed nature of roots was confirmed through PCR-based rol A gene amplification in genomic DNA of putative hairy roots. The extraction and quantification of reserpine in bioreactor grown roots has been done using monolithic reverse phase high-performance liquid chromatography (HPLC).

  16. “Agrolistic” transformation of plant cells: Integration of T-strands generated in planta

    PubMed Central

    Hansen, Geneviève; Chilton, Mary-Dell

    1996-01-01

    We describe a novel plant transformation technique, termed “agrolistic,” that combines the advantages of the Agrobacterium transformation system with the high efficiency of biolistic DNA delivery. Agrolistic transformation allows integration of the gene of interest without undesired vector sequence. The virulence genes virD1 and virD2 from Agrobacterium tumefaciens that are required in bacteria for excision of T-strands from the tumor-inducing plasmid were placed under the control of the CaMV35S promoter and codelivered with a target plasmid containing border sequences flanking the gene of interest. Transient expression assays in tobacco and in maize cells indicated that vir gene products caused strand-specific nicking in planta at the right border sequence, similar to VirD1/VirD2-catalyzed T-strand excision observed in Agrobacterium. Agrolistically transformed tobacco calli were obtained after codelivery of virD1 and virD2 genes together with a selectable marker flanked by border sequences. Some inserts exhibited right junctions with plant DNA that corresponded precisely to the sequence expected for T-DNA (portion of the tumor-inducing plasmid that is transferred to plant cells) insertion events. We designate these as “agrolistic” inserts, as distinguished from “biolistic” inserts. Both types of inserts were found in some transformed lines. The frequency of agrolistic inserts was 20% that of biolistic inserts. PMID:8962167

  17. Genetic transformation of black walnut (Juglans nigra)

    Treesearch

    Michael J. Bosela; Gurpreet S. Smagh; Charles H. Michler

    2004-01-01

    Disarmed Agrobacterium tumefaciens strains with binary vectors carrying transgenes for kanamycin resistance (npt II) and β-glucuronidase (GUS, uidA) were used for the genetic transformation of Eastern black walnut (Juglans nigra) somatic embryos. In total, explants from 16 embryo lines...

  18. A method of real-time fault diagnosis for power transformers based on vibration analysis

    NASA Astrophysics Data System (ADS)

    Hong, Kaixing; Huang, Hai; Zhou, Jianping; Shen, Yimin; Li, Yujie

    2015-11-01

    In this paper, a novel probability-based classification model is proposed for real-time fault detection of power transformers. First, the transformer vibration principle is introduced, and two effective feature extraction techniques are presented. Next, the details of the classification model based on support vector machine (SVM) are shown. The model also includes a binary decision tree (BDT) which divides transformers into different classes according to health state. The trained model produces posterior probabilities of membership to each predefined class for a tested vibration sample. During the experiments, the vibrations of transformers under different conditions are acquired, and the corresponding feature vectors are used to train the SVM classifiers. The effectiveness of this model is illustrated experimentally on typical in-service transformers. The consistency between the results of the proposed model and the actual condition of the test transformers indicates that the model can be used as a reliable method for transformer fault detection.

  19. A method of power analysis based on piecewise discrete Fourier transform

    NASA Astrophysics Data System (ADS)

    Xin, Miaomiao; Zhang, Yanchi; Xie, Da

    2018-04-01

    The paper analyzes the existing feature extraction methods. The characteristics of discrete Fourier transform and piecewise aggregation approximation are analyzed. Combining with the advantages of the two methods, a new piecewise discrete Fourier transform is proposed. And the method is used to analyze the lighting power of a large customer in this paper. The time series feature maps of four different cases are compared with the original data, discrete Fourier transform, piecewise aggregation approximation and piecewise discrete Fourier transform. This new method can reflect both the overall trend of electricity change and its internal changes in electrical analysis.

  20. Glass Bead-based Genetic Transformation:An Efficient Method for Transformation of Thraustochytrid Microorganisms.

    PubMed

    Adachi, Takumi; Sahara, Takehiko; Okuyama, Hidetoshi; Morita, Naoki

    2017-07-01

    Here, we describe a new method for genetic transformation of thraustochytrids, well-known producers of polyunsaturated fatty acids (PUFAs) like docosahexaenoic acid, by combining mild glass (zirconia) bead treatment and electroporation. Because the cell wall is a barrier against transfer of exogenous DNA into cells, gentle vortexing of cells with glass beads was performed prior to electroporation for partial cell wall disruption. G418-resistant transformants of thraustochytrid cells (Aurantiochytrium limacinum strain SR21 and thraustochytrid strain 12B) were successfully obtained with good reproducibility. The method reported here is simpler than methods using enzymes to generate spheroplasts and may provide advantages for PUFA production by using genetically modified thraustochytrids.

  1. Learning linear transformations between counting-based and prediction-based word embeddings

    PubMed Central

    Hayashi, Kohei; Kawarabayashi, Ken-ichi

    2017-01-01

    Despite the growing interest in prediction-based word embedding learning methods, it remains unclear as to how the vector spaces learnt by the prediction-based methods differ from that of the counting-based methods, or whether one can be transformed into the other. To study the relationship between counting-based and prediction-based embeddings, we propose a method for learning a linear transformation between two given sets of word embeddings. Our proposal contributes to the word embedding learning research in three ways: (a) we propose an efficient method to learn a linear transformation between two sets of word embeddings, (b) using the transformation learnt in (a), we empirically show that it is possible to predict distributed word embeddings for novel unseen words, and (c) empirically it is possible to linearly transform counting-based embeddings to prediction-based embeddings, for frequent words, different POS categories, and varying degrees of ambiguities. PMID:28926629

  2. Use of Ti plasmid DNA probes for determining tumorigenicity of agrobacterium strains

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Burr, T.J.; Norelli, J.L.; Katz, B.H.

    1990-06-01

    Probes consisting of T-DNA genes from the Ti plasmid of Agrobacterium tumefaciens were used for determining tumorigenicity of strains. Two {sup 32}P-labeled probes hybridized with 28 of 28 tumorigenic strains of the pathogen but not with 20 of 22 nontumorigenic strains. One probe, pTHE17, consists of all but the far left portion of the T-DNA of strain C58. Probe SmaI7 consists of SmaI fragment 7 of pTiC58, including onc genes 1, 4, and 6a and most of 2. Another probe, pAL4044, consisting of the vir region of strain Ach-5, hybridized with several nontumorigenic as well as tumorigenic strains. Colony hybridizationsmore » were done with 28 tumorigenic and 22 nontumorigenic Agrobacterium strains. About 10{sup 6} CFU of the different tumorigenic strains were detectable with this method. Southern analyses confirmed the presence or absence of Ti plasmids in strains for which tumorigenicity was questioned. Colony hybridization with the T-DNA probes provides a rapid and sensitive means for determining the tumorigenic nature of Agrobacterium strains.« less

  3. Surface plasmon resonance imaging reveals multiple binding modes of Agrobacterium transformation mediator VirE2 to ssDNA.

    PubMed

    Kim, Sanghyun; Zbaida, David; Elbaum, Michael; Leh, Hervé; Nogues, Claude; Buckle, Malcolm

    2015-07-27

    VirE2 is the major secreted protein of Agrobacterium tumefaciens in its genetic transformation of plant hosts. It is co-expressed with a small acidic chaperone VirE1, which prevents VirE2 oligomerization. After secretion into the host cell, VirE2 serves functions similar to a viral capsid in protecting the single-stranded transferred DNA en route to the nucleus. Binding of VirE2 to ssDNA is strongly cooperative and depends moreover on protein-protein interactions. In order to isolate the protein-DNA interactions, imaging surface plasmon resonance (SPRi) studies were conducted using surface-immobilized DNA substrates of length comparable to the protein-binding footprint. Binding curves revealed an important influence of substrate rigidity with a notable preference for poly-T sequences and absence of binding to both poly-A and double-stranded DNA fragments. Dissociation at high salt concentration confirmed the electrostatic nature of the interaction. VirE1-VirE2 heterodimers also bound to ssDNA, though by a different mechanism that was insensitive to high salt. Neither VirE2 nor VirE1-VirE2 followed the Langmuir isotherm expected for reversible monomeric binding. The differences reflect the cooperative self-interactions of VirE2 that are suppressed by VirE1. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  4. T-DNA transfer and T-DNA integration efficiencies upon Arabidopsis thaliana root explant cocultivation and floral dip transformation.

    PubMed

    Ghedira, Rim; De Buck, Sylvie; Van Ex, Frédéric; Angenon, Geert; Depicker, Ann

    2013-12-01

    T-DNA transfer and integration frequencies during Agrobacterium-mediated root explant cocultivation and floral dip transformations of Arabidopsis thaliana were analyzed with and without selection for transformation-competent cells. Based on the presence or absence of CRE recombinase activity without or with the CRE T-DNA being integrated, transient expression versus stable transformation was differentiated. During root explant cocultivation, continuous light enhanced the number of plant cells competent for interaction with Agrobacterium and thus the number of transient gene expression events. However, in transformation competent plant cells, continuous light did not further enhance cotransfer or cointegration frequencies. Upon selection for root transformants expressing a first T-DNA, 43-69 % of these transformants showed cotransfer of another non-selected T-DNA in two different light regimes. However, integration of the non-selected cotransferred T-DNA occurred only in 19-46 % of these transformants, indicating that T-DNA integration in regenerating root cells limits the transformation frequencies. After floral dip transformation, transient T-DNA expression without integration could not be detected, while stable T-DNA transformation occurred in 0.5-1.3 % of the T1 seedlings. Upon selection for floral dip transformants with a first T-DNA, 8-34 % of the transformants showed cotransfer of the other non-selected T-DNA and in 93-100 % of them, the T-DNA was also integrated. Therefore, a productive interaction between the agrobacteria and the female gametophyte, rather than the T-DNA integration process, restricts the floral dip transformation frequencies.

  5. Transformation of PRT6 RNAi construct into tomato (Solanum lycopersicum) cv. Micro-Tom

    NASA Astrophysics Data System (ADS)

    Suka, Intan Elya; Chew, Bee Lynn; Goh, Hoe-Han; Isa, Nurulhikma Md

    2018-04-01

    PROTEOLYSIS 6 plays major role in the N-end rule pathway as N-recognin which functions as E3 ligase enzyme. It mediates ubiquitin processes that lead to degradation of unstable substrate protein. The aim of the current study is to transform the PRT6 gene into tomato (Solanum lycopersicum) from the cultivar Micro-Tom and to investigate its function in regulating ripening in tomato fruits. The PRT6_RNAi construct was successfully transformed into Agrobacterium C58 via heat shock method and transformed into seven days old cotyledon explants. Factors affecting transformation efficiency such as co-cultivation time and type of plant growth regulator combination were evaluated. Results from this study found that pre-cultured cotyledons from seven days old seedlings incubated for 2 days in co-cultivation medium increased shoot regeneration. Plant growth hormones zeatin combine with auxin produced a higher number of callus formation but lower shoot proliferation and transformation frequency compared to treatments of single plant hormone in the selection medium. Polymerase chain reaction (PCR) was performed on the regenerated shoots to confirm the integration of PRT6 fragment into the genome of transgenic plants. Based on PCR analysis, all putative shoots were positive transformants.

  6. The Coordinate Transformation Method of High Resolution dem Data

    NASA Astrophysics Data System (ADS)

    Yan, Chaode; Guo, Wang; Li, Aimin

    2018-04-01

    Coordinate transformation methods of DEM data can be divided into two categories. One reconstruct based on original vector elevation data. The other transforms DEM data blocks by transforming parameters. But the former doesn't work in the absence of original vector data, and the later may cause errors at joint places between adjoining blocks of high resolution DEM data. In view of this problem, a method dealing with high resolution DEM data coordinate transformation is proposed. The method transforms DEM data into discrete vector elevation points, and then adjusts positions of points by bi-linear interpolation respectively. Finally, a TIN is generated by transformed points, and the new DEM data in target coordinate system is reconstructed based on TIN. An algorithm which can find blocks and transform automatically is given in this paper. The method is tested in different terrains and proved to be feasible and valid.

  7. Agrobacterium-transformed rice plants expressing synthetic cryIA(b) and cryIA(c) genes are highly toxic to striped stem borer and yellow stem borer.

    PubMed

    Cheng, X; Sardana, R; Kaplan, H; Altosaar, I

    1998-03-17

    Over 2,600 transgenic rice plants in nine strains were regenerated from >500 independently selected hygromycin-resistant calli after Agrobacterium-mediated transformation. The plants were transformed with fully modified (plant codon optimized) versions of two synthetic cryIA(b) and cryIA(c) coding sequences from Bacillus thuringiensis as well as the hph and gus genes, coding for hygromycin phosphotransferase and beta-glucuronidase, respectively. These sequences were placed under control of the maize ubiquitin promoter, the CaMV35S promoter, and the Brassica Bp10 gene promoter to achieve high and tissue-specific expression of the lepidopteran-specific delta-endotoxins. The integration, expression, and inheritance of these genes were demonstrated in R0 and R1 generations by Southern, Northern, and Western analyses and by other techniques. Accumulation of high levels (up to 3% of soluble proteins) of CryIA(b) and CryIA(c) proteins was detected in R0 plants. Bioassays with R1 transgenic plants indicated that the transgenic plants were highly toxic to two major rice insect pests, striped stem borer (Chilo suppressalis) and yellow stem borer (Scirpophaga incertulas), with mortalities of 97-100% within 5 days after infestation, thus offering a potential for effective insect resistance in transgenic rice plants.

  8. Identification of genes associated with asexual reproduction in Phyllosticta citricarpa mutants obtained through Agrobacterium tumefaciens transformation.

    PubMed

    Goulin, Eduardo Henrique; Savi, Daiani Cristina; Petters, Desirrê Alexia Lourenço; Kava, Vanessa; Galli-Terasawa, Lygia; Silva, Geraldo José; Glienke, Chirlei

    2016-11-01

    Phyllosticta citricarpa is the epidemiological agent of Citrus Black Spot (CBS) disease, which is responsible for large economic losses worldwide. CBS is characterized by the presence of spores (pycnidiospores) in dark lesions of fruit, which are also responsible for short distance dispersal of the disease. The identification of genes involved in asexual reproduction of P. citricarpa can be an alternative for directional disease control. We analyzed a library of mutants obtained through Agrobacterium tumefaciens transformation system, looking for alterations in growth and reproductive structure formation. Two mutant strains were found to have lost the ability to form pycnidia. The flanking T-DNA insertion regions were identified on P. citricarpa genome by using blast analysis and further gene prediction. The predicted genes containing the T-DNA insertions were identified as Spindle Poison Sensitivity Scp3, Ion Transport protein, and Cullin Binding proteins. The Ion Transport and Cullin Binding proteins are known to be correlated with sexual and asexual reproduction in fungi; however, the exact mechanism by which these proteins act on spore formation in P. citricarpa needs to be better characterized. The Scp3 proteins are suggested here for the first time as being associated with asexual reproduction in fungus. This protein is associated with microtubule formation, and as microtubules play an essential role as spindle machinery for chromosome segregation and cytokinesis, insertions in this gene can lead to abnormal formations, such as that observed here in P. citricarpa. We suggest these genes as new targets for fungicide development and CBS disease control, by iRNA. Copyright © 2016 Elsevier GmbH. All rights reserved.

  9. Improved full analytical polygon-based method using Fourier analysis of the three-dimensional affine transformation.

    PubMed

    Pan, Yijie; Wang, Yongtian; Liu, Juan; Li, Xin; Jia, Jia

    2014-03-01

    Previous research [Appl. Opt.52, A290 (2013)] has revealed that Fourier analysis of three-dimensional affine transformation theory can be used to improve the computation speed of the traditional polygon-based method. In this paper, we continue our research and propose an improved full analytical polygon-based method developed upon this theory. Vertex vectors of primitive and arbitrary triangles and the pseudo-inverse matrix were used to obtain an affine transformation matrix representing the spatial relationship between the two triangles. With this relationship and the primitive spectrum, we analytically obtained the spectrum of the arbitrary triangle. This algorithm discards low-level angular dependent computations. In order to add diffusive reflection to each arbitrary surface, we also propose a whole matrix computation approach that takes advantage of the affine transformation matrix and uses matrix multiplication to calculate shifting parameters of similar sub-polygons. The proposed method improves hologram computation speed for the conventional full analytical approach. Optical experimental results are demonstrated which prove that the proposed method can effectively reconstruct three-dimensional scenes.

  10. Agrobacterium and Tumor Induction: A Model System.

    ERIC Educational Resources Information Center

    Lennox, John E.

    1980-01-01

    The author offers laboratory procedures for experiments using the bacterium, Agrobacterium tumefaciens, which causes crown gall disease in a large number of plants. Three different approaches to growing a culture are given. (SA)

  11. Effective elimination of chimeric tissue in transgenics for the stable genetic transformation of lesquerella fendleri

    USDA-ARS?s Scientific Manuscript database

    In order to improve the potential of Lesquerella fendleri as a valuable industrial oilseed crop, a stable genetic transformation system was developed. Genetic transformation was performed by inoculating leaf segments with an Agrobacterium tumefaciens strain AGL1 carrying binary vector pCAMBIA 1301.1...

  12. SSAW: A new sequence similarity analysis method based on the stationary discrete wavelet transform.

    PubMed

    Lin, Jie; Wei, Jing; Adjeroh, Donald; Jiang, Bing-Hua; Jiang, Yue

    2018-05-02

    Alignment-free sequence similarity analysis methods often lead to significant savings in computational time over alignment-based counterparts. A new alignment-free sequence similarity analysis method, called SSAW is proposed. SSAW stands for Sequence Similarity Analysis using the Stationary Discrete Wavelet Transform (SDWT). It extracts k-mers from a sequence, then maps each k-mer to a complex number field. Then, the series of complex numbers formed are transformed into feature vectors using the stationary discrete wavelet transform. After these steps, the original sequence is turned into a feature vector with numeric values, which can then be used for clustering and/or classification. Using two different types of applications, namely, clustering and classification, we compared SSAW against the the-state-of-the-art alignment free sequence analysis methods. SSAW demonstrates competitive or superior performance in terms of standard indicators, such as accuracy, F-score, precision, and recall. The running time was significantly better in most cases. These make SSAW a suitable method for sequence analysis, especially, given the rapidly increasing volumes of sequence data required by most modern applications.

  13. Effect of Different Agrobacterium rhizogenes Strains on Hairy Root Induction and Phenylpropanoid Biosynthesis in Tartary Buckwheat (Fagopyrum tataricum Gaertn)

    PubMed Central

    Thwe, Aye; Valan Arasu, Mariadhas; Li, Xiaohua; Park, Chang Ha; Kim, Sun Ju; Al-Dhabi, Naif Abdullah; Park, Sang Un

    2016-01-01

    The development of an efficient protocol for successful hairy root induction by Agrobacterium rhizogenes is the key step toward an in vitro culturing method for the mass production of secondary metabolites. The selection of an effective Agrobacterium strain for the production of hairy roots is highly plant species dependent and must be determined empirically. Therefore, our goal was to investigate the transformation efficiency of different A. rhizogenes strains for the induction of transgenic hairy roots in Fagopyrum tataricum ‘Hokkai T10’ cultivar; to determine the expression levels of the polypropanoid biosynthetic pathway genes, such as ftpAL, FtC4H, Ft4CL, FrCHS, FrCH1, FrF3H, FtFLS1, FtFLS2, FtF3, H1, FtF3′H2, FtANS, and FtDFR; and to quantify the in vitro synthesis of phenolic compounds and anthocyanins. Among different strains, R1000 was the most promising candidate for hairy root stimulation because it induced the highest growth rate, root number, root length, transformation efficiency, and total anthocyanin and rutin content. The R1000, 15834, and A4 strains provided higher transcript levels for most metabolic pathway genes for the synthesis of rutin (22.31, 15.48, and 13.04 μg/mg DW, respectively), cyanidin 3-O-glucoside (800, 750, and 650 μg/g DW, respectively), and cyanidin 3-O-rutinoside (2410, 1530, and 1170 μg/g DW, respectively). A suitable A. rhizogenes strain could play a vital role in the fast growth of the bulk amount of hairy roots and secondary metabolites. Overall, R1000 was the most promising strain for hairy root induction in buckwheat. PMID:27014239

  14. Inactivation of the indole-diterpene biosynthetic gene cluster of Claviceps paspali by Agrobacterium-mediated gene replacement.

    PubMed

    Kozák, László; Szilágyi, Zoltán; Vágó, Barbara; Kakuk, Annamária; Tóth, László; Molnár, István; Pócsi, István

    2018-04-01

    The hypocrealean fungus Claviceps paspali is a parasite of wild grasses. This fungus is widely utilized in the pharmaceutical industry for the manufacture of ergot alkaloids, but also produces tremorgenic and neurotoxic indole-diterpene (IDT) secondary metabolites such as paspalitrems A and B. IDTs cause significant losses in agriculture and represent health hazards that threaten food security. Conversely, IDTs may also be utilized as lead compounds for pharmaceutical drug discovery. Current protoplast-mediated transformation protocols of C. paspali are inadequate as they suffer from inefficiencies in protoplast regeneration, a low frequency of DNA integration, and a low mitotic stability of the nascent transformants. We adapted and optimized Agrobacterium tumefaciens-mediated transformation (ATMT) for C. paspali and validated this method with the straightforward creation of a mutant strain of this fungus featuring a targeted replacement of key genes in the putative IDT biosynthetic gene cluster. Complete abrogation of IDT production in isolates of the mutant strain proved the predicted involvement of the target genes in the biosynthesis of IDTs. The mutant isolates continued to produce ergot alkaloids undisturbed, indicating that equivalent mutants generated in industrial ergot producers may have a better safety profile as they are devoid of IDT-type mycotoxins. Meanwhile, ATMT optimized for Claviceps spp. may open the door for the facile genetic engineering of these industrially and ecologically important organisms.

  15. [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.

  16. Comprehensive reliability allocation method for CNC lathes based on cubic transformed functions of failure mode and effects analysis

    NASA Astrophysics Data System (ADS)

    Yang, Zhou; Zhu, Yunpeng; Ren, Hongrui; Zhang, Yimin

    2015-03-01

    Reliability allocation of computerized numerical controlled(CNC) lathes is very important in industry. Traditional allocation methods only focus on high-failure rate components rather than moderate failure rate components, which is not applicable in some conditions. Aiming at solving the problem of CNC lathes reliability allocating, a comprehensive reliability allocation method based on cubic transformed functions of failure modes and effects analysis(FMEA) is presented. Firstly, conventional reliability allocation methods are introduced. Then the limitations of direct combination of comprehensive allocation method with the exponential transformed FMEA method are investigated. Subsequently, a cubic transformed function is established in order to overcome these limitations. Properties of the new transformed functions are discussed by considering the failure severity and the failure occurrence. Designers can choose appropriate transform amplitudes according to their requirements. Finally, a CNC lathe and a spindle system are used as an example to verify the new allocation method. Seven criteria are considered to compare the results of the new method with traditional methods. The allocation results indicate that the new method is more flexible than traditional methods. By employing the new cubic transformed function, the method covers a wider range of problems in CNC reliability allocation without losing the advantages of traditional methods.

  17. Advancing Crop Transformation in the Era of Genome Editing.

    PubMed

    Altpeter, Fredy; Springer, Nathan M; Bartley, Laura E; Blechl, Ann E; Brutnell, Thomas P; Citovsky, Vitaly; Conrad, Liza J; Gelvin, Stanton B; Jackson, David P; Kausch, Albert P; Lemaux, Peggy G; Medford, June I; Orozco-Cárdenas, Martha L; Tricoli, David M; Van Eck, Joyce; Voytas, Daniel F; Walbot, Virginia; Wang, Kan; Zhang, Zhanyuan J; Stewart, C Neal

    2016-07-01

    Plant transformation has enabled fundamental insights into plant biology and revolutionized commercial agriculture. Unfortunately, for most crops, transformation and regeneration remain arduous even after more than 30 years of technological advances. Genome editing provides novel opportunities to enhance crop productivity but relies on genetic transformation and plant regeneration, which are bottlenecks in the process. Here, we review the state of plant transformation and point to innovations needed to enable genome editing in crops. Plant tissue culture methods need optimization and simplification for efficiency and minimization of time in culture. Currently, specialized facilities exist for crop transformation. Single-cell and robotic techniques should be developed for high-throughput genomic screens. Plant genes involved in developmental reprogramming, wound response, and/or homologous recombination should be used to boost the recovery of transformed plants. Engineering universal Agrobacterium tumefaciens strains and recruiting other microbes, such as Ensifer or Rhizobium, could facilitate delivery of DNA and proteins into plant cells. Synthetic biology should be employed for de novo design of transformation systems. Genome editing is a potential game-changer in crop genetics when plant transformation systems are optimized. © 2016 American Society of Plant Biologists. All rights reserved.

  18. Establishment of an efficient plant regeneration culture protocol and achievement of successful genetic transformation in Jatropha curcas L.

    PubMed

    Liu, Ying; Liu, Guoxuan; Yang, Yali; Niu, Sufang; Yang, Fuguang; Yang, Shaoxia; Tang, Jianian; Chen, Jianping

    2017-12-01

    An efficient and reproducible protocol is described for shoot-bud regeneration and Agrobacterium tumefaciens-mediated genetic transformation of J. curcas. Treating the explants with high concentrations (5-120 mg/L) of TDZ for short durations (5-80 min) before inoculation culture increased significantly the regeneration frequency and improved the quality of the regenerated buds. The highest shoot-buds induction rate (87.35%) was achieved when petiole explants were treated with 20 mg/L TDZ solution for 20 min and inoculated on hormone-free MS medium for 30 days. Regenerated shoots of 0.5 cm or a little longer were isolated and grafted to seedling stocks of the same species, and then the grafted plantlets were planted on half-strength MS medium containing 0.1 mg/L IBA and 2 mg/L sodium nitroprusside (SNP). This grafting strategy was found to be very effective, to obtain that healthy grafted plantlets ready for acclimatization within 20 days. By the above mentioned protocol and with general Agrobacterium - mediated genetic transformation methods only 65 days were needed to obtain intact transgenic plants.

  19. Efficient and heritable transformation of Phalaenopsis orchids.

    PubMed

    Hsing, Hong-Xian; Lin, Yi-Jyun; Tong, Chii-Gong; Li, Min-Jeng; Chen, Yun-Jin; Ko, Swee-Suak

    2016-12-01

    Phalaenopsis orchid (Phal. orchid) is visually attractive and it is important economic floriculture species. Phal. orchids have many unique biological features. However, investigation of these features and validation on their biological functions are limited due to the lack of an efficient transformation method. We developed a heritable and efficient Agrobacterium- mediated transformation using protocorms derived from tetraploid or diploid Phal. orchids. A T-DNA vector construct containing eGFP driven by ubiquitin promoter was subjected to transformation. An approximate 1.2-5.2 % transformation rate was achieved. Genomic PCR confirmed that hygromycin selection marker, HptII gene and target gene eGFP were integrated into the orchid genome. Southern blotting indicated a low T-DNA insertion number in the orchid genome of the transformants. Western blot confirmed the expression of eGFP protein in the transgenic orchids. Furthermore, the GFP signal was detected in the transgenic orchids under microscopy. After backcrossing the pollinia of the transgenic plants to four different Phal. orchid varieties, the BC1 progenies showed hygromycin resistance and all surviving BC1 seedlings were HptII positive in PCR and expressed GFP protein as shown by western blot. This study demonstrated a stable transformation system was generated for Phal. orchids. This useful transformation protocol enables functional genomics studies and molecular breeding.

  20. An Agrobacterium-delivered CRISPR/Cas9 system for high-frequency targeted mutagenesis in maize.

    PubMed

    Char, Si Nian; Neelakandan, Anjanasree K; Nahampun, Hartinio; Frame, Bronwyn; Main, Marcy; Spalding, Martin H; Becraft, Philip W; Meyers, Blake C; Walbot, Virginia; Wang, Kan; Yang, Bing

    2017-02-01

    CRISPR/Cas9 is a powerful genome editing tool in many organisms, including a number of monocots and dicots. Although the design and application of CRISPR/Cas9 is simpler compared to other nuclease-based genome editing tools, optimization requires the consideration of the DNA delivery and tissue regeneration methods for a particular species to achieve accuracy and efficiency. Here, we describe a public sector system, ISU Maize CRISPR, utilizing Agrobacterium-delivered CRISPR/Cas9 for high-frequency targeted mutagenesis in maize. This system consists of an Escherichia coli cloning vector and an Agrobacterium binary vector. It can be used to clone up to four guide RNAs for single or multiplex gene targeting. We evaluated this system for its mutagenesis frequency and heritability using four maize genes in two duplicated pairs: Argonaute 18 (ZmAgo18a and ZmAgo18b) and dihydroflavonol 4-reductase or anthocyaninless genes (a1 and a4). T 0 transgenic events carrying mono- or diallelic mutations of one locus and various combinations of allelic mutations of two loci occurred at rates over 70% mutants per transgenic events in both Hi-II and B104 genotypes. Through genetic segregation, null segregants carrying only the desired mutant alleles without the CRISPR transgene could be generated in T 1 progeny. Inheritance of an active CRISPR/Cas9 transgene leads to additional target-specific mutations in subsequent generations. Duplex infection of immature embryos by mixing two individual Agrobacterium strains harbouring different Cas9/gRNA modules can be performed for improved cost efficiency. Together, the findings demonstrate that the ISU Maize CRISPR platform is an effective and robust tool to targeted mutagenesis in maize. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  1. A method based on IHS cylindrical transform model for quality assessment of image fusion

    NASA Astrophysics Data System (ADS)

    Zhu, Xiaokun; Jia, Yonghong

    2005-10-01

    Image fusion technique has been widely applied to remote sensing image analysis and processing, and methods for quality assessment of image fusion in remote sensing have also become the research issues at home and abroad. Traditional assessment methods combine calculation of quantitative indexes and visual interpretation to compare fused images quantificationally and qualitatively. However, in the existing assessment methods, there are two defects: on one hand, most imdexes lack the theoretic support to compare different fusion methods. On the hand, there is not a uniform preference for most of the quantitative assessment indexes when they are applied to estimate the fusion effects. That is, the spatial resolution and spectral feature could not be analyzed synchronously by these indexes and there is not a general method to unify the spatial and spectral feature assessment. So in this paper, on the basis of the approximate general model of four traditional fusion methods, including Intensity Hue Saturation(IHS) triangle transform fusion, High Pass Filter(HPF) fusion, Principal Component Analysis(PCA) fusion, Wavelet Transform(WT) fusion, a correlation coefficient assessment method based on IHS cylindrical transform is proposed. By experiments, this method can not only get the evaluation results of spatial and spectral features on the basis of uniform preference, but also can acquire the comparison between fusion image sources and fused images, and acquire differences among fusion methods. Compared with the traditional assessment methods, the new methods is more intuitionistic, and in accord with subjective estimation.

  2. [Genetic transformation of flax (Linum usitatissimum L.) with chimeric GFP-TUA6 gene for visualisation of microtubules].

    PubMed

    Shisha, E N; Korkhovoĭ, V I; Baer, G Ia; Guzenko, E V; Lemesh, V A; Kartel', N A; Emets, A I; Blium, Ia B

    2013-01-01

    The data of Agrobacterium-mediated transformation of some Linum usitatissimum cultivars zoned on the territories of Belarus and Ukraine with the plasmid carrying chimeric GFP-TUA6 gene and nptII gene as selectable marker conferring resistance to kanamycin are presented in this study. Transformation was affected by a number of factors including optical density (OD600), time of inoculation of explants with Agrobacterium and co-culture conditions. Transgenic nature of obtained lines was confirmed by PCR analysis. Expression of GFP-TUA6 gene was detected with confocal laser scanning microscopy. The obtained transgenic lines can be used for further functional studies the role of microtubules in the processes of building the flax fibres and resistance to wind.

  3. Combined chemical and physical transformation method with RbCl and sepiolite for the transformation of various bacterial species.

    PubMed

    Ren, Jun; Lee, Haram; Yoo, Seung Min; Yu, Myeong-Sang; Park, Hansoo; Na, Dokyun

    2017-04-01

    DNA transformation that delivers plasmid DNAs into bacterial cells is fundamental in genetic manipulation to engineer and study bacteria. Developed transformation methods to date are optimized to specific bacterial species for high efficiency. Thus, there is always a demand for simple and species-independent transformation methods. We herein describe the development of a chemico-physical transformation method that combines a rubidium chloride (RbCl)-based chemical method and sepiolite-based physical method, and report its use for the simple and efficient delivery of DNA into various bacterial species. Using this method, the best transformation efficiency for Escherichia coli DH5α was 4.3×10 6 CFU/μg of pUC19 plasmid, which is higher than or comparable to the reported transformation efficiencies to date. This method also allowed the introduction of plasmid DNAs into Bacillus subtilis (5.7×10 3 CFU/μg of pSEVA3b67Rb), Bacillus megaterium (2.5×10 3 CFU/μg of pSPAsp-hp), Lactococcus lactis subsp. lactis (1.0×10 2 CFU/μg of pTRKH3-ermGFP), and Lactococcus lactis subsp. cremoris (2.2×10 2 CFU/μg of pMSP3535VA). Remarkably, even when the conventional chemical and physical methods failed to generate transformed cells in Bacillus sp. and Enterococcus faecalis, E. malodoratus and E. mundtii, our combined method showed a significant transformation efficiency (2.4×10 4 , 4.5×10 2 , 2×10 1 , and 0.5×10 1 CFU/μg of plasmid DNA). Based on our results, we anticipate that our simple and efficient transformation method should prove usefulness for introducing DNA into various bacterial species without complicated optimization of parameters affecting DNA entry into the cell. Copyright © 2017. Published by Elsevier B.V.

  4. Advancing Crop Transformation in the Era of Genome Editing[OPEN

    PubMed Central

    Blechl, Ann E.; Brutnell, Thomas P.; Conrad, Liza J.; Gelvin, Stanton B.; Jackson, David P.; Kausch, Albert P.; Lemaux, Peggy G.; Medford, June I.; Orozco-Cárdenas, Martha L.; Tricoli, David M.; Van Eck, Joyce; Voytas, Daniel F.

    2016-01-01

    Plant transformation has enabled fundamental insights into plant biology and revolutionized commercial agriculture. Unfortunately, for most crops, transformation and regeneration remain arduous even after more than 30 years of technological advances. Genome editing provides novel opportunities to enhance crop productivity but relies on genetic transformation and plant regeneration, which are bottlenecks in the process. Here, we review the state of plant transformation and point to innovations needed to enable genome editing in crops. Plant tissue culture methods need optimization and simplification for efficiency and minimization of time in culture. Currently, specialized facilities exist for crop transformation. Single-cell and robotic techniques should be developed for high-throughput genomic screens. Plant genes involved in developmental reprogramming, wound response, and/or homologous recombination should be used to boost the recovery of transformed plants. Engineering universal Agrobacterium tumefaciens strains and recruiting other microbes, such as Ensifer or Rhizobium, could facilitate delivery of DNA and proteins into plant cells. Synthetic biology should be employed for de novo design of transformation systems. Genome editing is a potential game-changer in crop genetics when plant transformation systems are optimized. PMID:27335450

  5. Discrete fourier transform (DFT) analysis for applications using iterative transform methods

    NASA Technical Reports Server (NTRS)

    Dean, Bruce H. (Inventor)

    2012-01-01

    According to various embodiments, a method is provided for determining aberration data for an optical system. The method comprises collecting a data signal, and generating a pre-transformation algorithm. The data is pre-transformed by multiplying the data with the pre-transformation algorithm. A discrete Fourier transform of the pre-transformed data is performed in an iterative loop. The method further comprises back-transforming the data to generate aberration data.

  6. Superpixel Based Factor Analysis and Target Transformation Method for Martian Minerals Detection

    NASA Astrophysics Data System (ADS)

    Wu, X.; Zhang, X.; Lin, H.

    2018-04-01

    The Factor analysis and target transformation (FATT) is an effective method to test for the presence of particular mineral on Martian surface. It has been used both in thermal infrared (Thermal Emission Spectrometer, TES) and near-infrared (Compact Reconnaissance Imaging Spectrometer for Mars, CRISM) hyperspectral data. FATT derived a set of orthogonal eigenvectors from a mixed system and typically selected first 10 eigenvectors to least square fit the library mineral spectra. However, minerals present only in a limited pixels will be ignored because its weak spectral features compared with full image signatures. Here, we proposed a superpixel based FATT method to detect the mineral distributions on Mars. The simple linear iterative clustering (SLIC) algorithm was used to partition the CRISM image into multiple connected image regions with spectral homogeneous to enhance the weak signatures by increasing their proportion in a mixed system. A least square fitting was used in target transformation and performed to each region iteratively. Finally, the distribution of the specific minerals in image was obtained, where fitting residual less than a threshold represent presence and otherwise absence. We validate our method by identifying carbonates in a well analysed CRISM image in Nili Fossae on Mars. Our experimental results indicate that the proposed method work well both in simulated and real data sets.

  7. The Application of Continuous Wavelet Transform Based Foreground Subtraction Method in 21 cm Sky Surveys

    NASA Astrophysics Data System (ADS)

    Gu, Junhua; Xu, Haiguang; Wang, Jingying; An, Tao; Chen, Wen

    2013-08-01

    We propose a continuous wavelet transform based non-parametric foreground subtraction method for the detection of redshifted 21 cm signal from the epoch of reionization. This method works based on the assumption that the foreground spectra are smooth in frequency domain, while the 21 cm signal spectrum is full of saw-tooth-like structures, thus their characteristic scales are significantly different. We can distinguish them in the wavelet coefficient space easily and perform the foreground subtraction. Compared with the traditional spectral fitting based method, our method is more tolerant to complex foregrounds. Furthermore, we also find that when the instrument has uncorrected response error, our method can also work significantly better than the spectral fitting based method. Our method can obtain similar results with the Wp smoothing method, which is also a non-parametric method, but our method consumes much less computing time.

  8. Improving the french fry quality of russeted potatoes through transformation with the anti-sweetening gene (UgpA) from the Chipping cv. Snowden

    USDA-ARS?s Scientific Manuscript database

    Microtubers of two dual-purpose russeted potatoes were transformed with the anti-sweetening gene (UgpA) from the cv. Snowden using well know Agrobacterium tumifaciens mediated transformation system. Seventy-two and twenty-four distinct transformants of AOND95292-3Russ and ND7882b-7Russ, respectivel...

  9. An image hiding method based on cascaded iterative Fourier transform and public-key encryption algorithm

    NASA Astrophysics Data System (ADS)

    Zhang, B.; Sang, Jun; Alam, Mohammad S.

    2013-03-01

    An image hiding method based on cascaded iterative Fourier transform and public-key encryption algorithm was proposed. Firstly, the original secret image was encrypted into two phase-only masks M1 and M2 via cascaded iterative Fourier transform (CIFT) algorithm. Then, the public-key encryption algorithm RSA was adopted to encrypt M2 into M2' . Finally, a host image was enlarged by extending one pixel into 2×2 pixels and each element in M1 and M2' was multiplied with a superimposition coefficient and added to or subtracted from two different elements in the 2×2 pixels of the enlarged host image. To recover the secret image from the stego-image, the two masks were extracted from the stego-image without the original host image. By applying public-key encryption algorithm, the key distribution was facilitated, and also compared with the image hiding method based on optical interference, the proposed method may reach higher robustness by employing the characteristics of the CIFT algorithm. Computer simulations show that this method has good robustness against image processing.

  10. Advances in Maize Transformation Technologies and Development of Transgenic Maize

    PubMed Central

    Yadava, Pranjal; Abhishek, Alok; Singh, Reeva; Singh, Ishwar; Kaul, Tanushri; Pattanayak, Arunava; Agrawal, Pawan K.

    2017-01-01

    Maize is the principal grain crop of the world. It is also the crop where genetic engineering has been employed to a great extent to improve its various traits. The ability to transform maize is a crucial step for application of gene technology in maize improvement. There have been constant improvements in the maize transformation technologies over past several years. The choice of genotype and the explant material to initiate transformation and the different types of media to be used in various stages of tissue culture can have significant impact on the outcomes of the transformation efforts. Various methods of gene transfer, like the particle bombardment, protoplast transformation, Agrobacterium-mediated, in planta transformation, etc., have been tried and improved over years. Similarly, various selection systems for retrieval of the transformants have been attempted. The commercial success of maize transformation and transgenic development is unmatched by any other crop so far. Maize transformation with newer gene editing technologies is opening up a fresh dimension in transformation protocols and work-flows. This review captures the various past and recent facets in improvement in maize transformation technologies and attempts to present a comprehensive updated picture of the current state of the art in this area. PMID:28111576

  11. Advances in Maize Transformation Technologies and Development of Transgenic Maize.

    PubMed

    Yadava, Pranjal; Abhishek, Alok; Singh, Reeva; Singh, Ishwar; Kaul, Tanushri; Pattanayak, Arunava; Agrawal, Pawan K

    2016-01-01

    Maize is the principal grain crop of the world. It is also the crop where genetic engineering has been employed to a great extent to improve its various traits. The ability to transform maize is a crucial step for application of gene technology in maize improvement. There have been constant improvements in the maize transformation technologies over past several years. The choice of genotype and the explant material to initiate transformation and the different types of media to be used in various stages of tissue culture can have significant impact on the outcomes of the transformation efforts. Various methods of gene transfer, like the particle bombardment, protoplast transformation, Agrobacterium -mediated, in planta transformation, etc., have been tried and improved over years. Similarly, various selection systems for retrieval of the transformants have been attempted. The commercial success of maize transformation and transgenic development is unmatched by any other crop so far. Maize transformation with newer gene editing technologies is opening up a fresh dimension in transformation protocols and work-flows. This review captures the various past and recent facets in improvement in maize transformation technologies and attempts to present a comprehensive updated picture of the current state of the art in this area.

  12. 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.

  13. A Transfer Voltage Simulation Method for Generator Step Up Transformers

    NASA Astrophysics Data System (ADS)

    Funabashi, Toshihisa; Sugimoto, Toshirou; Ueda, Toshiaki; Ametani, Akihiro

    It has been found from measurements for 13 sets of GSU transformers that a transfer voltage of a generator step-up (GSU) transformer involves one dominant oscillation frequency. The frequency can be estimated from the inductance and capacitance values of the GSU transformer low-voltage-side. This observation has led to a new method for simulating a GSU transformer transfer voltage. The method is based on the EMTP TRANSFORMER model, but stray capacitances are added. The leakage inductance and the magnetizing resistance are modified using approximate curves for their frequency characteristics determined from the measured results. The new method is validated in comparison with the measured results.

  14. Wheat (Triticum aestivum L.) transformation using immature embryos.

    PubMed

    Ishida, Yuji; Tsunashima, Masako; Hiei, Yukoh; Komari, Toshihiko

    2015-01-01

    Wheat may now be transformed very efficiently by Agrobacterium tumefaciens. Under the protocol hereby described, immature embryos of healthy plants of wheat cultivar Fielder grown in a well-conditioned greenhouse were pretreated with centrifuging and cocultivated with A. tumefaciens. Transgenic wheat plants were obtained routinely from between 40 and 90 % of the immature embryos, thus infected in our tests. All regenerants were normal in morphology and fully fertile. About half of the transformed plants carried single copy of the transgene, which are inherited by the progeny in a Mendelian fashion.

  15. 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.

  16. Infrared fix pattern noise reduction method based on Shearlet Transform

    NASA Astrophysics Data System (ADS)

    Rong, Shenghui; Zhou, Huixin; Zhao, Dong; Cheng, Kuanhong; Qian, Kun; Qin, Hanlin

    2018-06-01

    The non-uniformity correction (NUC) is an effective way to reduce fix pattern noise (FPN) and improve infrared image quality. The temporal high-pass NUC method is a kind of practical NUC method because of its simple implementation. However, traditional temporal high-pass NUC methods rely deeply on the scene motion and suffer image ghosting and blurring. Thus, this paper proposes an improved NUC method based on Shearlet Transform (ST). First, the raw infrared image is decomposed into multiscale and multi-orientation subbands by ST and the FPN component mainly exists in some certain high-frequency subbands. Then, high-frequency subbands are processed by the temporal filter to extract the FPN due to its low-frequency characteristics. Besides, each subband has a confidence parameter to determine the degree of FPN, which is estimated by the variance of subbands adaptively. At last, the process of NUC is achieved by subtracting the estimated FPN component from the original subbands and the corrected infrared image can be obtained by the inverse ST. The performance of the proposed method is evaluated with real and synthetic infrared image sequences thoroughly. Experimental results indicate that the proposed method can reduce heavily FPN with less roughness and RMSE.

  17. Improved FFT-based numerical inversion of Laplace transforms via fast Hartley transform algorithm

    NASA Technical Reports Server (NTRS)

    Hwang, Chyi; Lu, Ming-Jeng; Shieh, Leang S.

    1991-01-01

    The disadvantages of numerical inversion of the Laplace transform via the conventional fast Fourier transform (FFT) are identified and an improved method is presented to remedy them. The improved method is based on introducing a new integration step length Delta(omega) = pi/mT for trapezoidal-rule approximation of the Bromwich integral, in which a new parameter, m, is introduced for controlling the accuracy of the numerical integration. Naturally, this method leads to multiple sets of complex FFT computations. A new inversion formula is derived such that N equally spaced samples of the inverse Laplace transform function can be obtained by (m/2) + 1 sets of N-point complex FFT computations or by m sets of real fast Hartley transform (FHT) computations.

  18. [A method to estimate the short-term fractal dimension of heart rate variability based on wavelet transform].

    PubMed

    Zhonggang, Liang; Hong, Yan

    2006-10-01

    A new method of calculating fractal dimension of short-term heart rate variability signals is presented. The method is based on wavelet transform and filter banks. The implementation of the method is: First of all we pick-up the fractal component from HRV signals using wavelet transform. Next, we estimate the power spectrum distribution of fractal component using auto-regressive model, and we estimate parameter 7 using the least square method. Finally according to formula D = 2- (gamma-1)/2 estimate fractal dimension of HRV signal. To validate the stability and reliability of the proposed method, using fractional brown movement simulate 24 fractal signals that fractal value is 1.6 to validate, the result shows that the method has stability and reliability.

  19. Performance comparison of ISAR imaging method based on time frequency transforms

    NASA Astrophysics Data System (ADS)

    Xie, Chunjian; Guo, Chenjiang; Xu, Jiadong

    2013-03-01

    Inverse synthetic aperture radar (ISAR) can image the moving target, especially the target in the air, so it is important in the air defence and missile defence system. Time-frequency Transform was applied to ISAR imaging process widely. Several time frequency transforms were introduced. Noise jamming methods were analysed, and when these noise jamming were added to the echo of the ISAR receiver, the image can become blur even can't to be identify. But the effect is different to the different time frequency analysis. The results of simulation experiment show the Performance Comparison of the method.

  20. Papaya (Carica papaya L.).

    PubMed

    Zhu, Yun J; Fitch, Maureen M M; Moore, Paul H

    2006-01-01

    Transgenic papaya plants were initially obtained using particle bombardment, a method having poor efficiency in producing intact, single-copy insertion of transgenes. Single-copy gene insertion was improved using Agrobacterium tumefaciens. With progress being made in genome sequencing and gene discovery, there is a need for more efficient methods of transformation in order to study the function of these genes. We describe a protocol for Agrobacterium-mediated transformation using carborundum-wounded papaya embryogenic calli. This method should lead to high-throughput transformation, which on average produced at least one plant that was positive in polymerase chain reaction (PCR), histochemical staining, or by Southern blot hybridization from 10 to 20% of the callus clusters that had been co-cultivated with Agrobacterium. Plants regenerated from the callus clusters in 9 to 13 mo.

  1. Endophytic occupation of root nodules and roots of Melilotus dentatus by Agrobacterium tumefaciens.

    PubMed

    Wang, Ling Ling; Wang, En Tao; Liu, Jie; Li, Ying; Chen, Wen Xin

    2006-10-01

    Agrobacterium strains have been frequently isolated from the root nodules of different legumes. Various possible mechanisms have been proposed to explain the existence of these bacteria in nodules, but there is no sufficient experimental evidence to support the estimations. In this work, we proved that the Agrobacterium strain CCBAU 81181, which was originally isolated from the root nodules of Onobrychis viciaefolia, and a symbiotic strain of Sinorhizobium meliloti CCBAU 10062 could coinhabit the root nodules of Melilotus dentatus. Analyses were performed by using a fluorescence marker, reisolation of bacteria from nodules, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of whole cellular proteins, and polymerase chain reaction amplification of symbiotic genes. The inoculation of A. tumefaciens CCBAU 81181 did not affect the growth and nodulation of plants. CCBAU 81181 and 24 other Agrobacterium strains isolated from nodules were incapable of nodulating on their original or alternative host and 22 strains of these strains were endophytes in the roots and stems of their hosts. Also, the tumor-inducing A. tumefaciens strains IAM 13129(T) and C58 were found capable of entering the roots of Glycyrrhiza pallidiflora, but did not cause pathogenic symptoms. With these results, we conclude that A. tumefaciens strains could be endophytic bacteria in the roots, stems, and root nodules. This finding partially explains why Agrobacterium strains were frequently isolated from the surface-sterilized nodules.

  2. [An automatic peak detection method for LIBS spectrum based on continuous wavelet transform].

    PubMed

    Chen, Peng-Fei; Tian, Di; Qiao, Shu-Jun; Yang, Guang

    2014-07-01

    Spectrum peak detection in the laser-induced breakdown spectroscopy (LIBS) is an essential step, but the presence of background and noise seriously disturb the accuracy of peak position. The present paper proposed a method applied to automatic peak detection for LIBS spectrum in order to enhance the ability of overlapping peaks searching and adaptivity. We introduced the ridge peak detection method based on continuous wavelet transform to LIBS, and discussed the choice of the mother wavelet and optimized the scale factor and the shift factor. This method also improved the ridge peak detection method with a correcting ridge method. The experimental results show that compared with other peak detection methods (the direct comparison method, derivative method and ridge peak search method), our method had a significant advantage on the ability to distinguish overlapping peaks and the precision of peak detection, and could be be applied to data processing in LIBS.

  3. [Application of optimized parameters SVM based on photoacoustic spectroscopy method in fault diagnosis of power transformer].

    PubMed

    Zhang, Yu-xin; Cheng, Zhi-feng; Xu, Zheng-ping; Bai, Jing

    2015-01-01

    In order to solve the problems such as complex operation, consumption for the carrier gas and long test period in traditional power transformer fault diagnosis approach based on dissolved gas analysis (DGA), this paper proposes a new method which is detecting 5 types of characteristic gas content in transformer oil such as CH4, C2H2, C2H4, C2H6 and H2 based on photoacoustic Spectroscopy and C2H2/C2H4, CH4/H2, C2H4/C2H6 three-ratios data are calculated. The support vector machine model was constructed using cross validation method under five support vector machine functions and four kernel functions, heuristic algorithms were used in parameter optimization for penalty factor c and g, which to establish the best SVM model for the highest fault diagnosis accuracy and the fast computing speed. Particles swarm optimization and genetic algorithm two types of heuristic algorithms were comparative studied in this paper for accuracy and speed in optimization. The simulation result shows that SVM model composed of C-SVC, RBF kernel functions and genetic algorithm obtain 97. 5% accuracy in test sample set and 98. 333 3% accuracy in train sample set, and genetic algorithm was about two times faster than particles swarm optimization in computing speed. The methods described in this paper has many advantages such as simple operation, non-contact measurement, no consumption for the carrier gas, long test period, high stability and sensitivity, the result shows that the methods described in this paper can instead of the traditional transformer fault diagnosis by gas chromatography and meets the actual project needs in transformer fault diagnosis.

  4. A direct method to transform between expansions in the configuration state function and Slater determinant bases

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Olsen, Jeppe, E-mail: jeppe@chem.au.dk

    2014-07-21

    A novel algorithm is introduced for the transformation of wave functions between the bases of Slater determinants (SD) and configuration state functions (CSF) in the genealogical coupling scheme. By modifying the expansion coefficients as each electron is spin-coupled, rather than performing a single many-electron transformation, the large transformation matrix that plagues previous approaches is avoided and the required number of operations is drastically reduced. As an example of the efficiency of the algorithm, the transformation for a configuration with 30 unpaired electrons and singlet spin is discussed. For this case, the 10 × 10{sup 6} coefficients in the CSF basismore » is obtained from the 150 × 10{sup 6} coefficients in the SD basis in 1 min, which should be compared with the seven years that the previously employed method is estimated to require.« less

  5. Genetic transformation of Begonia tuberhybrida by Ri rol genes.

    PubMed

    Kiyokawa, S; Kikuchi, Y; Kamada, H; Harada, H

    1996-04-01

    We have developed an Agrobacterium -mediated transformation system for commercial Begonia species. The leaf explants of Begonia semperflorens, Begonia x hiemalis and B. tuberhybrida were inoculated with Agrobacterium tumefaciens LBA4404 harboring a binary vector pBI121 which contains rolA, B and C genes of an agropine type Ri plasmid (pRiA4b). Kanamycin resistant shoots of B. tuberhybrida were obtained on MS agar medium supplemented with 0.1 mg/l NAA, 0.5 mg/l BA, 500 mg/l claforan and 100 mg/l kanamycin. These shoots exhibited GUS activity and Southern analysis showed a single copy insertion into the genome. When the transgenic plants were transferred to soil, they displayed the phenotype specific to the transgenic plants by A. rhizogenes such as dwarfness, delay of flowering, and wrinkled leaves and petals.

  6. Category's analysis and operational project capacity method of transformation in design

    NASA Astrophysics Data System (ADS)

    Obednina, S. V.; Bystrova, T. Y.

    2015-10-01

    The method of transformation is attracting widespread interest in fields such contemporary design. However, in theory of design little attention has been paid to a categorical status of the term "transformation". This paper presents the conceptual analysis of transformation based on the theory of form employed in the influential essays by Aristotle and Thomas Aquinas. In the present work the transformation as a method of shaping design has been explored as well as potential application of this term in design has been demonstrated.

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

    USDA-ARS?s Scientific Manuscript database

    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...

  8. Maize (Zea mays L.).

    PubMed

    Frame, Bronwyn; Warnberg, Katey; Main, Marcy; Wang, Kan

    2015-01-01

    Agrobacterium tumefaciens-mediated transformation is an effective method for introducing genes into maize. In this chapter, we describe a detailed protocol for genetic transformation of the maize genotype Hi II. Our starting plant material is immature embryos cocultivated with an Agrobacterium strain carrying a standard binary vector. In addition to step-by-step laboratory transformation procedures, we include extensive details in growing donor plants and caring for transgenic plants in the greenhouse.

  9. Impact of biological amendments on Agrobacterium tumefaciens soil survival

    USDA-ARS?s Scientific Manuscript database

    Paradox, the primary walnut rootstock used in California, is susceptible to Agrobacterium tumefaciens, which causes crown gall. While A. tumefaciens is susceptible to commonly used fumigants such as methyl bromide (MeBr) and Telone-C35 (1,3-dichloropropene and chloropicrin), these fumigants also sig...

  10. Textual blocks rectification method based on fast Hough transform analysis in identity documents recognition

    NASA Astrophysics Data System (ADS)

    Bezmaternykh, P. V.; Nikolaev, D. P.; Arlazarov, V. L.

    2018-04-01

    Textual blocks rectification or slant correction is an important stage of document image processing in OCR systems. This paper considers existing methods and introduces an approach for the construction of such algorithms based on Fast Hough Transform analysis. A quality measurement technique is proposed and obtained results are shown for both printed and handwritten textual blocks processing as a part of an industrial system of identity documents recognition on mobile devices.

  11. Agrobacterium rhizogenes - based transformation of soybean roots to form composite plants

    USDA-ARS?s Scientific Manuscript database

    Composite plants are a powerful tool to rapidly analyze the effects of gene overexpression, gene silencing, and examine test promoter expression in transgenic roots. No sterile tissue culture is needed. This avoids loss of valuable material due to contamination of sterile cultures. This method uses ...

  12. A method to analyze molecular tagging velocimetry data using the Hough transform.

    PubMed

    Sanchez-Gonzalez, R; McManamen, B; Bowersox, R D W; North, S W

    2015-10-01

    The development of a method to analyze molecular tagging velocimetry data based on the Hough transform is presented. This method, based on line fitting, parameterizes the grid lines "written" into a flowfield. Initial proof-of-principle illustration of this method was performed to obtain two-component velocity measurements in the wake of a cylinder in a Mach 4.6 flow, using a data set derived from computational fluid dynamics simulations. The Hough transform is attractive for molecular tagging velocimetry applications since it is capable of discriminating spurious features that can have a biasing effect in the fitting process. Assessment of the precision and accuracy of the method were also performed to show the dependence on analysis window size and signal-to-noise levels. The accuracy of this Hough transform-based method to quantify intersection displacements was determined to be comparable to cross-correlation methods. The employed line parameterization avoids the assumption of linearity in the vicinity of each intersection, which is important in the limit of drastic grid deformations resulting from large velocity gradients common in high-speed flow applications. This Hough transform method has the potential to enable the direct and spatially accurate measurement of local vorticity, which is important in applications involving turbulent flowfields. Finally, two-component velocity determinations using the Hough transform from experimentally obtained images are presented, demonstrating the feasibility of the proposed analysis method.

  13. Analysis of wave motion in one-dimensional structures through fast-Fourier-transform-based wavelet finite element method

    NASA Astrophysics Data System (ADS)

    Shen, Wei; Li, Dongsheng; Zhang, Shuaifang; Ou, Jinping

    2017-07-01

    This paper presents a hybrid method that combines the B-spline wavelet on the interval (BSWI) finite element method and spectral analysis based on fast Fourier transform (FFT) to study wave propagation in One-Dimensional (1D) structures. BSWI scaling functions are utilized to approximate the theoretical wave solution in the spatial domain and construct a high-accuracy dynamic stiffness matrix. Dynamic reduction on element level is applied to eliminate the interior degrees of freedom of BSWI elements and substantially reduce the size of the system matrix. The dynamic equations of the system are then transformed and solved in the frequency domain through FFT-based spectral analysis which is especially suitable for parallel computation. A comparative analysis of four different finite element methods is conducted to demonstrate the validity and efficiency of the proposed method when utilized in high-frequency wave problems. Other numerical examples are utilized to simulate the influence of crack and delamination on wave propagation in 1D rods and beams. Finally, the errors caused by FFT and their corresponding solutions are presented.

  14. Progress of tissue culture and genetic transformation research in pigeon pea [Cajanus cajan (L.) Millsp.].

    PubMed

    Krishna, Gaurav; Reddy, P Sairam; Ramteke, P W; Bhattacharya, P S

    2010-10-01

    Pigeon pea [Cajanus cajan (L.) Millsp.] (Family: Fabaceae) is an important legume crop cultivated across 50 countries in Asia, Africa, and the Americas; and ranks fifth in area among pulses after soybean, common bean, peanut, and chickpea. It is consumed as a major source of protein (21%) to the human population in many developing countries. In India, it is the second important food legume contributing to 80% of the global production. Several biotic and abiotic stresses are posing a big threat to its production and productivity. Attempts to address these problems through conventional breeding methods have met with partial success. This paper reviews the chronological progress made in tissue culture through organogenesis and somatic embryogenesis, including the influence of factors such as genotypes, explant sources, and culture media including the supplementation of plant growth regulators. Comprehensive lists of morphogenetic pathways involved in in vitro regeneration through organogenesis and somatic embryogenesis using different explant tissues of diverse pigeon pea genotypes are presented. Similarly, the establishment of protocols for the production of transgenics via particle bombardment and Agrobacterium-mediated transformation using different explant tissues, Agrobacterium strains, Ti plasmids, and plant selectable markers, as well as their interactions on transformation efficiency have been discussed. Future research thrusts on the use of different promoters and stacking of genes for various biotic and abiotic stresses in pigeon pea are suggested.

  15. Application of Tissue Culture and Transformation Techniques in Model Species Brachypodium distachyon.

    PubMed

    Sogutmaz Ozdemir, Bahar; Budak, Hikmet

    2018-01-01

    Brachypodium distachyon has recently emerged as a model plant species for the grass family (Poaceae) that includes major cereal crops and forage grasses. One of the important traits of a model species is its capacity to be transformed and ease of growing both in tissue culture and in greenhouse conditions. Hence, plant transformation technology is crucial for improvements in agricultural studies, both for the study of new genes and in the production of new transgenic plant species. In this chapter, we review an efficient tissue culture and two different transformation systems for Brachypodium using most commonly preferred gene transfer techniques in plant species, microprojectile bombardment method (biolistics) and Agrobacterium-mediated transformation.In plant transformation studies, frequently used explant materials are immature embryos due to their higher transformation efficiencies and regeneration capacity. However, mature embryos are available throughout the year in contrast to immature embryos. We explain a tissue culture protocol for Brachypodium using mature embryos with the selected inbred lines from our collection. Embryogenic calluses obtained from mature embryos are used to transform Brachypodium with both plant transformation techniques that are revised according to previously studied protocols applied in the grasses, such as applying vacuum infiltration, different wounding effects, modification in inoculation and cocultivation steps or optimization of bombardment parameters.

  16. Agrobacterium rhizogenes-mediated transformation of Superroot-derived Lotus corniculatus plants: a valuable tool for functional genomics.

    PubMed

    Jian, Bo; Hou, Wensheng; Wu, Cunxiang; Liu, Bin; Liu, Wei; Song, Shikui; Bi, Yurong; Han, Tianfu

    2009-06-25

    Transgenic approaches provide a powerful tool for gene function investigations in plants. However, some legumes are still recalcitrant to current transformation technologies, limiting the extent to which functional genomic studies can be performed on. Superroot of Lotus corniculatus is a continuous root cloning system allowing direct somatic embryogenesis and mass regeneration of plants. Recently, a technique to obtain transgenic L. corniculatus plants from Superroot-derived leaves through A. tumefaciens-mediated transformation was described. However, transformation efficiency was low and it took about six months from gene transfer to PCR identification. In the present study, we developed an A. rhizogenes-mediated transformation of Superroot-derived L. corniculatus for gene function investigation, combining the efficient A. rhizogenes-mediated transformation and the rapid regeneration system of Superroot. The transformation system using A. rhizogenes K599 harbouring pGFPGUSPlus was improved by validating some parameters which may influence the transformation frequency. Using stem sections with one node as explants, a 2-day pre-culture of explants, infection with K599 at OD(600) = 0.6, and co-cultivation on medium (pH 5.4) at 22 degrees C for 2 days enhanced the transformation frequency significantly. As proof of concept, Superroot-derived L. corniculatus was transformed with a gene from wheat encoding an Na+/H+ antiporter (TaNHX2) using the described system. Transgenic Superroot plants were obtained and had increased salt tolerance, as expected from the expression of TaNHX2. A rapid and efficient tool for gene function investigation in L. corniculatus was developed, combining the simplicity and high efficiency of the Superroot regeneration system and the availability of A. rhizogenes-mediated transformation. This system was improved by validating some parameters influencing the transformation frequency, which could reach 92% based on GUS detection. The combination

  17. 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.

  18. Sexually mature transgenic American chestnut trees via embryogenic suspension-based transformation.

    PubMed

    Andrade, Gisele M; Nairn, Campbell J; Le, Huong T; Merkle, Scott A

    2009-09-01

    The availability of a system for direct transfer of anti-fungal candidate genes into American chestnut (Castanea dentata), devastated by a fungal blight in the last century, would offer an alternative or supplemental approach to conventional breeding for production of chestnut trees resistant to the blight fungus and other pathogens. By taking advantage of the strong ability of embryogenic American chestnut cultures to proliferate in suspension, a high-throughput Agrobacterium tumefaciens-mediated transformation protocol for stable integration of foreign genes into the tree was established. Proembryogenic masses (PEMs) were co-cultivated with A. tumefaciens strain AGL1 harboring the plasmid pCAMBIA 2301, followed by stringent selection with 50 or 100 mg/l Geneticin. A protocol employing size-fractionation to enrich for small PEMs to use as target material and selection in suspension culture was applied to rapidly produce transgenic events with an average efficiency of four independent transformation events per 50 mg of target tissue and minimal escapes. Mature somatic embryos, representing 18 transgenic events and derived from multiple American chestnut target genotypes, were germinated and over 100 transgenic somatic seedlings were produced and acclimatized to greenhouse conditions. Multiple vigorous transgenic somatic seedlings produced functional staminate flowers within 3 years following regeneration.

  19. Genetic transformation of rare Verbascum eriophorum Godr. plants and metabolic alterations revealed by NMR-based metabolomics.

    PubMed

    Marchev, Andrey; Yordanova, Zhenya; Alipieva, Kalina; Zahmanov, Georgi; Rusinova-Videva, Snezhana; Kapchina-Toteva, Veneta; Simova, Svetlana; Popova, Milena; Georgiev, Milen I

    2016-09-01

    To develop a protocol to transform Verbascum eriophorum and to study the metabolic differences between mother plants and hairy root culture by applying NMR and processing the datasets with chemometric tools. Verbascum eriophorum is a rare species with restricted distribution, which is poorly studied. Agrobacterium rhizogenes-mediated genetic transformation of V. eriophorum and hairy root culture induction are reported for the first time. To determine metabolic alterations, V. eriophorum mother plants and relevant hairy root culture were subjected to comprehensive metabolomic analyses, using NMR (1D and 2D). Metabolomics data, processed using chemometric tools (and principal component analysis in particular) allowed exploration of V. eriophorum metabolome and have enabled identification of verbascoside (by means of 2D-TOCSY NMR) as the most abundant compound in hairy root culture. Metabolomics data contribute to the elucidation of metabolic alterations after T-DNA transfer to the host V. eriophorum genome and the development of hairy root culture for sustainable bioproduction of high value verbascoside.

  20. Nonuniform fast Fourier transform method for numerical diffraction simulation on tilted planes.

    PubMed

    Xiao, Yu; Tang, Xiahui; Qin, Yingxiong; Peng, Hao; Wang, Wei; Zhong, Lijing

    2016-10-01

    The method, based on the rotation of the angular spectrum in the frequency domain, is generally used for the diffraction simulation between the tilted planes. Due to the rotation of the angular spectrum, the interval between the sampling points in the Fourier domain is not even. For the conventional fast Fourier transform (FFT)-based methods, a spectrum interpolation is needed to get the approximate sampling value on the equidistant sampling points. However, due to the numerical error caused by the spectrum interpolation, the calculation accuracy degrades very quickly as the rotation angle increases. Here, the diffraction propagation between the tilted planes is transformed into a problem about the discrete Fourier transform on the uneven sampling points, which can be evaluated effectively and precisely through the nonuniform fast Fourier transform method (NUFFT). The most important advantage of this method is that the conventional spectrum interpolation is avoided and the high calculation accuracy can be guaranteed for different rotation angles, even when the rotation angle is close to π/2. Also, its calculation efficiency is comparable with that of the conventional FFT-based methods. Numerical examples as well as a discussion about the calculation accuracy and the sampling method are presented.

  1. Digital double random amplitude image encryption method based on the symmetry property of the parametric discrete Fourier transform

    NASA Astrophysics Data System (ADS)

    Bekkouche, Toufik; Bouguezel, Saad

    2018-03-01

    We propose a real-to-real image encryption method. It is a double random amplitude encryption method based on the parametric discrete Fourier transform coupled with chaotic maps to perform the scrambling. The main idea behind this method is the introduction of a complex-to-real conversion by exploiting the inherent symmetry property of the transform in the case of real-valued sequences. This conversion allows the encrypted image to be real-valued instead of being a complex-valued image as in all existing double random phase encryption methods. The advantage is to store or transmit only one image instead of two images (real and imaginary parts). Computer simulation results and comparisons with the existing double random amplitude encryption methods are provided for peak signal-to-noise ratio, correlation coefficient, histogram analysis, and key sensitivity.

  2. Morphogenic Regulators Baby boom and Wuschel Improve Monocot Transformation.

    PubMed

    Lowe, Keith; Wu, Emily; Wang, Ning; Hoerster, George; Hastings, Craig; Cho, Myeong-Je; Scelonge, Chris; Lenderts, Brian; Chamberlin, Mark; Cushatt, Josh; Wang, Lijuan; Ryan, Larisa; Khan, Tanveer; Chow-Yiu, Julia; Hua, Wei; Yu, Maryanne; Banh, Jenny; Bao, Zhongmeng; Brink, Kent; Igo, Elizabeth; Rudrappa, Bhojaraja; Shamseer, P M; Bruce, Wes; Newman, Lisa; Shen, Bo; Zheng, Peizhong; Bidney, Dennis; Falco, S Carl; RegisterIII, James C; Zhao, Zuo-Yu; Xu, Deping; Jones, Todd J; Gordon-Kamm, William James

    2016-09-06

    While transformation of the major monocot crops is currently possible, the process typically remains confined to one or two genotypes per species, often with poor agronomics, and efficiencies that place these methods beyond the reach of most academic laboratories. Here, we report a transformation approach involving overexpression of the maize (Zea mays) Baby boom (Bbm) and maize Wuschel2 (Wus2) genes, which produced high transformation frequencies in numerous previously non-transformable maize inbred lines. For example, the Pioneer inbred PHH5G is recalcitrant to biolistic and Agrobacterium transformation. However, when Bbm and Wus2 were expressed, transgenic calli were recovered from over 40% of the starting explants, with most producing healthy, fertile plants. Another limitation for many monocots is the intensive labor and greenhouse space required to supply immature embryos for transformation. This problem could be alleviated by using alternative target tissues that could be supplied consistently with automated preparation. As a major step toward this objective, we transformed Bbm and Wus2 directly into either embryo slices from mature seed or leaf segments from seedlings in a variety of Pioneer inbred lines, routinely recovering healthy, fertile T0 plants. Finally, we demonstrated that the maize Bbm and Wus2 genes stimulate transformation in sorghum (Sorghum bicolor) immature embryos, sugarcane (Saccharum officinarum) callus, and indica rice (Oryza sativa var. indica) callus. {copyright, serif} 2016 American Society of Plant Biologists. All rights reserved.

  3. Identification Method of Mud Shale Fractures Base on Wavelet Transform

    NASA Astrophysics Data System (ADS)

    Xia, Weixu; Lai, Fuqiang; Luo, Han

    2018-01-01

    In recent years, inspired by seismic analysis technology, a new method for analysing mud shale fractures oil and gas reservoirs by logging properties has emerged. By extracting the high frequency attribute of the wavelet transform in the logging attribute, the formation information hidden in the logging signal is extracted, identified the fractures that are not recognized by conventional logging and in the identified fracture segment to show the “cycle jump”, “high value”, “spike” and other response effect is more obvious. Finally formed a complete wavelet denoising method and wavelet high frequency identification fracture method.

  4. Morphogenic Regulators Baby boom and Wuschel Improve Monocot Transformation[OPEN

    PubMed Central

    Lowe, Keith; Wu, Emily; Cho, Myeong-Je; Lenderts, Brian; Chamberlin, Mark; Cushatt, Josh; Ryan, Larisa; Khan, Tanveer; Chow-Yiu, Julia; Hua, Wei; Banh, Jenny; Bao, Zhongmeng; Brink, Kent; Igo, Elizabeth; Rudrappa, Bhojaraja; Shamseer, PM; Shen, Bo; Zheng, Peizhong; Bidney, Dennis; Falco, Carl; Zhao, Zuo-Yu; Xu, Deping

    2016-01-01

    While transformation of the major monocot crops is currently possible, the process typically remains confined to one or two genotypes per species, often with poor agronomics, and efficiencies that place these methods beyond the reach of most academic laboratories. Here, we report a transformation approach involving overexpression of the maize (Zea mays) Baby boom (Bbm) and maize Wuschel2 (Wus2) genes, which produced high transformation frequencies in numerous previously nontransformable maize inbred lines. For example, the Pioneer inbred PHH5G is recalcitrant to biolistic and Agrobacterium tumefaciens transformation. However, when Bbm and Wus2 were expressed, transgenic calli were recovered from over 40% of the starting explants, with most producing healthy, fertile plants. Another limitation for many monocots is the intensive labor and greenhouse space required to supply immature embryos for transformation. This problem could be alleviated using alternative target tissues that could be supplied consistently with automated preparation. As a major step toward this objective, we transformed Bbm and Wus2 directly into either embryo slices from mature seed or leaf segments from seedlings in a variety of Pioneer inbred lines, routinely recovering healthy, fertile T0 plants. Finally, we demonstrated that the maize Bbm and Wus2 genes stimulate transformation in sorghum (Sorghum bicolor) immature embryos, sugarcane (Saccharum officinarum) callus, and indica rice (Oryza sativa ssp indica) callus. PMID:27600536

  5. A time-frequency analysis method to obtain stable estimates of magnetotelluric response function based on Hilbert-Huang transform

    NASA Astrophysics Data System (ADS)

    Cai, Jianhua

    2017-05-01

    The time-frequency analysis method represents signal as a function of time and frequency, and it is considered a powerful tool for handling arbitrary non-stationary time series by using instantaneous frequency and instantaneous amplitude. It also provides a possible alternative to the analysis of the non-stationary magnetotelluric (MT) signal. Based on the Hilbert-Huang transform (HHT), a time-frequency analysis method is proposed to obtain stable estimates of the magnetotelluric response function. In contrast to conventional methods, the response function estimation is performed in the time-frequency domain using instantaneous spectra rather than in the frequency domain, which allows for imaging the response parameter content as a function of time and frequency. The theory of the method is presented and the mathematical model and calculation procedure, which are used to estimate response function based on HHT time-frequency spectrum, are discussed. To evaluate the results, response function estimates are compared with estimates from a standard MT data processing method based on the Fourier transform. All results show that apparent resistivities and phases, which are calculated from the HHT time-frequency method, are generally more stable and reliable than those determined from the simple Fourier analysis. The proposed method overcomes the drawbacks of the traditional Fourier methods, and the resulting parameter minimises the estimation bias caused by the non-stationary characteristics of the MT data.

  6. Biolistic transformation of tobacco and maize suspension cells using bacterial cells as microprojectiles.

    PubMed

    Rasmussen, J L; Kikkert, J R; Roy, M K; Sanford, J C

    1994-01-01

    We have used both Escherichia coli cells and Agrobacterium tumefaciens cells as microprojectiles to deliver DNA into suspension-cultured tobacco (Nicotiana tabacum L. line NT1) cells using a helium powered biolistic device. In addition, E. coli cells were used as microprojectiles for the transformation of suspension-cultured maize (Zea mays cv. Black Mexican Sweet) cells. Pretreating the bacterial cells with phenol at a concentration of 1.0%, and combining the bacterial cells with tungsten particles increased the rates of transformation. In N. tabacum, we obtained hundreds of transient transformants per bombardment, but were unable to recover any stable transformants. In Z. mays we obtained thousands of transient transformants and an average of six stable transformants per bombardment. This difference is discussed.

  7. Optimization of conditions for transient Agrobacterium-mediated gene expression assays in Arabidopsis.

    PubMed

    Kim, Mi Jung; Baek, Kon; Park, Chung-Mo

    2009-08-01

    Transient genetic transformation of plant organs is an indispensable way of studying gene function in plants. This study was aimed to develop an optimized system for transient Agrobacterium-mediated transformation of the Arabidopsis leaves. The beta-glucuronidase (GUS) reporter gene was employed to evaluate growth and biochemical parameters that influence the levels of transient expression. The effects of plant culture conditions, Agrobacterial genetic backgrounds, densities of Agrobacterial cell suspensions, and of several detergents were analyzed. We found that optimization of plant culture conditions is the most critical factor among the parameters analyzed. Higher levels of transient expression were observed in plants grown under short day conditions (SDs) than in plants grown under long day conditions (LDs). Furthermore, incubation of the plants under SDs at high relative humidity (85-90%) for 24 h after infiltration greatly improved the levels of transient expression. Under the optimized culture conditions, expression of the reporter gene reached the peak 3 days after infiltration and was rapidly decreased after the peak. Among the five Agrobacterial strains examined, LAB4404 produced the highest levels of expression. We also examined the effects of detergents, including Triton X-100, Tween-20, and Silwet L-77. Supplementation of the infiltration media either with 0.01% Triton X-100 or 0.01% Tween-20 improved the levels of expression by approximately 1.6-fold. Our observations indicate that transient transformation of the Arabidopsis leaves in the infiltration media supplemented with 0.01% Triton X-100 and incubation of the infiltrated plants under SDs at high relative humidity are necessary for maximal levels of expression.

  8. A dynamic integrated fault diagnosis method for power transformers.

    PubMed

    Gao, Wensheng; Bai, Cuifen; Liu, Tong

    2015-01-01

    In order to diagnose transformer fault efficiently and accurately, a dynamic integrated fault diagnosis method based on Bayesian network is proposed in this paper. First, an integrated fault diagnosis model is established based on the causal relationship among abnormal working conditions, failure modes, and failure symptoms of transformers, aimed at obtaining the most possible failure mode. And then considering the evidence input into the diagnosis model is gradually acquired and the fault diagnosis process in reality is multistep, a dynamic fault diagnosis mechanism is proposed based on the integrated fault diagnosis model. Different from the existing one-step diagnosis mechanism, it includes a multistep evidence-selection process, which gives the most effective diagnostic test to be performed in next step. Therefore, it can reduce unnecessary diagnostic tests and improve the accuracy and efficiency of diagnosis. Finally, the dynamic integrated fault diagnosis method is applied to actual cases, and the validity of this method is verified.

  9. A Dynamic Integrated Fault Diagnosis Method for Power Transformers

    PubMed Central

    Gao, Wensheng; Liu, Tong

    2015-01-01

    In order to diagnose transformer fault efficiently and accurately, a dynamic integrated fault diagnosis method based on Bayesian network is proposed in this paper. First, an integrated fault diagnosis model is established based on the causal relationship among abnormal working conditions, failure modes, and failure symptoms of transformers, aimed at obtaining the most possible failure mode. And then considering the evidence input into the diagnosis model is gradually acquired and the fault diagnosis process in reality is multistep, a dynamic fault diagnosis mechanism is proposed based on the integrated fault diagnosis model. Different from the existing one-step diagnosis mechanism, it includes a multistep evidence-selection process, which gives the most effective diagnostic test to be performed in next step. Therefore, it can reduce unnecessary diagnostic tests and improve the accuracy and efficiency of diagnosis. Finally, the dynamic integrated fault diagnosis method is applied to actual cases, and the validity of this method is verified. PMID:25685841

  10. Susceptibility of Juglans Species andInterspecific Hybrids to Agrobacterium tumefaciens

    Treesearch

    James R. McKenna; Lynn Epstein

    2003-01-01

    Crown gall, caused by the common soil-borne bacterium Agrobacterium tumefaciens, can be an economic problem in walnut nurseries and production orchards in Caliiornia. The principal rootstocks used for commercial walnut production in California are the native Northern California black walnut, Juglans hindsii, and "Paradox,...

  11. Rapid DNA transformation in Salmonella Typhimurium by the hydrogel exposure method.

    PubMed

    Elabed, Hamouda; Hamza, Rim; Bakhrouf, Amina; Gaddour, Kamel

    2016-07-01

    Even with advances in molecular cloning and DNA transformation, new or alternative methods that permit DNA penetration in Salmonella enterica subspecies enterica serovar Typhimurium are required in order to use this pathogen in biotechnological or medical applications. In this work, an adapted protocol of bacterial transformation with plasmid DNA based on the "Yoshida effect" was applied and optimized on Salmonella enterica serovar Typhimurium LT2 reference strain. The plasmid transference based on the use of sepiolite as acicular materials to promote cell piercing via friction forces produced by spreading on the surface of a hydrogel. The transforming mixture containing sepiolite nanofibers, bacterial cells to be transformed and plasmid DNA were plated directly on selective medium containing 2% agar. In order to improve the procedure, three variables were tested and the transformation of Salmonella cells was accomplished using plasmids pUC19 and pBR322. Using the optimized protocol on Salmonella LT2 strain, the efficiency was about 10(5) transformed cells per 10(9) subjected to transformation with 0.2μg plasmid DNA. In summary, the procedure is fast, offers opportune efficiency and promises to become one of the widely used transformation methods in laboratories. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Hfq Influences Multiple Transport Systems and Virulence in the Plant Pathogen Agrobacterium tumefaciens

    PubMed Central

    Wilms, Ina; Möller, Philip; Stock, Anna-Maria; Gurski, Rosemarie; Lai, Erh-Min

    2012-01-01

    The Hfq protein mediates gene regulation by small RNAs (sRNAs) in about 50% of all bacteria. Depending on the species, phenotypic defects of an hfq mutant range from mild to severe. Here, we document that the purified Hfq protein of the plant pathogen and natural genetic engineer Agrobacterium tumefaciens binds to the previously described sRNA AbcR1 and its target mRNA atu2422, which codes for the substrate binding protein of an ABC transporter taking up proline and γ-aminobutyric acid (GABA). Several other ABC transporter components were overproduced in an hfq mutant compared to their levels in the parental strain, suggesting that Hfq plays a major role in controlling the uptake systems and metabolic versatility of A. tumefaciens. The hfq mutant showed delayed growth, altered cell morphology, and reduced motility. Although the DNA-transferring type IV secretion system was produced, tumor formation by the mutant strain was attenuated, demonstrating an important contribution of Hfq to plant transformation by A. tumefaciens. PMID:22821981

  13. Frequency-dependent FDTD methods using Z transforms

    NASA Technical Reports Server (NTRS)

    Sullivan, Dennis M.

    1992-01-01

    While the frequency-dependent finite-difference time-domain, or (FD)2TD, method can correctly calculate EM propagation through media whose dielectric properties are frequency-dependent, more elaborate applications lead to greater (FD)2TD complexity. Z-transform theory is presently used to develop the mathematical bases of the (FD)2TD method, simultaneously obtaining a clearer formulation and allowing researchers to draw on the existing literature of systems analysis and signal-processing.

  14. Multisource image fusion method using support value transform.

    PubMed

    Zheng, Sheng; Shi, Wen-Zhong; Liu, Jian; Zhu, Guang-Xi; Tian, Jin-Wen

    2007-07-01

    With the development of numerous imaging sensors, many images can be simultaneously pictured by various sensors. However, there are many scenarios where no one sensor can give the complete picture. Image fusion is an important approach to solve this problem and produces a single image which preserves all relevant information from a set of different sensors. In this paper, we proposed a new image fusion method using the support value transform, which uses the support value to represent the salient features of image. This is based on the fact that, in support vector machines (SVMs), the data with larger support values have a physical meaning in the sense that they reveal relative more importance of the data points for contributing to the SVM model. The mapped least squares SVM (mapped LS-SVM) is used to efficiently compute the support values of image. The support value analysis is developed by using a series of multiscale support value filters, which are obtained by filling zeros in the basic support value filter deduced from the mapped LS-SVM to match the resolution of the desired level. Compared with the widely used image fusion methods, such as the Laplacian pyramid, discrete wavelet transform methods, the proposed method is an undecimated transform-based approach. The fusion experiments are undertaken on multisource images. The results demonstrate that the proposed approach is effective and is superior to the conventional image fusion methods in terms of the pertained quantitative fusion evaluation indexes, such as quality of visual information (Q(AB/F)), the mutual information, etc.

  15. General method for designing wave shape transformers.

    PubMed

    Ma, Hua; Qu, Shaobo; Xu, Zhuo; Wang, Jiafu

    2008-12-22

    An effective method for designing wave shape transformers (WSTs) is investigated by adopting the coordinate transformation theory. Following this method, the devices employed to transform electromagnetic (EM) wave fronts from one style with arbitrary shape and size to another style, can be designed. To verify this method, three examples in 2D spaces are also presented. Compared with the methods proposed in other literatures, this method offers the general procedure in designing WSTs, and thus is of great importance for the potential and practical applications possessed by such kinds of devices.

  16. Fast data reconstructed method of Fourier transform imaging spectrometer based on multi-core CPU

    NASA Astrophysics Data System (ADS)

    Yu, Chunchao; Du, Debiao; Xia, Zongze; Song, Li; Zheng, Weijian; Yan, Min; Lei, Zhenggang

    2017-10-01

    Imaging spectrometer can gain two-dimensional space image and one-dimensional spectrum at the same time, which shows high utility in color and spectral measurements, the true color image synthesis, military reconnaissance and so on. In order to realize the fast reconstructed processing of the Fourier transform imaging spectrometer data, the paper designed the optimization reconstructed algorithm with OpenMP parallel calculating technology, which was further used for the optimization process for the HyperSpectral Imager of `HJ-1' Chinese satellite. The results show that the method based on multi-core parallel computing technology can control the multi-core CPU hardware resources competently and significantly enhance the calculation of the spectrum reconstruction processing efficiency. If the technology is applied to more cores workstation in parallel computing, it will be possible to complete Fourier transform imaging spectrometer real-time data processing with a single computer.

  17. An accurate and efficient reliability-based design optimization using the second order reliability method and improved stability transformation method

    NASA Astrophysics Data System (ADS)

    Meng, Zeng; Yang, Dixiong; Zhou, Huanlin; Yu, Bo

    2018-05-01

    The first order reliability method has been extensively adopted for reliability-based design optimization (RBDO), but it shows inaccuracy in calculating the failure probability with highly nonlinear performance functions. Thus, the second order reliability method is required to evaluate the reliability accurately. However, its application for RBDO is quite challenge owing to the expensive computational cost incurred by the repeated reliability evaluation and Hessian calculation of probabilistic constraints. In this article, a new improved stability transformation method is proposed to search the most probable point efficiently, and the Hessian matrix is calculated by the symmetric rank-one update. The computational capability of the proposed method is illustrated and compared to the existing RBDO approaches through three mathematical and two engineering examples. The comparison results indicate that the proposed method is very efficient and accurate, providing an alternative tool for RBDO of engineering structures.

  18. Agrobacterium tumefaciens Integrates Transfer DNA into Single Chromosomal Sites of Dimorphic Fungi and Yields Homokaryotic Progeny from Multinucleate Yeast

    PubMed Central

    Sullivan, Thomas D.; Rooney, Peggy J.; Klein, Bruce S.

    2002-01-01

    The dimorphic fungi Blastomyces dermatitidis and Histoplasma capsulatum cause systemic mycoses in humans and other animals. Forward genetic approaches to generating and screening mutants for biologically important phenotypes have been underutilized for these pathogens. The plant-transforming bacterium Agrobacterium tumefaciens was tested to determine whether it could transform these fungi and if the fate of transforming DNA was suited for use as an insertional mutagen. Yeast cells from both fungi and germinating conidia from B. dermatitidis were transformed via A. tumefaciens by using hygromycin resistance for selection. Transformation frequencies up to 1 per 100 yeast cells were obtained at high effector-to-target ratios of 3,000:1. B. dermatitidis and H. capsulatum ura5 lines were complemented with transfer DNA vectors expressing URA5 at efficiencies 5 to 10 times greater than those obtained using hygromycin selection. Southern blot analyses indicated that in 80% of transformants the transferred DNA was integrated into chromosomal DNA at single, unique sites in the genome. Progeny of B. dermatitidis transformants unexpectedly showed that a single round of colony growth under hygromycin selection or visible selection of transformants by lacZ expression generated homokaryotic progeny from multinucleate yeast. Theoretical analysis of random organelle sorting suggests that the majority of B. dermatitidis cells would be homokaryons after the ca. 20 generations necessary for colony formation. Taken together, the results demonstrate that A. tumefaciens efficiently transfers DNA into B. dermatitidis and H. capsulatum and has the properties necessary for use as an insertional mutagen in these fungi. PMID:12477790

  19. A new method of converter transformer protection without commutation failure

    NASA Astrophysics Data System (ADS)

    Zhang, Jiayu; Kong, Bo; Liu, Mingchang; Zhang, Jun; Guo, Jianhong; Jing, Xu

    2018-01-01

    With the development of AC / DC hybrid transmission technology, converter transformer as nodes of AC and DC conversion of HVDC transmission technology, its reliable safe and stable operation plays an important role in the DC transmission. As a common problem of DC transmission, commutation failure poses a serious threat to the safe and stable operation of power grid. According to the commutation relation between the AC bus voltage of converter station and the output DC voltage of converter, the generalized transformation ratio is defined, and a new method of converter transformer protection based on generalized transformation ratio is put forward. The method uses generalized ratio to realize the on-line monitoring of the fault or abnormal commutation components, and the use of valve side of converter transformer bushing CT current characteristics of converter transformer fault accurately, and is not influenced by the presence of commutation failure. Through the fault analysis and EMTDC/PSCAD simulation, the protection can be operated correctly under the condition of various faults of the converter.

  20. Efficient embryogenic suspension culturing and rapid transformation of a range of elite genotypes of sweet potato (Ipomoea batatas [L.] Lam.).

    PubMed

    Yang, Jun; Bi, Hui-Ping; Fan, Wei-Juan; Zhang, Min; Wang, Hong-Xia; Zhang, Peng

    2011-12-01

    Efficient Agrobacterium tumefaciens-mediated transformation was developed using embryogenic suspension cell cultures of elite sweet potato (Ipomoea batatas [L.] Lam.) cultivars, including Ayamurasaki, Sushu2, Sushu9, Sushu11, Wanshu1, Xushu18 and Xushu22. Embryogenic suspension cultures were established in LCP medium using embryogenic calli induced from apical or axillary buds on an induction medium containing 2 mg l(-1) 2,4-D. Suspension cultures were co-cultivated with A. tumefaciens strain LBA4404 harboring the binary plasmid pCAMBIA1301 with the hpt gene as a selectable marker and an intron-interrupted uidA gene as a visible marker. Several key steps of the sweet potato transformation system have been investigated and optimized, including the appropriate antibiotics and their concentrations for suppressing Agrobacterium growth and the optimal doses of hygromycin for transformant selection. A total of 485 putative transgenic plant lines were produced from the transformed calli via somatic embryogenesis and germination to plants under 10 mg l(-1) hygromycin and 200 mg l(-1) cefotaxime. PCR, GUS and Southern blot analyses of the regenerated plants showed that 92.35% of them were transgenic. The number of T-DNA insertions varied from one to three in most transgenic plant lines. Plants showed 100% survival when 308 transgenics were transferred to soil in the greenhouse and then to the field. Most of them were morphologically normal, with the production of storage roots after 3 months of cultivation in the greenhouse or fields. The development of such a robust transformation method suitable to a range of sweet potato genotypes not only provides a routine tool for genetic improvement via transgenesis but also allows us to conduct a functional verification of endogenous genes in sweet potato. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  1. Improved l1-SPIRiT using 3D walsh transform-based sparsity basis.

    PubMed

    Feng, Zhen; Liu, Feng; Jiang, Mingfeng; Crozier, Stuart; Guo, He; Wang, Yuxin

    2014-09-01

    l1-SPIRiT is a fast magnetic resonance imaging (MRI) method which combines parallel imaging (PI) with compressed sensing (CS) by performing a joint l1-norm and l2-norm optimization procedure. The original l1-SPIRiT method uses two-dimensional (2D) Wavelet transform to exploit the intra-coil data redundancies and a joint sparsity model to exploit the inter-coil data redundancies. In this work, we propose to stack all the coil images into a three-dimensional (3D) matrix, and then a novel 3D Walsh transform-based sparsity basis is applied to simultaneously reduce the intra-coil and inter-coil data redundancies. Both the 2D Wavelet transform-based and the proposed 3D Walsh transform-based sparsity bases were investigated in the l1-SPIRiT method. The experimental results show that the proposed 3D Walsh transform-based l1-SPIRiT method outperformed the original l1-SPIRiT in terms of image quality and computational efficiency. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. Agrobacterium tumefaciens Promotes Tumor Induction by Modulating Pathogen Defense in Arabidopsis thaliana[W

    PubMed Central

    Lee, Chil-Woo; Efetova, Marina; Engelmann, Julia C; Kramell, Robert; Wasternack, Claus; Ludwig-Müller, Jutta; Hedrich, Rainer; Deeken, Rosalia

    2009-01-01

    Agrobacterium tumefaciens causes crown gall disease by transferring and integrating bacterial DNA (T-DNA) into the plant genome. To examine the physiological changes and adaptations during Agrobacterium-induced tumor development, we compared the profiles of salicylic acid (SA), ethylene (ET), jasmonic acid (JA), and auxin (indole-3-acetic acid [IAA]) with changes in the Arabidopsis thaliana transcriptome. Our data indicate that host responses were much stronger toward the oncogenic strain C58 than to the disarmed strain GV3101 and that auxin acts as a key modulator of the Arabidopsis–Agrobacterium interaction. At initiation of infection, elevated levels of IAA and ET were associated with the induction of host genes involved in IAA, but not ET signaling. After T-DNA integration, SA as well as IAA and ET accumulated, but JA did not. This did not correlate with SA-controlled pathogenesis-related gene expression in the host, although high SA levels in mutant plants prevented tumor development, while low levels promoted it. Our data are consistent with a scenario in which ET and later on SA control virulence of agrobacteria, whereas ET and auxin stimulate neovascularization during tumor formation. We suggest that crosstalk among IAA, ET, and SA balances pathogen defense launched by the host and tumor growth initiated by agrobacteria. PMID:19794116

  3. A dual selection based, targeted gene replacement tool for Magnaporthe grisea and Fusarium oxysporum.

    PubMed

    Khang, Chang Hyun; Park, Sook-Young; Lee, Yong-Hwan; Kang, Seogchan

    2005-06-01

    Rapid progress in fungal genome sequencing presents many new opportunities for functional genomic analysis of fungal biology through the systematic mutagenesis of the genes identified through sequencing. However, the lack of efficient tools for targeted gene replacement is a limiting factor for fungal functional genomics, as it often necessitates the screening of a large number of transformants to identify the desired mutant. We developed an efficient method of gene replacement and evaluated factors affecting the efficiency of this method using two plant pathogenic fungi, Magnaporthe grisea and Fusarium oxysporum. This method is based on Agrobacterium tumefaciens-mediated transformation with a mutant allele of the target gene flanked by the herpes simplex virus thymidine kinase (HSVtk) gene as a conditional negative selection marker against ectopic transformants. The HSVtk gene product converts 5-fluoro-2'-deoxyuridine to a compound toxic to diverse fungi. Because ectopic transformants express HSVtk, while gene replacement mutants lack HSVtk, growing transformants on a medium amended with 5-fluoro-2'-deoxyuridine facilitates the identification of targeted mutants by counter-selecting against ectopic transformants. In addition to M. grisea and F. oxysporum, the method and associated vectors are likely to be applicable to manipulating genes in a broad spectrum of fungi, thus potentially serving as an efficient, universal functional genomic tool for harnessing the growing body of fungal genome sequence data to study fungal biology.

  4. Comparing transformation methods for DNA microarray data

    PubMed Central

    Thygesen, Helene H; Zwinderman, Aeilko H

    2004-01-01

    Background When DNA microarray data are used for gene clustering, genotype/phenotype correlation studies, or tissue classification the signal intensities are usually transformed and normalized in several steps in order to improve comparability and signal/noise ratio. These steps may include subtraction of an estimated background signal, subtracting the reference signal, smoothing (to account for nonlinear measurement effects), and more. Different authors use different approaches, and it is generally not clear to users which method they should prefer. Results We used the ratio between biological variance and measurement variance (which is an F-like statistic) as a quality measure for transformation methods, and we demonstrate a method for maximizing that variance ratio on real data. We explore a number of transformations issues, including Box-Cox transformation, baseline shift, partial subtraction of the log-reference signal and smoothing. It appears that the optimal choice of parameters for the transformation methods depends on the data. Further, the behavior of the variance ratio, under the null hypothesis of zero biological variance, appears to depend on the choice of parameters. Conclusions The use of replicates in microarray experiments is important. Adjustment for the null-hypothesis behavior of the variance ratio is critical to the selection of transformation method. PMID:15202953

  5. Comparing transformation methods for DNA microarray data.

    PubMed

    Thygesen, Helene H; Zwinderman, Aeilko H

    2004-06-17

    When DNA microarray data are used for gene clustering, genotype/phenotype correlation studies, or tissue classification the signal intensities are usually transformed and normalized in several steps in order to improve comparability and signal/noise ratio. These steps may include subtraction of an estimated background signal, subtracting the reference signal, smoothing (to account for nonlinear measurement effects), and more. Different authors use different approaches, and it is generally not clear to users which method they should prefer. We used the ratio between biological variance and measurement variance (which is an F-like statistic) as a quality measure for transformation methods, and we demonstrate a method for maximizing that variance ratio on real data. We explore a number of transformations issues, including Box-Cox transformation, baseline shift, partial subtraction of the log-reference signal and smoothing. It appears that the optimal choice of parameters for the transformation methods depends on the data. Further, the behavior of the variance ratio, under the null hypothesis of zero biological variance, appears to depend on the choice of parameters. The use of replicates in microarray experiments is important. Adjustment for the null-hypothesis behavior of the variance ratio is critical to the selection of transformation method.

  6. Invariant object recognition based on the generalized discrete radon transform

    NASA Astrophysics Data System (ADS)

    Easley, Glenn R.; Colonna, Flavia

    2004-04-01

    We introduce a method for classifying objects based on special cases of the generalized discrete Radon transform. We adjust the transform and the corresponding ridgelet transform by means of circular shifting and a singular value decomposition (SVD) to obtain a translation, rotation and scaling invariant set of feature vectors. We then use a back-propagation neural network to classify the input feature vectors. We conclude with experimental results and compare these with other invariant recognition methods.

  7. A New Instantaneous Frequency Measure Based on The Stockwell Transform

    NASA Astrophysics Data System (ADS)

    yedlin, M. J.; Ben-Horrin, Y.; Fraser, J. D.

    2011-12-01

    We propose the use of a new transform, the Stockwell transform[1], as a means of creating time-frequency maps and applying them to distinguish blasts from earthquakes. This new transform, the Stockwell transform can be considered as a variant of the continuous wavelet transform, that preserves the absolute phase.The Stockwell transform employs a complex Morlet mother wavelet. The novelty of this transform lies in its resolution properties. High frequencies in the candidate signal are well-resolved in time but poorly resolved in frequency, while the converse is true for low frequency signal components. The goal of this research is to obtain the instantaneous frequency as a function of time for both the earthquakes and the blasts. Two methods will be compared. In the first method, we will compute the analytic signal, the envelope and the instantaneous phase as a function of time[2]. The instantaneous phase derivative will yield the instantaneous angular frequency. The second method will be based on time-frequency analysis using the Stockwell transform. The Stockwell transform will be computed in non-redundant fashion using a dyadic representation[3]. For each time-point, the frequency centroid will be computed -- a representation for the most likely frequency at that time. A detailed comparison will be presented for both approaches to the computation of the instantaneous frequency. An advantage of the Stockwell approach is that no differentiation is applied. The Hilbert transform method can be less sensitive to edge effects. The goal of this research is to see if the new Stockwell-based method could be used as a discriminant between earthquakes and blasts. References [1] Stockwell, R.G., Mansinha, L. and Lowe, R.P. "Localization of the complex spectrum: the S transform", IEEE Trans. Signal Processing, vol.44, no.4, pp.998-1001, (1996). [2]Taner, M.T., Koehler, F. "Complex seismic trace analysis", Geophysics, vol. 44, Issue 6, pp. 1041-1063 (1979). [3] Brown, R

  8. Wearable Sensor-Based Human Activity Recognition Method with Multi-Features Extracted from Hilbert-Huang Transform.

    PubMed

    Xu, Huile; Liu, Jinyi; Hu, Haibo; Zhang, Yi

    2016-12-02

    Wearable sensors-based human activity recognition introduces many useful applications and services in health care, rehabilitation training, elderly monitoring and many other areas of human interaction. Existing works in this field mainly focus on recognizing activities by using traditional features extracted from Fourier transform (FT) or wavelet transform (WT). However, these signal processing approaches are suitable for a linear signal but not for a nonlinear signal. In this paper, we investigate the characteristics of the Hilbert-Huang transform (HHT) for dealing with activity data with properties such as nonlinearity and non-stationarity. A multi-features extraction method based on HHT is then proposed to improve the effect of activity recognition. The extracted multi-features include instantaneous amplitude (IA) and instantaneous frequency (IF) by means of empirical mode decomposition (EMD), as well as instantaneous energy density (IE) and marginal spectrum (MS) derived from Hilbert spectral analysis. Experimental studies are performed to verify the proposed approach by using the PAMAP2 dataset from the University of California, Irvine for wearable sensors-based activity recognition. Moreover, the effect of combining multi-features vs. a single-feature are investigated and discussed in the scenario of a dependent subject. The experimental results show that multi-features combination can further improve the performance measures. Finally, we test the effect of multi-features combination in the scenario of an independent subject. Our experimental results show that we achieve four performance indexes: recall, precision, F-measure, and accuracy to 0.9337, 0.9417, 0.9353, and 0.9377 respectively, which are all better than the achievements of related works.

  9. Wearable Sensor-Based Human Activity Recognition Method with Multi-Features Extracted from Hilbert-Huang Transform

    PubMed Central

    Xu, Huile; Liu, Jinyi; Hu, Haibo; Zhang, Yi

    2016-01-01

    Wearable sensors-based human activity recognition introduces many useful applications and services in health care, rehabilitation training, elderly monitoring and many other areas of human interaction. Existing works in this field mainly focus on recognizing activities by using traditional features extracted from Fourier transform (FT) or wavelet transform (WT). However, these signal processing approaches are suitable for a linear signal but not for a nonlinear signal. In this paper, we investigate the characteristics of the Hilbert-Huang transform (HHT) for dealing with activity data with properties such as nonlinearity and non-stationarity. A multi-features extraction method based on HHT is then proposed to improve the effect of activity recognition. The extracted multi-features include instantaneous amplitude (IA) and instantaneous frequency (IF) by means of empirical mode decomposition (EMD), as well as instantaneous energy density (IE) and marginal spectrum (MS) derived from Hilbert spectral analysis. Experimental studies are performed to verify the proposed approach by using the PAMAP2 dataset from the University of California, Irvine for wearable sensors-based activity recognition. Moreover, the effect of combining multi-features vs. a single-feature are investigated and discussed in the scenario of a dependent subject. The experimental results show that multi-features combination can further improve the performance measures. Finally, we test the effect of multi-features combination in the scenario of an independent subject. Our experimental results show that we achieve four performance indexes: recall, precision, F-measure, and accuracy to 0.9337, 0.9417, 0.9353, and 0.9377 respectively, which are all better than the achievements of related works. PMID:27918414

  10. Towards discrete wavelet transform-based human activity recognition

    NASA Astrophysics Data System (ADS)

    Khare, Manish; Jeon, Moongu

    2017-06-01

    Providing accurate recognition of human activities is a challenging problem for visual surveillance applications. In this paper, we present a simple and efficient algorithm for human activity recognition based on a wavelet transform. We adopt discrete wavelet transform (DWT) coefficients as a feature of human objects to obtain advantages of its multiresolution approach. The proposed method is tested on multiple levels of DWT. Experiments are carried out on different standard action datasets including KTH and i3D Post. The proposed method is compared with other state-of-the-art methods in terms of different quantitative performance measures. The proposed method is found to have better recognition accuracy in comparison to the state-of-the-art methods.

  11. A combinatorial filtering method for magnetotelluric time-series based on Hilbert-Huang transform

    NASA Astrophysics Data System (ADS)

    Cai, Jianhua

    2014-11-01

    Magnetotelluric (MT) time-series are often contaminated with noise from natural or man-made processes. A substantial improvement is possible when the time-series are presented as clean as possible for further processing. A combinatorial method is described for filtering of MT time-series based on the Hilbert-Huang transform that requires a minimum of human intervention and leaves good data sections unchanged. Good data sections are preserved because after empirical mode decomposition the data are analysed through hierarchies, morphological filtering, adaptive threshold and multi-point smoothing, allowing separation of noise from signals. The combinatorial method can be carried out without any assumption about the data distribution. Simulated data and the real measured MT time-series from three different regions, with noise caused by baseline drift, high frequency noise and power-line contribution, are processed to demonstrate the application of the proposed method. Results highlight the ability of the combinatorial method to pick out useful signals, and the noise is suppressed greatly so that their deleterious influence is eliminated for the MT transfer function estimation.

  12. Improving the local wavenumber method by automatic DEXP transformation

    NASA Astrophysics Data System (ADS)

    Abbas, Mahmoud Ahmed; Fedi, Maurizio; Florio, Giovanni

    2014-12-01

    In this paper we present a new method for source parameter estimation, based on the local wavenumber function. We make use of the stable properties of the Depth from EXtreme Points (DEXP) method, in which the depth to the source is determined at the extreme points of the field scaled with a power-law of the altitude. Thus the method results particularly suited to deal with local wavenumber of high-order, as it is able to overcome its known instability caused by the use of high-order derivatives. The DEXP transformation enjoys a relevant feature when applied to the local wavenumber function: the scaling-law is in fact independent of the structural index. So, differently from the DEXP transformation applied directly to potential fields, the Local Wavenumber DEXP transformation is fully automatic and may be implemented as a very fast imaging method, mapping every kind of source at the correct depth. Also the simultaneous presence of sources with different homogeneity degree can be easily and correctly treated. The method was applied to synthetic and real examples from Bulgaria and Italy and the results agree well with known information about the causative sources.

  13. Mutants of Agrobacterium tumefaciens with elevated vir gene expression

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pazour, G.J.; Ta, C.N.; Das, A.

    1991-08-15

    Expression of Agrobacterium tumefaciens virulence (vir) genes requires virA, virG, and a plant-derived inducing compound such as acetosyringone. To identify the critical functional domains of virA and virG, a mutational approach was used. Agrobacterium A136 harboring plasmid pGP159, which contains virA, virG, and a reporter virB:lacZ gene fusion, was mutagenized with UV light or nitrosoguanidine. Survivors that formed blue colonies on a plate containing 5-bromo-4-chloro-3-indolyl beta-D-galactoside were isolated and analyzed. Quantification of beta-galactosidase activity in liquid assays identified nine mutant strains. By plasmid reconstruction and other procedures, all mutations mapped to the virA locus. These mutations caused an 11- tomore » 560-fold increase in the vegetative level of virB:lacZ reporter gene expression. DNA sequence analysis showed that the mutations are located in four regions of VirA: transmembrane domain one, the active site, a glycine-rich region with homology to ATP-binding sites, and a region at the C terminus that has homology to the N terminus of VirG.« less

  14. 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.

  15. A Transform-Based Feature Extraction Approach for Motor Imagery Tasks Classification

    PubMed Central

    Khorshidtalab, Aida; Mesbah, Mostefa; Salami, Momoh J. E.

    2015-01-01

    In this paper, we present a new motor imagery classification method in the context of electroencephalography (EEG)-based brain–computer interface (BCI). This method uses a signal-dependent orthogonal transform, referred to as linear prediction singular value decomposition (LP-SVD), for feature extraction. The transform defines the mapping as the left singular vectors of the LP coefficient filter impulse response matrix. Using a logistic tree-based model classifier; the extracted features are classified into one of four motor imagery movements. The proposed approach was first benchmarked against two related state-of-the-art feature extraction approaches, namely, discrete cosine transform (DCT) and adaptive autoregressive (AAR)-based methods. By achieving an accuracy of 67.35%, the LP-SVD approach outperformed the other approaches by large margins (25% compared with DCT and 6 % compared with AAR-based methods). To further improve the discriminatory capability of the extracted features and reduce the computational complexity, we enlarged the extracted feature subset by incorporating two extra features, namely, Q- and the Hotelling’s \\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{upgreek} \\usepackage{mathrsfs} \\setlength{\\oddsidemargin}{-69pt} \\begin{document} }{}$T^{2}$ \\end{document} statistics of the transformed EEG and introduced a new EEG channel selection method. The performance of the EEG classification based on the expanded feature set and channel selection method was compared with that of a number of the state-of-the-art classification methods previously reported with the BCI IIIa competition data set. Our method came second with an average accuracy of 81.38%. PMID:27170898

  16. Agrobacterium-assisted selenium nanoparticles: molecular aspect of antifungal activity

    NASA Astrophysics Data System (ADS)

    Kumar, Anil; Bera, Smritilekha; Singh, Man; Mondal, Dhananjoy

    2018-03-01

    Selenium nanoparticles (SeNPs) were synthesized through the bioreduction of sodium selenite (Na2SeO3) using gram-negative agrobacterium (AGBT) species. Subsequently, their physicochemical properties (pH, viscosity and surface tension) and medicinal activities as anti-dermatophyte against soil keratinophilic fungi at the molecular level were assessed. UV-visible and FTIR spectroscopic data of the biologically synthesized SeNPs were then recorded for confirming the presence of native biological materials adhered to nanoparticles, which are inherently required to enhance the stability and solubility through inhibition of the nanoparticle’s natural aggregation and agglomeration. The λ max value between 290-300 nm in the absorption spectra of the biogenic materials in different concentrations of the Na2SeO3 corroborated the presence of SeNPs in the solution. The interaction of SeNPs in solution state was further studied through the determination of pH, viscosity and surface tension values of agrobacterium-derived SeNPs in different solvents. The pH value of SeNPs dispersed in water is reported as above 7.0 and the average viscosity, and surface tensions of the SeNPs are appeared as near to the water. The particle size distribution was further determined by DLS and the highest % of particle size of the synthesized SeNPs is found in between 200-300 nm. The anti-dermatophyte activity and molecular interaction with fungi DNA molecules were assessed providing the highest anti-dermatophyte activity at 0.1 M concentration and it is observed that the quantities and qualities of fungi DNA were affected by SeNPs. Considering all the outcomes of the studies together, our findings suggest that agrobacterium-mediated synthesis of SeNPs is dependent on bacterial metabolisms but not on the concentration of Na2SeO3 and are promising selenium-derived species with potential application in the prevention of fungal infection through denaturation of fungi DNA.

  17. A New Shape Description Method Using Angular Radial Transform

    NASA Astrophysics Data System (ADS)

    Lee, Jong-Min; Kim, Whoi-Yul

    Shape is one of the primary low-level image features in content-based image retrieval. In this paper we propose a new shape description method that consists of a rotationally invariant angular radial transform descriptor (IARTD). The IARTD is a feature vector that combines the magnitude and aligned phases of the angular radial transform (ART) coefficients. A phase correction scheme is employed to produce the aligned phase so that the IARTD is invariant to rotation. The distance between two IARTDs is defined by combining differences in the magnitudes and aligned phases. In an experiment using the MPEG-7 shape dataset, the proposed method outperforms existing methods; the average BEP of the proposed method is 57.69%, while the average BEPs of the invariant Zernike moments descriptor and the traditional ART are 41.64% and 36.51%, respectively.

  18. Regulation of hydantoin-hydrolyzing enzyme expression in Agrobacterium tumefaciens strain RU-AE01.

    PubMed

    Jiwaji, Meesbah; Dorrington, Rosemary Ann

    2009-10-01

    Optically pure D-: amino acids, like D-: hydroxyphenylglycine, are used in the semi-synthetic production of pharmaceuticals. They are synthesized industrially via the biocatalytic hydrolysis of p-hydroxyphenylhydantoin using enzymes derived from Agrobacterium tumefaciens strains. The reaction proceeds via a three-step pathway: (a) the ring-opening cleavage of the hydantoin ring by a D-: hydantoinase (encoded by hyuH), (b) conversion of the resultant D-: N-carbamylamino acid to the corresponding amino acid by a D-: N-carbamoylase (encoded by hyuC), and (c) chemical or enzymatic racemization of the un-reacted hydantoin substrate. While the structure and biochemical properties of these enzymes are well understood, little is known about their origin, their function, and their regulation in the native host. We investigated the mechanisms involved in the regulation of expression of the hydantoinase and N-carbamoylase enzyme activity in A. tumefaciens strain RU-AE01. We present evidence for a complex regulatory network that responds to the growth status of the cells, the presence of inducer, and nitrogen catabolite repression. Deletion analysis and site-directed mutagenesis were used to identify regulatory elements involved in transcriptional regulation of hyuH and hyuC expression. Finally, a comparison between the hyu gene clusters in several Agrobacterium strains provides insight into the function of D-: selective hydantoin-hydrolyzing enzyme systems in Agrobacterium species.

  19. The Use of Continuous Wavelet Transform Based on the Fast Fourier Transform in the Analysis of Multi-channel Electrogastrography Recordings.

    PubMed

    Komorowski, Dariusz; Pietraszek, Stanislaw

    2016-01-01

    This paper presents the analysis of multi-channel electrogastrographic (EGG) signals using the continuous wavelet transform based on the fast Fourier transform (CWTFT). The EGG analysis was based on the determination of the several signal parameters such as dominant frequency (DF), dominant power (DP) and index of normogastria (NI). The use of continuous wavelet transform (CWT) allows for better visible localization of the frequency components in the analyzed signals, than commonly used short-time Fourier transform (STFT). Such an analysis is possible by means of a variable width window, which corresponds to the scale time of observation (analysis). Wavelet analysis allows using long time windows when we need more precise low-frequency information, and shorter when we need high frequency information. Since the classic CWT transform requires considerable computing power and time, especially while applying it to the analysis of long signals, the authors used the CWT analysis based on the fast Fourier transform (FFT). The CWT was obtained using properties of the circular convolution to improve the speed of calculation. This method allows to obtain results for relatively long records of EGG in a fairly short time, much faster than using the classical methods based on running spectrum analysis (RSA). In this study authors indicate the possibility of a parametric analysis of EGG signals using continuous wavelet transform which is the completely new solution. The results obtained with the described method are shown in the example of an analysis of four-channel EGG recordings, performed for a non-caloric meal.

  20. 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. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  1. Adaptive pseudo-color enhancement method of weld radiographic images based on HSI color space and self-transformation of pixels.

    PubMed

    Jiang, Hongquan; Zhao, Yalin; Gao, Jianmin; Gao, Zhiyong

    2017-06-01

    The radiographic testing (RT) image of a steam turbine manufacturing enterprise has the characteristics of low gray level, low contrast, and blurriness, which lead to a substandard image quality. Moreover, it is not conducive for human eyes to detect and evaluate defects. This study proposes an adaptive pseudo-color enhancement method for weld radiographic images based on the hue, saturation, and intensity (HSI) color space and the self-transformation of pixels to solve these problems. First, the pixel's self-transformation is performed to the pixel value of the original RT image. The function value after the pixel's self-transformation is assigned to the HSI components in the HSI color space. Thereafter, the average intensity of the enhanced image is adaptively adjusted to 0.5 according to the intensity of the original image. Moreover, the hue range and interval can be adjusted according to personal habits. Finally, the HSI components after the adaptive adjustment can be transformed to display in the red, green, and blue color space. Numerous weld radiographic images from a steam turbine manufacturing enterprise are used to validate the proposed method. The experimental results show that the proposed pseudo-color enhancement method can improve image definition and make the target and background areas distinct in weld radiographic images. The enhanced images will be more conducive for defect recognition. Moreover, the image enhanced using the proposed method conforms to the human eye visual properties, and the effectiveness of defect recognition and evaluation can be ensured.

  2. Adaptive pseudo-color enhancement method of weld radiographic images based on HSI color space and self-transformation of pixels

    NASA Astrophysics Data System (ADS)

    Jiang, Hongquan; Zhao, Yalin; Gao, Jianmin; Gao, Zhiyong

    2017-06-01

    The radiographic testing (RT) image of a steam turbine manufacturing enterprise has the characteristics of low gray level, low contrast, and blurriness, which lead to a substandard image quality. Moreover, it is not conducive for human eyes to detect and evaluate defects. This study proposes an adaptive pseudo-color enhancement method for weld radiographic images based on the hue, saturation, and intensity (HSI) color space and the self-transformation of pixels to solve these problems. First, the pixel's self-transformation is performed to the pixel value of the original RT image. The function value after the pixel's self-transformation is assigned to the HSI components in the HSI color space. Thereafter, the average intensity of the enhanced image is adaptively adjusted to 0.5 according to the intensity of the original image. Moreover, the hue range and interval can be adjusted according to personal habits. Finally, the HSI components after the adaptive adjustment can be transformed to display in the red, green, and blue color space. Numerous weld radiographic images from a steam turbine manufacturing enterprise are used to validate the proposed method. The experimental results show that the proposed pseudo-color enhancement method can improve image definition and make the target and background areas distinct in weld radiographic images. The enhanced images will be more conducive for defect recognition. Moreover, the image enhanced using the proposed method conforms to the human eye visual properties, and the effectiveness of defect recognition and evaluation can be ensured.

  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. Fourier transform methods in local gravity modeling

    NASA Technical Reports Server (NTRS)

    Harrison, J. C.; Dickinson, M.

    1989-01-01

    New algorithms were derived for computing terrain corrections, all components of the attraction of the topography at the topographic surface and the gradients of these attractions. These algoriithms utilize fast Fourier transforms, but, in contrast to methods currently in use, all divergences of the integrals are removed during the analysis. Sequential methods employing a smooth intermediate reference surface were developed to avoid the very large transforms necessary when making computations at high resolution over a wide area. A new method for the numerical solution of Molodensky's problem was developed to mitigate the convergence difficulties that occur at short wavelengths with methods based on a Taylor series expansion. A trial field on a level surface is continued analytically to the topographic surface, and compared with that predicted from gravity observations. The difference is used to compute a correction to the trial field and the process iterated. Special techniques are employed to speed convergence and prevent oscillations. Three different spectral methods for fitting a point-mass set to a gravity field given on a regular grid at constant elevation are described. Two of the methods differ in the way that the spectrum of the point-mass set, which extends to infinite wave number, is matched to that of the gravity field which is band-limited. The third method is essentially a space-domain technique in which Fourier methods are used to solve a set of simultaneous equations.

  5. Infrared and visual image fusion method based on discrete cosine transform and local spatial frequency in discrete stationary wavelet transform domain

    NASA Astrophysics Data System (ADS)

    Jin, Xin; Jiang, Qian; Yao, Shaowen; Zhou, Dongming; Nie, Rencan; Lee, Shin-Jye; He, Kangjian

    2018-01-01

    In order to promote the performance of infrared and visual image fusion and provide better visual effects, this paper proposes a hybrid fusion method for infrared and visual image by the combination of discrete stationary wavelet transform (DSWT), discrete cosine transform (DCT) and local spatial frequency (LSF). The proposed method has three key processing steps. Firstly, DSWT is employed to decompose the important features of the source image into a series of sub-images with different levels and spatial frequencies. Secondly, DCT is used to separate the significant details of the sub-images according to the energy of different frequencies. Thirdly, LSF is applied to enhance the regional features of DCT coefficients, and it can be helpful and useful for image feature extraction. Some frequently-used image fusion methods and evaluation metrics are employed to evaluate the validity of the proposed method. The experiments indicate that the proposed method can achieve good fusion effect, and it is more efficient than other conventional image fusion methods.

  6. Fast heap transform-based QR-decomposition of real and complex matrices: algorithms and codes

    NASA Astrophysics Data System (ADS)

    Grigoryan, Artyom M.

    2015-03-01

    In this paper, we describe a new look on the application of Givens rotations to the QR-decomposition problem, which is similar to the method of Householder transformations. We apply the concept of the discrete heap transform, or signal-induced unitary transforms which had been introduced by Grigoryan (2006) and used in signal and image processing. Both cases of real and complex nonsingular matrices are considered and examples of performing QR-decomposition of square matrices are given. The proposed method of QR-decomposition for the complex matrix is novel and differs from the known method of complex Givens rotation and is based on analytical equations for the heap transforms. Many examples illustrated the proposed heap transform method of QR-decomposition are given, algorithms are described in detail, and MATLAB-based codes are included.

  7. [Agrobacterium rubi strains from blueberry plants are highly diverse].

    PubMed

    Abrahamovich, Eliana; López, Ana C; Alippi, Adriana M

    2014-01-01

    The diversity of a collection of Agrobacterium rubi strains isolated from blueberries from different regions of Argentina was studied by conventional microbiological tests and molecular techniques. Results from biochemical and physiological reactions, as well as from rep-PCR and RFLP analysis of PCR-amplified 23S rDNA showed high phenotypic and genotypic intraspecific variation. Copyright © 2014 Asociación Argentina de Microbiología. Publicado por Elsevier España. All rights reserved.

  8. Feedback regulation of an Agrobacterium catalase gene katA involved in Agrobacterium-plant interaction.

    PubMed

    Xu, X Q; Li, L P; Pan, S Q

    2001-11-01

    Catalases are known to detoxify H2O2, a major component of oxidative stress imposed on a cell. An Agrobacterium tumefaciens catalase encoded by a chromosomal gene katA has been implicated as an important virulence factor as it is involved in detoxification of H2O2 released during Agrobacterium-plant interaction. In this paper, we report a feedback regulation pathway that controls the expression of katA in A. tumefaciens cells. We observed that katA could be induced by plant tissue sections and by acidic pH on a minimal medium, which resembles the plant environment that the bacteria encounter during the course of infection. This represents a new regulatory factor for catalase induction in bacteria. More importantly, a feedback regulation was observed when the katA-gfp expression was studied in different genetic backgrounds. We found that introduction of a wild-type katA gene encoding a functional catalase into A. tumefaciens cells could repress the katA-gfp expression over 60-fold. The katA gene could be induced by H2O2 and the encoded catalase could detoxify H2O2. In addition, the katA-gfp expression of one bacterial cell could be repressed by other surrounding catalase-proficient bacterial cells. Furthermore, mutation at katA caused a 10-fold increase of the intracellular H2O2 concentration in the bacteria grown on an acidic pH medium. These results suggest that the endogenous H2O2 generated during A. tumefaciens cell growth could serve as the intracellular and intercellular inducer for the katA gene expression and that the acidic pH could pose an oxidative stress on the bacteria. Surprisingly, one mutated KatA protein, exhibiting no significant catalase activity as a result of the alteration of two important residues at the putative active site, could partially repress the katA-gfp expression. The feedback regulation of the katA gene by both catalase activity and KatA protein could presumably maintain an appropriated level of catalase activity and H2O2 inside A

  9. [Measurements of the concentration of atmospheric CO2 based on OP/FTIR method and infrared reflecting scanning Fourier transform spectrometry].

    PubMed

    Wei, Ru-Yi; Zhou, Jin-Song; Zhang, Xue-Min; Yu, Tao; Gao, Xiao-Hui; Ren, Xiao-Qiang

    2014-11-01

    The present paper describes the observations and measurements of the infrared absorption spectra of CO2 on the Earth's surface with OP/FTIR method by employing a mid-infrared reflecting scanning Fourier transform spectrometry, which are the first results produced by the first prototype in China developed by the team of authors. This reflecting scanning Fourier transform spectrometry works in the spectral range 2 100-3 150 cm(-1) with a spectral resolution of 2 cm(-1). Method to measure the atmospheric molecules was described and mathematical proof and quantitative algorithms to retrieve molecular concentration were established. The related models were performed both by a direct method based on the Beer-Lambert Law and by a simulating-fitting method based on HITRAN database and the instrument functions. Concentrations of CO2 were retrieved by the two models. The results of observation and modeling analyses indicate that the concentrations have a distribution of 300-370 ppm, and show tendency that going with the variation of the environment they first decrease slowly and then increase rapidly during the observation period, and reached low points in the afternoon and during the sunset. The concentrations with measuring times retrieved by the direct method and by the simulating-fitting method agree with each other very well, with the correlation of all the data is up to 99.79%, and the relative error is no more than 2.00%. The precision for retrieving is relatively high. The results of this paper demonstrate that, in the field of detecting atmospheric compositions, OP/FTIR method performed by the Infrared reflecting scanning Fourier transform spectrometry is a feasible and effective technical approach, and either the direct method or the simulating-fitting method is capable of retrieving concentrations with high precision.

  10. Ex vitro composite plants: an inexpensive, rapid method for root biology.

    PubMed

    Collier, Ray; Fuchs, Beth; Walter, Nathalie; Kevin Lutke, William; Taylor, Christopher G

    2005-08-01

    Plant transformation technology is frequently the rate-limiting step in gene function analysis in non-model plants. An important tool for root biologists is the Agrobacterium rhizogenes-derived composite plant, which has made possible genetic analyses in a wide variety of transformation recalcitrant dicotyledonous plants. The novel, rapid and inexpensive ex vitro method for producing composite plants described in this report represents a significant advance over existing composite plant induction protocols, which rely on expensive and time-consuming in vitro conditions. The utility of the new system is validated by expression and RNAi silencing of GFP in transgenic roots of composite plants, and is bolstered further by experimental disruption, via RNAi silencing, of endogenous plant resistance to the plant parasitic nematode Meloidogyne incognita in transgenic roots of Lycopersicon esculentum cv. Motelle composite plants. Critical parameters of the method are described and discussed herein.

  11. Multi-focus image fusion based on area-based standard deviation in dual tree contourlet transform domain

    NASA Astrophysics Data System (ADS)

    Dong, Min; Dong, Chenghui; Guo, Miao; Wang, Zhe; Mu, Xiaomin

    2018-04-01

    Multiresolution-based methods, such as wavelet and Contourlet are usually used to image fusion. This work presents a new image fusion frame-work by utilizing area-based standard deviation in dual tree Contourlet trans-form domain. Firstly, the pre-registered source images are decomposed with dual tree Contourlet transform; low-pass and high-pass coefficients are obtained. Then, the low-pass bands are fused with weighted average based on area standard deviation rather than the simple "averaging" rule. While the high-pass bands are merged with the "max-absolute' fusion rule. Finally, the modified low-pass and high-pass coefficients are used to reconstruct the final fused image. The major advantage of the proposed fusion method over conventional fusion is the approximately shift invariance and multidirectional selectivity of dual tree Contourlet transform. The proposed method is compared with wavelet- , Contourletbased methods and other the state-of-the art methods on common used multi focus images. Experiments demonstrate that the proposed fusion framework is feasible and effective, and it performs better in both subjective and objective evaluation.

  12. Application of succulent plant leaves for Agrobacterium infiltration-mediated protein production.

    PubMed

    Jones, Richard W

    2016-01-01

    When expressing plant cell wall degrading enzymes in the widely used tobacco (Nicotiana benthamiana) after Agrobacterium infiltration, difficulties arise due to the thin leaf structure. Thick leaved succulents, Kalanchoe blossfeldiana and Hylotelephium telephium, were tested as alternatives. A xyloglucanase, as well as a xyloglucanase inhibitor protein was successfully produced. Published by Elsevier B.V.

  13. Metabolic engineering of Agrobacterium sp. strain ATCC 31749 for production of an α-Gal epitope

    PubMed Central

    2010-01-01

    Background Oligosaccharides containing a terminal Gal-α1,3-Gal moiety are collectively known as α-Gal epitopes. α-Gal epitopes are integral components of several medical treatments under development, including flu and HIV vaccines as well as cancer treatments. The difficulty associated with synthesizing the α-Gal epitope hinders the development and application of these treatments due to the limited availability and high cost of the α-Gal epitope. This work illustrates the development of a whole-cell biocatalyst for synthesizing the α-Gal epitope, Gal-α1,3-Lac. Results Agrobacterium sp. ATCC 31749 was engineered to produce Gal-α1,3-Lac by the introduction of a UDP-galactose 4'-epimerase:α1,3-galactosyltransferase fusion enzyme. The engineered Agrobacterium synthesized 0.4 g/L of the α-Gal epitope. Additional metabolic engineering efforts addressed the factors limiting α-Gal epitope production, namely the availability of the two substrates, lactose and UDP-glucose. Through expression of a lactose permease, the intracellular lactose concentration increased by 60 to 110%, subsequently leading to an improvement in Gal-α1,3-Lac production. Knockout of the curdlan synthase gene increased UDP-glucose availability by eliminating the consumption of UDP-glucose for synthesis of the curdlan polysaccharide. With these additional engineering efforts, the final engineered strain synthesized approximately 1 g/L of Gal-α1,3-Lac. Conclusions The Agrobacterium biocatalyst developed in this work synthesizes gram-scale quantities of α-Gal epitope and does not require expensive cofactors or permeabilization, making it a useful biocatalyst for industrial production of the α-Gal epitope. Furthermore, the engineered Agrobacterium, with increased lactose uptake and improved UDP-glucose availability, is a promising host for the production of other medically-relevant oligosaccharides. PMID:20067629

  14. Plasmid-dependent attachment of Agrobacterium tumefaciens to plant tissue culture cells.

    PubMed

    Matthysse, A G; Wyman, P M; Holmes, K V

    1978-11-01

    Kinetic, microscopic, and biochemical studies show that virulent Ti (tumor inducing)-plasmid-containing strains of Agrobacterium attach to normal tobacco and carrot tissue culture cells. Kinetic studies showed that virulent strains of A. tumefaciens attach to the plant tissue culture cells in increasing numbers during the first 1 to 2 h of incubation of the bacteria with the plant cells. Five Ti-plasmid-containing virulent Agrobacterium strains showed greater attachment to tobacco cells than did five avirulent strains. Light and scanning electron microscopic observations confirmed that virulent strains showed little attachment. Bacterial attachment was blocked by prior incubation of the plant cells with lipopolysaccharide extracted from A. tumefaciens, but not from A. radiobacter, suggesting that bacterial lipopolysaccharide is one of the components involved in the attachment process. At least one other bacterial product may be required for attachment in tissue culture because the virulent A. tumefaciens NT1, which lacks the Ti plasmid, does not itself attach to tobacco cells, but its lipopolysaccharide does inhibit the attachment of virulent strains.

  15. Plasmid-dependent attachment of Agrobacterium tumefaciens to plant tissue culture cells.

    PubMed Central

    Matthysse, A G; Wyman, P M; Holmes, K V

    1978-01-01

    Kinetic, microscopic, and biochemical studies show that virulent Ti (tumor inducing)-plasmid-containing strains of Agrobacterium attach to normal tobacco and carrot tissue culture cells. Kinetic studies showed that virulent strains of A. tumefaciens attach to the plant tissue culture cells in increasing numbers during the first 1 to 2 h of incubation of the bacteria with the plant cells. Five Ti-plasmid-containing virulent Agrobacterium strains showed greater attachment to tobacco cells than did five avirulent strains. Light and scanning electron microscopic observations confirmed that virulent strains showed little attachment. Bacterial attachment was blocked by prior incubation of the plant cells with lipopolysaccharide extracted from A. tumefaciens, but not from A. radiobacter, suggesting that bacterial lipopolysaccharide is one of the components involved in the attachment process. At least one other bacterial product may be required for attachment in tissue culture because the virulent A. tumefaciens NT1, which lacks the Ti plasmid, does not itself attach to tobacco cells, but its lipopolysaccharide does inhibit the attachment of virulent strains. Images PMID:730370

  16. A Wavelet-based Fast Discrimination of Transformer Magnetizing Inrush Current

    NASA Astrophysics Data System (ADS)

    Kitayama, Masashi

    Recently customers who need electricity of higher quality have been installing co-generation facilities. They can avoid voltage sags and other distribution system related disturbances by supplying electricity to important load from their generators. For another example, FRIENDS, highly reliable distribution system using semiconductor switches or storage devices based on power electronics technology, is proposed. These examples illustrates that the request for high reliability in distribution system is increasing. In order to realize these systems, fast relaying algorithms are indispensable. The author proposes a new method of detecting magnetizing inrush current using discrete wavelet transform (DWT). DWT provides the function of detecting discontinuity of current waveform. Inrush current occurs when transformer core becomes saturated. The proposed method detects spikes of DWT components derived from the discontinuity of the current waveform at both the beginning and the end of inrush current. Wavelet thresholding, one of the wavelet-based statistical modeling, was applied to detect the DWT component spikes. The proposed method is verified using experimental data using single-phase transformer and the proposed method is proved to be effective.

  17. Better Rooting Procedure to Enhance Survival Rate of Field Grown Malaysian Eksotika Papaya Transformed with 1-Aminocyclopropane-1-Carboxylic Acid Oxidase Gene

    PubMed Central

    Sekeli, Rogayah; Abdullah, Janna Ong; Namasivayam, Parameswari; Muda, Pauziah; Abu Bakar, Umi Kalsom

    2013-01-01

    A high survival rate for transformed papaya plants when transferred to the field is useful in the quest for improving the commercial quality traits. We report in this paper an improved rooting method for the production of transformed Malaysian Eksotika papaya with high survival rate when transferred to the field. Shoots were regenerated from embryogenic calli transformed with antisense and RNAi constructs of 1-aminocyclopropane-1-carboxylic acid oxidase (ACO) genes using the Agrobacterium tumefaciens-mediated transformation method. Regenerated transformed shoots, each measuring approximately 3-4 cm in height, were cultured in liquid half-strength Murashige and Skoog (MS) medium or sterile distilled water, and with either perlite or vermiculite supplementation. All the culturing processes were conducted either under sterile or nonsterile condition. The results showed that rooting under sterile condition was better. Shoots cultured in half-strength MS medium supplemented with vermiculite exhibited a 92.5% rooting efficiency while perlite showed 77.5%. The survival rate of the vermiculite-grown transformed papaya plantlets after transfer into soil, contained in polybags, was 94%, and the rate after transfer into the ground was 92%. Morpho-histological analyses revealed that the tap roots were more compact, which might have contributed to the high survival rates of the plantlets. PMID:25969786

  18. Better rooting procedure to enhance survival rate of field grown malaysian eksotika papaya transformed with 1-aminocyclopropane-1-carboxylic Acid oxidase gene.

    PubMed

    Sekeli, Rogayah; Abdullah, Janna Ong; Namasivayam, Parameswari; Muda, Pauziah; Abu Bakar, Umi Kalsom

    2013-01-01

    A high survival rate for transformed papaya plants when transferred to the field is useful in the quest for improving the commercial quality traits. We report in this paper an improved rooting method for the production of transformed Malaysian Eksotika papaya with high survival rate when transferred to the field. Shoots were regenerated from embryogenic calli transformed with antisense and RNAi constructs of 1-aminocyclopropane-1-carboxylic acid oxidase (ACO) genes using the Agrobacterium tumefaciens-mediated transformation method. Regenerated transformed shoots, each measuring approximately 3-4 cm in height, were cultured in liquid half-strength Murashige and Skoog (MS) medium or sterile distilled water, and with either perlite or vermiculite supplementation. All the culturing processes were conducted either under sterile or nonsterile condition. The results showed that rooting under sterile condition was better. Shoots cultured in half-strength MS medium supplemented with vermiculite exhibited a 92.5% rooting efficiency while perlite showed 77.5%. The survival rate of the vermiculite-grown transformed papaya plantlets after transfer into soil, contained in polybags, was 94%, and the rate after transfer into the ground was 92%. Morpho-histological analyses revealed that the tap roots were more compact, which might have contributed to the high survival rates of the plantlets.

  19. Quantification of Agrobacterium tumefaciens C58 attachment to Arabidopsis thaliana roots.

    PubMed

    Petrovicheva, Anna; Joyner, Jessica; Muth, Theodore R

    2017-10-02

    Agrobacterium tumefaciens is the causal agent of crown gall disease and is a vector for DNA transfer in transgenic plants. The transformation process by A. tumefaciens has been widely studied, but the attachment stage has not been well characterized. Most measurements of attachment have used microscopy and colony counting, both of which are labor and time intensive. To reduce the time and effort required to analyze bacteria attaching to plant tissues, we developed a quantitative real-time PCR (qPCR) assay to quantify attached A. tumefaciens using the chvE gene as marker for the presence of the bacteria. The qPCR detection threshold of A. tumefaciens from pure culture was 104 cell equivalents/ml. The A. tumefaciens minimum threshold concentration from root-bound populations was determined to be 105 cell equivalents/ml inoculum to detect attachment above background. The qPCR assay can be used for measuring A. tumefaciens attachment in applications such as testing the effects of mutations on bacterial adhesion molecules or biofilm formation, comparing attachment across various plant species and ecotypes, and detecting mutations in putative attachment receptors expressed in plant roots. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. A novel analysis method for near infrared spectroscopy based on Hilbert-Huang transform

    NASA Astrophysics Data System (ADS)

    Zhou, Zhenyu; Yang, Hongyu; Liu, Yun; Ruan, Zongcai; Luo, Qingming; Gong, Hui; Lu, Zuhong

    2007-05-01

    Near Infrared Imager (NIRI) has been widely used to access the brain functional activity non-invasively. We use a portable, multi-channel and continuous-wave NIR topography instrument to measure the concentration changes of each hemoglobin species and map cerebral cortex functional activation. By extracting some essential features from the BOLD signals, optical tomography is able to be a new way of neuropsychological studies. Fourier spectral analysis provides a common framework for examining the distribution of global energy in the frequency domain. However, this method assumes that the signal should be stationary, which limits its application in non-stationary system. The hemoglobin species concentration changes are of such kind. In this work we develop a new signal processing method using Hilbert-Huang transform to perform spectral analysis of the functional NIRI signals. Compared with wavelet based multi-resolution analysis (MRA), we demonstrated the extraction of task related signal for observation of activation in the prefrontal cortex (PFC) in vision stimulation experiment. This method provides a new analysis tool for functional NIRI signals. Our experimental results show that the proposed approach provides the unique method for reconstructing target signal without losing original information and enables us to understand the episode of functional NIRI more precisely.

  1. Superfast algorithms of multidimensional discrete k-wave transforms and Volterra filtering based on superfast radon transform

    NASA Astrophysics Data System (ADS)

    Labunets, Valeri G.; Labunets-Rundblad, Ekaterina V.; Astola, Jaakko T.

    2001-12-01

    Fast algorithms for a wide class of non-separable n-dimensional (nD) discrete unitary K-transforms (DKT) are introduced. They need less 1D DKTs than in the case of the classical radix-2 FFT-type approach. The method utilizes a decomposition of the nD K-transform into the product of a new nD discrete Radon transform and of a set of parallel/independ 1D K-transforms. If the nD K-transform has a separable kernel (e.g., the case of the discrete Fourier transform) our approach leads to decrease of multiplicative complexity by the factor of n comparing to the classical row/column separable approach. It is well known that an n-th order Volterra filter of one dimensional signal can be evaluated by an appropriate nD linear convolution. This work describes new superfast algorithm for Volterra filtering. New approach is based on the superfast discrete Radon and Nussbaumer polynomial transforms.

  2. Plant phosphomannose isomerase as a selectable marker for rice transformation

    PubMed Central

    Hu, Lei; Li, Hao; Qin, Ruiying; Xu, Rongfang; Li, Juan; Li, Li; Wei, Pengcheng; Yang, Jianbo

    2016-01-01

    The E. coli phosphomannose isomerase (EcPMI) gene is widely used as a selectable marker gene (SMG) in mannose (Man) selection-based plant transformation. Although some plant species exhibit significant PMI activity and active PMIs were even identified in Man-sensitive plants, whether plant PMIs can be used as SMGs remains unclear. In this study, we isolated four novel PMI genes from Chlorella variabilis and Oryza sativa. Their isoenzymatic activities were examined in vitro and compared with that of EcPMI. The active plant PMIs were separately constructed into binary vectors as SMGs and then transformed into rice via Agrobacterium. In both Indica and Japonica subspecies, our results indicated that the plant PMIs could select and produce transgenic plants in a pattern similar to that of EcPMI. The transgenic plants exhibited an accumulation of plant PMI transcripts and enhancement of the in vivo PMI activity. Furthermore, a gene of interest was successfully transformed into rice using the plant PMIs as SMGs. Thus, novel SMGs for Man selection were isolated from plants, and our analysis suggested that PMIs encoding active enzymes might be common in plants and could potentially be used as appropriate genetic elements in cisgenesis engineering. PMID:27174847

  3. DETECTION AND IMPLICATIONS OF EARLY AGROBACTERIUM TUMEFACIENS INFECTION OF PARADOX SEEDS AND SEEDLINGS

    USDA-ARS?s Scientific Manuscript database

    Paradox (Juglans hindsii x J. regia), the dominant rootstock used in California, USA walnut production, has many desirable horticultural characteristics, but is highly susceptible to crown gall. Crown gall, caused by the soil-borne bacterium Agrobacterium tumefaciens, can not be consistently control...

  4. X-ray Structure of Imidazolonepropionase from Agrobacterium tumefaciens at 1.87 angstrom Resolution

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tyagi,R.; Kumaran, D.; Burley, S.

    2007-01-01

    Histidine degradation in agrobacterium tumefaciens involves four enzymes, including histidase, urocanase, imidazolonepropionase, and N-formylglutamate amido hydrolase. The third enzyme of the pathway, imidazolone-propionase, a 45.6 kDa protein, catalyzes conversion of imidazolone-5-propanoate to N-forminio-t-glutamate.

  5. Bioluminescence-based system for rapid detection of natural transformation.

    PubMed

    Santala, Ville; Karp, Matti; Santala, Suvi

    2016-07-01

    Horizontal gene transfer plays a significant role in bacterial evolution and has major clinical importance. Thus, it is vital to understand the mechanisms and kinetics of genetic transformations. Natural transformation is the driving mechanism for horizontal gene transfer in diverse genera of bacteria. Our study introduces a simple and rapid method for the investigation of natural transformation. This highly sensitive system allows the detection of a transformation event directly from a bacterial population without any separation step or selection of cells. The system is based on the bacterial luciferase operon from Photorhabdus luminescens The studied molecular tools consist of the functional modules luxCDE and luxAB, which involve a replicative plasmid and an integrative gene cassette. A well-established host for bacterial genetic investigations, Acinetobacter baylyi ADP1, is used as the model bacterium. We show that natural transformation followed by homologous recombination or plasmid recircularization can be readily detected in both actively growing and static biofilm-like cultures, including very rare transformation events. The system allows the detection of natural transformation within 1 h of introducing sample DNA into the culture. The introduced method provides a convenient means to study the kinetics of natural transformation under variable conditions and perturbations. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  6. A new stationary gridline artifact suppression method based on the 2D discrete wavelet transform

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tang, Hui, E-mail: corinna@seu.edu.cn; Key Laboratory of Computer Network and Information Integration; Centre de Recherche en Information Biomédicale sino-français, Laboratoire International Associé, Inserm, Université de Rennes 1, Rennes 35000

    2015-04-15

    Purpose: In digital x-ray radiography, an antiscatter grid is inserted between the patient and the image receptor to reduce scattered radiation. If the antiscatter grid is used in a stationary way, gridline artifacts will appear in the final image. In most of the gridline removal image processing methods, the useful information with spatial frequencies close to that of the gridline is usually lost or degraded. In this study, a new stationary gridline suppression method is designed to preserve more of the useful information. Methods: The method is as follows. The input image is first recursively decomposed into several smaller subimagesmore » using a multiscale 2D discrete wavelet transform. The decomposition process stops when the gridline signal is found to be greater than a threshold in one or several of these subimages using a gridline detection module. An automatic Gaussian band-stop filter is then applied to the detected subimages to remove the gridline signal. Finally, the restored image is achieved using the corresponding 2D inverse discrete wavelet transform. Results: The processed images show that the proposed method can remove the gridline signal efficiently while maintaining the image details. The spectra of a 1D Fourier transform of the processed images demonstrate that, compared with some existing gridline removal methods, the proposed method has better information preservation after the removal of the gridline artifacts. Additionally, the performance speed is relatively high. Conclusions: The experimental results demonstrate the efficiency of the proposed method. Compared with some existing gridline removal methods, the proposed method can preserve more information within an acceptable execution time.« less

  7. Iodate and nitrate transformation by Agrobacterium/Rhizobium related strain DVZ35 isolated from contaminated Hanford groundwater.

    PubMed

    Lee, Brady D; Ellis, Joshua T; Dodwell, Alex; Eisenhauer, Emalee E R; Saunders, Danielle L; Lee, M Hope

    2018-05-15

    Nitrate and radioiodine ( 129 I) contamination is widespread in groundwater underneath the Central Plateau of the Hanford Site. 129 I, a byproduct of nuclear fission, is of concern due to a 15.7 million year half-life, and toxicity. The Hanford 200 West Area contains plumes covering 4.3 km 2 with average 129 I concentrations of 3.5 pCi/L. Iodate accounts for 70.6% of the iodine present and organo-iodine and iodide make up 25.8% and 3.6%, respectively. Nitrate plumes encompassing the 129 I plumes have a surface area of 16 km 2 averaging 130 mg/L. A nitrate and iodate reducing bacterium closely related to Agrobacterium, strain DVZ35, was isolated from sediment incubated in a 129 I plume. Iodate removal efficiency was 36.3% in transition cultures, and 47.8% in anaerobic cultures. Nitrate (10 mM) was also reduced in the microcosm. When nitrate was spiked into the microcosms, iodate removal efficiency was 84.0% and 69.2% in transition and anaerobic cultures, respectively. Iodate reduction was lacking when nitrate was absent from the growth medium. These data indicate there is simultaneous reduction of nitrate and iodate by DVZ35, and iodate is reduced to iodide. Results provide the scientific basis for combined nitrogen and iodine cycling throughout the Hanford Site. Copyright © 2018. Published by Elsevier B.V.

  8. An efficient depth map preprocessing method based on structure-aided domain transform smoothing for 3D view generation

    PubMed Central

    Ma, Liyan; Qiu, Bo; Cui, Mingyue; Ding, Jianwei

    2017-01-01

    Depth image-based rendering (DIBR), which is used to render virtual views with a color image and the corresponding depth map, is one of the key techniques in the 2D to 3D conversion process. Due to the absence of knowledge about the 3D structure of a scene and its corresponding texture, DIBR in the 2D to 3D conversion process, inevitably leads to holes in the resulting 3D image as a result of newly-exposed areas. In this paper, we proposed a structure-aided depth map preprocessing framework in the transformed domain, which is inspired by recently proposed domain transform for its low complexity and high efficiency. Firstly, our framework integrates hybrid constraints including scene structure, edge consistency and visual saliency information in the transformed domain to improve the performance of depth map preprocess in an implicit way. Then, adaptive smooth localization is cooperated and realized in the proposed framework to further reduce over-smoothness and enhance optimization in the non-hole regions. Different from the other similar methods, the proposed method can simultaneously achieve the effects of hole filling, edge correction and local smoothing for typical depth maps in a united framework. Thanks to these advantages, it can yield visually satisfactory results with less computational complexity for high quality 2D to 3D conversion. Numerical experimental results demonstrate the excellent performances of the proposed method. PMID:28407027

  9. A method for encapsulating high voltage power transformers

    NASA Astrophysics Data System (ADS)

    Sanchez, Robert O.

    Voltage breakdowns become a major concern in reducing the size of high-voltage power converter transformers. Even the smallest of voids can provide a path for corona discharge which can cause a dielectric breakdown leading to a transformer failure. A method of encapsulating small high voltage transformers has been developed. The method virtually eliminates voids in the impregnation material, provides an exceptional dielectric between windings and provides a mechanically rugged package. The encapsulation material is a carboxyl terminated butadiene nitril (CTBN) modified mica filled epoxy. The method requires heat/vacuum to impregnate the coil and heat/pressure to cure the encapsulant. The transformer package utilizes a diallyl phthalate (DAP) contact assembly in which a coated core/coil assembly is mounted and soldered. This assembly is then loaded into an RTV mold and the encapsulation process begins.

  10. A method for surface topography measurement using a new focus function based on dual-tree complex wavelet transform

    NASA Astrophysics Data System (ADS)

    Li, Shimiao; Guo, Tong; Yuan, Lin; Chen, Jinping

    2018-01-01

    Surface topography measurement is an important tool widely used in many fields to determine the characteristics and functionality of a part or material. Among existing methods for this purpose, the focus variation method has proved high performance particularly in large slope scenarios. However, its performance depends largely on the effectiveness of focus function. This paper presents a method for surface topography measurement using a new focus measurement function based on dual-tree complex wavelet transform. Experiments are conducted on simulated defocused images to prove its high performance in comparison with other traditional approaches. The results showed that the new algorithm has better unimodality and sharpness. The method was also verified by measuring a MEMS micro resonator structure.

  11. Blueberry (Vaccinium corymbosum L.).

    PubMed

    Song, Guo-Qing; Sink, Kenneth C

    2006-01-01

    Recent advances in plant biotechnology have led to a reliable and reproductive method for genetic transformation of blueberry. These efforts built on previous attempts at transient and stable transformation of blueberry that demonstrated the potential of Agrobacterium tumefaciens-mediated transformation, and as well, the difficulties of selecting and regenerating transgenic plants. As a prerequisite for successful stable transformation, efficient regeneration systems were required despite many reports on factors controlling shoot regeneration from leaf explants. The A. tumefaciens-mediated transformation protocol described in this chapter is based on combining efficient regeneration methods and the results of A. tumefaciens-mediated transient transformation studies to optimize selected parameters for gene transfer. The protocol has led to successful regeneration of transgenic plants of four commercially important highbush blueberry cultivars.

  12. Novel primers for detection of genetically diverse virulent Agrobacterium tumefaciens bv1 strains

    USDA-ARS?s Scientific Manuscript database

    Novel primers were developed to amplify a 243 bp fragment of an intergenic region between gene5 and tms2 on the T-DNA of Agrobacterium tumefaciens. These primers exhibit 100% positive correlation with strain virulence, 100% negative correlation with avirulence and did not generate extraneous bands,...

  13. The Hilbert-Huang Transform-Based Denoising Method for the TEM Response of a PRBS Source Signal

    NASA Astrophysics Data System (ADS)

    Hai, Li; Guo-qiang, Xue; Pan, Zhao; Hua-sen, Zhong; Khan, Muhammad Younis

    2016-08-01

    The denoising process is critical in processing transient electromagnetic (TEM) sounding data. For the full waveform pseudo-random binary sequences (PRBS) response, an inadequate noise estimation may result in an erroneous interpretation. We consider the Hilbert-Huang transform (HHT) and its application to suppress the noise in the PRBS response. The focus is on the thresholding scheme to suppress the noise and the analysis of the signal based on its Hilbert time-frequency representation. The method first decomposes the signal into the intrinsic mode function, and then, inspired by the thresholding scheme in wavelet analysis; an adaptive and interval thresholding is conducted to set to zero all the components in intrinsic mode function which are lower than a threshold related to the noise level. The algorithm is based on the characteristic of the PRBS response. The HHT-based denoising scheme is tested on the synthetic and field data with the different noise levels. The result shows that the proposed method has a good capability in denoising and detail preservation.

  14. Deficiencies of the cryptography based on multiple-parameter fractional Fourier transform.

    PubMed

    Ran, Qiwen; Zhang, Haiying; Zhang, Jin; Tan, Liying; Ma, Jing

    2009-06-01

    Methods of image encryption based on fractional Fourier transform have an incipient flaw in security. We show that the schemes have the deficiency that one group of encryption keys has many groups of keys to decrypt the encrypted image correctly for several reasons. In some schemes, many factors result in the deficiencies, such as the encryption scheme based on multiple-parameter fractional Fourier transform [Opt. Lett.33, 581 (2008)]. A modified method is proposed to avoid all the deficiencies. Security and reliability are greatly improved without increasing the complexity of the encryption process. (c) 2009 Optical Society of America.

  15. Phase synchronization based on a Dual-Tree Complex Wavelet Transform

    NASA Astrophysics Data System (ADS)

    Ferreira, Maria Teodora; Domingues, Margarete Oliveira; Macau, Elbert E. N.

    2016-11-01

    In this work, we show the applicability of our Discrete Complex Wavelet Approach (DCWA) to verify the phenomenon of phase synchronization transition in two coupled chaotic Lorenz systems. DCWA is based on the phase assignment from complex wavelet coefficients obtained by using a Dual-Tree Complex Wavelet Transform (DT-CWT). We analyzed two coupled chaotic Lorenz systems, aiming to detect the transition from non-phase synchronization to phase synchronization. In addition, we check how good is the method in detecting periods of 2π phase-slips. In all experiments, DCWA is compared with classical phase detection methods such as the ones based on arctangent and Hilbert transform showing a much better performance.

  16. Use of Natural Transformation To Establish an Easy Knockout Method in Riemerella anatipestifer.

    PubMed

    Liu, MaFeng; Zhang, Li; Huang, Li; Biville, Francis; Zhu, DeKang; Wang, MingShu; Jia, RenYong; Chen, Shun; Sun, KunFeng; Yang, Qiao; Wu, Ying; Chen, XiaoYue; Cheng, AnChun

    2017-05-01

    Riemerella anatipestifer is a member of the family Flavobacteriaceae and a major causative agent of duck serositis. Little is known about its genetics and pathogenesis. Several bacteria are competent for natural transformation; however, whether R. anatipestifer is also competent for natural transformation has not been investigated. Here, we showed that R. anatipestifer strain ATCC 11845 can uptake the chromosomal DNA of R. anatipestifer strain RA-CH-1 in all growth phases. Subsequently, a natural transformation-based knockout method was established for R. anatipestifer ATCC 11845. Targeted mutagenesis gave transformation frequencies of ∼10 -5 transformants. Competition assay experiments showed that R. anatipestifer ATCC 11845 preferentially took up its own DNA rather than heterogeneous DNA, such as Escherichia coli DNA. Transformation was less efficient with the shuttle plasmid pLMF03 (transformation frequencies of ∼10 -9 transformants). However, the efficiency of transformation was increased approximately 100-fold using pLMF03 derivatives containing R. anatipestifer DNA fragments (transformation frequencies of ∼10 -7 transformants). Finally, we found that the R. anatipestifer RA-CH-1 strain was also naturally transformable, suggesting that natural competence is widely applicable for this species. The findings described here provide important tools for the genetic manipulation of R. anatipestifer IMPORTANCE Riemerella anatipestifer is an important duck pathogen that belongs to the family Flavobacteriaceae At least 21 different serotypes have been identified. Genetic diversity has been demonstrated among these serotypes. The genetic and pathogenic mechanisms of R. anatipestifer remain largely unknown because no genetic tools are available for this bacterium. At present, natural transformation has been found in some bacteria but not in R. anatipestifer For the first time, we showed that natural transformation occurred in R. anatipestifer ATCC 11845 and R

  17. A simple transformation independent method for outlier definition.

    PubMed

    Johansen, Martin Berg; Christensen, Peter Astrup

    2018-04-10

    Definition and elimination of outliers is a key element for medical laboratories establishing or verifying reference intervals (RIs). Especially as inclusion of just a few outlying observations may seriously affect the determination of the reference limits. Many methods have been developed for definition of outliers. Several of these methods are developed for the normal distribution and often data require transformation before outlier elimination. We have developed a non-parametric transformation independent outlier definition. The new method relies on drawing reproducible histograms. This is done by using defined bin sizes above and below the median. The method is compared to the method recommended by CLSI/IFCC, which uses Box-Cox transformation (BCT) and Tukey's fences for outlier definition. The comparison is done on eight simulated distributions and an indirect clinical datasets. The comparison on simulated distributions shows that without outliers added the recommended method in general defines fewer outliers. However, when outliers are added on one side the proposed method often produces better results. With outliers on both sides the methods are equally good. Furthermore, it is found that the presence of outliers affects the BCT, and subsequently affects the determined limits of current recommended methods. This is especially seen in skewed distributions. The proposed outlier definition reproduced current RI limits on clinical data containing outliers. We find our simple transformation independent outlier detection method as good as or better than the currently recommended methods.

  18. Crystal structure of the Agrobacterium virulence complex VirE1-VirE2 reveals a flexible protein that can accommodate different partners.

    PubMed

    Dym, Orly; Albeck, Shira; Unger, Tamar; Jacobovitch, Jossef; Branzburg, Anna; Michael, Yigal; Frenkiel-Krispin, Daphna; Wolf, Sharon Grayer; Elbaum, Michael

    2008-08-12

    Agrobacterium tumefaciens infects its plant hosts by a mechanism of horizontal gene transfer. This capability has led to its widespread use in artificial genetic transformation. In addition to DNA, the bacterium delivers an abundant ssDNA binding protein, VirE2, whose roles in the host include protection from cytoplasmic nucleases and adaptation for nuclear import. In Agrobacterium, VirE2 is bound to its acidic chaperone VirE1. When expressed in vitro in the absence of VirE1, VirE2 is prone to oligomerization and forms disordered filamentous aggregates. These filaments adopt an ordered solenoidal form in the presence of ssDNA, which was characterized previously by electron microscopy and three-dimensional image processing. VirE2 coexpressed in vitro with VirE1 forms a soluble heterodimer. VirE1 thus prevents VirE2 oligomerization and competes with its binding to ssDNA. We present here a crystal structure of VirE2 in complex with VirE1, showing that VirE2 is composed of two independent domains presenting a novel fold, joined by a flexible linker. Electrostatic interactions with VirE1 cement the two domains of VirE2 into a locked form. Comparison with the electron microscopy structure indicates that the VirE2 domains adopt different relative orientations. We suggest that the flexible linker between the domains enables VirE2 to accommodate its different binding partners.

  19. Determination of sulfur compounds in hydrotreated transformer base oil by potentiometric titration.

    PubMed

    Chao, Qiu; Sheng, Han; Cheng, Xingguo; Ren, Tianhui

    2005-06-01

    A method was developed to analyze the distribution of sulfur compounds in model sulfur compounds by potentiometric titration, and applied to analyze hydrotreated transformer base oil. Model thioethers were oxidized to corresponding sulfoxides by tetrabutylammonium periodate and sodium metaperiodate, respectively, and the sulfoxides were titrated by perchloric acid titrant in acetic anhydride. The contents of aliphatic thioethers and total thioethers were then determined from that of sulfoxides in solution. The method was applied to determine the organic sulfur compounds in hydrotreated transformer base oil.

  20. Physical methods for genetic plant transformation

    NASA Astrophysics Data System (ADS)

    Rivera, Ana Leonor; Gómez-Lim, Miguel; Fernández, Francisco; Loske, Achim M.

    2012-09-01

    Production of transgenic plants is a routine process for many crop species. Transgenes are introduced into plants to confer novel traits such as improved nutritional qualities, tolerance to pollutants, resistance to pathogens and for studies of plant metabolism. Nowadays, it is possible to insert genes from plants evolutionary distant from the host plant, as well as from fungi, viruses, bacteria and even animals. Genetic transformation requires penetration of the transgene through the plant cell wall, facilitated by biological or physical methods. The objective of this article is to review the state of the art of the physical methods used for genetic plant transformation and to describe the basic physics behind them.

  1. 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. © 2013.

  2. Theoretical Investigation of Thermo-Mechanical Behavior of Carbon Nanotube-Based Composites Using the Integral Transform Method

    NASA Technical Reports Server (NTRS)

    Pawloski, Janice S.

    2001-01-01

    This project uses the integral transform technique to model the problem of nanotube behavior as an axially symmetric system of shells. Assuming that the nanotube behavior can be described by the equations of elasticity, we seek a stress function x which satisfies the biharmonic equation: del(exp 4) chi = [partial deriv(r(exp 2)) + partial deriv(r) + partial deriv(z(exp 2))] chi = 0. The method of integral transformations is used to transform the differential equation. The symmetry with respect to the z-axis indicates that we only need to consider the sine transform of the stress function: X(bar)(r,zeta) = integral(from 0 to infinity) chi(r,z)sin(zeta,z) dz.

  3. The putative Agrobacterium transcriptional activator-like virulence protein VirD5 may target T-complex to prevent the degradation of coat proteins in the plant cell nucleus.

    PubMed

    Wang, Yafei; Peng, Wei; Zhou, Xu; Huang, Fei; Shao, Lingyun; Luo, Meizhong

    2014-09-01

    Agrobacterium exports at least five virulence proteins (VirE2, VirE3, VirF, VirD2, VirD5) into host cells and hijacks some host plant factors to facilitate its transformation process. Random DNA binding selection assays (RDSAs), electrophoretic mobility shift assays (EMSAs) and yeast one-hybrid systems were used to identify protein-bound DNA elements. Bimolecular fluorescence complementation, glutathione S-transferase pull-down and yeast two-hybrid assays were used to detect protein interactions. Protoplast transformation, coprecipitation, competitive binding and cell-free degradation assays were used to analyze the relationships among proteins. We found that Agrobacterium VirD5 exhibits transcriptional activation activity in yeast, is located in the plant cell nucleus, and forms homodimers. A specific VirD5-bound DNA element designated D5RE (VirD5 response element) was identified. VirD5 interacted directly with Arabidopsis VirE2 Interacting Protein 1 (AtVIP1). However, the ternary complex of VirD5-AtVIP1-VirE2 could be detected, whereas that of VirD5-AtVIP1-VBF (AtVIP1 Binding F-box protein) could not. We demonstrated that VirD5 competes with VBF for binding to AtVIP1 and stabilizes AtVIP1 and VirE2 in the cell-free degradation system. Our results indicated that VirD5 may act as both a transcriptional activator-like effector to regulate host gene expression and a protector preventing the coat proteins of the T-complex from being quickly degraded by the host's ubiquitin proteasome system (UPS). © 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.

  4. Coronary arteries segmentation based on the 3D discrete wavelet transform and 3D neutrosophic transform.

    PubMed

    Chen, Shuo-Tsung; Wang, Tzung-Dau; Lee, Wen-Jeng; Huang, Tsai-Wei; Hung, Pei-Kai; Wei, Cheng-Yu; Chen, Chung-Ming; Kung, Woon-Man

    2015-01-01

    Most applications in the field of medical image processing require precise estimation. To improve the accuracy of segmentation, this study aimed to propose a novel segmentation method for coronary arteries to allow for the automatic and accurate detection of coronary pathologies. The proposed segmentation method included 2 parts. First, 3D region growing was applied to give the initial segmentation of coronary arteries. Next, the location of vessel information, HHH subband coefficients of the 3D DWT, was detected by the proposed vessel-texture discrimination algorithm. Based on the initial segmentation, 3D DWT integrated with the 3D neutrosophic transformation could accurately detect the coronary arteries. Each subbranch of the segmented coronary arteries was segmented correctly by the proposed method. The obtained results are compared with those ground truth values obtained from the commercial software from GE Healthcare and the level-set method proposed by Yang et al., 2007. Results indicate that the proposed method is better in terms of efficiency analyzed. Based on the initial segmentation of coronary arteries obtained from 3D region growing, one-level 3D DWT and 3D neutrosophic transformation can be applied to detect coronary pathologies accurately.

  5. Multidimensional, mapping-based complex wavelet transforms.

    PubMed

    Fernandes, Felix C A; van Spaendonck, Rutger L C; Burrus, C Sidney

    2005-01-01

    Although the discrete wavelet transform (DWT) is a powerful tool for signal and image processing, it has three serious disadvantages: shift sensitivity, poor directionality, and lack of phase information. To overcome these disadvantages, we introduce multidimensional, mapping-based, complex wavelet transforms that consist of a mapping onto a complex function space followed by a DWT of the complex mapping. Unlike other popular transforms that also mitigate DWT shortcomings, the decoupled implementation of our transforms has two important advantages. First, the controllable redundancy of the mapping stage offers a balance between degree of shift sensitivity and transform redundancy. This allows us to create a directional, nonredundant, complex wavelet transform with potential benefits for image coding systems. To the best of our knowledge, no other complex wavelet transform is simultaneously directional and nonredundant. The second advantage of our approach is the flexibility to use any DWT in the transform implementation. As an example, we exploit this flexibility to create the complex double-density DWT: a shift-insensitive, directional, complex wavelet transform with a low redundancy of (3M - 1)/(2M - 1) in M dimensions. No other transform achieves all these properties at a lower redundancy, to the best of our knowledge. By exploiting the advantages of our multidimensional, mapping-based complex wavelet transforms in seismic signal-processing applications, we have demonstrated state-of-the-art results.

  6. Transformation-cost time-series method for analyzing irregularly sampled data

    NASA Astrophysics Data System (ADS)

    Ozken, Ibrahim; Eroglu, Deniz; Stemler, Thomas; Marwan, Norbert; Bagci, G. Baris; Kurths, Jürgen

    2015-06-01

    Irregular sampling of data sets is one of the challenges often encountered in time-series analysis, since traditional methods cannot be applied and the frequently used interpolation approach can corrupt the data and bias the subsequence analysis. Here we present the TrAnsformation-Cost Time-Series (TACTS) method, which allows us to analyze irregularly sampled data sets without degenerating the quality of the data set. Instead of using interpolation we consider time-series segments and determine how close they are to each other by determining the cost needed to transform one segment into the following one. Using a limited set of operations—with associated costs—to transform the time series segments, we determine a new time series, that is our transformation-cost time series. This cost time series is regularly sampled and can be analyzed using standard methods. While our main interest is the analysis of paleoclimate data, we develop our method using numerical examples like the logistic map and the Rössler oscillator. The numerical data allows us to test the stability of our method against noise and for different irregular samplings. In addition we provide guidance on how to choose the associated costs based on the time series at hand. The usefulness of the TACTS method is demonstrated using speleothem data from the Secret Cave in Borneo that is a good proxy for paleoclimatic variability in the monsoon activity around the maritime continent.

  7. Transformation-cost time-series method for analyzing irregularly sampled data.

    PubMed

    Ozken, Ibrahim; Eroglu, Deniz; Stemler, Thomas; Marwan, Norbert; Bagci, G Baris; Kurths, Jürgen

    2015-06-01

    Irregular sampling of data sets is one of the challenges often encountered in time-series analysis, since traditional methods cannot be applied and the frequently used interpolation approach can corrupt the data and bias the subsequence analysis. Here we present the TrAnsformation-Cost Time-Series (TACTS) method, which allows us to analyze irregularly sampled data sets without degenerating the quality of the data set. Instead of using interpolation we consider time-series segments and determine how close they are to each other by determining the cost needed to transform one segment into the following one. Using a limited set of operations-with associated costs-to transform the time series segments, we determine a new time series, that is our transformation-cost time series. This cost time series is regularly sampled and can be analyzed using standard methods. While our main interest is the analysis of paleoclimate data, we develop our method using numerical examples like the logistic map and the Rössler oscillator. The numerical data allows us to test the stability of our method against noise and for different irregular samplings. In addition we provide guidance on how to choose the associated costs based on the time series at hand. The usefulness of the TACTS method is demonstrated using speleothem data from the Secret Cave in Borneo that is a good proxy for paleoclimatic variability in the monsoon activity around the maritime continent.

  8. Use of ultrasonic array method for positioning multiple partial discharge sources in transformer oil.

    PubMed

    Xie, Qing; Tao, Junhan; Wang, Yongqiang; Geng, Jianghai; Cheng, Shuyi; Lü, Fangcheng

    2014-08-01

    Fast and accurate positioning of partial discharge (PD) sources in transformer oil is very important for the safe, stable operation of power systems because it allows timely elimination of insulation faults. There is usually more than one PD source once an insulation fault occurs in the transformer oil. This study, which has both theoretical and practical significance, proposes a method of identifying multiple PD sources in the transformer oil. The method combines the two-sided correlation transformation algorithm in the broadband signal focusing and the modified Gerschgorin disk estimator. The method of classification of multiple signals is used to determine the directions of arrival of signals from multiple PD sources. The ultrasonic array positioning method is based on the multi-platform direction finding and the global optimization searching. Both the 4 × 4 square planar ultrasonic sensor array and the ultrasonic array detection platform are built to test the method of identifying and positioning multiple PD sources. The obtained results verify the validity and the engineering practicability of this method.

  9. Single-image super-resolution based on Markov random field and contourlet transform

    NASA Astrophysics Data System (ADS)

    Wu, Wei; Liu, Zheng; Gueaieb, Wail; He, Xiaohai

    2011-04-01

    Learning-based methods are well adopted in image super-resolution. In this paper, we propose a new learning-based approach using contourlet transform and Markov random field. The proposed algorithm employs contourlet transform rather than the conventional wavelet to represent image features and takes into account the correlation between adjacent pixels or image patches through the Markov random field (MRF) model. The input low-resolution (LR) image is decomposed with the contourlet transform and fed to the MRF model together with the contourlet transform coefficients from the low- and high-resolution image pairs in the training set. The unknown high-frequency components/coefficients for the input low-resolution image are inferred by a belief propagation algorithm. Finally, the inverse contourlet transform converts the LR input and the inferred high-frequency coefficients into the super-resolved image. The effectiveness of the proposed method is demonstrated with the experiments on facial, vehicle plate, and real scene images. A better visual quality is achieved in terms of peak signal to noise ratio and the image structural similarity measurement.

  10. Genomic Species Are Ecological Species as Revealed by Comparative Genomics in Agrobacterium tumefaciens

    PubMed Central

    Lassalle, Florent; Campillo, Tony; Vial, Ludovic; Baude, Jessica; Costechareyre, Denis; Chapulliot, David; Shams, Malek; Abrouk, Danis; Lavire, Céline; Oger-Desfeux, Christine; Hommais, Florence; Guéguen, Laurent; Daubin, Vincent; Muller, Daniel; Nesme, Xavier

    2011-01-01

    The definition of bacterial species is based on genomic similarities, giving rise to the operational concept of genomic species, but the reasons of the occurrence of differentiated genomic species remain largely unknown. We used the Agrobacterium tumefaciens species complex and particularly the genomic species presently called genomovar G8, which includes the sequenced strain C58, to test the hypothesis of genomic species having specific ecological adaptations possibly involved in the speciation process. We analyzed the gene repertoire specific to G8 to identify potential adaptive genes. By hybridizing 25 strains of A. tumefaciens on DNA microarrays spanning the C58 genome, we highlighted the presence and absence of genes homologous to C58 in the taxon. We found 196 genes specific to genomovar G8 that were mostly clustered into seven genomic islands on the C58 genome—one on the circular chromosome and six on the linear chromosome—suggesting higher plasticity and a major adaptive role of the latter. Clusters encoded putative functional units, four of which had been verified experimentally. The combination of G8-specific functions defines a hypothetical species primary niche for G8 related to commensal interaction with a host plant. This supports that the G8 ancestor was able to exploit a new ecological niche, maybe initiating ecological isolation and thus speciation. Searching genomic data for synapomorphic traits is a powerful way to describe bacterial species. This procedure allowed us to find such phenotypic traits specific to genomovar G8 and thus propose a Latin binomial, Agrobacterium fabrum, for this bona fide genomic species. PMID:21795751

  11. Transformation of Althaea officinalis L. by Agrobacterium rhizogenes for the production of transgenic roots expressing the anti-HIV microbicide cyanovirin-N.

    PubMed

    Drake, Pascal M W; de Moraes Madeira, Luisa; Szeto, Tim H; Ma, Julian K-C

    2013-12-01

    The marshmallow plant (Althaea officinalis L.) has been used for centuries in medicine and other applications. Valuable secondary metabolites have previously been identified in Agrobacterium rhizogenes-generated transgenic 'hairy' roots in this species. In the present study, transgenic roots were produced in A. officinalis using A. rhizogenes. In addition to wild-type lines, roots expressing the anti-human immunodeficiency virus microbicide candidate, cyanovirin-N (CV-N), were generated. Wild-type and CV-N root lines were transferred to liquid culture and increased in mass by 49 and 19 % respectively over a 7 day culture period. In the latter, the concentration of CV-N present in the root tissue was 2.4 μg/g fresh weight, with an average secretion rate into the growth medium of 0.02 μg/ml/24 h. A. officinalis transgenic roots may therefore in the future be used not only as a source of therapeutic secondary metabolites, but also as an expression system for the production of recombinant pharmaceuticals.

  12. Multigrid method based on the transformation-free HOC scheme on nonuniform grids for 2D convection diffusion problems

    NASA Astrophysics Data System (ADS)

    Ge, Yongbin; Cao, Fujun

    2011-05-01

    In this paper, a multigrid method based on the high order compact (HOC) difference scheme on nonuniform grids, which has been proposed by Kalita et al. [J.C. Kalita, A.K. Dass, D.C. Dalal, A transformation-free HOC scheme for steady convection-diffusion on non-uniform grids, Int. J. Numer. Methods Fluids 44 (2004) 33-53], is proposed to solve the two-dimensional (2D) convection diffusion equation. The HOC scheme is not involved in any grid transformation to map the nonuniform grids to uniform grids, consequently, the multigrid method is brand-new for solving the discrete system arising from the difference equation on nonuniform grids. The corresponding multigrid projection and interpolation operators are constructed by the area ratio. Some boundary layer and local singularity problems are used to demonstrate the superiority of the present method. Numerical results show that the multigrid method with the HOC scheme on nonuniform grids almost gets as equally efficient convergence rate as on uniform grids and the computed solution on nonuniform grids retains fourth order accuracy while on uniform grids just gets very poor solution for very steep boundary layer or high local singularity problems. The present method is also applied to solve the 2D incompressible Navier-Stokes equations using the stream function-vorticity formulation and the numerical solutions of the lid-driven cavity flow problem are obtained and compared with solutions available in the literature.

  13. On a Chirplet Transform Based Method for Co-channel Voice Separation

    NASA Astrophysics Data System (ADS)

    Dugnol, B.; Fernández, C.; Galiano, G.; Velasco, J.

    We use signal and image theory based algorithms to produce estimations of the number of wolves emitting howls or barks in a given field recording as an individuals counting alternative to the traditional trace collecting methodologies. We proceed in two steps. Firstly, we clean and enhance the signal by using PDE based image processing algorithms applied to the signal spectrogram. Secondly, assuming that the wolves chorus may be modelled as an addition of nonlinear chirps, we use the quadratic energy distribution corresponding to the Chirplet Transform of the signal to produce estimates of the corresponding instantaneous frequencies, chirp-rates and amplitudes at each instant of the recording. We finally establish suitable criteria to decide how such estimates are connected in time.

  14. Real-Time PCR-Based Quantitation Method for the Genetically Modified Soybean Line GTS 40-3-2.

    PubMed

    Kitta, Kazumi; Takabatake, Reona; Mano, Junichi

    2016-01-01

    This chapter describes a real-time PCR-based method for quantitation of the relative amount of genetically modified (GM) soybean line GTS 40-3-2 [Roundup Ready(®) soybean (RRS)] contained in a batch. The method targets a taxon-specific soybean gene (lectin gene, Le1) and the specific DNA construct junction region between the Petunia hybrida chloroplast transit peptide sequence and the Agrobacterium 5-enolpyruvylshikimate-3-phosphate synthase gene (epsps) sequence present in GTS 40-3-2. The method employs plasmid pMulSL2 as a reference material in order to quantify the relative amount of GTS 40-3-2 in soybean samples using a conversion factor (Cf) equal to the ratio of the RRS-specific DNA to the taxon-specific DNA in representative genuine GTS 40-3-2 seeds.

  15. Transgenic cotton: from biotransformation methods to agricultural application.

    PubMed

    Zhang, Baohong

    2013-01-01

    Transgenic cotton is among the first transgenic plants commercially adopted around the world. Since it was first introduced into the field in the middle of 1990s, transgenic cotton has been quickly adopted by cotton farmers in many developed and developing countries. Transgenic cotton has offered many important environmental, social, and economic benefits, including reduced usage of pesticides, indirect increase of yield, minimizing environmental pollution, and reducing labor and cost. Agrobacterium-mediated genetic transformation method is the major method for obtaining transgenic cotton. However, pollen tube pathway-mediated method is also used, particularly by scientists in China, to breed commercial transgenic cotton. Although transgenic cotton plants with disease-resistance, abiotic stress tolerance, and improved fiber quality have been developed in the past decades, insect-resistant and herbicide-tolerant cotton are the two dominant transgenic cottons in the transgenic cotton market.

  16. [Transformation of antimicrobial peptide fusion gene of cecropin B and rabbit NP-1 to Houttuynia cordata].

    PubMed

    Dong, Yan; Zhang, Ying; Yi, Lang; Lai, Huili; Zhang, Yaming; Zhou, Lian; Wang, Peixun

    2010-07-01

    To transform the antimicrobial peptide fusion gene of cecropin B and rabbit NP-1(CN) into Houttuynia cordata to improve its antimicrobic capability. The fusion gene of CN designed and synthesized artificially was recombined with expression vector pBI121. The recombined vector was transformed to Agrobacterium tumefaciens LBA4404, by which CN gene was transformed to the explants of H. cordata. The transgenic regeneration plantlets were selected by kanamycin and rapid screening PCR. The transgenic plants were identified by PCR-Southern of genomic DNA and RT-PCR. The disease resistances were detected by antibacterial zone trail of leaf extracts to E. coli K12 and infection by Rhizoctonia solani. Gene of interesting CN was inserted into genomic DNA and expressed in transformed H, cordata, whose resistance to E. coli K12 and Rh. solani was stronger than that of the non-transformed control. The fusion gene CN can improve antimicrobic capability of transformed H. cordata.

  17. Finger millet [Eleusine coracana (L.) Gaertn].

    PubMed

    Ceasar, Stanislaus Antony; Ignacimuthu, Savarimuthu

    2015-01-01

    Millets are the primary food source for millions of people in tropical regions of the world supplying mineral nutrition and protein. In this chapter, we describe an optimized protocol for the Agrobacterium-mediated transformation of finger millet variety GPU 45. Agrobacterium strain LBA4404 harboring plasmid pCAMBIA1301 which contains hygromycin phosphotransferase (hph) as selectable marker gene and β-glucuronidase (GUS) as reporter gene has been used. This protocol utilizes the shoot apex explants for the somatic embryogenesis and regeneration of finger millet after the transformation by Agrobacterium. Desiccation of explants during cocultivation helps for the better recovery of transgenic plants. This protocol is very useful for the efficient production of transgenic plants in finger millet through Agrobacterium-mediated transformation.

  18. FFT-enhanced IHS transform method for fusing high-resolution satellite images

    USGS Publications Warehouse

    Ling, Y.; Ehlers, M.; Usery, E.L.; Madden, M.

    2007-01-01

    Existing image fusion techniques such as the intensity-hue-saturation (IHS) transform and principal components analysis (PCA) methods may not be optimal for fusing the new generation commercial high-resolution satellite images such as Ikonos and QuickBird. One problem is color distortion in the fused image, which causes visual changes as well as spectral differences between the original and fused images. In this paper, a fast Fourier transform (FFT)-enhanced IHS method is developed for fusing new generation high-resolution satellite images. This method combines a standard IHS transform with FFT filtering of both the panchromatic image and the intensity component of the original multispectral image. Ikonos and QuickBird data are used to assess the FFT-enhanced IHS transform method. Experimental results indicate that the FFT-enhanced IHS transform method may improve upon the standard IHS transform and the PCA methods in preserving spectral and spatial information. ?? 2006 International Society for Photogrammetry and Remote Sensing, Inc. (ISPRS).

  19. Genomic analysis of Agrobacterium radiobacter DSM 30147T and emended description of A. radiobacter (Beijerinck and van Delden 1902) Conn 1942 (Approved Lists 1980) emend. Sawada et al. 1993

    PubMed Central

    Zhang, Linshuang; Li, Xiangyang; Zhang, Feng; Wang, Gejiao

    2014-01-01

    Agrobacterium radiobacter is the only known non-phytopathogenic species in Agrobacterium genus. In this study, the whole-genome sequence of A. radiobacter type strain DSM 30147T was described and compared to the other available Agrobacterium genomes. This bacterium has a genome size of 7,122,065 bp distributed in 612 contigs, including 6,834 protein-coding genes and 41 RNA genes. It harbors a circular chromosome and a linear chromosome but not a tumor-inducing (Ti) plasmid. To the best of our knowledge, this is the first report of a genome from the A. radiobacter species. In addition, an emended description of A. radiobacter is described. This study reveals information that enhances the current understanding of its non-phytopathogenicity and its phylogenetic position within Agrobacterium genus. PMID:25197445

  20. Transformative, Mixed Methods Checklist for Psychological Research with Mexican Americans

    ERIC Educational Resources Information Center

    Canales, Genevieve

    2013-01-01

    This is a description of the creation of a research methods tool, the "Transformative, Mixed Methods Checklist for Psychological Research With Mexican Americans." For conducting literature reviews of and planning mixed methods studies with Mexican Americans, it contains evaluative criteria calling for transformative mixed methods, perspectives…

  1. A novel method for improving the accuracy of coordinate transformation in multiple measurement systems

    NASA Astrophysics Data System (ADS)

    Liu, W. L.; Li, Y. W.

    2017-09-01

    Large-scale dimensional metrology usually requires a combination of multiple measurement systems, such as laser tracking, total station, laser scanning, coordinate measuring arm and video photogrammetry, etc. Often, the results from different measurement systems must be combined to provide useful results. The coordinate transformation is used to unify coordinate frames in combination; however, coordinate transformation uncertainties directly affect the accuracy of the final measurement results. In this paper, a novel method is proposed for improving the accuracy of coordinate transformation, combining the advantages of the best-fit least-square and radial basis function (RBF) neural networks. First of all, the configuration of coordinate transformation is introduced and a transformation matrix containing seven variables is obtained. Second, the 3D uncertainty of the transformation model and the residual error variable vector are established based on the best-fit least-square. Finally, in order to optimize the uncertainty of the developed seven-variable transformation model, we used the RBF neural network to identify the uncertainty of the dynamic, and unstructured, owing to its great ability to approximate any nonlinear function to the designed accuracy. Intensive experimental studies were conducted to check the validity of the theoretical results. The results show that the mean error of coordinate transformation decreased from 0.078 mm to 0.054 mm after using this method in contrast with the GUM method.

  2. Method to eliminate flux linkage DC component in load transformer for static transfer switch.

    PubMed

    He, Yu; Mao, Chengxiong; Lu, Jiming; Wang, Dan; Tian, Bing

    2014-01-01

    Many industrial and commercial sensitive loads are subject to the voltage sags and interruptions. The static transfer switch (STS) based on the thyristors is applied to improve the power quality and reliability. However, the transfer will result in severe inrush current in the load transformer, because of the DC component in the magnetic flux generated in the transfer process. The inrush current which is always 2 ~ 30 p.u. can cause the disoperation of relay protective devices and bring potential damage to the transformer. The way to eliminate the DC component is to transfer the related phases when the residual flux linkage of the load transformer and the prospective flux linkage of the alternate source are equal. This paper analyzes how the flux linkage of each winding in the load transformer changes in the transfer process. Based on the residual flux linkage when the preferred source is completely disconnected, the method to calculate the proper time point to close each phase of the alternate source is developed. Simulation and laboratory experiments results are presented to show the effectiveness of the transfer method.

  3. The t-CWT: a new ERP detection and quantification method based on the continuous wavelet transform and Student's t-statistics.

    PubMed

    Bostanov, Vladimir; Kotchoubey, Boris

    2006-12-01

    This study was aimed at developing a method for extraction and assessment of event-related brain potentials (ERP) from single-trials. This method should be applicable in the assessment of single persons' ERPs and should be able to handle both single ERP components and whole waveforms. We adopted a recently developed ERP feature extraction method, the t-CWT, for the purposes of hypothesis testing in the statistical assessment of ERPs. The t-CWT is based on the continuous wavelet transform (CWT) and Student's t-statistics. The method was tested in two ERP paradigms, oddball and semantic priming, by assessing individual-participant data on a single-trial basis, and testing the significance of selected ERP components, P300 and N400, as well as of whole ERP waveforms. The t-CWT was also compared to other univariate and multivariate ERP assessment methods: peak picking, area computation, discrete wavelet transform (DWT) and principal component analysis (PCA). The t-CWT produced better results than all of the other assessment methods it was compared with. The t-CWT can be used as a reliable and powerful method for ERP-component detection and testing of statistical hypotheses concerning both single ERP components and whole waveforms extracted from either single persons' or group data. The t-CWT is the first such method based explicitly on the criteria of maximal statistical difference between two average ERPs in the time-frequency domain and is particularly suitable for ERP assessment of individual data (e.g. in clinical settings), but also for the investigation of small and/or novel ERP effects from group data.

  4. Maize transformation technology development for commercial event generation.

    PubMed

    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.

  5. 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

  6. Weighted least squares phase unwrapping based on the wavelet transform

    NASA Astrophysics Data System (ADS)

    Chen, Jiafeng; Chen, Haiqin; Yang, Zhengang; Ren, Haixia

    2007-01-01

    The weighted least squares phase unwrapping algorithm is a robust and accurate method to solve phase unwrapping problem. This method usually leads to a large sparse linear equation system. Gauss-Seidel relaxation iterative method is usually used to solve this large linear equation. However, this method is not practical due to its extremely slow convergence. The multigrid method is an efficient algorithm to improve convergence rate. However, this method needs an additional weight restriction operator which is very complicated. For this reason, the multiresolution analysis method based on the wavelet transform is proposed. By applying the wavelet transform, the original system is decomposed into its coarse and fine resolution levels and an equivalent equation system with better convergence condition can be obtained. Fast convergence in separate coarse resolution levels speeds up the overall system convergence rate. The simulated experiment shows that the proposed method converges faster and provides better result than the multigrid method.

  7. Foundation: Transforming data bases into knowledge bases

    NASA Technical Reports Server (NTRS)

    Purves, R. B.; Carnes, James R.; Cutts, Dannie E.

    1987-01-01

    One approach to transforming information stored in relational data bases into knowledge based representations and back again is described. This system, called Foundation, allows knowledge bases to take advantage of vast amounts of pre-existing data. A benefit of this approach is inspection, and even population, of data bases through an intelligent knowledge-based front-end.

  8. Feedback linearization of singularly perturbed systems based on canonical similarity transformations

    NASA Astrophysics Data System (ADS)

    Kabanov, A. A.

    2018-05-01

    This paper discusses the problem of feedback linearization of a singularly perturbed system in a state-dependent coefficient form. The result is based on the introduction of a canonical similarity transformation. The transformation matrix is constructed from separate blocks for fast and slow part of an original singularly perturbed system. The transformed singular perturbed system has a linear canonical form that significantly simplifies a control design problem. Proposed similarity transformation allows accomplishing linearization of the system without considering the virtual output (as it is needed for normal form method), a technique of a transition from phase coordinates of the transformed system to state variables of the original system is simpler. The application of the proposed approach is illustrated through example.

  9. Evaluating Pillar Industry's Transformation Capability: A Case Study of Two Chinese Steel-Based Cities.

    PubMed

    Li, Zhidong; Marinova, Dora; Guo, Xiumei; Gao, Yuan

    2015-01-01

    Many steel-based cities in China were established between the 1950s and 1960s. After more than half a century of development and boom, these cities are starting to decline and industrial transformation is urgently needed. This paper focuses on evaluating the transformation capability of resource-based cities building an evaluation model. Using Text Mining and the Document Explorer technique as a way of extracting text features, the 200 most frequently used words are derived from 100 publications related to steel- and other resource-based cities. The Expert Evaluation Method (EEM) and Analytic Hierarchy Process (AHP) techniques are then applied to select 53 indicators, determine their weights and establish an index system for evaluating the transformation capability of the pillar industry of China's steel-based cities. Using real data and expert reviews, the improved Fuzzy Relation Matrix (FRM) method is applied to two case studies in China, namely Panzhihua and Daye, and the evaluation model is developed using Fuzzy Comprehensive Evaluation (FCE). The cities' abilities to carry out industrial transformation are evaluated with concerns expressed for the case of Daye. The findings have policy implications for the potential and required industrial transformation in the two selected cities and other resource-based towns.

  10. Adaptive synchrosqueezing based on a quilted short-time Fourier transform

    NASA Astrophysics Data System (ADS)

    Berrian, Alexander; Saito, Naoki

    2017-08-01

    In recent years, the synchrosqueezing transform (SST) has gained popularity as a method for the analysis of signals that can be broken down into multiple components determined by instantaneous amplitudes and phases. One such version of SST, based on the short-time Fourier transform (STFT), enables the sharpening of instantaneous frequency (IF) information derived from the STFT, as well as the separation of amplitude-phase components corresponding to distinct IF curves. However, this SST is limited by the time-frequency resolution of the underlying window function, and may not resolve signals exhibiting diverse time-frequency behaviors with sufficient accuracy. In this work, we develop a framework for an SST based on a "quilted" short-time Fourier transform (SST-QSTFT), which allows adaptation to signal behavior in separate time-frequency regions through the use of multiple windows. This motivates us to introduce a discrete reassignment frequency formula based on a finite difference of the phase spectrum, ensuring computational accuracy for a wider variety of windows. We develop a theoretical framework for the SST-QSTFT in both the continuous and the discrete settings, and describe an algorithm for the automatic selection of optimal windows depending on the region of interest. Using synthetic data, we demonstrate the superior numerical performance of SST-QSTFT relative to other SST methods in a noisy context. Finally, we apply SST-QSTFT to audio recordings of animal calls to demonstrate the potential of our method for the analysis of real bioacoustic signals.

  11. Highly efficient preparation of sphingoid bases from glucosylceramides by chemoenzymatic method[S

    PubMed Central

    Gowda, Siddabasave Gowda B.; Usuki, Seigo; Hammam, Mostafa A. S.; Murai, Yuta; Igarashi, Yasuyuki; Monde, Kenji

    2016-01-01

    Sphingoid base derivatives have attracted increasing attention as promising chemotherapeutic candidates against lifestyle diseases such as diabetes and cancer. Natural sphingoid bases can be a potential resource instead of those derived by time-consuming total organic synthesis. In particular, glucosylceramides (GlcCers) in food plants are enriched sources of sphingoid bases, differing from those of animals. Several chemical methodologies to transform GlcCers to sphingoid bases have already investigated; however, these conventional methods using acid or alkaline hydrolysis are not efficient due to poor reaction yield, producing complex by-products and resulting in separation problems. In this study, an extremely efficient and practical chemoenzymatic transformation method has been developed using microwave-enhanced butanolysis of GlcCers and a large amount of readily available almond β-glucosidase for its deglycosylation reaction of lysoGlcCers. The method is superior to conventional acid/base hydrolysis methods in its rapidity and its reaction cleanness (no isomerization, no rearrangement) with excellent overall yield. PMID:26667669

  12. PET and MRI image fusion based on combination of 2-D Hilbert transform and IHS method.

    PubMed

    Haddadpour, Mozhdeh; Daneshvar, Sabalan; Seyedarabi, Hadi

    2017-08-01

    The process of medical image fusion is combining two or more medical images such as Magnetic Resonance Image (MRI) and Positron Emission Tomography (PET) and mapping them to a single image as fused image. So purpose of our study is assisting physicians to diagnose and treat the diseases in the least of the time. We used Magnetic Resonance Image (MRI) and Positron Emission Tomography (PET) as input images, so fused them based on combination of two dimensional Hilbert transform (2-D HT) and Intensity Hue Saturation (IHS) method. Evaluation metrics that we apply are Discrepancy (D k ) as an assessing spectral features and Average Gradient (AG k ) as an evaluating spatial features and also Overall Performance (O.P) to verify properly of the proposed method. In this paper we used three common evaluation metrics like Average Gradient (AG k ) and the lowest Discrepancy (D k ) and Overall Performance (O.P) to evaluate the performance of our method. Simulated and numerical results represent the desired performance of proposed method. Since that the main purpose of medical image fusion is preserving both spatial and spectral features of input images, so based on numerical results of evaluation metrics such as Average Gradient (AG k ), Discrepancy (D k ) and Overall Performance (O.P) and also desired simulated results, it can be concluded that our proposed method can preserve both spatial and spectral features of input images. Copyright © 2017 Chang Gung University. Published by Elsevier B.V. All rights reserved.

  13. Transformation of apple (Malus × domestica) using mutants of apple acetolactate synthase as a selectable marker and analysis of the T-DNA integration sites.

    PubMed

    Yao, Jia-Long; Tomes, Sumathi; Gleave, Andrew P

    2013-05-01

    Apple acetolactate synthase mutants were generated by site-specific mutagenesis and successfully used as selection marker in tobacco and apple transformation. T-DNA/Apple genome junctions were analysed using genome-walking PCR and sequencing. An Agrobacterium-mediated genetic transformation system was developed for apple (Malus × domestica), using mutants of apple acetolactate synthase (ALS) as a selectable marker. Four apple ALS mutants were generated by site-specific mutagenesis and subsequently cloned under the transcriptional control of the CaMV 35S promoter and ocs 3' terminator, in a pART27-derived plant transformation vector. Three of the four mutations were found to confer resistance to the herbicide Glean(®), containing the active agent chlorsulfuron, in tobacco (Nicotiana tabacum) transformation. In apple transformation, leaf explants infected with Agrobacterium tumefaciens EHA105 containing one of the three ALS mutants resulted in the production of shoots on medium containing 2-8 μg L(-1) Glean(®), whilst uninfected wild-type explants failed to regenerate shoots or survive on medium containing 1 and 3 μg L(-1) Glean(®), respectively. Glean(®)-resistant, regenerated shoots were further multiplied and rooted on medium containing 10 μg L(-1) Glean(®). The T-DNA and apple genome-DNA junctions from eight rooted transgenic apple plants were analysed using genome-walking PCR amplification and sequencing. This analysis confirmed T-DNA integration into the apple genome, identified the genome integration sites and revealed the extent of any vector backbone integration, T-DNA rearrangements and deletions of apple genome DNA at the sites of integration.

  14. 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

  15. Improved medical image fusion based on cascaded PCA and shift invariant wavelet transforms.

    PubMed

    Reena Benjamin, J; Jayasree, T

    2018-02-01

    In the medical field, radiologists need more informative and high-quality medical images to diagnose diseases. Image fusion plays a vital role in the field of biomedical image analysis. It aims to integrate the complementary information from multimodal images, producing a new composite image which is expected to be more informative for visual perception than any of the individual input images. The main objective of this paper is to improve the information, to preserve the edges and to enhance the quality of the fused image using cascaded principal component analysis (PCA) and shift invariant wavelet transforms. A novel image fusion technique based on cascaded PCA and shift invariant wavelet transforms is proposed in this paper. PCA in spatial domain extracts relevant information from the large dataset based on eigenvalue decomposition, and the wavelet transform operating in the complex domain with shift invariant properties brings out more directional and phase details of the image. The significance of maximum fusion rule applied in dual-tree complex wavelet transform domain enhances the average information and morphological details. The input images of the human brain of two different modalities (MRI and CT) are collected from whole brain atlas data distributed by Harvard University. Both MRI and CT images are fused using cascaded PCA and shift invariant wavelet transform method. The proposed method is evaluated based on three main key factors, namely structure preservation, edge preservation, contrast preservation. The experimental results and comparison with other existing fusion methods show the superior performance of the proposed image fusion framework in terms of visual and quantitative evaluations. In this paper, a complex wavelet-based image fusion has been discussed. The experimental results demonstrate that the proposed method enhances the directional features as well as fine edge details. Also, it reduces the redundant details, artifacts, distortions.

  16. Transformations Based on Continuous Piecewise-Affine Velocity Fields

    PubMed Central

    Freifeld, Oren; Hauberg, Søren; Batmanghelich, Kayhan; Fisher, Jonn W.

    2018-01-01

    We propose novel finite-dimensional spaces of well-behaved ℝn → ℝn transformations. The latter are obtained by (fast and highly-accurate) integration of continuous piecewise-affine velocity fields. The proposed method is simple yet highly expressive, effortlessly handles optional constraints (e.g., volume preservation and/or boundary conditions), and supports convenient modeling choices such as smoothing priors and coarse-to-fine analysis. Importantly, the proposed approach, partly due to its rapid likelihood evaluations and partly due to its other properties, facilitates tractable inference over rich transformation spaces, including using Markov-Chain Monte-Carlo methods. Its applications include, but are not limited to: monotonic regression (more generally, optimization over monotonic functions); modeling cumulative distribution functions or histograms; time-warping; image warping; image registration; real-time diffeomorphic image editing; data augmentation for image classifiers. Our GPU-based code is publicly available. PMID:28092517

  17. Transformations based on continuous piecewise-affine velocity fields

    DOE PAGES

    Freifeld, Oren; Hauberg, Soren; Batmanghelich, Kayhan; ...

    2017-01-11

    Here, we propose novel finite-dimensional spaces of well-behaved Rn → Rn transformations. The latter are obtained by (fast and highly-accurate) integration of continuous piecewise-affine velocity fields. The proposed method is simple yet highly expressive, effortlessly handles optional constraints (e.g., volume preservation and/or boundary conditions), and supports convenient modeling choices such as smoothing priors and coarse-to-fine analysis. Importantly, the proposed approach, partly due to its rapid likelihood evaluations and partly due to its other properties, facilitates tractable inference over rich transformation spaces, including using Markov-Chain Monte-Carlo methods. Its applications include, but are not limited to: monotonic regression (more generally, optimization overmore » monotonic functions); modeling cumulative distribution functions or histograms; time-warping; image warping; image registration; real-time diffeomorphic image editing; data augmentation for image classifiers. Our GPU-based code is publicly available.« less

  18. The optimal digital filters of sine and cosine transforms for geophysical transient electromagnetic method

    NASA Astrophysics Data System (ADS)

    Zhao, Yun-wei; Zhu, Zi-qiang; Lu, Guang-yin; Han, Bo

    2018-03-01

    The sine and cosine transforms implemented with digital filters have been used in the Transient electromagnetic methods for a few decades. Kong (2007) proposed a method of obtaining filter coefficients, which are computed in the sample domain by Hankel transform pair. However, the curve shape of Hankel transform pair changes with a parameter, which usually is set to be 1 or 3 in the process of obtaining the digital filter coefficients of sine and cosine transforms. First, this study investigates the influence of the parameter on the digital filter algorithm of sine and cosine transforms based on the digital filter algorithm of Hankel transform and the relationship between the sine, cosine function and the ±1/2 order Bessel function of the first kind. The results show that the selection of the parameter highly influences the precision of digital filter algorithm. Second, upon the optimal selection of the parameter, it is found that an optimal sampling interval s also exists to achieve the best precision of digital filter algorithm. Finally, this study proposes four groups of sine and cosine transform digital filter coefficients with different length, which may help to develop the digital filter algorithm of sine and cosine transforms, and promote its application.

  19. Transformational and Transactional Leadership: Association With Attitudes Toward Evidence-Based Practice

    PubMed Central

    Aarons, Gregory A.

    2006-01-01

    Objective Leadership in organizations is important in shaping workers’ perceptions, responses to organizational change, and acceptance of innovations, such as evidence-based practices. Transformational leadership inspires and motivates followers, whereas transactional leadership is based more on reinforcement and exchanges. Studies have shown that in youth and family service organizations, mental health providers’ attitudes toward adopting an evidence-based practice are associated with organizational context and individual provider differences. The purpose of this study was to expand these findings by examining the association between leadership and mental health providers’ attitudes toward adopting evidence-based practice. Methods Participants were 303 public-sector mental health service clinicians and case managers from 49 programs who were providing mental health services to children, adolescents, and their families. Data were gathered on providers’ characteristics, attitudes toward evidence-based practices, and perceptions of their supervisors’ leadership behaviors. Zero-order correlations and multilevel regression analyses were conducted that controlled for effects of service providers’ characteristics. Results Both transformational and transactional leadership were positively associated with providers’ having more positive attitudes toward adoption of evidence-based practice, and transformational leadership was negatively associated with providers’ perception of difference between the providers’ current practice and evidence-based practice. Conclusions Mental health service organizations may benefit from improving transformational and transactional supervisory leadership skills in preparation for implementing evidence-based practices. PMID:16870968

  20. Agrobacterium tumefaciens mutants affected in attachment to plant cells.

    PubMed Central

    Douglas, C J; Halperin, W; Nester, E W

    1982-01-01

    An analysis of Agrobacterium tumefaciens mutants with Tn5 insertions in chromosomal DNA showed that the chromosome of A. tumefaciens codes for a specific ability of this bacterium to attach to plant cells. This ability is associated with tumorigenesis by A. tumefaciens, the ability of avirulent A. tumefaciens to inhibit tumorigenesis, and the ability to adsorb certain phages. A second class of chromosomal mutations affects tumorigenesis without altering the ability to attach to plant cells. The attachment of A. tumefaciens to plant cells was assayed by mixing radiolabeled bacteria with suspensions of tobacco tissue culture cells or freshly isolated Zinnia leaf mesophyll cells. Under the conditions of this assay, an avirulent Ti plasmid-cured strain attached to the same extent as the same strain containing pTiB6806. Six of eight avirulent mutants with Tn5 insertions in chromosomal DNA showed defective attachment, whereas two retained wild-type attachment ability. In contrast to the strains showing wild-type attachment, the attachment-defective mutants failed to inhibit tumorigenesis when inoculated onto Jerusalem artichoke slices before inoculation of a virulent strain and also showed a loss of sensitivity to two Agrobacterium phages. The loss of phage sensitivity appeared to be due to a loss of ability to adsorb the phages. Staining with Calcofluor indicated that the mutants retained the ability to synthesize cellulose fibrils, which have been implicated in the attachment process. Southern filter hybridizations demonstrated that each mutant contained a single Tn5 insertion, and genetic linkage between the Tn5 insertion in one mutant and the attachment phenotype has also been demonstrated. Images PMID:6292165

  1. High efficiency transformation of banana [Musa acuminata L. cv. Matti (AA)] for enhanced tolerance to salt and drought stress through overexpression of a peanut salinity-induced pathogenesis-related class 10 protein.

    PubMed

    Rustagi, Anjana; Jain, Shalu; Kumar, Deepak; Shekhar, Shashi; Jain, Mukesh; Bhat, Vishnu; Sarin, Neera Bhalla

    2015-01-01

    Bananas and plantains (Musa spp. L.) are important subsistence crops and premium export commodity in several countries, and susceptible to a wide range of environmental and biotic stress conditions. Here, we report efficient, rapid, and reproducible Agrobacterium-mediated transformation and regeneration of an Indian niche cultivar of banana [M. acuminata cv. Matti (AA)]. Apical meristem-derived highly proliferative multiple shoot clump (MSC) explants were transformed with the Agrobacterium strain EHA105 harboring a binary vector pCAMBIA-1301 carrying hptII and uidA. Sequential agro-infiltration (10 min, 400 mmHg), infection (additional 35 min, Agrobacterium density A 600 = 0.8) and co-cultivation (18 h) regimen in 100 µM acetosyringone containing liquid medium were critical factors yielding high transformation efficiency (~81 %) corroborated by transient GUS expression assay. Stable transgenic events were recovered following two cycles of meristem initiation and selection on hygromycin containing medium. Histochemical GUS assay in several tissues of transgenic plants and molecular analyses confirmed stable integration and expression of transgene. The protocol described here allowed recovery of well-established putative transgenic plantlets in as little as 5 months. The transgenic banana plants could be readily acclimatized under greenhouse conditions, and were phenotypically similar to the wild-type untransformed control plants (WT). Transgenic plants overexpressing Salinity-Induced Pathogenesis-Related class 10 protein gene from Arachis hypogaea (AhSIPR10) in banana cv. Matti (AA) showed better photosynthetic efficiency and less membrane damage (P < 0.05) in the presence of NaCl and mannitol in comparison to WT plants suggesting the role of AhSIPR10 in better tolerance of salt stress and drought conditions.

  2. A Power Transformers Fault Diagnosis Model Based on Three DGA Ratios and PSO Optimization SVM

    NASA Astrophysics Data System (ADS)

    Ma, Hongzhe; Zhang, Wei; Wu, Rongrong; Yang, Chunyan

    2018-03-01

    In order to make up for the shortcomings of existing transformer fault diagnosis methods in dissolved gas-in-oil analysis (DGA) feature selection and parameter optimization, a transformer fault diagnosis model based on the three DGA ratios and particle swarm optimization (PSO) optimize support vector machine (SVM) is proposed. Using transforming support vector machine to the nonlinear and multi-classification SVM, establishing the particle swarm optimization to optimize the SVM multi classification model, and conducting transformer fault diagnosis combined with the cross validation principle. The fault diagnosis results show that the average accuracy of test method is better than the standard support vector machine and genetic algorithm support vector machine, and the proposed method can effectively improve the accuracy of transformer fault diagnosis is proved.

  3. Method to Eliminate Flux Linkage DC Component in Load Transformer for Static Transfer Switch

    PubMed Central

    2014-01-01

    Many industrial and commercial sensitive loads are subject to the voltage sags and interruptions. The static transfer switch (STS) based on the thyristors is applied to improve the power quality and reliability. However, the transfer will result in severe inrush current in the load transformer, because of the DC component in the magnetic flux generated in the transfer process. The inrush current which is always 2~30 p.u. can cause the disoperation of relay protective devices and bring potential damage to the transformer. The way to eliminate the DC component is to transfer the related phases when the residual flux linkage of the load transformer and the prospective flux linkage of the alternate source are equal. This paper analyzes how the flux linkage of each winding in the load transformer changes in the transfer process. Based on the residual flux linkage when the preferred source is completely disconnected, the method to calculate the proper time point to close each phase of the alternate source is developed. Simulation and laboratory experiments results are presented to show the effectiveness of the transfer method. PMID:25133255

  4. Quorum Sensing and Quorum Quenching in Agrobacterium: A "Go/No Go System"?

    PubMed

    Dessaux, Yves; Faure, Denis

    2018-04-16

    The pathogen Agrobacterium induces gall formation on a wide range of dicotyledonous plants. In this bacteria, most pathogenicity determinants are borne on the tumour inducing (Ti) plasmid. The conjugative transfer of this plasmid between agrobacteria is regulated by quorum sensing (QS). However, processes involved in the disturbance of QS also occur in this bacteria under the molecular form of a protein, TraM, inhibiting the sensing of the QS signals, and two lactonases BlcC (AttM) and AiiB that degrade the acylhomoserine lactone (AHL) QS signal. In the model Agrobacterium fabrum strain C58, several data, once integrated, strongly suggest that the QS regulation may not be reacting only to cell concentration. Rather, these QS elements in association with the quorum quenching (QQ) activities may constitute an integrated and complex “go/no go system” that finely controls the biologically costly transfer of the Ti plasmid in response to multiple environmental cues. This decision mechanism permits the bacteria to sense whether it is in a gall or not, in a living or decaying tumor, in stressed plant tissues, etc. In this scheme, the role of the lactonases selected and maintained in the course of Ti plasmid and agrobacterial evolution appears to be pivotal.

  5. Two-Stream Transformer Networks for Video-based Face Alignment.

    PubMed

    Liu, Hao; Lu, Jiwen; Feng, Jianjiang; Zhou, Jie

    2017-08-01

    In this paper, we propose a two-stream transformer networks (TSTN) approach for video-based face alignment. Unlike conventional image-based face alignment approaches which cannot explicitly model the temporal dependency in videos and motivated by the fact that consistent movements of facial landmarks usually occur across consecutive frames, our TSTN aims to capture the complementary information of both the spatial appearance on still frames and the temporal consistency information across frames. To achieve this, we develop a two-stream architecture, which decomposes the video-based face alignment into spatial and temporal streams accordingly. Specifically, the spatial stream aims to transform the facial image to the landmark positions by preserving the holistic facial shape structure. Accordingly, the temporal stream encodes the video input as active appearance codes, where the temporal consistency information across frames is captured to help shape refinements. Experimental results on the benchmarking video-based face alignment datasets show very competitive performance of our method in comparisons to the state-of-the-arts.

  6. A Rigid Image Registration Based on the Nonsubsampled Contourlet Transform and Genetic Algorithms

    PubMed Central

    Meskine, Fatiha; Chikr El Mezouar, Miloud; Taleb, Nasreddine

    2010-01-01

    Image registration is a fundamental task used in image processing to match two or more images taken at different times, from different sensors or from different viewpoints. The objective is to find in a huge search space of geometric transformations, an acceptable accurate solution in a reasonable time to provide better registered images. Exhaustive search is computationally expensive and the computational cost increases exponentially with the number of transformation parameters and the size of the data set. In this work, we present an efficient image registration algorithm that uses genetic algorithms within a multi-resolution framework based on the Non-Subsampled Contourlet Transform (NSCT). An adaptable genetic algorithm for registration is adopted in order to minimize the search space. This approach is used within a hybrid scheme applying the two techniques fitness sharing and elitism. Two NSCT based methods are proposed for registration. A comparative study is established between these methods and a wavelet based one. Because the NSCT is a shift-invariant multidirectional transform, the second method is adopted for its search speeding up property. Simulation results clearly show that both proposed techniques are really promising methods for image registration compared to the wavelet approach, while the second technique has led to the best performance results of all. Moreover, to demonstrate the effectiveness of these methods, these registration techniques have been successfully applied to register SPOT, IKONOS and Synthetic Aperture Radar (SAR) images. The algorithm has been shown to work perfectly well for multi-temporal satellite images as well, even in the presence of noise. PMID:22163672

  7. A rigid image registration based on the nonsubsampled contourlet transform and genetic algorithms.

    PubMed

    Meskine, Fatiha; Chikr El Mezouar, Miloud; Taleb, Nasreddine

    2010-01-01

    Image registration is a fundamental task used in image processing to match two or more images taken at different times, from different sensors or from different viewpoints. The objective is to find in a huge search space of geometric transformations, an acceptable accurate solution in a reasonable time to provide better registered images. Exhaustive search is computationally expensive and the computational cost increases exponentially with the number of transformation parameters and the size of the data set. In this work, we present an efficient image registration algorithm that uses genetic algorithms within a multi-resolution framework based on the Non-Subsampled Contourlet Transform (NSCT). An adaptable genetic algorithm for registration is adopted in order to minimize the search space. This approach is used within a hybrid scheme applying the two techniques fitness sharing and elitism. Two NSCT based methods are proposed for registration. A comparative study is established between these methods and a wavelet based one. Because the NSCT is a shift-invariant multidirectional transform, the second method is adopted for its search speeding up property. Simulation results clearly show that both proposed techniques are really promising methods for image registration compared to the wavelet approach, while the second technique has led to the best performance results of all. Moreover, to demonstrate the effectiveness of these methods, these registration techniques have been successfully applied to register SPOT, IKONOS and Synthetic Aperture Radar (SAR) images. The algorithm has been shown to work perfectly well for multi-temporal satellite images as well, even in the presence of noise.

  8. Development of an Agrobacterium-based transformation system for Rhizoctonia solani

    USDA-ARS?s Scientific Manuscript database

    A 8.7 kb binary vector containing the 1.9 kb hygromycin B phosphortransferase (hyg) gene was constructed with promoter and terminator regions from the glyceraldehyde-3-phosphate- dehydrogenase (gpd) gene of Rhizoctonia solani anastomosis group 3 (AG-3) at the 5'- and 3'- gene termini of hyg. Promot...

  9. Linear Chromosome-generating System of Agrobacterium tumefaciens C58: Protelomerase Generates and Protects Hairpin Ends

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Huang, Wai Mun; DaGloria, Jeanne; Fox, Heather

    2012-09-05

    Agrobacterium tumefaciens C58, the pathogenic bacteria that causes crown gall disease in plants, harbors one circular and one linear chromosome and two circular plasmids. The telomeres of its unusual linear chromosome are covalently closed hairpins. The circular and linear chromosomes co-segregate and are stably maintained in the organism. We have determined the sequence of the two ends of the linear chromosome thus completing the previously published genome sequence of A. tumefaciens C58. We found that the telomeres carry nearly identical 25-bp sequences at the hairpin ends that are related by dyad symmetry. We further showed that its Atu2523 gene encodesmore » a protelomerase (resolvase) and that the purified enzyme can generate the linear chromosomal closed hairpin ends in a sequence-specific manner. Agrobacterium protelomerase, whose presence is apparently limited to biovar 1 strains, acts via a cleavage-and-religation mechanism by making a pair of transient staggered nicks invariably at 6-bp spacing as the reaction intermediate. The enzyme can be significantly shortened at both the N and C termini and still maintain its enzymatic activity. Although the full-length enzyme can uniquely bind to its product telomeres, the N-terminal truncations cannot. The target site can also be shortened from the native 50-bp inverted repeat to 26 bp; thus, the Agrobacterium hairpin-generating system represents the most compact activity of all hairpin linear chromosome- and plasmid-generating systems to date. The biochemical analyses of the protelomerase reactions further revealed that the tip of the hairpin telomere may be unusually polymorphically capable of accommodating any nucleotide.« less

  10. Evaluating Pillar Industry’s Transformation Capability: A Case Study of Two Chinese Steel-Based Cities

    PubMed Central

    Li, Zhidong; Marinova, Dora; Guo, Xiumei; Gao, Yuan

    2015-01-01

    Many steel-based cities in China were established between the 1950s and 1960s. After more than half a century of development and boom, these cities are starting to decline and industrial transformation is urgently needed. This paper focuses on evaluating the transformation capability of resource-based cities building an evaluation model. Using Text Mining and the Document Explorer technique as a way of extracting text features, the 200 most frequently used words are derived from 100 publications related to steel- and other resource-based cities. The Expert Evaluation Method (EEM) and Analytic Hierarchy Process (AHP) techniques are then applied to select 53 indicators, determine their weights and establish an index system for evaluating the transformation capability of the pillar industry of China’s steel-based cities. Using real data and expert reviews, the improved Fuzzy Relation Matrix (FRM) method is applied to two case studies in China, namely Panzhihua and Daye, and the evaluation model is developed using Fuzzy Comprehensive Evaluation (FCE). The cities’ abilities to carry out industrial transformation are evaluated with concerns expressed for the case of Daye. The findings have policy implications for the potential and required industrial transformation in the two selected cities and other resource-based towns. PMID:26422266

  11. A Unified Method of Finding Laplace Transforms, Fourier Transforms, and Fourier Series. [and] An Inversion Method for Laplace Transforms, Fourier Transforms, and Fourier Series. Integral Transforms and Series Expansions. Modules and Monographs in Undergraduate Mathematics and Its Applications Project. UMAP Units 324 and 325.

    ERIC Educational Resources Information Center

    Grimm, C. A.

    This document contains two units that examine integral transforms and series expansions. In the first module, the user is expected to learn how to use the unified method presented to obtain Laplace transforms, Fourier transforms, complex Fourier series, real Fourier series, and half-range sine series for given piecewise continuous functions. In…

  12. Determination of accuracy of winding deformation method using kNN based classifier used for 3 MVA transformer

    NASA Astrophysics Data System (ADS)

    Ahmed, Mustafa Wasir; Baishya, Manash Jyoti; Sharma, Sasanka Sekhor; Hazarika, Manash

    2018-04-01

    This paper presents a detecting system on power transformer in transformer winding, core and on load tap changer (OLTC). Accuracy of winding deformation is determined using kNN based classifier. Winding deformation in power transformer can be measured using sweep frequency response analysis (SFRA), which can enhance the diagnosis accuracy to a large degree. It is suggested that in the results minor deformation faults can be detected at frequency range of 1 mHz to 2 MHz. The values of RCL parameters are changed when faults occur and hence frequency response of the winding will change accordingly. The SFRA data of tested transformer is compared with reference trace. The difference between two graphs indicate faults in the transformer. The deformation between 1 mHz to 1kHz gives winding deformation, 1 kHz to 100 kHz gives core deformation and 100 kHz to 2 MHz gives OLTC deformation.

  13. Stable Transformation of Ferns Using Spores as Targets: Pteris vittata and Ceratopteris thalictroides1[W][OPEN

    PubMed Central

    Muthukumar, Balasubramaniam; Joyce, Blake L.; Elless, Mark P.; Stewart, C. Neal

    2013-01-01

    Ferns (Pteridophyta) are very important members of the plant kingdom that lag behind other taxa with regards to our understanding of their genetics, genomics, and molecular biology. We report here, to our knowledge, the first instance of stable transformation of fern with recovery of transgenic sporophytes. Spores of the arsenic hyperaccumulating fern Pteris vittata and tetraploid ‘C-fern Express’ (Ceratopteris thalictroides) were stably transformed by Agrobacterium tumefaciens with constructs containing the P. vittata actin promoter driving a GUSPlus reporter gene. Reporter gene expression assays were performed on multiple tissues and growth stages of gametophytes and sporophytes. Southern-blot analysis confirmed stable transgene integration in recovered sporophytes and also confirmed that no plasmid from A. tumefaciens was present in the sporophyte tissues. We recovered seven independent transformants of P. vittata and four independent C. thalictroides transgenics. Inheritance analyses using β-glucuronidase (GUS) histochemical staining revealed that the GUS transgene was stably expressed in second generation C. thalictroides sporophytic tissues. In an independent experiment, the gusA gene that was driven by the 2× Cauliflower mosaic virus 35S promoter was bombarded into P. vittata spores using biolistics, in which putatively stable transgenic gametophytes were recovered. Transformation procedures required no tissue culture or selectable marker genes. However, we did attempt to use hygromycin selection, which was ineffective for recovering transgenic ferns. This simple stable transformation method should help facilitate functional genomics studies in ferns. PMID:23933990

  14. Analysis of photonic Doppler velocimetry data based on the continuous wavelet transform

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Liu Shouxian; Wang Detian; Li Tao

    2011-02-15

    The short time Fourier transform (STFT) cannot resolve rapid velocity changes in most photonic Doppler velocimetry (PDV) data. A practical analysis method based on the continuous wavelet transform (CWT) was presented to overcome this difficulty. The adaptability of the wavelet family predicates that the continuous wavelet transform uses an adaptive time window to estimate the instantaneous frequency of signals. The local frequencies of signal are accurately determined by finding the ridge in the spectrogram of the CWT and then are converted to target velocity according to the Doppler effects. A performance comparison between the CWT and STFT is demonstrated bymore » a plate-impact experiment data. The results illustrate that the new method is automatic and adequate for analysis of PDV data.« less

  15. Full-field optical coherence tomography image restoration based on Hilbert transformation

    NASA Astrophysics Data System (ADS)

    Na, Jihoon; Choi, Woo June; Choi, Eun Seo; Ryu, Seon Young; Lee, Byeong Ha

    2007-02-01

    We propose the envelope detection method that is based on Hilbert transform for image restoration in full-filed optical coherence tomography (FF-OCT). The FF-OCT system presenting a high-axial resolution of 0.9 μm was implemented with a Kohler illuminator based on Linnik interferometer configuration. A 250 W customized quartz tungsten halogen lamp was used as a broadband light source and a CCD camera was used as a 2-dimentional detector array. The proposed image restoration method for FF-OCT requires only single phase-shifting. By using both the original and the phase-shifted images, we could remove the offset and the background signals from the interference fringe images. The desired coherent envelope image was obtained by applying Hilbert transform. With the proposed image restoration method, we demonstrate en-face imaging performance of the implemented FF-OCT system by presenting a tilted mirror surface, an integrated circuit chip, and a piece of onion epithelium.

  16. Electrocardiogram ST-Segment Morphology Delineation Method Using Orthogonal Transformations

    PubMed Central

    2016-01-01

    Differentiation between ischaemic and non-ischaemic transient ST segment events of long term ambulatory electrocardiograms is a persisting weakness in present ischaemia detection systems. Traditional ST segment level measuring is not a sufficiently precise technique due to the single point of measurement and severe noise which is often present. We developed a robust noise resistant orthogonal-transformation based delineation method, which allows tracing the shape of transient ST segment morphology changes from the entire ST segment in terms of diagnostic and morphologic feature-vector time series, and also allows further analysis. For these purposes, we developed a new Legendre Polynomials based Transformation (LPT) of ST segment. Its basis functions have similar shapes to typical transient changes of ST segment morphology categories during myocardial ischaemia (level, slope and scooping), thus providing direct insight into the types of time domain morphology changes through the LPT feature-vector space. We also generated new Karhunen and Lo ève Transformation (KLT) ST segment basis functions using a robust covariance matrix constructed from the ST segment pattern vectors derived from the Long Term ST Database (LTST DB). As for the delineation of significant transient ischaemic and non-ischaemic ST segment episodes, we present a study on the representation of transient ST segment morphology categories, and an evaluation study on the classification power of the KLT- and LPT-based feature vectors to classify between ischaemic and non-ischaemic ST segment episodes of the LTST DB. Classification accuracy using the KLT and LPT feature vectors was 90% and 82%, respectively, when using the k-Nearest Neighbors (k = 3) classifier and 10-fold cross-validation. New sets of feature-vector time series for both transformations were derived for the records of the LTST DB which is freely available on the PhysioNet website and were contributed to the LTST DB. The KLT and LPT

  17. Phase-based motion magnification video for monitoring of vital signals using the Hermite transform

    NASA Astrophysics Data System (ADS)

    Brieva, Jorge; Moya-Albor, Ernesto

    2017-11-01

    In this paper we present a new Eulerian phase-based motion magnification technique using the Hermite Transform (HT) decomposition that is inspired in the Human Vision System (HVS). We test our method in one sequence of the breathing of a newborn baby and on a video sequence that shows the heartbeat on the wrist. We detect and magnify the heart pulse applying our technique. Our motion magnification approach is compared to the Laplacian phase based approach by means of quantitative metrics (based on the RMS error and the Fourier transform) to measure the quality of both reconstruction and magnification. In addition a noise robustness analysis is performed for the two methods.

  18. A method to perform a fast fourier transform with primitive image transformations.

    PubMed

    Sheridan, Phil

    2007-05-01

    The Fourier transform is one of the most important transformations in image processing. A major component of this influence comes from the ability to implement it efficiently on a digital computer. This paper describes a new methodology to perform a fast Fourier transform (FFT). This methodology emerges from considerations of the natural physical constraints imposed by image capture devices (camera/eye). The novel aspects of the specific FFT method described include: 1) a bit-wise reversal re-grouping operation of the conventional FFT is replaced by the use of lossless image rotation and scaling and 2) the usual arithmetic operations of complex multiplication are replaced with integer addition. The significance of the FFT presented in this paper is introduced by extending a discrete and finite image algebra, named Spiral Honeycomb Image Algebra (SHIA), to a continuous version, named SHIAC.

  19. Efficient transformation and expression of gfp gene in Valsa mali var. mali.

    PubMed

    Chen, Liang; Sun, Gengwu; Wu, Shujing; Liu, Huixiang; Wang, Hongkai

    2015-01-01

    Valsa mali var. mali, the causal agent of valsa canker of apple, causes great loss of apple production in apple producing regions. The pathogenic mechanism of the pathogen has not been studied extensively, thus a suitable gene marker for pathogenic invasion analysis and a random insertion of T-DNA for mutants are desirable. In this paper, we reported the construction of a binary vector pKO1-HPH containing a positive selective gene hygromycin phosphotransferase (hph), a reporter gene gfp conferring green fluorescent protein, and an efficient protocol for V. mali var. mali transformation mediated by Agrobacterium tumefaciens. A transformation efficiency up to about 75 transformants per 10(5) conidia was achieved when co-cultivation of V. mali var. mali and A. tumefaciens for 48 h in A. tumefaciens inductive medium agar plates. The insertions of hph gene and gfp gene into V. mali var. mali genome verified by polymerase chain reaction and southern blot analysis showed that 10 randomly-selected transformants exhibited a single, unique hybridization pattern. This is the first report of A. tumefaciens-mediated transformation of V. mali var mali carrying a 'reporter' gfp gene that stably and efficiently expressed in the transformed V. mali var. mali species.

  20. Methods for performing fast discrete curvelet transforms of data

    DOEpatents

    Candes, Emmanuel; Donoho, David; Demanet, Laurent

    2010-11-23

    Fast digital implementations of the second generation curvelet transform for use in data processing are disclosed. One such digital transformation is based on unequally-spaced fast Fourier transforms (USFFT) while another is based on the wrapping of specially selected Fourier samples. Both digital transformations return a table of digital curvelet coefficients indexed by a scale parameter, an orientation parameter, and a spatial location parameter. Both implementations are fast in the sense that they run in about O(n.sup.2 log n) flops for n by n Cartesian arrays or about O(N log N) flops for Cartesian arrays of size N=n.sup.3; in addition, they are also invertible, with rapid inversion algorithms of about the same complexity.

  1. Hough transform method for track finding in center drift chamber

    NASA Astrophysics Data System (ADS)

    Azmi, K. A. Mohammad Kamal; Wan Abdullah, W. A. T.; Ibrahim, Zainol Abidin

    2016-01-01

    Hough transform is a global tracking method used which had been expected to be faster approach for tracking the circular pattern of electron moving in Center Drift Chamber (CDC), by transforming the point of hit into a circular curve. This paper present the implementation of hough transform method for the reconstruction of tracks in Center Drift Chamber (CDC) which have been generated by random number in C language programming. Result from implementation of this method shows higher peak of circle parameter value (xc,yc,rc) that indicate the similarity value of the parameter needed for circular track in CDC for charged particles in the region of CDC.

  2. Exploring Possibilities for Transforming Established Subscription-based Scientific Journals into Open Access Journals. Present Situation, Transformation Criteria, and Exemplary Implementation within Trans-O-MIM.

    PubMed

    Haux, Reinhold; Kuballa, Stefanie; Schulze, Mareike; Böhm, Claudia; Gefeller, Olaf; Haaf, Jan; Henning, Peter; Mielke, Corinna; Niggemann, Florian; Schürg, Andrea; Bergemann, Dieter

    2016-12-07

    Based on today's information and communication technologies the open access paradigm has become an important approach for adequately communicating new scientific knowledge. Summarizing the present situation for journal transformation. Presenting criteria for adequate transformation as well as a specific approach for it. Describing our exemplary implementation of such a journal transformation. Studying the respective literature as well as discussing this topic in various discussion groups and meetings (primarily of editors and publishers, but also of authors and readers), with long term experience as editors and /or publishers of scientific publications as prerequisite. There is a clear will, particularly of political and funding organizations, towards open access publishing. In spite of this, there is still a large amount of scientific knowledge, being communicated through subscription-based journals. For successfully transforming such journals into open access, sixteen criteria for a goal-oriented, stepwise, sustainable, and fair transformation are suggested. The Tandem Model as transformation approach is introduced. Our exemplary implementation is done in the Trans-O-MIM project. It is exploring strategies, models and evaluation metrics for journal transformation. As instance the journal Methods of Information in Medicine will apply the Tandem Model from 2017 onwards. Within Trans-O-MIM we will reach at least nine of the sixteen criteria for adequate transformation. It was positive to implement Trans-O-MIM as international research project. After first steps for transforming Methods have successfully been made, challenges will remain, among others, in identifying appropriate incentives for open access publishing in order to support its transformation.

  3. Modelling of Singapore's topographic transformation based on DEMs

    NASA Astrophysics Data System (ADS)

    Wang, Tao; Belle, Iris; Hassler, Uta

    2015-02-01

    Singapore's topography has been heavily transformed by industrialization and urbanization processes. To investigate topographic changes and evaluate soil mass flows, historical topographic maps of 1924 and 2012 were employed, and basic topographic features were vectorized. Digital elevation models (DEMs) for the two years were reconstructed based on vector features. Corresponding slope maps, a surface difference map and a scatter plot of elevation changes were generated and used to quantify and categorize the nature of the topographic transformation. The surface difference map is aggregated into five main categories of changes: (1) areas without significant height changes, (2) lowered-down areas where hill ranges were cut down, (3) raised-up areas where valleys and swamps were filled in, (4) reclaimed areas from the sea, and (5) new water-covered areas. Considering spatial proximity and configurations of different types of changes, topographic transformation can be differentiated as either creating inland flat areas or reclaiming new land from the sea. Typical topographic changes are discussed in the context of Singapore's urbanization processes. The two slope maps and elevation histograms show that generally, the topographic surface of Singapore has become flatter and lower since 1924. More than 89% of height changes have happened within a range of 20 m and 95% have been below 40 m. Because of differences in land surveying and map drawing methods, uncertainties and inaccuracies inherent in the 1924 topographic maps are discussed in detail. In this work, a modified version of a traditional scatter plot is used to present height transformation patterns intuitively. This method of deriving categorical maps of topographical changes from a surface difference map can be used in similar studies to qualitatively interpret transformation. Slope maps and histograms were also used jointly to reveal additional patterns of topographic change.

  4. Study on Transformation of Ginsenosides in Different Methods

    PubMed Central

    Zheng, Meng-meng; Xu, Fang-xue; Li, Yu-juan; Xi, Xiao-zhi; Cui, Xiao-wei

    2017-01-01

    Ginseng is a traditional Chinese medicine and has the extensive pharmacological activity. Ginsenosides are the major constituent in ginseng and have the unique biological activity and medicinal value. Ginsenosides have the good effects on antitumor, anti-inflammatory, antioxidative and inhibition of the cell apoptosis. Studies have showed that the major ginsenosides could be converted into rare ginsenosides, which played a significant role in exerting pharmacological activity. However, the contents of some rare ginsenosides are very little. So it is very important to find the effective way to translate the main ginsenosides to rare ginsenosides. In order to provide the theoretical foundation for the transformation of ginsenoside in vitro, in this paper, many methods of the transformation of ginsenoside were summarized, mainly including physical methods, chemical methods, and biotransformation methods. PMID:29387726

  5. Transforming Mean and Osculating Elements Using Numerical Methods

    NASA Technical Reports Server (NTRS)

    Ely, Todd A.

    2010-01-01

    Mean element propagation of perturbed two body orbits has as its mathematical basis averaging theory of nonlinear dynamical systems. Averaged mean elements define the long-term evolution characteristics of an orbit. Using averaging theory, a near identity transformation can be found that transforms the mean elements back to the osculating elements that contain short period terms in addition to the secular and long period mean elements. The ability to perform the conversion is necessary so that orbit design conducted in mean elements can be converted back into osculating results. In the present work, this near identity transformation is found using the Fast Fourier Transform. An efficient method is found that is capable of recovering the osculating elements to first order

  6. 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

    Gossypium barbadense 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

  7. Choline Uptake in Agrobacterium tumefaciens by the High-Affinity ChoXWV Transporter▿

    PubMed Central

    Aktas, Meriyem; Jost, Kathinka A.; Fritz, Christiane; Narberhaus, Franz

    2011-01-01

    Agrobacterium tumefaciens is a facultative phytopathogen that causes crown gall disease. For successful plant transformation A. tumefaciens requires the membrane lipid phosphatidylcholine (PC), which is produced via the methylation and the PC synthase (Pcs) pathways. The latter route is dependent on choline. Although choline uptake has been demonstrated in A. tumefaciens, the responsible transporter(s) remained elusive. In this study, we identified the first choline transport system in A. tumefaciens. The ABC-type choline transporter is encoded by the chromosomally located choXWV operon (ChoX, binding protein; ChoW, permease; and ChoV, ATPase). The Cho system is not critical for growth and PC synthesis. However, [14C]choline uptake is severely reduced in A. tumefaciens choX mutants. Recombinant ChoX is able to bind choline with high affinity (equilibrium dissociation constant [KD] of ≈2 μM). Since other quaternary amines are bound by ChoX with much lower affinities (acetylcholine, KD of ≈80 μM; betaine, KD of ≈470 μM), the ChoXWV system functions as a high-affinity transporter with a preference for choline. Two tryptophan residues (W40 and W87) located in the predicted ligand-binding pocket are essential for choline binding. The structural model of ChoX built on Sinorhizobium meliloti ChoX resembles the typical structure of substrate binding proteins with a so-called “Venus flytrap mechanism” of substrate binding. PMID:21803998

  8. Segmentation-based wavelet transform for still-image compression

    NASA Astrophysics Data System (ADS)

    Mozelle, Gerard; Seghier, Abdellatif; Preteux, Francoise J.

    1996-10-01

    In order to address simultaneously the two functionalities, content-based scalability required by MPEG-4, we introduce a segmentation-based wavelet transform (SBWT). SBWT takes into account both the mathematical properties of multiresolution analysis and the flexibility of region-based approaches for image compression. The associated methodology has two stages: 1) image segmentation into convex and polygonal regions; 2) 2D-wavelet transform of the signal corresponding to each region. In this paper, we have mathematically studied a method for constructing a multiresolution analysis (VjOmega)j (epsilon) N adapted to a polygonal region which provides an adaptive region-based filtering. The explicit construction of scaling functions, pre-wavelets and orthonormal wavelets bases defined on a polygon is carried out by using scaling functions is established by using the theory of Toeplitz operators. The corresponding expression can be interpreted as a location property which allow defining interior and boundary scaling functions. Concerning orthonormal wavelets and pre-wavelets, a similar expansion is obtained by taking advantage of the properties of the orthogonal projector P(V(j(Omega )) perpendicular from the space Vj(Omega ) + 1 onto the space (Vj(Omega )) perpendicular. Finally the mathematical results provide a simple and fast algorithm adapted to polygonal regions.

  9. Content Based Image Retrieval based on Wavelet Transform coefficients distribution

    PubMed Central

    Lamard, Mathieu; Cazuguel, Guy; Quellec, Gwénolé; Bekri, Lynda; Roux, Christian; Cochener, Béatrice

    2007-01-01

    In this paper we propose a content based image retrieval method for diagnosis aid in medical fields. We characterize images without extracting significant features by using distribution of coefficients obtained by building signatures from the distribution of wavelet transform. The research is carried out by computing signature distances between the query and database images. Several signatures are proposed; they use a model of wavelet coefficient distribution. To enhance results, a weighted distance between signatures is used and an adapted wavelet base is proposed. Retrieval efficiency is given for different databases including a diabetic retinopathy, a mammography and a face database. Results are promising: the retrieval efficiency is higher than 95% for some cases using an optimization process. PMID:18003013

  10. Chemometric Analysis of Multicomponent Biodegradable Plastics by Fourier Transform Infrared Spectrometry: The R-Matrix Method

    USDA-ARS?s Scientific Manuscript database

    A new chemometric method based on absorbance ratios from Fourier transform infrared spectra was devised to analyze multicomponent biodegradable plastics. The method uses the BeerLambert law to directly compute individual component concentrations and weight losses before and after biodegradation of c...

  11. Identification of material constants for piezoelectric transformers by three-dimensional, finite-element method and a design-sensitivity method.

    PubMed

    Joo, Hyun-Woo; Lee, Chang-Hwan; Rho, Jong-Seok; Jung, Hyun-Kyo

    2003-08-01

    In this paper, an inversion scheme for piezoelectric constants of piezoelectric transformers is proposed. The impedance of piezoelectric transducers is calculated using a three-dimensional finite element method. The validity of this is confirmed experimentally. The effects of material coefficients on piezoelectric transformers are investigated numerically. Six material coefficient variables for piezoelectric transformers were selected, and a design sensitivity method was adopted as an inversion scheme. The validity of the proposed method was confirmed by step-up ratio calculations. The proposed method is applied to the analysis of a sample piezoelectric transformer, and its resonance characteristics are obtained by numerically combined equivalent circuit method.

  12. Transformation of standardized clinical models based on OWL technologies: from CEM to OpenEHR archetypes

    PubMed Central

    Legaz-García, María del Carmen; Menárguez-Tortosa, Marcos; Fernández-Breis, Jesualdo Tomás; Chute, Christopher G; Tao, Cui

    2015-01-01

    Introduction The semantic interoperability of electronic healthcare records (EHRs) systems is a major challenge in the medical informatics area. International initiatives pursue the use of semantically interoperable clinical models, and ontologies have frequently been used in semantic interoperability efforts. The objective of this paper is to propose a generic, ontology-based, flexible approach for supporting the automatic transformation of clinical models, which is illustrated for the transformation of Clinical Element Models (CEMs) into openEHR archetypes. Methods Our transformation method exploits the fact that the information models of the most relevant EHR specifications are available in the Web Ontology Language (OWL). The transformation approach is based on defining mappings between those ontological structures. We propose a way in which CEM entities can be transformed into openEHR by using transformation templates and OWL as common representation formalism. The transformation architecture exploits the reasoning and inferencing capabilities of OWL technologies. Results We have devised a generic, flexible approach for the transformation of clinical models, implemented for the unidirectional transformation from CEM to openEHR, a series of reusable transformation templates, a proof-of-concept implementation, and a set of openEHR archetypes that validate the methodological approach. Conclusions We have been able to transform CEM into archetypes in an automatic, flexible, reusable transformation approach that could be extended to other clinical model specifications. We exploit the potential of OWL technologies for supporting the transformation process. We believe that our approach could be useful for international efforts in the area of semantic interoperability of EHR systems. PMID:25670753

  13. Multilayer modal actuator-based piezoelectric transformers.

    PubMed

    Huang, Yao-Tien; Wu, Wen-Jong; Wang, Yen-Chieh; Lee, Chih-Kung

    2007-02-01

    An innovative, multilayer piezoelectric transformer equipped with a full modal filtering input electrode is reported herein. This modal-shaped electrode, based on the orthogonal property of structural vibration modes, is characterized by full modal filtering to ensure that only the desired vibration mode is excited during operation. The newly developed piezoelectric transformer is comprised of three layers: a multilayered input layer, an insulation layer, and a single output layer. The electrode shape of the input layer is derived from its structural vibration modal shape, which takes advantage of the orthogonal property of the vibration modes to achieve a full modal filtering effect. The insulation layer possesses two functions: first, to couple the mechanical vibration energy between the input and output, and second, to provide electrical insulation between the two layers. To meet the two functions, a low temperature, co-fired ceramic (LTCC) was used to provide the high mechanical rigidity and high electrical insulation. It can be shown that this newly developed piezoelectric transformer has the advantage of possessing a more efficient energy transfer and a wider optimal working frequency range when compared to traditional piezoelectric transformers. A multilayer piezoelectric, transformer-based inverter applicable for use in LCD monitors or portable displays is presented as well.

  14. Hough transform method for track finding in center drift chamber

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Azmi, K. A. Mohammad Kamal, E-mail: khasmidatul@siswa.um.edu.my; Wan Abdullah, W. A. T., E-mail: wat@um.edu.my; Ibrahim, Zainol Abidin

    Hough transform is a global tracking method used which had been expected to be faster approach for tracking the circular pattern of electron moving in Center Drift Chamber (CDC), by transforming the point of hit into a circular curve. This paper present the implementation of hough transform method for the reconstruction of tracks in Center Drift Chamber (CDC) which have been generated by random number in C language programming. Result from implementation of this method shows higher peak of circle parameter value (xc,yc,rc) that indicate the similarity value of the parameter needed for circular track in CDC for charged particlesmore » in the region of CDC.« less

  15. Analysis of temperature rise for piezoelectric transformer using finite-element method.

    PubMed

    Joo, Hyun-Woo; Lee, Chang-Hwan; Rho, Jong-Seok; Jung, Hyun-Kyo

    2006-08-01

    Analysis of heat problem and temperature field of a piezoelectric transformer, operated at steady-state conditions, is described. The resonance frequency of the transformer is calculated from impedance and electrical gain analysis using a finite-element method. Mechanical displacement and electric potential of the transformer at the calculated resonance frequency are used to calculate the loss distribution of the transformer. Temperature distribution using discretized heat transfer equation is calculated from the obtained losses of the transformer. Properties of the piezoelectric material, dependent on the temperature field, are measured to recalculate the losses, temperature distribution, and new resonance characteristics of the transformer. Iterative method is adopted to recalculate the losses and resonance frequency due to the changes of the material constants from temperature increase. Computed temperature distributions and new resonance characteristics of the transformer at steady-state temperature are verified by comparison with experimental results.

  16. A host-specific biological control of grape crown gall by Agrobacterium vitis strain F2/5: its regulation and population dynamics.

    PubMed

    Kaewnum, Supaporn; Zheng, Desen; Reid, Cheryl L; Johnson, Kameka L; Gee, Jodi C; Burr, Thomas J

    2013-05-01

    Nontumorigenic Agrobacterium vitis strain F2/5 is able to prevent crown gall caused by tumorigenic A. vitis on grape but not on other plant species such as tobacco. Mutations in a quorum-sensing transcription factor, aviR, and in caseinolytic protease (clp) component genes clpA and clpP1 resulted in reduced or loss of biological control. All mutants were complemented; however, restoration of biological control by complemented clpA and clpP1 mutants was dependent on the copy number of vector that was used as well as timing of application of the complemented mutants to grape wounds in relation to inoculation with pathogen. Mutations in other quorum-sensing and clp genes and in a gene associated with polyketide synthesis did not affect biological control. It was determined that, although F2/5 inhibits transformation by tumorigenic A. vitis strains on grape, it does not affect growth of the pathogen in wounded grape tissue over time.

  17. Transformation of pecan and regeneration of transgenic plants.

    PubMed

    McGranahan, G H; Leslie, C A; Dandekar, A M; Uratsu, S L; Yates, I E

    1993-09-01

    A gene transfer system developed for walnut (Juglans regia L.) was successfully applied to pecan (Carya illinoensis [Wang] K. Koch). Repetitively embryogenic somatic embryos derived from open-pollinated seed of 'Elliott', 'Wichita', and 'Schley' were co-cultivated with Agrobacterium strain EHA 101/pCGN 7001, which contains marker genes for beta-glucuronidase activity and resistance to kanamycin. Several modifications of the standard walnut transformation techniques were tested, including a lower concentration of kanamycin and a modified induction medium, but these treatments had no measurable effect on efficiency of transformation. Nineteen of the 764 viable inoculated embryos produced transgenic subclones; 13 of these were from the line 'Elliott'6, 3 from 'Schley'5/3, and 3 from 'Wichita'9. Transgenic embryos of 'Wichita'9 germinated most readily and three subclones were successfully micropropagated. Three transgenic plants of one of these subclones were obtained by grafting the tissue cultured shoots to seedling pecan rootstock in the greenhouse. Gene insertion, initially detected by GUS activity, was confirmed by detection of integrated T-DNA sequences using Southern analysis.

  18. Boundary particle method for Laplace transformed time fractional diffusion equations

    NASA Astrophysics Data System (ADS)

    Fu, Zhuo-Jia; Chen, Wen; Yang, Hai-Tian

    2013-02-01

    This paper develops a novel boundary meshless approach, Laplace transformed boundary particle method (LTBPM), for numerical modeling of time fractional diffusion equations. It implements Laplace transform technique to obtain the corresponding time-independent inhomogeneous equation in Laplace space and then employs a truly boundary-only meshless boundary particle method (BPM) to solve this Laplace-transformed problem. Unlike the other boundary discretization methods, the BPM does not require any inner nodes, since the recursive composite multiple reciprocity technique (RC-MRM) is used to convert the inhomogeneous problem into the higher-order homogeneous problem. Finally, the Stehfest numerical inverse Laplace transform (NILT) is implemented to retrieve the numerical solutions of time fractional diffusion equations from the corresponding BPM solutions. In comparison with finite difference discretization, the LTBPM introduces Laplace transform and Stehfest NILT algorithm to deal with time fractional derivative term, which evades costly convolution integral calculation in time fractional derivation approximation and avoids the effect of time step on numerical accuracy and stability. Consequently, it can effectively simulate long time-history fractional diffusion systems. Error analysis and numerical experiments demonstrate that the present LTBPM is highly accurate and computationally efficient for 2D and 3D time fractional diffusion equations.

  19. 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.

  20. Gaussian Quadrature is an efficient method for the back-transformation in estimating the usual intake distribution when assessing dietary exposure.

    PubMed

    Dekkers, A L M; Slob, W

    2012-10-01

    In dietary exposure assessment, statistical methods exist for estimating the usual intake distribution from daily intake data. These methods transform the dietary intake data to normal observations, eliminate the within-person variance, and then back-transform the data to the original scale. We propose Gaussian Quadrature (GQ), a numerical integration method, as an efficient way of back-transformation. We compare GQ with six published methods. One method uses a log-transformation, while the other methods, including GQ, use a Box-Cox transformation. This study shows that, for various parameter choices, the methods with a Box-Cox transformation estimate the theoretical usual intake distributions quite well, although one method, a Taylor approximation, is less accurate. Two applications--on folate intake and fruit consumption--confirmed these results. In one extreme case, some methods, including GQ, could not be applied for low percentiles. We solved this problem by modifying GQ. One method is based on the assumption that the daily intakes are log-normally distributed. Even if this condition is not fulfilled, the log-transformation performs well as long as the within-individual variance is small compared to the mean. We conclude that the modified GQ is an efficient, fast and accurate method for estimating the usual intake distribution. Copyright © 2012 Elsevier Ltd. All rights reserved.

  1. The method of parallel-hierarchical transformation for rapid recognition of dynamic images using GPGPU technology

    NASA Astrophysics Data System (ADS)

    Timchenko, Leonid; Yarovyi, Andrii; Kokriatskaya, Nataliya; Nakonechna, Svitlana; Abramenko, Ludmila; Ławicki, Tomasz; Popiel, Piotr; Yesmakhanova, Laura

    2016-09-01

    The paper presents a method of parallel-hierarchical transformations for rapid recognition of dynamic images using GPU technology. Direct parallel-hierarchical transformations based on cluster CPU-and GPU-oriented hardware platform. Mathematic models of training of the parallel hierarchical (PH) network for the transformation are developed, as well as a training method of the PH network for recognition of dynamic images. This research is most topical for problems on organizing high-performance computations of super large arrays of information designed to implement multi-stage sensing and processing as well as compaction and recognition of data in the informational structures and computer devices. This method has such advantages as high performance through the use of recent advances in parallelization, possibility to work with images of ultra dimension, ease of scaling in case of changing the number of nodes in the cluster, auto scan of local network to detect compute nodes.

  2. A novel rail defect detection method based on undecimated lifting wavelet packet transform and Shannon entropy-improved adaptive line enhancer

    NASA Astrophysics Data System (ADS)

    Hao, Qiushi; Zhang, Xin; Wang, Yan; Shen, Yi; Makis, Viliam

    2018-07-01

    Acoustic emission (AE) technology is sensitive to subliminal rail defects, however strong wheel-rail contact rolling noise under high-speed condition has gravely impeded detecting of rail defects using traditional denoising methods. In this context, the paper develops an adaptive detection method for rail cracks, which combines multiresolution analysis with an improved adaptive line enhancer (ALE). To obtain elaborate multiresolution information of transient crack signals with low computational cost, lifting scheme-based undecimated wavelet packet transform is adopted. In order to feature the impulsive property of crack signals, a Shannon entropy-improved ALE is proposed as a signal enhancing approach, where Shannon entropy is introduced to improve the cost function. Then a rail defect detection plan based on the proposed method for high-speed condition is put forward. From theoretical analysis and experimental verification, it is demonstrated that the proposed method has superior performance in enhancing the rail defect AE signal and reducing the strong background noise, offering an effective multiresolution approach for rail defect detection under high-speed and strong-noise condition.

  3. Purification, crystallization and preliminary X-ray diffraction analysis of a novel keto-deoxy-d-galactarate (KDG) dehydratase from Agrobacterium tumefaciens

    PubMed Central

    Taberman, Helena; Andberg, Martina; Parkkinen, Tarja; Richard, Peter; Hakulinen, Nina; Koivula, Anu; Rouvinen, Juha

    2014-01-01

    d-Galacturonic acid is the main component of pectin. It could be used to produce affordable renewable fuels, chemicals and materials through biotechnical conversion. Keto-deoxy-d-galactarate (KDG) dehydratase is an enzyme in the oxidative pathway of d-galacturonic acid in Agrobacterium tumefaciens (At). It converts 3-deoxy-2-keto-l-threo-hexarate to α-ketoglutaric semialdehyde. At KDG dehydratase was crystallized by the hanging-drop vapour-diffusion method. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 169.1, b = 117.8, c = 74.3 Å, β = 112.4° and an asymmetric unit of four monomers. X-ray diffraction data were collected to 1.9 Å resolution using synchrotron radiation. The three-dimensional structure of At KDG dehydratase will provide valuable information on the function of the enzyme and will allow it to be engineered for biorefinery-based applications. PMID:24419616

  4. Short-term data forecasting based on wavelet transformation and chaos theory

    NASA Astrophysics Data System (ADS)

    Wang, Yi; Li, Cunbin; Zhang, Liang

    2017-09-01

    A sketch of wavelet transformation and its application was given. Concerning the characteristics of time sequence, Haar wavelet was used to do data reduction. After processing, the effect of “data nail” on forecasting was reduced. Chaos theory was also introduced, a new chaos time series forecasting flow based on wavelet transformation was proposed. The largest Lyapunov exponent was larger than zero from small data sets, it verified the data change behavior still met chaotic behavior. Based on this, chaos time series to forecast short-term change behavior could be used. At last, the example analysis of the price from a real electricity market showed that the forecasting method increased the precision of the forecasting more effectively and steadily.

  5. Functional Genomic Analysis of Cotton Genes with Agrobacterium-Mediated Virus-Induced Gene Silencing

    PubMed Central

    Gao, Xiquan; Shan, Libo

    2015-01-01

    Cotton (Gossypium spp.) is one of the most agronomically important crops worldwide for its unique textile fiber production and serving as food and feed stock. Molecular breeding and genetic engineering of useful genes into cotton have emerged as advanced approaches to improve cotton yield, fiber quality, and resistance to various stresses. However, the understanding of gene functions and regulations in cotton is largely hindered by the limited molecular and biochemical tools. Here, we describe the method of an Agrobacterium infiltration-based virus-induced gene silencing (VIGS) assay to transiently silence endogenous genes in cotton at 2-week-old seedling stage. The genes of interest could be readily silenced with a consistently high efficiency. To monitor gene silencing efficiency, we have cloned cotton GrCla1 from G. raimondii, a homolog gene of Arabidopsis Cloroplastos alterados 1 (AtCla1) involved in chloroplast development, and inserted into a tobacco rattle virus (TRV) binary vector pYL156. Silencing of GrCla1 results in albino phenotype on the newly emerging leaves, serving as a visual marker for silencing efficiency. To further explore the possibility of using VIGS assay to reveal the essential genes mediating disease resistance to Verticillium dahliae, a fungal pathogen causing severe Verticillium wilt in cotton, we developed a seedling infection assay to inoculate cotton seedlings when the genes of interest are silenced by VIGS. The method we describe here could be further explored for functional genomic analysis of cotton genes involved in development and various biotic and abiotic stresses. PMID:23386302

  6. Functional genomic analysis of cotton genes with agrobacterium-mediated virus-induced gene silencing.

    PubMed

    Gao, Xiquan; Shan, Libo

    2013-01-01

    Cotton (Gossypium spp.) is one of the most agronomically important crops worldwide for its unique textile fiber production and serving as food and feed stock. Molecular breeding and genetic engineering of useful genes into cotton have emerged as advanced approaches to improve cotton yield, fiber quality, and resistance to various stresses. However, the understanding of gene functions and regulations in cotton is largely hindered by the limited molecular and biochemical tools. Here, we describe the method of an Agrobacterium infiltration-based virus-induced gene silencing (VIGS) assay to transiently silence endogenous genes in cotton at 2-week-old seedling stage. The genes of interest could be readily silenced with a consistently high efficiency. To monitor gene silencing efficiency, we have cloned cotton GrCla1 from G. raimondii, a homolog gene of Arabidopsis Cloroplastos alterados 1 (AtCla1) involved in chloroplast development, and inserted into a tobacco rattle virus (TRV) binary vector pYL156. Silencing of GrCla1 results in albino phenotype on the newly emerging leaves, serving as a visual marker for silencing efficiency. To further explore the possibility of using VIGS assay to reveal the essential genes mediating disease resistance to Verticillium dahliae, a fungal pathogen causing severe Verticillium wilt in cotton, we developed a seedling infection assay to inoculate cotton seedlings when the genes of interest are silenced by VIGS. The method we describe here could be further explored for functional genomic analysis of cotton genes involved in development and various biotic and abiotic stresses.

  7. Transforming han: a correlational method for psychology and religion.

    PubMed

    Oh, Whachul

    2015-06-01

    Han is a destructive feeling in Korea. Although Korea accomplished significant exterior growth, Korean society is still experiencing the dark aspects of transforming han as evidenced by having the highest suicide rate in Asia. Some reasons for this may be the fragmentation between North and South Korea. If we can transform han then it can become constructive. I was challenged to think of possibilities for transforming han internally; this brings me to the correlational method through psychological and religious interpretation. This study is to challenge and encourage many han-ridden people in Korean society. Through the psychological and religious understanding of han, people suffering can positively transform their han. They can relate to han more subjectively, and this means the han-ridden psyche has an innate sacredness of potential to transform.

  8. The genome of cultivated sweet potato contains Agrobacterium T-DNAs with expressed genes: An example of a naturally transgenic food crop

    PubMed Central

    Kyndt, Tina; Quispe, Dora; Zhai, Hong; Jarret, Robert; Ghislain, Marc; Liu, Qingchang; Gheysen, Godelieve

    2015-01-01

    Agrobacterium rhizogenes and Agrobacterium tumefaciens are plant pathogenic bacteria capable of transferring DNA fragments [transfer DNA (T-DNA)] bearing functional genes into the host plant genome. This naturally occurring mechanism has been adapted by plant biotechnologists to develop genetically modified crops that today are grown on more than 10% of the world’s arable land, although their use can result in considerable controversy. While assembling small interfering RNAs, or siRNAs, of sweet potato plants for metagenomic analysis, sequences homologous to T-DNA sequences from Agrobacterium spp. were discovered. Simple and quantitative PCR, Southern blotting, genome walking, and bacterial artificial chromosome library screening and sequencing unambiguously demonstrated that two different T-DNA regions (IbT-DNA1 and IbT-DNA2) are present in the cultivated sweet potato (Ipomoea batatas [L.] Lam.) genome and that these foreign genes are expressed at detectable levels in different tissues of the sweet potato plant. IbT-DNA1 was found to contain four open reading frames (ORFs) homologous to the tryptophan-2-monooxygenase (iaaM), indole-3-acetamide hydrolase (iaaH), C-protein (C-prot), and agrocinopine synthase (Acs) genes of Agrobacterium spp. IbT-DNA1 was detected in all 291 cultigens examined, but not in close wild relatives. IbT-DNA2 contained at least five ORFs with significant homology to the ORF14, ORF17n, rooting locus (Rol)B/RolC, ORF13, and ORF18/ORF17n genes of A. rhizogenes. IbT-DNA2 was detected in 45 of 217 genotypes that included both cultivated and wild species. Our finding, that sweet potato is naturally transgenic while being a widely and traditionally consumed food crop, could affect the current consumer distrust of the safety of transgenic food crops. PMID:25902487

  9. Physical methods for genetic transformation of fungi and yeast

    NASA Astrophysics Data System (ADS)

    Rivera, Ana Leonor; Magaña-Ortíz, Denis; Gómez-Lim, Miguel; Fernández, Francisco; Loske, Achim M.

    2014-06-01

    The production of transgenic fungi is a routine process. Currently, it is possible to insert genes from other fungi, viruses, bacteria and even animals, albeit with low efficiency, into the genomes of a number of fungal species. Genetic transformation requires the penetration of the transgene through the fungal cell wall, a process that can be facilitated by biological or physical methods. Novel methodologies for the efficient introduction of specific genes and stronger promoters are needed to increase production levels. A possible solution to this problem is the recently discovered shock-wave-mediated transformation. The objective of this article is to review the state of the art of the physical methods used for genetic fungi transformation and to describe some of the basic physics and molecular biology behind them.

  10. Transformational Teaching: Theoretical Underpinnings, Basic Principles, and Core Methods

    PubMed Central

    Slavich, George M.; Zimbardo, Philip G.

    2012-01-01

    Approaches to classroom instruction have evolved considerably over the past 50 years. This progress has been spurred by the development of several learning principles and methods of instruction, including active learning, student-centered learning, collaborative learning, experiential learning, and problem-based learning. In the present paper, we suggest that these seemingly different strategies share important underlying characteristics and can be viewed as complimentary components of a broader approach to classroom instruction called transformational teaching. Transformational teaching involves creating dynamic relationships between teachers, students, and a shared body of knowledge to promote student learning and personal growth. From this perspective, instructors are intellectual coaches who create teams of students who collaborate with each other and with their teacher to master bodies of information. Teachers assume the traditional role of facilitating students’ acquisition of key course concepts, but do so while enhancing students’ personal development and attitudes toward learning. They accomplish these goals by establishing a shared vision for a course, providing modeling and mastery experiences, challenging and encouraging students, personalizing attention and feedback, creating experiential lessons that transcend the boundaries of the classroom, and promoting ample opportunities for preflection and reflection. We propose that these methods are synergistically related and, when used together, maximize students’ potential for intellectual and personal growth. PMID:23162369

  11. Mellin Transform-Based Correction Method for Linear Scale Inconsistency of Intrusion Events Identification in OFPS

    NASA Astrophysics Data System (ADS)

    Wang, Baocheng; Qu, Dandan; Tian, Qing; Pang, Liping

    2018-05-01

    For the problem that the linear scale of intrusion signals in the optical fiber pre-warning system (OFPS) is inconsistent, this paper presents a method to correct the scale. Firstly, the intrusion signals are intercepted, and an aggregate of the segments with equal length is obtained. Then, the Mellin transform (MT) is applied to convert them into the same scale. The spectral characteristics are obtained by the Fourier transform. Finally, we adopt back-propagation (BP) neural network to identify intrusion types, which takes the spectral characteristics as input. We carried out the field experiments and collected the optical fiber intrusion signals which contain the picking signal, shoveling signal, and running signal. The experimental results show that the proposed algorithm can effectively improve the recognition accuracy of the intrusion signals.

  12. Embryo-specific expression of a visual reporter gene as a selection system for citrus transformation

    PubMed Central

    Zambon, Flavia T.; Erpen, Lígia; Soriano, Leonardo; Grosser, Jude

    2018-01-01

    The embryo-specific Dc3 gene promoter driving the VvMybA1 anthocyanin regulatory gene was used to develop a visual selection system for the genetic transformation of citrus. Agrobacterium-mediated transformation of cell suspension cultures resulted in the production of purple transgenic somatic embryos that could be easily separated from the green non-transgenic embryos. The somatic embryos produced phenotypically normal plants devoid of any visual purple coloration. These results were also confirmed using protoplast transformation. There was minimal gene expression in unstressed one-year-old transgenic lines. Cold and drought stress did not have any effect on gene expression, while exogenous ABA and NaCl application resulted in a minor change in gene expression in several transgenic lines. When gas exchange was measured in intact leaves, the transgenic lines were similar to controls under the same environment. Our results provide conclusive evidence for the utilization of a plant-derived, embryo-specific visual reporter system for the genetic transformation of citrus. Such a system could aid in the development of an all-plant, consumer-friendly GM citrus tree. PMID:29293649

  13. 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.

  14. Mutational analysis of the transcriptional activator VirG of Agrobacterium tumefaciens.

    PubMed Central

    Scheeren-Groot, E P; Rodenburg, K W; den Dulk-Ras, A; Turk, S C; Hooykaas, P J

    1994-01-01

    To find VirG proteins with altered properties, the virG gene was mutagenized. Random chemical mutagenesis of single-stranded DNA containing the Agrobacterium tumefaciens virG gene led with high frequency to the inactivation of the gene. Sequence analysis showed that 29% of the mutants contained a virG gene with one single-base-pair substitution somewhere in the open reading frame. Thirty-nine different mutations that rendered the VirG protein inactive were mapped. Besides these inactive mutants, two mutants in which the vir genes were active even in the absence of acetosyringone were found on indicator plates. A VirG protein with an N54D substitution turned out to be able to induce a virB-lacZ reporter gene to a high level even in the absence of the inducer acetosyringone. A VirG protein with an I77V substitution exhibited almost no induction in the absence of acetosyringone but showed a maximum induction level already at low concentrations of acetosyringone. Images PMID:7961391

  15. Pepper, sweet (Capsicum annuum).

    PubMed

    Heidmann, Iris; Boutilier, Kim

    2015-01-01

    Capsicum (pepper) species are economically important crops that are recalcitrant to genetic transformation by Agrobacterium (Agrobacterium tumefaciens). A number of protocols for pepper transformation have been described but are not routinely applicable. The main bottleneck in pepper transformation is the low frequency of cells that are both susceptible for Agrobacterium infection and have the ability to regenerate. Here, we describe a protocol for the efficient regeneration of transgenic sweet pepper (C. annuum) through inducible activation of the BABY BOOM (BBM) AP2/ERF transcription factor. Using this approach, we can routinely achieve a transformation efficiency of at least 0.6 %. The main improvements in this protocol are the reproducibility in transforming different genotypes and the ability to produce fertile shoots. An added advantage of this protocol is that BBM activity can be induced subsequently in stable transgenic lines, providing a novel regeneration system for clonal propagation through somatic embryogenesis.

  16. 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. © 2014 German Botanical Society and The Royal Botanical Society of the Netherlands.

  17. Retinal identification based on an Improved Circular Gabor Filter and Scale Invariant Feature Transform.

    PubMed

    Meng, Xianjing; Yin, Yilong; Yang, Gongping; Xi, Xiaoming

    2013-07-18

    Retinal identification based on retinal vasculatures in the retina provides the most secure and accurate means of authentication among biometrics and has primarily been used in combination with access control systems at high security facilities. Recently, there has been much interest in retina identification. As digital retina images always suffer from deformations, the Scale Invariant Feature Transform (SIFT), which is known for its distinctiveness and invariance for scale and rotation, has been introduced to retinal based identification. However, some shortcomings like the difficulty of feature extraction and mismatching exist in SIFT-based identification. To solve these problems, a novel preprocessing method based on the Improved Circular Gabor Transform (ICGF) is proposed. After further processing by the iterated spatial anisotropic smooth method, the number of uninformative SIFT keypoints is decreased dramatically. Tested on the VARIA and eight simulated retina databases combining rotation and scaling, the developed method presents promising results and shows robustness to rotations and scale changes.

  18. Retinal Identification Based on an Improved Circular Gabor Filter and Scale Invariant Feature Transform

    PubMed Central

    Meng, Xianjing; Yin, Yilong; Yang, Gongping; Xi, Xiaoming

    2013-01-01

    Retinal identification based on retinal vasculatures in the retina provides the most secure and accurate means of authentication among biometrics and has primarily been used in combination with access control systems at high security facilities. Recently, there has been much interest in retina identification. As digital retina images always suffer from deformations, the Scale Invariant Feature Transform (SIFT), which is known for its distinctiveness and invariance for scale and rotation, has been introduced to retinal based identification. However, some shortcomings like the difficulty of feature extraction and mismatching exist in SIFT-based identification. To solve these problems, a novel preprocessing method based on the Improved Circular Gabor Transform (ICGF) is proposed. After further processing by the iterated spatial anisotropic smooth method, the number of uninformative SIFT keypoints is decreased dramatically. Tested on the VARIA and eight simulated retina databases combining rotation and scaling, the developed method presents promising results and shows robustness to rotations and scale changes. PMID:23873409

  19. Sweet Potato [Ipomoea batatas (L.) Lam].

    PubMed

    Song, Guo-qing; Yamaguchi, Ken-ichi

    2006-01-01

    Among the available transformation methods reported on sweet potato, Agrobacterium tumefaciens-mediated transformation is more successful and desirable. Stem explants have shown to be ideal for the transformation of sweet potato because of their ready availability as explants, the simple transformation process, and high-frequency-regeneration via somatic embryogenesis. Under the two-step kanamycin-hygromycin selection method and using the appropriate explants type (stem explants), the efficiency of transformation can be considerably improved in cv. Beniazuma. The high efficiency in the transformation of stem explants suggests that the transformation protocol described in this chapter warrants testing for routine stable transformation of diverse varieties of sweet potato.

  20. Historical account on gaining insights on the mechanism of crown gall tumorigenesis induced by Agrobacterium tumefaciens

    PubMed Central

    Kado, Clarence I.

    2014-01-01

    The plant tumor disease known as crown gall was not called by that name until more recent times. Galls on plants were described by Malpighi (1679) who believed that these extraordinary growth are spontaneously produced. Agrobacterium was first isolated from tumors in 1897 by Fridiano Cavara in Napoli, Italy. After this bacterium was recognized to be the cause of crown gall disease, questions were raised on the mechanism by which it caused tumors on a variety of plants. Numerous very detailed studies led to the identification of Agrobacterium tumefaciens as the causal bacterium that cleverly transferred a genetic principle to plant host cells and integrated it into their chromosomes. Such studies have led to a variety of sophisticated mechanisms used by this organism to aid in its survival against competing microorganisms. Knowledge gained from these fundamental discoveries has opened many avenues for researchers to examine their primary organisms of study for similar mechanisms of pathogenesis in both plants and animals. These discoveries also advanced the genetic engineering of domesticated plants for improved food and fiber. PMID:25147542

  1. Detection technology of polarization target based on curvelet transform in turbid liquid

    NASA Astrophysics Data System (ADS)

    Zhang, Su; Duan, Jin; Fu, Qiang; Zhan, Juntong; Ma, Wanzhuo

    2015-08-01

    To suppress the interference of the target detecting in the turbid medium, a kind of polarization detection technology based on Curvelet transform was applied. This method firstly adjusts the angles of polarizing film to get the intensity images of the situations at 0°,60° and 120°, then deduces the images of Stokes vectors, degree of polarization (DOP) and polarization angle (PA) according to the Mueller matrix. At last the DOP and intensity images can be decomposed by Curvelet transform to realize the fusion of the high and low coefficients respectively, after the processed coefficients are reconstructed, the target which is easier to detect can be achieved. To prove this method, many targets in turbid medium have been detected by polarization method and fused their DOP and intensity images with Curvelet transform algorithm. As an example screws in moderate and high concentration liquid are presented respectively, from which we can see the unpolarized targets are less obvious in higher concentration liquid. When the DOP and intensity images are fused by Curvelet transform, the targets are emerged clearly out of the turbid medium, and the values of the quality evaluation parameters in clarity, degree of contract and spatial frequency are prominently enhanced comparing with the unpolarized images, which can show the feasibility of this method.

  2. X-ray structure of imidazolonepropionase from Agrobacterium tumefaciens at 1.87 Å resolution

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tyagi, Rajiv; Kumaran, Desigan; Burley, Stephen K.

    2010-01-12

    Histidine degradation in Agrobacterium tumefaciens involves four enzymes, including histidase (EC 4.3.1.3), urocanase (EC 4.2.1.49), imidazolonepropionase (EC 3.5.2.7), and N-formylglutamate amidohydrolase (EC 3.5.3.8). The third enzyme of the pathway, imidazolone-propionase, a 45.6 kDa protein, catalyzes conversion of imidazolone-5-propanoate to N-forminio-L-glutamate. Initial studies of the role of imidazolonepropionase in histidine degradation were published in 1953. Subsequent publications have been limited to enzyme kinetics, crystallization, and a recently reported structure determination. The imidazolonepropionases are members of metallodepenent-hydrolases (or amidohydroase) superfamily, which includs ureases, adenosine deaminases, phosphotriesterases, dihydroorotases, allantoinases, hydantoinases, adenine and cytosine deaminases, imidazolonepropionases, aryldial-kylphosphatases, chlorohydrolases, and formylmethanofuran dehydroases. Proteins belonging tomore » this large group share a common three-dimensional structural motif (an eightfold {alpha}/{beta} or TIM barrel) with similar active sites. Most superfamily members also share a conserved metal binding site, involving four histidine residues and one aspartic acid. Imidazolonepropionase is one of the targets selected for X-ray crystallpgrahpic structure determination by the New York Structural GenomiX Research Consortium (NYSGXRC) Target ID: 9252b to correlate the structure function relationship of poorly studied by important enzyme. Here they report the crystal structure of imidazolonepropionase from Agrobacterium tumefaciens determined at 1.87 {angstrom} resolution.« less

  3. An actuator extension transformation for a motion simulator and an inverse transformation applying Newton-Raphson's method

    NASA Technical Reports Server (NTRS)

    Dieudonne, J. E.

    1972-01-01

    A set of equations which transform position and angular orientation of the centroid of the payload platform of a six-degree-of-freedom motion simulator into extensions of the simulator's actuators has been derived and is based on a geometrical representation of the system. An iterative scheme, Newton-Raphson's method, has been successfully used in a real time environment in the calculation of the position and angular orientation of the centroid of the payload platform when the magnitude of the actuator extensions is known. Sufficient accuracy is obtained by using only one Newton-Raphson iteration per integration step of the real time environment.

  4. Application of differential transformation method for solving dengue transmission mathematical model

    NASA Astrophysics Data System (ADS)

    Ndii, Meksianis Z.; Anggriani, Nursanti; Supriatna, Asep K.

    2018-03-01

    The differential transformation method (DTM) is a semi-analytical numerical technique which depends on Taylor series and has application in many areas including Biomathematics. The aim of this paper is to employ the differential transformation method (DTM) to solve system of non-linear differential equations for dengue transmission mathematical model. Analytical and numerical solutions are determined and the results are compared to that of Runge-Kutta method. We found a good agreement between DTM and Runge-Kutta method.

  5. A progressive data compression scheme based upon adaptive transform coding: Mixture block coding of natural images

    NASA Technical Reports Server (NTRS)

    Rost, Martin C.; Sayood, Khalid

    1991-01-01

    A method for efficiently coding natural images using a vector-quantized variable-blocksized transform source coder is presented. The method, mixture block coding (MBC), incorporates variable-rate coding by using a mixture of discrete cosine transform (DCT) source coders. Which coders are selected to code any given image region is made through a threshold driven distortion criterion. In this paper, MBC is used in two different applications. The base method is concerned with single-pass low-rate image data compression. The second is a natural extension of the base method which allows for low-rate progressive transmission (PT). Since the base method adapts easily to progressive coding, it offers the aesthetic advantage of progressive coding without incorporating extensive channel overhead. Image compression rates of approximately 0.5 bit/pel are demonstrated for both monochrome and color images.

  6. Direct Linear Transformation Method for Three-Dimensional Cinematography

    ERIC Educational Resources Information Center

    Shapiro, Robert

    1978-01-01

    The ability of Direct Linear Transformation Method for three-dimensional cinematography to locate points in space was shown to meet the accuracy requirements associated with research on human movement. (JD)

  7. Effect of pre-plant soil fumigants on Agrobacterium tumefaciens, pythiaceous species, and subsequent soil recolonization by A. tumefaciens

    USDA-ARS?s Scientific Manuscript database

    Paradox (Juglans hindsii x J. regia), the dominant rootstock used in the California walnut industry, is susceptible to crown gall, caused by Agrobacterium tumefaciens. In practice, soil fumigation has been a common preplant management strategy for crown gall, but even an industry standard, methyl b...

  8. OBS Data Denoising Based on Compressed Sensing Using Fast Discrete Curvelet Transform

    NASA Astrophysics Data System (ADS)

    Nan, F.; Xu, Y.

    2017-12-01

    OBS (Ocean Bottom Seismometer) data denoising is an important step of OBS data processing and inversion. It is necessary to get clearer seismic phases for further velocity structure analysis. Traditional methods for OBS data denoising include band-pass filter, Wiener filter and deconvolution etc. (Liu, 2015). Most of these filtering methods are based on Fourier Transform (FT). Recently, the multi-scale transform methods such as wavelet transform (WT) and Curvelet transform (CvT) are widely used for data denoising in various applications. The FT, WT and CvT could represent signal sparsely and separate noise in transform domain. They could be used in different cases. Compared with Curvelet transform, the FT has Gibbs phenomenon and it cannot handle points discontinuities well. WT is well localized and multi scale, but it has poor orientation selectivity and could not handle curves discontinuities well. CvT is a multiscale directional transform that could represent curves with only a small number of coefficients. It provide an optimal sparse representation of objects with singularities along smooth curves, which is suitable for seismic data processing. As we know, different seismic phases in OBS data are showed as discontinuous curves in time domain. Hence, we promote to analysis the OBS data via CvT and separate the noise in CvT domain. In this paper, our sparsity-promoting inversion approach is restrained by L1 condition and we solve this L1 problem by using modified iteration thresholding. Results show that the proposed method could suppress the noise well and give sparse results in Curvelet domain. Figure 1 compares the Curvelet denoising method with Wavelet method on the same iterations and threshold through synthetic example. a)Original data. b) Add-noise data. c) Denoised data using CvT. d) Denoised data using WT. The CvT can well eliminate the noise and has better result than WT. Further we applied the CvT denoise method for the OBS data processing. Figure 2a

  9. Sound Source Localization Using Non-Conformal Surface Sound Field Transformation Based on Spherical Harmonic Wave Decomposition

    PubMed Central

    Zhang, Lanyue; Ding, Dandan; Yang, Desen; Wang, Jia; Shi, Jie

    2017-01-01

    Spherical microphone arrays have been paid increasing attention for their ability to locate a sound source with arbitrary incident angle in three-dimensional space. Low-frequency sound sources are usually located by using spherical near-field acoustic holography. The reconstruction surface and holography surface are conformal surfaces in the conventional sound field transformation based on generalized Fourier transform. When the sound source is on the cylindrical surface, it is difficult to locate by using spherical surface conformal transform. The non-conformal sound field transformation by making a transfer matrix based on spherical harmonic wave decomposition is proposed in this paper, which can achieve the transformation of a spherical surface into a cylindrical surface by using spherical array data. The theoretical expressions of the proposed method are deduced, and the performance of the method is simulated. Moreover, the experiment of sound source localization by using a spherical array with randomly and uniformly distributed elements is carried out. Results show that the non-conformal surface sound field transformation from a spherical surface to a cylindrical surface is realized by using the proposed method. The localization deviation is around 0.01 m, and the resolution is around 0.3 m. The application of the spherical array is extended, and the localization ability of the spherical array is improved. PMID:28489065

  10. Environmental transformations and ecological effects of iron-based nanoparticles.

    PubMed

    Lei, Cheng; Sun, Yuqing; Tsang, Daniel C W; Lin, Daohui

    2018-01-01

    The increasing application of iron-based nanoparticles (NPs), especially high concentrations of zero-valent iron nanoparticles (nZVI), has raised concerns regarding their environmental behavior and potential ecological effects. In the environment, iron-based NPs undergo physical, chemical, and/or biological transformations as influenced by environmental factors such as pH, ions, dissolved oxygen, natural organic matter (NOM), and biotas. This review presents recent research advances on environmental transformations of iron-based NPs, and articulates their relationships with the observed toxicities. The type and extent of physical, chemical, and biological transformations, including aggregation, oxidation, and bio-reduction, depend on the properties of NPs and the receiving environment. Toxicities of iron-based NPs to bacteria, algae, fish, and plants are increasingly observed, which are evaluated with a particular focus on the underlying mechanisms. The toxicity of iron-based NPs is a function of their properties, tolerance of test organisms, and environmental conditions. Oxidative stress induced by reactive oxygen species is considered as the primary toxic mechanism of iron-based NPs. Factors influencing the toxicity of iron-based NPs are addressed and environmental transformations play a significant role, for example, surface oxidation or coating by NOM generally lowers the toxicity of nZVI. Research gaps and future directions are suggested with an aim to boost concerted research efforts on environmental transformations and toxicity of iron-based NPs, e.g., toxicity studies of transformed NPs in field, expansion of toxicity endpoints, and roles of laden contaminants and surface coating. This review will enhance our understanding of potential risks of iron-based NPs and proper uses of environmentally benign NPs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  11. Local CC2 response method based on the Laplace transform: analytic energy gradients for ground and excited states.

    PubMed

    Ledermüller, Katrin; Schütz, Martin

    2014-04-28

    A multistate local CC2 response method for the calculation of analytic energy gradients with respect to nuclear displacements is presented for ground and electronically excited states. The gradient enables the search for equilibrium geometries of extended molecular systems. Laplace transform is used to partition the eigenvalue problem in order to obtain an effective singles eigenvalue problem and adaptive, state-specific local approximations. This leads to an approximation in the energy Lagrangian, which however is shown (by comparison with the corresponding gradient method without Laplace transform) to be of no concern for geometry optimizations. The accuracy of the local approximation is tested and the efficiency of the new code is demonstrated by application calculations devoted to a photocatalytic decarboxylation process of present interest.

  12. Laplace Transform Based Radiative Transfer Studies

    NASA Astrophysics Data System (ADS)

    Hu, Y.; Lin, B.; Ng, T.; Yang, P.; Wiscombe, W.; Herath, J.; Duffy, D.

    2006-12-01

    Multiple scattering is the major uncertainty for data analysis of space-based lidar measurements. Until now, accurate quantitative lidar data analysis has been limited to very thin objects that are dominated by single scattering, where photons from the laser beam only scatter a single time with particles in the atmosphere before reaching the receiver, and simple linear relationship between physical property and lidar signal exists. In reality, multiple scattering is always a factor in space-based lidar measurement and it dominates space- based lidar returns from clouds, dust aerosols, vegetation canopy and phytoplankton. While multiple scattering are clear signals, the lack of a fast-enough lidar multiple scattering computation tool forces us to treat the signal as unwanted "noise" and use simple multiple scattering correction scheme to remove them. Such multiple scattering treatments waste the multiple scattering signals and may cause orders of magnitude errors in retrieved physical properties. Thus the lack of fast and accurate time-dependent radiative transfer tools significantly limits lidar remote sensing capabilities. Analyzing lidar multiple scattering signals requires fast and accurate time-dependent radiative transfer computations. Currently, multiple scattering is done with Monte Carlo simulations. Monte Carlo simulations take minutes to hours and are too slow for interactive satellite data analysis processes and can only be used to help system / algorithm design and error assessment. We present an innovative physics approach to solve the time-dependent radiative transfer problem. The technique utilizes FPGA based reconfigurable computing hardware. The approach is as following, 1. Physics solution: Perform Laplace transform on the time and spatial dimensions and Fourier transform on the viewing azimuth dimension, and convert the radiative transfer differential equation solving into a fast matrix inversion problem. The majority of the radiative transfer

  13. Chaotic Signal Denoising Based on Hierarchical Threshold Synchrosqueezed Wavelet Transform

    NASA Astrophysics Data System (ADS)

    Wang, Wen-Bo; Jing, Yun-yu; Zhao, Yan-chao; Zhang, Lian-Hua; Wang, Xiang-Li

    2017-12-01

    In order to overcoming the shortcoming of single threshold synchrosqueezed wavelet transform(SWT) denoising method, an adaptive hierarchical threshold SWT chaotic signal denoising method is proposed. Firstly, a new SWT threshold function is constructed based on Stein unbiased risk estimation, which is two order continuous derivable. Then, by using of the new threshold function, a threshold process based on the minimum mean square error was implemented, and the optimal estimation value of each layer threshold in SWT chaotic denoising is obtained. The experimental results of the simulating chaotic signal and measured sunspot signals show that, the proposed method can filter the noise of chaotic signal well, and the intrinsic chaotic characteristic of the original signal can be recovered very well. Compared with the EEMD denoising method and the single threshold SWT denoising method, the proposed method can obtain better denoising result for the chaotic signal.

  14. A T-DNA gene required for agropine biosynthesis by transformed plants is functionally and evolutionarily related to a Ti plasmid gene required for catabolism of agropine by Agrobacterium strains.

    PubMed Central

    Hong, S B; Hwang, I; Dessaux, Y; Guyon, P; Kim, K S; Farrand, S K

    1997-01-01

    The mechanisms that ensure that Ti plasmid T-DNA genes encoding proteins involved in the biosynthesis of opines in crown gall tumors are always matched by Ti plasmid genes conferring the ability to catabolize that set of opines on the inducing Agrobacterium strains are unknown. The pathway for the biosynthesis of the opine agropine is thought to require an enzyme, mannopine cyclase, coded for by the ags gene located in the T(R) region of octopine-type Ti plasmids. Extracts prepared from agropine-type tumors contained an activity that cyclized mannopine to agropine. Tumor cells containing a T region in which ags was mutated lacked this activity and did not contain agropine. Expression of ags from the lac promoter conferred mannopine-lactonizing activity on Escherichia coli. Agrobacterium tumefaciens strains harboring an octopine-type Ti plasmid exhibit a similar activity which is not coded for by ags. Analysis of the DNA sequence of the gene encoding this activity, called agcA, showed it to be about 60% identical to T-DNA ags genes. Relatedness decreased abruptly in the 5' and 3' untranslated regions of the genes. ags is preceded by a promoter that functions only in the plant. Expression analysis showed that agcA also is preceded by its own promoter, which is active in the bacterium. Translation of agcA yielded a protein of about 45 kDa, consistent with the size predicted from the DNA sequence. Antibodies raised against the agcA product cross-reacted with the anabolic enzyme. These results indicate that the agropine system arose by a duplication of a progenitor gene, one copy of which became associated with the T-DNA and the other copy of which remained associated with the bacterium. PMID:9244272

  15. WE-AB-202-06: Correlating Lung CT HU with Transformation-Based and Xe-CT Derived Ventilation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Du, K; Patton, T; Bayouth, J

    Purpose: Regional lung ventilation is useful to reduce radiation-induced function damage during lung cancer radiation therapy. Recently a new direct HU (Hounsfield unit)-based method was proposed to estimate the ventilation potential without image registration. The purpose of this study is to examine if there is a functional dependence between HU values and transformation-based or Xe-CT derived ventilation. Methods: 4DCT images acquired from 13 patients prior to radiation therapy and 4 mechanically ventilated sheep subjects which also have associated Xe-CT images were used for this analysis. Transformation-based ventilation was computed using Jacobian determinant of the transformation field between peak-exhale and peak-inhalemore » 4DCT images. Both transformation and Xe-CT derived ventilation was computed for each HU bin. Color scatter plot and cumulative histogram were used to compare and validate the direct HU-based method. Results: There was little change of the center and shape of the HU histograms between free breathing CT and 4DCT average, with or without smoothing, and between the repeated 4DCT scans. HU of −750 and −630 were found to have the greatest transformation-based ventilation for human and sheep subjects, respectively. Maximum Xe-CT derived ventilation was found to locate at HU of −600 in sheep subjects. The curve between Xe-CT ventilation and HU was noisy for tissue above HU −400, possibly due to less intensity change of Xe gas during wash-out and wash-in phases. Conclusion: Both transformation-based and Xe-CT ventilation demonstrated that lung tissues with HU values in the range of (-750, −600) HU have the maximum ventilation potential. The correlation between HU and ventilation suggests that HU might be used to help guide the ventilation calculation and make it more robust to noise and image registration errors. Research support from NIH grants CA166703 and CA166119 and a gift from Roger Koch.« less

  16. Evaluations and modifications of semi-selective media for improved isolation of Agrobacterium tumefaciens biovar 1 from cultivated walnut

    USDA-ARS?s Scientific Manuscript database

    Agrobacterium tumefaciens, the causal agent of crown gall of walnut, is an aerobic, Gram negative bacterium belonging to the family Rhizobiaceae. Like many in this group, A. tumefaciens is a common inhabitant of soil and plant host tissue. Isolation from these complex environments is difficult even ...

  17. Efficient transformation of oil palm protoplasts by PEG-mediated transfection and DNA microinjection.

    PubMed

    Masani, Mat Yunus Abdul; Noll, Gundula A; Parveez, Ghulam Kadir Ahmad; Sambanthamurthi, Ravigadevi; Prüfer, Dirk

    2014-01-01

    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. 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. 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.

  18. 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

  19. MEMS based digital transform spectrometers

    NASA Astrophysics Data System (ADS)

    Geller, Yariv; Ramani, Mouli

    2005-09-01

    Earlier this year, a new breed of Spectrometers based on Micro-Electro-Mechanical-System (MEMS) engines has been introduced to the commercial market. The use of these engines combined with transform mathematics, produces powerful spectrometers at unprecedented low cost in various spectral regions.

  20. Fourier-transform and global contrast interferometer alignment methods

    DOEpatents

    Goldberg, Kenneth A.

    2001-01-01

    Interferometric methods are presented to facilitate alignment of image-plane components within an interferometer and for the magnified viewing of interferometer masks in situ. Fourier-transforms are performed on intensity patterns that are detected with the interferometer and are used to calculate pseudo-images of the electric field in the image plane of the test optic where the critical alignment of various components is being performed. Fine alignment is aided by the introduction and optimization of a global contrast parameter that is easily calculated from the Fourier-transform.

  1. Non-contact radio frequency shielding and wave guiding by multi-folded transformation optics method

    PubMed Central

    Madni, Hamza Ahmad; Zheng, Bin; Yang, Yihao; Wang, Huaping; Zhang, Xianmin; Yin, Wenyan; Li, Erping; Chen, Hongsheng

    2016-01-01

    Compared with conventional radio frequency (RF) shielding methods in which the conductive coating material encloses the circuits design and the leakage problem occurs due to the gap in such conductive material, non-contact RF shielding at a distance is very promising but still impossible to achieve so far. In this paper, a multi-folded transformation optics method is proposed to design a non-contact device for RF shielding. This “open-shielded” device can shield any object at a distance from the electromagnetic waves at the operating frequency, while the object is still physically open to the outer space. Based on this, an open-carpet cloak is proposed and the functionality of the open-carpet cloak is demonstrated. Furthermore, we investigate a scheme of non-contact wave guiding to remotely control the propagation of surface waves over any obstacles. The flexibilities of such multi-folded transformation optics method demonstrate the powerfulness of the method in the design of novel remote devices with impressive new functionalities. PMID:27841358

  2. A Method for Extracting Suspected Parotid Lesions in CT Images using Feature-based Segmentation and Active Contours based on Stationary Wavelet Transform

    NASA Astrophysics Data System (ADS)

    Wu, T. Y.; Lin, S. F.

    2013-10-01

    Automatic suspected lesion extraction is an important application in computer-aided diagnosis (CAD). In this paper, we propose a method to automatically extract the suspected parotid regions for clinical evaluation in head and neck CT images. The suspected lesion tissues in low contrast tissue regions can be localized with feature-based segmentation (FBS) based on local texture features, and can be delineated with accuracy by modified active contour models (ACM). At first, stationary wavelet transform (SWT) is introduced. The derived wavelet coefficients are applied to derive the local features for FBS, and to generate enhanced energy maps for ACM computation. Geometric shape features (GSFs) are proposed to analyze each soft tissue region segmented by FBS; the regions with higher similarity GSFs with the lesions are extracted and the information is also applied as the initial conditions for fine delineation computation. Consequently, the suspected lesions can be automatically localized and accurately delineated for aiding clinical diagnosis. The performance of the proposed method is evaluated by comparing with the results outlined by clinical experts. The experiments on 20 pathological CT data sets show that the true-positive (TP) rate on recognizing parotid lesions is about 94%, and the dimension accuracy of delineation results can also approach over 93%.

  3. Novel image encryption algorithm based on multiple-parameter discrete fractional random transform

    NASA Astrophysics Data System (ADS)

    Zhou, Nanrun; Dong, Taiji; Wu, Jianhua

    2010-08-01

    A new method of digital image encryption is presented by utilizing a new multiple-parameter discrete fractional random transform. Image encryption and decryption are performed based on the index additivity and multiple parameters of the multiple-parameter fractional random transform. The plaintext and ciphertext are respectively in the spatial domain and in the fractional domain determined by the encryption keys. The proposed algorithm can resist statistic analyses effectively. The computer simulation results show that the proposed encryption algorithm is sensitive to the multiple keys, and that it has considerable robustness, noise immunity and security.

  4. Establishment of an efficient virus-induced gene silencing (VIGS) assay in Arabidopsis by Agrobacterium-mediated rubbing infection.

    PubMed

    Manhães, Ana Marcia E de A; de Oliveira, Marcos V V; Shan, Libo

    2015-01-01

    Several VIGS protocols have been established for high-throughput functional genomic screens as it bypasses the time-consuming and laborious process of generation of transgenic plants. The silencing efficiency in this approach is largely hindered by a technically demanding step in which the first pair of newly emerged true leaves at the 2-week-old stage are infiltrated with a needleless syringe. To further optimize VIGS efficiency and achieve rapid inoculation for a large-scale functional genomic study, here we describe a protocol of an efficient VIGS assay in Arabidopsis using Agrobacterium-mediated rubbing infection. The Agrobacterium inoculation is performed by simply rubbing the leaves with Filter Agent Celite(®) 545. The highly efficient and uniform silencing effect was indicated by the development of a visibly albino phenotype due to silencing of the Cloroplastos alterados 1 (CLA1) gene in the newly emerged leaves. In addition, the albino phenotype could be observed in stems and flowers, indicating its potential application for gene functional studies in the late vegetative development and flowering stages.

  5. Image restoration method based on Hilbert transform for full-field optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Na, Jihoon; Choi, Woo June; Choi, Eun Seo; Ryu, Seon Young; Lee, Byeong Ha

    2008-01-01

    A full-field optical coherence tomography (FF-OCT) system utilizing a simple but novel image restoration method suitable for a high-speed system is demonstrated. An en-face image is retrieved from only two phase-shifted interference fringe images through using the mathematical Hilbert transform. With a thermal light source, a high-resolution FF-OCT system having axial and transverse resolutions of 1 and 2.2 μm, respectively, was implemented. The feasibility of the proposed scheme is confirmed by presenting the obtained en-face images of biological samples such as a piece of garlic and a gold beetle. The proposed method is robust to the error in the amount of the phase shift and does not leave residual fringes. The use of just two interference images and the strong immunity to phase errors provide great advantages in the imaging speed and the system design flexibility of a high-speed high-resolution FF-OCT system.

  6. Alfalfa (Medicago sativa L.).

    PubMed

    Fu, Chunxiang; Hernandez, Timothy; Zhou, Chuanen; Wang, Zeng-Yu

    2015-01-01

    Alfalfa (Medicago sativa L.) is a high-quality forage crop widely grown throughout the world. This chapter describes an efficient protocol that allows for the generation of large number of transgenic alfalfa plants by sonication-assisted Agrobacterium-mediated transformation. Binary vectors carrying different selectable marker genes that confer resistance to phosphinothricin (bar), kanamycin (npt II), or hygromycin (hph) were used to generate transgenic alfalfa plants. Intact trifoliates collected from clonally propagated plants in the greenhouse were sterilized with bleach and then inoculated with Agrobacterium strain EHA105. More than 80 % of infected leaf pieces could produce rooted transgenic plants in 4-5 months after Agrobacterium-mediated transformation.

  7. A contourlet transform based algorithm for real-time video encoding

    NASA Astrophysics Data System (ADS)

    Katsigiannis, Stamos; Papaioannou, Georgios; Maroulis, Dimitris

    2012-06-01

    In recent years, real-time video communication over the internet has been widely utilized for applications like video conferencing. Streaming live video over heterogeneous IP networks, including wireless networks, requires video coding algorithms that can support various levels of quality in order to adapt to the network end-to-end bandwidth and transmitter/receiver resources. In this work, a scalable video coding and compression algorithm based on the Contourlet Transform is proposed. The algorithm allows for multiple levels of detail, without re-encoding the video frames, by just dropping the encoded information referring to higher resolution than needed. Compression is achieved by means of lossy and lossless methods, as well as variable bit rate encoding schemes. Furthermore, due to the transformation utilized, it does not suffer from blocking artifacts that occur with many widely adopted compression algorithms. Another highly advantageous characteristic of the algorithm is the suppression of noise induced by low-quality sensors usually encountered in web-cameras, due to the manipulation of the transform coefficients at the compression stage. The proposed algorithm is designed to introduce minimal coding delay, thus achieving real-time performance. Performance is enhanced by utilizing the vast computational capabilities of modern GPUs, providing satisfactory encoding and decoding times at relatively low cost. These characteristics make this method suitable for applications like video-conferencing that demand real-time performance, along with the highest visual quality possible for each user. Through the presented performance and quality evaluation of the algorithm, experimental results show that the proposed algorithm achieves better or comparable visual quality relative to other compression and encoding methods tested, while maintaining a satisfactory compression ratio. Especially at low bitrates, it provides more human-eye friendly images compared to

  8. A dark incubation period is important for Agrobacterium-mediated transformation of mature internode explants of sweet orange, grapefruit, citron, and a citrange rootstock.

    PubMed

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

    2012-01-01

    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. 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. 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.

  9. Osmoregulation in Agrobacterium tumefaciens: accumulation of a novel disaccharide is controlled by osmotic strength and glycine betaine.

    PubMed Central

    Smith, L T; Smith, G M; Madkour, M A

    1990-01-01

    We have investigated the mechanism of osmotic stress adaptation (osmoregulation) in Agrobacterium tumefaciens biotype I (salt-tolerant) and biotype II (salt-sensitive) strains. Using natural-abundance 13C nuclear magnetic resonance spectroscopy, we identified all organic solutes that accumulated to significant levels in osmotically stressed cultures. When stressed, biotype I strains (C58, NT1, and A348) accumulated glutamate and a novel disaccharide, beta-fructofuranosyl-alpha-mannopyranoside, commonly known as mannosucrose. In the salt-sensitive biotype II strain K84, glutamate was observed but mannosucrose was not. We speculate that mannosucrose confers the extra osmotic tolerance observed in the biotype I strains. In addition to identifying the osmoregulated solutes that this species synthesizes, we investigated the ability of A. tumefaciens to utilize the powerful osmotic stress protectant glycine betaine when it is supplied in the medium. Results from growth experiments, nuclear magnetic resonance spectroscopy, and a 14C labeling experiment demonstrated that in the absence of osmotic stress, glycine betaine was metabolized, while in stressed cultures, glycine betaine accumulated intracellularly and conferred enhanced osmotic stress tolerance. Furthermore, when glycine betaine was taken up in stressed cells, its accumulation caused the intracellular concentration of mannosucrose to drop significantly. The possible role of osmoregulation of A. tumefaciens in the transformation of plants is discussed. PMID:2254260

  10. Artificial retina model for the retinally blind based on wavelet transform

    NASA Astrophysics Data System (ADS)

    Zeng, Yan-an; Song, Xin-qiang; Jiang, Fa-gang; Chang, Da-ding

    2007-01-01

    Artificial retina is aimed for the stimulation of remained retinal neurons in the patients with degenerated photoreceptors. Microelectrode arrays have been developed for this as a part of stimulator. Design such microelectrode arrays first requires a suitable mathematical method for human retinal information processing. In this paper, a flexible and adjustable human visual information extracting model is presented, which is based on the wavelet transform. With the flexible of wavelet transform to image information processing and the consistent to human visual information extracting, wavelet transform theory is applied to the artificial retina model for the retinally blind. The response of the model to synthetic image is shown. The simulated experiment demonstrates that the model behaves in a manner qualitatively similar to biological retinas and thus may serve as a basis for the development of an artificial retina.

  11. A Method for Transforming Existing Web Service Descriptions into an Enhanced Semantic Web Service Framework

    NASA Astrophysics Data System (ADS)

    Du, Xiaofeng; Song, William; Munro, Malcolm

    Web Services as a new distributed system technology has been widely adopted by industries in the areas, such as enterprise application integration (EAI), business process management (BPM), and virtual organisation (VO). However, lack of semantics in the current Web Service standards has been a major barrier in service discovery and composition. In this chapter, we propose an enhanced context-based semantic service description framework (CbSSDF+) that tackles the problem and improves the flexibility of service discovery and the correctness of generated composite services. We also provide an agile transformation method to demonstrate how the various formats of Web Service descriptions on the Web can be managed and renovated step by step into CbSSDF+ based service description without large amount of engineering work. At the end of the chapter, we evaluate the applicability of the transformation method and the effectiveness of CbSSDF+ through a series of experiments.

  12. The backward phase flow and FBI-transform-based Eulerian Gaussian beams for the Schroedinger equation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Leung Shingyu, E-mail: masyleung@ust.h; Qian Jianliang, E-mail: qian@math.msu.ed

    2010-11-20

    We propose the backward phase flow method to implement the Fourier-Bros-Iagolnitzer (FBI)-transform-based Eulerian Gaussian beam method for solving the Schroedinger equation in the semi-classical regime. The idea of Eulerian Gaussian beams has been first proposed in . In this paper we aim at two crucial computational issues of the Eulerian Gaussian beam method: how to carry out long-time beam propagation and how to compute beam ingredients rapidly in phase space. By virtue of the FBI transform, we address the first issue by introducing the reinitialization strategy into the Eulerian Gaussian beam framework. Essentially we reinitialize beam propagation by applying themore » FBI transform to wavefields at intermediate time steps when the beams become too wide. To address the second issue, inspired by the original phase flow method, we propose the backward phase flow method which allows us to compute beam ingredients rapidly. Numerical examples demonstrate the efficiency and accuracy of the proposed algorithms.« less

  13. Disassembly of synthetic Agrobacterium T-DNA–protein complexes via the host SCFVBF ubiquitin–ligase complex pathway

    PubMed Central

    Zaltsman, Adi; Lacroix, Benoît; Gafni, Yedidya; Citovsky, Vitaly

    2013-01-01

    One the most intriguing, yet least studied, aspects of the bacterium–host plant interaction is the role of the host ubiquitin/proteasome system (UPS) in the infection process. Increasing evidence indicates that pathogenic bacteria subvert the host UPS to facilitate infection. Although both mammalian and plant bacterial pathogens are known to use the host UPS, the first prokaryotic F-box protein, an essential component of UPS, was identified in Agrobacterium. During its infection, which culminates in genetic modification of the host cell, Agrobacterium transfers its T-DNA—as a complex (T-complex) with the bacterial VirE2 and host VIP1 proteins—into the host cell nucleus. There the T-DNA is uncoated from its protein components before undergoing integration into the host genome. It has been suggested that the host UPS mediates this uncoating process, but there is no evidence indicating that this activity can unmask the T-DNA molecule. Here we provide support for the idea that the plant UPS uncoats synthetic T-complexes via the Skp1/Cullin/F-box protein VBF pathway and exposes the T-DNA molecule to external enzymatic activity. PMID:23248273

  14. Transformation and Evaluation of Cry1Ac+Cry2A and GTGene in Gossypium hirsutum L.

    PubMed Central

    Puspito, Agung N.; Rao, Abdul Q.; Hafeez, Muhammad N.; Iqbal, Muhammad S.; Bajwa, Kamran S.; Ali, Qurban; Rashid, Bushra; Abbas, Muhammad A.; Latif, Ayesha; Shahid, Ahmad A.; Nasir, Idrees A.; Husnain, Tayyab

    2015-01-01

    More than 50 countries around the globe cultivate cotton on a large scale. It is a major cash crop of Pakistan and is considered “white gold” because it is highly important to the economy of Pakistan. In addition to its importance, cotton cultivation faces several problems, such as insect pests, weeds, and viruses. In the past, insects have been controlled by insecticides, but this method caused a severe loss to the economy. However, conventional breeding methods have provided considerable breakthroughs in the improvement of cotton, but it also has several limitations. In comparison with conventional methods, biotechnology has the potential to create genetically modified plants that are environmentally safe and economically viable. In this study, a local cotton variety VH 289 was transformed with two Bt genes (Cry1Ac and Cry2A) and a herbicide resistant gene (cp4 EPSPS) using the Agrobacterium mediated transformation method. The constitutive CaMV 35S promoter was attached to the genes taken from Bacillus thuringiensis (Bt) and to an herbicide resistant gene during cloning, and this promoter was used for the expression of the genes in cotton plants. This construct was used to develop the Glyphosate Tolerance Gene (GTGene) for herbicide tolerance and insecticidal gene (Cry1Ac and Cry2A) for insect tolerance in the cotton variety VH 289. The transgenic cotton variety performed 85% better compared with the non-transgenic variety. The study results suggest that farmers should use the transgenic cotton variety for general cultivation to improve the production of cotton. PMID:26617613

  15. Bioassays of quorum sensing compounds using Agrobacterium tumefaciens and Chromobacterium violaceum.

    PubMed

    Chu, Weihua; Vattem, Dhiraj A; Maitin, Vatsala; Barnes, Mary B; McLean, Robert J C

    2011-01-01

    In most bacteria, a global level of regulation exists involving intercellular communication via the production and response to cell density-dependent signal molecules. This cell density-dependent regulation has been termed quorum sensing (QS). QS is a global regulator, which has been associated with a number of important features in bacteria including virulence regulation and biofilm formation. Consequently, there is considerable interest in understanding, detecting, and inhibiting QS. Acyl homoserine lactones (acyl HSLs) are used as extracellular QS signals by a variety of Gram-negative bacteria. Chromobacterium violaceum, a Gram-negative bacterium commonly found in soil and water, produces the characteristic purple pigment violacein, the production of which is regulated by acyl HSL-mediated QS. Based on this readily observed pigmentation phenotype, C. violaceum strains can be used to detect various aspects of acyl HSL-mediated QS activity. In another commonly used bioassay organism, Agrobacterium tumefaciens, QS can be detected by the use of a reporter gene such as lacZ. Here, we describe several commonly used approaches incorporating C. violaceum and A. tumefaciens that can be used to detect acyl HSLs and QS inhibition.

  16. Influence of volatile organic compounds emitted by Pseudomonas and Serratia strains on Agrobacterium tumefaciens biofilms.

    PubMed

    Plyuta, Vladimir; Lipasova, Valentina; Popova, Alexandra; Koksharova, Olga; Kuznetsov, Alexander; Szegedi, Erno; Chernin, Leonid; Khmel, Inessa

    2016-07-01

    The ability to form biofilms plays an important role in bacteria-host interactions, including plant pathogenicity. In this work, we investigated the action of volatile organic compounds (VOCs) produced by rhizospheric strains of Pseudomonas chlororaphis 449, Pseudomonas fluorescens B-4117, Serratia plymuthica IC1270, as well as Serratia proteamaculans strain 94, isolated from spoiled meat, on biofilms formation by three strains of Agrobacterium tumefaciens which are causative agents of crown-gall disease in a wide range of plants. In dual culture assays, the pool of volatiles emitted by the tested Pseudomonas and Serratia strains suppressed the formation of biofilms of A. tumefaciens strains grown on polycarbonate membrane filters and killed Agrobacterium cells in mature biofilms. The individual VOCs produced by the tested Pseudomonas strains, that is, ketones (2-nonanone, 2-heptanone, 2-undecanone), and dimethyl disulfide (DMDS) produced by Serratia strains, were shown to kill A. tumefaciens cells in mature biofilms and suppress their formation. The data obtained in this study suggest an additional potential of some ketones and DMDS as protectors of plants against A. tumefaciens strains, whose virulence is associated with the formation of biofilms on the infected plants. © 2016 APMIS. Published by John Wiley & Sons Ltd.

  17. Non-singular acoustic cloak derived by the ray tracing method with rotationally symmetric transformations

    PubMed Central

    Wu, Linzhi

    2016-01-01

    Recently, the ray tracing method has been used to derive the non-singular cylindrical invisibility cloaks for out-of-plane shear waves, which is impossible via the transformation method directly owing to the singular push-forward mapping. In this paper, the method is adopted to design a kind of non-singular acoustic cloak. Based on Hamilton's equations of motion, eikonal equation and pre-designed ray equations, we derive several constraint equations for bulk modulus and density tensor. On the premise that the perfect matching conditions are satisfied, a series of non-singular physical profiles can be obtained by arranging the singular terms reasonably. The physical profiles derived by the ray tracing method will degenerate to the transformation-based solutions when taking the transport equation into consideration. This illuminates the essence of the newly designed cloaks that they are actually the so-called eikonal cloaks that can accurately control the paths of energy flux but with small disturbance in energy distribution along the paths. The near-perfect invisible performance has been demonstrated by the numerical ray tracing results and the pressure distribution snapshots. Finally, a kind of reduced cloak is conceived, and the good invisible performance has been measured quantitatively by the normalized scattering width. PMID:27118884

  18. Abiotic and biotic factors responsible for antimonite oxidation in Agrobacterium tumefaciens GW4

    PubMed Central

    Li, Jingxin; Yang, Birong; Shi, Manman; Yuan, Kai; Guo, Wei; Wang, Qian; Wang, Gejiao

    2017-01-01

    Antimonite [Sb(III)]-oxidizing bacteria can transform the toxic Sb(III) into the less toxic antimonate [Sb(V)]. Recently, the cytoplasmic Sb(III)-oxidase AnoA and the periplasmic arsenite [As(III)] oxidase AioAB were shown to responsible for bacterial Sb(III) oxidation, however, disruption of each gene only partially decreased Sb(III) oxidation efficiency. This study showed that in Agrobacterium tumefaciens GW4, Sb(III) induced cellular H2O2 content and H2O2 degradation gene katA. Gene knock-out/complementation of katA, anoA, aioA and anoA/aioA and Sb(III) oxidation and growth experiments showed that katA, anoA and aioA were essential for Sb(III) oxidation and resistance and katA was also essential for H2O2 resistance. Furthermore, linear correlations were observed between cellular H2O2 and Sb(V) content in vivo and chemical H2O2 and Sb(V) content in vitro (R2 = 0.93 and 0.94, respectively). These results indicate that besides the biotic factors, the cellular H2O2 induced by Sb(III) also catalyzes bacterial Sb(III) oxidation as an abiotic oxidant. The data reveal a novel mechanism that bacterial Sb(III) oxidation is associated with abiotic (cellular H2O2) and biotic (AnoA and AioAB) factors and Sb(III) oxidation process consumes cellular H2O2 which contributes to microbial detoxification of both Sb(III) and cellular H2O2. PMID:28252030

  19. Abiotic and biotic factors responsible for antimonite oxidation in Agrobacterium tumefaciens GW4

    NASA Astrophysics Data System (ADS)

    Li, Jingxin; Yang, Birong; Shi, Manman; Yuan, Kai; Guo, Wei; Wang, Qian; Wang, Gejiao

    2017-03-01

    Antimonite [Sb(III)]-oxidizing bacteria can transform the toxic Sb(III) into the less toxic antimonate [Sb(V)]. Recently, the cytoplasmic Sb(III)-oxidase AnoA and the periplasmic arsenite [As(III)] oxidase AioAB were shown to responsible for bacterial Sb(III) oxidation, however, disruption of each gene only partially decreased Sb(III) oxidation efficiency. This study showed that in Agrobacterium tumefaciens GW4, Sb(III) induced cellular H2O2 content and H2O2 degradation gene katA. Gene knock-out/complementation of katA, anoA, aioA and anoA/aioA and Sb(III) oxidation and growth experiments showed that katA, anoA and aioA were essential for Sb(III) oxidation and resistance and katA was also essential for H2O2 resistance. Furthermore, linear correlations were observed between cellular H2O2 and Sb(V) content in vivo and chemical H2O2 and Sb(V) content in vitro (R2 = 0.93 and 0.94, respectively). These results indicate that besides the biotic factors, the cellular H2O2 induced by Sb(III) also catalyzes bacterial Sb(III) oxidation as an abiotic oxidant. The data reveal a novel mechanism that bacterial Sb(III) oxidation is associated with abiotic (cellular H2O2) and biotic (AnoA and AioAB) factors and Sb(III) oxidation process consumes cellular H2O2 which contributes to microbial detoxification of both Sb(III) and cellular H2O2.

  20. IMPROVING PLANT GENETIC ENGINEERING BY MANIPULATING THE HOST. (R829479C001)

    EPA Science Inventory

    Agrobacterium-mediated transformation is a major technique for the genetic engineering of plants. However, there are many economically important crop and tree species that remain highly recalcitrant to Agrobacterium infection. Although attempts have been made to ...

  1. Potato (Solanum tuberosum L.).

    PubMed

    Chetty, Venkateswari J; Narváez-Vásquez, Javier; Orozco-Cárdenas, Martha L

    2015-01-01

    Agrobacterium-mediated transformation is the most common method for the incorporation of foreign genes into the genome of potato as well as many other species in the Solanaceae family. This chapter describes protocols for the genetic transformation of three species of potato: Solanum tuberosum subsp. tuberosum (Desiréé), S. tuberosum subsp. andigenum (Blue potato), and S. tuberosum subsp. andigena using internodal segments as explants.

  2. Development of a Coordinate Transformation method for direct georeferencing in map projection frames

    NASA Astrophysics Data System (ADS)

    Zhao, Haitao; Zhang, Bing; Wu, Changshan; Zuo, Zhengli; Chen, Zhengchao

    2013-03-01

    This paper develops a novel Coordinate Transformation method (CT-method), with which the orientation angles (roll, pitch, heading) of the local tangent frame of the GPS/INS system are transformed into those (omega, phi, kappa) of the map projection frame for direct georeferencing (DG). Especially, the orientation angles in the map projection frame were derived from a sequence of coordinate transformations. The effectiveness of orientation angles transformation was verified through comparing with DG results obtained from conventional methods (Legat method and POSPac method) using empirical data. Moreover, the CT-method was also validated with simulated data. One advantage of the proposed method is that the orientation angles can be acquired simultaneously while calculating position elements of exterior orientation (EO) parameters and auxiliary points coordinates by coordinate transformation. These three methods were demonstrated and compared using empirical data. Empirical results show that the CT-method is both as sound and effective as Legat method. Compared with POSPac method, the CT-method is more suitable for calculating EO parameters for DG in map projection frames. DG accuracy of the CT-method and Legat method are at the same level. DG results of all these three methods have systematic errors in height due to inconsistent length projection distortion in the vertical and horizontal components, and these errors can be significantly reduced using the EO height correction technique in Legat's approach. Similar to the results obtained with empirical data, the effectiveness of the CT-method was also proved with simulated data. POSPac method: The method is presented by Applanix POSPac software technical note (Hutton and Savina, 1997). It is implemented in the POSEO module of POSPac software.

  3. Joint sparse reconstruction of multi-contrast MRI images with graph based redundant wavelet transform.

    PubMed

    Lai, Zongying; Zhang, Xinlin; Guo, Di; Du, Xiaofeng; Yang, Yonggui; Guo, Gang; Chen, Zhong; Qu, Xiaobo

    2018-05-03

    Multi-contrast images in magnetic resonance imaging (MRI) provide abundant contrast information reflecting the characteristics of the internal tissues of human bodies, and thus have been widely utilized in clinical diagnosis. However, long acquisition time limits the application of multi-contrast MRI. One efficient way to accelerate data acquisition is to under-sample the k-space data and then reconstruct images with sparsity constraint. However, images are compromised at high acceleration factor if images are reconstructed individually. We aim to improve the images with a jointly sparse reconstruction and Graph-based redundant wavelet transform (GBRWT). First, a sparsifying transform, GBRWT, is trained to reflect the similarity of tissue structures in multi-contrast images. Second, joint multi-contrast image reconstruction is formulated as a ℓ 2, 1 norm optimization problem under GBRWT representations. Third, the optimization problem is numerically solved using a derived alternating direction method. Experimental results in synthetic and in vivo MRI data demonstrate that the proposed joint reconstruction method can achieve lower reconstruction errors and better preserve image structures than the compared joint reconstruction methods. Besides, the proposed method outperforms single image reconstruction with joint sparsity constraint of multi-contrast images. The proposed method explores the joint sparsity of multi-contrast MRI images under graph-based redundant wavelet transform and realizes joint sparse reconstruction of multi-contrast images. Experiment demonstrate that the proposed method outperforms the compared joint reconstruction methods as well as individual reconstructions. With this high quality image reconstruction method, it is possible to achieve the high acceleration factors by exploring the complementary information provided by multi-contrast MRI.

  4. Peach (Prunus persica L.).

    PubMed

    Sabbadini, Silvia; Pandolfini, Tiziana; Girolomini, Luca; Molesini, Barbara; Navacchi, Oriano

    2015-01-01

    Until now, the application of genetic transformation techniques in peach has been limited by the difficulties in developing efficient regeneration and transformation protocols. Here we describe an efficient regeneration protocol for the commercial micropropagation of GF677 rootstock (Prunus persica × Prunus amygdalus). The method is based on the production, via organogenesis, of meristematic bulk tissues characterized by a high competence for shoot regeneration. This protocol has also been used to obtain GF677 plants genetically engineered with an empty hairpin cassette (hereafter indicated as hp-pBin19), through Agrobacterium tumefaciens-mediated transformation. After 7-8 months of selection on media containing kanamycin, we obtained two genetically modified GF677 lines. PCR and Southern blot analyses were performed to confirm the genetic status.

  5. Rolling Bearing Fault Diagnosis Based on an Improved HTT Transform

    PubMed Central

    Tang, Guiji; Tian, Tian; Zhou, Chong

    2018-01-01

    When rolling bearing failure occurs, vibration signals generally contain different signal components, such as impulsive fault feature signals, background noise and harmonic interference signals. One of the most challenging aspects of rolling bearing fault diagnosis is how to inhibit noise and harmonic interference signals, while enhancing impulsive fault feature signals. This paper presents a novel bearing fault diagnosis method, namely an improved Hilbert time–time (IHTT) transform, by combining a Hilbert time–time (HTT) transform with principal component analysis (PCA). Firstly, the HTT transform was performed on vibration signals to derive a HTT transform matrix. Then, PCA was employed to de-noise the HTT transform matrix in order to improve the robustness of the HTT transform. Finally, the diagonal time series of the de-noised HTT transform matrix was extracted as the enhanced impulsive fault feature signal and the contained fault characteristic information was identified through further analyses of amplitude and envelope spectrums. Both simulated and experimental analyses validated the superiority of the presented method for detecting bearing failures. PMID:29662013

  6. Pepper, chili (Capsicum annuum).

    PubMed

    Min, Jung; Shin, Sun Hee; Jeon, En Mi; Park, Jung Mi; Hyun, Ji Young; Harn, Chee Hark

    2015-01-01

    Pepper is a recalcitrant plant for Agrobacterium-mediated genetic transformation. Several obstacles to genetic transformation remain such as extremely low transformation rates; the choice of correct genotype is critical; and there is a high frequency of false positives due to direct shoot formation. Here, we report a useful protocol with a suitable selection method. The most important aspect of the pepper transformation protocol is selecting shoots growing from the callus, which is referred to as callus-mediated shoot formation. This protocol is a reproducible and reliable system for pepper transformation.

  7. [Research on spatially modulated Fourier transform imaging spectrometer data processing method].

    PubMed

    Huang, Min; Xiangli, Bin; Lü, Qun-Bo; Zhou, Jin-Song; Jing, Juan-Juan; Cui, Yan

    2010-03-01

    Fourier transform imaging spectrometer is a new technic, and has been developed very rapidly in nearly ten years. The data catched by Fourier transform imaging spectrometer is indirect data, can not be used by user, and need to be processed by various approaches, including data pretreatment, apodization, phase correction, FFT, and spectral radicalization calibration. No paper so far has been found roundly to introduce this method. In the present paper, the author will give an effective method to process the interfering data to spectral data, and with this method we can obtain good result.

  8. Tensor-Train Split-Operator Fourier Transform (TT-SOFT) Method: Multidimensional Nonadiabatic Quantum Dynamics.

    PubMed

    Greene, Samuel M; Batista, Victor S

    2017-09-12

    We introduce the "tensor-train split-operator Fourier transform" (TT-SOFT) method for simulations of multidimensional nonadiabatic quantum dynamics. TT-SOFT is essentially the grid-based SOFT method implemented in dynamically adaptive tensor-train representations. In the same spirit of all matrix product states, the tensor-train format enables the representation, propagation, and computation of observables of multidimensional wave functions in terms of the grid-based wavepacket tensor components, bypassing the need of actually computing the wave function in its full-rank tensor product grid space. We demonstrate the accuracy and efficiency of the TT-SOFT method as applied to propagation of 24-dimensional wave packets, describing the S 1 /S 2 interconversion dynamics of pyrazine after UV photoexcitation to the S 2 state. Our results show that the TT-SOFT method is a powerful computational approach for simulations of quantum dynamics of polyatomic systems since it avoids the exponential scaling problem of full-rank grid-based representations.

  9. Methods for transforming and expression screening of filamentous fungal cells with a DNA library

    DOEpatents

    Teter, Sarah; Lamsa, Michael; Cherry, Joel; Ward, Connie

    2015-06-02

    The present invention relates to methods for expression screening of filamentous fungal transformants, comprising: (a) isolating single colony transformants of a DNA library introduced into E. coli; (b) preparing DNA from each of the single colony E. coli transformants; (c) introducing a sample of each of the DNA preparations of step (b) into separate suspensions of protoplasts of a filamentous fungus to obtain transformants thereof, wherein each transformant contains one or more copies of an individual polynucleotide from the DNA library; (d) growing the individual filamentous fungal transformants of step (c) on selective growth medium, thereby permitting growth of the filamentous fungal transformants, while suppressing growth of untransformed filamentous fungi; and (e) measuring activity or a property of each polypeptide encoded by the individual polynucleotides. The present invention also relates to isolated polynucleotides encoding polypeptides of interest obtained by such methods, to nucleic acid constructs, expression vectors, and recombinant host cells comprising the isolated polynucleotides, and to methods of producing the polypeptides encoded by the isolated polynucleotides.

  10. A singlechip-computer-controlled conductivity meter based on conductance-frequency transformation

    NASA Astrophysics Data System (ADS)

    Chen, Wenxiang; Hong, Baocai

    2005-02-01

    A portable conductivity meter controlled by singlechip computer was designed. The instrument uses conductance-frequency transformation method to measure the conductivity of solution. The circuitry is simple and reliable. Another feature of the instrument is that the temperature compensation is realised by changing counting time of the timing counter. The theoretical based and the usage of temperature compensation are narrated.

  11. Space-time least-squares finite element method for convection-reaction system with transformed variables

    PubMed Central

    Nam, Jaewook

    2011-01-01

    We present a method to solve a convection-reaction system based on a least-squares finite element method (LSFEM). For steady-state computations, issues related to recirculation flow are stated and demonstrated with a simple example. The method can compute concentration profiles in open flow even when the generation term is small. This is the case for estimating hemolysis in blood. Time-dependent flows are computed with the space-time LSFEM discretization. We observe that the computed hemoglobin concentration can become negative in certain regions of the flow; it is a physically unacceptable result. To prevent this, we propose a quadratic transformation of variables. The transformed governing equation can be solved in a straightforward way by LSFEM with no sign of unphysical behavior. The effect of localized high shear on blood damage is shown in a circular Couette-flow-with-blade configuration, and a physiological condition is tested in an arterial graft flow. PMID:21709752

  12. Smeared spectrum jamming suppression based on generalized S transform and threshold segmentation

    NASA Astrophysics Data System (ADS)

    Li, Xin; Wang, Chunyang; Tan, Ming; Fu, Xiaolong

    2018-04-01

    Smeared Spectrum (SMSP) jamming is an effective jamming in countering linear frequency modulation (LFM) radar. According to the time-frequency distribution difference between jamming and echo, a jamming suppression method based on Generalized S transform (GST) and threshold segmentation is proposed. The sub-pulse period is firstly estimated based on auto correlation function firstly. Secondly, the time-frequency image and the related gray scale image are achieved based on GST. Finally, the Tsallis cross entropy is utilized to compute the optimized segmentation threshold, and then the jamming suppression filter is constructed based on the threshold. The simulation results show that the proposed method is of good performance in the suppression of false targets produced by SMSP.

  13. Iris Location Algorithm Based on the CANNY Operator and Gradient Hough Transform

    NASA Astrophysics Data System (ADS)

    Zhong, L. H.; Meng, K.; Wang, Y.; Dai, Z. Q.; Li, S.

    2017-12-01

    In the iris recognition system, the accuracy of the localization of the inner and outer edges of the iris directly affects the performance of the recognition system, so iris localization has important research meaning. Our iris data contain eyelid, eyelashes, light spot and other noise, even the gray transformation of the images is not obvious, so the general methods of iris location are unable to realize the iris location. The method of the iris location based on Canny operator and gradient Hough transform is proposed. Firstly, the images are pre-processed; then, calculating the gradient information of images, the inner and outer edges of iris are coarse positioned using Canny operator; finally, according to the gradient Hough transform to realize precise localization of the inner and outer edge of iris. The experimental results show that our algorithm can achieve the localization of the inner and outer edges of the iris well, and the algorithm has strong anti-interference ability, can greatly reduce the location time and has higher accuracy and stability.

  14. A transformation method for constrained-function minimization

    NASA Technical Reports Server (NTRS)

    Park, S. K.

    1975-01-01

    A direct method for constrained-function minimization is discussed. The method involves the construction of an appropriate function mapping all of one finite dimensional space onto the region defined by the constraints. Functions which produce such a transformation are constructed for a variety of constraint regions including, for example, those arising from linear and quadratic inequalities and equalities. In addition, the computational performance of this method is studied in the situation where the Davidon-Fletcher-Powell algorithm is used to solve the resulting unconstrained problem. Good performance is demonstrated for 19 test problems by achieving rapid convergence to a solution from several widely separated starting points.

  15. Comparison of methods applied in photoinduced transient spectroscopy to determining the defect center parameters: The correlation procedure and the signal analysis based on inverse Laplace transformation

    NASA Astrophysics Data System (ADS)

    Suproniuk, M.; Pawłowski, M.; Wierzbowski, M.; Majda-Zdancewicz, E.; Pawłowski, Ma.

    2018-04-01

    The procedure for determination of trap parameters by photo-induced transient spectroscopy is based on the Arrhenius plot that illustrates a thermal dependence of the emission rate. In this paper, we show that the Arrhenius plot obtained by the correlation method is shifted toward lower temperatures as compared to the one obtained with the inverse Laplace transformation. This shift is caused by the model adequacy error of the correlation method and introduces errors to a calculation procedure of defect center parameters. The effect is exemplified by comparing the results of the determination of trap parameters with both methods based on photocurrent transients for defect centers observed in tin-doped neutron-irradiated silicon crystals and in gallium arsenide grown with the Vertical Gradient Freeze method.

  16. Fast and secure encryption-decryption method based on chaotic dynamics

    DOEpatents

    Protopopescu, Vladimir A.; Santoro, Robert T.; Tolliver, Johnny S.

    1995-01-01

    A method and system for the secure encryption of information. The method comprises the steps of dividing a message of length L into its character components; generating m chaotic iterates from m independent chaotic maps; producing an "initial" value based upon the m chaotic iterates; transforming the "initial" value to create a pseudo-random integer; repeating the steps of generating, producing and transforming until a pseudo-random integer sequence of length L is created; and encrypting the message as ciphertext based upon the pseudo random integer sequence. A system for accomplishing the invention is also provided.

  17. Effectiveness of Discovery Learning-Based Transformation Geometry Module

    NASA Astrophysics Data System (ADS)

    Febriana, R.; Haryono, Y.; Yusri, R.

    2017-09-01

    Development of transformation geometry module is conducted because the students got difficulties to understand the existing book. The purpose of the research was to find out the effectiveness of discovery learning-based transformation geometry module toward student’s activity. Model of the development was Plomp model consisting preliminary research, prototyping phase and assessment phase. The research was focused on assessment phase where it was to observe the designed product effectiveness. The instrument was observation sheet. The observed activities were visual activities, oral activities, listening activities, mental activities, emotional activities and motor activities. Based on the result of the research, it is found that visual activities, learning activities, writing activities, the student’s activity is in the criteria very effective. It can be concluded that the use of discovery learning-based transformation geometry module use can increase the positive student’s activity and decrease the negative activity.

  18. Linear Transformation Method for Multinuclide Decay Calculation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Ding Yuan

    2010-12-29

    A linear transformation method for generic multinuclide decay calculations is presented together with its properties and implications. The method takes advantage of the linear form of the decay solution N(t) = F(t)N{sub 0}, where N(t) is a column vector that represents the numbers of atoms of the radioactive nuclides in the decay chain, N{sub 0} is the initial value vector of N(t), and F(t) is a lower triangular matrix whose time-dependent elements are independent of the initial values of the system.

  19. Transformer partial discharge monitoring based on optical fiber sensing

    NASA Astrophysics Data System (ADS)

    Wang, Kun; Tong, Xinglin; Zhu, Xiaolong

    2014-06-01

    The power transformer is the most important equipment of the high voltage power grid, however, some traditional methods of online partial discharge monitoring have some limitations. Based on many advantages of the optical fiber sensing technology, we have done some research on fiber optics Fabry-Perot (FP) sensing which can be useful for the transformer on online partial discharge monitoring. This research aimed at improving the reliability of power system safety monitoring. We have done some work as follows: designing a set for fiber optics FP sensor preparation, according to the fabrication procedure strictly making out the sensors, building a reasonable signal demodulation system for fiber optics FP sensing, doing a preliminary analysis about online partial discharge signal monitoring, including the research on different discharge intensities with the same measuring distance and different measuring distances with the same discharge intensity, and then making a detailed analysis of the experimental results.

  20. 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