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Sample records for generate transgenic animals

  1. The methods to generate transgenic animals and to control transgene expression.

    Science.gov (United States)

    Houdebine, Louis-Marie

    2002-09-25

    Transgenic animals have been used for years to study gene function and to create models for the study of human diseases. This approach has become still more justified after the complete sequencing of several genomes. Transgenic animals are ready to become industrial bioreactors for the preparation of pharmaceuticals in milk and probably in the future in egg white. Improvement of animal production by transgenesis is still in infancy. Despite its intensive use, animal transgenesis is still suffering from technical limitations. The generation of transgenics has recently become easier or possible for different species thanks to the use of transposons or retrovirus, to incubation of sperm which DNA followed by fertilization by intracellular sperm injection or not and to the use of the cloning technique using somatic cells in which genes have been added or inactivated. The Cre-LoxP system is more and more used to withdraw a given sequence from the genome or to target the integration of a foreign DNA. The tetracycline system has been improved and can more and more frequently be used to obtain faithful expression of transgenes. Several tools: RNA forming a triple helix with DNA, antisense RNA including double strand RNA inducing RNA interference and ribozymes, and also expression of proteins having a negative transdominant effect, are tentatively being improved to inhibit specifically the expression of host or viral genes.All these techniques are expected to offer experimenters new and more precise models to study gene function even in large animals. Improvement of breeding by transgenesis has become more plausible including through the precise allele replacement in farm animals.

  2. [Transgenic animals bioreactors].

    Science.gov (United States)

    Gou, Ke-Mian; An, Xiao-Rong; Tian, Jian-Hui; Chen, Yong-Fu

    2002-01-01

    The production of human recombinant proteins in milk of transgenic farm animals offers a safe, very cost-effective source of commercially important proteins that cannot be produced as efficiently in adequate quantities by other methods. This review has summarized the current status of gene selection, vector construct, transgenic methods, economics, and obvious potential in transgenic animals bioreactors. Recently, a more powerful approach was adopted in the transgenic animals founded on the application of nuclear transfer. As we will illustrate, this strategy presents a breakthrough in the overall efficiency of generating transgenic farm animals, product consistency, and time of product development. The successful adaptation of Cre-/lox P-mediated site-specific DNA recombination systems in farm animals will offer unprecedented possibilities for generating transgenic animals.

  3. Transgenic animal bioreactors.

    Science.gov (United States)

    Houdebine, L M

    2000-01-01

    The production of recombinant proteins is one of the major successes of biotechnology. Animal cells are required to synthesize proteins with the appropriate post-translational modifications. Transgenic animals are being used for this purpose. Milk, egg white, blood, urine, seminal plasma and silk worm cocoon from transgenic animals are candidates to be the source of recombinant proteins at an industrial scale. Although the first recombinant protein produced by transgenic animals is expected to be in the market in 2000, a certain number of technical problems remain to be solved before the various systems are optimized. Although the generation of transgenic farm animals has become recently easier mainly with the technique of animal cloning using transfected somatic cells as nuclear donor, this point remains a limitation as far as cost is concerned. Numerous experiments carried out for the last 15 years have shown that the expression of the transgene is predictable only to a limited extent. This is clearly due to the fact that the expression vectors are not constructed in an appropriate manner. This undoubtedly comes from the fact that all the signals contained in genes have not yet been identified. Gene constructions thus result sometime in poorly functional expression vectors. One possibility consists in using long genomic DNA fragments contained in YAC or BAC vectors. The other relies on the identification of the major important elements required to obtain a satisfactory transgene expression. These elements include essentially gene insulators, chromatin openers, matrix attached regions, enhancers and introns. A certain number of proteins having complex structures (formed by several subunits, being glycosylated, cleaved, carboxylated...) have been obtained at levels sufficient for an industrial exploitation. In other cases, the mammary cellular machinery seems insufficient to promote all the post-translational modifications. The addition of genes coding for enzymes

  4. Generation of transgenic frogs.

    Science.gov (United States)

    Loeber, Jana; Pan, Fong Cheng; Pieler, Tomas

    2009-01-01

    The possibility of generating transgenic animals is of obvious advantage for the analysis of gene function in development and disease. One of the established vertebrate model systems in developmental biology is the amphibian Xenopus laevis. Different techniques have been successfully applied to create Xenopus transgenics; in this chapter, the so-called meganuclease method is described. This technique is not only technically simple, but also comparably efficient and applicable to both Xenopus laevis and Xenopus tropicalis. The commercially available endonuclease I-SceI (meganuclease) mediates the integration of foreign DNA into the frog genome after coinjection into fertilized eggs. Tissue-specific gene expression, as well as germline transmission, has been observed.

  5. Transgenic Farm Animals

    Science.gov (United States)

    The development of recombinant DNA technology has enabled scientists to isolate single genes, analyze and modify their nucleotide structure(s), make copies of these isolated genes, and insert copies of these genes into the genome of plants and animals. The transgenic technology of adding genes to li...

  6. Genetically engineered livestock for agriculture: a generation after the first transgenic animal research conference.

    Science.gov (United States)

    Murray, James D; Maga, Elizabeth A

    2016-06-01

    At the time of the first Transgenic Animal Research Conference, the lack of knowledge about promoter, enhancer and coding regions of genes of interest greatly hampered our efforts to create transgenes that would express appropriately in livestock. Additionally, we were limited to gene insertion by pronuclear microinjection. As predicted then, widespread genome sequencing efforts and technological advancements have profoundly altered what we can do. There have been many developments in technology to create transgenic animals since we first met at Granlibakken in 1997, including the advent of somatic cell nuclear transfer-based cloning and gene editing. We can now create new transgenes that will express when and where we want and can target precisely in the genome where we want to make a change or insert a transgene. With the large number of sequenced genomes, we have unprecedented access to sequence information including, control regions, coding regions, and known allelic variants. These technological developments have ushered in new and renewed enthusiasm for the production of transgenic animals among scientists and animal agriculturalists around the world, both for the production of more relevant biomedical research models as well as for agricultural applications. However, even though great advancements have been made in our ability to control gene expression and target genetic changes in our animals, there still are no genetically engineered animal products on the market for food. World-wide there has been a failure of the regulatory processes to effectively move forward. Estimates suggest the world will need to increase our current food production 70 % by 2050; that is we will have to produce the total amount of food each year that has been consumed by mankind over the past 500 years. The combination of transgenic animal technology and gene editing will become increasingly more important tools to help feed the world. However, to date the practical benefits of

  7. Transgenic animals resistant to infectious diseases.

    Science.gov (United States)

    Tiley, L

    2016-04-01

    The list of transgenic animals developed to test ways of producing livestock resistant to infectious disease continues to grow. Although the basic techniques for generating transgenic animals have not changed very much in the ten years since they were last reviewed for the World Organisation for Animal Health, one recent fundamental technological advance stands to revolutionise genome engineering. The advent of technically simple and efficient site-specific gene targeting has profound implications for genetically modifying livestock species.

  8. Transgenic Animal Mutation Assays

    Institute of Scientific and Technical Information of China (English)

    Tao Chen; Ph.D.D.A.B.T.

    2005-01-01

    @@ The novel transgenic mouse and rat mutation assays have provided a tool for analyzing in vivo mutation in any tissue, thus permitting the direct comparison of cancer incidence with mutant frequency.

  9. Transgenic animals and their application in medicine

    Directory of Open Access Journals (Sweden)

    Bagle TR, Kunkulol RR, Baig MS, More SY

    2013-01-01

    Full Text Available Transgenic animals are animals that are genetically altered to have traits that mimic symptoms of specific human pathologies. They provide genetic models of various human diseases which are important in understanding disease and developing new targets. In early 1980 Gordon and co-workers described the first gene addition experiment using the microinjection technology and since then the impact of transgenic technology on basic research has been significant. Within 20 years of its inception, ATryn the first drug approved by USFDA from transgenic animals was developed and it has opened door to drugs from transgenic animals. In addition, they are looked upon as potential future donors for xenotransplantation. With increasing knowledge about the genetics and improvements in the transgenetic technology numerous useful applications like biologically safe new-generation drugs based on human regulatory proteins are being developed.Various aspects of concern in the coming years are the regulatory guidelines, ethical issues and patents related to the use of transgenic animals. This modern medicine is on the threshold of a pharmacological revolution. Use of transgenic animals will provide solutions for drug research, xenotransplantation, clinical trials and will prove to be a new insight in drug development.

  10. Cancer immunotherapy : insights from transgenic animal models

    NARCIS (Netherlands)

    McLaughlin, PMJ; Kroesen, BJ; Harmsen, MC; de Leij, LFMH

    2001-01-01

    A wide range of strategies in cancer immunotherapy has been developed in the last decade, some of which are currently being used in clinical settings. The development of these immunotherapeutical strategies has been facilitated by the generation of relevant transgenic animal models. Since the

  11. Cancer immunotherapy : insights from transgenic animal models

    NARCIS (Netherlands)

    McLaughlin, PMJ; Kroesen, BJ; Harmsen, MC; de Leij, LFMH

    2001-01-01

    A wide range of strategies in cancer immunotherapy has been developed in the last decade, some of which are currently being used in clinical settings. The development of these immunotherapeutical strategies has been facilitated by the generation of relevant transgenic animal models. Since the differ

  12. Expression systems and species used for transgenic animal bioreactors.

    Science.gov (United States)

    Wang, Yanli; Zhao, Sihai; Bai, Liang; Fan, Jianglin; Liu, Enqi

    2013-01-01

    Transgenic animal bioreactors can produce therapeutic proteins with high value for pharmaceutical use. In this paper, we compared different systems capable of producing therapeutic proteins (bacteria, mammalian cells, transgenic plants, and transgenic animals) and found that transgenic animals were potentially ideal bioreactors for the synthesis of pharmaceutical protein complexes. Compared with other transgenic animal expression systems (egg white, blood, urine, seminal plasma, and silkworm cocoon), the mammary glands of transgenic animals have enormous potential. Compared with other mammalian species (pig, goat, sheep, and cow) that are currently being studied as bioreactors, rabbits offer many advantages: high fertility, easy generation of transgenic founders and offspring, insensitivity to prion diseases, relatively high milk production, and no transmission of severe diseases to humans. Noticeably, for a small- or medium-sized facility, the rabbit system is ideal to produce up to 50 kg of protein per year, considering both economical and hygienic aspects; rabbits are attractive candidates for the mammary-gland-specific expression of recombinant proteins. We also reviewed recombinant proteins that have been produced by targeted expression in the mammary glands of rabbits and discussed the limitations of transgenic animal bioreactors.

  13. Expression Systems and Species Used for Transgenic Animal Bioreactors

    Directory of Open Access Journals (Sweden)

    Yanli Wang

    2013-01-01

    Full Text Available Transgenic animal bioreactors can produce therapeutic proteins with high value for pharmaceutical use. In this paper, we compared different systems capable of producing therapeutic proteins (bacteria, mammalian cells, transgenic plants, and transgenic animals and found that transgenic animals were potentially ideal bioreactors for the synthesis of pharmaceutical protein complexes. Compared with other transgenic animal expression systems (egg white, blood, urine, seminal plasma, and silkworm cocoon, the mammary glands of transgenic animals have enormous potential. Compared with other mammalian species (pig, goat, sheep, and cow that are currently being studied as bioreactors, rabbits offer many advantages: high fertility, easy generation of transgenic founders and offspring, insensitivity to prion diseases, relatively high milk production, and no transmission of severe diseases to humans. Noticeably, for a small- or medium-sized facility, the rabbit system is ideal to produce up to 50 kg of protein per year, considering both economical and hygienic aspects; rabbits are attractive candidates for the mammary-gland-specific expression of recombinant proteins. We also reviewed recombinant proteins that have been produced by targeted expression in the mammary glands of rabbits and discussed the limitations of transgenic animal bioreactors.

  14. Expression Systems and Species Used for Transgenic Animal Bioreactors

    OpenAIRE

    Yanli Wang; Sihai Zhao; Liang Bai; Jianglin Fan; Enqi Liu

    2013-01-01

    Transgenic animal bioreactors can produce therapeutic proteins with high value for pharmaceutical use. In this paper, we compared different systems capable of producing therapeutic proteins (bacteria, mammalian cells, transgenic plants, and transgenic animals) and found that transgenic animals were potentially ideal bioreactors for the synthesis of pharmaceutical protein complexes. Compared with other transgenic animal expression systems (egg white, blood, urine, seminal plasma, and silkworm ...

  15. Expression Systems and Species Used for Transgenic Animal Bioreactors

    OpenAIRE

    Yanli Wang; Sihai Zhao; Liang Bai; Jianglin Fan; Enqi Liu

    2013-01-01

    Transgenic animal bioreactors can produce therapeutic proteins with high value for pharmaceutical use. In this paper, we compared different systems capable of producing therapeutic proteins (bacteria, mammalian cells, transgenic plants, and transgenic animals) and found that transgenic animals were potentially ideal bioreactors for the synthesis of pharmaceutical protein complexes. Compared with other transgenic animal expression systems (egg white, blood, urine, seminal plasma, and silkworm ...

  16. Generation of Transgenic Hydra by Embryo Microinjection

    Science.gov (United States)

    Juliano, Celina E.; Lin, Haifan; Steele, Robert E.

    2014-01-01

    As a member of the phylum Cnidaria, the sister group to all bilaterians, Hydra can shed light on fundamental biological processes shared among multicellular animals. Hydra is used as a model for the study of regeneration, pattern formation, and stem cells. However, research efforts have been hampered by lack of a reliable method for gene perturbations to study molecular function. The development of transgenic methods has revitalized the study of Hydra biology1. Transgenic Hydra allow for the tracking of live cells, sorting to yield pure cell populations for biochemical analysis, manipulation of gene function by knockdown and over-expression, and analysis of promoter function. Plasmid DNA injected into early stage embryos randomly integrates into the genome early in development. This results in hatchlings that express transgenes in patches of tissue in one or more of the three lineages (ectodermal epithelial, endodermal epithelial, or interstitial). The success rate of obtaining a hatchling with transgenic tissue is between 10% and 20%. Asexual propagation of the transgenic hatchling is used to establish a uniformly transgenic line in a particular lineage. Generating transgenic Hydra is surprisingly simple and robust, and here we describe a protocol that can be easily implemented at low cost. PMID:25285460

  17. Generation of bovine transgenics using somatic cell nuclear transfer

    Directory of Open Access Journals (Sweden)

    Stice Steven L

    2003-11-01

    Full Text Available Abstract The ability to produce transgenic animals through the introduction of exogenous DNA has existed for many years. However, past methods available to generate transgenic animals, such as pronuclear microinjection or the use of embryonic stem cells, have either been inefficient or not available in all animals, bovine included. More recently somatic cell nuclear transfer has provided a method to create transgenic animals that overcomes many deficiencies present in other methods. This review summarizes the benefits of using somatic cell nuclear transfer to create bovine transgenics as well as the possible opportunities this method creates for the future.

  18. Transgenic farm animals: applications in agriculture and biomedicine.

    Science.gov (United States)

    Yang, X; Tian, X C; Dai, Y; Wang, B

    2000-01-01

    During the last decade, tremendous progress has been made in the area of transgenic farm animals. While there are many important transgenic farm animal applications in agriculture, funding has been very limited and progress has been rather slow in this area. Encouragingly, the potential applications of transgenic farm animals as bioreactors for producing human therapeutic proteins and as organ donors for transplantations in humans have attracted vast funding from the private sectors. Several transgenic animal products are already in various phases of clinical trials. Estimates are, that in the near future, the worlds demands on human pharmaceutical proteins may largely be met by transgenic farm animals. While there are still major challenges ahead in the area of xenotransplantation using transgenic animal organs, transgenic tissues or cells have demonstrated promising results as a potential tool for gene therapy. Recent development on cloning, embryonic stem cells and alternative transgenic methods may further expand the transgenic applications in both agriculture and biomedicine.

  19. Generation of stable Xenopus laevis transgenic lines expressing a transgene controlled by weak promoters.

    Science.gov (United States)

    L'hostis-Guidet, Anne; Recher, Gaëlle; Guillet, Brigitte; Al-Mohammad, Abdulrahim; Coumailleau, Pascal; Tiaho, François; Boujard, Daniel; Madigou, Thierry

    2009-10-01

    Combining two existing protocols of trangenesis, namely the REMI and the I-SceI meganuclease methods, we generated Xenopus leavis expressing a transgene under the control of a promoter that presented a restricted pattern of activity and a low level of expression. This was realized by co-incubating sperm nuclei, the I-SceI enzyme and the transgene prior to transplantation into unfertilized eggs. The addition of the woodchuck hepatitis virus posttranscriptional regulatory element in our constructs further enhanced the expression of the transgene without affecting the tissue-specificity of the promoter activity. Using this combination of methods we produced high rates of fully transgenic animals that stably transmitted the transgene to the next generations with a transmission rate of 50% indicating a single integration event.

  20. [Transgenic animals: uses and limitations in the 21st century medicine].

    Science.gov (United States)

    Cavagnari, Brian M

    2010-08-01

    One of the most important advances in biotechnology during the last decades was the development of transgenic animals. In this article, I discuss why transgenic animals are excellent models to analyze gen function and regulation, and to look for new therapeutic strategies for human diseases. Moreover, their use as bioreactors to produce pharmaceutical products for the treatment of human diseases, and the possibility of generating transgenic pigs as an alternative source of organ donors for humans is also discussed.

  1. A simplified method of generating transgenic Xenopus

    OpenAIRE

    Sparrow, Duncan B.; Latinkic, Branko; Mohun, Tim J.

    2000-01-01

    Currently transgenic frog embryos are generated using restriction-enzyme-mediated integration (REMI) on decondensed sperm nuclei followed by nuclear transplantation into unfertilized eggs. We have developed a simplified version of this protocol that has the potential to increase the numbers of normally developing transgenic embryos.

  2. Using transgenic reporter assays to functionally characterize enhancers in animals.

    Science.gov (United States)

    Kvon, Evgeny Z

    2015-09-01

    Enhancers or cis-regulatory modules play an instructive role in regulating gene expression during animal development and in response to the environment. Despite their importance, we only have an incomplete map of enhancers in the genome and our understanding of the mechanisms governing their function is still limited. Recent advances in genomics provided powerful tools to generate genome-wide maps of potential enhancers. However, most of these methods are based on indirect measures of enhancer activity and have to be followed by functional testing. Animal transgenesis has been a valuable method to functionally test and characterize enhancers in vivo. In this review I discuss how different transgenic strategies are utilized to characterize enhancers in model organisms focusing on studies in Drosophila and mouse. I will further discuss recent large-scale transgenic efforts to systematically identify and catalog enhancers as well as highlight the challenges and future directions in the field.

  3. WP1: transgenic opto-animals

    Science.gov (United States)

    UŻarowska, E.; Czajkowski, Rafał; Konopka, W.

    2014-11-01

    We aim to create a set of genetic tools where permanent opsin expression (ChR or NpHR) is precisely limited to the population of neurons that express immediate early gene c-fos during a specific temporal window of behavioral training. Since the c-fos gene is only expressed in neurons that form experience-dependent ensemble, this approach will result in specific labeling of a small subset of cells that create memory trace for the learned behavior. To this end we employ two alternative inducible gene expression systems: Tet Expression System and Cre/lox System. In both cases, the temporal window for opsin induction is controlled pharmacologically, by doxycycline or tamoxifen, respectively. Both systems will be used for creating lines of transgenic animals.

  4. Generation of red fluorescent protein transgenic dogs.

    Science.gov (United States)

    Hong, So Gun; Kim, Min Kyu; Jang, Goo; Oh, Hyun Ju; Park, Jung Eun; Kang, Jung Taek; Koo, Ok Jae; Kim, Teoan; Kwon, Mo Sun; Koo, Bon Chul; Ra, Jeong Chan; Kim, Dae Yong; Ko, CheMyong; Lee, Byeong Chun

    2009-05-01

    Dogs (Canis familiaris) share many common genetic diseases with humans and development of disease models using a transgenic approach has long been awaited. However, due to the technical difficulty in obtaining fertilizable eggs and the unavailability of embryonic stem cells, no transgenic dog has been generated. Canine fetal fibroblasts were stably transfected with a red fluorescent protein (RFP) gene-expressing construct using retrovirus gene delivery method. Somatic cell nuclear transfer was then employed to replace the nucleus of an oocyte with the nucleus of the RFP-fibroblasts. Using this approach, we produced the first generation of transgenic dogs with four female and two male expressing RFP.

  5. Recent advances in the development of new transgenic animal technology.

    Science.gov (United States)

    Miao, Xiangyang

    2013-03-01

    Transgenic animal technology is one of the fastest growing biotechnology areas. It is used to integrate exogenous genes into the animal genome by genetic engineering technology so that these genes can be inherited and expressed by offspring. The transgenic efficiency and precise control of gene expression are the key limiting factors in the production of transgenic animals. A variety of transgenic technologies are available. Each has its own advantages and disadvantages and needs further study because of unresolved technical and safety issues. Further studies will allow transgenic technology to explore gene function, animal genetic improvement, bioreactors, animal disease models, and organ transplantation. This article reviews the recently developed animal transgenic technologies, including the germ line stem cell-mediated method to improve efficiency, gene targeting to improve accuracy, RNA interference-mediated gene silencing technology, zinc-finger nuclease gene targeting technology and induced pluripotent stem cell technology. These new transgenic techniques can provide a better platform to develop transgenic animals for breeding new animal varieties and promote the development of medical sciences, livestock production, and other fields.

  6. An Efficient Method for Generation of Transgenic Rats Avoiding Embryo Manipulation

    Directory of Open Access Journals (Sweden)

    Bhola Shankar Pradhan

    2016-01-01

    Full Text Available Although rats are preferred over mice as an animal model, transgenic animals are generated predominantly using mouse embryos. There are limitations in the generation of transgenic rat by embryo manipulation. Unlike mouse embryos, most of the rat embryos do not survive after male pronuclear DNA injection which reduces the efficiency of generation of transgenic rat by this method. More importantly, this method requires hundreds of eggs collected by killing several females for insertion of transgene to generate transgenic rat. To this end, we developed a noninvasive and deathless technique for generation of transgenic rats by integrating transgene into the genome of the spermatogonial cells by testicular injection of DNA followed by electroporation. After standardization of this technique using EGFP as a transgene, a transgenic disease model displaying alpha thalassemia was successfully generated using rats. This efficient method will ease the generation of transgenic rats without killing the lives of rats while simultaneously reducing the number of rats used for generation of transgenic animal.

  7. Size matters: use of YACs, BACs and PACs in transgenic animals.

    Science.gov (United States)

    Giraldo, P; Montoliu, L

    2001-04-01

    In 1993, several groups, working independently, reported the successful generation of transgenic mice with yeast artificial chromosomes (YACs) using standard techniques. The transfer of these large fragments of cloned genomic DNA correlated with optimal expression levels of the transgenes, irrespective of their location in the host genome. Thereafter, other groups confirmed the advantages of YAC transgenesis and position-independent and copy number-dependent transgene expression were demonstrated in most cases. The transfer of YACs to the germ line of mice has become popular in many transgenic facilities to guarantee faithful expression of transgenes. This technique was rapidly exported to livestock and soon transgenic rabbits, pigs and other mammals were produced with YACs. Transgenic animals were also produced with bacterial or P1-derived artificial chromosomes (BACs/PACs) with similar success. The use of YACs, BACs and PACs in transgenesis has allowed the discovery of new genes by complementation of mutations, the identification of key regulatory sequences within genomic loci that are crucial for the proper expression of genes and the design of improved animal models of human genetic diseases. Transgenesis with artificial chromosomes has proven useful in a variety of biological, medical and biotechnological applications and is considered a major breakthrough in the generation of transgenic animals. In this report, we will review the recent history of YAC/BAC/PAC-transgenic animals indicating their benefits and the potential problems associated with them. In this new era of genomics, the generation and analysis of transgenic animals carrying artificial chromosome-type transgenes will be fundamental to functionally identify and understand the role of new genes, included within large pieces of genomes, by direct complementation of mutations or by observation of their phenotypic consequences.

  8. The changing role of cell culture in the generation of transgenic livestock

    OpenAIRE

    Whitelaw, C B; Farini, E; Webster, J

    1999-01-01

    Transgenesis may allow the generation of farm animals with altered phenotype, animal models for research and animal bioreactors. Although such animals have been produced, the time and expense involved in generating transgenic livestock and then evaluating the transgene expression pattern is very restrictive. If questions about the ability and efficiency of expression could be asked solely in vitro rapid progress could be achieved. Unfortunately, experiments addressing transcriptional control ...

  9. The changing role of cell culture in the generation of transgenic livestock

    OpenAIRE

    Whitelaw, C. B.; Farini, E.; Webster, J

    1999-01-01

    Transgenesis may allow the generation of farm animals with altered phenotype, animal models for research and animal bioreactors. Although such animals have been produced, the time and expense involved in generating transgenic livestock and then evaluating the transgene expression pattern is very restrictive. If questions about the ability and efficiency of expression could be asked solely in vitro rapid progress could be achieved. Unfortunately, experiments addressing transcriptional control ...

  10. Generation of BAC transgenic epithelial organoids.

    Directory of Open Access Journals (Sweden)

    Gerald Schwank

    Full Text Available Under previously developed culture conditions, mouse and human intestinal epithelia can be cultured and expanded over long periods. These so-called organoids recapitulate the three-dimensional architecture of the gut epithelium, and consist of all major intestinal cell types. One key advantage of these ex vivo cultures is their accessibility to live imaging. So far the establishment of transgenic fluorescent reporter organoids has required the generation of transgenic mice, a laborious and time-consuming process, which cannot be extended to human cultures. Here we present a transfection protocol that enables the generation of recombinant mouse and human reporter organoids using BAC (bacterial artificial chromosome technology.

  11. Influence of Species Differences on the Neuropathology of Transgenic Huntington's Disease Animal Models

    Institute of Scientific and Technical Information of China (English)

    Xiao-Jiang Li; Shihua Li

    2012-01-01

    Transgenic animal models have revealed much about the pathogenesis of age-dependent neurodegenerative diseases and proved to be a useful tool for uncovering therapeutic targets.Huntington's disease is a well-characterized neurodegenerative disorder that is caused by expansion of a CAG repeat,which results in expansion of a polyglutamine tract in the N-terminal region of huntingtin (HTT).Similar CAG/glutamine expansions are also found to cause eight other neurodegenerative diseases that affect distinct brain regions in an agedependent manner.Identification of this CAG/glutamine expansion has led to the generation of a variety of transgenic animal models.Of these different animal models,transgenic mice have been investigated extensively,and they show similar neuropathology and phenotypes as seen in their respective diseases.The common pathological hallmark of age-dependent neurodegeneration is the formation of aggregates or inclusions consisting of misfolded proteins in the affected brain regions; however,overt or striking neurodegeneration and apoptosis have not been reported in most transgenic mouse models for age-dependent diseases,including HD.By comparing the neuropathology of transgenic HD mouse,pig,and monkey models,we found that mutant HTT is more toxic to larger animals than mice,and larger animals also show neuropathology that has not been uncovered by transgenic mouse models.This review will discuss the importancc of transgenic large animal models for analyzing the pathogenesis of neurodegenerative diseases and developing effective treatments.

  12. The potential of transgenic animals for improved agricultural productivity.

    Science.gov (United States)

    Ward, K A; Nancarrow, C D; Byrne, C R; Shanahan, C M; Murray, J D; Leish, Z; Townrow, C; Rigby, N W; Wilson, B W; Hunt, C L

    1990-09-01

    The techniques involved in the transfer of foreign DNA to domestic animals have advanced to the stage where transgenic animals that express foreign genes can be reliably produced, albeit still at low efficiency. This paper reviews the current status of some of the more important areas in agriculture where this technology is being applied. Numerous attempts have been made to modify the growth performance characteristics of domestic animals by the introduction of metallothionein/growth hormone fusion genes. A summary of our work with transgenic sheep is presented. The results demonstrate that the unregulated production of growth hormone in transgenic sheep reduces carcass fat, elevates metabolic rate and heat production, causes skeletal abnormalities and impairs survival. The introduction of new metabolic pathways to domestic animals offers an attractive approach to improved animal productivity. This paper summarises recent results of research directed towards the introduction of a cysteine biosynthetic pathway and the glyoxylate cycle to transgenic sheep. So far, the genes encoding the enzymes have been isolated and expressed both in cells in culture and in transgenic mice. The results of work currently in progress demonstrate that some modification of the fusion genes is required to enhance their expression in transgenic animals.

  13. Generation of the regulatory protein rtTA transgenic mice

    Institute of Scientific and Technical Information of China (English)

    Kang Xu; Xin-Yan Deng; Ying Yue; Zhong-Min Guo; Bing Huang; Xun Hong; Dong Xiao; Xi-Gu Chen

    2005-01-01

    AIM: To translate Tet-on system into a conditional mouse model, in which hepatitis B or C virus (HBV or HCV) gene could be spatiotemporally expressed to overcome "immune tolerance" formed during the embryonic development and "immune escape" against hepatitis virus antigen(s), an effector mouse, carrying the reverse tetracycline-responsive transcriptional activator (rtTA) gene under the tight control of liver-specific human apoE promoter, is required to be generated. METHODS: To address this end, rtTA fragment amplified by PCR was effectively inserted into the vector of pLiv.7 containing apoE promoter to create the rtTA expressing vector, I.e., pApoE-rtTA. ApoE-rtTA transgenic fragment (-6.9 kb) released from pApoE-rtTA was transferred into mice by pronucleus injection, followed by obtaining one transgene (+) founder animal from microinjection through PCR and Southern blot analysis.RESULTS: rtTA transgene which could be transmitted to subsequent generation (F1) derived from founder was expressed in a liver-specific fashion. CONCLUSION: Taken together, these findings demonstrate that rtTA transgenic mice, in which rtTA expression is appropriately targeted to the murine liver, are successfully produced, which lays a solid foundation to 'off-on-off' regulate expression of target gene (s) (e.g., HBV and/or HCV) in transgenic mice mediated by Tet-on system.

  14. Containment and competition: transgenic animals in the One Health agenda.

    Science.gov (United States)

    Lezaun, Javier; Porter, Natalie

    2015-03-01

    The development of the One World, One Health agenda coincides in time with the appearance of a different model for the management of human-animal relations: the genetic manipulation of animal species in order to curtail their ability as carriers of human pathogens. In this paper we examine two examples of this emergent transgenic approach to disease control: the development of transgenic chickens incapable of shedding avian flu viruses, and the creation of transgenic mosquitoes refractory to dengue or malaria infection. Our analysis elaborates three distinctions between the One World, One Health agenda and its transgenic counterpoint. The first concerns the conceptualization of outbreaks and the forms of surveillance that support disease control efforts. The second addresses the nature of the interspecies interface, and the relative role of humans and animals in preventing pathogen transmission. The third axis of comparison considers the proprietary dimensions of transgenic animals and their implications for the assumed public health ethos of One Health programs. We argue that the fundamental difference between these two approaches to infectious disease control can be summarized as one between strategies of containment and strategies of competition. While One World, One Health programs seek to establish an equilibrium in the human-animal interface in order to contain the circulation of pathogens across species, transgenic strategies deliberately trigger a new ecological dynamic by introducing novel animal varieties designed to out-compete pathogen-carrying hosts and vectors. In other words, while One World, One Health policies focus on introducing measures of inter-species containment, transgenic approaches derive their prophylactic benefit from provoking new cycles of intra-species competition between GM animals and their wild-type counterparts. The coexistence of these divergent health protection strategies, we suggest, helps to elucidate enduring tensions and

  15. High-level expressing YAC vector for transgenic animal bioreactors.

    Science.gov (United States)

    Fujiwara, Y; Miwa, M; Takahashi, R; Kodaira, K; Hirabayashi, M; Suzuki, T; Ueda, M

    1999-04-01

    The position effect is one major problem in the production of transgenic animals as mammary gland bioreactors. In the present study, we introduced the human growth hormone (hGH) gene into 210-kb human alpha-lactalbumin position-independent YAC vectors using homologous recombination and produced transgenic rats via microinjection of YAC DNA into rat embryos. The efficiency of producing transgenic rats with the YAC vector DNA was the same as that using plasmid constructs. All analyzed transgenic rats had one copy of the transgene and produced milk containing a high level of hGH (0.25-8.9 mg/ml). In transgenic rats with the YAC vector in which the human alpha-lactalbumin gene was replaced with the hGH gene, tissue specificity of hGH mRNA was the same as that of the endogenous rat alpha-lactalbumin gene. Thus, the 210-kb human alpha-lactalbumin YAC is a useful vector for high-level expression of foreign genes in the milk of transgenic animals.

  16. Prospect of creating transgenic animals by using spermatogonial transplantation

    Institute of Scientific and Technical Information of China (English)

    WU Yingji; LUO Fenhua; BOU Shorgan

    2005-01-01

    Transgenic animal technology is a powerful tool for researching bioscience, biomedicine, bioreactor, and agriculture. There are various ways to produce transgenic animals. The most common ways currently available are pronuclear microinjection and nuclear transfer techniques. However, these methods usually result in low efficiency, causing mosaic (in pronuclear microinjection), or developmental abnormalities (in nuclear transfer). In 1994, Brinster and his colleagues reported an original method to transfer spermatogonial stem cells from donor to recipient mice. The donor spermatogonia were able to form spermatozoa in recipient testes, and to produce progeny carrying the donor's genetic characters. Since then, a series of novel methods were invented by using spermatogonia transplantation. These new methods facilitate the research and application of spermatogonia. Some of these methods, when combining with genetic modification methods, will form a novel methodology for creating transgenic animals. The present paper reviews the achievements of research on spermatogonia transplantation related to creating transgenic animal. Such as, transplantation techniques, cryopreservation of spermatogonia, preparation of recipients, long-term proliferation of spermatogonia in culture, genetic modification of spermatogonia, and characterization of germ line transmission of the modified gene, etc. Furthermore the methodologies for creating transgenic animals by using spermatogonia transplantation were described. Based on the difference between donors and recipients used, the methodology is categorized into two groups: allogeneic transplantation, and autologous transplantation. Although progress in this research area has been swift, potential difficulties remain to be overcome in each approach. The advantages and existing problems in the methodology are discussed.

  17. ChR2 transgenic animals in peripheral sensory system: Sensing light as various sensations.

    Science.gov (United States)

    Ji, Zhi-Gang; Wang, Hongxia

    2016-04-01

    Since the introduction of Channelrhodopsin-2 (ChR2) to neuroscience, optogenetics technology was developed, making it possible to activate specific neurons or circuits with spatial and temporal precision. Various ChR2 transgenic animal models have been generated and are playing important roles in revealing the mechanisms of neural activities, mapping neural circuits, controlling the behaviors of animals as well as exploring new strategy for treating the neurological diseases in both central and peripheral nervous system. An animal including humans senses environments through Aristotle's five senses (sight, hearing, smell, taste and touch). Usually, each sense is associated with a kind of sensory organ (eyes, ears, nose, tongue and skin). Is it possible that one could hear light, smell light, taste light and touch light? When ChR2 is targeted to different peripheral sensory neurons by viral vectors or generating ChR2 transgenic animals, the animals can sense the light as various sensations such as hearing, touch, pain, smell and taste. In this review, we focus on ChR2 transgenic animals in the peripheral nervous system. Firstly the working principle of ChR2 as an optogenetic actuator is simply described. Then the current transgenic animal lines where ChR2 was expressed in peripheral sensory neurons are presented and the findings obtained by these animal models are reviewed.

  18. Highly efficient generation of transgenic sheep by lentivirus accompanying the alteration of methylation status.

    Directory of Open Access Journals (Sweden)

    Chenxi Liu

    Full Text Available BACKGROUND: Low efficiency of gene transfer and silence of transgene expression are the critical factors hampering the development of transgenic livestock. Recently, transfer of recombinant lentivirus has been demonstrated to be an efficient transgene delivery method in various animals. However, the lentiviral transgenesis and the methylation status of transgene in sheep have not been well addressed. METHODOLOGY/PRINCIPLE FINDINGS: EGFP transgenic sheep were generated by injecting recombinant lentivirus into zygotes. Of the 13 lambs born, 8 carried the EGFP transgene, and its chromosomal integration was identified in all tested tissues. Western blotting showed that GFP was expressed in all transgenic founders and their various tissues. Analysis of CpG methylation status of CMV promoter by bisulfate sequencing unraveled remarkable variation of methylation levels in transgenic sheep. The average methylation levels ranged from 37.6% to 79.1% in the transgenic individuals and 34.7% to 83% in the tested tissues. Correlative analysis of methylation status with GFP expression revealed that the GFP expression level was inversely correlated with methylation density. The similar phenomenon was also observed in tested tissues. Transgene integration determined by Southern blotting presented multiple integrants ranging from 2 to 6 copies in the genome of transgenic sheep. CONCLUSIONS/SIGNIFICANCE: Injection of lentiviral transgene into zygotes could be a promising efficient gene delivery system to generate transgenic sheep and achieved widespread transgene expression. The promoter of integrants transferred by lentiviral vector was subjected to dramatic alteration of methylation status and the transgene expression level was inversely correlative with promoter methylation density. Our work illustrated for the first time that generation of transgenic sheep by injecting recombinant lentivirus into zygote could be an efficient tool to improve sheep performance by

  19. Transgenic animal bioreactors in biotechnology and production of blood proteins.

    Science.gov (United States)

    Lubon, H

    1998-01-01

    The regulatory elements of genes used to target the tissue-specific expression of heterologous human proteins have been studied in vitro and in transgenic mice. Hybrid genes exhibiting the desired performance have been introduced into large animals. Complex proteins like protein C, factor IX, factor VIII, fibrinogen and hemoglobin, in addition to simpler proteins like alpha 1-antitrypsin, antithrombin III, albumin and tissue plasminogen activator have been produced in transgenic livestock. The amount of functional protein secreted when the transgene is expressed at high levels may be limited by the required posttranslational modifications in host tissues. This can be overcome by engineering the transgenic bioreactor to express the appropriate modifying enzymes. Genetically engineered livestock are thus rapidly becoming a choice for the production of recombinant human blood proteins.

  20. [Generation of transgenic mice expressing human lysozyme in mammary gland].

    Science.gov (United States)

    Yan, Hua; Li, Guo-cai; Sun, Huai-chang

    2005-10-01

    To evaluate the feasibility of generating animal mammary gland bioreactors expressing human lysozyme (hLYZ). The recombinant vector p205C3-hLYZ, as a result of connecting the hLYZ cDNA with the mammry gland expression vector p205C3, was used to generate transfer genic mice by microinjection. A total of 136 F0 mice were obtained, of which 7 (2 females and 5 males) and 4 (1 females and 3 males) were found to contain the transfer-gene by PCR and Southern blotting respectively. The results of Western blotting indicated that the expressed protein had the same molecular weight as that of normal hLYZ. From the F1 generation on, the mice mated only with their brothers or sisters and a colony of F7 transgenic mice was obtained. Among the offspring, the female transgenic mice maintained and expressed the transfer-gene stably with an expression level as high as 750 mg/L. The expressed protein had strong tissue specificity, and in addition to the mammary glands, some degree of ectropic expression in the spleens and intestines of the transgenic mice was confirmed by dot blotting assay. These data indicate that the mice mammary gland bioreactors expressing hLYZ have been successfully generated.

  1. Generating a transgenic mouse line stably expressing human MHC surface antigen from a HAC carrying multiple genomic BACs.

    Science.gov (United States)

    Hasegawa, Yoshinori; Ishikura, Tomoyuki; Hasegawa, Takanori; Watanabe, Takashi; Suzuki, Junpei; Nakayama, Manabu; Okamura, Yoshiaki; Okazaki, Tuneko; Koseki, Haruhiko; Ohara, Osamu; Ikeno, Masashi; Masumoto, Hiroshi

    2015-03-01

    The human artificial chromosome (HAC) vector is a promising tool to improve the problematic suppression and position effects of transgene expression frequently seen in transgenic cells and animals produced by conventional plasmid or viral vectors. We generated transgenic mice maintaining a single HAC vector carrying two genomic bacterial artificial chromosomes (BACs) from human HLA-DR loci (DRA and DRB1). Both transgenes on the HAC in transgenic mice exhibited tissue-specific expression in kidney, liver, lung, spleen, lymph node, bone marrow, and thymus cells in RT-PCR analysis. Stable functional expression of a cell surface HLA-DR marker from both transgenes, DRA and DRB1 on the HAC, was detected by flow cytometric analysis of splenocytes and maintained through at least eight filial generations. These results indicate that the de novo HAC system can allow us to manipulate multiple BAC transgenes with coordinated expression as a surface antigen through the generation of transgenic animals.

  2. Producing proteins in transgenic plants and animals.

    Science.gov (United States)

    Larrick, J W; Thomas, D W

    2001-08-01

    The requirement for large quantities of therapeutic proteins has fueled interest in the production of recombinant proteins in plants and animals. The first commercial products to be made in this way have experienced much success, and it is predicted that in the future a plethora of protein products will be made using these 'natural' bioreactors.

  3. Oral immunization of animals with transgenic cherry tomatillo expressing HBsAg

    Institute of Scientific and Technical Information of China (English)

    Yi Gao; Yina Ma; Mei Li; Tonq Chenq; Shao-Wei Li; Jun Zhang; Ning-Shao Xia

    2003-01-01

    AIM: To investigate the expression of recombinant HBsAg (rHBsAg) in transgenic cherry tomatillo in order to explore the feasibility of producing HBV oral vaccine with cherry tomatillo by animal immune tests.METHODS: The recombinant plant expression vector containing HBsAg gene was constructed. Mediated with Agrobacterium tumefaciens, HBsAg gene was transferred into cotyledons of cherry tomatillo. Transformed cherry tomatillos were obtained through hygromycin delay-selection. Integrated DNA in transgenic cherry tomatillo was confirmed by hygromycin resistance selection, Gus detection, polymerase chain reaction (PCR) and dot blotting analysis. Antigenicity of rHBsAg was examined by ELISA and the immunogenicity of rHBsAg derived from transgenic cherry tomatillo tissues was confirmed by oral feed of transformed tissues to BALB/c mice primed with commercial HBV vaccines. Specific antibody titers in mice's serum were examined by ELISA every week.RESULTS: By far, 10 positive lines of transgenic cherry tomatillos containing HBsAg gene were obtained. Among different organs of the same transgenic cherry tomatillo,level of rHBsAg expressed in leaves was the highest with the yield up to 300ng/g fresh weight. And the rHBsAg expression level in fruits was about 10 ng/g fresh weight.In animal immune tests, oral delivery with transgenic tissues to mice primed with commercial vaccine instead of naive mice resulted in significant immune response.CONCLUSION: The result of this animal immune test indicated the rHBsAg derived from transgenic cherry tomatillo possessed normal immunogenicity. This work demonstrated the feasibility to generate oral immunogenic rHBsAg in transgenic cherry tomatillo, and would provide some experimental approach for the production of low-cost oral vaccines using transgenic cherry tomatillo in large scale.

  4. Effective generation of transgenic pigs and mice by linker based sperm-mediated gene transfer.

    Directory of Open Access Journals (Sweden)

    Shih Ping Yao

    2002-04-01

    Full Text Available Abstract Background Transgenic animals have become valuable tools for both research and applied purposes. The current method of gene transfer, microinjection, which is widely used in transgenic mouse production, has only had limited success in producing transgenic animals of larger or higher species. Here, we report a linker based sperm-mediated gene transfer method (LB-SMGT that greatly improves the production efficiency of large transgenic animals. Results The linker protein, a monoclonal antibody (mAb C, is reactive to a surface antigen on sperm of all tested species including pig, mouse, chicken, cow, goat, sheep, and human. mAb C is a basic protein that binds to DNA through ionic interaction allowing exogenous DNA to be linked specifically to sperm. After fertilization of the egg, the DNA is shown to be successfully integrated into the genome of viable pig and mouse offspring with germ-line transfer to the F1 generation at a highly efficient rate: 37.5% of pigs and 33% of mice. The integration is demonstrated again by FISH analysis and F2 transmission in pigs. Furthermore, expression of the transgene is demonstrated in 61% (35/57 of transgenic pigs (F0 generation. Conclusions Our data suggests that LB-SMGT could be used to generate transgenic animals efficiently in many different species.

  5. Sperm cells as vectors in the production of transgenic animals

    Energy Technology Data Exchange (ETDEWEB)

    Prince, R.M.

    1993-04-28

    Transgenic animals are used in industry and in biomedical research in order to provide in vivo experimental model systems. Sperm cells have been reported used as vectors in the production of transgenic animals before, however no approach has of yet proven to be successful. Fertilizing eggs with genetically modified sperm would be advantageous in that sperm are readily accessible and stable, and eggs can be fertilized by modified sperm cells in vivo. Recent elucidations regarding the unique manner of DNA packaging in sperm chromatin by protamines has provided us with the insight for developing a method of introducing foreign DNA into sperm which is likely to succeed where others have failed. We have developed a method for mimicking the in vivo system of sperm chromatin toroid subunits in vitro, concentrating these toroids, and fluorescent visualization. Our present work concerns development of a method to successfully deliver DNA across the cell membranes and into the nucleus.

  6. TBASE: a computerized database for transgenic animals and targeted mutations.

    Science.gov (United States)

    Woychik, R P; Wassom, J S; Kingsbury, D; Jacobson, D A

    1993-05-27

    A computerized database, called TBASE, has been developed to organize and make available information on transgenic animals and targeted mutations by using resources at the Oak Ridge National Laboratory (ORNL) and Johns Hopkins University (JHU). The database is available through the JHU Computational Biology Gopher Server. To ensure that all interested users have access, several mechanisms will be installed to accommodate varying levels of telecommunication network connectivity.

  7. Transgenic animal models for study of the pathogenesis of Huntington's disease and therapy.

    Science.gov (United States)

    Chang, Renbao; Liu, Xudong; Li, Shihua; Li, Xiao-Jiang

    2015-01-01

    Huntington's disease (HD) is caused by a genetic mutation that results in polyglutamine expansion in the N-terminal regions of huntingtin. As a result, this polyQ expansion leads to the misfolding and aggregation of mutant huntingtin as well as age-dependent neurodegeneration. The genetic mutation in HD allows for generating a variety of animal models that express different forms of mutant huntingtin and show differential pathology. Studies of these animal models have provided an important insight into the pathogenesis of HD. Mouse models of HD include transgenic mice, which express N-terminal or full-length mutant huntingtin ubiquitously or selectively in different cell types, and knock-in mice that express full-length mutant Htt at the endogenous level. Large animals, such as pig, sheep, and monkeys, have also been used to generate animal HD models. This review focuses on the different features of commonly used transgenic HD mouse models as well as transgenic large animal models of HD, and also discusses how to use them to identify potential therapeutics. Since HD shares many pathological features with other neurodegenerative diseases, identification of therapies for HD would also help to develop effective treatment for different neurodegenerative diseases that are also caused by protein misfolding and occur in an age-dependent manner.

  8. A protocol for generation of transgenic mice by manipulating spermatogonial stem cells in vivo.

    OpenAIRE

    sprotocols

    2015-01-01

    Authors: Lalit Sehgal, Rahul Thorat, Nileema Khapare, Amitabha Mukhopadhaya, Mugdha Sawant & Sorab Dalal ### Abstract This protocol describes a technique for the generation of transgenic mice by in-vivo manipulation of spermatogonial stem cells (SSCs) with a high rate of success. In this study SSCs in pre-pubescent animals were infected in vivo with recombinant lentiviruses expressing EGFP-f and mated with normal females. All male pre-founder mice produced transgenic pups with an ...

  9. Robust generation of transgenic mice by simple hypotonic solution mediated delivery of transgene in testicular germ cells

    Science.gov (United States)

    Usmani, Abul; Ganguli, Nirmalya; Jain, Subodh K; Ganguli, Nilanjana; Sarkar, Rajesh Kumar; Choubey, Mayank; Shukla, Mansi; Sarkar, Hironmoy; Majumdar, Subeer S

    2016-01-01

    Our ability to decipher gene sequences has increased enormously with the advent of modern sequencing tools, but the ability to divulge functions of new genes have not increased correspondingly. This has caused a remarkable delay in functional interpretation of several newly found genes in tissue and age specific manner, limiting the pace of biological research. This is mainly due to lack of advancements in methodological tools for transgenesis. Predominantly practiced method of transgenesis by pronuclear DNA-microinjection is time consuming, tedious, and requires highly skilled persons for embryo-manipulation. Testicular electroporation mediated transgenesis requires use of electric current to testis. To this end, we have now developed an innovative technique for making transgenic mice by giving hypotonic shock to male germ cells for the gene delivery. Desired transgene was suspended in hypotonic Tris-HCl solution (pH 7.0) and simply injected in testis. This resulted in internalization of the transgene in dividing germ-cells residing at basal compartment of tubules leading to its integration in native genome of mice. Such males generated transgenic progeny by natural mating. Several transgenic animals can be generated with minimum skill within short span of time by this easily adaptable novel technique. PMID:27933305

  10. Transgenic animal models for study of the pathogenesis of Huntington’s disease and therapy

    Directory of Open Access Journals (Sweden)

    Chang RB

    2015-04-01

    Full Text Available Renbao Chang,1 Xudong Liu,1 Shihua Li,2 Xiao-Jiang Li1,2 1State Key Laboratory of Molecular Developmental Biology, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, People’s Republic of China; 2Department of Human Genetics, Emory University School of Medicine, Atlanta, GA, USA Abstract: Huntington’s disease (HD is caused by a genetic mutation that results in polyglutamine expansion in the N-terminal regions of huntingtin. As a result, this polyQ expansion leads to the misfolding and aggregation of mutant huntingtin as well as age-dependent neurodegeneration. The genetic mutation in HD allows for generating a variety of animal models that express different forms of mutant huntingtin and show differential pathology. Studies of these animal models have provided an important insight into the pathogenesis of HD. Mouse models of HD include transgenic mice, which express N-terminal or full-length mutant huntingtin ubiquitously or selectively in different cell types, and knock-in mice that express full-length mutant Htt at the endogenous level. Large animals, such as pig, sheep, and monkeys, have also been used to generate animal HD models. This review focuses on the different features of commonly used transgenic HD mouse models as well as transgenic large animal models of HD, and also discusses how to use them to identify potential therapeutics. Since HD shares many pathological features with other neurodegenerative diseases, identification of therapies for HD would also help to develop effective treatment for different neurodegenerative diseases that are also caused by protein misfolding and occur in an age-dependent manner. Keywords: transgenic animal models, Huntington’s disease, pathogenesis, therapy

  11. Biosafety assessment of transgenic Bt cotton on model animals

    Directory of Open Access Journals (Sweden)

    Sadia Bano

    2016-05-01

    Full Text Available Abstract Background: To know the effects of transgenic crops on soil microorganisms, animals and other expected hazards due to the introduction of GM crops into the environment is critical both scientifically and environmentally. The work was conducted to study the effect of insecticidal Bt protein on Rats and Earthworms. Methods: For this purpose, animals like rat and soil organisms like Earthworm were selected. Rats were selected on the basis of its 95% homology on genomic, cellular and enzymatic level with human while earthworm were preferred on the basis of their direct contact with soil to evaluate the impact of Bt (Cry1AC crop field soil on earthworm, secreted by root exudates of Bt cotton. Several physical, molecular, biochemical and histological analyses were performed on both Rats/Earthworms fed on standard diet (control group as well containing Bt protein (experimental group. Results: Molecular analyses such as immune Dot blot, SDS-PAGE, ELISA and PCR, confirmed the absence of Cry1Ac protein in blood and urine samples of rats, which were fed with Bt protein in their diet. Furthermore, histological studies showed that there was no difference in cellular architecture in liver, heart, kidney and intestine of Bt and non-Bt diet fed rats. To see the effect of Bt on earthworm two different groups were studied, one with transgenic plant field soil supplemented with grinded leaves of cotton and second group with non-Bt field soil. Conclusions: No lethal effects of transgenic Bt protein on the survival of earthworm and rats were observed. Bradford assay, Dipstick assay ELISA demonstrated the absence of Cry1Ac protein in the mid-gut epithelial tissue of earthworm. The results of present study will be helpful in successful deployment and commercial release of genetically modified crop in Pakistan.

  12. [Transgenic animals as bioreactors in the service of the pharmaceutical industry].

    Science.gov (United States)

    Honscha, W

    1999-10-01

    A modern pharmacotherapy without the use of proteins as drugs is not more practicable. In the clinic blood coagulation factors (factor VIII and IX), growth hormones (human growth hormone), enzymes (1-antitrysin, insulin), and cytokines are currently used. At the beginning, the proteins were isolated from biological sources or expressed in vitro by the use of E. coli or eukaryotic cell lines. At the moment efforts were undertaken to express these proteins in the milk of transgenic animals. This review article describes the methods for the generation of transgenic animals, the benefits and drawbacks of this new technique in comparison to established methods for the production of proteins as pharmaceuticals. At the end of the review possible improvements of the method are described.

  13. Molecular characterization of transgene integration by next-generation sequencing in transgenic cattle.

    Directory of Open Access Journals (Sweden)

    Ran Zhang

    Full Text Available As the number of transgenic livestock increases, reliable detection and molecular characterization of transgene integration sites and copy number are crucial not only for interpreting the relationship between the integration site and the specific phenotype but also for commercial and economic demands. However, the ability of conventional PCR techniques to detect incomplete and multiple integration events is limited, making it technically challenging to characterize transgenes. Next-generation sequencing has enabled cost-effective, routine and widespread high-throughput genomic analysis. Here, we demonstrate the use of next-generation sequencing to extensively characterize cattle harboring a 150-kb human lactoferrin transgene that was initially analyzed by chromosome walking without success. Using this approach, the sites upstream and downstream of the target gene integration site in the host genome were identified at the single nucleotide level. The sequencing result was verified by event-specific PCR for the integration sites and FISH for the chromosomal location. Sequencing depth analysis revealed that multiple copies of the incomplete target gene and the vector backbone were present in the host genome. Upon integration, complex recombination was also observed between the target gene and the vector backbone. These findings indicate that next-generation sequencing is a reliable and accurate approach for the molecular characterization of the transgene sequence, integration sites and copy number in transgenic species.

  14. Animales transgénicos: pasado, presente y futuro Transgenic animals: past, present and future

    Directory of Open Access Journals (Sweden)

    R. Felmer

    2004-12-01

    Full Text Available Desde la demostración del primer animal transgénico en 1980, la ingeniería genética ha revolucionado todos los aspectos de la investigación biológica y biomédica. Desde entonces, se ha logrado la generación de varios tipos de animales transgénicos, incluyendo vacas, cerdos, ovejas, cabras y conejos. Hasta muy recientemente, la generación de animales transgénicos involucraba la microinyección de pequeñas cantidades de ADN en el pronúcleo de un embrión al estado de dos células, técnica conocida como microinyección pronuclear. El descubrimiento de que los animales podrían ser clonados mediante transferencia nuclear de células mantenidas en cultivo abrió las puertas para realizar recombinación homóloga en estas especies. Esto podría tener importantes implicaciones especialmente para los propósitos del descubrimiento de nuevas drogas, para mejorar características productivas de los animales, en la clonación de cerdos como fuentes de órganos para trasplantes y en la producción de proteínas farmacéuticas. En esta revisión se discuten los avances que ha tenido la tecnología para la generación de animales transgénicos y los beneficios de la transferencia nuclear como una nueva ruta para la generación de animales transgénicos de granja.Since the initial demonstration in 1980 that a transgenic animal could be generated harbouring a transgene from a different species, genetic engineering has revolutionized all aspects of fundamental biological and biomedical research. Since then, much has been accomplished in the generation of various types of transgenic animals, including cows, pigs, sheeps, goats and rabbits. Until recently, genetically modified livestock could only be generated by pronuclear injection. The discovery that animals can be cloned by nuclear transfer from cultured somatic cells means that it is now possible to achieve gene targeting in these species. This may have important implications for the purposes of

  15. The use of transgenic animals to study lipoprotein metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Rubin, E.M.; Plump, A.S.

    1993-12-01

    The application of transgenic technology to lipoprotein metabolism and atherosclerosis was first reported in 1988. Today, a large percentage of the genes involved in lipoprotein metabolism have been overexpressed in mice, and a substantial number of these same genes have been disrupted by homologous recombination in embryonic stem (ES) cells. The utility of animal models of lipoprotein metabolism and atherosclerosis is far-reaching given the complex nature of these systems. There are at least 17 known genes directly involved in lipoprotein metabolism and likely dozens more may be involved. This massive network of interacting factors has necessitated the development of in vivo systems which can be subject to genetic manipulation. The power of overexpression is obvious: elucidating function in a relatively controlled genetic environment in which the whole system is present and operational. The not-so-obvious problem with transgenics is ``background,`` or for purposes of the current discussion, the mouse`s own lipoprotein system. With the advent of gene knockout, we have been given the ability to overcome ``background.`` By recreating the genetic complement of the mouse we can alter a system in essentially any manner desired. As unique tools, and in combination with one another, the overexpression of foreign genes and the targeted disruption or alteration of endogenous genes has already and will continue to offer a wealth of information on the biology of lipoprotein metabolism and its effect on atherosclerosis susceptibility.

  16. Generation and characterization of human heme oxygenase-1 transgenic pigs.

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    Hye-Jung Yeom

    Full Text Available Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1, an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs. Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation.

  17. Generation and characterization of human heme oxygenase-1 transgenic pigs.

    Science.gov (United States)

    Yeom, Hye-Jung; Koo, Ok Jae; Yang, Jaeseok; Cho, Bumrae; Hwang, Jong-Ik; Park, Sol Ji; Hurh, Sunghoon; Kim, Hwajung; Lee, Eun Mi; Ro, Han; Kang, Jung Taek; Kim, Su Jin; Won, Jae-Kyung; O'Connell, Philip J; Kim, Hyunil; Surh, Charles D; Lee, Byeong-Chun; Ahn, Curie

    2012-01-01

    Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1), an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs). Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation.

  18. Cloned animal and transgenics; Kuron gijutsu to dobutsu kojo

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, S. [National Inst. of Animal Industry, Ibaraki (Japan)

    1998-10-10

    Somatic cell cloning technologies are explained, and their use in the manufacture of biologically active substances such as pharmaceuticals is described. In general a cloning technology means a technology of nucleus transfer in which the nucleus of an ovum is removed and the nucleus of another cell is transferred. Since a mammalian ovum is but 100-150{mu}m in diameter, the removal or transfer of a nucleus has to be performed under an inverted microscope provided with a micromanipulator. In 1986, this method was employed for the transfer of a cell nucleus at its early division stage in a sheep embryo, and a sheep was cloned. The production of useful substances such as pharmaceuticals is performed through gene recombination for coli bacteria, yeast fungi, and cultured mammalian cells, but there are some problems to be solved about the amount produced, cost of production, biological activity, etc. Under such circumstances, a concept of animal factory has been invented in which Tg (transgenic) animals themselves are utilized as bioreactors, and several venture organizations have already been incorporated. 3 refs., 1 fig., 1 tab.

  19. Generation of transgenic Wuzhishan miniature pigs expressing monomeric red fluorescent protein by somatic cell nuclear transfer.

    Science.gov (United States)

    Lu, Yue; Kang, Jin-Dan; Li, Suo; Wang, Wei; Jin, Jun-Xue; Hong, Yu; Cui, Cheng-du; Yan, Chang-Guo; Yin, Xi-Jun

    2013-08-01

    Red fluorescent protein and its variants enable researchers to study gene expression, localization, and protein-protein interactions in vitro in real-time. Fluorophores with higher wavelengths are usually preferred since they efficiently penetrate tissues and produce less toxic emissions. A recently developed fluorescent protein marker, monomeric red fluorescent protein (mRFP1), is particularly useful because of its rapid maturation and minimal interference with green fluorescent protein (GFP) and GFP-derived markers. We generated a pCX-mRFP1-pgk-neoR construct and evaluated the ability of mRFP1 to function as a fluorescent marker in transgenic Wuzhishan miniature pigs. Transgenic embryos were generated by somatic cell nuclear transfer (SCNT) of nuclei isolated from ear fibroblasts expressing mRFP1. Embryos generated by SCNT developed into blastocysts in vitro (11.65%; 31/266). Thereafter, a total of 685 transgenic embryos were transferred into the oviducts of three recipients, two of which became pregnant. Of these, one recipient had six aborted fetuses, whereas the other recipient gave birth to four offspring. All offspring expressed the pCX-mRFP1-pgk-neoR gene as shown by PCR and fluorescence in situ hybridization analysis. The transgenic pigs expressed mRFP1 in all organs and tissues at high levels. These results demonstrate that Wuzhishan miniature pigs can express mRFP1. To conclude, this transgenic animal represents an excellent model with widespread applications in medicine and agriculture.

  20. Every silver lining has a cloud: the scientific and animal welfare issues surrounding a new approach to the production of transgenic animals.

    Science.gov (United States)

    Combes, Robert D; Balls, Michael

    2014-05-01

    The scientific basis and advantages of using recently developed CRISPR/Cas-9 technology for transgenesis have been assessed with respect to other production methods, laboratory animal welfare, and the scientific relevance of transgenic models of human diseases in general. As the new technology is straightforward, causes targeted DNA double strand breaks and can result in homozygous changes in a single step, it is more accurate and more efficient than other production methods and speeds up transgenesis. CRISPR/Cas-9 also obviates the use of embryonic stem cells, and is being used to generate transgenic non-human primates (NHPs). While the use of this method reduces the level of animal wastage resulting from the production of each new strain, any long-term contribution to reduction will be offset by the overall increase in the numbers of transgenic animals likely to result from its widespread usage. Likewise, the contribution to refinement of using a more-precise technique, thereby minimising the occurrence of unwanted genetic effects, will be countered by a probable substantial increase in the production of transgenic strains of increasingly sentient species. For ethical and welfare reasons, we believe that the generation of transgenic NHPs should be allowed only in extremely exceptional circumstances. In addition, we present information, which, on both welfare and scientific grounds, leads us to question the current policy of generating ever-more new transgenic models in light of the general failure of many of them, after over two decades of ubiquitous use, to result in significant advances in the understanding and treatment of many key human diseases. Because this unsatisfactory situation is likely to be due to inherent, as well as possibly avoidable, limitations in the transgenic approach to studying disease, which are briefly reviewed, it is concluded that a thorough reappraisal of the rationale for using genetically-altered animals in fundamental research and

  1. Transgenic bioreactors.

    Science.gov (United States)

    Jänne, J; Alhonen, L; Hyttinen, J M; Peura, T; Tolvanen, M; Korhonen, V P

    1998-01-01

    Since the generation of the first transgenic mice in 1980, transgene technology has also been successfully applied to large farm animals. Although this technology can be employed to improve certain production traits of livestock, this approach has not been very successful so far owing to unwanted effects encountered in the production animals. However, by using tissue-specific targeting of the transgene expression, it is possible to produce heterologous proteins in the extracellular space of large transgenic farm animals. Even though some recombinant proteins, such as human hemoglobin, have been produced in the blood of transgenic pigs, in the majority of the cases mammary gland targeted expression of the transgene has been employed. Using production genes driven by regulatory sequences of milk protein genes a number of valuable therapeutic proteins have been produced in the milk of transgenic bioreactors, ranging from rabbits to dairy cattle. Unlike bacterial fermentors, the mammary gland of transgenic bioreactors appear to carry out proper postsynthetic modifications of human proteins required for full biological activity. In comparison with mammalian cell bioreactors, transgenic livestock with mammary gland targeted expression seems to be able to produce valuable human therapeutic proteins at very low cost. Although not one transgenically produced therapeutic protein is yet on the market, the first such proteins have recently entered or even completed clinical trials required for their approval.

  2. [Effects of transgenic Bt crops on non-target soil animals].

    Science.gov (United States)

    Yuan, Yi-gang; Ge, Feng

    2010-05-01

    Transgenic Bt crops are widely planted around the world. With the quick development and extension of genetically modified crops, it is needed to make a deep study on the effects of Bt crops on soil ecosystem. This paper reviewed the research progress on the effects of transgenic Bt crops on the population dynamics and community structure of soil animals, e.g., earthworm, nematode, springtail, mite, and beetle, etc. The development history of Bt crops was introduced, the passway the Bt protein comes into soil as well as the residual and degradation of Bt protein in soil were analyzed, and the critical research fields about the ecological risk analysis of transgenic Bt crops on non-target soil animals in the future were approached, which would provide a reference for the research of the effects of transgenic Bt crops on non-target soil animals.

  3. Generation and characterization of a transgenic pig carrying a DsRed-monomer reporter gene.

    Directory of Open Access Journals (Sweden)

    Chih-Jen Chou

    Full Text Available Pigs are an optimal animal for conducting biomedical research because of their anatomical and physiological resemblance to humans. In contrast to the abundant resources available in the study of mice, few fluorescent protein-harboring porcine models are available for preclinical studies. In this paper, we report the successful generation and characterization of a transgenic DsRed-Monomer porcine model.The transgene comprised a CMV enhancer/chicken-beta actin promoter and DsRed monomeric cDNA. Transgenic pigs were produced by using pronuclear microinjection. PCR and Southern blot analyses were applied for identification of the transgene. Histology, blood examinations and computed tomography were performed to study the health conditions. The pig amniotic fluid progenitor/stem cells were also isolated to examine the existence of red fluorescence and differentiation ability.Transgenic pigs were successfully generated and transmitted to offspring at a germ-line transmission rate of 43.59% (17/39. Ubiquitous expression of red fluorescence was detected in the brain, eye, tongue, heart, lung, liver, pancreas, spleen, stomach, small intestine, large intestine, kidney, testis, and muscle; this was confirmed by histology and western blot analyses. In addition, we confirmed the differentiation potential of amniotic fluid progenitor stem cells isolated from the transgenic pig.This red fluorescent pig can serve as a host for other fluorescent-labeled cells in order to study cell-microenvironment interactions, and can provide optimal red-fluorescent-labeled cells and tissues for research in developmental biology, regenerative medicine, and xenotransplantation.

  4. Role of papillomavirus oncogenes in human cervical cancer: Transgenic animal studies

    Energy Technology Data Exchange (ETDEWEB)

    Griep, A.E.; Lambert, P.F. [Univ. of Wisconsin School of Medicine, Madison, WI (United States)

    1994-05-01

    Human papillomaviruses are believed to be etiologic agents for the majority of human cervical carcinoma, a common cancer that is a leading cause of death by cancer among women worldwide. In cervical carcinoma, a subset of papillomaviral genes, namely E6 and E7, are expressed. In vitro tissue culture studies indicate that HPV E6 and E7 are oncogenes, and that their oncogenicity is due in part to their capacity to inactivate cellular tumor suppressor genes. The behavior of E6 and E7 in vitro and the genetic evidence from analysis of human cancers suggest that the E6 and E7 genes play a significant role in the development of cervical cancer. This hypothesis is now being tested using animal models. In this review, we summarize our current knowledge of the oncogenicity of papillomavirus genes that has been generated through their study in transgenic mice. 82 refs., 4 figs., 1 tab.

  5. Chinese Experts Successfully Produced Transgenic Animals from Haploid Embryonic Stem Cells

    Institute of Scientific and Technical Information of China (English)

    2012-01-01

    Individual animals produced by haploid stem cells are ideal models for studying recessive genes. Hap- loid stem cells not only can maintain haploidy, but also are capable of replicating themselves infinitely. Modified genes can be passed on to future generations through genetic engineering of haploid embryonic stem cells, which thus avoids the germlinechimerism caused by other transgenic methods and greatly im- proves the analysis efficiency of the function of gene modification. However, natural haploids are only re- stricted to germline cells in mammals. Currently in mammals, only the embryonic stem cells in rats and mice can be used as the carrier of gene modification, but the embryonic stem cells of other mammals, in- eluding primates, cannot guarantee germline transmission, which has seriously hindered the establishment of disease models by using these species.

  6. Animal feed compositions containing phytase derived from transgenic alfalfa and methods of use thereof

    Science.gov (United States)

    Austin-Phillips, Sandra; Koegel, Richard G.; Straub, Richard J.; Cook, Mark

    1999-01-01

    A value-added composition of matter containing plant matter from transgenic alfalfa which expresses exogenous phytase activity is disclosed. The phytase activity is a gene product of an exogenous gene encoding for phytase which has been stably incorporated into the genome of alfalfa plants. The transgenic alfalfa expresses phytase activity in nutritionally-significant amounts, thereby enabling its use in animal feeds to eliminate the need for phosphorous supplementation of livestock, poultry, and fish feed rations.

  7. Animal feed compositions containing phytase derived from transgenic alfalfa and methods of use thereof

    Energy Technology Data Exchange (ETDEWEB)

    Austin-Phillips, Sandra (Madison, WI); Koegel, Richard G. (Madison, WI); Straub, Richard J. (Brooklyn, WI); Cook, Mark (Madison, WI)

    2001-01-01

    A value-added composition of matter containing plant matter from transgenic alfalfa which expresses exogenous phytase activity is disclosed. The phytase activity is a gene product of an exogenous gene encoding for phytase which has been stably incorporated into the genome of alfalfa plants. The transgenic alfalfa expresses phytase activity in nutritionally-significant amounts, thereby enabling its use in animal feeds to eliminate the need for phosphorous supplementation of livestock, poultry, and fish feed rations.

  8. Tissue-specific posttranscriptional downregulation of expression of the S100A4(mts1) gene in transgenic animals

    DEFF Research Database (Denmark)

    Ambartsumian, N; Klingelhöfer, Jörg; Grigorian, M

    1998-01-01

    The S100A4(mts1) is a gene associated with generation of metastatic disease. In order to analyze the consequences of alteration of the pattern of expression of the S100A4(mts1) gene we obtained strains of transgenic mice bearing the S100A4(mts1) gene under the control of a ubiquitous...... and constitutive 3-hydroxy-3-methylglutaryl CoA reductase (HMGCR) gene promoter. In transgenic animals the expression of the transgene RNA was detected in all organs, but only some of the organs showed elevated levels of the protein. Expression of the S100A4(Mts1) protein was downregulated in the organs...... that normally do not express the gene in the wild-type animal. The transgene RNA is detected in the polysomes indicating that it could be translated into the S100A4(Mts1) protein. The specificity of the S100A4(Mts1) protein expression is determined by a complex mechanism including regulation of translation and...

  9. Rapid cohort generation and analysis of disease spectrum of large animal model of cone dystrophy.

    Directory of Open Access Journals (Sweden)

    Corinne Kostic

    Full Text Available Large animal models are an important resource for the understanding of human disease and for evaluating the applicability of new therapies to human patients. For many diseases, such as cone dystrophy, research effort is hampered by the lack of such models. Lentiviral transgenesis is a methodology broadly applicable to animals from many different species. When conjugated to the expression of a dominant mutant protein, this technology offers an attractive approach to generate new large animal models in a heterogeneous background. We adopted this strategy to mimic the phenotype diversity encounter in humans and generate a cohort of pigs for cone dystrophy by expressing a dominant mutant allele of the guanylate cyclase 2D (GUCY2D gene. Sixty percent of the piglets were transgenic, with mutant GUCY2D mRNA detected in the retina of all animals tested. Functional impairment of vision was observed among the transgenic pigs at 3 months of age, with a follow-up at 1 year indicating a subsequent slower progression of phenotype. Abnormal retina morphology, notably among the cone photoreceptor cell population, was observed exclusively amongst the transgenic animals. Of particular note, these transgenic animals were characterized by a range in the severity of the phenotype, reflecting the human clinical situation. We demonstrate that a transgenic approach using lentiviral vectors offers a powerful tool for large animal model development. Not only is the efficiency of transgenesis higher than conventional transgenic methodology but this technique also produces a heterogeneous cohort of transgenic animals that mimics the genetic variation encountered in human patients.

  10. Transgenesis and animal welfare : implications of transgenic procedures for the well-being of the laboratory mice

    NARCIS (Netherlands)

    Meer, Miriam van der

    2002-01-01

    Transgenic animals play an important role in biomedical research. Their use as animal model is still increasing. Although the process of transgenesis may contribute to refinement of animal use, the application of the biotechnological procedures that are involved in the production of transgenic anima

  11. GPU Generation of Large Varied Animated Crowds

    OpenAIRE

    Isaac Rudomin; Benjamín Hernández; Oriam de Gyves; Leonel Toledo; Ivan Rivalcoba; Sergio Ruiz

    2013-01-01

    ..We discuss several steps in the process of simulating and visualizing large and varied crowds in real time for consumer-level computers and graphic cards (GPUs). Animating varied crowds using a diversity of models and animations (assets) is complex and costly. One has to use models that are expensive if bought, take a long time to model, and consume too much memory and computing resources. We discuss methods for simulating, generating, animating and rendering crowds of varied aspect and a d...

  12. Generation of transgenic cattle expressing human β-defensin 3 as an approach to reducing susceptibility to Mycobacterium bovis infection.

    Science.gov (United States)

    Su, Feng; Wang, Yongsheng; Liu, Guanghui; Ru, Kun; Liu, Xin; Yu, Yuan; Liu, Jun; Wu, Yongyan; Quan, Fusheng; Guo, Zekun; Zhang, Yong

    2016-03-01

    Bovine tuberculosis results from infection with Mycobacterium bovis, a member of the Mycobacterium tuberculosis family. Worldwide, M. bovis infections result in economic losses in the livestock industry; cattle production is especially hard-hit by this disease. Generating M. bovis-resistant cattle may potentially mitigate the impact of this disease by reducing M. bovis infections. In this study, we used transgenic somatic cell nuclear transfer to generate cattle expressing the gene encoding human β-defensin 3 (HBD3), which confers resistance to mycobacteria in vitro. We first generated alveolar epithelial cells expressing HBD3 under the control of the bovine MUC1 promoter, and confirmed that these cells secreted HBD3 and possessed anti-mycobacterial capacity. We then generated and identified transgenic cattle by somatic cell nuclear transfer. The cleavage and blastocyst formation rates of genetically modified embryos provided evidence that monoclonal transgenic bovine fetal fibroblast cells have an integral reprogramming ability that is similar to that of normal cells. Five genetically modified cows were generated, and their anti-mycobacterial capacities were evaluated. Alveolar epithelial cells and macrophages from these cattle expressed higher levels of HBD3 protein compared with non-transgenic cells and possessed effective anti-mycobacterial capacity. These results suggest that the overall risk of M. bovis infection in transgenic cattle is efficiently reduced, and support the development of genetically modified animals as an effective tool to reduce M. bovis infection.

  13. Novel method of generating non-human transgenic animals, and transgenic animals thus obtained

    OpenAIRE

    Ventura Oliveira, Pedro Nuno; GUTIÉRREZ-ADÁN, Alfonso; Montoliu, Lluís

    2005-01-01

    Fecha de presentación internacional: 04.04.2005. - Titulares: Consejo Superior de Investigaciones Científicas (CSIC). - Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (INIA)

  14. Generation of doubled haploid transgenic wheat lines by microspore transformation.

    Science.gov (United States)

    Brew-Appiah, Rhoda A T; Ankrah, Nii; Liu, Weiguo; Konzak, Calvin F; von Wettstein, Diter; Rustgi, Sachin

    2013-01-01

    Microspores can be induced to develop homozygous doubled haploid plants in a single generation. In the present experiments androgenic microspores of wheat have been genetically transformed and developed into mature homozygous transgenic plants. Two different transformation techniques were investigated, one employing electroporation and the other co-cultivation with Agrobacterium tumefaciens. Different tissue culture and transfection conditions were tested on nine different wheat cultivars using four different constructs. A total of 19 fertile transformants in five genotypes from four market classes of common wheat were recovered by the two procedures. PCR followed by DNA sequencing of the products, Southern blot analyses and bio/histo-chemical and histological assays of the recombinant enzymes confirmed the presence of the transgenes in the T0 transformants and their stable inheritance in homozygous T1∶2 doubled haploid progenies. Several decisive factors determining the transformation and regeneration efficiency with the two procedures were determined: (i) pretreatment of immature spikes with CuSO4 solution (500 mg/L) at 4°C for 10 days; (ii) electroporation of plasmid DNA in enlarged microspores by a single pulse of ∼375 V; (iii) induction of microspores after transfection at 28°C in NPB-99 medium and regeneration at 26°C in MMS5 medium; (iv) co-cultivation with Agrobacterium AGL-1 cells for transfer of plasmid T-DNA into microspores at day 0 for co-cultivation with timentin (200-400 mg/L).

  15. Efficient generation of marker-free transgenic rice plants using an improved transposon-mediated transgene reintegration strategy.

    Science.gov (United States)

    Gao, Xiaoqing; Zhou, Jie; Li, Jun; Zou, Xiaowei; Zhao, Jianhua; Li, Qingliang; Xia, Ran; Yang, Ruifang; Wang, Dekai; Zuo, Zhaoxue; Tu, Jumin; Tao, Yuezhi; Chen, Xiaoyun; Xie, Qi; Zhu, Zengrong; Qu, Shaohong

    2015-01-01

    Marker-free transgenic plants can be developed through transposon-mediated transgene reintegration, which allows intact transgene insertion with defined boundaries and requires only a few primary transformants. In this study, we improved the selection strategy and validated that the maize (Zea mays) Activator/Dissociation (Ds) transposable element can be routinely used to generate marker-free transgenic plants. A Ds-based gene of interest was linked to green fluorescent protein in transfer DNA (T-DNA), and a green fluorescent protein-aided counterselection against T-DNA was used together with polymerase chain reaction (PCR)-based positive selection for the gene of interest to screen marker-free progeny. To test the efficacy of this strategy, we cloned the Bacillus thuringiensis (Bt) δ-endotoxin gene into the Ds elements and transformed transposon vectors into rice (Oryza sativa) cultivars via Agrobacterium tumefaciens. PCR assays of the transposon empty donor site exhibited transposition in somatic cells in 60.5% to 100% of the rice transformants. Marker-free (T-DNA-free) transgenic rice plants derived from unlinked germinal transposition were obtained from the T1 generation of 26.1% of the primary transformants. Individual marker-free transgenic rice lines were subjected to thermal asymmetric interlaced-PCR to determine Ds(Bt) reintegration positions, reverse transcription-PCR and enzyme-linked immunosorbent assay to detect Bt expression levels, and bioassays to confirm resistance against the striped stem borer Chilo suppressalis. Overall, we efficiently generated marker-free transgenic plants with optimized transgene insertion and expression. The transposon-mediated marker-free platform established in this study can be used in rice and possibly in other important crops.

  16. Spontaneous generation of infectious prion disease in transgenic mice.

    Science.gov (United States)

    Torres, Juan-María; Castilla, Joaquín; Pintado, Belén; Gutiérrez-Adan, Alfonso; Andréoletti, Olivier; Aguilar-Calvo, Patricia; Arroba, Ana-Isabel; Parra-Arrondo, Beatriz; Ferrer, Isidro; Manzanares, Jorge; Espinosa, Juan-Carlos

    2013-12-01

    We generated transgenic mice expressing bovine cellular prion protein (PrP(C)) with a leucine substitution at codon 113 (113L). This protein is homologous to human protein with mutation 102L, and its genetic link with Gerstmann-Sträussler-Scheinker syndrome has been established. This mutation in bovine PrP(C) causes a fully penetrant, lethal, spongiform encephalopathy. This genetic disease was transmitted by intracerebral inoculation of brain homogenate from ill mice expressing mutant bovine PrP to mice expressing wild-type bovine PrP, which indicated de novo generation of infectious prions. Our findings demonstrate that a single amino acid change in the PrP(C) sequence can induce spontaneous generation of an infectious prion disease that differs from all others identified in hosts expressing the same PrP(C) sequence. These observations support the view that a variety of infectious prion strains might spontaneously emerge in hosts displaying random genetic PrP(C) mutations.

  17. Production of human lactoferrin and lysozyme in the milk of transgenic dairy animals: past, present, and future.

    Science.gov (United States)

    Cooper, Caitlin A; Maga, Elizabeth A; Murray, James D

    2015-08-01

    Genetic engineering, which was first developed in the 1980s, allows for specific additions to animals' genomes that are not possible through conventional breeding. Using genetic engineering to improve agricultural animals was first suggested when the technology was in the early stages of development by Palmiter et al. (Nature 300:611-615, 1982). One of the first agricultural applications identified was generating transgenic dairy animals that could produce altered or novel proteins in their milk. Human milk contains high levels of antimicrobial proteins that are found in low concentrations in the milk of ruminants, including the antimicrobial proteins lactoferrin and lysozyme. Lactoferrin and lysozyme are both part of the innate immune system and are secreted in tears, mucus, and throughout the gastrointestinal (GI) tract. Due to their antimicrobial properties and abundance in human milk, multiple lines of transgenic dairy animals that produce either human lactoferrin or human lysozyme have been developed. The focus of this review is to catalogue the different lines of genetically engineered dairy animals that produce either recombinant lactoferrin or lysozyme that have been generated over the years as well as compare the wealth of research that has been done on the in vitro and in vivo effects of the milk they produce. While recent advances including the development of CRISPRs and TALENs have removed many of the technical barriers to predictable and efficient genetic engineering in agricultural species, there are still many political and regulatory hurdles before genetic engineering can be used in agriculture. It is important to consider the substantial amount of work that has been done thus far on well established lines of genetically engineered animals evaluating both the animals themselves and the products they yield to identify the most effective path forward for future research and acceptance of this technology.

  18. Generation of doubled haploid transgenic wheat lines by microspore transformation.

    Directory of Open Access Journals (Sweden)

    Rhoda A T Brew-Appiah

    Full Text Available Microspores can be induced to develop homozygous doubled haploid plants in a single generation. In the present experiments androgenic microspores of wheat have been genetically transformed and developed into mature homozygous transgenic plants. Two different transformation techniques were investigated, one employing electroporation and the other co-cultivation with Agrobacterium tumefaciens. Different tissue culture and transfection conditions were tested on nine different wheat cultivars using four different constructs. A total of 19 fertile transformants in five genotypes from four market classes of common wheat were recovered by the two procedures. PCR followed by DNA sequencing of the products, Southern blot analyses and bio/histo-chemical and histological assays of the recombinant enzymes confirmed the presence of the transgenes in the T0 transformants and their stable inheritance in homozygous T1∶2 doubled haploid progenies. Several decisive factors determining the transformation and regeneration efficiency with the two procedures were determined: (i pretreatment of immature spikes with CuSO4 solution (500 mg/L at 4°C for 10 days; (ii electroporation of plasmid DNA in enlarged microspores by a single pulse of ∼375 V; (iii induction of microspores after transfection at 28°C in NPB-99 medium and regeneration at 26°C in MMS5 medium; (iv co-cultivation with Agrobacterium AGL-1 cells for transfer of plasmid T-DNA into microspores at day 0 for <24 hours; and (v elimination of AGL-1 cells after co-cultivation with timentin (200-400 mg/L.

  19. Transgenic and cloned animals in the food chain--are we prepared to tackle it?

    Science.gov (United States)

    Jagadeesan, Premanandh; Bin Salem, Samara

    2015-11-01

    Transgenic and cloned animal production for various purposes has been increasing rapidly in recent times. While the actual impact of these animals in the food chain is unknown, the significance of tracking and monitoring measures to curb accidental and or deliberate release has been discussed. Religious perspectives from different faiths and traditions have been presented. Although the concept of substantial equivalence satisfies the technical and nutritional requirements of these products when assessed against comparators, public opinion and religious concerns should also be considered by the regulators while developing policy regulations. In conclusion, measures to prevent real or perceived risks of transgenic and cloned animals in food production require global coordinated action. It is worthwhile to consider establishing effective tracking systems and analytical procedures as this will be a valuable tool if a global consensus is not reached on policy regulation.

  20. Approaches for Generating Animations for Lectures.

    Science.gov (United States)

    Roessling, Guido; Freisleben, Bernd

    This paper provides a short review of the following basic approaches for generating animations so that teachers can determine the way best suited for them and be better prepared to select a tool addressing their needs: (1) using classical presentation tools such as PowerPoint; (2) visual editing using drag and drop or selection of options; (3)…

  1. [The use of transgenic animals in biomedical research in Germany. Part 2: Ethical evaluation of the use of transgenic animals in biomedical research and perspectives for the changeover in research to research animal-free methods].

    Science.gov (United States)

    Sauer, Ursula G; Kolar, Roman; Rusche, Brigitte

    2006-01-01

    As a rule, transgenic animals are being used in in vivo experiments to examine gene functions, their regulation or the contribution of genetic alterations to the development of diseases. Many transgenic animals already are affected in their wellbeing due to the genetic modification alone regardless of the procedures performed with them. Moreover, it is to be questioned whether the experimental use of transgenic animals led to results that were of such outstanding scientific relevance that they legitimated the suffering of the animals. In order to point to possible approaches to avoiding the use of transgenic animals in the areas of research identified, subsequent investigations aimed at collecting information on non-animal test methods that might be applied in pursuing the aforesaid questions. In particular, these were non-animal test methods that make use of genetic techniques. Amongst these are in vitro cell culture methods with genetically modified cells, such as the so called Transfected Cell Array, as well as in vitro test methods, in which specifically targeted genes can be turned on or off selectively for example by the so-called RNA interference technique or by antisense oligonucleotide genes. Since such technologies can also be applied to cell cultures with human cells, investigations with these methods enable direct information on the function of human genes. Even though a one to one replacement of animal experiments with transgenic animals by non-animal test methods is considered unlikely, from the point of view of animal welfare the broad spectrum of already available non animal test methods with which to study the function of genes and genetically caused pathophysiological reactions proves that waiving of animal tests with transgenic animals is possible without impeding biomedical research. Even if it cannot be totally excluded that some very specific questions linked to the respective animal experiment might not be pursued for the time being

  2. Big Animal Cloning Using Transgenic Induced Pluripotent Stem Cells: A Case Study of Goat Transgenic Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Song, Hui; Li, Hui; Huang, Mingrui; Xu, Dan; Wang, Ziyu; Wang, Feng

    2016-02-01

    Using of embryonic stem cells (ESCs) could improve production traits and disease resistance by improving the efficiency of somatic cell nuclear transfer (SCNT) technology. However, robust ESCs have not been established from domestic ungulates. In the present study, we generated goat induced pluripotent stem cells (giPSCs) and transgenic cloned dairy goat induced pluripotent stem cells (tgiPSCs) from dairy goat fibroblasts (gFs) and transgenic cloned dairy goat fibroblasts (tgFs), respectively, using lentiviruses that contained hOCT4, hSOX2, hMYC, and hKLF4 without chemical compounds. The giPSCs and tgiPSCs expressed endogenous pluripotent markers, including OCT4, SOX2, MYC, KLF4, and NANOG. Moreover, they were able to maintain a normal karyotype and differentiate into derivatives from all three germ layers in vitro and in vivo. Using SCNT, tgFs and tgiPSCs were used as donor cells to produce embryos, which were named tgF-Embryos and tgiPSC-Embryos. The fusion rates and cleavage rates had no significant differences between tgF-Embryos and tgiPSC-Embryos. However, the expression of IGF-2, which is an important gene associated with embryonic development, was significantly lower in tgiPSC-Embryos than in tgF-Embryos and was not significantly different from vivo-Embryos.

  3. Generation of transgenic dogs that conditionally express green fluorescent protein.

    Science.gov (United States)

    Kim, Min Jung; Oh, Hyun Ju; Park, Jung Eun; Kim, Geon A; Hong, So Gun; Jang, Goo; Kwon, Mo Sun; Koo, Bon Chul; Kim, Teoan; Kang, Sung Keun; Ra, Jeong Chan; Ko, Chemyong; Lee, Byeong Chun

    2011-06-01

    We report the creation of a transgenic dog that conditionally expresses eGFP (enhanced green fluorescent protein) under the regulation of doxycycline. Briefly, fetal fibroblasts infected with a Tet-on eGFP vector were used for somatic cell nuclear transfer. Subsequently reconstructed oocytes were transferred to recipients. Three clones having transgenes were born and one dog was alive. The dog showed all features of inducible expression of eGFP upon doxycycline administration, and successful breeding resulted in eGFP-positive puppies, confirming stable insertion of the transgene into the genome. This inducible dog model will be useful for a variety of medical research studies.

  4. Ethical issues of transplanting organs from transgenic animals into human beings.

    Science.gov (United States)

    Behnam Manesh, Shima; Omani Samani, Reza; Behnam Manesh, Shayan

    2014-01-01

    One of the most important applications of transgenic animals for medical purposes is to transplant their organs into human's body, an issue which has caused a lot of ethical and scientific discussions. we can divide the ethical arguments to two comprehensive groups; the first group which is known as deontological critiques (related to the action itself regardless of any results pointing the human or animal) and the second group, called the consequentialist critiques (which are directly pointing the consequences of the action). The latter arguments also can be divided to two subgroups. In the first one which named anthropocentrism, just humankind has inherent value in the moral society, and it studies the problem just from a human-based point of view while in second named, biocentrism all the living organism have this value and it deals specially with the problem from the animal-based viewpoint. In this descriptive-analytic study, ethical issues were retrieved from books, papers, international guidelines, thesis, declarations and instructions, and even some weekly journals using keywords related to transgenic animals, organ, and transplantation. According to the precautionary principle with the strong legal and ethical background, due to lack of accepted scientific certainties about the safety of the procedure, in this phase, transplanting animal's organs into human beings have the potential harm and danger for both human and animals, and application of this procedure is unethical until the safety to human will be proven.

  5. Generation of human MHC (HLA-A11/DR1) transgenic mice for vaccine evaluation

    Science.gov (United States)

    Zeng, Yang; Gao, Tongtong; Zhao, Guangyu; Jiang, Yuting; Yang, Yi; Yu, Hong; Kou, Zhihua; Lone, Yuchun; Sun, Shihui; Zhou, Yusen

    2016-01-01

    ABSTRACT The rapid occurrence of emerging infectious diseases demonstrates an urgent need for a new preclinical experimental model that reliably replicates human immune responses. Here, a new homozygous humanized human leukocyte antigen (HLA)-A11/DR1 transgenic mouse (HLA-A11+/+/DR01+/+/H-2-β2m−/−/IAβ−/−) was generated by crossing HLA-A11 transgenic (Tg) mice with HLA-A2+/+/DR01+/+/H-2-β2m−/−/IAβ−/− mice. The HLA-A11-restricted immune response of this mouse model was then examined. HLA-A11 Tg mice expressing a chimeric major histocompatibility complex (MHC) molecule comprising the α1, α2, and β2m domains of human HLA-A11 and the α3 transmembrane and cytoplasmic domains of murine H-2Db were generated. The correct integration of HLA-A11 and HLA-DR1 into the genome of the HLA-A11/DR1 Tg mice (which lacked the expression of endogenous H-2-I/II molecules) was then confirmed. Immunizing mice with a recombinant HBV vaccine or a recombinant HIV-1 protein resulted in the generation of IFN-γ-producing cytotoxic T lymphocyte (CTL) and antigen-specific antibodies. The HLA-A11-restricted CTL response was directed at HLA immunodominant epitopes. These mice represent a versatile animal model for studying the immunogenicity of HLA CTL epitopes in the absence of a murine MHC response. The established animal model will also be useful for evaluating and optimizing T cell-based vaccines and for studying differences in antigen processing between mice and humans. PMID:26479036

  6. Ethical Issues of Transplanting Organs from Transgenic Animals into Human Beings

    Directory of Open Access Journals (Sweden)

    Shima Behnam Manesh

    2014-04-01

    Full Text Available One of the most important applications of transgenic animals for medical purposes is to transplant their organs into human’s body, an issue which has caused a lot of ethical and scientific discussions. we can divide the ethical arguments to two comprehensive groups; the first group which is known as deontological critiques (related to the action itself regardless of any results pointing the human or animal and the second group, called the consequentialist critiques (which are directly pointing the consequences of the action. The latter arguments also can be divided to two subgroups. In the first one which named anthropocentrism, just humankind has inherent value in the moral society, and it studies the problem just from a human-based point of view while in second named, biocentrism all the living organism have this value and it deals specially with the problem from the animal-based viewpoint. In this descriptive-analytic study, ethical issues were retrieved from books, papers, international guidelines, thesis, declarations and instructions, and even some weekly journals using keywords related to transgenic animals, organ, and transplantation. According to the precautionary principle with the strong legal and ethical background, due to lack of accepted scientific certainties about the safety of the procedure, in this phase, transplanting animal’s organs into human beings have the potential harm and danger for both human and animals, and application of this procedure is unethical until the safety to human will be proven.

  7. Modes of thrust generation in flying animals

    Science.gov (United States)

    Luo, Haoxiang; Song, Jialei; Tobalske, Bret; Luo Team; Tobalske Team

    2016-11-01

    For flying animals in forward flight, thrust is usually much smaller as compared with weight support and has not been given the same amount of attention. Several modes of thrust generation are discussed in this presentation. For insects performing slow flight that is characterized by low advance ratios (i.e., the ratio between flight speed and wing speed), thrust is usually generated by a "backward flick" mode, in which the wings moves upward and backward at a faster speed than the flight speed. Paddling mode is another mode used by some insects like fruit flies who row their wings backward during upstroke like paddles (Ristroph et al., PRL, 2011). Birds wings have high advance ratios and produce thrust during downstroke by directing aerodynamic lift forward. At intermediate advance ratios around one (e.g., hummingbirds and bats), the animal wings generate thrust during both downstroke and upstroke, and thrust generation during upstroke may come at cost of negative weight support. These conclusions are supported by previous experiment studies of insects, birds, and bats, as well as our recent computational modeling of hummingbirds. Supported by the NSF.

  8. Transgenic mammalian species, generated by somatic cell cloning, in biomedicine, biopharmaceutical industry and human nutrition/dietetics--recent achievements.

    Science.gov (United States)

    Samiec, M; Skrzyszowska, M

    2011-01-01

    Somatic cell cloning technology in mammals promotes the multiplication of productively-valuable genetically engineered individuals, and consequently allows also for standardization of transgenic farm animal-derived products, which, in the context of market requirements, will have growing significance. Gene farming is one of the most promising areas in modern biotechnology. The use of live bioreactors for the expression of human genes in the lactating mammary gland of transgenic animals seems to be the most cost-effective method for the production/processing of valuable recombinant therapeutic proteins. Among the transgenic farm livestock species used so far, cattle, goats, sheep, pigs and rabbits are useful candidates for the expression of tens to hundreds of grams of genetically-engineered proteins or xenogeneic biopreparations in the milk. At the beginning of the new millennium, a revolution in the treatment of disease is taking shape due to the emergence of new therapies based on recombinant human proteins. The ever-growing demand for such pharmaceutical or nutriceutical proteins is an important driving force for the development of safe and large-scale production platforms. The aim of this paper is to present an overall survey of the state of the art in investigations which provide the current knowledge for deciphering the possibilities of practical application of the transgenic mammalian species generated by somatic cell cloning in biomedicine, the biopharmaceutical industry, human nutrition/dietetics and agriculture.

  9. [Generation of sugar beet transgenic plants expressing bar gene].

    Science.gov (United States)

    Mishutkina, Ia V; Kamionskaia, A M; Skriabin, K G

    2010-01-01

    The parameters of transformation using Agrobacterium tumefaciens EHA 105 for 5 domestic sorts and lines of sugar beet (Beta vulgaris L. var. saccharifera (Alef) Krass) were optimized. The system of transgenic tissue selection based on resistance to phosphinothricin, allowing to avoid the appearing of chimeric shoots among initial transformants was developed. The transgenic plants of sugar beet sorts Ramonskaya single seed 47, L'govskaya single seed 52 and RMS 73, and LBO 17 and LBO 19 lines expressing the gene of phosphinothricin acetyl transferase bar have been obtained. The resistance of these sorts and lines to the effect of phosphinothricin in vitro has been shown.

  10. Analysis on virus resistance and fruit quality for T4 generation of transgenic papaya

    Institute of Scientific and Technical Information of China (English)

    WEI Xiangdong; LAN Congyu; LU Zhijing; YE Changming

    2007-01-01

    Molecular biological characterization,fruit characters,and nutrients were analyzed for T4 generation of transgenic papaya.All transgenic papaya plants with the mutated replicase (RP) gene from papaya ringspot virus (PRSV) showed high resistance or immunity against PRSV in the field.The RP transgene can be steadily inherited to,and expressed at RNA level,the progenies.The growth characteristics of transgenic papaya were much better than nontransgenic papaya in the field.The non-transgenic papaya seedlings began to show typical symptoms caused by PRSV after being inoculated with PRSV.They died quickly and never grew to produce fruit.The adult trees developed yellow leaves and produced smaller fruits and were doomed to a slow death after some time,while most oftransgenic papaya plants (about 91.8%) did not show any symptoms caused by PRSV,and produced more,bigger,and high quality fruits.Compared with non-transgenic plants,the fresh fruit length of T4 generation of transgenic papaya increased 2.6%-5%,and the diameter decreased 0.6%-1.5%.The flesh thickness of fresh fruit increased 12%-15%,which made it fitter for eating.Although the fresh fruit quality changed,there was no significant difference between transgenic and non-transgenic papaya.The quality characteristics of dry fruit including the contents of water,lipid,N,protein,reduced sugar,vitamin A,vitamin C,and carotene in the T4 generation of transgenic papaya were all the same as their non-transgenic parents.This means that transgenic plants and non-transgenic plants are substantially equivalent,and the transgene has no effect on dry fruit quality.In this study,we found that vitamin A and vitamin C in red-fleshed papaya were 1.4-1.8 and 1.78-2.07 times more than the yellow-fleshed ones,respectively,while N and protein were only 84.2%-92.1% and 82.1%-98.9% of the yellow-fleshed ones.

  11. The possibilities of practical application of transgenic mammalian species generated by somatic cell cloning in pharmacology, veterinary medicine and xenotransplantology.

    Science.gov (United States)

    Samiec, M; Skrzyszowska, M

    2011-01-01

    Cloning of genetically-modified mammals to produce: 1) novel animal bioreactors expressing human genes in rens, urinary bladder and the male accessory sex glands, as well as 2) porcine organs suitable in pig-to-human xenotransplantology, could offer new advantages for biomedical purposes. So too does the generation and/or multiplication of genetically-engineered cloned animals in order to produce: 3) physiologically-relevant animal models of serious monogenic human diseases and 4) prion disease-resistant small as well as large animals (i.e., rodents, ruminants). The basic purpose of this paper is to overview current knowledge deciphering the possibilities of using transgenic specimens created by somatic cell nuclear transfer in medical pharmacology, veterinary medicine, agriculture, transplantational medicine and immunology.

  12. Tol2-mediated generation of a transgenic haplochromine cichlid, Astatotilapia burtoni.

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    Scott A Juntti

    Full Text Available Cichlid fishes represent one of the most species-rich and rapid radiations of a vertebrate family. These ~2200 species, predominantly found in the East African Great Lakes, exhibit dramatic differences in anatomy, physiology, and behavior. However, the genetic bases for this radiation, and for the control of their divergent traits, are unknown. A flood of genomic and transcriptomic data promises to suggest mechanisms underlying the diversity, but transgenic technology will be needed to rigorously test the hypotheses generated. Here we demonstrate the successful use of the Tol2 transposon system to generate transgenic Astatotilapia burtoni, a haplochromine cichlid from Lake Tanganyika, carrying the GFP transgene under the control of the ubiquitous EF1α promoter. The transgene integrates into the genome, is successfully passed through the germline, and the widespread GFP expression pattern is stable across siblings and multiple generations. The stable inheritance and expression patterns indicate that the Tol2 system can be applied to generate A. burtoni transgenic lines. Transgenesis has proven to be a powerful technology for manipulating genes and cells in other model organisms and we anticipate that transgenic A. burtoni and other cichlids will be used to test the mechanisms underlying behavior and speciation.

  13. Generation of a new bioluminescent model for visualisation of mammary tumour development in transgenic mice

    Directory of Open Access Journals (Sweden)

    Zagozdzon Agnieszka M

    2012-05-01

    Full Text Available Abstract Background Numerous transgenic models have been generated to study breast cancer. However, despite many advantages, traditional transgenic models for breast cancer are also burdened with difficulties in early detection and longitudinal observation of transgene-induced tumours, which in most cases are randomly located and occur at various time points. Methods such as palpation followed by mechanical measurement of the tumours are of limited value in transgenic models. There is a crucial need for making these previously generated models suitable for modern methods of tumour visualisation and monitoring, e.g. by bioluminescence-based techniques. This approach was successfully used in the current study. Results A new mouse strain (MMTV-Luc2 mice expressing Luc2 luciferase primarily in mammary tissue in females, with low-level background expression in internal organs, was generated and bred to homozygosity. After these mice were intercrossed with MMTV-PyVT mice, all double transgenic females developed mammary tumours by the age of 10 weeks, the localisation and progression of which could be effectively monitored using the luminescence-based in vivo imaging. Luminescence-based readout allowed for early visualisation of the locally overgrown mammary tissue and for longitudinal evaluation of local progression of the tumours. When sampled ex vivo at the age of 10 weeks, all tumours derived from MMTV-Luc2PyVT females displayed robust bioluminescent signal. Conclusions We have created a novel transgenic strain for visualisation and longitudinal monitoring of mammary tumour development in transgenic mice as an addition and/or a new and more advanced alternative to manual methods. Generation of this mouse strain is vital for making many of the existing mammary tumour transgenic models applicable for in vivo imaging techniques.

  14. Generation of a new bioluminescent model for visualisation of mammary tumour development in transgenic mice

    LENUS (Irish Health Repository)

    Zagozdzon, Agnieszka M

    2012-05-30

    AbstractBackgroundNumerous transgenic models have been generated to study breast cancer. However, despite many advantages, traditional transgenic models for breast cancer are also burdened with difficulties in early detection and longitudinal observation of transgene-induced tumours, which in most cases are randomly located and occur at various time points. Methods such as palpation followed by mechanical measurement of the tumours are of limited value in transgenic models. There is a crucial need for making these previously generated models suitable for modern methods of tumour visualisation and monitoring, e.g. by bioluminescence-based techniques. This approach was successfully used in the current study.ResultsA new mouse strain (MMTV-Luc2 mice) expressing Luc2 luciferase primarily in mammary tissue in females, with low-level background expression in internal organs, was generated and bred to homozygosity. After these mice were intercrossed with MMTV-PyVT mice, all double transgenic females developed mammary tumours by the age of 10 weeks, the localisation and progression of which could be effectively monitored using the luminescence-based in vivo imaging. Luminescence-based readout allowed for early visualisation of the locally overgrown mammary tissue and for longitudinal evaluation of local progression of the tumours. When sampled ex vivo at the age of 10 weeks, all tumours derived from MMTV-Luc2PyVT females displayed robust bioluminescent signal.ConclusionsWe have created a novel transgenic strain for visualisation and longitudinal monitoring of mammary tumour development in transgenic mice as an addition and\\/or a new and more advanced alternative to manual methods. Generation of this mouse strain is vital for making many of the existing mammary tumour transgenic models applicable for in vivo imaging techniques.

  15. Generation and characterization of transgenic mice expressing tamoxifen-inducible cre-fusion protein specifically in mouse liver.

    Science.gov (United States)

    Zhu, Huan-Zhang; Chen, Jian-Quan; Cheng, Guo-Xiang; Xue, Jing-Lun

    2003-08-01

    To establish transgenic mice expressing tamoxifen-inducible Cre-ERt recombinase specifically in the liver and to provide an efficient animal model for studying gene function in the liver and creating various mouse models mimicking human diseases. Alb-Cre-ERt transgenic mice were produced by microinjecting the construct with Cre-ERt fusion gene of DNA fragments into fertilized eggs derived from inbred C57BL/6 strain. Transgenic mice were identified by using PCR and Southern blotting. Expression of Cre-ERt fusion gene was analyzed in the liver, kidney, brain and lung from F1 generation transgenic mice at 8 weeks of age by reverse transcription (RT)-PCR. Four hundred and fourteen fertilized eggs of C57 BL/6 mice were microinjected with recombinant Alb-Cre-ERt DNA fragments, and 312 survival eggs injected were transferred to the oviducts of 12 pseudopregnant recipient mice, 6 of 12 recipient mice became pregnant and gave birth to 44 offsprings. Of the 44 offsprings, two males and one female carried the hybrid Cre-ERt fusion gene. Three mice were determined as founders, and were back crossed to set up F1 generations with other inbred C57BL/6 mice. Transmission of Cre-ERt fusion gene in F1 offspring followed Mendelian rules. The expression of Cre-ERt mRNA was detected only in the liver of F1 offspring from two of three founder mice. Transgenic mice expressing tamoxifen-inducible Cre-ERt recombinase under control of the liver-specific promoter are preliminary established.

  16. Cellular reprogramming in farm animals: an overview of iPSC generation in the mammalian farm animal species.

    Science.gov (United States)

    Ogorevc, J; Orehek, S; Dovč, P

    2016-01-01

    Establishment of embryonic stem cell (ESC) lines has been successful in mouse and human, but not in farm animals. Development of direct reprogramming technology offers an alternative approach for generation of pluripotent stem cells, applicable also in farm animals. Induced pluripotent stem cells (iPSCs) represent practically limitless, ethically acceptable, individuum-specific source of pluripotent cells that can be generated from different types of somatic cells. iPSCs can differentiate to all cell types of an organism's body and have a tremendous potential for numerous applications in medicine, agriculture, and biotechnology. However, molecular mechanisms behind the reprogramming process remain largely unknown and hamper generation of bona fide iPSCs and their use in human clinical practice. Large animal models are essential to expand the knowledge obtained on rodents and facilitate development and validation of transplantation therapies in preclinical studies. Additionally, transgenic animals with special traits could be generated from genetically modified pluripotent cells, using advanced reproduction techniques. Despite their applicative potential, it seems that iPSCs in farm animals haven't received the deserved attention. The aim of this review was to provide a systematic overview on iPSC generation in the most important mammalian farm animal species (cattle, pig, horse, sheep, goat, and rabbit), compare protein sequence similarity of pluripotency-related transcription factors in different species, and discuss potential uses of farm animal iPSCs. Literature mining revealed 32 studies, describing iPSC generation in pig (13 studies), cattle (5), horse (5), sheep (4), goat (3), and rabbit (2) that are summarized in a concise, tabular format.

  17. Recombineering strategies for developing next generation BAC transgenic tools for optogenetics and beyond

    Directory of Open Access Journals (Sweden)

    Jonathan T Ting

    2014-04-01

    Full Text Available The development and application of diverse BAC transgenic rodent lines has enabled rapid progress for precise molecular targeting of genetically-defined cell types in the mammalian central nervous system. These transgenic tools have played a central role in the optogenetic revolution in neuroscience. Indeed, an overwhelming proportion of studies in this field have made use of BAC transgenic cre driver lines to achieve targeted expression of optogenetic probes in the brain. In addition, several BAC transgenic mouse lines have been established for direct cell-type specific expression of Channelrhodopsin-2 (ChR2. While the benefits of these new tools largely outweigh any accompanying challenges, many available BAC transgenic lines may suffer from confounds due in part to increased gene dosage of one or more ‘extra’ genes contained within the large BAC DNA sequences. Here we discuss this under-appreciated issue and propose strategies for developing the next generation of BAC transgenic lines that are devoid of extra genes. Furthermore, we provide evidence that these strategies are simple, reproducible, and do not disrupt the intended cell-type specific transgene expression patterns for several distinct BAC clones. These strategies may be widely implemented for improved BAC transgenesis across diverse disciplines.

  18. Generation of β-lactoglobulin-modified transgenic goats by homologous recombination.

    Science.gov (United States)

    Zhu, Hongmei; Hu, Linyong; Liu, Jun; Chen, Huatao; Cui, Chenchen; Song, Yujie; Jin, Yaping; Zhang, Yong

    2016-12-01

    β-Lactoglobulin (BLG) is a dominant allergen present in the milk of goats and other ungulates, although it is not found in human breast milk. Thus, the presence of BLG restricts the consumption of goat's milk by humans. In the present study, we examined whether the disruption of the BLG gene in goats by homologous recombination (HR) reduced BLG content in goat's milk and decreased the allergic response to milk. In one approach, exon 2 of the BLG gene was efficiently targeted using HR with a BLG knockout vector. In a second approach to disrupt BLG gene expression and drive exogenous human α-lactalbumin (hLA) gene expression, two hLA knock-in constructs were used to target exons 1-4 of the BLG gene via HR, and expression of hLA was then confirmed in goat mammary epithelial cells in vitro. The recombinant clones from both approaches were then used for somatic cell nuclear transfer, generating two transgenic goats possessing a BLG knockout allele or site-specific hLA integration allele. Milk assays demonstrated a reduction in BLG levels in both the BLG knockout and hLA knock-in goats; furthermore, hLA was present in the hLA knock-in goat's milk. Allergenic analysis in mice indicated that the transgenic goat's milk was less allergenic than wild-type goat's milk. These results support the development of gene-targeted animals as an effective tool for reducing allergic reactions to milk and improving nutrition. © 2016 Federation of European Biochemical Societies.

  19. Pancreatic acinar cells: molecular insight from studies of signal-transduction using transgenic animals.

    Science.gov (United States)

    Yule, David I

    2010-11-01

    Pancreatic acinar cells are classical exocrine gland cells. The apical regions of clusters of coupled acinar cells collectively form a lumen which constitutes the blind end of a tube created by ductal cells - a structure reminiscent of a "bunch of grapes". When activated by neural or hormonal secretagogues, pancreatic acinar cells are stimulated to secrete a variety of proteins. These proteins are predominately inactive digestive enzyme precursors called "zymogens". Acinar cell secretion is absolutely dependent on secretagogue-induced increases in intracellular free Ca(2+). The increase in [Ca(2+)](i) has precise temporal and spatial characteristics as a result of the exquisite regulation of the proteins responsible for Ca(2+) release, Ca(2+) influx and Ca(2+) clearance in the acinar cell. This brief review discusses recent studies in which transgenic animal models have been utilized to define in molecular detail the components of the Ca(2+) signaling machinery which contribute to these characteristics.

  20. From Gene Targeting to Genome Editing: Transgenic animals applications and beyond

    Directory of Open Access Journals (Sweden)

    MAURÍCIO ROCHA-MARTINS

    2015-08-01

    Full Text Available ABSTRACTGenome modification technologies are powerful tools for molecular biology and related areas. Advances in animal transgenesis and genome editing technologies during the past three decades allowed systematic interrogation of gene function that can help model how the genome influences cellular physiology. Genetic engineering via homologous recombination (HR has been the standard method to modify genomic sequences. Nevertheless, nuclease-guided genome editing methods that were developed recently, such as ZFN, TALEN and CRISPR/Cas, opened new perspectives for biomedical research. Here, we present a brief historical perspective of genome modification methods, focusing on transgenic mice models. Moreover, we describe how new techniques were discovered and improved, present the paradigm shifts and discuss their limitations and applications for biomedical research as well as possible future directions.

  1. Stable Skin-specific Overexpression of Human CTLA4-Ig in Transgenic Mice through Seven Generations

    Institute of Scientific and Technical Information of China (English)

    Yong WANG; Yong NI; Hong WEI; Feng-Chao WANG; Liang-Peng GE; Xiang GAO

    2006-01-01

    Skin graft rejection is a typical cellular immune response, mainly mediated by T cells. Cytotoxic T lymphocyte associated antigen 4-immunoglobin (CTLA4-Ig) extends graft survival by blocking the T cell co-stimulation pathway and inhibiting T cell activation. To investigate the efficacy of CTLA4-Ig in prolonging skin graft survival, human CTLA4-Ig (hCTLA4-Ig) was engineered to overexpress in mouse skin by transgenesis using the K14 promoter. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot assay indicated that the expression of CTLA4-Ig remained skin-specific and relatively constant compared to the internal control protein, AKT, through seven generations. The presence and concentration of the hCTLA4-Ig protein in transgenic mouse sera was determined by enzyme-linked immunosorbent assay (ELISA), and the results indicated that the serum CTLA4-Ig concentration also remained constant through generations. Survival of transgenic mouse skins grafted onto rat wounds was remarkably prolonged compared to that of wild-type skins from the same mouse strain, and remained comparable among all seven generations. This suggested that the bioactive hCTLA4-Ig protein was stably expressed in transgenical mice through at least seven generations, which was consistent with the stable skin-specific CTLA4-Ig expression.The results demonstrated that the transgenic expression of hCTLA4-Ig in skin driven by the K14 promoter remained constant through generations, and a transgenic line can be established to provide transgenic skin with extended survival reproducibly.

  2. Analysis of T-DNA integration and generative segregation in transgenic winter triticale (x Triticosecale Wittmack

    Directory of Open Access Journals (Sweden)

    Hensel Goetz

    2012-09-01

    Full Text Available Abstract Background While the genetic transformation of the major cereal crops has become relatively routine, to date only a few reports were published on transgenic triticale, and robust data on T-DNA integration and segregation have not been available in this species. Results Here, we present a comprehensive analysis of stable transgenic winter triticale cv. Bogo carrying the selectable marker gene HYGROMYCIN PHOSPHOTRANSFERASE (HPT and a synthetic green fluorescent protein gene (gfp. Progeny of four independent transgenic plants were comprehensively investigated with regard to the number of integrated T-DNA copies, the number of plant genomic integration loci, the integrity and functionality of individual T-DNA copies, as well as the segregation of transgenes in T1 and T2 generations, which also enabled us to identify homozygous transgenic lines. The truncation of some integrated T-DNAs at their left end along with the occurrence of independent segregation of multiple T-DNAs unintendedly resulted in a single-copy segregant that is selectable marker-free and homozygous for the gfp gene. The heritable expression of gfp driven by the maize UBI-1 promoter was demonstrated by confocal laser scanning microscopy. Conclusions The used transformation method is a valuable tool for the genetic engineering of triticale. Here we show that comprehensive molecular analyses are required for the correct interpretation of phenotypic data collected from the transgenic plants.

  3. Generating of rice OsCENH3-GFP transgenic plants and their genetic applications

    Institute of Scientific and Technical Information of China (English)

    YU HengXiu; WANG Xin; GONG ZhiYun; TANG Ding; GU MingHong; CHENG ZhuKuan

    2008-01-01

    In order to investigate rice functional centromeres, OsCENH3-GFP chimeric gene was constructed and transformed into the indica rice variety, Zhongxian 3037, mediated by Agrobacturium. The integration of the exogenous genes in the transgenic plants was confirmed by PCR and Southern blotting. The transgenic plants grow normally during their whole life time, just like Zhongxian 3037. No significant defects were detected in either mitosis or meiosis of the transgenic plants. The overlapping of GFP signals and anti-CENH3 foci in both mitotic and meiotic cells from To and T1 generation plants indicated that GFP had been successfully fused with CENH3, so the GFP signals can well represent the CENH3 locations on each chromosome. To evaluate the applicability of the transgenic plants to other genetic studies, fluorescence in situ hybridization (FISH) using rice centromeric tandem repetitive sequence CentO as the probe was conducted on the zygotene chromosomes of pollen mother cells (PMCs). It has been revealed that the GFP signals are overlapping with CentO FISH signals, showing that CentO is one of the key elements constituting rice functional centromeres. Immunofluorescent staining using anti-α-tublin antibody and anti-PAIR2 antibody on the chromosomes during mitosis and meiosis stages of the transgenic plants further reveals that OsCENH3-GFP transgenic plants can be widely used for studying rice molecular biology, especially for tagging functional centromeres in both living cells and tissues.

  4. Generation of marker- and/or backbone-free transgenic wheat plants via Agrobacterium-mediated transformation

    Directory of Open Access Journals (Sweden)

    Wang Genping

    2016-09-01

    Full Text Available 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.

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

    Science.gov (United States)

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

    2016-01-01

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

  6. Illustrating Language through Computer Generated Animation.

    Science.gov (United States)

    Withrow, Margaret

    1979-01-01

    Originally part of a symposium on educational media for the deaf, the paper discusses the use of animated sequences which illustrate linguistic principles. The work of the Computer Graphics Research Group at Ohio State University is highlighted. It has been discovered that computer-assisted instructional systems enhance learning through greater…

  7. Generation and characterization of gsuα:EGFP transgenic zebrafish for evaluating endocrine-disrupting effects

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Xiaoxia [Key Laboratory of Aquatic Biodiversity and Conservation of the Chinese Academy of Sciences, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei (China); University of Chinese Academy of Sciences, Beijing (China); Chen, Xiaowen; Jin, Xia; He, Jiangyan [Key Laboratory of Aquatic Biodiversity and Conservation of the Chinese Academy of Sciences, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei (China); Yin, Zhan, E-mail: zyin@ihb.ac.cn [Key Laboratory of Aquatic Biodiversity and Conservation of the Chinese Academy of Sciences, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan, Hubei (China); Ningbo Laboratory, State Key Laboratory of Freshwater Ecology and Biotechnology (China)

    2014-07-01

    The glycoprotein subunit α (gsuα) gene encodes the shared α subunit of the three pituitary heterodimeric glycoprotein hormones: follicle-stimulating hormone β (Fshβ), luteinizing hormone β (Lhβ) and thyroid stimulating hormone β (Tshβ). In our current study, we identified and characterized the promoter region of zebrafish gsuα and generated a stable gsuα:EGFP transgenic line, which recapitulated the endogenous gsuα expression in the early developing pituitary gland. A relatively conserved regulatory element set is presented in the promoter regions of zebrafish and three other known mammalian gsuα promoters. Our results also demonstrated that the expression patterns of the gsuα:EGFP transgene were all identical to those expression patterns of the endogenous gsuα expression in the pituitary tissue when our transgenic fish were treated with various endocrine chemicals, including forskolin (FSK), SP600125, trichostatin A (TSA), KClO{sub 4}, dexamethasone (Dex), β-estradiol and progesterone. Thus, this gsuα:EGFP transgenic fish reporter line provides another valuable tool for investigating the lineage development of gsuα-expressing gonadotrophins and the coordinated regulation of various glycoprotein hormone subunit genes. These reporter fish can serve as a novel platform to perform screenings of endocrine-disrupting chemicals (EDCs) in vivo as well. - Highlights: • Identification of the promoter of zebrafish glycoprotein subunit α (gsuα) gene • Generation of stable transmission gsuα:EGFP transgenic zebrafish reporter • Demonstration of the recapitulation of the gsuα:EGFP and endogenous gsuα expression • Suggestion of the gsuα:EGFP transgenic zebrafish as a novel platform for EDC study.

  8. Generation of transgenic pigs by cytoplasmic injection of piggyBac transposase-based pmGENIE-3 plasmids.

    Science.gov (United States)

    Li, Zicong; Zeng, Fang; Meng, Fanming; Xu, Zhiqian; Zhang, Xianwei; Huang, Xiaoling; Tang, Fei; Gao, Wenchao; Shi, Junsong; He, Xiaoyan; Liu, Dewu; Wang, Chong; Urschitz, Johann; Moisyadi, Stefan; Wu, Zhenfang

    2014-05-01

    The process of transgenesis involves the introduction of a foreign gene, the transgene, into the genome of an animal. Gene transfer by pronuclear microinjection (PNI) is the predominant method used to produce transgenic animals. However, this technique does not always result in germline transgenic offspring and has a low success rate for livestock. Alternate approaches, such as somatic cell nuclear transfer using transgenic fibroblasts, do not show an increase in efficiency compared to PNI, while viral-based transgenesis is hampered by issues regarding transgene size and biosafety considerations. We have recently described highly successful transgenesis experiments with mice using a piggyBac transposase-based vector, pmhyGENIE-3. This construct, a single and self-inactivating plasmid, contains all the transpositional elements necessary for successful gene transfer. In this series of experiments, our laboratories have implemented cytoplasmic injection (CTI) of pmGENIE-3 for transgene delivery into in vivo-fertilized pig zygotes. More than 8.00% of the injected embryos developed into transgenic animals containing monogenic and often single transgenes in their genome. However, the CTI technique was unsuccessful during the injection of in vitro-fertilized pig zygotes. In summary, here we have described a method that is not only easy to implement, but also demonstrated the highest efficiency rate for nonviral livestock transgenesis.

  9. Transgenic plants and animals: Altered organisms from recombinant DNA technology. (Latest citations from the Life Sciences Collection data base). Published Search

    Energy Technology Data Exchange (ETDEWEB)

    1992-06-01

    The bibliography contains citations concerning the development and use of transgenic plants and animals. Transgenic plants and animals are organisms with foreign genes inserted into their cells. Topics include methods of induction of new genes and transgenetic expression in the organism, development of animal models of human diseases, and design of insect tolerant plants. Examples of transgenic organisms include mice, fish, chickens, pigs, rye, maize, tobacco, tomatoes, lettuce, and cotton. This information is of value for the increased production of food from animals by producing animal carcasses with reduced fat content. The information is also valuable for production of herbicide tolerant, virus resistant, and insect resistant crop plants, as well as the rapid production of transgenic plants with flowers and seeds. (Contains 250 citations and includes a subject term index and title list.)

  10. Implementation and enforcement of the 3Rs principle in the field of transgenic animals used for scientific purposes. Report and recommendations of the BfR expert workshop, May 18-20, 2009, Berlin, Germany.

    Science.gov (United States)

    Kretlow, Ariane; Butzke, Daniel; Goetz, Mario E; Grune, Barbara; Halder, Marlies; Henkler, Frank; Liebsch, Manfred; Nobiling, Rainer; Oelgeschlaeger, Michael; Reifenberg, Kurt; Schaefer, Bernd; Seiler, Andrea; Luch, Andreas

    2010-01-01

    In 2007, 2.7 million vertebrates were used for animal experiments and other scientific purposes in Germany alone. Since 1998 there has been an increase in the number of animals used for research purposes, which is partly attributable to the growing use of transgenic animals. These animals are, for instance, used as in vivo models to mimic human diseases like diabetes, cancer or Alzheimer's disease. Here, transgenic model organisms serve as valuable tools, being instrumental in facilitating the analysis of the molecular mechanisms underlying human diseases, and might contribute to the development of novel therapeutic approaches. Due to variable and, sometimes low, efficiency (depending on the species used), however, the generation of such animals often requires a large number of embryo donors and recipients. The experts evaluated methods that could possibly be utilised to reduce, refine or even replace experiments with transgenic vertebrates in the mid-term future. Among the promising alternative model organisms available at the moment are the fruit fly Drosophila melanogaster and the roundworm Caenorhabditis elegans. Specific cell culture experiments or three-dimensional (3D) tissue models also offer valuable opportunities to replace experiments with transgenic animals or reduce the number of laboratory animals required by assisting in decision-making processes. Furthermore, at the workshop an in vitro technique was presented which permits the production of complete human antibodies without using genetically modified ("humanised") animals. Up to now, genetically modified mice are widely used for this purpose. Improved breeding protocols, enhanced efficiency of mutagenesis as well as training of laboratory personnel and animal keepers can also help to reduce the numbers of laboratory animals. Well-trained staff in particular can help to minimise the pain, suffering and discomfort of animals and, at the same time, improve the quality of data obtained from animal

  11. Generation of a fluorescent transgenic zebrafish for detection of environmental estrogens

    Energy Technology Data Exchange (ETDEWEB)

    Chen Hao; Hu Jingying; Yang Jian; Wang Yuexiang; Xu Hui; Jiang Qiu; Gong Yuebo; Gu Yinliang [Department of Molecular Genetics, Shanghai Medical School and Key Laboratory of Molecular Medicine, Ministry of Education, Fudan University, 200032 (China); Song Houyan, E-mail: hysong@shmu.edu.cn [Department of Molecular Genetics, Shanghai Medical School and Key Laboratory of Molecular Medicine, Ministry of Education, Fudan University, 200032 (China)

    2010-01-21

    To establish a novel in vivo test system for rapid detection of environmental estrogens, an ere-zvtg1: gfp transgenic zebrafish line has been generated. In this transgenic line, under control conditions, GFP was exclusively expressed in the liver of mature adult female fish. Male and larval transgenic fish did not express GFP but could be induced to express GFP in the liver after exposure to 17-{alpha}-ethynylestradiol (EE{sub 2}). Concurrent accumulation of zvtg1 and gfp mRNAs in embryos and larvae after EE{sub 2} exposure was observed, which indicated that the expression of gfp transgene was driven by the zvtg1 promoter. Green fluorescence was first observed in the liver at 53, 74, 100 or 131 h post-fertilization (hpf) after exposure to 100, 10, 1 or 0.1 ng/L EE{sub 2} from 1 to 2 cell stage, respectively. As for mature male transgenic zebrafish, green fluorescence was observed after exposure to 100, 10, 1 or 0.1 ng/L EE{sub 2} for 2, 3, 4 or 7 days, respectively; as for mature female, fluorescence was increased after exposure to relatively high concentrations of EE{sub 2} (10 and 100 ng/L). Green fluorescence in the liver was increased with prolonging of exposure time and was repeatedly induced after removal and re-addition of EE{sub 2}. We also demonstrated that GFP expression could be induced by other estrogenic compounds, including {beta}-estradiol (E{sub 2}, 0.1 {mu}g/L), cadmium chloride (CdCl{sub 2}, 10 {mu}g/L), zearalenone (50 {mu}g/L), estriol (E{sub 3}, 1 {mu}g/L), diethylstilbestrol (DES, 50 ng/L) bisphenol A (BPA, 1 mg/L) but not by weakly estrogenic compounds such as nonylphenol (NP, up to 10 mg/L), or non-estrogenic steroid hormones such as progesterone (up to 100 mg/L) and 17-hydroxysteroid (up to 50 mg/L). These data suggest the transgenic zebrafish is sensitive and specific for detection of estrogenic compounds. Because the observed-effect concentrations are as low as those of environment and the observed-effect exposure times are very short

  12. Generation of transgenic papaya with double resistance to Papaya ringspot virus and Papaya leaf-distortion mosaic virus.

    Science.gov (United States)

    Kung, Yi-Jung; Bau, Huey-Jiunn; Wu, Yi-Ling; Huang, Chiung-Huei; Chen, Tsui-Miao; Yeh, Shyi-Dong

    2009-11-01

    During the field tests of coat protein (CP)-transgenic papaya lines resistant to Papaya ringspot virus (PRSV), another Potyvirus sp., Papaya leaf-distortion mosaic virus (PLDMV), appeared as an emerging threat to the transgenic papaya. In this investigation, an untranslatable chimeric construct containing the truncated CP coding region of the PLDMV P-TW-WF isolate and the truncated CP coding region with the complete 3' untranslated region of PRSV YK isolate was transferred into papaya (Carica papaya cv. Thailand) via Agrobacterium-mediated transformation to generate transgenic plants with resistance to PLDMV and PRSV. Seventy-five transgenic lines were obtained and challenged with PRSV YK or PLDMV P-TW-WF by mechanical inoculation under greenhouse conditions. Thirty-eight transgenic lines showing no symptoms 1 month after inoculation were regarded as highly resistant lines. Southern and Northern analyses revealed that four weakly resistant lines have one or two inserts of the construct and accumulate detectable amounts of transgene transcript, whereas nine resistant lines contain two or three inserts without significant accumulation of transgene transcript. The results indicated that double virus resistance in transgenic lines resulted from double or more copies of the insert through the mechanism of RNA-mediated posttranscriptional gene silencing. Furthermore, three of nine resistant lines showed high levels of resistance to heterologous PRSV strains originating from Hawaii, Thailand, and Mexico. Our transgenic lines have great potential for controlling a number of PRSV strains and PLDMV in Taiwan and elsewhere.

  13. Cytoplasmic injection of murine zygotes with Sleeping Beauty transposon plasmids and minicircles results in the efficient generation of germline transgenic mice.

    Science.gov (United States)

    Garrels, Wiebke; Talluri, Thirumala R; Ziegler, Maren; Most, Ilka; Forcato, Diego O; Schmeer, Marco; Schleef, Martin; Ivics, Zoltán; Kues, Wilfried A

    2016-01-01

    Transgenesis in the mouse is an essential tool for the understanding of gene function and genome organization. Here, we describe a simplified microinjection protocol for efficient germline transgenesis and sustained transgene expression in the mouse model employing binary Sleeping Beauty transposon constructs of different topology. The protocol is based on co-injection of supercoiled plasmids or minicircles, encoding the Sleeping Beauty transposase and a transposon construct, into the cytoplasm of murine zygotes. Importantly, this simplified injection avoids the mechanical penetration of the vulnerable pronuclear membrane, resulting in higher survival rates of treated embryos and a more rapid pace of injections. Upon translation of the transposase, transposase-catalyzed transposition into the genome results in stable transgenic animals carrying monomeric transgenes. In summary, cytoplasmic injection of binary transposon constructs is a feasible, plasmid-based, and simplified microinjection method to generate genetically modified mice. The modular design of the components allows the multiplexing of different transposons, and the generation of multi-transposon transgenic mice in a single step.

  14. Realistic facial animation generation based on facial expression mapping

    Science.gov (United States)

    Yu, Hui; Garrod, Oliver; Jack, Rachael; Schyns, Philippe

    2014-01-01

    Facial expressions reflect internal emotional states of a character or in response to social communications. Though much effort has been taken to generate realistic facial expressions, it still remains a challenging topic due to human being's sensitivity to subtle facial movements. In this paper, we present a method for facial animation generation, which reflects true facial muscle movements with high fidelity. An intermediate model space is introduced to transfer captured static AU peak frames based on FACS to the conformed target face. And then dynamic parameters derived using a psychophysics method is integrated to generate facial animation, which is assumed to represent natural correlation of multiple AUs. Finally, the animation sequence in the intermediate model space is mapped to the target face to produce final animation.

  15. Generation of marker-free Bt transgenic indica rice and evaluation of its yellow stem borer resistance.

    Science.gov (United States)

    Kumar, S; Arul, L; Talwar, D

    2010-01-01

    We report on generation of marker-free (‘clean DNA’) transgenic rice (Oryza sativa), carrying minimal gene-expression-cassettes of the genes of interest, and evaluation of its resistance to yellow stem borer Scirpophaga incertulas (Lepidoptera: Pyralidae). The transgenic indica rice harbours a translational fusion of 2 different Bacillus thuringiensis (Bt) genes, namely cry1B-1Aa, driven by the green-tissue-specific phosphoenol pyruvate carboxylase (PEPC) promoter. Mature seed-derived calli of an elite indica rice cultivar Pusa Basmati-1 were co-bombarded with gene-expression-cassettes (clean DNA fragments) of the Bt gene and the marker hpt gene, to generate marker-free transgenic rice plants. The clean DNA fragments for bombardment were obtained by restriction digestion and gel extraction. Through biolistic transformation, 67 independent transformants were generated. Transformation frequency reached 3.3%, and 81% of the transgenic plants were co-transformants. Stable integration of the Bt gene was confirmed, and the insert copy number was determined by Southern analysis. Western analysis and ELISA revealed a high level of Bt protein expression in transgenic plants. Progeny analysis confirmed stable inheritance of the Bt gene according to the Mendelian (3:1) ratio. Insect bioassays revealed complete protection of transgenic plants from yellow stem borer infestation. PCR analysis of T2 progeny plants resulted in the recovery of up to 4% marker-free transgenic rice plants.

  16. Isolation of yellow catfish β-actin promoter and generation of transgenic yellow catfish expressing enhanced yellow fluorescent protein.

    Science.gov (United States)

    Ge, Jiachun; Dong, Zhangji; Li, Jingyun; Xu, Zhiqiang; Song, Wei; Bao, Jie; Liang, Dong; Li, Junbo; Li, Kui; Jia, Wenshuang; Zhao, Muzi; Cai, Yongxiang; Yang, Jiaxin; Pan, Jianlin; Zhao, Qingshun

    2012-10-01

    Yellow catfish (Pelteobagrus fulvidraco Richardson) is one of the most important freshwater farmed species in China. However, its small size and slow growth rate limit its commercial value. Because genetic engineering has been a powerful tool to develop and improve fish traits for aquaculture, we performed transgenic research on yellow catfish in order to increase its size and growth rate. Performing PCR with degenerate primers, we cloned a genomic fragment comprising 5'-flanking sequence upstream of the initiation codon of β-actin gene in yellow catfish. The sequence is 1,017 bp long, containing the core sequence of proximal promoter including CAAT box, CArG motif and TATA box. Microinjecting the transgene construct Tg(beta-actin:eYFP) of the proximal promoter fused to enhanced yellow fluorescent protein (eYFP) reporter gene into zebrafish and yellow catfish embryos, we found the promoter could drive the reporter to express transiently in both embryos at early development. Screening the offspring of five transgenic zebrafish founders developed from the embryos microinjected with Tg(ycbeta-actin:mCherry) or 19 yellow catfish founders developed from the embryos microinjected with Tg(beta-actin:eYFP), we obtained three lines of transgenic zebrafish and one transgenic yellow catfish, respectively. Analyzing the expression patterns of the reporter genes in transgenic zebrafish (Tg(ycbeta-actin:mCherry)nju8/+) and transgenic yellow catfish (Tg(beta-actin:eYFP)nju11/+), we found the reporters were broadly expressed in both animals. In summary, we have established a platform to make transgenic yellow catfish using the proximal promoter of its own β-actin gene. The results will help us to create transgenic yellow catfish using "all yellow catfish" transgene constructs.

  17. Protection by nitric oxide against liver inflammatory injury in animals carrying a nitric oxide synthase-2 transgene.

    Science.gov (United States)

    Mojena, M; Hortelano, S; Castrillo, A; Diaz-Guerra, M J; Garcia-Barchino, M J; Saez, G T; Bosca, L

    2001-03-01

    The effect of pre-existent hepatic NO synthesis on liver injury induced by lipopolysaccharide was studied in animals carrying a nitric oxide synthase-2 (NOS-2) transgene under the control of the phosphoenolpyruvate carboxykinase (PEPCK) promoter. These animals expressed NOS-2 in liver cells under fasting conditions. Lipopolysaccharide-induced liver injury in D-galactosamine-conditioned mice, which enhanced notably the effect of the endotoxin on the liver, was impaired in animals expressing NOS-2. This protection against inflammatory liver damage was dependent on NO synthesis and was caused by an inhibition of nuclear factor kB (NF-kB) activity and an impairment of the synthesis of the proinflammatory cytokines tumor necrosis factor a and interleukin 1b. These data indicate that intrahepatic synthesis of NO protects liver by inhibiting the release of cascades of proinflammatory mediators and suggest a beneficial role for local delivery of NO in the control of liver injury.

  18. Small-Animal PET Imaging of Tau Pathology with 18F-THK5117 in 2 Transgenic Mouse Models.

    Science.gov (United States)

    Brendel, Matthias; Jaworska, Anna; Probst, Federico; Overhoff, Felix; Korzhova, Viktoria; Lindner, Simon; Carlsen, Janette; Bartenstein, Peter; Harada, Ryuichi; Kudo, Yukitsuka; Haass, Christian; Van Leuven, Fred; Okamura, Nobuyuki; Herms, Jochen; Rominger, Axel

    2016-05-01

    Abnormal accumulation of tau aggregates in the brain is one of the hallmarks of Alzheimer disease neuropathology. We visualized tau deposition in vivo with the previously developed 2-arylquinoline derivative (18)F-THK5117 using small-animal PET in conjunction with autoradiography and immunohistochemistry gold standard assessment in 2 transgenic mouse models expressing hyperphosphorylated tau. Small-animal PET recordings were obtained in groups of P301S (n = 11) and biGT mice (n = 16) of different ages, with age-matched wild-type (WT) serving as controls. After intravenous administration of 16 ± 2 MBq of (18)F-THK5117, a dynamic 90-min emission recording was initiated for P301S mice and during 20-50 min after injection for biGT mice, followed by a 15-min transmission scan. After coregistration to the MRI atlas and scaling to the cerebellum, we performed volume-of-interest-based analysis (SUV ratio [SUVR]) and statistical parametric mapping. Small-animal PET results were compared with autoradiography ex vivo and in vitro and further validated with AT8 staining for neurofibrillary tangles. SUVRs calculated from static recordings during the interval of 20-50 min after tracer injection correlated highly with estimates of binding potential based on the entire dynamic emission recordings (R = 0.85). SUVR increases were detected in the brain stem of aged P301S mice (+11%; P parametric mapping analysis. Saturable binding of the tracer was verified by autoradiographic blocking studies. In the first dedicated small-animal PET study in 2 different transgenic tauopathy mouse models using the tau tracer (18)F-THK5117, the temporal and spatial progression could be visualized in good correlation with gold standard assessments of tau accumulation. The serial small-animal PET method could afford the means for preclinical testing of novel therapeutic approaches by accommodating interanimal variability at baseline, while detection thresholds in young animals have to be considered.

  19. 转基因动物在毒理学中的应用%Use of transgeneic animals in toxicology

    Institute of Scientific and Technical Information of China (English)

    高虹; 乔红伟

    2011-01-01

    转基因动物作为研究代谢调节、分化、发育中有关基因功能的工具已在各领域发挥了巨大的作用.在新药开发和研究中,因转基因动物的准确、经济、实验次数少和显著缩短实验时间等优点,现已成为一种进行“快速筛选”和药物非临床前安全评价的的手段.本文将对毒理学研究方面常用的转基因小鼠和大鼠进行阐述,旨在更全面的了解其在毒理学研究中的重要作用,为利用转基因技术技术培育应用于毒理学研究方面的转基因小鼠和大鼠的制备提供参考性的依据.%Transgeneie animals play an important role in researches of gene functions related to regulation of metabolism, cell differentiation and growth, etc. In the new drug development and investigation, transgenic animal models have been a useful tool of non-clinic safety evaluation and rapid screening. In this article we will review the commonly used transgenic mice and rats in toxicology, and to briefly discuss the significance of transgenic animals in toxicological studies.

  20. Efficient and stable generation of transgenic silkworm, Bombyx mori with the lepidopteran derived transposon piggyBac

    Institute of Scientific and Technical Information of China (English)

    DAI Hongjiu; XU Guojiang; Thomas Jean-luc; WANG Zhugang; JIANG Rongjing; FEI Jian

    2005-01-01

    The piggyBac transposable element was successfully used for generation of the transgenic silkworm, Bombyx mori. The EGFP was adopted in this study as a screening marker in transgenic vector under the control of an artificial promoter containing Pax-6 binding sites that can drive eye-specific genes expression in various insect species including B. mori and Drosophila. 111 independent transgenic lines were obtained among 700 fertile G0 moths. Most of the transgenic lines contained two or more chromosomal insertions of the EGFP marker, which were stably inherited over more than six generations during the time of this project. PiggyBac-mediated transposition was confirmed by identifying the characteristic TTAA duplication sequence at the insertion sites. The stable and high efficient transmission of a genetic marker into B. mori confirms the usage of this vector-marker system for industrial production and theoretical research.

  1. Generation of a Stable Transgenic Swine Model Expressing a Porcine Histone 2B-eGFP Fusion Protein for Cell Tracking and Chromosome Dynamics Studies

    Science.gov (United States)

    Simpson, Sean; Collins, Bruce; Sommer, Jeff; Petters, Robert M.; Caballero, Ignacio; Platt, Jeff L.

    2017-01-01

    Transgenic pigs have become an attractive research model in the field of translational research, regenerative medicine, and stem cell therapy due to their anatomic, genetic and physiological similarities with humans. The development of fluorescent proteins as molecular tags has allowed investigators to track cell migration and engraftment levels after transplantation. Here we describe the development of two transgenic pig models via SCNT expressing a fusion protein composed of eGFP and porcine Histone 2B (pH2B). This fusion protein is targeted to the nucleosomes resulting a nuclear/chromatin eGFP signal. The first model (I) was generated via random insertion of pH2B-eGFP driven by the CAG promoter (chicken beta actin promoter and rabbit Globin poly A; pCAG-pH2B-eGFP) and protected by human interferon-β matrix attachment regions (MARs). Despite the consistent, high, and ubiquitous expression of the fusion protein pH2B-eGFP in all tissues analyzed, two independently generated Model I transgenic lines developed neurodegenerative symptoms including Wallerian degeneration between 3–5 months of age, requiring euthanasia. A second transgenic model (II) was developed via CRISPR-Cas9 mediated homology-directed repair (HDR) of IRES-pH2B-eGFP into the endogenous β-actin (ACTB) locus. Model II transgenic animals showed ubiquitous expression of pH2B-eGFP on all tissues analyzed. Unlike the pCAG-pH2B-eGFP/MAR line, all Model II animals were healthy and multiple pregnancies have been established with progeny showing the expected Mendelian ratio for the transmission of the pH2B-eGFP. Expression of pH2B-eGFP was used to examine the timing of the maternal to zygotic transition after IVF, and to examine chromosome segregation of SCNT embryos. To our knowledge this is the first viable transgenic pig model with chromatin-associated eGFP allowing both cell tracking and the study of chromatin dynamics in a large animal model. PMID:28081156

  2. Evidence for the accumulation of Abeta immunoreactive material in the human brain and in transgenic animal models.

    Science.gov (United States)

    Cuello, A Claudio; Allard, Simon; Ferretti, Maria Teresa

    2012-12-10

    In this review we highlight the evidence for an intracellular origin of Abeta (Aβ) amyloid peptides as well as the observations for a pathological accumulation of these peptides in Alzheimer's disease and Down syndrome, as well as in transgenic animal models. We deliberate on the controversy as to whether the intracellular Aβ immunoreactive material is simply an accumulation of unprocessed full length amyloid precursor protein (APP) or a mix of processed APP fragments including Aβ. Finally, we discuss the possible pathological significance of these intracellular APP fragments and the expected future research directions regarding this thought-provoking problem. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. Transgenic mice in developmental toxicology

    Energy Technology Data Exchange (ETDEWEB)

    Woychik, R.P.

    1992-01-01

    Advances in molecular biology and embryology are being utilized for the generation of transgenic mice, animals that contain specific additions, deletions, or modifications of genes or sequences in their DNA. Mouse embryonic stem cells and homologous recombination procedures have made it possible to target specific DNA structural alterations to highly localized region in the host chromosomes. The majority of the DNA structural rearrangements in transgenic mice can be passed through the germ line and used to establish new genetic traits in the carrier animals. Since the use of transgenic mice is having such an enormous impact on so many areas of mammalian biological research, including developmental toxicology, the objective of this review is to briefly describe the fundamental methodologies for generating transgenic mice and to describe one particular application that has direct relevance to the field of genetic toxicology.

  4. Transgenic mice in developmental toxicology

    Energy Technology Data Exchange (ETDEWEB)

    Woychik, R.P.

    1992-12-31

    Advances in molecular biology and embryology are being utilized for the generation of transgenic mice, animals that contain specific additions, deletions, or modifications of genes or sequences in their DNA. Mouse embryonic stem cells and homologous recombination procedures have made it possible to target specific DNA structural alterations to highly localized region in the host chromosomes. The majority of the DNA structural rearrangements in transgenic mice can be passed through the germ line and used to establish new genetic traits in the carrier animals. Since the use of transgenic mice is having such an enormous impact on so many areas of mammalian biological research, including developmental toxicology, the objective of this review is to briefly describe the fundamental methodologies for generating transgenic mice and to describe one particular application that has direct relevance to the field of genetic toxicology.

  5. PPG: online generation of protein pictures and animations.

    Science.gov (United States)

    Binisti, Cédric; Salim, Ahmed Ali; Tufféry, Pierre

    2005-07-01

    The protein picture generator (PPG) is an online service to generate pictures of a protein structure. Its design was conceived as an answer to the need expressed by a part of the community to have some means to produce simply complex pictures to insert in publications or in presentations. PPG can produce static or animated pictures. It can be accessed at http://bioserv.rpbs.jussieu.fr/cgi-bin/PPG.

  6. Efficient generation of transgenic chickens using the spermatogonial stem cells in vivo and ex vivo transfection

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    -pression became readily detectable in the frozen whole mount tissue sections of recepient testes. Moreover, sperms carrying GFP could be produced normally. The results of artificially inseminating wild-type females with these sperms showed 12.5% (8/64) of offspring embryo expressed GFP and 11.1% (2/18) hatched chicks were tested transgenic. Our data therefore suggest TMGT and TTSSCs are the feasible methods for the generation of transgenic chickens.

  7. Efficient generation of transgenic chickens using the spermatogonial stem cells in vivo and ex vivo transfection

    Institute of Scientific and Technical Information of China (English)

    LI BiChun; YU Fei; WANG KeHua; CHEN GuoHong; SUN GuoBo; SUN HuaiChang; XU Qi; GAO Bo; ZHOU GuanYue; ZHAO WenMing; WU XinSheng; BAO WenBin

    2008-01-01

    -pression became readily detectable in the frozen whole mount tissue sections of recepient testes. Moreover, sperms carrying GFP could be produced normally. The results of artificially inseminating wild-type females with these sperms showed 12.5% (8/64) of offspring embryo expressed GFP and 11.1% (2/18) hatched chicks were tested transgenic. Our data therefore suggest TMGT and TTSSCs are the feasible methods for the generation of transgenic chickens.

  8. 外源基因在转基因动物中遗传和表达的稳定性%Inheritance and expression stability of transgene in transgenic animals

    Institute of Scientific and Technical Information of China (English)

    孔庆然; 刘忠华

    2011-01-01

    转基因技术经过近半个世纪的发展,已成为当今生物技术研究的热点.近10多年来,与核移植技术的结合,转基因效率大大提高,携带有不同外源基因的不同种类的转基因动物迅速增加.但是,成功获得转基因动物并不是转基因动物研究的最终日的,如何利用转基因技术为人类的需求服务才是科研人员始终面对的课题.在畜牧生产领域,通过转基因技术培育家畜新品种是转基因技术应用的重要体现,在我国这方面已经引起了广泛关注.但迄今为止,外源基因在转基因动物中遗传和表达的稳定性仍然是亟待解决的问题,究其原因,这主要与位置效应,外源基因的表观遗传学修饰和遗传效率相关,文章结合目前的研究进展和本实验室的研究结果,从这3方面阐述其作用机制,期望为转基因动物遗传育种向产业化的迈进提供一定的理论探讨.%Transgenic technology is one of the most hotspots in biology.In the past decade, the progress in animal cloning has provided an alternative method to improve transgenic efficiency.Many kinds of transgenic animals have been successfully produced via the combination of transfection and nuclear transfer.However, the ultimate aim of transgenesis is not to produce several transgenic animals, but to service for the needs of human.In animal production, transgenic technology has been used to breed new livestock, which has received a lot of attention in China.It has been evidenced that inheritance and expression instability of transgene in transgenic animals is still the major limitation, which is attributed to position effect,epigenetic modification, and hereditary efficiency of transgene.In this review, we discussed the three points for promoting the industrialization of animal transgenic breeding.

  9. Transgenic plants and animals: Altered organisms from recombinant DNA technology. July 1982-July 1989 (Citations from the Life Sciences Collection data base). Report for July 1982-July 1989

    Energy Technology Data Exchange (ETDEWEB)

    1990-01-01

    This bibliography contains citations concerning the development and use of transgenic plants and animals. Topics include methods of induction of new genes and transgenetic expression in the organism, development of animal models of human diseases, and design of insect tolerant plants. Examples of transgenic organisms include mice, fish, chicken, pigs, rye, maize, tobacco, tomatoes, lettuce, and cotton. This information is of value for the increased production of food from animals by producing animal carcasses with reduced fat content. The information is also valuable for production of herbicide tolerant, virus resistant, and insect resistant crop plants, as well as the rapid production of transgenic plants with flowers and seeds. (Contains 383 citations fully indexed and including a title list.)

  10. Building a Better Bean: Prospects for the Next Generation of Transgenic Soybeans

    Institute of Scientific and Technical Information of China (English)

    T. Jones; P. Olhoft; M. Cheng; H.P. Hong; L. Mankin; S. Hill

    2007-01-01

    @@ The first transgenic soybeans, with the glyphosate-tolerant trait, were introduced just over a decade ago in 1996. The rapid rate of adoption and global cultivation of transgenic soybeans since then is an unqualified success story.

  11. A three generation reproduction study with Sprague-Dawley rats consuming high-amylose transgenic rice.

    Science.gov (United States)

    Zhou, Xing Hua; Dong, Ying; Zhao, Yan Sheng; Xiao, Xiang; Wang, Yun; He, Yuan Qing; Liu, Qiao Quan

    2014-12-01

    The transgenic rice line (TRS) enriched with amylose and resistant starch (RS) was developed by antisense RNA inhibition of starch-branching enzymes. Cereal starch with high amylose has a great benefit on human health through its resistant starch. In order to evaluate the effect of transgenic rice on rats, the rats were fed diets containing 70% TRS rice flour, its near-isogenic rice flour or the standard diet as the control through three generations. In the present study, clinical performance, reproductive capacity and pathological responses including body weight, food consumption, reproductive data, hematological parameters, serum chemistry components, organ relative weights and histopathology were examined. Some statistically significant differences were observed in rats consuming the high amylose rice diet when compared to rats fed the near-isogenic control rice diet or the conventional (non-rice) standard diet. These differences were generally of small magnitude, appeared to be random in nature, and were within normal limits for the strain of rat used, and were therefore not considered to be biologically meaningful or treatment related.

  12. Generation of NSE-MerCreMer transgenic mice with tamoxifen inducible Cre activity in neurons.

    Directory of Open Access Journals (Sweden)

    Mandy Ka Man Kam

    Full Text Available To establish a genetic tool for conditional deletion or expression of gene in neurons in a temporally controlled manner, we generated a transgenic mouse (NSE-MerCreMer, which expressed a tamoxifen inducible type of Cre recombinase specifically in neurons. The tamoxifen inducible Cre recombinase (MerCreMer is a fusion protein containing Cre recombinase with two modified estrogen receptor ligand binding domains at both ends, and is driven by the neural-specific rat neural specific enolase (NSE promoter. A total of two transgenic lines were established, and expression of MerCreMer in neurons of the central and enteric nervous systems was confirmed. Transcript of MerCreMer was detected in several non-neural tissues such as heart, liver, and kidney in these lines. In the background of the Cre reporter mouse strain Rosa26R, Cre recombinase activity was inducible in neurons of adult NSE-MerCreMer mice treated with tamoxifen by intragastric gavage, but not in those fed with corn oil only. We conclude that NSE-MerCreMer lines will be useful for studying gene functions in neurons for the conditions that Cre-mediated recombination resulting in embryonic lethality, which precludes investigation of gene functions in neurons through later stages of development and in adult.

  13. A further review of inhalation studies with cigarette smoke and lung cancer in experimental animals, including transgenic mice.

    Science.gov (United States)

    Coggins, C R E

    2010-10-01

    The lack of an effective animal model for pulmonary carcinogenesis in smokers is a continuing problem for researchers trying to design Potentially Reduced Risk Products for those smokers who are either unwilling or unable to quit smoking. The major failing of inhalation assays with cigarette smoke in laboratory animals is that these assays produce only small percentages of animals with pulmonary tumors (e.g. adenomas, with the occasional adenocarcinoma), as opposed to the highly invasive carcinomas (e.g. small cell and squamous cell) seen in smokers. To update previous reviews on animal models, and to add different types of transgenic (Tg) mice to the review. Reviews were made of articles retrieved from PubMed and elsewhere. The addition of Tg mice to the arsenal of tests used for the evaluation of the carcinogenic potential of cigarettes did not result in any better understanding of the inability of such testing to reflect the epidemiological evidence for lung cancer in smokers. As in previous reviews on the subject, the best assay providing support for the epidemiology data is still the 5-month whole-body exposure of male A/J mice to a combination of mainstream/sidestream smoke, followed by a 4-month recovery.

  14. 动物转基因新技术研究进展%Progress in Research of Animal Transgenic Technology

    Institute of Scientific and Technical Information of China (English)

    陈青; 曹文广

    2011-01-01

    动物转基因技术是将外源基因导入动物体内,外源基因随机整合或定点整合(打靶)在染色体基因组上并得到表达和遗传的生物技术.该技术自发现以来,在畜牧业、医药产业、环境保护以及新型生物材料等领域已得到了广泛应用.本文主要就近年来迅速发展起来的慢病毒载体导入法,精原干细胞法、锌指核酸酶介导的基因打靶、RNA干扰介导的基因沉默等新的高效转基因技术进行了概述.%Animal transgenic technology refers to the transfer of an exogenous gene into animal genome by random or site-specific integration and the foreign gene can be expressed and inherited to the offspring. Various technologies in animal transgenesis have now been widely applied to areas such as animal husbandry, medical industry, environmental protection and new biological materials production. This article reviews the recently developed technologies based on lentivectors, spermatogonial stem cells, zinc finger nucleases and RNA interference.

  15. The Advance in Research on Transgenic Animal Technical Application to the Transgenic Cattle%动物转基因技术在转基因牛中的研究进展

    Institute of Scientific and Technical Information of China (English)

    张兆顺; 成功; 昝林森

    2012-01-01

    本研究综述了目前应用于转基因牛中原核显微注射法、体细胞克隆法、病毒载体法等传统动物转基因技术以及具有应用前景的ZFN技术、TALEN技术和iPS技术等.概述了20年来国内外转基因牛重要的研究进展,并就目前转基因牛在乳腺生物反应器、抗病育种、品种改良主要3个方面的应用进行了介绍.在此基础上,就目前转基因牛育种中存在的问题及今后发展趋势进行了讨论,以促进转基因牛技术的发展.%The article summarized the transgenic animal technology used in transgenic cattle research, such as microinjection, somatic cell nuclear transfer, virus and the more prospects technology such as ZFN, TALEN and iPS. Reviewed the progress of transgenic cattle research in Domestic and foreign, and the application of transgenic cattle research in mammary bioreactor, breeding for disease resistance and breeding improvement. Finally, transgenic cattle breeding problems and future development trends were discussed to improve the research of transgenic cattle.

  16. Generation of minipigs with targeted transgene insertion by recombinase-mediated cassette exchange (RMCE) and somatic cell nuclear transfer (SCNT)

    DEFF Research Database (Denmark)

    Jakobsen, Jannik Ejnar; Johansen, Marianne G; Schmidt, Mette;

    2013-01-01

    Targeted transgenesis using site-specific recombinases is an attractive method to create genetically modified animals as it allows for integration of the transgene in a pre-selected transcriptionally active genomic site. Here we describe the application of recombinase-mediated cassette exchange...... (RMCE) in cells from a Göttingen minipig with four RMCE acceptor loci, each containing a green fluorescence protein (GFP) marker gene driven by a human UbiC promoter. The four RMCE acceptor loci segregated independent of each other, and expression profiles could be determined in various tissues. Using...... minicircles in RMCE in fibroblasts with all four acceptor loci and followed by SCNT, we produced piglets with a single copy of a transgene incorporated into one of the transcriptionally active acceptor loci. The transgene, consisting of a cDNA of the Alzheimer’s disease-causing gene PSEN1M146I driven...

  17. Identification of the genomic insertion site of Pmel-1 TCR α and β transgenes by next-generation sequencing.

    Directory of Open Access Journals (Sweden)

    Yun Ji

    Full Text Available The pmel-1 T cell receptor transgenic mouse has been extensively employed as an ideal model system to study the mechanisms of tumor immunology, CD8+ T cell differentiation, autoimmunity and adoptive immunotherapy. The 'zygosity' of the transgene affects the transgene expression levels and may compromise optimal breeding scheme design. However, the integration sites for the pmel-1 mouse have remained uncharacterized. This is also true for many other commonly used transgenic mice created before the modern era of rapid and inexpensive next-generation sequencing. Here, we show that whole genome sequencing can be used to determine the exact pmel-1 genomic integration site, even with relatively 'shallow' (8X coverage. The results were used to develop a validated polymerase chain reaction-based genotyping assay. For the first time, we provide a quick and convenient polymerase chain reaction method to determine the dosage of pmel-1 transgene for this freely and publically available mouse resource. We also demonstrate that next-generation sequencing provides a feasible approach for mapping foreign DNA integration sites, even when information of the original vector sequences is only partially known.

  18. Symmetry in locomotor central pattern generators and animal gaits

    Science.gov (United States)

    Golubitsky, Martin; Stewart, Ian; Buono, Pietro-Luciano; Collins, J. J.

    1999-10-01

    Animal locomotion is controlled, in part, by a central pattern generator (CPG), which is an intraspinal network of neurons capable of generating a rhythmic output. The spatio-temporal symmetries of the quadrupedal gaits walk, trot and pace lead to plausible assumptions about the symmetries of locomotor CPGs. These assumptions imply that the CPG of a quadruped should consist of eight nominally identical subcircuits, arranged in an essentially unique matter. Here we apply analogous arguments to myriapod CPGs. Analyses based on symmetry applied to these networks lead to testable predictions, including a distinction between primary and secondary gaits, the existence of a new primary gait called `jump', and the occurrence of half-integer wave numbers in myriapod gaits. For bipeds, our analysis also predicts two gaits with the out-of-phase symmetry of the walk and two gaits with the in-phase symmetry of the hop. We present data that support each of these predictions. This work suggests that symmetry can be used to infer a plausible class of CPG network architectures from observed patterns of animal gaits.

  19. Induced Pluripotent Stem Cells Generated from P0-Cre;Z/EG Transgenic Mice.

    Directory of Open Access Journals (Sweden)

    Yasuhiro Ogawa

    Full Text Available Neural crest (NC cells are a migratory, multipotent cell population that arises at the neural plate border, and migrate from the dorsal neural tube to their target tissues, where they differentiate into various cell types. Abnormal development of NC cells can result in severe congenital birth defects. Because only a limited number of cells can be obtained from an embryo, mechanistic studies are difficult to perform with directly isolated NC cells. Protein zero (P0 is expressed by migrating NC cells during the early embryonic period. In the P0-Cre;Z/EG transgenic mouse, transient activation of the P0 promoter induces Cre-mediated recombination, indelibly tagging NC-derived cells with enhanced green fluorescent protein (EGFP. Induced pluripotent stem cell (iPSC technology offers new opportunities for both mechanistic studies and development of stem cell-based therapies. Here, we report the generation of iPSCs from the P0-Cre;Z/EG mouse. P0-Cre;Z/EG mouse-derived iPSCs (P/G-iPSCs exhibited pluripotent stem cell properties. In lineage-directed differentiation studies, P/G-iPSCs were efficiently differentiated along the neural lineage while expressing EGFP. These results suggest that P/G-iPSCs are useful to study NC development and NC-associated diseases.

  20. Impact of intracellular beta-amyloid in transgenic animals and cell models.

    Science.gov (United States)

    Cuello, A Claudio; Canneva, Fabio

    2008-01-01

    The present commentary based on cell and animal models of intracellular beta-amyloid (iAbeta) expression indicates that low levels of microscopically undetectable iAbeta could have a physiological role in the modulation of the cyclic AMP response element (CRE)-dependent gene expression and, as a consequence, a positive influence on synaptic plasticity (the 'good' Abeta?). On the other hand, high levels of iAbeta resembling the pathological and microscopically visible accumulation of this amyloid peptide, akin to that observed in Down syndrome and Alzheimer's disease, disrupt CRE-regulated gene expression, therefore compromising the protein synthesis-dependent component of long-term potentiation (the 'bad' Abeta?). Moreover, intracellular pathology would be independent and additive to the toxic effects of the extracellular Abeta burden. 2008 S. Karger AG, Basel

  1. Generation of fad2 transgenic mice that produce omega-6 fatty acids

    Institute of Scientific and Technical Information of China (English)

    CHEN Qing; LIU Qing; WU ZhiFang; WANG ZongYi; GOU KeMian

    2009-01-01

    Fatty acid desaturase-2 (FAD2)introduces a double bond in position △12 in oleic acid (18:1)to form linoleic acid (18:2 n-6)in higher plants and microbes.A new transgenic expression cassette,containing CMV promoter/fad2 cDNA/SV40 polyA,was constructedto produce transgenic mice.Among 63 healthy offspring,10 founders (15.9%)integrated the cotton fad2 transgene into their genomes,as demonstrated by PCR and Southern blotting analysis.All founder mice were fertile and heterozygous fad2 female and nontransgenic littermates were used for fatty acid analysis using gas chromatography.One fad2 transgenic line showed substantial differences in the fatty acid profiles and the level of linoleic acid was increased 19% (P<0.05)in transgenic muscles compared to their nontransgenic littermates.Moreover,it exhibited an 87% and a 9% increase (P<0.05)in arachidonic acid (20:4 n-6)in muscles and liver,compared to their nontransgenic littermates.The results indicate that the plant fad2 gene can be functionally expressed in transgenic mice and may playan active role in conversion of oleic acid into linoleic acid.

  2. Generation of fad2 transgenic mice that produce omega-6 fatty acids

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Fatty acid desaturase-2 (FAD2) introduces a double bond in position 12 in oleic acid (18:1) to form linoleic acid (18:2 n-6) in higher plants and microbes. A new transgenic expression cassette, containing CMV promoter/fad2 cDNA/SV40 polyA, was constructedto produce transgenic mice. Among 63 healthy offspring, 10 founders (15.9%) integrated the cotton fad2 transgene into their genomes, as demonstrated by PCR and Southern blotting analysis. All founder mice were fertile and heterozygous fad2 female and nontransgenic littermates were used for fatty acid analysis using gas chromatography. One fad2 transgenic line showed substantial differences in the fatty acid profiles and the level of linoleic acid was increased 19% (P<0.05) in transgenic muscles compared to their nontransgenic littermates. Moreover, it exhibited an 87% and a 9% increase (P<0.05) in arachidonic acid (20:4 n-6) in muscles and liver, compared to their nontransgenic littermates. The results indicate that the plant fad2 gene can be functionally expressed in transgenic mice and may playan active role in conversion of oleic acid into linoleic acid.

  3. Retention of the ability to synthesize HIV-1 and HBV antigens in generations of tomato plants transgenic for the TBI-HBS gene

    Science.gov (United States)

    In development of new types of edible vaccines on the basis of transgenic plants, the ability of the latter to retain the synthesis of foreign antibodies in a series of generations is of great importance. For this purpose, the goal of this study was to investigate the ability of transgenic tomato pl...

  4. Generation and Characterization of a Transgenic Mouse Carrying a Functional Human β -Globin Gene with the IVSI-6 Thalassemia Mutation.

    Science.gov (United States)

    Breveglieri, Giulia; Mancini, Irene; Bianchi, Nicoletta; Lampronti, Ilaria; Salvatori, Francesca; Fabbri, Enrica; Zuccato, Cristina; Cosenza, Lucia C; Montagner, Giulia; Borgatti, Monica; Altruda, Fiorella; Fagoonee, Sharmila; Carandina, Gianni; Rubini, Michele; Aiello, Vincenzo; Breda, Laura; Rivella, Stefano; Gambari, Roberto; Finotti, Alessia

    2015-01-01

    Mouse models that carry mutations causing thalassemia represent a suitable tool to test in vivo new mutation-specific therapeutic approaches. Transgenic mice carrying the β-globin IVSI-6 mutation (the most frequent in Middle-Eastern regions and recurrent in Italy and Greece) are, at present, not available. We report the production and characterization of a transgenic mouse line (TG-β-IVSI-6) carrying the IVSI-6 thalassemia point mutation within the human β-globin gene. In the TG-β-IVSI-6 mouse (a) the transgenic integration region is located in mouse chromosome 7; (b) the expression of the transgene is tissue specific; (c) as expected, normally spliced human β-globin mRNA is produced, giving rise to β-globin production and formation of a human-mouse tetrameric chimeric hemoglobin (mu) α-globin2/(hu) β-globin2 and, more importantly, (d) the aberrant β-globin-IVSI-6 RNAs are present in blood cells. The TG-β-IVSI-6 mouse reproduces the molecular features of IVSI-6 β-thalassemia and might be used as an in vivo model to characterize the effects of antisense oligodeoxynucleotides targeting the cryptic sites responsible for the generation of aberrantly spliced β-globin RNA sequences, caused by the IVSI-6 mutation. These experiments are expected to be crucial for the development of a personalized therapy for β-thalassemia.

  5. Use of the 2A peptide for generation of multi-transgenic pigs through a single round of nuclear transfer.

    Directory of Open Access Journals (Sweden)

    Wei Deng

    Full Text Available Multiple genetic modifications in pigs can essentially benefit research on agriculture, human disease and xenotransplantation. Most multi-transgenic pigs have been produced by complex and time-consuming breeding programs using multiple single-transgenic pigs. This study explored the feasibility of producing multi-transgenic pigs using the viral 2A peptide in the light of previous research indicating that it can be utilized for multi-gene transfer in gene therapy and somatic cell reprogramming. A 2A peptide-based double-promoter expression vector that mediated the expression of four fluorescent proteins was constructed and transfected into primary porcine fetal fibroblasts. Cell colonies (54.3% formed under G418 selection co-expressed the four fluorescent proteins at uniformly high levels. The reconstructed embryos, which were obtained by somatic cell nuclear transfer and confirmed to express the four fluorescent proteins evenly, were transplanted into seven recipient gilts. Eleven piglets were delivered by two gilts, and seven of them co-expressed the four fluorescent proteins at equivalently high levels in various tissues. The fluorescence intensities were directly observed at the nose, hoof and tongue using goggles. The results suggest that the strategy of combining the 2A peptide and double promoters efficiently mediates the co-expression of the four fluorescent proteins in pigs and is hence a promising methodology to generate multi-transgenic pigs by a single nuclear transfer.

  6. Generation of Transgene-free Induced Pluripotent Stem Cells with Non-viral Methods

    Institute of Scientific and Technical Information of China (English)

    Tao Wang; Hua-shan Zhao; Qiu-ling Zhang; Chang-lin Xu; Chang-bai Liu

    2013-01-01

    Induced pluripotent stem (iPS) cells were originally generated from mouse fibroblasts by enforced expression of Yamanaka factors (Oct3/4,Sox2,Klf4,and c-Myc). The technique was quickly re-produced with human fibroblasts or mesenchymal stem cells. Although having been showed therapeutic po-tential in animal models of sickle cell anemia and Parkinson's disease,iPS cells generated by viral methods do not suit all the clinical applications. Various non-viral methods have appeared in recent years for application of iPS cells in cell transplantation therapy. These methods mainly include DNA vector-based approaches,transfection of mRNA,and transduction of reprogramming proteins. This review summarized these non-viral methods and compare the advantages,disadvantages,efficiency,and safety of these methods.

  7. Assessment of fetal cell chimerism in transgenic pig lines generated by Sleeping beauty transposition.

    Science.gov (United States)

    Garrels, Wiebke; Holler, Stephanie; Taylor, Ulrike; Herrmann, Doris; Niemann, Heiner; Ivics, Zoltan; Kues, Wilfried A

    2014-01-01

    Human cells migrate between mother and fetus during pregnancy and persist in the respective host for long-term after birth. Fetal microchimerism occurs also in twins sharing a common placenta or chorion. Whether microchimerism occurs in multiparous mammals such as the domestic pig, where fetuses have separate placentas and chorions, is not well understood. Here, we assessed cell chimerism in litters of wild-type sows inseminated with semen of transposon transgenic boars. Segregation of three independent monomeric transposons ensured an excess of transgenic over non-transgenic offspring in every litter. Transgenic siblings (n = 35) showed robust ubiquitous expression of the reporter transposon encoding a fluorescent protein, and provided an unique resource to assess a potential cell trafficking to non-transgenic littermates (n = 7) or mothers (n = 4). Sensitive flow cytometry, fluorescence microscopy, and real-time PCR provided no evidence for microchimerism in porcine littermates, or piglets and their mothers in both blood and solid organs. These data indicate that the epitheliochorial structure of the porcine placenta effectively prevents cellular exchange during gestation.

  8. Assessment of fetal cell chimerism in transgenic pig lines generated by Sleeping beauty transposition.

    Directory of Open Access Journals (Sweden)

    Wiebke Garrels

    Full Text Available Human cells migrate between mother and fetus during pregnancy and persist in the respective host for long-term after birth. Fetal microchimerism occurs also in twins sharing a common placenta or chorion. Whether microchimerism occurs in multiparous mammals such as the domestic pig, where fetuses have separate placentas and chorions, is not well understood. Here, we assessed cell chimerism in litters of wild-type sows inseminated with semen of transposon transgenic boars. Segregation of three independent monomeric transposons ensured an excess of transgenic over non-transgenic offspring in every litter. Transgenic siblings (n = 35 showed robust ubiquitous expression of the reporter transposon encoding a fluorescent protein, and provided an unique resource to assess a potential cell trafficking to non-transgenic littermates (n = 7 or mothers (n = 4. Sensitive flow cytometry, fluorescence microscopy, and real-time PCR provided no evidence for microchimerism in porcine littermates, or piglets and their mothers in both blood and solid organs. These data indicate that the epitheliochorial structure of the porcine placenta effectively prevents cellular exchange during gestation.

  9. Generation of transgenic watermelon resistant to Zucchini yellow mosaic virus and Papaya ringspot virus type W.

    Science.gov (United States)

    Yu, Tsong-Ann; Chiang, Chu-Hui; Wu, Hui-Wen; Li, Chin-Mei; Yang, Ching-Fu; Chen, Jun-Han; Chen, Yu-Wen; Yeh, Shyi-Dong

    2011-03-01

    Zucchini yellow mosaic virus (ZYMV) and Papaya ringspot virus type W (PRSV W) are major limiting factors for production of watermelon worldwide. For the effective control of these two viruses by transgenic resistance, an untranslatable chimeric construct containing truncated ZYMV coat protein (CP) and PRSV W CP genes was transferred to commercial watermelon cultivars by Agrobacterium-mediated transformation. Using our protocol, a total of 27 putative transgenic lines were obtained from three cultivars of 'Feeling' (23 lines), 'China baby' (3 lines), and 'Quality' (1 line). PCR and Southern blot analyses confirmed that the chimeric construct was incorporated into the genomic DNA of the transformants. Greenhouse evaluation of the selected ten transgenic lines of 'Feeling' cultivar revealed that two immune lines conferred complete resistance to ZYMV and PRSV W, from which virus accumulation were not detected by Western blotting 4 weeks after inoculation. The transgenic transcript was not detected, but small interfering RNA (siRNA) was readily detected from the two immune lines and T(1) progeny of line ZW 10 before inoculation, indicating that RNA-mediated post-transcriptional gene silencing (PTGS) is the underlying mechanism for the double-virus resistance. The segregation ratio of T(1) progeny of the immune line ZW10 indicated that the single inserted transgene is nuclearly inherited and associated with the phenotype of double-virus resistance as a dominant trait. The transgenic lines derived from the commercial watermelon cultivars have great potential for control of the two important viruses and can be implemented directly without further breeding.

  10. Computer-Generated, Three-Dimensional Character Animation.

    Science.gov (United States)

    Van Baerle, Susan Lynn

    This master's thesis begins by discussing the differences between 3-D computer animation of solid three-dimensional, or monolithic, objects, and the animation of characters, i.e., collections of movable parts with soft pliable surfaces. Principles from two-dimensional character animation that can be transferred to three-dimensional character…

  11. Characterization of Growth and Reproduction Performance, Transgene Integration, Expression, and Transmission Patterns in Transgenic Pigs Produced by piggyBac Transposition-Mediated Gene Transfer.

    Science.gov (United States)

    Zeng, Fang; Li, Zicong; Cai, Gengyuan; Gao, Wenchao; Jiang, Gelong; Liu, Dewu; Urschitz, Johann; Moisyadi, Stefan; Wu, Zhenfang

    2016-10-01

    Previously we successfully produced a group of EGFP-expressing founder transgenic pigs by a newly developed efficient and simple pig transgenesis method based on cytoplasmic injection of piggyBac plasmids. In this study, we investigated the growth and reproduction performance and characterized the transgene insertion, transmission, and expression patterns in transgenic pigs generated by piggyBac transposition. Results showed that transgene has no injurious effect on the growth and reproduction of transgenic pigs. Multiple copies of monogenic EGFP transgene were inserted at noncoding sequences of host genome, and passed from founder transgenic pigs to their transgenic offspring in segregation or linkage manner. The EGFP transgene was ubiquitously expressed in transgenic pigs, and its expression intensity was associated with transgene copy number but not related to its promoter DNA methylation level. To the best of our knowledge, this is first study that fully described the growth and reproduction performance, transgene insertion, expression, and transmission profiles in transgenic pigs produced by piggyBac system. It not only demonstrates that piggyBac transposition-mediated gene transfer is an effective and favorable approach for pig transgenesis, but also provides scientific information for understanding the transgene insertion, expression and transmission patterns in transgenic animals produced by piggyBac transposition.

  12. Modification of bacterial artificial chromosomes (BACs) and preparation of intact BAC DNA for generation of transgenic mice.

    Science.gov (United States)

    Gong, Shiaoching; Yang, X William

    2005-05-01

    BAC transgenesis is a powerful tool for the study of gene expression and gene function in the mouse in vivo. In this unit, detailed protocols are provided for modification (i.e., marker gene insertion, deletion, or point mutation) of BACs by homologous recombination in E. coli. This method utilizes a shuttle vector that allows transient expression of the E. coli RecA gene to support homologous recombination in the BAC host bacteria. In addition, two protocols are provided for purification of BAC DNA for microinjection to generate transgenic mice. Since BAC DNA is prone to degradation, which may introduce positional effects in transgenic mice, two methods are given for purification of intact BAC DNA for subsequent microinjection.

  13. Bacterial composting of animal fleshing generated from tannery industries.

    Science.gov (United States)

    Ravindran, B; Sekaran, G

    2010-12-01

    Animal fleshing (ANFL) is the major proteinaceous solid waste generated during the manufacture of leather, which requires to be disposed of by environmentally sound manner. This study reports about the treatment of ANFL into an organic compost and its effects on physiological parameters of different crops in a laboratory study. The ANFL was hydrolysed using Selenomonas ruminantium HM000123 and then the hydrolysed ANFL was mixed with cow dung and leaf litter for producing composted organic fertilizer (COF). The COF was characterized for pH, electrical conductivity (EC), total Kjeldhal nitrogen (TKN), and total organic carbon (TOC). The composting resulted in a significant reduction in pH, TOC and C:N ratio and an increase in TKN after 49 days in a compost reactor. Scanning electron microscope and FT-IR were used to analyse the hydrolysis of intra structural ANFL matrix and changes in the functional groups, respectively, in initial and final day COF. Thermogravimetry (TG) analysis was carried out for the raw mixture and COF samples to identify the weight loss under the nitrogen environment. The relative seed germination was found to be 94% in tomato (Lycopersicon esculentum), 92% in green gram (Vigna radiata), 86% in bottle gourd (Lagenaria siceraria (Mol.) Standl.) and 84% in cucumber (Cucumis sativus L.) using the extracts of COF. The results indicate that the combination of both hydrolysis and bacterial composting reduced the overall time required for composting and producing a nutrient-enriched compost product.

  14. Generation and characterization of neurogenin1-GFP transgenic medaka with potential for rapid developmental neurotoxicity screening

    Energy Technology Data Exchange (ETDEWEB)

    Fan Chunyang [Integrated Systems Toxicology and Toxicity Assessment Divisions, National Health and Environmental Effects Research Laboratory, US EPA, Research Triangle Park, NC 27711 (United States); Curriculum in Toxicology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599 (United States); Simmons, Steven O. [Integrated Systems Toxicology and Toxicity Assessment Divisions, National Health and Environmental Effects Research Laboratory, US EPA, Research Triangle Park, NC 27711 (United States); Law, Sheran H.W. [Environmental Sciences and Policy Division, Nicholas School of the Environment and Earth Sciences, Duke University, Durham, NC 27708 (United States); Jensen, Karl; Cowden, John [Integrated Systems Toxicology and Toxicity Assessment Divisions, National Health and Environmental Effects Research Laboratory, US EPA, Research Triangle Park, NC 27711 (United States); Hinton, David [Environmental Sciences and Policy Division, Nicholas School of the Environment and Earth Sciences, Duke University, Durham, NC 27708 (United States); Padilla, Stephanie [Integrated Systems Toxicology and Toxicity Assessment Divisions, National Health and Environmental Effects Research Laboratory, US EPA, Research Triangle Park, NC 27711 (United States); Ramabhadran, Ram, E-mail: Ram.Ramabhadran@gmail.com [Integrated Systems Toxicology and Toxicity Assessment Divisions, National Health and Environmental Effects Research Laboratory, US EPA, Research Triangle Park, NC 27711 (United States)

    2011-09-15

    Fish models such as zebrafish and medaka are increasingly used as alternatives to rodents in developmental and toxicological studies. These developmental and toxicological studies can be facilitated by the use of transgenic reporters that permit the real-time, noninvasive observation of the fish. Here we report the construction and characterization of transgenic medaka lines expressing green fluorescent protein (GFP) under the control of the zebrafish neurogenin 1 (ngn1) gene promoter. Neurogenin (ngn1) is a helix-loop-helix transcription factor expressed in proliferating neuronal progenitor cells early in neuronal differentiation and plays a crucial role in directing neurogenesis. GFP expression was detected from 24 h post-fertilization until hatching, in a spatial pattern consistent with the previously reported zebrafish ngn1 expression. Temporal expression of the transgene parallels the expression profile of the endogenous medaka ngn1 transcript. Further, we demonstrate that embryos from the transgenic line permit the non-destructive, real-time screening of ngn1 promoter-directed GFP expression in a 96-well format, enabling higher throughput studies of developmental neurotoxicants. This strain has been deposited with and maintained by the National BioResource Project and is available on request ( (http://www.shigen.nig.ac.jp/medaka/strainDetailAction.do?quickSearch=true and strainId=5660)).

  15. Increased Survival of Western Corn Rootworm on Transgenic Corn Within Three Generations of Onplant Greenhouse Selection

    Science.gov (United States)

    The agricultural industry has adopted a high dose/refuge strategy as a means of delaying the onset of insect resistance to transgenic crops. Recently, Bt corn products developed for control of western corn rootworm (WCR), Diabrotica virgifera virgifera, have been introduced with less than high-dose...

  16. Transgenic biofortification of the starchy staple cassava (Manihot esculenta) generates a novel sink for protein.

    Science.gov (United States)

    Abhary, Mohammad; Siritunga, Dimuth; Stevens, Gene; Taylor, Nigel J; Fauquet, Claude M

    2011-01-25

    Although calorie dense, the starchy, tuberous roots of cassava provide the lowest sources of dietary protein within the major staple food crops (Manihot esculenta Crantz). (Montagnac JA, Davis CR, Tanumihardjo SA. (2009) Compr Rev Food Sci Food Saf 8:181-194). Cassava was genetically modified to express zeolin, a nutritionally balanced storage protein under control of the patatin promoter. Transgenic plants accumulated zeolin within de novo protein bodies localized within the root storage tissues, resulting in total protein levels of 12.5% dry weight within this tissue, a fourfold increase compared to non-transgenic controls. No significant differences were seen for morphological or agronomic characteristics of transgenic and wild type plants in the greenhouse and field trials, but relative to controls, levels of cyanogenic compounds were reduced by up to 55% in both leaf and root tissues of transgenic plants. Data described here represent a proof of concept towards the potential transformation of cassava from a starchy staple, devoid of storage protein, to one capable of supplying inexpensive, plant-based proteins for food, feed and industrial applications.

  17. Transgenic Farm Animals

    Science.gov (United States)

    Solomon, Morse B.; Eastridge, Janet S.; Paroczay, Ernest W.

    Conventional science to improve muscle and meat parameters has involved breeding strategies, such as selection of dominant traits or selection of preferred traits by cross breeding, and the use of endogenous and exogenous hormones. Improvements in the quality of food products that enter the market have largely been the result of postharvest intervention strategies. Biotechnology is a more extreme scientific method that offers the potential to improve the quality, yield, and safety of food products by direct genetic manipulation. In the December 13, 2007 issue of the Southeast Farm Press, an article by Roy Roberson pointed out that biotechnology is driving most segments of U.S. farm growth. He indicated that nationwide, the agriculture industry is booming and much of that growth is the result of biotechnology advancements.

  18. The generation of animated sequences from state transition systems.

    OpenAIRE

    Clark, Tony; Palmer, I. J.

    1995-01-01

    We address the issue of the construction of a computer animation system by composing together a collection of behaviours for the individual objects which are to be animated. The behaviours are developed using a formal notation for state transition systems. The notation is particularly flexible, allowing both a constructive and constraining aproach to behaviour representation. The behaviour necessary for animating Newton's Cradle for an arbitrary number of spheres is developed as an example of...

  19. Cellular reprogramming in farm animals: an overview of iPSC generation in the mammalian farm animal species

    OpenAIRE

    Ogorevc, J.; Orehek, S.; Dovč, P.

    2016-01-01

    Establishment of embryonic stem cell (ESC) lines has been successful in mouse and human, but not in farm animals. Development of direct reprogramming technology offers an alternative approach for generation of pluripotent stem cells, applicable also in farm animals. Induced pluripotent stem cells (iPSCs) represent practically limitless, ethically acceptable, individuum-specific source of pluripotent cells that can be generated from different types of somatic cells. iPSCs can differentiate to ...

  20. Making recombinant proteins in animals--different systems, different applications.

    Science.gov (United States)

    Dyck, Michael K; Lacroix, Dan; Pothier, François; Sirard, Marc-André

    2003-09-01

    Transgenic animal bioreactors represent a powerful tool to address the growing need for therapeutic recombinant proteins. The ability of transgenic animals to produce complex, biologically active recombinant proteins in an efficient and economic manner has stimulated a great deal of interest in this area. As a result, genetically modified animals of several species, expressing foreign proteins in various tissues, are currently being developed. However, the generation of transgenic animals is a cumbersome process and remains problematic in the application of this technology. The advantages and disadvantages of different transgenic systems in relation to other bioreactor systems are discussed.

  1. A pre-breeding screening program for transgenic boars based on fluorescence in situ hybridization assay.

    Science.gov (United States)

    Bou, Gerelchimeg; Sun, Mingju; Lv, Ming; Zhu, Jiang; Li, Hui; Wang, Juan; Li, Lu; Liu, Zhongfeng; Zheng, Zhong; He, Wenteng; Kong, Qingran; Liu, Zhonghua

    2014-08-01

    For efficient transgenic herd expansion, only the transgenic animals that possess the ability to transmit transgene into next generation are considered for breeding. However, for transgenic pig, practically lacking a pre-breeding screening program, time, labor and money is always wasted to maintain non-transgenic pigs, low or null transgenic transmission pigs and the related fruitless gestations. Developing a pre-breeding screening program would make the transgenic herd expansion more economical and efficient. In this technical report, we proposed a three-step pre-breeding screening program for transgenic boars simply through combining the fluorescence in situ hybridization (FISH) assay with the common pre-breeding screening workflow. In the first step of screening, combined with general transgenic phenotype analysis, FISH is used to identify transgenic boars. In the second step of screening, combined with conventional semen test, FISH is used to detect transgenic sperm, thus to identify the individuals producing high quality semen and transgenic sperm. In the third step of screening, FISH is used to assess the in vitro fertilization embryos, thus finally to identify the individuals with the ability to produce transgenic embryos. By this three-step screening, the non-transgenic boars and boars with no ability to produce transgenic sperm or transgenic embryos would be eliminated; therefore only those boars could produce transgenic offspring are maintained and used for breeding and herd expansion. It is the first time a systematic pre-breeding screening program is proposed for transgenic pigs. This program might also be applied in other transgenic large animals, and provide an economical and efficient strategy for herd expansion.

  2. Generation of an Oocyte-Specific Cas9 Transgenic Mouse for Genome Editing.

    Directory of Open Access Journals (Sweden)

    Linlin Zhang

    Full Text Available The CRISPR/Cas9 system has been developed as an easy-handle and multiplexable approach for engineering eukaryotic genomes by zygote microinjection of Cas9 and sgRNA, while preparing Cas9 for microinjection is laborious and introducing inconsistency into the experiment. Here, we describe a modified strategy for gene targeting through using oocyte-specific Cas9 transgenic mouse. With this mouse line, we successfully achieve precise gene targeting by injection of sgRNAs only into one-cell-stage embryos. Through comprehensive analysis, we also show allele complexity and off-target mutagenesis induced by this strategy is obviously lower than Cas9 mRNA/sgRNA injection. Thus, injection of sgRNAs into oocyte-specific Cas9 transgenic mouse embryo provides a convenient, efficient and reliable approach for mouse genome editing.

  3. Next-generation transgenic cotton: pyramiding RNAi and Bt counters insect resistance.

    Science.gov (United States)

    Ni, Mi; Ma, Wei; Wang, Xiaofang; Gao, Meijing; Dai, Yan; Wei, Xiaoli; Zhang, Lei; Peng, Yonggang; Chen, Shuyuan; Ding, Lingyun; Tian, Yue; Li, Jie; Wang, Haiping; Wang, Xiaolin; Xu, Guowang; Guo, Wangzhen; Yang, Yihua; Wu, Yidong; Heuberger, Shannon; Tabashnik, Bruce E; Zhang, Tianzhen; Zhu, Zhen

    2017-02-15

    Transgenic crops producing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) are extensively cultivated worldwide. To counter rapidly increasing pest resistance to crops that produce single Bt toxins, transgenic plant 'pyramids' producing two or more Bt toxins that kill the same pest have been widely adopted. However, cross-resistance and antagonism between Bt toxins limit the sustainability of this approach. Here we describe development and testing of the first pyramids of cotton combining protection from a Bt toxin and RNA interference (RNAi). We developed two types of transgenic cotton plants producing double-stranded RNA (dsRNA) from the global lepidopteran pest Helicoverpa armigera designed to interfere with its metabolism of juvenile hormone (JH). We focused on suppression of JH acid methyltransferase (JHAMT), which is crucial for JH synthesis, and JH-binding protein (JHBP), which transports JH to organs. In 2015 and 2016, we tested larvae from a Bt-resistant strain and a related susceptible strain of H. armigera on seven types of cotton: two controls, Bt cotton, two types of RNAi cotton (targeting JHAMT or JHBP) and two pyramids (Bt cotton plus each type of RNAi). Both types of RNAi cotton were effective against Bt-resistant insects. Bt cotton and RNAi acted independently against the susceptible strain. In computer simulations of conditions in northern China, where millions of farmers grow Bt cotton as well as abundant non-transgenic host plants of H. armigera, pyramided cotton combining a Bt toxin and RNAi substantially delayed resistance relative to using Bt cotton alone.

  4. RNA Sequencing Identifies Upregulated Kyphoscoliosis Peptidase and Phosphatidic Acid Signaling Pathways in Muscle Hypertrophy Generated by Transgenic Expression of Myostatin Propeptide

    OpenAIRE

    Yuanxin Miao; Jinzeng Yang; Zhong Xu; Lu Jing; Shuhong Zhao; Xinyun Li

    2015-01-01

    Myostatin (MSTN), a member of the transforming growth factor-β superfamily, plays a crucial negative role in muscle growth. MSTN mutations or inhibitions can dramatically increase muscle mass in most mammal species. Previously, we generated a transgenic mouse model of muscle hypertrophy via the transgenic expression of the MSTN N-terminal propeptide cDNA under the control of the skeletal muscle-specific MLC1 promoter. Here, we compare the mRNA profiles between transgenic mice and wild-type li...

  5. RNA Sequencing Identifies Upregulated Kyphoscoliosis Peptidase and Phosphatidic Acid Signaling Pathways in Muscle Hypertrophy Generated by Transgenic Expression of Myostatin Propeptide

    OpenAIRE

    Yuanxin Miao; Jinzeng Yang; Zhong Xu; Lu Jing; Shuhong Zhao; Xinyun Li

    2015-01-01

    Myostatin (MSTN), a member of the transforming growth factor-β superfamily, plays a crucial negative role in muscle growth. MSTN mutations or inhibitions can dramatically increase muscle mass in most mammal species. Previously, we generated a transgenic mouse model of muscle hypertrophy via the transgenic expression of the MSTN N-terminal propeptide cDNA under the control of the skeletal muscle-specific MLC1 promoter. Here, we compare the mRNA profiles between transgenic mice and wild-type li...

  6. Ethical issues in transgenics.

    Science.gov (United States)

    Sherlock, R; Morrey, J D

    2000-01-01

    The arguments of critics and concerns of the public on generating transgenic cloned animals are analyzed for the absence or presence of logical structure. Critics' arguments are symbolically compared with "genetic trespassing," "genetic speeding," or "going the wrong way," and responses are provided to these arguments. Scientists will be empowered to participate in the public discussion and to engage the critics on these issues as they consider thoughtful, plausible responses to their concerns. Temporary moratoriums are recognized as a plausible approach to dealing with possible concerns of new scientific advancements.

  7. Subroutine functions for generating animation data; Animation yo gazo data seisei subroutine kino ni tsuite

    Energy Technology Data Exchange (ETDEWEB)

    Suematsu, K. [National Aerospace Laboratory, Tokyo (Japan)

    1992-04-01

    Animations play an important role as a means to visualize results of computer numerical simulations. This report describes various subroutine functions for animations developed for the purpose of arranging software for animations. The functions are as follows; a graphic-data transforming function, a through-vision displaying function, an image-creating function for animations, and a logograph and character data output function. The graphic-data transforming function enables such processings as clipping, shading, and through-vision transformaing, as well as transformation processing to rotate, transfer, enlarge, or reduce polygonal data. Subroutines for preparing data of oil flowchart and streamline chart, for line drawing, and for creating grid diagram for scope are provided as subroutines for creating animations. These subroutines are applied to the simulation of HOPE, a spacecraft, to verify their effectiveness. 5 refs., 9 figs., 4 tabs.

  8. Subroutine functions for generating animation data. Animation yo gazo data seisei subroutine kino ni tsuite

    Energy Technology Data Exchange (ETDEWEB)

    Suematsu, K. (National Aerospace Laboratory, Tokyo (Japan))

    1992-04-01

    Animations play an important role as a means to visualize results of computer numerical simulations. This report describes various subroutine functions for animations developed for the purpose of arranging software for animations. The functions are as follows; a graphic-data transforming function, a through-vision displaying function, an image-creating function for animations, and a logograph and character data output function. The graphic-data transforming function enables such processings as clipping, shading, and through-vision transformaing, as well as transformation processing to rotate, transfer, enlarge, or reduce polygonal data. Subroutines for preparing data of oil flowchart and streamline chart, for line drawing, and for creating grid diagram for scope are provided as subroutines for creating animations. These subroutines are applied to the simulation of HOPE, a spacecraft, to verify their effectiveness. 5 refs., 9 figs., 4 tabs.

  9. A high efficiency technique for the generation of transgenic sugar beets from stomatal guard cells.

    Science.gov (United States)

    Hall, R D; Riksen-Bruinsma, T; Weyens, G J; Rosquin, I J; Denys, P N; Evans, I J; Lathouwers, J E; Lefèbvre, M P; Dunwell, J M; van Tunen, A; Krens, F A

    1996-09-01

    An optimized protocol has been developed for the efficient and rapid genetic modification of sugar beet (Beta vulgaris L.). A polyethylene glycol-mediated DNA transformation technique could be applied to protoplast populations enriched specifically for a single totipotent cell type derived from stomatal guard cells, to achieve high transformation frequencies. Bialaphos resistance, conferred by the pat gene, produced a highly efficient selection system. The majority of plants were obtained within 8 to 9 weeks and were appropriate for plant breeding purposes. All were resistant to glufosinate-ammonium-based herbicides. Detailed genomic characterization has verified transgene integration, and progeny analysis showed Mendelian inheritance.

  10. Quantitative analysis of tetracycline-inducible expression of the green fluorescent protein gene in transgenic chickens.

    Science.gov (United States)

    Koo, Bon Chul; Kwon, Mo Sun; Roh, Ji Yeol; Kim, Minjee; Kim, Jin-Hoi; Kim, Teoan

    2012-01-01

    The use of transgenic farm animals as "bioreactors" to address the growing demand for biopharmaceuticals, both in terms of increased quantity and greater number, represents a key development in the advancement of medical science. However, the potential for detrimental side-effects as a result of uncontrolled constitutive expression of foreign genes in transgenic animals is a well-recognized limitation of such systems. Previously, using a tetracycline-inducible expression system, we demonstrated the induction of expression of a transgene encoding green fluorescent protein (GFP) in transgenic chickens by feeding with doxycycline, a tetracycline derivative; expression of GFP reverted to pre-induction levels when the inducer was removed from the diet. As a proof of principle study, however, quantitative assessment of expression was not possible, as only one G0 and one G1 transgenic chicken was obtained. In the current study, a sufficient number of G2 and G3 transgenic chickens were obtained, and quantification analysis demonstrated up to a 20-fold induction of expression by doxycycline. In addition, stable transmission of the transgene without any apparent genetic modifications was observed through several generations. The use of an inducible expression system that can be regulated by dietary supplementation could help mitigate the physiological disruption that can occur in transgenic animals as a result of uncontrolled constitutive expression of a transgene. Importantly, these results also support the use of the retroviral system for generating transgenic animals with minimal risk in terms of biosafety.

  11. Animation Model to Conceptualize ATP Generation: A Mitochondrial Oxidative Phosphorylation

    Science.gov (United States)

    Jena, Ananta Kumar

    2015-01-01

    Adenosine triphosphate (ATP) is the molecular unit of intracellular energy and it is the product of oxidative phosphorylation of cellular respiration uses in cellular processes. The study explores the growth of the misconception levels amongst the learners and evaluates the effectiveness of animation model over traditional methods. The data…

  12. Transgene Stacking in Cotton Improvement

    Institute of Scientific and Technical Information of China (English)

    YANG Ye-hua; WANG Xue-kui; YAO Ming-jing; FAN Yu-peng; GAO Da-yu

    2008-01-01

    @@ To date,more and more transgenic varieties of upland cotton (Gossypium hirsuturn L.) generated with transgenes,which derived from varies of alien species,are playing important role in agricultural production.Stacking of multi-transgenes has a potential for combining all the merits of distinct transgenic lines in a cultivar and possibly makes a significant contribution to cultivar improvement.

  13. Generation of stable 'low phytic acid' transgenic rice through antisense repression of the 1D-myo-inositol 3-phosphate synthase gene (RINO1) using the 18-kDa oleosin promoter.

    Science.gov (United States)

    Kuwano, Mio; Mimura, Tetsuro; Takaiwa, Fumio; Yoshida, Kaoru T

    2009-01-01

    Phytic acid acts as the major storage form of phosphorus in plant seeds and is poorly digested by monogastric animals. The degradation of phytic acid in animal diets is necessary to overcome both environmental and nutritional issues. The enzyme 1D-myo-inositol 3-phosphate [Ins(3)P(1)] synthase (EC 5.5.1.4) catalyses the first step of myo-inositol biosynthesis and directs phytic acid biosynthesis in seeds. The rice Ins(3)P(1) synthase gene (RINO1) is highly expressed in developing seed embryos and in the aleurone layer, where phytic acid is synthesized and stored. In rice seeds, 18-kDa oleosin (Ole18) is expressed in a seed-specific manner, and its transcripts are restricted to the embryo and the aleurone layer. Therefore, to effectively suppress phytic acid biosynthesis, antisense RINO1 cDNA was expressed under the control of the Ole18 promoter, directing the same spatial pattern in seeds as RINO1 in transgenic rice plants. The generated transgenic rice plants showed strong 'low phytic acid' (lpa) phenotypes, in which seed phytic acid was reduced by 68% and free available phosphate was concomitantly increased. No negative effects on seed weight, germination or plant growth were observed. The available phosphate levels of the stable transgenic plants surpassed those of currently available rice lpa mutants.

  14. Reactivation of Endogenous Genes and Epigenetic Remodeling Are Barriers for Generating Transgene-Free Induced Pluripotent Stem Cells in Pig.

    Directory of Open Access Journals (Sweden)

    Kwang-Hwan Choi

    Full Text Available Cellular reprogramming of committed cells into a pluripotent state can be induced by ectopic expression of genes such as OCT4, SOX2, KLF4, and MYC. Reprogrammed cells can be maintained by activating endogenous pluripotent networks without transgene expression. Although various research groups have attempted to generate pig induced pluripotent stem cells (iPSCs, authentic iPSCs have not be obtained, instead showing dependence on transgene expression. In this study, iPSCs were derived from porcine fetal fibroblasts via drug-inducible vectors carrying human transcription factors (OCT4, SOX2, KLF4, and MYC. Therefore, this study investigated characteristics of iPSCs and reprogramming mechanisms in pig. The iPSCs were stably maintained over an extended period with potential in vitro differentiation into three germ layers. In addition, the pluripotent state of iPSCs was regulated by modulating culture conditions. They showed naive- or primed-like pluripotent states in LIF or bFGF supplemented culture conditions, respectively. However, iPSCs could not be maintained without ectopic expression of transgenes. The cultured iPSCs expressed endogenous transcription factors such as OCT4 and SOX2, but not NANOG (a known gateway to complete reprogramming. Endogenous genes related to mesenchymal-to-epithelial transition (DPPA2, CDH1, EPCAM, and OCLN were not sufficiently reactivated, as measured by qPCR. DNA methylation analysis for promoters of OCT4, NANOG, and XIST showed that epigenetic reprogramming did not occur in female iPSCs. Based on our results, expression of exogenous genes could not sufficiently activate the essential endogenous genes and remodel the epigenetic milieu to achieve faithful pluripotency in pig. Accordingly, investigating iPSCs could help us improve and develop reprogramming methods by understanding reprogramming mechanisms in pig.

  15. Identification of the MUC2 Promoter as a Strong Promoter for Intestinal Gene Expression through Generation of Transgenic Quail Expressing GFP in Gut Epithelial Cells

    Directory of Open Access Journals (Sweden)

    Rachel M. Woodfint

    2017-01-01

    Full Text Available Identification of tissue- and stage-specific gene promoters is valuable for delineating the functional roles of specific genes in genetically engineered animals. Here, through the comparison of gene expression in different tissues by analysis of a microarray database, the intestinal specificity of mucin 2 (MUC2 expression was identified in mice and humans, and further confirmed in chickens by RT-PCR (reverse transcription-PCR analysis. An analysis of cis-acting elements in avian MUC2 gene promoters revealed conservation of binding sites, within a 2.9 kb proximal promoter region, for transcription factors such as caudal type homeobox 2 (CDX2, GATA binding protein 4 (GATA4, hepatocyte nuclear factor 4 α (HNF4A, and transcription factor 4 (TCF4 that are important for maintaining intestinal homeostasis and functional integrity. By generating transgenic quail, we demonstrated that the 2.9 kb chicken MUC2 promoter could drive green fluorescent protein (GFP reporter expression exclusively in the small intestine, large intestine, and ceca. Fluorescence image analysis further revealed GFP expression in intestine epithelial cells. The GFP expression was barely detectable in the embryonic intestine, but increased during post-hatch development. The spatiotemporal expression pattern of the reporter gene confirmed that the 2.9 kb MUC2 promoter could retain the regulatory element to drive expression of target genes in intestinal tissues after hatching. This new transgene expression system, using the MUC2 promoter, will provide a new method of overexpressing target genes to study gene function in the avian intestine.

  16. Identification of the MUC2 Promoter as a Strong Promoter for Intestinal Gene Expression through Generation of Transgenic Quail Expressing GFP in Gut Epithelial Cells

    Science.gov (United States)

    Woodfint, Rachel M.; Chen, Paula R.; Ahn, Jinsoo; Suh, Yeunsu; Hwang, Seongsoo; Lee, Sang Suk; Lee, Kichoon

    2017-01-01

    Identification of tissue- and stage-specific gene promoters is valuable for delineating the functional roles of specific genes in genetically engineered animals. Here, through the comparison of gene expression in different tissues by analysis of a microarray database, the intestinal specificity of mucin 2 (MUC2) expression was identified in mice and humans, and further confirmed in chickens by RT-PCR (reverse transcription-PCR) analysis. An analysis of cis-acting elements in avian MUC2 gene promoters revealed conservation of binding sites, within a 2.9 kb proximal promoter region, for transcription factors such as caudal type homeobox 2 (CDX2), GATA binding protein 4 (GATA4), hepatocyte nuclear factor 4 α (HNF4A), and transcription factor 4 (TCF4) that are important for maintaining intestinal homeostasis and functional integrity. By generating transgenic quail, we demonstrated that the 2.9 kb chicken MUC2 promoter could drive green fluorescent protein (GFP) reporter expression exclusively in the small intestine, large intestine, and ceca. Fluorescence image analysis further revealed GFP expression in intestine epithelial cells. The GFP expression was barely detectable in the embryonic intestine, but increased during post-hatch development. The spatiotemporal expression pattern of the reporter gene confirmed that the 2.9 kb MUC2 promoter could retain the regulatory element to drive expression of target genes in intestinal tissues after hatching. This new transgene expression system, using the MUC2 promoter, will provide a new method of overexpressing target genes to study gene function in the avian intestine. PMID:28106824

  17. Generation of Triple-Transgenic Forsythia Cell Cultures as a Platform for the Efficient, Stable, and Sustainable Production of Lignans.

    Science.gov (United States)

    Murata, Jun; Matsumoto, Erika; Morimoto, Kinuyo; Koyama, Tomotsugu; Satake, Honoo

    2015-01-01

    Sesamin is a furofuran lignan biosynthesized from the precursor lignan pinoresinol specifically in sesame seeds. This lignan is shown to exhibit anti-hypertensive activity, protect the liver from damages by ethanol and lipid oxidation, and reduce lung tumor growth. Despite rapidly elevating demand, plant sources of lignans are frequently limited because of the high cost of locating and collecting plants. Indeed, the acquisition of sesamin exclusively depends on the conventional extraction of particular Sesamum seeds. In this study, we have created the efficient, stable and sustainable sesamin production system using triple-transgenic Forsythia koreana cell suspension cultures, U18i-CPi-Fk. These transgenic cell cultures were generated by stably introducing an RNAi sequence against the pinoresinol-glucosylating enzyme, UGT71A18, into existing CPi-Fk cells, which had been created by introducing Sesamum indicum sesamin synthase (CYP81Q1) and an RNA interference (RNAi) sequence against pinoresinol/lariciresinol reductase (PLR) into F. koreanna cells. Compared to its transgenic prototype, U18i-CPi-Fk displayed 5-fold higher production of pinoresinol aglycone and 1.4-fold higher production of sesamin, respectively, while the wildtype cannot produce sesamin due to a lack of any intrinsic sesamin synthase. Moreover, red LED irradiation of U18i-CPi-Fk specifically resulted in 3.0-fold greater production in both pinoresinol aglycone and sesamin than production of these lignans under the dark condition, whereas pinoresinol production was decreased in the wildtype under red LED. Moreover, we developed a procedure for sodium alginate-based long-term storage of U18i-CPi-Fk in liquid nitrogen. Production of sesamin in U18i-CPi-Fk re-thawed after six-month cryopreservation was equivalent to that of non-cryopreserved U18i-CPi-Fk. These data warrant on-demand production of sesamin anytime and anywhere. Collectively, the present study provides evidence that U18i-CP-Fk is an

  18. Generation of Triple-Transgenic Forsythia Cell Cultures as a Platform for the Efficient, Stable, and Sustainable Production of Lignans.

    Directory of Open Access Journals (Sweden)

    Jun Murata

    Full Text Available Sesamin is a furofuran lignan biosynthesized from the precursor lignan pinoresinol specifically in sesame seeds. This lignan is shown to exhibit anti-hypertensive activity, protect the liver from damages by ethanol and lipid oxidation, and reduce lung tumor growth. Despite rapidly elevating demand, plant sources of lignans are frequently limited because of the high cost of locating and collecting plants. Indeed, the acquisition of sesamin exclusively depends on the conventional extraction of particular Sesamum seeds. In this study, we have created the efficient, stable and sustainable sesamin production system using triple-transgenic Forsythia koreana cell suspension cultures, U18i-CPi-Fk. These transgenic cell cultures were generated by stably introducing an RNAi sequence against the pinoresinol-glucosylating enzyme, UGT71A18, into existing CPi-Fk cells, which had been created by introducing Sesamum indicum sesamin synthase (CYP81Q1 and an RNA interference (RNAi sequence against pinoresinol/lariciresinol reductase (PLR into F. koreanna cells. Compared to its transgenic prototype, U18i-CPi-Fk displayed 5-fold higher production of pinoresinol aglycone and 1.4-fold higher production of sesamin, respectively, while the wildtype cannot produce sesamin due to a lack of any intrinsic sesamin synthase. Moreover, red LED irradiation of U18i-CPi-Fk specifically resulted in 3.0-fold greater production in both pinoresinol aglycone and sesamin than production of these lignans under the dark condition, whereas pinoresinol production was decreased in the wildtype under red LED. Moreover, we developed a procedure for sodium alginate-based long-term storage of U18i-CPi-Fk in liquid nitrogen. Production of sesamin in U18i-CPi-Fk re-thawed after six-month cryopreservation was equivalent to that of non-cryopreserved U18i-CPi-Fk. These data warrant on-demand production of sesamin anytime and anywhere. Collectively, the present study provides evidence that U18i

  19. Characterization and multi-generational stability of the growth hormone transgene (EO-1alpha) responsible for enhanced growth rates in Atlantic Salmon.

    Science.gov (United States)

    Yaskowiak, Edward S; Shears, Margaret A; Agarwal-Mawal, Alka; Fletcher, Garth L

    2006-08-01

    Transgenic technologies provide a promising means by which desirable traits can be introduced into cultured fish species within a single generation thus accelerating the production of genetically superior broodstock for aquaculture. However, before such fish are allowed to be marketed as food they must receive government regulatory approval. Two pivotal regulatory requirements are: (1) complete characterization of the genomically integrated transgene and, (2) demonstration that the transgene remains stable over multiple generations. We have generated a stable line of growth hormone (GH) transgenic Atlantic salmon (Salmo salar) using an "all fish" gene construct (opAFP-GHc2) containing a growth hormone cDNA from chinook salmon whose expression is regulated by the 5' promoter and 3' termination regions derived from an ocean pout antifreeze protein (AFP) gene. In this study we show that a reorganized form of the opAFP-GHc2 construct (termed EO-1alpha) integrated as a single functional copy into a 35 bp repeat region of the genomic DNA. PCR based mapping revealed that the linear sequence of the EO-1alpha integrant was organized as follows: base pairs 1580-2193 of the ocean pout promoter region followed by the intact chinook salmon GH cDNA, the complete ocean pout antifreeze 3' region, and the first 1678 bp of the ocean pout antifreeze 5' region. Sequence analysis of the EO-1alpha integrant and genomic flanking regions in F2 and F4 generation salmon revealed that they were identical. In addition, apart from the disruption at the integration sites, the consensus sequences of the integrant in these two generations of salmon were identical to the sequence of the opAFP-GHc2 construct. These results indicate that the EO-1alpha transgene codes for the chinook salmon GH, and that the transgene and the integration site have remained stable over multiple generations.

  20. Generation of Peroxisome-Deficient Somatic Animal Cell Mutants.

    Science.gov (United States)

    Okumoto, Kanji; Fujiki, Yukio

    2017-01-01

    Cell mutants with a genetic defect affecting various cellular phenotypes are widely utilized as a powerful tool in genetic, biochemical, and cell biological research. More than a dozen complementation groups of animal somatic mutant cells defective in peroxisome biogenesis have been successfully isolated in Chinese hamster ovary (CHO) cells and used as a model system reflecting fatal human severe genetic disorders named peroxisome biogenesis disorders (PBD). Isolation and characterization of peroxisome-deficient CHO cell mutants has allowed the identification of PEX genes and the gene products peroxins, which directly leads to the accomplishment of isolation of pathogenic genes responsible for human PBDs, as well as elucidation of their functional roles in peroxisome biogenesis. Here, we describe the procedure to isolate peroxisome-deficient mammalian cell mutants from CHO cells, by making use of an effective, photo-sensitized selection method.

  1. mlo-based powdery mildew resistance in hexaploid bread wheat generated by a non-transgenic TILLING approach.

    Science.gov (United States)

    Acevedo-Garcia, Johanna; Spencer, David; Thieron, Hannah; Reinstädler, Anja; Hammond-Kosack, Kim; Phillips, Andrew L; Panstruga, Ralph

    2017-03-01

    Wheat is one of the most widely grown cereal crops in the world and is an important food grain source for humans. However, wheat yields can be reduced by many abiotic and biotic stress factors, including powdery mildew disease caused by Blumeria graminis f.sp. tritici (Bgt). Generating resistant varieties is thus a major effort in plant breeding. Here, we took advantage of the non-transgenic Targeting Induced Lesions IN Genomes (TILLING) technology to select partial loss-of-function alleles of TaMlo, the orthologue of the barley Mlo (Mildew resistance locus o) gene. Natural and induced loss-of-function alleles (mlo) of barley Mlo are known to confer durable broad-spectrum powdery mildew resistance, typically at the expense of pleiotropic phenotypes such as premature leaf senescence. We identified 16 missense mutations in the three wheat TaMlo homoeologues, TaMlo-A1, TaMlo-B1 and TaMlo-D1 that each lead to single amino acid exchanges. Using transient gene expression assays in barley single cells, we functionally analysed the different missense mutants and identified the most promising candidates affecting powdery mildew susceptibility. By stacking of selected mutant alleles we generated four independent lines with non-conservative mutations in each of the three TaMlo homoeologues. Homozygous triple mutant lines and surprisingly also some of the homozygous double mutant lines showed enhanced, yet incomplete, Bgt resistance without the occurrence of discernible pleiotropic phenotypes. These lines thus represent an important step towards the production of commercial non-transgenic, powdery mildew-resistant bread wheat varieties.

  2. A developmentally regulated Cre-lox system to generate marker-free transgenic Brassica napus plants.

    Science.gov (United States)

    Kopertekh, Lilya; Broer, Inge; Schiemann, Joachim

    2012-01-01

    In this chapter, a strategy for engineering marker-free Brassica napus plants is described. It is based on the Cre-lox site-specific recombination system and includes three essential steps. At first, the binary vector pLH-nap-lx-cre-35S-bar-lx-vst has been designed. In this vector, the cre gene and the bar expression cassette are flanked by two lox sites in direct orientation. The lox-flanked sequence is placed between a seed-specific napin promoter and a coding region for the vstI gene. At the second step, the cre-bar vector was transferred into B. napus hypocotyl explants by Agrobacterium tumefaciens-mediated transformation. Finally, T1 progeny was tested for excision of the marker gene at phenotypic and molecular levels. PCR, sequencing, and Southern blot analysis confirmed complete and precise deletion of the lox-flanked DNA region. This developmentally regulated Cre-lox system can be applied to remove undesirable DNA in transgenic plants propagated by seeds.

  3. The potential impact of new generation transgenic methods on creating rabbit models of cardiac diseases.

    Science.gov (United States)

    Bősze, Z; Major, P; Baczkó, I; Odening, K E; Bodrogi, L; Hiripi, L; Varró, A

    2016-07-01

    Since the creation of the first transgenic rabbit thirty years ago, pronuclear microinjection remained the single applied method and resulted in numerous important rabbit models of human diseases, including cardiac deficiencies, albeit with low efficiency. For additive transgenesis a novel transposon mediated method, e.g., the Sleeping Beauty transgenesis, increased the efficiency, and its application to create cardiac disease models is expected in the near future. The targeted genome engineering nuclease family, e.g., the zink finger nuclease (ZFN), the transcription activator-like effector nuclease (TALEN) and the newest, clustered regularly interspaced short palindromic repeats (CRISPR) with the CRISPR associated effector protein (CAS), revolutionized the non-mouse transgenesis. The latest gene-targeting technology, the CRISPR/CAS system, was proven to be efficient in rabbit to create multi-gene knockout models. In the future, the number of tailor-made rabbit models produced with one of the above mentioned methods is expected to exponentially increase and to provide adequate models of heart diseases.

  4. A strategy for the generation, characterization and distribution of animal models by The Michael J. Fox Foundation for Parkinson’s Research

    Directory of Open Access Journals (Sweden)

    Marco A. S. Baptista

    2013-11-01

    Full Text Available Progress in Parkinson’s disease (PD research and therapeutic development is hindered by many challenges, including a need for robust preclinical animal models. Limited availability of these tools is due to technical hurdles, patent issues, licensing restrictions and the high costs associated with generating and distributing these animal models. Furthermore, the lack of standardization of phenotypic characterization and use of varying methodologies has made it difficult to compare outcome measures across laboratories. In response, The Michael J. Fox Foundation for Parkinson’s Research (MJFF is directly sponsoring the generation, characterization and distribution of preclinical rodent models, enabling increased access to these crucial tools in order to accelerate PD research. To date, MJFF has initiated and funded the generation of 30 different models, which include transgenic or knockout models of PD-relevant genes such as Park1 (also known as Park4 and SNCA, Park8 (LRRK2, Park7 (DJ-1, Park6 (PINK1, Park2 (Parkin, VPS35, EiF4G1 and GBA. The phenotypic characterization of these animals is performed in a uniform and streamlined manner at independent contract research organizations. Finally, MJFF created a central repository at The Jackson Laboratory (JAX that houses both non-MJFF and MJFF-generated preclinical animal models. Funding from MJFF, which subsidizes the costs involved in transfer, rederivation and colony expansion, has directly resulted in over 2500 rodents being distributed to the PD community for research use.

  5. Mutant human FUS Is ubiquitously mislocalized and generates persistent stress granules in primary cultured transgenic zebrafish cells.

    Science.gov (United States)

    Acosta, Jamie Rae; Goldsbury, Claire; Winnick, Claire; Badrock, Andrew P; Fraser, Stuart T; Laird, Angela S; Hall, Thomas E; Don, Emily K; Fifita, Jennifer A; Blair, Ian P; Nicholson, Garth A; Cole, Nicholas J

    2014-01-01

    FUS mutations can occur in familial amyotrophic lateral sclerosis (fALS), a neurodegenerative disease with cytoplasmic FUS inclusion bodies in motor neurons. To investigate FUS pathology, we generated transgenic zebrafish expressing GFP-tagged wild-type or fALS (R521C) human FUS. Cell cultures were made from these zebrafish and the subcellular localization of human FUS and the generation of stress granule (SG) inclusions examined in different cell types, including differentiated motor neurons. We demonstrate that mutant FUS is mislocalized from the nucleus to the cytosol to a similar extent in motor neurons and all other cell types. Both wild-type and R521C FUS localized to SGs in zebrafish cells, demonstrating an intrinsic ability of human FUS to accumulate in SGs irrespective of the presence of disease-associated mutations or specific cell type. However, elevation in relative cytosolic to nuclear FUS by the R521C mutation led to a significant increase in SG assembly and persistence within a sub population of vulnerable cells, although these cells were not selectively motor neurons.

  6. Mutant human FUS Is ubiquitously mislocalized and generates persistent stress granules in primary cultured transgenic zebrafish cells.

    Directory of Open Access Journals (Sweden)

    Jamie Rae Acosta

    Full Text Available FUS mutations can occur in familial amyotrophic lateral sclerosis (fALS, a neurodegenerative disease with cytoplasmic FUS inclusion bodies in motor neurons. To investigate FUS pathology, we generated transgenic zebrafish expressing GFP-tagged wild-type or fALS (R521C human FUS. Cell cultures were made from these zebrafish and the subcellular localization of human FUS and the generation of stress granule (SG inclusions examined in different cell types, including differentiated motor neurons. We demonstrate that mutant FUS is mislocalized from the nucleus to the cytosol to a similar extent in motor neurons and all other cell types. Both wild-type and R521C FUS localized to SGs in zebrafish cells, demonstrating an intrinsic ability of human FUS to accumulate in SGs irrespective of the presence of disease-associated mutations or specific cell type. However, elevation in relative cytosolic to nuclear FUS by the R521C mutation led to a significant increase in SG assembly and persistence within a sub population of vulnerable cells, although these cells were not selectively motor neurons.

  7. Human lysozyme expressed in the mammary gland of transgenic dairy goats can inhibit the growth of bacteria that cause mastitis and the cold-spoilage of milk.

    Science.gov (United States)

    Maga, Elizabeth A; Cullor, James S; Smith, Wayne; Anderson, Gary B; Murray, James D

    2006-01-01

    The addition of human milk components with intrinsic antimicrobial activity to livestock milk by genetic engineering has the potential to benefit milk safety and production as well as the health of the lactating animal. As a model for the dairy cow, we generated transgenic goats that expressed human lysozyme in their milk at 68% of the levels found in human milk. Milk from these transgenic animals had a bacteriostatic effect on both in vitro and in vivo growth of several microorganisms important to the dairy industry. In vitro, milk from transgenic animals was capable of slowing the growth of mastitis-causing strains of Escherichia coli (P transgenic animals. In vivo, milk from transgenic animals supported less bacterial growth than control milk. This transgenic model demonstrates the possibilities offered by genetic engineering to enhance the antimicrobial nature of milk and the udder.

  8. Generation of transgenic wheat resistant to wheat yel-low mosaic virus and identifi-cation of gene silence induced by virus infection

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The plasmid containing the promoter Act1, the coat protein (cp) gene of wheat yellow mosaic virus (WYMV) and the selectable bar gene, was delivered via particle bombardment, directly into immature embryos of a wheat cultivars. PCR and PCR-RFLP were employed to screen the existence of the cp gene in T0 and T1 generations. Seeds from the positive T1 plants were sowed in fields heavily contaminated with WYMV to detect their resistance. In field trial of virus infection, one of the transgenic wheat lines, P8-T2, exhibited highly disease-resistance. Western blot and RT-PCR analysis showed that the expression level of cp gene in the resistant transgenic line was reduced greatly compared to those susceptible to WYMV infection. This provided evidence to presume that the resistance obtained by the transgenic wheat line was stimulated by the mechanism of the virus induced gene silencing.

  9. Efficient generation of knock-in transgenic zebrafish carrying reporter/driver genes by CRISPR/Cas9-mediated genome engineering.

    Science.gov (United States)

    Kimura, Yukiko; Hisano, Yu; Kawahara, Atsuo; Higashijima, Shin-ichi

    2014-10-08

    The type II bacterial CRISPR/Cas9 system is rapidly becoming popular for genome-engineering due to its simplicity, flexibility, and high efficiency. Recently, targeted knock-in of a long DNA fragment via homology-independent DNA repair has been achieved in zebrafish using CRISPR/Cas9 system. This raised the possibility that knock-in transgenic zebrafish could be efficiently generated using CRISPR/Cas9. However, how widely this method can be applied for the targeting integration of foreign genes into endogenous genomic loci is unclear. Here, we report efficient generation of knock-in transgenic zebrafish that have cell-type specific Gal4 or reporter gene expression. A donor plasmid containing a heat-shock promoter was co-injected with a short guide RNA (sgRNA) targeted for genome digestion, a sgRNA targeted for donor plasmid digestion, and Cas9 mRNA. We have succeeded in establishing stable knock-in transgenic fish with several different constructs for 4 genetic loci at a frequency being exceeding 25%. Due to its simplicity, design flexibility, and high efficiency, we propose that CRISPR/Cas9-mediated knock-in will become a standard method for the generation transgenic zebrafish.

  10. RNA sequencing identifies upregulated kyphoscoliosis peptidase and phosphatidic acid signaling pathways in muscle hypertrophy generated by transgenic expression of myostatin propeptide.

    Science.gov (United States)

    Miao, Yuanxin; Yang, Jinzeng; Xu, Zhong; Jing, Lu; Zhao, Shuhong; Li, Xinyun

    2015-04-09

    Myostatin (MSTN), a member of the transforming growth factor-β superfamily, plays a crucial negative role in muscle growth. MSTN mutations or inhibitions can dramatically increase muscle mass in most mammal species. Previously, we generated a transgenic mouse model of muscle hypertrophy via the transgenic expression of the MSTN N-terminal propeptide cDNA under the control of the skeletal muscle-specific MLC1 promoter. Here, we compare the mRNA profiles between transgenic mice and wild-type littermate controls with a high-throughput RNA sequencing method. The results show that 132 genes were significantly differentially expressed between transgenic mice and wild-type control mice; 97 of these genes were up-regulated, and 35 genes were down-regulated in the skeletal muscle. Several genes that had not been reported to be involved in muscle hypertrophy were identified, including up-regulated myosin binding protein H (mybph), and zinc metallopeptidase STE24 (Zmpste24). In addition, kyphoscoliosis peptidase (Ky), which plays a vital role in muscle growth, was also up-regulated in the transgenic mice. Interestingly, a pathway analysis based on grouping the differentially expressed genes uncovered that cardiomyopathy-related pathways and phosphatidic acid (PA) pathways (Dgki, Dgkz, Plcd4) were up-regulated. Increased PA signaling may increase mTOR signaling, resulting in skeletal muscle growth. The findings of the RNA sequencing analysis help to understand the molecular mechanisms of muscle hypertrophy caused by MSTN inhibition.

  11. RNA Sequencing Identifies Upregulated Kyphoscoliosis Peptidase and Phosphatidic Acid Signaling Pathways in Muscle Hypertrophy Generated by Transgenic Expression of Myostatin Propeptide

    Directory of Open Access Journals (Sweden)

    Yuanxin Miao

    2015-04-01

    Full Text Available Myostatin (MSTN, a member of the transforming growth factor-β superfamily, plays a crucial negative role in muscle growth. MSTN mutations or inhibitions can dramatically increase muscle mass in most mammal species. Previously, we generated a transgenic mouse model of muscle hypertrophy via the transgenic expression of the MSTN N-terminal propeptide cDNA under the control of the skeletal muscle-specific MLC1 promoter. Here, we compare the mRNA profiles between transgenic mice and wild-type littermate controls with a high-throughput RNA sequencing method. The results show that 132 genes were significantly differentially expressed between transgenic mice and wild-type control mice; 97 of these genes were up-regulated, and 35 genes were down-regulated in the skeletal muscle. Several genes that had not been reported to be involved in muscle hypertrophy were identified, including up-regulated myosin binding protein H (mybph, and zinc metallopeptidase STE24 (Zmpste24. In addition, kyphoscoliosis peptidase (Ky, which plays a vital role in muscle growth, was also up-regulated in the transgenic mice. Interestingly, a pathway analysis based on grouping the differentially expressed genes uncovered that cardiomyopathy-related pathways and phosphatidic acid (PA pathways (Dgki, Dgkz, Plcd4 were up-regulated. Increased PA signaling may increase mTOR signaling, resulting in skeletal muscle growth. The findings of the RNA sequencing analysis help to understand the molecular mechanisms of muscle hypertrophy caused by MSTN inhibition.

  12. Generative Technologies for Model Animation in the TopCased Platform

    Science.gov (United States)

    Crégut, Xavier; Combemale, Benoit; Pantel, Marc; Faudoux, Raphaël; Pavei, Jonatas

    Domain Specific Modeling Languages (DSML) are more and more used to handle high level concepts, and thus bring complex software development under control. The increasingly recurring definition of new languages raises the problem of the definition of support tools such as editor, simulator, compiler, etc. In this paper we propose generative technologies that have been designed to ease the development of model animation tools inside the TopCased platform. These tools rely on the automatically generated graphical editors of TopCased and provide additional generators for building model animator graphical interface. We also rely on an architecture for executable metamodel (i.e., the TopCased model execution metamodeling pattern) to bind the behavioral semantics of the modeling language. These tools were designed in a pragmatic manner by abstracting the various model animators that had been hand-coded in the TopCased project, and then validated by refactoring these animators.

  13. Bone Marrow Transplantation in Mice as a Tool to Generate Genetically Modified Animals

    Science.gov (United States)

    Rőszer, Tamás; Pintye, Éva; Benkő, Ilona

    2008-12-01

    Transgenic mice can be used either as models of known inherited human diseases or can be applied to perform phenotypic tests of genes with unknown function. In some special applications of gene modification we have to create a tissue specific mutation of a given gene. In some cases however the gene modification can be lethal in the intrauterine life, therefore we should engraft the mutated cells in the postnatal life period. After total body irradiation transplantation of bone marrow cells can be a solution to introduce mutant hematopoietic stem cells into a mature animal. Bone marrow transplantation is a useful and novel tool to study the role of hematopoietic cells in the pathogenesis of inflammation, autoimmune syndromes and many metabolic alterations coupled recently to leukocyte functions.

  14. Transgene Stacking in Cotton Improvement

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    To date,more and more transgenic varieties of upland cotton(Gossypium hirsutum L.) generated with transgenes,which derived from varies of alien species,are playing important role in agricultural production.Stacking of multi-transgenes has a potential for combining all the merits of distinct

  15. Acetylcholinesterase (AChE) gene modification in transgenic animals: functional consequences of selected exon and regulatory region deletion.

    Science.gov (United States)

    Camp, Shelley; Zhang, Limin; Marquez, Michael; de la Torre, Brian; Long, Jeffery M; Bucht, Goran; Taylor, Palmer

    2005-12-15

    AChE is an alternatively spliced gene. Exons 2, 3 and 4 are invariantly spliced, and this sequence is responsible for catalytic function. The 3' alternatively spliced exons, 5 and 6, are responsible for AChE disposition in tissue [J. Massoulie, The origin of the molecular diversity and functional anchoring of cholinesterases. Neurosignals 11 (3) (2002) 130-143; Y. Li, S. Camp, P. Taylor, Tissue-specific expression and alternative mRNA processing of the mammalian acetylcholinesterase gene. J. Biol. Chem. 268 (8) (1993) 5790-5797]. The splice to exon 5 produces the GPI anchored form of AChE found in the hematopoietic system, whereas the splice to exon 6 produces a sequence that binds to the structural subunits PRiMA and ColQ, producing AChE expression in brain and muscle. A third alternative RNA species is present that is not spliced at the 3' end; the intron 3' of exon 4 is used as coding sequence and produces the read-through, unanchored form of AChE. In order to further understand the role of alternative splicing in the expression of the AChE gene, we have used homologous recombination in stem cells to produce gene specific deletions in mice. Alternatively and together exon 5 and exon 6 were deleted. A cassette containing the neomycin gene flanked by loxP sites was used to replace the exon(s) of interest. Tissue analysis of mice with exon 5 deleted and the neomycin cassette retained showed very low levels of AChE expression, far less than would have been anticipated. Only the read-through species of the enzyme was produced; clearly the inclusion of the selection cassette disrupted splicing of exon 4 to exon 6. The selection cassette was then deleted in exon 5, exon 6 and exons 5 + 6 deleted mice by breeding to Ella-cre transgenic mice. AChE expression in serum, brain and muscle has been analyzed. Another AChE gene targeted mouse strain involving a region in the first intron, found to be critical for AChE expression in muscle cells [S. Camp, L. Zhang, M. Marquez, B

  16. Generation of selectable marker-free transgenic rice resistant to chewing insects using two co-transformation systems

    Institute of Scientific and Technical Information of China (English)

    Hengxiu Yu; Quanhong Yao; Ling Wang; Zhipeng Zhao; Zhiyun Gong; Shuzhu Tang; Qiaoquan Liu; Minghong Gua

    2009-01-01

    To produce selectable marker-free (SMF) transgenic rice resistant to chewing insects, the Bacillus thuringiensis crylA (c) gene (Bt) was introduced into two efitejaponica rice varieties by using two Agrobacterium-mediated co-transformation systems. One system is with a single mini-twin T-DNA binary vector in one Agrobacterium strain, which consists of two separate T-DNA regions, one carrying the Bt while the other contains the selectable marker gene, hygromycin resistant gene (HPT). The other system uses two separate binary vectors in two separate Agrobacterium cultures, containing the Bt or HPT gene on individual plasmids. A lot of independent transgenic rice lines harboring both Bt and selectable marker genes were obtained. The results showed that the co-transformation frequency of the Bt gene and HPTgene was much higher by using the mini-twin T-DNA vector system (29.87%) than that by the two separate binary vector sys-tems (4.52%). However, the frequency of the SMF transgenic rice plants obtained from the offspring of co-transgenic plants (21.74%) was lower for the mini-twin T-DNA vector system than that for the latter (50-60%). The data of ELISA implied that the expressed Bt pro-teins were quantitated as 0.025-0.103% of total leaf soluble proteins in the transgenic plant. Therefore, several elite transgenic rice lines, free of the selectable marker gene, were chosen. The results from both in vitro and in vivo insect bioassays indicated that the SMF trans-genic rice was shown to be highly resistant to the striped stem borer and rice leaf folder. Moreover, in a natural field condition without any insecticide applied, all the transgenic rice plants were found to be not injured by the rice leaf folder, whereas the wild types were impaired seriously.

  17. Consumption of milk from transgenic goats expressing human lysozyme in the mammary gland results in the modulation of intestinal microflora.

    Science.gov (United States)

    Maga, Elizabeth A; Walker, Richard L; Anderson, Gary B; Murray, James D

    2006-08-01

    Lysozyme is a key antimicrobial component of human milk that has several health-promoting functions including the development of a healthy intestinal tract. However, levels of lysozyme in the milk of dairy animals are negligible. We have generated transgenic dairy goats that express human lysozyme (HLZ) in their milk in an attempt to deliver the benefits of human milk in a continual fashion. To test the feasibility of this transgenic approach to achieve a biological impact at the level of the intestine, feeding trials were conducted in two animal models. Pasteurized milk from HLZ transgenic animals was fed to both kid goats (ruminant model) and young pigs (human model), and the numbers of total coliforms and Escherichia coli present in the small intestine were determined. Data from this proof-of-principle study demonstrate that milk from transgenic animals was capable of modulating the bacterial population of the gut in both animal models. Pigs that consumed pasteurized milk from HLZ transgenic goats had fewer numbers of coliforms and E. coli in their intestine than did those receiving milk from non-transgenic control animals. The opposite effect was seen in goats. Milk from these transgenic animals not only represent one of the first transgenic food products with the potential of benefiting human health, but are also a unique model to study the development and role of intestinal microflora on health, well-being and resistance to disease.

  18. The extinct animal show: the paleoimagery tradition and computer generated imagery in factual television programs.

    Science.gov (United States)

    Campbell, Vincent

    2009-03-01

    Extinct animals have always been popular subjects for the media, in both fiction, and factual output. In recent years, a distinctive new type of factual television program has emerged in which computer generated imagery is used extensively to bring extinct animals back to life. Such has been the commercial audience success of these programs that they have generated some public and academic debates about their relative status as science, documentary, and entertainment, as well as about their reflection of trends in factual television production, and the aesthetic tensions in the application of new media technologies. Such discussions ignore a crucial contextual feature of computer generated extinct animal programs, namely the established tradition of paleoimagery. This paper examines a selection of extinct animal shows in terms of the dominant frames of the paleoimagery genre. The paper suggests that such an examination has two consequences. First, it allows for a more context-sensitive evaluation of extinct animal programs, acknowledging rather than ignoring relevant representational traditions. Second, it allows for an appraisal and evaluation of public and critical reception of extinct animal programs above and beyond the traditional debates about tensions between science, documentary, entertainment, and public understanding.

  19. Generation and Characterization of a Transgenic Mouse Carrying a Functional Human β-Globin Gene with the IVSI-6 Thalassemia Mutation

    Directory of Open Access Journals (Sweden)

    Giulia Breveglieri

    2015-01-01

    Full Text Available Mouse models that carry mutations causing thalassemia represent a suitable tool to test in vivo new mutation-specific therapeutic approaches. Transgenic mice carrying the β-globin IVSI-6 mutation (the most frequent in Middle-Eastern regions and recurrent in Italy and Greece are, at present, not available. We report the production and characterization of a transgenic mouse line (TG-β-IVSI-6 carrying the IVSI-6 thalassemia point mutation within the human β-globin gene. In the TG-β-IVSI-6 mouse (a the transgenic integration region is located in mouse chromosome 7; (b the expression of the transgene is tissue specific; (c as expected, normally spliced human β-globin mRNA is produced, giving rise to β-globin production and formation of a human-mouse tetrameric chimeric hemoglobin αmu-globin2/βhu-globin2 and, more importantly, (d the aberrant β-globin-IVSI-6 RNAs are present in blood cells. The TG-β-IVSI-6 mouse reproduces the molecular features of IVSI-6 β-thalassemia and might be used as an in vivo model to characterize the effects of antisense oligodeoxynucleotides targeting the cryptic sites responsible for the generation of aberrantly spliced β-globin RNA sequences, caused by the IVSI-6 mutation. These experiments are expected to be crucial for the development of a personalized therapy for β-thalassemia.

  20. Generation and Characterization of a Transgenic Mouse Carrying a Functional Human β-Globin Gene with the IVSI-6 Thalassemia Mutation

    Science.gov (United States)

    Mancini, Irene; Lampronti, Ilaria; Salvatori, Francesca; Fabbri, Enrica; Zuccato, Cristina; Cosenza, Lucia C.; Montagner, Giulia; Borgatti, Monica; Altruda, Fiorella; Fagoonee, Sharmila; Carandina, Gianni; Aiello, Vincenzo; Breda, Laura; Rivella, Stefano; Gambari, Roberto

    2015-01-01

    Mouse models that carry mutations causing thalassemia represent a suitable tool to test in vivo new mutation-specific therapeutic approaches. Transgenic mice carrying the β-globin IVSI-6 mutation (the most frequent in Middle-Eastern regions and recurrent in Italy and Greece) are, at present, not available. We report the production and characterization of a transgenic mouse line (TG-β-IVSI-6) carrying the IVSI-6 thalassemia point mutation within the human β-globin gene. In the TG-β-IVSI-6 mouse (a) the transgenic integration region is located in mouse chromosome 7; (b) the expression of the transgene is tissue specific; (c) as expected, normally spliced human β-globin mRNA is produced, giving rise to β-globin production and formation of a human-mouse tetrameric chimeric hemoglobin mu α-globin2/hu β-globin2 and, more importantly, (d) the aberrant β-globin-IVSI-6 RNAs are present in blood cells. The TG-β-IVSI-6 mouse reproduces the molecular features of IVSI-6 β-thalassemia and might be used as an in vivo model to characterize the effects of antisense oligodeoxynucleotides targeting the cryptic sites responsible for the generation of aberrantly spliced β-globin RNA sequences, caused by the IVSI-6 mutation. These experiments are expected to be crucial for the development of a personalized therapy for β-thalassemia. PMID:26097845

  1. The effect of MT gene on heavy metal accumulations in transgenic common carp and experimental animal%金属硫蛋白基因对转基因鲤及实验动物体内重金属累积的影响

    Institute of Scientific and Technical Information of China (English)

    闫学春; 孙效文; 梁利群; 曹顶臣

    2005-01-01

    One of the most important things for industry production of transgenic fish is whether the fish meal is safe to the health of human beings, which means the foreign genes and their expressed products might be harmful to fish and the animal eating this fish. For the transgenic common carp produced by injecting fusion genes which included chum salmon' s growth hormone gene and common carp's metallothionine promoter, two things should be investigated. One is whether the transgenic common carp will produce excessive amounts of growth hormone in its body, and the foreign hormone how to affect the animal eating this fish. The other thing is whether this fish will accumulate excessive heavy metal in its body, what effect will be on the animal eating this fish. The investigation of the concentrations of the foreign growth hormone in transgenic carp is reported. This article examines the effect of a metallothionine transgene on the residual heavy metal concentrations in the transgenic common carp and the cat eating the transgenic carp. Using flame atomic absorption spectrophotometry, we measured the concentrations of the heavy mental elements including Zn, Cu, Cd, and Pb in the muscle, liver, kidney and gills of the transgenic common carp and non-transgenic carp. Also measured were the concentrations of Zn and Cd in the muscle, liver and kidney of the cat fed with transgenic and non-transgenic common carps. In the cat, there was no significant difference (P > 0.05) in tissue concentrations of heavy metals between the experimental group and the control group. The heavy metal concentrations in the non-transgenic common carp and transgenic common carp were statistically similar. The result showed that metallothionine gene as a promoter for aquaculture fish genetransfer would not bring about the heavy metal accumulations in transgenic fish and animal eating them。

  2. Lactogenic immunity in transgenic mice producing recombinant antibodies neutralizing coronavirus.

    Science.gov (United States)

    Castilla, J; Sola, I; Pintado, B; Sánchez-Morgado, J M; Enjuanes, L

    1998-01-01

    Protection against coronavirus infections can be provided by the oral administration of virus neutralizing antibodies. To provide lactogenic immunity, eighteen lines of transgenic mice secreting a recombinant IgG1 monoclonal antibody (rIgG1) and ten lines of transgenic mice secreting recombinant IgA monoclonal antibodies (rIgA) neutralizing transmissible gastroenteritis coronavirus (TGEV) into the milk were generated. Genes encoding the light and heavy chains of monoclonal antibody (MAb) 6A.C3 were expressed under the control of regulatory sequences derived from the mouse genomic DNA encoding the whey acidic protein (WAP) and beta-lactoglobulin (BLG), which are highly abundant milk proteins. The MAb 6A.C3 binds to a highly conserved epitope present in coronaviruses of several species. This MAb does not allow the selection of neutralization escaping virus mutants. The antibody was expressed in the milk of transgenic mice with titers of one million as determined by RIA, and neutralized TGEV infectivity by one million fold corresponding to immunoglobulin concentrations of 5 to 6 mg per ml. Matrix attachment regions (MAR) sequences were not essential for rIgG1 transgene expression, but co-microinjection of MAR and antibody genes led to a twenty to ten thousand-fold increase in the antibody titer in 50% of the rIgG1 transgenic animals generated. Co-microinjection of the genomic BLG gene with rIgA light and heavy chain genes led to the generation of transgenic mice carrying the three transgenes. The highest antibody titers were produced by transgenic mice that had integrated the antibody and BLG genes, although the number of transgenic animals generated does not allow a definitive conclusion on the enhancing effect of BLG co-integration. Antibody expression levels were transgene copy number independent and integration site dependent. The generation of transgenic animals producing virus neutralizing antibodies in the milk could be a general approach to provide protection

  3. 多囊卵巢综合征转基因动物模型的研究进展%Transgenic animal model of polycystic ovary syndrome

    Institute of Scientific and Technical Information of China (English)

    许婷; 李昕; 冯异

    2013-01-01

    Polycystic ovary syndrome is the most common endocrinal and metabolic disease in reproductive female. The etiology is still unclear. Building an appropriate animal model will improve the anderstanding of etiology, path-ogenesis, clinical strategy and the prevention performance for complication of PCOS. The previous animal models are induced by estrogen, androgen or constant illumination. Recently, new genetically modified animal models, such as transgenic, generalized knock-out or tissue specific knock-out mice were applied in the research of PCOS. The present review summarizes the origin action, features, research progression, application and advantages/disadvantages of several PCOS-like mice, providing optimal model for research.%多囊卵巢综合征(PCOS)是育龄期妇女常见的生殖内分泌及代谢紊乱性疾病,病因不明.建立合适的动物模型对研究其发病机制、病程发展、临床治疗及并发症的预防有着重要的意义.既往的动物模型为雌、雄激素或灯光控制诱导.近年来,转基因动物模型、基因敲除动物及组织特异性基因敲除动物被广泛应用于PCOS的研究.本综述总结了几种通过基因手段复制的啮齿类PCOS动物模型的来源、特点、研究进展、应用及优缺点,为实验研究选择相应的动物模型提供参考.

  4. Animator

    Science.gov (United States)

    Tech Directions, 2008

    2008-01-01

    Art and animation work is the most significant part of electronic game development, but is also found in television commercials, computer programs, the Internet, comic books, and in just about every visual media imaginable. It is the part of the project that makes an abstract design idea concrete and visible. Animators create the motion of life in…

  5. Animals

    Energy Technology Data Exchange (ETDEWEB)

    Skuterud, L.; Strand, P. [Norwegian Radiation Protection Authority (Norway); Howard, B.J. [Inst. of Terrestrial Ecology (United Kingdom)

    1997-10-01

    The radionuclides of most concern with respect to contamination of animals after a nuclear accident are radioiodine, radiocaesium and radiostrontium (ICRP 30, 1979). Of the other significant anthropogenic radionuclides likely to be released in most accidents, only small proportions of that ingested will be absorbed in an animals gut, and the main animal products, milk and meat, will not normally be contaminated to a significant extent. Animal products will mostly be contaminated as a result of ingestion of contaminated feed and possibly, but to a much lesser extent, from inhalation (for radioiodine only). Direct external contamination of animals is of little or no consequence in human food production. Radioiodine and radiostrontium are important with respect to contamination of milk; radiocaesium contaminates both milk and meat. The physical and chemical form of a radionuclide can influence its absorption in the animal gut. For example, following the Chernobyl accident radiocaesium incorporated into vegetation by root uptake was more readily absorbed than that associated with the original deposit. The transfer of radiocaesium and radiostrontium to animals will be presented both as transfer coefficients and aggregated transfer coefficients. For most animal meat products, only radiocaesium is important as other radionuclides do not significantly contaminate muscle. Farm animal products are the most important foodstuff determining radiocaesium intake by the average consumer in the Nordic countries. The major potential source of radioiodine and radiostrontium to humans is milk and milk products. Of the different species, the smaller animals have the highest transfer of radiocaesium from fodder to meat and milk. (EG). 68 refs.

  6. Transgenic dairy cattle: genetic engineering on a large scale.

    Science.gov (United States)

    Wall, R J; Kerr, D E; Bondioli, K R

    1997-09-01

    Amid the explosion of fundamental knowledge generated from transgenic animal models, a small group of scientists has been producing transgenic livestock with goals of improving animal production efficiency and generating new products. The ability to modify mammary-specific genes provides an opportunity to pursue several distinctly different avenues of research. The objective of the emerging gene "pharming" industry is to produce pharmaceuticals for treating human diseases. It is argued that mammary glands are an ideal site for producing complex bioactive proteins that can be cost effectively harvested and purified. Consequently, during the past decade, approximately a dozen companies have been created to capture the US market for pharmaceuticals produced from transgenic bioreactors estimated at $3 billion annually. Several products produced in this way are now in human clinical trials. Another research direction, which has been widely discussed but has received less attention in the laboratory, is genetic engineering of the bovine mammary gland to alter the composition of milk destined for human consumption. Proposals include increasing or altering endogenous proteins, decreasing fat, and altering milk composition to resemble that of human milk. Initial studies using transgenic mice to investigate the feasibility of enhancing manufacturing properties of milk have been encouraging. The potential profitability of gene "pharming" seems clear, as do the benefits of transgenic cows producing milk that has been optimized for food products. To take full advantage of enhanced milk, it may be desirable to restructure the method by which dairy producers are compensated. However, the cost of producing functional transgenic cattle will remain a severe limitation to realizing the potential of transgenic cattle until inefficiencies of transgenic technology are overcome. These inefficiencies include low rates of gene integration, poor embryo survival, and unpredictable transgene

  7. Generation of hermaphrodite transgenic papaya lines with virus resistance via transformation of somatic embryos derived from adventitious roots of in vitro shoots.

    Science.gov (United States)

    Kung, Yi-Jung; Yu, Tsong-Ann; Huang, Chiung-Huei; Wang, Hui-Chin; Wang, Shin-Lan; Yeh, Shyi-Dong

    2010-08-01

    Papaya production is seriously limited by Papaya ringspot virus (PRSV) worldwide and Papaya leaf-distortion mosaic virus (PLDMV) in Eastern Asia. An efficient transformation method for developing papaya lines with transgenic resistance to these viruses and commercially desirable traits, such as hermaphroditism, is crucial to shorten the breeding program for this fruit crop. In this investigation, an untranslatable chimeric construct pYP08 containing truncated PRSV coat protein (CP) and PLDMV CP genes coupled with the 3' untranslational region of PLDMV, was generated. Root segments from different portions of adventitious roots of in vitro multiple shoots of hermaphroditic plants of papaya cultivars 'Tainung No. 2', 'Sunrise', and 'Thailand' were cultured on induction medium for regeneration into somatic embryos. The highest frequency of somatic embryogenesis was from the root-tip segments of adventitious roots developed 2-4 weeks after rooting in perlite medium. After proliferation, embryogenic tissues derived from somatic embryos were wounded in liquid-phase by carborundum and transformed by Agrobacterium carrying pYP08. Similarly, another construct pBG-PLDMVstop containing untranslatable CP gene of PLDMV was also transferred to 'Sunrise' and 'Thailand', the parental cultivars of 'Tainung No. 2'. Among 107 transgenic lines regenerated from 349 root-tip segments, nine lines of Tainung No. 2 carrying YP08 were highly resistant to PRSV and PLDMV, and 9 lines (8 'Sunrise' and 1 'Thailand') carrying PLDMV CP highly resistant to PLDMV, by a mechanism of post-transcriptional gene silencing. The hermaphroditic characteristics of the transgenic lines were confirmed by PCR with sex-linked primers and phenotypes of flower and fruit. Our approach has generated transgenic resistance to both PRSV and PLDMV with commercially desirable characters and can significantly shorten the time-consuming breeding programs for the generation of elite cultivars of papaya hybrids.

  8. Generation of selectable marker-free transgenic eggplant resistant to Alternaria solani using the R/RS site-specific recombination system.

    Science.gov (United States)

    Darwish, Nader Ahmed; Khan, Raham Sher; Ntui, Valentine Otang; Nakamura, Ikuo; Mii, Masahiro

    2014-03-01

    Marker-free transgenic eggplants, exhibiting enhanced resistance to Alternaria solani , can be generated on plant growth regulators (PGRs)- and antibiotic-free MS medium employing the multi-auto-transformation (MAT) vector, pMAT21 - wasabi defensin , wherein isopentenyl transferase ( ipt ) gene is used as a positive selection marker. Use of the selection marker genes conferring antibiotic or herbicide resistance in transgenic plants has been considered a serious problem for environment and the public. Multi-auto-transformation (MAT) vector system has been one of the tools to excise the selection marker gene and produce marker-free transgenic plants. Ipt gene was used as a selection marker gene. Wasabi defensin gene, isolated from Wasabia japonica (a Japanese horseradish which has been a potential source of antimicrobial proteins), was used as a gene of interest. Wasabi defensin gene was cloned from the binary vector, pEKH-WD, to an ipt-type MAT vector, pMAT21, by gateway cloning technology and transferred to Agrobacterium tumefaciens strain EHA105. Infected cotyledon explants of eggplant were cultured on PGRs- and antibiotic-free MS medium. Extreme shooty phenotype/ipt shoots were produced by the explants infected with the pMAT21-wasabi defensin (WD). The same PGRs- and antibiotic-free MS medium was used in subcultures of the ipt shoots. Subsequently, morphologically normal shoots emerged from the Ipt shoots. Molecular analyses of genomic DNA from transgenic plants confirmed the integration of the WD gene and excision of the selection marker (ipt gene). Expression of the WD gene was confirmed by RT-PCR and Northern blot analyses. In vitro whole plant and detached leaf assay of the marker-free transgenic plants exhibited enhanced resistance against Alternaria solani.

  9. Efficient generation of transgene-free induced pluripotent stem cells from normal and neoplastic bone marrow and cord blood mononuclear cells.

    Science.gov (United States)

    Hu, Kejin; Yu, Junying; Suknuntha, Kran; Tian, Shulan; Montgomery, Karen; Choi, Kyung-Dal; Stewart, Ron; Thomson, James A; Slukvin, Igor I

    2011-04-07

    Reprogramming blood cells to induced pluripotent stem cells (iPSCs) provides a novel tool for modeling blood diseases in vitro. However, the well-known limitations of current reprogramming technologies include low efficiency, slow kinetics, and transgene integration and residual expression. In the present study, we have demonstrated that iPSCs free of transgene and vector sequences could be generated from human BM and CB mononuclear cells using non-integrating episomal vectors. The reprogramming described here is up to 100 times more efficient, occurs 1-3 weeks faster compared with the reprogramming of fibroblasts, and does not require isolation of progenitors or multiple rounds of transfection. Blood-derived iPSC lines lacked rearrangements of IGH and TCR, indicating that their origin is non-B- or non-T-lymphoid cells. When cocultured on OP9, blood-derived iPSCs could be differentiated back to the blood cells, albeit with lower efficiency compared to fibroblast-derived iPSCs. We also generated transgene-free iPSCs from the BM of a patient with chronic myeloid leukemia (CML). CML iPSCs showed a unique complex chromosomal translocation identified in marrow sample while displaying typical embryonic stem cell phenotype and pluripotent differentiation potential. This approach provides an opportunity to explore banked normal and diseased CB and BM samples without the limitations associated with virus-based methods.

  10. Lentiviral-Mediated Transgene Expression Can Potentiate Intestinal Mesenchymal-Epithelial Signaling

    Directory of Open Access Journals (Sweden)

    Dismuke Adria D

    2009-07-01

    Full Text Available Abstract Mesenchymal-epithelial signaling is essential for the development of many organs and is often disrupted in disease. In this study, we demonstrate the use of lentiviral-mediated transgene delivery as an effective approach for ectopic transgene expression and an alternative to generation of transgenic animals. One benefit to this approach is that it can be used independently or in conjunction with established transgenic or knockout animals for studying modulation of mesenchymal-epithelial interactions. To display the power of this approach, we explored ectopic expression of a Wnt ligand in the mouse intestinal mesenchyme and demonstrate its functional influence on the adjacent epithelium. Our findings highlight the efficient use of lentiviral-mediated transgene expression for modulating mesenchymal-epithelial interactions in vivo.

  11. Lentiviral-Mediated Transgene Expression Can Potentiate Intestinal Mesenchymal-Epithelial Signaling

    Directory of Open Access Journals (Sweden)

    Kohn Aimee

    2009-01-01

    Full Text Available Abstract Mesenchymal-epithelial signaling is essential for the development of many organs and is often disrupted in disease. In this study, we demonstrate the use of lentiviral-mediated transgene delivery as an effective approach for ectopic transgene expression and an alternative to generation of transgenic animals. One benefit to this approach is that it can be used independently or in conjunction with established transgenic or knockout animals for studying modulation of mesenchymal-epithelial interactions. To display the power of this approach, we explored ectopic expression of a Wnt ligand in the mouse intestinal mesenchyme and demonstrate its functional influence on the adjacent epithelium. Our findings highlight the efficient use of lentiviral-mediated transgene expression for modulating mesenchymal-epithelial interactions in vivo.

  12. Aging-related correlation of insulin-degrading enzyme with gamma-secretase-generated products involving insulin and glucose levels in transgenic mice.

    Science.gov (United States)

    Hwang, Dae Y; Cho, Jung S; Kim, Chuel K; Shim, Sun B; Jee, Seung W; Lee, Su H; Seo, Su J; Cho, Joon Y; Lee, Seok H; Kim, Yong K

    2005-09-01

    Insulin-degrading enzyme (IDE) is a 110-kDa thiol zinc-methalloendopeptidase that can cleave small Abeta peptides and the APP intracellular domain (AICD). The aim of this study was to examine aging-related correlation of IDE with gamma-secretase-generated products involving insulin and glucose levels in transgenic brains expressing neuron-specific enolase (NSE)-controlled human mutant presenilin-2 (hPS2m). Herein, we concluded that the levels of IDE expression in transgenic brains were decreased relative to those of control mice at 15 months of age. In parallel, inhibition in the IDE expression at this age underlies to the levels-up of Abeta-42, AICD, gamma-secretase, and glucose with a level-down of insulin. Thus, IDE expression is critical target for the therapeutic trials.

  13. Generation of the bovine viral diarrhea virus e0 protein in transgenic astragalus and its immunogenicity in sika deer.

    Science.gov (United States)

    Gao, Yugang; Zhao, Xueliang; Zang, Pu; Liu, Qun; Wei, Gongqing; Zhang, Lianxue

    2014-01-01

    The bovine viral diarrhea virus (BVDV), a single-stranded RNA virus, can cause fatal diarrhea syndrome, respiratory problems, and reproductive disorders in herds. Over the past few years, it has become clear that the BVDV infection rates are increasing and it is likely that an effective vaccine for BVDV will be needed. In this study, transgenic Astragalus was used as an alternative productive platform for the expression of glycoprotein E0. The immunogenicity of glycoprotein E0 expressed in transgenic Astragalus was detected in deer. The presence of pBI121-E0 was confirmed by polymerase chain reaction (PCR), transcription was verified by reverse transcription- (RT-) PCR, and recombinant protein expression was confirmed by ELISA and Western blot analyses. Deer that were immunized subcutaneously with the transgenic plant vaccine developed specific humoral and cell-mediated immune responses against BVDV. This study provides a new method for a protein with weak immunogenicity to be used as part of a transgenic plant vaccine.

  14. Generation of the Bovine Viral Diarrhea Virus E0 Protein in Transgenic Astragalus and Its Immunogenicity in Sika Deer

    Directory of Open Access Journals (Sweden)

    Yugang Gao

    2014-01-01

    Full Text Available The bovine viral diarrhea virus (BVDV, a single-stranded RNA virus, can cause fatal diarrhea syndrome, respiratory problems, and reproductive disorders in herds. Over the past few years, it has become clear that the BVDV infection rates are increasing and it is likely that an effective vaccine for BVDV will be needed. In this study, transgenic Astragalus was used as an alternative productive platform for the expression of glycoprotein E0. The immunogenicity of glycoprotein E0 expressed in transgenic Astragalus was detected in deer. The presence of pBI121-E0 was confirmed by polymerase chain reaction (PCR, transcription was verified by reverse transcription- (RT- PCR, and recombinant protein expression was confirmed by ELISA and Western blot analyses. Deer that were immunized subcutaneously with the transgenic plant vaccine developed specific humoral and cell-mediated immune responses against BVDV. This study provides a new method for a protein with weak immunogenicity to be used as part of a transgenic plant vaccine.

  15. ANIMALES TRANSGÉNICOS COMO ORGANISMOS ARTIFICIALES ANIMAIS TRANSGÊNICOS COMO ORGANISMOS ARTIFICIAIS TRANSGENIC ANIMALS AS ARTIFICIAL ORGANISMS

    Directory of Open Access Journals (Sweden)

    Alexandre Bota Arqué

    2007-06-01

    Full Text Available La modificación genética de organismos vivos ha generado interrogantes éticos de difícil solución. La búsqueda de marcos conceptuales lleva a plantear si ha habido una modificación del estatuto ontológico de los organismos alterados por la biotecnología. Este artículo realiza una aproximación al organismo biotecnológico a partir del pensamiento de los filósofos José Ortega y Gasset y Xavier Zubiri acerca del hombre y de la técnica, conceptualizando el objeto artificial biotecnológico como "organismo artificial", con la particularidad de ser el primer objeto producido por el hombre que se reproduce independientementeA modificação genética de organismos vivos gerou interrogativos éticos de difícil solução. A busca de marcos conceituais leva a questionar se ocorreu uma modificação do estatuto ontológico dos organismos modificados pela biotecnologia. Este artigo realiza uma aproximação ao organismo biotecnológico a partir do pensamento dos filósofos José Ortega y Gasset e Xavier Zubiri a respeito do homem e da técnica. Conceptualiza o objeto artificial biotecnológico como um "organismo artificial", com a particularidade de ser o primeiro objeto produzido pelo homem que se reproduz independentementeThe possibility of genetic modification of living organisms has generated ethical questions of difficult solution. The searching for conceptual frames has put into question whether the ontological status of organisms altered by biotechnology has been changed. This paper approaches the biotechnological organism following the thinking of philosophers Ortega y Gasset and Xavier Zubiri, based on their view on anthropology and technology conceptualizing the biotechnological artificial object as an "artificial organism" with the particularity that it is the first object produced by human beings which reproduces independently

  16. Animals

    Institute of Scientific and Technical Information of China (English)

    杨光

    2000-01-01

    The largest animal ever to live on the earth is the blue whale(蓝鲸)It weighs about 80 tons--more than 24 elephants. It is more than 30 metres long. A newborn baby whale weighs as much as a big elephant.

  17. Alzheimer’s disease transgenic animal models:How to get more similar pathological characteristics?%阿尔茨海默病转基因动物模型:如何更接近病理特征?

    Institute of Scientific and Technical Information of China (English)

    董贤慧; 柴锡庆

    2013-01-01

    BACKGROUND:Alzheimer’s disease causes and pathogenesis remain unclear, which greatly restrict the screening of drugs. And the main reason is lack of suitable animal models. The developing transgenic animal technology al ows studying the role of certain pathogenic gene in vivo, and has regarded the ideal animal models for Alzheimer’s disease. OBJECTIVE:To summarize the research advance of Alzheimer’s disease transgenic animal models. METHODS:Using“Alzheimer’s disease, transgenic mouse, animal model, dementia”in Chinese and English as the key words, the first author retrieved PubMed and CNKI databases published before July 2013. Final y, 41 articles were included in result analysis. RESULTS AND CONCLUSION:The etiology of Alzheimer’s disease is diverse, and genetic factor is one important factor. The existing transgenic animal models of Alzheimer’s disease include single genetical y modified models, double genetical y modified models and multiple transgenic models. Single transgenic animal models can make a kind of mutated exogenous gene integrate into the genomes of animals by using recombinant DNA technology. This kind of models can be applied to only study one specific pathological change of Alzheimer’s disease. Double transgenic animal models can make two kinds of mutated exogenous gene integrate into the genomes of animals and simultaneously transfect animals by using recombinant DNA technology. This kind of models is closer to the pathological changes of Alzheimer’s disease than single transgenic animal models, but stil cannot simulate Alzheimer’s disease. Multiple genetical y modified models are obtained with different transgenic mice hybridization or several genes transfection, which are most similar to clinical process and pathological features of Alzheimer’s disease. However, this kind of models may develop a decline in consanguinity. Each kind of animal model has their advantages and shortcomings, and a better transgenic animal

  18. Bridging the generation gap: flowering plant gametophytes and animal germlines reveal unexpected similarities.

    Science.gov (United States)

    Dickinson, Hugh G; Grant-Downton, Robert

    2009-11-01

    Alternation of generations underpins all plant life histories and is held to possess important adaptive features. A wide range of data have accumulated over the past century which suggest that alternation from sporophyte to gametophyte in angiosperms includes a significant phase of 'informational reprogramming', leaving the founder cells of the gametophyte developmentally uncommitted. This review attempts to bring together results from these historic studies with more recent data on molecular and epigenetic events which accompany alternation, gametophyte development and gametogenesis in angiosperms. It is striking that most members of the other principal group of multicellular eukaryotes--the animals--have a completely different a life history: animals generate their gametes directly from diploid germlines, often set aside early in development. Nevertheless, a comparison between animal germlines and angiosperm gametophyte development reveals a number of surprising similarities at the cytological and molecular levels. This difference in life history but similarity in developmental process is reviewed in the context of the very different life strategies adopted by plants and animals, and particularly the fact that plants do not set aside diploid germlines early in development.

  19. ANIMALS

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Mammals(哺乳动物)Mammals are the world's most dominant(最占优势的)animal.They are extremely(非常)diverse(多种多样的)creatures(生物,动物)that include(包括)the biggest ever animal (the blue whale鲸,which eats up to 6 tons every day),the smallest(leaf-nosed bat小蹄蝠) and the laziest(sloth树獭,who spends 80% of their time sleeping).There are over 4,600 kinds of mammals and they live in very different environments(环境)—oceans(海洋),rivers,the jungle(丛林),deserts,and plains(平原).

  20. Generation and characterization of transgenic mice expressing mitochondrial targeted red fluorescent protein selectively in neurons: modeling mitochondriopathy in excitotoxicity and amyotrophic lateral sclerosis

    Directory of Open Access Journals (Sweden)

    Wang Yi

    2011-11-01

    Full Text Available Abstract Background Mitochondria have roles or appear to have roles in the pathogenesis of several chronic age-related and acute neurological disorders, including Charcot-Marie-Tooth disease, amyotrophic lateral sclerosis, Parkinson's disease, and cerebral ischemia, and could be critical targets for development of rational mechanism-based, disease-modifying therapeutics for treating these disorders effectively. A deeper understanding of neural tissue mitochondria pathobiologies as definitive mediators of neural injury, disease, and cell death merits further study, and the development of additional tools to study neural mitochondria will help achieve this unmet need. Results We created transgenic mice that express the coral (Discosoma sp. red fluorescent protein DsRed2 specifically in mitochondria of neurons using a construct engineered with a Thy1 promoter, specific for neuron expression, to drive expression of a fusion protein of DsRed2 with a mitochondrial targeting sequence. The biochemical and histological characterization of these mice shows the expression of mitochondrial-targeted DsRed2 to be specific for mitochondria and concentrated in distinct CNS regions, including cerebral cortex, hippocampus, thalamus, brainstem, and spinal cord. Red fluorescent mitochondria were visualized in cerebral cortical and hippocampal pyramidal neurons, ventrobasal thalamic neurons, subthalamic neurons, and spinal motor neurons. For the purpose of proof of principle application, these mice were used in excitotoxicity paradigms and double transgenic mice were generated by crossing Thy1-mitoDsRed2 mice with transgenic mice expressing enhanced-GFP (eGFP under the control of the Hlxb9 promoter that drives eGFP expression specifically in motor neurons and by crossing Thy1-mitoDsRed2 mice to amyotrophic lateral sclerosis (ALS mice expressing human mutant superoxide dismutase-1. Conclusions These novel transgenic mice will be a useful tool for better understanding

  1. V-Man Generation for 3-D Real Time Animation. Chapter 5

    Science.gov (United States)

    Nebel, Jean-Christophe; Sibiryakov, Alexander; Ju, Xiangyang

    2007-01-01

    The V-Man project has developed an intuitive authoring and intelligent system to create, animate, control and interact in real-time with a new generation of 3D virtual characters: The V-Men. It combines several innovative algorithms coming from Virtual Reality, Physical Simulation, Computer Vision, Robotics and Artificial Intelligence. Given a high-level task like "walk to that spot" or "get that object", a V-Man generates the complete animation required to accomplish the task. V-Men synthesise motion at runtime according to their environment, their task and their physical parameters, drawing upon its unique set of skills manufactured during the character creation. The key to the system is the automated creation of realistic V-Men, not requiring the expertise of an animator. It is based on real human data captured by 3D static and dynamic body scanners, which is then processed to generate firstly animatable body meshes, secondly 3D garments and finally skinned body meshes.

  2. Physical Exercise Restores the Generation of Newborn Neurons in an Animal Model of Chronic Epilepsy

    Science.gov (United States)

    Mendonça, Fabricio N.; Santos, Luiz E. C.; Rodrigues, Antônio M.; Gomes da Silva, Sérgio; Arida, Ricardo M.; da Silveira, Gilcélio A.; Scorza, Fulvio A.; Almeida, Antônio-Carlos G.

    2017-01-01

    Neurogenesis impairment is associated with the chronic phase of the epilepsy in humans and also observed in animal models. Recent studies with animal models have shown that physical exercise is capable of improving neurogenesis in adult subjects, alleviating cognitive impairment and depression. Here, we show that there is a reduction in the generation of newborn granule cells in the dentate gyrus of adult rats subjected to a chronic model of epilepsy during the postnatal period of brain development. We also show that the physical exercise was capable to restore the number of newborn granule cells in this animals to the level observed in the control group. Notably, a larger number of newborn granule cells exhibiting morphological characteristics indicative of correct targeting into the hippocampal circuitry and the absence of basal dendrite projections was also observed in the epileptic animals subjected to physical exercise compared to the epileptic animals. The results described here could represent a positive interference of the physical exercise on the neurogenesis process in subjects with chronic epilepsy. The results may also help to reinterpret the benefits of the physical exercise in alleviating symptoms of depression and cognitive dysfunction. PMID:28298884

  3. Plasma adiponectin levels are increased despite insulin resistance in corticotropin-releasing hormone transgenic mice, an animal model of Cushing syndrome.

    Science.gov (United States)

    Shinahara, Masayuki; Nishiyama, Mitsuru; Iwasaki, Yasumasa; Nakayama, Shuichi; Noguchi, Toru; Kambayashi, Machiko; Okada, Yasushi; Tsuda, Masayuki; Stenzel-Poore, Mary P; Hashimoto, Kozo; Terada, Yoshio

    2009-01-01

    Adiponectin (AdN), an adipokine derived from the adipose tissue, has an insulin-sensitizing effect, and plasma AdN is shown to be decreased in obesity and/or insulin resistant state. To clarify whether changes in AdN are also responsible for the development of glucocorticoid-induced insulin resistance, we examined AdN concentration in plasma and AdN expression in the adipose tissue, using corticotropin-releasing hormone (CRH) transgenic mouse (CRH-Tg), an animal model of Cushing syndrome. We found, unexpectedly, that plasma AdN levels in CRHTg were significantly higher than those in wild-type littermates (wild-type: 19.7+/-2.5, CRH-Tg: 32.4+/-3.1 microg/mL, pAdN mRNA and protein levels were significantly decreased in the adipose tissue of CRH-Tg. Bilateral adrenalectomy in CRH-Tg eliminated both their Cushing's phenotype and their increase in plasma AdN levels (wild-type/sham: 9.4+/-0.5, CRH-Tg/sham: 15.7+/-2.0, CRH-Tg/ADX: 8.5+/-0.4 microg/mL). These results strongly suggest that AdN is not a major factor responsible for the development of insulin resistance in Cushing syndrome. Our data also suggest that glucocorticoid increases plasma AdN levels but decreases AdN expression in adipocytes, the latter being explained possibly by the decrease in AdN metabolism in the Cushing state.

  4. Transgenic animals and its applications in transfusion medicine%转基因动物在输血医学中的应用

    Institute of Scientific and Technical Information of China (English)

    章金刚; 卜凤荣

    2001-01-01

    There has been a considerable amount of work done on integrating the transgenic animal technology into transfusion medicine. We would like to review some of these areas in turn: the preparations of plasma proteins and hemoglobin by transgenesis bioreactors, the model systems for human diseases (blood transmitted viral diseases and blood related genetic diseases), as well as the transgenesis in basic research of blood transfusion medicine.%转基因动物技术是在动物整体水平研究和表达目的基因的生物技术,其基本特点是:分子及细胞水平操作,组织及整体水平表达,是常规分子生物学理论和技术的拓展和延伸,也是现代生物高技术研究和开发的热点之一。本文简述了转基因动物在输血医学领域的应用及其发展前景,包括利用转基因动物生物反应器制备人血浆蛋白和人血红蛋白、建立血传病毒的感染模型和血液相关遗传病模型以及转基因动物与输血医学的基础研究等。

  5. Two-photon microscopy imaging of thy1GFP-M transgenic mice: a novel animal model to investigate brain dendritic cell subsets in vivo.

    Directory of Open Access Journals (Sweden)

    Claudia Laperchia

    Full Text Available Transgenic mice expressing fluorescent proteins in specific cell populations are widely used for in vivo brain studies with two-photon fluorescence (TPF microscopy. Mice of the thy1GFP-M line have been engineered for selective expression of green fluorescent protein (GFP in neuronal populations. Here, we report that TPF microscopy reveals, at the brain surface of these mice, also motile non-neuronal GFP+ cells. We have analyzed the behavior of these cells in vivo and characterized in brain sections their immunophenotype.With TPF imaging, motile GFP+ cells were found in the meninges, subarachnoid space and upper cortical layers. The striking feature of these cells was their ability to move across the brain parenchyma, exhibiting evident shape changes during their scanning-like motion. In brain sections, GFP+ cells were immunonegative to antigens recognizing motile cells such as migratory neuroblasts, neuronal and glial precursors, mast cells, and fibroblasts. GFP+ non-neuronal cells exhibited instead the characteristic features and immunophenotype (CD11c and major histocompatibility complex molecule class II immunopositivity of dendritic cells (DCs, and were immunonegative to the microglial marker Iba-1. GFP+ cells were also identified in lymph nodes and blood of thy1GFP-M mice, supporting their identity as DCs. Thus, TPF microscopy has here allowed the visualization for the first time of the motile behavior of brain DCs in situ. The results indicate that the thy1GFP-M mouse line provides a novel animal model for the study of subsets of these professional antigen-presenting cells in the brain. Information on brain DCs is still very limited and imaging in thy1GFP-M mice has a great potential for analyses of DC-neuron interaction in normal and pathological conditions.

  6. Generation of an optimized lentiviral vector encoding a high-expression factor VIII transgene for gene therapy of hemophilia A.

    Science.gov (United States)

    Johnston, J M; Denning, G; Doering, C B; Spencer, H T

    2013-06-01

    We previously compared the expression of several human factor VIII (fVIII) transgene variants and demonstrated the superior expression properties of B domain-deleted porcine fVIII. Subsequently, a hybrid human/porcine fVIII molecule (HP-fVIII) comprising 91% human amino-acid sequence was engineered to maintain the high-expression characteristics of porcine fVIII. The bioengineered construct then was used effectively to treat knockout mice with hemophilia A. In the current study, we focused on optimizing self-inactivating (SIN) lentiviral vector systems by analyzing the efficacy of various lentiviral components in terms of virus production, transduction efficiency and transgene expression. Specifically, three parameters were evaluated: (1) the woodchuck hepatitis post-transcriptional regulatory element (WPRE), (2) HIV versus SIV viral vector systems and (3) various internal promoters. The inclusion of a WPRE sequence had negligible effects on viral production and HP-fVIII expression. HIV and SIV vectors were compared and found to be similar with respect to transduction efficiency in both K562s and HEK-293T cells. However, there was an enhanced expression of HP-fVIII by the SIV system, which was evident in both K562 and BHK-M cell lines. To further compare expression of HP-fVIII from an SIV-based lentiviral system, we constructed expression vectors containing the high expression transgene and a human elongation factor-1 alpha, cytomegalovirus (CMV) or phosphoglycerate kinase promoter. Expression was significantly greater from the CMV promoter, which also yielded therapeutic levels of HP-fVIII in hemophilia A mice. Based on these studies, an optimized vector contains the HP-fVIII transgene driven by a CMV internal promoter within a SIV-based lentiviral backbone lacking a WPRE.

  7. Establishment and characterization of fetal fibroblast cell lines for generating human lysozyme transgenic goats by somatic cell nuclear transfer.

    Science.gov (United States)

    Liu, Jun; Luo, Yan; Zheng, Liming; Liu, Qingqing; Yang, Zhongcai; Wang, Yongsheng; Su, Jianmin; Quan, Fusheng; Zhang, Yong

    2013-10-01

    This study was performed to qualify goat fetal fibroblast (GFF) cell lines for genetic modification and somatic cell nuclear transfer (SCNT) to produce human lysozyme (hLYZ) transgenic goats. Nine GFF cell lines were established from different fetuses, and the proliferative lifespan and chromosomal stability were analyzed. The results suggested that cell lines with a longer lifespan had stable chromosomes compared with those of cells lines with a shorter lifespan. According to the proliferative lifespan, we divided GFF cell lines into two groups: cell lines with a long lifespan (GFF1/2/7/8/9; group L) and cell lines with a short lifespan (GFF3/4/5/6; group S). Next, a hLYZ expression vector was introduced into these cell lines by electroporation. The efficiencies of colony formation, expansion in culture, and the quality of transgenic clonal cell lines were significant higher in group L than those in group S. The mean fusion rate and blastocyst rate in group L were higher than those in group S (80.3 ± 1.7 vs. 65.1 ± 4.2 % and 19.5 ± 0.6 vs. 15.1 ± 1.1 %, respectively, P embryos into the oviducts of recipient goats, three live kids were born. PCR and Southern blot analyses confirmed integration of the transgene in cloned goats. In conclusion, the lifespan of GFF cell lines has a major effect on the efficiency to produce transgenic cloned goats. Therefore, the proliferative lifespan of primary cells may be used as a criterion to characterize the quality of cell lines for genetic modification and SCNT.

  8. Generation of the Bovine Viral Diarrhea Virus E0 Protein in Transgenic Astragalus and Its Immunogenicity in Sika Deer

    OpenAIRE

    Yugang Gao; Xueliang Zhao; Pu Zang; Qun Liu; Gongqing Wei; Lianxue Zhang

    2014-01-01

    The bovine viral diarrhea virus (BVDV), a single-stranded RNA virus, can cause fatal diarrhea syndrome, respiratory problems, and reproductive disorders in herds. Over the past few years, it has become clear that the BVDV infection rates are increasing and it is likely that an effective vaccine for BVDV will be needed. In this study, transgenic Astragalus was used as an alternative productive platform for the expression of glycoprotein E0. The immunogenicity of glycoprotein E0 expressed in tr...

  9. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm

    Energy Technology Data Exchange (ETDEWEB)

    Harholt, Jesper; Bach, Inga C; Lind-Bouquin, Solveig; Nunan, Kylie J.; Madrid, Susan M.; Brinch-Pedersen, Henrik; Holm, Preben B.; Scheller, Henrik V.

    2009-12-08

    Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal KDEL were used. Transgenic plants were recovered in all four cases but no qualitative differences could be observed whether KDEL was added or not. Endo-xylanase activity in transgenic grains was increased between two and three fold relative to wild type. The grains were shriveled and had a 25-33% decrease in mass. Extensive analysis of the cell walls showed a 10-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water extractable arabinoxylan, and a shift in the MW of the water extractable arabinoxylan from being mainly larger than 85 kD to being between 2 kD and 85 kD. Ferulic acid esterase expressing grains were also shriveled and the seed weight was decreased by 20-50%. No ferulic acid esterase activity could be detected in wild type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15-40% increase in water unextractable arabinoxylan and a decrease in monomeric ferulic acid between 13 and 34%. In all the plants the observed changes are consistent with a plant response that serves to minimize the effect of the heterologously expressed enzymes by increasing arabinoxylan biosynthesis and cross-linking.

  10. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm.

    Science.gov (United States)

    Harholt, Jesper; Bach, Inga C; Lind-Bouquin, Solveig; Nunan, Kylie J; Madrid, Susan M; Brinch-Pedersen, Henrik; Holm, Preben B; Scheller, Henrik V

    2010-04-01

    Endo-xylanase (from Bacillus subtilis) or ferulic acid esterase (from Aspergillus niger) were expressed in wheat under the control of the endosperm-specific 1DX5 glutenin promoter. Constructs both with and without the endoplasmic reticulum retention signal (Lys-Asp-Glu-Leu) KDEL were used. Transgenic plants were recovered in all four cases but no qualitative differences could be observed whether KDEL was added or not. Endo-xylanase activity in transgenic grains was increased between two and threefold relative to wild type. The grains were shrivelled and had a 25%-33% decrease in mass. Extensive analysis of the cell walls showed a 10%-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water-extractable arabinoxylan, and a shift in the MW of the water-extractable arabinoxylan from being mainly larger than 85 kD to being between 2 and 85 kD. Ferulic acid esterase-expressing grains were also shrivelled, and the seed weight was decreased by 20%-50%. No ferulic acid esterase activity could be detected in wild-type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15%-40% increase in water-unextractable arabinoxylan and a decrease in monomeric ferulic acid between 13% and 34%. In all the plants, the observed changes are consistent with a plant response that serves to minimize the effect of the heterologously expressed enzymes by increasing arabinoxylan biosynthesis and cross-linking.

  11. Clinical assessment of freezing of gait in Parkinson's disease from computer-generated animation.

    Science.gov (United States)

    Morris, Tiffany R; Cho, Catherine; Dilda, Valentina; Shine, James M; Naismith, Sharon L; Lewis, Simon J G; Moore, Steven T

    2013-06-01

    The current 'gold standard' for clinical evaluation of freezing of gait (FOG) in Parkinson's disease (PD) is determination of the number of FOG episodes from video by independent raters. We have previously described a robust technique for objective FOG assessment from lower-limb acceleration. However, there is no existing method for validation of autonomous FOG measures in the absence of video documentation. In this study we compared the results of clinical evaluation of FOG from computer-generated animations (derived from body-mounted inertial sensors) during a timed up and go test with the 'gold standard' of clinical video assessment, utilizing a cohort of 10 experienced raters from four PD centers. Agreement between the 10 clinical observers for scoring of FOG from computer animations was more robust for the relative duration of freeze events (percent time frozen; intraclass correlation coefficient of 0.65) than number of FOG episodes, and was comparable with clinical evaluation of the patient from video (intraclass correlation coefficient 0.73). This result suggests that percent time frozen should be considered (along with number of FOG events) to better convey FOG severity. The ability of clinical observers to quantify FOG from computer-generated animation derived from lower-limb motion data provides a potential approach to validation of accelerometry-based FOG identification outside of the clinic. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Application of a nuclear localization signal gene in transgene mice

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Efficient gene transfer by cytoplasm co-injec- tion will offer a powerful means for transgenic animals. Using co-injection in cytoplasm, two independent gene constructs, including bovine (?-s1-casein-hG-CSF and a mammal expression vector expressing a nuclear localization signal (mNLS), were introduced into fertilized mouse eggs. The target gene construct was docked into host nucleus probably by the nuclear localization signal. Transgene mice have been obtained at 58% (29/50) of integration ratio. Expression level of the positive transgene mice was detected by Western blotting. Maximal expression of human G-CSF was estimated about 540 mg/L of milk. The expression ratio was up to 75% (9/12). The results here have important practical implications for the generation of mammary gland bioreactors and other transgene studies. Co-injection of a target gene with an expression vector of a mammal nuclear localization signal by cytoplasm appears to be a useful, efficient and easy strategy for generating transgenic animals, which may be able to substitute the routine method of pronucleus-injection of fertilized eggs.

  13. Transgenesis may affect farm animal welfare: a case for systematic risk assessment.

    Science.gov (United States)

    Van Reenen, C G; Meuwissen, T H; Hopster, H; Oldenbroek, K; Kruip, T H; Blokhuis, H J

    2001-07-01

    This paper considers (potentially) harmful consequences of transgenesis for farm animal welfare and examines the strategy of studying health and welfare of transgenic farm animals. Evidence is discussed showing that treatments imposed in the context of farm animal transgenesis are by no means biologically neutral and may compromise animal health and welfare. Factors posing a risk for the welfare of transgenic farm animals include integration of a transgene within an endogenous gene with possible loss of host gene function (insertional mutations), inappropriate transgene expression and exposure of the host to biologically active transgene-derived proteins, and in vitro reproductive technologies employed in the process of generating transgenic farm animals that may result in an increased incidence of difficult parturition and fetal and neonatal losses and the development of unusually large or otherwise abnormal offspring (large offspring syndrome). Critical components of a scheme for evaluating welfare of transgenic farm animals are identified, related to specific characteristics of transgenic animals and to factors that may interact with the effects of transgenesis. The feasibility of an evaluation of welfare of transgenic farm animals in practice is addressed against the background of the objectives and conditions of three successive stages in a long-term transgenic program. Concrete steps with regard to breeding and testing of transgenic farm animals are presented, considering three technologies to generate transgenic founders: microinjection, electroporation and nuclear transfer, and gene targeting including gene knockout. The proposed steps allow for unbiased estimations of the essential treatment effects, including hemi- and homozygous transgene effects as well as effects of in vitro reproductive technologies. It is suggested that the implementation of appropriate breeding and testing procedures should be accompanied by the use of a comprehensive welfare

  14. Optimization of embryo culture conditions for increasing efficiency of cloning in buffalo (Bubalus bubalis) and generation of transgenic embryos via cloning.

    Science.gov (United States)

    Wadhwa, Neerja; Kunj, Neetu; Tiwari, Shuchita; Saraiya, Megha; Majumdar, Subeer S

    2009-09-01

    Cloning in bovine species is marred by low efficiency of blastocyst formation. Any increase in the efficiency of blastocyst formation upon nuclear transfer will greatly enhance the efficiency of cloning. In the present study, the effect of various media, protein sources, and growth factors on the development of cloned buffalo embryos was evaluated. Among various combinations tested, culture of cloned embryos in TCM-199 media on the feeder layer of Buffalo Oviductal Epithelial Cells (BOEC) in the presence of bovine serum albumin-free fatty acid (BSA-FFA) and leukemia inhibitory factor (LIF) provided most suitable environment for efficient development of cloned blastocysts. Under these conditions, we achieved a blastocyst formation rate of 43%, which is better than those reported previously. Because preimplantation embryonic development, in vivo, occurs in an environment of oviductal cells, the blastocysts generated by this method may presumably be more suitable for implantation and further development. Additionally, we generated green blastocysts from enucleated oocytes by transfer of nuclei from cells transfected with EGFP transgene, showing possibility of transgenesis via cloning in this species. To our knowledge, this is the first report regarding the production of transgenic cloned buffalo embryos and their developmental competence with respect to various media, cocultures, and supplements.

  15. Artificial MicroRNA-Based Specific Gene Silencing of Grain Hardness Genes in Polyploid Cereals Appeared to Be Not Stable Over Transgenic Plant Generations

    Science.gov (United States)

    Gasparis, Sebastian; Kała, Maciej; Przyborowski, Mateusz; Orczyk, Waclaw; Nadolska-Orczyk, Anna

    2017-01-01

    Gene silencing by RNA interference is a particularly important tool in the study of gene function in polyploid cereal species for which the collections of natural or induced mutants are very limited. Previously we have been testing small interfering RNA-based approach of gene silencing in wheat and triticale. In this research, artificial microRNAs (amiRs) were studied in the same species and the same target genes to compare effectiveness of both gene silencing pathways. amiR cassettes were designed to silence Puroindoline a (Pina) and Puroindoline b (Pinb) hardness genes in wheat and their orthologues Secaloindoline a (Sina) and Secaloindoline b (Sinb) genes in triticale. Each of the two cassettes contained 21 nt microRNA (miR) precursor derived from conserved regions of Pina/Sina or Pinb/Sinb genes, respectively. Transgenic plants were obtained with high efficiency in two cultivars of wheat and one cultivar of triticale after using the Pinb-derived amiR vector for silencing of Pinb or Sinb, respectively. Lack of transgenic plants in wheat or very low transformation efficiency in triticale was observed using the Pina-derived amiR cassette, despite large numbers of embryos attempted. Silencing of Pinb in wheat and Sinb in triticale was highly efficient in the T1 generation. The transcript level of Pinb in wheat was reduced up to 92% and Sinb in triticale was reduced up to 98%. Moreover, intended silencing of Pinb/Sinb with Pinb-derived amiR cassette was highly correlated with simultaneous silencing of Pina/Sina in the same transgenic plants. High downregulation of Pinb/Pina genes in T1 plants of wheat and Sinb/Sina genes in T1 plants of triticale was associated with strong expression of Pinb-derived amiR. Silencing of the target genes correlated with increased grain hardness in both species. Total protein content in the grains of transgenic wheat was significantly lower. Although, the Pinb-derived amiR cassette was stably inherited in the T2 generation of wheat and

  16. The development of a modular system to burn farm animal waste to generate heat and power

    Energy Technology Data Exchange (ETDEWEB)

    Virr, M.J. [Spinheat Ltd., Pottsville, PA (United States)

    2002-07-01

    This paper presents an internally circulating fluid bed (ICFB) boiler that can burn poultry litter or other waste fuels. The disposal of animal waste is a significant problem for many poultry producers in the Chesapeake Bay area where poultry waste has contaminated local watersheds and the unpleasant odour has offended the local population. ICFB boilers can be made in the range of 3,000 to 23,000 kg/hr. Spinheat Ltd. has designed a complete modular small co-generating power plant in the range of 100 to 1,000 kW electrical generation with equivalent steam output. The co-generating power unit fits on the processor's premises to supply electricity and steam for process use by burning the poultry litter. The unit has been tested for poultry litter combustion and for emissions. This paper illustrates the complete design of the modular plants in the 100, 150, 200, 580 kW range as well as the 1 MW size. This new co-generating unit solves the waste management problem for poultry producers, as it can result in half the amount of poultry litter being spread on land as fertilizer. The cost of building the co-generation plant was reported in this paper along with the running costs of the plant. 1 tab., 8 figs.

  17. Screening and Integration Analysis of OsDREB1A BC4F2 and BC5F1 Generations of Transgenic Ciherang Rice (Oryza sativa L. for High-Salinity Tolerance

    Directory of Open Access Journals (Sweden)

    Dika Migi Priyono

    2014-06-01

    Full Text Available Salinity is one of a number of abiotic stresses that threaten rice production in Indonesia. To support food security programs, BB-Biogen has developed rice lines derived from OsDREB1A transformed Ciherang up to BC4F2 and BC5F1 generations. To verify the salinity tolerance and the stability of transgene integration, the BC4F2 and BC5F1 generations of Ciherang-OsDREB1A transgenic were screened to select high-salinity tolerant lines. Second, molecular analysis using the primers hptII-F/hptII-R and 35S-496-F/OsDREB1A-R was conducted to identify the existence and the stability of the transgene integration in the BC4F2 and BC5F1 generations. Screening 543 BC4F2 and BC5F1 Ciherang-OsDREB1A transgenic lines in a nutrient solution with a final electrical conductivity (EC of approximately 18 mS/cm for 26 days yielded 134 putative transgenic plants. Integration analysis using the hptII-F/hptII-R primers showed that 73 of the 134 putative transgenic plants had positive PCR products, indicating the presence of the transgene in those plants. All the 73 plants also produced PCR products when tested with the specific primer 35S-496- F/OsDREB1A-R, indicating that transgene integration was maintained during the development of BC4F2 and BC5F1.

  18. Production of germline transgenic chickens expressing enhanced green fluorescent protein using a MoMLV-based retrovirus vector.

    Science.gov (United States)

    Koo, Bon Chul; Kwon, Mo Sun; Choi, Bok Ryul; Kim, Jin-Hoi; Cho, Seong-Keun; Sohn, Sea Hwan; Cho, Eun Jung; Lee, Hoon Taek; Chang, Wonkyung; Jeon, Iksoo; Park, Jin-Ki; Park, Jae Bok; Kim, Teoan

    2006-11-01

    The Moloney murine leukemia virus (MoMLV) -based retrovirus vector system has been used most often in gene transfer work, but has been known to cause silencing of the imported gene in transgenic animals. In the present study, using a MoMLV-based retrovirus vector, we successfully generated a new transgenic chicken line expressing high levels of enhanced green fluorescent protein (eGFP). The level of eGFP expression was conserved after germline transmission and as much as 100 microg of eGFP could be detected per 1 mg of tissue protein. DNA sequencing showed that the transgene had been integrated at chromosome 26 of the G1 and G2 generation transgenic chickens. Owing to the stable integration of the transgene, it is now feasible to produce G3 generation of homozygous eGFP transgenic chickens that will provide 100% transgenic eggs. These results will help establish a useful transgenic chicken model system for studies of embryonic development and for efficient production of transgenic chickens as bioreactors.

  19. Analysis of the APX, PGD1 and R1G1B constitutive gene promoters in various organs over three homozygous generations of transgenic rice plants.

    Science.gov (United States)

    Park, Su-Hyun; Bang, Seung Woon; Jeong, Jin Seo; Jung, Harin; Redillas, Mark Christian Felipe Reveche; Kim, Hyung Il; Lee, Kang Hyun; Kim, Youn Shic; Kim, Ju-Kon

    2012-06-01

    We have previously characterized the constitutively active promoters of the APX, PGD1 and R1G1B genes in rice (Park et al. 2010 in J Exp Bot 61:2459-2467). To have potential crop biotechnology applications, gene promoters must be stably active over many generations. In our current study, we report our further detailed analysis of the APX, PGD1 and R1G1B gene promoters in various organs and tissues of transgenic rice plants for three (T₃₋₅) homozygous generations. The copy numbers in 37 transgenic lines that harbor promoter:gfp constructs were determined and promoter activities were measured by real-time qPCR. Analysis of the 37 lines revealed that 15 contained a single copy of one of the three promoter:gfp chimeric constructs. The promoter activity levels were generally higher in multi-copy lines, whereas variations in these levels over the T₃₋₅ generations studied were observed to be smaller in single-copy than in multi-copy lines. The three promoters were further found to be highly active in the whole plant body at both the vegetative and reproductive stages of plant growth, with the exception of the APX in the ovary and R1G1B in the pistil and filaments where zero or very low levels of activity were detected. Of note, the spatial activities of the PGD1 promoter were found to be strikingly similar to those of the ZmUbi1, a widely used constitutive promoter. Our comparison of promoter activities between T₃, T₄ and T₅ plants revealed that the APX, PGD1 and R1G1B promoters maintained their activities at comparable levels in leaves and roots over three homozygous generations and are therefore potentially viable alternative promoters for crop biotechnology applications.

  20. Remote automated multi-generational growth and observation of an animal in low Earth orbit.

    Science.gov (United States)

    Oczypok, Elizabeth A; Etheridge, Timothy; Freeman, Jacob; Stodieck, Louis; Johnsen, Robert; Baillie, David; Szewczyk, Nathaniel J

    2012-03-07

    The ultimate survival of humanity is dependent upon colonization of other planetary bodies. Key challenges to such habitation are (patho)physiologic changes induced by known, and unknown, factors associated with long-duration and distance space exploration. However, we currently lack biological models for detecting and studying these changes. Here, we use a remote automated culture system to successfully grow an animal in low Earth orbit for six months. Our observations, over 12 generations, demonstrate that the multi-cellular soil worm Caenorhabditis elegans develops from egg to adulthood and produces progeny with identical timings in space as on the Earth. Additionally, these animals display normal rates of movement when fully fed, comparable declines in movement when starved, and appropriate growth arrest upon starvation and recovery upon re-feeding. These observations establish C. elegans as a biological model that can be used to detect changes in animal growth, development, reproduction and behaviour in response to environmental conditions during long-duration spaceflight. This experimental system is ready to be incorporated on future, unmanned interplanetary missions and could be used to study cost-effectively the effects of such missions on these biological processes and the efficacy of new life support systems and radiation shielding technologies.

  1. Generation of transgenic wheat (Triticum aestivum L.) accumulating heterologous endo-xylanase or ferulic acid esterase in the endosperm

    DEFF Research Database (Denmark)

    Harholt, Jesper; Bach, Inga Christensen; Lind Bouquin, Solveig

    2010-01-01

    . Extensive analysis of the cell walls showed a 10%-15% increase in arabinose to xylose ratio, a 50% increase in the proportion of water-extractable arabinoxylan, and a shift in the MW of the water-extractable arabinoxylan from being mainly larger than 85 kD to being between 2 and 85 kD. Ferulic acid esterase......-expressing grains were also shrivelled, and the seed weight was decreased by 20%-50%. No ferulic acid esterase activity could be detected in wild-type grains whereas ferulic acid esterase activity was detected in transgenic lines. The grain cell walls had 15%-40% increase in water-unextractable arabinoxylan...

  2. Transgenic Primate Research Paves the Path to a Better Animal Model: Are We a Step Closer to Curing Inherited Human Genetic Disorders?

    Institute of Scientific and Technical Information of China (English)

    Anthony W.S. Chan

    2009-01-01

    While the advancement of transgenic primate models has led to a new era in modeling human conditions and has a clear impact on elucidating the mechanism of human genetic diseases, some thoughts should be considered if non-human primates are the appropriate model.

  3. Visualization and live imaging analysis of a mosquito saliva protein in host animal skin using a transgenic mosquito with a secreted luciferase reporter system.

    Science.gov (United States)

    Yamamoto, D S; Yokomine, T; Sumitani, M; Yagi, K; Matsuoka, H; Yoshida, S

    2013-12-01

    Mosquitoes inject saliva into a vertebrate host during blood feeding. The analysis of mosquito saliva in host skin is important for the elucidation of the inflammatory responses to mosquito bites, the development of antithrombotic drugs, and the transmission-blocking of vector-borne diseases. We produced transgenic Anopheles stephensi mosquitoes expressing the secretory luciferase protein (MetLuc) fused to a saliva protein (AAPP) in the salivary glands. The transgene product (AAPP-MetLuc) of transgenic mosquitoes exhibited both luciferase activity as a MetLuc and binding activity to collagen as an AAPP. The detection of luminescence in the skin of mice bitten by transgenic mosquitoes showed that AAPP-MetLuc was injected into the skin as a component of saliva via blood feeding. AAPP-MetLuc remained at the mosquito bite site in host skin with luciferase activity for at least 4 h after blood feeding. AAPP was also suspected of remaining at the site of injury caused by the mosquito bite and blocking platelet aggregation by binding to collagen. These results demonstrated the establishment of visualization and time-lapse analysis of mosquito saliva in living vertebrate host skin. This technique may facilitate the analysis of mosquito saliva after its injection into host skin, and the development of new drugs and disease control strategies.

  4. Production of transgenic pigs mediated by pseudotyped lentivirus and sperm.

    Directory of Open Access Journals (Sweden)

    Yongliang Zhang

    Full Text Available Sperm-mediated gene transfer can be a very efficient method to produce transgenic pigs, however, the results from different laboratories had not been widely repeated. Genomic integration of transgene by injection of pseudotyped lentivirus to the perivitelline space has been proved to be a reliable route to generate transgenic animals. To test whether transgene in the lentivirus can be delivered by sperm, we studied incubation of pseudotyped lentiviruses and sperm before insemination. After incubation with pig spermatozoa, 62±3 lentiviral particles were detected per 100 sperm cells using quantitative real-time RT-PCR. The association of lentivirus with sperm was further confirmed by electron microscopy. The sperm incubated with lentiviral particles were artificially inseminated into pigs. Of the 59 piglets born from inseminated 5 sows, 6 piglets (10.17% carried the transgene based on the PCR identification. Foreign gene and EGFP was successfully detected in ear tissue biopsies from two PCR-positive pigs, revealed via in situ hybridization and immunohistochemistry. Offspring of one PCR-positive boar with normal sows showed PCR-positive. Two PCR-positive founders and offsprings of PCR-positive boar were further identified by Southern-blot analysis, out of which the two founders and two offsprings were positive in Southern blotting, strongly indicating integration of foreign gene into genome. The results indicate that incubation of sperm with pseudotyped lentiviruses can incorporated with sperm-mediated gene transfer to produce transgenic pigs with improved efficiency.

  5. [Progress on transgenic mosquitoes].

    Science.gov (United States)

    Yang, Pin

    2011-04-30

    The genetically modified mosquitoes have been developed aiming to control mosquito-borne diseases by either reducing population sizes or replacing existing populations with vectors unable to transmit the disease. introduces some progress on the generation of transgenic mosquitoes and their fitness in wild population. This paper

  6. Transgenic mouse models generated by hydrodynamic transfection for genetic studies of liver cancer and preclinical testing of anti-cancer therapy.

    Science.gov (United States)

    Ju, Hye-Lim; Han, Kwang-Hyub; Lee, Jong Doo; Ro, Simon Weonsang

    2016-04-01

    Hepatocellular carcinoma (HCC) is one of the most lethal cancers worldwide; however, the genetic mechanisms underlying its pathogenesis are incompletely understood. Genetically engineered mouse (GEM) models of HCC have been developed to elucidate the role of individual cancer-related genes in hepatocarcinogenesis. However, the expensive and time-consuming processes related to generating a GEM model discourage the development of diverse genotype models. Recently, a simple and inexpensive liver-specific transgenic approach was developed, in which a hydrodynamics-based transfection (HT) method was coupled with the Sleeping Beauty transposase system. Various HT models in which different oncogenic pathways are activated and/or tumor-suppressing pathways inactivated have been developed in recent years. The applicability of HT models in liver cancer research is expected to broaden and ultimately elucidate the cooperation between oncogenic signaling pathways and aid in designing molecular therapy to target altered pathways.

  7. Toward an ideal animal model to trace donor cell fates after stem cell therapy: production of stably labeled multipotent mesenchymal stem cells from bone marrow of transgenic pigs harboring enhanced green fluorescence protein gene.

    Science.gov (United States)

    Hsiao, F S H; Lian, W S; Lin, S P; Lin, C J; Lin, Y S; Cheng, E C H; Liu, C W; Cheng, C C; Cheng, P H; Ding, S T; Lee, K H; Kuo, T F; Cheng, C F; Cheng, W T K; Wu, S C

    2011-11-01

    The discovery of postnatal mesenchymal stem cells (MSC) with their general multipotentiality has fueled much interest in the development of cell-based therapies. Proper identification of transplanted MSC is crucial for evaluating donor cell distribution, differentiation, and migration. Lack of an efficient marker of transplanted MSC has precluded our understanding of MSC-related regenerative studies, especially in large animal models such as pigs. In the present study, we produced transgenic pigs harboring an enhanced green fluorescent protein (EGFP) gene. The pigs provide a reliable and reproducible source for obtaining stable EGFP-labeled MSC, which is very useful for donor cell tracking after transplantation. The undifferentiated EGFP-tagged MSC expressed a greater quantity of EGFP while maintaining MSC multipotentiality. These cells exhibited homogeneous surface epitopes and possessed classic trilineage differentiation potential into osteogenic, adipogenic, and chondrogenic lineages, with robust EGFP expression maintained in all differentiated progeny. Injection of donor MSC can dramatically increase the thickness of infarcted myocardium and improve cardiac function in mice. Moreover, the MSC, with their strong EGFP expression, can be easily distinguished from the background autofluorescence in myocardial infarcts. We demonstrated an efficient, effective, and easy way to identify MSC after long-term culture and transplantation. With the transgenic model, we were able to obtain stem or progenitor cells in earlier passages compared with the transfection of traceable markers into established MSC. Because the integration site of the transgene was the same for all cells, we lessened the potential for positional effects and the heterogeneity of the stem cells. The EGFP-transgenic pigs may serve as useful biomedical and agricultural models of somatic stem cell biology.

  8. A Simple Line Drawing Definition and Transfer Model for Facial Animation Generation

    Directory of Open Access Journals (Sweden)

    Qingxiang Wang

    2013-11-01

    Full Text Available The Line Drawing Animation is an active research area in Non-Photorealistic Rendering. Many researchs are focused on the skech abastract, like such as portrait drawing of human and animation generation. However most of the model are too complex to calculate or pay attention to the details which are not stable that are not suitable for realtime transfer for continuous sequence of video. This paper proposes a simple line drawing definition and transfer model with Bézier Curves and the core of the AAM fit parameters. The facial line drawings have seven basic emotions include neutral, happiness, anger, disgust, fear, sadness and surprised. Each of the drawing in a specific model is consisted of a same set of cubic Bézier curves. The proposed model is suitabl for shape conbination anmition. In the experiment, the AAM method is used to get the facial features of the face and then find the nearest combination of the emotion to transfer to the line drawing model. The result shows that the method is simple and fast. Only a few of the parameters are needed to transfer that is suitable to record and communication.

  9. Fast calculation method of computer generated hologram animation for viewpoint parallel shift and rotation using Fourier transform optical system.

    Science.gov (United States)

    Watanabe, Ryosuke; Yamaguchi, Kazuhiro; Sakamoto, Yuji

    2016-01-20

    Computer generated hologram (CGH) animations can be made by switching many CGHs on an electronic display. Some fast calculation methods for CGH animations have been proposed, but one for viewpoint movement has not been proposed. Therefore, we designed a fast calculation method of CGH animations for viewpoint parallel shifts and rotation. A Fourier transform optical system was adopted to expand the viewing angle. The results of experiments were that the calculation time of our method was over 6 times faster than that of the conventional method. Furthermore, the degradation in CGH animation quality was found to be sufficiently small.

  10. Selection of in vitro produced, transgenic embryos by nested PCR for efficient production of transgenic goats.

    Science.gov (United States)

    Huang, S Z; Huang, Y; Chen, M J; Zeng, F Y; Ren, Z R; Zeng, Y T

    2001-09-01

    The production of valuable pharmaceutical proteins using transgenic animals as bioreactors has become one of the goals of biotechnology. However, the efficiency of producing transgenic animals by means of pronuclear microinjection is low. This may be attributed in part to the low integration rate of foreign DNA. Therefore, a large number of recipients are required to produce transgenic animals. We recently developed a transgenic procedure that combined the techniques of goat oocyte in vitro maturation (IVM), in vitro fertilization (IVF), microinjection, preimplantation selection of the transgenic embryos with nested PCR and transferring the transgenic embryos into the recipient goat uterus to produce transgenic goats. Thirty-seven transgenic embryos determined by nested PCR were transferred to thirty-two recipient goats. In the end, four live-born kids were produced. As predicted, all the live kids were transgenic as identified by PCR as well as Southern blot hybridization, The integration rate was 100% (4/4) which was completely in accordance with the results of embryo preimplantation detection. The results showed a significant decrease in the number of recipients required as only 8 recipients (32/4) were needed to obtain one live transgenic goat. We suggest that the transgenic system described herein may provide an improved way to efficiently produce transgenic goats on a large scale.

  11. Overview of Animal Models of Obesity

    Science.gov (United States)

    Lutz, Thomas A.; Woods, Stephen C.

    2012-01-01

    This is a review of animal models of obesity currently used in research. We have focused upon more commonly utilized models since there are far too many newly created models to consider, especially those caused by selective molecular genetic approaches modifying one or more genes in specific populations of cells. Further, we will not discuss the generation and use of inducible transgenic animals (induced knock-out or knock-in) even though they often bear significant advantages compared to traditional transgenic animals; influences of the genetic modification during the development of the animals can be minimized. The number of these animal models is simply too large to be covered in this chapter. PMID:22948848

  12. Transgenic rabbits that overexpress the neonatal Fc receptor (FcRn generate higher quantities and improved qualities of anti-thymocyte globulin (ATG.

    Directory of Open Access Journals (Sweden)

    Mária Baranyi

    Full Text Available Immune suppression with rabbit anti-thymocyte globulin (rATG is a well-established therapeutic concept for preventing host rejection of transplanted organs and graft versus host disease. Increasing the efficiency of rATG production by reducing the number of animals would be highly beneficial to lower cost and to improve quality standards. We have developed transgenic (Tg mice and rabbits that overexpress the neonatal Fc receptor (FcRn and have shown an augmented humoral immune response in these animals. To test whether our FcRn Tg rabbits produced rATG more efficiently, we immunized them and their New Zealand White controls with live Jurkat cells. By day 21 after immunization, Tg animals produced significantly, 1.5 times higher amount of total IgG compared to their wt littermates. Also, the binding efficiency of Tg sera to Jurkat cells and their complement-mediated cytotoxicity was significantly higher. The purified Tg IgG preparation contained 2.6 the amount of Jurkat specific IgG as the wt preparation analyzed by complement-mediated lysis, suggesting greater antigen-specific B cell activation in the Tg rabbits. To test this hypothesis, immunization with ovalbumin and human α1-antitrypsin was performed, resulting in significantly greater numbers of antigen-specific B-cells in the FcRn Tg rabbits as compared with wt controls. The shift towards significantly larger populations of antigen-specific B cells relative to the non-specific B cell pool is further corroborated by our previous findings in FcRn Tg mice. Consequently, our FcRn Tg rabbits have the potential to offer substantial qualitative and quantitative improvements for the production of rATG and other polyclonal or monoclonal antibodies.

  13. Hepatitis B Virus Splice-Generated Protein Induces T-Cell Responses in HLA-Transgenic Mice and Hepatitis B Virus-Infected Patients▿

    Science.gov (United States)

    Mancini-Bourgine, Maryline; Bayard, Florence; Soussan, Patrick; Deng, Qiang; Lone, Yu-Chun; Kremsdorf, Dina; Michel, Marie-Louise

    2007-01-01

    Hepatitis B virus splice-generated protein (HBSP), encoded by a spliced hepatitis B virus RNA, was recently identified in liver biopsy specimens from patients with chronic active hepatitis B. We investigated the possible generation of immunogenic peptides by the processing of this protein in vivo. We identified a panel of potential epitopes in HBSP by using predictive computational algorithms for peptide binding to HLA molecules. We used transgenic mice devoid of murine major histocompatibility complex (MHC) class I molecules and positive for human MHC class I molecules to characterize immune responses specific for HBSP. Two HLA-A2-restricted peptides and one immunodominant HLA-B7-restricted epitope were identified following the immunization of mice with DNA vectors encoding HBSP. Most importantly, a set of overlapping peptides covering the HBSP sequence induced significant HBSP-specific T-cell responses in peripheral blood mononuclear cells from patients with chronic hepatitis B. The response was multispecific, as several epitopes were recognized by CD8+ and CD4+ human T cells. This study provides the first evidence that this protein generated in vivo from an alternative reading frame of the hepatitis B virus genome activates T-cell responses in hepatitis B virus-infected patients. Given that hepatitis B is an immune response-mediated disease, the detection of T-cell responses directed against HBSP in patients with chronic hepatitis B suggests a potential role for this protein in liver disease progression. PMID:17360751

  14. Efficient generation of transgene-free human induced pluripotent stem cells (iPSCs) by temperature-sensitive Sendai virus vectors.

    Science.gov (United States)

    Ban, Hiroshi; Nishishita, Naoki; Fusaki, Noemi; Tabata, Toshiaki; Saeki, Koichi; Shikamura, Masayuki; Takada, Nozomi; Inoue, Makoto; Hasegawa, Mamoru; Kawamata, Shin; Nishikawa, Shin-Ichi

    2011-08-23

    After the first report of induced pluripotent stem cells (iPSCs), considerable efforts have been made to develop more efficient methods for generating iPSCs without foreign gene insertions. Here we show that Sendai virus vector, an RNA virus vector that carries no risk of integrating into the host genome, is a practical solution for the efficient generation of safer iPSCs. We improved the Sendai virus vectors by introducing temperature-sensitive mutations so that the vectors could be easily removed at nonpermissive temperatures. Using these vectors enabled the efficient production of viral/factor-free iPSCs from both human fibroblasts and CD34(+) cord blood cells. Temperature-shift treatment was more effective in eliminating remaining viral vector-related genes. The resulting iPSCs expressed human embryonic stem cell markers and exhibited pluripotency. We suggest that generation of transgene-free iPSCs from cord blood cells should be an important step in providing allogeneic iPSC-derived therapy in the future.

  15. Full life-cycle assessment of gene flow consistent with fitness differences in transgenic and wild-type Japanese medaka fish (Oryzias latipes).

    Science.gov (United States)

    Pennington, Kelly M; Kapuscinski, Anne R; Morton, Michael S; Cooper, Anne M; Miller, Loren M

    2010-01-01

    Transgenic fish in development for aquaculture could escape from farms and interbreed with wild relatives in the nearby environment. Predicting whether escapes would result in transgene introgression is a major challenge in assessing environmental risks of transgenic fish. Previous studies have simulated gene flow from transgenic fish using mathematical modeling of fitness traits to predict the relative selective value of transgenic genotypes. Here, we present the first study of gene flow over the full life cycle in openly-breeding populations of transgenic animals, along with measurement of fitness traits. We conducted two invasion experiments in which we released two lines of growth-enhanced transgenic fish (T67 and T400), Japanese medaka (Oryzias latipes), into populations of wild-type (W) medaka in structured mesocosms. After several generations, the frequency of transgenic fish varied across replicates in the first invasion experiment (6 months), but the frequency of transgenic fish decreased in the second experiment (19 months). We also measured selected fitness traits in transgenic and wild-type medaka because these traits could be used to predict the relative selective value of a genotype. We found that: T400 males were more fertile than W males; offspring of W females lived longer than those with transgenic mothers; and W and T67 females reached sexual maturity sooner than T400 females. In contrast with other research that reported larger transgenic males had a mating advantage, we found that W males obtained more matings with females than T males; genetic background effects may account for our differing results as we compared W and T fish derived from different strains. The decreasing frequency of transgenic fish in the second invasion experiment suggests that transgenic fish had a selective disadvantage in the experimental environment. Our finding of transgenic advantage of some fitness traits and wild-type advantage in others is consistent with our

  16. Tetracycline-inducible Expression Systems: New Strategies and Practices in the Transgenic Mouse Modeling

    Institute of Scientific and Technical Information of China (English)

    Yan SUN; Xigu CHEN; Dong XIAO

    2007-01-01

    To accurately analyze the function of transgene(s) of interest in transgenic mice, and to generate credible transgenic animal models for multifarious human diseases to precisely mimic human disease states, it is critical to tightly regulate gene expression in the animals in a conditional manner. The ability to turn gene expression on or off in the restricted cells or tissues at specific time permits unprecedented flexibility in dissecting gene functions in health and disease. Pioneering studies in conditional transgene expression have brought about the development of a wide variety of controlled gene expression systems, which meet this criterion. Among them, the tetracycline-controlled expression systems (e.g. Tet-off system and Tet-on system) have been used extensively in vitro and in vivo. In recent years, some strategies derived from tetracycline-inducible system alone, as well as the combined use of Tet-based systems and Cre/lox P switching gene expression system, have been newly developed to allow more flexibility for exploring gene functions in health and disease, and produce credible transgenic animal models for various human diseases. In this review these newly developed strategies are discussed.

  17. Effects of antagonists of growth hormone-releasing hormone (GHRH) on GH and insulin-like growth factor I levels in transgenic mice overexpressing the human GHRH gene, an animal model of acromegaly.

    Science.gov (United States)

    Kovacs, M; Kineman, R D; Schally, A V; Zarandi, M; Groot, K; Frohman, L A

    1997-11-01

    Transgenic mice overexpressing the human GH-releasing hormone (hGHRH) gene, an animal model of acromegaly, were used to investigate the effects of potent GHRH antagonists MZ-4-71 and MZ-5-156 on the excessive GH and insulin-like growth factor I (IGF-I) secretion caused by overproduction of hGHRH. Because metallothionein (MT)-GHRH mice express the hGHRH transgene in various tissues, including the pituitary and hypothalamus, initial experiments focused on the effectiveness of the GHRH antagonists in blocking basal and stimulated GH secretion from pituitary cells in vitro. Both MZ-4-71 and MZ-5-156 suppressed basal release of GH from superfused MT-GHRH pituitary cells, apparently by blocking the action of endogenously produced hGHRH. In addition, these antagonists effectively eliminated the response to stimulatory action of exogenous hGHRH(1-29)NH2 (30 and 100 nM). To ascertain whether MZ-4-71 and MZ-5-156 could antagonize the effect of hGHRH hyperstimulation in vivo, each antagonist was administered to MT-GHRH transgenic mice in a single iv dose of 10-200 microg. Both compounds decreased serum GH levels in transgenic mice by 39-72% at 1 h after injection. The inhibitory effect of 50 microg MZ-5-156 was maintained for 5 h. Twice daily ip administration of 100 microg MZ-5-156 for 3 days suppressed the highly elevated serum GH and IGF-I concentrations in transgenic mice by 56.8% and 39.0%, respectively. This treatment also reduced IGF-I messenger RNA levels in the liver by 21.8% but did not affect the level of GH messenger RNA in the pituitary. Our results demonstrate that GHRH antagonists MZ-4-71 and MZ-5-156 can inhibit elevated GH levels caused by overproduction of hGHRH. The suppression of circulating GH concentrations induced by the antagonists seems to be physiologically relevant, because both IGF-I secretion and synthesis also were reduced. Our findings, showing the suppression of GH and IGF-I secretion with GHRH antagonists, suggest that this class of analogs

  18. The challenges of commercializing second-generation transgenic crop traits necessitate the development of international public sector research infrastructure.

    Science.gov (United States)

    Rothstein, Steven J; Bi, Yong-Mei; Coneva, Viktoriya; Han, Mei; Good, Allen

    2014-10-01

    It has been 30 years since the first transformation of a gene into a plant species, and since that time a number of biotechnology products have been developed, with the most important being insect- and herbicide-resistant crops. The development of second-generation products, including nutrient use efficiency and tolerance to important environmental stressors such as drought, has, up to this time, been less successful. This is in part due to the inherent complexities of these traits and in part due to limitations in research infrastructure necessary for public sector researchers to test their best ideas. Here we discuss lessons from previous work in the generation of the first-generation traits, as well as work from our labs and others on identifying genes for nitrogen use efficiency. We then describe some of the issues that have impeded rapid progress in this area. Finally, we propose the type of public sector organization that we feel is necessary to make advances in important second-generation traits such as nitrogen use efficiency.

  19. Targeted therapy of animal eyes with tumors by laser-generated focused ultrasound (Conference Presentation)

    Science.gov (United States)

    Lee, Taehwa; Luo, Wei; Demirci, Hakan; Guo, L. Jay

    2016-03-01

    Cavitation therapy based on high-amplitude focused ultrasound (e.g., Histotripsy) has shown great promise in clinical trials. The technique realizes localized treatments of tissues and diseased cells by controlling cavitation zones, which can be even smaller than its acoustic spot sizes. Also, the short pressure pulse used in the technique can minimize the unwanted heat accumulation, which the conventional piezoelectric transducers suffer from due to low operating frequencies and relatively long acoustic pulses. However, this modality requires bulky system composed of array of piezoelectric elements and electric amplifiers in order to obtain high pressure amplitude. Moreover, especially when treating an area much smaller than the acoustic spot size, this approach may be vulnerable to nucleation sites within the focal volume, which can potentially induce cavitation and thus enlarge the total treatment area. Here, we show targeted cell-level therapy by using laser generated ultrasound. By employing a concave lens coated by a carbon nanotube (CNT)-polymer composite, high-amplitude acoustic pressure can be obtained at a tight focal spot (<100 um). The small focal spot, comparable to cavitation zone, lead to controlled cavitation treatment. Such feature can be exploited for treating intraocular tumors but without harming other parts of the eye (e.g. healthy retina and choroid) and therefore preserve the vision of the patients. We demonstrate that the localized disruption effects can be used for cell-level surgery to remove cells and to kill cells. Some experimental examples are shown using animal eyeballs.

  20. Observation of tendon repair in animal model using second-harmonic-generation microscopy

    Science.gov (United States)

    Hase, Eiji; Minamikawa, Takeo; Sato, Katsuya; Takahashi, Mitsuhiko; Yasui, Takashi

    2016-03-01

    Tendon rupture is a trauma difficult to recover the condition before injury. In previous researches, tensile test and staining method have been widely used to elucidate the mechanism of the repair process from the viewpoints of the mechanical property and the histological findings. However, since both methods are destructive and invasive, it is difficult to obtain both of them for the same sample. If both the mechanical property and the histological findings can be obtained from the same sample, one may obtain new findings regarding mechanisms of tendon repairing process. In this paper, we used second-harmonic-generation (SHG) microscopy, showing high selectivity and good image contrast to collagen molecules as well as high spatial resolution, optical three-dimensional sectioning, deep penetration, and without additional staining. Since SHG light intensity sensitively reflects the structural maturity of collagen molecule and its aggregates, it will be a good indicator for the repairing degree of the ruptured tendon. From comparison of SHG images between the 4-weeks-repaired tendon and the sound tendon in the animal model, we confirmed that SHG light intensity of the repaired tendon was significantly lower than that of the sound tendon, indicating that the collagen structure in the repaired tendon is still immature. Furthermore, we performed both SHG imaging and the tensile test for the same sample, and confirmed a correlation between them. This result shows a potential of SHG light for an indicator of the histological and mechanical recovery of the ruptured tendon.

  1. COMPARISON TRANSGENIC AND NON-TRANSGENIC MILK QUALITY

    Directory of Open Access Journals (Sweden)

    Peter Chrenek

    2012-02-01

    Full Text Available Transgenic founder rabbits carrying a gene construct consisting of a 2.5 kb murine whey acidic protein promoter (mWAP, 7.2 kb of the human clotting factor VIII (hFVIII cDNA and 4.6 kb of 3’ flanking sequences of mWAP gene were crossed for five generations. Transgenic females showed high level of recombinant hFVIII (rhFVIII mRNA expression in biopsed mammary gland tissues. The presence of the mWAP-hFVIII transgene in rabbit genome and secretion of rhFVIII into milk of transgenic females (F1, F2, F3, F4 and F5 generation did not have any adverse phenotypic effect on milk quality.

  2. 转基因技术:糖尿病的动物模型构建及治疗%Transgenic Technology:Establishment of Animal Models and Treatment of Diabetes Mellitus

    Institute of Scientific and Technical Information of China (English)

    王鹤霏; 刘东军

    2015-01-01

    糖尿病是严重危害人类健康的疾病,近年来,随着转基因理论及技术的发展,很多科学家都致力于应用转基因技术治疗糖尿病,并取得了显著成效。主要综述了目前在转基因糖尿病动物模型构建和糖尿病治疗方面取得的诸多成果,以及面临的挑战。提出了今后在建立高效的胰岛素基因转移体系、选择与β细胞生理特点相似又免受自身免疫系统攻击的靶细胞、基因表达的持续性和糖尿病易感基因的鉴定等方向的突破,可以加快糖尿病的基因治疗用于临床的步伐。%Diabetes mellitus endangers human health seriously. In recent years, with the development of transgenic theory and technology, many researchers are devoted to treat diabetes by applying transgenic technology. Therefore, some remarkable achievements have been achieved. This review describes the current achievements and challenges in the establishment of animal models and treatment ofD. mellitus by transgenic technology. In future the breakthroughs from following aspects may accelerate the gene therapy ofD. mellitus in clinical practice, such as the establishment of effective insulin gene transfer system, selection of target cells resembling physiological characteristics ofβ cells and preventing them from the attack of own immune system, sustainability of gene expression and identification ofD. mellitus susceptible genes.

  3. Handmade cloned transgenic sheep rich in omega-3 Fatty acids.

    Science.gov (United States)

    Zhang, Peng; Liu, Peng; Dou, Hongwei; Chen, Lei; Chen, Longxin; Lin, Lin; Tan, Pingping; Vajta, Gabor; Gao, Jianfeng; Du, Yutao; Ma, Runlin Z

    2013-01-01

    Technology of somatic cell nuclear transfer (SCNT) has been adapted worldwide to generate transgenic animals, although the traditional procedure relies largely on instrumental micromanipulation. In this study, we used the modified handmade cloning (HMC) established in cattle and pig to produce transgenic sheep with elevated levels of omega-3 (n-3) fatty acids. Codon-optimized nematode mfat-1 was inserted into a eukaryotic expression vector and was transferred into the genome of primary ovine fibroblast cells from a male Chinese merino sheep. Reverse transcriptase PCR, gas chromatography, and chromosome analyses were performed to select nuclear donor cells capable of converting omega-6 (n-6) into n-3 fatty acids. Blastocysts developed after 7 days of in vitro culture were surgically transplanted into the uterus of female ovine recipients of a local sheep breed in Xinjiang. For the HMC, approximately 8.9% (n  =925) of reconstructed embryos developed to the blastocyst stage. Four recipients became pregnant after 53 blastocysts were transplanted into 29 naturally cycling females, and a total of 3 live transgenic lambs were produced. Detailed analyses on one of the transgenic lambs revealed a single integration of the modified nematode mfat-1 gene at sheep chromosome 5. The transgenic sheep expressed functional n-3 fatty acid desaturase, accompanied by more than 2-folds reduction of n-6/n-3 ratio in the muscle (psheep produced by the HMC. Compared to the traditional SCNT method, HMC showed an equivalent efficiency but proved cheaper and easier in operation.

  4. Transgenics, agroindustry and food sovereignty

    Directory of Open Access Journals (Sweden)

    Xavier Alejandro León Vega

    2014-10-01

    Full Text Available Food sovereignty has been implemented constitutionally in Ecuador; however, many of the actions and policies are designed to benefit the dominant model of food production, based in agroindustry, intensive monocultures, agrochemicals and transgenics. This article reflects upon the role of family farming as a generator of food sovereignty, and secondly the threat to them by agroindustry agriculture based in transgenic. The role played by food aid in the introduction of transgenic in Latin America and other regions of the world is also analyzed.

  5. On the Legal Supervision of Transgenic Animal Bioreactor Pharmacy in China%论我国转基因动物生物反应器制药的法律监管

    Institute of Scientific and Technical Information of China (English)

    刘旭霞; 沈大力; 耿宁

    2014-01-01

    In recent years,it has become a hot spot to produce medicines with transgenic animal bioreactors in the development and utilization of biotechnology .But the potential benefits and risks coexist in the application of transgenic animal biotechnology .There are no special provisions in the present "Pharmaceutical Administration Law of the People's Republic of China"and its implementing regulations , and other related pharmacy management measures.There are obvious loopholes in the legal system .Therefore,it is necessary to establish a comprehensive system including a preclinical trial system , a clinical quality management system , new pharmacy licensing system , pharmaceutical production and market access system ,and post-marketing adverse reaction reporting and recalling system,to legalize the development of transgenic animal bioreacotrs pharmacy .%近年来,通过转基因动物生物反应器生产药物成为生物技术开发利用的热点。但转基因动物生物技术应用的潜在收益和风险并存,现有《药品管理法》及其实施条例,以及其他相关药品管理办法均未对转基因动物生物反应器药品监管进行专门规定,制度上存在明显漏洞。因此,有必要建立转基因动物生物反应器制药临床前实验制度、临床实验质量管理制度、新药上市许可制度、药品生产与市场准入制度,及上市后不良反应报告和召回制度,促使转基因动物生物反应器制药走上法制化发展轨道。

  6. Welfare assessment in transgenic pigs expressing green fluorescent protein (GFP)

    DEFF Research Database (Denmark)

    Huber, Reinhard C.; Remuge, Liliana; Carlisle, Ailsa

    2012-01-01

    Since large animal transgenesis has been successfully attempted for the first time about 25 years ago, the technology has been applied in various lines of transgenic pigs. Nevertheless one of the concerns with the technology—animal welfare—has not been approached through systematic assessment...... and statements regarding the welfare of transgenic pigs have been based on anecdotal observations during early stages of transgenic programs. The main aim of the present study was therefore to perform an extensive welfare assessment comparing heterozygous transgenic animals expressing GFP with wildtype animals...... months. The absence of significant differences between GFP and wildtype animals in the parameters observed suggests that the transgenic animals in question are unlikely to suffer from deleterious effects of transgene expression on their welfare and thus support existing anecdotal observations of pigs...

  7. Efficient Generation of Dancing Animation Synchronizing with Music Based on Meta Motion Graphs

    Science.gov (United States)

    Xu, Jianfeng; Takagi, Koichi; Sakazawa, Shigeyuki

    This paper presents a system for automatic generation of dancing animation that is synchronized with a piece of music by re-using motion capture data. Basically, the dancing motion is synthesized according to the rhythm and intensity features of music. For this purpose, we propose a novel meta motion graph structure to embed the necessary features including both rhythm and intensity, which is constructed on the motion capture database beforehand. In this paper, we consider two scenarios for non-streaming music and streaming music, where global search and local search are required respectively. In the case of the former, once a piece of music is input, the efficient dynamic programming algorithm can be employed to globally search a best path in the meta motion graph, where an objective function is properly designed by measuring the quality of beat synchronization, intensity matching, and motion smoothness. In the case of the latter, the input music is stored in a buffer in a streaming mode, then an efficient search method is presented for a certain amount of music data (called a segment) in the buffer with the same objective function, resulting in a segment-based search approach. For streaming applications, we define an additional property in the above meta motion graph to deal with the unpredictable future music, which guarantees that there is some motion to match the unknown remaining music. A user study with totally 60 subjects demonstrates that our system outperforms the stat-of-the-art techniques in both scenarios. Furthermore, our system improves the synthesis speed greatly (maximal speedup is more than 500 times), which is essential for mobile applications. We have implemented our system on commercially available smart phones and confirmed that it works well on these mobile phones.

  8. Generation of a Focused Poly(amino ether Library: Polymer-mediated Transgene Delivery and Gold-Nanorod based Theranostic Systems

    Directory of Open Access Journals (Sweden)

    Lucas Vu, James Ramos, Thrimoorthy Potta, Kaushal Rege

    2012-01-01

    Full Text Available A focused library of twenty-one cationic poly(amino ethers was synthesized following ring-opening polymerization of two diglycidyl ethers by different oligoamines. The polymers were screened in parallel for plasmid DNA (pDNA delivery, and transgene expression efficacies of individual polymers were compared to those of 25 kDa polyethylenimine (PEI, a current standard for polymer-mediated transgene delivery. Seven lead polymers that demonstrated higher transgene expression than PEI in pancreatic and prostate cancer cells lines were identified from the screen. All seven lead polymers showed highest transgene expression at a polymer:pDNA weight ratio of 5:1 in the MIA PaCa-2 pancreatic cancer cell line. Among the conditions studied, transgene expression efficacy correlated with minimal polymer cytotoxicity but not polyplex sizes. In addition, this study indicated that methylene spacing between amine centers in the monomers, amine content, and molecular weight of the polymers are all significant factors and should be considered when designing polymers for transgene delivery. A lead effective polymer was employed for coating gold nanorods, leading to theranostic nanoassemblies that possess combined transgene delivery and optical imaging capabilities, leading to potential theranostic systems.

  9. The temporal expression pattern of alpha-synuclein modulates olfactory neurogenesis in transgenic mice.

    Directory of Open Access Journals (Sweden)

    Sebastian R Schreglmann

    Full Text Available Adult neurogenesis mirrors the brain´s endogenous capacity to generate new neurons throughout life. In the subventricular zone/ olfactory bulb system adult neurogenesis is linked to physiological olfactory function and has been shown to be impaired in murine models of neuronal alpha-Synuclein overexpression. We analyzed the degree and temporo-spatial dynamics of adult olfactory bulb neurogenesis in transgenic mice expressing human wild-type alpha-Synuclein (WTS under the murine Thy1 (mThy1 promoter, a model known to have a particularly high tg expression associated with impaired olfaction.Survival of newly generated neurons (NeuN-positive in the olfactory bulb was unchanged in mThy1 transgenic animals. Due to decreased dopaminergic differentiation a reduction in new dopaminergic neurons within the olfactory bulb glomerular layer was present. This is in contrast to our previously published data on transgenic animals that express WTS under the control of the human platelet-derived growth factor β (PDGF promoter, that display a widespread decrease in survival of newly generated neurons in regions of adult neurogenesis, resulting in a much more pronounced neurogenesis deficit. Temporal and quantitative expression analysis using immunofluorescence co-localization analysis and Western blots revealed that in comparison to PDGF transgenic animals, in mThy1 transgenic animals WTS is expressed from later stages of neuronal maturation only but at significantly higher levels both in the olfactory bulb and cortex.The dissociation between higher absolute expression levels of alpha-Synuclein but less severe impact on adult olfactory neurogenesis in mThy1 transgenic mice highlights the importance of temporal expression characteristics of alpha-Synuclein on the maturation of newborn neurons.

  10. A modified RMCE-compatible Rosa26 locus for the expression of transgenes from exogenous promoters.

    Science.gov (United States)

    Tchorz, Jan S; Suply, Thomas; Ksiazek, Iwona; Giachino, Claudio; Cloëtta, Dimitri; Danzer, Claus-Peter; Doll, Thierry; Isken, Andrea; Lemaistre, Marianne; Taylor, Verdon; Bettler, Bernhard; Kinzel, Bernd; Mueller, Matthias

    2012-01-01

    Generation of gain-of-function transgenic mice by targeting the Rosa26 locus has been established as an alternative to classical transgenic mice produced by pronuclear microinjection. However, targeting transgenes to the endogenous Rosa26 promoter results in moderate ubiquitous expression and is not suitable for high expression levels. Therefore, we now generated a modified Rosa26 (modRosa26) locus that combines efficient targeted transgenesis using recombinase-mediated cassette exchange (RMCE) by Flipase (Flp-RMCE) or Cre recombinase (Cre-RMCE) with transgene expression from exogenous promoters. We silenced the endogenous Rosa26 promoter and characterized several ubiquitous (pCAG, EF1α and CMV) and tissue-specific (VeCad, αSMA) promoters in the modRosa26 locus in vivo. We demonstrate that the ubiquitous pCAG promoter in the modRosa26 locus now offers high transgene expression. While tissue-specific promoters were all active in their cognate tissues they additionally led to rare ectopic expression. To achieve high expression levels in a tissue-specific manner, we therefore combined Flp-RMCE for rapid ES cell targeting, the pCAG promoter for high transgene levels and Cre/LoxP conditional transgene activation using well-characterized Cre lines. Using this approach we generated a Cre/LoxP-inducible reporter mouse line with high EGFP expression levels that enables cell tracing in live cells. A second reporter line expressing luciferase permits efficient monitoring of Cre activity in live animals. Thus, targeting the modRosa26 locus by RMCE minimizes the effort required to target ES cells and generates a tool for the use exogenous promoters in combination with single-copy transgenes for predictable expression in mice.

  11. Potential molecular consequences of transgene integration: The R6/2 mouse example

    Science.gov (United States)

    Jacobsen, Jessie C.; Erdin, Serkan; Chiang, Colby; Hanscom, Carrie; Handley, Renee R.; Barker, Douglas D.; Stortchevoi, Alex; Blumenthal, Ian; Reid, Suzanne J.; Snell, Russell G.; MacDonald, Marcy E.; Morton, A. Jennifer; Ernst, Carl; Gusella, James F.; Talkowski, Michael E.

    2017-01-01

    Integration of exogenous DNA into a host genome represents an important route to generate animal and cellular models for exploration into human disease and therapeutic development. In most models, little is known concerning structural integrity of the transgene, precise site of integration, or its impact on the host genome. We previously used whole-genome and targeted sequencing approaches to reconstruct transgene structure and integration sites in models of Huntington’s disease, revealing complex structural rearrangements that can result from transgenesis. Here, we demonstrate in the R6/2 mouse, a widely used Huntington’s disease model, that integration of a rearranged transgene with coincident deletion of 5,444 bp of host genome within the gene Gm12695 has striking molecular consequences. Gm12695, the function of which is unknown, is normally expressed at negligible levels in mouse brain, but transgene integration has resulted in cortical expression of a partial fragment (exons 8–11) 3’ to the transgene integration site in R6/2. This transcript shows significant expression among the extensive network of differentially expressed genes associated with this model, including synaptic transmission, cell signalling and transcription. These data illustrate the value of sequence-level resolution of transgene insertions and transcription analysis to inform phenotypic characterization of transgenic models utilized in therapeutic research. PMID:28120936

  12. T-Cell Mediated Immune Responses Induced in ret Transgenic Mouse Model of Malignant Melanoma

    Energy Technology Data Exchange (ETDEWEB)

    Abschuetz, Oliver [Skin Cancer Unit, German Cancer Research Center (DKFZ), Heidelberg and Department of Dermatology, Venereology and Allergology, University Medical Center Mannheim, Ruprecht-Karl University of Heidelberg, Mannheim , Heidelberg 69120 (Germany); Osen, Wolfram [Division of Translational Immunology, German Cancer Center, Heidelberg 69120 (Germany); Frank, Kathrin [Skin Cancer Unit, German Cancer Research Center (DKFZ), Heidelberg and Department of Dermatology, Venereology and Allergology, University Medical Center Mannheim, Ruprecht-Karl University of Heidelberg, Mannheim , Heidelberg 69120 (Germany); Kato, Masashi [Unit of Environmental Health Sciences, Department of Biomedical Sciences, College of Life and Health Sciences, Chubu University, Aichi 487-8501 (Japan); Schadendorf, Dirk [Department of Dermatology, University Hospital Essen, Essen 45122 (Germany); Umansky, Viktor, E-mail: v.umansky@dkfz.de [Skin Cancer Unit, German Cancer Research Center (DKFZ), Heidelberg and Department of Dermatology, Venereology and Allergology, University Medical Center Mannheim, Ruprecht-Karl University of Heidelberg, Mannheim , Heidelberg 69120 (Germany)

    2012-04-26

    Poor response of human malignant melanoma to currently available treatments requires a development of innovative therapeutic strategies. Their evaluation should be based on animal models that resemble human melanoma with respect to genetics, histopathology and clinical features. Here we used a transgenic mouse model of spontaneous skin melanoma, in which the ret transgene is expressed in melanocytes under the control of metallothionein-I promoter. After a short latency, around 25% mice develop macroscopic skin melanoma metastasizing to lymph nodes, bone marrow, lungs and brain, whereas other transgenic mice showed only metastatic lesions without visible skin tumors. We found that tumor lesions expressed melanoma associated antigens (MAA) tyrosinase, tyrosinase related protein (TRP)-1, TRP-2 and gp100, which could be applied as targets for the immunotherapy. Upon peptide vaccination, ret transgenic mice without macroscopic melanomas were able to generate T cell responses not only against a strong model antigen ovalbumin but also against typical MAA TRP-2. Although mice bearing macroscopic primary tumors could also display an antigen-specific T cell reactivity, it was significantly down-regulated as compared to tumor-free transgenic mice or non-transgenic littermates. We suggest that ret transgenic mice could be used as a pre-clinical model for the evaluation of novel strategies of melanoma immunotherapy.

  13. Applications of Somatic Cell Nuclear Transfer Technology in Transgenic Animal Research%体细胞核移植技术在转基因动物研究中的应用

    Institute of Scientific and Technical Information of China (English)

    马毅; 曹斌云

    2001-01-01

    利用基因工程技术将目的基因整合入动物体细胞染色体中,并将其作为供体核移植入受体——去核卵母细胞构成重建胚,然后将其移植入假孕母体,待其妊娠、分娩,便可得到经定向遗传修饰的转基因克隆动物。本文就这一领域的发展历史、研究现状、应用前景及存在的问题等方面作一概述。%Transgenic techmology has developed quickly in recent years.Thegene of interest could be integrated into the chromosome of donor-somatic cell of animal.After donor is transferred into the recipient-enucleated oocyte,the reconstructed embryo is transplanted into the female animal to establish pregency until the birth of offspring.Thus,we can get atransgenic clone animal,which is genetically modified.This paper will review the history of development,the status of research,the future of applications and the existing problems in this field.

  14. Generating knock-in parasites: integration of an ornithine decarboxylase transgene into its chromosomal locus in Leishmania donovani.

    Science.gov (United States)

    Roberts, Sigrid C; Kline, Chelsey; Liu, Wei; Ullman, Buddy

    2011-06-01

    Leishmania null mutants created by targeted gene replacement are typically complemented with chimeric episomes harboring the replaced gene in order to validate that the observed phenotype is due to the specific gene deletion. However, the current inventory of available episomes for complementation of genetic lesions in Leishmania is unstable in the absence of drug selection, and levels of gene expression cannot be controlled, especially in vivo. To circumvent this impediment, a strategy to re-introduce the targeted gene into the original chromosomal locus to generate "knock-in" parasites within selectable null backgrounds has been developed. A genomic fragment encompassing the ornithine decarboxylase locus and lacking heterologous DNA sequences was transfected into ornithine decarboxylase-deficient Leishmania donovani. The construct randomly integrated into either chromosomal allele by homologous recombination restoring polyamine prototrophy and revealing that LdODC was functionally expressed in the knock-in clones. This strategy offers a mechanism for complementing a genetic lesion amenable to positive selection in a manner that facilitates stable gene expression from its original locus in the absence of continuous drug pressure.

  15. Rational design, fabrication, characterization and in vitro testing of biodegradable microparticles that generate targeted and sustained transgene expression in HepG2 liver cells

    Science.gov (United States)

    Intra, Janjira; Salem, Aliasger K.

    2017-01-01

    was similar to PLGA particles loaded with ApoE pDNA alone. The transfection efficiency of all particle formulations prepared with ApoE pDNA was significantly higher in HepG2 cells when compared to HEK293 and COS7 cell lines. The release of PEI-pDNA complexes from particles prepared with different PLGA polymer compositions including PLGA 50-50, PLGA 75-25 and PLGA 85-15 was sustained in all cases but the release profile was dependent on the polymer composition. Transmission electron microscopy images showed that PEI-pDNA complexes remained structurally intact after release. The optimum formulation for PLGA particles loaded with PEI-ApoE pDNA complexes was prepared using 2% PVA, 50-50 PLGA compositions and N/P ratios of 5 to 10. Strong sustained transgene expression in HepG2 cells was generated by PLGA PEI ApoE pDNA particles up to the full 13 days tested. PMID:20681752

  16. Integration mechanisms of transgenes and population fitness of GH transgenic fish

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    It has been more than 20 years since the first batch of transgenic fish was produced. Five stable germ-line transmitted growth hormone (GH) transgenic fish lines have been generated. This paper reviews the mechanisms of integration and gene targeting of the transgene, as well as the viability, reproduction and transgenic approaches for the reproductive containment of GH-transgenic fish. Further, we propose that it should be necessary to do the following studies, in particularly, of the breeding of transgenic fish: to assess the fitness of transgenic fish in an aqueous environment with a large space and a complex structure; and to develop a controllable on-off strategy of reproduction in transgenic fish.

  17. Design and Management of a Transgenic Facility

    Institute of Scientific and Technical Information of China (English)

    Bob Springsteen

    2001-01-01

    @@ In 1965, I was given the opportunity to manage a research animal colony. At that time, the animal colony consisted of numerous species, such as primates, dogs, cats, rab bits, guinea pigs, hamsters, rats, mice, and some farm animals as well. Over the years,this menagerie was reduced to mice and rab bits. The animal facility now houses 8 000mice, of which 80% are transgenics. In approximately six years, transgenic mice have become the mainstay of the Berkeley Lab animal facility, and this population continues to grow.

  18. Evaluating the fitness of human lysozyme transgenic dairy goats: growth and reproductive traits

    OpenAIRE

    2010-01-01

    While there are many reports in the literature describing the attributes of specific applications of transgenic animals for agriculture, there are relatively few studies focusing on the fitness of the transgenic animals themselves. This work was designed to gather information on genetically modified food animals to determine if the presence of a transgene can impact general animal production traits. More specifically, we used a line of transgenic dairy goats expressing human lysozyme in their...

  19. The past, present and future of transgenic bioreactors.

    Science.gov (United States)

    Drohan, W N

    1997-07-01

    Hybrid genes can control the tissue-specific synthesis of human proteins in transgenic animals. Thus, it is now possible to produce proteins of biomedical value in the body fluids or cells of transgenic livestock. In fact, the first transgenically produced protein, antithrombin III, is now in clinical trials and others will soon follow.

  20. Tumorigenic potential of pituitary tumor transforming gene (PTTG in vivo investigated using a transgenic mouse model, and effects of cross breeding with p53 (+/− transgenic mice

    Directory of Open Access Journals (Sweden)

    Fong Miranda Y

    2012-11-01

    Full Text Available Abstract Background Pituitary tumor-transforming gene (PTTG is an oncogene that is overexpressed in variety of tumors and exhibits characteristics of a transforming gene. Previous transgenic mouse models to access the tumorigenic potential in the pituitary and ovary have resulted in dysplasia without formation of visible tumors, possibly due to the insufficient expression of PTTG. PTTG expression level is critical for ovarian tumorigenesis in a xenograft model. Therefore, the tumorigenic function of PTTG in vivo remains unclear. We generated a transgenic mouse that overexpresses PTTG driven by the CMV promoter to determine whether PTTG functions as a transforming oncogene that is capable of initiating tumorigenesis. Methods Transgenic animals were generated by microinjection of PTTG transgene into the male pronucleus of FVB 0.5 day old embryos. Expression levels of PTTG in tissues of transgenic animals were analyzed using an immunohistochemical analysis. H&E staining and immunohistostaining were performed to examine the type of tumor in transgenic and PTTG transgenic/p53+/- animals. Results PTTG transgenic offspring (TgPTTG were monitored for tumor development at various ages. H&E analysis was performed to identify the presence of cancer and hyperplastic conditions verified with the proliferation marker PCNA and the microvessel marker CD31. Immunohistochemistry was performed to determine transgene expression, revealing localization to the epithelium of the fallopian tube, with more generalized expression in the liver, lung, kidney, and spleen. At eight months of age, 2 out of 15 TgPTTG developed ovarian cancer, 2 out of 15 developed benign tumors, 2 out of 15 developed cervical dysplasia, and 3 out of 15 developed adenomyosis of the uterus. At ten months of age, 2 out of 10 TgPTTG developed adenocarcinoma of the ovary, 1 out of 10 developed a papillary serous adenocarcinoma, and 2 out of 10 presented with atypia of ovarian epithelial cells

  1. Comparative systems biology between human and animal models based on next-generation sequencing methods.

    Science.gov (United States)

    Zhao, Yu-Qi; Li, Gong-Hua; Huang, Jing-Fei

    2013-04-01

    Animal models provide myriad benefits to both experimental and clinical research. Unfortunately, in many situations, they fall short of expected results or provide contradictory results. In part, this can be the result of traditional molecular biological approaches that are relatively inefficient in elucidating underlying molecular mechanism. To improve the efficacy of animal models, a technological breakthrough is required. The growing availability and application of the high-throughput methods make systematic comparisons between human and animal models easier to perform. In the present study, we introduce the concept of the comparative systems biology, which we define as "comparisons of biological systems in different states or species used to achieve an integrated understanding of life forms with all their characteristic complexity of interactions at multiple levels". Furthermore, we discuss the applications of RNA-seq and ChIP-seq technologies to comparative systems biology between human and animal models and assess the potential applications for this approach in the future studies.

  2. Knockdown of myostatin expression by RNAi enhances muscle growth in transgenic sheep.

    Directory of Open Access Journals (Sweden)

    Shengwei Hu

    Full Text Available Myostatin (MSTN has been shown to be a negative regulator of skeletal muscle development and growth. MSTN dysfunction therefore offers a strategy for promoting animal growth performance in livestock production. In this study, we investigated the possibility of using RNAi-based technology to generate transgenic sheep with a double-muscle phenotype. A shRNA expression cassette targeting sheep MSTN was used to generate stable shRNA-expressing fibroblast clones. Transgenic sheep were further produced by somatic cell nuclear transfer (SCNT technology. Five lambs developed to term and three live lambs were obtained. Integration of shRNA expression cassette in three live lambs was confirmed by PCR. RNase protection assay showed that the shRNAs targeting MSTN were expressed in muscle tissues of three transgenic sheep. MSTN expression was significantly inhibited in muscle tissues of transgenic sheep when compared with control sheep. Moreover, transgenic sheep showed a tendency to faster increase in body weight than control sheep. Histological analysis showed that myofiber diameter of transgenic sheep M17 were bigger than that of control sheep. Our findings demonstrate a promising approach to promoting muscle growth in livestock production.

  3. The effectiveness of computer-generated 3D animations in inquiry chemistry laboratory

    Science.gov (United States)

    Theall, Rachel Morgan

    It has been shown that students need a molecular-level understanding of substances in order to comprehend chemistry. For solid structures, atomic-level understanding requires students to learn additional and different concepts than for other states of matter. To aid understanding, animations were created to model unit cell structures and depict the properties of unit cells. In order to determine if these animations are helpful to students, they were tested during a laboratory exercise in which students had previously been using model kits and images from textbooks to learn about solid structures. Students evaluated in this study were from two lecture sections of general chemistry, one that routinely used animations during lecture and one that used a more traditional lecture format that did not include animations or models. Twelve laboratory sections of these lectures, taught by six different instructors each teaching two sections, were chosen for participation. One section for each instructor was given the animations as an optional tool for completing the laboratory assignment, which consisted of questions about unit cells and crystal structures. The results of the study indicate that students who looked at the animations performed significantly better on the assignment. For the control group, students who routinely viewed multiple representations of chemistry in lecture performed significantly better on the lab assignment than students in the lecture section where chemistry concepts were only presented on the chalkboard and overhead projector. Students in the traditional lecture section also had significantly less appreciation for the model kits used in the laboratory than students in the other lecture section. Observations of students in the lab combined with statistical results led to the revision of the solid structures investigation. Additional animations were created and inserted into the module that covered areas where students indicated more help was needed

  4. Genetically modified animals and pharmacological research.

    Science.gov (United States)

    Wells, Dominic J

    2010-01-01

    This chapter reviews the use of genetically modified animals and the increasingly detailed knowledge of the genomes of the domestic species. The different approaches to genetic modification are outlined as are the advantages and disadvantages of the techniques in different species. Genetically modified mice have been fundamental in understanding gene function and in generating affordable models of human disease although these are not without their drawbacks. Transgenic farm animals have been developed for nutritionally enhanced food, disease resistance and xenografting. Transgenic rabbits, goats, sheep and cows have been developed as living bioreactors producing potentially high value biopharmaceuticals, commonly referred to as "pharming". Domestic animals are also important as a target as well as for testing genetic-based therapies for both inherited and acquired disease. This latter field may be the most important of all, in the future development of novel therapies.

  5. Overexpression of carnation S-adenosylmethionine decarboxylase gene generates a broad-spectrum tolerance to abiotic stresses in transgenic tobacco plants.

    Science.gov (United States)

    Wi, Soo Jin; Kim, Woo Taek; Park, Ky Young

    2006-10-01

    Polyamines (PAs), such as putrescine, spermidine, and spermine, are present in all living organism and implicate in a wide range of cellular physiological processes. We have used transgenic technology in an attempt to evaluate their potential for mitigating the adverse effects of several abiotic stresses in plants. Sense construct of full-length cDNA for S-adenosylmethionine decarboxylase (SAMDC), a key enzyme in PA biosynthesis, from carnation (Dianthus caryophyllus L.) flower was introduced into tobacco (Nicotiana tabacum L.) by Agrobacterium tumefaciens-mediated transformation. Several transgenic lines overexpressing SAMDC gene under the control of cauliflower mosaic virus 35S promoter accumulated soluble total PAs by 2.2 (S16-S-4) to 3.1 (S16-S-1) times than wild-type plants. The transgenic tobacco did not show any difference in organ phenotype compared to the wild-type. The number and weight of seeds increased, and net photosynthetic rate also increased in transgenic plants. Stress-induced damage was attenuated in these transgenic plants, in the symptom of visible yellowing and chlorophyll degradation after all experienced stresses such as salt stress, cold stress, acidic stress, and abscisic acid treatment. H2O2-induced damage was attenuated by spermidine treatment. Transcripts for antioxidant enzymes (ascorbate peroxidase, manganase superoxide dismutase, and glutathione S-transferase) in transgenic plants and GUS activity transformed with SAMDC promoter::GUS fusion were induced more significantly by stress treatment, compared to control. These results that the transgenic plants with sense SAMDC cDNA are more tolerant to abiotic stresses than wild-type plants suggest that PAs may play an important role in contributing stress tolerance in plants.

  6. The generation algorithm of arbitrary polygon animation based on dynamic correction

    Directory of Open Access Journals (Sweden)

    Hou Ya Wei

    2016-01-01

    Full Text Available This paper, based on the key-frame polygon sequence, proposes a method that makes use of dynamic correction to develop continuous animation. Firstly we use quadratic Bezier curve to interpolate the corresponding sides vector of polygon sequence consecutive frame and realize the continuity of animation sequences. And then, according to Bezier curve characteristic, we conduct dynamic regulation to interpolation parameters and implement the changing smoothness. Meanwhile, we take use of Lagrange Multiplier Method to correct the polygon and close it. Finally, we provide the concrete algorithm flow and present numerical experiment results. The experiment results show that the algorithm acquires excellent effect.

  7. Progress Toward a Human CD4/CCR5 Transgenic Rat Model for De Novo Infection by Human Immunodeficiency Virus Type 1

    Science.gov (United States)

    Keppler, Oliver T.; Welte, Frank J.; Ngo, Tuan A.; Chin, Peggy S.; Patton, Kathryn S.; Tsou, Chia-Lin; Abbey, Nancy W.; Sharkey, Mark E.; Grant, Robert M.; You, Yun; Scarborough, John D.; Ellmeier, Wilfried; Littman, Dan R.; Stevenson, Mario; Charo, Israel F.; Herndier, Brian G.; Speck, Roberto F.; Goldsmith, Mark A.

    2002-01-01

    The development of a permissive small animal model for the study of human immunodeficiency virus type (HIV)-1 pathogenesis and the testing of antiviral strategies has been hampered by the inability of HIV-1 to infect primary rodent cells productively. In this study, we explored transgenic rats expressing the HIV-1 receptor complex as a susceptible host. Rats transgenic for human CD4 (hCD4) and the human chemokine receptor CCR5 (hCCR5) were generated that express the transgenes in CD4+ T lymphocytes, macrophages, and microglia. In ex vivo cultures, CD4+ T lymphocytes, macrophages, and microglia from hCD4/hCCR5 transgenic rats were highly susceptible to infection by HIV-1 R5 viruses leading to expression of abundant levels of early HIV-1 gene products comparable to those found in human reference cultures. Primary rat macrophages and microglia, but not lymphocytes, from double-transgenic rats could be productively infected by various recombinant and primary R5 strains of HIV-1. Moreover, after systemic challenge with HIV-1, lymphatic organs from hCD4/hCCR5 transgenic rats contained episomal 2–long terminal repeat (LTR) circles, integrated provirus, and early viral gene products, demonstrating susceptibility to HIV-1 in vivo. Transgenic rats also displayed a low-level plasma viremia early in infection. Thus, transgenic rats expressing the appropriate human receptor complex are promising candidates for a small animal model of HIV-1 infection. PMID:11901198

  8. Handmade cloned transgenic sheep rich in omega-3 Fatty acids.

    Directory of Open Access Journals (Sweden)

    Peng Zhang

    Full Text Available Technology of somatic cell nuclear transfer (SCNT has been adapted worldwide to generate transgenic animals, although the traditional procedure relies largely on instrumental micromanipulation. In this study, we used the modified handmade cloning (HMC established in cattle and pig to produce transgenic sheep with elevated levels of omega-3 (n-3 fatty acids. Codon-optimized nematode mfat-1 was inserted into a eukaryotic expression vector and was transferred into the genome of primary ovine fibroblast cells from a male Chinese merino sheep. Reverse transcriptase PCR, gas chromatography, and chromosome analyses were performed to select nuclear donor cells capable of converting omega-6 (n-6 into n-3 fatty acids. Blastocysts developed after 7 days of in vitro culture were surgically transplanted into the uterus of female ovine recipients of a local sheep breed in Xinjiang. For the HMC, approximately 8.9% (n  =925 of reconstructed embryos developed to the blastocyst stage. Four recipients became pregnant after 53 blastocysts were transplanted into 29 naturally cycling females, and a total of 3 live transgenic lambs were produced. Detailed analyses on one of the transgenic lambs revealed a single integration of the modified nematode mfat-1 gene at sheep chromosome 5. The transgenic sheep expressed functional n-3 fatty acid desaturase, accompanied by more than 2-folds reduction of n-6/n-3 ratio in the muscle (p<0.01 and other major organs/tissues (p<0.05. To our knowledge, this is the first report of transgenic sheep produced by the HMC. Compared to the traditional SCNT method, HMC showed an equivalent efficiency but proved cheaper and easier in operation.

  9. Transgenic Plants as Sensors of Environmental Pollution Genotoxicity

    Science.gov (United States)

    Kovalchuk, Igor; Kovalchuk, Olga

    2008-01-01

    Rapid technological development is inevitably associated with many environmental problems which primarily include pollution of soil, water and air. In many cases, the presence of contamination is difficult to assess. It is even more difficult to evaluate its potential danger to the environment and humans. Despite the existence of several whole organism-based and cell-based models of sensing pollution and evaluation of toxicity and mutagenicity, there is no ideal system that allows one to make a quick and cheap assessment. In this respect, transgenic organisms that can be intentionally altered to be more sensitive to particular pollutants are especially promising. Transgenic plants represent an ideal system, since they can be grown at the site of pollution or potentially dangerous sites. Plants are ethically more acceptable and esthetically more appealing than animals as sensors of environmental pollution. In this review, we will discuss various transgenic plant-based models that have been successfully used for biomonitoring genotoxic pollutants. We will also discuss the benefits and potential drawbacks of these systems and describe some novel ideas for the future generation of efficient transgenic phytosensors.

  10. The influence of matrix attachment regions on transgene expression in Arabidopsis thaliana wild type and gene silencing mutants.

    Science.gov (United States)

    De Bolle, Miguel F C; Butaye, Katleen M J; Goderis, Inge J W M; Wouters, Piet F J; Jacobs, Anni; Delauré, Stijn L; Depicker, Ann; Cammue, Bruno P A

    2007-03-01

    Many studies in both animal and plant systems have shown that matrix attachment regions (MARs) can increase the expression of flanking transgenes. However, our previous studies revealed no effect of the chicken lysozyme MARs (chiMARs) on transgene expression in the first generation transgenic Arabidopsis thaliana plants transformed with a beta-glucuronidase gene (uidA) unless gene silencing mutants were used as genetic background for transformation. In the present study, we investigated why chiMARs do not influence transgene expression in transgenic wild-type Arabidopsis plants. We first studied the effect of chiMARs on transgene expression in the progeny of primary transformants harboring chiMAR-flanked T-DNAs. Our data indicate that chiMARs do not affect transgene expression in consecutive generations of wild-type A. thaliana plants. Next, we examined whether these observed results in A. thaliana transformants are influenced by the applied transformation method. The results from in vitro transformed A. thaliana plants are in accordance with those from in planta transformed A. thaliana plants and again reveal no influence of chiMARs on transgene expression in A. thaliana wild-type transformants. The effect of chi-MARs on transgene expression is also examined in in vitro transformed Nicotiana tabacum plants, but as for A. thaliana, the transgene expression in tobacco transformants is not altered by the presence of chi-MARs. Taken together, our results show that the applied method or the plant species used for transformation does not influence whether and how chiMARs have an effect on transgene expression. Finally, we studied the effect of MARs (tabMARs) of plant origin (tobacco) on the transgene expression in A. thaliana wild-type plants and suppressed gene silencing (sgs2) mutants. Our results clearly show that similar to chiMARs, the tobacco-derived MARs do not enhance transgene expression in a wild-type background but can be used to enhance transgene expression

  11. TRANSGENIC PLANTS RESISTANT TO INSECTS

    Directory of Open Access Journals (Sweden)

    S. Kereša

    2009-09-01

    Full Text Available Proteinase inhibitors are secondary metabolites present in all plants and it seems that their major role is protection of plants against attacks of animals, insects and microorganisms. One of the family of proteinase inhibitors are squash inhibitors of serine proteinases purified from seeds belonging to genera Cucurbita, Cucumis and Momordica. Squash inhibitors consist of 29-32 amino acid residues and are considered to be the smallest inhibitors of the serine proteinases known. Because of shortness, genes for these inhibitors could be synthesised and modified at different ways. Modifications could lead to changes in inhibitor activity. Tobacco as a model plant was transformed with 12 different genes of squash inhibitors. Stable integration of transgenes in putative transgenic plants was determined by PCR analysis using genomic DNA and primers that anneal to promoter and terminator region. The first step of proteinase inhibitor gene expression in transgenic plants was revealed by RT-PCR analysis. In entomological tests where larvae were fed with leaves, influence of transgenic T0 plants, as well as non-transgenic control plants on retardation of larval growth of S. littoralis was examined. Results of entomological tests showed that it is possible to express squash proteinase inhibitors in plants at level that significantly reduces S. littoralis larval growth.

  12. Evaluation Of Electricity Generation From Animal Based Wastes In A Microbial Fuel Cell

    Directory of Open Access Journals (Sweden)

    Duduyemi Oladejo

    2015-04-01

    Full Text Available ABSTRACT Electric current from organic waste of poultry droppings were generated with A Microbial Fuel Cell MFC technology to evaluate affects of temperature 30 to 50oC 100gl 300gl and 500gl slurry concentrations prepared with the distilled water and inoculated when introduced into the anodic chamber. A constant concentration of 50gl of the oxidizing agent Potassium ferricyanide at the cathode chamber was prepared to evaluate the voltage and current generated by the set up for 7 days in each case. Higher slurry concentrations were observed to generate higher initial current and voltage than in lower concentrations. Higher slurry concentrations also demonstrated sustained power generation up to the day 6 before decline. A maximum current of 1.1V and 0.15 mA was achieved while the temperature variation was observed to have minimal effect within the range considered at low concentration. A MFC is a biochemical-catalyzed system capable of generating electricity as a by-product also providing an alternative method of waste treatment. Application Alternative power source and waste treatment.

  13. Locus-specific integration of extrachromosomal transgenes in C. elegans with the CRISPR/Cas9 system

    Directory of Open Access Journals (Sweden)

    Sawako Yoshina

    2016-03-01

    Full Text Available We established a method to generate integration from extrachromosomal arrays with the CRISPR/Cas9 system. Multi-copy transgenes were integrated into the defined loci of chromosomes by this method, while a multi-copy transgene is integrated into random loci by previous methods, such as UV- and gamma-irradiation. The effects of a combination of sgRNAs, which define the cleavage sites in extrachromosomes and chromosomes, and the copy number of potential cleavable sequences were examined. The relative copy number of cleavable sequences in extrachromosomes affects the frequency of fertile F1 transgenic animals. The expression levels of the reporter gene were almost proportional to the copy numbers of the integrated sequences at the same integration site. The technique is applicable to the transgenic strains abundantly stored and shared among the C. elegans community, particularly when researchers use sgRNAs against common plasmid sequences such as β-lactamase.

  14. Expression of human apolipoprotein B and assembly of lipoprotein(a) in transgenic mice

    Energy Technology Data Exchange (ETDEWEB)

    Callow, M.J.; Stoltzfus, L.J.; Rubin, E.M. [Lawrence Berkeley Lab., CA (United States); Lawn, R.M. [Stanford Univ., CA (United States)

    1994-03-15

    The atherogenic macromolecule lipoprotein(a) [Lp(a)] has resisted in vivo analyses partly because it is found in a limited number of experimental animals. Although transgenic mice expressing human apolipoprotein (a) [apo(a)] have previously been described, they failed to assemble Lp(a) particles because of the inability of human apo(a) to associate with mouse apolipoprotein B (apoB). The authors isolated a 90-kilobase P1 phagemid containing the human apoB gene and with this DNA generated 13 lines of transgenic mice of which 11 expressed human apoB. The human apoB transcript was expressed and edited in the liver of the transgenic mice. Plasma concentrations of human apoB, as well as low density lipoprotein (LDL), were related to transgene copy number; the transgenic line with the most copies of human apoB had a >4-fold increase in LDL cholesterol compared with nontransgenics and a lipoprotein profile similar to that of humans. When human apoB and apo(a) transgenic mice were bred together, plasma apo(a) in mice expressing both human proteins was tightly associated with lipoproteins in the LDL density region. These studies demonstrate the successful expression of human apoB and the efficient assembly of Lp(a) in mice.

  15. Expression of GFP in nuclear transplants generated by transplantation of embryonic cell nuclei from GFP-transgenic fish into nonenucleated eggs of medaka, Oryzias latipes.

    Science.gov (United States)

    Niwa, K; Kani, S; Kinoshita, M; Ozato, K; Wakamatsu, Y

    2000-01-01

    In order to investigate whether foreign genes can be used as genetic markers of donor nuclei in fish nuclear transplantation, expression of the GFP gene derived from donor nuclei was examined in nuclear transplants in medaka (Oryzias latipes). Embryonic nuclei were obtained from blastula embryos produced by crossing of transgenic fish of the wild-type strain heterozygous for the GFP gene with nontransgenic ones or by mutual crossing between transgenic fish. The GFP gene was driven by the promoter of the medaka elongation factor gene, EF-1alpha-A, which is known to induce GFP expression in many tissues except for the muscle in the transgenic fish. The nuclei were transplanted into nonenucleated unfertilized eggs of the orange-red strain. Adult nuclear transplants were successfully obtained at the rate of about 2% of the operated eggs. They were triploid and had no reproductive potential. The GFP gene was expressed in embryos, fry, and adults of nuclear transplants in a pattern similar to that in the transgenic fish. These results indicate that GFP is useful as a foreign genetic marker of donor nuclei in fish nuclear transplantation.

  16. Interspecies chimeric complementation for the generation of functional human tissues and organs in large animal hosts.

    Science.gov (United States)

    Wu, Jun; Izpisua Belmonte, Juan Carlos

    2016-06-01

    The past decade's rapid progress in human pluripotent stem cell (hPSC) research has generated hope for meeting the rising demand of organ donation, which remains the only effective cure for end-stage organ failure, a major cause of death worldwide. Despite the potential, generation of transplantable organs from hPSCs using in vitro differentiation is far-fetched. An in vivo interspecies chimeric complementation strategy relying on chimeric-competent hPSCs and zygote genome editing provides an auspicious alternative for providing unlimited organ source for transplantation.

  17. An efficient genotyping method for genome-modified animals and human cells generated with CRISPR/Cas9 system.

    Science.gov (United States)

    Zhu, Xiaoxiao; Xu, Yajie; Yu, Shanshan; Lu, Lu; Ding, Mingqin; Cheng, Jing; Song, Guoxu; Gao, Xing; Yao, Liangming; Fan, Dongdong; Meng, Shu; Zhang, Xuewen; Hu, Shengdi; Tian, Yong

    2014-09-19

    The rapid generation of various species and strains of laboratory animals using CRISPR/Cas9 technology has dramatically accelerated the interrogation of gene function in vivo. So far, the dominant approach for genotyping of genome-modified animals has been the T7E1 endonuclease cleavage assay. Here, we present a polyacrylamide gel electrophoresis-based (PAGE) method to genotype mice harboring different types of indel mutations. We developed 6 strains of genome-modified mice using CRISPR/Cas9 system, and utilized this approach to genotype mice from F0 to F2 generation, which included single and multiplexed genome-modified mice. We also determined the maximal detection sensitivity for detecting mosaic DNA using PAGE-based assay as 0.5%. We further applied PAGE-based genotyping approach to detect CRISPR/Cas9-mediated on- and off-target effect in human 293T and induced pluripotent stem cells (iPSCs). Thus, PAGE-based genotyping approach meets the rapidly increasing demand for genotyping of the fast-growing number of genome-modified animals and human cell lines created using CRISPR/Cas9 system or other nuclease systems such as TALEN or ZFN.

  18. The next generation of disease risk: are the effects of prenatal nutrition transmitted across generations? Evidence from animal and human studies.

    Science.gov (United States)

    Roseboom, T J; Watson, E D

    2012-11-01

    Suboptimal intrauterine conditions, including poor nutrition, during critical periods of growth may lead to lifelong changes in the body's organs and tissues, thus providing a physiological basis for adult-onset disease. Remarkably, recent evidence suggests that the long-term consequences of adverse conditions during early development may not be limited to one generation, but may lead to poor health in the generations to follow, even if these individuals develop in normal conditions themselves. For example, the diet of a pregnant mother may affect the development and disease risk of her children and even her grandchildren. There is limited evidence for this in humans since studies of multiple generations are difficult to maintain. However, recent animal models have been generated to investigate this phenomenon and will be instrumental in the future for assessing the underlying mechanisms of intergenerational and transgenerational transmission of disease. These mechanisms remain unclear, though environmental, metabolic and epigenetic factors are likely involved. Researchers have begun to address how changes in metabolism and epigenetic regulation of gene expression caused by poor nutrition can be passed from one generation to the next. Ultimately, these findings will shed light on the transmission of diabetes, obesity and cardiovascular disease that are rapidly expanding in Western countries. Public health strategies that focus on improved maternal nutrition may provide a means of promoting cardiovascular and metabolic health. However, the full impact of these strategies may not be apparent for decades.

  19. 204 POTENTIAL OF GREEN FLUORESCENT PROTEIN LOCUS FOR GENE EDITING IN DNA TRANSPOSON-PRODUCED TRANSGENIC CATTLE.

    Science.gov (United States)

    Yum, S-Y; Lee, S-J; Kim, H-M; Lee, C-I; Kim, H-S; Kim, H-J; Choi, W-J; Hahn, S-E; Lee, J-H; Kim, S-J; Jang, G

    2016-01-01

    Recently, we published on the efficient production of transgenic cattle using the DNA transposon system (Yum et al. 2016 Sci. Rep. 6, 27185). In that study, 8 transgenic cattle were born following transposon-mediated gene delivery system (Sleeping Beauty and Piggybac transposon) via microinjection of zygotes. In the analysis of their genomic stability using next-generation sequencing, there was no significant difference in the number of genomic variants between transgenic and nontransgenic cattle. In this study, we have described current status of those transgenic cattle in term of health, germ-line transmission, and application. All the transgenic cattle have grown up to date (the oldest being 30 months old, the youngest being 12 months old) without any health issue. In general blood analysis, there were not any significant changes between transgenic cattle and wild type. Because the transgene (green fluorescent protein; GFP) expression is constitutively active and has strong expression, it could be visualised without fluorescence equipment. One of transgenic male cattle reached puberty and semen was collected. Over 200 frozen semen straws were produced and some were used for IVF. In every IVF replication, around 80% blastocysts expressed the GFP. Over 36 GFP blastocysts were frozen for embryo transfer in the future, and we are planning to crossbreed for generating homozygotic transgenic cattle. Another application is to use the GFP locus to gene-edit the transgenic cattle, as long-term expression of transgene did not affect their health. In 1 cell stage, embryos produced using GFP frozen-thawed semen, single guide RNA for GFP, Cas9, together with donor DNA that included RFP and homology arms to link the double-strand break of single guide RNA target site, were co-injected and RFP was observed. Knockout/-in for editing GFP locus using CRISPR-Cas9 might be a valuable approach for the next generation of transgenic models by microinjection. In conclusion, we

  20. Generation and characterization of RAG2 knockout pigs as animal model for severe combined immunodeficiency.

    Science.gov (United States)

    Suzuki, Shunichi; Iwamoto, Masaki; Hashimoto, Michiko; Suzuki, Misae; Nakai, Michiko; Fuchimoto, Daiichiro; Sembon, Shoichiro; Eguchi-Ogawa, Tomoko; Uenishi, Hirohide; Onishi, Akira

    2016-10-01

    Pigs with severe combined immunodeficiency (SCID) are versatile animal models for human medical research because of their biological similarities to humans, suitable body size, and longevity for practical research. SCID pigs with defined mutation(s) can be an invaluable tool for research on porcine immunity. In this study, we produced RAG2-knockout pigs via somatic cell nuclear transfer and analyzed their phenotype. The V(D)J recombination processes were confirmed as being inactivated. They consistently lacked mature T and B cells but had substantial numbers of cells considered to be T- or B-cell progenitors as well as NK cells. They also lacked thymic medulla and lymphoid aggregations in the spleen, mesenteric lymph nodes, and ileal Peyer's patches. We showed more severe immunological defects in the RAG2 and IL2RG double-knockout pig through this study. Thus, SCID pigs could be promising animal models not only for translational medical research but also for immunological studies of pigs themselves.

  1. Transposon-mediated transgenesis, transgenic rescue, and tissue-specific gene expression in rodents and rabbits.

    Science.gov (United States)

    Katter, Katharina; Geurts, Aron M; Hoffmann, Orsolya; Mátés, Lajos; Landa, Vladimir; Hiripi, László; Moreno, Carol; Lazar, Jozef; Bashir, Sanum; Zidek, Vaclav; Popova, Elena; Jerchow, Boris; Becker, Katja; Devaraj, Anantharam; Walter, Ingrid; Grzybowksi, Michael; Corbett, Molly; Filho, Artur Rangel; Hodges, Matthew R; Bader, Michael; Ivics, Zoltán; Jacob, Howard J; Pravenec, Michal; Bosze, Zsuzsanna; Rülicke, Thomas; Izsvák, Zsuzsanna

    2013-03-01

    Germline transgenesis is an important procedure for functional investigation of biological pathways, as well as for animal biotechnology. We have established a simple, nonviral protocol in three important biomedical model organisms frequently used in physiological studies. The protocol is based on the hyperactive Sleeping Beauty transposon system, SB100X, which reproducibly promoted generation of transgenic founders at frequencies of 50-64, 14-72, and 15% in mice, rats, and rabbits, respectively. The SB100X-mediated transgene integrations are less prone to genetic mosaicism and gene silencing as compared to either the classical pronuclear injection or to lentivirus-mediated transgenesis. The method was successfully applied to a variety of transgenes and animal models, and can be used to generate founders with single-copy integrations. The transposon vector also allows the generation of transgenic lines with tissue-specific expression patterns specified by promoter elements of choice, exemplified by a rat reporter strain useful for tracking serotonergic neurons. As a proof of principle, we rescued an inborn genetic defect in the fawn-hooded hypertensive rat by SB100X transgenesis. A side-by-side comparison of the SB100X- and piggyBac-based protocols revealed that the two systems are complementary, offering new opportunities in genome manipulation.

  2. ADVANCES IN TRANSGENIC MAIZE FOR QUALITY IMPROVEMENT

    Directory of Open Access Journals (Sweden)

    M.Rajendar Reddy

    2015-12-01

    Full Text Available Maize (Zea mays is a major food and animal feed worldwide and occupies a relevant place in the world economy and trade as an industrial grain crop. Currently more than 70% of maize production is used for food and feed; therefore, knowledge of genes involved in grain structure and chemical is important for improving the nutritional and food-making properties of maize. It is a good source of carbohydrates, fats, proteins, vitamins and minerals but deficient in two essential amino acids, Viz., lysine and tryptophan. To overcome this problem and to improve the above quality characters the maize breeders have followed different strategies like opaque 2, QPM and development of transgenic maize with improved quality characters. Finally we can conclude that the conventional breeding techniques and now plant biotechnology are helping meet the growing demand for food production, nutrition security while preserving our environment for future generations

  3. Visualization of C. elegans transgenic arrays by GFP

    Directory of Open Access Journals (Sweden)

    Sternberg Paul W

    2006-06-01

    Full Text Available Abstract Background Targeting the green fluorescent protein (GFP via the E. coli lac repressor (LacI to a specific DNA sequence, the lac operator (lacO, allows visualization of chromosomes in yeast and mammalian cells. In principle this method of visualization could be used for genetic mosaic analysis, which requires cell-autonomous markers that can be scored easily and at single cell resolution. The C. elegans lin-3 gene encodes an epidermal growth factor family (EGF growth factor. lin-3 is expressed in the gonadal anchor cell and acts through LET-23 (transmembrane protein tyrosine kinase and ortholog of EGF receptor to signal the vulval precursor cells to generate vulval tissue. lin-3 is expressed in the vulval cells later, and recent evidence raises the possibility that lin-3 acts in the vulval cells as a relay signal during vulval induction. It is thus of interest to test the site of action of lin-3 by mosaic analysis. Results We visualized transgenes in living C. elegans by targeting the green fluorescent protein (GFP via the E. coli lac repressor (LacI to a specific 256 sequence repeat of the lac operator (lacO incorporated into transgenes. We engineered animals to express a nuclear-localized GFP-LacI fusion protein. C. elegans cells having a lacO transgene result in nuclear-localized bright spots (i.e., GFP-LacI bound to lacO. Cells with diffuse nuclear fluorescence correspond to unbound nuclear localized GFP-LacI. We detected chromosomes in living animals by chromosomally integrating the array of the lacO repeat sequence and visualizing the integrated transgene with GFP-LacI. This detection system can be applied to determine polyploidy as well as investigating chromosome segregation. To assess the GFP-LacI•lacO system as a marker for mosaic analysis, we conducted genetic mosaic analysis of the epidermal growth factor lin-3, expressed in the anchor cell. We establish that lin-3 acts in the anchor cell to induce vulva development

  4. Generation of cloned and chimeric embryos/offspring using the new methods of animal biotechnology.

    Science.gov (United States)

    Skrzyszowska, Maria; Karasiewicz, Jolanta; Bednarczyk, Marek; Samiec, Marcin; Smorag, Zdzisław; Waś, Bogusław; Guszkiewicz, Andrzej; Korwin-Kossakowski, Maciej; Górniewska, Maria; Szablisty, Ewa; Modliński, Jacek A; Łakota, Paweł; Wawrzyńska, Magdalena; Sechman, Andrzej; Wojtysiak, Dorota; Hrabia, Anna; Mika, Maria; Lisowski, Mirosław; Czekalski, Przemysław; Rzasa, Janusz; Kapkowska, Ewa

    2006-01-01

    The article summarizes results of studies concerning: 1/ qualitative evaluation of pig nuclear donor cells to somatic cell cloning, 2/ developmental potency of sheep somatic cells to create chimera, 3/ efficient production of chicken chimera. The quality of nuclear donor cells is one of the most important factors to determine the efficiency of somatic cell cloning. Morphological criteria commonly used for qualitative evaluation of somatic cells may be insufficient for practical application in the cloning. Therefore, different types of somatic cells being the source of genomic DNA in the cloning procedure were analyzed on apoptosis with the use of live-DNA or plasma membrane fluorescent markers. It has been found that morphological criteria are a sufficient selection factor for qualitative evaluation of nuclear donor cells to somatic cell cloning. Developmental potencies of sheep somatic cells in embryos and chimeric animals were studied using blastocyst complementation test. Fetal fibroblasts stained with vital fluorescent dye and microsurgically placed in morulae or blastocysts were later identified in embryos cultured in vitro. Transfer of Polish merino blastocysts harbouring Heatherhead fibroblasts to recipient ewes brought about normal births at term. Newly-born animals were of merino appearance with dark patches on their noses, near the mouth and on their clovens. This overt chimerism shows that fetal fibroblasts introduced to sheep morulae/blastocysts revealed full developmental plasticity. To achieve the efficient production of chicken chimeras, the blastodermal cells from embryos of the donor breeds, (Green-legged Partridgelike breed or GPxAraucana) were transferred into the embryos of the recipient breed (White Leghorn), and the effect of chimerism on the selected reproductive and physiological traits of recipients was examined. Using the model which allowed identification of the chimerism at many loci, it has been found that 93.9% of the examined birds

  5. Virus factories, double membrane vesicles and viroplasm generated in animal cells.

    Science.gov (United States)

    Netherton, Christopher L; Wileman, Tom

    2011-11-01

    Many viruses reorganise cellular membrane compartments and the cytoskeleton to generate subcellular microenvironments called virus factories or 'viroplasm'. These create a platform to concentrate replicase proteins, virus genomes and host proteins required for replication and also protect against antiviral defences. There is growing interest in understanding how viruses induce such large changes in cellular organisation, and recent studies are beginning to reveal the relationship between virus factories and viroplasm and the cellular structures that house them. In this review, we discuss how three supergroups of (+)RNA viruses generate replication sites from membrane-bound organelles and highlight research on perinuclear factories induced by the nucleocytoplasmic large DNA viruses. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Advancing environmental risk assessment for transgenic biofeedstock crops

    OpenAIRE

    2009-01-01

    Abstract Transgenic modification of plants is a key enabling technology for developing sustainable biofeedstocks for biofuels production. Regulatory decisions and the wider acceptance and development of transgenic biofeedstock crops are considered from the context of science-based risk assessment. The risk assessment paradigm for transgenic biofeedstock crops is fundamentally no different from that of current generation transgenic crops, except that the focus of the assessment must consider t...

  7. Evaluating the fitness of human lysozyme transgenic dairy goats: growth and reproductive traits.

    Science.gov (United States)

    Jackson, Kathryn A; Berg, Jolene M; Murray, James D; Maga, Elizabeth A

    2010-12-01

    While there are many reports in the literature describing the attributes of specific applications of transgenic animals for agriculture, there are relatively few studies focusing on the fitness of the transgenic animals themselves. This work was designed to gather information on genetically modified food animals to determine if the presence of a transgene can impact general animal production traits. More specifically, we used a line of transgenic dairy goats expressing human lysozyme in their mammary gland to evaluate the reproductive fitness and growth and development of these animals compared to their non-transgenic counterparts and the impact of consuming a transgenic food product, lysozyme-containing milk. In males, none of the parameters of semen quality, including semen volume and concentration, total sperm per ejaculate, sperm morphology, viability and motility, were significantly different between transgenic bucks and non-transgenic full-sib controls. Likewise, transgenic females of this line did not significantly differ in the reproductive traits of gestation length and litter size compared to their non-transgenic counterparts. To evaluate growth, transgenic and non-transgenic kid goats received colostrum and milk from either transgenic or non-transgenic does from birth until weaning. Neither the presence of the transgene nor the consumption of milk from transgenic animals significantly affected birth weight, weaning weight, overall gain and post-wean gain. These results indicate that the analyzed reproductive and growth traits were not regularly or substantially impacted by the presence or expression of the transgene. The evaluation of these general parameters is an important aspect of defining the safety of applying transgenic technology to animal agriculture.

  8. Transgenic mice for MTCP1 develop T-cell prolymphocytic leukemia.

    Science.gov (United States)

    Gritti, C; Dastot, H; Soulier, J; Janin, A; Daniel, M T; Madani, A; Grimber, G; Briand, P; Sigaux, F; Stern, M H

    1998-07-15

    T-cell prolymphocytic leukemia (T-PLL) is a rare form of mature T-cell leukemia associated with chromosomal rearrangements implicating MTCP1 or TCL1 genes. These genes encode two homologous proteins, p13(MTCP1) and p14(TCL1), which share no similarity with other known protein. To determine the oncogenic role of MTCP1, mice transgenic for MTCP1 under the control of CD2 regulatory regions (CD2-p13 mice) were generated. No abnormality was detected during the first year after birth. A late effect of the transgene was searched for in a cohort of 48 CD2-p13 mice aged 15 to 20 months, issued from 3 independent founders. Lymphoid hemopathies, occurring in the three transgenic lines, were characterized by lymphoid cells with an irregular nucleus, a unique and prominent nucleolus, condensed chromatin, a basophilic cytoplasm devoid of granules, and an immunophenotype of mature T cells. The molecular characterization of Tcrb rearrangements demonstrated the monoclonal origin of these populations. Histopathological analysis of the cohort demonstrated early splenic and hepatic infiltrations, whereas lymphocytosis and medullar infiltrations were found infrequently. The engraftment of these proliferations in H2-matched animals demonstrated their malignant nature. Cumulative incidence of the disease at 20 months was 100%, 50%, and 21% in F3, F4, and F7 lines, respectively, and null in the control group. The level of expression of the transgene, as estimated by Western blotting in the transgenic lines correlated with the tumoral incidence, with the highest expression of p13(MTCP1) being found in F3 mice. CD2-p13 transgenic mice developed an hemopathy similar to human T-PLL. These data demonstrate that p13(MTCP1) is an oncoprotein and that CD2-p13 transgenic mice represent the first animal model for mature T-PLL.

  9. Subcutaneous implants for long-acting drug therapy in laboratory animals may generate unintended drug reservoirs

    Directory of Open Access Journals (Sweden)

    Michael Guarnieri

    2014-01-01

    Full Text Available Background: Long-acting therapy in laboratory animals offers advantages over the current practice of 2-3 daily drug injections. Yet little is known about the disintegration of biodegradable drug implants in rodents. Objective: Compare bioavailability of buprenorphine with the biodegradation of lipid-encapsulated subcutaneous drug pellets. Methods: Pharmacokinetic and histopathology studies were conducted in BALB/c female mice implanted with cholesterol-buprenorphine drug pellets. Results: Drug levels are below the level of detection (0.5 ng/mL plasma within 4-5 days of implant. However, necroscopy revealed that interstitial tissues begin to seal implants within a week. Visual inspection of the implant site revealed no evidence of inflammation or edema associated with the cholesterol-drug residue. Chemical analyses demonstrated that the residues contained 10-13% of the initial opiate dose for at least two weeks post implant. Discussion: The results demonstrate that biodegradable scaffolds can become sequestered in the subcutaneous space. Conclusion: Drug implants can retain significant and unintended reservoirs of drugs.

  10. Transgenic expression of an expanded (GCG)13 repeat PABPN1 leads to weakness and coordination defects in mice.

    Science.gov (United States)

    Dion, Patrick; Shanmugam, Vijayalakshmi; Gaspar, Claudia; Messaed, Christiane; Meijer, Inge; Toulouse, André; Laganiere, Janet; Roussel, Julie; Rochefort, Daniel; Laganiere, Simon; Allen, Carol; Karpati, George; Bouchard, Jean-Pierre; Brais, Bernard; Rouleau, Guy A

    2005-04-01

    Oculopharyngeal muscular dystrophy (OPMD) is a late-onset disorder caused by a (GCG)n trinucleotide repeat expansion in the poly(A) binding protein nuclear-1 (PABPN1) gene, which in turn leads to an expanded polyalanine tract in the protein. We generated transgenic mice expressing either the wild type or the expanded form of human PABPN1, and transgenic animals with the expanded form showed clear signs of abnormal limb clasping, muscle weakness, coordination deficits, and peripheral nerves alterations. Analysis of mitotic and postmitotic tissues in those transgenic animals revealed ubiquitinated PABPN1-positive intranuclear inclusions (INIs) in neuronal cells. This latter observation led us to test and confirm the presence of similar INIs in postmortem brain sections from an OPMD patient. Our results indicate that expanded PABPN1, presumably via the toxic effects of its polyalanine tract, can lead to inclusion formation and neurodegeneration in both the mouse and the human.

  11. Contrasting on Hydrocarbon Generation Model and Characteristics of Pyrolysis of Modern Aquatic Plant (Gloeocapsa) vs Marine Animal (Mantis Shrimp)

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    A hydrocarbon model of the modern aquatic plant Gloeocapsa and the aquatic animal mantis shrimp was found in thermal simulating experiment. The results show that the modern aquatic plant Gloeocapsa is characterized by late generation, late termination and a long duration of oil generation, while the aquatic animal mantis shrimp is characterized by early generation, early termination and a short duration of oil generation. The n-alkanes from Gloeocapsa and mantis shrimp are characterized by peak carbon C15-C17. With increasing thermal simulating temperature, the peak carbon changes from C17 to C15, and the odd-even predominance of n-allanes becomes less clear. The products from Gloeocapsa contain abundant phenanthrene and naphthalene compounds,and even a little retene, while those from mantis shrimp are dominated by naphthalene compounds, and are poor in phenanthrene compounds. Gloeocapsa and mantis shrimp are rich in C27sterane, relatively rich in tricyclic terpones (C1,-C2,) and hopanes (C27-C35), poor in 5α,14β17β sterane, and coprostane does not disappear until 450 ℃. The tricyclic terpanes in Gloeocapsaare characterized by peak carbon C23 and C2>C21, and in mantis shrimp by C21 and C21>C23. The content of pregnanes is very low at low temperature of thermal simulation, while above 360 ℃,high levels of pregnane series compounds and even degraded C2s and C26 steranes occur in thermal simulation products.

  12. Transgenic chickens as bioreactors for protein-based drugs.

    Science.gov (United States)

    Lillico, Simon G; McGrew, Michael J; Sherman, Adrian; Sang, Helen M

    2005-02-01

    The potential of using transgenic animals for the synthesis of therapeutic proteins was suggested over twenty years ago. Considerable progress has been made in developing methods for the production of transgenic animals and specifically in the expression of therapeutic proteins in the mammary glands of cows, sheep and goats. Development of transgenic hens for protein production in eggs has lagged behind these systems. The positive features associated with the use of the chicken in terms of cost, speed of development of a production flock and potentially appropriate glycosylation of target proteins have led to significant advances in transgenic chicken models in the past few years.

  13. TRANSGENIC FISH MODEL IN ENVIRONMENTAL TOXICOLOGY

    Directory of Open Access Journals (Sweden)

    Madhuri Sharma

    2012-05-01

    Full Text Available A number of experiments and the use of drugs have been performed in fish. The fish may be used as model organism in various biological experiments, including environmental toxicology. Aquatic animals are being engineered to increase aquaculture production, for medical and industrial research, and for ornamental reasons. Fish have been found to play an important role in assessing potential risks associated with exposure to toxic substances in aquatic environment. Hence, it has been thought that the development of transgenic fish can enhance the use of fish in environmental toxicology. India has developed experimental transgenics of rohu fish, zebra fish, cat fish and singhi fish. Genes, promoters and vectors of indigenous origin are now available for only two species namely rohu and singhi for engineering growth. Development of fish model carrying identical transgenes to those found in rodents is beneficial and has shown that several aspects of in vivo mutagenesis are similar between the two classes of vertebrates. Fish shows the frequencies of spontaneous mutations similar to rodents and respond to mutagen exposure consistent with known mutagenic mechanisms. The feasibility of in vivo mutation analysis using transgenic fish has been demonstrated and the potential value of transgenic fish as a comparative animal model has been illustrated. Therefore, the transgenic fish can give the significant contribution to study the environmental toxicity in animals as a whole.

  14. Animal research

    DEFF Research Database (Denmark)

    Olsson, I.A.S.; Sandøe, Peter

    2012-01-01

    in research is analyzed from the viewpoint of three distinct ethical approaches: contractarianism, utilitarianism, and animal rights view. On a contractarian view, research on animals is only an ethical issue to the extent that other humans as parties to the social contract care about how research animals......This article presents the ethical issues in animal research using a combined approach of ethical theory and analysis of scientific findings with bearing on the ethical analysis. The article opens with a general discussion of the moral acceptability of animal use in research. The use of animals...... are faring. From the utilitarian perspective, the use of sentient animals in research that may harm them is an ethical issue, but harm done to animals can be balanced by benefit generated for humans and other animals. The animal rights view, when thoroughgoing, is abolitionist as regards the use of animals...

  15. A novel transgenic mouse model of Chinese Charcot-Marie-Tooth disease type 2L

    Institute of Scientific and Technical Information of China (English)

    Ruxu Zhang; Qian Pan; Beisha Tang; Fufeng Zhang; Xiaobo Li; Shunxiang Huang; Xiaohong Zi; Ting Liu; Sanmei Liu; Xuning Li; Kun Xia

    2014-01-01

    We previously found that the K141N mutation in heat shock protein B8 (HSPB8) was respon-sible for Charcot-Marie-Tooth disease type 2L in a large Chinese family. The objective of the present study was to generate a transgenic mouse model bearing the K141N mutation in the human HSPB8 gene, and to determine whether this K141NHSPB8 transgenic mouse model would manifest the clinical phenotype of Charcot-Marie-Tooth disease type 2L, and consequently be suitable for use in studies of disease pathogenesis. Transgenic mice overexpressing K141NHSPB8 were generated using K141N mutant HSPB8 cDNA cloned into a pCAGGS plasmid driven by a human cytomegalovirus expression system. PCR and western blot analysis conifrmed integra-tion of the K141NHSPB8 gene and widespread expression in tissues of the transgenic mice. The K141NHSPB8 transgenic mice exhibited decreased muscle strength in the hind limbs and impaired motor coordination, but no obvious sensory disturbance at 6 months of age by behavioral assess-ment. Electrophysiological analysis showed that the compound motor action potential amplitude in the sciatic nerve was signiifcantly decreased, but motor nerve conduction velocity remained normal at 6 months of age. Pathological analysis of the sciatic nerve showed reduced myelinated ifber density, notable axonal edema and vacuolar degeneration in K141NHSPB8 transgenic mice, suggesting axonal involvement in the peripheral nerve damage in these animals. These ifndings indicate that the K141NHSPB8 transgenic mouse successfully models Charcot-Marie-Tooth disease type 2L and can be used to study the pathogenesis of the disease.

  16. Neuroanatomy and transgenic technologies

    Science.gov (United States)

    This is a short review that introduces recent advances of neuroanatomy and transgenic technologies. The anatomical complexity of the nervous system remains a subject of tremendous fascination among neuroscientists. In order to tackle this extraordinary complexity, powerful transgenic technologies a...

  17. Inducible gene manipulations in brain serotonergic neurons of transgenic rats.

    Directory of Open Access Journals (Sweden)

    Tillmann Weber

    Full Text Available The serotonergic (5-HT system has been implicated in various physiological processes and neuropsychiatric disorders, but in many aspects its role in normal and pathologic brain function is still unclear. One reason for this might be the lack of appropriate animal models which can address the complexity of physiological and pathophysiological 5-HT functioning. In this respect, rats offer many advantages over mice as they have been the animal of choice for sophisticated neurophysiological and behavioral studies. However, only recently technologies for the targeted and tissue specific modification of rat genes - a prerequisite for a detailed study of the 5-HT system - have been successfully developed. Here, we describe a rat transgenic system for inducible gene manipulations in 5-HT neurons. We generated a Cre driver line consisting of a tamoxifen-inducible CreERT2 recombinase under the control of mouse Tph2 regulatory sequences. Tissue-specific serotonergic Cre recombinase expression was detected in four transgenic TPH2-CreERT2 rat founder lines. For functional analysis of Cre-mediated recombination, we used a rat Cre reporter line (CAG-loxP.EGFP, in which EGFP is expressed after Cre-mediated removal of a loxP-flanked lacZ STOP cassette. We show an in-depth characterisation of this rat Cre reporter line and demonstrate its applicability for monitoring Cre-mediated recombination in all major neuronal subpopulations of the rat brain. Upon tamoxifen induction, double transgenic TPH2-CreERT2/CAG-loxP.EGFP rats show selective and efficient EGFP expression in 5-HT neurons. Without tamoxifen administration, EGFP is only expressed in few 5-HT neurons which confirms minimal background recombination. This 5-HT neuron specific CreERT2 line allows Cre-mediated, inducible gene deletion or gene overexpression in transgenic rats which provides new opportunities to decipher the complex functions of the mammalian serotonergic system.

  18. Evolution of somatic mutations in mammary tumors in transgenic mice is influenced by the inherited genotype

    Directory of Open Access Journals (Sweden)

    Li Yi

    2004-06-01

    Full Text Available Abstract Background MMTV-Wnt1 transgenic mice develop mammary hyperplasia early in development, followed by the appearance of solitary mammary tumors with a high proportion of cells expressing early lineage markers and many myoepithelial cells. The occurrence of tumors is accelerated in experiments that activate FGF proto-oncogenes or remove the tumor suppressor genes Pten or P53, implying that secondary oncogenic events are required for progression from mammary hyperplasia to carcinoma. It is not known, however, which oncogenic pathways contribute to Wnt1-induced tumorigenesis – further experimental manipulation of these mice is needed. Secondary events also appear to be required for mammary tumorigenesis in MMTV-Neu transgenic mice because the transgene in the tumors usually contains an acquired mutation that activates the Neu protein-tyrosine kinase. Methods cDNA or DNA from the mammary glands and mammary tumors from MMTV-Wnt1, MMTV-Wnt1/p53-/-, MMTV-Neu transgenic mice, and newly generated MMTV-Wnt1/MMTV-Neu bitransgenic mice, was sequenced to seek activating mutations in H-Ras, K-Ras, and N-Ras genes, or in the MMTV-Neu transgene. In addition, tumors from bitransgenic animals were examined to determine the cellular phenotype. Results We found activating mutations at codons 12, 13, and 61 of H-Ras in just over half of the mammary tumors in MMTV-Wnt1 transgenic mice, and we confirmed the high frequency of activating mutations of Neu in tumors in MMTV-Neu transgenic mice. Tumors appeared earlier in bitransgenic MMTV-Wnt1/MMTV-Neu mice, but no Ras or MMTV-Neu mutations were found in these tumors, which were phenotypically similar to those arising in MMTV-Wnt1 mice. In addition, no Ras mutations were found in the mammary tumors that arise in MMTV-Wnt1 transgenic mice lacking an intact P53 gene. Conclusions Tumorigenic properties of cells undergoing functionally significant secondary mutations in H-Ras or the MMTV-Neu transgene allow selection

  19. Energy generation for an ad hoc wireless sensor network-based monitoring system using animal head movement

    DEFF Research Database (Denmark)

    S. Nadimi, Esmaeil; Blanes-Vidal, Victoria; Jørgensen, Rasmus Nyholm

    2011-01-01

    The supply of energy to electronics is an imperative constraining factor to be considered during the design process of mobile ad hoc wireless sensor networks (MANETs). This influence is especially important when the MANET is deployed unattended or the wireless modules within the MANET are not eas......The supply of energy to electronics is an imperative constraining factor to be considered during the design process of mobile ad hoc wireless sensor networks (MANETs). This influence is especially important when the MANET is deployed unattended or the wireless modules within the MANET...... accessible; however, animal movement can be potentially used to generate energy. In this study, the head movements of individual sheep in a flock during grazing were monitored in order to investigate the amount of energy that can be generated by these movements. By applying the Lagrange–d’Alembert Principle...... to this problem, the equations of motion from each neck-mounted sensor as well as the amount of mechanical energy generated per time instant (each second) during upward and downward head movements were calculated. This resulted in the production of 857 mW and 1660 mW during the downward and upward movements...

  20. Generation and characterization of a stable red fluorescent ...

    African Journals Online (AJOL)

    AJL

    2012-04-12

    Apr 12, 2012 ... such as the fluorescent transgenic zebrafish (Danio rerio) and Japanese medaka ..... number in transgenic animals. Biotechniques, 37: ... Development of transgenic fish for ornamental and bioreactor by strong expression.

  1. Production of functional human nerve growth factor from the saliva of transgenic mice by using salivary glands as bioreactors

    OpenAIRE

    Fang Zeng; Zicong Li; Qingchun Zhu; Rui Dong; Chengcheng Zhao; Guoling Li; Guo Li; Wenchao Gao; Gelong Jiang; Enqin Zheng; Gengyuan Cai; Stefan Moisyadi; Johann Urschitz; Huaqiang Yang; Dewu Liu

    2017-01-01

    The salivary glands of animals have great potential to act as powerful bioreactors to produce human therapeutic proteins. Human nerve growth factor (hNGF) is an important pharmaceutical protein that is clinically effective in the treatment of many human neuronal and non-neuronal diseases. In this study, we generated 18 transgenic (TG) founder mice each carrying a salivary gland specific promoter-driven hNGF transgene. A TG mouse line secreting high levels of hNGF protein in its saliva (1.36 μ...

  2. Modification of the Genome of Domestic Animals.

    Science.gov (United States)

    Lotti, Samantha N; Polkoff, Kathryn M; Rubessa, Marcello; Wheeler, Matthew B

    2017-01-19

    In the past few years, new technologies have arisen that enable higher efficiency of gene editing. With the increase ease of using gene editing technologies, it is important to consider the best method for transferring new genetic material to livestock animals. Microinjection is a technique that has proven to be effective in mice but is less efficient in large livestock animals. Over the years, a variety of methods have been used for cloning as well as gene transfer including; nuclear transfer, sperm mediated gene transfer (SMGT), and liposome-mediated DNA transfer. This review looks at the different success rate of these methods and how they have evolved to become more efficient. As well as gene editing technologies, including Zinc-finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and the most recent clustered regulatory interspaced short palindromic repeats (CRISPRs). Through the advancements in gene-editing technologies, generating transgenic animals is now more accessible and affordable. The goals of producing transgenic animals are to 1) increase our understanding of biology and biomedical science; 2) increase our ability to produce more efficient animals; and 3) produce disease resistant animals. ZFNs, TALENs, and CRISPRs combined with gene transfer methods increase the possibility of achieving these goals.

  3. Identification of short hairpin RNA targeting foot-and-mouth disease virus with transgenic bovine fetal epithelium cells.

    Directory of Open Access Journals (Sweden)

    Hongmei Wang

    Full Text Available BACKGROUND: Although it is known that RNA interference (RNAi targeting viral genes protects experimental animals, such as mice, from the challenge of Foot-and-mouth disease virus (FMDV, it has not been previously investigated whether shRNAs targeting FMDV in transgenic dairy cattle or primary transgenic bovine epithelium cells will confer resistance against FMDV challenge. PRINCIPAL FINDING: Here we constructed three recombinant lentiviral vectors containing shRNA against VP2 (RNAi-VP2, VP3 (RNAi-VP3, or VP4 (RNAi-VP4 of FMDV, and found that all of them strongly suppressed the transient expression of a FLAG-tagged viral gene fusion protein in 293T cells. In BHK-21 cells, RNAi-VP4 was found to be more potent in inhibition of viral replication than the others with over 98% inhibition of viral replication. Therefore, recombinant lentiviral vector RNAi-VP4 was transfected into bovine fetal fibroblast cells to generate transgenic nuclear donor cells. With subsequent somatic cell cloning, we generated forty transgenic blastocysts, and then transferred them to 20 synchronized recipient cows. Three transgenic bovine fetuses were obtained after pregnant period of 4 months, and integration into chromosome in cloned fetuses was confirmed by Southern hybridization. The primary tongue epithelium cells of transgenic fetuses were isolated and inoculated with 100 TCID(50 of FMDV, and it was observed that shRNA significantly suppressed viral RNA synthesis and inhibited over 91% of viral replication after inoculation of FMDV for 48 h. CONCLUSION: RNAi-VP4 targeting viral VP4 gene appears to prevent primary epithelium cells of transgenic bovine fetus from FMDV infection, and it could be a candidate shRNA used for cultivation of transgenic cattle against FMDV.

  4. Doxycycline-Induced Expression of Transgenic Human Tumor Necrosis Factor α in Adult Mice Results in Psoriasis-like Arthritis

    Science.gov (United States)

    Retser, Eugen; Schied, Tanja; Skryabin, Boris V; Vogl, Thomas; Kanczler, Janos M; Hamann, Nina; Niehoff, Anja; Hermann, Sven; Eisenblätter, Michel; Wachsmuth, Lydia; Pap, Thomas; van Lent, Peter L E M; Loser, Karin; Roth, Johannes; Zaucke, Frank; Ludwig, Stephan; Wixler, Viktor

    2013-01-01

    Objective To generate doxycycline-inducible human tumor necrosis factor α (TNFα)–transgenic mice to overcome a major disadvantage of existing transgenic mice with constitutive expression of TNFα, which is the limitation in crossing them with various knockout or transgenic mice. Methods A transgenic mouse line that expresses the human TNFα cytokine exclusively after doxycycline administration was generated and analyzed for the onset of diseases. Results Doxycycline-inducible human TNFα–transgenic mice developed an inflammatory arthritis– and psoriasis-like phenotype, with fore and hind paws being prominently affected. The formation of “sausage digits” with characteristic involvement of the distal interphalangeal joints and nail malformation was observed. Synovial hyperplasia, enthesitis, cartilage and bone alterations, formation of pannus tissue, and inflammation of the skin epidermis and nail matrix appeared as early as 1 week after the treatment of mice with doxycycline and became aggravated over time. The abrogation of human TNFα expression by the removal of doxycycline 6 weeks after beginning stimulation resulted in fast resolution of the most advanced macroscopic and histologic disorders, and 3–6 weeks later, only minimal signs of disease were visible. Conclusion Upon doxycycline administration, the doxycycline-inducible human TNFα–transgenic mouse displays the major features of inflammatory arthritis. It represents a unique animal model for studying the molecular mechanisms of arthritis, especially the early phases of disease genesis and tissue remodeling steps upon abrogation of TNFα expression. Furthermore, unlimited crossing of doxycycline-inducible human TNFα–transgenic mice with various knockout or transgenic mice opens new possibilities for unraveling the role of various signaling molecules acting in concert with TNFα. PMID:23740547

  5. A comparative study between xerographic, computer-assisted overlay generation and animated-superimposition methods in bite mark analyses.

    Science.gov (United States)

    Tai, Meng Wei; Chong, Zhen Feng; Asif, Muhammad Khan; Rahmat, Rabiah A; Nambiar, Phrabhakaran

    2016-09-01

    This study was to compare the suitability and precision of xerographic and computer-assisted methods for bite mark investigations. Eleven subjects were asked to bite on their forearm and the bite marks were photographically recorded. Alginate impressions of the subjects' dentition were taken and their casts were made using dental stone. The overlays generated by xerographic method were obtained by photocopying the subjects' casts and the incisal edge outlines were then transferred on a transparent sheet. The bite mark images were imported into Adobe Photoshop® software and printed to life-size. The bite mark analyses using xerographically generated overlays were done by comparing an overlay to the corresponding printed bite mark images manually. In computer-assisted method, the subjects' casts were scanned into Adobe Photoshop®. The bite mark analyses using computer-assisted overlay generation were done by matching an overlay and the corresponding bite mark images digitally using Adobe Photoshop®. Another comparison method was superimposing the cast images with corresponding bite mark images employing the Adobe Photoshop® CS6 and GIF-Animator©. A score with a range of 0-3 was given during analysis to each precision-determining criterion and the score was increased with better matching. The Kruskal Wallis H test showed significant difference between the three sets of data (H=18.761, poverlay generation and lastly the xerographic method. The superior precision contributed by digital method is discernible despite the human skin being a poor recording medium of bite marks. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  6. Developments in transgenic technology: applications for medicine.

    Science.gov (United States)

    Hunter, Cheryl V; Tiley, Laurence S; Sang, Helen M

    2005-06-01

    Recent advances in the efficiency of transgenic technology have important implications for medicine. The production of therapeutic proteins from animal bioreactors is well established and the first products are close to market. The genetic modification of pigs to improve their suitability as organ donors for xenotransplantation has been initiated, but many challenges remain. The use of transgenesis, in combination with the method of RNA interference to knock down gene expression, has been proposed as a method for making animals resistant to viral diseases, which could reduce the likelihood of transmission to humans. Here, the latest developments in transgenic technology and their applications relevant to medicine and human health will be discussed.

  7. Pharming and transgenic plants.

    Science.gov (United States)

    Liénard, David; Sourrouille, Christophe; Gomord, Véronique; Faye, Loïc

    2007-01-01

    Plant represented the essence of pharmacopoeia until the beginning of the 19th century when plant-derived pharmaceuticals were partly supplanted by drugs produced by the industrial methods of chemical synthesis. In the last decades, genetic engineering has offered an alternative to chemical synthesis, using bacteria, yeasts and animal cells as factories for the production of therapeutic proteins. More recently, molecular farming has rapidly pushed towards plants among the major players in recombinant protein production systems. Indeed, therapeutic protein production is safe and extremely cost-effective in plants. Unlike microbial fermentation, plants are capable of carrying out post-translational modifications and, unlike production systems based on mammalian cell cultures, plants are devoid of human infective viruses and prions. Furthermore, a large panel of strategies and new plant expression systems are currently developed to improve the plant-made pharmaceutical's yields and quality. Recent advances in the control of post-translational maturations in transgenic plants will allow them, in the near future, to perform human-like maturations on recombinant proteins and, hence, make plant expression systems suitable alternatives to animal cell factories.

  8. “全鱼”转生长激素基因黄颡鱼首建者的建立%Generation of "all fish" growth hormone gene transgenic yellow catfish founders

    Institute of Scientific and Technical Information of China (English)

    葛家春; 宋伟; 董张及; 许志强; 鲍洁; 周国勤; 潘建林; 杨家新; 赵庆顺

    2013-01-01

    黄颡鱼(Pelteobagrus fulvidraco)是一种重要的淡水名优经济鱼类,然而较小的体型和较慢的生长速率极大地降低了它的经济价值.为获得大规格且快速生长的黄颡鱼,开展了转生长激素基因黄颡鱼的研究.运用RT PCR和RACE-PCR技术,克隆了黄颡鱼生长激素基因的603 bp(碱基对)的编码序列和485 bp的3′UTR(非翻译区)序列.通过重叠PCR和限制性内切酶酶切连接,组建了一个长为2105 bp的“全鱼”转生长激素基因构件,该构件含1017 bp的黄颡鱼β-肌动蛋白近端启动子、603 bp的生长激素基因编码序列和485 bp的3′-UTR序列.采用显微注射将转生长激素基因构件导入黄颡鱼受精卵,从301尾由注射胚胎发育而来的黄颡鱼中共筛选获得40尾转生长激素基因黄颡鱼首建者.成功地建立了黄颡鱼基因组改造的技术平台,为黄颡鱼的基因工程育种奠定了基础.%Yellow catfish (Pelteobagrus fulvidraco Richardson) is one of the most important freshwater farmed species in China. However, its small size and slow growth rate limit its economic value. Growth hormone is a protein secreted by pituitary. It promotes animal growth. Overexpression of growth hormone in fishes results in the growth hormone gene transgenic fishes with enlarged body size and accelerated growth. In order to increase its body size and growth rate, we performed transgenic research on yellow catfish. Employing RT-PCR and RACE(rapid amplification of cDNA ends)-PCR, we cloned 603 bp (base pairs) complete coding sequence and 485 bp 3'-UTR (untranslated region) of yellow catfish growth hormone gene, respectively. Performing overlapping PCR and molecular cloning with restriction endonucleases, we constructed an "all fish" growth hormone gene transgenic construct with 2105 bp in length. The construct comprises 1017 bp of yellow catfish β-actin proximal promoter, 603 bp complete coding sequence and 485 bp 3'-UTR of yellow catfish growth hormone

  9. Permanent neonatal diabetes in INS(C94Y) transgenic pigs.

    Science.gov (United States)

    Renner, Simone; Braun-Reichhart, Christina; Blutke, Andreas; Herbach, Nadja; Emrich, Daniela; Streckel, Elisabeth; Wünsch, Annegret; Kessler, Barbara; Kurome, Mayuko; Bähr, Andrea; Klymiuk, Nikolai; Krebs, Stefan; Puk, Oliver; Nagashima, Hiroshi; Graw, Jochen; Blum, Helmut; Wanke, Ruediger; Wolf, Eckhard

    2013-05-01

    Mutations in the insulin (INS) gene may cause permanent neonatal diabetes mellitus (PNDM). Ins2 mutant mouse models provided important insights into the disease mechanisms of PNDM but have limitations for translational research. To establish a large animal model of PNDM, we generated INS(C94Y) transgenic pigs. A line expressing high levels of INS(C94Y) mRNA (70-86% of wild-type INS transcripts) exhibited elevated blood glucose soon after birth but unaltered β-cell mass at the age of 8 days. At 4.5 months, INS(C94Y) transgenic pigs exhibited 41% reduced body weight, 72% decreased β-cell mass (-53% relative to body weight), and 60% lower fasting insulin levels compared with littermate controls. β-cells of INS(C94Y) transgenic pigs showed a marked reduction of insulin secretory granules and severe dilation of the endoplasmic reticulum. Cataract development was already visible in 8-day-old INS(C94Y) transgenic pigs and became more severe with increasing age. Diabetes-associated pathological alterations of kidney and nervous tissue were not detected during the observation period of 1 year. The stable diabetic phenotype and its rescue by insulin treatment make the INS(C94Y) transgenic pig an attractive model for insulin supplementation and islet transplantation trials, and for studying developmental consequences of maternal diabetes mellitus.

  10. Mating animals by minimising the covariance between ancestral contributions generates less inbreeding without compromising genetic gain in breeding schemes with truncation selection

    DEFF Research Database (Denmark)

    Henryon, M; Berg, P; Sørensen, A C

    2009-01-01

    We reasoned that mating animals by minimising the covariance between ancestral contributions (MCAC mating) will generate less inbreeding and at least as much genetic gain as minimum-coancestry mating in breeding schemes where the animals are truncation-selected. We tested this hypothesis...... they can be achieved without extra costs or practical constraints. MCAC mating merely uses pedigree information to pair the animals more appropriately and is clearly a worthy alternative to minimum-coancestry mating and probably any other mating criterion. We believe, therefore, that MCAC mating should...... by stochastic simulation and compared the mating criteria in hierarchical and factorial breeding schemes, where the animals were selected based on breeding values predicted by animal-model BLUP. Random mating was included as a reference-mating criterion. We found that MCAC mating generated 4% to 8% less...

  11. Production of recombinant proteins in milk of transgenic and non-transgenic goats

    Directory of Open Access Journals (Sweden)

    Raylene Ramos Moura

    2011-10-01

    Full Text Available Among all the transgenic mammalians produced so far, goats have represented an excellent model of transgenesis when considering the factors such as the market demand for protein, volume of milk produced per lactation and reproductive rate. Various recombinant proteins have been obtained from the transgenic and non-transgenic goats, and among these, human antithrombin, produced by the transgenic goats, was the first recombinant protein of animal origin to be released as a drug for the clinical use in humans. This review reports the aspects inherent to the production of recombinant proteins in the goats, from the production of the animal bioreactors up to the expression of these proteins in their milk.

  12. Transgenic plants with enhanced growth characteristics

    Energy Technology Data Exchange (ETDEWEB)

    Unkefer, Pat J.; Anderson, Penelope S.; Knight, Thomas J.

    2016-09-06

    The invention relates to transgenic plants exhibiting dramatically enhanced growth rates, greater seed and fruit/pod yields, earlier and more productive flowering, more efficient nitrogen utilization, increased tolerance to high salt conditions, and increased biomass yields. In one embodiment, transgenic plants engineered to over-express both glutamine phenylpyruvate transaminase (GPT) and glutamine synthetase (GS) are provided. The GPT+GS double-transgenic plants of the invention consistently exhibit enhanced growth characteristics, with T0 generation lines showing an increase in biomass over wild type counterparts of between 50% and 300%. Generations that result from sexual crosses and/or selfing typically perform even better, with some of the double-transgenic plants achieving an astounding four-fold biomass increase over wild type plants.

  13. Development of transgenic rats producing human β-amyloid precursor protein as a model for Alzheimer's disease: Transgene and endogenous APP genes are regulated tissue-specifically

    Directory of Open Access Journals (Sweden)

    Chan Anthony WS

    2008-02-01

    Full Text Available Abstract Background Alzheimer's disease (AD is a devastating neurodegenerative disorder that affects a large and growing number of elderly individuals. In addition to idiopathic disease, AD is also associated with autosomal dominant inheritance, which causes a familial form of AD (FAD. Some instances of FAD have been linked to mutations in the β-amyloid protein precursor (APP. Although there are numerous mouse AD models available, few rat AD models, which have several advantages over mice, have been generated. Results Fischer 344 rats expressing human APP driven by the ubiquitin-C promoter were generated via lentiviral vector infection of Fischer 344 zygotes. We generated two separate APP-transgenic rat lines, APP21 and APP31. Serum levels of human amyloid-beta (Aβ40 were 298 pg/ml for hemizygous and 486 pg/ml for homozygous APP21 animals. Serum Aβ42 levels in APP21 homozygous rats were 135 pg/ml. Immunohistochemistry in brain showed that the human APP transgene was expressed in neurons, but not in glial cells. These findings were consistent with independent examination of enhanced green fluorescent protein (eGFP in the brains of eGFP-transgenic rats. APP21 and APP31 rats expressed 7.5- and 3-times more APP mRNA, respectively, than did wild-type rats. Northern blots showed that the human APP transgene, driven by the ubiquitin-C promoter, is expressed significantly more in brain, kidney and lung compared to heart and liver. A similar expression pattern was also seen for the endogenous rat APP. The unexpected similarity in the tissue-specific expression patterns of endogenous rat APP and transgenic human APP mRNAs suggests regulatory elements within the cDNA sequence of APP. Conclusion This manuscript describes the generation of APP-transgenic inbred Fischer 344 rats. These are the first human AD model rat lines generated by lentiviral infection. The APP21 rat line expresses high levels of human APP and could be a useful model for AD. Tissue

  14. Generation of transgenic rice lines with reduced contents of multiple potential allergens using a null mutant in combination with an RNA silencing method.

    Science.gov (United States)

    Wakasa, Yuhya; Hirano, Kana; Urisu, Atsuo; Matsuda, Tsukasa; Takaiwa, Fumio

    2011-12-01

    Rice seed proteins are known to be a causative antigen in some patients with food allergy, especially cereal allergy, with clinical symptoms such as eczema and dermatitis. The α-amylase/trypsin inhibitors (14-16 kDa), α-globulin (26 kDa) and β-glyoxalase I (33 kDa) are regarded as major potential allergens of rice (Oryza sativa L.) seed based on specific recognition by serum IgE from allergy patients. In order to suppress the production of these major allergens in rice grains, a mutant in the 'Koshihikari' background lacking the 26 kDa allergen (GbN-1) was used as a host for RNA silencing. A binary vector harboring two RNA interference (RNAi) gene cassettes for suppression of 14-16 kDa and 33 kDa allergens driven by the 13 kDa and 10 kDa prolamin endosperm-specific promoters, respectively, was introduced into the GbN-1 genome by Agrobacterium-mediated transformation. In the most promising transgenic line, the content of the three potential allergens was remarkably reduced to a very faint level without a change in seed phenotype. IgE binding of 15 patients' sera to the transgenic rice seed mostly deficient in the three major allergens was on average only about 10% that of the control wild-type rice, suggesting that these three accounted for the great majority of rice seed causative allergens recognized by patients' IgE and that the sequential allergen deletion/reduction strategy works in the development of hypo-allergenic rice lines.

  15. Characterization of transgene integration pattern in F4 hGH-transgenic common carp (Cyprinus carpio L.)

    Institute of Scientific and Technical Information of China (English)

    Bo WU; Yong Hua SUN; Yan Wu WANG; Ya Ping WANG; Zuo Yan ZHU

    2005-01-01

    The integration pattern and adjacent host sequences of the inserted pMThGH-transgene in the F4 hGH-transgenic common carp were extensively studied. Here we show that each F4 transgenic fish contained about 200 copies of the pMThGH-transgene and the transgenes were integrated into the host genome generally with concatemers in a head-totail arrangement at 4-5 insertion sites. By using a method of plasmid rescue, four hundred copies of transgenes from two individuals of F4 transgenic fish, A and B, were recovered and clarified into 6 classes. All classes of recovered transgenes contained either complete or partial pMThGH sequences. The class Ⅰ, which comprised 83% and 84.5% respectively of the recovered transgene copies from fish A and B, had maintained the original configuration, indicating that most transgenes were faithfully inherited during the four generations of reproduction. The other five classes were different from the original configuration in both molecular weight and restriction map, indicating that a few transgenes had undergone mutation, rearrangement or deletion during integration and germline transmission. In the five types of aberrant transgenes, three flanking sequences of the host genome were analyzed. These sequences were common carp β-actin gene, common carp DNA sequences homologous to mouse phosphoglycerate kinase-1 and human epidermal keratin 14, respectively.

  16. Generation of Five Human Lactoferrin Transgenic Cloned Goats Using Fibroblast Cells and Their Methylation Status of Putative Differential Methylation Regions of IGF2R and H19 Imprinted Genes

    NARCIS (Netherlands)

    Meng, L.; Wan, Y.; Sun, Y.; Zhang, Y.; Wang, Z.; Song, Y.; Wang, F.

    2013-01-01

    Background - Somatic cell nuclear transfer (SCNT) is a promising technique to produce transgenic cloned mammalian, including transgenic goats which may produce Human Lactoferrin (hLF). However, success percentage of SCNT is low, because of gestational and neonatal failure of transgenic embryos.

  17. Strategies to enable the adoption of animal biotechnology to sustainably improve global food safety and security.

    Science.gov (United States)

    Tizard, Mark; Hallerman, Eric; Fahrenkrug, Scott; Newell-McGloughlin, Martina; Gibson, John; de Loos, Frans; Wagner, Stefan; Laible, Götz; Han, Jae Yong; D'Occhio, Michael; Kelly, Lisa; Lowenthal, John; Gobius, Kari; Silva, Primal; Cooper, Caitlin; Doran, Tim

    2016-10-01

    The ability to generate transgenic animals has existed for over 30 years, and from those early days many predicted that the technology would have beneficial applications in agriculture. Numerous transgenic agricultural animals now exist, however to date only one product from a transgenic animal has been approved for the food chain, due in part to cumbersome regulations. Recently, new techniques such as precision breeding have emerged, which enables the introduction of desired traits without the use of transgenes. The rapidly growing human population, environmental degradation, and concerns related to zoonotic and pandemic diseases have increased pressure on the animal agriculture sector to provide a safe, secure and sustainable food supply. There is a clear need to adopt transgenic technologies as well as new methods such as gene editing and precision breeding to meet these challenges and the rising demand for animal products. To achieve this goal, cooperation, education, and communication between multiple stakeholders-including scientists, industry, farmers, governments, trade organizations, NGOs and the public-is necessary. This report is the culmination of concepts first discussed at an OECD sponsored conference and aims to identify the main barriers to the adoption of animal biotechnology, tactics for navigating those barriers, strategies to improve public perception and trust, as well as industry engagement, and actions for governments and trade organizations including the OECD to harmonize regulations and trade agreements. Specifically, the report focuses on animal biotechnologies that are intended to improve breeding and genetics and currently are not routinely used in commercial animal agriculture. We put forward recommendations on how scientists, regulators, and trade organizations can work together to ensure that the potential benefits of animal biotechnology can be realized to meet the future needs of agriculture to feed the world.

  18. Delayed allograft rejection in mice transgenic for a soluble form of the IL-4 receptor.

    Science.gov (United States)

    Maliszewski, C R; Morrissey, P J; Fanslow, W C; Sato, T A; Willis, C; Davison, B

    1992-09-01

    Accumulating evidence suggests that in serum and other biological fluids, cytokine binding is a property associated with soluble proteins, including a high-affinity soluble version of the IL-4 receptor (sIL-4R). While it is tempting to speculate that sIL-4R might act as a serum carrier protein or serve to inhibit or modulate IL-4 action, specific biological roles for sIL-4R remain to be established. To further assess the immunoregulatory and therapeutic potential of sIL-4R and other soluble receptors, we have created transgenic mice which constitutively express elevated levels of biologically active sIL-4R. Phenotypic characterization of lymphoid organs in sIL-4R transgenic mice revealed normal numbers of B and T cells and normal surface marker expression. Splenic lymphocytes displayed normal in vitro activities as measured by the PFC response and generation of cytotoxic T cells. In addition, antigen-specific IgE and IgG1 in vivo responses were similar in control and transgenic mice. Despite the apparent developmental normality of the sIL-4R transgenic mice, these animals were markedly deficient in the ability to reject cardiac allografts, suggesting that IL-4 is critical for the generation of alloreactivity. The results further suggest that the ability of sIL-4R to regulate IL-4 activities may be under the control of complex interactions that remain to be elucidated.

  19. Assessment of novel foods in animal nutrition.

    Science.gov (United States)

    Flachowsky, Gerhard; Aulrich, Karen

    2003-01-01

    Composition of feeds from GMO crops were determined as well as digestion and feeding experiments were carried out with broilers (Bt-corn), layers (Bt-Corn, Pat-corn), pigs (Bt-corn, Pat-sugar beet, soybeans), sheep (Bt-corn silage, Pat-corn silage), growing bulls (Bt-corn silage) and fistulated cows (Bt-corn silage). Up to now, no significant differences in nutritional value between feeds from isogenic and transgenic plants of the first generation were observed. The so-called substantial equivalence, but also the nutritional equivalence of transgenic hybrids could be demonstrated. Recombinant plant DNA constructs were not detected in animal tissues samples. In the future, long term feeding experiments for nutritional assessment of novel feeds should be combined with risk assessment studies. Proposal for discussion has been submitted.

  20. Exploring Shifts in Middle School Learners' Modeling Activity While Generating Drawings, Animations, and Computational Simulations of Molecular Diffusion

    Science.gov (United States)

    Wilkerson-Jerde, Michelle H.; Gravel, Brian E.; Macrander, Christopher A.

    2015-04-01

    Modeling and using technology are two practices of particular interest to K-12 science educators. These practices are inextricably linked among professionals, who engage in modeling activity with and across a variety of representational technologies. In this paper, we explore the practices of five sixth-grade girls as they generated models of smell diffusion using drawing, stop-motion animation, and computational simulation during a multi-day workshop. We analyze video, student discourse, and artifacts to address the questions: In what ways did learners' modeling practices, reasoning about mechanism, and ideas about smell shift as they worked across this variety of representational technologies? And, what supports enabled them to persist and progress in the modeling activity? We found that the girls engaged in two distinct modeling cycles that reflected persistence and deepening engagement in the task. In the first, messing about, they focused on describing and representing many ideas related to the spread of smell at once. In the second, digging in, they focused on testing and revising specific mechanisms that underlie smell diffusion. Upon deeper analysis, we found these cycles were linked to the girls' invention of "oogtom," a representational object that encapsulated many ideas from the first cycle and allowed the girls to restart modeling with the mechanistic focus required to construct simulations. We analyze the role of activity design, facilitation, and technological infrastructure in this pattern of engagement over the course of the workshop and discuss implications for future research, curriculum design, and classroom practice.

  1. Estimated Seed Shadow Generated by Japanese Martens(Martes melampus): Comparison with Forest-Dwelling Animals in Japan.

    Science.gov (United States)

    Tsuji, Yamato; Okumura, Tadanobu; Kitahara, Masahiko; Jiang, Zhaowen

    2016-08-01

    To evaluate the seed shadow generated by wild Japanese martens (Martes melampus), we combined data on their ranging behavior from the northern foot of Mt. Fuji, central Japan (seven males and three females) with data on gut passage time obtained from martens in Toyama Municipal Family Park Zoo (three males and one female). The movement distances varied, and mean distances for 0-1, 2-3, and 4-5 h intervals were 152.4, 734.7, and 1,162.4 m, respectively, with no significant sex difference. The mean gut passage time of ingested seeds was 7.4 h (range: 0.6-51.7 h), and two-thirds were defecated within 12 h. Seeds of fleshy fruits was frequently transported to 501-1,000 m, and 20% of ingested seeds were transported > 1,000 m from feeding sites. We found positive correlations between body size and home range of the animals in Japan and their seed dispersal distances. We conclude that Japanese martens are medium-range dispersers that can transport seeds from the source to open habitats conducive for germination and/or growth, partly due to scent marking behaviors.

  2. Dendritic poly(L-lysine)-b-Poly(L-lactide)-b-dendritic poly(L-lysine) amphiphilic gene delivery vectors: roles of PLL dendritic generation and enhanced transgene efficacies via termini modification.

    Science.gov (United States)

    Li, Yang; Zhu, Yingdan; Xia, Kejia; Sheng, Ruilong; Jia, Lin; Hou, Xiaodong; Xu, Yuhong; Cao, Amin

    2009-08-10

    As an effort to prepare new efficient gene delivery vectors, we have recently developed and reported an amphiphilic dendritic poly(L-lysine)-b-poly(L-lactide)-b-dendritic poly(L-lysine) D(2)-PLLA-D(2) with two-generation PLL dendrons and a PLLA block. In this work, we continued to explore the roles of dendritic PLL generation in DNA binding and intracellular delivery of gene, and a new series of amphiphilic dendritic poly(L-lysine)-b-poly(L-lactide)-b-dendritic poly(L-lysine)s D(n)-PLLA-D(n) (n = 3-5) were synthesized and were structurally characterized. Furthermore, plasmid DNA binding affinity for these cationic amphiphiles was examined by agarose gel electrophoresis and fluorescence titration assay in pure water and PBS buffer solution containing 150 mM NaCl (pH = 7.4), respectively. By dynamic light scattering (DLS) and transmission electronic microscopy (TEM), the interaction and complexation in between were investigated, concerning the DNA/vector polyplex particle morphologies and zeta potentials. Utilizing a human hepatocellular carcinoma cell-line SMMC-7721, cell toxicity, and gene transfection in vitro were explored. To further improve transgene efficiency for these synthetic cationic amphiphiles as gene delivery vectors, new structural DE(n)-PLLA-DE(n) (n = 2-3) were prepared through an amino termini modification of the D(n)-PLLA-D(n) (n = 2-3) with less toxic 4,7,10,13-tetraazatridecanoic acids, and gene transfection with these DE(n)-PLLA-DE(n) (n = 2-3) was examined with an alternative human gastric carcinoma cell-line HGC-27. As a result, the high plasmid DNA binding affinity, low cytotoxicity, and much enhanced transgene efficacy suggest a new possible clue to design effective synthetic gene delivery vectors with amphiphilic skeleton and less toxic polyamine building blocks.

  3. A germline GFP transgenic axolotl and its use to track cell fate: dual origin of the fin mesenchyme during development and the fate of blood cells during regeneration.

    Science.gov (United States)

    Sobkow, Lidia; Epperlein, Hans-Henning; Herklotz, Stephan; Straube, Werner L; Tanaka, Elly M

    2006-02-15

    The development of transgenesis in axolotls is crucial for studying development and regeneration as it would allow for long-term cell fate tracing as well as gene expression analysis. We demonstrate here that plasmid injection into the one-cell stage axolotl embryo generates mosaic transgenic animals that display germline transmission of the transgene. The inclusion of SceI meganuclease in the injections (Thermes, V., Grabher, C., Ristoratore, F., Bourrat, F., Choulika, A., Wittbrodt, J., Joly, J.S., 2002. I-SceI meganuclease mediates highly efficient transgenesis in fish. Mech. Dev. 118, 91-98) resulted in a higher percentage of F0 animals displaying strong expression throughout the body. This represents the first demonstration in the axolotl of germline transmission of a transgene. Using this technique we have generated a germline transgenic animal expressing GFP ubiquitously in all tissues examined. We have used this animal to study cell fate in the dorsal fin during development. We have uncovered a contribution of somite cells to dorsal fin mesenchyme in the axolotl, which was previously assumed to derive solely from neural crest. We have also studied the role of blood during tail regeneration by transplanting the ventral blood-forming region from GFP+ embryos into unlabeled hosts. During tail regeneration, we do not observe GFP+ cells contributing to muscle or nerve, suggesting that during tail regeneration blood stem cells do not undergo significant plasticity.

  4. Subchronic toxicity study of GH transgenic carp.

    Science.gov (United States)

    Yong, Ling; Liu, Yu-Mei; Jia, Xu-Dong; Li, Ning; Zhang, Wen-Zhong

    2012-11-01

    A subchronic toxicity study of GH (growth hormone) transgenic carp was carried out with 60 SD rats aged 4 weeks, weight 115∼125 g. Ten male and 10 female rats were allotted into each group. Animals of the three groups (transgenic carp group (GH-TC), parental carp group (PC) and control group) were fed soy- and alfalfa-free diet (SAFD) with 10% GH transgenic carp powder, 10% parental carp powder or 10% common carp powder for 90 consecutive days, respectively. In the end of study, animals were killed by exsanguination via the carotid artery under diethyl ether anesthesia, then weights of heart, liver, kidneys, spleen, thymus, brain, ovaries and uterus/testis were measured. Pathological examination of organs was determined. Endocrine hormones of triiodothyronine (T3), thyroid hormone (T4), follicle-stimulating hormone (FSH), 17β-estradiol (E2), progesterone (P) and testosterone (T) levels were detected by specific ELISA kit. Parameters of blood routine and blood biochemical were measured. The weights of the body and organs of the rats, food intake, blood routine, blood biochemical test and serum hormones showed no significant differences among the GH transgenic carp-treated, parental carp-treated and control groups (P>0.05). Thus, it was concluded that at the dose level of this study, GH transgenic carp showed no subchronic toxicity and endocrine disruption to SD rats.

  5. [An estimation of morphofunctional state of the animals of the first generation from parents long time exposed in hazardous area of Chernobyl NPS].

    Science.gov (United States)

    Konoplia, E F; Verezhchako, G G; Andronova, E V; Zaĭtseva, O A; Fedosenko, O L

    2009-01-01

    The morphofunctional state offspring of rats first generation in age 3 and 6 monthes from long time exposed males and females in hazardous area of Chernobyl NPS was studied (registration point Masany). At experimental animals the damages in blood, in sex systems, in heredetic apparatus, in morrow cells and in blood lymphocytes were shown, that testify about the effects of low-intensive long-term radiation on organism. Obtained data indicate remote consequences of radiation-ecological factors of environmental surroundings at posterity animal of generation F1 breeding in zone of enhanced radioactive contamination.

  6. The effect of Bt-transgene introgression on plant growth and reproduction in wild Brassica juncea.

    Science.gov (United States)

    Liu, Yong-Bo; Darmency, Henry; Stewart, C Neal; Wei, Wei; Tang, Zhi-Xi; Ma, Ke-Ping

    2015-06-01

    This study aims to investigate the relative plant growth and reproduction of insect-resistant and susceptible plants following the introgression of an insect-resistance Bt-transgene from Brassica napus, oilseed rape, to wild Brassica juncea. The second backcrossed generation (BC2) from a single backcross family was grown in pure and mixed stands of Bt-transgenic and non-transgenic siblings under two insect treatments. Various proportions of Bt-transgenic plants were employed in mixed stands to study the interaction between resistant and susceptible plants. In the pure stands, Bt-transgenic BC2 plants performed better than non-transgenic plants with or without insect treatments. In mixed stands, Bt-transgenic BC2 plants produced fewer seeds than their non-Bt counterparts at low proportions of Bt-transgenic BC2 plants in the absence of insects. Reproductive allocation of non-transgenic plants marginally increased with increasing proportions of Bt-transgenic plants under herbivore pressure, which resulted in increased total biomass and seed production per stand. The results showed that the growth of non-transgenic plants was protected by Bt-transgenic plants under herbivore pressure. The Bt-transgene might not be advantageous in mixed stands of backcrossed hybrids; thus transgene introgression would not be facilitated when herbivorous insects are not present. However, a relatively large initial population of Bt-transgenic plants might result in transgene persistence when target herbivores are present.

  7. Recombinant Human Factor IX Produced from Transgenic Porcine Milk

    OpenAIRE

    Meng-Hwan Lee; Yin-Shen Lin; Ching-Fu Tu; Chon-Ho Yen

    2014-01-01

    Production of biopharmaceuticals from transgenic animal milk is a cost-effective method for highly complex proteins that cannot be efficiently produced using conventional systems such as microorganisms or animal cells. Yields of recombinant human factor IX (rhFIX) produced from transgenic porcine milk under the control of the bovine α-lactalbumin promoter reached 0.25 mg/mL. The rhFIX protein was purified from transgenic porcine milk using a three-column purification scheme after a precipitat...

  8. Transgenes in F4 pMThGH- transgenic common carp (Cy- prinus carpio L.) are highly polymorphic

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    To gain information on the integration pattern of pMThGH-tansgene, 50 transgenes were recovered from F4 generation of pMThGH transgenic common carp (Cyprinus carpio L.) and 33 recovered genes were analyzed. The restriction maps of these recovered genes were constructed by digestion with five kinds of enzymes. These transgenes can be classified into 4 types according to their restriction maps. Only one type of transgenes maintains its original molecular form, whereas the other three types are very different from the original one and vary each other on both molecular weight and restriction maps. This implies that the sequences of most transgenes have been deleted and/or rearranged during integration and inheritance. The results of PCR am-plification and Southern blot hybridization indicate that MThGH in TypeI transgene keeps intact but most of its se-quence has been deleted in other three types. All these results suggest that transgenes in F4 generation of transgenic carp are highly polymorphic. Two DNA fragments concerning integration site of transgenes were cloned from recovered transgenes, and found to be homologous to the 5′UTR of β-actin gene of common carp and mouse mRNA for receptor tyrosine kinase (RTK), respectively.

  9. Germline modification of domestic animals.

    Science.gov (United States)

    Tang, L; González, R; Dobrinski, I

    2015-01-01

    Genetically-modified domestic animal models are of increasing significance in biomedical research and agriculture. As authentic ES cells derived from domestic animals are not yet available, the prevailing approaches for engineering genetic modifications in those animals are pronuclear microinjection and somatic cell nuclear transfer (SCNT, also known as cloning). Both pronuclear microinjection and SCNT are inefficient, costly, and time-consuming. In animals produced by pronuclear microinjection, the exogenous transgene is usually inserted randomly into the genome, which results in highly variable expression patterns and levels in different founders. Therefore, significant efforts are required to generate and screen multiple founders to obtain animals with optimal transgene expression. For SCNT, specific genetic modifications (both gain-of-function and loss-of-function) can be engineered and carefully selected in the somatic cell nucleus before nuclear transfer. SCNT has been used to generate a variety of genetically modified animals such as goats, pigs, sheep and cattle; however, animals resulting from SCNT frequently suffer from developmental abnormalities associated with incomplete nuclear reprogramming. Other strategies to generate genetically-modified animals rely on the use of the spermatozoon as a natural vector to introduce genetic material into the female gamete. This sperm mediated DNA transfer (SMGT) combined with intracytoplasmatic sperm injection (ICSI) has relatively high efficiency and allows the insertion of large DNA fragments, which, in turn, enhance proper gene expression. An approach currently being developed to complement SCNT for producing genetically modified animals is germ cell transplantation using genetically modified male germline stem cells (GSCs). This approach relies on the ability of GSCs that are genetically modified in vitro to colonize the recipient testis and produce donor derived sperm upon transplantation. As the genetic change

  10. Improved generation of rat gene knockouts by target-selected mutagenesis in mismatch repair-deficient animals

    Directory of Open Access Journals (Sweden)

    van Roekel Henk S

    2008-10-01

    Full Text Available Abstract Background The laboratory rat (Rattus norvegicus is one of the preferred model organisms in physiological and pharmacological research, although the availability of specific genetic models, especially gene knockouts, is limited. N-ethyl-N-nitrosourea (ENU-driven target-selected mutagenesis is currently the most successful method in rats, although it is still very laborious and expensive. Results As ENU-induced DNA damage is normally recognized by the mismatch repair (MMR system, we hypothesized that the effectiveness of the target-selected mutagenesis approach could be improved by using a MMR-deficient genetic background. Indeed, Msh6 knockout rats were found to be more sensitive to ENU treatment and the germ line mutation rate was boosted more than two-fold to 1 mutation per 585 kb. In addition, the molecular mutation spectrum was found to be changed in favor of generating knockout-type alleles by ~20%, resulting in an overall increase in efficiency of ~2.5 fold. The improved effectiveness was demonstrated by high throughput mutation discovery in 70 Mb of sequence in a set of only 310 mutant F1 rats. This resulted in the identification of 89 mutations of which four introduced a premature stopcodon and 64 resulted in amino acid changes. Conclusion Taken together, we show that the use of a MMR-deficient background considerably improves ENU-driven target-selected mutagenesis in the rat, thereby reducing animal use as well as screening costs. The use of a mismatch repair-deficient genetic background for improving mutagenesis and target-selected knockout efficiency is in principle applicable to any organism of interest.

  11. Improved generation of rat gene knockouts by target-selected mutagenesis in mismatch repair-deficient animals

    Science.gov (United States)

    van Boxtel, Ruben; Toonen, Pim W; Verheul, Mark; van Roekel, Henk S; Nijman, Isaac J; Guryev, Victor; Cuppen, Edwin

    2008-01-01

    Background The laboratory rat (Rattus norvegicus) is one of the preferred model organisms in physiological and pharmacological research, although the availability of specific genetic models, especially gene knockouts, is limited. N-ethyl-N-nitrosourea (ENU)-driven target-selected mutagenesis is currently the most successful method in rats, although it is still very laborious and expensive. Results As ENU-induced DNA damage is normally recognized by the mismatch repair (MMR) system, we hypothesized that the effectiveness of the target-selected mutagenesis approach could be improved by using a MMR-deficient genetic background. Indeed, Msh6 knockout rats were found to be more sensitive to ENU treatment and the germ line mutation rate was boosted more than two-fold to 1 mutation per 585 kb. In addition, the molecular mutation spectrum was found to be changed in favor of generating knockout-type alleles by ~20%, resulting in an overall increase in efficiency of ~2.5 fold. The improved effectiveness was demonstrated by high throughput mutation discovery in 70 Mb of sequence in a set of only 310 mutant F1 rats. This resulted in the identification of 89 mutations of which four introduced a premature stopcodon and 64 resulted in amino acid changes. Conclusion Taken together, we show that the use of a MMR-deficient background considerably improves ENU-driven target-selected mutagenesis in the rat, thereby reducing animal use as well as screening costs. The use of a mismatch repair-deficient genetic background for improving mutagenesis and target-selected knockout efficiency is in principle applicable to any organism of interest. PMID:18840264

  12. Production of transgenic medaka with increased resistance to bacterial pathogens.

    Science.gov (United States)

    Sarmasik, Aliye; Warr, Gregory; Chen, Thomas T

    2002-06-01

    Cecropins, first identified in silk moth (Hyalophora cecropia), are a group of antimicrobial peptides with bactericidal activity against a broad spectrum of bacteria. In this study we investigated whether (1) this group of antimicrobial peptides could exhibit bactericidal activity toward known fish bacterial pathogens and (2) expression of cecropin transgenes in transgenic medaka (Oryzias latipas) could result in increasing resistance of the transgenic fish to infection by fish bacterial pathogens. Cecropin gene construct containing silk moth preprocecropin B, procecropin B and cecropin B, and porcine cecropin P1 driven by a cytomegalovirus (CMV) promoter were transfected into chinook salmon embryonic cells (CHSE-214) by lipofection, and the resulting permanent transformants were collected. In an "inhibition zone" assay medium isolated from each transformant exhibited strong bactericidal activity toward known fish bacterial pathogens such as Pseudomonas fluorescens, Aeromonas hydrophila, and Vibrio anguillarum. The same cecropin transgene constructs were introduced into newly fertilized medaka eggs by electroporation to produce transgenic fish. About 40% to 60% of the embryos survived from electroporation, and about 5% to 11% of the surviving fish were shown to contain cecropin transgenes by polymerase chain reaction analysis of genomic DNA samples isolated from presumptive transgenic fish. These P1 transgenic fish were used as founder stocks, and following generations of successive breeding, a total of 20 F2 families of transgenic fish were established. Expression of cecropin transgenes was detected in the F2 transgenics by reverse transcriptase polymerase chain reaction analysis. Southern blot analysis of genomic DNA isolated from different F2 fish showed that cecropin transgenes were integrated into the genomes of F2 transgenic fish. To determine whether transgenic fish carrying cecropin transgenes could exhibit resistance to infection by known fish bacterial

  13. A simplified, robust, and streamlined procedure for the production of C. elegans transgenes via recombineering

    Directory of Open Access Journals (Sweden)

    Fisher Alfred L

    2008-12-01

    Full Text Available Abstract Background The nematode Caenorhabditis elegans has emerged as a powerful system to study biologic questions ranging from development to aging. The generation of transgenic animals is an important experimental tool and allows use of GFP fusion proteins to study the expression of genes of interest or generation of epitope tagged versions of specific genes. Transgenes are often generated by placing a promoter upstream of a reporter gene or cDNA. This often produces a representative expression pattern, but important exceptions have been observed. To better capture the genuine expression pattern and timing, several investigators have modified large pieces of DNA carried by BACs or fosmids for use in the construction of transgenic animals via recombineering. However, these techniques are not in widespread use despite the advantages when compared to traditional approaches. Additionally, some groups have encountered problems with employing these techniques. Hence, we sought identify ways to improve the simplicity and reliability of the procedure. Results We describe here several important modifications we have made to existing protocols to make the procedure simpler and more robust. Among these are the use of galK gene as a selection marker for both the positive and negative selection steps in recombineering, the use of R6K based plasmids which eliminate the need for extensive PCR product purification, a means to integrate the unc-119 marker on to the fosmid backbone, and placement of homology arms to commonly used GFP and TAP fusion genes flanking the galK cassette which reduces the cost of oligos by 50%. Conclusion We have made several significant changes that allow the production of C. elegans transgenes from a commercially available fosmid library in a robust and streamlined manner. These changes make the technique more attractive especially to small academic labs unfamiliar with recombineering.

  14. Transgenic rats with green, red, and blue fluorescence: powerful tools for bioimaging, cell trafficking, and differentiation

    Science.gov (United States)

    Murakami, Takashi; Kobayashi, Eiji

    2005-04-01

    The rat represents a perfect animal for broadening medical experiments, because its physiology has been well understood in the history of experimental animals. In addition, its larger body size takes enough advantage for surgical manipulation, compared to the mouse. Many rat models mimicking human diseases, therefore, have been used in a variety of biomedical studies including physiology, pharmacology, transplantation, and immunology. In an effort to create the specifically designed rats for biomedical research and regenerative medicine, we have developed the engineered rat system on the basis of transgenic technology and succeeded in establishing various transgenic rat strains. The transgenic rats with green fluorescent protein (GFP) were generated in the two different strains (Wistar and Lewis), in which GFP is driven under the chicken beta-actin promoter and cytomegalovirus enhancer (CAG promoter). Their GFP expression levels were different in each organ, but the Lewis line expressed GFP strongly and ubiquitously in most of the organs compared with that of Wistar. For red fluorescence, DsRed2 was transduced to the Wistar rats: one line specifically expresses DsRed2 in the liver under the mouse albumin promoter, another is designed for the Cre/LoxP system as the double reporter rat (the initial DsRed2 expression turns on GFP in the presence of Cre recombinase). LacZ-transgenic rats represent blue color, and LacZ is driven the CAG (DA) or ROSA26 promoter (Lewis). Our unique transgenic rats" system highlights the powerful performance for the elucidation of many cellular processes in regenerative medicine, leading to innovative medical treatments.

  15. Exogenous enzymes upgrade transgenesis and genetic engineering of farm animals.

    Science.gov (United States)

    Bosch, Pablo; Forcato, Diego O; Alustiza, Fabrisio E; Alessio, Ana P; Fili, Alejandro E; Olmos Nicotra, María F; Liaudat, Ana C; Rodríguez, Nancy; Talluri, Thirumala R; Kues, Wilfried A

    2015-05-01

    Transgenic farm animals are attractive alternative mammalian models to rodents for the study of developmental, genetic, reproductive and disease-related biological questions, as well for the production of recombinant proteins, or the assessment of xenotransplants for human patients. Until recently, the ability to generate transgenic farm animals relied on methods of passive transgenesis. In recent years, significant improvements have been made to introduce and apply active techniques of transgenesis and genetic engineering in these species. These new approaches dramatically enhance the ease and speed with which livestock species can be genetically modified, and allow to performing precise genetic modifications. This paper provides a synopsis of enzyme-mediated genetic engineering in livestock species covering the early attempts employing naturally occurring DNA-modifying proteins to recent approaches working with tailored enzymatic systems.

  16. Antifungal activity of a virally encoded gene in transgenic wheat.

    Science.gov (United States)

    Clausen, M; Kräuter, R; Schachermayr, G; Potrykus, I; Sautter, C

    2000-04-01

    The cDNA encoding the antifungal protein KP4 from Ustilago maydis-infecting virus was inserted behind the ubiquitin promoter of maize and genetically transferred to wheat varieties particularly susceptible to stinking smut (Tilletia tritici) disease. The transgene was integrated and inherited over several generations. Of seven transgenic lines, three showed antifungal activity against U. maydis. The antifungal activity correlated with the presence of the KP4 transgene. KP4-transgenic, soil-grown wheat plants exhibit increased endogenous resistance against stinking smut.

  17. Single-copy insertion of transgenes in Caenorhabditis elegans

    DEFF Research Database (Denmark)

    Frøkjaer-Jensen, Christian; Davis, M Wayne; Hopkins, Christopher E;

    2008-01-01

    At present, transgenes in Caenorhabditis elegans are generated by injecting DNA into the germline. The DNA assembles into a semistable extrachromosomal array composed of many copies of injected DNA. These transgenes are typically overexpressed in somatic cells and silenced in the germline. We have...

  18. Development and application of transgenic technologies in cassava

    NARCIS (Netherlands)

    Taylor, N.; Chavarriaga, P.; Raemakers, C.J.J.M.; Sititunga, D.; Zhang, P.

    2004-01-01

    The capacity to integrate transgenes into the tropical root crop cassava (Manihot esculenta Crantz) is now established and being utilized to generate plants expressing traits of agronomic interest. The tissue culture and gene transfer systems currently employed to produce these transgenic cassava

  19. Development and application of transgenic technologies in cassava

    NARCIS (Netherlands)

    Taylor, N.; Chavarriaga, P.; Raemakers, C.J.J.M.; Sititunga, D.; Zhang, P.

    2004-01-01

    The capacity to integrate transgenes into the tropical root crop cassava (Manihot esculenta Crantz) is now established and being utilized to generate plants expressing traits of agronomic interest. The tissue culture and gene transfer systems currently employed to produce these transgenic cassava ha

  20. Genetic Stability Analysis of the Fifth Generation of Transgenic Soybeans Expressing γ-linolenic Acid%T5代γ-亚麻酸转基因大豆的遗传稳定性分析

    Institute of Scientific and Technical Information of China (English)

    陈晟; 郭丽琼; 宋景深; 林俊芳

    2012-01-01

    利用PCR检测和标记基因抗性检测对表达γ-亚麻酸转基因大豆第5代植株的外源基因遗传稳定性及目的基因和抗性基因的分离情况进行了分析.结果表明:在20个转基因大豆株系中,有2个转基因大豆株系(TS824和TS825)只含有Δ6-fad基因而没有bar基因;7个转基因大豆株系(TS81、TS83、TS85、TS87、TS811、TS814、TS818)只含有bar基因而没有Δ6-fad基因;9个转基因大豆株系(TS88、TS89、TS810、TS813、TS815、TS816、TS817、TS819和TS820)既有Δ6 -fad基因也有bar基因;2个转基因大豆株系(TS82和TS86)既没有Δ6-fad基因也没有bar基因.因此,Δ6-fad基因在部分株系中获得了稳定的遗传,同时有2个株系目的基因和抗性基因在后代中分离.%The genetic stability of foreign gene and separation of Δ6-fad and bar gene were observed in the 5th generation of transgenic soybeans expressing -γ-linolenic acid. By PCR amplification and resistance testing of selective marker gene, the ge nome of transgenic soybeans TS824 and TS825 were integrated Δ6-fad gene(marker free) ;TS81 ,TS83,TS85,TS87,TS814, TS816 and TS818 were integrated bar gene;TS88 ,TS89 ,TS810,TS813 ,TS815 ,TS816.TS817 ,TS819 and TS820 were integrat ed both Δ6-fad gene and bar gene;neither Δ6-fad gene nor bar gene was detected in TS82 and TS86. The results showed that Δ6-fad gene was stably maintained in the genome of part of transgenic soybeans,and Δ6-fad and bar gene of transgenic soy beans TS824 and TS825 were separated.

  1. Translational research challenges: finding the right animal models.

    Science.gov (United States)

    Prabhakar, Sharma

    2012-12-01

    Translation of scientific discoveries into meaningful human applications, particularly novel therapies of human diseases, requires development of suitable animal models. Experimental approaches to test new drugs in preclinical phases often necessitated animal models that not only replicate human disease in etiopathogenesis and pathobiology but also biomarkers development and toxicity prediction. Whereas the transgenic and knockout techniques have revolutionized manipulation of rodents and other species to get greater insights into human disease pathogenesis, we are far from generating ideal animal models of most human disease states. The challenges in using the currently available animal models for translational research, particularly for developing potentially new drugs for human disease, coupled with the difficulties in toxicity prediction have led some researchers to develop a scoring system for translatability. These aspects and the challenges in selecting an animal model among those that are available to study human disease pathobiology and drug development are the topics covered in this detailed review.

  2. Transgenic expression of human INS gene in Ins1/Ins2 double knockout mice leads to insulin underproduction and diabetes in some male mice.

    Science.gov (United States)

    Karaca, Melis; Durel, Béatrice; Languille, Laëtitia; Lamotte, Luciane; Tourrel-Cuzin, Cécile; Leroux, Loïc; Abou Sleymane, Gretta; Saint-Just, Susan; Bucchini, Danielle; Ktorza, Alain; Joshi, Rajiv L

    2007-01-01

    We have generated transgenic mouse lines expressing exclusively a human INS transgene on an Ins1/Ins2 double knockout (mIKO) background. The transgene expression was driven by either a 4000 bp or a 353 bp promoter. These transgenic lines, designated mIKO:INS4000 and mIKO:INS353, were viable and fertile. Determination of the amounts of insulin transcripts and total pancreatic insulin content revealed relative insulin underproduction in both lines, from birth to adulthood. Total pancreatic insulin stores in mIKO:INS4000 and mIKO:INS353 mice represented only about 50% and 27%, respectively, as compared to wild-type mice. Morphometric analysis of pancreas did not show any compensatory beta-cell hyperplasia. The majority of animals in both lines remained normoglycemic throughout their lives. Nevertheless, glucose tolerance tests revealed glucose intolerance in nearly half of mIKO:INS4000 male mice, likely due to impaired insulin secretion detected in those animals. In addition, a small fraction (2-4%) of male mice in both lines spontaneously developed diabetes with very distinct pathophysiological features. Diabetes was never seen in female animals. The diabetes developed by mIKO:INS353 mice was rapidly lethal, accompanied by a dramatic depletion of pancreatic insulin stores whereas the mIKO:INS4000 diabetic animals could live for several months. This suggests a possible link between the structure of the human INS gene promoter and the type of diabetes developed in these lines.

  3. Research advances on transgenic plant vaccines.

    Science.gov (United States)

    Han, Mei; Su, Tao; Zu, Yuan-Gang; An, Zhi-Gang

    2006-04-01

    In recent years, with the development of genetics molecular biology and plant biotechnology, the vaccination (e.g. genetic engineering subunit vaccine, living vector vaccine, nucleic acid vaccine) programs are taking on a prosperous evolvement. In particular, the technology of the use of transgenic plants to produce human or animal therapeutic vaccines receives increasing attention. Expressing vaccine candidates in vegetables and fruits open up a new avenue for producing oral/edible vaccines. Transgenic plant vaccine disquisitions exhibit a tempting latent exploiting foreground. There are a lot of advantages for transgenic plant vaccines, such as low cost, easiness of storage, and convenient immune-inoculation. Some productions converged in edible tissues, so they can be consumed directly without isolation and purification. Up to now, many transgenic plant vaccine productions have been investigated and developed. In this review, recent advances on plant-derived recombinant protein expression systems, infectious targets, and delivery systems are presented. Some issues of high concern such as biosafety and public health are also discussed. Special attention is given to the prospects and limitations on transgenic plant vaccines.

  4. Production of pigs expressing a transgene under the control of a tetracycline-inducible system.

    Directory of Open Access Journals (Sweden)

    Yong-Xun Jin

    Full Text Available Pigs are anatomically and physiologically closer to humans than other laboratory animals. Transgenic (TG pigs are widely used as models of human diseases. The aim of this study was to produce pigs expressing a tetracycline (Tet-inducible transgene. The Tet-on system was first tested in infected donor cells. Porcine fetal fibroblasts were infected with a universal doxycycline-inducible vector containing the target gene enhanced green fluorescent protein (eGFP. At 1 day after treatment with 1 µg/ml doxycycline, the fluorescence intensity of these cells was increased. Somatic cell nuclear transfer (SCNT was then performed using these donor cells. The Tet-on system was then tested in the generated porcine SCNT-TG embryos. Of 4,951 porcine SCNT-TG embryos generated, 850 were cultured in the presence of 1 µg/ml doxycycline in vitro. All of these embryos expressed eGFP and 15 embryos developed to blastocyst stage. The remaining 4,101 embryos were transferred to thirty three surrogate pigs from which thirty eight cloned TG piglets were obtained. PCR analysis showed that the transgene was inserted into the genome of each of these piglets. Two TG fibroblast cell lines were established from these TG piglets, and these cells were used as donor cells for re-cloning. The re-cloned SCNT embryos expressed the eGFP transgene under the control of doxycycline. These data show that the expression of transgenes in cloned TG pigs can be regulated by the Tet-on/off systems.

  5. The development and application of a cold atmospheric plasma generator for treatment of skin and soft-tissue injuries in animals

    Science.gov (United States)

    Emelyanov, O. A.; Petrova, N. O.; Smirnova, N. V.; Shemet, M. V.

    2017-08-01

    We describe a device for obtaining cold plasma in air at atmospheric pressure using a system of positive high-voltage pin electrodes, which is intended for the treatment of skin and soft-tissue injuries in animals. Plasma is generated due to the development of periodic pulsed discharge of nanosecond duration at current pulse amplitudes 10-20 mA, characteristic frequencies 10-20 kHz, and applied voltages within 8-10 kV. The high efficacy of the proposed device and method is confirmed by the good clinical results of treating large domestic animals with traumatic injuries.

  6. Weeding with transgenes.

    Science.gov (United States)

    Duke, Stephen O

    2003-05-01

    Transgenes promise to reduce insecticide and fungicide use but relatively little has been done to significantly reduce herbicide use through genetic engineering. Recently, three strategies for transgene utilization have been developed that have the potential to change this. These are the improvement of weed-specific biocontrol agents, enhancement of crop competition or allelopathic traits, and production of cover crops that will self-destruct near the time of planting. Failsafe risk mitigation technologies are needed for most of these strategies.

  7. Development of cystic glandular hyperplasia of the endometrium in Mullerian inhibitory substance type II receptor-pituitary tumor transforming gene transgenic mice.

    Science.gov (United States)

    El-Naggar, Shahenda M; Malik, Mohammad T; Martin, Alvin; Moore, Joseph P; Proctor, Mary; Hamid, Tariq; Kakar, Sham S

    2007-07-01

    The pituitary tumor transforming gene (PTTG)/securin is an oncogene that is involved in cell cycle regulation and sister chromatid separation. PTTG is highly expressed in various tumors including ovarian tumors, suggesting that PTTG may play a role in ovarian tumorigenesis. Overexpression of PTTG resulted in induction of cellular transformation in vitro and tumor formation in nude mice. To ascertain PTTG function in ovarian tumorigenesis, we generated a transgenic mouse model of PTTG by cloning PTTG cDNA downstream of Mullerian inhibitory substance type II receptor gene promoter (MISIIR) in order to target the ovarian surface epithelium. By screening of transgenic animals, we identified five founders (four males and one female). Using the four male founders, we developed four transgenic lines. PTTG expression was increased in ovarian surface epithelium, ovarian granulosa cells, as well as in the pituitary gland. Transgenic females did not develop any visible ovarian tumors at 8-10 months of age; however, there was an overall increase in the corpus luteum mass in transgenic ovary, suggesting increased luteinization. These changes were associated with an increase in serum LH and testosterone levels. In addition, there was a generalized hypertrophy of the myometrium of MISIIR-PTTG transgenic uteri with cystic glandular and hyperplasia of the endometrium. Based on these results, we conclude that the overexpression of PTTG may be required to initiate precancerous conditions but is not sufficient to induce ovarian tumorigenesis and may require another partner to initiate cellular transformation.

  8. Assessment of Fecundity and Germ Line Transmission in Two Transgenic Pig Lines Produced by Sleeping Beauty Transposition

    Directory of Open Access Journals (Sweden)

    Wilfried A. Kues

    2012-10-01

    Full Text Available Recently, we described a simplified injection method for producing transgenic pigs using a non-autonomous Sleeping Beauty transposon system. The founder animals showed ubiquitous expression of the Venus fluorophore in almost all cell types. To assess, whether expression of the reporter fluorophore affects animal welfare or fecundity, we analyzed reproductive parameters of two founder boars, germ line transmission, and organ and cell specific transgene expression in animals of the F1 and F2 generation. Molecular analysis of ejaculated sperm cells suggested three monomeric integrations of the Venus transposon in both founders. To test germ line transmission of the three monomeric transposon integrations, wild-type sows were artificially inseminated. The offspring were nursed to sexual maturity and hemizygous lines were established. A clear segregation of the monomeric transposons following the Mendelian rules was observed in the F1 and F2 offspring. Apparently, almost all somatic cells, as well as oocytes and spermatozoa, expressed the Venus fluorophore at cell-type specific levels. No detrimental effects of Venus expression on animal health or fecundity were found. Importantly, all hemizygous lines expressed the fluorophore in comparable levels, and no case of transgene silencing or variegated expression was found after germ line transmission, suggesting that the insertions occurred at transcriptionally permissive loci. The results show that Sleeping Beauty transposase-catalyzed transposition is a promising approach for stable genetic modification of the pig genome.

  9. Assessment of fecundity and germ line transmission in two transgenic pig lines produced by sleeping beauty transposition.

    Science.gov (United States)

    Garrels, Wiebke; Holler, Stephanie; Cleve, Nicole; Niemann, Heiner; Ivics, Zoltan; Kues, Wilfried A

    2012-10-12

    Recently, we described a simplified injection method for producing transgenic pigs using a non-autonomous Sleeping Beauty transposon system. The founder animals showed ubiquitous expression of the Venus fluorophore in almost all cell types. To assess, whether expression of the reporter fluorophore affects animal welfare or fecundity, we analyzed reproductive parameters of two founder boars, germ line transmission, and organ and cell specific transgene expression in animals of the F1 and F2 generation. Molecular analysis of ejaculated sperm cells suggested three monomeric integrations of the Venus transposon in both founders. To test germ line transmission of the three monomeric transposon integrations, wild-type sows were artificially inseminated. The offspring were nursed to sexual maturity and hemizygous lines were established. A clear segregation of the monomeric transposons following the Mendelian rules was observed in the F1 and F2 offspring. Apparently, almost all somatic cells, as well as oocytes and spermatozoa, expressed the Venus fluorophore at cell-type specific levels. No detrimental effects of Venus expression on animal health or fecundity were found. Importantly, all hemizygous lines expressed the fluorophore in comparable levels, and no case of transgene silencing or variegated expression was found after germ line transmission, suggesting that the insertions occurred at transcriptionally permissive loci. The results show that Sleeping Beauty transposase-catalyzed transposition is a promising approach for stable genetic modification of the pig genome.

  10. Transgenic soybean plants overexpressing O-acetylserine sulfhydrylase accumulate enhanced levels of cysteine and Bowman-Birk protease inhibitor in seeds.

    Science.gov (United States)

    Kim, Won-Seok; Chronis, Demosthenis; Juergens, Matthew; Schroeder, Amy C; Hyun, Seung Won; Jez, Joseph M; Krishnan, Hari B

    2012-01-01

    Soybeans provide an excellent source of protein in animal feed. Soybean protein quality can be enhanced by increasing the concentration of sulfur-containing amino acids. Previous attempts to increase the concentration of sulfur-containing amino acids through the expression of heterologous proteins have met with limited success. Here, we report a successful strategy to increase the cysteine content of soybean seed through the overexpression of a key sulfur assimilatory enzyme. We have generated several transgenic soybean plants that overexpress a cytosolic isoform of O-acetylserine sulfhydrylase (OASS). These transgenic soybean plants exhibit a four- to tenfold increase in OASS activity when compared with non-transformed wild-type. The OASS activity in the transgenic soybeans was significantly higher at all the stages of seed development. Unlike the non-transformed soybean plants, there was no marked decrease in the OASS activity even at later stages of seed development. Overexpression of cytosolic OASS resulted in a 58-74% increase in protein-bound cysteine levels compared with non-transformed wild-type soybean seeds. A 22-32% increase in the free cysteine levels was also observed in transgenic soybeans overexpressing OASS. Furthermore, these transgenic soybean plants showed a marked increase in the accumulation of Bowman-Birk protease inhibitor, a cysteine-rich protein. The overall increase in soybean total cysteine content (both free and protein-bound) satisfies the recommended levels required for the optimal growth of monogastric animals.

  11. Promoter interference mediated by the U3 region in early-generation HIV-1-derived lentivirus vectors can influence detection of transgene expression in a cell-type and species-specific manner.

    Science.gov (United States)

    Ginn, Samantha L; Fleming, Jane; Rowe, Peter B; Alexander, Ian E

    2003-08-10

    In a previous study using an early-generation VSV-G-pseudotyped lentivirus vector encoding enhanced green fluorescent protein (EGFP) under the transcriptional control of a human cytomegalovirus (CMV) immediate-early promoter, we examined transduction efficiency in dissociated dorsal root ganglia (DRG) cultures. In cultures of murine origin, transgene expression was observed solely in the sensory neurons with the stromal cell population failing to show evidence of transduction. In contrast, efficient and sustained transduction of both sensory neurons and the stromal cell population was observed in cultures of human origin. Given the widespread use of murine models in preclinical gene therapy studies, in the current study we investigated the basis of this apparent neuron specificity of lentivirus-mediated transduction in murine DRG cultures. The interspecies differences persisted at high multiplicities of infection, and irrespective of whether lentiviral vector stocks were packaged in the presence or absence of human immunodeficiency virus type 1 (HIV-1) accessory proteins. Cell-type specificity of CMV promoter expression, tropism of the VSV-G envelope, and blocks to molecular transduction were also precluded as possible mechanisms, thereby implicating transcriptional repression of the internal heterologous promoter. This promoter interference effect was found to be mediated by cis-acting sequences upstream of the core promoter elements located in the U3 region of the proviral long terminal repeats (LTRs). Deletion of this region, as in late-generation self-inactivating (SIN) lentivirus vectors, relieves this effect. This provides a basis for reevaluating data produced using early-generation U3-bearing lentivirus vectors and for reconciling these with results obtained using more contemporary SIN lentivirus vectors carrying a U3 deletion.

  12. Animal pharming, two decades on.

    Science.gov (United States)

    Kind, Alexander; Schnieke, Angelika

    2008-12-01

    Since its inception 20 years ago, the animal pharming industry has promoted transgenic animals as a cost-effective method of biopharmaceutical production. However, it took until 2006 for the first therapeutic product to gain regulatory approval. This was an important milestone, but scepticism still abounds. Can pharming regain investor confidence, and will society accept transgenic livestock as a production method? There is some cause for optimism, biopharmaceuticals are a large, expanding market and animal pharming has already made considerable strides. A novel production platform has been established, groundbreaking technologies developed, a necessary regulatory framework put in place. Nevertheless, despite cost advantages, pharming has become a niche production method and its long term success may depend on products unique to transgenic animals.

  13. Bacterial xylanase expression in mammalian cells and transgenic mice.

    Science.gov (United States)

    Fontes, C M; Ali, S; Gilbert, H J; Hazlewood, G P; Hirst, B H; Hall, J

    1999-06-11

    The energy which simple-stomached livestock can derive from dietary plant material is limited by the lack of plant polysaccharide degrading enzymes in their gastro-intestinal (GI) tract and the inefficient microbial fermentation of such material in their hind-gut. In poultry the non-starch polysaccharides found in cereal grains can also impair normal digestive function as they form viscous gels in the GI tract inhibiting the breakdown and absorption of nutrients. The nutrition of such livestock could, therefore, be improved by the introduction of enzymes able to degrade plant polysaccharides in the small intestine. We describe the expression of a xylanase, XYLY', from the bacterium Clostridium thermocellum in mammalian cells and the exocrine pancreas of transgenic mice. The enzyme is synthesised, secreted and functionally active in the eukaryote system. This work demonstrates the feasibility of generating animals with the endogenous capacity to depolymerise the xylan component of hemi-cellulose.

  14. Lysostaphin expression in mammary glands confers protection against staphylococcal infection in transgenic mice.

    Science.gov (United States)

    Kerr, D E; Plaut, K; Bramley, A J; Williamson, C M; Lax, A J; Moore, K; Wells, K D; Wall, R J

    2001-01-01

    Infection of the mammary gland, in addition to causing animal distress, is a major economic burden of the dairy industry. Staphylococcus aureus is the major contagious mastitis pathogen, accounting for approximately 15-30% of infections, and has proved difficult to control using standard management practices. As a first step toward enhancing mastitis resistance of dairy animals, we report the generation of transgenic mice that secrete a potent anti-staphylococcal protein into milk. The protein, lysostaphin, is a peptidoglycan hydrolase normally produced by Staphylococcus simulans. When the native form is secreted by transfected eukaryotic cells it becomes glycosylated and inactive. However, removal of two glycosylation motifs through engineering asparagine to glutamine codon substitutions enables secretion of Gln(125,232)-lysostaphin, a bioactive variant. Three lines of transgenic mice, in which the 5'-flanking region of the ovine beta-lactoglobulin gene directed the secretion of Gln(125,232)-lysostaphin into milk, exhibit substantial resistance to an intramammary challenge of 104 colony-forming units (c.f.u.) of S. aureus, with the highest expressing line being completely resistant. Milk protein content and profiles of transgenic and nontransgenic mice are similar. These results clearly demonstrate the potential of genetic engineering to combat the most prevalent disease of dairy cattle.

  15. Transgenic rabbits as therapeutic protein bioreactors and human disease models.

    Science.gov (United States)

    Fan, Jianglin; Watanabe, Teruo

    2003-09-01

    Genetically modified laboratory animals provide a powerful approach for studying gene expression and regulation and allow one to directly examine structure-function and cause-and-effect relationships in pathophysiological processes. Today, transgenic mice are available as a research tool in almost every research institution. On the other hand, the development of a relatively large mammalian transgenic model, transgenic rabbits, has provided unprecedented opportunities for investigators to study the mechanisms of human diseases and has also provided an alternative way to produce therapeutic proteins to treat human diseases. Transgenic rabbits expressing human genes have been used as a model for cardiovascular disease, AIDS, and cancer research. The recombinant proteins can be produced from the milk of transgenic rabbits not only at lower cost but also on a relatively large scale. One of the most promising and attractive recombinant proteins derived from transgenic rabbit milk, human alpha-glucosidase, has been successfully used to treat the patients who are genetically deficient in this enzyme. Although the pronuclear microinjection is still the major and most popular method for the creation of transgenic rabbits, recent progress in gene targeting and animal cloning has opened new avenues that should make it possible to produce transgenic rabbits by somatic cell nuclear transfer in the future. Based on a computer-assisted search of the studies of transgenic rabbits published in the English literature here, we introduce to the reader the achievements made thus far with transgenic rabbits, with emphasis on the application of these rabbits as human disease models and live bioreactors for producing human therapeutic proteins and on the recent progress in cloned rabbits.

  16. Position-independent expression of transgenes in zebrafish.

    Science.gov (United States)

    Caldovic, L; Agalliu, D; Hackett, P B

    1999-10-01

    The variability in expression patterns of transgenes, caused by the influence of neighboring chromatin, is called 'position effect'. Border elements are DNA sequences, which have the ability to alleviate position effects. The abilities of two types of border elements, scs/scs' from the D. melanogaster 87A7 heat shock locus and the A-element from the chicken lysozyme gene, to protect transgenes from position effects were quantified in developing zebrafish embryos. The transgenic construct used was FV3CAT, which consists of the carp beta-actin transcriptional regulatory region, the chloramphenicol acetyltransferase (CAT) gene and the 3'-untranslated region from the Chinook salmon growth hormone gene. FV3CAT constructs flanked by either scs/scs'-elements or A-elements were introduced into zebrafish chromosomes and the spatial and temporal expression patterns of the transgenes were quantified in multiple generations of transgenic zebrafish. Levels of transgene expression were uniform in the pre-differentiated and fully differentiated populations of cells present during embryonic development. Levels of transgene expression were proportional to the numbers of integrated transgenes. Expression of transgenes per cell varied less than two-fold in different transgenic lines. Both types of border elements were able to prevent the influences of neighboring chromatin on transgene expression through three generations of fish. The results are consistent with the ability of border elements to function with equal efficiencies in the many cell types found in vertebrates. Thus, inclusion of border elements in genetic constructs can provide reliable and reproducible levels of gene expression in multiple lines of fish.

  17. Production of transgenic calves by somatic cell nuclear transfer

    Institute of Scientific and Technical Information of China (English)

    GONG Guochun; WAN Rong; HUANG Yinghua; LI Ning; DAI Yunping; FAN Baoliang; ZHU Huabing; WANG Lili; WANG Haiping; TANG Bo; LIU Ying; LI Rong

    2004-01-01

    Bovine fetal oviduct epithelial cells were transfected with constructed double marker selective vector (pCE-EGFP-IRES-Neo-dNdB) containing the enhanced green fluorescent protein (EGFP) and neomycin-resistant (Neor) genes by electroporation, and a transgenic cell line was obtained. Somatic cell nuclear transfer (SCNT) was carried out using the transgenic cells as nuclei donor. A total of 424 SCNT embryos were reconstructed and 208 (49.1%) of them developed to blastocyst stage. 17 blastocysts on D 7 after reconstruction were transferred to 17 surrogate calves, and 5 (29.4%) recipients were found to be pregnant. Three of them maintained to term and delivered three cloned calves. PCR and Southern blot analysis confirmed the integration of transgene in all of the three cloned calves. In addition, expression of EGFP was detected in biopsy isolated from the transgenic cloned calves and fibroblasts derived from the biopsy. Our results suggest that transgenic calves could be efficiently produced by SCNT using transgenic cells as nuclei donor. Furthermore, all cloned animals could be ensured to be transgenic by efficiently pre-screening transgenic cells and SCNT embryos using the constructed double marker selective vector.

  18. Apoptosis of transgenic cloned and recloned bovine blastocysts

    Institute of Scientific and Technical Information of China (English)

    Guojie Sun; Rong Li; Yunping Dai; Haiping Wang; Lili Wang; Ying Liu; Fangrong Ding; Hengxi Wei; Ning Li

    2009-01-01

    Apoptosis plays an important role in preimplantation embryonic development. Investigating mechanisms of apoptosis can provide useful information for obtaining high-quality embryos and help to improve cloning efficiency. Here, we investigated the incidence of blastomere apoptosis in transgenic blastocysts generated by somatic cell nuclear transfer (SCNT) and recloning using a terminal deoxy-nucleotidyl transferase-mediated d-UTP nick end-labeling (TUNEL) assay. Transgenic recloned embryos were the second generation SCNT embryos derived from the somatic cells of a transgenic SCNT calf. The blastocyst rate of transgenic SCNT embryos was lower than that of nontransgenic SCNT embryos. The incidence of apoptosis in transgenic SCNT embryos was higher than that of nontrans-genie SCNT embryos. The blastocyst rate and the incidence of apoptosis in transgenic recloned embryos were similar to nontransgenic SCNT embryos. The process of donor cell transfection and drug selection may decrease the developmental capacity of transgenic SCNT embryos. Serial cloning did not influence the developmental capacity of transgenic recloned embryos.

  19. Manipulation of the repertoire of digestive enzymes secreted into the gastrointestinal tract of transgenic mice.

    Science.gov (United States)

    Hall, J; Ali, S; Surani, M A; Hazlewood, G P; Clark, A J; Simons, J P; Hirst, B H; Gilbert, H J

    1993-03-01

    In non-ruminant livestock the energy which can be derived from dietary cellulose and xylan is limited by the inefficient microbial fermentation of these polymers in the hind-gut. Furthermore, in poultry, cereal-derived plant structural polysaccharides impair normal digestive function through the formation of gel-like structures, which trap nutrients rendering them unavailable to the animal. The nutrition of non-ruminant livestock could be significantly improved by the depolymerization of plant structural polysaccharides, through the introduction of cellulase activity into the small intestines of these animals. Here we describe the expression of Clostridium thermocellum endoglucanase E in the exocrine pancreas of transgenic mice. A non-glycosylated active enzyme is secreted into the small intestines, and is resistant to proteolytic inactivation, demonstrating the feasibility of generating non-ruminant animals with the endogenous capacity to depolymerize plant structural polysaccharides in the small intestines.

  20. Expression of recombinant human lysozyme in transgenic chicken promotes the growth of Bifidobacterium in the intestine and improves postnatal growth of chicken

    OpenAIRE

    Wang, Hai; Wu, Hongping; Wang, Kejun; Cao, Zhichen; Yu, Kun; Lian, Ling; Lian, Zhengxing

    2016-01-01

    Lysozyme is one kind of antimicrobial proteins and often used as feed additive which can defend against pathogenic bacteria and enhance immune function of animals. In this study, we have injected the lentiviral vector expressing recombinant human lysozyme (rhLZ) gene into the blastoderm of chicken embryo to investigate the effect of recombinant human lysozyme on postnatal intestinal microbiota distribution and growth performance of chicken. Successfully, we generated 194 transgenic chickens i...

  1. Transgenic studies on homeobox genes in nervous system development: spina bifida in Isl1 transgenic mice.

    Science.gov (United States)

    Kappen, Claudia; Yaworsky, Paul J; Muller, Yunhua L; Salbaum, J Michael

    2013-04-01

    To develop in vivo assays for homeobox gene function in neural development, we generated transgenic mice in which the expression of a homeobox gene is altered only within the nervous system, in neurons or neuronal precursor cells. Transgenic expression of Hoxc8 did not result in gross abnormalities, while a Hoxd4 transgene caused death shortly after birth. In neural progenitor cells, the motorneuron-specific homeodomain transcription factor Isl1 induced early developmental defects, including absence of anterior neural structures, profound defects in the neuroepithelium and defective neural tube closure. A fraction of Isl1 transgenic mice exhibited spina bifida. Isl1 transgene expression was also associated with decreased proliferation and increased Pbx1 expression in the ventral neural tube. Our results suggest a function for some homeobox genes in development of the nervous system, and that cell-type- and region-specific transgenic models will be useful to identify the cellular and molecular targets of homeobox transcription factors in nervous system development.

  2. T cell receptor (TCR-transgenic CD8 lymphocytes rendered insensitive to transforming growth factor beta (TGFβ signaling mediate superior tumor regression in an animal model of adoptive cell therapy

    Directory of Open Access Journals (Sweden)

    Quatromoni Jon G

    2012-06-01

    Full Text Available Abstract Tumor antigen-reactive T cells must enter into an immunosuppressive tumor microenvironment, continue to produce cytokine and deliver apoptotic death signals to affect tumor regression. Many tumors produce transforming growth factor beta (TGFβ, which inhibits T cell activation, proliferation and cytotoxicity. In a murine model of adoptive cell therapy, we demonstrate that transgenic Pmel-1 CD8 T cells, rendered insensitive to TGFβ by transduction with a TGFβ dominant negative receptor II (DN, were more effective in mediating regression of established B16 melanoma. Smaller numbers of DN Pmel-1 T cells effectively mediated tumor regression and retained the ability to produce interferon-γ in the tumor microenvironment. These results support efforts to incorporate this DN receptor in clinical trials of adoptive cell therapy for cancer.

  3. High-throughput expression of animal venom toxins in Escherichia coli to generate a large library of oxidized disulphide-reticulated peptides for drug discovery.

    Science.gov (United States)

    Turchetto, Jeremy; Sequeira, Ana Filipa; Ramond, Laurie; Peysson, Fanny; Brás, Joana L A; Saez, Natalie J; Duhoo, Yoan; Blémont, Marilyne; Guerreiro, Catarina I P D; Quinton, Loic; De Pauw, Edwin; Gilles, Nicolas; Darbon, Hervé; Fontes, Carlos M G A; Vincentelli, Renaud

    2017-01-17

    Animal venoms are complex molecular cocktails containing a wide range of biologically active disulphide-reticulated peptides that target, with high selectivity and efficacy, a variety of membrane receptors. Disulphide-reticulated peptides have evolved to display improved specificity, low immunogenicity and to show much higher resistance to degradation than linear peptides. These properties make venom peptides attractive candidates for drug development. However, recombinant expression of reticulated peptides containing disulphide bonds is challenging, especially when associated with the production of large libraries of bioactive molecules for drug screening. To date, as an alternative to artificial synthetic chemical libraries, no comprehensive recombinant libraries of natural venom peptides are accessible for high-throughput screening to identify novel therapeutics. In the accompanying paper an efficient system for the expression and purification of oxidized disulphide-reticulated venom peptides in Escherichia coli is described. Here we report the development of a high-throughput automated platform, that could be adapted to the production of other families, to generate the largest ever library of recombinant venom peptides. The peptides were produced in the periplasm of E. coli using redox-active DsbC as a fusion tag, thus allowing the efficient formation of correctly folded disulphide bridges. TEV protease was used to remove fusion tags and recover the animal venom peptides in the native state. Globally, within nine months, out of a total of 4992 synthetic genes encoding a representative diversity of venom peptides, a library containing 2736 recombinant disulphide-reticulated peptides was generated. The data revealed that the animal venom peptides produced in the bacterial host were natively folded and, thus, are putatively biologically active. Overall this study reveals that high-throughput expression of animal venom peptides in E. coli can generate large

  4. Intra-bone marrow-bone marrow transplantation slows disease progression and prolongs survival in G93A mutant SOD1 transgenic mice, an animal model mouse for amyotrophic lateral sclerosis.

    Science.gov (United States)

    Ohnishi, Shizuo; Ito, Hidefumi; Suzuki, Yasuhiro; Adachi, Yasushi; Wate, Reika; Zhang, Jianhua; Nakano, Satoshi; Kusaka, Hirofumi; Ikehara, Susumu

    2009-11-03

    It has been reported that bone marrow transplantation (BMT) has clinical effects on not only hematopoietic diseases and autoimmune diseases but also solid malignant tumors and metabolic diseases. We have found that intra-bone marrow-bone marrow transplantation (IBM-BMT) is superior to conventional intravenous BMT, since IBM-BMT enables rapid recovery of donor hematopoiesis and reduces the extent of graft-versus-host disease (GVHD). In this experiment, we examined the effects of IBM-BMT on symptomatic G93A mutant SOD1 transgenic mice (mSOD1 Tg mice), a model mouse line for amyotrophic lateral sclerosis (ALS). Symptomatic mSOD1 Tg mice (12 weeks old) were irradiated with 6Gyx2 at a 4-hour interval, one day before IBM-BMT. The mice were transplanted with bone marrow cells (BMCs) from 12-wk-old eGFP-transgenic C57BL/6 mice (eGFP Tg mice) or BMCs from 12-wk-old mSOD1 Tg mice. The ALS model mice transplanted with BMCs from eGFP Tg mice showed longer survival and slower disease progression than those transplanted with BMCs from mSOD1 Tg mice or untreated mSOD1 Tg mice. There was a significantly high number of eGFP(+) cells in the anterior horn of the spinal cord of the mSOD1 Tg mice transplanted with BMCs of eGFP Tg mice, some of which expressed Iba-1, a marker of microglia, although they did not differentiate into neural cells. These results suggest that the replacement with normal hematopoietic cells improved the neural cell environment, thereby slowing the progression of the disease.

  5. The Establishment of Double-Transgenic Mice that Co-Express the appA and MxA Genes Mediated by Type A Spermatogonia In vivo

    Institute of Scientific and Technical Information of China (English)

    BAI Li-jing; JU Hui-ming; MU Yu-lian; YANG Shu-lin; REN Hong-yan; AO Hong; WANG Chu-duan; LI Kui

    2014-01-01

    Type A spermatogonial stem cells are the only immortal diploid cells in the postnatal animal that undergo self-renewal through the lifetime of an animal and transmit genes to subsequent generations. In this paper, the generation and characterization of double-transgenic mice co-expressing the Escherichia coli appA gene and human MxA gene generated via the in vivo transfection of type A spermatogonial cells were reported for the ifrst time. The dicistronic expression vector pcDNA-appA-MxA(AMP) and ExGen500 transfection reagent were injected into the testicular tissue of 7-d-old male ICR mice. The mice that underwent testis-mediated gene transfer were mated with wild-type female mice, and the integration and expression of the foreign genes in the offspring were evaluated. Transgenic mice that co-expressed appA and MxA showed a gene integration rate of 8.89%(16/180). The transgenic mice were environmentally friendly, as the amount of phosphorous remaining in the manure was reduced by as much as 11.1%by the appA gene (P<0.05);these animals also exhibited a strong anti-viral phenotype.

  6. Somatic cell nuclear transfer and transgenesis in large animals: current and future insights.

    Science.gov (United States)

    Galli, C; Lagutina, I; Perota, A; Colleoni, S; Duchi, R; Lucchini, F; Lazzari, G

    2012-06-01

    Somatic cell nuclear transfer (SCNT) was first developed in livestock for the purpose of accelerating the widespread use of superior genotypes. Although many problems still exist now after fifteen years of research owing to the limited understanding of genome reprogramming, SCNT has provided a powerful tool to make copies of selected individuals in different species, to study genome pluripotency and differentiation, opening new avenues of research in regenerative medicine and representing the main route for making transgenic livestock. Besides well-established methods to deliver transgenes, recent development in enzymatic engineering to edit the genome provides more precise and reproducible tools to target-specific genomic loci especially for producing knockout animals. The interest in generating transgenic livestock lies in the agricultural and biomedical areas and it is, in most cases, at the stage of research and development, with few exceptions that are making the way into practical applications.

  7. Production of transgenic pigs over-expressing the antiviral gene Mx1.

    Science.gov (United States)

    Yan, Quanmei; Yang, Huaqiang; Yang, Dongshan; Zhao, Bentian; Ouyang, Zhen; Liu, Zhaoming; Fan, Nana; Ouyang, Hongsheng; Gu, Weiwang; Lai, Liangxue

    2014-01-01

    The myxovirus resistance gene (Mx1) has a broad spectrum of antiviral activities. It is therefore an interesting candidate gene to improve disease resistance in farm animals. In this study, we report the use of somatic cell nuclear transfer (SCNT) to produce transgenic pigs over-expressing the Mx1 gene. These transgenic pigs express approximately 15-25 times more Mx1 mRNA than non-transgenic pigs, and the protein level of Mx1 was also markedly enhanced. We challenged fibroblast cells isolated from the ear skin of transgenic and control pigs with influenza A virus and classical swine fever virus (CFSV). Indirect immunofluorescence assay (IFA) revealed a profound decrease of influenza A proliferation in Mx1 transgenic cells. Growth kinetics showed an approximately 10-fold reduction of viral copies in the transgenic cells compared to non-transgenic controls. Additionally, we found that the Mx1 transgenic cells were more resistant to CSFV infection in comparison to non-transgenic cells. These results demonstrate that the Mx1 transgene can protect against viral infection in cells of transgenic pigs and indicate that the Mx1 transgene can be harnessed to develop disease-resistant pigs.

  8. Advancing environmental risk assessment for transgenic biofeedstock crops

    Directory of Open Access Journals (Sweden)

    Wolt Jeffrey D

    2009-11-01

    Full Text Available Abstract Transgenic modification of plants is a key enabling technology for developing sustainable biofeedstocks for biofuels production. Regulatory decisions and the wider acceptance and development of transgenic biofeedstock crops are considered from the context of science-based risk assessment. The risk assessment paradigm for transgenic biofeedstock crops is fundamentally no different from that of current generation transgenic crops, except that the focus of the assessment must consider the unique attributes of a given biofeedstock crop and its environmental release. For currently envisioned biofeedstock crops, particular emphasis in risk assessment will be given to characterization of altered metabolic profiles and their implications relative to non-target environmental effects and food safety; weediness and invasiveness when plants are modified for abiotic stress tolerance or are domesticated; and aggregate risk when plants are platforms for multi-product production. Robust risk assessments for transgenic biofeedstock crops are case-specific, initiated through problem formulation, and use tiered approaches for risk characterization.

  9. [Allergic risk of transgenic food: prevention strategies].

    Science.gov (United States)

    Moneret-Vautrin, Denise-Anne

    2002-01-01

    Numerous allergens proceed from foods. The allergic risk of transgenic foods needs to be evaluated according recommendations from the Joint Expert Committee FAO/WHO. Potential issues are the risk of cross reactivity with existing allergens, the modification of allergenicity of the transgenic protein induced by a modified metabolism in the host, the modified allergenicity of the proteins of the transgenic plant, a potential neo-allergenicity of the transgenic protein, and the risk of dissemination through pollens, inducing a respiratory sensitization then a cross food allergy. The algorithm includes three steps for evaluation: first the search for significant homology of the protein with allergens listed in allergen databanks, or the identity of a sequence of six aminoacids with known allergens, then a cross reactivity explored through the binding to IgEs from patients allergic to the source of the gene, or allergic to organisms of the same group or botanical family, and finally the extent of the pepsine resistance. The risk of immunogenicity has to be studied with appropriate animal models. A post-marketing surveillance is recommended for monitoring of adverse effects. The structure of an Allergo-Vigilance Network, the tools for efficiency and the groups at higher risk will be discussed.

  10. Generation of a recombinant chimeric Newcastle disease virus vaccine that allows serological differentiation between vaccinated and infected animals

    NARCIS (Netherlands)

    Peeters, B.P.; Leeuw, de O.S.; Verstegen, I.; Koch, G.; Gielkens, A.L.

    2001-01-01

    Using a recently developed reverse genetics system, we have generated a recombinant Newcastle disease virus (NDV) vaccine in which the gene encoding the hemagglutinin-neuraminidase (HN) has been replaced by a hybrid HN gene consisting of the cytoplasmic domain, transmembrane region, and stalk region

  11. Creating a YouTube-Like Collaborative Environment in Mathematics: Integrating Animated Geogebra Constructions and Student-Generated Screencast Videos

    Science.gov (United States)

    Lazarus, Jill; Roulet, Geoffrey

    2013-01-01

    This article discusses the integration of student-generated GeoGebra applets and Jing screencast videos to create a YouTube-like medium for sharing in mathematics. The value of combining dynamic mathematics software and screencast videos for facilitating communication and representations in a digital era is demonstrated herein. We share our…

  12. Transgenic nonhuman primates for neurodegenerative diseases

    Directory of Open Access Journals (Sweden)

    Chan Anthony WS

    2004-06-01

    Full Text Available Abstract Animal models that represent human diseases constitute an important tool in understanding the pathogenesis of the diseases, and in developing effective therapies. Neurodegenerative diseases are complex disorders involving neuropathologic and psychiatric alterations. Although transgenic and knock-in mouse models of Alzheimer's disease, (AD, Parkinson's disease (PD and Huntington's disease (HD have been created, limited representation in clinical aspects has been recognized and the rodent models lack true neurodegeneration. Chemical induction of HD and PD in nonhuman primates (NHP has been reported, however, the role of intrinsic genetic factors in the development of the diseases is indeterminable. Nonhuman primates closely parallel humans with regard to genetic, neuroanatomic, and cognitive/behavioral characteristics. Accordingly, the development of NHP models for neurodegenerative diseases holds greater promise for success in the discovery of diagnoses, treatments, and cures than approaches using other animal species. Therefore, a transgenic NHP carrying a mutant gene similar to that of patients will help to clarify our understanding of disease onset and progression. Additionally, monitoring disease onset and development in the transgenic NHP by high resolution brain imaging technology such as MRI, and behavioral and cognitive testing can all be carried out simultaneously in the NHP but not in other animal models. Moreover, because of the similarity in motor repertoire between NHPs and humans, it will also be possible to compare the neurologic syndrome observed in the NHP model to that in patients. Understanding the correlation between genetic defects and physiologic changes (e.g. oxidative damage will lead to a better understanding of disease progression and the development of patient treatments, medications and preventive approaches for high risk individuals. The impact of the transgenic NHP model in understanding the role which

  13. Transgenic Crops for Herbicide Resistance

    Science.gov (United States)

    Since their introduction in 1995, crops made resistant to the broad-spectrum herbicides glyphosate and glufosinate with transgenes are widely available and used in much of the world. As of 2008, over 80% of the transgenic crops grown world-wide have this transgenic trait. This technology has had m...

  14. The effect of mammary gland-specific transgene expression on rabbit reproductive gland structure.

    Science.gov (United States)

    Tvrdá, Eva; Massányi, Peter; Lukác, Norbert; Danko, Ján; Schlarmannova, Janka; Chrenek, Peter

    2014-01-01

    Transgenic rabbits are excellent animal models for human diseases and suitable bioreactors for the production of recombinant proteins on an experimental and commercial scale. The aim of this study was to compare the structure of the mWAP-hFVIII transgenic and non-transgenic rabbit ovarian and testicular tissue. Ovarian and testicular tissue samples were taken from transgenic and non-transgenic New Zealand White rabbits, examined by optical microscopy and analyzed morphometrically. An increase of the relative volume of primary follicles and a decrease of the relative volume of antral follicles was detected in the transgenic ovarian structure (P 0.05). In the testes a significant decrease (P transgenic testicular structure, but the relative volume of all basic structures (germinal epithelium, interstitium and lumen) was unaltered (P > 0.05). Generally, this study demonstrates a weak negative effect of mWAP-hFVIII transgenesis on rabbit gonadal structure.

  15. Composite potato plants with transgenic roots on non-transgenic shoots: a model system for studying gene silencing in roots

    DEFF Research Database (Denmark)

    Horn, Patricia; Santala, Johanna; Nielsen, Steen Lykke;

    2014-01-01

    Composite plants, with transgenic roots on a non-transgenic shoot, can be obtained by shoot explant transformation with Agrobacterium rhizogenes. The aim of this study was to generate composite potato plants (Solanum tuberosum) to be used as a model system in future studies on root...... of composite potato plants expressed significantly higher amounts of β-glucuronidase (GUS) than the roots of a GUS-transgenic potato line event. Silencing of the uidA transgene (GUS) was tested by inducing roots on the GUS-transgenic cv. Albatros event with strains of A. rhizogenes over-expressing either......-mediated silencing (co-suppression) was not functional in roots. The results suggest that composite plants offer a useful experimental system for potato research, which has gained little previous attention....

  16. Generation of human/rat xenograft animal model for the study of human donor stem cell behaviors in vivo

    Institute of Scientific and Technical Information of China (English)

    Yan Sun; Dong Xiao; Xing-Hua Pan; Ruo-Shuang Zhang; Guang-Hui Cui; Xi-Gu Chen

    2007-01-01

    AIM: To accurately and realistically elucidate human stem cell behaviors In vivo and the fundamental mechanisms controlling human stem cell fates in vivo, which is urgently required in regenerative medicine and treatments for some human diseases, a surrogate human-rat chimera model was developed.METHODS: Human-rat chimeras were achieved by in utero transplanting low-density mononuclear cells from human umbilical cord blood into the fetal rats at 9-11 d of gestation, and subsequently, a variety of methods, including flow cytometry, PCR as well as immunohistochemical assay, were used to test the human donor contribution in the recipients.RESULTS: Of 29 live-born recipients, 19 had the presence of human CD45+ cells in peripheral blood (PB) detected by flow cytometry, while PCR analysis on genomic DNA from 11 different adult tissues showed that 14 selected from flow cytometry-positive 19 animals possessed of donor-derived human cell engraftment in multiple tissues (i.e. liver, spleen, thymus, heart, kidney, blood, lung, muscle, gut and skin) examined at the time of tissue collection, as confirmed by detecting human β2-microglobulin expression using immunohistochemistry.In this xenogeneic system, the engrafted donor-derived human cells persisted in multiple tissues for at least 6 mo after birth. Moreover, transplanted human donor cells underwent site-specific differentiation into CK18-positive human cells in chimeric liver and CD45-positive human cells in chimeric spleen and thymus of recipients.CONCLUSION: Taken together, these findings suggest that we successfully developed human-rat chimeras, in which xenogeneic human cells exist up to 6 mo later. This humanized small animal model, which offers an in vivo environment more closely resembling to the situations in human, provides an invaluable and effective approach for in vivo investigating human stem cell behaviors, and further in vivo examining fundamental mechanisms controlling human stem cell fates in the future

  17. Generations.

    Science.gov (United States)

    Chambers, David W

    2005-01-01

    Groups naturally promote their strengths and prefer values and rules that give them an identity and an advantage. This shows up as generational tensions across cohorts who share common experiences, including common elders. Dramatic cultural events in America since 1925 can help create an understanding of the differing value structures of the Silents, the Boomers, Gen Xers, and the Millennials. Differences in how these generations see motivation and values, fundamental reality, relations with others, and work are presented, as are some applications of these differences to the dental profession.

  18. Efficient and Rapid Development of Transgenic Hamster Models of TSEs Using a Radical New Technology

    Science.gov (United States)

    2007-09-01

    by TOSK Inc. to introduce the human, sheep, and other prion genes into golden Syrian hamsters and to use the new transgenic animals both to measure...Transgenic Hamster Models of TSEs Using a Radical New Technology PRINCIPAL INVESTIGATOR: Robert G. Rohwer, Ph.D. Irena Alexeeva, Ph.D...From - To) 1 SEP 2003 - 31 AUG 2007 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Efficient and Rapid Development of Transgenic Hamster Models of

  19. Study on the Handle of Keratin Transgenic Cotton Fabric

    Institute of Scientific and Technical Information of China (English)

    蒋培清; 严文源; 严灏景

    2004-01-01

    Gene of animal keratin can be inoculated into cotton fiber and thus get the keratin transgenic cotton fiber through transgenic technology. Handle of two kinds of pure cotton poplin, one of which is made of the keratin transgenic cotton while the other is made of the ordinary cotton of the same breed as control group and both with absolutely identical spinning, weaving, and dyeing process, was objectively evaluated with KES system. The result of analysis indicates that the principal changes of keratin transgenic cotton fabric are that the bending and shearing property of the fabric are considerably enhanced, KOSHI (Stiffness) and HARI (Anti-drape stiffness) of the fabric are good, while SHINAYAKASA (Flexibility with soft feeling) and SHARI (Crispness) decline.

  20. Growth enhancement in transgenic Atlantic salmon by the use of an "all fish" chimeric growth hormone gene construct.

    Science.gov (United States)

    Du, S J; Gong, Z Y; Fletcher, G L; Shears, M A; King, M J; Idler, D R; Hew, C L

    1992-02-01

    We have developed an "all fish" growth hormone (GH) chimeric gene construct by using an antifreeze protein gene (AFP) promoter from ocean pout linked to a chinook salmon GH cDNA clone. After microinjection into fertilized, nonactivated Atlantic salmon eggs via the micropyle, transgenic Atlantic salmon were generated. The presence of the transgene was detected by polymerase chain reaction (PCR) using specific oligonucleotide primers. A number of these transgenic fish showed dramatic increases in their growth rate. At one year old, the average increase of the transgenic fish was 2 to 6 fold and the largest transgenic fish was 13 times that of the average non-transgenic control.

  1. CROP OF TRANSGENIC SOY-BEAN. PEASANT AND INDIGENOUS COMMUNITY EFFECTS OF EUSTERN PARAGUAY

    Directory of Open Access Journals (Sweden)

    Hugo Florencio Centurión Mereles

    2011-09-01

    Full Text Available The Word offers us a critical glance about the economic and socio-cultural impacts of the transgenic crops in peasant and indigenous communities of the eastern region of Paraguay, it is given special attention to the use of Roundup, with the undoubted environment cost and the uncertain risks to human and animal health. The impacts and interactions that the techno-commoditization of the organisms genetically modified OGM have on the affected populations and the environment-species, soil, water, woods, flora, fauna is discussed in the Word.The extent of use of glyphosate on transgenic crops would involve not only the environment of the crop, but go to generate profound cultural changes, technological, of management, environmental, economical, social and legal, whose effects we already see them come with the decline of peasant and indigenous communities, that at not finding adequate strategies to face them, collapse as collectivity. The Word contains important elements to renew the debate and the critical thought in relation to the problematic of transgenic crop and its impact in indigenous and peasant populations.

  2. Genotype-independent transmission of transgenic fluorophore protein by boar spermatozoa.

    Directory of Open Access Journals (Sweden)

    Wiebke Garrels

    Full Text Available Recently, we generated transposon-transgenic boars (Sus scrofa, which carry three monomeric copies of a fluorophore marker gene. Amazingly, a ubiquitous fluorophore expression in somatic, as well as in germ cells was found. Here, we characterized the prominent fluorophore load in mature spermatozoa of these animals. Sperm samples were analyzed for general fertility parameters, sorted according to X and Y chromosome-bearing sperm fractions, assessed for potential detrimental effects of the reporter, and used for inseminations into estrous sows. Independent of their genotype, all spermatozoa were uniformly fluorescent with a subcellular compartmentalization of the fluorophore protein in postacrosomal sheath, mid piece and tail. Transmission of the fluorophore protein to fertilized oocytes was shown by confocal microscopic analysis of zygotes. The monomeric copies of the transgene segregated during meiosis, rendering a certain fraction of the spermatozoa non-transgenic (about 10% based on analysis of 74 F1 offspring. The genotype-independent transmission of the fluorophore protein by spermatozoa to oocytes represents a non-genetic contribution to the mammalian embryo.

  3. Co-expression of NCED and ALO improves vitamin C level and tolerance to drought and chilling in transgenic tobacco and stylo plants.

    Science.gov (United States)

    Bao, Gegen; Zhuo, Chunliu; Qian, Chunmei; Xiao, Ting; Guo, Zhenfei; Lu, Shaoyun

    2016-01-01

    Abscisic acid (ABA) regulates plant adaptive responses to various environmental stresses, while L-ascorbic acid (AsA) that is also named vitamin C is an important antioxidant and involves in plant stress tolerance and the immune system in domestic animals. Transgenic tobacco (Nicotiana tabacum L.) and stylo [Stylosanthes guianensis (Aublet) Swartz], a forage legume, plants co-expressing stylo 9-cis-epoxycarotenoid dioxygenase (SgNCED1) and yeast D-arabinono-1,4-lactone oxidase (ALO) genes were generated in this study, and tolerance to drought and chilling was analysed in comparison with transgenic tobacco overexpressing SgNCED1 or ALO and the wild-type plants. Compared to the SgNCED1 or ALO transgenic plants, in which only ABA or AsA levels were increased, both ABA and AsA levels were increased in transgenic tobacco and stylo plants co-expressing SgNCED1 and ALO genes. Compared to the wild type, an enhanced drought tolerance was observed in SgNCED1 transgenic tobacco plants with induced expression of drought-responsive genes, but not in ALO plants, while an enhanced chilling tolerance was observed in ALO transgenic tobaccos with induced expression of cold-responsive genes, but not in SgNCED1 plants. Co-expression of SgNCED1 and ALO genes resulted in elevated tolerance to both drought and chilling in transgenic tobacco and stylo plants with induced expression of both drought and cold-responsive genes. Our result suggests that co-expression of SgNCED1 and ALO genes is an effective way for use in forage plant improvement for increased tolerance to drought and chilling and nutrition quality.

  4. Toxicity assessment of transgenic papaya ringspot virus of 823-2210 line papaya fruits.

    Science.gov (United States)

    Lin, Hsin-Tang; Yen, Gow-Chin; Huang, Ting-Tzu; Chan, Lit-Fu; Cheng, Ying-Huey; Wu, Jhaol-Huei; Yeh, Shyi-Dong; Wang, Sheng-Yang; Liao, Jiunn-Wang

    2013-02-20

    The transgenic papaya is a valuable strategy for creating plants resistant to papaya ringspot virus (PRSV) infection and increasing production. This study was further performed to evaluate the comparative toxicity effects of the newly developed transgenic line of the fruits of two backcross transgenic papaya lines (2210 and 823) and one hybrid line (823-2210) and compare to their parent non-transgenic (TN-2) counterparts. The stability analysis of coat protein (CP) of PRSV was investigated using the digestion stability assays in simulated gastric fluid (SGF), simulated intestinal fluid (SIF), and bile salts to detect the CP fragments. Results revealed that the CP fragments were rapidly hydrolyzed in SGF and were undetectable in organs and gastrointestinal contents in rats. For the genotoxicity, three in vitro assays were conducted and exhibited that non-transgenic and backcross transgenic papaya fruits were negative. Moreover, a repeated animal feeding study was conducted by feeding 2 g/kg of body weight (bw) of non-transgenic and backcross transgenic papaya fruits for 28 days in rats. There were no biological or toxicological significances between non-transgenic and backcross transgenic papaya fruits in rats. The results demonstrated that the backcross transgenic papaya fruit can be recognized as an equivalent substitution for traditional papaya in food safety.

  5. Pollutant generation coefficient and discharge coefficient in animal production%畜禽养殖业产污系数和排污系数计算方法

    Institute of Scientific and Technical Information of China (English)

    董红敏; 朱志平; 黄宏坤; 陈永杏; 尚斌; 陶秀萍; 周忠凯

    2011-01-01

    Pollutant generation and discharge coefficients in animal production are the key parameters for animal environment engineering research and animal waste treatment projects. There were no authoritative pollutant generation and discharge coefficients available in China because the research about animal husbandry environmental engineering started late. Based on the characteristics of China's animal industry, generation and discharge coefficients of pollutants definitions, calculation methods, application scope and application of methods were discussed combined with typical cases analysis. Under the application of a typical pig farm in Beijing, three stage (nursery, fattening and gestation)generation coefficient for one pig were calculated, which were 252.8, 479.6, 493.4 g/d for COD, 20.4, 33.2, 43.7 g/d for TN, 3.48、 6.06、 9.93 g/d for TP, respectively. Under the pig waste treatment system operation condition, three stage discharge coefficients were calculated, these were 44.9, 64.1, 22.5 g/d for COD, 14.1, 20.9, 36.3 g/d for TN, 1.0, 1.8, 0.4 g/d for TP, respectively. The results can provide a reference for the pollution source census, animal waste treatment system operation.%畜禽养殖业产污系数与排污系数是畜牧环境研究和粪便处理工程设计的基础指标,但由于中国畜牧业环境工作起步晚,还没有根据中国畜牧业生产特性确定的产污系数和排污系数.该文根据中国畜禽养殖业的特点,提出了畜禽养殖业产污系数和排污系数的定义、计算方法,并结合典型猪场进行了案例分析.对于北京市某养猪场进行分析,结果表明:该猪场保育、育肥和妊娠母猪3个阶段的COD产污系数分别为每头252.8、479.6、493.4 g/d,全氮分别为每头20.4、33.2、43.7 g/d,全磷分别为每头3.48、6.06、9.93 g/d,在该猪场废弃物处理系统的运行情况下,计算得出了该场保育、育肥和妊娠母猪3个阶段的COD排污系数分别为每头44.9、64.1

  6. Efficient production by sperm-mediated gene transfer of human decay accelerating factor (hDAF) transgenic pigs for xenotransplantation

    Science.gov (United States)

    Lavitrano, Marialuisa; Bacci, Maria Laura; Forni, Monica; Lazzereschi, Davide; Di Stefano, Carla; Fioretti, Daniela; Giancotti, Paola; Marfé, Gabriella; Pucci, Loredana; Renzi, Luigina; Wang, Hongjun; Stoppacciaro, Antonella; Stassi, Giorgio; Sargiacomo, Massimo; Sinibaldi, Paola; Turchi, Valeria; Giovannoni, Roberto; Della Casa, Giacinto; Seren, Eraldo; Rossi, Giancarlo

    2002-01-01

    A large number of hDAF transgenic pigs to be used for xenotransplantation research were generated by using sperm-mediated gene transfer (SMGT). The efficiency of transgenesis obtained with SMGT was much greater than with any other method. In the experiments reported, up to 80% of pigs had the transgene integrated into the genome. Most of the pigs carrying the hDAF gene transcribed it in a stable manner (64%). The great majority of pigs that transcribed the gene expressed the protein (83%). The hDAF gene was transmitted to progeny. Expression was stable and found in caveolae as it is in human cells. The expressed gene was functional based on in vitro experiments performed on peripheral blood mononuclear cells. These results show that our SMGT approach to transgenesis provides an efficient procedure for studies involving large animal models. PMID:12393815

  7. Towards Transgenic Primates: What can we learn from mouse genetics?

    Institute of Scientific and Technical Information of China (English)

    KUANG Hui; WANG Phillip L.; TSIEN Joe Z.

    2009-01-01

    Considering the great physiological and behavioral similarities with humans, monkeys represent the ideal models not only for the study of complex cognitive behavior but also for the precUnical research and development of novel therapeutics for treating human diseases. Various powerful genetic tech-nologies initially developed for making mouse models are being explored for generating transgenic primate models. We review the latest genetic engineering technologies and discuss the potentials and limitations for systematic production of transgenic primates.

  8. Virtual Transgenics: Using a Molecular Biology Simulation to Impact Student Academic Achievement and Attitudes

    Science.gov (United States)

    Shegog, Ross; Lazarus, Melanie M.; Murray, Nancy G.; Diamond, Pamela M.; Sessions, Nathalie; Zsigmond, Eva

    2012-01-01

    The transgenic mouse model is useful for studying the causes and potential cures for human genetic diseases. Exposing high school biology students to laboratory experience in developing transgenic animal models is logistically prohibitive. Computer-based simulation, however, offers this potential in addition to advantages of fidelity and reach.…

  9. Maternal endometrial edema may increase perinatal mortality of cloned and transgenic piglets

    DEFF Research Database (Denmark)

    Schmidt, Mette; Winter, Kjeld Dahl; Dantzer, Vibeke

    2011-01-01

    The perinatal mortality of cloned animals is a well-known problem. In the present retrospective study, we report on mortality of cloned transgenic or non-transgenic piglets produced as part of several investigations. Large White (LW) sows (n = 105) received hand-made cloned LW or minipig blastocy...

  10. [Effects of phytase transgenic corn planting on soil nematode community].

    Science.gov (United States)

    Zhao, Zong-Chao; Su, Ying; Mou, Wen-Ya; Liu, Man-Qiang; Chen, Xiao-Yun; Chen, Fa-Jun

    2014-04-01

    A healthy soil ecosystem is essential for nutrient cycling and energy conversion, and the impact of exogenous genes from genetically modified crops had aroused wide concerns. Phytase transgenic corn (i. e., the inbred line BVLA430101) was issued a bio-safety certificate on 27 September 2009 in China, which could improve the efficiency of feed utilization, reduce environmental pollution caused by animal manure. In this study, the abundance of trophic groups, community structure and ecological indices of soil nematodes were studied over the growing cycle of phytase transgenic corn (ab. transgenic corn) and control conventional parental corn (ab. control corn) in the field. Totally 29 and 26 nematode genera were isolated from transgenic corn and control corn fields, respectively. The abundances of bacterivores and omnivores-predators, the total number of soil nematodes, and the Shannon index (H) were significantly greater under transgenic corn than under control corn, while the opposite trend was found for the relative abundance of herbivores and the maturity index (Sigma MI) of soil nematodes. Repeated-measures analysis of variance (ANOVA) did not detect any significant effects of transgenic corn on the composition and abundance of nematode trophic groups and ecological indices of soil nematodes. Furthermore, the Student-T test showed that the abundances of bacterivores and omnivores-predators and the total number of soil nematodes during the milk-ripe stage were significant higher in the transgenic corn field than in the control corn field. The effects of transgenic corn planting on soil nematodes might be related to the increase in the nitrogen content of field soil under transgenic corn compared to control corn.

  11. Large genetic animal models of Huntington's Disease.

    Science.gov (United States)

    Morton, A Jennifer; Howland, David S

    2013-01-01

    The dominant nature of the Huntington's disease gene mutation has allowed genetic models to be developed in multiple species, with the mutation causing an abnormal neurological phenotype in all animals in which it is expressed. Many different rodent models have been generated. The most widely used of these, the transgenic R6/2 mouse, carries the mutation in a fragment of the human huntingtin gene and has a rapidly progressive and fatal neurological phenotype with many relevant pathological changes. Nevertheless, their rapid decline has been frequently questioned in the context of a disease that takes years to manifest in humans, and strenuous efforts have been made to make rodent models that are genetically more 'relevant' to the human condition, including full length huntingtin gene transgenic and knock-in mice. While there is no doubt that we have learned, and continue to learn much from rodent models, their usefulness is limited by two species constraints. First, the brains of rodents differ significantly from humans in both their small size and their neuroanatomical organization. Second, rodents have much shorter lifespans than humans. Here, we review new approaches taken to these challenges in the development of models of Huntington's disease in large brained, long-lived animals. We discuss the need for such models, and how they might be used to fill specific niches in preclinical Huntington's disease research, particularly in testing gene-based therapeutics. We discuss the advantages and disadvantages of animals in which the prodromal period of disease extends over a long time span. We suggest that there is considerable 'value added' for large animal models in preclinical Huntington's disease research.

  12. Development of a Conditional Transgenic Mouse Model to Test the Fallopian Tube Origin of Serous Ovarian Cancer

    Science.gov (United States)

    2009-03-01

    transgene plasmid to follow up on this next strategy. KEY RESEARCH ACCOMPLISHMENTS: • molecular cloning of Ovgp1-lacZ reporter gene plasmid...high-expressing 242 line • molecular cloning of Ovgp1-Cre transgene plasmid REPORTABLE OUTCOMES: • generation of two Ovgp1-lacZ transgenic

  13. Resistance of Antimicrobial Peptide Gene Transgenic Rice to Bacterial Blight

    Institute of Scientific and Technical Information of China (English)

    WANG Wei; WU Chao; LIU Mei; LIU Xu-ri; Hu Guo-cheng; SI Hua-min; SUN Zong-xiu; LIU Wen-zhen; Fu Ya-ping

    2011-01-01

    Antimierobial peptide is a polypeptide with antimicrobial activity.Antimicrobial peptide genes Np3 and Np5 from Chinese shrimp (Fenneropenaeus Chinensis) were integrated into Oryza sativa L.subsp.japonica cv.Aichi ashahi by Agrobacterium mediated transformation system.PCR analysis showed that the positive ratios of Np3 and Np5 were 36% and 45% in T0 generation,respectively.RT-PCR analysis showed that the antimicrobial peptide genes were expressed in T1 generation,and there was no obvious difference in agronomic traits between transgenic plants and non-transgenic plants.Four Np3 and Np5 transgenic lines in T1 generation were inoculated with ×anthomonas oryzae pv.oryzae strain CR4,and all the four transgenic lines had significantly enhanced resistance to bacterial blight caused by the strain CR4.The Np5 transgenic lines also showed higher resistance to bacterial blight caused by strains JS97-2,Zhe 173 and OS-225.It is suggested that transgenic lines with Np5 gene might possess broad spectrum resistance to rice bacterial blight.

  14. Transcription activator-like effector nuclease (TALEN-mediated CLYBL targeting enables enhanced transgene expression and one-step generation of dual reporter human induced pluripotent stem cell (iPSC and neural stem cell (NSC lines.

    Directory of Open Access Journals (Sweden)

    Trevor Cerbini

    Full Text Available Targeted genome engineering to robustly express transgenes is an essential methodology for stem cell-based research and therapy. Although designer nucleases have been used to drastically enhance gene editing efficiency, targeted addition and stable expression of transgenes to date is limited at single gene/locus and mostly PPP1R12C/AAVS1 in human stem cells. Here we constructed transcription activator-like effector nucleases (TALENs targeting the safe-harbor like gene CLYBL to mediate reporter gene integration at 38%-58% efficiency, and used both AAVS1-TALENs and CLYBL-TALENs to simultaneously knock-in multiple reporter genes at dual safe-harbor loci in human induced pluripotent stem cells (iPSCs and neural stem cells (NSCs. The CLYBL-TALEN engineered cell lines maintained robust reporter expression during self-renewal and differentiation, and revealed that CLYBL targeting resulted in stronger transgene expression and less perturbation on local gene expression than PPP1R12C/AAVS1. TALEN-mediated CLYBL engineering provides improved transgene expression and options for multiple genetic modification in human stem cells.

  15. Transcription activator-like effector nuclease (TALEN)-mediated CLYBL targeting enables enhanced transgene expression and one-step generation of dual reporter human induced pluripotent stem cell (iPSC) and neural stem cell (NSC) lines.

    Science.gov (United States)

    Cerbini, Trevor; Funahashi, Ray; Luo, Yongquan; Liu, Chengyu; Park, Kyeyoon; Rao, Mahendra; Malik, Nasir; Zou, Jizhong

    2015-01-01

    Targeted genome engineering to robustly express transgenes is an essential methodology for stem cell-based research and therapy. Although designer nucleases have been used to drastically enhance gene editing efficiency, targeted addition and stable expression of transgenes to date is limited at single gene/locus and mostly PPP1R12C/AAVS1 in human stem cells. Here we constructed transcription activator-like effector nucleases (TALENs) targeting the safe-harbor like gene CLYBL to mediate reporter gene integration at 38%-58% efficiency, and used both AAVS1-TALENs and CLYBL-TALENs to simultaneously knock-in multiple reporter genes at dual safe-harbor loci in human induced pluripotent stem cells (iPSCs) and neural stem cells (NSCs). The CLYBL-TALEN engineered cell lines maintained robust reporter expression during self-renewal and differentiation, and revealed that CLYBL targeting resulted in stronger transgene expression and less perturbation on local gene expression than PPP1R12C/AAVS1. TALEN-mediated CLYBL engineering provides improved transgene expression and options for multiple genetic modification in human stem cells.

  16. Limited fitness advantages of crop-weed hybrid progeny containing insect-resistant transgenes (Bt/CpTI in transgenic rice field.

    Directory of Open Access Journals (Sweden)

    Xiao Yang

    Full Text Available BACKGROUND: The spread of insect-resistance transgenes from genetically engineered (GE rice to its coexisting weedy rice (O. sativa f. spontanea populations via gene flow creates a major concern for commercial GE rice cultivation. Transgene flow to weedy rice seems unavoidable. Therefore, characterization of potential fitness effect brought by the transgenes is essential to assess environmental consequences caused by crop-weed transgene flow. METHODOLOGY/PRINCIPAL FINDINGS: Field performance of fitness-related traits was assessed in advanced hybrid progeny of F(4 generation derived from a cross between an insect-resistant transgenic (Bt/CpTI rice line and a weedy strain. The performance of transgene-positive hybrid progeny was compared with the transgene-negative progeny and weedy parent in pure and mixed planting of transgenic and nontransgenic plants under environmental conditions with natural vs. low insect pressure. Results showed that under natural insect pressure the insect-resistant transgenes could effectively suppress target insects and bring significantly increased fitness to transgenic plants in pure planting, compared with nontransgenic plants (including weedy parent. In contrast, no significant differences in fitness were detected under low insect pressure. However, such increase in fitness was not detected in the mixed planting of transgenic and nontransgenic plants due to significantly reduced insect pressure. CONCLUSIONS/SIGNIFICANCE: Insect-resistance transgenes may have limited fitness advantages to hybrid progeny resulted from crop-weed transgene flow owning to the significantly reduced ambient target insect pressure when an insect-resistant GE crop is grown. Given that the extensive cultivation of an insect-resistant GE crop will ultimately reduce the target insect pressure, the rapid spread of insect-resistance transgenes in weedy populations in commercial GE crop fields may be not likely to happen.

  17. Research of human hair animation generation technology in cartoon%一种卡通动画中人物头发运动生成技术研究

    Institute of Scientific and Technical Information of China (English)

    欧训勇

    2014-01-01

    This paper introduces a new hybrid technique for human hair animation in cartoon , which allows the animators to create attractive and controllable hair animations without having to draw everything by hand except a sparse set of keyframes. This method demonstrates how to give a cel-animation character accentuated hair motion. The novelty of this approach is that we neither simply interpolate the key frames nor generate the movement of the hair only using physical simulations. From a small number of rough sketches we prepare key frames that are used as indicators of hair motion. The hair movements are created based on a hair motion database built from physical simulations custom-designed by the animator. Hair animations with constraints from the key frames can be generated in two stages: a matching process to search for the desired motion sequences from the database and then smoothly connect them; the discrepancies between the database sequences and the key frames are interpolated throughout the animation using transition function.%介绍了一种新的卡通动画中人物头发运动的混合技术,使动画师们能方便地创作出有吸引力和可控性的头发运动效果,而不需要手工对头发运动表现过程进行逐帧绘制,仅设置好所需的一些关键帧画面。该技术方法演示如何根据给出的手绘卡通人物角色动作配以相应的头发运动效果。这种方法的新颖之处是,既不是简单地插入关键帧来获得头发运动效果,也不是通过物理模拟的方式来生成头发飘逸的动画,而是从动画师预先准备好的少量的作为关键帧草图的头发运动效果,建立物理模拟动画数据库,根据此数据库中头发的运动效果来创建出特定头发运动动画。此方法生成的头发动画分两个阶段完成,首先是在数据库中搜索对应的运动效果画面,然后顺利连接;其次数据库序列的关键帧之间的过渡帧由在动画中运

  18. Application of Echocardiography on Transgenic Mice with Cardiomyopathies

    Directory of Open Access Journals (Sweden)

    G. Chen

    2012-01-01

    Full Text Available Cardiomyopathies are common cardiac disorders that primarily affect cardiac muscle resulting in cardiac dysfunction and heart failure. Transgenic mouse disease models have been developed to investigate the cellular mechanisms underlying heart failure and sudden cardiac death observed in cardiomyopathy cases and to explore the therapeutic outcomes in experimental animals in vivo. Echocardiography is an essential diagnostic tool for accurate and noninvasive assessment of cardiac structure and function in experimental animals. Our laboratory has been among the first to apply high-frequency research echocardiography on transgenic mice with cardiomyopathies. In this work, we have summarized our and other studies on assessment of systolic and diastolic dysfunction using conventional echocardiography, pulsed Doppler, and tissue Doppler imaging in transgenic mice with various cardiomyopathies. Estimation of embryonic mouse hearts has been performed as well using this high-resolution echocardiography. Some technical considerations in mouse echocardiography have also been discussed.

  19. Applying digital particle image velocimetry to animal-generated flows : Traps, hurdles and cures in mapping steady and unsteady flows in Re regimes between 10(-2) and 10(5)

    NARCIS (Netherlands)

    Stamhuis, EJ; Videler, JJ; van Duren, LA; Muller, UK

    2002-01-01

    Digital particle image velocimetry (DPIV) has been applied to animal-generated flows since 1993 to map the flow patterns and vortex wakes produced by a range of feeding and swimming aquatic animals, covering a Re range of 10(-2)-10(5). In this paper, the special circumstances, problems and some solu

  20. Transgenic barley: a prospective tool for biotechnology and agriculture.

    Science.gov (United States)

    Mrízová, Katarína; Holasková, Edita; Öz, M Tufan; Jiskrová, Eva; Frébort, Ivo; Galuszka, Petr

    2014-01-01

    Barley (Hordeum vulgare L.) is one of the founder crops of agriculture, and today it is the fourth most important cereal grain worldwide. Barley is used as malt in brewing and distilling industry, as an additive for animal feed, and as a component of various food and bread for human consumption. Progress in stable genetic transformation of barley ensures a potential for improvement of its agronomic performance or use of barley in various biotechnological and industrial applications. Recently, barley grain has been successfully used in molecular farming as a promising bioreactor adapted for production of human therapeutic proteins or animal vaccines. In addition to development of reliable transformation technologies, an extensive amount of various barley genetic resources and tools such as sequence data, microarrays, genetic maps, and databases has been generated. Current status on barley transformation technologies including gene transfer techniques, targets, and progeny stabilization, recent trials for improvement of agricultural traits and performance of barley, especially in relation to increased biotic and abiotic stress tolerance, and potential use of barley grain as a protein production platform have been reviewed in this study. Overall, barley represents a promising tool for both agricultural and biotechnological transgenic approaches, and is considered an ancient but rediscovered crop as a model industrial platform for molecular farming.

  1. Sperm Nuclear Transfer and Transgenic Production in the Fish Medaka

    Directory of Open Access Journals (Sweden)

    Tongming Liu, Ling Liu, Qiwei Wei, Yunhan Hong

    2011-01-01

    Full Text Available Sperm nuclear transfer or intracytoplasmic sperm injection (ICSI is a powerful assisted reproductive technology (ART for treating human male infertility. Controversial reports of increased birth defects have raised concerns about the ART's safety. The cause for birth defects, however, has remained elusive for analysis in human because of the sample size, male infertility genetics, physiological heterogeneity and associated procedures such as embryo manipulations. Animal models are required to evaluate factors leading to the increased birth defects. Here we report the establishment of medakafish model for ICSI and transgenic production. This small laboratory fish has high fecundity and easy embryology. We show that ICSI produced a 5% high percentage of fertile animals that exhibited both paternal and maternal contribution as evidenced by the pigmentation marker. Furthermore, when sperm were pre-incubated with a plasmid ubiquitously expressing RFP and subjected to ICSI, 50% of sperm nuclear transplants showed germline transmission. We conclude that medaka is an excellent model for ICSI to evaluate birth defects and that sperm nuclear transfer can mediate stable gene transfer at high efficiency. Although more demanding for experimentation, sperm-mediated transgenesis should be particularly applicable for aquaculture species with a lengthy generation time and/or a large adult body size.

  2. Cross-generational trans fat intake modifies BDNF mRNA in the hippocampus: Impact on memory loss in a mania animal model.

    Science.gov (United States)

    Trevizol, Fabíola; Dias, Verônica T; Roversi, Katiane; Barcelos, Raquel C S; Kuhn, Fábio T; Roversi, Karine; Pase, Camila S; Golombieski, Ronaldo; Veit, Juliana C; Piccolo, Jaqueline; Emanuelli, Tatiana; Rocha, João B T; Bürger, Marilise E

    2015-05-01

    Recently, we have described the influence of dietary fatty acids (FA) on mania-like behavior of first generation animals. Here, two sequential generations of female rats were supplemented with soybean oil (SO, rich in n-6 FA, control group), fish oil (FO, rich in n-3 FA) and hydrogenated vegetable fat (HVF, rich in trans FA) from pregnancy and during lactation. In adulthood, half of each group was exposed to an amphetamine (AMPH)-induced mania animal model for behavioral, biochemical and molecular assessments. FO supplementation was associated with lower reactive species (RS) generation and protein carbonyl (PC) levels and increased dopamine transporter (DAT) levels, while HVF increased RS and PC levels, thus decreasing catalase (CAT) activity and DAT levels in hippocampus after AMPH treatment. AMPH impaired short- (1 h) and long- (24 h) term memory in the HVF group. AMPH exposure was able to reduce hippocampal BDNF- mRNA expression, which was increased in FO. While HVF was related to higher trans FA (TFA) incorporation in hippocampus, FO was associated with increased percentage of n-3 polyunsaturated FA (PUFA) together with lower n-6/n-3 PUFA ratio. Interestingly, our data showed a positive correlation between brain-derived neurotrophic factor (BDNF) mRNA and short- and long-term memory (r(2)  = 0.53; P = 0.000/r(2)  = 0.32; P = 0.011, respectively), as well as a negative correlation between PC and DAT levels (r(2)  = 0.23; P = 0.015). Our findings confirm that provision of n-3 or TFA during development over two generations is able to change the neuronal membrane lipid composition, protecting or impairing the hippocampus, respectively, thus affecting neurothrophic factor expression such as BDNF mRNA. In this context, chronic consumption of trans fats over two generations can facilitate the development of mania-like behavior, so leading to memory impairment and emotionality, which are related to neuropsychiatric conditions.

  3. Transgenic algae engineered for higher performance

    Energy Technology Data Exchange (ETDEWEB)

    Unkefer, Pat J; Anderson, Penelope S; Knight, Thomas J

    2014-10-21

    The present disclosure relates to transgenic algae having increased growth characteristics, and methods of increasing growth characteristics of algae. In particular, the disclosure relates to transgenic algae comprising a glutamine phenylpyruvate transaminase transgene and to transgenic algae comprising a glutamine phenylpyruvate transaminase transgene and a glutamine synthetase.

  4. Institute of Laboratory Animal Resources

    Science.gov (United States)

    1992-06-01

    country of Central and South America, as well as to the Caribbean, and Mexico and published notices in newsletters. Young scientists from Mexico , Peru, and...Transgenic Animals. J. Gordon. 30 4 F 88 The Spontaneously Hypercholesterolemic Pig as an Animal Model for Human Atherosclerosis . A. Attie & M...and Potential Approaches. An NIH Conference. The Contributions of Nonhuman Primates to Understanding Coronary Artery Atherosclerosis in Humans. T

  5. Transgenic overexpression of LARGE induces α-dystroglycan hyperglycosylation in skeletal and cardiac muscle.

    Directory of Open Access Journals (Sweden)

    Martin Brockington

    Full Text Available BACKGROUND: LARGE is one of seven putative or demonstrated glycosyltransferase enzymes defective in a common group of muscular dystrophies with reduced glycosylation of α-dystroglycan. Overexpression of LARGE induces hyperglycosylation of α-dystroglycan in both wild type and in cells from dystroglycanopathy patients, irrespective of their primary gene defect, restoring functional glycosylation. Viral delivery of LARGE to skeletal muscle in animal models of dystroglycanopathy has identical effects in vivo, suggesting that the restoration of functional glycosylation could have therapeutic applications in these disorders. Pharmacological strategies to upregulate Large expression are also being explored. METHODOLOGY/PRINCIPAL FINDINGS: In order to asses the safety and efficacy of long term LARGE over-expression in vivo, we have generated four mouse lines expressing a human LARGE transgene. On observation, LARGE transgenic mice were indistinguishable from the wild type littermates. Tissue analysis from young mice of all four lines showed a variable pattern of transgene expression: highest in skeletal and cardiac muscles, and lower in brain, kidney and liver. Transgene expression in striated muscles correlated with α-dystroglycan hyperglycosylation, as determined by immunoreactivity to antibody IIH6 and increased laminin binding on an overlay assay. Other components of the dystroglycan complex and extracellular matrix ligands were normally expressed, and general muscle histology was indistinguishable from wild type controls. Further detailed muscle physiological analysis demonstrated a loss of force in response to eccentric exercise in the older, but not in the younger mice, suggesting this deficit developed over time. However this remained a subclinical feature as no pathology was observed in older mice in any muscles including the diaphragm, which is sensitive to mechanical load-induced damage. CONCLUSIONS/SIGNIFICANCE: This work shows that

  6. Methodological strategies for transgene copy number quantification in goats (Capra hircus) using real-time PCR.

    Science.gov (United States)

    Batista, Ribrio I T P; Luciano, Maria C S; Teixeira, Dárcio I A; Freitas, Vicente J F; Melo, Luciana M; Andreeva, Lyudmila E; Serova, Irina A; Serov, Oleg L

    2014-01-01

    Taking into account the importance of goats as transgenic models, as well as the rarity of copy number (CN) studies in farm animals, the present work aimed to evaluate methodological strategies for accurate and precise transgene CN quantification in goats using quantitative polymerase chain reaction (qPCR). Mouse and goat lines transgenic for human granulocyte-colony stimulating factor were used. After selecting the best genomic DNA extraction method to be applied in mouse and goat samples, intra-assay variations, accuracy and precision of CN quantifications were assessed. The optimized conditions were submitted to mathematical strategies and used to quantify CN in goat lines. The findings were as follows: validation of qPCR conditions is required, and amplification efficiency is the most important. Absolute and relative quantifications are able to produce similar results. For normalized absolute quantification, the same plasmid fragment used to generate goat lines must be mixed with wild-type goat genomic DNA, allowing the choice of an endogenous reference gene for data normalization. For relative quantifications, a resin-based genomic DNA extraction method is strongly recommended when using mouse tail tips as calibrators to avoid tissue-specific inhibitors. Efficient qPCR amplifications (≥95%) allow reliable CN measurements with SYBR technology. TaqMan must be used with caution in goats if the nucleotide sequence of the endogenous reference gene is not yet well understood. Adhering to these general guidelines can result in more exact CN determination in goats. Even when working under nonoptimal circumstances, if assays are performed that respect the minimum qPCR requirements, good estimations of transgene CN can be achieved.

  7. Persistent amyloidosis following suppression of Abeta production in a transgenic model of Alzheimer disease.

    Directory of Open Access Journals (Sweden)

    2005-12-01

    Full Text Available BACKGROUND: The proteases (secretases that cleave amyloid-beta (Abeta peptide from the amyloid precursor protein (APP have been the focus of considerable investigation in the development of treatments for Alzheimer disease. The prediction has been that reducing Abeta production in the brain, even after the onset of clinical symptoms and the development of associated pathology, will facilitate the repair of damaged tissue and removal of amyloid lesions. However, no long-term studies using animal models of amyloid pathology have yet been performed to test this hypothesis. METHODS AND FINDINGS: We have generated a transgenic mouse model that genetically mimics the arrest of Abeta production expected from treatment with secretase inhibitors. These mice overexpress mutant APP from a vector that can be regulated by doxycycline. Under normal conditions, high-level expression of APP quickly induces fulminant amyloid pathology. We show that doxycycline administration inhibits transgenic APP expression by greater than 95% and reduces Abeta production to levels found in nontransgenic mice. Suppression of transgenic Abeta synthesis in this model abruptly halts the progression of amyloid pathology. However, formation and disaggregation of amyloid deposits appear to be in disequilibrium as the plaques require far longer to disperse than to assemble. Mice in which APP synthesis was suppressed for as long as 6 mo after the formation of Abeta deposits retain a considerable amyloid load, with little sign of active clearance. CONCLUSION: This study demonstrates that amyloid lesions in transgenic mice are highly stable structures in vivo that are slow to disaggregate. Our findings suggest that arresting Abeta production in patients with Alzheimer disease should halt the progression of pathology, but that early treatment may be imperative, as it appears that amyloid deposits, once formed, will require additional intervention to clear.

  8. Persistent amyloidosis following suppression of Abeta production in a transgenic model of Alzheimer disease.

    Directory of Open Access Journals (Sweden)

    Joanna L Jankowsky

    2005-12-01

    Full Text Available The proteases (secretases that cleave amyloid-beta (Abeta peptide from the amyloid precursor protein (APP have been the focus of considerable investigation in the development of treatments for Alzheimer disease. The prediction has been that reducing Abeta production in the brain, even after the onset of clinical symptoms and the development of associated pathology, will facilitate the repair of damaged tissue and removal of amyloid lesions. However, no long-term studies using animal models of amyloid pathology have yet been performed to test this hypothesis.We have generated a transgenic mouse model that genetically mimics the arrest of Abeta production expected from treatment with secretase inhibitors. These mice overexpress mutant APP from a vector that can be regulated by doxycycline. Under normal conditions, high-level expression of APP quickly induces fulminant amyloid pathology. We show that doxycycline administration inhibits transgenic APP expression by greater than 95% and reduces Abeta production to levels found in nontransgenic mice. Suppression of transgenic Abeta synthesis in this model abruptly halts the progression of amyloid pathology. However, formation and disaggregation of amyloid deposits appear to be in disequilibrium as the plaques require far longer to disperse than to assemble. Mice in which APP synthesis was suppressed for as long as 6 mo after the formation of Abeta deposits retain a considerable amyloid load, with little sign of active clearance.This study demonstrates that amyloid lesions in transgenic mice are highly stable structures in vivo that are slow to disaggregate. Our findings suggest that arresting Abeta production in patients with Alzheimer disease should halt the progression of pathology, but that early treatment may be imperative, as it appears that amyloid deposits, once formed, will require additional intervention to clear.

  9. Vaccine generated immunity targets an HPV16 E7 HLA-A2.1-restricted CD8(+) T cell epitope relocated to an early gene or a late gene of the cottontail rabbit papillomavirus (CRPV) genome in HLA-A2.1 transgenic rabbits.

    Science.gov (United States)

    Bounds, Callie E; Hu, Jiafen; Cladel, Nancy M; Balogh, Karla; Christensen, Neil D

    2011-02-01

    The newly established HLA-A2.1 transgenic rabbit model has proven useful for testing the immunogenicity of well known and computer-predicted A2-restricted epitopes. In the current study we compared the protective immunity induced to a preferred HPV16 E7 A2-restricted epitope that has been relocated to positions within the CRPV E7 gene and the CRPV L2 gene. Epitope expression from both the E7 protein and the L2 protein resulted in increased protection against viral DNA challenge of the HLA-A2.1 transgenic rabbits as compared to control-vaccinated rabbit groups. These data indicate that proteins expressed at both early and late time points during a natural papillomavirus infection can be targeted by epitope-specific immunity and indicate this immunity is increased to early rather than late expressed proteins of papillomaviruses. This study also highlights the broad utility of the HLAA2.1 transgenic rabbit model for testing numerous immunological factors involved in vaccine generated protective immunity.

  10. Comparison of Model Predictions and Laboratory Observations of Transgene Frequencies in Continuously-Breeding Mosquito Populations.

    Science.gov (United States)

    Valerio, Laura; North, Ace; Collins, C Matilda; Mumford, John D; Facchinelli, Luca; Spaccapelo, Roberta; Benedict, Mark Q

    2016-09-22

    The persistence of transgenes in the environment is a consideration in risk assessments of transgenic organisms. Combining mathematical models that predict the frequency of transgenes and experimental demonstrations can validate the model predictions, or can detect significant biological deviations that were neither apparent nor included as model parameters. In order to assess the correlation between predictions and observations, models were constructed to estimate the frequency of a transgene causing male sexual sterility in simulated populations of a malaria mosquito Anopheles gambiae that were seeded with transgenic females at various proportions. Concurrently, overlapping-generation laboratory populations similar to those being modeled were initialized with various starting transgene proportions, and the subsequent proportions of transgenic individuals in populations were determined weekly until the transgene disappeared. The specific transgene being tested contained a homing endonuclease gene expressed in testes, I-PpoI, that cleaves the ribosomal DNA and results in complete male sexual sterility with no effect on female fertility. The transgene was observed to disappear more rapidly than the model predicted in all cases. The period before ovipositions that contained no transgenic progeny ranged from as little as three weeks after cage initiation to as long as 11 weeks.

  11. Comparison of Model Predictions and Laboratory Observations of Transgene Frequencies in Continuously-Breeding Mosquito Populations

    Directory of Open Access Journals (Sweden)

    Laura Valerio

    2016-09-01

    Full Text Available The persistence of transgenes in the environment is a consideration in risk assessments of transgenic organisms. Combining mathematical models that predict the frequency of transgenes and experimental demonstrations can validate the model predictions, or can detect significant biological deviations that were neither apparent nor included as model parameters. In order to assess the correlation between predictions and observations, models were constructed to estimate the frequency of a transgene causing male sexual sterility in simulated populations of a malaria mosquito Anopheles gambiae that were seeded with transgenic females at various proportions. Concurrently, overlapping-generation laboratory populations similar to those being modeled were initialized with various starting transgene proportions, and the subsequent proportions of transgenic individuals in populations were determined weekly until the transgene disappeared. The specific transgene being tested contained a homing endonuclease gene expressed in testes, I-PpoI, that cleaves the ribosomal DNA and results in complete male sexual sterility with no effect on female fertility. The transgene was observed to disappear more rapidly than the model predicted in all cases. The period before ovipositions that contained no transgenic progeny ranged from as little as three weeks after cage initiation to as long as 11 weeks.

  12. Transgenic fish systems and their application in ecotoxicology.

    Science.gov (United States)

    Lee, Okhyun; Green, Jon M; Tyler, Charles R

    2015-02-01

    The use of transgenics in fish is a relatively recent development for advancing understanding of genetic mechanisms and developmental processes, improving aquaculture, and for pharmaceutical discovery. Transgenic fish have also been applied in ecotoxicology where they have the potential to provide more advanced and integrated systems for assessing health impacts of chemicals. The zebrafish (Daniorerio) is the most popular fish for transgenic models, for reasons including their high fecundity, transparency of their embryos, rapid organogenesis and availability of extensive genetic resources. The most commonly used technique for producing transgenic zebrafish is via microinjection of transgenes into fertilized eggs. Transposon and meganuclease have become the most reliable methods for insertion of the genetic construct in the production of stable transgenic fish lines. The GAL4-UAS system, where GAL4 is placed under the control of a desired promoter and UAS is fused with a fluorescent marker, has greatly enhanced model development for studies in ecotoxicology. Transgenic fish have been developed to study for the effects of heavy metal toxicity (via heat-shock protein genes), oxidative stress (via an electrophile-responsive element), for various organic chemicals acting through the aryl hydrocarbon receptor, thyroid and glucocorticoid response pathways, and estrogenicity. These models vary in their sensitivity with only very few able to detect responses for environmentally relevant exposures. Nevertheless, the potential of these systems for analyses of chemical effects in real time and across multiple targets in intact organisms is considerable. Here we illustrate the techniques used for generating transgenic zebrafish and assess progress in the development and application of transgenic fish (principally zebrafish) for studies in environmental toxicology. We further provide a viewpoint on future development opportunities.

  13. The Molecular Detection and Functions Identification of T1 Generation of A tA MT1;1 Transgenic Sugar Beets%转AtAMT1;1基因甜菜T1代基因表达及其功能鉴定

    Institute of Scientific and Technical Information of China (English)

    张少英; 段娜; 孙亚卿; 于超; 张永丰

    2013-01-01

    Sugar beet is an important sugar crop, to improve beet nitrogen use efficiency, its genetic transform-ation of the high-affinity ammonium transporter gene was done. This experiment takes a molecule detection and physiological characteristics analysis to the T1 generation of A tA MT1;1 transgenic sugar beet. The results show that the PCR detection and RT-PCR detection of T1 generation of transgenic sugar beet is positive. In the control of nitrogen supply conditions, transgenic plants chlorophyll content and ammonia nitrogen content enhance than the control of wild-type sugar beet, and the difference was significant. Transgenic sugar beet yield and sugar content higher than the wild type, preliminary evidence A tA MT1;1 transgenic sugar beet can increase the efficiency of nitrogen utilization.%甜菜是中国重要的糖料作物,为提高甜菜氮素利用效率,对其进行了高亲和铵转运蛋白基因的遗传转化。本研究主要对转A tA MT1;1基因甜菜T1代植株进行了PCR、RT-PCR检测和相关生理特性的分析。结果表明,PCR和RT-PCR检测的T1代转基因甜菜呈阳性,并在根中表达;在控制氮素供应的条件下,转基因植株的叶绿素和氨态氮含量显著高于野生型甜菜,块根产量和含糖率也高于对照,初步证明转A tA MT1;1基因甜菜可提高氮素的利用效率。

  14. Green fluorescent protein (GFP) transgenic pig produced by somatic cell nuclear transfer

    Institute of Scientific and Technical Information of China (English)

    LIU ZhongHua; SUN Shuang; LI YuTian; WANG HongBin; R S PRATHER; SONG Jun; WANG ZhenKun; TIAN JiangTian; KONG QingRan; ZHENG Zhong; YIN Zhi; GAO Li; MA HaiKun

    2008-01-01

    Transgenic somatic cell nuclear transfer is a very promising route for producing transgenic farm ani-mals. Research on GFP transgenic pigs can provide useful information for breeding transgenic pigs, human disease models and human organ xenotransplantation. In this study, a liposomal transfection system was screened and transgenic embryos were reconstructed by nuclear transfer of GFP positive cells into enucleated in vitro matured oocytes. The development of reconstructed embryos both in vitro and in vivo was observed, and GFP expression was determined. The results showed that porcine fe-tal-derived fibroblast cells cultured with 4.0 plJmL liposome and 1.6 pg/mL plasmid DNA for 6 h re-sulted in the highest transfection rate (3.6%). The percentage of GFP reconstructed embryos that de-veloped in vitro to the blastocyst stage was 10%. Of those the GFP positive percentage was 48%. Re-constructed transgenic embryos were transferred to 10 recipients. 5 of them were pregnant, and 3 de-livered 6 cloned piglets in which 4 piglets were transgenic for the GFP as verified by both GFP protein expression and GFP DNA sequence analysis. The percentage of reconstructed embryos that resulted in cloned piglets was 1.0%; while the percentage of piglets that were transgenic was 0.7%. This is the first group of transgenic cloned pigs born in China, marking a great progress in Chinese transgenic cloned pig research.

  15. Expression of recombinant human lysozyme in the milk of transgenic mice

    Institute of Scientific and Technical Information of China (English)

    YU Zhengquan; FAN Baoliang; DAI Yunping; ZHENG Ming; NIU Huiling; WANG Meili; WANG Lili; FEI Jing; LI Ning

    2003-01-01

    Human lysozyme is a 130-aa (amino acid) alkaline polypeptide, and has both anti-bacterial and anti-viral properties which make it an important component of human natural immunity system. As a first step toward the ultimate goal ofimproving the anti-bacterial properties of bovine and ovine milk, a transgenic mouse that contains the genomic DNA sequence of the human lysozme gene has been generated for the first time. From 83 mice generated by microinjection, a total of 6 positive transgenic mice were identified by PCR and Southern blot. F1 mice positive for transgene in lines were also detected by PCR. This shows that transgene could be transmitted from founder transgenic mice to their offspring. Recombinant human lysozyme (rHlys) was found in the whey of 3 female positive transgenic mice by Western blot. The highest concentration of rHlys for transgenic micewas 0.2 mg/mL. The antibacterial activity of the whey for transgenic mice was highly enhanced up to 0.4 times as much as that of human, while that of non-transgenic mouse was very low. Although the lysozyme activity of transgenic mice is still lower than that of human, the rHlys exhibits the same specific activity as that of human lysozyme. It provides a strong basis for further studies into the possible application of rHlys express in mammary gland.

  16. Expression of human erythropoietin directed by mWAP promoter in mammary gland of transgenic mice

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The present work has generated transgenic mice with a hybrid gene construct consisting of genomic sequences encoding human erythropoietin (hEPO) and governed by regulatory sequences of mouse whey acidic protein (mWAP). The construct proved effective by transient expression in lactating animal. After introducing hybrid gene construct into single-cell embryo via pronuclear microinjection, surviving embryo are reimplanted into pseudopregnant foster mother mouse. 58 mice of 86 generation zero mice obtained were identified to be positive by PCR-Southern blot and genomic DNA Southern blot methods. The integration rate is 67%. hEPO was expressed in the milk of 16 mice of 39 mice measured by hEPO ELISA kit .The expression level gets over 15 m g/mL.

  17. Transgenic cloned sheep overexpressing ovine toll-like receptor 4.

    Science.gov (United States)

    Deng, Shoulong; Li, Guiguan; Zhang, Jinlong; Zhang, Xiaosheng; Cui, Maosheng; Guo, Yong; Liu, Guoshi; Li, Guangpeng; Feng, Jianzhong; Lian, Zhengxing

    2013-07-01

    An ovine fetal fibroblast cell line highly expressing TLR4 was established by inserting TLR4 into a reconstructive p3S-LoxP plasmid. Transgenic sheep overexpressing TLR4 were produced by transferring TLR4-transfected fetal fibroblasts into metaphase (M)II-stage enucleated oocytes (using SCNT). Because reconstructed embryos derived from MII-stage enucleated oocytes matured in vivo using a delayed-activated method had a higher pregnancy rate (18.52%) than that from MII-stage enucleated oocytes matured in vitro, the former procedure was used. Nine TLR4-transgenic live births were confirmed using polymerase chain reaction and Southern blot analysis. Increased expression of TLR4 at mRNA and protein levels in ear tissues of transgenic lambs were verified using reverse transcription polymerase chain reaction and immunohistochemistry, respectively. More toll-like receptor 4 protein was expressed by peripheral blood monocytes and/or macrophages collected from 3-month-old TLR4-transgenic than nontransgenic lambs at 0, 1, and 4 hours after lipopolysaccharide stimulation. Furthermore, interferon-γ and tumor necrosis factor α secreted by monocytes and/or macrophages of TLR4-transgenic lambs were significantly higher at 1 hour. Therefore, lipopolysaccharide-induced inflammatory responses from monocytes and/or macrophages occurred sooner in TLR4-transgenic lambs, consistent with an enhanced host immune response. In conclusion, transgenic sheep overexpressing TLR4 are a primary model to investigate the role of transgenic animals in disease resistance and have potential for breeding sheep with disease resistance.

  18. Epigenetic silencing in transgenic plants

    Directory of Open Access Journals (Sweden)

    Sarma eRajeev Kumar

    2015-09-01

    Full Text Available Epigenetic silencing is a natural phenomenon in which the expression of gene is regulated through modifications of DNA, RNA or histone proteins. It is a mechanism for defending host genomes against the effects of transposable element, viral infection and acts as a modulator of expression of duplicated gene family members and as a silencer of transgenes. A major breakthrough in understanding the mechanism of epigenetic silencing was discovery of silencing in transgenic tobacco plants due to interaction between two homologous promoters. The molecular mechanism of epigenetic mechanism is highly complicated and it is not completely understood yet. Two different molecular routes have been proposed for this, i.e. transcriptional gene silencing (TGS, which is associated with heavy methylation of promoter regions and blocks the transcription of transgene. The basic mechanism underlying post-transcriptional gene silencing (PTGS is degradation of the cytosolic mRNA of transgenes or endogenous genes. Undesired transgene silencing is of a major concern in transgenic technology used in crop improvement. A complete understanding of this phenomenon will be very useful for transgenic applications, where silencing of specific genes are required. The current status of epigenetic silencing in transgenic technology has been discussed and summarized in this mini-review.

  19. Epigenetic silencing in transgenic plants

    Science.gov (United States)

    Rajeevkumar, Sarma; Anunanthini, Pushpanathan; Sathishkumar, Ramalingam

    2015-01-01

    Epigenetic silencing is a natural phenomenon in which the expression of genes is regulated through modifications of DNA, RNA, or histone proteins. It is a mechanism for defending host genomes against the effects of transposable elements and viral infection, and acts as a modulator of expression of duplicated gene family members and as a silencer of transgenes. A major breakthrough in understanding the mechanism of epigenetic silencing was the discovery of silencing in transgenic tobacco plants due to the interaction between two homologous promoters. The molecular mechanism of epigenetic mechanism is highly complicated and it is not completely understood yet. Two different molecular routes have been proposed for this, that is, transcriptional gene silencing, which is associated with heavy methylation of promoter regions and blocks the transcription of transgenes, and post-transcriptional gene silencing (PTGS), the basic mechanism is degradation of the cytosolic mRNA of transgenes or endogenous genes. Undesired transgene silencing is of major concern in the transgenic technologies used in crop improvement. A complete understanding of this phenomenon will be very useful for transgenic applications, where silencing of specific genes is required. The current status of epigenetic silencing in transgenic technology is discussed and summarized in this mini-review. PMID:26442010

  20. Transgenic Mouse Model for Reducing Oxidative Damage in Bone

    Science.gov (United States)

    Schreurs, A.-S.; Torres, S.; Truong, T.; Kumar, A.; Alwood, J. S.; Limoli, C. L.; Globus, R. K.

    2014-01-01

    parameters. Taken together, this typically catabolic treatment (disuse and irradiation) appeared to stimulate cortical expansion in MCAT mice but not WT mice. In conclusion, these results reveal the importance of mitochondrial ROS generation in skeletal remodeling and show that MCAT mice provide a useful animal model for bone studies.

  1. Derivation of Transgene-Free Rat Induced Pluripotent Stem Cells Approximating the Quality of Embryonic Stem Cells.

    Science.gov (United States)

    Li, Shuping; Lan, He; Men, Hongsheng; Wu, Yuanyuan; Li, Ning; Capecchi, Mario R; Bryda, Elizabeth C; Wu, Sen

    2016-09-13

    : Although a variety of reprogramming strategies have been reported to create transgene-free induced pluripotent stem (iPS) cells from differentiated cell sources, a fundamental question still remains: Can we generate safe iPS cells that have the full spectrum of features of corresponding embryonic stem (ES) cells? Studies in transgene-free mouse iPS cells have indicated a positive answer to this question. However, the reality is that no other species have a derived transgene-free iPS cell line that can truly mimic ES cell quality. Specifically, critical data for chimera formation and germline transmission are generally lacking. To date, the rat is the only species, other than the mouse, that has commonly recognized authentic ES cells that can be used for direct comparison with measure features of iPS cells. To help find the underlying reasons of the current inability to derive germline-competent ES/iPS cells in nonrodent animals, we first used optimized culture conditions to isolate and establish rat ES cell lines and demonstrated they are fully competent for chimeric formation and germline transmission. We then used episomal vectors bearing eight reprogramming genes to improve rat iPS (riPS) cell generation from Sprague-Dawley rat embryonic fibroblasts. The obtained transgene-free riPS cells exhibit the typical characteristics of pluripotent stem cells; moreover, they are amenable to subsequent genetic modification by homologous recombination. Although they can contribute significantly to chimeric formation, no germline transmission has been achieved. Although this partial success in achieving competency is encouraging, it suggests that more efforts are still needed to derive ground-state riPS cells. To date, no species other than the mouse have derived transgene-free induced pluripotent stem (iPS) cells that can truly mimic ES cell quality. In the current study, episomal vectors were used to obtain rat transgene-free iPS cells, which contributed to chimeric

  2. Recombinant human factor IX produced from transgenic porcine milk.

    Science.gov (United States)

    Lee, Meng-Hwan; Lin, Yin-Shen; Tu, Ching-Fu; Yen, Chon-Ho

    2014-01-01

    Production of biopharmaceuticals from transgenic animal milk is a cost-effective method for highly complex proteins that cannot be efficiently produced using conventional systems such as microorganisms or animal cells. Yields of recombinant human factor IX (rhFIX) produced from transgenic porcine milk under the control of the bovine α-lactalbumin promoter reached 0.25 mg/mL. The rhFIX protein was purified from transgenic porcine milk using a three-column purification scheme after a precipitation step to remove casein. The purified protein had high specific activity and a low ratio of the active form (FIXa). The purified rhFIX had 11.9 γ-carboxyglutamic acid (Gla) residues/mol protein, which approached full occupancy of the 12 potential sites in the Gla domain. The rhFIX was shown to have a higher isoelectric point and lower sialic acid content than plasma-derived FIX (pdFIX). The rhFIX had the same N-glycosylation sites and phosphorylation sites as pdFIX, but had a higher specific activity. These results suggest that rhFIX produced from porcine milk is physiologically active and they support the use of transgenic animals as bioreactors for industrial scale production in milk.

  3. Recombinant Human Factor IX Produced from Transgenic Porcine Milk

    Directory of Open Access Journals (Sweden)

    Meng-Hwan Lee

    2014-01-01

    Full Text Available Production of biopharmaceuticals from transgenic animal milk is a cost-effective method for highly complex proteins that cannot be efficiently produced using conventional systems such as microorganisms or animal cells. Yields of recombinant human factor IX (rhFIX produced from transgenic porcine milk under the control of the bovine α-lactalbumin promoter reached 0.25 mg/mL. The rhFIX protein was purified from transgenic porcine milk using a three-column purification scheme after a precipitation step to remove casein. The purified protein had high specific activity and a low ratio of the active form (FIXa. The purified rhFIX had 11.9 γ-carboxyglutamic acid (Gla residues/mol protein, which approached full occupancy of the 12 potential sites in the Gla domain. The rhFIX was shown to have a higher isoelectric point and lower sialic acid content than plasma-derived FIX (pdFIX. The rhFIX had the same N-glycosylation sites and phosphorylation sites as pdFIX, but had a higher specific activity. These results suggest that rhFIX produced from porcine milk is physiologically active and they support the use of transgenic animals as bioreactors for industrial scale production in milk.

  4. Animal use for science in Europe.

    Science.gov (United States)

    Daneshian, Mardas; Busquet, Francois; Hartung, Thomas; Leist, Marcel

    2015-01-01

    To investigate long-term trends of animal use, the EU animal use statistics from the 15 countries that have been in the EU since 1995 plus respective data from Switzerland were analyzed. The overall number of animals used for scientific purposes in these countries, i.e., about 11 million/year, remained relatively constant between 1995 and 2011, with net increases in Germany and the UK and net decreases in Belgium, Denmark, Italy, Finland, the Netherlands and Sweden. The relatively low and constant numbers of experimental animals used for safety assessment (toxicology, 8%) may be due to the particularly intensive research on alternative methods in this area. The many efficiently working NGOs, multiple initiatives of the European Parliament, and coordinated activities of industry and the European Commission may have contributed to keeping the animal numbers in this field in check. Basic biological science, and research and development for medicine, veterinary and dentistry together currently make up 65% of animal use in science. Although the total numbers have remained relatively constant, consumption of transgenic animals has increased drastically; in Germany transgenic animals accounted for 30% of total animal use in 2011. Therefore, more focus on alternatives to the use of animals in biomedical research, in particular on transgenic animals, will be important in the future. One initiative designed to provide inter-sector information exchange for future actions is the "MEP - 3Rs scientists pairing scheme" initiated in 2015 by CAAT-Europe and MEP Pietikäinen.

  5. Generation of HIV-1 Gag VLPs by transient transfection of HEK 293 suspension cell cultures using an optimized animal-derived component free medium.

    Science.gov (United States)

    Cervera, Laura; Gutiérrez-Granados, Sonia; Martínez, Marta; Blanco, Julià; Gòdia, Francesc; Segura, María Mercedes

    2013-07-20

    Virus-like particles (VLPs) offer great promise as candidates for new vaccine strategies. Large-scale approaches for the manufacturing of HIV-1 Gag VLPs have mainly focused on the use of the baculovirus expression system. In this work, the development and optimization of an HIV-1 Gag VLP production protocol by transient gene expression in mammalian cell suspension cultures is reported. To facilitate process optimization, a Gag-GFP fusion construct enabling the generation of fluorescent VLPs was used. The great majority of Gag-GFP present in cell culture supernatants was shown to be correctly assembled into virus-like particles of the expected size and morphology consistent with immature HIV-1 particles. Medium optimization was performed using design of experiments (DoE). Culture medium supplementation with non-animal derived components including recombinant proteins and lipids of synthetic or non-animal-derived origin resulted in improved HEK 293 cell growth and VLP production. The maximum cell density attained using the optimized Freestyle culture medium was 5.4×10(6)cells/mL in batch mode, almost double of that observed using the unsupplemented medium (2.9×10(6)cells/mL). Best production performance was attained when cells were transfected at mid-log phase (2-3×10(6)cells/mL) with medium exchange at the time of transfection using standard amounts of plasmid DNA and polyethylenimine. By using an optimized production protocol, VLP titers were increased 2.4-fold obtaining 2.8μg of Gag-GFP/mL or 2.7×10(9)VLPs/mL according to ELISA and nanoparticle tracking quantification analyses, respectively. Copyright © 2013 Elsevier B.V. All rights reserved.

  6. Characterization of Fam20C expression in odontogenesis and osteogenesis using transgenic mice

    Institute of Scientific and Technical Information of China (English)

    Er-Xia Du; Xiao-Fang Wang; Wu-Chen Yang; Deborah Kaback; Siu-Pok Yee; Chun-Lin Qin; Anne George; Jian-Jun Hao

    2015-01-01

    Our previous studies have demonstrated that Fam20C promotes differentiation and mineralization of odontoblasts, ameloblasts, osteoblasts and osteocytes during tooth and bone development. Ablation of the Fam20C gene inhibits bone and tooth growth by increasing fibroblast growth factor 23 in serum and causing hypophosphatemia in conditional knockout mice. However, control and regulation of the expression of Fam20C are still unknown. In this study, we generated a transgenic reporter model which expresses green fluorescence protein (GFP) driven by the Fam20C promoter. Recombineering was used to insert a 16 kb fragment of the mouse Fam20C gene (containing the 15 kb promoter and 1.1 kb of exon 1) into a pBluescript SK vector with the topaz variant of GFP and a bovine growth hormone polyadenylation sequence. GFP expression was subsequently evaluated by histomorphometry on cryosections from E14 to adult mice. Fluorescence was evident in the bone and teeth as early as E17.5. The GFP signal was maintained stably in odontoblasts and osteoblasts until 4 weeks after birth. The expression of GFP was significantly reduced in teeth, alveolar bone and muscle by 8 weeks of age. We also observed colocalization of the GFP signal with the Fam20C antibody in postnatal 1-and 7-day-old animals. Successful generation of Fam20C-GFP transgenic mice will provide a unique model for studying Fam20C gene expression and the biological function of this gene during odontogenesis and osteogenesis.

  7. Expression of functional G protein-coupled receptors in photoreceptors of transgenic Xenopus laevis.

    Science.gov (United States)

    Zhang, Li; Salom, David; He, Jianhua; Okun, Alex; Ballesteros, Juan; Palczewski, Krzysztof; Li, Ning

    2005-11-08

    G protein-coupled receptors (GPCRs) constitute the largest superfamily of transmembrane signaling proteins; however, the only known GPCR crystal structure is that of rhodopsin. This disparity reflects the difficulty in generating purified GPCR samples of sufficient quantity and quality. Rhodopsin, the light receptor of retinal rod neurons, is produced in large amounts of homogeneous quality in the vertebrate retina. We used transgenic Xenopus laevis to convert these retina rod cells into bioreactors to successfully produce 20 model GPCRs. The receptors accumulated in rod outer segments and were homogeneously glycosylated. Ligand and [(35)S]GTPgammaS binding assays of the 5HT(1A) and EDG(1) GPCRs confirmed that they were properly folded and functional. 5HT(1A)R was highly purified by taking advantage of the rhodopsin C-terminal immunoaffinity tag common to all GPCR constructs. We have also developed an automated system that can generate hundreds of transgenic tadpoles per day. This expression approach could be extended to other animal model systems and become a general method for the production of large numbers of GPCRs and other membrane proteins for pharmacological and structural studies.

  8. Characterization of Fam20C expression in odontogenesis and osteogenesis using transgenic mice.

    Science.gov (United States)

    Du, Er-Xia; Wang, Xiao-Fang; Yang, Wu-Chen; Kaback, Deborah; Yee, Siu-Pok; Qin, Chun-Lin; George, Anne; Hao, Jian-Jun

    2015-06-26

    Our previous studies have demonstrated that Fam20C promotes differentiation and mineralization of odontoblasts, ameloblasts, osteoblasts and osteocytes during tooth and bone development. Ablation of the Fam20C gene inhibits bone and tooth growth by increasing fibroblast growth factor 23 in serum and causing hypophosphatemia in conditional knockout mice. However, control and regulation of the expression of Fam20C are still unknown. In this study, we generated a transgenic reporter model which expresses green fluorescence protein (GFP) driven by the Fam20C promoter. Recombineering was used to insert a 16 kb fragment of the mouse Fam20C gene (containing the 15 kb promoter and 1.1 kb of exon 1) into a pBluescript SK vector with the topaz variant of GFP and a bovine growth hormone polyadenylation sequence. GFP expression was subsequently evaluated by histomorphometry on cryosections from E14 to adult mice. Fluorescence was evident in the bone and teeth as early as E17.5. The GFP signal was maintained stably in odontoblasts and osteoblasts until 4 weeks after birth. The expression of GFP was significantly reduced in teeth, alveolar bone and muscle by 8 weeks of age. We also observed colocalization of the GFP signal with the Fam20C antibody in postnatal 1- and 7-day-old animals. Successful generation of Fam20C-GFP transgenic mice will provide a unique model for studying Fam20C gene expression and the biological function of this gene during odontogenesis and osteogenesis.

  9. A novel transgenic rat model with a full Alzheimer's-like amyloid pathology displays pre-plaque intracellular amyloid-beta-associated cognitive impairment.

    Science.gov (United States)

    Leon, Wanda Carolina; Canneva, Fabio; Partridge, Vanessa; Allard, Simon; Ferretti, Maria Teresa; DeWilde, Arald; Vercauteren, Freya; Atifeh, Ramtin; Ducatenzeiler, Adriana; Klein, William; Szyf, Moshe; Alhonen, Leena; Cuello, A Claudio

    2010-01-01

    Alzheimer's disease (AD) is a neurodegenerative pathology in which amyloid-beta (Abeta) peptide accumulates in different brain areas leading to deposition of plaques and a progressive decline of cognitive functions. After a decade in which a number of transgenic (Tg) mouse models mimicking AD-like amyloid-deposition pathology have been successfully generated, few rat models have been reported that develop intracellular and extracellular Abeta accumulation, together with impairment of cognition. The generation of a Tg rat reproducing the full AD-like amyloid pathology has been elusive. Here we describe the generation and characterization of a new transgenic rat line, coded McGill-R-Thy1-APP, developed to express the human amyloid-beta precursor protein (AbetaPP) carrying both the Swedish and Indiana mutations under the control of the murine Thy1.2 promoter. The selected mono-transgenic line displays an extended phase of intraneuronal Abeta accumulation, already apparent at 1 week after birth, which is widespread throughout different cortical areas and the hippocampus (CA1, CA2, CA3, and dentate gyrus). Homozygous Tg animals eventually produce extracellular Abeta deposits and, by 6 months of age, dense, thioflavine S-positive, amyloid plaques are detected, associated with glial activation and surrounding dystrophic neurites. The cognitive functions in transgenic McGill-R-Thy1-APP rats, as assessed using the Morris water maze task, were found already altered as early as at 3 months of age, when no CNS plaques are yet present. The spatial cognitive impairment becomes more prominent in older animals (13 months), where the behavioral performance of Tg rats positively correlates with the levels of soluble Abeta (trimers) measured in the cortex.

  10. Generation of Aphid Resistant Transgenic Wheat with aha from Arisaema heterophyllum by Particle Bombardment%用基因枪法获得转异天南星基因aha抗蚜虫小麦

    Institute of Scientific and Technical Information of China (English)

    张彦; 喻修道; 唐克轩; 夏兰琴

    2012-01-01

    Agglutinin is a class of mannose-binding protein, which has detrimental effect on aphids and other pests. In this study, the vector harboring aha cloned from Arisaema heterophyllum directed by rice Rubisco small subunit promoter (rbcS) was constructed. By co-transformed with pAHC20 that harbors bar selection gene, the aha gene was transferred into wheat variety Kenong 199 via bombardment. After induction, regeneration, two rounds of selection and with the conformation by PCR analysis, 42 transgenic plants with foreign aha gene were obtained, average co-transformation efficiency of 2.41%. According to PCR analysis, the segregation of the T1 plants was basically consistent with Mendel's separation. Aphid resistance bioassay was carried out using eight randomly selected transgenic lines by multiple discrimination method. One high-resistant and three low-resistant transgenic lines were identified, accounting for 44.4% of the tested materials. This study has laid a basis for application of aha gene in development of aphid resistant transgenic wheat.%凝集素是一类具有特异糖结合活性的蛋白,对蚜虫等害虫有很强的抗杀作用.利用异天南星凝集素基因aha(Arisaema heterophyllum agglutinin)以及水稻Rubisco小亚基启动子,构建了aha基因植物绿色组织特异性表达载体,并采用基因枪转化方法,与携带bar基因的pAHC20载体共转化到小麦品种科农199中.经过愈伤诱导、再生和筛选以及PCR鉴定,获得aha转基因植株42株,平均转化效率为2.41%.对转aha基因植株后代PCR鉴定表明,T1代转基因植株的分离比例基本符合孟德尔遗传规律.利用室内多目标综合判别法评定抗蚜虫特性,8个T1代转基因株系中有高抗材料1份,低抗材料3份,占参试比例44.4%.本研究为获得抗蚜虫转基因小麦新材料奠定了基础.

  11. [Nuclear transfer of goat somatic cells transgenic for human lactoferrin].

    Science.gov (United States)

    Li, Lan; Shen, Wei; Pan, Qing-Yu; Min, Ling-Jiang; Sun, Yu-Jiang; Fang, Yong-Wei; Deng, Ji-Xian; Pan, Qing-Jie

    2006-12-01

    Transgenic animal mammary gland bioreactors are being used to produce recombinant proteins with appropriate post-translational modifications, and nuclear transfer of transgenic somatic cells is a more powerful method to produce mammary gland bioreactor. Here we describe efficient gene transfer and nuclear transfer in goat somatic cells. Gene targeting vector pGBC2LF was constructed by cloning human lactoferrin (LF) gene cDNA into exon 2 of the milk goat beta-casein gene, and the endogenous start condon was replaced by that of human LF gene. Goat fetal fibroblasts were transfected with linearized pGBC2LF and 14 cell lines were positive according to PCR and Southern blot. The transgenic cells were used as donor cells of nuclear transfer, and some of reconstructed embryos could develop to blastocyst in vitro.

  12. [Safety evaluation of food from transgenic fish and the molecular biological mechanism].

    Science.gov (United States)

    Zhang, Xichun; Yang, Xiaoguang

    2004-03-01

    More progresses have been made in the studying of transgenic fish in China, but the studying work of safety evaluation of food from transgenic fish are started up just now. Compared to plants and animals on the land, it is more difficult to control the mobility of fish and fish can give birth to a large number of offsprings, so the ecological risk or hazard about transgenic fish is more critical than others. Another safety problem is the chimerism which is initiated by the gene transfer methods used in the transgenic fish. Getting sterile triploid transgenic fish and fixed point integration are efficient to solve the two problems above respectively. The solution of the two problems are also the basis of safety evaluation and detection of food from transgenic fish. Up to now, there are little reports on the safety evaluation of transgenic fish including nutritional evaluation and allergic reaction, and there are no basic research on the detection of transgenic fish for the aim of food safety. In brief, it is very urgent to start up the research on the safety evaluation and detection of transgenic fish for the control of food safety.

  13. Mosquito transgenic technologies to reduce Plasmodium transmission.

    Science.gov (United States)

    Fuchs, Silke; Nolan, Tony; Crisanti, Andrea

    2013-01-01

    The ability to introduce genetic constructs of choice into the genome of Anopheles mosquitoes provides a valuable tool to study the molecular interactions between the Plasmodium parasite and its insect host. In the long term, this technology could potentially offer new ways to control vector-borne diseases through the suppression of target mosquito populations or through the introgression of traits that preclude pathogen transmission. Here, we describe in detail protocols for the generation of transgenic Anopheles gambiae mosquitoes based on germ-line transformation using either modified transposable elements or the site-specific PhiC31 recombinase.

  14. Approaches of Establishment of Technological System of Transgenic Sheep%绵羊转基因技术体系构建及研究进展

    Institute of Scientific and Technical Information of China (English)

    刘明军; 张雪梅; 李文蓉; 黄俊成; 汪立芹

    2014-01-01

    Transgenic approach can break-through the barrier of breeds, integrate multiple gene effects and modify the genome based on the requirement of mankind, therefore it exhibits great potentials. Since the first transgenic animal born in 1982, more than ten species of transgenic animals have been reported. The methods to generate transgenic animals were developed from original pronuclear microinjection to nuclear transfer, viral vector transgenesis, and concurrent genome editing. The aims of the transgenic animal are also expanded from establishment of transgenic animal technique to generation of transgenic model, improvement of the productive performance or product quality, as bioreactor to produce high value pharmaceutical proteins, and breeding new animal breeds. Hereby the worldwide approaches, the faced problems, and the tendency to produce transgenic sheep were summarized. The proceedings of generation of transgenic novel breeds or animal models, and the characteristics of transgenic techniques were reviewed. The barrier and bottleneck of transgenic techniques were also addressed. Particularly, the feature, tendency and potential of the concurrently highlighted genome editing technique were comprehensively documented. Moreover, the necessity to establish a technological system of transgenic sheep was elucidated, and the present status, gain of innovation and breakthrough, highlighted target of transgenic sheep in domestic in future were reviewed. Meanwhile, the outlook of the economic, ecological and social effects resulted from transgenic sheep were further analyzed and previewed as well.%转基因技术能够突破种间隔离、实现多基因聚合、按照人类需要改造生物基因组,因而具有广阔的应用发展前景。自从1982年首例转基因动物诞生以来,已先后报道了10余种转基因动物。转基因方法也由早期单一的原核显微注射发展到核移植转基因、病毒载体转基因和基因组编辑技术。转

  15. Tongue Epithelium Cells from shRNA Mediated Transgenic Goat Show High Resistance to Foot and Mouth Disease Virus.

    Science.gov (United States)

    Li, Wenting; Wang, Kejun; Kang, Shimeng; Deng, Shoulong; Han, Hongbing; Lian, Ling; Lian, Zhengxing

    2015-12-16

    Foot and mouth disease induced by foot and mouth disease virus (FMDV) is severe threat to cloven-hoofed domestic animals. The gene 3Dpol in FMDV genome encodes the viral RNA polymerase, a vital element for FMDV replication. In this study, a conserved 3D-7414shRNA targeting FMDV-3Dpol gene was designed and injected into pronuclear embryos to produce the transgenic goats. Sixty-one goats were produced, of which, seven goats positively integrated 3D-7414shRNA. Loss of function assay demonstrated that siRNA effectively knockdown 3Dpol gene in skin epithelium cells of transgenic goats. Subsequently, the tongue epithelium cells from transgenic and non-transgenic goats were infected with FMDV O/YS/CHA/05 strain. A significant decrease of virus titres and virus copy number was observed in cells of transgenic goats compared with that of non-transgenic goats, which indicated that 3D-7414siRNA inhibited FMDV replication by interfering FMDV-3Dpol gene. Furthermore, we found that expression of TLR7, RIG-I and TRAF6 was lower in FMDV infected cells from transgenic goats compared to that from non-transgenic goats, which might result from lower virus copy number in transgenic goats' cells. In conclusion, we successfully produced transgenic goats highly expressing 3D-7414siRNA targeting 3Dpol gene, and the tongue epithelium cells from the transgenic goats showed effective resistance to FMDV.

  16. Heterologous expression in transgenic mosquitoes

    Institute of Scientific and Technical Information of China (English)

    Santhosh P K; Yu hua Deng; Weidong Gu; Xiaoguang Chen

    2010-01-01

    Arthropod-borne diseases such as malaria and dengue virus afflict billions of people worldwide imposing major economic and social burdens. Control of such pathogens is mainly performed by vector management and treatment of affected individuals with drugs. The failure of these conventional approaches due to emergence of insecticide-resistant insects and drug-resistant parasites demonstrate the need of novel and efficacious control strategies to combat these diseases. Genetic modification(GM) of mosquito vectors to impair their ability to be infected and transmit pathogens has emerged as a new strategy to reduce transmission of many vector-borne diseases and deliver public health gains. Several advances in developing transgenic mosquitoes unable to transmit pathogens have gained support, some of them attempt to manipulate the naturally occurring endogenous refractory mechanisms, while others initiate the identification of an exogenous foreign gene which disrupt the pathogen development in insect vectors. Heterologous expression of transgenes under a native or heterologous promoter is important for the screening and effecting of the transgenic mosquitoes. The effect of the transgene on mosquito fitness is a crucial parameter influencing the success of this transgenic approach. This review examines these two aspects and describes the basic research work that has been accomplished towards understanding the complex relation between the parasite and its vector and focuses on recent advances and perspectives towards construction of transgenic mosquitoes refractory to vector-borne disease transmission.

  17. Expression of spearmint limonene synthase in transgenic spike lavender results in an altered monoterpene composition in developing leaves.

    Science.gov (United States)

    Muñoz-Bertomeu, Jesús; Ros, Roc; Arrillaga, Isabel; Segura, Juan

    2008-01-01

    We generated transgenic spike lavender (Lavandula latifolia) plants constitutively expressing the limonene synthase (LS) gene from spearmint (Mentha spicata), encoding the LS enzyme that catalyzes the synthesis of limonene from geranyl diphosphate. Overexpression of the LS transgene did not consistently affect monoterpene profile in pooled leaves or flowers from transgenic T(0) plants. Analyses from cohorts of leaves sampled at different developmental stages showed that essential oil accumulation in transgenic and control plants was higher in developing than in mature leaves. Furthermore, developing leaves of transgenic plants contained increased limonene contents (more than 450% increase compared to controls) that correlated with the highest transcript accumulation of the LS gene. The levels of other monoterpene pathway components were also significantly altered. T(0) transgenic plants were grown for 2 years, self-pollinated, and the T(1) seeds obtained. The increased limonene phenotype was maintained in the progenies that inherited the LS transgene.

  18. The transgenic rabbit as model for human diseases and as a source of biologically active recombinant proteins.

    Science.gov (United States)

    Bosze, Zs; Hiripi, L; Carnwath, J W; Niemann, H

    2003-10-01

    Until recently, transgenic rabbits were produced exclusively by pronuclear microinjection which results in additive random insertional transgenesis; however, progress in somatic cell cloning based on nuclear transfer will soon make it possible to produce rabbits with modifications to specific genes by the combination of homologous recombination and subsequent prescreening of nuclear donor cells. Transgenic rabbits have been found to be excellent animal models for inherited and acquired human diseases including hypertrophic cardiomyopathy, perturbed lipoprotein metabolism and atherosclerosis. Transgenic rabbits have also proved to be suitable bioreactors for the production of recombinant protein both on an experimental and a commercial scale. This review summarizes recent research based on the transgenic rabbit model.

  19. Neuron Loss in Transgenic Mouse Models of Alzheimer's Disease

    Directory of Open Access Journals (Sweden)

    Oliver Wirths

    2010-01-01

    Full Text Available Since their initial generation in the mid 1990s, transgenic mouse models of Alzheimers's disease (AD have been proven to be valuable model systems which are indispensable for modern AD research. Whereas most of these models are characterized by extensive amyloid plaque pathology, inflammatory changes and often behavioral deficits, modeling of neuron loss was much less successful. The present paper discusses the current achievements of modeling neuron loss in transgenic mouse models based on APP/Aβ and Tau overexpression and provides an overview of currently available AD mouse models showing these pathological alterations.

  20. Human lactoferrin transgenic rabbits produced efficiently using dimethylsulfoxide-sperm-mediated gene transfer.

    Science.gov (United States)

    Li, Lan; Shen, Wei; Min, Lingjiang; Dong, Huansheng; Sun, Yujiang; Pan, Qingjie

    2006-01-01

    Transgenic animal mammary gland bioreactors are used to produce recombinant proteins. However, it is difficult to validate whether these transgenic domestic animals are able to express the recombinant protein efficiently in their mammary glands before the birth of transgenic offspring. In the present study, a simple and efficient method was established to evaluate the functionality of animal mammary gland tissue-expressed cassettes. The gene transfer vector pGBC2LF was constructed, and the expression of human lactoferrin (LF) gene was controlled by the goat beta-casein gene 5' flanking sequence. To obtain the most efficient transfection, the influence of DNA concentration, dimethylsulfoxide (DMSO) concentration, and the ratio of linear-to-circular DNA required for associating DNA with spermatozoa were evaluated. Transfection of exogenous DNA into rabbit spermatozoa was found to be efficient using 30 microg mL(-1) DNA, DMSO at a final concentration of 3%, and a 3 : 1 ratio of linear-to-circular DNA, with 29 of 85 (34.1%) in vitro-fertilised embryos being transgenic. Using DMSO-sperm-mediated gene transfer (DMSO-SMGT), 89 rabbit offspring were produced, with 46 of these (57.1%) being transgenic. As mammary gland bioreactor models, 17 of 21 (81%) transgenic female rabbits could express human LF protein in their glands. During lactation of the transgenic rabbits, the highest level of human LF protein expressed was 153 +/- 31 microg mL(-1), and the mean expression level in all of the transgenic rabbits was 103 +/- 20 microg mL(-1) in the third week, declining gradually after this time. Our results demonstrate that transgenic rabbits produced by DMSO-SMGT were able to express human LF protein in the correct tissue.

  1. Modeling Alzheimer's disease in transgenic rats.

    Science.gov (United States)

    Do Carmo, Sonia; Cuello, A Claudio

    2013-10-25

    Alzheimer's disease (AD) is the most common form of dementia. At the diagnostic stage, the AD brain is characterized by the accumulation of extracellular amyloid plaques, intracellular neurofibrillary tangles and neuronal loss. Despite the large variety of therapeutic approaches, this condition remains incurable, since at the time of clinical diagnosis, the brain has already suffered irreversible and extensive damage. In recent years, it has become evident that AD starts decades prior to its clinical presentation. In this regard, transgenic animal models can shed much light on the mechanisms underlying this "pre-clinical" stage, enabling the identification and validation of new therapeutic targets. This paper summarizes the formidable efforts to create models mimicking the various aspects of AD pathology in the rat. Transgenic rat models offer distinctive advantages over mice. Rats are physiologically, genetically and morphologically closer to humans. More importantly, the rat has a well-characterized, rich behavioral display. Consequently, rat models of AD should allow a more sophisticated and accurate assessment of the impact of pathology and novel therapeutics on cognitive outcomes.

  2. Animal-derived pharmaceutical proteins.

    Science.gov (United States)

    Redwan, el-Rashdy M

    2009-01-01

    Livestock animals have made a significant contribution to human health and well-being throughout humankind's history. A significant contribution of farm animals to human health are the longstanding use of bovine and porcine for production of insulin (for treatment of diabetes), gelatin (for pharmaceutical and other purposes), as well as horse and sheep antibody against natural venoms, toxins, drugs and microbial peptides. Gelatin being the biggest animal protein consumed in human health, follows with antibodies fragments. The chronic problem of animal-derived therapeutics, especially those of high molecular weight, is the immunogenicity induction in addition to their biosafety. However, the invertebrates and lower vertebrates donate the human being a several crucial emergency saving life small-peptides or their analogs such as Refludan, Prialt, Exendin. Not only, but the farm animals are enormously using as models for novel surgical strategies, testing of biodegradable implants and sources of tissue replacements, such as skin and heart valves. Recently, they are being harnessing as bioreactor for production of biopharmaceutical related products through gene farming with efficiency far greater than any conventional microbial or cell-culture production systems. Only 16 transgenic cows would be covering the worldwide needs from human growth hormone. The transgenic, especially animal, technology would be solving a several biopharmaceutical products disadvantages, such as cost, biosafety, immunogenicity and the availability dimensions.

  3. Transgenic Mice for cGMP Imaging

    Science.gov (United States)

    Thunemann, Martin; Wen, Lai; Hillenbrand, Matthias; Vachaviolos, Angelos; Feil, Susanne; Ott, Thomas; Han, Xiaoxing; Fukumura, Dai; Jain, Rakesh K.; Russwurm, Michael; de Wit, Cor; Feil, Robert

    2014-01-01

    Rationale Cyclic GMP (cGMP) is an important intracellular signaling molecule in the cardiovascular system, but its spatiotemporal dynamics in vivo is largely unknown. Objective To generate and characterize transgenic mice expressing the fluorescence resonance energy transfer–based ratiometric cGMP sensor, cGMP indicator with an EC50 of 500 nmol/L (cGi500), in cardiovascular tissues. Methods and Results Mouse lines with smooth muscle–specific or ubiquitous expression of cGi500 were generated by random transgenesis using an SM22α promoter fragment or by targeted integration of a Cre recombinase–activatable expression cassette driven by the cytomegalovirus early enhancer/chicken β-actin/β-globin promoter into the Rosa26 locus, respectively. Primary smooth muscle cells isolated from aorta, bladder, and colon of cGi500 mice showed strong sensor fluorescence. Basal cGMP concentrations were 3 µmol/L could also be monitored in blood vessels of the isolated retina and in the cremaster microcirculation of anesthetized mice. Moreover, with the use of a dorsal skinfold chamber model and multiphoton fluorescence resonance energy transfer microscopy, nitric oxide–stimulated vascular cGMP signals associated with vasodilation were detected in vivo in an acutely untouched preparation. Conclusions These cGi500 transgenic mice permit the visualization of cardiovascular cGMP signals in live cells, tissues, and mice under normal and pathological conditions or during pharmacotherapy with cGMP-elevating drugs. PMID:23801067

  4. Lessons Learned from the Transgenic Huntington's Disease Rats

    Directory of Open Access Journals (Sweden)

    Rinske Vlamings

    2012-01-01

    Full Text Available Huntington's disease (HD is a fatal inherited disorder leading to selective neurodegeneration and neuropsychiatric symptoms. Currently, there is no treatment to slow down or to stop the disease. There is also no therapy to effectively reduce the symptoms. In the investigation of novel therapies, different animal models of Huntington's disease, varying from insects to nonhuman primates, have been created and used. Few years ago, the first transgenic rat model of HD, carrying a truncated huntingtin cDNA fragment with 51 CAG repeats under control of the native rat huntingtin promoter, was introduced. We have been using this animal model in our research and review here our experience with the behavioural, neurophysiological, and histopathological phenotype of the transgenic Huntington's disease rats with relevant literature.

  5. Glyphosate drift promotes changes in fitness and transgene gene flow in canola (Brassica napus) and hybrids

    Science.gov (United States)

    Londo, Jason P.; Bautista, Nonnatus S.; Sagers, Cynthia L.; Lee, E. Henry; Watrud, Lidia S.

    2010-01-01

    Background and Aims With the advent of transgenic crops, genetically modified, herbicide-resistant Brassica napus has become a model system for examining the risks and potential ecological consequences of escape of transgenes from cultivation into wild compatible species. Escaped transgenic feral B. napus and hybrids with compatible weedy species have been identified outside of agriculture and without the apparent selection for herbicide resistance. However, herbicide (glyphosate) exposure can extend beyond crop field boundaries, and a drift-level of herbicide could function as a selective agent contributing to increased persistence of transgenes in the environment. Methods The effects of a drift level (0·1 × the field application rate) of glyphosate herbicide and varied levels of plant competition were examined on plant fitness-associated traits and gene flow in a simulated field plot, common garden experiment. Plants included transgenic, glyphosate-resistant B. napus, its weedy ancestor B. rapa, and hybrid and advanced generations derived from them. Key Results The results of this experiment demonstrate reductions in reproductive fitness for non-transgenic genotypes and a contrasting increase in plant fitness for transgenic genotypes as a result of glyphosate-drift treatments. Results also suggest that a drift level of glyphosate spray may influence the movement of transgenes among transgenic crops and weeds and alter the processes of hybridization and introgression in non-agronomic habitats by impacting flowering phenology and pollen availability within the community. Conclusions The results of this study demonstrate the potential for persistence of glyphosate resistance transgenes in weedy plant communities due to the effect of glyphosate spray drift on plant fitness. Additionally, glyphosate drift has the potential to change the gene-flow dynamics between compatible transgenic crops and weeds, simultaneously reducing direct introgression into weedy species

  6. Standing genetic variation and compensatory evolution in transgenic organisms: a growth-enhanced salmon simulation.

    Science.gov (United States)

    Ahrens, Robert N M; Devlin, Robert H

    2011-06-01

    Genetically modified strains usually are generated within defined genetic backgrounds to minimize variation for the engineered characteristic in order to facilitate basic research investigations or for commercial application. However, interactions between transgenes and genetic background have been documented in both model and commercial agricultural species, indicating that allelic variation at transgene-modifying loci are not uncommon in genomes. Engineered organisms that have the potential to allow entry of transgenes into natural populations may cause changes to ecosystems via the interaction of their specific phenotypes with ecosystem components and services. A transgene introgressing through natural populations is likely to encounter a range of natural genetic variation (among individuals or sub-populations) that could result in changes in phenotype, concomitant with effects on fitness and ecosystem consequences that differ from that seen in the progenitor transgenic strain. In the present study, using a growth hormone transgenic salmon example, we have modeled selection of modifier loci (single and multiple) in the presence of a transgene and have found that accounting for genetic background can significantly affect the persistence of transgenes in populations, potentially reducing or reversing a "Trojan gene" effect. Influences from altered life history characteristics (e.g., developmental timing, age of maturation) and compensatory demographic/ecosystem controls (e.g., density dependence) also were found to have a strong influence on transgene effects. Further, with the presence of a transgene in a population, genetic backgrounds were found to shift in non-transgenic individuals as well, an effect expected to direct phenotypes away from naturally selected optima. The present model has revealed the importance of understanding effects of selection for background genetics on the evolution of phenotypes in populations harbouring transgenes.

  7. Production of transgenic mice by random recombination of targeted genes in female germline stem cells

    Institute of Scientific and Technical Information of China (English)

    Yong Zhang; Ji Xiong; Jie Xiang; Ji Wu; Zhaojuan Yang; Yunze Yang; Shuzeng Wang; Lingjun Shi; Wenhai Xie; Kejing Sun; Kang Zou; Lei Wang

    2011-01-01

    Oocyte production in most mammalian species is believed to cease before birth. However, this idea has been challenged with the finding that postnatal mouse ovaries possess mitotically active germ cells. A recent study showed that female germline stem cells (FGSCs) from adult mice were isolated, cultured long term and produced oocytes and progeny after transplantation into infertile mice. Here, we demonstrate the successful generation of transgenic or gene knock-down mice using FGSCs. The FGSCs from ovaries of 5-day-old and adult mice were isolated and either infected with recombinant viruses carrying green fluorescent protein, Oocyte-G1 or the mouse dynein axonemal intermediate chain 2 gene, or transfected with the Oocyte-G1 specific shRNA expression vector (pRS shOocyte-G1 vector), and then transplanted into infertile mice. Transplanted cells in the ovaries underwent oogenesis and produced heterozygous offspring after mating with wild-type male mice. The offspring were genetically characterized and the biological functions of the transferred or knock-down genes were investigated. Efficiency of genetransfer or gene knock-down was 29%-37% and it took 2 months to produce transgenic offspring. Gene manipulation of FGSCs is a rapid and efficient method of animal transgenesis and may serve as a powerful tool for biomedical science and biotechnology.

  8. Transgenic Tobacco Expressing a Modified VP6 Gene Protects Mice Against Rotavirus Infection

    Institute of Scientific and Technical Information of China (English)

    Jiang-Li DONG; Bo ZHOU; Gang SHENG; Tao WANG

    2005-01-01

    Elevated expression of the rotavirus VP6 antigen in transgenic plants is a critical factor in the development of a safe and effective rotavirus vaccine. Using codon optimization, a gene that encodes the inner capsid protein VP6 of the human group A rotavirus was synthesized (sVP6). The VP6 and sVp6genes were transformed into tobacco (Nicotiana tabacum L.) plants using Agrobacterium tumefaciens. The expression level of the sVP6 gene in transgenic plants was 3.8-34-fold higher than that of controls containing the non-modified VP6 gene, accounting for up to 0.34% of the total soluble protein (TSP). Then, BALB/c female mice that had been gavaged weekly with 10 mg TSP containing 34 μg VP6 protein, in which VP6-specific serum IgG and mucosal IgA antibodies were investigated. The severity and duration of diarrhea caused by simian rotavirus SA-11 challenge were reduced significantly in passively immunized pups, which indicates that anti-VP6 antibodies generated in orally immunized female mice can be passed onto pups and provide heterotypic protection. An edible vaccine based on the VP6 of human rotavirus group A could provide a means to protect children and young animals from severe acute diarrhea.

  9. Characterization of expression of Puumala virus nucleocapsid protein in transgenic plants.

    Science.gov (United States)

    Khattak, Shahryar; Darai, Gholamreza; Süle, Sandor; Rösen-Wolff, Angela

    2002-01-01

    Transgenic plants expressing a foreign gene are a suitable system for the production of relevant immunogens in high amounts that can be used for the development of a new generation of vaccines against a variety of infectious diseases. In the present study, the expression of the nucleocapsid (N) protein of hantavirus serotype Puumala in tobacco and potato plants was investigated. Transgenic tobacco and potato plants were generated and established. These transgenic plants expressed the N protein of Puumala virus strain CG-1820. No major differences were observed when the phenotype and growth rates of transgenic plants were compared to those of normal plants. However, it was found that the leaves of transgenic tobacco plants were more slender and the tubers of transgenic potato plants were smaller than those in normal plants. In order to investigate the distribution of the expression of the foreign gene in transgenic plants, the proteins of leaves and roots of the individual transgenic tobacco and potato plants were examined by Western blot analyses. It was found that all transgenic tobacco and potato plants expressed the N protein in the leaves, whereas transgenic potato plants are able to significantly express the viral proteins also in the tubers and roots. The antigens were expressed at a level of 1 ng of protein/5 microg of dried leaves. The hantaviral recombinant N proteins obtained from transgenic tobacco and potato plants were able to elicit specific humoral and mucosal immune responses when administered intraperitoneally or orally to rabbits and mice. The expression of viral proteins in plants has two major advantages compared to other expression systems: firstly, there is no risk of contamination with mammalian viruses or other pathogens, and secondly, the production of high amounts of antigens is cheap and therefore of great economic interest.

  10. Mutagenicity testing with transgenic mice. Part I: Comparison with the mouse bone marrow micronucleus test

    OpenAIRE

    Wahnschaffe U; Bitsch A; Kielhorn J; Mangelsdorf I

    2005-01-01

    Abstract As part of a larger literature study on transgenic animals in mutagenicity testing, test results from the